CHEMICAL ANALYSIS OF FOOD

Upon completion of this course, the students will be

able to:
i) Describes the properties of moisture, protein,
carbohydrates, lipids, fiber, ash, minerals and
vitamins.
ii) Explain the principle and applications of analytical
methods for food component.
iii)Analyze and evaluate the compositions of food
products .
iv)Interpret the results and express scientific ideas in
written form
WHY DO WE NEED TO
UNDERSTAND FOOD
COMPOSITION ?
 WHAT MUST A FOOD
SCIENTIST BE ABLE TO
DO ?
THE 6 BASIC TYPES OF
ANALYSIS
 Proximate Analysis
● Refers to analysis of foodstuffs to establish their
composition.

● It provides information related to the content of:

1. Moisture
2. Crude protein (Nitrogen)
3. Crude fat
4. Ash
5. Carbohydrate
__________________
Total = 100%
Sampling, Preparation
and Preservation of
Food Samples
● The objectives:

i) able to understand definition of sampling

ii) able to describe different sampling
methods
iii)able to describe the preservation of food
sample prior to analysis.
iv)able to analyse the data using the
suitable statistical tools
 Introduction

!To control food quality and acceptance to

satisfactory limits, it is important to monitor
the crucial characteristics of raw products,
ingredients and processed foods.

!This can be done by evaluating all foods or

ingredients from a particular lot provided the
analytical technique is rapid and non-
destructive.
● How, this is practically impossible because
many analytical techniques used are time
consuming, expensive or labor intensive and
so it is not economically feasible to analyze
large amounts of material.

● It is therefore, a normal practice to select a

fraction/portion from the whole material for
analysis, and to assume that its properties are
representative of the whole material (quality
of selected portion is similar of the whole lot).
 SAMPLING

What is sampling?

A process of obtaining a portion or

sample that is representative of the
total quantity from which the sample is
obtained.
● The main objective of sample selection is to ensure
that the properties of the laboratory sample are
representative of the properties of the population,
otherwise erroneous results will be obtained.

● Selection of a limited number of samples for analysis is

of great benefit because it allows in reduction in time,
expense and personnel required to carry out the
analytical procedure, while still providing useful
information about the properties of the population.
● To ensure that the estimated value obtained from the
laboratory sample is a good representation of the true
value of the population it is necessary to develop a
sampling plan.

● Sampling plan- written document that contains

precise details of sample size, the locations from
which the sample should be selected, the method
used to collect the sample, and the method used to
preserve them prior to analysis.

● It should also specify the required documentation of

procedures carried out during the sampling process.
 Purpose of Analysis
! Official sample
official or legal requirements by government
laboratories -to ensure that manufacturers are
supplying safe foods that meet legal and labeling
requirements.

! Raw materials
Prior acceptance by a factory, or before use in a
particular manufacturing process, to ensure that
they are of an appropriate quality.
● Process control sample
A food is often analysed during processing to ensure
that the process is operating in an efficient manner.
Thus if a problem develops during processing it can
be quickly detected and the process adjusted so
that the properties of the sample are not adversely
effected.

● Finished product
Samples of the final product are usually selected and
tested to ensure that the food is safe, meets legal
and labeling requirements, and is of a high and
consistent quality.
! Research and development
Samples are analyzed by food scientists involved in
fundamental research or in product development
 Process in sampling involves three main stages
which are:

1.Identifying the population from which the

sample will be obtained (homogeneous or
heterogeneous)
2.Obtaining gross samples from total population
by using different sampling methods (random,
stratified, systematic, clusters etc.)
3.Reduce the bulk sample to a laboratory-size
suitable for analysis
A. Identify the population from which the
sample will be obtained.

Define the nature of the population from

which samples are to be selected before
deciding on the type of sampling plan to
use.
Homogenous or Heterogeneous Populations

a) Homogeneous population
● The properties of the individual samples are the same at every
location within the material .

● Sample can be taken from any location and analytical data

obtained will be representative of the whole.

● However, most population that are sampled are heterogeneous

b) Heterogeneous population
! The properties of the individual samples vary
with location (e.g. a truck full of potatoes,
some of which are bad).

! Therefore, the location within population

where a sample is taken will affect the
subsequent data obtained.
B. Sampling methods
! Several sampling methods/techniques in
common use are probability sampling and
non-probability sampling:

a) Probability sampling
Probability sampling is used when a
representative sample is desired. It uses
principle of statistical sampling and probability
to eliminate human bias.
 Probability sampling
1. Random sampling

● Each unit in the population is assigned a number.

● Samples are drawn randomly using either a

random number tables or computer generated
random numbers.

● Every sample has an equal chance of being

selected and included in the sample.

● Units corresponding to the random numbers are

then analysed as an estimate of the population.
 2. Systematic sampling
" Systematic sampling is used when a complete list of sample units
is not available, but when samples are distributed evenly over
time or space, such as on a production line.

" The first sample is selected at random and then every nth unit
after that are drawn systematically with location or time .

● An example, every 10th box in a truck may be analyzed,

or a sample may be chosen from a conveyor belt every 1
minute.
 3. Stratified sampling

" Involves dividing the population into subgroups so

that each subgroups is as homogeneous as possible.

" Random samples are then taken from each

subgroup. The procedure provides a representative
sample because no part of the population is
excluded.

" Subgroups may be regional, seasonal, retail sale

point, etc., as defined by knowledge of the food
being studied.
 4. Cluster sampling

! The entire population is divided into pre-existing

segments called so that cluster’s characteristics
are as identical as possible. Most often these
clusters are geographic.

! Clusters should be small and having a similar

number of units in each cluster.

! The clusters are randomly selected, and every

member of each selected cluster is included in
the sample.
 Non-probability sampling

" Non-probability sampling is used when it is not

possible to collect a representative sample, or a
representative sample is not desired.

" For example, in case of adulteration such as rodent

contamination, the objective of the sampling plan
may be to highlight the adulteration rather than
collect a representative sample of the population.

" The sample collector uses judgement rather than

statistical considerations in the selection of the
sample.
 1. Convenience sampling

"Convenience is performed when ease of sampling is

the key factor. The first pallet in a lot or the sample that
is most accessible is selected.

"This type of sampling will not be representative of the

population, and therefore is not recommended.
 2. Judgement sampling

! Judgement sampling is solely at the discretion of the

sampler and therefore is highly dependent on the
person taking the sample.

! This method is used when it is the only practical way

of obtaining the sample.
 C. Preparation of laboratory samples
Sample Definition
Primary sample The collection of one or more units
initially taken from the total population
of the food
Reduced sample A representative part of the primary
sample obtained by a division or
reduction process
Laboratory sample The sample sent to or received by the
laboratory
Analytical sample The portion prepared from the
laboratory sample from which the
portions for analysis are taken

Analytical portion The quantity of food of the proper

weight for each analytical measurement
 Preparation of laboratory samples

Before sample blending is done, it is necessary to

wash, remove or drain irrelevant extraneous
matter. An examples as follows:

i) soil or sand that adheres to fresh fruit or

vegetables can be removed by washing or wiping
the outer surface.
ii)shells are usually separated from nut kernels and
pits from stone fruits.
iii) Large fish are cleaned, scaled, and eviscerated,
while small fish can be blended whole.

iv) Shellfish are shucked, eggs are broken to isolate the

liquid interior.

v) Meat is removed as completely as possible from

bone.
vi) suspended matter or sediment present in
liquids such as, juice and cooking oil is removed
by filtration or separated by centrifugation.

vii) canned fruit and vegetable products may be

drained through screens if it is not necessary to
analyse the composite sample
 Reducing of particle size
● The food material within the sample population or
within individual units are heterogeneous. For this
reason it is usually necessary to make samples
homogeneous before they are analysed, otherwise
it would be difficult to select a representative
laboratory sample from the sample.

● A number of mechanical devices have been

developed for homogenizing foods. The type used
depends on the properties of the food being
analyzed (e.g., solid, semi-solid, liquid).
" Homogenization can be achieved using
mechanical devices for examples: grinders, mixers,
slicers, blenders.

" Liquid samples can be mixed using magnetic stirrer

or sonic oscillators.
 Reducing Sample Size

● Once the sample has been made homogeneous,

it will have properties which are representative of
the population from which it was originally
selected.

● There are a few methods for reducing the size of a

sample in order to obtain reliable and repeatable
results.
 A. Powder or ground materials

1. Coning and Quartering (Manually)

●form a conical heap of a granular or powdered

and spread it into a circular, flat cake.

●the cake is divided radially into quarters and two

opposite quarters are combined. The other two
quarters are discarded.

● if the sample is too large, then it can be coned

and quartered again until the desired sample size
is obtained.
 2. Mechanically
Using mills having stream diverters.
An example: Rotary Riffle
• In these riffles, the material to be sampled is fed from
a feed hopper to a feeder.
• The feeder drops the material at a uniform rate into
a series of bins on a rotating table.
• The turntable speed is set so that each sample
container will pass under the end of the feeder
numerous times before the feed hopper is emptied
into a different container.
 B. Liquids

! Liquid samples are thoroughly mixed before a

subsample is removed with a syringe-type sampler
or by submerging a container under the liquid’s
surface (a so-called “grab” sample)

! Frozen, crystalised fluids must be liquefied

completely and mixed
! Laboratory sample will have properties which are
representative of the population from which it was
originally selected.

! It will be homogenous sample of about 250-500 g/

mL which will be sufficient for analysis.
 Sample Identification

Laboratory samples should always be labeled carefully

so that if any problem develops its origin can easily be
identified. The information used to identify a sample
includes:

a) Sample description
b) Time sample was taken
c) Location sample was taken from
d) Person who took the sample
e) Method used to select the sample
Hammer mill

Pin mill

Vertical mill
 Problems in sampling
! Difficult in obtaining representative samples.

! Loss of plant material during sampling processes

and unnable to avoid the removal of extraneous
material from plants without removal of the
constituents.

! Enzymatic changes which have effects on the

food composition.

! Metal contamination during grinding.

 Preventing changes in food samples.

! Once sample have been selected, ensure that

samples does not undergo any significant changes
in its properties from the moment of sampling to the
time when the actual analysis is carried out.

! There are many possible ways for changes in food

composition which are due to enzymatic, chemical,
microbial or physical changes.

! A number of methods can be applied to minimise

these changes prior to analysis
 The changes that might occurs due to the factors
involved and possible methods to prevent
these changes from happening are as
follows:

1. Enzymatic Changes
Problem:
Many foods contain active enzymes that can cause changes in the
properties of the food prior to analysis.

Examples of the enzymes are proteases, cellulases, lipases, etc.

These enzymes alters the characteristics of the analysed

compound for examples protein, carbohydrates and lipids which
lead to erroneous data. Therefore, these enzymes should be
inactivated or eliminated.
 Proposed Method

✓ Freezing, drying (lowering the water activity)

✓ Inactivated using heat treatment (denatured the
enzymes)
✓ chemical preservatives (or a combination) are often
used to control enzyme activity.
✓ stored at low temperature (-20oC to -30oC)

Method used depending on the type of food being

analyzed and the purpose of the analysis.
2. Lipid Changes

Problem:

The content of unsaturated lipids in food may

be altered or reduced by various oxidation
reactions. Examples are exposure to light,
elevated temperatures, oxygen or pro-oxidants
can increase the rate of fats oxidation.

.
 Proposed Method

✓ Samples that have high unsaturated lipid

contents should be stored under nitrogen or
some other inert gas and placed in dark
rooms or covered bottles with aluminium foil
and stored at refrigerated temperatures.

✓ Antioxidants may be added to retard

oxidation process provided that the
antioxidant will not interfere with the
analysis.
3. Microbial Growth and Contamination

Problem

Microorganisms are present naturally in many

foods and if they are not controlled they can
alter the composition of the sample to be
analyzed.
 Proposed Method

✓ Freezing, drying (lowering the water activity of the

food and retard the growth of microbial spoilage)
✓ heat treatment (kill some of the microorganisms)
✓ chemical preservatives like sodium benzoate (or a
combination) are often used to control the growth
of microbes in foods.
4. Physical Changes

Problem

A number of physical changes may occur in a

sample, for examples, water may be lost due to
evaporation or water gained due to
condensation.

Other changes include fat or ice may melt or

crystallize resulted in changes of sample
structural properties.

.
Proposed Method

✓ Controlling the storage temperature of the

sample.
 Storage of the analytical samples

! Sampling preparation usually mean that

analytical samples stored at proper storage
conditions prior to analysis. At least three replicate
samples should be stored.

● Samples are frozen storage at –20 or even –30 °C,

which is a current common practice.
! The samples must be closely sealed in a container
with minimum of headspace.

! Air-dried samples should be stored in such a way as

to prevent uptake of water or contamination by
insects or mites.
 Food commodities

! Roasted coffee / Tea ! Eggs

! Chocolate products ! Meat and poultry
! Flour ! Fish and seafood
! Bread ! Fruits
! Dairy products ! Vegetables
!Liquid milk ! Nuts
!Powdered milk
!Condensed milk ! Sugar and
!Butter confectionary
!Cheese ! Spices
!Ice cream
! Frozen food products
Evaluation of
Analytical Data
 Data Analysis

!Food analysis usually involves making a number

of repeated measurements on the same sample
to provide confidence that the analysis was
carried out correctly and to obtain best estimate
of the value being measured and a statistical
indication of a reliability of the value.

A variety of statistical techniques are available

that enable us to obtain this information about
the laboratory sample from multiple (2 or more)
measurements.
● A minimum of three assays are typically
performed

● Varieties of statistical analysis techniques are

available to obtain statistical indication of
the reliability of the value:
 Mean (Measures of central tendency)

Mean is the best experimental estimate of the value that

can be obtained from the measurements.

It does not necessarily have to correspond to the true value

of the parameter one is trying to measure.

mean = x1 + x2 + x3 + … + xn = ∑ xi
n n

∑, represents the summation

xi, represents scores
n, represents number of scores.
 Accuracy and precision

! Accuracy - the degree of

conformity of a measured/
calculated quantity to its actual
(true) value

! Precision - the degree to which

further measurements or
calculations will show the same or
similar results
! There may be some form of systematic error in our
analytical method in which the measured value is
not the same as the true value (see below).

! Accuracy refers to how closely the measured value

agrees with the true value. The problem with
determining the accuracy is that the true value of
the parameter being measured is often not known.

! Nevertheless, it is sometimes possible to purchase or

prepare standards that have known properties and
analyze these standards using the same analytical
technique as used for the unknown food samples.
2. Absolute Error (Eabs)
! Can be obtained by the difference
between the true value (xtrue) and the
measured value (xi):
Eabs = (xi - xtrue).

! For these reasons, analytical instruments

should be carefully maintained and
frequently calibrated to ensure that they
are operating correctly.
3. Standard Deviation:
● Measures the spread of the experimental
values

● Gives a good indication of how close the

values are to each other

● SD = √ ∑ (xi - x )2
n -1
Coefficient of variation (CV)

CV = Std. deviation x 100

mean
CV = 0.42 x 100 = 0.58 %
72.14
!Another parameter that is commonly used
to provide an indication of the relative
spread of the data around the mean is the
coefficient of variation,

! CV = [SD /χ ] ´ 100%.

! CV below than 5% is considered

acceptable
 Sources of errors
A. Systematic errors
● Produce results that consistently deviate from
expected results (true value). 
● Sources of errors due to inaccurate instruments or
measuring device. Eg: a pipette that constantly
delivers wrong volume of reagent will produce
high degree of precision but low accuracies.
● Overcome by proper calibration of instrument or
using different analytical method.
 
B. Random Errors

! Misjudging the color of the indicator near the end

point - this is probably the most common one. Not
only color change is sometimes very delicate and
slow, but different people have different sensitivity to
colors. This is not the same as being color blind,
although these things are related.
! Misreading the volume - at any moment, and due to
whatever reason.
C. Blunders error
! Eg using the wrong reagent or instrument or sloppy
technique.

! Experimental data are scattered. Fortunately,

blunders are easily identified and corrected.
Rounding off numbers

! Premature or incorrect rounding off

can produced serious error in the final
results.

! It is usually desirable to carry extra

numbers during calculations and
perform the rounding off on the final
answer
! e.g 62.722 = 62.72; 62.727 = 62.73
 Standard Curve
! Standard curves use to determine concentrations of
an unknown sample

! A curve (usually straight line) relate to the

concentration versus to the observation values

! Estimation of the unknown concentration of samples

can be obtained by extrapolation from the
standard curve:
Linear regression; y = ax + b
! Correlation coefficient is used to know how well
the data fit into a straight line. An ideal situation is
whereby the data points lies perfectly on a straight
line.

! However, this is never the case, because errors are

introduced in making standards and reading
absorbance values.
! The correlation coefficient determination is as
follow:
! r = ΣXY
√ (ΣX2) (ΣY2)

! r value should be closed to 1.000 because this

value is a perfect correlation.

! In analytical work, r should be 0.9970 or better.

 Rejecting data
! When you came across a value that does not match
the others, can you reject this value and not include
this value in your data analysis?
! Q test can be used to reject aberrant (strange/
outliers) value.
Q value = X2 – X1
W
!X2 = the questionable value
!X1= the next closest value to X1
!W = the total spread of all values
(highest value – lowest value)
Q-values for rejection of results

Number of Q of rejection
observations (90% level)
3 0.94
4 0.76
5 0.64
6 0.56
7 0.51
8 0.47
9 0.44
10 0.41
● If the Q-value same or higher than the value given
in a Q-test table for the n samples being analyzed,
then that value can be rejected at 90% confidence
level.
● If the Q-value is less than the value given in a Q-test
table for the n samples being analyzed, then that
value can be accepted at 90% confidence level.

● An example, if five measurements were carried out

and one measurement was very different from
others (e.g., 20,22,25,50,21), having a Q-value of
0.84, then that particular value can be safely
rejected (because it is higher than the value of 0.64
given in the Q-test table for five observations).