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Year : 2011  |  Volume : 1  |  Issue : 2  |  Page : 174-178

Impact of morin (a bioflavonoid) on ammonium chloride-mediated oxidative damage in rat kidney

Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalainagar, Tamil Nadu, India

Date of Submission01-Jan-2011
Date of Acceptance02-Apr-2011
Date of Web Publication23-Aug-2011

Correspondence Address:
Perumal Subramanian
Department of Biochemistry and Biotechnology, Faculty of Science, Annamalai University, Annamalainagar - 608 002, Tamil Nadu
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Source of Support: Indian Council of Medical Research (ICMR), New Delhi., Conflict of Interest: None

DOI: 10.4103/2231-0738.84210

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Introduction : Hyperammonemia is a major contributing factor to neurological abnormalities observed in hepatic encephalopathy and in congenital defects of ammonia detoxification. Ammonia toxicity results in free radical generation that leads to oxidative stress and tissue damage. Morin is a bioflavonoid, a constituent of many herbs and fruits that are used as herbal medicines and also several biological activities. Our aim is to investigate the effect of morin on blood ammonia and plasma urea as well as kidney lipid peroxidation, and the antioxidant status in ammonium chloride-induced hyperammonemic rats. Materials and Methods: Male albino Wistar rats weighing 180 - 200 g were used for the study. Hyperammonemia was induced by interaperitonial injection of ammonium chloride (100 mg / kg body weight). The rats were treated with morin (30 mg / kg body weight) via oral administration. Administration of morin in hyperammonemic rats reduced the levels of ammonia and urea. The antioxidant property of morin was studied by assessing the activities of thiobarbituric acid reactive substances (TBARS), hydroperoxides (HP), conjugated dienes (CD), and antioxidants superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and reduced glutathione (GSH) in ammonium chloride-treated rats. Results and Conclusions: Kidney oxidative stress was effectively modulated by morin administration. Morin significantly improved the status of kidney antioxidants and decreased the levels of ammonia, urea, TBARS, HP, and CD, as compared to the ammonium chloride-treated group. The study offers evidence for the antihyperammonemic and antioxidant effects of morin against oxidative stress in the kidney, induced by ammonium chloride.

Keywords: Ammonium chloride, antioxidants, kidney, lipid peroxidation, morin

How to cite this article:
Subash S, Subramanian P. Impact of morin (a bioflavonoid) on ammonium chloride-mediated oxidative damage in rat kidney. Int J Nutr Pharmacol Neurol Dis 2011;1:174-8

How to cite this URL:
Subash S, Subramanian P. Impact of morin (a bioflavonoid) on ammonium chloride-mediated oxidative damage in rat kidney. Int J Nutr Pharmacol Neurol Dis [serial online] 2011 [cited 2022 Aug 11];1:174-8. Available from:

   Introduction Top

The neurological complications of hyperammonemia in the central nervous system (CNS) are now receiving more attention. Ammonia is a neurotoxin that has been strongly implicated in the pathogenesis of hepatic encephalopathy. [1] Ammonia has also been a major pathogenetic factor associated with inborn errors of urea cycle, Reye's syndrome, organic acidurias, and disorders of fatty acid oxidation. [2] Ammonia-induced neurotoxicity has been reported to include a dysfunction of multiple neurotransmitter system glutamate-mediated excitotoxicity, electrophysiological disturbances, and defects in brain bioenergetics. [1],[3] In spite of extensive investigations, the precise mechanisms involved in ammonia neurotoxicity are not completely understood.

Oxidative stress is an evolving concept in ammonia neurotoxicity. Its effect on oxidative and nitrosative stress in the CNS has been recently reviewed. [1] Recent studies have reported an increased production of free radicals in cultured astrocytes after treatment with pathophysiological concentrations of ammonia. [4] A concurrent increase of superoxide production and a reduction in the activities of various antioxidant enzymes have been seen in animal models with acute ammonia toxicity. [5] Oxidative stress-mediated lipid peroxidation has also been seen as one of the characteristic features of hyperammonemia. [6],[7],[8]

The greatest disadvantage of the currently available potent conventional or synthetic antihyperammonemic agents / therapies lies in their toxicity and reappearance of symptoms after discontinuation. Furthermore, these drugs can cause serious adverse effects. [9] Hence, the screening and development of therapeutic agents from traditional medicinal plants (active components) for their antihyperammonemic activity is in progress worldwide.

Flavonoids are a group of polyphenolic compounds diverse in chemical structure and characteristics. They are widely distributed in foods of plant origin such as vegetables, fruits, tea, and wine. Scavenging of free radicals seems to play a considerable part in the antioxidant activity of flavonoid compounds. Regular ingestion of flavonoid-containing foods may protect against the adverse effect of ammonia toxicity. Phytopharmaceuticals are gaining importance in allopathic as well as traditional medicine owing to their non-addictive and non-toxic nature. Novel antioxidants may offer an effective and safe means of counteracting some of the problems and bolster the body's defenses against free radicals and hyperammonemia.

Morin (3, 5, 7, 2',4'-pentahydroxyflavone; a yellowish pigment) is a flavonoid constituent of vegetables, berries, and fruits, [10] for example, mulberries and osage orange (Maclura pomifera), as well as in many Chinese herbs, used as herbal medicines. It exhibits various biological activities including antioxidation, cytoprotection, antimutagenesis, antidiabetic, [11],[12] and anti-inflammation activities. [13] It also rescues neurons from cell death in acute injury and reduces neurological deficits caused by ischemic damage to the brain, thereby providing neuroprotection. [14]

Furthermore, morin is a well-known therapeutic agent for a number of diseases. To our knowledge, this report is the first study to investigate the effect of morin on kidney lipid peroxidation and antioxidant status in ammonium chloride (AC)-induced hyperammonemic rats. Therefore, the objective of the present study is to investigate the influence of morin on blood ammonia and plasma urea as well as kidney thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides (LOOH), and conjugated dienes (CD), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and reduced glutathione (GSH) in an animal model of AC-induced hyperammonemia.

   Materials and Methods Top

Experimental animals

All the experiments were carried out with male albino Wistar rats weighing 180 - 200 g, obtained from the Central Animal House, Rajah Muthiah Institute of Health Sciences, Annamalai University, Tamil Nadu, India. They were housed in polypropylene cages (47 cm Χ 34 cm Χ 20 cm) lined with husk, renewed every 24 hours under a 12 : 12 hour light / dark cycle at around 22° C and had free access to tap water and food. The rats were fed on a standard pellet diet (Pranav Agro Industries Ltd., Maharashtra, India). The experiment was carried out according to the guidelines of the Committee for the ( ) Purpose of Control and Supervision of Experiments on Animals (CPCSEA), New Delhi, India, and approved by the Animal Ethical Committee of Annamalai University (Approval no. 424; dated 21 / 03 / 2007).

Drugs and chemicals

Morin, thiobarbituric acid, butylated hydroxy toluene, nitroblue tetrazolium, phenazine methosulfate, and glutathione were purchased from Sigma Chemical Company, St. Louis, MO, USA. Ammonium chloride was purchased from Sisco Research Laboratories, Mumbai, India. All other biochemicals and chemicals used in the study were of analytical grade.

Preparation of morin

Morin was freshly dissolved in a small amount of ethanol and then diluted with physiological saline. [15]

Induction of experimental hyperammonemia

Hyperammonemia was induced in Wistar rats by intraperitoneal injections of ammonium chloride at a dose of 100 mg / kg body weight, thrice a week, for eight consecutive weeks. [8],[16]

Experimental design

In the experiment, a total of 32 rats were used. The rats were divided into four groups of eight rats each. Group I: Rats received physiological saline and were considered as control; Group II: Rats were orally administered with morin (30 mg / kg) using an intragastric tube; [17] Group III: Rats were treated with AC (100 mg / kg; i.p); Group IV: Rats were treated with AC + morin thrice a week, for eight weeks.

At the end of eight weeks, all the animals were killed by decapitation after overnight fasting. Blood samples were collected and analyzed for ammonia [18] and plasma urea by the diacetyl monoxime method. [19] The kidney tissues were excised immediately and rinsed in ice-chilled normal saline. Known weights of the tissues were homogenized in 5.0 ml of 0.1 M Tris-HCl buffer (pH, 7.4). The homogenate was centrifuged and the supernatant was used for the estimation of TBARS, [20] lipid hydroperoxides, [21] CD, [22] SOD, [23] CAT, [24] GPx, [25] and GSH. [26]

Statistical analysis

All data were expressed as mean ± S.D. The statistical significance was evaluated by one-way analysis of variance (ANOVA) using the SPSS version 9.5 (SPSS, Cary, NC, USA) and individual comparison was done by Duncan's multiple range test (DMRT).

   Results Top

[Table 1] shows the levels of blood ammonia and plasma urea as well as kidney TBARS, lipoprotein lipid hydroperoxides (LOOH), and CD of control and experimental animals. Hyperammonemia indicated by ammonia and uric acid and lipid peroxidation indicated by TBARS, LOOH, and CD were significantly higher in AC-administered animals as compared to those of normal control rats. Ammonia, uric acid, TBARS, LOOH, and CD levels were lowered significantly in AC-administered animals treated with morin (group IV).
Table 1: Effect of morin on changes in the levels of blood ammonia and plasma urea, and kidney TBARS, HP, and CD of normal and experimental rats

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[Table 2] shows the activities of SOD, CAT, GPx, and GSH in the kidneys of the control and experimental animals. SOD, CAT, GPx, and GSH activities in the kidneys of AC-administered rats (group III) were significantly lower than those in the control rats (group I). Treatment with morin in AC-administered rats (group IV) significantly elevated SOD, CAT, GPx, and GSH activities, as compared to those animals on AC treatment alone (group III).
Table 2: Effects of morin on changes in the levels of SOD, CAT, GPX, and GSH in kidneys of normal and experimental rats

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   Discussion Top

This study is one of the series of studies showing that chronic hyperammonemia causes an imbalance in the oxidative status of the renal tissue and that the resulting free radicals damage the kidney through a peroxidative mechanism. The elevated levels of ammonia and urea may indicate a hyperammonemic condition in rats treated with ammonium chloride. [6],[16],[17] A reduction in the levels of ammonia and urea during morin treatment shows a significant antihyperammonemic activity of this bioflavonoid. [17] The reduction in their levels thus show the antihyperammonemic nature of this bioflavonoid, along with the antioxidant [13] and neuroprotective efficacy. [14] Furthermore, morin has the ability to normalize the levels of urea and uric acid during ischemic conditions. [27] Our results corroborate these previous findings.

Many studies have shown that oxidative stress and free radical production-mediated lipid peroxidation could be involved in the mechanism of ammonia toxicity. [5],[6],[8],[16] Elevated levels of ammonia in the blood and kidney result in the impairment of renal function. A marked elevation in the concentration of TBARS, LOOH, and CD are observed in the kidney of hyperammonemic rats. Excess ammonia induces nitric oxide synthase, which leads to an enhanced production of nitric oxide and other toxic-free radicals, as well as thiobarbituric acid-positive compounds, in the kidney, and leads to oxidative stress and tissue damage. [5],[7],[28] Administration of morin significantly decreases the levels of TBARS, LOOH, and CD in the kidneys of group IV rats. Morin can reduce the levels of lipid peroxidation products during hyperammonemia. This may be due to the flavonoids, which have been shown to inhibit lipid peroxidation in rat tissues and also to inhibit the free radical production in the cells of various enzymes. In this context, Bartosikova et al., (2003), have reported that morin treatment reduced the levels of TBARS in alloxan-induced diabetes mellitus, which may corroborate our present findings.

The level of lipid peroxidation in cells is controlled by various cellular defense mechanisms consisting of enzymatic and non-enzymatic scavenger systems, [29] the levels of which are altered in hyperammonemia. [16],[30] This might have decreased the levels of antioxidants in the kidney such as SOD, CAT, GPx, and GSH in the AC-treated group rats. Increased superoxide production and reduced activities of antioxidant enzymes have been reported in kidneys of rats subjected to ammonia toxicity. [7] In our investigations, the levels of both enzymatic and non-enzymatic antioxidants, which declined in the kidney of hyperammonemic animals, were significantly restored to near normal after treatment with morin (3, 5, 7, 2',4'- pentahydroxyflavone). It possesses superoxide-scavenging and powerful antioxidant activities, [31] therefore, it is possible that morin modulates kidney lipid peroxidation and the antioxidant status during a hyperammonemic condition, and this could be attributed to the natural antioxidants, ammonia lowering effect, and its free radical scavenging properties. However, the exact mechanism is still unclear and further research on the action of morin is underway.

   Conclusions Top

The study offers evidence of the antihyperammonemic and antioxidant effects of morin against oxidative stress in kidneys, induced by ammonium chloride.

   Acknowledegment Top

We thank the Indian Council of Medical Research (ICMR), New Delhi, for funding support in the form of Senior Research Fellowship (SRF) to S. Subash.

   References Top

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