← Back
Reduced amino acid Schiff base containing ruthenium(III) complexes: Synthesis, characterization, DNA interaction, and in vitro cytotoxicity.
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
Iraqi Journal of Veterinary Sciences
www.vetmedmosul.com
Effect of sodium benzoate on some biochemical, physiological and
histopathological aspects in adult male rats
S.A. Al-Ameen1 , E.H. Jarjees2
and F.Kh. Tawfeeq3
1
Department of Chemistry, Science College, University of Mosul, 2Department of Dental Industry Techniques, Al-Noor
University College, 3Department of Biochemistry, Physiology and Pharmacology, College of Veterinary Medicine, University
of Mosul, Mosul, Iraq
Article information
Article history:
Received April 14, 2021
Accepted July 11, 2021
Available online March 3, 2022
Keywords:
Sodium benzoate
Growth hormone
Nitric oxide
Correspondence:
E.H. Jarjees
eman.hazim@alnoor.edu.iq
Abstract
Sodium benzoate (SB) is a white powder, used as preservative and food additive.
Biochemical, physiological and histopathological effects of SB been tested in adult male
rats. Twenty-four adult albino male rats aged100 day and weighted 250-350 g were used.
Animals were divided into four groups. The first group considered as control, which
received normal saline orally, other groups treated with SB by 300, 400 and 500 mg/kg of
body weight respectively for 30 days. At the end of experiment, blood samples were
collected from retro orbital sinus. Heart, liver, spleen, kidneys and brain were obtained for
weight recording. The results indicated a significant decrease of super oxide dismutase
SOD activity and a significant increase of nitric oxide NO level of treated group 500
mg/kg of body weight. Moreover, findings revealed that there are no significant changes
in growth hormone GH activity and body weight. A significant reduction of heart weight
of treated group 500 mg/kg of body weight were observed. The histopathological changes
ranged from mild to severe in the brain cortex, as focal gliosis, satellitosis, mild
vacuolation and vasogenic edema in treated groups with SB by different doses. Also, some
changes were observed in liver represented by congestion of portal vein, mild hydropic
degeneration of hepatocytes, stenosis of sinusoids, steatosis and necrosis of hepatocytes in
treated groups with SB compared to control group. It concluded that short-term exposure
to high doses of SB may be considered an oxidant substance that caused oxidative stress.
Furthermore, SB can harm various organs in the body.
DOI: 10.33899/ijvs.2021.129935.1705, ©Authors, 2022, College of Veterinary Medicine, University of Mosul.
This is an open access article under the CC BY 4.0 license (http://creativecommons.org/licenses/by/4.0/).
important one is sodium benzoate (3). Sodium benzoate SB,
is white in color, odorless crystallized and found as a grain
or powder easily crystalline in appearance and found as
powder or grain, water-soluble and in ethanol, it is slightly
dissolved. (4). It can be used in a number of foods,
including fruit juices, fruit-based fillings, pickles, salad
dressings, jams, and carbonated beverages, as well as
cosmetics (5). SB has been used as a therapeutic agent in
medical applications. (6). Sodium benzoate has the E
number E211 and is used as a preservative and food
additive. Under acidic conditions, it is bacteriostatic and
fungi static. When this compound is consumed
inadvertently by the human as food additives, it has harmful
Introduction
Many people, during the huge advances which have
happened in technology and daily lifestyle as they are in
age of speed, turn to use fast and prepackaged foods which
is sold in markets rather than preparing them at homes with
no consideration to what they contain of added materials,
which are changing or damaging their qualities, also they
generally cause health problems (1). On the other hand,
these materials play an important role in keeping these
types of foods for long time without being damaged (2).
These materials include antioxidants, food coloring agents,
anti-infectious agents, flavoring agents and the most
267
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
effects on the body (7). Benzoic acid and sodium benzoate
are on the Food and Drug Administration's FDA list of
generally accepted as safe drugs in the United States
GRAS. With a current maximum level of 0.1 percent in
food, both can be used as antimicrobial agents, flavoring
agents, and adjuvants (7). The FDA hasn't decided whether
or not substantially different terms of use are GRAS (3).
While sodium benzoate is considered a healthy
preservative, excessive ingestion of these preservatives
could be harmful to consumers. Excessive use of SB has
been linked to changes in serum clinical parameters,
according to many studies, research has declared that when
SB combines with vitamin C in soft drinks to create
benzene, sodium benzoate has been linked to cancer. Also,
the study observed that giving this material to the pregnant
female rats by mouth led to change in the hypothalamus of
the new generation and this case causes change in secretion
of growth hormone, thus inverse some effects on the
animal’s weights (8). Moreover, SB can cause
neurotoxicity, nephrotoxicity, and teratogenicity in zebra
fish larvae during early embryogenesis (9). The aim of this
research was to see how long-term SB administration
affects health on some biochemical, physiological and
histopathological parameters in adult male albino rats.
hormone (GH) level was measured using COBAS e 411
analyzer- Roche diagnostics.
Materials and Methods
Results
Experimental animals
In this study, 24 male albino rats have been used at age
of 100 days and their weights 250 to 350 g. They have been
obtained from animal house in the college of Veterinary
Medicine/University of Mosul. Feeding add libtum.
Biochemical test
The results indicated a significant reduction in super
oxide dismutase activity in treated groups with increasing
sodium benzoate dose compared to the control group.
Moreover, the study revealed that there were no significant
changes in the growth hormone level of all groups (Table
1).
Physiological and histopathological examination
After blood collection, internal organs weight liver,
heart, kidneys, spleen and brain were recorded. Then the
Liver and Brain was placed in solution of 10% formalin for
a period of 24 hours for fixation, after that the tissues were
dehydrated using a gradual concatenations of alcohol
solution 50% - 100% for 5 minute each. Then the tissue
samples were cleared in 2 separated xylene changes prior to
placing them in paraffin wax for final sectioning. Later the
samples were sectioned at 5 μm thickness, then it stained by
hematoxylin and eosin stain to study the histological
changes compared to the control group using light
microscope (11).
Statistical analysis
The results were expressed as Mean±SEM standard
error of the mean. Data were analyzed by one-way analysis
of variance ANOVA and were performed using the minitab
18 program, The Tukey’s test was used to compare
significance between groups. The significance level was
accepted at P<0.05 (12).
Experimental protocol
Twenty-four adult male rats were divided into four
groups. Each group included 6 rats. All treatments were
given orally daily by gavage needle for 30 days as T1
control group given normal saline, T2, T3 and T4 treatment
with sodium benzoate prepared by dissolving it in normal
saline at concentration 300, 400 and 500 mg/kg
respectively.
Table 1: Effect of sodium benzoate on growth hormone
(GH) level, SOD activity and NO level
Mean ± SE (n=6) for 30 days
GH pg/ml
SOD µmol/L
NO µmol/L
T1
32.51±2.03A
183.38±5.53A 107.09±0.520B
T2
51.01±8.16A
149.27±4.02B
112.17±1.42B
T3
43.60±5.06A
125.15±2.00C 113.14±0.623B
T4
41.01±5.86A
109.92±1.34D
141.44±5.80A
Different letters vertically mean difference between groups
at probability level P<0.05.
Blood samples collection
After 30 days of treatment, Blood samples were
obtained from retro orbital sinus by glass capillaries and
collected in plain tubes and allow the sample for
coagulation then centrifuged at 3000 rpm for 15 min.
Serum samples were immediately stored at -20ºc in
eppendorf tubes till used for analysis of biochemical
parameters (10).
The present study revealed a non-significant changes in
body weight of treated group with sodium benzoate 300
mg/kg bw in comparison with control and other treated
groups, and a significant decrease in heart weight of treated
group with sodium benzoate 500 mg/kg bw in comparison
with the control and other treated groups. While no
significant changes can be observed in the liver, spleen,
kidneys and brain weights (Table 2).
Biochemical analyses
Blood Serum nitric oxide NO and super oxide dismutase
SOD activity, markers of oxidative stress were estimated
using kits from Al-shkairate establishment for medical
supply. Sweilleh Amman 11910 Jordon. serum growth
268
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
Table 2: Sodium benzoate's effect on body, liver, heart, kidney, spleen and brain weights in adult male rats
Weight mean(mg/100 g b.w.) ± SE (n=6) for 30 days
Body weight
Liver
Heart
Kidney
Spleen
T1
334±14.55A
10.71±0.70A
1.13±0.07A
1.13±0.08A
1.15±0.11A
T2
376.5±8.88A
12.30±0.84A
1.09±0.03A
1.24±0.06A
1.11±0.13A
T3
342.6±7.61A
12.33±0.90A
1.07±0.04A
1.24±0.06A
1.09±0.09A
T4
328.8±16.61A
10.57±0.59A
0.89±0.04B
1.21±0.09A
0.96±0.12A
Different letters vertically mean difference between groups at probability level P<0.05.
Brain
1.99±0.13A
2.11±0.12A
1.91±0.11A
1.82±0.08A
The histopathological results
The study revealed in comparison with control group
(Figures 1 and 2) to normal architecture of the cortex of
brain with mild vasogenic edema (Figure 3) and shows
focal gliosis and satellitosis in the cortex of brain (Figure 4)
in group treated with 300 mg/kg.bw of sodium benzoate. In
addition, observed mild vacuolation, gliosis and satellitosis
in the cortex of brain (Figure 5), and mild vacuolation,
gliosis and vasogenic edema in the cortex of brain (Figure
6) in group treated with 400 mg/kg.bw of sodium benzoate.
Figure 3: photomicrograph of rat’s brain of group T2 shows
normal architecture of the cortex with mild vasogenic
edema (A).
A
C
Figure 1: photomicrograph of rat’s brain of control group
shows normal architecture of the cortex of brain.
B
Figure 4: photomicrograph of rat’s brain of group T2 shows
focal gliosis (A) and satellitosis (B) in the cortex.
A
Moreover, showed diffuse gliosis, vacuolation,
satellitosis and vasogenic edema in the cortex of brain
(Figure 7) in group treated with 500 mg/kg bw of sodium
benzoate. On the other hand noticed some changes in the
liver tissue that was investigating in congestion of portal
vein, mild hydropic degeneration of hepatocytes and
stenosis of sinusoids (Figure 8) in group treated with
300mg/kg.bw of sodium benzoate, and shows congestion of
portal vein, hydropic degeneration of hepatocytes, necrosis
of others and stenosis of sinusoids (Figure 9) in group
treated with 400 mg/kg.bw of sodium benzoate, in addition
shows severe hydropic degeneration, fatty change steatosis,
A
Figure 2: photomicrograph of rat’s brain of control group
shows normal architecture of the thalamus region of brain.
269
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
necrosis of hepatocytes and congestion of portal vein
(Figure 10) in group treated with 500 mg/kg bw of sodium
benzoate, in comparison of control group (Figure 11).
B
A
Figure 8: photomicrograph of rat’s liver of group T2 shows
congestion of portal vein (A), mild hydropic degeneration
of hepatocytes (B) and stenosis of sinusoids (C).
Figure 5: photomicrograph of rat’s brain of group T3 shows
mild vacuolation (A), gliosis (B) and satellitosis (C) in the
cortex.
D
A
B
A
C
C
B
Figure 9: photomicrograph of rats liver of group T3 shows
congestion of portal vein (A), hydropic degeneration of
hepatocytes (B), necrosis of others (C) and stenosis of
sinusoids (D).
Figure 6: photomicrograph of rat’s brain of group T3 shows
mild vacuolation (A), gliosis (B) and vasogenic edema (C)
in the cortex.
B
A
C
B
D
C
D
A
Figure 10: photomicrograph of rat’s liver of group T4
shows severe hydropic degeneration (A), fatty change
(steatosis) (B), necrosis of hepatocytes (C) and congestion
of portal vein (D).
Figure 7: photomicrograph of rat’s brain of group T4 shows
diffuse gliosis (A), vacuolation (B), satellitosis (C) and
vasogenic edema (D) in the cortex.
270
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
the changes ranging from mild to severe. Also observed
some changes in liver represented by portal vein
congestion, mild hydropic degeneration of hepatocytes,
stenosis of sinusoids, fatty change steatosis and hepatocytes
necrosis with sodium benzoate treatment groups in
compared to control group the moderate to severe
vacculation of seminiferous tubule of testes in treated
groups with sodium benzoate (19).
Salah et al. (20) that showed vacuolation of hepatocytes,
congestion of the central vein. Moreover, apoptosis, and
necrosis of some hepatocytes in pregnant mice injected (IP)
with a Platinum drug at the dose of 3 mg/kg of bw.
Conclusion
Figure 11: photomicrograph of rat’s liver of control group
T1 shows normal architecture of liver tissue representing by
central veins (A), hepatocytes (B) and sinusoids (C). H&E
stain, 100X.
Short term adult male rat’s exposure to high doses of
sodium benzoate might act as oxidant material, inducing
oxidative stress and cell damage. The preservative material
can cause defects in different organs of the body.
Discussion
Acknowledgments
While sodium benzoate is considered a safe chemical,
short-term exposure can cause health problems as defect in
different organs of the body. Moreover, sodium considers
as a basic component in all cell physiology. The dis
advantages and side effects of sodium benzoate on human
health, including cell damage, have been approved (13),
and this research was designed to observe these effects.
The present study revealed no significant changes in
body, liver, spleen, kidneys and brain weights, with a
significant reduction in heart weight in treated group with
500 mg/kg sodium benzoate. This result agrees with (8,14),
benzoate treatment did not alter body weight when
compared to control. While the current result didn’t agree
with (15,16), who found a significant decrease in rat’s body
weight treated with sodium benzoate. The obtained results
may be attributed to a non-significant alteration in growth
hormone levels that determined in this research.
This result was not consistent with Hela et al. (16), who
showed the administration of food additives mixture to rats
increased serum thyroid hormones T3 and T4 levels, may
be due to alteration in thyroid hormones might result in
pituitary-thyroid axis.
On the other hand, the present study refers to a
significant decrease in the superoxide dismutase activity in
sodium benzoate treated groups and this reduction reverses
in a manner proportional to the increasing of sodium
benzoate doses.
On the other hands, nitric oxide levels were increased in
group treated with 500 mg/kg.bw of sodium benzoate,
suggested that sodium benzoate acts as oxidant material and
induced oxidative stress (8,17,18).
The histological effects, the outcomes refer to changes
occurred in the brain cortex such as focal gliosis,
satellitosis, mild vacculation and vasogenic edema in
treated groups with sodium benzoate in different doses and
The authors are very grateful to the University of
Mosul, Veterinary College, animal house staff for their
provided facilities which helped us to completion this work.
Conflicts of interest
No conflicts
References
1.
2.
3.
4.
5.
6.
7.
8.
271
Tawfeeq FK, Jirjees IH. Effect on monosodium glutamate on pregnant
female rats and embryo. Al- Anbar J Vet Sci. 2013;6(2):100-107.
[available at]
Makwana S, Choudhary R, Dogra N, Kohli P, Haddock J. Nano
encapsulation and immobilization of cinnamaldehyde for developing
antimicrobial food packaging material. Food Sci Technol.
2014;57(2):470-6. DOI: 10.1016/j.lwt.2014. 01.043
Piper JD, Piper PW. Benzoate and sorbate salts: A Systematic review
of the potential hazards of these invaluable preservatives and the
expanding spectrum of clinical uses for sodium benzoate. Comp Rev
Food Sci Foof Safty. 2017;16:868-880. DOI: 10.1111/15414337.12284
Hadi MM, Mahdi WTh. The effect of sodium benzoate as a
preservative on the reproductive system of male rats. Int J Res Sci.
2019;10(1):98-105. DOI: 10.26452/ijrps.v10i1.1785
Noorafshan A, Erfanizadeh M, Karbalay SD. Sodium benzoate, a food
preservative, induces anxiety and motor impairment in rats. Neuro Sci.
2014;19 (1):24-28. [available at]
Inetianbor JE, Yakubu JM, Ezeonu SC. Effects of food additives and
preservatives on man- a review. Asia J Sci Technol. 2015;6(2):11181135. [available at]
Shahid M, Alwan NA, Al-Masoudi EA. A study of toxic effect of
sodium benzoate, vitamin C alone and their combination on
reproductive functions of adult male rabbits. Bas J Vet Res.
2018;17(3):533-543. [available at]
Khoshnoud MJ, Siavashpour A, Bakhshizadeh M, Rashedinia M.
Effects of sodium benzoate, a commonly used food preservative on
learning memory and oxidative stress in brain of mice. J Biochem Mol
Toxicol. 2017;94(01):1-7. DOI: 10.1002/jbt.22022
Iraqi Journal of Veterinary Sciences, Vol. 36, No. 2, 2022 (267-272)
تأثير بنزوات الصوديوم على بعض الجوانب
الكيموحيوية والفسلجية والنسيجية في ذكور الجرذان
البالغة
9.
Tsay HJ, Wang YH, Chen WL, Huang MY, Chen YH. Treatment with
sodium benzoate leads to malformation of zebra fish larvae.
Neurotoxicol
Teratol.
2007;29(5):562-9.
DOI:
10.1016/j.ntt.2007.05.001
10. Ahmed MA. Protective effect of aqueous extract of Alhagi maurorum
in spermatogenesis and antioxidant status of adult rats exposed to
carbon tetrachloride. Iraqi J Vet Sci. 2019;33(1):1-7. DOI:
10.33899/ijvs.2019.125509.1031
11. Al-Jammas S, Al-Saraj A. The histological changes induced by
cytarabine on rabbits livers (with and without vitamin E
administration). Iraqi J Vet Sci. 2020;34(1):9-13. DOI:
10.33899/ijvs.2020.163564
12. Hamzah FZ, Al-Sharafi NM, Kasim SF. Effect of aqueous rosemary
extract on some sexual hormones in male rats with high thyroxine
level.
Iraqi
J
Vet
Sci.
2021;35(2):369-373.
DOI:
10.33899/ijvs.2020.126872.1404
13. Hartwig A. Benzoic acid and alkali benzoates. Occupat Hlth Safety.
2018;3(4):1758-1828. DOI: 10.1002/3527600418.mb6585e6318
14. Kehinde OS, Christianah OI, Oyetunji OA. Ascorbic acid and sodium
benzoate synergistically aggravates testicular dysfunction in adult
Wistar rats. Int J Physiol Pathophysiol Pharmacol. 2018;10(1):39-46.
[available at]
15. Helal EE, El-Sayed RA, Hedeab GM, El-Gamal MS. Effects of some
food additives on some biochemical parameters in young male albino
rats and the ameliorative role of royal jelly. Egypt J Hospit Med.
2017;67(2):605- 613. DOI: 10. 12816/0037812
16. Helal EG, Barayan AW, Abdelaziz MA, EL-Shenawe NS. Adverse
effects of mono sodium glutamate, sodium benzoate and
chlorophyllins on some physiological parameters in male albino rats.
Egypt
J
Hospit
Med.
2019;74(8):18571864.
DOI:
10.21608/ejhm.2019.28865
17. Ansari FA, Ali SN, Arif H, Khan AA, Mahmood R. Acute oral dose
of sodium nitrite induces redox imbalance, DNA damage, metabolic
and histological changes in rat intestine. PLoS ONE. 2017;12(4):175196. DOI: 10.1371/journal. pone.0175196
18. Moeen SS, Elhalwagy MA, Ayaz NO. Alterations in oxidative stress
and antioxidantin albino rats treated with individual and combined
various food additives. Int J Adv Res Biol Sci. 2018;5(6):118-123.
DOI: 10.22192/ijarbs.2018.05.06.011
19. Oladele JO, Oladele OT, Ademiluyi AO, Oyeleke OM, Awosanya
OO, Oyewole OI. Chaya (Jatropha tanjorensis) leafs protect against
sodium benzoate mediated renal dysfunction and hepatic damage in
rats. Clin Phytosci. 2020;6:13. DOI: 10.1186/s40816-020-00160-5
20. Salah BA, Sadoon HS. Histopathological and some biochemical
effects of platinum drug on the liver and kidney of pregnant mice Mus
musculus and their embryos. Iraqi J Vet Sci. 2021;35(2):291-300.
DOI: 10.33899/ijvs.2020.126706.1364
2 إيمان حازم جرجيس،1صفاء عبد العزيز االمين
3و فدوى خالد توفيق
قسم تقنيات صناعة2 ، جامعة الموصل، كلية العلوم،قسم الكيمياء1
كلية،فرع الفسلجة والكيمياء الحياتية3 ، كلية النور الجامعة،أسنان
العراق، الموصل، جامعة الموصل،الطب البيطري
الخالصة
تستخدم بنزوات الصوديوم بصورة واسعة كمادة حافظة ومضاف
غذائي للكثير من األطعمة ويعتقد أن التعرض لفترة طويلة لهذه المادة
وزيادة تراكيزها في الدم يؤدي إلى تأثيرات جانبية على صحة اإلنسان
تضمن البحث إمكانية اختبار التأثيرات الكيموحيوية.وأعضاءه الحيوية
،والفسلجية والنسيجية لبنزوات الصوديوم في ذكور الجرذان البالغة
تم، غم350-250 منها تتراوح أوزانها بين24 حيث تم استخدام
اعتبرت المجموعة األولى مجموعة سيطرة. مجاميع4 تقسيمها إلى
تلقت المحلول الملحي عن طريق الفم في حين أخضعت المجاميع الثالثة
500 و400 ،300 األخرى للمعاملة ببنزوات الصوديوم بالجرع
ثم جمعت عينات الدم من محجر، يوما30 كغم على التوالي لمدة/ملغم
العين لتقدير أوكسيد النتريك وفعالية إنزيم فوق أوكسيد الدسميوتيز
أظهرت النتائج وجود زيادة.ونشاط هرمون النمو في مصل الدم
معنوية في تركيز أوكسيد النتريك وانخفاض معنوي في فعالية إنزيم
فوق أوكسيد الدسميوتيز في مصل الدم للمجموعة المعاملة بالجرعة
بينما لم تظهر النتائج وجود،كغم مقارنة بمجموعة السيطرة/ ملغم500
كذلك أشارت النتائج.فرق معنوي في فعالية هرمون النمو ووزن الجسم
إلى وجود انخفاض معنوي مقارنة بمجموعة السيطرة في وزن القلب
عالوة.كغم/ ملغم500 للمجموعة المعاملة ببنزوات الصوديوم بالجرعة
اظهر الفحص النسجي تغيرات تتراوح بين خفيفة إلى شديدة،على ذلك
في قشرة الدماغ مثل الدباق البؤري والتساتل والوذمة الوعائية في
500 و400 ،300 المجموعات المعاملة ببنزوات الصوديوم بالجرع
أخيرا البد من اإلشارة إلى وجود بعض التغيرات في الكبد.كغم/ملغم
متمثلة باحتقان الوريد البابي والتنكس المائي وتضيق الجيوب والتنكس
الدهني والنخر في خالياه للمجاميع الثالثة المعاملة ببنزوات الصوديوم
.
مقارنة بمجموعة السيطرة
272