INFLUENCE and histological analysis were carried out after

 INFLUENCE OF ELAESIS GUINEENSIS LEAF IN DIET ON PETROLEUM-MEDIATED KIDNEY DAMAGE
IN RAT

Achuba ,F. I .
Department of Biochemistry, Delta State University, PMB 1, Abraka Nigeria. [email protected]

 

Abstract

The toxicity of petroleum hydrocarbon across the
living systems is now a common knowledge among the scientific community. What
is lacking is a mini-scale antidote that can be adopted by the inhabitants of
crude oil producing areas of the world. This was the reason for this study. The
study is comprised forty eight female rats divided into six groups of eight
rats each. Rats in control group were fed with diet without any treatment while
rats in groups 2 and 3 were fed with diets treated with known amount of Elaesis guineensis leaf respectively.
Rats in group 4 were fed with crude oil contaminated diet. Rats in groups 5 and
6 were fed with contaminated diet mixed with known amount of ground Elaesis guineensis leaf. Biochemical and
histological analysis were carried out after three and six months respectively.
The results show that pretreatment of crude oil contaminated diet with Elaesis guineensis leaf tend to restore values of lipid peroxidation, xanthine oxidase
activity, superoxide dismutase activity and catalase activity close to control
values. Histological examination indicates
protective effect of Elaesis guineensis leaf against deleterious effect of
crude oil on the kidney.   Thus, it is pertinent to state that
there exist potentials in the use Elaesis
guineensis leaf in the treatment of crude oil toxicity. And indeed setting
a fresh agenda for further serious scientific investigations

 

 

 

Keywords: Catalase, Crude oil, Kidney, Lipid
peroxidation, Elaesis guineensis,
Superoxide dismutase .Xanthine oxidase,

 

 

 

 1.0 Introduction

 Humans and animals get exposed to crude oil or
its byproducts when these chemicals are released into the surroundings during
oil exploration activities, equipment failures, corrosion, illegal bunkering,
usage, oil theft and illicit refining 1-3. Crude oil stimulates oxidative
stress in animals 4, 5. Lipid peroxidation, xanthine oxidase superoxide
dismutase (SOD) and catalase activities are part of oxidative stress indices
6. Lipid peroxidation elicits oxidative damage in plants and animals and its
value in conjunction with alterations in the level of antioxidants represent a
measure of oxidative stress. Similarly, the activity of xanthine oxidase is a
defense mechanism as well as measure of oxidative stress in animals 6. Report
has it that the deleterious action of crude oil on the kidney is based on
oxidative stress 7.

 Byproducts of the Elaesis guineensis tree are
important medicinally. This is because the leaf juice have wound healing
property while the sap is used as laxative 8.This is due to   compounds rich in medicinal and antioxidant
properties inherent in Elaesis guineensis
leaf 9, 10.  The antioxidant action is
attributed to the presence of phytochemicals (flavonoid, tannin and phenols) in
the leaves of Elaesis guineensis tree
11. In fact, Elaesis guineensis leaf extract contains more antioxidative
phenolic compounds than various green tea extracts 12. Therefore, Elaesis
guineensis leaf extract is a potential source of functional food ingredient,
based on reports of its health benefit 13 .This study is aimed at evaluating
the protective potentials of Elaesis
guineensis leaf against crude oil contaminated diet induced nephrotoxicity
in rats.

2.0 Materials
and methods

The crude oil used for this study was obtained from
Nigeria National Petroleum Corporation (NNPC) Warri, Delta State, Nigeria. The
palm leaf used was obtained from Elaeis guineensis tree in Obiaruku,
Delta state, Nigeria Forty eight (48) female albino wistar rats with weights
ranging from 0.088kg to 0.182 kg obtained from the animal house of Department
of Anatomy, Delta State University, Abraka, Nigeria were used for this study.
The rats were housed in a standard wooden cage made up of wire gauze, net and
solid woods and left to acclimatize for one week on grower’s marsh and tap
water at laboratory temperature of 28o C and
12 hour day/ night regime. After the acclimatization period, the rats
were weighed and grouped.

2.1 Preparation
of leaf powder.

The
leaves of Elaeis guineensis were isolated from the stock and
sun- dried. The dried leaf was then ground with domestic kitchen blender into a
fine powder and stored in a clean and sealed plastic container

2.2 Treatment
of animals

The forty eight (48) female albino wistar rats were
assigned to six (6) groups according to their weights, with eight rats in each
group. Rats in the control, Group 1 were fed with grower’s marsh only. Rats in
Group 2 were fed with grower’s marsh treated with 5g of powdered Elaeis
guineensis leaf. Group 3 rats were fed with grower’s marsh treated 10g of powdered
Elaeis guineensis leaf. Group 4 rats were fed with grower’s marsh
contaminated with crude oil (4ml per 100g of feed).This
concentration of crude oil in diet was established to be tolerated by the rats
over a long period in a preliminary study. Rats in Group 5 were fed
grower’s marsh contaminated with crude oil (4ml per 100g of feed) plus 5g of
powdered palm fronds. While rats in Group 6 were fed with crude oil
contaminated marsh (4ml per 100g of feed) plus 10g of powdered palm leaves. The
rats in each group were allowed access to clean drinking water while the
experiment lasted. The feeds were prepared fresh daily and stale feed remnants
were discarded regularly. This was done every morning
between the hours of 8 am – 9 am and each group provided with 400 g of the
respective diet. The animals in each group were exposed to their
respective diets for three and six months respectively. The National Institute of health guide for the care and use
of laboratory animals (NIH, 1985) was adhered to in the course of the
experiment

 

 

2.3 Collection
of samples

After three months, four rats were sacrificed in each
group and the kidneys collected. Five grams (5.0 g) of the kidneys were weighed
in chilled conditions and homogenized with 5ml of normal saline in a mortar. The mixture was diluted with 45 ml of buffered saline (pH
7.4) before it was subjected to centrifugation at 2, 500 rpm and the
supernatant was transferred into plastic tubes and stored at – 4o C in the refrigerator before
used for analysis within forty eight hours. This same procedure was adopted
after six months exposure period.

2.4 Determination of lipid peroxidation and xanthine oxidase activity

The activity of
xanthine oxidase in the kidney of rats was measured using the method of
Bergmeyer et. al. 14, a reaction based on the oxidation of xanthine to uric
acid, a molecule that absorbs light maximally at 290 nm. A unit of activity is
that forming one micromole of uric acid per minute at 25oC. Lipid
peroxidation in the kidney of rats was measured by the thiobarbituric acid
reacting substances TBARS, method of Gutteridge and Wilkins 15.Total
superoxide dismutase activity was assayed using the method of Misra and
Fredorich 16. Catalase was assayed as reported by Rani et al. 17  

2.5Statistical Analysis

Analysis
of variance (ANOVA) and post Hoc Fisher’s test for multiple comparison
were carried out using version 20 of   statistical package for social science (SPSS) to
determine statistical significant differences between means. P values <0.05 were taken as being significantly different   3.0 Results and Discussion The effects of Elaeis guineensis leaf on kidney lipid peroxidation and xanthine oxidase activity against crude oil induced nephrotoxicity in rats after three and six months are shown in tables 1 and 2.  Lipid peroxidation in the kidney of rats exposed to crude oil contaminated diet (group 4) was significantly (P<0.05) higher in comparison with the control (group 1). Rats fed palm leaf pretreated diets (Group 2 and 3) showed significantly lower kidney levels of lipid peroxidation when compared with the control (group 4). Moreover, rats fed crude oil contaminated diets that was pretreated with various amounts of Elaesis guineensis leaf (Group 5 and 6) exhibited significantly lower kidney lipid peroxidation level when compared with the control (group 1) and  rats fed crude oil contaminated diet alone (group 4) .  Lipid peroxidation, is an index of oxidative stress, induces disturbance of functional loss of biomembranes, that results in inactivation of membrane bound receptors and enzymes 17, 18, 19. The present study shows that the consumption of crude oil treated diet increased the level of lipid peroxidation in rats.  This study shows that exposure to crude oil leads to oxidative damage of the kidney as evident by the rise in renal level of lipid peroxidation. This is based on the premise that metabolism of hydrocarbons generates free radicals 20. This is in line with earlier studies 5, 6, 7, 21. Elaesis guineensis leaf is rich in bioactive phytochemicals whose antioxidant activity is several folds higher than that of vitamins C and E 22, 23, 24.This may be the basis for the decreased level of lipid peroxidation in the kidney of rats exposed to crude oil that was treated with Elaesis guineensis leaf The kidney oxidative stress enzyme (xanthine oxidase, Sod and catalase) activities were significantly (P<0.05) lower in rats fed crude oil contaminated diets (group 4) in comparison with all the experimental groups (Tables 1 and 2).Rats fed with Elaesis guineensis leaf treated crude oil contaminated diet (Groups 5 and 6) have significantly higher xanthine oxidase activities in the kidney when compared with rats fed with crude oil contaminated diet only (group 4). However, rats fed with only Elaesis guineensis leaf treated diets (Groups 2 and 3) have significantly higher  oxidative stress marker enzyme  activities when compared with rats fed with only crude oil contaminated diet (group 1). Xanthine oxidase is involved in phase one process in the inactivation of xenobiotics in animals 25. The increase in the activity of xanthine oxidase in rats exposed to Elaesis guineensis leaf treated diet indicates response of the enzyme to enhance the metabolism of endogenous xanthine. This is in a bid to increase the production of uric acid, a potent antioxidant 7, 25, 26. The decrease in activity of xanthine oxidase in rats exposed to crude oil contaminated diet alone shows that the metabolism of crude oil leads to a reduced ability to produce uric acid Nevertheless, the alteration in the activity of oxidative enzymes had been reported as a measure of oxidative stress 27. However, addition of ground Elaesis guineensis leaf resulted in decrease in toxic effects of crude oil. This is exhibited in the increase in activities of oxidative stress marker enzymes towards control values in rats fed with crude oil contaminated diets that were pretreated with Elaesis guineensis leaf. This is due to the ability of Elaesis guineensis leaf to act as an antioxidant, protecting endothelial cells of the kidney against reactive free radicals thereby restoring the level of antioxidant enzymes 11, 13. Substances with antioxidant potentials possess health promoting properties, since they quench free radicals which are involved in many diseases processes 13, 28, 29, 30. Generally, the deleterious action of crude oil on kidney tissue and the protective influence of the Elaesis guineensis leaf is further highlighted by histological examination of the kidney tissue (Figure 1).Previous study had shown that plant materials with antioxidant properties can attenuate the negative effect of crude oil on animals 31 4.0 Conclusion This study has indicated that the ingestion of crude oil treated diet can result in increase in oxidative stress and consequent kidney damage. However, the crude oil toxicities were reversed by the consumption of diets that were pretreated with Elaesis guineensis leaf. This study, therefore, shows possible protective role of Elaesis guineensis leaf against crude oil induced nephrotoxicity.   5.0   References 1  Otitoju O, Onwurah .INE. Preliminary investigation into the possible endocrine disrupting activity of bonny light crude oil contaminated diet on wistar rats. Biokemistri 2007; 19(2):23-28       2   Ovuru SS, Ekweozor IKE. 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In: Methods of Enzymatic Analysis, HV Bergmeyer    (eds).New York: Academic Press. 1974; 2:  428–429. 15 Guttridge JMC, Wilkins C. Copper dependent hydroxyl radical damage to ascorbic acid formation of thiobarbituric acid reactive products. FEBS Lett. 1982; 137: 327-340.  16   Misra HP, Fridovich I. The role of superoxide ion in the autoxidation of epinephrine and a simple assay for superoxide dismutase. J Biol Chem 1972 (247): 3170 – 3175. 17    Rani P, Meena UK, Karthikeyan J. Evaluation of antioxidant properties of berries. India J Clin Biochem 2004; 19 (2) 103-110. 18    Halliwell B. Free radicals and antioxidants: a personal view. Nutr Rev1994; 5:253-265. 19    Niki E. Lipid peroxidation products as oxidative stress biomarkers. Biofactors. 2008;34(2):171-180 20    Greenberg ME, Li XM, Giugiu BG, Gu X, Qin J, Salomon RG, Hazen S.The lipid whisker model of the structure of oxidized cell membranes. J Biol Chem 2008; 283:2385-239 21    Achuba FI. Spent engine oil mediated oxidative stress in cowpea (Vigna unguiculata) seedlings. EJEAFChe. 2010; 9(5): 910-917 22    Alisi CS., Ojiako AO., Osuagwu CG, Onyeze GOC (2011) Response pattern of antioxidants in carbon tetrachloride-induced hepatoxicity is tightly logistic in rabbits. Eur J Med Plants. 2011;1:118-129 23    Cowan MM (1999) Plant products as antimicrobial agents. Clin. Microbial. Rev. 1999; 12(4):564-582 24    Lee YL, Jian SY, Lian PY, Mau JL. Antioxidant properties of extract from a white mutant of the mushroom Hypsizigusmarmoreus. J. Food Compos Anal. 2008;21:116-124 25    Jaffri JM, Mohamed S, Ahmad IN, Mustapha NM, Manap YA, Rohimi N. Effects of catechin-rich Elaesis guineensis leaf extract on normal and hypertensive rats kidney and liver. Food Chem.2011; 128:433–441 26    Ezedom T, Asagba SO. Effect of a controlled food-chain mediated exposure to cadmium and arsenic on oxidative enzymes in the tissues of rat Toxicol Reports 2016 ;(3) :708–715 27    Achuba FI. African land snail Achatina marginatus, as bioindicator of environmental pollution. North- Western J  Zool 2008; 4 (1): 1-5 28    Förstermann U, Xia N,   Li. H. Roles of Vascular Oxidative Stress and Nitric Oxide in the Pathogenesis of Atherosclerosis. Circulation Res.2017; 120:713-735 29    Hybertson BM, Gao, B, Bose, SK., McCord JM. Oxidative Stress in health and disease: The therapeutic potential of Nrf2 activation. Mol Asp Med 2011;32(4):234-246 30    Galli F, Piroddi M., Annetti C, Aisa C,  Floridi E.,  Floridi A (2005) Oxidative stress and reactive oxygen species. Contrib Nephrol 2005; 149: 240-260 31    Achuba FI, Ubogu LA, Ekute BO. Moringa oleifera attenuates crude  oil  contaminated diet induced biochemical effects in wistar albino rats UK J Pharm Biosci 2016;  4(5) 70-77   Table 1.  The effect of  Elaeis guineensis leaf on the level of  oxidative stress indicators in the kidney of rats after three months of exposure to crude oil contaminated diet. Groups Lipid peroxidation    Xanthine oxidase activity (nmol/g  tissue)           (units/g  tissue) SOD activity            Catalase activity  (units/g  tissue)         (nmol/g tissue) Group 1 0.35±  0.05 a                     60.04 ±  4.28 a                                                          26.75 ±  2.21 a                   54.53± 2.55 a Group 2 0.14 ±  0.02 b                    60.83 ±  1.76 a 28.63 ±  3.62 a                   51.33± 3.61 b Group 3 0.10 ±   0.03 b                   69.28 ±  3.34 b                                  29.44 ±  1.47 b                   52.1 2± 1.15 b Group 4 0.76 ±   0.10 c                   42.43 ± 1.78 c 20.10 ± 1.66 c                    46.42± 2.11 c Group 5 0.52  ±  0.01 d                   51.09 ±  2.70 d 22.22 ±  1.80 d                   49.44± 1.52 d Group 6 0.34  ±  0.01 a                   57.05 ±   5.89 a 24.52 ±   1.33 a                  50. 33± 1.66 b   Each value represents mean ± standard deviation. n = 4 in each group. Values not sharing a common superscript letter in the same column differ significantly at (P < 0.05). Group 1: ((Normal Control). Group 2:   feed mixed with 5.0g Elaesis guineensis leaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf. Group 4: Feed mixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixed with 5.0 g of Elaesis guineensis leaf. Group 6: contaminated diet mixed with 10.0 g of Elaesis guineensis leaf.                           Table 2.  The effect of  Elaeis guineensis leaf on the level of  oxidative stress indicators in the kidney of rats after six months of exposure to crude oil contaminated diet Groups Lipid peroxidation    Xanthine oxidase activity (nmol/g  tissue)           (units/g  tissue) SOD activity            Catalase activity  (units/g  tissue)         (nmol/g tissue) Group 1 0.42±  0.08 a                     62.04 ±  3.80 a                                                          28.88 ±  1.11 a                   53.97± 1.45 a Group 2 0.22 ±  0.01 b                    61.41 ±  2.64 a 27.96 ±  3.62 a                   52.36± 2.55 a Group 3 0.11 ±   0.04 b                   68.24 ±  2.22 b                                  29.55 ±  2.81 a                   52.66± 1.22 a Group 4 0.89 ±   0.11 c                   38.43 ± 2.66 c 18.33 ± 1.88c                    43.31± 1.53 c Group 5 0.66  ±  0.12 d                   54.11 ±  3.50 d 23.43 ±  1.92 d                   50.02± 1.68 b Group 6 0.53  ±  0.06 a                   55.44 ±   6.70 a 24.99 ±   1.63 a                  50. 91± 1.74 b   Each value represents mean ± standard deviation. n = 4 in each group. Values not sharing a common superscript letter in the same column differ significantly at (P < 0.05). Group 1: ((Normal Control). Group 2:   feed mixed with 5.0g Elaesis guineensis leaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf. Group 4: Feed mixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixed with 5.0 g of Elaesis guineensis leaf. Group 6: contaminated diet mixed with 10.0 g of Elaesis guineensis leaf.                                                                   Group 1: ((Normal Control). Group 2:   feed mixed with 5.0g Elaesis guineensis leaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf. Group 4: Feed mixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixed with 5.0 g of Elaesis guineensis leaf. Group 6: contaminated diet mixed with 10.0 g of Elaesis guineensis leaf.     Figure 1: Photomicrographs of kidney section of rats fed crude oil contaminated diet and diets pretreated with different amount of ground Elaesis guineensis leaf