EJSS - Volume 64 - Issue 2 - Pages 631-642
EJSS - Volume 64 - Issue 2 - Pages 631-642
EJSS - Volume 64 - Issue 2 - Pages 631-642
Emam Nofal 1, Fardous A. Menesy1, Samia Z. Elbably2 Manal E. Abd-El Rahman 2, Hassan
El-Ramady 3 and József Prokisch 4
(1)
Horticulture Department, Faculty of Agriculture, Kafrelsheikh University, Kafr El-Sheikh, Egypt
(2)
Ornamental Department, Horticulture Research Station, Sakha, Kafr El-Sheikh, Egypt
(3)
Soil and Water Department, Faculty of Agriculture, Kafrelsheikh University, Kafr El-Sheikh, Egypt
(4)
Institute of Animal Science, Biotechnology and Nature Conservation, Faculty of Agricultural and Food
Sciences and Environmental Management, University of Debrecen, 138 Böszörményi Street, 4032 Debrecen,
Hungary
S OIL salinity is considered a serious and limiting factor reducing the crop productivity in particular
under arid, and semi-arid conditions. Pot experiments were conducted at the Ornamental
Department of the Horticulture Research Station in Sakha, Kafr El-Sheikh Governorate, Egypt during
the season of 2021 and 2022. This study aimed to evaluate the effect of foliar application with nano-
NPK and nano-sulfur using different doses on the growth and chemical compounds of Khaya
senegalensis L. under salinity stress. The studied treatments were mineral NPK fertilizer at 5 g L-1 (as
a control), applied nano-NPK at 1, 2 and 3 ml L-1, besides three combinations of nano NPK (2 ml L-1)
and applied nano-sulfur at 1, 2 and 3 ml L-1, separately. The obtained results showed that the
treatment of nano-NPK 2 ml L-1 + nano-sulfur 2 and/or 3 ml L-1 significantly increased plant height
(cm), stem diameter (cm), fresh and dry weights of vegetative parts per seedling (g), membrane
stability index, as well as root length (cm), number of roots and root fresh and dry weights per plant.
The superiority of the nano-NPK at 2 ml L-1 + nano-sulfur at 2 ml L-1 treatment comparing with the
control in both seasons. Most treatments recorded significantly higher chlorophyll a, b, carotenoids,
catalase and peroxidase activity in the leaves than control. A superiority was found after applying
nano-NPK 2 ml L-1 + nano-sulfur 2 ml L-1 in both seasons for nitrogen, phosphorus and potassium
content in the leaves, which was significantly increased over the control.
Table 2. Physical and chemical analysis of the used soil the 1st and 2nd two Experimental season 2021 and 2022.
Soil Physical properties Soil chemical properties
Available nutrients
Season Coarse Fine Texture EC**,
Silt Clay pH* -1 OM, % (ppm)
sand sand class dS m
N P K
Frist season 2.53 22.05 37.79 37.63 Clay loam 8.12 4.56 1.02 37.06 7.03 212.4
Second season 2.71 21.59 35.28 40.42 Clayey 8.03 4.66 1.15 42.16 8.34 209.8
In both season pH* was determined in soil suspension (1: 5) and Soil Electrical Conductivity (EC) ** was
determined in saturated soil paste extract.
Fig. 1. An overview on the main treatments and measurements during the study.
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Egypt. J. Soil Sci. 64, No. 2 (2024)
634 EMAM NOFAL, et al.,
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Total carotenoids (mg/gm fw)= (1000A470 - 1.82
Chl. a – 85.02Chl. b)/198
Where, A = Absorbance at respective wave length,
Chl. a= chlorophyll a and Chl. b= chlorophyll-b.
2.6 Membrane stability index
Membrane stability index (MSI) or electrolyte
leakage (EL) was determined according to Sairam
et al. (1997). Leaf disks (200 mg) were taken in
two sets of test tubes containing 10 ml of distilled
water. One set was kept at 40ºC in a water bath for
30 min and electrical conductivity (C1) was
measured. The second set was incubated at 100ºC
for 15 min and electrical conductivity (C2) was
measured. MSI was calculated according to the
following formula:
MSI (%) = (1- C1/C2) ×100
Fig. 2. Transmission Electron Microscope 2.7 Leaf chemical composition
(TEM) imaging of nano-sulfur. The content of nitrogen (N), phosphorus (P) and
potassium (K) in the dried leaves were achieved
2.3 Vegetative growth parameters using 0.2 gm were digested according to
At the end of experimental seasons on November Peterburgski, (1968) as digested solution was used
10th to 15th 2021 and 2022 respectively, the to determine N, P and K by Kjeldahl device,
following data were recorded including plant height spectrophotometer (Jenway 6405) and flame
(cm), stem diameter (cm), fresh and dry weights of photometer (Jenway PFP7, Staffordshire, UK) of
stem per plant (g), as well as root length (cm), the three elements, respectively (Mertens, 2005).
number of roots and root fresh and dry weights (g)
per plant. 2.8 Determining of enzyme activities
The crude enzymatic extracts for determining the
2.4 Estimation of chlorophyll content activities of catalase and peroxidase were obtained
Chlorophyll content was measured using 20 ml of by soaking 0.3 g of leaves in liquid nitrogen and
acetone 80% + 1 g leaf tissue and 0.5 g of (MgCO3) then adding 2 mL of homogenized medium;
powder was added and further grinded gently. The followed by centrifugation at 12,000×g for 15 min
mixture was then incubated at 4ºC for 3 hrs. The at 4 °C. The homogenization media were 0.1 M
mixture was centrifuged at 2500 rpm for 5 min and potassium phosphate buffer, pH 6.8, 0.1 Mm
the supernatant was transferred to a 100 ml EDTA, 1 mM phenyl methyl sulfonyl fluoride
volumetric flask and the volume was made up to (PMSF), and 1 % poly vinyl pyrrolidone (PVPP)
100 ml with the addition of acetone 80% and the (w/v) (Peixoto et al. 1999) for the enzymes CAT,
solution was used for estimation chlorophyll a and POX 50 mM potassium phosphate buffer, pH 7.0, 1
b (Chl. a and Chl. b) were calculated using the mM ascorbate, and 1 mM EDTA (Nakano and
following equation according to Rajalakshmi and Asada, 1981).
Banu (2014): The catalase (EC 1.11.1.6) activity was determined
Chlorophyll a (mg/gm fw)= [12.7(A663) – 2.69 by the addition of 0.1 mL of enzymatic extract to
(A645)] V/1000W 2.9 mL of reaction solution that was composed of
Chlorophyll b (mg/gm fw)= [22.9(A645) – 4.68 50 mM potassium phosphate buffer at pH 7.0 and
(A663)] V/1000W 12.5 mM H2O2 (Havir and McHale 1987). The
Where A = Absorbance of specific wavelength, V = decreasing absorbance during the first minute of
Final volume of chlorophyll extract in 80% acetone reaction was measured at 240 nm at 25 °C. The
and W = Fresh weight of tissue extract. enzymatic activity was calculated using a molar
extinction coefficient of 36 mol L-1 cm-1 and
2.5 Estimation of total carotenoids expressed by μmol of H2O2 min-1 mg-1 fw
The content of carotenoids was estimated according (Anderson et al., 1995). Catalase and peroxidase
to Sumanta et al. (2014) using the same chlorophyll were determined by spectrophotometer (Jenway
extract as measured at 470 nm in spectrophotometer 6405).
(Jenway 6405, the UK) to estimate the total content The POX activity (EC 1.11.1.7) was assessed
of carotenoids (including both xanthophylls + through the production rate of purpurogallin at 420
carotene), as follows: nm according to the proposed method of Nakano
and Asada (1981) with a molar extinction
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coefficient of 2.47 mmol-1 L cm-1. The enzymatic (S-NPs) at 2 ml L-1 gave the tallest plants (69.36
activity was expressed in μmol purpurogallin min- cm) compared to the control followed by the
1 -1
g fw (Chance and Maehley 1955). increase in seedling treated with nano-NPK at 2 ml
L-1 plus nano-sulfur at 3 ml-1 then nano-NPK at 2
2.9 Statistical analyses
Data were statically analyzed by analysis of ml L-1 plus nano-sulfur at 1 ml L-1 while the control
variance (ANOVA) using Costat (Version 6.303, treatment gave the lowest value (63.88 cm).
Co Hort, USA, 1998–2004)] program for data set of Nano-NPK fertilizer under different doses
the two independent experimental and combined significantly increased in the length as compared to
analysis was carried out after checking the control. Data listed in Table (3) showed that
homogeneity of the variance Bartlett's test. spraying plants with nano-NPK at different doses or
Duncan`s multiple range test (Duncan, 1955) was nano-NPK at 2 ml L-1 plus nano-sulfur at 1, 2, 3 ml
used in order to compare the mean at the P ≤ 0.05% L-1 precipitate the mineral fertilizer (control). Nano-
probability level according to Gomez and Gomez
NPK plus nano-sulfur at 2 ml L-1 gave the highest
(1984). Resulted were presented as average mean
value of stem diameter (13.0 cm) followed in terms
of the two independent seasons ±SE.
of the increase treating with NPK-NPs at 2 ml L-1
3. Results plus nano-sulfur at 2 ml L-1 which gave average
3.1 Vegetative characteristics stem diameter compared to the control treatment
In general, all studied values were recorded that which resulted the lowest stem diameter per plant
highest values of vegetative parameters comparing (8.98 cm). Data resulted in Table (3) showed
with the control with superiority to nano-form than significantly superiority in fresh and dry weight of
mineral form. Data of Khaya growth parameters plants treated with NPK-NPs and nano-NPK plus
included, plant height stem diameter, fresh and dry sulfur at different doses compared to the control
weight, beside the leaves number, are listed in (mineral fertilizer). Adding sulfur-NPs at different
Table (3). The treated plants with N, P and K nano- doses along with the nano-NPK (2 ml L-1) caused
fertilizer at different levels or plus nano-sulfur at all increase in fresh and dry weights of plant with
levels were significantly superior in all traits increasing the applied dose of S-NPs.
compared to control (mineral fertilizer) treatment.
Nano-NPK (NPK-NPs) at 2 ml L-1 plus nano-sulfur
Table 3. Effect of nano NPK and nano sulfur foliar application on stem growth of Khaya senegalensis L
plants (average of two seasons, 2021 and 2022).
Plant height Stem diameter Fresh weight of Dry weight of
Treatments
(cm) (cm) stem (g) stem (g)
(T1) Control (mineral NPK 5 g L-1) 63.88d 8.98g 43.85e 11.05g
(T2) NPK-NPs 1 ml L-1 64.15d 9.83f 46.20d 13.74f
(T3) NPK-NPs 2 ml L-1 64.81cd 11.19d 51.00c 15.78d
(T4) NPK-NPs 3 ml L-1 64.57cd 10.44e 46.59d 14.10e
(T5) NPK-NPs 2 ml L-1 + S-NPs 1 ml L-1 65.47c 11.58c 55.76b 16.82c
(T6) NPK-NPs 2 ml L-1+ S-NPs 2 ml L-1 69.36a 13.00a 65.20a 20.23a
(T7) NPK-NPs 2 ml L-1+ S-NPs 3 ml L-1 66.83b 12.26b 64.17a 19.36b
F-test ** ** ** **
Mean followed by the same letter in the same column do not differ significantly by Duncan's multiple range test at 5% level.
3.4 Membrane stability index (MSI) ml L-1 plus nano-sulfur at dose of 2 ml L-1 gave the
Data presented Figure (3D) showed that there were highest value of MSI (60.9 %) compared the
non-significant differences between all treatment control treating with mineral fertilizer, which
and control in membrane stability index (MSI) in recorded the lowest value (58.9%).
leaves but the treating seedlings with nano-NPK 2
Fig 3. Effect of nano-NPK and nano-sulfur foliar application on photosynthetic pigments and Membrane
stability index (MSI) of Khaya senegalensis L. as (A) chlorophyll a (mg g-1 fw), (B) chlorophyll b
(mg g-1 fw), (C), carotenoids (mg g-1 fw) and (D) membrane stability index (%). Data are mean
value ± SE. Bars at the same letter are not significant at P≤0.05 level. (T1) Control (mineral NPK 5
g L-1), (T2) NPK-NPs 1 ml L-1, (T3) NPK-NPs 2 ml L-1, (T4) NPK-NPs 3 ml L-1, (T5) NPK-NPs 2 ml
L-1 + S-NPs 1 ml L-1, (T6) NPK-NPs 2 ml L-1 + S-NPs 2 ml L-1, (T7) NPK-NPs 2 ml L-1 + S-NPs 3 ml
L-1 (average of two seasons, 2021 and 2022).
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3.5 Leaf chemical composition
An increase was observed in all treatments after NPK along with nano-sulfur at 2 ml L-1 followed by
applying different doses of both NPK-NPs and S- treated plants with nano-NPK at 2 ml L-1 plus nano-
NPs regarding the content of N, P, and K in the sulfur using 2 or 1 ml L-1 whereas, the control
seedling leaves (Figure 4). The highest value of N, treatment gave the lowest values (2.04, 0.48, and
P, and K content (2.62, 0.56 and 1.8 %) in plant 1.22 %).
leaves achieved after foliar application of nano-
Fig. 4. Effect of nano-NPK and nano-sulfur foliar application on chemical composition nitrogen,
phosphorus and potassium of Khaya senegalensis L. in leaves (%) for N, P, K (A), (B) and (C),
respectively. Data are mean value ± SE. Bars at the same letter are not significant at P≤0.05 level.
(T1) Control (mineral NPK 5 g L-1), (T2) NPK-NPs 1 ml L-1, (T3) NPK-NPs 2 ml L-1, (T4) NPK-
NPs 3 ml L-1, (T5) NPK-NPs 2 ml L-1 + S-NPs 1 ml L-1, (T6) NPK-NPs 2 ml L-1+ S-NPs 2 ml L-1,
(T7) NPK-NPs 2 ml L-1+ S-NPs 3 ml L-1 (average of two seasons, 2021 and 2022).
3.6 Antioxidant activity
Data illustrated in Figure (5A) revealed that the 30.02) which gave the equivalent values in
treated plants with NPK-NP fertilizer at different significant, followed by nano-NPK at 2 ml L-1 plus
doses or plus nano-sulfur at all levels were nano-sulfur at 2 ml L-1, whereas the lowest value
significantly superior in the catalase and peroxidase was recorded from the control (24.54). For
activities. Concerning catalase activity (μmol of peroxidase activity (μmol purpurogallin min-1 g-1
H2O2 min-1 mg-1 fw) data showed that the treating fw), the same previous trend that recorded by
plants with nano-NPK at 2 ml L-1 along with nano- catalase (Figure 5B).
sulfur at either 2 or 3 ml L-1 resulted the highest
significant values of catalase activity (30.52, and
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638 EMAM NOFAL, et al.,
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Catalase activity
A Peroxidase activity AB
40 (μmol of H2O2 min-1 mg-1 fw) 1.5 (μmol purpurogallin min-1 g-1 fw)
b a a
30 e de c cd a b
1 c
f d e
20 f
10 0.5
0
T1 T2 T3 T4 T5 T6 T7 0
T1 T2 T3 T4 T5 T6 T7
Fig. 5. Effect of nano-NPK and nano-sulfur foliar application on antioxidant activity of Khaya senegalensis
L. as (A) catalase activity (μmol of H2O2 min-1 mg-1 fw) and (B) peroxidase activity (μmol
purpurogallin min-1 g-1 fw). Data are mean value ± SE. Bars at the same letter are not significant
at P≤0.05 level. (T1) Control (mineral NPK 5 g L-1), (T2) NPK-NPs 1 ml L-1, (T3) NPK-NPs 2 ml
L-1, (T4) NPK-NPs 3 ml L-1, (T5) NPK-NPs 2 ml L-1 + S-NPs 1 ml L-1, (T6) NPK-NPs 2 ml L-1+
S-NPs 2 ml L-1, (T7) NPK-NPs 2 ml L-1+ S-NPs 3 ml L-1 (average of two seasons, 2021 and 2022).
4. Discussion
This section will answer the following main combined foliar spraying of NPK-NPs fertilizer
questions in the present study: and/or with nano-sulfur at different applied rates had
1- What are the NPK-NP and S-NP-fertilizers? significantly positive effects on the morphological
2- What is the suggested mechanism of such nano- parameters including seedling height, stem diameter,
fertilizers? fresh and dry weights and leaf number comparing
3- What is the expected combined application of the with the control. This effect significantly increased
studied nanofertilizers? with increasing the added dose of nano-NPK and/or
4- To what extent can consider the studied candidates nano-S up to 2 ml L-1 comparing with the control.
are promising nanofertilizers under salinity This may be attributed to the role of nano-fertilizers
stress? in ameliorating salinity stress by modulating the
5- Are Khaya seedlings tolerant plants to salinity? oxidative and salt stress, as well as promoting the
antioxidative role under such stress (Singh et al.
The reason beyond the selection of this tree for the 2023). The increase in leaves number almost resulted
current study may back to its importance in many from the physiological role of studied nutrients (N, P,
therapeutic activities in both people and animals by K, and S), which are essential and responsible for
treat a variety of diseases including dermatoses, improving the shoot growth and the accumulation of
malaria, jaundice, leprosy, syphilis, and mental the carbohydrate substance and stimulate the
illness (Amang et al. 2023). The growing of such tree vegetative growth as well (Bang et al. 2020). It could
depends on the growing media and environmental notice that applied NPK-NPs depends on applying
stresses like soil salinity under arid and semi-arid soluble forms of NPK mineral fertilizers in a
zones (Table 1). The mitigation of soil salinity was polymer, which let these NPK nutrients release
investigated by applying many materials and/or slowly to plants as -slow-release fertilizer. The
certain framing practices including nanomaterials applied nano-fertilizers are a source of needed
(NMs). Under forestry cultivation, the potential nutrients (N, P, K, and S), which are essential for all
application of nanotechnology or nanofertilizers still morphological (all vegetative parameters),
require research and evidence particularly in physiological (photosynthetic pigments), and
producing good quality transplants under nursery biochemical attributes (enzymatic antioxidants) of
growing conditions (Singh et al. 2021). studied seedlings.
The influence of nano-fertilizers through foliar The role of nano-NPK and or nano-S for increasing
spraying has been suggested for amplifying plant total fresh and dry weight and elevated percent of
growth of different horticultural species under chlorophyll a, b and carotenoids content in this study
environmental stress particularly seedling stage such may be due to the beneficial effect of nano fertilizers
as banana (Shalaby et al. 2022b), and strawberry (El- in increasing the bioavailability of such necessary
Bialy et al. 2023). In the current investigation, the nutrients to the growing plants leading to increase
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chlorophyll forming, dry matter production and Nanoparticles of sulfur hold promise for enhancing
improved overall growth of the plant under such plant nutrition when incorporated into saline/alkaline
studied stress (Saad-Allah and Ragab 2020; EL- soil due to their significantly increased surface area,
Madah et al. 2024) facilitating the transformation into soluble sulfur
compounds. These compounds are readily absorbed
How can foliar nanofertilizers attribute to the
by plant roots, thereby augmenting the plant's sulfur
previous impact on mahogany seedlings? Foliar
uptake. However, when applied via foliar spraying,
sprayed nanoparticles to plant may enter stomata by
sulfur nanoparticles may encounter limitations,
increasing the osmatic pressure of the stomata cell,
which needs more studies for more details. The
the frequently opening the stomatal cells and enabled
insoluble nature of sulfur granules can hinder their
the plant to receive more nutrients loaded on the
incorporation into plant metabolites, potentially
surface of NPs as indicated by Qureshi et al. (2018).
resulting in reduced efficacy. Moreover, foliar
These results an agreement with those obtained by
application may lead to sulfur deposition on leaf
Haggag (2018), (Shalaby et al. 2022b), and (El-Bialy
surfaces, limiting stomatal exchange and impeding
et al. 2023), who observed that nanofertilizers
nutrient absorption. Therefore, while nano-sulfur
significantly enhanced the uptake of nitrogen,
shows potential for soil application to improve plant
phosphorus and potassium in olive, banana and
nutrition, its effectiveness via foliar application
strawberry seedlings, respectively.
warrants further investigation to optimize its delivery
Data of the study showed that adding nano-sulfur at and utilization in agricultural practices.
different levels can play an important role in the
Based on the increasing possibility of global soil
growth of Khaya senegalensis seedlings under
salinization due to climate change, more studies on
salinity stress compared to the control as these
soil salinity and its mitigation are needed in
treatments augmented the morphological traits
particular under nano-farming system and modern
including plant height, stem diameter, number of
agriculture. Agrican mahogany proved from the
leaves fresh and dry weight as in Table (3) also
current study its promising candidate to be
increased root parameters as root length, number of
successfully cultivated under saline conditions with
roots, root fresh and dry weight as recorded in Table
more opening questions can be answered in the
(4). These results are confirmed with those obtained
upcoming studies.
by Thirunavukkarasu et al. (2023). Chlorophyll a, b
and carotenoids content as well as catalase and Conclusions
peroxidase activities were recorded the highest African mahogany is important tree that has many
values in the seedling leaves after applying nano- benefits for the human and animal as well. It can be
NPK plus nano-sulfur using 2 ml L-1 from both concluded that the combined application of both
nanofertilizers as compared to untreated plants. nano-NPK fertilizer and nano-sulfur at 2 ml L-1
every 15 days interval through the growth season
Therefore, application of such nanofertilizers can
positively gained the best morphological,
play a promising role in increasing studied seedlings
biochemical, and physiological parameters. These
to be tolerant against soil salinity. The role of S-NPs
parameters included mainly the content of nutrients
as a nanofertilizer is distinguished for promoting the
in leave, the content of chlorophyll a and b,
plant chlorophyll content due to sulfur is a part of
carotenoids, catalase, peroxidase activity. The
carbon compounds found in two amino acids and is a
suggested mechanism of cultivated seedlings to be
precursor for many coenzymes and vitamins essential
tolerant to soil salinity may back to promoting the
for metabolism (Taiz and Zeiger 2002). Nano-S can
defense system. This system was supported by
also improve plant growth by increasing the
enzymatic antioxidants, and bioavailability of
efficiency of chemical energy production in the
nutrients leading to be tolerant the oxidative and salt
photosynthesis system and also improving plant
stress. More studies are needed to answer more open
growth and biomass production and many studies
questions regarding this important ornamental crop.
have showed that the application of nanofertilizer led
to an increase in the content of photosynthetic Conflicts of interest: There are no conflicts to
pigments when using appropriate dose of sulfur-NPs declare.
which plays antioxidant role or modulate the Funding: No external fund for this work.
antioxidant defense system (Najafi et al. 2020). Acknowledgments: The authors would like to thank
their institutions for providing the needed support.
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640 EMAM NOFAL, et al.,
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