Hepatoprotective and Nephroprotective Effects of ...€¦ · 2 Methanolic Extracts of Different...

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Hepatoprotective and Nephroprotective Effects of 1

Methanolic Extracts of Different Parts of Tamarindus 2

Indica Linn in Rats Following Acute and Chronic 3

Carbon Tetrachloride Intoxication 4

Mubarak L. Liman*and Sunday E. Atawodi** 5

6

Department of Biochemistry, Ahmadu Bello University Zaria, Nigeria 7

^Current address: Deptof Science Laboratory Technology, NuhuBamalli Polytechnic Zaria,Nigeria 8

9

ABSTRACT 10

Aims: To investigate the hepatoprotective and nephroprotective potential of the methanolic extracts of 11

the leaves, stem bark, seeds, fruit pulp, fruit bark and roots of Tamarindus indicaLinn in acute and chronic 12

rat model of organ injuries. 13

Study design:The acute-injury model involved intraperitoneal pre-treatment with 10 mg/kg body weight of 14

the extract for two days followed by intoxication with carbon tetrachloride at 0.6ml/kg, while the chronic 15

injury model involved repeated intoxication with carbon tetrachloride (0.3ml/kg) at every 72 hourly 16

intervals together with a concomitant 24 hourly administration of the extracts (5mg/kg) for twelve days, 17

following initial CCl4 intoxication at 0.6ml/kg. 18

Place and Duration of Study: Department of Biochemistry, Faculty of Science, Ahmadu Bello University 19

Zaria, Kaduna State, Nigeria. January 2011-June 2011. 20

Methodology: In both acute and chronic experimental model, the rats were sacrificed at the end of the 21

each experiment. Bilirubin,aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were 22

determined from serum as indices of hepatic injuries while urea and creatinine were determined as 23

markers of kidney damage. 24

Results: Extract treatment caused a significant (p<0.05) decrease in the activities of ALT from 25

61.04±2.77U/I in CCl4 control group to between 15.82±2.63 and 50.67±3.44U/I while AST activities were 26

similarly lowered from 100.57±3.27U/I to between 25.10±1.48U/I and 53.45±3.19 U/I. There was 27

alsosignificant (p<0.05) decrease in the levels of bilirubin, urea and creatinine when compared to the CCl4 28

control. In general, extracts from the fruit pulp, the stem bark and fruit bark demonstrated better 29

hepatoprotective and nephroprotective potential than those of the seed, root and leaves. 30


Conclusions:Various parts of Tamarindus indica possess hepatoprotective and nephroprotective 31

properties to justify their usage in traditional medicine in Nigeria and some other developing countries. 32

Keywords:Tamarindus indica;hepatoprotective effect; nephroprotective effect;kidney damage; liver 33

damage. 34

1. INTRODUCTION 35

Since pre-historical times, plant materials have widely been used for treatment of illness and diseases 36

[16] in traditional societies of Africa and elsewhere [10]. Traditional medical practices on the African 37

continent date as far back as 4000 years and were the sole medical system for healthcare delivery before 38

the advent of orthodox or modern medicine. Even today, traditional medicine is still the predominant 39

means of health care in developing countries where larger percentage of the total population is poor. 40

Even in modern medicine, plants are the basis for the development of drugs because of their component 41

phytochemicals such as alkaloids, tannins, flavonoids and other phenolic compounds that produce 42

definite physiological and pharmacological action in the body of living organisms [15]. Thus, a systematic 43

search for useful bioactivities from medicinal plants such as Tamarindus indica Linn is now considered to 44

be a rational approach in nutraceutical and drug development [11]. 45

Tamarind (Tamarindus indica Linn) is a perennial commercial and ornamental herb belonging to the 46

dicotyledonous family of Leguminosae which is known by several names such as Indian date (English), 47

tsamiya(Hausa) and tamrhind(Arabic). It is a slow growing but long living plant (80-200 years) that 48

averages 20-25 m in height, 1m in diameter, and has a wide spreading crown with a short, stout trunk. It 49

grows wild in many tropical and sub-tropical regions of the world, as it is well adapted to semi-arid tropical 50

as well as humid tropical areas with seasonally high rainfall. Tamarindus indica is widely used in 51

traditional medicine in Africa for the treatment of many diseases such as fever, dysentery, jaundice, 52

gonorrheal and gastrointestinal disorders [15]. For example, the pulp has been reported to be useful in 53

the management of a number of ailments including the alleviation of sunstroke, Daturapoisoning and the 54

intoxicating effects of alcohol and cannabis [12]. It can be gargled for sore throats, dressing of wounds, is 55

said to aid in the cure of malarial feverand in the restoration of sensation in cases of paralysis. Also, the 56

fruits are reported to have anti-fungal and anti-bacterial properties [14] in many countries, while the 57

powdered seed husks or seed extracts is used in the treatment of boils, ulcer, diabetes and dysentery in 58

Cambodia and India. 59

Scientific investigations have indeed confirmed the antibacterial, antifungal, hypoglycemic, anti-60

hypercholesterolemic and cytotoxic effects [15, 16, 21] of the T. Indica seeds and fruit pulp, which have 61

also been demonstrated to enhance the bioavailability of drugs like ibuprofen in humans [11]. 62

Furthermore, it has been found that ethanol and ethyl acetate extracts prepared from the seed coat 63

exhibited anti-oxidative activity as measured by the thiocyanate and thiobarbituric acid (TBA) methods 64

[18]. Similarly, Komutarinet al [16] reported the in vitro and in vivo anti-inflammatory capacities of the pulp 65


and seed extracts through modulation of nitric oxide production. However, there appears no systematic 66

and comprehensive study evaluating the different parts of the plant for in vivo hepatoprotective and 67

nephroprotective effects. Therefore this work seeks to establish the in vivo hepatoprotective and 68

nephroprotective effects of different parts of Tamarindus indica in acute and chronic animal models of 69

organ toxicity. 70

71

2. MATERIALS AND METHODS 72

2.1 Plant material collection and extraction 73

The various parts of Tamarindus indica (leaves, stem bark, root bark, and whole fruits) were carefully 74

collected from a tree in Zaria Local Government area of Kaduna State, Nigeria. The plant was 75

authenticated at the Herbarium Section of the Biological Science Department, Ahmadu Bello University, 76

Zaria, Nigeria where a voucher number 900265 was assigned.The whole fruit was then carefully peeled 77

off to obtain the fruit bark and subsequently the seeds separated from the pulp. Samples of leaves, stem 78

bark, root bark, fruit bark and seeds were separately dried at room temperature and pulverized using 79

mortar and pestle. They were then defatted with petroleum ether for 6 hrs and then extracted with 80

methanol (4hrs x 2 times) using Soxhlex extractor. The combined methanol portions were taken to 81

dryness in vacuo in a desiccator and kept at -4oC until required. 82

83

The fruit pulp extracts were obtained by gentle warm maceration over a bath at 50°C. A weighed amount 84

of the fruits were put into a beaker with 300ml methanol and allowed to stand for 2hrs. The mixture was 85

then thoroughly macerated, and the seeds and debris picked out with the aid of laboratory tongs. 86

Petroleum ether (300ml) was then added and the entire mixture was shaken intermittently for 2 hours. 87

The liquid suspension was then carefully decanted into a separating funnel while the remaining debris 88

was discarded. The mixture in the separating funnel was then allowed to stand for clear separation of the 89

two immiscible layers. The individual layers were then carefully run out and collected in a pre-weighed 90

bottle. The process was repeated and the methanolic layer was combined and then dried in vacuo to 91

obtain the dried extracts which were then stored in air tight dark glass bottles in a refrigerator at -4°C until 92

required [4].This elaborate procedure was necessitated by the fact that fruit pulp and the seed are so 93

waxed together in a sticky manner that physical separation was otherwise, practically impossible. 94

95

2.2 Chemicals and reagents 96

Aspartate aminotransferase (AST) , alanine aminotransferase (ALT) , bilirubin , urea and creatinine assay 97

kits were obtained from Randox Laboratories Ltd., Ardmore, Antrim, United Kingdom. Other reagents and 98

chemicals were of analytical grade obtained from Sigma-Aldrich Company Ltd (USA). 99


2.3 Experimental Animals 100

Male albino rats weighing 150-200g (7-8weeks old) were obtained from the animal house of the National 101

Research Institute for Chemical Technology (NARICT), Basawa, Zaria, Nigeria. They were acclimatized in 102

a well ventilated room within the animal facility of the Department of Biochemistry, Ahmadu Bello 103

University Zaria for two weeks before the commencement of the study. They were allowed free access to 104

rat feeds (obtained from ECWA feed Ltd, Bukuru, Jos, Nigeria) and tap water ad libitum through the 105

course of experiment. Animals were weighed and randomly assigned to each of 17 treatment groups 106

(n=5). Permission was obtained from the University’s Ethical Committee for laboratory use of the animals. 107

2.4 Animal treatments 108

The study was carried out in two phases. The first phase involved the assessment of preventive potential 109

of the extracts of different parts of Tamarindus indica in acute liver injury model. In this acute 110

experimental model, the effects of the extracts were investigated in rats first by pre-treatment 111

intraperitoneally with 10mg/kg body weight of the extract for two days followed by intoxication with carbon 112

tetrachloride (CCl4) at 0.6ml/kg on the third day. This dose was selected because the study considered 113

that to recommend any new compound as commercially and clinically useful as hepatoprotective agent, it 114

must have be effective at lower or comparable doses to the antioxidant supplements that are currently 115

available. Besides, previous sub-acute toxicity studies reveal safety of dose up to 500mg/kg weight, in 116

addition to the fact that most part of the plants are consumed either as food or as traditional medicinal 117

preparations. 118

In the second phase, the possible therapeutic potential of the extract in chronic liver injury was evaluated. 119

This was carried out by repeated intoxication of rats with carbon tetrachloride (0.3ml/Kg) at every 72 120

hourly intervals with concomitant daily administration of the extracts (5mg/Kg) for twelve days, following 121

initial CCl4 intoxication at 0.6ml/kg. In all cases control groups treated with vitamin E alone, vitamin E + 122

CCl4, CCl4 only or solvent alone were also included [2,4,5,20]. 123

For each phase, treatment for each plant part extract was divided into the following groups: solvent only 124

(corn oil); vitamin E only, Vitamin E + CCl4; extract only, extract + CCl4 and CCl4 only groups with 5 rats 125

in each group. An untreated control group was also included. For the two acute intoxication, vitamin E as 126

an oil gel was administered at a dose of 50mg/kg, while for low level chronic CCl4 intoxication, vitamin E 127

was administered at 10mg/kg, similar to the dose used in human subjects.These groupings were 128

necessary to establish the preventive or therapeutic effect of the plant extracts on the organ damage 129

caused by CCl4 intoxication. 130

2.5 Animal sacrifice and tissue collection 131


Twenty four hours (24hrs) after the last treatment, all rats were sacrificed to permit maximum collection of 132

blood that allowed triplicate analysis of all parameters for a statistically valid assessment. The sacrifice 133

was performed under mild chloroform anaesthesia.Blood was collected directly at sacrifice; serum was 134

separated after coagulating and centrifuging at 3000 rpm for 15 min and then stored at -20oC until 135

required for analysis. 136

2.5.1 Determination of Aspartate aminotransferase (AST) and Alanine aminotransferase (ALT) 137

Aspartate aminotransferase (AST) and alanine aminotransferase (ALT), were determined independently 138

as described by Akramet al. [1], using assay kits (Randox Laboratories Ltd). Exactly, 0.5µl of reagent 1 139

(which is made up of 100 mM phosphate buffer pH 7.4, 100mM L-aspartate or 200 mML-alanine and 140

2mM α-oxoglutarate) was measured into a clean test tube containing 0.1ml of serum, mixed and 141

incubated for 30 minutes at 37oC. Exactly, 0.5ml of reagent 2 (made up of 2 mM 2,4-142

dinitrophenylhydrazine) was added, mixed and allowed to stand for 20 minutes at 25oC before 0.5ml of 143

sodium hydroxide (0.4M) was added, mixed and absorbance was read against the reagent blank at 144

540nm after 5 minutes. The AST and ALT activities were determined from the standard calibration curve 145

prepared using the kits pre-validation data as provided by the manufacturers. 146

2.5.2 Determination of Total, Conjugated and Unconjugated Bilirubin 147

Conjugated (indirect), unconjugated (direct) and total bilirubins were estimated as described by Sheirine 148

and Safinaz [22] using assay kits (Randox Laboratories Ltd). These determinations were carried out 149

based on the principles that bilirubin reacts with diazotised sulphanilic acid to form a blue coloured 150

complex. Hence direct bilirubin was determined by its reaction with diazotised sulphanilic acid while total 151

bilirubin is determined in the presence of caffeine, which aids release of albumin bound bilirubin. For total 152

bilirubin (mg/dl), the absorbance of the sample against sample blank (ATB) was read at 560 nm and 153

multiplied by a factor of 10.8, while for direct bilirubin the absorbance was read against the sample blank 154

(ADB) at 530 nm and multiplied by a factor of 14.4. Indirect bilirubin was obtained by difference between 155

the total and direct bilirubin. 156

2.5.3 Determination of urea 157

Urea was analysed based on the principle that in the presence of urease, urea in the serum is hydrolysed 158

to ammonia which is trapped by Berthelot’s reaction using analytical kits (Randox Laboratories), and 159

measured spectrophotometrically as described by Stephen et al. [23]. In the method, 100µl of reagent 1 160

(which is made up of 116 mM EDTA, 6mM sodium nitroprusside and 1g/l urease) was added into a clean 161

test tube containing 10µl of serum. They were mixed and incubated at 37oc for 10 minutes. Then, 2.50ml 162

of reagent 2 (made up of 120 mM phenol) and 2.50ml of reagent 3 (made up of 27mM sodium 163

hypochlorite and 0.14N sodium hydroxide) were added, mixed immediately and incubated at 37oc for 15 164


minutes before absorbance was taken at 540nm. Urea concentration was then calculated by simple 165

proportion using the absorbance of a known concentration of the standard. 166

2.5.4 Determination of Creatinine 167

Creatinine level was assayed using assay kits as described by Stephen et al. [23]. The analysis is based 168

on the principle that creatinine in alkaline solution reacts with picric acid to form a coloured complex 169

whose intensity is directly proportional to the creatinine concentration. In the method used, 0.2ml of 170

serum was added into a clean test tube containing 2.0 ml of creatinine working reagent (made up of picric 171

acid and sodium hydroxide). The mixture was shaken and the absorbance (A1) at 510nm was read after 172

30 seconds and at exactly 2 minutes later (A2). The creatinine concentration (mg/dL) was calculated by 173

simple proportion from the standard using the difference between A1 andA2 in both cases. 174

2.6 Statistical analysis 175

Results obtained were expressed as mean ± SD. All analyses performed were in triplicates and data 176

analyzed by analysis of variance (ANOVA) using the Statistical Package for Social Sciences (SPSS) 177

version 14 software with the confidence level set at 95% (p<0.05). 178

3. RESULTSAND DISCUSSION 179

3.1 Results 180

Figures 1-7 compares the effects of the extracts on the levels of some serum biochemical markers in rat’s 181

serum following two day pre-treatment with extracts (5mg/kg) while Figures 8-14 compare the levels of 182

biochemical parameters following twelve days therapeutic trials on chronic liver injury model. 183

In the acute model experiment, the activities of AST and ALT in the CCl4 only-treated control were 184

statistically elevated (p<0.05) above the untreated control and the extract pre-treated groups. Similarly, 185

statistically significant lowering (p<0.05) of AST and ALT levels were observed in the vitamin E - treated 186

control as compared to the CCl4 control (Figures 1& 2). There was however no statistical significant 187

change in the activity of these enzymes in the extract treated groups as compared to the solvent-treated 188

group and the untreated control group (Figures 1&2). 189

On the other hand, in the chronic model experiment, the levels of AST and ALT in the CCl4 only group 190

were significantly (p<0.05) elevated above the levels in the untreated control group. No such statistical 191

(p>0.05) differences exist when the untreated control group is compared with the extract-treated groups 192

except groups treated with the leaves and root extracts (Figures 8 &9). However, in most of the groups 193

intoxicated with CCl4 but administered methanolic extracts of different parts of Tamarindus indica, there 194

was a significant (p<0.05) decrease in the levels of the liver marker enzymes to levels that were not 195

significantly (p>0.05) different from that of the untreated control, but in some cases, even significantly 196


(p<0.05) lower than the untreated control, especially, in the case of stem and pulp extracts treated groups 197

(Figure 9). Similarly, the liver enzyme levels in the solvent and vitamin E - treated control showed no 198

significant (p>0.05) difference with the untreated control group. In general, the fruit pulps extract (Figures 199

1 & 8) exhibited the best lowering effect in the levels of AST and ALT, while the leaves extract (Figures 200

2& 9) lowered the AST and ALT levels to the least extent. 201

The levels of total, conjugated and unconjugated bilirubin were significantly (p<0.05) elevated in the 202

CCl4control group above the untreated group in the acute model experiment (Figures 3, 4&5). The 203

reverse is however the case when the various extract or vitamin pre-treated groups were compared with 204

the untreated control group (Figures 3&5), where the bilirubin levels were not significantly elevated 205

(p>0.05).The observed bilirubin lowering effect in the acute model experiment was also observed in the 206

experimental model of chronic liver injury (Figure 10, 11&12). However, the decrease caused by the 207

extracts in the levels of unconjugated bilirubin in the chronic injury model experiment was much more 208

pronounced (Figure 12), but the vitamin E treated control group showed the best bilirubin lowering 209

potential, followed closely by the fruit pulp (Figure 10) and stem bark (Figure 12) extract treated groups, 210

while the leaves treated group (Figures 10,11&12) showed the least potential. 211

The kidney function indicators, urea and creatinine also showed varied responses to the different 212

treatment across the groups in the acute model experiments. There were elevations in levels of urea in 213

the CCl4 only treated group when compared to the untreated control (Figures 6&7). However, in the 214

extract pre-treated groups, there were no such significant (p>0.05) elevation when compared with the 215

untreated control, except in the case of the leaves and root extract treated groups which showed 216

significant (p<0.05) elevation in the level of urea (Figure 6). 217

In the chronic injury model experiment, the CCl4 only control group showed statistically (p<0.05) elevated 218

levels of urea and creatinine when compared to the untreated control (Figures 13&14). However, no such 219

elevations existed when the CCl4 intoxicated group treated with extracts were compared with the 220

untreated or solvent-treated group. Groups treated with only stem (Figure 14), seed (Figure 13), pulp 221

(Figure 14) or fruit bark (Figure 13) extracts were particularly and significantly (p<0.05) lower than the 222

untreated control and the vitamin E-treated group. 223

224

4.0DISCUSSION 225

The results obtained generally showed a statistically significant (p<0.05) lowering in the levels of 226

aspartate aminotransferase, alanine aminotransferase, bilirubin, urea and creatinine in the extracts-227

treated groups as compared to the CCl4 control, both in the ameliorative (chronic) and in the preventive 228

(acute) model experiments. In contrast, no statistical (p>0.05) difference were observed in most instances 229

between the extract-treated groups and the vitamin E-treated group or the untreated control group 230


(Figures 3-7, 12-14), suggesting that the extracts were able to counteract the hepatotoxic effects of 231

CCl4by restoring the functional integrity of the liver and the kidney. 232

This is because cellular damages are usually accompanied with leakage of intracellular enzymes into the 233

blood, and as such the levels of these enzymes in the serum can be measured as indicators of cell 234

damage [22]. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) are among such 235

enzymes. Alanine aminotransferase is found in the liver and is markedly elevated in hepatitis and other 236

liver diseases. Oxidants like free radicals are capable of causing peroxidative degradation of cellular 237

membranes and endoplasmic reticulum which are rich in polyunsaturated fatty acids. Hence, oxidative 238

damage is implicated in most disease processes such as cardiovascular disease, cancer, inflammatory 239

conditions, asthma, liver diseases and muscular degeneration [6] through destruction of the hepatic 240

cellular membrane by altering the cellular permeability of hepatocytes leading to elevated levels of serum 241

biochemical parameters like ALT, AST and bilirubin. Similarly these radicals can cause oxidative damage 242

of the kidney leading to elevated levels of urea and creatinine [3]. Hence many workers consider the 243

reverse of this phenomenon as indices of hepatoprotective and nephroprotective activities [2,3]. 244

245

Thus, the significant (p<0.05) elevation in the levels of ALT and AST ( Figures 1,2,8 & 9)observed in the 246

CCl4 control when compared to the untreated group can be attributed to cellular damages caused by CCl4 247

intoxication. These elevations were clearly more evident in the chronic model experiment than it was in 248

the acute model experiment, probably because of the repeated long term CCl4 intoxication in the latter. In 249

the acute model experiment, the extract-treated and the vitamin-E treated groups showed significant 250

decrease compared to the CCl4 control, indicating the protective effects of the extracts and vitamin E 251

against CCl4-induced organ damages (Figures 1 & 2). Similarly, the reversal and maintenance of lowered 252

AST and ALT levels even after chronic intoxication with CCl4 in the chronic model experiment suggests 253

strong hepatoprotective andtherapeutic effects of the extracts. Comparatively the effects exhibited by the 254

stem, pulp, fruit bark and seeds extracts were more pronounced than those shown by the root and leaves 255

(Figures 1,2, 7 & 8). 256

Bilirubin is excreted by the liver, and hence any interference with the normal liver functions affects its rate 257

of excretion. Thus, elevated levels of bilirubin is used as an index of liver function as high levels of 258

unconjugated bilirubin suggests liver malfunction or there is excessive breakdown of haemoglobin [8,25]. 259

The results obtained showed a clearly significant (p<0.05) increase in the levels of total and unconjugated 260

bilirubin in CCl4 treated animals, which are markedly reduced by pre-treatment (Figures 3, 4 & 5) or 261

concomitant treatment (Figures 10, 11&12) with the methanolic extract of the stem, pulp, fruit bark or 262

seed. Although this was true and consistent for both the acute and preventive model experiments, it can 263

be saliently observed that the effect on the unconjugated bilirubin in the chronic (curative) model 264

experimentwas more pronounced (Figure 10). This appears to demonstrate the abilities of the extracts to 265

restore and maintain liver functions even under chronic toxicological condition. In comparative terms, the 266


stem bark, fruit pulp and fruit bark extracts exhibited the best bilirubin lowering potential and hence 267

possibly better hepatoprotective and curative capacity. 268

The kidney is a target organ for xenobiotics metabolism, second only to the liver, and hence substances 269

like carbon tetrachloride that are toxic to the liver are also known to be potentially toxic to the kidney 270

[8,23], whose principal function is the excretion and elimination of wastes like urea and creatinine. Thus, 271

impairment of the kidney function leads to decreased clearance rate of urea and creatinine which 272

conversely results in their elevated levels within the serum [2, 3, 4, 5, 20]. The study revealed a 273

significantly (p<0.05) elevated levelsof urea and creatinine in the CCl4 groups which were significantly 274

(p<0.05) reduced by pre-treatment or administration of either stem, fruit pulp or fruit bark extracts strongly 275

suggesting their nephroprotective abilities (Figures 6 & 7). Interestingly, these significant (p<0.05) 276

decreases in urea and creatinine levels in the extracts-treated groups were more clearly observed in the 277

chronic experimental model (Figures 13 & 14).In the acute experimental model, the urea and creatinine 278

levels were markedly decreased only in the stem bark, fruit pulp and fruit bark extracts treated groups 279

(Figures 6 & 7), suggesting better that these parts contained substances with better ameliorative 280

capacities, while in the chronic experimental model, all the extracts except the leaves demonstrated good 281

capacity for protection and amelioration of kidney damage (Figures 13 & 14). 282

Researches into plant foods and plants with medicinal properties have received increased interest in 283

recent years. For instance, various reports have indicated that many plant foods and medicinal plants 284

including Vernoniaamygdalina [3], Labisiapumila [19], Spirulina maxima [13], Hibiscus esculentus [6], 285

Moringaoleifera [7], Anogeissusleiocarpus [8]are known to contain antioxidant constituents with 286

demonstrated capacity to chemoprevent oxidative stress-related diseases. Other workers have shown 287

that under in vitro conditions seeds of T. indicaexhibited antioxidant effect [24]. The present findings have 288

thus proven that Tamarindus indica has potent capacity to prevent and ameliorate oxidative damage to 289

the liver and kidney. 290

The organ protective effect of Tamarindus indica may be related to the presence of antioxidant 291

compounds in the different methanolic extracts of the plant parts. For instance, polar solvent extracts of 292

tamarind seeds and pulp revealed the presence of compounds with possible antioxidant potential and 293

demonstrable anti-microbial, anti-diabetic and anti-cancer potential[4,9,16,18,21]. Methanolic extracts of 294

plants have been reported to have phenolics, which have several biological activities and health benefits, 295

including antioxidant roles as a major constituent [16]. Similarly, the presence of polyphenols was 296

reported in Tamarindus indica fruits [15,21]with a profile dominated by proanthocyanidins in various 297

forms, catechin, procyanidin B2, epicatechin, procyanidintrimer, procyanidin tetramer, 298

procyanidinpentamer, procyanidinhexamer and other compounds in less quantity [16, 24]. Hence, the 299

organ protective and therapeutic effects exhibited by different parts of Tamarindus indica may be ascribed 300

to these phytoconstituents. 301


Considered together, the reversal of elevated levels of AST, ALT, bilirubin, urea and creatinine by the 302

extracts of various parts of Tamarindus indica clearly illustrates the organ-protective and therapeutic 303

potential of different parts of the plant. It is worth noting however, that the stem and fruit extracts showed 304

strongest organ protective activities against CCl4 induced damage while the roots and leaves showed the 305

least potency. This observation appears to justify the use of the stem, seed and fruit to treat jaundice boils 306

and stroke respectively in traditional medicine, while the low activity of the roots and leaves might explain 307

the reason for their poor usage in traditional medical practice[15]. 308

4. CONCLUSION 309

In conclusion amongst the various parts, the fruit pulp, stem bark, fruit bark and seeds extracts had the 310

most effective hepatoprotective and nephroprotective potentials while the roots and the leaves are the 311

least respectively.The findings suggest that Tamarindus indica, a widely available plant resource in 312

tropical Africa and Asia could serve as a cheap source of naturally occurring drugs and antioxidants for 313

pharmaceutical and nutraceutical industries, but the exact bioactive substance responsible for the 314

observed activity as well as the mechanism of the biological protection and therapeutic capacities would 315

require further elaboration. 316

COMPETING INTERESTS 317

We declare that no competing interests exist. 318

AUTHORS’ CONTRIBUTIONS 319

S.E. Atawodi designed the study, wrote the protocol and interpreted the datawhileM.L.Liman managed 320

the literature searches, managed the laboratory analyses of the study and wrote the first draft of the 321

manuscript. All authors read and approved the final manuscript. 322

ETHICAL APPROVAL 323

All authors hereby declare that "Principles of laboratory animal care" (NIH publication No. 85-23, revised 324

1985) were followed, as well as specific national laws where applicable. All experiments have been 325

examined and approved by the appropriate ethics committee of the University 326

327

ACKNOWLEDGEMENTS 328

We gratefully acknowledge Alexander Von Humboldt Foundation of Germany for material donation in 329

support of Professor Atawodi’s Research. 330

331

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399

400

Figure 1: Mean aspartate aminotransferase (AST) activity in serum of rats intoxicated with carbon 401

tetrachloride (0.6ml/kg) following three days pre-treatment with methanol extract of Tamarindus indica 402

(10.0 mg/kg). 403

404

Figure 2: Mean alanine aminotransferase (ALT) activity in serum of rats intoxicated with carbon 405

tetrachloride (0.6ml/kg) following three days pre-treatment with methanol extract of Tamarindus indica 406

(10.0 mg/kg). 407

408

0

10

20

30

40

50

60A

ST

act

ivit

y

U/)

Treatment

0

5

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25

30

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50

ALT

a

ctiv

ity

U/I

Treatment


- 409

Figure 3: Mean total bilirubin in serum of rats intoxicated with carbon tetrachloride (0.6ml/kg) following 410

three days pre-treatment with methanol extract of Tamarindus indica (10.0 mg/kg). 411

412

413

Figure 4: Mean conjugated bilirubin in serum of rats intoxicated with carbon tetrachloride (0.6ml/kg) 414

following three days pre-treatment with methanol extract of Tamarindus indica (10.0mg/kg). 415

416

0

0.2

0.4

0.6

0.8

1

1.2

1.4

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To

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bil

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bin

mg

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Treatment

0

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0.1

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0.2

0.25

0.3

0.35

0.4

0.45

Co

nju

ga

ted

bil

iru

bin

mg

/dl

Treatment


417

Figure 5: Mean unconjugated bilirubin in serum of rats intoxicated with carbon tetrachloride (0.6ml/kg) 418

following three days pre-treatment with methanol extract of Tamarindus indica (10.0 mg/kg). 419

420

421

Figure 6: Mean urea concentration in serum of rats intoxicated with carbon tetrachloride (0.6ml/kg) 422

following three days pre-treatment with methanol extract of Tamarindus indica (10.0 mg/kg). 423

424

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

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0

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Ure

a C

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cen

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n m

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425

Figure 7: Mean creatinine concentration in serum of rats intoxicated with carbon tetrachloride (0.6ml/kg) 426

following three days pre-treatment with methanol extract of Tamarindus indica (10.0 mg/kg) 427

428

429

Figure 8:Meanaspartate aminotransferase (AST) activity in serum of ratsfollowing daily intraperitoneal 430

administration of Tamarindus indicaextract (5mg/kg) with 72 hourly injection of carbon tetrachloride 431

(0.3ml/kg) for 12 days.432

0

0.5

1

1.5

2

2.5

Cre

ati

nin

e c

on

cen

tra

tio

n m

g/d

l

Treatment

0

20

40

60

80

100

120

AS

T a

ctiv

ity

U

/)

Treatment

17

433

Figure 9: Mean alanine aminotransferase (ALT) activity in serum of ratsfollowing daily intraperitoneal 434

administration of Tamarindus indicaextract (5mg/kg) with 72 hourly injection of carbon tetrachloride 435

(0.3ml/kg) for 12 days. 436

437

438

Figure 10: Mean total bilirubin in serum of ratsfollowing daily intraperitoneal administration 439

ofTamarindusindicaextract (5mg/kg) with 72 hourly injection of carbon tetrachloride (0.3ml/kg) for 12 days. 440

0

10

20

30

40

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60

70

ALT

a

ctiv

ity

U/I

Treatment

0

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1.8

2

To

tal

bil

iru

bin

mg

/dl

Treatment

18

441

442

Figure 11:Mean conjugated bilirubin in serum of ratsfollowing daily intraperitoneal administration of 443

Tamarindus indicaextract (5mg/kg) with 72 hourly injection of carbon tetrachloride (0.3ml/kg) for 12 days. 444

445

446

447

Figure 12: Mean unconjugated bilirubin in serum of ratsfollowing daily intraperitoneal administration of 448


450

0

0.05

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0.15

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Treatment

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Un

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ub

in m

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l

Treatment

19

451

Figure 13: Mean urea concentration in serum of ratsfollowing daily intraperitoneal administration of 452


454

455

456

Figure 14: Mean creatinine concentration in serum of ratsfollowing daily intraperitoneal administration of 457


0

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Ure

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n m

g/d

l

Treatment

Hepatoprotective and Nephroprotective Effects of ...€¦ · 2 Methanolic Extracts of Different...

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