INFLUENCE OF ELAESIS GUINEENSIS LEAF IN DIET ON PETROLEUM-MEDIATED KIDNEY DAMAGEIN RAT Achuba ,F. I .Department of Biochemistry, Delta State University, PMB 1, Abraka Nigeria. [email protected]  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 forty eight female rats divided into six groups of eightrats 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 respectively.Rats in group 4 were fed with crude oil contaminated diet. Rats in groups 5 and6 were fed with contaminated diet mixed with known amount of ground Elaesis guineensis leaf. Biochemical andhistological 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 oxidaseactivity, superoxide dismutase activity and catalase activity close to controlvalues.

Histological examination indicatesprotective effect of Elaesis guineensis leaf against deleterious effect ofcrude oil on the kidney.   Thus, it is pertinent to state thatthere exist potentials in the use Elaesisguineensis leaf in the treatment of crude oil toxicity. And indeed settinga fresh agenda for further serious scientific 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 superoxidedismutase (SOD) and catalase activities are part of oxidative stress indices6. Lipid peroxidation elicits oxidative damage in plants and animals and itsvalue in conjunction with alterations in the level of antioxidants represent ameasure of oxidative stress. Similarly, the activity of xanthine oxidase 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 Forty eight (48) female albino wistar rats with weightsranging from 0.088kg to 0.182 kg obtained from the animal house of Departmentof 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 andsolid woods and left to acclimatize for one week on grower’s marsh and tapwater 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 forty eight (48) female albino wistar rats wereassigned to six (6) 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 theexperiment   2.3 Collectionof samplesAfter three months, four rats were sacrificed in eachgroup and the kidneys collected. Five grams (5.

0 g) of the kidneys were weighedin chilled conditions and homogenized with 5ml of normal saline in a mortar. The mixture was diluted with 45 ml of buffered saline (pH7.4) 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 lipid peroxidation and xanthine oxidase activityThe activity ofxanthine oxidase in the kidney of rats was measured using the method ofBergmeyer et.

al. 14, 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 15.Totalsuperoxide dismutase activity was assayed using the method of Misra andFredorich 16. Catalase was assayed as reported by Rani et al. 17  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) todetermine statistical significant differences between means. P values <0.05were taken as being significantly different 3.0 Results and DiscussionThe effects of Elaeisguineensis leaf on kidney lipid peroxidation and xanthine oxidase activityagainst crude oil induced nephrotoxicity in rats after three and six months areshown in tables 1 and 2.

 Lipidperoxidation in the kidney of rats exposed to crude oil contaminated diet (group4) was significantly (P<0.05) higher in comparison with the control (group1). Rats fed palm leaf pretreated diets (Group 2 and 3) showed significantlylower kidney levels of lipid peroxidation when compared with the control (group4). Moreover, rats fed crude oil contaminated diets that was pretreated withvarious amounts of Elaesis guineensisleaf (Group 5 and 6) exhibited significantly lower kidney lipid peroxidation levelwhen compared with the control (group 1) and rats fed crude oil contaminated diet alone (group 4) .  Lipid peroxidation, is an index of oxidativestress, induces disturbance of functional loss of biomembranes, that results ininactivation of membrane bound receptors and enzymes 17, 18, 19.

The presentstudy shows that the consumption of crude oil treated diet increased the levelof lipid peroxidation in rats.  Thisstudy shows that exposure to crude oil leads to oxidative damage of the kidneyas evident by the rise in renal level of lipid peroxidation. This is based onthe premise that metabolism of hydrocarbons generates free radicals 20. Thisis in line with earlier studies 5, 6, 7, 21. Elaesis guineensis leaf is rich in bioactive phytochemicals whoseantioxidant 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 inthe kidney of rats exposed to crude oil that was treated with Elaesis guineensis leafThe kidney oxidative stressenzyme (xanthine oxidase, Sod and catalase) activities were significantly(P<0.05) lower in rats fed crude oil contaminated diets (group 4) incomparison with all the experimental groups (Tables 1 and 2).

Rats fed with Elaesis guineensis leaf treated crude oilcontaminated diet (Groups 5 and 6) have significantly higher xanthine oxidaseactivities in the kidney when compared with rats fed with crude oilcontaminated 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 contaminateddiet (group 1).Xanthineoxidase is involved in phase one process in the inactivation of xenobiotics inanimals 25. The increase in the activity ofxanthine oxidase in rats exposed to Elaesisguineensis leaf treated diet indicates response of the enzyme to enhancethe metabolism of endogenous xanthine. This is in a bid to increase theproduction of uric acid, a potent antioxidant 7, 25, 26. The decrease inactivity of xanthine oxidase in rats exposed to crude oil contaminated dietalone shows that the metabolism of crude oil leads to a reduced ability to produceuric acidNevertheless,the alteration in the activity of oxidative enzymes had been reported as ameasure of oxidative stress 27. However, addition of ground Elaesis guineensis leaf resulted indecrease in toxic effects of crude oil. This is exhibited in the increase inactivities of oxidative stress marker enzymes towards control values in ratsfed with crude oil contaminated diets that were pretreated with Elaesisguineensis leaf.

This is due to the ability of Elaesis guineensis leaf to act as an antioxidant, protectingendothelial cells of the kidney against reactive free radicals therebyrestoring the level of antioxidant enzymes 11, 13. Substances withantioxidant potentials possess health promoting properties, since they quenchfree radicals which are involved in many diseases processes 13, 28, 29, 30. 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 propertiescan attenuate the negative effect of crude oil on animals 314.0 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. Preliminaryinvestigation into the possible endocrine disrupting activity of bonny lightcrude oil contaminated diet on wistar rats. Biokemistri 2007; 19(2):23-28      2   OvuruSS, Ekweozor IKE.

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  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

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