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BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

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 BIOCHEMICAL AND
ENVIRONMENTAL STUDIES ON
NEEM LEAVES AND SEED

Nadia, Y.A.Attia1; Ibrahem, [Link] El-Aleem1;

Journal Kareem, [Link].2 and Nesrein, S.Saliem1.
1
J. Biol. Chem. Department of Biochemistry Faculty of Agriculture, Benha
Environ. Sci., 2015, University, Egypt.
2
Vol. 10 (3):373- 397 Horticulture Research Institute, Agricultural Research
[Link] Center Giza, Egypt.

ABSTRACT
Neem seed (Azadirachta indica) was used to study the chemical
composition, physicochemical characteristics and fatty acids
composition of neem oil, amino acids composition of neem meal,
effect of different treatments on the removal of antinutritional factors
from neem meal, determination of neem protein subunits molecular
weight by using SDS-PAGE and digestability index of neem protein.
Neem leaves were also used to investigate the protective effect of
neem leaves (ethanol extracts) against the injury of alloxan on; blood
suger levels, liver, kidney functions and lipid profile in rats. Results
showed that oil was the major component of seed (46.4%) followed by
protein (19.44%) and carbohydrate (17.16%).
The antinutrative factors; trypsin inhibitor, total cyanogenic,
phytic acid and tannins in neem seed and meal were 5.13 and 12.36,
0.13 and 0.25, 0.35 and 0.51 and 0.17and 0.32g/100g respectively.
Neem seed oil contained high amounts of unsaturated fatty acids
(67.07%). The major unsaturated fatty acid was oleic acid (46.85%)
followed by linoleic (19.21%) and linolenic (0.90%) while, total
saturated fatty acids content was 31.70%.Neem meal protein essential
amino acids content was 36%. The optimum conditions for maximum
digestibility (88.0%) was heating after soaking in Na2SO3.
Autoclaving and roasting had great effect on protein subunits
especially those of high molecular weights .Heating neem meal after
soaking in sulfite showed the best treatment for the removal of
antinutritional factors. Gas Chromatography Mass Analysis of neem
leaves ethanol extract showed the presence of 21 components.
Administration of neem leaves ethanol extract in diabetic rats showed
374 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

significant decrement in blood glucose and significant increment in

insulin. Histology studies showed marked improvement in diabetic
groups treated with neem leaves extracts.
Key words: Azadirachta indica, Diabetic, Liver and kidney
Functions,

INTRODUCTION
The neem plant (Azadirachta indica) belongs to the family
Meliaceae. Neem is a fast growing evergreen plant found in most parts
of Africa including Kenya, Nigeria and Ghana (Schmutterer, 1988).
The tree grows in a wide range of soils and it is a sturdy tree which
can be established in poor dry soils without irrigation (Margraf,
1990). Neem kernel cake (NKC) obtained from the neem seed oil
industry is a potential alternative source of non-conventional feedstuff
(Odunsi et al., 2009). The protein content of NKC (CP: 30-40% dry
weight) is close to that of soya bean meal SBM (CP: 44%), a highly
valued protein ingredient used in poultry diet (Reddy et al., 1988).
Margraf (1990) reported that the neem plant has many uses; for
example, the bark, leaves, fruits and sap help to cure various skin
diseases, verneral diseases (e.g. syphilis) and tuberculosis. It can also
be used as antiseptics, remedy against rheumatism and antidote to
snake or scorpion bite. Neem was also used for various purposes
including pest control, manufacture of shampoos, soaps and tooth
paste (Saxene, 1984). Neem seed is a part of the neem tree wich has
high concentration of oil (35-45%). It is also can be used as an organic
bio-pesticide repellant against insects such as meal worms and aphids.
Neem oil is used for treating many skin diseases such as eczema,
psoriasis (skin disease) and skin allergies. Neem oil is considered as
one of the most important of the commercially available product of
neem for medicine and cosmetics, Orhevba et al., (2013). One of the
major problems in the utilization of neem seed as a food is the
presence of antinutritional factors such as phytic acid, tannin, HCN
and trypsin inhibitor activity (Mubarak et al., 2012). Ogbuewu et al.,
(2009) concluded that inclusion of neem leaves meal up to 15% in the
diets of rabbits resulted in significant reductions in serum cholesterol
and glucose levels. The reduction in serum cholesterol value of the
rabbit bucks fed neem leaf meal based diets is an indication that neem
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 375

leaves could reduce the deposition of cholesterol in the skin and

muscles. The reduction in serum cholesterol is a positive development
since low cholesterol meats command high market price.
Daniel et al., (2012) revealed that the [Link] leaves and bark
extracts contained saponins, steroids, tannins, phenols,
anthraquinones, lactones, flavonoids, cardiac and cyanogenic
glycosides, but showed absence of alkaloids. The [Link] seed
extract was positive for all the above mentioned components including
alkaloid. Omnia Abdel Hamid (2014) evaluated the effects of neem
leaves aqueous extract on streptozotocin (STZ) experimental diabetic
rats .Results revealed significant increase in glucose, cholesterol,
triglycerides, insulin and hydroxyl methyl glutaryl coenzyme A
reductase in diabetic rats when compared to control. The treatment
with neem extract resulted in significant decrement in all these
parameters where as serum cortisol and HDL has no significant
change in all groups.
The comparative analysis on the effects of ethanol leaves-
extracts of Vernonia amygdalina and Azadirachta indica on lipid
profile were conducted using albino rats. The serum levels (mg/dl) of
total cholesterol, LDL- cholesterol, HDL-Cholesterol and
triacylglycerol were studied spectrophtometricaly. Total cholesterol,
triacylglycerol and LDL- cholesterol levels were significantly
(P<0.05) reduced by both extracts while, HDL-cholesterol levels were
increased significantly (Offor et al., 2014).
The main goal of this investigation include the following points:
Determination of the chemical composition of neem seed,
physical and chemical properties of neem oil, amino acids
composition of neem meal, effect of different treatments on the
removal of antinutritional factors, determination of neem protein
subunits molecular weight by using SDS-PAGE and digestability
index of neem protein. Also to evaluate the biological effects of neem
leaves ethanol extract on diabetic rats.

MATERIALS AND METHODS

Plant Material
Neem leaves were obtained from Horticulture Research Institute,
Agricultural Research Center Giza, Egypt (July 2013).
376 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Proximate analyses:
Moisture, ash, crud protein, crud fiber, total lipids and its
physicochemical properties, and HCN were determined according to
the method of the association of official analytical chemists
(A.O.A.C., 2005). Carbohydrates were determined according to
Bernfeld (1955) and Miller (1959). The trypsin inhibitor activity was
measured according to the method described by Hamerstrand et al.,
(1981). Tanines were determined by the method of Mangan (1988).
Phenolic compounds was determined by the method of Gutfinger
(1981). Phytic acid was determined by the method of Mohammed
(1986). Fatty acids were determined by the method of Vogel (1975).
Amino acids were determined by the method of Moore et al., (1958).
Protein subunits molecular weights were determined by using
polyacrylamide gel electrophoresis according to Laemmli (1970).
Removal of antinutritional compounds:
1) The neem seed meal was soaked in water for 24h and heated
at 100°C for 1h according to Odunsi et al., (2009).
2) The neem seed meal was soaked in 0.1% Na2SO3 sodium meta
bisulfite for 24h and heated at 100°C for 1h according to the
method of Friedman and Gumbmann (1986).
3) The neem seed meal was soaked in 0.1% Na2CO3 sodium
carbonate for 24h and heated at 100°C for 1h according to
Abd El- Aleem and Mohamed (2005).
4) The neem seed meal was soaked in 10% NaCl sodium chlorid
for 24h and heated at 100°C for 1h according to the method
reported by Akande and Fabiyi., (2010).
5) The neem seed meal was soaked in 25% ethanol for 24h and
heated at 100°C for 1h according to the method of James et
al., (2009).
6) The neem seed meal was soaked in 75% methanol for 24h and
heated at 100°C for 1h according to James et al., (2009).
7) The meal was autoclaved at 121°C for 30 min according to
Mohamed and Girgis (2005).
8) The meal was roasted at 140°C for 30 min as reported by Abd
El-Aleem and Mohamed (2005).
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 377

Preparation of neem leaves extract:

The dried leaves were powdered mechanically and soaked in
95% ethanol in brown bottles at room temperature (25 -30 oC) in dark
place for 5 days and was shaken gently every day. The mixture was
filtrated by suction pump in bokhnnar cone throw filter paper. The
ethanolic extract was concentrated by heating under vacuum at 62o C
(Bhandar et al., 1998).
Biological evaluation of neem leaves ethanol extract:
Experimental animals
A total of 24 adult male albino rats (Wister Strain) weighted 90
to 100g were obtained from Organization of Biological Product and
Vaccines (Helwan Farm), Egypt. They were kept in wire –bottom
stainless steel cage in a room temperature maintained 25 ± 2 oc .Rats
were kept under normal healthy condition for 14 days and fed on a
basal diet. The diet contained of 65% starch, 15% casein, 10% corn
oil, 5% cellulose, 4% minerals and 1% vitamins (Philip et al .1993)
Dosage and administration of decoction: The decoction was
administered at a doses of 100 ml /100g boody weight and 200 ml
/100g boody weight neem leaves ethanol extract (Ayon Bhattachary
et al., 2014) using a Sondineedel by gastric gavage (Iddamaldeniya
et al., 2006). After that animals were divided into two main groups.
Firstmain group (6 rats): negative control (healthy control) without
any treatment.
The Second main group (18 rats): was the diabatic group. Rats
were injected with a single dose of alloxan solution 150 mg/kg body
weight (Buko, et al., 1996). After 24 hours of alloxan injection, the
presence of diabetes was confirmed (glucose blood was higher than
180 mg/dl). Rats were left for one week for stabilize diabetes, and
then rats were divided into three subgroups (6 rats per each).
First subgroup was kept as diabetic (positive control) and fed
on basil diet for eight weeks.
Second subgroup was fed a basil diet and received orally neem
ethanol extract 100ml/100g body weight/day for eight weeks.
Third subgroup was fed a basil diet and received orally neem
ethanol extract 200 ml/100g body weight/day for eight weeks.
378 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Blood samples:
At the end of experiment blood was collected in tubes from reto-
orbital vein and was centrifuged at 3000 rpm for 20 min, for serum
preparation.
Serum analysis
Serum parameter were determined by enzymatic colorimetric
methods, the glucose concentration was determined according to the
method of Trinder (1969). Serum triglycerides and total cholesterol
were determined according to the method of Fossati and Precipe,
(1982) and Finely (1978). High density lipoprotein HDL- cholesterol
and low density lipoprotein (LDL- cholesterol) were determined
according to the method of Lopes-Virella et al., (1977). Alanine
transaminase (ALT) and aspartate transaminase (AST) were
determined according to Reitman and Frankel (1957). Alkaline
phosphastase (ALP) in serum was determined according to the method
Young et al., (1972). Albumin was determined in serum according to
the method described by Doumas et al. (1971). Serum total proteins
was determined using Biuret method performed by Doumas (1975).
Uric acid, serum urea and Creatinine were determined according to
Haisman and Muller (1977), Tabacco et al., (1979) and Henery et
al., (1974).
Tissue samples:
At the end of the experiment rats were sacrificed and the post
mortem findings were recorded. Tissues specimens from pancreas,
liver and kidney organs were obtained for weighting, fixed in 10%
formalin saline for histopathological examination. Routine
histopathological procedures were carried out according Schermer
(1967)
The histopathological examination:
According to Drury and Wallington (1986), these histopathological
was determined at Faculty of Vet. Med. Benha University.
Statistical analysis:
Statistical analysis was carried out using ANOVA with one
factor under significance level of 0.05 for the whole results using
SPSS var.19 and data were treated as complete randomization design
according to Steel et al., (1997). Multiple comparisons were carried
out applying LSD and Takey.
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 379

RESULTS AND DISCUSSION

Chemical composition:
Data concerning neem seed and meal chemical composition are
shown in Table (1)
Table (1): Chemical analysis of neem seed and meal calaulatedon
dry weight basis.

Results show that oil was the major component of seed (46.4%)
followed by protein (19.44%) and carbohydrate (17.16%). The
antinutrative factors; trypsin inhibitor, total cyanogenic, phytic acid
and tannins in neem seed and meal were 5.13 and 12.36; 0.13 and
0.25; 0.35 and0.51; 0.17and 0.32 respectively. The obtained results
are in agreement with those reported by Yashim et al., (2008) and
Harry-Asobala et al., (2014).
Fatty acids composition of neem seed oil:
Gas liquid chromatography analysis of neem oil Table, (2)
showed that the oil contained high amounts of unsaturated fatty acid
(67.07%). The major unsaturated fatty acids was oleic acid (46.85 %)
followed by linoleic (19.21%) and linolenic (0.90 %) while, total
saturated fatty acids content was 31.70% (palmitic acid 15.57% and
stearic acids 14.50 %).The obtained results are in agreement with
those reported by Sergio et al., (2007) and Djenontin et al., (2012).
380 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Table (2): Physiochemical characteristics and fatty acid analysis

of neem seed oil

Amino acids composition of neem meal:

Data concerning amino acids composition of neem meal are
reported in Table (3)
Data indicate that total essential amino acids ratio was 36.10 %
while, total non- essential amino acids ratio was 59.58 %.Glutamic
and arginine were the most abundant amino acids followed by aspartic
acids. Sulfur amino acids cystine and methionine were the most
limiting amino acids. Cystine and methionine were presented in
minute quantities (2.17 – 1.33) g/100g protein. These results are in
agreement with the data reported by Bodwell and Hopkins (1985)
and James et al., (2004). They found that essential amino acids of
neem protein ranged from 35 to 45% of their total amino acids. Amino
acid scored pattern proposed by FAO and WHO (1973) qualified an
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 381

ideal protein as one in which 36% of the total residues are essential
amino acids.
Table (3) Amino acids composition of neem seed meal (g\100g
protein)

Effect of pH on neem meal protein isolation:

Results indicate that the maximum neem protein extraction
(94.35%.) was achieved at pH 12 , and in the acidic pH range the
percentage of the extracted protein was very lower than alkaline and
reached its lowest amount at pH 4.0 (isoelectric point). These results
could be explained on the basis of the exhibited role of ionogenic
groups in protein molecules in lyophilic colloidal systems of protein
solutions. These proteins might be positively or negatively charged
depending on the hydrogen ion concentration of the medium. The
amount of NaOH bounds by the protein molecules depends on the
equilibrium of hydrogen, Wu and Sexson (1979).
Effect of different treatments on neem meal protein
382 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Table (4): Effect of different treatments on the removal of

antinutritional factors from neem meal

a) Effect of different treatments on neem meal protein

digestibility and the removal of antinutritional factors
Data in Table (4) show effect of different treatments on neem
meal protein digestibility. Results indicate that digestibility of the
native meal was 72.35% then improved by increasing heat
temperature. The optimum conditions for maximum digestibility
(88.0%) was heating after soaking in Na2SO3. Heating neem meal
after soaking in H2O, Na2SO3 and Na2CO3 was more effective in the
destruction of phytic acid comparing with autoclaving and roasting.
The cyanogenic glycosides was totally destroyed after all different
treatments and the obtained results are in agreement with those
reported by Bahlol and Abd El-Aleem (2004). Also, results in Table
(4) show that treatment of neem meal protein with methanol solution
gave the highest removal of tannins (89.8 %) and the obtained results
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 383

in agreement with that reported by Nadia Attia et al., (2004). Results

concerning the effect of different heating treatments on inactivation of
trypsin inhibitor of neem meal indicated that soaking with Na2SO3 and
heating for 1h inactivated trypsin inhibitor to its lowest amount
(11.98%). Inactivation of trypsin inhibitor is difficult to achieve by
heat alone and soaking in sulfite showed high effect in facilitating
inactivation of trypsin inhibitor in neem meal .Therefor, it could be
concluded that heating neem meal after soaking in sulfite is the best
[Link] obtained results are in agreement with those reported by
Friedman and Gumbmann (1986).
b) Effect of different treatments on neem meal protein subunits by
using SDS- PAGE
Polyacrylamide Gel Electrophoresis in the presence of detergent
Sodium Dodecyl Sulphate (SDS-PAGE) was used for determining the
subunit molecular weights (M.W.) of protein extracted by alkaline
solutions from neem meal after different treatments. Results showed
that neem meal protein (without heating) dissociated into 7 subunits
with molecular weights (MW) ranging from 94 to 19 KD. Different
tretments showed that heating resulted slight reduction in some high
molecular weights but autoclaving and roasting had great effect on
protein subunits especially those of high molecular weights which
disappeared and might be dissociated to low molecular weights
subunits. These finding are in agreement with those reported by Nadia
Attia et al., (2004).
Gas Chromatography - Mass analysis of neem leaves ethanol
extract
Results revealed the presence of 21components. The identified
components were 6- Propyl-2- thiouracil (1.98 % ), 2- Deoxy-D-
galactose (1.91%), Imidazole, 2-amino-5-[(2-carboxy)vinyl]- (1.39 %)
, 2-Dodecenoic acid (2.67 %) , 2-Hexadecanol (2.35 %) , 2-Amino-
4,6-dihydroxypyrimidine (8.52%) , Hexanoic acid, 3-hydroxy-, ethyl
ester (1.65 %) , Indole-4-carboxaldehyde (43.91 %) , Benzoic acid
(1.99 %), Megastigma-3,7(E),9-triene (5.7 %) , Dl –Metanephrine
(2.51 %) , Pseudo solasodine diacetate (0.66 %) , 4′-Hydroxy-3′-
methyl acetophenone( 3.6 %), Carvacrol (4.03 %) , Gedunin (0.45%)
, 2,4-Quinolinediol (1.4 %) , solasodine –diacetate (1.86 %) ,
Solasodine (2.34 %) , Butylate dhydroxy toluene (3.51 %) , Guanine
(1.09 %), Glycitein (1.32 %). Results of the identified components are
384 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

in agreement with those reported by Akapuka et al., (2013), and

Jessinata et al., (2014).
Biological effects:
Effect of neem leaves ethanol extracts on blood serum glucose,
insulin, and lipid profil:
Data in Table (5) show that in diabetic group there was
significant increment in glucose level and significant decrement in
insulin level comparing with control group (negative). Administration
with neem leaves ethanol extract at 200ml/100g boody weight caused
significant increment in insulin and significant decrement in glucose
level .Data in Table (5) indicate that in the diabetic group there was
significant increment in total lipid, tri-glycerides, total cholesterol and
LDL, and significant decrement in HDL comparing with control group
(negative). Administration with neem leaves ethanol extract at 100 ml
and 200ml/100g boody weight caused significant decrement in total
lipid, tri-glycerides, total cholesterol and LDL, and significant
increment in HDL comparing with diabetic [Link] obtained results
of lipid profil are in agreement with those reported by Offor et al.,
(2014), but Omina Abd el Hamid (2014) reported no change in HDL.

Table (5) The effect of neem leaves ethanol extract on blood serum
glucose, insulin, and lipid profil.
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 385

Liver functions
Data in Table (6) show the effect of neem leaves ethanol
extracts at different doses on liver functions. There was significant
increment in total protein, albumin globulin and liver enzymes (ALT,
AST, ALP) in diabetic group comparing with control group.
Administration of neem leaves ethanol extract at 100ml /100g boody
weight showed significant decrement in total protein, albumin,
globulin and liver enzymes ALT, and AST. Neem leaves ethanol
extract at 200ml /100g boody weight caused significant decrement in
all parameter comparing with diabetic group. These results are in
agreement with those reported by EmanTaha et al., (2010).
Table (6): Effect of neem leaves ethanol extract at different doses
on liver functions

kidney functions:
Data reported in Table (7) show the effect of neem leaves
ethanol extract at different doses on kidney functions. Results
indicated that the diabetic group showed significant increment in uric
acid, creatinine and urea comparing to normal group (control).
Administration of neem leaves ethanol extract at 100 and 200 ml
/100g boody weight showed significant decrement in all parameter
386 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

comparing with the diabetic group .The obtained results are in

agreement with those reported by Shravan Kumar et al, (2011).
Table (7) Effect ofneem leaves ethanol extract at different dose on
kidney functions

Histopathological studies
Pancreas of normal rats showed no histoligical changes Fig (1 A,
B). Diabetic rats, Fig (2c, d) show necerosis of islets of Langerhans
and apoptosis of acinar epithelium. Treatment of diabetic rats with
neem leaves extracts (100 and 200 ml) in Fig (3, E, F) show that islet
is well – vascularized, with viable. Liver in diabetic group showed
severe congestion in the central vein, sinusoids and portal blood
vessels Fig (4,b) comparing with control group Fig (4,a) while kidney
in this group showed severe congestion and dilatation of the renal
blood vessels, intertubular blood capillaries and glomerular tube Fig
(5 ,a,b,c) comparing with control group Fig (5,d). Liver and kidney in
neem treated group showed nearly normal histological structure
without any degenerative changes in the hepatic or vascular changes.
Fig (6-9).
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 387

Fig(1) . Pancreas of normal rats (A&B) control. A, islet appears

normal; H&E, x40. B, fibrosis and necrosis are absent;
trichromex 20.

Fig (2) (C&D) Diabetic group. C, islet is diminuted with paucity of

cells and necrosis (arrow); (Inset) exocrine pancreas of
diabetic rats. Darkly-pigmented bodies are seen in many
acinar cells. Eosinophilia of exocrine pancreas has
increased; H&E, x60. D, there is paucity of islet cells and
presence of fibrous spots at islet periphery (arrow)
trichrome, x60.
388 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Fig (3) (E&F) Neem-treated diabetic group. E, islet is well-

vascularised, with viable cells; H&E, x60. F, viable cells are
observable in the central part of the islet; necrosis and
fibrosis (arrows) are confined to the periphery; trichrome,
x60.

Fig (4) a. control group liver of rat fed on basal diet showing
normal histologic structure of hepatic parenchyma. H&E
stain x 200) .[Link] in diabetic group, show sever
congestion in the central vein M&F×100.
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 389

Fig (5) kidney in diabetic group, A, showing severe congestion and

dilatation in the renal blood vessels, intertubular blood capillaries
H&E × 300. B showing severe degenerative changes in the
epithelial cell lining H&E × 300.C showing leukocytes ibiltravion
replacing the renal tubuls H&E ×100. (d): Kidney of rat fed on
basal diet showing normal histological structure of renal tubules
(T) and glomeruli (G). H & E stain x 400.

Fig (6) liver in Diabetic and neem leave extract (100 ml) group,
showed mild vaculation in the hepocytes H&E × 300
390 BIOCHEMICAL AND ENVIRONMENTAL STUDIES ON NEEM LEAVES AND SEED

Fig (7) kidney in Diabetic and neem leave extract (100 ml) group
showed mild vacuolation at epithelial cell lining at the renal
tubules H&E × 600

Fig (8) liver in Diabetic and neem leave extract (200 ml) group A showed
nearly normal histological structure without any degenerative
H&E × 300. B mononuclear lelluer infiltration H&E × 200.

Fig (9) kidney in Diabetic and neem leave extract (200 ml ) group A
showed mild degeneration changes and mild degeneration at
the epithelial cell living at the renal tubuls , B mild congestion
peoislmrvrheoneye H&E × 200.
 J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397 391

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‫دراﺳﺎت آﻴﻤﻴﺎﺋﻴﺔ وﺑﻴﺌﻴﺔ ﻋﻠﻰ أوراق وﺑﺬور اﻟﻨﻴﻢ‬

‫‪،2‬‬
‫ﻥﺎدیﺔ یﺤﻴﻲ أﺣﻤﺪ ﻋﻄﻴﺔ ‪ ، 1‬إﺑﺮاهﻴﻢ ﻡﺤﻤﺪ ﻋﺒﺪ اﻟﻌﻠﻴﻢ ‪ ، 1‬آﺮیﻢ ﻡﺼﻄﻔﻰ اﻟﻄﻮﺑﺠﻲ‬
‫‪1‬‬
‫ﻥﺴﺮیﻦ ﺳﻌﺪ ﺳﻼﻡﺔ‬
‫‪ 1‬ﻗﺴﻢ اﻟﻜﻴﻤﻴﺎء اﻟﺤﻴﻮﻳﺔ – آﻠﻴﺔ اﻟﺰراﻋﺔ – ﺟﺎﻡﻌﺔ ﺑﻨﻬﺎ‪-‬ﻡﺼﺮ‪ 2 ،‬ﻡﻌﻬﺪ ﺑﺤﻮث اﻟﺒﺴﺎﺗﻴﻦ –‬
‫ﻡﺮآﺰ اﻟﺒﺤﻮث اﻟﺰراﻋﻴﺔ – اﻟﺠﻴﺰة‬

‫ﺗﻌﺘﺒﺮ ﺷﺠﺮﻩ اﻟﻨﻴﻢ اﺡﺪى اﻻﺷﺠﺎر اﻟﺘﻰ ﺧﻠﻘﻬﺎ اﷲ ﻟﻠﻔﺎﺋﺪﻩ ﻓﻬﻰ ﺗﺸﻔﻰ اﻟﻜﺜﻴﺮ ﻡﻦ اﻻﻡﺮاض‬
‫اﻟﻤﻌﺮوﻓﻪ وﺗﻨﻤﻮ ﻓﻰ ﺑﻠﺪان ﺷﺮق اﺱﻴﺎ وﻓﻰ اﻟﺒﻠﺪان اﻻﺱﺘﻮاﺋﻴﻪ واﻟﻤﻤﻄﺮﻩ ﺡﻴﺚ ﻳﺴﺘﺨﺪم ﺟﻤﻴﻊ‬
‫اﺟﺰاﺋﻬﺎ ﻓﻰ ذﻟﻚ ‪.‬‬
‫وﺗﻬﺪف هﺬﻩ اﻟﺪراﺱﻪ اﻟﻰ ﻡﻌﺮﻓﺔ ﺗﺎﺙﻴﺮ اﻟﻤﺴﺘﺨﻠﺺ اﻟﻜﺤﻮﻟﻰ ﻋﻠﻰ ﻓﺌﺮان اﻟﺘﺠﺎرب اﻟﻤﻌﺎﻡﻠﻪ‬
‫ﺑـ ‪alloxan‬‬
‫وﻓﻴﻤﺎ یﻠﻰ اهﻢ اﻟﻨﺘﺎﺋﺞ اﻟﻤﺘﺤﺼﻞ ﻋﻠﻴﻬﺎ ‪:‬‬
‫اﻟﺘﺮآﻴﺐ اﻟﻜﻴﻤﺎوى ﻟﻠﺒﺬور‪:‬‬
‫‪ -‬اوﺿﺢ اﻟﺘﺮآﻴﺐ اﻟﻜﻴﻤﺎوى ﻟﻠﺒﺬور اﻥﻬﺎ ﺗﺤﺘﻮى ﻋﻠﻰ ‪ % 15‬رﻃﻮﺑﻪ و ‪ % 16,6‬اﻟﻴﺎف و‪2‬‬
‫‪ %‬رﻡﺎد و ‪ %19,44‬ﺑﺮوﺗﻴﻦ و‪ % 46,4‬دهﻮن آﻠﻴﻪ و ‪ 17,16‬آﺮﺑﻮهﻴﺪرات ‪.‬‬
‫‪ -‬ووﺟﺪ اﻳﻀﺎ ان اﻟﺒﺬور ﺗﺤﺘﻮى ﻋﻠﻰ ﺗﺮآﻴﺰ ﻋﺎل ﻡﻦ ﻋﻨﺎﺹﺮ ‪ ، Ca ، K‬ﺙﻢ ‪Mg , Na‬‬
‫اﻟﺘﺮآﻴﺐ اﻟﻜﻴﻤﺎوى ﻟﻜﺴﺐ اﻟﻨﻴﻢ‪:‬‬
‫‪ -‬اوﺿﺢ اﻟﺘﺮآﻴﺐ اﻟﻜﻴﻤﺎوى ﻟﻠﻜﺴﺐ اﻥﻪ ﻳﺤﺘﻮى ﻋﻠﻰ ‪ % 12‬رﻃﻮﺑﻪ و ‪ % 12‬اﻟﻴﺎف و‪4,30‬‬
‫‪ %‬رﻡﺎد و ‪ %36,5‬ﺑﺮوﺗﻴﻦ و‪ %6,3‬دهﻮن آﻠﻴﻪ و ‪40,9‬آﺮﺑﻮهﻴﺪرات‪.‬‬
‫‪ -‬ووﺟﺪ اﻳﻀﺎ اﻥﻪ ﻳﺤﺘﻮى ﻋﻠﻰ ﺗﺮآﻴﺰ ﻋﺎل ﻡﻦ ﻋﻨﺎﺹﺮ ‪Mg ، Ca ، K‬‬
‫اﻟﺘﺎﺙﻴﺮ اﻟﺒﻴﻮﻟﻮﺟﻰ ‪:‬‬
 ‫‪J. Biol. Chem. Environ. Sci., 2015, 10 (3), 373-397‬‬ ‫‪397‬‬

‫اﺟﺮﻳﺖ اﻟﺘﺠﺮﺑﻪ ﻋﻠﻰ ‪ 24‬ﻓﺄرا ﻡﻦ ذآﻮر ﻓﺌﺮان اﻟﺘﺠﺎرب اﻟﺒﻴﻀﺎء )اﻻﻟﺒﻴﻨﻮ ( ﺡﻴﺚ ﺗﻢ‬
‫ﺗﻐﺬﻳﻪ اﻟﻔﺌﺮان ﻋﻠﻰ ﻋﻠﻴﻘﻪ ﻋﺎدﻳﻪ ﻟﻤﺪﻩ اﺱﺒﻮﻋﻴﻦ ﻗﺒﻞ ﺑﺪاﻳﻪ اﻟﺘﺠﺮﺑﻪ ﺙﻢ ﻗﺴﻤﺖ اﻟﻰ ارﺑﻊ‬
‫ﻡﺠﻤﻮﻋﺎت ﺡﻴﺚ ﺗﺤﺘﻮى آﻞ ﻡﺠﻤﻮﻋﻪ ﻋﻠﻰ ﺱﺘﻪ ﻓﺌﺮان ‪.‬‬
‫اﻟﻤﺠﻤﻮﻋﻪ اﻻوﻟﻰ ‪ :‬ﺗﻢ ﺗﻐﺬﻳﺘﻬﺎ ﻋﻠﻰ اﻟﻌﻠﻴﻘﻪ اﻻﺱﺎﺱﻴﻪ ) اﻟﻤﺠﻤﻮﻋﻪ اﻟﻀﺎﺑﻄﻪ اﻟﺴﺎﻟﺒﻪ (‪.‬‬
‫اﻟﻤﺠﻤﻮﻋﻪ اﻟﺘﺎﻥﻴﻪ ‪ :‬ﺗﻢ ﺡﻘﻨﻬﺎ ﺑﺎﻻﻟﻮآﺴﺎن ﻓﻘﻂ )اﻟﻤﺠﻤﻮﻋﻪ اﻟﻀﺎﺑﻄﻪ اﻟﻤﻮﺟﺒﻪ (‪.‬‬
‫اﻟﻤﺠﻤﻮﻋﻪ اﻟﺜﺎﻟﺜﻪ ‪ :‬ﺗﻢ اﻋﻄﺎءهﺎ ‪ 100‬ﻡﻠﻠﻴﺠﻢ ‪ /‬آﺠﻢ ﻡﺴﺘﺨﻠﺺ آﺤﻮﻟﻰ ﻻوراق اﻟﻨﻴﻢ ‪+‬‬
‫اﻻﻟﻮآﺴﺎن ‪.‬‬
‫اﻟﻤﺠﻤﻮﻋﻪ اﻟﺮاﺑﻌﻪ ‪ :‬ﺗﻢ اﻋﻄﺎءهﺎ ‪ 200‬ﻡﻠﻠﻴﺠﻢ ‪ /‬آﺠﻢ ﻡﺴﺘﺨﻠﺺ آﺤﻮﻟﻰ ﻻوراق اﻟﻨﻴﻢ ‪+‬‬
‫اﻻﻟﻮآﺴﺎن ‪.‬‬
‫اﻟﺪراﺱﺎت اﻟﻜﻴﻤﻴﺎﺋﻴﻪ اﻟﺤﻴﻮﻳﻪ‬
‫‪ .1‬اﻟﺘﺎﺙﻴﺮ ﻋﻠﻰ ﻡﺴﺘﻮى ﺱﻜﺮ اﻟﺪم ‪:‬‬
‫ﺡﺪوث ﻥﻘﺺ ﻡﻌﻨﻮى ﻓﻰ ﻥﺴﺒﻪ اﻟﺴﻜﺮ ﻓﻰ ﺱﻴﺮم اﻟﺪم ﻓﻰ اﻟﻤﺠﻤﻮﻋﻪ اﻟﺘﻰ ﺗﻌﺎﻥﻰ ﻡﻦ‬
‫ارﺗﻔﺎع اﻟﺴﻜﺮ ﻥﺘﻴﺠﻪ اﻟﻤﻌﺎﻡﻠﻪ ﺑﺘﺮآﻴﺰ ‪ 200‬ﻡﻠﻠﻴﺠﻢ ﻡﻦ اﻟﻤﺴﺘﺨﻠﺺ اﻟﻜﺤﻮﻟﻰ ﻟﻼوراق ‪.‬‬
‫‪ .2‬ﻡﺪﻟﻮﻻت اﻟﺪهﻮن ‪:‬‬
‫اﻟﻤﻌﺎﻡﻠﻪ ﺑﻜﻼ اﻟﺘﺮآﻴﺰﻳﻦ ادت اﻟﻰ ﻥﻘﺺ ﻡﻌﻨﻮى ﻓﻰ اﻟﺠﻠﺴﻴﺮﻳﺪات اﻟﺜﻼﺙﻴﻪ واﻟﻜﻮﻟﻴﺴﺘﺮول‬
‫اﻟﻜﻠﻰ واﻟﻠﻴﺒﻮﺑﺮوﺗﻴﻨﺎت اﻟﻤﻨﺨﻔﻀﻪ ﻓﻰ اﻟﻜﺜﺎﻓﻪ ﺑﻴﻨﻤﺎ ﺡﺪوث زﻳﺎدﻩ ﻓﻰ اﻟﻠﻴﺒﻮﺑﺮوﺗﻴﻨﺎت اﻟﻤﺮﺗﻔﻌﻪ‬
‫ﻓﻰ اﻟﻜﺜﺎﻓﻪ‪.‬‬
‫‪ .3‬وﻇﺎﺋﻒ اﻟﻜﺒﺪ ‪:‬‬
‫ﻡﻌﺎﻡﻠﻪ اﻟﻤﺠﻤﻮﻋﺎت اﻟﺘﻰ ﺗﻌﺎﻥﻰ ﻡﻦ ارﺗﻔﺎع ﻥﺴﺒﻪ اﻟﺴﻜﺮ ﺑﻤﺴﺘﺨﻠﺺ اوراق اﻟﻨﻴﻢ ادى اﻟﻰ‬
‫ﺡﺪوث ﻥﻘﺺ ﻓﻰ اﻥﺰﻳﻤﻰ اﺱﺒﺎرﺗﻴﺖ ﺗﺮاﻥﺲ اﻡﻴﻨﻴﺰ و اﻻﻻﻥﻴﻦ ﺗﺮاﻥﺲ اﻡﻴﻨﻴﺰ وآﺬﻟﻚ اﻟﻔﻮﺱﻔﺎﺗﻴﺰ‬
‫اﻟﻘﺎﻋﺪى وذﻟﻚ ﻋﻨﺪ اﻟﻤﻘﺎرﻥﻪ ﺑﺎﻟﻤﺠﻤﻮﻋﻪ اﻟﻜﻨﺘﺮول اﻟﻤﺼﺎﺑﻪ ﺑﺎرﺗﻔﺎع ﻥﺴﺒﻪ اﻟﺴﻜﺮ ‪.‬‬
‫‪ .4‬وﻇﺎﺋﻒ اﻟﻜﻠﻰ ‪:‬‬
‫اﻟﻤﻌﺎﻡﻠﻪ ﺑﻤﺴﺘﺨﻠﺺ اوراق اﻟﻨﻴﻢ ادى اﻟﻰ ﺡﺪوث ﻥﻘﺺ ﻡﻌﻨﻮى ﻓﻰ اﻟﻴﻮرﻳﺎ وﺡﻤﺾ‬
‫اﻟﻴﻮرﻳﻚ واﻟﻜﺮﻳﺎﺗﻴﻨﻴﻦ وذﻟﻚ ﻋﻨﺪ اﻟﻤﻘﺎرﻥﻪ ﺑﺎﻟﻤﺠﻤﻮﻋﻪ اﻟﻜﻨﺘﺮول اﻟﻤﺼﺎﺑﻪ ﺑﺎرﺗﻔﺎع ﻥﺴﺒﻪ اﻟﺴﻜﺮ ‪.‬‬
‫‪ -5‬اﻟﻔﺤﺺ اﻟﻬﺴﺘﻮﻟﻮﺟﻰ‬
‫اﻇﻬﺮت ﻥﺘﺎﺋﺞ اﻟﻔﺤﺺ اﻟﻬﺴﺘﻮﻟﻮﺟﻰ ان ﻡﺴﺘﺨﻠﺺ اوراق اﻟﻨﻴﻢ ادت اﻟﻰ ﺗﻌﺰﻳﺰ ﺧﻼﻳﺎ‬
‫اﻟﺒﻨﻜﺮﻳﺎس ﻓﻰ اﻟﻔﺌﺮان اﻟﻤﺼﺎﺑﻪ ﺑﻤﺮض اﻟﺴﻜﺮ وﺗﺤﺴﻴﻦ ﻓﻰ اﺑﻨﻴﻪ اﻟﺒﻨﻜﺮﻳﺎس وﻇﻬﺮ اﻟﻜﺒﺪ‬
‫واﻟﻜﻠﻰ ﻓﻰ وﺿﻌﻬﺎ اﻟﻄﺒﻴﻌﻰ وﺧﺎﺹﻪ ﻓﻰ ﺗﺮآﻴﺰ ‪ 200‬ﻡﻠﻠﻴﺠﻢ ‪.‬‬

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