EFFECT leaf (5.0 g and 10.0 g respectively).

EFFECT OF PRETREATMENT OF CRUDE OIL CONTAMINATED DIET WITH OIL PALM
LEAF ON LIPID PEROXIDATION AND XANTHINE OXIDASE ACTIVITY IN THE KIDNEY OF 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 was comprised forty eight female Wister rats divided into six groups of
eight rats each. The rats were fed as described thus. Group 1: ((normal
Control). Group 2:   feed mixed with 5.0g
oil palm leaf. Group 3: feed mixed with 10.0g oil palm leaf. Group 4: Feed
mixed with 4ml crude oil (Crude oil Control). Groups 5 and 6: Contaminated diet
mixed with ground oil palm leaf (5.0 g and 10.0 g respectively). At the end of
exposure periods (three and six months respectively), the rats were sacrificed
and the kidney used to prepare supernatant used for the determinations oxidative
stress indices (lipid peroxidation and xanthine oxidase activity). The results
show that pretreatment of crude oil contaminated diet with oil palm leaf tend
to restore values of lipid peroxidation and xanthine oxidase activity close to
control values. Thus, it is pertinent to state that there exist potentials in
the use oil palm leaf in the treatment of crude oil toxicity. And indeed
setting a fresh agenda for further serious scientific investigations

 

 

 

Keywords: Crude oil, Kidney, Lipid peroxidation, Oil
palm, .Xanthine oxidase,

 

 

 1.0 Introduction

The
impact of crude oil spillage on the ecosystem as a result of oil exploration
activities, equipment failures, corrosion, illegal bunkering, oil theft and
illicit refining are well documented 1-3.

Crude oil
has been proved to alter levels of oxidative stress markers in animals 4, 5.
These oxidative stress markers include lipid peroxidation and changes in the
activities of xanthine oxidase. The process of lipid peroxidation precedes
oxidative damage in plants and animals and alterations in the level of
antioxidants represent a measure of oxidative stress. In addition, the activity
of xanthine oxidase is an example of defense mechanism as well as a measure of
oxidative stress 6. Exposure to petroleum hydrocarbons can cause impairment
of renal function via a mechanism dependent on oxidative stress 7.

Various
parts of oil palm tree such as the trunk, fronds, leaves, mesocarp fibres,
shells and empty fruit bunches which are disposed as waste can be used
medicinally. The juice squeezed out from palm leaves can be applied to enhance
wound healing while the sap can be used as laxative 8.

The oil
palm tree contains biologically active compounds with medicinal and antioxidant
properties 9, 10. It has been confirmed that flavonoid, tannin and phenolic
are the main phytochemical constituents in oil palm leaves, hence its ability
as an effective antioxidant 11. Oil palm fronds extract has 8% higher
contents of non-toxic antioxidative phenolic compounds than various green tea
extracts 12. Oil palm frond extract may be a potential new source of
functional food ingredient, based on reports of its benefit on health 13 .The
aim of this study was to determine the effects of Oil palm leave treated crude
oil contaminated diet on the level of lipid peroxidation and xanthine oxidase
activity in the kidney of 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 frond 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 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 28C and 12 hour day/ night
regime. After the acclimatization period, the rats were weighed and
grouped.

2.1 Preparation of leaf powder.

The
leaves 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

2.2Treatment 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 which is Group 1 were fed with grower’s marsh
only. Rats in Group 2 were fed with grower’s marsh and 5g of powdered palm
frond. Group 3 rats were fed with grower’s marsh and 10g of powdered palm leaves.
Group 4 rats were fed with grower’s marsh contaminated with crude oil (4ml per
100g of feed). 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. 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
adopted all through the experiment

 

2.3 Collection
of samples

After the exposure period, all the rats were
sacrificed and the kidneys were harvested. 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 known amount of
buffered saline before being centrifuged and the supernatant was transferred into
plastic tubes and stored at – 4C before used for analysis within forty eight
hours.

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, 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.

2.5Statistical Analysis

Analysis
of variance (ANOVA) and post
Hoc Fisher’s test for multiple comparison was performed using statistical
package for social science (SPSS), version 20  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 rats exposed to crude oil contaminated ( group 4) was significantly (P<0.05) higher in comparison with the control (group 1). Rats fed palm leave 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 oil palm leaves (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, which is a potential marker of oxidative stress, induce disturbance of cell membrane, and functional loss of biomembranes, that results in inactivation of membrane bound receptors and enzymes 16, 17, 18. The present study shows that the consumption of crude oil contaminated diet increased the level of lipid peroxidation in rats.  This study shows that exposure to hydrocarbons present in crude oil can lead 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 present in crude oil generate free radicals 19. This is in consonance with previous studies by 5, 6, 7, 20. Oil palm (Elaeis guineensis) frond is rich in bioactive phytochemicals such as polyphenols and these polyphenolic compounds are considered to have antioxidant activity that is several folds higher than that of vitamins C and E 21, 22, 23.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 oil palm leaf The  kidney xanthine oxidase activities were significantly (P<0.05) lower  in the rats fed crude oil contaminated diets (group 4) in comparison with all the experimental groups( Tables 1 and 2).Rats fed with oil palm 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 oil palm leaf treated diets (Groups 2 and 3) have significantly higher kidney xanthine oxidase activity 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 24. The increase in the activity of xanthine oxidase in rats exposed to oil palm 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, 24, 25. The decrease in activity of xanthine oxidase in rats exposed to crude oil contaminated diet alone shows that the metabolism of crude oil hydrocarbons leads to a reduced ability to produce uric acid Nevertheless, the increase in the activity of oxidative enzymes had been reported as a measure of oxidative stress 26. However, addition of ground oil palm leaves resulted in decrease in toxic effects of crude oil. This is exhibited in the decrease in xanthine oxidase activity in rats fed with crude oil contaminated diets that were pretreated with oil palm leaves. This is due to the ability of oil palm leaves to act as an antioxidant, protecting endothelial cells of the kidney against reactive free radicals thereby decreasing 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, 27, 28, 29 This study has indicated that the consumption of crude oil contaminated diet can result in increase in oxidative stress which causes corresponding increases in lipid peroxidation levels and xanthine oxidase activity. However, the crude oil toxicities were reversed by the consumption of diets that were pretreated with oil palm leaves. 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Nat Rev Drug Discovery. 3:205-214                     Table 1.  The effect of  Elaeis guineensis leaf on the level of lipid peroxidation and xanthine oxidase activity in the kidney of rats after three months of exposure to crude oil contaminated diet. Groups Lipid peroxidation (nmol/g  tissue)  Xanthine oxidase activity (units/g  tissue) Group 1 0.35±  0.05 a 60.04 ±  4.28 a Group 2 0.14 ±  0.02 b 60.83 ±  1.76 a Group 3 0.10 ±   0.03 b 69.28 ±  3.34 b Group 4 0.76 ±   0.10 c 42.43 ± 1.78 c Group 5 0.52  ±  0.01 d 51.09 ±  2.70 d Group 6 0.34  ±  0.01 a 57.05 ±   5.89 a   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 oil palm leaf. Group 3: feed mixed with 10.0g oil palm leaf. Group 4: Feed mixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixed with 5.0 g of oil palm leaf. Group 6: contaminated diet mixed with 10.0 g of oil palm leaf.                     Table 2.  The effect of Elaeis guineensis leaf on the level of lipid peroxidation and xanthine oxidase activity in the kidney of rats after six months of exposure to crude oil contaminated diet. Groups Lipid peroxidation (nmol/g  tissue)  Xanthine oxidase activity (units/g  tissue) Group 1 0.42±  0.08 a 62.04 ±  3.80 a Group 2 0.22 ±  0.01 b 61.41 ±  2.64 a Group 3 0.11 ±   0.04 b 68.24 ±  2.22 b Group 4 0.89 ±   0.11 c 38.43 ± 2.66 c Group 5 0.66  ±  0.12 d 54.11 ±  3.50 d Group 6 0.53  ±  0.06 a 55.44 ±   6.70 a   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 oil palm leaf. Group 3: feed mixed with 10.0g oil palm leaf. Group 4: Feed mixed with 4ml crude oil (Crude oil Control). Group 5: Contaminated diet mixed with 5.0 g of oil palm leaf. Group 6: contaminated diet mixed with 10.0 g of oil palm leaf.