130S PDF

Effect of Wheat Flour Extraction Rates
on Physico-Chemical Characteristics of
Sourdough Flat Bread
By

GHULAM MUEEN-UD-DIN
M.Sc. (Hons.) Food Technology
A dissertation submitted in partial fulfillment of the requirements

for the degree of
Doctor of Philosophy
In
Food Technology
NATIONAL INSTITUTE OF FOOD SCIENCE AND TECHNOLOGY

UNIVERSITY OF AGRICULTURE
FAISALABAD-PAKISTAN
2009
To
The Controller of Examinations,
University of Agriculture,
Faisalabad-Pakistan
“We, the Supervisory Committee, certify that the contents and form of thesis
submitted by Ghulam Mueen-ud-Din, Regd. No. 96-ag-1723, have been found
satisfactory and recommend that it be processed for evaluation, by the External
Examiner(s) for the award of degree”.
Supervisory Committee
1. Chairman _________________________________
(Prof. Dr. Salim-ur-Rehman)
2. Member _________________________________
(Prof. Dr. Faqir Muhammad Anjum)
3. Member _________________________________
(Dr. Haq Nawaz)
CONTENTS
No. Title Page
Abstract i
Acknowledgments ii
List of Tables iii
List of Figures v
1 INTRODUCTION 01
2 REVIEW OF LITERATURE 07
2.1 Flat breads 08
2.2 Wheat and its composition 09
2.3 Wheat flour extraction rate 11
2.4 Sourdoughs 13
2.4.1 Types and classification of sourdough 14
2.4.2Use of sourdough in cereal products 15
2.4.3Sourdough fermenting microorganisms 16
2.4.4Yeast in sourdough 18
2.5 Beneficial effects of sourdough fermentation 20
2.5.1Heath benefits of sourdough bread 20
2.5.2Improving taste and nutritional quality 21
2.5.3Dough structure and bread characteristics 22
2.5.4Microbial spoilage 24
2.6 Flavoring compounds and sourdough fermentation 25
2.6.1Solid Phase Micro extraction (SPME) 27
2.7 Organic acids in sourdough 28
2.8 Sensory evaluation of bread 29
3 MATERIALS AND METHODS 32
3.1 Procurement of wheat varieties 32
3.1.1 Physical characteristics of wheat 32
3.1.2 Thousand kernel weights 32
3.1.3 Test weight 32
3.2 Milling characteristics of wheat 33
3.2.1 Milling of wheat 33
3.2.2 Wet and dry gluten 33
3.2.3 Falling Number (FN) 34
3.2.4 Particle size index (PSI) 34
3.2.5 SDS - sedimentation value 34
3.2.6 Pelshenke value 35
3.3 Proximate analysis of wheat flour 35
3.3.1 Moisture Content 35
3.3.2 Crude protein content 36
No. Title Page
3.3.3 Crude fat content 36
3.3.4 Crude fiber content 37
3.3.5 Ash content 37
3.3.6 Nitrogen Free Extract (NFE) 38
3.4 Rheological characteristics 38
3.4.1 Water absorption 38
3.4.2 Arrival time 38
3.4.3 Dough development time 38
3.4.4 Dough stability 39
3.4.5 Departure time/dough resistance 39
3.4.6 Tolerance index (TI) 39
3.4.7 Softening of the dough 39
3.5 Preparation of flat breads (Naan) 39
3.6 Sensory evaluation 40
3.7 Selection of wheat variety 40
3.8 Flour milling for different extraction rate flours. 40
3.9 Proximate analysis of wheat flours 41
3.10 Rheological characteristics 41
3.11 Sourdough starter cultures 41
3.11.1 Mother sponge 41
3.11.2 Sourdough preparation 42
3.11.3 Acid content analysis 42
3.11.4 Microbial content 42
3.12 Sourdough volatile compounds 43
3.12.1 Description of Dynamic Headspace analysis 44
3.12.2 Solid Phase Microextraction (SPME) method 45
3.13 Preparation of sourdough naan 46
3.14 Organic acids in Sourdough naan 46
3.15 Volatile compounds in sourdough naan 46
3.16 Minerals content of sourdough naan 47
3.16.1 Phosphorous content in sourdough naan 47
3.17 Sensory evaluation 48
3.18 Statistical analysis 48
4 RESULTS AND DISCUSSION 49
4.1. Suitability of different wheat varieties for the production
of yeast leavened flat bread (naan) 49
4.1.1. Objective 49
4.1.2. Results 50
4.1.2.1. Physico-chemical characteristics of wheat
varieties 50
No. Title Page
4.1.2.1.a. Thousand kernel weight and test 50
weight of wheat varieties
4.1.2.1.b. Milling of wheat 50
4.1.2.1.c. Wet gluten (WG) and dry gluten (DG)
of flour 50
4.1.2.1.d. Falling number (FN) of flour 53
4.1.2.1.e. Particle size index (PSI) of flour 53
4.1.2.1.f. SDS-Sedimentation (SDS-S) value of
flour 53
4.1.2.1.g. Pelshenke value (PV) of flour 54
4.1.2.1.h. Proximate analysis of straight grade
flour (SGF) of wheat varieties 54
4.1.2.2. Farinographic characteristics of straight grade
flour (SGF) of wheat varieties 56
4.1.2.3. Sensory evaluation of flat breads (naan)
prepared from straight grade flour (SGF) of
wheat varieties 60
4.1.3. Discussion 63
4.1.4. Conclusion 71
4.2. Effect of flour extraction rates on the quality of
sourdough 71
4.2.1. Objective 71
4.2.2. Results 71
4.2.2.1. Proximate composition of different extraction
rate flours 71
4.2.2.1.a. Moisture content of flour 72
4.2.2.1.b. Crude protein of flour 72
4.2.2.1.c. Crude fat of flour 72
4.2.2.1.d. Crude fiber of flour 72
4.2.2.1.e. Ash content of flour 74
4.2.2.1.f. Nitrogen free extract (NFE) of flour 74
4.2.2.2. Farinographic characteristics of different
extraction rate flours 74
4.2.2.3. Sourdough Preparation 76
4.2.2.3.a. pH and acidity of sourdough prepared
with different extraction rate flours and
cultures 77
4.2.2.3.b. Bacterial and yeast counts of sourdough
prepared with different extraction rate
flours and cultures 81
No. Title Page
4.3. Sourdough volatile compounds 87
4.3.1. Objective 87
4.3.2. Results 87
4.3.2.1. Sourdough volatile compounds extracted
with SPME technique 87
4.3.2.2. Sourdough volatile compounds extracted
with Dynamic Headspace technique 90
4.3.2.3. Comparison of SPME and DHT for
Sourdough volatile compounds extraction 93
4.4. Analysis of sourdough naan 96
4.4.1. Objective 96
4.4.2. Results 96
4.4.2.1. pH and acidity of sourdough naan 96
4.4.2.2. Organic acids of sourdough naan 100
4.4.2.3. Mineral contents in sourdough naan 105
4.4.2.4. Volatile compounds of sourdough naan 109
4.4.2.4.a. Sourdough naan volatile compounds
extracted with SPME technique 109
4.4.2.4.b. Sourdough naan volatile compounds
extracted with Dynamic Headspace
technique 109
4.4.2.5.c. Comparison of SPME and DHT for
Sourdough naan volatile compounds
extraction 114
4.5. Sensory evaluation of sourdough naan 120
4.5.1. Objective 120
4.5.2. Results 120
4.5.2.1. Color of sourdough naan 120
4.5.2.2. Flavor of sourdough naan 124
4.5.2.3. Taste of sourdough naan 124
4.5.2.4. Texture of sourdough naan 124
4.5.2.5. Flexibility of sourdough naan 125
4.5.2.6. Chewability and overall acceptability of
sourdough naan 125
5 SUMMARY 129
LITERATURE CITED 135
Appendix-I 167
ACKNOWLEDGMENTS
All praises and thanks are for the Almighty Allah, the Merciful, the only creator of
the universe and source of all knowledge and wisdom, who blessed me with health,
thoughts, nice teachers, helping friends and afforded opportunity to complete this study. I
offer my humblest thanks to the Holy Prophet Hazrat Muhammad (Peace be Upon Him),
whose moral and spiritual teachings enlightened my heart, mind and flourished my
thoughts towards achieving high ideals of life.
I am grateful to my supervisor, Professor Dr. Salim-ur-Rehman for his supervision
in planning, execution and scholarly ideas that beautified the scientific nature of the
research work carried out. He always directed to enlighten the ways of life as well.
I am thankful to the members of my supervisory committee Professor
Dr. Faqir Muhammad Anjum, Director General National, Institute of Food Science and
Technology, University of Agriculture, Faisalabad and Dr. Haq Nawaz, Associate
Professor, Institute of Animal Nutrition and Feed Technology for their kind help and
constructive criticism during the course of study for the accomplishment of the work
presented in this manuscript. My special thanks to Prof. Dr. Åse Solvejg Hansen,
Department of Food Science, University of Copenhagen, Copenhagen, Denmark for her
valuable advice and support during the research work. She is such a nice lady.
I am also very grateful to my respectable teacher Dr. Nuzhat Huma, Associate
Professor, National, Institute of Food Science and Technology, University of Agriculture,
Faisalabad and my friend cum teachers Dr. Anjum Murtaza and Dr. Omer Mukhtar Tarar
for their, technical guidance, moral support and prayers to accomplish this study. My
sincere gratitude is to all my loving friends especially Shahid, Asim, Farhan, Haroon,
Anjum, Shehbaz, Niazi, Azhar, Abrar and Waseem.
Last but not least, I wish to express my sincere feelings of gratitude to my loving
and caring parents for their support, moral strength and material welfare through out my
life. I would like to express my deepest gratitude for my affectionate brothers, sisters,
bhabis for their help and support.
Ghulam Mueen-ud-Din
LIST OF TABLES
Tab. No. Title Page

Mean sum of squares for physical characteristics of wheat
1 51
varieties
Mean values for physical characteristics of different wheat
2 51
varieties
Mean sum of squares for general characteristics of SGF from
3 52
different wheat varieties
Mean values for general characteristics of SGF from different
4 52
wheat varieties
Mean sum of squares for proximate composition of SGF from
5 55
Mean values for proximate composition of SGF from different
6 55
wheat varieties
Mean sum of squares for farinographic properties of SGF from
7 57
Mean values for farinographic characteristics of SGF from
8 58
Mean sum of squares for sensory characteristics of flat bread
9 61
naan prepared from SGF of different wheat varieties
Mean values for sensory evaluation of flat breads prepared
10 61
from SGF of different wheat varieties
Mean sum of squares for proximate composition of different
11 73
extraction rate flours
Mean values for proximate composition of different extraction
12 73
rate flours
Mean sum of squares for farinographic characteristics of
13 75
different extraction rate flours
Mean values for farinographic characteristics of different
14 75
Mean sum of squares for pH and acidity of sourdough
15 78
prepared with different extraction rate flours and cultures
Mean values for pH and acidity of sourdough prepared with
16 78
different extraction rate flours and cultures
Mean sum of squares for bacterial and yeast count of
17 sourdough prepared with different extraction rate flours and 82
cultures
Mean values for bacterial and yeast count of sourdough
18 82
Volatile compounds produced by LA-1 and LA-5 cultures in
19 sourdough prepared from flours with different extraction rates 88
(SPME technique)
Volatile compounds produced by LA-1 and La-5 cultures in
20 sourdough prepared from flours with different extraction rates 91
(Dynamic Headspace technique)
Mean sum of squares for pH and acidity of sourdough naan
21 97
Mean values for pH and acidity of sourdough naan prepared
22 97
with different extraction rate flours and cultures
Mean sum of squares for organic acids of sourdough naan
23 101
Mean values for organic acids of sourdough naan prepared
24 101
Mean sum of squares for minerals of sourdough naan prepared
25 106
Mean values for minerals of sourdough naan prepared with
26 107
27 sourdough naan prepared from flours with different extraction 110
rates (SPME technique)
28 sourdough naan prepared from flours with different extraction 112
rates (Dynamic Headspace technique)
Mean sum of squares for sensory characteristics of sourdough
29 121
naan prepared with different extraction rate flours and cultures
Mean values for sensory characteristics of sourdough naan
30 122
LIST OF FIGURES
Fig. No. Title Page
1 Farinograms for different Pakistani wheat varieties 59

Sensory scores for flat breads prepared from different
2 62
wheat varieties
pH of sourdough prepared from flours with different
3 79
extraction rates as influenced by various starter
Acidity of sourdough prepared from flours with different
4 80
pH of sourdough naan prepared from flours with different
5 98
Acidity of sourdough naan prepared from flours with
6 different extraction rates as influenced by various starter 99
cultures
Lactic acid contents of sourdough naan prepared from
7 flours with different extraction rates as influenced by 102
various starter cultures
Acetic acid contents of sourdough naan prepared from
Citric acid contents of sourdough naan prepared from
Sensory characteristics of sourdough naan prepared with
10 123
Abstract
The project was designed to determine the suitability of different wheat varieties
for the production of naan and to evaluate the performance of mixed starter cultures of
yeast and LAB and wheat flour extraction rates on the quality of sourdough naan. On the
basis of physico-chemical, rheological characteristics of wheat flour and sensory
characteristics of the naan, wheat variety AS-2002 were found the most suitable for the
production of naan. Consequently, this variety was milled to get 64, 76, 88 and 100%
extraction rates flour for subsequent studies. The extraction rates significantly influenced
the proximate composition and rheological characteristics of the flour. The sourdoughs
were prepared from different extraction rates flour using freeze dried mixed cultures
namely; LA-1 containing homo-fermentative strain Pediococcus acidilacti and yeast
Saccharomyces cerevisiae and LA-5, containing hetero-fermentative strain L. brevis and
homo-fermentative strain L. casei and yeast Saccharomyces cerevisiae. The pH of the
sourdoughs decreased and acidity increased significantly with an increase in extraction
rate of flour. The sourdoughs were analyzed for volatiles; the compounds extracted were
alcohols, esters, acids, carbonyls and aromatic compounds. The sourdough naans were
prepared by using different extraction rates flour. The production of organic acids
increased with an increase in extraction rate and freeze dried cultures containing hetero-
fermentative strains of LAB showed better performance than homo-fermentative in the
production of organic acids. The sourdough naans prepared with different extraction rates
flour showed highly significant effect on minerals contents. The volatile compounds
extracted from the sourdough naans using SPME and Dynamic Headspace techniques
were mainly identified as alcohols, esters, carbonyls. It was found that hetero-
fermentative strains of LAB produced more volatile compounds than homo-fermentive
and SPME technique extracted volatile compounds with higher relative peak areas than
Dynamic Headspace. The sensory evaluation of naans revealed that the sourdough naans
were preferred over yeast leavened due to their better taste; flavor and chewability. The
sourdough naans prepared from 76% extraction rate flour were awarded the highest
scores for most of the sensory attribute.
Chapter-1
INTRODUCTION
Bread is consumed in large quantity in the world in different types and

forms depending on cultural habits (Cayot, 2007). Flat breads are oldest, most
diverse, and most popular product in the world. It is estimated that over 1.8
billion people consume various kinds of flat breads all over the world. The
popularity of these traditional breads is growing due to ethnic population,
higher demand for exotic, healthy and natural breads (Qarooni, 1996).
Wheat (Triticum aestivum) is the most important crop for making bread,
due to its absolute baking performance in comparison to all other cereals
(Dewettinck et al., 2008). Flat breads produced from wheat flour are served as
source of dietary energy and protein for people, whose daily diet consists of
cereal products. In Pakistan, the annual wheat production stood at 21.74 million
tones during the year 2007-08 (GOP, 2008). Almost 80% of this grain is consumed
in the form of flat breads locally known as chapattis, rotis and naan (Anjum et al.,
1991). It is a staple food and contributes 68-75% of total food intake in daily diet
for the people of Pakistan (Aslam et al., 1982).
Bread products and their production techniques differ widely around the
world (Dewettinck et al., 2008). The objective of bread making is to convert cereal
flours into attractive, palatable and digestible food. The earliest breads were
unleavened or flat, but the first major technical innovation was the introduction
of leavening processes, which yielded breads of superior palatability
(Chamberlain, 1975). The foremost quality characteristics of leavened wheat
breads are high volume, soft and elastic crumb structure, good shelf life and
microbiological safety of the product (Cauvain, 2003)
Based on the cross section of baked products, flat breads can be divided
into two groups; single-layered and double-layered. Single layered flat breads
are the most diverse products which could be either leavened or unleavened. The
unleavened group includes arepa, corn and wheat flour tortillas, paratha, puri,
chapatti and Scandinavian flat breads. The leavened, single layered flat breads
are divided into dough and batter based products. barbari, naan, pizza crust, and
tanoor are the diverse and widely consumed dough based products. The batter
based flat breads include blintzes, crepes, dosai and kisra. The double-layered
flat breads include arabic and baladi bread (Qarooni, 1996).
Naan is the usual commercial flat bread of Pakistan. It is mostly prepared
from lower extraction rate wheat flour and dough is leavened either with yeast
or sodium bicarbonate or both. It is baked mostly in the country restaurants and
small bake shops known as tanoor shops. The baking technique of naan is to
plaster the flattened dough disc on the inside surface of a deep-dug earthen
oven, called a “tanoor” which is heated internally by burning firewood or natural
gas. It is comparatively more nutritious than chapatti/roti, as it is prepared from
fermented dough.
The bacterial fermented products are more palatable having wide
diversity of flavor, aroma, texture and acquire their characteristic properties by
the action of lactic acid bacteria (Hammes, 1990). The microorganisms of genera
Lactococcus, Lactobacillus, Leuconostoc, Streptococcus and Pediococcus are involved in
these fermentations. Lactic acid bacteria (LAB) are the most important bacteria
used for the production of many fermented foods. These foods include pickles,
sausages, cheese, yoghurt and sourdough breads.
The use of sourdough process as a mean of leavening is one of the oldest
biotechnological processes in cereal food production (Rocken and Voysey, 1995).
The sourdough bread is prepared from a mixture of flour and water that is
fermented with (LAB), mainly hetero-fermentative strains, elaborating lactic acid
and acetic acid in the mixture, and hence, resulting a pleasant sour taste end
product (De Vuyst and Neysens, 2004).
The flavor of sourdough wheat bread is richer and more aromatic than
yeast leavened bread (Brummer and Lorenz, 1991). The use of hetero-
fermentative LAB results in the production of flavoring compounds like ethyl
acetate, alcohols and aldehydes, whereas, homo-fermentative LAB produce
diacetyl and other carbonyls. However, on the other hand, yeast fermentation
results in the production of iso-alcohols only, which may contribute little
towards final bread flavor (Rehman et al., 2006). Although, wheat and rye flours
are mostly used for sourdough making but maize and rice flours could also be
used (Buttery and Ling, 1999). The sourdough bread is characterized by a typical
aroma, taste, texture and shelf life. It differs from conventional yeasted bread,
because it contains a predominant bacterial flora (De Vuyst and Neysens, 2004).
Presently, consumers intend to like a wide range of foods that are more
nutritious and flavorful and have longer shelf life having no added preservatives
(Hansen and Hansen, 1996; Collar et al., 1994a). The sourdough fermentation
have a number of beneficial effects that include, a prolong shelf, accelerated
volume, delayed staling, improved bread flavor and nutritional value (Thiele et
al., 2002; Lavermicocca et al., 2000). It has been reported that sourdough
fermentation increases folate and thiamin contents, and decreases tocopherol and
tocotrienol contents depending upon the process of production (Kariluoto et al.,
2004; Liukkonen et al., 2003). It also improves sensory characteristics such as loaf
volume, evenness of baking, color, aroma, taste and texture of breads with
extended shelf life by inhibiting mold growth (Rehman et al., 2007).
The mold growth is the most common cause of microbial spoilage in
bread that results in huge economic losses. Another concern is the chances of
production of mycotoxins produced by the molds which may cause public health
problems. Certain sourdough LAB and their components have antifungal effects
against various fungal species which are isolated from flour and bakery products
(Legan, 1993).
The LAB has a long history of use in foods which are generally recognized
as safe organisms. The representative’s genera of LAB are Lactobacillus,
Leuconostoc, Pediococcus, and Weissella (De Vuyst and Neysens, 2004).
Lactobacillus plantarum, Lactobacillus sanfranciscensis and Lactobacillus brevis are
most widely used LAB in sourdough fermentation. Beside this, several species of
yeast, Sacchromyces cerevisiae and Candida humilis are also added in sourdough.
Like other fermented foods produced by mixed micro flora, sensory, health and
nutritional properties of baked sourdough products depend on joint activity of
yeast and LAB (Gobbetti, 1998). The former is mainly responsible for the
leavening, while the latter are the main cause of dough acidification (Boraam et
al., 1993).
The quality of sourdough bread also depends upon several other factors
such as source of acidification, processes (sourdough or straight) and flour
extraction rates (Martinez-Anaya, 1994). The extraction rate of the flour is one of
the most important factors for determining the properties of sourdough
(Salovaara and Valjakka, 1987).
Wheat flours are characterized by the flour extraction, which is the
proportion of flour by weight, derived by milling from a known quantity of
wheat (Slavin et al., 2000). White flour is refined form having extraction rate of
60-75 %. Brown flour has a higher extraction rate (85%), containing more bran
and this may impart a darker color, strong flavor and aroma to the products.
With an increase in exaction rate, the protein content, fiber, sugar, lipids and
mineral matters increase, whereas, the starch decreases (Kent and Amos, 1967).
Moreover, flour extraction rate affects the protein content, water absorption and
gluten strength (Orth and Mander, 1975). The extraction rate of wheat flour has
marked effect upon its nutritional content. As a result of milling process, nutrient
level of high extraction rate flour increases directly with increase in flour ash
values (Matthews and Workman, 1977). However, on the other hand, the
minerals content is reduced to 30% than the whole wheat flour and also the
concentration of essential nutrients is decreased with lowering the extraction rate
(Pederson and O’Eggum, 1983).
The most common source of dietary fiber in wheat products is bran.
Although, the addition of bran in appropriate level have health benefits but it
causes severe problems in the flavor and texture. The bran fraction is rich in
spoilage organisms such as rope forming bacteria Bacillus subtilis which increases
the challenge to retain microbiological safety of wheat breads. Another example
of the potential of sourdough is the ability to modify the bran fraction of the
grain (rich in fiber) in such a way that higher quantities of bran can be utilized in
the breads (Nielson et al., 2007).
Addition of LAB into dough in a freeze-dried state is one of the preferred
methods used to initiate sourdough fermentation. Freezing or freeze drying has
some advantages over the use of fresh cultures. The most important advantages
of the freeze-dried starter preparations are the excellent storage stability, easy
handling during storage, distribution and other biotechnological properties
(Cossignani et al., 1996). The freeze- dried mixed cultures of LAB and yeasts vary
in their composition in sourdough sponges. In such sponges, yeasts act mainly as
leavening agents, while LAB contributes mainly to the flavoring compounds of
bread. The use of mixed cultures has a number of advantages, including
improvements in flavor and texture and retention of freshness compared to
baker’s yeast bread (Meignen et al., 2001).
Rye sourdough bread production has been studied extensively in Europe,
but no study has been made on the preparation of wheat sourdough naan, a
popular staple food of the people of Afghanistan, Iran, India and Pakistan.
Moreover, no study has been undertaken to determine the flavor profile of naan
bread prepared with different extraction rate flours and LAB.
Freeze-dried cultures containing homo-fermentative strain Pediococcus
acidilacti along with yeast Saccharomyces cerevisiae and mixed cultures containing
hetero-fermentative strain L. brevis and homo-fermentative strain L. casei, along
with yeast Saccharomyces cerevisiae, were used to prepare sourdough naan using
different extraction rate flours. Therefore, this study was planned to investigate
the effect of different extraction rates and mixed cultures of LAB and yeast on the
quality characteristics of sourdough naan.
Objectives:
The main objectives of the study were:-

• To determine the suitability of different wheat varieties for the production
of leavened flat bread (naan) through physico-chemical, rheological and
sensory analyses.
• To determine the effect of extraction rates on the quality of sourdough
naan.
• To evaluate the performance of mixed cultures of yeast and LAB in
sourdough naan.
• To determine organic acids, mineral contents and volatile compounds in
sourdough naan.
• To study the acceptability of sourdough naan through sensory evaluation.
Chapter-2
REVIEW OF LITERATURE
Wheat (Triticum aestivum) is a staple food in many parts of the world. In

Pakistan, 80% of the total wheat produced is used for making chapattis, rotis,
and naan. These are primary and the cheapest source of protein and calories for
the population. (Anjum and Walker, 1991). Its portentous portion named as
gluten, which assists to convert it into a variety of popular baked products. The
naan is leavened flat bread which is prepared from flour of fine granulation by
the addition of yeast alone or with chemical leavening agents. Sourdough
process may be used to prepare various types of good quality breads. However,
quality of sourdough bread depends upon several factors such as source of
acidification, the form of starter addition and flour extraction rates (Martinez-
Anaya, 1994; Brummer and Lorenz, 1991). The sourdough bread is preferred over
yeast leavened bread due to production of organic acids during fermentation
which may exert beneficial effects on sensory properties and keeping quality of
the breads. Moreover, sourdough fermentation may contribute towards
production of flavoring compounds in the baked goods, thus enhancing their
popularity among consumers. The literature regarding these aspects has been
reviewed under following headings:
2.1 Flat breads
2.2 Wheat and its composition

2.3 Wheat flour extraction rate
2.4 Sourdoughs
2.4.1 Types and classification of sourdough
2.4.2 Use of sourdough in cereal products
2.4.3 Sourdough fermenting microorganisms
2.4.4 Yeast in sourdough
2.5 Beneficial effects of sourdough fermentation
2.5.1 Heath benefits of sourdough bread
2.5.2 Improving taste and nutritional quality
2.5.3 Dough structure and bread characteristics
2.5.4 Microbial spoilage
2.6 Flavoring compounds and sourdough fermentation
2.6.1 Solid Phase Micro extraction (SPME)
2.7 Organic acids in sourdough
2.8 Sensory evaluation of bread
2.1 Flat breads

Flat breads are probably the oldest, most diverse, and popular products in
the world with estimated two billions consumers all over the world (Qarooni,
1996). These are prepared from wide varieties of cereal grains that are grounded
into flour, mixed with water into dough, and then usually baked into loaves or
cakes. The flour may be originated from grass seeds such as wheat and rye, from
legumes and tubers, and even from such exotic items as artichoke (Encyclopedia
American, 1994). The craft of bread making existed as evidenced by excavations
undertaken in many parts of the world (Spicher, 1975). The earliest breads were
likely unleavened or flat (Quail, 1996), but the first major technical innovation
was the introduction of leavening, which yielded breads of superior palatability
(Chamberlain, 1975).
Among cereals, wheat is extensively used for the production of flat breads
(Shewry and Tatham, 1994). The unique bread making properties of wheat flour
are mainly due to its gluten proteins to form a viscoelastic network when mixed
with water (Fincher and Stone, 1986; Hoseney et al., 1988). The suitability of
wheat cultivars for application is influenced by genetic, seasonal and
environmental factors. Various wheat varieties have been found suitable for the
preparation of flat breads (Finney et al., 1980). The quality of flat breads is
subjected to a number of variables, such as wheat quality, milling practice,
ingredients, and processing methods.
Naan is leavened flat bread prepared from essential ingredients like
flour, water, salt and yeast (Aidoo et al., 2006). Therefore, the naan is made from
flour of finer granulation than that used for chapattis because finer the
granulation, the more rapid is the process of fermentation (Qarooni, 1996). For its
preparation, the ingredients may include up to 25% yogurt and 6% shortening
and the amount of water may be decreased to 35% (Faridi, 1988).
2.2 Wheat and its composition

Wheat is grown throughout the world and is adaptable to the wide range
of environmental conditions (Kent and Evers, 1994). Wheat based foods are
major source of nutrients in many regions of the world. Although, wheat is
generally used as a source of carbohydrates; but its products are also a
substantive source of protein, vitamins, and minerals, when consumed as a major
component of a diet. It is used extensively in many parts of the world for the
preparation of different types of bread and many other bakery products
(Hoseney et al., 1988; Fincher and Stone, 1986).
Wheat quality is influenced by both genotypic and environmental factors
(Loffler and Busch, 1982) and its flour is analyzed for chemical composition to
evaluate its quality, processing and storage life. However, moisture has
significant effect on the its keeping quality. The flour produced from dry and
sound grains can be kept for longer periods if properly stored, but that from wet
grains deteriorates dramatically within a few days (Pomranz, 1988).
Protein content varies significantly in different wheat varieties. It is the
best single test that can be applied to determine the quality of flour, because it
has a direct correlation with baking quality (Stone and Savin, 1999; Matz, 1996).
The refining of flour greatly affects the protein content as it decreases from 14.2%
at 100% extraction to 12.7% at 66% extraction of flour. It is the response to the
removal of germ and aleurone layer as bran; both are relatively rich in protein
(Pedersen, 1994).
Anjum and Walker (2000) reported protein contents, in six most prevalent
Pakistani bread wheat cultivars that range from 11.99 to 13.80%. Ahmed (1993)
found 12.13% crude protein in whole wheat flour obtained from chakkies.
Rehman et al. (2001) analyzed a few wheat varieties of Sindh province for
composition and found protein that ranges from 11.86 to 11.95%. Butt et al. (2001)
estimated protein content in 30 spring wheat cultivars and reported 10.06 to
11.89% in white flour. Farooq et al. (2001) noted significant (p≤0.01) effect of
varieties and lines on chemical composition of wheat, especially on protein
fractions.
Most of the flours are mixture of soft and hard wheat. Flour from two
manufacturers may consist of different blend, which could produce varying
results in the kitchen (Kent and Evers 1994). Various types of wheat (soft and
semi hard) are also used for the preparation of flat bread (Williams et al., 1986).
Finney et al. (1987) suggested that soft wheat’s are suitable for Egyptian, Iranian
and Indian flat breads. Rao et al. (1986) used flour containing 8.0% moisture,
10.6% protein and 1.45% ash contents for the preparation of chapattis. Hansen
and Ross (1996) revealed that better flat breads could be produced from soft and
medium hard wheat, which contain low to medium protein content with weak
gluten and baking strength.
Ash content is important to determine flour grade and extraction rate,
since the ash content of the bran is about twenty times that of endosperm. It is a
reliable indicator of the efficiency with which the bran and germ are separated
from the endosperm (Matz 1996). The ash predicts the level of bran in the flour.
The ratio of bran to endosperm is higher in small kernels (Posner, 2000). It varies
from one variety to another and in many cases within the same variety grown at
different locations assuming all other variables relevant to kernel structure and
milling technology are constant. Wheat with higher endosperm ash is inherently
bound to give higher flour ash values.
2.3 Wheat flour extraction rate

Wheat flours are characterized by the flour extraction, which may be
defined as the proportion of flour by weight, derived by milling from a known
quantity of wheat (Slavin et al., 2000). The extraction rate is used to define
different types of wheat flours. The flour with less than 75% extraction rate is
typical white. If the extraction rate exceeds 80%, the flour will contain non
endosperm particles, and if the extraction rate approaches 100%, whole meal
flour can be made (Clydesdale, 1994). In the milling process, the grains may be
fractionated into different types of flour. With decreasing extraction rates in
milling, more and more of the outer grain layers are removed resulting in the
loss of dietary fiber and associated bioactive compounds such as vitamins B
group, tryptophan and lignin (Nilsson et al., 1997).
Flat bread quality depends upon a number of variables such as wheat
quality, flour types, extraction rate, ingredients and processing methods.
Moreover, Inherit characteristics of wheat, tempering conditions, and milling
practices cause a significant difference in flour quality (Qarooni, 1996). Also,
extraction rate of wheat flour has marked effect on its nutritional content
(Matthews and Workman, 1977). However, nutrient level increases directly with
flour ash values and nutritional quality of flours produced worldwide is lower
than the wheat as a result of milling process (Barret et al., 1980). The minerals
content reduced to 30% of that in whole wheat and also the concentration of
essential nutrients decreased with lowering the extraction rate (Pederson and
O’Eggum, 1983). However, the principal nutritional benefit of processing is to
increase the bioavailability of the nutrients present in the grain, making the
cereal grain a better substrate for digestive enzymes (Michalska et al., 2007). Also,
the flour extraction rate affects the protein content, farinographic water
absorption and gluten strength (Orth and Mander, 1975). With an increase in
exaction rate, the protein content, fiber, sugar, lipids and mineral matter increase,
whereas the starch decreases (Kent and Amos, 1967). Although, wheat milling
products differ in amino acid composition, but the rate of extraction have no
major effect on protein quality of flours (Pederson et al., 1989). The dietary fiber
content remains constant for extraction rate between 65% and 80% and increases
linearly with extraction rate (Nyman et al., 1984).
With a high extraction rate, the content of nutrients such as B vitamins
and minerals increases. It is due to the buffering capacity of the flour contributed
by phytic acid from the aleurone layer of grains, stimulates the growth and
biochemical activity of microflora in the sourdough, resulting in higher
production of organic acid and flavoring compounds (Hansen and Hansen,
1994). Higher extraction rate of flour results in higher production of lactic and
acetic acids. The final acid values in sourdough made from whole meal flour are
almost double the values in sourdough made from straight grade flours
(Brummer and Lorentz, 1991). Faridi et al. (1981) assessed five wheat varieties of
United States in preparing flat breads like barbri, lavash, tanoor, and sangak with
flour levels of extraction rates i.e. 77%, 82%, 84%, and 87%. Soft wheat varieties
with 77% extraction rate have been found suitable for Iranian flat breads.
It has been observed that by increasing the extraction rate, the amount of
protein, fat, fiber, ash and wet gluten increase and water absorption and color of
wheat flour improve whereas, moisture content, sedimentation value and falling
number decrease. The rheological studies showed a reduction in dough stability,
whereas the dough mixing tolerance index improved with the increase in
extraction rate. Overall quality of Iranian flat breads was affected by flour
extraction rate as indicated by the sensory acceptability test (Azizi et al, 2006).
Increase in the extraction rate from straight grade flour to 80% results in an
average increase of 0.4 g/100g in protein content, 0.2 g/100g in ash and 1.1
g/100g in farinograph water absorption whereas, averages decrease of 1.3 in L-
value of color and 37.4 units of relative viscosity was noticed (Qarooni et al,
1994).
In a similar study conducted by Michalska et al. (2007), the relation
between flour extraction rates and content of bioactive compounds was
investigated. Rye flours with extraction rate of 100% (whole meal flour), 95%
(brown flour), 90% (brown flour) and 70% (light flour) were prepared. It is
concluded that milling process causes a decrease in all bioactive components i.e.
total phenolic content, total flavonoids, inositol hexaphosphate, reduced and
oxidized glutathione, tocopherols and tocotrienols. Brown flour is suggested
most resistant against oxidation processes followed by light and whole meal
flour. The ratio of tocotrienols to tocopherols is the highest in rye flours with
extraction rate of 100–90% whereas light flour was the poorest source of
tocopherols and tocotrienols.
The wheat flour types influence the production of flavoring compounds in
wheat sourdough. It has a significant effect on the production of ethyl acetate
and ethanol in sourdoughs fermented with hetero-fermentative cultures, with
the highest amount detected in sourdoughs made from whole meal flour and
low grade flour (Hansen and Hansen, 1994). A high ash content of flour has been
reported to increase the amount of volatile compounds in mixed fermentations
(Czerny and Schieberle 2002; Hansen and Hansen 1994).
2.4 Sourdough
Fermented foods are of great importance because they provide and
preserve large quantity of nutritious foods with improved aroma and texture.
These foods include alcoholic beverages, vinegar, pickles, sausages, cheese,
yoghurts and sourdough breads. In recent years, sourdough bread has enjoyed
renowned success due to increasing consumer demand in Europe for its natural
taste and good health effects (Brummer and Lorenz, 1991). A mixture of cereals
in water, result in the formation of dough, characterized by sour aroma when left
alone for a while, is the first example of fermented food employed by mankind
(Hammes and Ganzle, 1998). The sourdough is a mixture of flour and water that
is fermented with lactic acid bacteria (LAB), mainly hetero-fermentative strains,
elaborating lactic acid and acetic acid in the mixture, and hence, resulting in a
pleasant sour taste of end product (De Vyust and Neysens, 2004).
The sourdough fermentation is a traditional process for improving the
bread quality and producing different wheat and rye breads (Thiele et al., 2002).
At present the sourdough is employed in the manufacture of breads, cakes and
crackers (Ottogalli, et al., 1996). The typical characteristic of sourdough is mainly
due to its microflora, basically represented by lactic acid bacteria (LAB) and
yeasts. Due to microbial community such dough is metabolically active and can
be reactivated. These microorganisms ensure acid production and leavening
upon addition of flour and water (Anon., 1994).
The mechanisms of sourdough are complex (Hammes and Ganzle, 1998).
Various flour characteristics and process parameters contribute to exercise very
particular effects on the metabolic activity of the sourdough microflora. During
fermentation, biochemical changes occur in the carbohydrate and protein
components of the flour due to the action of microbial and indigenous enzymes.
Moreover, fermentation temperature also influences the growth and metabolism
of LAB and yeast (Spicher et al., 1980).
2.4.1 Types and classification of sourdough

Specifically, sourdoughs are grouped into three types, on the basis of the
technology applied i.e. type I, type II and type III sourdough. The first type
sourdoughs (Type I) are produced in traditional way. To keep the
microorganisms in an active state, these are characterized by continuous, daily
increments. Type II sourdoughs are used as dough-souring supplements during
bread making. These are semi-fluid silo preparations prepared in long
fermentation periods (from 2 to 5 days) and at elevated fermentation
temperature often >30˚C to speed up the activity. Type III sourdoughs are in
dried form, having LAB resistant to the drying process (Hammes and Ganzle,
1998; Bocker et al., 1995).
The dough’s of types II and III require the addition of baker’s yeast
(Saccharomyces cerevisiae) as leavening agent whereas type I sourdoughs does not
require this addition. Sourdough LAB, consisting of obligate and facultative
hetero-fermentative and obligate homo-fermentative species associated with
type I, type II and type III sourdoughs. Type 0 dough, for which baker’s yeast is
the main fermenting agent, is not made with sourdough technology. Bacterial
isolates from, a mature sourdough or other natural environment are selected
and tested for their suitability for being employed as sourdough starters and
their viability after drying. Lyophilized strains of Lactobacillus delbrueckii,
Lactobacillus fructivorans, Lactobacillus plantarum, and Lactobacillus brevis have
been established as sough dough LAB. (Hammes and Ganzle, 1998). In contrast
to the type-1 sourdough starters, frequent inoculation of these strains is required
as these are not well adapted to the cereal environment (Roecken and Voysey,
1995). Due to the selective pressures that results from the environmental
conditions of sourdough preparation, Lactobacillus sanfranciscensis dominates
type I sourdough fermentations (Corsetti et al., 2001; Foschino et al., 1999).
2.4.2 Use of sourdough in cereal products

Sourdough process has been used as a form of leavening is the oldest
biotechnological processes in food production (Roecken and Voysey, 1995). The
use of sourdough in wheat breads has gained popularity as a mean to improve
the quality and flavor of wheat breads (Corsetti et al., 2000; Thiele et al., 2002;
Brummer and Lorenz, 1991) To facilitate continuous production, one could save
a portion of ripe sourdough to seed subsequent dough, a process that continued
into the nineteenth century (Williams and Pullen, 1998). A vast array of
traditional products relies on the use of sourdough fermentation to yield baked
goods with particular quality characteristics. Some examples include the well
known Italian products associated with Crhistmas, Panettone, which originated in
Milan (Sugihara, 1977). San Francisco sourdough French breads (Kline et al.,
1970) and soda crackers (Sugihara, 1985) are other examples of wheat products
that rely on the process of souring. The same process is also used in the
production of a number of flat breads, a typical example of which is the Egyptian
baladi bread (Qarooni, 1996).
2.4.3 Sourdough fermenting microorganisms

So far, a few less than 50 different species of LAB isolated from sourdough
have been reported (Hammes et al., 2005). Numerous species of lactic acid
bacteria occur naturally in wheat flour, including members of the genera
Lactobacillus, Pediococcus, Enterococcus, Lactococcus, and Leuconostoc (Hammes and
Vogel, 1997). Most of the species of lactic acid bacteria of the genus Lactobacillus
are isolated from sourdoughs (Corsetti et al., 2001; Ottogalli et al., 1996).
Lactobacillus sanfranciscensis, Lb. brevis and Lb. plantarum are the most frequent
lactobacilli isolated from sourdoughs (Bocker, Valmorri et al., 2006; Corsetti et al.,
2003; Corsetti et al., 2001; Gobbetti, 1998; Vogel and Hammes, 1990).
Homo-fermentative species do not produce any carbon dioxide; their
function is acidification and flavor development. Although, homo-fermentative
species of LAB may be used in the majority of fermented food applications but
hetrofermentative species play a major role in sourdough fermentation
(Salovaara, 1998), especially when sourdoughs are prepared in a traditional
manner (Corsetti et al., 2003; Corsetti et al., 2001). The hetero-fermentative LAB
results in better taste and flavor of the sourdough breads, because only hetero-
fermentative lactic acid bacteria can produce the considerable amount of acetic
acid under anaerobic conditions which are desired in sourdough (Kosmina,
1977). On the other hand fermentation with homo-fermentative lactic acid
bacteria results in high concentration of lactic acid, relative to acetic acid, results
in mild and flat sour taste in bread (Spicher and Rabe, 1981). Oura et al. (1982)
prepared rye bread with pure culture of hetero-fermentative bacteria L. brevis,
this provided the rye bread with desirable aroma but not an elastic crumb. They
observed an opposite effect when they used homo-fermentative bacteria (L.
plantarum). It was concluded that in order to get satisfactory aroma and crumb
characteristics, both bacterial species must incorporated.
Cossignani et al. (1996) used Lactobacillus sanfranciscensis, L. plantarum and
Saccharomyces cerevisiae for leavening wheat sourdoughs. They found that the
dough’s fermented with starters had more balanced microbiological and
biochemical characteristics than dough’s started with Saccharomyces cerevisiae, in
which alcoholic fermentation end products largely predominated. By using
starters, the greatest lactic acid bacteria cell number and acetic acid production
was achieved. The starters resulted in more complete profiles of volatile
compounds and greater structural stability.
Starting from glucose, homo-fermentative LAB mainly produce lactic acid
through glycolysis (homolactic fermentation) while hetero-fermentative LAB
produce, besides lactic acid, CO2, acetic acid and/or ethanol (depending on the
presence of additional substrates acting as electron acceptors (Axelsson, 1998).
The LAB, both homo-fermentative and hetero-fermentative species, contribute
most to the process of dough acidification, while yeasts are primarily responsible
for the leavening however, the hetero-fermentative LAB also contribute partly to
the leavening process (Gobbetti et al., 1995a; Spicher, 1983).
Gobbetti (1998) reported Lb. sanfranciscensis and Lb. plantarum association
in Italian wheat sourdough. Lb. plantarum may be superseded by another
facultative hetero-fermentative species, Lactobacillus alimentarius in its association
with Lb. sanfranciscensis in sourdough made from durum wheat (Corsetti et al.,
2001). Lactobacillus alimentarius is capable of fermenting all four flour soluble
carbohydrates (maltose, sucrose, glucose and fructose) and it is possible that this
reduces direct metabolic competition with Lb. sanfranciscensis. Most of the Lb.
alimentarius strains, due to a phenotypical misidentification, probably belong to
Lb. paralimentarius, a facultatively hetero-fermentative species first isolated from
Japanese sourdough (Cai et al., 1999). Lactobacillus brevis and Lb. plantarum have
generally been found associated with Lactobacillus fermentum in Russian
sourdoughs (Kazanskaya, et al.,, 1983). Gobbetti et al. (1994a) reported that Lb.
acidophilus is common in Umbrian (Italian region) sourdoughs, even though it is
rarely isolated from sourdoughs of different origin (Infantes and Tourneur,
1991). Corsetti et al. (2005) described a new sourdough associated species, Lb.
rossiae, that seems to be wide diffused in sourdoughs of southern and central
Italy (Settanni, et al., 2005). Lb. rossiae is often associated with the key sourdough
Lb. sanfranciscensis. Lb. rossiae has been found in environments other than
sourdough (De Angelis et al., 2006a), while no other habitat is known for Lb.
sanfranciscensis (Hammes et al., 2005). However, occurrence of lactic acid bacteria
and yeasts in sourdoughs and the association between acidification and bacterial
metabolism was first demonstrated in 1894 (Hammes and Ganzle, 1998).
Association of yeasts and lactic acid bacteria are often used in the
production of beverages and fermented foods (Gobbetti, 1998). The vast majority
of yeasts found in sourdoughs have been allotted to the species Candida milleri,
Candida holmii, Saccharomyces exiguous and Saccharomyces cerevisiae (Hammes and
Ganzle, 1998). Most of yeast preparations often contain LAB, especially
lactobacilli rather than Pediococcus, Lactococcus and Leuconostoc spp. (Jenson, 1998),
which contributes a little to the aroma development acidification and of dough
because of the limbed processing time (Rothe and Ruttloff, 1983).
2.4.4 Yeast in sourdough

The microorganisms are mostly found on the glumes and glumules of
caryopsis, which is usually removed during milling or during the other
treatments of the grain before milling, hence, yeasts are not very abundant in
refined flour (Galli and Franzetti, 1987). The sourdough fermentation is a
complex process caused by the combined effects of the metabolism of yeasts and
lactic acid bacteria. The former is mainly responsible for the leavening, while the
latter acidify it. The dough in special conditions, some yeast and lactic acid
bacteria act synergistically. Along with fermentation of sugars to carbon dioxide
and ethanol, the yeast also produces some by-products which impart taste and
flavor to the bread (Boraam et al., 1993).
These yeasts are often associated with LAB in sourdough and yeasts/LAB
ratio is generally 1:100 (Ottogalli et al., 1996; Gobbetti et al., 1994). The yeasts
found in sourdoughs belong to more than 20 species (Gulloet al., 2002; Stolz,
1999; Rossi, 1996). Typical yeasts associated with LAB in sourdoughs are
Saccharomyces exiguus, Candida humilis (formerly described as Candida milleri) and
Issatchenkia orientalis (Candida krusei) (Succi et al., 2003; Gobbetti et al., 1994a;
Spicher and Schroder, 1978). Other yeast species detected in sourdough
ecosystem are: Pichia anomala as Hansenula anomala, Saturnispora saitoi as Pichia
saitoi, Torulaspora delbrueckii, Debaryomyces hansenii and Pichia membranifaciens
(Succi et al., 2003; Foschino and Galli, 1997; Gobbetti et al., 1994a). The presence of
Candida humilis in sourdough was reported by Barnett et al. (2000), however the
dominance of Candida humilis in sourdough is a new observation.
The variability in the number and type of yeasts species in dough are
affected by many factors like dough hydration, level the type of cereal used, the
leavening temperature and the sourdough maintenance temperature (Gobbetti et
al., 1994; Hardy, 1982). Among yeasts, S. cerevisiae is most sensitive, while S.
exiguous, or its imperfect form Torlupsis holmii is very resistant. This depends
upon pH of the medium and the presence of undissociated form of acetic acid
(Lodder, 1974). The titratable acidity and pH of the dough are important during
sourdough fermentation. In the initial phase, both acidity and pH remain
constant, whereas, during the intermediate phase titratable acidity increases due
to the presence of yeast. During long term fermentation phase the yeast presence
becomes negative and titratable acidity and pH of the dough depend mainly on
the lactic acid bacteria introduced into the system (Mascaros et al., 1994). The
yeasts present in sourdough are only slightly influenced by lactic acid, but much
more effected by acetic acid (Schulz, 1972).
The available carbohydrates in wheat flour are maltose followed by
sucrose, glucose and fructose, along with some trisacchrides like maltotriose and
reffinose. The glucose increases during fermentation, whereas sucrose decreases
in the presence of yeast due the reaction of invertase (Gobbetti et al., 1994) .The
yeasts present in sourdoughs are not able to ferment maltose, a sugar common in
flour. However, it can nevertheless develop because of glucose released into the
medium by some lactic acid bacteria species, e.g. L. sanfranciscensis (Foschino and
Galli, 1997; Boraam et al., 1993).
2.5 Beneficial effects of sourdough fermentation
2.5.1 Heath benefits of sourdough bread

Cereal foods in its various forms are essential components of the daily diet
and nutritionally, they are an important source of carbohydrates, protein, dietary
fiber and many vitamins and non-nutrients (Liu et al., 2000; Pereira et al., 2002).
The whole grains intake is reported to be associated with health benefits,
including improved regulation of blood glucose level and decreased risk of
diabetes cardiovascular disease, and certain cancers (Jacobs et al., 1998). Bakery
products, particularly sourdough breads produced from high extraction rate
flours are the best potential source of fiber to reduce cardio-vascular disease,
gastrointestinal disorders and diabetes (Pomeranz et al., 1977)
The sourdough provides aromatic and pleasing flavor, and improves
overall quality and shelf life of whole grain breads (Katina et al., 2005). The
sourdough fermentation has been reported to increase folate content (Kariluoto
et al., 2004; Liukkonen et al., 2003), decrease tocopherol and tocotrienol content
(Liukkonen et al., 2003),
Whole meal cereals are an important source of minerals such as K, P, Mg,
or Zn, but mineral utilization is limited by the presence of phytic acid (Lopez et
al., 1998). Wheat and rye contain about 2–58 mg/g phytic acid, which is localized
in the aleurone layer of a kernel (Garcia-Estepa et al., 1999). The degradation of
phytate, has repeatedly been reported in sourdough processes (Angelis et al.,
2003). Reduction of phytic acid content during bread making depends on
phytase action, meal particle size, pH, temperature, water content and
fermentation time (Angelis et al., 2003). Phytate-degrading enzymes exist in
cereals, yeast and lactic acid bacteria isolated from sourdoughs (Lopez et al.,
2000). Generally, low pH favors degradation of phytic acid and optimal pH value
for hydrolysis is 4.5 in wheat and rye doughs. Use of sourdoughs or acidified
sponges can be adjusted to improve mineral bioavailability by increasing phytic
acid hydrolysis (Fretzdorff and Brummer, 1992).
The sourdough has also great potential to modify the macromolecules in
the dough, the most well known examples being the ability of sourdoughs to
reduce digestibility of starch (Liljeberg et al., 1995). The presence of lactic acid in
bread, either added or formed during sourdough fermentation, has also been
reported to reduce acute glyceamic and insulinaemic responses (Liljeberg et al.,
1995).
2.5.2 Improving nutritional and sensory quality

LAB has a long history of use in food and is generally regarded as safe
organisms (Magnusson et al., 2003). Cereal grains like wheat from which most
breads are produced, are low in some of the essential amino acids, i.e., lysine,
threonine, methionine, tryptophan and isoleucine. Such cereal grains can be
considered low in quality protein. Thus, cereal grain-based diets, prevalent in
many areas of the world, may be deficient in some essential amino acids.
Lyophilized cultures of the microorganism may be added to cereal grains such as
wheat in bulk to increase the basic nutritive protein quality of the wheat (El-
Megeed et al., 1989).
The mineral availability of sourdough baked goods is also improved
(Larsson and Sandberg, 1991; Lopez et al., 2003). The low pH values associated
with chemically or microbiologically acidified wheat dough’s lead to
solubilization of the phytate complex, thus increasing mineral bioavailability.
Exopolysaccharides produced by L. sanfranciscensis improve the nutritional
properties of sourdough fermented products in view of the fact that they may be
metabolized by bifidobacteria (Korakli et al., 2002). Some sourdough lactic acid
bacteria have shown hydrolyzing activities towards prolamin peptides involved
in human cereal intolerance (Di Cagno et al., 2002).
2.5.3 Dough structure and bread characteristics

The sourdough fermentation affects the dough rheology at two levels, in
sourdough itself, and in bread dough containing sourdough. In dough,
fermentation decreases elasticity and viscosity, whereas the addition of
sourdough to final bread dough results in less elastic and softer dough’s. The
level of rheological changes taking place in these doughs and its influences on
bread quality can be controlled by adjusting fermentation time and the ash
content of flour during the pre-fermentation process (Clarke et al., 2004).
Di-Cagno et al. (2002) measured the rheology of fermented dough’s by
using empirical techniques and found a decrease in resistance to extension and
an increase in both extensibility and degree of softening. During the sourdough
fermentation different organic acids are produced. These organic acids improve
the flavor of bread, help the swelling of gluten and increase gases retention,
which result in products with good texture and massive volume and also
function as natural dough conditioner (Park et al., 2006)
The exopolysacchardies are microbial polysaccharides secreted
extracellularly, the amount and structure of which depend on the particular
microorganisms and the available carbon substrate (Korakli et al., 2001). These
are produced by lactic acid bacteria during fermentation is one of the aspects of
sourdough technology with the potential for the replacement of hydrocolloids.
These compounds, commonly named as gums, are used as texturizing,
antistaling, or prebiotic additives in bread production (Tieking et al., 2003).
Though the pH of a ripe sourdough varies with the nature of the process
and starter culture used, but for wheat sourdoughs, it ranges from 3.5 to 4.3. The
nature of the flour, in particular its ash content, has a considerable effect on
acidification (Clarke et al., 2002; Collar et al., 1994). The acids strongly influence
the mixing behavior of dough and the doug with lower pH value requires a
slightly shorter mixing time (Hoseney, 1994). This acid increases the solubility of
the glutenin fraction extracted from wheat flour and also affects the swelling
power of gluten (Axford et al., 1979).
Medium strength flours are generally superior to either strong or weak
flours for double layered flat breads. Strong flours result in superior dough
handling properties, but result in unsatisfactory crumb texture and separation
quality. On the other hand, durum wheat flour results in softer product. The
softer the wheat, the harder and less flexible the final product (Williams et al.,
1988)
In comparison to bread prepared with bakers yeast, the sourdough breads
are characterized by moist, dense grains and rather chewy texture (Qarooni,
1996). The application of sourdough to wheat breads has a positive impact on
bread volume (Clarke et al., 2004; Corsetti et al., 1998a; Collar et al. 1994a; Barber
et al., 1992) which is a primary quality characteristic of bread (Maleki et al., 1980).
The larger loaf size produces softer bread and the sourdough breads have been
shown to have lower crumb firmness values (Clarke et al., 2004; Corsetti et al.,
2000; Collar, 1994a). The holes of relatively small size (~1 or 2 mm) are required
in bakery products, whereas large voids or irregular crumb distributions are
undesirable (Cauvain, 1998). An increase in the mean cell area has been
demonstrated via addition of 20% sourdough (Crowley et al., 2002).
The optimal use of sourdough can improve the taste and flavor of the
bread (Rehman et al., 2006). The flavor of sourdough wheat bread is richer and
more aromatic than wheat bread, a factor that can be attributed to the long
fermentation time of sourdough (Brummer and Lorenz, 1991). The concentration
of 2-phyenylethanol, one of the most potent odorants of wheat bread crumb
increases in sourdough bread crumb (Gassenmeier and Schieberle, 1995). The
production of volatile flavor components in sourdough is strongly dependent on
the starter culture, but the role played by the flour used has also been recognized
(Hansen and Hansen, 1994). The main influence of microorganisms on
sourdough flavor has been identified as their ability to enhance or reduce the
amount of specific volatiles already present in the flour (Czerny and Schieberle,
2002).
In bakery products, staling indicates decreasing consumer acceptance
which is caused by changes in crumb other than those resulting from the action
of spoilage organisms (Bechtel et al., 1953). The application of lactic acid bacteria
in the form of sourdough have positive effect on bread staling. One such effect is
an improvement in loaf specific volume, which is associated with the reduction
in the rate of staling (Maleki et al., 1980; Axford et al., 1968). The breads
containing sourdough can decrease the staling rate as measured by differential
scanning calorimetric (Corsetti et al., 1998a; 2000; Barber et al., 1992).
2.5.4 Microbial spoilage

The sourdough inhibits the growth of the pathogens by synthesizing the
antimicrobial compounds, like lactic acid, acetic acid, benzoic acid and hydrogen
peroxide (Park et al., 2006). Mold growth is the most common cause of microbial
spoilage and deterioration in the quality of bread during storage. Certain
sourdough lactic acid bacteria and their components have an antifungal effect
against various fungal species due to the production of organic acids,
particularly acetic acid (Roecken, 1996). The use of lactic acid bacteria as a means
of biopreservation, that is, control of one organism by another has shown very
good results (Magnusson et al., 2003). Positive effects of the use of sourdough on
the mold free shelf life of wheat bread have also been reported by Barber et al,
(1992).
In order to reduce the use of preservatives and treatments that may affect
healthy attributes of food has led to attempts to improve bread quality and shelf
life through formulation with compounds naturally occurring in foods
(Lavermicocca et al., 2000). There is no correlation between bread shelf-life and
pH level, but the type and amount of acid present may have an effect on other
micro static agents (Barber et al., 1992). Employing an agar well diffusion assay it
was found that the individual organic acid (acetic, caproic, proionic, butyric, n-
valeric, and formic) produced by L. sanfrancisco inhibits Fusarium graminearum
(Corsetti et al., 1998b). Other substances contributing to activity of this nature
include returein, hydroxyl fatty acids, proteinaceous compounds, cyclic
dipeptides, 3-phenyllactic acid, caproic acid, and diacetyl hydrogen peroxide
(Magnusson, 2003). Although, antimold activities may vary greatly among the
strains and are mainly detected within obligately hetero-fermentative
Lactobacillus species but rate of inhibition of number of fungal species is highly
strain dependent. (Corsetti et al., 1998a).
2.6 Flavoring compounds and sourdough fermentation

Flavor is simultaneous perception of taste, odors and trigeminal nerve
response (Lawless and Heymann, 1999). The aroma of bread is one of the most
important characters influencing the acceptance of a consumer (Schiebeler, 1996).
The ingredients play a vital role in the development of characteristic flavor of the
bread. Other main sources are dough fermentation and the baking process
(Hansen and Schieberle, 2005). The flavor of bread crumb is mainly affected by
enzymatic activity during the dough fermentation, whereas the flavor of bread
crust is more influenced by the thermal reactions during the baking process
(Rothe, 1974). The sourdough is mainly used to improve flavor in wheat breads
(Hansen and Hansen, 1996). The flavor of sourdough wheat bread is richer and
more aromatic than yeast fermented breads, due to long fermentation time of
sourdough (Brummer and Lorenz, 1991). Some other factors also influence the
production of volatile compound during production of sourdough bread
including free amino acids formed during fermentation through proteolysis
which influence volatile compound profile of the products (Gobbetti et a.l, 1994b;
Collar and Martinez, 1993; Spicher and Nierle, 1984). The production of volatiles
is also influenced by proofing and baking processes (Hansen et al., 1989b).
The bread flavor is composed of hundreds of volatile and non-volatile
compounds, i.e. many alcohols, ketones, aldehydes, acids, esters, furan derivates,
ether derivates, hydrocarbons, ketones, lactones, prazines, pyrrol derivates and
sulphur compounds which serve as flavor stimuli (Maga, 1974; Schieberle, 1996).
Chemical analyses of flavor compounds can be combined with sensory analysis
of bread. The compounds that have been positively correlated with flavor of
wheat crumb are acetaldehyde, 2-methylpropanoic acid, 2/3-methyl-1-butanol,
3-methylbutanoic acid, isopentanal, 2-nonenal, benzylethanol, 2- phenylethanol,
2,3-butandione and 3-hydoxy-2-butanone, dimethyl sulphide and 2-furfural
(Hansen and Hansen, 1996; Rothe, 1974).
Abde-el-Malek et al (1974) concluded that micro flora involved in
fermentation of bladi bread is lactic acid bacteria L. brevis and L. fermenti and
yeast. The lactobacillus replaces are responsible for the typical flavor of bladi
bread. In sourdough fermentation, organic acetic acid and volatile flavoring
compounds produced are dependent upon the microorganisms in the dough
(Hansen and Hansen, 1994a). The volatile compounds are produced both in lactic
acid fermentation and in alcoholic fermentation, but the levels of these
compounds are much higher in yeast fermentation (Hansen and Hansen, 1994b;
Meignen et al., 2001). However, Schieberle (1996) concluded that volatiles formed
do not affect the final flavor of the bread. The compounds having a high flavor
dilution factor would have a significant impact on the final odor. Salovaara and
Valjakka (1987) found that flour type has a considerable influence on the
production of acids. The concentration of acetic acids in whole wheat flour is
almost double the concentration in bread made from straight grade flour. The
concentration of lactic acid is 30 to 50% higher in bread made from whole wheat
flour as compared to bread made from straight grade flour. Moreover, addition
of sucrose to wheat dough stimulates both yeast and LAB growth and increases
bacterial production of lactic and acetic acids (Corsetti et al., 1994).
There is also a lack of competition between Lb. sanfranciscensis and S.
exiguous exist for maltose consumption which is elementary for the stability of
both the microorganisms for the production of good quality San Francisco
French bread (Sugihara et al.,, 1970).
Optimum temperature for co-culture of C. humilis and Lb. sanfranciscensis
are 28 and 32 ºC, respectively, but reduced production of acetate by Lb.
sanfranciscensis has been observed at 35 °C, although, both lactate and ethanol
formations are not affected at this temperature (Brandt et al., 2004). In a French
yeast sourdough, more than 40 flavoring components are identified: 20 alcohols,
7 esters, 6 lactones, 6 aldehydes, 3 alkanes and a single sulphur compound
(Frasse et al., 1993).
2.6.1 Solid Phase Microextraction (SPME)

Generally, bread quality is judged on the basis of its volume, color and
flavor, but bread aroma is the most important factor influencing the consumer
acceptance (Caul, 1972). Previously, different extraction techniques have been
introduced in flavor analysis. These techniques are organic solvent extraction,
distillation and purge and trap extraction method. All theses techniques are
complex, time consuming and expensive (Ruiz et al., 2003). Solid-phase micro-
extraction (SPME) is a modern, solvent–free sample preparation technique
(Arthur and Pawliszyn, 1990). This technique has been developed to combine
sampling and sample preparation in one step. SPME technique along with other
application is used to measure the volatile flavor profiles of food stuff (Young
and Pepard, 1994). The SPME is fast, sensitive, solvent less and economical
method of sample preparation before analysis through gas chromatography and
in some cases, high-performance liquid chromatography (Hook et al., 2002).
The effectiveness of analyte pre-concentration using the SPME technique
depends on many factors such as type of fiber, sample volume, temperature and
extraction time, salting, mode of extraction, desorption of analyses from the fiber
and derivatisation (Muller et al., 1999). The SPME uses a short length of fused
silica coated with adsorbent called as fiber. This fused silica coated fiber is
immersed directly into an aqueous sample or into the head space above a liquid
or solid sample with a general rule that similar is dissolved in similar applies to
the SPME too, i.e. polar compounds are adsorbed on polar fibres and nonpolar
on nonpolar ones (Pawliszyn, 1997). However, the efficiency of pre-concentration
depends not only on the type of fibre but also on its thickness (coating volume).
The type of fibre affects the amount and character of adsorbed species. A
Polydimethylsiloxane (PDMS) fibre of 100 µm thickness is most frequently used
in the analysis of fragrances and impurities in food products (Go´ recki et al.,
1999).
Ruiz et al (2003) developed a solid phase micrextraction method to analyze
volatile compounds in bread crumb. They used three different fibers to
determine the volatile compounds. Carboxen/Polydimethylsiloxane showed the
best extraction efficiency.
2.7 Organic acids in sourdough

Mold growth being the most important cause of bread spoilage, could be
prevented by the use of homo and hetero-fermentative LAB (Spicher, 1983). This
fungistatic effect is due to the production of acetic acid by LAB (Roecken, 1996).
Corsetti et al. (1998) studied the antifungal activity of sourdough LAB. They
concluded that a mixture of acetic acid, propionic acid, caproic acid and butyric
acid are responsible for the activity. The caproic acid along with acetic acid plays
an important role in inhibiting fungal growth. They also found that sourdough
bread made from low pH and high concentration of acetic acid has large volume
and lower rate of staling.
Hansen et al. (1989) studied the effect of three hetero-fermentative and two
homo-fermentative LAB strains on the production of organic acids in sourdough.
The results show that acetic acid content is much higher in the dough acidified
with hetero-fermentative strains as compared to homo-fermentative strains. The
homo-fermentative strain produces only L- lactic acid, whereas other cultures
produce both L- lactic acid and D- lactic acid. In another study (Hansen and
Hansen, 1994), the effect of wheat flour types on the production of organic acid
was also observed. The wheat flours have significant effect on the production of
lactic and acetic acids. The highest lactic acid contents are observed in low grade
and whole meal flour. The production of acetic acid depends on the starter
culture used. The acetic acid is found to be 12% of the total acid contents in
hetero-fermentative cultures, but it has not been detected in homo-fermentative
cultures but sourdough made from straight grade flour fermented with homo-
fermentative strain L. plantarum shows small amount of acetic acid. Vernochi et
al. (2004) found that sourdough fermentation with C. milleri results in high
amounts of acetic acid and mannitol which improves the qualitative
characteristics of leavened dough and the baked products.
2.8 Sensory evaluation of bread

Sensory evaluation of food and food products is one of the most important
aspects of quality control. Two primary sensory characteristics of flat bread are
considered to be color and texture (Farooq et al., 20001). The acceptability of any
food product like bread is subjected to a number of factors such as effect of
climate, geographical location, consumer’s age and level of income along with
other factors. Flat breads are mostly made from flour of high extraction rate. So,
the sensory qualities of flat breads including color, texture, chewability may be
closely examined (Qarooni, 1996) because as the time passes, breads loose their
commercial value through sensory changes which are due to physical and
chemical deterioration (Park et al., 2006).
The acceptability of flat breads and chapatti can be related to a number of
factors, such as behavior during preparation, rheological characteristics and
sensory qualities. The flat breads characteristics such as color and appearance of
dough, percentage of water absorption and puffing, appearance, texture and
taste are affected due to differences in wheat varieties (Austin and Ram, 1971). It
has been reported that though grain protein increases with fertilizer increase but
no changes are observed in chapatti quality characters such as water absorption,
puffing, texture and taste (Austin and Jhamb, 1964).
Azizi et al, (2006) studied the effect of wheat flour extraction rate on
sensory characteristics of Iranian flat breads. It has been observed that by
increasing the extraction rate, the quality of breads is improved up to 90%
extraction rate. Thus overall quality of breads is affected by flour extraction rate.
In another study (Park et al., 2006), it was found that the texture profile was
lower in sourdough bread as compared to controlled bread. There has been no
significant difference in the sensory characteristics of bread present except for the
shape of the bread. The results suggests that the quality of bread improved by
the use of sourdough.
Lotong et al. (2000) developed a lexicon for describing the flavor of wheat
sourdough bread. A highly trained description sensory panel identified and
defined referenced 32 flavors attributes for wheat sourdough bread. Crumb and
crust of wheat sourdough were evaluated differently. All 37 wheat sourdough
breads showed different flavor attributes and intensities in both crumb and crust
samples. The principal component analysis for crumb and crust indicated that
number of attributes could not be reduced into a smaller set of components that
completely described the wide range of breads used in that study.
The crumb of wheat bread made from sourdough fermented with the
hetero-fermentative Lb. sanfranciscensis shows a pleasant, mild, sour odor and
taste, whereas sourdough bread fermented with the homo-fermentative Lb.
plantarum has an unpleasant metallic sour taste. However, sourdough wheat
bread supplemented with sourdough yeast S. cerevisiae illustrates a more
aromatic flavor (Katina et al., 2006). Sensory evaluation of sourdough bread
crumb suggests that a most intense and bread-like flavor relates to propanone, 3-
methylbutanal, benzyl alcohol and 2-phenylethanol (Hansen et al., 1989).
Chapter-3
MATERIALS AND METHODS
The research was carried out to study the effect of wheat flour extraction
rates on physico-chemical characteristics of sourdough flat bread (naan). The
work was conducted at National Institute of Food Science and Technology,
University of Agriculture, Faisalabad-Pakistan and Department of Food Sciences,
University of Copenhagen, Denmark.
Details of raw materials used and the procedures employed are as follow:
3.1 Procurement of wheat varieties
Wheat varieties namely Inqulab-91, Auqab-2000, Iqbal 2000, and AS-2002

were procured from Wheat Research Institute, Ayub Agriculture Research
Institute, Faisalabad-Pakistan.
3.1.1 Physical characteristics of wheat
3.1.2 Thousand kernel weights

Hundred grams representative sample of each wheat sample was taken
and thousand kernel weights were recorded by counting clean, sound and
unbroken kernels. The kernel weight was calculated as grams per thousand
kernels (Farooq et al., 2001).
3.1.3 Test weight

Test weight (Kg/hl) of each sample was determined according to method
No. 55-10 as described in AACC, 2000.
3.2 Milling characteristics of wheat
3.2.1 Milling of wheat
One kg of each sample of wheat was cleaned and tempered to 14%

moisture level and kept for 24 h at ambient temperature in a close plastic jar. The
amount of water required for tempering was calculated according to following
expression given in method No. 26-29 as described in AACC, 2000.
100-original moisture%
Water to be added (ml) = ------------------------------ - 1 x wt. of sample (g)
100-desired moisture%
After tempering, wheat samples were milled by using Quadrumate Senior
Mill (C. W. Brabender, Duisburg, Germany). Four milling fractions namely break
flour, reduction flour, bran and shorts were obtained. Break and reduction flours
were mixed together to obtain straight grade flour for the preparation of product.
3.2.2 Wet and dry gluten
Wet and dry gluten contents of flour were estimated by following the
method No. 38-10 as described in AACC, 2000. 25 g of flour was taken in a
porcelain cup. Sufficient water was added to form firm dough. Dough was hand
kneaded into ball and placed it in water at room temperature for 60 min. Dough
was kneaded gently in stream of tap water over bolting cloth until starch and
soluble matter were removed. To determine whether gluten was starch free, one
or two drops of wash water, obtained by squeezing, were dropped into clear
water. The gluten thus obtained was pressed as dry as possible between the
hands, and weighed as wet gluten. Then it was transferred to an air oven to a
constant weight at 100 °C for 24 hours and cooled. It was weighed as dry gluten.
3.2.3 Falling Number (FN)
Falling numbers were determined using Pertin Falling Number Apparatus

1900 (Pertin Instruments AB, SE 1405, Huddinge, Sweden) by following the
method No. 56-81 as described in AACC, 2000. 7 g of flour was taken into dry FN
tube. 25 ml water was added at 22±2°C. Rubber stopper was inserted and tube
was shaken in upright position for 30 times up and down until mixed.
Viscometer stirrer was used to scrape down slurry coating upper part of tube,
and all slurry scraped from stopper. Tube and viscometer stirrer were placed
into water bath after mixing. At the end of test, time was recorded in sec. FNs
were calculated using following formula:
FN (14% moisture basis) = FN as is x (100-14) / (100-moisture of sample in %)
3.2.4 Particle size index (PSI)
The particle size index of each sample was determined by sieving through
sieve shaker model RX-86-2 using U.S. standard sieve No. 70 using the method
No. 55-30 as described in AACC, 2000. 10 g of flour were weighed and poured
over the top of the sieve. Weighing pan was cleaned thoroughly and then fixed
under the sieve. After fixing sieve over the pan, the sieve shaker was run for ten
min. Five min were given to settle the thrus after completion of the shaking time.
The material passed through the sieve (thrus) was collected from the pan. Any
material adhering to the bottom of the sieve was also added to the pan and
collected. The collected material was weighed in an electric balance and the
results were expressed as particle size index in percentage.
3.2.5 SDS - sedimentation value

The sedimentation value was determined by following the procedure
outlined in method No. 56-61-3 as described in AACC, 2000. 3 g of whole wheat
flour were taken in a graduated cylinder in which 50 ml bromophenol blue dye
was added and the cylinder was shaken for specific intervals. Then 50 ml SDS
reagent was added and the cylinder was inverted for the specific time and
allowed to stand to sediment the contents. After 15 min the volume of the
sediment was recorded as SDS sedimentation value in ml.
3.2.6 Pelshenke value
The pelshenke value of each wheat variety was determined according to

the method No. 56-50 as described in AACC, 2000. 2.25 ml of yeast suspension
were added to 4 g of flour and it was kneaded to make a dough ball with clean,
smooth and uniform surface. The dough ball was placed in 200 ml beaker
containing 100 ml distill water (30oC). The time of immersion of dough ball was
recorded. The time taken by the dough ball to disintegrate was noted. The
difference between immersion and disintegration times of dough ball was
expressed as pelshenke value or whole meal fermentation time in min.
3.3 Proximate analysis of wheat flour

All samples were analyzed for moisture, crude protein, crude fat, crude
fiber, total ash and Nitrogen Free Extract (NFE) contents according to their
respective standard methods as described in AACC, 2000.
3.3.1 Moisture Content

The moisture content of flour samples was determined according to
method No. 44-15 A as described in AACC, 2000. 5 g flour was taken in a tarred
crucible and dried in hot air oven (Memmert Model 200) at 100±5ºC till a
constant weight. The moisture contents were calculated by the formula given
below.
Wt. of original sample – Wt. of dried sample
Moisture (%) ═ ------------------------------------------------------- x 100
Wt. of original sample
3.3.2 Crude protein content

Crude protein was determined by the Kjeldahl method according to
method No. 46-10 as described in AACC, 2000. The samples were first digested
in digestion flask with H2SO4 in the presence of a digestion mixture for 3-4 h till
the contents of digestion flask get transparent color. Samples were then diluted
with distilled water up to 250 ml in a volumetric flask. The ammonia from the
samples trapped in H2SO4 was liberated through distillation after adding 40%
NaOH solution and collected in a flask containing 4% boric acid solution using
methyl red as an indicator to determine nitrogen content in a sample by titrating
against standard 0.1N H2SO4 solution. The crude protein percentage was
calculated by using following formula
0.0014 × Vol. of 0.1N H2SO4× 250 ml
N (%) = ------------------------------------------------------------ x 100
Vol. of diluted sample × Wt. of original sample
Protein (%) was then calculated by multiplying N (%) with factor 5.7.
3.3.3 Crude fat content

The method employed was that of solvent extraction using a Soxhlet
extraction as described in method No. 30-10 (AACC 2000). 2 g of flour were taken
in a thimble and placed in extraction tube of Soxhlet apparatus. About 250 ml of
Hexane were added in 500 ml bottom flask of the apparatus and connected to the
Soxhlet apparatus. The fat was extracted by running Hexane over the sample at
the rate of 3-4 drops per sec for about 5 h. The content of the flask was
transferred to a pre-weighed petri dish and dried on a hot plate for 10 min at a
temperature of 40-50ºC. The petri dish was cooled in desiccator and weighed. Fat
percent age was calculated according to the following formula.
Weight of fat in sample
Crude fat (%) = ----------------------------- x 100
Weight of sample
3.3.4 Crude fiber content

Crude fiber content was determined by following the method No. 32-10 as
described in AACC, 2000. 2 g fat and moisture free sample was taken and placed
in 1000 ml beaker. 200 ml solution of 1.25 % H2SO4 was added in the beaker. The
sample was then digested by boiling for 30 min. Then it was filtered by using
suction apparatus. The residue was washed with hot water until become acid
free. The residue was then again transferred to 1000 ml beaker and boiled with
200 ml solution of 1.25 % NaOH for 30 min. It was again filtered and the residue
was transferred to pre-weighed crucible and dried in an oven at 100 ºC for 24 h
till constant weight was obtained. Then the dried residue was charred on a
burner and ignited into muffle furnace at 550-600ºC for 5-6 hours, cooled in
desiccators and weighed. The loss in weight during incineration represents the
weight of crude fiber in sample. The crude fiber % age was calculated by using
the following formula.
Weight of residue – Weight of ash
Crude fiber (%) = ----------------------------------------------- x 100
Weight of sample
3.3.5 Ash content

Ash content was determined by incineration of the sample at 600ºC
according to method No. 08-01 as described in AACC, 2000. 5 g oven dried
sample was taken in a pre-weighed crucible and charred on a burner. Then it
was ignited in a muffle furnace at 550ºC till constant weight of grayish ash was
obtained. The ash of sample was calculated by using following formula.
Weight of ash
Ash (%) = ---------------------- x 100
Weight of sample
3.3.6 Nitrogen Free Extract (NFE)

NFE contents of flour samples were calculated by subtracting the
percentages of moisture, ash, crude protein, crude fat, and crude fiber from 100
as below
NFE % ═ 100 – (moisture % + ash % + crude protein % + crude fat % + crude

fiber %)
3.4 Rheological characteristics

Rheological characteristics of flour samples were determined with
Brabender Farinograph (C. W. Brabender, Duisburg, Germany) according to the
method No. 54-21 as described in AACC, 2000. Dough characteristics such as
water absorption, dough stability, dough development time, tolerance index and
softening of dough were interpreted from farinogram.
3.4.1 Water absorption

The water absorbed by the flour was the percent of water required to
reach the center of curve on the 500 line.
3.4.2 Arrival time

The arrival time (min) was recorded by measuring the time required to
reach the top of curve on the 500 B.U line from the start of a curve. It gives the
rate of hydration of flour.
3.4.3 Dough development time

The dough development time (min) was determined from fiarinogram as
the time required for a curve to reach at its full development or maximum
consistency showing the highest peak. High peak value is associated with strong
wheat which has long mixing time.
3.4.4 Dough stability

This is the time (min) that top of the curve remains above the 500 B.U line
and was measured from the arrival time to the departure time. Greater tolerance
would indicate that the flour can stand for more mixing and the longest
fermentation conditions.
3.4.5 Departure time/dough resistance

The time required for the curve to reach the point where the top of the
curve left the 500 B.U line.
3.4.6 Tolerance index (TI)

This indicates how fast a flour will breakdown after it has reached its full
development. It was measured in Brabender Units. It was expressed as the
difference between the center of the curve at the peak and center of the curve five
min after the peak.
3.4.7 Softening of the dough

Degree of softening was measured at the difference between the center of
the curve at peak and centre of the curve at 12 min after the peak. It was
expressed in Brabender Units (B.U.).
3.5 Preparation of flat breads (Naan)

The production of traditional naan was based on yeast fermentation
process. Flour (300 g), yeast (3.5 g), sugar (10 g), sodium bicarbonate (3 g), salt (4
g) and water (200 ml) were mixed into dough. The amount of water required was
determined by farinographic water absorption .The dough was kneaded for 10
min (4 min slow + 6 min fast) with a Hobart mixer (model A-200, H-194–268,
Hobart Corp., Troy, OH) equipped with a dough-kneading arm. After mixing,
the dough was allowed to ferment for 1 h at 30ºC. A dough ball of 100 g was
taken and sheeted into a disk of 7-in. diameter with rolling pin. The disk was
pressed with the fingertips in the center covering 5-in diameter. It was allowed to
proof for 30 min and baked in an oven at 315ºC for 3 min. The naans were
packed in a polyethylene bag at 30ºC and stored in a refrigerator for different
physico-chemical analysis (Farooq et al., 2001).
3.6 Sensory evaluation

Freshly prepared naans were evaluated for sensory characteristics like
color, texture, texture, flexibility, chewability and overall acceptability at room
temperature in sensory evaluation laboratory by a panel of 10 judges on 9-point
Hedonic Scale (Land and Shepherd, 1988).
3.7 Selection of wheat variety
On the basis of physico-chemical, a rheological and sensory characteristic,

the best suited wheat variety was selected for further studies.
3.8 Flour milling for different extraction rate flours.

Selected wheat variety (AS-2002) was milled to get different extraction
rate flours from 64% to 100%.
Wheat tempered to 15% moisture was milled by using Quadrumate Senior
Mill (C. W. Brabender, Duisburg, Germany). Four milling fractions namely break
flour, reduction flour, bran and shorts were obtained. Break and reduction flours
were mixed to get straight grade flour. The by-products obtained during milling
such as bran and shorts were milled in a hammer mill and blended with 64%
extraction rate flour by passing through a 40-mesh sieve (610 μ). The 64%, 76%,
88% and 100% extraction rates flours were obtained by proportional mixing of
the by-product of wheat with 64% extraction rate flour.
3.9 Proximate analysis of wheat flours
Prepared wheat flours with different extraction rates were analyzed for
moisture, crude protein, crude fat, crude fiber, total ash and NFE contents
according to their respective standard methods (AACC 2000).
3.10 Rheological characteristics

Rheological behavior of different flour samples was evaluated by running
flour samples through Brabender Farinograph (C. W. Brabender, Duisburg,
Germany) (AACC 2000). Dough characteristics such as water absorption, dough
stability, dough development time and tolerance index of dough were
interpreted from Farinogram.
3.11 Sourdough starter cultures

Freeze dried cultures were procured from Lallemand Baking Solutions,
Montreal, Canada. Freeze dried cultures namely, LA-1 containing homo-
fermentative strain Pediococcus acidilacti along with yeast Saccharomyces cerevisiae
and LA-5, mixed culture containing hetero-fermentative strain L. brevis and
homo-fermentative strain L. casei, along with yeast Saccharomyces cerevisiae, was
used.
3.11.1 Mother sponge

Mother Sponge was prepared from flour (200g), tap water (200ml at 30°C)
and freeze dried starter culture was added to give colony-forming units (cfu) of
108 bacteria per gram and yeast count as 106 cfu per gram of sponge. This was
mixed by hand with a spoon and incubated for 20 h at 30 °C (Hansen and
Hansen, 1994).
3.11.2 Sourdough preparation
Sourdough was prepared from flour with 64, 76, 88 and 100% extraction
rates. The flour (200 g), water (100 ml) and fermented sponge (70 g) were mixed
together into dough and incubated for 20 h at 30°C.
3.11.3 Acid content analysis

pH and total titratable acidity (TTA) of dough samples were determined
according to method given by Lefebvre et al. (2002), using the Inolab WTW Series
720 pH meter. A 10g sample was blended with 90 ml distill water and the
suspension was then titrated with a 0.1 mol/L NaOH to a final pH of 8.5. The
TTA was expressed as the amount (ml) of NaOH used.
3.11.4 Microbial content

Samples (10 g) of the sourdough were homogenized with 90 ml NaCl
(0.99% w/v). Colony forming units were determined by plating serial dilutions
on MYPG-Agar for yeasts and on modified MRS agar (containing maltose) for
LAB (Volgel et al., 1994). Incubation temperature was 25 °C for yeasts and 30 °C
for LAB.
LAB count
The modified MRS Agar containing following ingredients was used.
Ingredients Quantity
Trypton 10 g
Meat extract 10 g
Yeast extract 5g
Glucose 7g
Fructose 7g
Maltose 7g
Sodium
2g
glyconate
Tween 80 1 ml
K2HPO4 2g
Sodium
5g
acetate
Triammonium
2g
citrate
MgSO4. 7 H2O 0.2 g
MnSO4. 4
0.05 g
H2O
Cystein HCl 0.5 g
Agar 15 g
Water 1000 ml
The pH was adjusted at 6.3. Media was autoclaved at 121 °C for 15 min.
Yeast count
MYPG-Agar medium containing following ingredients was used
Malt extracts 3g
Yeast extracts 3g
Glucose monohydrate 10 g
Peptone 5g
Agar 20 g
Water 1000 ml
The pH was adjusted at 5.6. Media was autoclaved at 121 °C for 15 min.
3.12 Sourdough volatile compounds

The sourdough volatiles compounds were collected by both Dynamic
Headspace (DH) and Solid Phase Microextraction (SPME) Techniques and
analysed by gas chromatography with identification based on GC-retention
times for reference compounds and GC-mass spectrometry.
3.12.1 Description of Dynamic Headspace analysis

A 10 g sourdough sample was mixed with 30 ml (20%) NaCl solution (pH
3 with 0.05 citric acid) and 100 µl 4-methyl-1-pentanol (50 mg L-1) was added as
internal standard. Volatile compounds were collected on a Tenax-TA trap. The
trap contained 250 mg of Tenax-TA with mesh size 60/80 and a density of 0.37 g
ml-1 (Buchem bv, Apeldoorn, Netherlands). The sample/suspension was
equilibrated to 30±1°C in a circulating water bath and then purged with nitrogen
(100 ml min-1) for 60 min.
a) GC-MS analysis
The trapped volatiles were desorbed using an automatic thermal
desorption unit (ATD 400, Perkin Elmer, Norwalk, USA). Primary desorption
was carried out by heating the trap to 250°C with a flow (60 ml min-1) of carrier
gas (He) for 15 min. The stripped volatiles were trapped in a Tenax TA cold trap
(30 mg held at 5°C), which was subsequently heated at 300°C for 4 min
(secondary desorption, outlet split 1:10). This allowed for rapid transfer of
volatiles to a gas chromatograph-mass spectrometer (GC-MS, G1800A GCD
System, Hewlett-Packard, Palo Alto, CA, USA) through a heated (225°C) transfer
line.
Separation of volatiles was carried out on a DB-Wax capillary column 30
m long x 0.25 mm internal diameter, 0.25 µm film thickness. The column flow
rate was 1.0 ml min-1 using helium as a carrier gas. The column temperature
program was: 10 min at 45°C, from 45°C to 240°C at 6°C min-1, and finally 10 min
at 240°C. The GC was equipped with a mass spectrometric detector operating in
the electron ionization mode at 70 eV. Mass-to-charge ratios between 15 and 300
were scanned. The volatile compounds were identified by matching their mass
spectra with those of a commercial database (Wiley 275.L, HP product no.
G1035A). The software program, GCD plus ChemStation G1074B (Version
A.01.00, Hewlett Packard, Palo Alto, California), was used for data analysis.
b) Concentrations
Concentrations are presented as relative areas calculated as peak area of
the volatile compound divided by the peak area of internal standard
3.12.2 Solid Phase Microextraction (SPME) method

The sourdough volatile compounds were analyzed according to the
method proposed by Ruiz et al, (2003). A 2 g of dough sample was weighed into
a 20-ml vial. Then, 8.0 ml of a 20% NaCl solution (pH 3 with 0.05 citric acid) and
100 µl of internal standard solution (4-methyl-1-pentanol at 50 ppm) were added
to the vial. Then, the capped vial with silicone septa was placed in tray 1 of CTC
Combi Pal automatic sampler attached with GMS. Pre incubation time 300 sec
and incubation temperature 50oC with an extraction time of 1 h were used. The
SPME fibers used were 85-μm carboxen/polydimethylsiloxane (PDMS) and 100-
μm PDMS. The sample was continuously agitated at a speed of 500 rpm during
the extraction process in CTC Combi Pal automatic sampler. After 1 h extraction
automatic sampler inserted the fiber into the GC–MS Chromatograph (Agilent
Technologies 6890 N Network GC System with 5973 inert mass selective
detector) injector port for thermal desorption of the extracted volatiles for 5 min.
The column used was DB-WAX (Agilent Technologies, USA) with a 0.25-
μm film thickness, 30 m length and 0.25 mm I.D. The temperature program was
35oC maintained for 5 min, then increased by 5oC /min to 50oC, where it was
held again for 5 min, then increased by 5.5oC /min to 230oC, where it was held
again for 5 min.
Helium was used as carrier gas, with a column flow of 2.0 ml/min. The
injector was at 300 oC in splitless mode for 5 min using an inlet of 0.75 mm I. D.
The concentrations are again presented as relative areas calculated as peak area
of the volatile compound divided by the peak area of internal standard.
3.13 Preparation of sourdough naan

The sourdough naans were prepared by using wheat flours of different
extraction rates. Dough was prepared by mixing flour (250 g), fermented sponge
(100 g), sugar (10 g), salt (6 g), and water as determined by farinographic water
absorption and incubated for 20 h at 30 °C. A dough ball of 100 g was taken and
sheeted into a disk of 7-in. diameter with rolling pin. The disk was pressed with
the fingertips in the center covering 5-in. diameter. Put it in proofing chamber at
37°C and 85% RH for 45 min. Finally it was baked in oven at 315ºC for 3 min. The
sourdough breads were packed in a polyethylene bag and kept at -20°C for
further analysis.
3.14 Organic acids in Sourdough naan

Organic acids (lactic acid, acetic acid and citric acid) contents in
sourdough naans were determined using High Performance Liquid
Chromatography (HPLC) by following the method of Vernocchi et al. (2004).
HPLC (Model 1050, Hewlett-Packard, Waldbronn Germany) equipped
with a Bio-Rad Aminex (Bio-Rad Laboratories, Hertfordshire, UK) HPX-87H
column (300×7.8 mm) was used for organic acids determination. The following
conditions were used: mobile phase, 0.08 M H2SO4; flow rate, 0.60 ml min_1;
temperature, 65°C; the column was connected with a UV detector (210 nm).
For HPLC analysis, 100 ml of 0.1 N sulfuric solution was added to each 10
g samples, the mixture was homogenized for 5 min by an Omni Mixer
Homogeniser (Omni International, Warrengton, VA), filtered through a 0.22-mm
membrane filter and analyzed as described above
3.15 Volatile compounds in sourdough naan

The sourdough naan volatiles compounds were collected by both
Dynamic Headspace and Solid Phase Microextraction Techniques and analyzed
with Gas Chromatography with identification based on GC-retention times for
reference compounds and GC-mass spectrometry as described earlier in 3.12.
3.16 Minerals content of sourdough naan
The minerals such as Fe, Ca, Zn and Cu were estimated according to the
respective method as described in AOAC (2000) using Atomic Absorption
Spectrophotometer (Varian, AA240, Victoria, Australia). A 0.5 g sample was
digested separately by using wet digestion method. The sample was first
digested with 10 ml HNO3 at a temperature of 60-70°C for 20 min and then
digested with HClO4 at a temperature of 190°C till the solution become clear. The
digested sample was transferred to 250 ml volumetric flask and volume was
made with distilled water and then filtered. The solution was loaded into atomic
absorption spectrophotometer apparatus. The standard curve was prepared by
running samples of known strength through atomic absorption
spectrophotometer. The mineral contents of unknown samples were estimated
by using the respective standard curve prepared for each mineral.
3.16.1 Phosphorous content in sourdough naan
Ammonium vandate was used to determine phosphorus content by

aminonaphthol sulphonic acid method as described by Kitson and Mellon (1992).
Reagents
Preparation of 2.5% ammonium molybdate:

2.5 g Ammonium molybdate was dissolved in 3 ml conc. H2SO4 and
volume was made to 100 ml with distilled water.
Preparation of aminonaphthol sulphonic acid:
0.5 g Aminonaphthol sulphonic acid was dissolved into 15% Sodium

bisulphate solution (195 ml) with added 20% Na2SO3 (5 ml) and dissolve.
Preparation of phosphorus standard solution:
0.351 g of KH2PO4 was dissolved in distilled water to make 100 ml.

Concentration of phosphorous achieved was 0.8 mg/ml.
Procedure
In a test tube, add ammonium molybdate solution (1 ml),

amminonaphthol sulphonic acid solutions (0.4 ml), and 0.1 ml of sample or
standard at various dilutions. Distilled water was added to make up the volume
(10 ml). Solution was shaken and allowed to stand for 5 min to develop color.
Blank was used to set the zero absorbance at 720 nm wavelength.
3.17 Sensory evaluation

Sourdough naans prepared with different extraction rate flours were
evaluated for sensory characteristics like, color, flavor, taste, texture,
foldingability, chewability and overall acceptability at room temperature in
sensory evaluation laboratory by a panel of judges on 9-point Hedonic Scale
(Appendix-I) as described in 3.6. (Land and Shepherd, 1988).
3.18 Statistical analysis

Statistical analysis was carried out using Minitab (V.13.1, Minitab Inc., PA
16801-3008, USA). Duncan’s Multiple Range Test was applied to calculate the
level of significance (Steel et al., 1997).
Chapter-4
RESULTS AND DISCUSSION
The present research work consisted of two phases. During the first phase,
four Pakistani wheat varieties were analyzed for their suitability for the
production of naans. On the basis of physico-chemical, rheological and sensory
attributes, best suited wheat variety was selected for further studies.
During the second phase, selected wheat variety was milled to get
different extraction rate flours. Sourdough naans were prepared from different
extraction rate flours using two different freeze dried mixed starter cultures. The
sourdough naans prepared from different extraction rate flours were analyzed
for different quality attributes.
4.1. SUITABILITY OF DIFFERENT WHEAT VARIETIES FOR THE

PRODUCTION OF YEAST LEAVENED FLAT BREAD (NAAN)
4.1.1. OBJECTIVE
The objective of this study was to determine the suitability of different
wheat varieties for the production of yeast leavened naan through physico-
chemical, rheological and sensory analyses. Four Pakistani wheat varieties
namely Inqulab-91, Auqab-2000, Iqbal-2000, and AS-2002 were investigated for
their suitability for the production of naans. Before milling, wheat grains were
tested for their physical characteristics such as test weight and thousand kernel
weight.
4.1.2. RESULTS
4.1.2.1. Physico-chemical characteristics of wheat varieties
4.1.2.1.a. Thousand kernel weight and Test weight of wheat varieties

The analyses of variances for TKW and TW for four different wheat
varieties are given in Table 1. Both TKW and TW were highly significantly
(P<0.01) affected by varieties. The mean values for TKW and TW are given in
Table 2. The variety AS-2002 showed the highest TKW (43.16 g) and TW (79.43
kg/hL), while Inqulab-91, Auqab-2000 and Iqbal-2000 showed 42.06, 40.39 and
39.22 g for TKW and 77.35, 71.50 and 71.28 kg/hL for TW, respectively.
4.1.2.1.b. Milling of wheat
All wheat varieties were tempered to 15% moisture level and then milled
by using Quardrumate Senior Mill. Four milling fractions namely break flour,
reduction flour, bran and shorts were obtained. The break and reduction flours
were mixed together to get straight grade flours. The straight grade flours (SGF)
were analyzed for characteristics such as wet and dry gluten contents, falling
number (FN), particle size index (PSI), SDS–sedimentation (SDS-S) and
pelshenke values (PV).
4.1.2.1.c. Wet gluten (WG) and dry gluten (DG) of flour

The gluten proteins impart unique bread making properties to wheat.
Wheat varieties varied significantly (P<0.05) for their wet and dry gluten
contents as shown in Table 3. Mean values for wet and dry gluten are given in
Table 4. The values for WG ranged from 29.45 to 33.56%. The highest amount of
WG was found in straight grade flour of wheat variety AS-2002, whereas the
lowest was found in Auqab-2000.
The mean values for DG ranged from 8.72 to 10.69%. Flour from Inqulab-
91 showed the highest DG content of 10.69% and the Auqab-2000 gave the lowest
DG content of 8.72%.
4.1.2.1.d. Falling number (FN) of flour
Highly significant differences were notice in respect of FN in wheat

varieties (Table 3). Mean values for falling number are given in Table 4 showing
that FN values ranged from 490.67 to 532.67 sec. The highest FN value was
observed in wheat variety Inqulab-91 (532.67 sec), whereas wheat variety AS-
2002 exhibited the lowest FN value (490.67 sec).
4.1.2.1.e. Particle size index (PSI) of flour
Particle size index (PSI) is a measure of kernel hardness and is positively

correlated with test weight and thousand kernel weight. The wheat varieties
differed significantly with regards to PSI. The statistical results are shown in
Table 3. This indicates highly significant effect on PSI.
The mean values for PSI is given in Table 4. The PSI ranged from 19.00 to
20.17% in four wheat varieties. The highest PSI was found in wheat variety
Inqulab-91 having PSI 20.17%, whereas wheat variety Iqbal-2000 showed the
lowest PSI 19.00%.
4.1.2.1.f. SDS-Sedimentation (SDS-S) value of flour

The statistical analysis for SDS-S value of different wheat varieties is given
in Table 3. It is evident from the results that SDS-S values of four wheat varieties
were found highly significant (P<0.01).
The mean values for SDS-S value are given in Table 4, indicating that SDS-
S values of different wheat varieties ranged from 26.39 to 34.04 ml. The highest
SDS-S value was observed in wheat varieties AS-2002 as 34.04 ml, whereas wheat
variety Auqab-2000 showed the lowest SDS-S value as 26.39 ml.
4.1.2.1.g. Pelshenke value (PV) of flour
PV is also an estimation of wheat protein quality and can be related to

SDS-S value. The statistical results of the PVs of four different wheat varieties are
given in Table 3. The wheat varieties showed highly significant (P< 0.01) effect
with regards to PV.
The means for the PVs are given in Table 4. The PVs were in the range of
172.33 to 186.00 min. The highest PV was found in wheat variety AS-2002 as
186.00 min, whereas Auqab-2000 showed the lowest value as 172.33 min.
4.1.2.1.h. Proximate analysis of straight grade flour (SGF) of wheat varieties
Statistical analysis for proximate composition of straight grade flour (SGF)

is given in Table 5. It is evident from sum of squares table that wheat varieties
significantly (P<0.05) differed in respect of moisture, protein and nitrogen free
extract (NFE), whereas as crude fiber, crude fat and total ash were found non-
significant (P>0.05).
Mean values for proximate composition of SGF are given in Table 6. The
moisture content was found in the range of 11.78 to 12.09%. The highest moisture
(12.09%) was found in SGF from Iqbal-2000, whereas SGF from variety AS-2002
showed the lowest moisture content (11.78%). The crude protein was found the
highest (12.05%) in flour from AS-2002, while the lowest in (11.71%) in Iqbal-
2000. The highest value (1.40%) of crude fat was estimated in AS-2002 and the
lowest (1.29%) in the Iqbal-2000. The results regarding crude fiber revealed that
all the varieties had almost the similar crude fiber contents. The highest value
was found (0.44%) in SGF from variety Inqulab-91. Total ash varied very little
similar that of crude fiber and the highest value (0.57%) was found in Iqbal-2000.
The Nitrogen free extract (NFE) contents were in the range of 73.66 to 74.04%.
The highest percentage of NFE (74.04%) was found in SGF from variety Auqab-
2000, while the lowest (73.66%) was in variety Inqulab-91.
4.1.2.2. Farinographic characteristics of straight grade flour (SGF) of wheat

varieties
Farinograms of SGF of different wheat varieties determined with
Brabender farinograph are given in Fig 1. Statistical analysis regarding the
farinographic characteristics of flours from different wheat varieties is given in
Table 7, indicating highly significant difference (P<0.01) among all farinographic
characteristics.
Mean values for different farinographic characteristics are given in Table
8. Water absorption (WA) is an important characteristic of wheat. Flour having
low WA produces dough low in moisture which on baking may produce dry and
stiff breads of poor quality. The WA of different wheat varieties were in the
range of 53.33 to 55.83%. The highest WA was shown by AS-2002 having 55.83%,
whereas, variety Auqab-2000 showed the lowest WA with value of 53.33%.
Arrival time (AT) varied from 1.47 to 2.47 min. Dough development
times (DDT) were in the range of 3.53 to 4.63 min and the highest DDT was
produced by variety Inqulab-91. Dough stability (DS) of different varieties were
recorded from 6.27 to 8.13 min. Wheat variety Iqbal-2000 showed the highest DS
with a value of 8.13 min. Departure time (DT) were in the range of 8.17 to 9.50
min and the variety Iqbal-2000 showed the highest DT. Tolerance index (TI)
ranged from 21.00 to 64.00 BU and softening of dough (SD) varied from 59.00 to
80.33 BU.
4.1.2.3. Sensory evaluation of flat breads (naan) prepared from straight grade
flour (SGF) of wheat varieties
Yeast leavened flat breads (naan) were prepared using SGF from different
wheat varieties. The yeast leavened naan was subjected to sensory evaluation by
a panel of judges. The objective of the sensory evaluation was the selection of
best wheat variety for the production of yeast leavened naan. The sensory
evaluation parameters included were color, texture, flexibility, chewability and
overall acceptability.
Statistical analysis for sensory characteristics is presented in Table 9. It is
evident from the results that sensory attributes like color, texture, chewability
and overall acceptability were found significant (P<0.05). The flexibility was
found highly significant (P<0.01) among all the naans.
The mean values for the sensory parameters are given in Table 10. The
color got scores from 6.20 to 7.20. The naans prepared from variety AS-2002 got
(7.20) the highest scores in terms of color, whereas, naans prepared from variety
Auqab-2000 got (6.20) the lowest scores. Sensory scores for flat breads (naan)
prepared from different wheat varieties is also shown in Fig 2.
The texture of naans was found significantly different for all varieties. The
texture got scores in the range of 6.60 to 7.40. The naans prepared from AS-2002
got the highest scores (7.40) and naans from Iqbal-2000 got just 6.60 scores.
The flexibility was the only sensory parameters that showed significant
variation among all the treatments. The naans prepared from AS-2002 got the
highest scores (7.20), whereas naans from Iqbal-2000 got the lowest (5.80) scores.
The chewability and overall acceptability were also found significant. In
terms of chewability, the naans prepared from Inqulab-91 were ranked first with
mean scores of (7.00) followed by AS-2002 with mean scores of (6.80). The naans
prepared from AS-2002 got the highest scores (7.40) for overall acceptability,
followed by Inqulab-91 scores (6.80).
4.1.3. DISCUSSION
Both thousand kernel weight (TKW) and test weight (TW), are closely
associated with each other and affected flour yield and protein contents. These
are considered important tools in wheat grading systems. Generally, greater the
TKW and TW, better will be the quality and yield of flour.
Pakistani wheat varieties have been investigated for TKW and TW by
many researchers. Rehman et al. (2001) evaluated wheat varieties Mehran-89 and
Anmol-91 and found TKW 39.30 and 42.20 g and TW 75.90 and 74.40 kg/hL,
respectively. Randhawa et al. (2002) estimated TKW and TW of Inqulab-91,
Chenab-2000, Iqbal-2000 and Uqab-2000. Their investigations showed 36.29,
40.89, 37.79, 39.60 g TKW and 69.83, 69.83, 70.00 and 69.67 kg/hL TW of these
varieties, respectively. Butt et al. (2001) tested 30 wheat varieties for their TW
and found that TW varied from 69.33 to 80.42 kg/hL, which are similar to the
results of present studies. The wheat with lower test weight generally gives poor
extraction rate (Cauvain, 1998). Anjum et al. (2002) reported that Pakistani wheat
varieties posses more than 30 g of TKW. All these findings are almost similar to
the results of present studies. The TKW exhibited higher values than other
varieties except Inqulab-91. However, present studies showed higher TW values
ranging from 71.28 to 79.43 kg/hL. The variations existed may be due to genetic
makeup, environment, soil, location and frequency of application of fertilizers
(Rehman et al., 2001; Farooq et al., 2001 and Anjum et al., 2002). However, TW
may be affected more by environment than genotype (Kent and Evers 1994).
Finney et al. (1987) also reported wide variations in TKW among various wheat
varieties grown in different locations
Kent and Evers (1994) reported that for flat breads flour, wheat should be
of high thousand kernels and test weights. The results suggested that wheat
variety AS-2002, showed the highest TKW and TW. Thus, this variety could be
utilized for the preparation of flat breads (naan).
The flour gluten content is one of the most important factors responsible
for quality and baking strength of wheat flour (Maleki and Parchami, 1976). The
results of present studies indicated significant differences among wheat varieties
that might be due to difference in their protein contents. The protein content has
been found to be correlated with the gluten content (Anjum and Walker, 2000).
The differences in wet and dry gluten contents of different wheat varieties are
reflected by the variation in moisture and protein contents (Corbellini et al.,
1999). The results of present studies are in line with the findings of Farooq et al.
(2001) who noted significant effect of varieties and lines on wet and dry gluten
contents of wheat.
The research work previously conducted on Pakistani wheat varieties has
revealed that the differences in WG due to wheat varieties were significant as
reported by Ahmad (2001) who found significant variation for wet and dry
gluten contents of Pakistani wheat varieties. The results of present studies are
also in conformity with the results reported by Miralbes (2003) who observed
WG contents in the range of 15.6 to 39.3%. Wrigley et al, (1982) concluded that
gluten contents may be affected by wheat varieties. The result regarding gluten
contents are also confirmed by the results of Line et al, (2003) who reported WG
contents in the range of 20.5 to 45.6% and DG contents in the range of 8.44 to
11.77%. Ahmad (2001) reported that the DG contents present in the range of 7.51
to 13.52 in the Pakistani wheat varieties. The difference in gluten contents of
different wheat varieties may be due to the environmental conditions like
temperature, rainfall and their genotypic difference.
The FNs have indirect relation with α-amylase activity; high the FN value,
lower will be enzyme activity. The present studies have indicated that Pakistani
wheat varieties are low in enzyme activities which may suggest that wheat flour
from Pakistani wheat varieties should be supplemented with malt to get
desirable enzyme activities for the production of quality bread. These results are
also supported by the study of Anjum and Walker (1991) who reported low
alpha amylase activity in Pakistani wheat. Zahoor (2003) analyzed 44 Pakistani
wheat varieties and reported FN in the range of 277.83-1065 sec. Moreover, the
differences in FN values in different wheat varieties are reflected by the variation
in moisture and protein contents (Corbellini et al., 1999).
The results for PSI indicated that all these wheat varieties fell under the
category of medium hard wheat according to AACC (2000) hardness scale for
wheat varieties. Butt et al. (2001) reported the results of PSI for 30 Pakistani
wheat varieties in the range of 17.32-42.41%. The results of the present studies for
PSI are also confirmed by the findings of Anjum et al. (1993) who categorized the
Pakistani wheat varieties as medium and hard groups. In the present studies, the
highest PSI was found in wheat variety Inqulab-91 and considered harder than
other varieties. Difference in hardness might b due to environmental and
conditions to which cultivar is subjected during growth as reported by Bushuk
(1998)
The SDS-S value represents the strength of wheat proteins. More is the
SDS-S value i.e. ≥ 30 ml, higher will the protein strength (William et al., 1986).
The results of present studies are confirmed by the findings of Corbellini et al.,
(1999) who found the SDS-S values in the range of 20 to 50 ml. The difference in
SDS-S value may be due to the difference in genetic make up of the wheat
varieties. Two wheat varieties, As-2002 and Iqbal 2000 showed SDS-S values
higher than 30 ml, indicating that their protein strength was more than two
varieties and might be fit for production of naan.
The PV relates to the strength of flour and is independent on variation in
protein contents and gives better understanding in respect of relation between
diversity of protein and the quality of wheat (Branlad and Dardevet, 1985).
The present studies showed that results for PVs were in line with that of
SDS-S test as it showed the similar pattern of increase or decrease for varieties.
Branlad and Dardevet (1985) showed low values for PV ranging from 150 to 170
min which might be due to varietals, soil, cultivation practices and
environmental factors.
Quality of breads depends upon the quality of flour from which it is

made. Ideal quality flour for one type of product may not supremely be
performed for others (Finney, 1987). The previous investigations on the
proximate composition of flours revealed that whole wheat flours contain
moisture in the range of 7.68 to 9.32%, crude protein 12.18 to 14.10% and ash 1.32
to 1.72% (Butt et al., 2001). The wheat flours (75% extraction rate) obtained from
Punjab wheat varieties contain moisture 12.50%, crude protein 12.00%, crude fat
1.30%, crude fiber 0.38% and ash 0.58% (Farooq et al., 2001). Yamamoto et al.
(1996) found 0.27 to 0.40% ash contents in soft wheat and 1.6% minerals in the
wheat grains of USA origin. There are many genetic and non-genetic factors
which may alter the composition of the wheat grains. Among these are
environmental conditions and storage conditions, location, soil and fertilizers
(Finney et al., 1987; Anjum and Walker, 2000; Butt et al., 2001).
The moisture content in flour determines its stability. Higher the moisture
in flour, the lower would be the shelf life. It is affected by the type of milling and
moisture percentage used for wheat conditioning. The moisture content in SGF is
higher than whole wheat flour due to tempering of wheat before milling.
The protein is the main ingredient on which flour specification and quality
of the product depends. It varies from 6-20% in different wheat varieties (Kent,
1983). The protein contents are lower in straight grade flour than that of whole
wheat flour due to the removal of bran and aleurone layer which contains
appreciable amount of protein (Alvi, 1994). Various Indian wheat’s have been
evaluated to determine their suitability for different bakery products, including
flat breads (Austin and Ram, 1971). Mixtures of medium and soft wheat are
proved to be more suitable for the flat breads than that of soft wheat’s, which
contain low to medium protein contents with weak gluten and baking strength
(Ahmed et al., 2001; Hansen and Poll, 1997). Finney et al. (1987) reported 13.4%
protein and 0.45% ash in straight grade flour from wheat grown in Kansas. The
results of the present studies have shown that wheat varieties were medium hard
and more suitable for naan production.
The fat percentage is higher in whole wheat flour than that of SGF because
germ is ground along with endosperm during milling resulting in flour with
higher fat contents than other flours (Rao et al., 1983; Farooq et al., 2001). The fat
content showed negative correlations with test weight and thousand kernel
weight. With an increase in test and thousand kernel weights, a reduction in fat
content was happened.
Ash is a mineral residue remaining after a sample has been completely
oxidized in a manner such that all organic volatile material is driven off, while
preventing any mineral from being lost (Posner, 1991). The purity of flour can be
assessed by the amount of ash contents. The SGF contains less ash content due to
the fact that 80% of the total amount of minerals is concentrated in the aleurone
layer of pericarp (bran) which is removed during milling process and remaining
20% minerals are present in endosperm (Lopez et al., 2003). Moreover, the ash
content has been found to increase with an increase in fat content (Kent, 1983).
The SGF also showed less fiber content in present studies because wheat
bran was removed during milling which decreased the percentage of fiber in
flour. The results are in line with the findings of Kent (1983) that fiber is
concentrated mainly in bran and only 7% of total fiber is present in endosperm
and embryo. The nitrogen free extract values are higher in SGF than that of
whole wheat flour due to the fact that NFE is calculated by difference and these
increased proportionately. The NFE has a positive correlation with moisture of
flour.
According to the specifications of previous studies and from the analysis
of present investigations, it was concluded that all wheat varieties might be
suitable for the production of naan. However, wheat variety AS-2002 was found
higher in protein and could be more suitable for making flat bread (naan).
Rheological methods are useful in the study of dough properties.
Rheometery evaluates important functional properties of flour including
viscosity, elasticity and plasticity which can be related with dough behavior
during processing and end product quality (Bloksma and Bushuk, 1988). The
parameters such as water absorption, arrival time, dough development time,
dough stability, departure time, tolerance index and softening of dough are key
factors in rheological properties (Sollars and Rubenthaler, 1975). Good quality
flat breads can be produced from flour of high WA (Kent and Evers 1994).
Variation in WA depends not only on protein quality but also on damage starch
of wheat flour (Pyne et al., 1987; Anjum and Walker 1991). The results of the
present studies are in line with the findings of Borghi et al. (1996), in which WA
ranged from 53 to 60%. Moreover, higher WA is an indication of better dough
development for naan preparation (Farooq et al., 2001). Simon (1987) reported
that the flour with higher water absorption can produce bread which may
remain soft for a long time. This also suggests that flours possessing higher WA
may be more suitable for yeast leavened products and texture and grain of bread
may be improved.
Farooq et al. (2001) also found highly significant difference among various
farinographic characteristics. They determined the positive correlation (r=0.67**)
of WA with protein. Variations in WA and AT depend upon protein quality and
damaged starch content (Anjum and Walker, 1991).
The WA, DDT and DS of whole wheat flour are higher than that of
straight grade (Rao and Others 1983). So the results of the present studies
showed the similar patterns. The WA and DDT of whole wheat are higher than
that of SGF, perhaps due to the presence of bran particles in whole wheat flour
which may interfere in quick development of gluten. The results of the DDT are
in line with the findings of Corbellini et al. (1999), in which the DDT ranged from
90 to 300 sec.
The slight variation in the AT might be due to the variation in the protein
content of wheat varieties (Siddique, 1989). The shorter DDT can be associated
with reduced farmographic stability and a high degree of softening (Borghi et al.
1996). The DDT may be associated with farinographic stability and degree of
softening (Borghi et al., 1996) and as the dough development time, dough
stability and tolerance index increase the softening of dough decreases (Anjum
and Walker, 2000).
. In general, the flours which have low softening values are strong and the
ones having high softening values are weak. Similarly, flours which have good
tolerance to mixing have low tolerance index and the higher are the tolerance
index value; the weaker would be the flour (Farooq et al., 2001).
So, from the farinographic studies, it was concluded that two wheat
varieties AS-2002 and Inqulab-91 were more suitable for the production of naan.
They had better water absorption and dough development time.
The color of naan bread is an important parameter for the consumer’s
acceptance. The significant difference was found in the color of all naan breads.
The difference in color of all the naans maybe due to the differences in
hardness/softness of wheat grains (Farooq et al., 2001). The range of scores for
color is in agreement with the findings of Siddique (1989) who has reported the
scores range from 7.11 to 8.28 for flat bread chapattis.
The results for the texture of naans are in line with findings of Farooq et al,
(2001) in which flat breads got scores in the range of 6.53 to 8.20. Siddique (1989)
proved that softness in chapatti texture was highly correlated with flour color
and consequently bran content which might be due to more water absorption.
They also proved that texture of flat breads improves with the increase in protein
and fiber contents. The results of the present studies have shown that naans
prepared from AS-2002 got the highest scores in terms of texture; this has already
been proved by physico-chemical analysis that variety AS-2002 might be good
for flat bread production.
Flexibility or foldingability is important sensory parameters for naan
breads. It is the folding of naan bread piece into scoop for picking the curry.
Highly significant variations were observed among the treatments. Flexibility
depends upon many factors like protein content of flour, farinographic water
absorption and damaged starch but wheat starches didn’t affect the flexibility
(Wang and Flores, 1999). The present studies have already shown that variety
AS-2002 had the highest water absorption, more damaged starch and good
protein content that might have resulted in better flexibility of the naan bread.
Freshly prepared naans had maximum chewability that gradually
decreased with the passage of time during storage which might be caused due to
loss in moisture content. The chewability is related to hardness or softness of the
naan bread (Farooq et al., 2001). All naan breads showed significant difference
(P<0.05) in terms of chewability, that might be due to difference in protein
contents and water absorption of flour. Results for overall acceptability showed
that naan breads prepared from AS-2002 got the highest scores.
The difference in sensory acceptability of all the flat breads may be due to
the difference in the hardness of wheat grains and several other factors like
wheat varieties and milling characteristics (Farooq et al., 2001). Although, there
are many factors responsible for the quality of flat bread (naan) such as method
of preparation of dough, protein content and quality, diastatic activity, water
absorption, damaged starch etc., but it was observed from the study that the
differences are mainly due to the variation in the genetic make up of wheat
varieties as other conditions were identical for all the naan breads.
The sensory evaluation of the naan breads prepared with straight grade
flour from different varieties showed that variety AS-2002 got maximum scores
for most of sensory parameters.
4.1.4. CONCLUSION
It is concluded from the above results and discussion regarding the
physico-chemical characteristics of different wheat varieties, that wheat variety
AS-2002 showed better performance for most of the parameters and suitable for
the production of leavened flat bread (naan). So, on the basis of physico-chemical
tests and sensory evaluation wheat variety, AS-2002 was selected for further
studies.
4.2. EFFECT OF FLOUR EXTRACTION RATES ON THE QUALITY OF

SOURDOUGH
4.2.1. OBJECTIVE
This study was undertaken to determine the effect of extraction rates on the
quality of sourdough.
Selected wheat variety (AS-2002) was milled to get different extraction
rate flours ranged from 64% to 100%. The 64% extraction rate flour was straight
grade flour, whereas 76%, 88% and 100% extraction rates flours were obtained by
adding bran portions of wheat into 64% extraction rate flour and then
pulverizing with grinder equipped with 1 mm sieve. The prepared flours were
analyzed for proximate composition and farinographic characteristics and used
for making sourdough.
4.2.2. RESULTS
4.2.2.1. Proximate composition of different extraction rate flours
Different extraction rate flours were analyzed for proximate composition.
The statistical analysis of proximate composition of different extraction rate
flours is given in Table 11. All the parameters showed highly significant (P<0.01)
differences from each other.
4.2.2.1.a. Moisture content of flour

Moisture varied highly significantly among all the flours (Table 11). The
mean values for moisture contents of different extraction rate flours are
presented in Table 12 which ranged from 9.8 to 11.78%. The highest moisture
level (11.78%) was found in 64% extraction rate flour which decreased gradually
with an increase in extraction rate and the lowest (9.80%) moisture was found in
100% extraction rate flour.
4.2.2.1.b. Crude protein of flour

The statistical analysis for crude protein is presented in Table 11. The
protein contents for all flours varied significantly (P<0.05). The mean values for
protein content in different extraction rate flours are given in Table 12, indicating
that protein contents increased with an increase in extraction rate. The 64%
extraction rate flour showed the lowest crude protein (12.05%) whereas, 100%
extraction rate flour showed the highest (13.36%) protein contents.
4.2.2.1.c. Crude fat of flour

The statistical analysis for crude fiber content is shown in Table 11,
indicating that crude fat showed highly significant difference (P<0.01) among all
flours.
The mean values for fat content of different extraction rate flours are
presented in Table 12. The fat content increased with an increase in flour
extraction rate. The highest fat content (2.76%) was found in 100% extraction rate
flour, whereas, the lowest (1.40%) was found in 64% extraction rate flour.
4.2.2.1.d. Crude fiber of flour
Results for mean sum of squares for crude fiber contents of different
extraction rate flours are given in Table 11. The statistical analysis showed highly
significant (P<0.01) effect on the quantity of crude fiber.
The mean values for crude fiber of different extraction rate flours are
given in Table 12. The crude fiber contents ranged from 0.40 to 2.14%. The 100%
extraction rate flour exhibited the highest crude fiber (2.14%), whereas, 64%
extraction rate flour contained the lowest crude fiber (0.40%). The crude fiber
increased with an increase in flour extraction rate.
4.2.2.1.e. Ash content of flour

Ash is the mineral residue remaining after a sample has been completely
oxidized in a manner such that all organic volatile material is driven off, while
preventing any mineral from being lost (Posner, 1991). The statistical analysis
(Table11) showed highly significant (P<0.01) effect on total ash contents.
The mean values for ash contents are given in Table 12 indicating that ash
content ranged from 0.56 to 1.55%. The highest ash content (1.55%) was found in
100% extraction rate flour, whereas, 64% extraction rate flour showed the lowest
(0.56%) ash content.
4.2.2.1.f. Nitrogen free extract (NFE) of flour

The statistical analysis for NFE showed highly significant (P<0.01)
variation among all these flours. The mean values for NFE are given in Table 12.
The NFE was found in the range of 70.39 to 73.81 with the highest NFE value
(73.81) was observed in 64% extraction rate flour and the lowest was found in
The nitrogen free extract values were found higher in 64% extraction rate
flour than other flours due to the fact that NFE is calculated by difference and
these increased proportionately. Moreover, NFE has a positive correlation with
moisture content.
4.2.2.2. Farinographic characteristics of different extraction rate flours

The different extraction rate flours were analyzed for their farinographic
characteristics using Brabender farinograph. The statistical analysis regarding
farinographic properties is given in Table 13. The farinographic parameters like
Water absorption, dough development time, dough stability and tolerance index
were found highly significant (P<0.01) among all the flours.
The mean values for farinographic characteristics of different extraction
rate flours are given in Table 14. WA is an important characteristic of wheat. The
WA was found in the range of 55.83 to 64.72%. It increased with an increase in
flour extraction rate and was observed the highest (64.72%) in flour with 100%
extraction rate and gradually decreased to 55.83% in 64% extraction rate flour.
The DDT also increased with an increase in flour extraction rate. The
mean values of DDT (Table 14) were found in the range of 4.53 to 6.58 min. The
highest DDT (6.58 min) was given by 100% extraction rate flour. The flour with
64% extraction rate showed DDT 4.53 min.
Dough stabilities (DS) and tolerance indices (TIs) were reduced with an
increase in flour extraction rates. The DSs were in the range of 1.80 to 6.27 min,
the highest DS was (6.27 min) found in 64% extraction rate flour and it gradually
decreased with flour extraction rate and 100% extraction rate flour gave DS just
1.80 min (Table14). The TIs were found in the range of 45 to 64 BU. The highest
TI (64 BU) was shown by 64% extraction rate flour and it reduced to 45 BU with
4.2.2.3. Sourdough Preparation

Sourdoughs were prepared from different extraction rate flours using
freeze dried lactic acid bacteria (LAB) cultures LA-1 containing homo-
fermentative strain Pediococcus acidilacti along with yeast Saccharomyces cerevisiae
and LA-5, mixed culture containing hetero-fermentative strain L. brevis and
homo-fermentative strain L. casei, along with yeast Saccharomyces cerevisiae.
Dough prepared from straight grade flour and fermented with yeast
Saccharomyces cerevisiae alone was chosen as control treatment
4.2.2.3.a. pH and acidity of sourdough prepared with different extraction rate

flours and cultures
Statistical analyses for pH and acidity of doughs prepared with different
extraction rate flours and cultures are given in Table 15. It is evident from the
table that treatment had highly significant effect (P<0.01) on pH and acidity of
the dough. Highly significant results were obtained when LAB fermented dough
were compared with control treatment. Two LAB types showed highly
significant effect (P<0.01) in terms of acidity of the dough, whereas pH of the
dough fermented with different LAB showed non-significant effect. When
sourdoughs were prepared from different extraction rate flours and LAB, it
resulted in highly significant differences in terms of acidity, whereas pH of the
dough was affected non-significantly.
The mean values for pH and acidity are given in Table 16. The control
treatment showed the highest pH (4.8). The pH of the sourdoughs prepared with
different extraction flours decreased with an increase in flour extraction rate (Fig.
3). The pH reduced from 3.6 to 3.2. The 64% extraction rate flour showed the
highest pH (3.6), whereas 100% extraction rate flour showed the lowest pH (3.2).
The mean values for acidity are shown in Table 16 (Fig. 4). The control
treatment gave acidity 6.8 ml. It is evident that with the increase in extraction
rate, the acidity increased. The acidity values for sourdough prepared from
different extraction rate flours and culture LA- were in the range of 9.5 to 20.8 ml.
Sourdough prepared with 64% extraction rate flour showed the lowest acidity
(9.5 ml) and sourdough prepared from 100% extraction rate flour gave the
highest acidity (20.8 ml). The LA-5 culture resulted in more acidity than that of
LA-1. The 64 % extraction flour showed the lowest acidity value (10.1 ml) that
gradually increased to 22.3 ml in dough prepared with 100% extraction rate
flour.
4.2.2.3.b. Bacterial and yeast counts of sourdough prepared with different

extraction rate flours and cultures
Sourdoughs prepared from different extraction rate flours and cultures
were evaluated for bacterial and yeast counts. Dough prepared from SGF and
fermented with yeast Saccharomyces cerevisiae alone was selected as control
treatment.
The statistical analyses for bacterial and yeast counts in different
sourdoughs and dough fermented with yeast Saccharomyces cerevisiae is given in
Table 17.
All the treatments varied highly significantly (P<0.01) with respect to
bacterial and yeast counts. Comparison of yeast fermented dough with
sourdoughs also showed highly significant effect (P<0.01). The LAB cultures also
varied significantly from each other with regards to yeast and bacterial counts.
Flour extraction rate showed significant effect (P<0.01) with respect to bacterial
count while they varied non- significantly for yeast counts. Similarly, comparison
of LAB cultures with extraction rates resulted in significant results for bacterial
count but varied non-significantly with yeast count.
The mean values for bacterial and yeast count are given in Table 18.
Bacterial count increased with an increase in flour extraction rate for both the
cultures. Bacterial count for sourdough made from different extraction rate flours
using culture LA-1 ranged from 2.2×108 to 2.0×109 colony forming units per gram
(cfu/g), with highest bacterial count (2.0×109 cfu/g) in sourdough made from
100% extraction rate flour. The lowest bacterial count was in yeast leavened
dough (3.2×102 cfu/g) as expected.
Bacterial count for sourdough made from different extraction rate flours
using culture LA-5 increased with an increase in flour extraction rate. Bacterial
counts ranged from 1.4×109 to 2.1×109 cfu/g and the highest bacterial count
(2.1×109 cfu/g) was observed in sourdough made from 100% extraction rate
flour, whereas the lowest (1.4×109 cfu/g) was in sourdough made from 64%
extraction rate flour.
The mean values for yeast count are given in Table 18 indicating the
highest yeast count (1.6×109 cfu/g) was in dough fermented with yeast (control).
Sourdoughs made from different extraction rate flours using culture LA-1
resulted yeast counts in the range of 1.9×107 to 3.1×107 cfu/g while yeast count
for sourdoughs made from LA-5 culture was lower yeast count as compared to
that from LA-1. However, these were not differed significantly.
4.2.3. DISCUSSION
Moisture contents affect the water absorption and storage quality of
flours. It was observed that moisture content decreased with the increase in flour
extraction rate. The results of present study are agreed with findings of Butt et al,
(1997) and Qamar (2002), who found moisture contents in different wheat flours
range from 7.8 to 14.8%. Similar results were reported by Azizi et al. (2006) where
moisture varied from 9.88 to 12.30% in different extraction rate flours. The
decrease in flour moisture contents might be due to the higher amount of bran in
higher extraction rate flours.
The flour protein content in the present study increased as the extraction
rate increased (Dewettinck et al., 2008). Results of the present study are in line
with the results of Azizi et al. (2006) who reported increase in protein content
with an increase in extraction rate. This is a consequence of higher proportion of
aleuronic layer and peripheral endosperm in the flour (Ramirez-Wong et al.,
2007). The results are also supported by the study of Orth and Mander (1975)
who concluded that flour extraction rate affects the protein content, and gluten
strength. Kent and Amos (1967) reported that with an increase in exaction rate,
the protein content, fiber, sugar, lipids and mineral matter increase, whereas the
starch decreases. Alvi (1994) also reported low quantity of protein in straight
grade flour than that of whole wheat flour due to the removal of bran and
aleurone layer which contains appreciable amount of protein.
The fat percentage was higher in 100% extraction rate or (whole wheat
flour) than that of 64% extraction rate or white flour because germ is ground
along with endosperm during milling, results in flour with higher fat content
than other flours (Rao et al., 1983; Farooq et al., 2001). The results of fat contents
are in agreement with the findings of Azizi et al, (2006) who reported an increase
in fat contents of flour with an increase in flour extraction rates.
The results for crude fiber contents in different extraction rate flours
showed that crude fiber increased with an increase in flour extraction rate. The
64% extraction rate flour showed less fiber contents because wheat bran was
removed during milling process which decreased the amount of fiber in flour.
Azizi et al, (2006) reported crude fiber in the range of 0.30 to 2.24% in different
extraction rate flours. The results of present studies are also supported by Kent
(1983) that fiber is concentrated entirely in bran and only 7% of total fiber is in
endosperm and embryo. Kent and Amos (1967) also reported increase in fiber
content with an increase in flour exaction rate.
The ash content increased as the extraction rate increased. This increase
might be due to higher proportion of pericarp and aleuronic layer (Ramirez-
Wong et al., 2007). The results for ash contents in the present studies are in
agreement with the findings of Vetrimani et al. (2005), who reported that the ash
contents increased from 0.52 to 1.32%, with an increase in extraction rate from
66% to 100%. The 64% extraction rate flour contained less ash content due to the
fact that 80% of the total amounts of minerals are concentrated in the aleurone
layer of pericerap (bran) which was removed during milling process while only
20% minerals are present in endosperm (Lopez et al., 2003; Kent (1983)).
The farinograph commonly provide empirical information about the
mixing behavior of dough (Arendt et al., 2007). The results of farinographic
characteristics of present studies showed that water absorption increased as the
extraction rate increased. The water absorption influences the handling
properties of the dough and is related to the finished baked product (Catterall,
1998). The water absorption increased with the increase in flour extraction rate.
The increase in WA might be due higher protein and complex carbohydrate
contents contributed from bran (Pomeranz, 1988). The results of present studies
are supported by Azizi et al. (2006) who reported WA of different extraction rate
flours in the range of 56 to 66%. Farooq et al. (2001) reported that higher water
absorption is an indication of better dough development for naan preparation.
Simon (1987) showed that the flour with higher water absorption produces bread
of good characteristics which remains soft for a long time, exhibiting
improvement in texture and grain.
The Present studies showed variation in dough development times for
extraction rates. Vetrimani et al. (2005) reported similar results for DDT in which
it increased from 2.5 to 3.5 min and DS was decreased from 4.5 to 2.5 min. High
DDT maybe due to the presence of increased amount of bran particles in high
extraction rate flours, which may interfere in the quick development of gluten
and hydration of endosperm. Orth and Mander (1975) concluded that high
extraction rate flours produced doughs with less gluten strength might be due to
the presence of non gluten protein. Dexter and Matsuo (1978) also reported the
similar results. The variations in the farinographic characteristics might be due to
difference in endosperm portion among the different extraction rate flours
(Vetrimani et al., 2005). With the increase in extraction rate resistance to extension
and extensibility reduced, which might be due to reduction in gluten network
and presence of bran particles in flour (Haridas and Rao, 1991)
The pH of sourdough varies with the kind of starter culture and nature of
the process used but for wheat sourdoughs it ranges from 3.5 to 4.3 (Collar et al.,
1994; Thiele et al., 2002). The results of pH and titratable acidity (TTA) of
sourdoughs in the present studies ranged within typical values. Flour extraction
rate showed highly significant effect (P<0.01) on TTA and with the increase in
flour extraction rate TTA increased. Flour ash content might have influenced the
acidification of sourdough (Arendt et al., 2007). The sourdough made from whole
meal flour showed almost double TTA than sourdough made from straight
grade flour. The increase in TTA might be due to the presence of more quantity
of fermentable carbohydrates in whole meal flour. The starter culture also had an
effect on TTA, but this effect was small in comparison to flour extraction rates
(Roecken and Voysey, 1995).
The use of mixed culture resulted in higher TTA compared to yeast
leavened dough (Plessas et al., 2008). Weustink (1989) reported a linear
relationship for ash contents (0.55 to 1.65 %) and the production of TTA in the
sourdoughs. In the present studies, the results for ash contents were also in
similar range and TTA increased with an increase in ash content. The pH values
were found in the range of 3.5 to 3.8, when sourdoughs were prepared with
different extraction flours (Hansen and Hansen, 1994) which had considerable
influence on the production of acids (Salovaara and Valjakka, 1987) and
increased to almost double the values in sourdough made from straight grade
flour (Brummer and Lorenz, 1991).
The bacterial cfu increased gradually with an increase in flour extraction
rate. The bacterial counts (109 cfu/g dough) in sourdough made from whole meal
flour were significantly higher than sourdoughs made from straight grade (108
cfu/g dough) (Hansen and Hansen, 1994). The yeast counts, in contrast with
bacterial counts were maximum in straight grade flour (107 cfu/g dough) which
decreased gradually to 105 cfu/g dough with increase in flour extraction rate. The
decrease in yeast count with the increase in extraction rate might be due to
increase in buffering capacity by the use of higher extraction rate flour that
resulted in higher acid contents (Rohrlich, 1960). The results are in line with the
findings of Hansen and Hansen (1994) who reported yeast count of 105 cfu/g
dough in sourdoughs made from different flours.
The results for bacterial and yeast counts are in accordance with the
definition of sourdough as stated by Loenner and Preve (1988) that sourdough
should contain at least 108 cfu/g dough bacterial and 105 cfu/g dough yeast
counts.
4.2.4. CONCLUSION
The studies suggest that flour of high extraction rates produce more
acidity in sourdough as compared to low extraction rate flours. The higher water
absorption and better dough development contributed by high extraction reflect
the suitability of flour for the preparation of sourdough and production of naans.
4.3. SOURDOUGH VOLATILE COMPOUNDS
4.3.1. OBJECTIVE
This piece of study was carried out to evaluate the performance of mixed
cultures of yeast and LAB in sourdough volatile production. Volatiles
compounds produced by sourdough made from different extraction rate flours
and starter culture LA-1 and LA-5 were extracted by both Solid Phase
Microextraction (SPME) and Dynamic Headspace Technique (DHT) and
analyzed with gas chromatography and identification of compounds based on
GC-retention times for reference compounds and GC-mass spectrometry.
4.3.2. RESULTS
4.3.2.1. Sourdough volatile compounds extracted with SPME technique

SPME is a fast, modern, solvent free and economical method of sample
preparation (Arthur and Pawliszyn, 1990; Hook et al., 2002). This technique
results in sampling and sample preparation in one step. SPME is extensively
used to measure the volatile flavor profiles of food stuff.
The volatile compounds extracted using SPME technique were alcohols,
esters, acids, carbonyls and aromatic compounds. The details of the volatile
compounds are given in Table 19. Different alcohols produced by LA-1 and LA-5
starter cultures (Table 19) were mainly ethanol, 1-propanol, 2-methyl, 1-butanol,
2-methyl-, 1-butanol, 3-methyl, 1-pentanol, 1-hexanol heptanol. Ethanol was
found in both LA-1 and LA-5 fermented sourdoughs, the yield of which was
mainly influenced by starter cultures.
The esters identified include ethyl acetate and ethyl lactate. The ethyl
acetate was present in both LA-1 and LA-5 fermented doughs, whereas ethyl
lactate was not observed in LA-5 fermented dough. Production of ethyl acetate
increased with an increase in flour extraction rate.
The production of acetic acid was more in sourdough made from LA-5
starter culture, whereas LA-1 resulted in more production of hexanoic and
propionic acids.
The carbonyl identified were acetaldehydes, benzaldehydes and N-
hexanal. The production of hexanal was more effected by starter cultures rather
than flour extraction rate. Both acetaldehydes and hexanal were produced in
higher amounts in LA-1 fermented dough as compared to the one fermented
with LA-5.
4.3.2.2 Sourdough volatile compounds extracted with Dynamic

Headspace technique
Dynamic Headspace has been a very popular technique proven for its
performance and used in a number of studies (Hansen and Hansen, 1994;
Aparicio and Morales, 1994; Hansen and Hansen, 1996). The sourdoughs
prepared with different extraction rate flours and starter cultures (LA-1 and LA-
5) were analyzed for volatiles production using DHT and analyzed by gas
chromatography.
The results for sourdough volatiles using DHT are shown in Table 20. The
sourdoughs volatiles belong to alcohols, acids, esters carbonyls and many other
aromatic compounds.
Alcohols produced were mainly ethanol, Iso-amyl alcohol, 2-Butanol, 1-
butanol, 3-methyl, 1-pentanol, 1-hexanol, heptanol, propanol, 1-octanol , N-
octanol and nonanol. The production of 2- butanol was observed more in LA-1
sourdough as compared to LA-5 sourdough, but flour type did not have too
much effect on its production. Iso-amyl alcohol was only found in LA-1
sourdough having high relative peak areas. The production of 1-butanol, 3-
methyl was observed in all doughs including control due to the presence of yeast
Saccharomyces cerevisiae in all these doughs. Hexanol, propanol and heptanol
were observed in all sourdoughs with very little variation. Nonanol was
observed in LA-5 sourdough only.
Ethyl acetate was found as dominating ester followed by ethyl lactate
depending on type of cultures. Although, it was found in LA-1 sourdough but
flour extraction rate did not affect it too much.
The acids detected were acetic acid, butanoic acid, propanoic acid and
hexanoic acid. The acetic acid was found in LA-5 sourdough only, whereas
butanoic acid and propanoic acid were found in both sourdoughs.
The production of carbonyls was the highest in LA-1 sourdoughs. Other
carbonyls like nonanal and decanal were not found abundantly. The productions
of other aromatic compounds were random except benzene ethanol which was
found in more quantity in LA-1 sourdough.
4.3.2.3. Comparison of SPME and DHT for Sourdough volatile

compounds extraction
The comparison of sourdough volatiles extracted using SPME and DHT
techniques revealed that DHT resulted in more number of volatile compounds
than SPME, but compounds extracted with SPME showed higher relative peak
areas for most of the compounds. With regards to different alcohols produced,
iso-amyl alcohol, 2- butanol, propanol, octanol were detected with DHT but were
found missing with the SPME technique. The ethanol extracted with SPME
showed much higher relative peak area than DHT (Table 19). The hetero-
fermentative LA-5 showed more relative peaks areas for ethanol that increased
with increase in flour extraction rate, when extracted with SPME.
Among different acids detected in sourdough hexanoic acid was detected
with DHT but was found missing with SPME technique. Different acids showed
greater relative peak areas with SPME, compared with DHT. The homo-
fermentative strain LA-1 resulted in greater acetic acid production using SPME.
The carbonyls like butane, 3-methyl and decanal were only detected with
DHT (Table 20). Different other aromatic compounds were randomly detected
with both DHT and SPME techniques without ant specific pattern.
4.3.3. DISCUSSION
The production of volatile compounds is desired character of sourdough
fermentation that results in pleasant taste and aroma (Kim et al., 2008). The
sourdough volatile compounds like alcohols, aldehydes, ketones, esters and
sulphur are produced during fermentation process as a result of biological and
biochemical reactions and contribute to the flavor (Spicher, 1983). In present
studies, alcohols like ethanol was found in both LA-1 and LA-5 fermented
sourdoughs and the yield was mainly influenced by starter cultures. These
results are in line with the findings of Hansen and Hansen, (1994) who reported
that the starter cultures have more impact on production of alcohols in
sourdoughs. The sourdough produced more alcohols than control. The dough
fermented with LA-1 culture produced more alcohols with the highest amount
detected in sourdough made from whole meal flour.
Hetero-fermentative LAB can be differentiated from homo-fermentative
on the basis of volatile compounds production (Rehman et al., 2006). Ethyl
acetate was found as dominating ester followed by ethyl lactate depending on
type of cultures. Although, it was found in LA-1 sourdoughs but flour extraction
rate did not affect it too much (Hansen and Hansen, 1994). The results are
supported by the study of Hansen et al. (1989) that homo-fermentative LAB are
primary contributors of ethyl acetate and diacetyl. The results of present study
are also in accordance with the findings of Hansen and Hansen (1994) which
resulted in more ethyl acetate production with an increase in flour extraction
rate. The sourdoughs containing hetero-fermentative strain resulted in more
ethanol and ethyl actetate production. Gobbetti et al. (1995a) reported hetero-
fermentative Lb. brevis, as the most appropriate profiles of flavor compounds.
Non-volatile compounds including organic acids produced by both homo
and hetero-fermentative LAB decrease the dough pH and give aroma to bread
dough (Gobbetti et al., 1995a). Among non-volatiles, acetic acid, butanoic acid
and propionic acid are reported as important sourdough flavoring compounds
(Czerny and Schieberle, 2002). The acids detected were acetic acid, butanoic acid,
propanoic acid and hexanoic acid. The acetic acid was found in LA-5 sourdough
only, whereas butanoic acid and propanoic acid were found in both sourdoughs.
Corsetti et al. (1998) reported that these two acids along with other acids are
responsible for the anti-mould activity of sourdoughs. The production of acetic
acid was more in sourdough made from LA-5 starter culture, whereas LA-1
resulted in more production of hexanoic and propionic acids.
The productions of different cabonyls in the present study were the
highest in LA-1 sourdoughs. The results are supported by Rehman et al. (2006)
that homo-fermentative LAB synthesizes carbonyls and diacetyls. The results are
in line with the findings of Hansen and Hansen (1994) who reported that
carbonyls are present in homofermentative strains.
4.3.4. CONCLUSION
The analysis of volatile compounds with both SPME and DHT revealed
that DHT extracted more volatile compounds compared with SPME but relative
peak areas of volatile compounds were higher with SPME technique. The SPME
technique for the determination of volatile was found faster and simpler than
DHT. The generation of volatile compounds in the sourdoughs was influenced
by types of LAB and sourdough yeast. It may be concluded that with an increase
in flour extraction rate, concentration of volatile compounds increases in the
sourdoughs. Freeze dried cultures containing hetero-fermentative strains of LAB
show better performance than homo-fermentative in the production of volatile
compounds.
4.4. ANALYSIS OF SOURDOUGH NAAN
4.4.1. OBJECTIVE
This part of study was undertaken to determine organic acids, mineral
contents and volatile compounds of sourdough naan. The sourdough naans were
prepared from different extraction rate flours using both LA-1 and LA-5 mixed
bacterial and yeast cultures. Yeast leavened naan made from straight grade flour
was considered as control treatment.
4.4.2. RESULTS
4.4.2.1. pH and acidity of sourdough naan

Statistical analysis for pH and acidity of naan is given in Table 21. It is
evident that the treatments differed significantly (P<0.01) in respect of both pH
and acidity of the naans. The comparison of yeast leavened naans with
sourdough naans also resulted in significant differences. Two different LAB
types varied significantly (P<0.01) with regards to acidity of sourdough naan but
showed non-significant differences for pH. The flour extraction rates also
showed similar results that acidity was significantly affected, whereas pH was
found to be non-significantly effected. The interaction of LAB types with flour
extraction rates resulted in significant effect (P<0.01) for acidity of sourdough
naan but pH of the sourdough naan were non-significantly affected.
The mean values for the pH and acidity of naan are given in Table 22,
showing that yeast leavened naan showed the highest pH (5.2) and the lowest
acidity (3.5) values. The sourdough naans prepared from different extraction rate
flours using LA-1 starter culture showed acidity in the range of 3.5 to 9.5 ml,
which increased with an increase in flour extraction rate. The sourdough naan
with 100% extraction rate using LA-1 as starter culture showed the highest
acidity (9.5 ml). The pH of sourdough naan changed little with an increase in
flour extraction rate and was in the range of 4.6 to 4.8.
The mean values for pH and acidity of sourdough naan prepared from
different flours using LA-5 as starter culture are given in Table 22. The graphical
presentation of pH and acidity of sourdough naan is shown in Fig. 5 and 6
respectively. The acidity of the sourdough naan increased with an increase in
flour extraction rate. The acidity was in the range of 3.5 to 9.7 ml. The sourdough
naan prepared with 100% extraction rate flour showed the highest acidity (9.7
ml) whereas, 64% extraction rate flour sourdough naan resulted in the lowest
acidity (3.5).
4.4.2.2. Organic acids of sourdough naan

Organic acids (lactic acid, acetic acid and citric acid) in sourdough naan
were determined using HPLC. The statistical analysis for organic acids produced
by sourdough naans prepared from different extraction rate flours and cultures
is given in Table 23. The treatments showed highly significant effect (P<0.01) on
organic acids of sourdough naan. When control was compared with LAB
fermented, resulted in highly significant differences as well. Two LAB cultures
showed highly significant differences (P<0.01) for lactic acid, acetic acid and
citric acid. Different flour extraction rates resulted in highly significant effect on
organic acid production. The interaction of LAB cultures with flour extraction
rates showed highly significant (P<0.01) results as well.
The mean values for organic acids (lactic acid, acetic acid and citric acid)
in sourdough naan are presented in Table 24. Control bread showed the lowest
organic acids production for all three organic acids. The sourdough naans
prepared with different extraction rate flours and LA-1 starter culture produced
lactic acid in the range of 7975 to 16800 ppm (Fig. 7), acetic acid in the range of
515 to 1075 ppm (Fig. 8) and citric acid in the range of 450 to 950 ppm (Fig. 9). As
a result of LA-5 starter culture, lactic acid in the range of 8606 to 22600 ppm,
acetic acid in the range of 1105 to 2050 ppm and citric acid in the range of 510 to
1360 ppm were produced.
For LA-1 starter culture, the lactic acid production increased with an
increase in flour extraction rates with highest amounts of lactic acid (16800 ppm)
and citric acid (950 ppm) contents were found in 100% extraction rate sourdough
naan. However, the acetic acid content was high (1075 ppm) in 64% extraction
rate sourdough naan.
For LA-5 starter culture, the lactic acid production increased with an
increase in flour extraction rates with the highest lactic acid (22600 ppm), acetic
acid (2050 ppm) and citric acid (1360 ppm) contents were found in 100%
extraction rate sourdough naan.
4.4.2.3. Mineral contents in sourdough naan

Statistical analysis for minerals content in sourdough naan prepared with
different extraction rate flours are given in Table 25. The treatments showed
highly significant effect (P<0.01) with regards to minerals like Ca, Fe, Zn, Cu and
P. When sourdough naans were compared with control (yeast leavened naan),
resulted in highly significant (P<0.01) differences for all the minerals studied.
The LAB types did not affect the minerals content but flour extraction rates
exhibited highly significant influence on the quantities of Ca, Fe, Zn, Cu and P.
However, the interaction of LAB types with flour extraction rates are non-
significantly (P>0.05) affected the minerals.
The mean values for sourdough naan made from different extraction rate
flours are given in Table 26. The control treatment showed lower mineral
contents as compared to sourdough naan.
The sourdough naans made from different extraction rate flours and
starter culture LA-1 resulted in Ca in the range of 0.22 to 0.44 mg/g. The highest
Ca content (0.44 mg/g) was observed in sourdough naan made from 100%
extraction rate flour. The Fe contents were in the range of 19.2 to 48.4 ppm. The
100% extraction rate sourdough naan contained the highest amount of Fe (48.4
ppm). The Zn contents were found in the range of 8.5 to 28.8 ppm and the
highest Zn content (28.8 ppm) was found in 100% extraction rate sourdough
naan.
The Cu contents varied from 1.5 to 3.8 ppm and it increased with an
increase in flour extraction rate with the highest (3.8 ppm) found in 100%
extraction rate sourdough naan. The Fe contents also increased with an increase
in flour extraction rate and were found in the range of 1.44 to 4.15 mg/g. The
highest Fe contents were found (4.15 ppm) in 100% extraction rate sourdough
naan.
The sourdough naan prepared with different extraction rate flours and
LA-5 starter culture also showed increase in minerals contents with an increase
in flour extraction rate. The Ca ranged from 0.22 to 0.44 mg/g, with highest Ca
content observed in 100% extraction rate sourdough naan. The Fe content also
increased with an increase in flour extraction rate and the highest Fe content was
found (47.9 ppm) in 100% extraction rate sourdough naan, whereas the lowest Fe
content (19.7) was found in 64% extraction rate flour naan. Similarly, Zn content
increased with an increase in flour extraction rate and ranged from 8.2 to 28.5
ppm with highest (28.5 ppm) was found in 100% extraction rate sourdough naan.
The Cu and P contents ranged from 1.40 to 3.8 ppm and 1.46 to 4.20 mg/g,
respectively, with the highest Cu (3.8 ppm) and P (4.20 mg/g) contents were
found in 100% extraction rate sourdough naan.
4.4.2.4. Volatile compounds of sourdough naan
The sourdough naans prepared with different extraction rate flours and
starter culture LA-1 and LA-5 were analyzed for volatiles production. Both
SMPE and DH techniques were used for this purpose
4.4.2.4.a. Sourdough naan volatile compounds extracted with SPME technique

The volatile compounds extracted from sourdough naan using SPME
technique are given in Table 27. The volatile compounds identified were
alcohols, esters, carbonyls and other aromatic compounds. Non-volatile extracted
was acetic acid.
The alcohols included ethanol, 1-propanol, 2-methyl, 1-butanol, 2-methyl-,
1-butanol, 3-methyl and 1-hexanol. The ethanol increased with an increase in
flour extraction rates and was observed high in LA-5.
The esters identified were ethyl acetate and ethyl lactate. The carbonyls
detected were benzaldehyde, hexanal, n-hexanal, furancarboxaldehyde,
propanal, 2-methyl, butanal, 2-methyl, butanal, 3-methyl and 5-methylfurfural.
Other aromatic compound detected was benzene ethanol. The relative
peak area for benzene ethanol increased with an increase in flour extraction rates
and found randomly in all the samples.
4.4.2.4.b. Sourdough naan volatile compounds extracted with Dynamic

Headspace technique
Sourdough naans analyzed for volatile production, using DHT are given
in Table 28. Both volatile and non-volatile compounds were extracted. The
volatiles compounds identified in sourdough naans were alcohols, esters,
carbonyls and other aromatic compounds, whereas non-volatile detected was
acetic acid alone.
The alcohols identified were ethanol, 2-butanol, iso amyl alcohol, 1-
butanol, 3-methyl, 1-pentanol, 1-hexanol, heptanol, propanol and 1-octanol. The
ethanol was more abundantly produced in LA-5 sourdough naans as compared
to LA-1 sourdough naan. The production of 2-butanol was more obvious in LA-1
sourdough naan than LA-5. Similarly, iso-amyl alcohol was abundantly found
only in LA-1 sourdough naans. The alcohols like 1-pentanol, 1-hexanol, heptanol,
propanol and 1-octanol were randomly found in all the samples and were found
increasing with an increase in flour extraction rate.
The esters found in sourdough naans were ethyl acetate and ethyl lactate.
Ethyl lactate was randomly found in all the sourdough naans but was absent in
control sample. The production of carbonyls was dominant in homo-
fermentative than hetro-fermentative LAB. Acetaldehydes were found only in
LA-5 sourdough naans but the rest of the carbonyls were found in high quantity
in LA-1 sourdough naan.
Non-volatile compounds produced were acids like acetic acid, butanoic
acid, propanoic acid and hexanoic acid. The acetic and butanoic acid were
observed in LA-5 sourdough naans only. Many important aromatic compounds
were also detected in both LA-1 and LA-5 sourdough naans. The compounds
found were benzene ethanol, 2-buten-1-ol, 2-methyl, 1-octen-3-ol and 3-buten-1-
ol, 3-methyl. All these were randomly found in all the samples.
4.4.2.4.c. Comparison of SPME and DHT for Sourdough naan volatile

compounds extraction
DHT resulted in extraction of more number of volatile compounds from
sourdough naans as compared to SPME. The relative peak areas of compounds
varied randomly for both the techniques.
Among alcohols 2-butanol, 1-pentanol, heptanol, propanol and 1-octanol
were missing when sourdough naans analyzed with SPME. Iso amyl alcohol
extracted with DHT only showed very high relative peak area and was found
only in homo-fermentative made sourdough naans. Similarly esters identified
like ethyl acetate showed higher relative peak area with DHT compared to SPME
technique. Ethyl acetate showed more relative area with hetero-fermentative
strains when extracted with DHT (Table 28).
Similarly propionic acid, hexanoic acid were missing with SPME but were
extracted with DHT. Only acetic acid was extracted with both SPME and DHT.
The relative areas for acids in sourdough naans were less compared to their
respective sourdoughs.
SPME resulted in more extraction of carbonyls than DHT. Among
cabonyls benzaldehyde, 2-furancarboxaldehyde, propanal, 2-methyl, 5-
methylfurfural were extracted with SPME technique only. Carbonyls like
acetaldehydes, decanal and nonanol were found missing with SPME and were
extracted only with DHT.
Among different aromatic compounds only benzene ethanol was
extracted with both the techniques. Aromatic compounds like 2-buten-1-ol, 2-
methyl, 1-octen-3-ol and 3-buten-1-ol, 3-methyl were extracted with DHT only
(Table 28).
4.4.3. DISCUSSION
The acidities of the sourdough naans made using starters were
significantly higher compared to the control. The pH of sourdough naans
changed little with an increase in the flour extraction rates. With the increase in
flour extraction rate lower pH and higher TTA were achieved. However,
addition of starter and strain significantly influence acidity related parameters
and increase the acidity of bread (Rouzaud and Martinez-Anaya, 1997)
The results for pH and acidity are in accordance with findings of Park et
al. (2007) who reported increased acidity and lower pH values of sourdough
breads in contrast to yeast leavened bread which showed less acidity and higher
pH values. The results are also in line with the findings of Hansen and Hansen
(1996) who reported the pH of sourdough bread in the range of 5.9 (control
bread) to 4.5 in bread having 20% sourdough, and the corresponding breads
resulted in TTA values ranged between 3.3 and 4.6 ml.
The results showed that TTA value of 100% extraction rate flour
sourdough naan was almost double than sourdough naan made from SGF.
Similar results were reported by Brummer and Lorenz (1991) where acidity
values of sourdough bread made from whole meal flour were almost double the
values in sourdough bread made from straight grade flour. Brummer (1989)
suggested TTA values between 3.5 and 4.0 in order to get good taste in wheat
bread. Similarly Martinez-Anaya et al. (1990) reported lower pH and higher TTA
in breads fermented with LAB than yeast fermented breads. The pH values
ranged from 4.92 to 5.33 and TTA values for breads ranged from 3.38 to 5.13 ml.
Hansen and Hansen (1994) reported pH values in the range of 3.5 to 3.8, when
sourdoughs were prepared with different extraction rate flours. Highly
significant effect (P<0.01) on TTA was also observed and concluded that with the
increase in flour extraction rate, TTA increased.
Mould growth is a most important cause of bread spoilage, which can be
prevented by the use of homo and hetro-fermentative LAB (Spicher, 1983). This
fungistatic effect is due to the production of acetic acid by lactic acid bacteria
(Roecken, 1996). The production of organic acids by homo and hetero-
fermentative results in lowering pH and contributes to bread aroma (Gobbetti et
al., 1995a). The findings for acetic acid in the present study showed that homo-
fermentative strain LA-1 produced only small quantity of this acid, whereas
hetero-fermentative strain LA-5 produced acetic acid in appreciable amount. The
results are in line with the findings of Hansen and Hansen (1994) which showed
that acetic acid production is strain dependent and homo-fermentative strains
did not produce acetic acid except for sourdough made from straight grade flour.
The production of lactic acid and citric acids increased with an increase in flour
extraction rate in the present study indicating that lactic acid and citric acids
production is flour extraction rate dependent and their total contents were
independent of starter cultures. The use of mixed culture resulted in higher lactic
acid concentrations compared to yeast leavened bread (Plessas et al., 2008).
The results of present study are also supported by findings of Hansen et
al. (1989) in which the production of acetic acid was much higher in the dough
acidified with hetero-fermentative strains as compared to homo-fermentative
strains which produced only lactic and citric acids. The increased production of
acetic acid may improve the qualitative characteristics of sourdough naan.
The results of the present study indicated that with the increase in flour
extraction rate, minerals content of sourdough naans increased appreciably (Kent
and Amos, 1967). The flour extraction rate is the most important factor for
controlling the amount of minerals (Pederson et al., 1989). The results are in
accordance with the findings of Nielsen et al. (2007) which showed high content
of dietary fiber, Mg, P, Fe, Cu and Zn, in bread made from whole wheat flour.
With the increase in flour extraction rate minerals content increase, as
does the buffering capacity of the flour due to the phytic acid from the aleurone
layer of the cereal (Hansen, 2006). The utilization of minerals such as Fe, P, Cu
and Zn is limited due to the presence of phytic acid. The phytic acid contents are
decreased during sourdough bread making due to the presence of phytase
enzyme in sourdough (Nielsen et al., 2007). Lopez et al. (2001) reported that
sourdough fermentation reduced phytic acid content up to 62%, whereas
conventional yeast fermentation reduced it only by 38%. Since sourdough has
low pH (3.7-4.5) and high acidity, so it is more effective in reducing phytic acid
contents and increased phytase activity. This is in accordance with the study of
Fretzdorff and Brummer (1992) who found that pH was the most important
factor in reducing the content of phytic acid during bread making as phytic acid
in dough’s with pH 4.0-4.3.6 was more effectively reduced than in dough’s with
higher pH.
From the health point of view sourdough fermentation result in decrease
of phytic acid by phytate hydrolysis, thus increase mineral bioavailability and
would be beneficial and attractive in improving mineral status (Plessas et al.,
2008).
The volatile compounds of different breads have investigated widely
during the past few years (Rehman et al., 2006). These studies established that
bread volatiles belong to several chemical classes like alcohols, aldehydes,
ketones etc, depending upon the bread kind (Bianchi et al., 2008).
The results of volatile compounds in sourdough naans revealed that
amount of volatile compounds was increased in sourdough breads compared
with control. Both non-volatiles and volatiles compound were produced in
sourdough naans. Non volatiles included organic acids, which were produced by
homo and hetero-fermentative bacteria whereas, volatiles like alcohols, esters,
aldehydes, cabonyls, ketones were produced due to different biochemical
reactions during fermentation (Rehman et al., 2006). Spicher (1983) reported that
most of the volatiles are produced during baking. The production of volatile
compounds was mostly starter cultures dependent and for some compounds the
interaction of starter culture and flour type was also observed as reported by
Hansen and Hansen (1994)
The ethanol contents in sourdough naans increased with an increase in
flour extraction rates and were found high in LA-5. This increase might be due to
increase in ash contents, which increased the amount of volatile compounds
(Hansen and Schieberle, 2005). Rehman et al. (2006) reported that the interactions
between starter cultures used and flour types always play an important role in
the production of flavor compounds in the sourdough breads. The present study
resulted in increased production of alcohols with the use of mixed culture of LAB
and yeast in comparison to fermentation with yeast alone. The more alcohols
production might be due to synergetic effect of LAB and yeast. Ethyl acetates
were produced more in LA-5 which is might be a character of hetrofermentative
strain as reported by Damiani et al. (1996).
The production of alcohols and esters were much lower in sourdough
naans than corresponding sourdoughs as reported by (Hansen and Hansen,
1996; Plessas et al., 2008). This might be due to their evaporation during baking
(Hansen et al., 1989a). Similarly iso-amyl alcohol was abundantly found only in
LA-1 sourdough naans as reported by Damiani et al. (1996).
The ethyl acetate and majority of Carbonyls were detected in homo-
fermentative LA-1 sourdough naans. The results are in line with the findings of
Hansen and Hansen (1994) which reported that ethyl acetate and carbonyls are
mostly found in homo-fermentative sourdoughs. The ethyl acetate content was
much lower in sourdough naans compared with their respective sourdoughs.
Acetic and butanoic acids were found in LA-5 sourdough naans only.
Corsetti et al. (1998) reported that these two acids along with other acids are
responsible for the antimould activity of sourdoughs. This quality makes the LA-
5 sourdough naans more effective against mould development than LA-1
sourdough naans.
4.4.4. CONCLUSION
The analysis of sourdough naans revealed that with the increase in flour
extraction rate, organic acids and mineral contents increased. The analysis of
volatile compounds using both SPME and DH techniques showed that the
sourdough naans had higher content of volatiles as compared to yeast leavened,
however, their concentrations were lower than corresponding sourdoughs. It
was noticed that increase in flour extraction rates enhanced the concentration of
volatile compounds in sourdough naans. Freeze dried cultures containing
hetero-fermentative strains of LAB showed better performance in the production
of volatile compounds as compared to homo-fermentative.
4.5. SENSORY EVALUATION OF SOURDOUGH NAAN
4.5.1. OBJECTIVE
This study was carried out to determine acceptability of sourdough naan
prepared with different extraction rate flours and starter cultures through
sensory evaluation. The Sourdough naan were evaluated for their sensory
characteristics such as color, flavor, taste, texture, flexibility, chewability and
overall acceptability immediately after baking by a panel of 10 judges.
4.5.2. RESULTS
Statistical analysis for sensory evaluation of sourdough naan is given in
Table 29. All treatments differed significantly (P<0.01) with regards to all sensory
parameters. Comparison of control with sourdough naan prepared from LAB
cultures resulted in significant effect (P<0.05) on color and taste, whereas flavor,
texture, flexibility, chew ability and overall acceptability were effected highly
significantly (P<0.01). LAB cultures showed significant effect (P<0.05) on taste,
but showed highly significant results tor flavor, whereas color, texture,
flexibility, chew ability and overall acceptability varied non-significantly. The
flour extraction rates showed highly significant (P<0.01) effect on all sensory
characteristics, whereas the interaction of flour extraction rate with LAB types
showed non-significant (P>0.05) results for sensory evaluation parameters.
4.5.2.1. Color of sourdough naan

The color of naan bread is an important parameter for the consumer’s
acceptance. The sourdough naan prepared with different extraction rate flours
and starter culture LA-1 got scores for color in the range of 5.0 to 7.8 with the
highest scores (7.8) for 64% extraction rate sourdough naan (Table 30, Fig. 10).
The sourdough naans prepared with LA-5 starter culture got color scores in the
range of 4.85 to 7.85, 64% extraction rate sourdough naan stood first having
highest (7.85) scores.
4.5.2.2. Flavor of sourdough naan
Flavor is an instantaneous perception of taste and odor and difference in
taste is due to the volatile compounds in a product. The flavor of naan was also
differed significantly (P<0.01) (Table 29, Fig. 10). Mean scores for flavor were in
the range of 6.0 to 6.14. The 88% extraction rate sourdough naans got the highest
scores (6.14) as shown in Table 30. Flavor of LA-5 sourdough naan got scores in
the range of 6.14 to 7.14 and 88% extraction rate naan got first position with
highest (7.85) scores. The flavor scores of control were found the lowest.
4.5.2.3. Taste of sourdough naan

Taste of is an important sensory parameter which plays an important role
in consumer’s acceptance of the product. The mean values for taste are given in
Table 30. The taste of sourdough naan with different extraction rate flours using
LA-1 starter culture got scores in the range of 6.0 to 7.0 and the highest score (7.0)
was given to 64% extraction rate sourdough naan. The taste of sourdough naan
made from 76% extraction rate flour with having LA-5 starter culture got the
highest scores (7.85) followed by 64% extraction rate naan. The scores for taste
decreased with an increase in flour extraction rate due to the addition of more
bran particle which might have affected the taste of sourdough naan.
4.5.2.4. Texture of sourdough naan

Texture is also considered as one of the most important sensory attributes
for sourdough naan. Mean values for texture are given in Table 30. The texture
of sourdough naan made from different extraction rate flours with LA-1 culture
got scores from 5.14 to 7.00 and 64% extraction rate sourdough got the highest
(7.00) scores. The texture of sourdough naan made from 76% extraction rate flour
and fermented with LA-5 culture got the highest scores (7.85) followed by 64%
extraction rate naan. The comparison of LA-1 and LA-5 starter culture showed
that LA-5 made sourdough naan got better scores for texture as compared to LA-
1 starter culture. .
4.5.2.5. Flexibility of sourdough naan
Flexibility is the folding of naan bread piece into scoop for picking the
curry. Mean values for flexibility are given in Table 30. The flexibility or
foldingability scores of sourdough naan using LA-1 starter culture were in the
range of 5.0 to 7.85 with 64% extraction rate sourdough naan got first position,
while flexibility of sourdough naan using LA-5 starter culture got scores in the
range of 5.0 to 7.85, and 64% extraction rate sourdough naan got the highest
(7.85) flexibility scores.
4.5.2.6. Chewability and overall acceptability of sourdough naan

Although, color and other sensory attribute of naan bread are important, it
was chewability and overall acceptability which clearly differentiated sourdough
naan made from different extraction rate flours and starter cultures.
Mean values for chewability and overall acceptability are given in Table
30. For LA-1 culture, mean values for chewability and overall acceptability were
in the range of 5.0 to 7.85 and 5.14 to 7.85, respectively and 76% extraction rate
sourdough got the highest score (7.85) for chewability and overall acceptability.
Whereas for LA-5 culture chewability and overall acceptability got scores in the
range of 5.0 to 7.85 and 5.42 to 8.0, respectively and 76% extraction rate
sourdough naan got the highest scores for both chewability and overall
acceptability.
4.5.3. DISCUSSION
In the present study scores for color of sourdough naans were higher than
color scores shown by control bread, might be due to the production of lactic
acid which improved bread color (Tarar, 1999). The results of the present study
are in line with the findings of Farooq et al. (2001) who reported a decrease in
color scores of naans with the increase in flour extraction rate. The color scores
decreased with an increase in flour extraction rate due to the increase in bran
portion, which masked the color of sourdough naan at higher extraction rate
(Siddique, 1989).
Bread flavor is a key element for consumer acceptance and product
identification (Rehman et al., 2006). The results for flavor scores of sourdough
naan were much higher than that of control breads, which suggested that the
addition of sourdough improved the bread’s flavor. The scores for flavor in the
present study are in agreement with the findings of Siddique (1989) who
reported the scores of flavor range from 7.11 to 8.28 for flat bread. Sourdough
added bread was higher in sensory attributes like flavor than bread prepared
with yeast fermentation (Shim et al., 1996).
There is considerable improvement in bread flavor due to sourdough
addition (Thiele et al., 2002). The improvement in the flavor of sourdough might
be due to the production of lactic acid and acetic acid during fermentation, this
production of acids improved the volatile compounds profile thus, enhanced the
flavor of bread. The sourdough naan flavor was improved up to 88% extraction
rate flour. The higher ash content flour strongly increased the metabolic activities
like formation of acids and volatile compounds but resulting stronger flavor of
bread was not necessarily accepted and liked by the consumer (Rouzaud and
Martinez-Anaya, 1997).
The present study showed that sourdough naan made from both LA-1
and LA-5 starter culture got higher scores for taste as compared to control. The
control bread produced sweet yeasty odour and sweet bread taste, whereas
sourdough naans resulted in mild pickling and sour odour and had home made
bread taste. The taste of sourdough naans were improved up to 88% extraction
rate, higher extraction resulted in sourdough naan with extreme sour taste,
disliked by the panel. This extreme sour taste might be due to higher TTA.
Brummer and Lorenz (2003) reported that sourdough fermentation has a well-
established role in improving flavor and taste of rye and wheat breads.
Javanainen and Linko (1993) found that sourdough with pH range of 4.5 to 5.0
produce acceptable light sour taste bread. The results are in line with the findings
of Tarar (1999) who reported improvement in taste of breads with the addition of
lactic acid.
Quality of bread depends upon volume, taste and texture of breads
(William et al., 1988). The texture is a group of physical characteristics, sensed by
feeling of touch and is a multi parameter attribute (Arendt et al., 2007). The
sourdough addition resulted in softer naans than control. The increase in softness
showed the same trend as Shim et al. (1996). Production of organic acids during
sourdough fermentation helps in the swelling of gluten and increases gases
retention which produces products with good texture and increased volume
(Park et al., 2006). The addition of sourdough had a positive effect on the quality
of bread due to the production of number of metabolites by LAB (Arendt et al.,
2007). The results for the texture of sourdough naan showed improvement in
texture and palatability of bread (Plessas et al., 2008). The results in present study
are in agreement with findings of Farooq et al, (2001) who reported the naan
scores in the range of 6.53 to 8.20.
Flexibility of naan decreased with an increase in flour extraction rate due
to the increase of bran particle. The difference in flexibility of all the flat breads
may be due to the difference in the hardness of wheat grains and several other
factors like flour types and milling characteristics (Farooq et al., 2001). Similar
results were reported by Wang and Flores (1999) when tortilla made from
different wheat flours were offered to panel of judges for evaluation of
foldingability, exhibited positive correlation with flour protein, damage starch
and water absorption.
The chewability is related to hardness and softness of the naan bread
(Shim et al., 1996). The present study showed that sourdough naans had less
hardness, chewiness, compared to control. The scores for chewability indicated
that judges gave maximum scores to 76% extraction rate sourdough naan
followed by 64% extraction rate sourdough naan which were little sticky due to
the absence of bran particles.. Freshly prepared naan had maximum chewability
that gradually decreased with the passage of time due to the loss of moisture
(Farooq et al., 2001)
Color and taste of the bread are primary factors in determining the overall
acceptability (Rashid et al., 1982). The flour of high extraction rate imparts darker
color which was not attractive and got les scores. Also with the increase in flour
extraction rate, taste and other sensory attributes were also affected significantly.
The results of present study suggest that the addition of sourdough improve the
overall acceptability of sourdough naans. This is similar to the results of Shim et
al. (1996) that sourdough bread was higher in sensory tests than ordinary yeast
leavened bread. Increase in flour extraction rate up to 76% was acceptable and
got maximum scores for most of the sensory parameters. On the basis of sensory
evaluation it was concluded that sourdough naan prepared from 76% extraction
rate flour and starter culture ranked first for overall acceptability.
4.5.4. CONCLUSION
On the basis of sensory evaluation, it was concluded that sourdough naan
were preferred over control because of its better taste, flavor and other sensory
attributes. Hence, the quality of sourdough naans might be enhanced depending
upon the types of flour and starter culture used. The overall quality of
sourdough naans were affected by flour extraction rates. The sensory analysis
revealed that sourdough naan made from 76% extraction rate flour stood first for
most of the sensory attributes and hence, declared as best sourdough naan.
Table 1: Mean sum of squares for physical characteristics of wheat
varieties
SOV df Kernel Weight Test Weight
Varieties 3 9.145** 51.187**
Error 8 0.097 0.242
Total 11
** = Highly significant (P<0.01)
Table 2: Mean values for physical characteristics of different wheat

varieties
Characteristics Inqulab-91 Auqab-2000 Iqbal-2000 AS-2002
1000 kernel
42.06±0.20b 40.39±0.45c 39.22±0.28d 43.16±0.28a
weight (g)
Test weight
77.35±0.40b 71.50±0.64c 71.28±0.460c 79.43±0.43a
(Kg/hL)
Table 3: Mean sum of squares for general characteristics of SGF from
Particle
Wet Dry Falling Sedimentation Pelshenke
SOV df size
gluten gluten number value value
index
Varieties 3 13.235** 1.976* 891.63** 0.926** 36.296** 97.63**
Error 8 0.208 0.272 36.5 0.095 0.134 11.5
Total 11
* = Significant (P<0.05); ** = Highly significant (P<0.01)

SGF: Straight grade flour
Table 4: Mean values for general characteristics of SGF from different

wheat varieties
WG (%) 33.41±0.379a 29.45±0.487c 30.36±0.525b 33.56±0.423a
DG (%) 10.69±0.450a 8.72±0.851c 9.55±0.340b 9.85±0.221b
FN (sec) 532.67±6.807a 514.33±6.028b 510.00±6.557b 490.67±4.509c
PSI (%) 20.17±0.351a 19.97±0.306b 19.00±0.265c 19.27±0.306c
SDS-S (ml) 28.17±0.444c 26.39±0.445d 31.92±0.226b 34.04±0.306a
PV (min) 180.33±1.528b 172.33±4.509c 181.67±3.215b 186.00±3.606a

WG: Wet gluten
DG: Dry gluten
FN: Falling number
PSI: Particle size index
SDS-S: SDS-Sedimentation
PV: Pelshenke value
Table 5: Mean sum of squares for proximate composition of SGF from
Crude Crude Crude Total
SOV df Moisture NFE
protein fat fiber ash
Varietie
3 0.076* 0.073* 0.0056NS 0.0010NS 0.0008NS 0.080*
s
Error 8 0.034 0.027 0.091 0.0012 0.0042 0.028
Total 11
* = Significant (P<0.05); NS = Non-significant

Table 6: Mean values for proximate composition of SGF from different

wheat varieties
Moisture (%) 12.07±0.155a 11.83±0.185b 12.09±0.236a 11.78±0.151b
Crude protein (%) 11.95±0.127ab 11.77±0.125b 11.71±0.205b 12.05±0.191a
Crude fat (%) 1.33±0.070 1.35±0.133 1.29±0.101 1.40±0.060
Crude fiber (%) 0.44±0.049 0.41±0.020 0.40±0.025 0.40±0.040
Ash (%) 0.56±0.075 0.60±0.078 0.57±0.046 0.56±0.056
NFE (%) 73.66±0.295bc 74.04±0.101a 73.94±0.114b 73.83±0.061c

Table 7: Mean sum of squares for farinographic properties of SGF from different wheat varieties
Dough
Water Arrival Departure Dough Tolerance Softening
SOV Df development
absorption time time Stability index of dough
time
Varieties 3 3.600** 0.658** 1.129** 0.77** 2.698** 996.75** 327.555**
Error 8 0.1533 0.0233 0.037 0.038 0.033 11.33 8.5
Total 11
** = Highly significant (P<0.01)

Table 8: Mean values for farinographic characteristics of SGF from
WA (%) 55.33±0.473ab 53.33±0.379c 54.47±0.351b 55.83±0.351a
AT (min.) 2.47±0.153a 1.53±0.153c 1.47±0.153c 2.03±0.153b
DDT (min.) 4.63±0.153a 3.53±0.252c 4.43±0.208b 4.53±0.153ab
DS (min.) 6.67±0.153b 8.03±0.153a 8.13±0.153a 6.27±0.252b
DT (min.) 9.07±0.208b 9.43±0.208a 9.50±0.200a 8.17±0.153c
TI (BU) 39.67±2.517c 21.00±3.606d 51.00±3.606b 64.00±3.606a
SOD (BU) 59.00±3.606c 78.81±3.512b 79.67±1.528b 80.33±2.517a

WA: Water absorption
AT: Arrival time
DDT: Dough development time
DS: Dough stability
DT: Departure time
TI: Tolerance index
SOD: Softening of dough
Table 9: Mean sum of squares for sensory characteristics of flat bread
naan prepared from SGF of different wheat varieties
Overall
SOV df Color Texture Flexibility Chewability
Acceptability
Varieties 3 0.85* 0.583* 1.933** 0.5833 1.25*
Error 8 0.25 0.325 0.2 0.175 0.25
Total 11

Table 10: Mean values for sensory evaluation of flat breads prepared from
SGF of different wheat varieties
Color 6.60±0.548b 6.20±0.447c 6.60±0.548b 7.20±0.447a
Texture 7.00±0.707b 6.80±0.447c 6.60±0.548d 7.40±0.548a
Flexibility 6.80±0.447b 6.20±0.447c 5.80±0.447d 7.20±0.447a
Chewability 7.00±0.707a 6.60±0.548c 6.20±0.447d 6.80±0.447b
Overall 6.80±0.447b 6.60±0.548c 6.20±0.447d 7.40±0.548a

Acceptability
Table 11: Mean sum of squares for proximate composition of different
Crude Crude Crude Total
SOV df Moisture NFE
protein fat fiber ash
Treatments 3 2.180** 0.955* 1.023** 1.675** 0.552** 6.489**
Error 8 0.177 0.137 0.0042 0.0045 0.0022 0.267
Total 11

NFE: Nitrogen free extract
Table 12: Mean values for proximate composition of different extraction

rate flours
Wheat flour with different extraction rates

Parameters
64% 76% 88% 100%
Moisture 11.78±0.70a 11.08±0.22ab 10.42±0.26bc 9.80±0.32c
Crude protein 12.05±0.74c 12.48±0.46bc 12.92±0.44ab 13.36±0.29a
Crude fat 1.40±0.06d 1.90±0.01c 2.34±0.07b 2.76±0.09a
Crude fiber 0.40±0.04d 1.06±0.08c 1.62±0.10b 2.14±0.01a
Total ash 0.56±0.03d 0.86±0.04c 1.21±0.07b 1.55±0.04a
NFE 73.81±0.16a 72.62±0.75b 71.49±0.41c 70.39±0.56d

Table 13: Mean sum of squares for farinographic characteristics of different
SOV df WA DDT DS TI
Treatments 3 44.09* 2.388** 12.140** 217.59**
Error 8 6.15 0.090 0.072 6.86
Total 11
Table 14: Mean values for farinographic characteristics of different

Wheat flour with different extraction rates

Parameters
64% 76% 88% 100%
WA (%) 55.83±0.14c 58.30±0.23bc 61.24±4.54ab 64.72±1.98a
DDT (min) 4.53±0.09c 5.47±0.10b 6.14±0.56a 6.58±0.17a
DS (min) 6.27±0.22a 3.70±0.46b 2.26±0.15c 1.80±0.08c
TI (BU) 64.00±2.33a 62.10±1.02a 56.24±3.59b 45.08±2.84c
WA: Water absorption

DDT: Dough development time
DS: Dough stability
TI: Tolerance index
Table 15: Mean sum of squares for pH and acidity of sourdough prepared
SOV df pH Acidity
Treatments 8 0.6883** 98.880**
Control V/S LAB fermented 1 5.0417** 283.594**
LAB types 1 0.0150NS 24.604**
Extraction rates 3 0.1450** 158.204**
LAB types x Extraction rates 3 0.0050NS 2.744**
Error 18 0.0117 0.527
Total 26
** = Highly significant (P<0.01); NS = Non-significant
Table16: Mean values for pH and acidity of sourdough prepared with

Acidity (ml 0.1N
Starter cultures Extraction rates pH
NaOH)
Control 4.8±0.046a 6.8±0.162f
64% 3.6±0.023b 9.5±0.138e
76% 3.5±0.115bc 16.5±0.537d

LA-1
88% 3.4±0.035bcd 17.5±0.162cd
100% 3.3±0.052cd 20.8±0.439b
LA-5 64% 3.6±0.023b 10.1±0.335e

76% 3.5±0.046bc 18.7±0.266c
88% 3.3±0.109cd 21.4±0.843ab
100% 3.2±0.023d 22.3±0.369a

Table 17: Mean sum of squares for bacterial and yeast count of sourdough
SOV df Bacterial count Yeast count
Treatments 8 170.490** 840225.00**
LAB types 1 265.335** 3310.00**
Extraction rates 3 168.295** 44.00NS
LAB types x Extraction rates 3 37.695** 37.00NS
Error 18 1.051 45.00
Total 26
** = Highly significant (P<0.01); NS = Non-significant
Table 18: Mean values for bacterial and yeast count of sourdough prepared
Starter cultures Extraction rates Bacterial count Yeast count
Control 3.2x102g 1.6x109a
64% 2.2x108f 2.2x107b
76% 6.2x108e 3.1x107b

LA-1
88% 1.2x109d 1.9x107b
100% 2.0x109a 2.6x107b
LA-5 64% 1.4x109c 1.0x106c

76% 1.6x109b 1.5x106c
88% 1.8x109b 9.0x105c
100% 2.1x109a 6.5x105c

Table 19: Volatile compounds produced by LA-1 and LA-5 cultures in sourdough prepared from flours with
different extraction rates (SPME technique)
LA-1 LA-5
64% 76% 88% 100% 64% 76% 88% 100%
Compounds Control
ER Flour ER Flour ER Flour ER Flour ER Flour ER Flour ER Flour ER Flour
Alcohols
Ethanol 1200 1552 1562 1597 1683 1780 1876 1987 2200
1-propanol, 2-methyl 31 110 100 105 80 - - - -
1-butanol, 2-methyl- - 440 378 356 299 20 40 23 22
1-butanol, 3-methyl 77 36 94 116 129 196 376 110 175
1-pentanol 31 - 51 32 35 31 52 36 41
1-hexanol 124 92 254 170 159 150 203 142 137
Heptanol - - 25 23 26 15 23 13 17
Esters
Ethyl acetate 231 233 244 231 278 194 444 473 480
Ethyl lactate - 85 117 116 121 - - - -
Acids
Acetic acid - 163 849 251 421 224 339 491 427
Hexanoic acid - - - 18 25 - - - -
Propanoic acid - 85 91 116 - - 24 21 20
Carbonyls
Acetaldehyde 41 17 30 - 16 15 19 27 47
Benzaldehyde - - - 3
N-Hexanal - 13 15 18 23 - - - -
Aromatic compounds
Benzene ethanol - 178 212 72 81 - - 20 43
Furan, 2-pentyl - - - - - - - 3
1-octen-3-ol - - 29 - -
3-buten-1-ol, 3-methyl - - - - - - - 10 4
Control Naan bread made from Saccharomyces cerevisiae alone

LA-1 Starter culture containing Pediococcus acidilacti + Saccharomyces cerevisiae.
LA-5 Starter culture L .brevis + L .casei + Saccharomyces cerevisiae
ER Extraction rate
The amounts of volatile compounds are expressed as Relative peak are = (peak area of compound/peak area of internal standard) × 100
Table 20: Volatile compounds produced by LA-1 and La-5 cultures in sourdough prepared from flours with
different extraction rates (Dynamic Headspace technique)
LA-1 LA-5
64% ER 76% ER 88% ER 100% ER 64% ER 76% ER 88% ER 100% ER
Compounds Control
Flour Flour Flour Flour Flour Flour Flour Flour
Alcohols
Ethanol 57 107 80 71 62 41 53 55 186
2-Butanol - 2021 1344 1025 714 103 230 84 109
Iso amyl alcohol 4565 5824 - 2449 - - - - -
1-butanol, 3-methyl 6 25 67 41 32 570 1767 843 424
1-pentanol 15 21 92 44 37 58 171 78 116
1-hexanol 123 103 319 146 102 207 326 230 297
Heptanol - 11 31 16 11 12 46 16 49
Propanol 56 88 77 56 40 17 39 17 44
1-Octanol - - 16 8 7 - 27 10 -
N-octanol - - 16 8 7 - - - -
Nonanol - - - - - - 10 5 7
Esters
Ethyl acetate 920 1058 1034 826 708 120 425 730 850
Ethyl lactate - 7 9 5 - - - - -
Acids
Acetic acid - 15 19 23 4 49 10 10 16
Butanoic acid - 12 12 18 18 28 53 41 238
Propanoic acid, - 169 151 43 124 43 124 58 171
Hexanoic acid - - 38 4 - - - - -
Carbonyls
Acetaldehyde 4 78 234 220 340 55 34 72 50
Nonanal - 7 8 - 4 - 10 11 -
Butanal, 3-methyl - 15 12 -
Decanal - - 7 - - - - 10 -
Aromatic compounds
Benzene ethanol 44 74 47 22 20 - 13 - -
2-Buten-1-ol, 2-methyl - - - - - 9 16 9 -
1-octen-3-ol - - 10 5 4 8 16 7 10
3-buten-1-ol, 3-methyl - - - - - 18 32 17 29

ER Extraction rate
The amounts of volatile compounds are expressed as Relative peak are = (peak area of compound/peak area of internal standard) × 100
Table 21: Mean sum of squares for pH and acidity of sourdough naan
Acidity
SOV df pH
(ml 0.1N NaOH)
Treatments 8 0.0858* 17.143**
LAB types 1 0.0037NS 1.654**
Extraction rates 3 0.0337NS 31.914**
LAB types x Extraction rates 3 0.0037NS 2.594**
Error 18 0.0249 0.132
Total 26
* = Significant (P<0.05); ** = Highly significant (P<0.01); NS = Non-significant
Table 22: Mean values for pH and acidity of sourdough naan prepared with
Acidity
Starter cultures Extraction rates pH
(ml 0.1N NaOH)
Control 5.2±0.144a 3.5±0.046f
64% 4.8±0.063b 4.7±0.092e
76% 4.8±0.104b 5.7±0.196d

LA-1
88% 4.7±0.052b 6.9±0.069c
100% 4.6±0.052b 9.5±0.387a
LA-5 64% 4.8±0.052b 3.5±0.087f

76% 4.8±0.081b 7.2±0.248c
88% 4.7±0.156b 8.5±0.179b
100% 4.7±0.017b 9.7±0.300a
Table 23: Mean sum of squares for organic acids of sourdough naan
SOV df Lactic acid Acetic acid Citric acid
Treatments 8 86896899** 665421** 278738**
Control V/S LAB fermented 1 191032480** 319704** 349209**
LAB types 1 56742975** 3003338** 267759**
Extraction rates 3 142372240** 54113* 493134**
LAB types x Extraction rates 3 6761005** 612663** 44509**
Error 18 1087903 12797 1913
Total 26
Table 24: Mean values for organic acids of sourdough naan prepared with
Starter cultures Extraction rates Lactic acid Acetic acid Citric acid
Control 5025±253.9e 850±18.52e 380±11.63e

64% 7975±181.1d 1075±16.57d 450±14.86de
76% 9820±474.3d 920±29.48de 520±8.52d

LA-1
88% 13210±1133.8c 860±85.99e 670±23.84c
100% 16800±339.7b 515±36.89f 905±40.66b
64% 8606±133.8d 1105±42.11d 510±29.44d
76% 12960±138.4c 1365±63.05c 650±26.68c

LA-5
88% 15940±108.4b 1680±66.64b 870±32.16b
100% 22600±1221.4a 2050±134.02a 1360±21.79a

Table 25: Mean sum of squares for minerals content of sourdough naan prepared with different extraction rate
flours and cultures
SOV df Ca Fe Zn Cu P
Treatments 8 0.020** 399.195** 225.936** 2.845** 3.729**
Control V/S LAB fermented 1 0.025** 430.954** 183.707** 2.802** 3.089**
LAB types 1 0.001NS 0.004NS 0.015NS 0.001NS 0.004NS
Extraction rates 3 0.045** 920.414** 540.970** 6.645** 8.911**
LAB types x Extraction rates 3 0.001NS 0.454NS 0.285NS 0.010NS 0.002NS
Error 18 0.001 4.924 1.295 0.067 0.038
Total 26
Table 26: Mean values for minerals content of sourdough naan prepared
Starter Extraction Ca Fe Zn Cu P
cultures rates (mg/g) (ppm) (ppm) (ppm) (mg/g)
Control 0.21±0.006d 19.5±0.13d 8.3±0.18c 1.4±0.01d 1.48±0.06cd
64% 0.22±0.006d 19.2±0.61d 8.5±0.19c 1.5±0.08d 1.44±0.04d
76% 0.26±0.011c 26.5±0.32c 9.6±0.13c 1.8±0.12cd 1.82±0.06c
LA-1
88% 0.33±0.012b 34.8±1.49b 19.4±1.06b 2.7±0.17b 2.76±0.12b
100% 0.42±0.006a 48.4±1.35a 28.8±0.96a 3.8±0.17a 4.15±0.16a
64% 0.22±0.005d 19.7±0.38d 8.2±0.12c 1.4±0.03d 1.46±0.02cd
76% 0.27±0.012c 26.0±1.58c 9.8±0.39c 1.9±0.11c 1.80±0.09cd
LA-5
88% 0.32±0.011b 35.2±0.22b 20.0±0.12b 2.6±0.01b 2.82±0.16b
100% 0.42±0.029a 47.9±2.75a 28.5±1.25a 3.8±0.33a 4.20±0.17a
Table 27: Volatile compounds produced by LA-1 and LA-5 cultures in
sourdough naan prepared from flours with different extraction
rates (SPME technique)
LA-1 LA-5
76% 88% 76% 88%
64% 100% 64% 100%
Cont ER ER ER ER
Compounds ER ER ER ER
rol Flou Flou Flou Flou
Flour Flour Flour Flour
r r r r
Alcohols
Ethanol 130 50 196 118 171 170 185 245 301
1-propanol, 2-
methyl 22 - - - - - - - -
1-butanol, 2-
methyl- 348 50 196 118 171 9 - 9 -
1-butanol, 3-
methyl 152 24 97 59 78 71 75 85 40
1-hexanol 7 6 11 - 14 7 16 29 20
Esters
Ethyl acetate 10 8 9 10 12 20 26 30 34
Ethyl lactate 4 12 14 18 20 - - - -
Acids
Acetic acid 12 5 7 12 16 12 18 27 40
Carbonyls
Benzaldehyde 2 4 6 10 - 2 4 6 -
Hexanal - 82 11 10 18 10 14 27 20
N-Hexanal - - - - - 10 14 27 20
2-
furancarboxal
dehyde 26 32 114 77 35 28 75 44 68
Propanal, 2-
methyl 4 6 15 17 16 6 21 19 23
Butanal, 2-
methyl - 8 30 25 33 8 39 42 47
Butanal, 3-
methyl - 18 70 57 87 24 39 111 118
5-
methylfurfura
l - - - 9 - 16 - - -
Aromatic
compounds
Benzene
ethanol 34 45 50 55 78 20 25 34 56

ER Extraction rate
The amounts of volatile compounds are expressed as Relative peak are = (peak area of
compound/peak area of internal standard) × 100
Table 28: Volatile compounds produced by LA-1 and LA-5 cultures in
sourdough naan prepared from flours with different extraction
rates (Dynamic Headspace technique)
LA-1 LA-5
64% 76% 88% 100% 64% 76% 88% 100%
Compounds Control ER ER ER ER ER ER ER ER
Flour Flour Flour Flour Flour lour Flour Flour
Alcohols
Ethanol 127 152 80 71 58 43 50 58 531
2-Butanol - 2021 1344 1025 674 109 134 528 -
Iso amyl
4321 5824 2449 - - - - -
alcohol
1-butanol, 3-
7 2883 2242 3448 3505 602 1880 887 1448
methyl
1-pentanol 16 21 92 44 35 85 100 81 192
1-hexanol 84 103 319 146 96 300 314 242 408
Heptanol - 11 31 16 12 13 27 17 15
Propanol 57 88 77 56 38 18 23 17 80
1-Octanol - - 16 - - - 16 10 15
Esters
Ethyl acetate 22 18 39 44 53 765 826 1034 1058
Ethyl lactate - 7 18 20 8 12 14 9 -
Acids
Acetic acid 4 - 5 6 8 10 13 17 24
Butanoic acid - - - - - 23 17 42 46
Propanoic
- 182 214 169 142 52 72 61 388
acid,
Hexanoic acid - - 38 4 -
Carbonyls
Acetaldehyde 57 - - - - 76 116 76 455
Nonanal - 7 8 - 4 - 6 11 16
Butanal, 3-
- 13 10 9 - - - - -
methyl
Decanal - - 7 - - - - 10 -
Aromatic
compounds
Benzene
34 74 47 22 19 14 9 12 15
ethanol
2-Buten-1-ol, 2-
- - - - - - 9 11 -
methyl
1-octen-3-ol - - 10 5 4 9 9 8 11
3-buten-1-ol, 3-
- - - - - 28 19 18 63
methyl

ER Extraction rate
Relative peak are = (peak area of compound/peak area of internal standard) × 100
Table 29: Mean sum of squares for sensory characteristics of sourdough naan prepared with different extraction
rate flours and cultures
Overall-
SOV Df Color Flavor Taste Texture Flexibility Chewability
acceptability
Treatments 8 8.49** 7.97** 3.10** 8.09** 11.09** 7.84** 4.65**
Control V/S LAB fermented 1 1.93* 40.35** 3.03* 1.84** 4.31** 2.21** 3.38**
LAB types 1 0.01NS 7.43** 3.55* 0.004NS 0.006NS 0.001NS 0.008NS
Extraction rates 3 22.0** 4.63** 4.95** 20.94** 28.14** 20.15** 11.25**
LAB types x Extraction rates 3 0.01NS 0.71NS 1.12NS 0.004NS 0.006NS 0.009NS 0.025NS
Error 54 0.29 0.36 0.52 0.26 0.30 0.16 0.21
Total 62
Table 30: Mean values for sensory characteristics of sourdough naan prepared with different extraction rate flours
and cultures
Starter Extraction Overall-

Color Flavor Taste Texture Flexibility Chewability
cultures rates acceptability
Control 7.14±0.16b 4.14±0.16d 6.00±0.31c 7.00±0.23b 7.14±0.31b 7.00±0.25b 5.85±0.23c
64% 7.85±0.09a 6.14±0.08c 7.00±0.23b 7.00±0.14b 7.85±0.14a 7.42±0.08b 7.00±0.27b
76% 7.00±0.25b 6.00±0.25c 6.85±0.32b 7.85±0.15a 7.00±0.31b 7.85±0.14a 7.85±0.07a
LA-1
88% 6.28±0.19c 6.85±0.33b 6.28±0.21c 6.00±0.15c 5.57±0.24c 6.14±0.12c 6.28±0.15c
100% 5.00±0.32d 6.00±0.22c 6.00±0.25c 5.14±0.12d 5.00±0.17c 5.00±0.08d 5.14±0.13c
64% 7.85±0.07a 7.14±0.22b 7.00±0.30b 7.00±0.24b 7.85±0.09a 7.00±0.22b 7.14±0.21b
76% 7.00±0.23b 7.00±0.22b 7.85±0.07a 7.85±0.09a 7.14±0.20b 7.85±0.10a 8.00±0.10a
LA-5
88% 6.14±0.22c 7.85±0.14a 7.14±0.31b 6.14±0.19c 6.00±0.13c 6.57±0.12c 6.57±0.16c
100% 4.85±0.19d 6.14±0.31c 6.14±0.34c 5.00±0.31d 5.00±0.18c 5.00±0.14d 5.42±0.16c
Fig.1: Farinograms for different Pakistani wheat varieties
Inqulab-91 Auqab-2000 Iqbal-2000 AS-2002
9.0
8.0
7.0
Sensory scores
6.0
5.0
4.0
3.0
2.0
1.0
0.0
Color Texture Flexibility Chewability Overall
Accept.
Sensory characteristics
Fig. 2: Sensory scores for flat breads prepared from different

wheat varieties
Fig. 3 pH of sourdough prepared from flours with different
extraction rates as influenced by various starter cultures
3.7
LA-1 LA-5
3.6
3.5
3.4
pH value
3.3
3.2
3.1
3.0
2.9
64% 76% 88% 100%
Extraction rates
Fig. 4 Acidity of sourdough prepared from flours with
different extraction rates as influenced by various starter
25.0 cultures
LA-1 LA-5
Acidity (ml 0.1N NaOH)
20.0
15.0
10.0
5.0
0.0
64% 76% 88% 100%
Extraction rates
Fig. 5 pH of sourdough naan prepared from flours with
cultures
4.9
LA-1 LA-5
4.8
pH value
4.7
4.6
4.5
4.4
64% 76% 88% 100%
Extraction rates
Fig. 6 Acidity of sourdough naan prepared from flours with
cultures
12.0
LA-1 LA-5
10.0
Acidity (ml 0.1N NaOH)
8.0
6.0
4.0
2.0
0.0
64% 76% 88% 100%
Extraction rates
Fig. 7 Lactic acid contents of sourdough naan prepared from
flours with different extraction rates as influenced by various
starter cultures
25000.0
LA-1 LA-5
22500.0
20000.0
Lactic acid (ppm)
17500.0
15000.0
12500.0
10000.0
7500.0
5000.0
64% 76% 88% 100%
Extraction rates
Fig. 8 Acetic acid contents of sourdough naan prepared from
starter cultures
2500.0
LA-1 LA-5
2250.0
2000.0
Acetic acid (ppm)
1750.0
1500.0
1250.0
1000.0
750.0
500.0
250.0
0.0
64% 76% 88% 100%
Extraction rates
Fig. 9 Citric acid contents of sourdough naan prepared from
starter cultures
1600.0
LA-1 LA-5
1400.0
1200.0
Citric acid (ppm)
1000.0
800.0
600.0
400.0
200.0
0.0
64% 76% 88% 100%
Extraction rates
Fig.10 Sensory characteristics of sourdough naan prepared
9
8
7
6
Scores
5
4
3
2
1
0
Color Flavor Taste Texture Flexibility Chewability Overall-
acceptability
Sensory attributes
Control LA-1 64% LA-1 76% LA-1 88% LA-1 100% LA-5 64%
LA-5 76% LA-5 88% LA-5 100%
Chapter-5
SUMMARY
Naan is a yeast/chemical leavened commercial flat bread of Pakistan,

prepared from lower extraction rate wheat flour. Sourdough breads are
produced in the world but previously, no work has been done on the production
of sourdough naan. This project was designed to determine the suitability of
different Pakistani wheat varieties for the production of naan and to evaluate the
effect of wheat flour extraction rates and mixed starter cultures of yeast and LAB
on the quality of sourdough naan.
Pakistani wheat varieties namely Inqulab-91, Auqab-2000, Iqbal 2000, and
AS-2002 were analyzed for thousand kernel weights, test weight, wet and dry
gluten, falling numbers, particle size index, SDS-sedimentation value, pelshenke
value, proximate composition and rheological characteristics. The naans were
prepared following the traditional method based on yeast fermentation process
and evaluated for various sensory attributes like color, texture, flexibility,
chewability and overall acceptability by a panel of 10 judges on 9-point hedonic
scale.
Wheat varieties were found to be significantly different with regards to
physical, milling and rheological characteristics. The variety; AS-2002 showed
the highest thousand kernel and test weights as well as wet gluten content, SDS-
sedimentation and pelshenke values. Among the proximate composition and
rheological characteristics, AS-2002 had the highest crude protein, water
absorption, tolerance index and softening of the dough. The naans prepared from
AS-2002 were awarded the highest scores for color, texture, flexibility and overall
acceptability.
On the basis of physico-chemical, rheological and sensory characteristics,
the wheat variety AS-2002 was selected for further studies. It was milled to get
64, 76, 88 and 100% extraction rates flour by proportional mixing of the by-
product of wheat with 64% extraction rate flour. These flours were analyzed for
proximate composition and rheological characteristics.
The extraction rates significantly influenced the proximate composition
and rheological characteristics of the flours. The flour having 100% extraction
rate had the highest protein, fat, fiber and ash contents. The water absorption
and dough development time were increased while the dough stability and
tolerance index were decreased with an increase in the extract rate. The water
absorption and dough development time were found to be the highest in flour
with 100% extraction rate.
Freeze dried cultures namely; LA-1, containing homo-fermentative strain
Pediococcus acidilacti and yeast Saccharomyces cerevisiae and LA-5, mixed culture
containing hetero-fermentative strain L. brevis and homo-fermentative strain L.
casei with yeast Saccharomyces cerevisiae were procured from Lallemand Baking
Solutions, Montreal, Canada for sourdough naan preparation.
Sourdoughs were prepared from flours with 64, 76, 88 and 100%
extraction rates using both LA-1 and LA-5 cultures. Acid contents, bacterial and
yeast counts of sourdough samples were determined. The pH of the sourdoughs
decreased and acidity increased significantly with an increase in flour extraction
rate. Highly significant results were obtained when LAB fermented dough was
compared with yeast fermented. Regarding the bacterial and yeast counts, all the
sourdough samples varied significantly. The bacterial count increased with an
increase in flour extraction rate for both the cultures.
The sourdough volatiles compounds were collected by both Solid Phase
Microextraction (SPME) and Dynamic Headspace (DH) techniques and analyzed
by gas chromatography with identification based on GC-retention times for
reference compounds and GC-mass spectrometry.
The volatile compounds extracted using SPME technique were alcohols,
esters, acids, carbonyls and aromatic compounds. The alcohols produced were
mainly ethanol, 1-propanol, 2-methyl, 1-butanol, 2-methyl-, 1-butanol, 3-methyl,
1-pentanol, 1-hexanol heptanol. The ethanol was found in both LA-1 and LA-5
fermented sourdoughs. The esters identified include ethyl acetate and ethyl
lactate. The ethyl acetate was present in both LA-1 and LA-5 fermented dough,
whereas ethyl lactate was not observed in LA-5 fermented dough. The
production of ethyl acetate increased with an increase in flour extraction rate.
The carbonyl identified were acetaldehydes, benzaldehydes and N-hexanal. The
productions of hexanal were more effected by starter cultures rather than flour
extraction rate. Both acetaldehydes and hexanal were produced in higher
amounts in LA-1 fermented dough as compared to the one fermented with LA-5.
The volatiles produced using headspace technique were alcohols, acids,
esters carbonyls and many other aromatic compounds. Alcohols produced were
mainly ethanol, Isoamyl alcohol, 2-Butanol, 1-butanol, 3-methyl, 1-pentanol, 1-
hexanol, heptanol, propanol, 1-octanol, N-octanol and nonanol. Ethyl acetate was
found as dominating ester followed by ethyl lactate depending on type of the
cultures. Although, it was found in LA-1 sourdoughs but flour extraction rate
did not affect it too much. The acids detected were acetic acid, butanoic acid,
propanoic acid and hexanoic acid. The acetic acid was found in LA-5 sourdough
only, whereas butanoic acid and propanoic acid were found in both sourdoughs.
The production of cabonyls was the highest in LA-1 sourdoughs.
The sourdough naans were prepared using different extraction rate flours.
Organic acids (lactic acid, acetic acid and citric acid) of sourdough naan were
determined using HPLC and volatile compounds collected by DM and SPME
techniques, using gas chromatography. The sourdough naans were also analyzed
for minerals contents as well as sensory perception.
All types of sourdough naans differed significantly (P<0.01) in case of pH
and acidity. Starter cultures as well as flour extraction rates significantly
influenced the production of organic acids in sourdough naans. The naans
prepared with yeast fermented dough showed the lowest organic acids
production. The sourdough naans prepared with different extraction rate flours
and LA-1 starter culture produced lactic acid in the range of 7975 to 16800 ppm,
acetic acid in the range of 515 to 1075 ppm and citric acid in the range of 450 to
950 ppm. The LA-5 starter culture produced lactic acid in the range of 8606 to
22600 ppm, acetic acid in the range of 1105 to 2050 ppm and citric acid in the
range of 510 to 1360 ppm. On the whole, the production of organic acids
increased with an increase in flour extraction rate and freeze dried cultures
containing hetero-fermentative strains of LAB showed better performance than
homo-fermentative in the production of organic acids.
The sourdough naans prepared with different extraction rate flours
showed highly significant effect on minerals contents. When sourdough naans
were compared with that produced with yeast leavened exhibited highly
significant differences in all the minerals. Although, LAB types did not influence
the minerals content but flour extraction rates exhibited highly significant effect
on Ca, Fe, Zn, Cu and P contents.
The volatile compounds extracted from sourdough naans using SPME
technique were identified as alcohols, esters, carbonyls and other aromatic
compounds. Non-volatile extracted was acetic acid. The alcohols included
ethanol, 1-propanol, 2-methyl, 1-butanol, 2-methyl-, 1-butanol, 3-methyl and 1-
hexanol. The ethanol increased with an increase in flour extraction rates and was
found high in LA-5. The esters identified were ethyl acetate and ethyl lactate.
The ethyl acetate was produced more dough fermented with LA-5. Majority of
carbonyls were detected in homo-fermentative LA-1 sourdough naans. The
carbonyls varied randomly in samples with regards to relative peak areas but
majority of them were found absent in yeast bread. The other aromatic
compound detected was benzene ethanol.
The volatiles compounds identified using DHT were alcohols, esters,
carbonyls and other aromatic compounds whereas non-volatile detected was
acetic acid alone. The ethanol was more abundantly produced in LA-5
sourdough naans as compare to LA-1 sourdough naans. The production of 2-
butanol was more obvious in LA-1 sourdough naans than that produce with LA-
5. Similarly, iso-amyl alcohol was abundantly found only in LA-1 sourdough
naans. The esters found in sourdough naans were ethyl acetate and ethyl lactate.
The LA-5 sourdough naans abundantly produced ethyl acetate. Non-volatile
compounds produced were acetic acid, butanoic acid, propanoic acid and
hexanoic acid. The acetic and butanoic acids were only found in LA-5 sourdough
naans.
On sensory evaluation, a significant difference was found in all types of
sourdough naans for all the parameters. The sourdough naans were preferred
over control because of its better taste, flavor and chewability. The sourdough
naans prepared from 76% extraction rate flour were awarded the highest scores
for most of the sensory attributes and hence declared as best sourdough naan.
Conclusions:
Following conclusions are drawn from these studies:
• Wheat variety AS-2002 is found better for the production of the naans.
• Increase in extraction rate enhances the nutritional value of the naans.
• Wheat flour of 76% extraction rate is found better than straight grade and
whole wheat flours for the production of the sourdough naans on the
basis of composition and sensory acceptability.
• Increase in flour extraction rates enhances the concentration of organic
acids and volatile compounds in the sourdough naans.
• SPME technique is found faster and simpler in the determination of
volatiles than Dynamic Headspace but, the later shows more efficiency.
• Freeze dried cultures containing hetero-fermentative strains of LAB show
better performance than homo-fermentative in the production of organic
acids and volatile compounds as well as sensory characteristics.
Recommendations:
Hence, it is recommended that:
• Sourdough naans of high extract rate flour should preferably be
consumed as compared to traditional yeast leavened naan due to their
better flavor, taste, softness and health benefits.
• LAB strains should be isolated and produced indigenously for commercial
production of sourdough naans which results in the saving of foreign
exchange.
• Mixed cultures of yeast and LAB should be employed for the commercial
production of sourdough naans.
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Fig. 9 Citric acid contents of sourdough naan prepared from
starter cultures
1600.0
LA-1 LA-5
1400.0
1200.0
Citric acid (ppm)
1000.0
800.0
600.0
400.0
200.0
0.0
64% 76% 88% 100%
Extraction rates
Appendix-I
Sensory evaluation score card for flat bread (Naan)
Name of the Judge
Date & Time
Signature
Parameters A B C D E F G H I
Color
Flavor
Taste
Texture
Flexibility
Chewability
Overall
Acceptability
9-Points Hedonic Scale
9. Like extremely 8. Like very much

7. Like moderately 6. Like slightly
5. Neither like nor dislike 4. Dislike slightly
3. Dislike moderately 2. Dislike very much
1. Dislike extremely
Directions
Evaluate these samples for given sensory parameters and score according
to 9-point Hedonic Scale
Rinse mouth each time with distilled water

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Effect of Wheat Flour Extraction Rates

A dissertation submitted in partial fulfillment of the requirements

NATIONAL INSTITUTE OF FOOD SCIENCE AND TECHNOLOGY

Tab. No. Title Page

Fig. No. Title Page

1 Farinograms for different Pakistani wheat varieties 59

Bread is consumed in large quantity in the world in different types and

The main objectives of the study were:-

Wheat (Triticum aestivum) is a staple food in many parts of the world. In

2.2 Wheat and its composition

2.1 Flat breads

2.2 Wheat and its composition

2.3 Wheat flour extraction rate

2.4.1 Types and classification of sourdough

2.4.2 Use of sourdough in cereal products

2.4.3 Sourdough fermenting microorganisms

2.4.4 Yeast in sourdough

2.5 Beneficial effects of sourdough fermentation

2.5.1 Heath benefits of sourdough bread

2.5.2 Improving nutritional and sensory quality

2.5.3 Dough structure and bread characteristics

2.5.4 Microbial spoilage

2.6 Flavoring compounds and sourdough fermentation

2.6.1 Solid Phase Microextraction (SPME)

2.7 Organic acids in sourdough

2.8 Sensory evaluation of bread

MATERIALS AND METHODS

3.1 Procurement of wheat varieties

Wheat varieties namely Inqulab-91, Auqab-2000, Iqbal 2000, and AS-2002

3.1.1 Physical characteristics of wheat

3.1.2 Thousand kernel weights

3.1.3 Test weight

3.2 Milling characteristics of wheat

3.2.1 Milling of wheat

One kg of each sample of wheat was cleaned and tempered to 14%

3.2.2 Wet and dry gluten

Falling numbers were determined using Pertin Falling Number Apparatus

FN (14% moisture basis) = FN as is x (100-14) / (100-moisture of sample in %)

3.2.4 Particle size index (PSI)

3.2.5 SDS - sedimentation value

3.2.6 Pelshenke value

The pelshenke value of each wheat variety was determined according to

3.3 Proximate analysis of wheat flour

3.3.1 Moisture Content

3.3.2 Crude protein content

3.3.3 Crude fat content

3.3.4 Crude fiber content

3.3.5 Ash content

3.3.6 Nitrogen Free Extract (NFE)

NFE % ═ 100 – (moisture % + ash % + crude protein % + crude fat % + crude

3.4 Rheological characteristics

3.4.1 Water absorption

3.4.2 Arrival time

3.4.3 Dough development time

3.4.4 Dough stability

3.4.5 Departure time/dough resistance

3.4.6 Tolerance index (TI)

3.4.7 Softening of the dough

3.5 Preparation of flat breads (Naan)

Varieties 3 9.145 51.187