Evaluation of Antidiabetic and Antioxidant Activity of Praecitrullus fistulosus Fruits in STZ Induced Diabetic Rats
Arpita Karandikar, G. Sriram Prasath and S. Subramanian*
Department of Biochemistry, University of Madras, Guindy Campus, Chennai 600 025, India
*Corresponding Author E-mail: subbus2020@yahoo.co.in
ABSTRACT:
Praecitrullus fistulosus, commonly known as round melon or Tinda is a climbing herb known for its edible fruits. The entire immature fruits are rich in phytochemicals of both beneficial as well as pharmacological activities. The present study was aimed to evaluate the antidiabetic and antioxidant properties of Praecitrullus fistulosus fruit extract in STZ-induced experimental diabetic rats. The phytochemical screening of the Praecitrullus fistulosus fruit extract revealed the presence of alkaloids, flavonoids, saponins, tannins, phytosterol and diterpenes. The effect of oral administration of Praecitrullus fistulosus fruit extract (300mg/kg b.w.) on the levels of biochemical parameters were determined in experimental groups of rats. The altered levels of biochemical parameters in the diabetic rats were significantly reverted back to near basal values upon treatment with the fruit extract. The levels of glycogen content, the activities of glycogen metabolizing enzymes were improved upon the extract treatment. The altered activities of serum aminotransferases and alkaline phosphatase were restored to normalcy. The levels of lipid peroxides in the plasma and pancreatic tissues of diabetic rats were elevated significantly and were normalized by the administration of fruit extract. The activities of pancreatic antioxidants and the levels of plasma non-enzymatic antioxidants were markedly declined in the diabetic rats. Upon treatment with fruit extract to diabetic rats, decreased levels were elevated to near normal values. The results of the study indicate that Praecitrullus fistulosus fruit extract possesses antidiabetic and antioxidant property. The results are comparable with gliclazide, an oral standard hypoglycemic drug. The phytochemicals present in the Praecitrullus fistulosus fruit extract may account for the observed pharmacological properties.
KEYWORDS: Praecitrullus Fistulosus; STZ-diabetes; antidiabetic; antioxidant.
INTRODUCTION:
Diabetes mellitus (DM) is a chronic metabolic disorder that has emerged as one of the main alarms to human health in the 21st century [1]. It is characterized by persistent hyperglycemia resulting from defects in insulin secretion, insulin action, or both. DM may be suspected or recognized clinically by the onset of one or more of the characteristic symptoms such as polyuria, polydipsia, polyphagia and unusual weight loss. Under normal physiological conditions, the blood glucose levels are controlled by insulin which lowers the blood glucose level by facilitating the entry of glucose in to the cells for energy production[2].
About 80% of people with diabetes are in developing countries, of which India and China share the larger contribution.
The pathogenesis, progress and the possibility of diabetes management by oral antidiabetic medications have stimulated great interest in recent decades. Numerous therapies designed for the treatment of DM have proven to be fairly effective, but none is ideal due to undesirable side effects and diminution after prolonged use. Management of diabetes without any side effects is still a challenge to the medical system. This leads to increasing demand for an antidiabetic medicinal plant which has comparatively less side effects.
According to the recommendation of the WHO expert committee on Diabetes mellitus, an investigation of hypoglycaemic agents of plant origin used in traditional medicine have become more important. In the series of medicinal plants that lacks scientific scrutiny is Praecitrullus fistulosus, which belongs to the family Cucurbitaceae. It is commonly known as “Tinda”and the other vernacular names include Indian roung guord, apple gourd, and Indian Baby pumpkin [3]. It is one of the excellent plants gifted by nature having the composition of essential constituents that are required for normal and good human health [4]. It has been cultivated in Asia since ancient times and has been considered as an under exploited crop in the western world [5]. The tender fruits are picked at the immature stage and cooked as a cooked vegetable. Though, the plant has morphological features similar to watermelon both have different monoploid chromosome number and pollen morphology. The plant prefers sandy soils where its roots can penetrate easily and grown in dry season. The fruits are ready to harvest in 13-15 weeks from sowing, depending on the temperature and other growing conditions. The tender fruits are green in colour and have a diameter of 10-12cm and the seeds are still soft. Up to 6 fruits of about 500g each can be harvested per plant.
The fruit is a type of berry called a pepo by Gerald Carr. The fruit is approximately spherical, and 10–12 cm in diameter [6]. Fruit is about the size of a small turnip, depressed at each end, hispid when young afterwards glabrous [7]. The fruit is a berry that has a fleshy fruit with several seeds each with hard coat (Joseph),smooth(wealth of India), black in which all parts of the pericarp are pulpy or fleshy except the exocarp which is often skin-light. They are green coloured, apple-sized fruits, spherical in shape and 50 to 60 grams in weight. Plants are vigorous, productive and begin to bear fruits in 70 days [8] . Praecitrullus fistulosus possess a wide range of pharmacological properties such as a antioxidant activity (fruits), antimicrobial activity (seeds), [9, 10]. The leaves are cooked as vegetable and taken for blood pressure [11].
In the absence of systematic studies in literature, the present study is aimed to evaluate the antidiabetic and antioxidant Praecitrullus fistulosus fruits in STZ-induced experimental rats.
MATERIALS AND METHODS:
Plant Material:
The fruits of Praecitrullus fistulosus were collected from Indore District (M.P). The leaves were identified and authenticated and a voucher specimen was deposited at the Department of Botany, University of Madras, Chennai.
Preparation of Plant Extract:
The fruits of Praecitrullus fistulosus were dried at room temperature and powdered in an electrical grinder, which was then stored in an airtight container at 5° C until further use. The powdered fruits were delipidated with petroleum ether (60 - 80° C) for overnight. It was then filtered and soxhalation was performed with 95% Ethanol. Ethanol was evaporated in a rotary evaporator at 40 – 50° C under reduced pressure. The yield was (18.4g).
Preliminary Phytochemical Screening:
The ethanolic extract of Praecitrullus fistulosus fruits were subjected to preliminary phytochemical screening of various plant constituents [12, 13].
Experimental Animals:
Male albino Wistar rats (150-180 g) were purchased from TANUVAS, Madavaram, Chennai. The rats were housed in polypropylene cages lined with husk and kept in Animal house, Department of Biochemistry. It was renewed every 24 hours. The rats were fed with commercial pellated rats chow (VRK Nutritional Solutions, Maharashtra, India) and had free access to water. The experimental rats were maintained in a controlled environment (12:12 hours light/dark cycle) and temperature (30 ± 2°C). The experiments were designed and conducted in accordance with the ethical norms approved by Ministry of Social Justices and Empowerment, Government of India and Institutional Animal Ethics Committee Guidelines for the investigation of experimental pain in conscious rats. The rats were acclimatized for one week before starting the experiments.
Induction of Diabetes Mellitus:
Experimental diabetes was induced in overnight fasted rats by single intraperitoneal injection of streptozotocin (45 mg/kg b.w) dissolved in freshly prepared 0.1M of cold citrate buffer (pH 4.5) [14]. Since, STZ is capable of inducing fatal hypoglycemia due to massive pancreatic insulin release, the rats were provided with 10% glucose solution after 6 h of STZ administration for the next 24 h to overcome drug induced hypoglycemia [15]. Neither death nor any other adverse effect was observed. After a week time, for the development and aggravation of diabetes, rats with moderate diabetes (i.e. fasting blood glucose concentration, >250 mg/dl) that exhibited hyperglycemia and glycosuria were selected for further experimentation.
Experimental Design:
The rats were grouped into 4 groups, comprising of 6 rats in each group as follows:
Group I Control rats
Group II STZ induced diabetic Rats.
Group III Diabetic Rats treated with Praecitrullus fistulosus fruits extract (300 mg/Kg Body weight/day) in aqueous solution orally for 30 days.
Group IV Diabetic Rats treated with gliclazide (5mg/Kg body weight/day) in aqueous solution orally for 30 days.
During the experimental period, body weight and blood glucose levels of all the rats were determined at regular intervals. At the end of the experimental period, the rats were fasted over night, anaesthetized, and sacrificed by cervical decapitation. The blood was collected with or without anticoagulant for plasma or serum separation respectively.
Preparation of tissue homogenate:
The liver and pancreatic tissues were excised, rinsed in icecold saline. Known amount of the tissues were homogenized in Tris–HCl buffer (100 mM, pH 7.4) at 4°C, in a Potter–Elvehjem homogenizer with a Teflon pestle at 600 rpm for 3 min. The homogenate was then centrifuged at 12,000×g for 30 min at 4°C. The supernatant was collected as tissue homogenate, which was used to assay of various parameters. The protein content in the tissue homogenate was estimated by the method of Lowry et al (1951) [16]. A portion of wet liver tissue was used for the estimation of glycogen content [17]. Glycogen synthase [18], glycogen phosphorylase [19] were assayed in liver tissues.
Oral Glucose Tolerance Test (OGTT):
At the end of the experimental period, fasting blood samples were taken from all the groups of rats to perform oral glucose tolerance test. Four more blood samples were collected at 30, 60, 90 and 120 min intervals after an oral administration of glucose solution at a dosage of 2 g kg-1 body weight. All the blood samples were collected with EDTA for the estimation of glucose.
Biochemical parameters:
Blood glucose level was estimated by the method of glucose oxidase/peroxidase method as described by Trinder (1969) [20] using a commercial kit (Span Diagnostic Chemicals, India). and urea [21] estimation. Glycosylated hemoglobin was estimated according to method of Nayak and Pattabiraman, (1981) [22]. Plasma was used for protein assay (Lowry et al., 1951) [16]. Urine sugar was detected using urine strip. Serum was used for the determination of creatinine [23] and uric acid [24]. Aspartate transaminase(AST), Alanine transaminase(ALT)and Alkaline phosphatase (ALP) were assayed by the method of King (1965(a, b)) [25, 26].
Assay of antioxidant status:
The levels of lipid peroxides were determined in plasma and tissue homogenate [27, 28] . The activities of enzymatic antioxidants such as SOD, catalase and GPx [29, 30, 31] were assayed in the tissue homogenate of control and experimental groups of rats. The levels of nonenzymatic antioxidants, vitamin C, vitamin E, and GSH were determined[32, 33, 34].
Statistical Analysis:
The values were expressed as mean ± S.D for six rats in each group. All data were analyzed with SPSS/16.0 student software. Hypothesis testing method included one way analysis of variance (ANOVA) followed by post hoc testing performed with least significant difference (LSD) test. A Value of P < 0.05 was considered as significant.
RESULTS:
Table 1 shows the qualitative analysis of phytochemicals present in the ethanolic extract of Praecitrullus fistulosus fruits. From the preliminary phytochemical evaluation, it was found that the Praecitrullus fistulosus fruits extract found to contain alkaloids, flavonoids, saponins, tannins, phytosterol and diterpenes
Table 1 Phytochemical screening of Praecitrullus fistulosus fruits extract:
Phytoconstituents |
Inference |
Alkaloids |
+ |
Flavonoids |
+ |
Saponins |
+ |
Tannins |
+ |
Phytosterol |
+ |
Diterpenes |
+ |
Triterpenoids |
- |
Glycosides |
+ |
Anthraquinones |
- |
Phenols |
+ |
Table 2 shows the changes in body weight in control and experimental group of animals. The body weight of control rats was normal whereas there was a significant decrease in the body weight of STZ induced diabetic rats. Diabetic rats treated with Praecitrullus fistulosus fruits extract and gliclazide for 30 days showed a significant improvement in body weight.
Table 2 Effect of Praecitrullus fistulosus extract on changes in body weight of experimental groups of rats after 30 days treatment.
Groups |
Body weight (g) |
|
Initial |
Final |
|
Control |
174.38 ± 2.79 |
235.39 ± 4.99 |
Diabetic |
167.91 ± 3.55 |
144.76± 5.14* |
Diabetic + Praecitrullus fistulosus extract |
175.12 ± 3.29 |
195.19 ± 4.37@ |
Diabetic + gliclazide |
169.81 ± 3.11 |
192.80 ± 4.86@ |
Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 3 depicts the levels of blood glucose in certain durations after the oral administration of glucose (2g/Kg body weight) in normal and experimental group of rats. In control rats, the blood glucose level reached the maximum peak at 60 min after an oral glucose load and then it was gradually reverted back to near normal levels after 120 min.
Table 3 Effect of Praecitrullus fistulosus extract on the blood glucose level (mg/dl) in the experimental groups of rats receiving an oral glucose load.
Groups |
Fasting |
30 min |
60 min |
90 min |
120 min |
Control |
81.64 ± 10.82 |
182.35 ± 15.17 |
210.39 ± 18.99 |
161.48 ± 15.90 |
105.47 ± 16.78 |
Diabetic |
290.35 ± 22.94* |
327.49 ± 25.07* |
391.62 ± 31.49* |
355.14 ± 26.38* |
315.99 ± 24.60* |
Diabetic + Praecitrullus fistulosus extract |
169.12 ± 17.16@ |
212.45 ± 18.94@ |
281.65 ± 28.72@ |
201.56 ± 19.74@ |
172.92 ± 18.12@ |
Diabetic + gliclazide |
157.39 ± 15.98@ |
224.70 ± 21.68@ |
276.92 ± 27.32@ |
195.36 ± 21.56@ |
165.30 ± 17.82@ |
Unit: mg/dL; Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 4 depicts the levels of blood glucose, glycosylated hemoglobin and urine sugar. STZ induced diabetic rats showed a significant elevation in the levels of blood glucose, presence of urine sugar and a simultaneous increase in glycosylated hemoglobin. Oral administration of ethanolic extract of Praecitrullus fistulosus fruits to the diabetic group of rats significantly reduced the levels of blood glucose and glycosylated hemoglobin. Urine sugar which was present in the diabetic group of rats was found to be absent in rats treated with the extract.
Table 4 Effect of Praecitrullus fistulosus extract on the levels of blood glucose, glycosylated hemoglobin, and urine sugar in the experimental groups of rats.
Groups |
Glucose (mg/dl) |
Glycosylated hemoglobin (%) |
Urine sugar |
Control |
90.65 ± 13.84 |
6.74 ± 1.82 |
Nil |
Diabetic |
288.92 ± 24.29* |
12.95 ± 3.14* |
+++ |
Diabetic + Praecitrullus fistulosus extract |
158.92 ± 20.12@ |
8.42 ± 1.84@ |
Nil |
Diabetic + gliclazide |
162.13 ± 18.58@ |
8.09 ± 1.95@ |
Nil |
Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 5 depicts the levels of total protein, blood urea, serum creatinine and uric acid in control and experimental group of rats. In STZ induced diabetic rats, there was a significant decrease in the total protein and increase in the levels of urea, uric acid and creatinine when compared with the control group of rats. Administration of an ethanolic extract of Praecitrullus fistulosus fruits as well as the standard drug, gliclazide to the diabetic group of rats significantly decreased the levels of blood urea, uric acid, serum creatinine and increased the levels of total protein.
Table 6 depicts the levels the levels of glycogen content and activities of glycogen synthase and glycogen phosphorylase in liver tissues control and experimental groups of rats. A significant decline in the glycogen level as well as in the glycogen synthase activity and a concomitant increase in the activity of glycogen phosphorylase were noted in the liver of diabetic rats. Oral administration of Praecitrullus fistulosus fruits extract to diabetic rats restored the level of glycogen and the activities of glycogen synthase, glycogen phosphorylase to near normalcy when compared to control group of rats.
Table 7 depicts the levels of aspartate transaminase, alanine transaminase and alkaline phosphatase in the control and experimental group of rats. Diabetic rats showed a significant elevation in the levels of aspartate transaminase, alanine transaminase and alkaline phosphatase when compared with the control group of rats. Administration of Praecitrullus fistulosus fruits extract and gliclazide to the diabetic rats resulted in a significant decrease in the levels of these markers.
Table 5 Effect of Praecitrullus fistulosus extract on the levels of protein, urea, creatinine and uric acid in plasma of experimental groups of rats.
Groups |
Protein (g/dl) |
Urea (mg/dl) |
Creatinine (mg/dl) |
Uric acid (mg/dl) |
Control |
8.59 ± 1.99 |
28.35 ± 3.91 |
0.86 ± 0.09 |
2.94 ± 0.85 |
Diabetic |
5.82 ± 1.10* |
50.59 ± 6.22* |
2.48 ± 0.51* |
5.38 ± 1.66* |
Diabetic + Praecitrullus fistulosus extract |
7.15 ± 1.31@ |
32.74 ± 5.81@ |
1.14 ± 0.25@ |
3.62 ± 1.31@ |
Diabetic + gliclazide |
7.28 ± 1.42@ |
35.36 ± 4.85@ |
1.32 ± 0.41@ |
3.89 ± 1.22@ |
Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 6 Effect of Praecitrullus fistulosus extract on the levels of liver glycogen content, and the activities of glycogen metabolizing enzymes in the experimental groups of rats.
Groups |
Glycogen |
Glycogen synthase |
Glycogen phosporylase |
Control |
49.13 ± 6.75 |
782.79 ± 54.26 |
591.57 ± 32.14 |
Diabetic |
28.14 ± 4.81* |
526.31 ± 35.19* |
891.24 ± 61.97 * |
Diabetic + Praecitrullus fistulosus extract |
41.02 ± 4.94@ |
685.41 ± 40.92 @ |
689.38 ± 45.09 @ |
Diabetic + gliclazide |
35.48 ± 5.63@ |
671.94 ± 45.61 @ |
675.99 ± 48.24@ |
Units are expressed as: mg/g wet tissue for glycogen, μmoles of UDP formed/h/mg protein for glycogen synthase and μmoles of Pi liberated/h/mg protein for glycogen phosphorylase. Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 7 Effect of Praecitrullus fistulosus extract on the activity of AST, ALT and ALP in the serum of experimental groups of rats.
Groups |
AST |
ALT |
ALP |
Control |
41.35 ± 12.71 |
22.74 ± 3.86 |
76.24 ± 12.88 |
Diabetic |
122.69 ± 19.28* |
65.81 ± 17.25* |
190.14 ± 22.82* |
Diabetic + Praecitrullus fistulosus extract |
75.34 ± 16.28@ |
36.95 ± 8.26@ |
95.44 ± 18.17@ |
Diabetic + gliclazide |
80.55 ± 15.91@ |
31.67 ± 10.38@ |
105.28 ± 16.93@ |
The enzyme activities are expressed as: AST and ALT µmoles of pyruvate liberated /h/mg of protein; ALP µmoles of phenol liberated/min/mg of protein. Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
The levels of TBARS in the plasma and pancreas of control and experimental group of rats are presented in Table 8. STZ induced diabetic rats showed marked increase in the levels of TBARS when compared to control rats. Treatment of Praecitrullus fistulosus fruits extract to diabetic rats showed a significant decrease in the levels of TBARS.
Table 8 Effect of Praecitrullus fistulosus extract on the level of TBARS in plasma and pancreas, of experimental groups of rats.
Groups |
TBARS |
|
Plasma |
Pancreas |
|
Control |
3.57 ± 0.83 |
32.81 ± 6.79 |
Diabetic |
7.25 ± 2.16* |
70.64 ± 16.71* |
Diabetic + Praecitrullus fistulosus extract |
5.12 ± 1.21@ |
42.86 ± 12.94@ |
Diabetic + gliclazide |
4.86 ± 1.54@ |
45.37 ± 14.96@ |
Units: mM/100 g in tissues; nM/ml in plasma. Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 9 and 10 illustrates the activities of enzymatic and non enzymatic antioxidants in pancreas and plasma of control and experimental group of rats. In STZ induced diabetic rats, there was a significant reduction in the activities of enzymatic and non enzymatic antioxidants in pancreas and plasma respectively Treatment of Praecitrullus fistulosus fruits extract to the diabetic rats showed improvement in the activities of enzymatic and non enzymatic antioxidants.
DISCUSSION:
Streptozotocin is specifically cytotoxic to β-cells of the pancreas. The mechanism behind its action is that STZ is preferentially up taken by pancreatic beta cell via GLUT2 transporter and causes DNA alkylation followed by the activation of poly ADP ribose polymerase leading to depletion of NAD+ and ATP. Enhanced ATP dephosporylation after STZ treatment supplies substrate for xanthine oxidase resulting in the formation of superoxide radicals and also nitric oxide moiety is liberated. As a net result, destruction of β-cells occurs by necrosis [35]. However, the dose of streptozotocin required for inducing
diabetes depends on the animal species, route of administration, dosage, duration and nutritional status.
Phytochemical analysis on the Praecitrullus fistulosus fruits extract revealed the presence of biologically active ingredients such as alkaloids, flavonoids, saponins, tannins, phytosterol and diterpenes. Phytochemicals are bioactive non-nutrient plant derivatives that have preventive properties. They confer plants with odor (terpenoids), pigmentation (tannins and quinines) and antioxidants (flavonoids)[36]. These bioactive components are said to be responsible for the broad spectrum of pharmacological activities of medicinal plants.
Diabetic rats showed a clear muscle atrophy involving a decrease in both skeletal muscle mass and protein content. This was accompanied by marked loss of total carcass nitrogen. These changes are related to important alterations in the protein turnover in skeletal muscle [37]. Diabetic rats treated with Praecitrullus fistulosus fruits extract and gliclazide for 30 days showed a significant improvement in body weight indicating the beneficial effect of the fruits extract in controlling muscle wasting.
Diabetes mellitus is characterized by decreased glucose tolerance due to low secretion of insulin or its action. This is manifested by elevated blood glucose levels and glucosuria, which may be accompanied by changes in lipid and protein metabolism. When the diabetic rats were challenged with an oral glucose load, the blood glucose levels reached a peak at 60 minutes and gradually decreased to pre-glucose load level around 120 minutes, the present study revealed that oral administration of the ethanolic extract of Praecitrullus fistulosus fruits improved the glucose tolerance in STZ - induced diabetic rats.
Table 9 Effect of Praecitrullus fistulosus extract on the activity of SOD, Catalase and GPx, and the level of GSH in pancreatic tissues of experimental groups of rats.
Groups |
SOD |
Catalase |
GPx |
GSH |
Control |
4.36 ± 1.59 |
19.74 ± 4.12 |
7.12 ± 1.86 |
23.94 ± 3.12 |
Diabetic |
1.86 ± 0.91* |
7.26 ± 2.19* |
3.71 ± 0.99* |
12.86 ± 2.74* |
Diabetic + Praecitrullus fistulosus extract |
3.51 ± 1.62@ |
16.88 ± 4.05@ |
4.99 ± 1.38@ |
18.62 ± 2.98@ |
Diabetic + gliclazide |
3.37 ± 1.72@ |
15.36 ± 3.19@ |
4.67 ± 1.69@ |
19.15 ± 3.18@ |
Activity is expressed as: 50% of inhibition of epinephrine autooxidation/min/mg of protein for SOD; µmoles of hydrogen peroxide decomposed/min/mg of protein for catalase; µmoles of glutathione oxidized/min/mg of protein for GPx; mg/100 g tissue for GSH. Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Table 10 Effect of Praecitrullus fistulosus extract on the levels of vitamin C, vitamin E, ceruloplasmin and GSH in plasma of experimental groups of rats.
Groups |
Vitamin C |
Vitamin E |
Ceruloplasmin |
GSH |
Control |
1.86 ± 0.37 |
0.81 ± 0.12 |
13.75 ± 2.14 |
22.94 ± 4.75 |
Diabetic |
0.61 ± 0.14* |
0.41 ± 0.07* |
5.26 ± 1.39* |
13.76 ± 2.81* |
Diabetic + Praecitrullus fistulosus extract |
1.42 ± 0.29@ |
0.66 ± 0.14@ |
8.94 ± 2.10@ |
17.91 ± 3.15@ |
Diabetic + gliclazide |
1.38 ± 0.31@ |
0.62 ± 0.15@ |
7.68 ± 1.76@ |
18.24 ± 3.17@ |
Units: mg/dl. Values are given as mean ± SD for groups of six rats in each. Values are statistically significant at p < 0.05. Statistical significance was compared within the groups as follows: *compared with control, @ compared with diabetic rats.
Blood glucose is an index for the diagnosis of diabetes mellitus. Liver functions as a primary organ for glucose homeostasis and plays a vital role in the maintenance of blood glucose levels. STZ administration induces pronounced increase in the concentrations of blood glucose. Blood glucose levels are maintained mainly by insulin that facilitates the uptake, utilization and storage of glucose. During diabetes, the blood glucose levels are drastically increased which results from reduced glucose utilization by various tissues, which is a typical condition of insulinopenic diabetes. The elevated blood glucose level observed in diabetic rats was almost normalized upon extract treatment which may be due to stimulation of glucose utilization by the peripheral tissues.
Persistent hyperglycemia results in increased glycation of a number of proteins including haemoglobin and alpha crystalline or lens [38]. This glycation alters the structure and function of haemoglobin resulting in Hb desaturation and precipitation of red blood cells as Heing bodies [39]. Oral administration of Praecitrullus fistulosus fruits extract to diabetic group of rats reduced the formation of glycosylated hemoglobin by virtue of its normoglycemic activity. Also, oral administration of Praecitrullus fistulosus fruits to diabetic rats showed decreased urine sugar excretion due to the normalization of blood glucose levels.
Protein serves as a source of nutrition for the tissues and its synthesis and regulation determines normal function. Insulin plays a pivotal role in protein synthesis. Diabetes mellitus shows profound changes in circulating aminoacids and hepatic aminoacid uptake [40]. The significant decrease of plasma protein in STZ-induced diabetic rats could be attributed to suppressed protein synthesis. Urea is a non-protein nitrogenous waste product whose level reflects a normal and continued protein metabolism. Diabetes mellitus is associated with changes in negative nitrogen balance and loss of nitrogen from most organ systems. The increase in synthesis of urea in diabetic rats may be due to the enhanced catabolism of both liver and plasma proteins. There was increased protein catabolism with flow of aminoacids into the liver, which feeds glyconeogenesis during diabetes [41]. Dighe et al. reported that accelerated proteolysis of uncontrolled diabetes occurs as a result of deranged glucagon mediated regulation of cAMP formation in insulin deficiency [42]. This accounts for the observed decrease in the total protein content in STZ induced diabetic rats. Oral administration of the fruit extract to diabetic rats significantly inhibits proteolysis caused by insulin deficiency and thus increases the level of plasma proteins to near normal levels.
Persistent hyperglycemia causes severe derangement in protein metabolism that result in the development of negative nitrogen balance. This in turn elevates urea and creatinine level [43], which act as a biochemical diagnostic marker for assessing renal impairment and drug-induced toxicity. The observed alteration in the levels of blood urea and serum creatinine in group of diabetic rats reverted to near normalcy by treatment with the fruit extract, indicating its renal protective nature.
Uric acid is the main catabolic product from purine nucleotides by xanthine oxidase enzymatic system. It is a biomarker for the development of diabetic complications. Costa et al. (2002) suggested increased uric acid concentration to be a risk factor for cardiovascular diseases[44]. The levels of serum uric acid in diabetes induced rats were found to be increased. Treatment with the fruit extract significantly ameliorated the toxic effects of STZ as indicated by the restoration of serum uric acid levels.
Glycogen is a branched polymer of glucose residues which is synthesized by the enzyme glycogen synthase [45]. Glycogen synthase and glycogen phosphorylase are the rate-limiting enzymes in glycogen metabolism. During diabetic conditions, the glycogen levels, glycogen synthase activity and responsiveness to insulin signaling are diminished and glycogen phosphorylase activity is significantly increased [46]. Oral administration of fruit extract to diabetic rats restored the glycogen content and the activities of glycogen metabolizing enzymes demonstrating the possible role on the utilization and storage of glucose in the hepatic tissues of the experimental diabetic rats.
ALT and AST are the intracellular enzymes serve as a clinical index of tissue injury chiefly hepatocyte as well as renal injury. ALP acts as a marker of biliary function and cholestasis. The elevation in the levels of these hepatic marker enzymes indicates hepatocyte injury[47]. The increased activities of ALT, AST and ALP in the serum of diabetic rats may be primarily due to the leakage of these enzymes from liver as well as kidney into blood stream [48]. Oral administration of fruits extract to diabetic group of rats showed a notable decline in the activity of these enzymes to their basal levels, signifying its non toxic as well as tissue protective nature.
Diabetes is usually accompanied by increased production of free radicals or impaired antioxidant defenses. Excessive generation of free radicals cause damage to cellular proteins, membrane lipids and nucleic acids and eventually cell death. Decreased plasma insulin in diabetic conditions increases fatty acyl coenzyme A oxidase activity, which initiates β-oxidation of fatty acids, resulting in lipid peroxidation. Increased lipid peroxidation impairs membrane activity by decreasing membrane fluidity and altering activity of membrane bound enzymes and receptors. The products of lipid peroxidation are injurious to most cells in the body and are associated with a variety of diseases, such as atherosclerosis and brain damage. The significant increase in levels of TBARS observed in plasma and pancreas of diabetic rats were decreased to near normalcy upon oral administration of Praecitrullus fistulosus fruits extract to diabetic rats which could be a result of improved antioxidant status.
Hyperglycemia mediated oxidative stress contributes to the development and progression of most of the diabetes-associated complications. Oxidative stress definitely refers to the situation of an imbalance between the production of Reactive Oxygen Species (ROS) and antioxidant defense. There is emerging evidence that the formation of ROS is a direct consequence of hyperglycemia [49]. The reduced antioxidant capacity potentially makes pancreatic β-cells sensitive to ROS mediated signal transduction and cellular response. Thus, maintenance of β-cell oxidant status and their protection against oxidative damage might delay the onset of diabetes as well as the progression of its complications.
SOD is an important defense enzyme which catalyses the dismutation of superoxide radicals to hydrogen peroxide thereby reducing the likelihood of superoxide anion interacting with NO to form reactive peroxynitrite [50].The so formed hydrogen peroxide is successively metabolized into water and non-reactive oxygen species by the activities of catalase and GPx. The compromises in enzymatic antioxidant defense systems and alterations in their activities have been implicated in the mechanisms of abnormal tissue functions observed in diabetes mellitus. The levels of nonenymatic antioxidants were decreased in diabetic rats. However, oral administration of Praecitrullus fistulosus fruit extract to diabetic rats significantly improved the activities of enzymatic antioxidants and non enzymatic antioxidants, which in turn reflects its antioxidant property.
CONCLUSION:
Oral administration of the fruit extract resulted in the normalization of biochemical parameters and improved the antioxidant status. Also the levels of glycogen content and the activities of glycogen metabolizing enzymes were normalized upon treatment with the fruit extract. The antioxidant competence of the pancreatic tissues was improved by the fruit extract indicating its antioxidant potential. The observed activities of the extract may be attributed to the presence of various biologically active secondary metabolites. Further studies are in progress on the isolation and characterization of the active ingredients in the Praecitrullus fistulosus fruit extract.
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Received on 04.12.2013 Modified on 06.01.2014
Accepted on 14.01.2014 © RJPT All right reserved
Research J. Pharm. and Tech. 7(2): Feb. 2014; Page 196-203