PROTECTIVE The toxicity of petroleum hydrocarbon across the

PROTECTIVE INFLUENCEOF ELAESIS GUINEENSIS LEAF IN DIET ONPETROLEUM-MEDIATED KIDNEY DAMAGE IN RAT  AbstractThe toxicity of petroleum hydrocarbon across theliving systems is now a common knowledge among the scientific community. Whatis lacking is a mini-scale antidote that can be adopted by the inhabitants ofcrude oil producing areas of the world.

This was the reason for this study. Thestudy is comprised ninety six female rats divided into six groups of sixteenrats each. Rats in control group were fed with diet without any treatment whilerats in groups 2 and 3 were fed with diets treated with known amount of Elaesis guineensis leaf. Rats in group 4were fed with crude oil contaminated diet. Rats in groups 5 and 6 were fed withcontaminated diet mixed with known amount of ground Elaesis guineensis leaf.

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Biochemical and histological analysis werecarried out after three and six months respectively. The results show thatpretreatment of crude oil contaminated diet with Elaesis guineensis leaf tend to restore valuesof lipid peroxidation, xanthine oxidase activity (XO), superoxide dismutaseactivity (SOD) and catalase activity (CAT) close to control values. Histological examination indicates protective effect of Elaesisguineensis 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 treatmentof crude oil toxicity. And indeed setting a fresh agenda for further seriousscientific investigations   Keywords: Catalase, Crude oil, Kidney, Lipidperoxidation, Elaesis guineensis,Superoxide dismutase .

Xanthine oxidase,     1.0 Introduction Humans and animals get exposed to crude oil orits byproducts when these chemicals are released into the surroundings duringoil exploration activities, equipment failures, corrosion, illegal bunkering,usage, oil theft and illicit refining 1-3. Crude oil stimulates oxidativestress in animals 4, 5. Lipid peroxidation, xanthine oxidase (XO), superoxidedismutase (SOD) and catalase (CAT) activities are part of oxidative stressindices 6. Lipid peroxidation elicits oxidative damage in plants and animalsand its value in conjunction with alterations in the level of antioxidantsrepresent a measure of oxidative stress. Similarly, the activity of XO is adefense mechanism as well as measure of oxidative stress in animals 6. Reporthas it that the deleterious action of crude oil on the kidney is based onoxidative stress 7.

 Byproducts of the Elaesis guineensis tree areimportant medicinally. This is because the leaf juice have wound healingproperty while the sap is used as laxative 8.This is due to   compounds rich in medicinal and antioxidantproperties inherent in Elaesis guineensisleaf 9, 10.

  The antioxidant action isattributed to the presence of phytochemicals (flavonoid, tannin and phenols) inthe leaves of Elaesis guineensis tree11. In fact, Elaesis guineensis leaf extract contains more antioxidativephenolic compounds than various green tea extracts 12. Therefore, Elaesisguineensis leaf extract is a potential source of functional food ingredient,based on reports of its health benefit 13 .This study is aimed at evaluatingthe protective potentials of Elaesisguineensis leaf against crude oil contaminated diet induced nephrotoxicityin rats.

2.0 Materialsand methodsThe crude oil used for this study was obtained fromNigeria National Petroleum Corporation (NNPC) Warri, Delta State, Nigeria. Thepalm leaf used was obtained from Elaeis guineensis tree in Obiaruku,Delta state, Nigeria Ninety six female albino wistar rats with weights ranging from178 g to 182 g obtained from the animal house of Department of Anatomy, DeltaState University, Abraka, Nigeria were used for this study. The rats werehoused in a standard wooden cage made up of wire gauze, net and solid woods andleft to acclimatize for one week on grower’s marsh and tap water at laboratory temperature of 28o C and12 hour day/ night regime. After the acclimatization period, the ratswere weighed and grouped.2.1 Preparationof leaf powder.Theleaves of Elaeis guineensis were isolated from the stock andsun- dried.

The dried leaf was then ground with domestic kitchen blender into afine powder and stored in a clean and sealed plastic container2.2 Treatmentof animalsThe ninety six female albino wistar rats wereassigned to six groups according to their weights, with eight rats in eachgroup. Rats in the control, Group 1 were fed with grower’s marsh only. Rats inGroup 2 were fed with grower’s marsh treated with 5g of powdered Elaeisguineensis leaf. Group 3 rats were fed with grower’s marsh treated 10g of powderedElaeis guineensis leaf. Group 4 rats were fed with grower’s marshcontaminated with crude oil (4ml per 100g of feed).Thisconcentration of crude oil in diet was established to be tolerated by the ratsover a long period in a preliminary study. Rats in Group 5 were fedgrower’s marsh contaminated with crude oil (4ml per 100g of feed) plus 5g ofpowdered palm fronds.

While rats in Group 6 were fed with crude oilcontaminated marsh (4ml per 100g of feed) plus 10g of powdered palm leaves. Therats in each group were allowed access to clean drinking water while theexperiment lasted. The feeds were prepared fresh daily and stale feed remnantswere discarded regularly. This was done every morningbetween the hours of 8 am – 9 am and each group provided with 400 g of therespective diet. The animals in each group were exposed to theirrespective diets for three and six months respectively. The National Institute of health guide for the care and useof laboratory animals (NIH, 1985) was adhered to in the course of the experiment14  2.3 Collectionof samplesAfter three months, eight rats were sacrificed ineach group and the kidneys collected. One gram of the kidneys were weighed inchilled conditions and homogenized with 5ml of normal saline in a mortar.

The mixture was diluted with 5 ml of buffered saline (pH 7.4)before it was subjected to centrifugation at 2, 500 rpm and thesupernatant was transferred into plastic tubes and stored at – 4o C in the refrigerator beforeused for analysis within forty eight hours. This same procedure was adoptedafter six months exposure period.

2.4 Determination of biochemical and histological analysisThe activity ofxanthine oxidase in the kidney of rats was measured using the method ofBergmeyer et. al.

15, a reaction based on the oxidation of xanthine to uricacid, a molecule that absorbs light maximally at 290 nm. A unit of activity isthat forming one micromole of uric acid per minute at 25oC. Lipidperoxidation in the kidney of rats was measured by the thiobarbituric acidreacting substances (TBARS) method of Gutteridge and Wilkins 16.Totalsuperoxide dismutase activity was assayed using the method of Misra andFredorich 17. Catalase was assayed as reported by Rani et al.

18. The of method of Drury and Wallington 19 wasadopted for the histological study 2.5Statistical AnalysisAnalysisof variance (ANOVA) and post Hoc Fisher’s test for multiple comparisonwere carried out using version 20 of   statistical package for social science (SPSS) to determinestatistical significant differences between means.

P values <0.05 were takenas being significantly different   3.0 Results and DiscussionThe effects of Elaeisguineensis leaf on kidney lipid peroxidation and activities of oxidativestress marker enzymes against crude oil induced nephrotoxicity in rats afterthree and six months are shown in tables 1 and 2.  Lipid peroxidation in the kidney of ratsexposed to crude oil contaminated diet (group 4) was significantly (P<0.05)higher in comparison with the control (group 1). Rats fed palm leaf pretreateddiets (Group 2 and 3) showed significantly lower kidney levels of lipidperoxidation when compared with the control (group 4).

Moreover, rats fed crudeoil contaminated diets that was pretreated with various amounts of Elaesis guineensis leaf (Group 5 and 6)exhibited significantly lower kidney lipid peroxidation level when comparedwith   rats fed crude oil contaminateddiet alone (group 4) .  Lipid peroxidation is an index of oxidativestress, it induces functional loss of biomembranes, that results ininactivation of membrane bound receptors and enzymes 20, 21, 22. This studyshows that exposure to crude oil leads to oxidative damage of the kidney asevident by the rise in renal level of lipid peroxidation. This is based on thepremise that metabolism of hydrocarbons generates free radicals, that is inline with earlier studies 5, 6, 7, 23, 24. Elaesis guineensis leaf is rich in bioactive phytochemicals whoseantioxidant activity is several folds higher than that of vitamins C and E 25,26, 27.This may be the basis for the decreased level of lipid peroxidation inthe kidney of rats exposed to crude oil that was treated with Elaesis guineensis leafThe kidney oxidative stressenzyme (XO, SOD and CAT) activities were significantly (P<0.05) lower inrats fed crude oil contaminated diets (group 4) in comparison with all theexperimental groups (Tables 1 and 2). XO isinvolved in phase one process in the inactivation of xenobiotics in animals 28.

The increase in the activity of XO in rats exposed toElaesis guineensis leaf treated dietindicates response of the enzyme to enhance the metabolism of endogenousxanthine. This is in a bid to increase the production of uric acid, a potentantioxidant 7, 25, 29. The decrease in activity of XO in rats exposed tocrude oil contaminated diet alone shows that the metabolism of crude oil leadsto a reduced ability to produce uric acid. Nevertheless, the alteration in theactivity of oxidative enzymes had been reported as a measure of oxidative stress27. However, addition of ground Elaesisguineensis leaf resulted in decrease in toxic effects of crude oil. This isexhibited in the increase in activities of oxidative stress marker enzymestowards control values in rats fed with crude oil contaminated diets that werepretreated with Elaesis guineensis leaf. This is due to the ability of Elaesis guineensis leaf to act as anantioxidant, protecting endothelial cells of the kidney against reactive freeradicals thereby restoring the level of antioxidant enzymes 11, 13.

This isattributed to the presence of substances with antioxidant potentials and healthpromoting properties, which quench free radicals that are, involved in manydiseases processes 13, 29, 30, 32. Generally,the deleterious action of crude oil on kidney tissue and the protectiveinfluence of the Elaesis guineensisleaf is further highlighted by histological examination of the kidney tissue (Figure1).Previous study had shown that plant materials with antioxidant properties canattenuate the negative effect of crude oil on animals 334.

0 ConclusionThisstudy has indicated that the ingestion of crude oil treated diet can result inincrease in oxidative stress and consequent kidney damage. However, the crudeoil toxicities were reversed by the consumption of diets that were pretreatedwith Elaesis guineensis leaf. This study, therefore, shows possible protectiverole of Elaesis guineensis leaf against crude oil induced nephrotoxicity. 5.0   References1  Otitoju O, Onwurah .INE.

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, Annetti C, Aisa C,  Floridi E., Floridi A (2005) Oxidative stress and reactive oxygen species. ContribNephrol 2005; 149: 240-26033    Achuba FI, Ubogu LA, Ekute BO. Moringaoleifera attenuates crude  oil  contaminated diet induced biochemical effectsin wistar albino rats UK J PharmBiosci 2016;  4(5) 70-77 Table 1.  The effect of Elaeis guineensis leaf on the level of  oxidative stress indicators in the kidney ofrats 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 ineach group. Values not sharing a common superscript letter in the same columndiffer significantly at (P < 0.05).Group 1: ((Normal Control).

Group 2:   feed mixed with 5.0g Elaesis guineensisleaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf. Group 4: Feedmixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixedwith 5.0 g of Elaesis guineensis leaf.

Group 6: contaminated diet mixed with10.0 g of Elaesis guineensis leaf.             Table 2.  The effect of Elaeis guineensis leaf on the level of  oxidative stress indicators in the kidney ofrats 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 samecolumn differ significantly at (P < 0.05).

Group 1: ((Normal Control). Group 2:   feed mixed with 5.0g Elaesis guineensisleaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf.

Group 4: Feedmixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixedwith 5.0 g of Elaesis guineensis leaf. Group 6: contaminated diet mixed with10.0 g of Elaesis guineensis leaf.                                 Group 1: ((Normal Control). Group 2:   feed mixed with 5.

0g Elaesis guineensisleaf. Group 3: feed mixed with 10.0g Elaesis guineensis leaf.

Group 4: Feedmixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixedwith 5.0 g of Elaesis guineensis leaf. Group 6: contaminated diet mixed with10.0 g of Elaesis guineensis leaf.  Figure 1: Photomicrographs of kidney section of rats fed crude oil contaminated diet and dietspretreated with different amount of ground Elaesis guineensis leaf