The present study was aimed to investigate the antioxidant potential of
In mammals, ammonia is an important nitrogen substrate in several reactions and plays an important role in nitrogen homeostasis of mammalian cells. Ammonia is produced by amino acid and protein catabolism and is toxic to brain and muscles. Ammonia toxicity results in free-radical generation that leads to oxidative stress and tissue damage [
The greatest disadvantage of the currently available potent conventional or synthetic antihyperammonemic agents/therapies lies in their toxicity and reappearance of symptoms after discontinuation. Valporic acid, phenobarbitol and carbamazepine are some of the currently used antiseizure and antihyperammonemic drugs. These drugs or therapies are sometimes inadequate and can have serious adverse effects [
Although various phytochemical constituents and diverse medicinal activities have been attributed to this plant, no biochemical studies have been carried out to shed light on the role of
The mature green
Alcoholic extract of the fruit was prepared according to the method developed by Shibib et al. [
Ammonium chloride was purchased from Sisco Research Laboratories, Mumbai, India. All the other chemicals used in the study were of analytical grade.
Adult male albino Wistar rats, weighing 180–200 g bred in the Central Animal House, Rajah Muthiah Medical College, Annamalai University, were used for the experiment. The animals were housed in polycarbonate cages in a room with a 12-h day-night cycle, at a temperature of 22 ± 2°C and humidity of 45–64%. The animals were fed with a standard pellet diet (Hindustan Lever Ltd, Mumbai, India) and water
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 1% (w/v) CMC and were considered as control, Group II normal rats were administered with MCE (300 mg kg−1 body weight) using an intragastric tube [
At the end of eighth week, the rats were fasted overnight and killed by cervical dislocation after anesthetizing with ketamine hydrochloride (30 mg kg−1 body weight; im). Blood was collected, and plasma and serum were separated by centrifugation. The liver and brain tissues were excised immediately and rinsed in ice-chilled normal saline. About 500 mg 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 various biochemical parameters.
Blood ammonia was determined by the enzymatic kinetic colorimetric assay developed by Wolheim [
Plasma thiobarbituric acid reactive substances (TBARS) were estimated by the method developed by Yagi [
Statistical analysis was performed using one-way analysis of variance (ANOVA), followed by Duncan’s multiple range test (DMRT) using SPSS software package 13.0. The results were expressed as mean ± SD from eight rats in each group, and
Table
Effect of MCE on changes in the blood ammonia and plasma urea, serum AST, ALT and ALP of normal and experimental rats.
Groups | Blood ammonia ( | Urea (mg dl−1) | AST (IU l−1) | ALT (IU l−1) | ALP (IU l−1) |
---|---|---|---|---|---|
Normal | 89.54 ± 4.47a | 10.28 ± 2.99a | 72.10 ± 6.26a | 24.27 ± 2.08a | 74.42 ± 6.14a |
Normal + MCE (300 mg kg−1) | 85.32 ± 8.94a | 11.40 ± 0.77a | 70.24 ± 6.28a | 22.01 ± 1.78a | 73.96 ± 4.50a |
AC (100 mg kg−1) | 327.15 ± 26.83b | 22.93 ± 1.78b | 118.41 ± 10.73b | 60.34 ± 5.36b | 141.72 ± 12.52b |
MCE (300 mg kg−1) + AC | 140.26 ± 17.88c | 13.15 ± 0.89c | 85.82 ± 8.74c | 32.74 ± 2.68c | 85.71 ± 7.15c |
Each value is mean ± SD for 8 rats in each group. Values not sharing a common superscript (a, b and c) differ significantly at
Table
Effect of MCE on the levels of TBARS and HP in plasma, liver and brain in normal and experimental rats.
Groups | Plasma TBARS (nM ml−1) | Plasma HP (values × 10−5 mM dL−1) | Liver TBARS (mM per 100 g wet tissue) | Liver HP (mM per 100 g wet tissue) | Brain TBARS (mM per 100 g wet tissue) | Brain HP (mM per 100 g wet tissue) |
---|---|---|---|---|---|---|
Normal | 2.74 ± 0.17a | 8.40 ± 0.11a | 0.86 ± 0.07a | 66.08 ± 5.21a | 1.06 ± 0.08a | 112.00 ± 8.50a |
Normal + MCE (300 mg kg−1) | 2.93 ± 0.17a | 8.25 ± 0.54a | 0.80 ± 0.06a | 64.03 ± 5.12a | 0.96 ± 0.07a | 111.28 ± 8.48a |
AC (100 mg kg−1) | 4.56 ± 0.35b | 13.16 ± 1.01b | 2.16 ± 0.14b | 97.86 ± 7.03b | 1.97 ± 0.15b | 135.24 ± 10.30b |
MCE (300 mg kg−1) + AC | 3.10 ± 0.17c | 10.20 ± 0.89c | 1.15 ± 0.11c | 76.56 ± 5.81c | 1.36 ± 0.11c | 118.44 ± 2.76c |
Each value is mean ± SD for 8 rats in each group. Values not sharing a common superscript (a, b and c) differ significantly at
The activities of SOD, CAT, GSH and GPx in the liver and brain of the normal and experimental rats are shown in Tables
Effect of MCE on the activities of SOD and CAT in the liver and brain of normal and experimental rats.
Groups | Liver SOD (Ua mg−1 protein) | Liver catalase (Ub mg−1 protein) | Brain SOD (Ua mg−1 protein) | Brain catalase (Ub mg−1 protein) |
---|---|---|---|---|
Normal | 9.01 ± 0.58a | 84.02 ± 6.30a | 7.01 ± 0.47a | 3.14 ± 0.23a |
Normal + MCE (300 mg kg−1) | 9.70 ± 0.73a | 85.04 ± 6.45a | 7.02 ± 0.48a | 3.27 ± 0.19a |
AC (100 mg kg−1) | 3.78 ± 0.29b | 40.98 ± 3.11b | 5.28 ± 0.30b | 0.87 ± 0.07b |
MCE (300 mg kg−1) + AC | 6.41 ± 0.38c | 70.60 ± 5.37c | 7.40 ± 0.35c | 2.74 ± 0.21c |
Each value is mean ± SD for 8 rats in each group. Values not sharing a common superscript (a, b and c) differ significantly at
Ua − Ub is defined as the enzyme concentration required to inhibit the OD at 560 nm of chromogen production by 50% in 1 min.
Effect of MCE on the activities of GSH and the levels of GPx normal and experimental rats.
Groups | Liver GSH (mg per 100 g wet tissue) | Liver GPx (Ud mg−1 protein) | Brain GSH (mg per 100 g wet tissue) | Brain GPx (Ud mg−1 protein) |
---|---|---|---|---|
Normal | 49.28 ± 3.02a | 9.39 ± 0.70a | 35.45 ± 2.69a | 3.38 ± 0.25a |
Normal + MCE (30 mg kg−1) | 50.46 ± 4.31a | 9.85 ± 0.74a | 37.40 ± 2.76a | 3.57 ± 0.27a |
AC (100 mg kg−1) | 24.86 ± 1.93b | 5.06 ± 0.43b | 20.94 ± 1.74b | 1.19 ± 0.09b |
MCE (30 mg kg−1) + AC | 42.79 ± 3.20c | 7.57 ± 0.72c | 27.62 ± 2.16c | 2.69 ± 0.20c |
Each value is mean ± SD for 8 rats in each group. Values not sharing a common superscript (a, b and c) differ significantly at
Ammonia is present in all living organisms as a product of degradation of proteins and other nitrogenous compounds. At higher levels, ammonia is toxic, leading to functional disturbances in the central nervous system that could lead to coma and death. To avoid the deleterious effects of ammonia, ureotelic animals detoxify ammonia by incorporating it into urea that is eliminated in urine [
Numerous investigations have documented that plant extracts containing phenolic compounds and flavonoids offer ammonia detoxification by removing excess ammonia, urea, uric acid and creatinine during various disease conditions, such as hyperammonemia, nephrotoxicity, and so forth [
The elevated levels of circulatory liver markers and lipid peroxidation products in AC rats might be due to the liver damage caused by ammonia-induced free radical generation [
MCE attenuates the AC-induced oxidative damages. Thick line represents inhibition.
Under normal conditions, a dynamic equilibrium exists between the production of reactive oxygen species (ROS) and the antioxidant capacity of the cell [
ROS may attack any type of molecules, but their main target appears to be polyunsaturated fatty acids (PUFAs), the precursors of lipid peroxide formation [
Serum AST, ALT and ALP are the most sensitive markers employed in the diagnosis of liver diseases. When the liver cell plasma membrane is damaged, numerous enzymes normally located in the cytosol are released into the blood stream [
The ROS generation in tissues is efficiently scavenged by the enzymatic and nonenzymatic antioxidants. The decrease in the activities of antioxidant enzymes is in close relationship with the induction of lipid peroxidation [
We observed decreased levels of GSH along with decreased activity of GPx in the liver and brain tissues of hyperammonemic rats. GSH is a major endogenous antioxidant, which counteracts free-radical-mediated damage [
The biochemical findings of our present study indicate that MCE exerts protection to AC-induced hyperammonemic rats against oxidative stress. This could be due to the prevention or inhibition of lipid peroxidative system by its antioxidant, maintenance of cellular integrity and hepatoprotective effect. However, the exact mechanism is still unclear and further research on the effect of the constituents of this plant is needed.