International Journal of Scientific Research in
Environmental Science and Toxicology

Cocos nucifera (Coconut) water is one of the world natural products with increasing scientific evidence that support its role in health and medicinal application. This study was undertaken to investigate the therapeutic potential of Cocos nucifera water on cadmium-induced toxicity in male rat. Sixty-four animals were randomly grouped into five, groups 3, 4, 5 (n=16) were exposed to 100, 200, and 300 ppm cadmium doses for 6 weeks in their drinking water as cadmium chloride. At the end of 6 weeks, eight from each group were sacrificed and the remaining was administered with 10 % coconut water for a week. Control animals (n=8; group 1) received distilled water and Cocos nucifera water (n=8; group 2) for the same six week period. Lipid profile analysis revealed that cadmiuminduction exhibited different dyslipidemia patterns. Significant dose-dependent hypocholesterolemia, hypotriacylglyceridemia and hypophospholipidemia characterized the effect of cadmium exposure at all doses while Cocos nucifera water treatment for a week reverse these effects by 4, 24 and 22 % in plasma and 16, 15 and 2 % in the erythrocytes respectively. The exposure to cadmium resulted in reduced cholesterol, triacylglyceride concentration in the brain and cholesterol, phospholipids concentrations in the hepatic compartments. Also, the exposure caused an increase in brain phospholipids and hepatic triacylglycerides concentrations. The administration of Cocos nucifera water significantly (p < 0.05) reverses these effects. Our findings indicate that although the cadmium-exposure at different doses resulted in down and upregulation of lipid profile, the administration of Cocos nucifera water was able to reverse the damage caused by cadmium-induced toxicity.

Keywords: Cadmium; Cocos nucifera water; Lipid profile; Brain; Therapeutic; Dyslipidaemia

The Cocos nucifera (Coconut) plant has long been recognized as a valuable source of various commodities for human life. The water of the endosperm, known as coconut water, is the most nutritious, wholesome beverage that the nature has provided for the people to fight the sultry heat and other therapeutic purposes [1]. In recent years, coconut water has also assumed great nutritional significance owing to its physio-chemical nature. It satisfies thirst and as well invigorates the body and brings about a sense of wellbeing and renewed health [2]. Coconut water has been used as an oral rehydration in patients with diarrhoea to replace fluid loss from the gastrointestinal tract, as a home glucose electrolyte solution for well-nourished children with mild diarrhoea and in an extreme situation such as short-term intravenous hydration fluid in a patient [3, 4].

Research has shown that coconut water contains a variety of nutrients like vitamins, antioxidants, amino acids, enzymes and growth factors. Also, as a good source of the major minerals like calcium, magnesium, and potassium and some trace elements like zinc, selenium, iodine, manganese, boron, and molybdenum [5]. Its nutritive properties are widely recognized to be an electrolyte, antidote, anti-oxidant, cardio-protective, hepatoprotective, anti-thrombotic, anticancer, anti-cholecystitic, anti-bacterial, anti-caries, antiprotozoal, antiviral, anti-diabetic, immune-stimulatory and anti-atherosclerosis activities [6]. Some of the diseases which the coconut ameliorates or reverse might have been generated from the accumulation of heavy metals.

Heavy metals are also considered as trace elements because of their presence in trace concentrations (Parts per Billion (ppb) ranges to less than 10ppm) in various environmental matrices. Cadmium (Cd) is one of the most toxic heavy metals present in the environment and has been involved in poisoning cases of human and animal population, thus becoming a serious threat to living organisms [7]. Some regulations and recommendations doses for cadmium include 0.04 mg/L in drinking water (Environmental Protection Agency), 0.005mg/L in bottled water (Food and Drug Administration). These regulations and recommendations can be expressed as “not-to-exceed” levels [8]. The exposure to cadmium dose that is higher than recommended doses causes various forms of diseases such as osteomalacia and osteoporosis, hypertension, arteriosclerosis, anaemia and cancer [9, 10]; inhibit the activities of various pancreatic proteases [11] and antioxidant enzymes which release free oxygen radicals [10].

Cadmium stimulates the formation of metallothioneins (a family of low molecular weight metal binding proteins unique in their high cysteine content) and reactive oxygen species, thus causing oxidative damage resulting in loss of membrane functions, causing dyslipidemia [12, 13]. Earlier research conducted in our laboratory reported the changes in the plasma, erythrocyte, brain and hepatic cell lipid profile of rat after low-level exposure to cadmium through drinking water [13]. The therapeutic benefits of plant foods have been the focus of many extensive dietary studies. This prompted the present study to continue our efforts to investigate the therapeutic effect of coconut water on low-level cadmium-induced toxicity in the rat.

Cadmium chloride was procured from Sigma-Aldrich, Missouri, USA. All other chemicals used in this study were of the purest grade available and were obtained from the British Drug House (BDH) Chemicals Limited, Poole, Dodset, England

Matured peeled coconut was purchased from the Badagry Central market, Badagry Local Government, Lagos, Nigeria, botanically authenticated by the Department of Botany, Lagos State University, Ojo-Lagos. They were cut open at the ‘eye’ of the coconut to extract the water every day of the research. 10% of the collected coconut water was then administered to the animals daily.

Sixty-four (64) male Wistar albino rats with a mean weight of 190 g bred in the Animal House of the Department of Biochemistry; Faculty of Sciences; Lagos State University, Ojo- Lagos, Nigeria was used for the experiment. They were housed in animal stainless cages with a 12 h light / dark cycle and allow free access to food and water for 14 days prior to the experiment.

They were randomly and equally distributed into eight groups with eight rats in each group. The cadmium doses were selected based on the research of Yapping et al [14]. Distilled water was administered to the control group because its demineralized water and coconut water which consists of minerals was given to the positive control animals.

Group 1: Given distilled water for 6 weeks (Control).

Group 2: Given 10 % coconut water and feed for 6 weeks (Positive control).

Group 3a: Administered 100 ppm Cd as cadmium chloride for 6 weeks.

Group 3b: Administered 100 ppm Cd as cadmium chloride

for 6 weeks and 10 % coconut water for 1 week.

Group 4a: Administered 200 ppm Cd as cadmium chloride for 6 weeks.

Group 4b: Administered 200 ppm Cd as cadmium chloride for 6 weeks and 10 % coconut water for 1 week.

Group 5a: Administered 300 ppm Cd as cadmium chloride for 6 weeks.

Group 5b: Administered 300 pm Cd as cadmium chloride for 6 weeks and 10 % coconut water for 1 week.

All groups were fed ad libitum with grower mash (product of Animal care, Lagos, Nigeria). At the end of the treatment period, the rats were weighed and sacrificed under light ether anesthesia. The blood was collected, centrifuge into plasma was collected while the brain was quickly excised, weighed and placed on ice until required for biochemical analysis. All experiments on the rats were approved by the Animal Ethical Committee of the Department of Biochemistry, Lagos State University, Ojo-Lagos, Nigeria, in compliance with the Ethical guide for the care and use of laboratory animals (NIH Publication No. 85-93, revised 1985) [15].

Plasma concentrations of total cholesterol and triglycerides were determined with commercial kits (Spin React S.A., Santa Colona, Sant Esteve De Bas, and Spain). Phospholipids in plasma were extracted with chloroform–methanol mixture (2:1, v/v) as described by Folch et al [16] and the extraction of lipids from erythrocytes was done using chloroform-isopropanol 7:11 v/v method [17]. The extracts were stored at -20°C for further analysis. Determination of lipids was performed as described by Stewart [18]. Briefly, 100 μl of lipid extracts as well as 100 μl of chloroform were evaporated to dryness over a hot water bath at 60oC and allowed to cool. 2.0 ml chloroform and 2.0 ml of ammonium ferrothiocyanate were added, vortexed and allow the phases to separate. The chloroform layer was removed and the absorbance read at 488nm against blank using UV/ Visible Spectrophotometer (Model SM 755s), a product of Surgien field Instrument, Zhejiang, China (Mainland).

Lipids were extracted from the brain and liver as described by Folch et al [16]. 10 % brain and liver homogenate was prepared with chloroform-methanol mixture (2:1, v/v), washed with 0.05M KCl solution. An aliquot of the chloroform-methanol extracts was then used for the determination of cholesterol, triglyceride and phospholipid concentrations. Cholesterol was determined as described by Allain et al [19], triacylglyceride concentrations determined to follow the procedure described by Kriketos et al [20] while determination of phospholipids followed the same procedure as described for plasma.

Data are expressed as mean ± S.E.M. One way analysis of variance (ANOVA) followed by Tukey’s Test was used to analyze the results with p < 0.05 considered significant.

Summaries of cadmium exposure and administration of Cocos nucifera water on the lipid profile of plasma, erythrocytes brain and hepatic compartments of the animals are depicted [Figures 1-4]. Cadmium exposure resulted in a significant (p< 0.05) dose – dependent decrease in cholesterol, triacylglyceride and phospholipids concentrations of plasma and erythrocytes

[Figures.1 and 2] whereas administration of Cocos nucifera water reversed these effect by 50, 1 and 3% in plasma and 11, 12 and 25% in the erythrocyte compared to control respectively. In the brain [Figure. 3], cadmium exposure resulted in significant (p < 0.05) dose-dependent decrease in cholesterol (33%), triacylglyceride (34%) and increase in phospholipid (25%) concentrations while the administration of Cocos nucifera water reverses these effects. In fig. 4 the exposure of cadmium resulted in a significant (p < 0.05) decrease in cholesterol (35%), phospholipid (50%) contents and 25% increase in triacylglyceride content. Whereas, administration of Cocos nucifera water lead to decrease in

cholesterol, phospholipid and increase in triacylcglyceride concentration.

Lipids are used for energy storage to form cell membranes, precursors for hormones, have a prominent role as messengers and regulators of inflammation and other cellular functions [21]. Lipid profiles have been shown to be the important predictors for the metabolic disturbances, including dyslipidemia, hypertension, diabetes, cardiovascular disease and liver dysfunction [22]. Evidence abounds that cadmium exposure alters the lipid profile [13, 23], as observed in this study [Figures 1-4]. The hypolipidemia observed in this study might be due to genetic mutations causing an underproduction or increased clearance of low density lipoproteins by cadmium toxicity and result in lipid levels low enough to cause significant consequences [24]. The defect lipid metabolism might lead to structural and functional central nervous system diseases, by affecting different metabolic pathways such as (1) cholesterol biosynthesis, (2) lipid transport and lipoprotein assembly, (3) receptors that mediate the cellular uptake of lipids, and (4) signaling molecules [25]. The mechanisms underlying the accumulation of liver triacyglycerides include increased liver fatty acid mobilization and delivery to the liver, coupled with increased hepatic lipogenesis and decreased secretion of very low density lipoprotein [26].

The administration of fresh coconut water after induction of Cd showed a significant reversal therapeutic potential on the Cdinduced lipid toxicity. Coconut liquid endosperm called coconut water, with its many applications, is one of the world’s most versatile natural products that have multifarious utility. The observed medicinal effect of coconut water may be associated with the presence of bioactive components such as free amino acid L-arginine, vitamins like water-soluble B-vitamins and ascorbic acid, inorganic ions, such as calcium, magnesium and potassium [27, 28]. L-arginine functions as an intermediate in the urea cycle, which takes place mainly in hepatocytes in mitochondria and continues in the cytosol. Different type of metabolic and physiological conditions, triggered by stress may result in partial deficiencies of enzyme production in the urea cycle. These disturbances can be reducing the activity of the enzymes, which disrupts the urea cycle and prevents the body from processing nitrogen effectively. In particular, deficiency of argininosuccinate synthase can trigger the development of dyslipidemia and fatty liver [29] as observed in this study.

The therapeutic explanation could be that the inorganic ions present in the coconut water are required for the normal cellular function and are critical for bone formation, hemoglobin function, gene expression, and enzyme activation of lipid biosynthesis [27, 30-32]. The concentration of inorganic ions known as electrolytes in coconut water generates an osmotic pressure similar to that observed in blood [27]. As a result, coconut water can be used as a short term intravenous hydration fluid under certain emergency situations. Therefore, the basic ion composition of coconut water can replenish the cadmium-induced toxicity, due to the water-soluble B vitamins and minerals which are required as coenzymes for enzymatic reactions essential for cellular function such as prevent mitochondrial toxicity and the transamination and decarboxylation reactions (vitamins B6) [33-35]

Thiamine (Vitamin B1) is a necessary step in the synthesis of fatty acids, steroids, and the aromatic amino acid precursors to a range of neurotransmitters and other bioactive compounds essential for brain function [36], and a cofactor during metabolic processes and contributes to the structure and function of cellular membranes [37]. The flavoproteins (B2) are also co-factors in the metabolism of essential fatty acids in brain lipids [38]. A vast array of processes and enzymes involved in every aspect of peripheral and brain cell function are dependent on vitamin B3 known as niacin and anti-atherogenic lipolysis in adipose tissue [39].

Pantothenic acid (vitamin B5) is a substrate for the synthesis of the ubiquitous coenzyme A which contributes to the structure and function of brain cells via its involvement in the synthesis of cholesterol, amino acids, phospholipids, and fatty acids [40]. In addition to vitamin B, coconut water also contains vitamin C (ascorbic acid), which is an important dietary antioxidant [33]. Previous studies showed that coconut water showed the free radical scavenging ability that may be partly attributed to the ascorbic acid [41].

The present study reveals that fresh matured coconut water has a therapeutic potential on lipid profile in cadmium exposed male rat.

We declare that there is no conflict of interest.

The authors appreciate the technical staff of the Cell and Tissue Culture Research Laboratory (Drug Discovery Unit); Department of Biochemistry; Lagos State University; Ojo; Lagos; Badagry Expressway; Lagos - Nigeria.

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