|Year : 2018 | Volume
| Issue : 5 | Page : 251-259
Antidiabetic potential of active fraction obtained from methanolic extract of Ichnocarpus frutescens: A possible herbal remedy
Mallu Srujana, Ranjana Ramesh, Lakshmi Devi Nanjaiah
Department of Studies in Microbiology, University of Mysore, Manasa Gangothri, Mysore; Skanda Life sciences Pvt. Ltd., Bengaluru, Karnataka, India
|Date of Submission||12-Jan-2018|
|Date of Acceptance||08-Jun-2018|
|Date of Web Publication||14-Dec-2018|
Dr. Lakshmi Devi Nanjaiah
Department of Studies in Microbiology, University of Mysore, Mysore - 570 016, Karnataka
Source of Support: None, Conflict of Interest: None
OBJECTIVES: Ichnocarpus frutescens is a common plant used by tribal people and in Ayurveda for its high medicinal value. The purpose of the study was to investigate whether I. frutescens has any persuasive medicinal property to manage diabetes mellitus.
MATERIALS AND METHODS: Initially, male albino Wistar rats were given intraperitoneal injection of streptozotocin-nicotinamide to induce diabetes, followed with the administration of active fraction obtained from the methanolic extract of I. frutescens for the next 28 consecutive days. Glibenclamide (25 mg/kg) was used as positive control.
RESULTS: According to the results obtained, active fraction at a dose of 50 mg/kg body weight exhibited significant antihyperglycemic activity, which was evident with reduced blood glucose level up to 58.84%. The active fraction also showed improvement in serum lipid profile as well as regeneration of pancreatic β-cells in diabetic rats. Concurrent histopathological studies reinforce the effect of active fraction in healing pancreas, thus justifying the possible mechanism of its antidiabetic activity.
CONCLUSION: The results of the present investigation lead credence to the use of I. frutescens in ameliorating the diabetic condition.
Keywords: Diabetes mellitus, glibenclamide, hepatic glycogen, Ichnocarpus frutescens, plasma insulin
|How to cite this article:|
Srujana M, Ramesh R, Nanjaiah LD. Antidiabetic potential of active fraction obtained from methanolic extract of Ichnocarpus frutescens: A possible herbal remedy. Indian J Pharmacol 2018;50:251-9
|How to cite this URL:|
Srujana M, Ramesh R, Nanjaiah LD. Antidiabetic potential of active fraction obtained from methanolic extract of Ichnocarpus frutescens: A possible herbal remedy. Indian J Pharmacol [serial online] 2018 [cited 2020 Jul 5];50:251-9. Available from: http://www.ijp-online.com/text.asp?2018/50/5/251/247532
| » Introduction|| |
Humanity has exploited many medicinal plants from time immemorial to protect himself/herself against several diseases and also to improve his/her lifestyle. In the current era, medicinal plants are gaining immense importance because of their various phytoconstituents with distinctive properties.
Diabetes mellitus is a common endocrine-based disease with microvascular and macrovascular complications characterized with retinopathy, neuropathy, nephropathy, and atherosclerosis is a result of hyperglycemia and hyperlipidemia. There is an explosive increase in diabetic patients since the last two decades and is estimated to be the seventh major cause of death by 2030. Further, it occupies the third position when its fatal complications are considered. Diabetes mellitus is characterized by several complications, basically chronic hyperglycemia with altered metabolism of carbohydrate, fat, and protein, which results from fault insulin secretion, inefficient insulin action, or both. Such deficiency results in damage of many body systems, in particular the blood vessels and nerves. Diabetes has negative impact on modern lifestyle with direct consequence of increased overweight and sedentary population. The epidemiological status of diabetes has stimulated the researchers for new innovations and targets for the eradication of this incurable disease.
Currently, available synthetic antidiabetic drugs include insulin and other glucose level controlling agents such as alpha-glucosidase inhibitors, biguanides, sulfonylureas, and gliptins. These drugs are advised individually or in combination to achieve better glycemic regulation. However, the above-said drugs are known to cause side effects such as hypoglycemic coma and hepatorenal disturbance; further, they are not advisable during pregnancy. For example, “acarbose” is reported to have low efficacy in reducing the serum glucose levels. Lipase inhibitor causes reduction in patient's body weight (b.w.) and other drugs are known to be the reason for abdominal pain, hepatotoxicity, diarrhea, flatulence, and hypoglycemia. Hence, it is a puzzling task to identify a better lead molecule which is devoid of unwanted long-term adverse effects. In this regard, search for safer and effective hypoglycemic agents is a continuous challenge in managing diabetes. In this context, medicinal plants are reconsidered as effective and safe alternative natural remedy to treat diabetes mellitus as they contain a diverse group of compounds. For example, “metformin” derived from medicinal plants is an efficient oral glucose-lowering agent.
Available literature justifies that there are more than 800 plants species showing hypoglycemic activity. American Ginseng, green tea, and Astragalus are reported to have antidiabetic effect. The plant products with antidiabetic activity are in great demand due to low cost, limited side effects, and easy accessibility. Hence, various plants are being scrutinized for their effect on glycemic index. However, in several such plants, an organized clinical experiment is lacking to assess their health benefits. The WHO recommends the investigation on favorable effect of plant products in treating diabetes mellitus.
India is one of the eight important versatile centers enormously rich in medicinal and aromatic plants in diverse ecosystems. A large majority of Indian population is treated by conventional systems of remedy such as Ayurveda, Unani, and Siddha. Ichnocarpus frutescens R. Br., belonging to family Apocynaceae, is used from centuries as an important ingredient in traditional Indian medicine. It is commonly designated as “Black Sariva,” “black creeper,” or “dudhilata.” I. frutescens is a plant where all the plant parts are effectively utilized for various medicinal properties. It is used as folk medicine and as component in Ayurveda and Unani medicines against diseases related with blood, skin, and inflammation. The tribals of Karnataka and Uttar Pradesh used this plant for treating diabetes. The plant is enriched with several chemical constituents such as phenolic acids, phenylpropanoids, flavonoids, coumarines, sterols, and pentacyclic triterpenoids, i.e., Δ12-dehydrolupanyl-3 β-palmitate, Δ12-dehydrolupeol, 5-hydroxy octacosan-25-one, lupeol acetate, friedelin, oleanolic acid, friedelinol, nonane, sitosterol dotriacontanoic acid, and sitosterol palmitate.
Tribal people used this whole plant as medicine in treating atrophy, cough, convulsions, measles, bleeding gums, simple fevers, dysentery, and liver disorder. Savithramma et al. have done a survey among local population of Kailasakona, a sacred grove of Chittoor district, Andhra Pradesh, India, and explored 31 medicinal plants, wherein they have documented I. frutescens leaf powder having medicinal value to treat diabetes. Barik et al. reported promising antidiabetic activity of the aqueous root extract of I. frutescens in Type-2 diabetic-induced rats. Bioactive fractions of I. frutescens have been extracted using various solvent systems which are reported to possess the hepatoprotective effect and antioxidant, anti-inflammatory, analgesic, antitumor, and α-glucosidase inhibitory activities., Kumarappan et al. reported an extensive review on complete ethnomedical and ethnopharmacological properties of various parts of I. frutescens, wherein traditional usage to analysis of individual component with pharmaceutical properties has been described.
In light of above information, the present investigation was designed to evaluate the effect of the active fraction obtained from the methanolic extract of I. frutescens on hyperglycemia status.
| » Materials and Methods|| |
Drugs and chemicals
Streptozotocin (STZ) and nicotinamide (NA) were procured from Sisco Research Laboratories, Mumbai, India. Fasting blood glucose levels were determined using glucose oxidase-peroxidase reactive strips from Agappe Diagnostics Pvt. Ltd., Kerala, India, and insulin assay kit was purchased from BARC, Mumbai, Maharashtra, India. All other chemicals and solvents were of highest analytical grade.
The fresh leaves, stems, and flowers of I. frutescens (L.) were collected from areas surrounding Mysore (Karnataka, India) in November 2014 and preserved at JSS College of Pharmacy, Mysore, India (accession number: Jsscp-Pcog-18). No specific permission was obtained for the collection of the plant material as the plant is available in plenty around Mysore, and it is not an endangered or protected species. The location is not privately owned or protected in any way. The leaves, stems, and flowers were dried under shade and powdered by the help of mechanical process. The coarse powder was stored in an airtight container for further studies. Further phytoconstituents were extracted according to the method described by Markhan. Briefly, 100 g of dry powder was extracted using 500 mL of methanol and filtered through Whatman No. 1 filter paper, and the solvent containing the phytochemical constituents was concentrated using a rotary flash evaporator. To this, methanol: hexane mixture was added in 2:1 ratio and centrifuged at 2000 rpm for 10 min, to obtain brown-colored precipitate. This active fraction dissolved in water was used for feeding the experimental rats at desired concentration.
Animal care and ethical approval
In the present study, 10-week-old male Wistar albino rats, weighing about 150–170 g b.w., were used in the present study. Animals were obtained from the Central Animal Facility, University of Mysore, Mysore, and acclimatized to the laboratory conditions for 2 weeks. All animals received regular human care. They were randomly distributed into different groups consisting of six per cage and fed standard laboratory diet provided by the Central Animal Facility, University of Mysore, Mysore. Clean drinking water (ad libitum) was supplied throughout the study period. The room temperature was maintained at 22°C ± 3°C and the relative humidity at 30%–70%, with 12-h light and dark cycle. Experiments were complied with the rulings of the “Committee for the Purpose of Control and Supervision of Experiments on Animals” (CPCSEA) Mysore, India (122/GO/ReBi/1999/CPCSEA dated June 3, 2015). The study was permitted by the Institutional Ethical Committee of Mysore, India (UOM/IAEC/15/2016). The investigation was carried under the supervision of expertise in animal handling and care.
Oral glucose tolerance test in nondiabetic rats
The oral glucose tolerance test was executed in nondiabetic rats according to the method described by Barik et al. The rats were fasted overnight (16 h) before the test. Fasting blood glucose level in each rat was tested before the test. Overnight-fasted rats (n = 24) were divided into four groups. Control (Group 1) was provided with an equal volume of distilled water. Group 2 and 3 rats were administered with active fraction at doses of 25 and 50 mg/kg b.w. through oral gavage. Group 4 rats were fed with glibenclamide at a dose of 25 mg/kg b.w. Glucose (2 g/kg b.w.) was fed 30 min after the administration of extracts. Blood was drawn from the retro-orbital plexus at 30, 60, 90, and 120 min of extract administration, and plasma glucose level was determined by a blood glucose meter. All the data were expressed as the average level in six experimental animals in one group.
Induction of diabetes
Noninsulin-dependent diabetes mellitus (Type-2) was induced by intraperitoneal injection of NA at 230 mg/kg b.w. in saline. After 15 min, a freshly prepared STZ at a concentration of 65 mg/kg b.w. dissolved in 0.1 M citrated buffer (pH 4.5) was intraperitoneally injected. After 8 h of STZ-NA administration, the rats were kept on 15% glucose solution bottles for the next 24 h in their cages to prevent hypoglycemia., After 48 h of STZ-NA administration, the diabetic state was assessed by measuring the fasting blood glucose level using a glucometer (Glucocard Vital Strip Method). The rats with serum glucose above 230 mg/dL, as well as with polydipsia, polyuria, and polyphagia, were selected and equally distributed into different groups, except to G1 and G6.
A total of 36 rats were divided into six groups as follows:
- Group I: Normal untreated rats
- Group II: Diabetic untreated rats
- Group III: Diabetic rats treated with 25 mg active fraction/kg b.w./day for 28 days
- Group IV: Diabetic rats treated with 50 mg active fraction/kg b.w./day for 28 days
- Group V: Diabetic rats treated with 25 mg glibenclamide/kg b.w./day for 28 days
- Group VI: Normal rats treated with 50 mg active fraction/kg b.w./day for 28 days.
Active fraction of I. frutescens was dissolved freshly in 0.5% carboxy methyl cellulose to get the desired concentration as per the dose level and administered to animals daily through oral route by gavage for a period of 28 days. During the study, daily feed intake and weekly b.w. variations were monitored for all the experimental rats.
Analysis of hematological parameters
During the experimental trial, blood samples were collected humanely from rats treated with mild ether anesthesia by retro-orbital plexus puncture method using a fine capillary tube. Blood was collected in tubes containing dipotassium ethylene di-amide tetra acetic acid anticoagulant and without anticoagulant for clinical chemistry. The blood samples collected in the tubes without anticoagulant were centrifuged at 3000 rpm for 10 min to obtain serum. Fasting blood glucose levels were checked using the glucometer (Glucocard Vital Strip Method) from all the animals on day 1 (48 h after the STZ administration and before the test sample administration), 7, 14, 21, and 28 days of the study period. Blood plasma was recovered for the determination of plasma insulin levels. Glycosylated hemoglobin was estimated according to the method described by Sudhakar and Pattabiraman.
On completion of the study period, on day 28, overnight-fasted rats were humanely sacrificed by exposing them to excess carbon dioxide in a gas chamber. Blood was collected by a cardiac puncture, and the serum was separated for the estimation total cholesterol (TC), triglycerides (TGs), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and very LDL (VLDL) using commercially available kits (Agappe, Kerala, India). Assay kits from Agappe, Kerala, India, was used for the estimation of serum glutamic oxaloacetic transaminase (SGOT) and serum glutamic pyruvic transaminase (SGPT). Pancreas was carefully observed for any lesions and collected for histopathological studies. Small weighed portion of liver was immediately processed for the measurement of hepatic glycogen levels.
Excised pancreas and liver samples were washed in ice-cold normal saline, patted dry, and immediately preserved in 10% Neutral buffered formalin (NBF). They were processed in an automatic tissue processor and embedded in paraffin wax. Sections of 5 μm were cut and stained with hematoxylin and eosin, and later, the microscopic slides were photographed using a light microscope.
The raw data obtained from the present study were subjected to one-way analysis of variance with Duncan multiple range test for the data on b.w., and clinical chemistry parameters were analyzed using GraphPad Prism Software, Inc (version 5.01). All analyses and comparisons were evaluated at the 95% level of confidence (P < 0.05). The data generated were compared with the control group animals.
| » Results|| |
The oral glucose tolerance test was carried out to study the effect of active fraction on glucose metabolism. Administration of distilled water and glibenclamide was considered as negative and positive control, respectively. Animals treated with 25 and 50 mg/kg b.w. of extracts and glibenclamide (25 mg/kg b.w.) showed a decrease in blood glucose level (149.33 mg/dL, 136.5 mg/dL, and 128.5 mg/dL, respectively) when compared with the control group (151.33 mg/dL) in 30 min after the administration of glucose. After 1 h, a noticeable decrease was observed in groups treated with active fraction and glibenclamide, which shows that they have played a significant role in synthesis of glycogen from glucose. After 120 min, blood glucose level of rats treated with active fraction at 50 mg/kg b.w was approximately similar to the control group (81.5 mg/dL) without causing a hypoglycemic state.
Hypoglycemic activity in normal and streptozotocin-induced diabetic rats
According to the results obtained, 2.39-fold increase (P < 0.05) in the fasting blood glucose levels was observed in STZ-NA-induced diabetic untreated rats when compared to diabetic rats treated with active fraction at 50 mg/kg b.w. [Table 1]. Oral administration of the purified fraction at 25 and 50 mg/kg b.w. to the diabetic rats gradually lowered the blood glucose level after the 7th day of administration, reaching a level of 192.5 and 166.0 mg/dL, respectively, after 28th day. The results indicate 40.18% and 55.98% fall in blood glucose levels of diabetic rats on administration of 25 and 50 mg/kg b.w. of purified fraction, respectively. Glibenclamide-treated diabetic rats showed 63.03% fall in blood glucose levels at 25 mg/kg b.w.
Effect of purified fraction on body weight of normal and diabetic rats
The summary of b.w. (g) of experimental animals is presented in [Table 2]. Diabetic untreated rats showed significant (P < 0.05) reduction in b.w. when compared to normal control rats. In glibenclamide-fed positive control rats, b.w. increased from 157.5 g to 167.08 g in 28 days of the experimental period. Similarly, on administration of active fractions, a gradual increase in b.w. was observed in diabetic-induced rats. Normal rats fed with active fraction showed b.w. almost similar to control rats.
Effect on plasma insulin, glycosylated hemoglobin, and hepatic glycogen levels
[Figure 1]a depicts the plasma insulin levels of the control and experimental groups of rats. There was a significant variation in the plasma insulin levels of diabetic control group as compared to normal rats. After 28 days, plasma insulin levels of diabetic control rats were almost 2.7 times lesser than the control rats. After treatment with active fraction at 25 and 50 mg/kg b.w., plasma insulin levels of diabetic rats were increased from 7.18 ± 0.32 to 13.39 ± 0.51 IU/ml and 7.39 ± 0.54 to 15.91 ± 0.47 IU/ml, respectively. Treatment of diabetic rats with glibenclamide at 25 mg/kg b.w. results in the increase of plasma insulin levels from 7.38 ± 0.45 to 17.81 ± 0.67 IU/ml.
|Figure 1: (a) Plasma insulin level, (b) glycosylated hemoglobin, (c) hepatic glycogen level. Values are mean ± standard deviation (n = 6). Mean values with different lowercase letter are significantly different (P < 0.05)|
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Further, glycosylated hemoglobin (HbA1c) (%) level was significantly elevated in diabetic rats (8.6%) as compared to control (6.6%) [Figure 1]b. In experimental rats fed with glibenclamide and the active fraction of I. frutescens, HbA1c (%) level was reduced as compared to diabetes-induced rats.
The effects of oral administration of purified fraction for 28 days on hepatic glycogen levels of control and experimental groups of rats are depicted in [Figure 1]c. A significant decline in glycogen levels was observed in the liver tissue of diabetic control rats. Restoration of glycogen levels almost to the near-normal levels were observed in diabetic rats treated with purified fraction at 50 mg/kg b.w. and glibenclamide. Significant variations were not found in normal rats treated with purified fraction alone.
Effect on serum lipid profile
Analysis of serum lipid profile in control rats showed the normal range of HDL, LDL, TG, and TC. On induction of diabetes, TC significantly (P < 0.05) increased from 102.5 to 138.0 mg/dL. Similarly, LDL, VLDL, and TG levels were also found to elevate on induction of diabetes, while HDL level decreased from 49.67 to 39.83 mg/dL. Treatment of diabetic rats with active fraction and glibenclamide resulted in the significant increase in HDL-cholesterol (HDL-c) level and decrease in elevated TC, TG, and LDL cholesterol (LDL-c) level, when compared to diabetic rats, which is explained by the increase in plasma insulin level after treatment [Table 3].
Effect on serum glutamic oxaloacetic transaminase and serum glutamic pyruvic transaminase
As per the results obtained, a significant increase in serum SGOT and SGPT levels was observed in STZ-induced diabetic control rats [Table 3]. In glibenclamide-fed positive control rats, SGOT and SGPT levels increased in diabetic rats. Similarly, administration of active fraction at 50 mg/kg b.w. to the diabetic rats for 28 days maintained SGOT and SGPT levels in normal range [Table 3].
In the normal control rats, the Islets of Langerhans More Details in pancreas depicted normal acini and normal cellular population [Figure 2]a. However, in diabetic control rats, minute and reduced number of islet cells was observed [Figure 2]b. Pancreatic section from diabetic rats administered with active fraction showed the regeneration of the normal size and cell count of islet cells, especially in the central β-cell region [Figure 2]c and [Figure 2]d. Administration of glibenclamide at 25 mg/kg b.w. also resulted in the restoration of normal cell count and size of islets with hyperplasia [Figure 2]e. In the normal group administered with active fraction (group 6) no significant change was observed [Figure 2]f.
|Figure 2: Histopathological section of pancreas of representative rats from respective group (H and E). (a) Group 1 (×10); (b) Group 2 showing vacuolar change minimal (arrows) (×40); (c) Group 3 showing exocrine area, mononuclear cell infiltration (arrows) minimal (×10); (d) Group 4 (×10); (e) Group 5 (×10); (f) Group 6 (×10)|
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| » Discussion|| |
The incidence of diabetes has continued to prevail despite large number of discoveries and invention of newer drugs to treat or prevent the condition. The persistent hyperglycemia causes long-term dysfunction and damage of various organs, especially the tissues requiring insulin for glucose uptake. Management of diabetes is therefore a major challenging task because of adverse effects associated with synthetic drugs. Over the years, many plants are being generally used to treat diabetes mellitus. Plants are known to exhibit hypoglycemic, hypolipidemic, and antioxidant activity due to the presence of flavonoids, ellagic acids, phenolic acids, phytosterols, gallotannins, and other related polyphenols. Earlier literature reveals the utilization of I. frutescens leaves in the treatment of diabetes and jaundice. However, scientific validation for their therapeutic efficacy is limited. In this context, the present investigation was carried out to study the anti-diabetic effect of I. frutescens in diabetic-induced albino Wister rats.
Oral glucose tolerance test was carried out initially to provide the evidence that active fraction obtained from I. frutescens has the ability to lower blood glucose, especially in normal glycemic rats. Accordingly, the results clearly indicate that after 120 min, blood glucose level of rats treated with active fraction was similar to the control group without causing any hypoglycemic state. Polyphenol extract of I. frutescens has shown to reduce blood glucose within 1 h at a concentration of 200 mg/kg.
In the present investigation, diabetes in rats was induced by STZ which is a nitrosourea complex obtained from Streptomyces achromogenes which induces DNA strand breakage causing discerning obliteration of β-cells of the islets of Langerhans, is less toxic than alloxan, and also allows a consistent maintenance of diabetes. Oral administration of the purified fraction to the diabetic-induced rats showed 40.18% and 55.98% reduction in blood glucose levels, indicating hypoglycemic activity of active fraction. Similarly, Barik et al. evaluated the aqueous root extract of I. frutescens and reported its significant effect in reducing fasting blood sugar level in STZ-NA-induced diabetic rats on the 10th and 15th day. Dash et al. reported significant (P < 0.01) reduction in the blood glucose levels on administration of chloroform and methanol extract of I. frutescens whole plant in STZ-induced diabetic rats at the dose of 200 and 400 mg/kg b.w. However, in the present study, 50 mg/kg b.w. of purified fraction of I. frutescens is found to reduce 55.98% of glucose level as compared to diabetic-induced rats. Ethanol extract of Symplocos cochinchinensis in STZ-induced diabetic model resulted in attenuation of postprandial hyperglycemia, compared to diabetic control animals. They reported the efficiency of S. cochinchinensis extract in inhibiting alpha-glucosidase that modifies sucrose hydrolysis in small intestine in normal and diabetic rats, thus resulting in reducing sucrose level.
Hypoglycemic effect of certain plants has been attributed to insulin effect either by increasing the pancreatic secretion of insulin from β-cells of islets of Langerhans or its release from bound insulin. Some plants extracts are known to inhibit hepatic glucose production or correct insulin resistance.
Further, the administration of purified fraction to diabetic-induced rats showed the significant increase in b.w., which indirectly suggests to beneficial potential of active fraction of I frutescens. In people suffering from diabetes mellitus, excessive loss in b.w. is observed especially by depletion of body proteins. Wasting of protein-energy accompanying high glucose level has been attributed to change glucose metabolism. In diabetes mellitus, cells become inefficient to utilize glucose as energy source which leads to increased consumption and decreased storage of protein, leading to consequent loss in b.w. of diabetic patients. When glucose level of the blood exceeds the renal threshold, the kidney removes the excess sugar from the blood and excretes it in the urine. In the present study, there was a marked reduction in the b.w. of diabetic rats. This condition was alleviated by the treatment of the diabetic rats with administration of purified fraction. The efficacy of purified fraction in preventing loss of b.w. could be as a result of its ability to increase the plasma insulin levels, thereby improving glycemic control mechanisms.
Inefficiency of pancreatic β-cells to balance the plasma insulin directs to increase concentration of glucose, leading to diabetes. Insulin is secreted in response to high blood sugar levels. It balances glucose output from the liver and glucose uptake in the skeletal muscle and adipose tissue. In the present study, diabetic rats treated with glibenclamide showed an increase in plasma insulin level compared to the diabetic control rats. A significant rise in the secretion of plasma insulin levels was observed in diabetic rats treated with active fraction at 50 mg/kg b.w. for 28 days. This suggests that the active fraction might trigger the secretion of insulin from the existing β-cells of the pancreas and might regenerate the β-cells of the pancreas destructed by STZ. An ideal antidiabetic drug alleviates the condition of hyperglycemia and improves pancreatic cell function, while not causing hypoglycemia. Accordingly, normal rats treated with active fraction at 50 mg/kg b.w. resulted in slight raise in the plasma insulin levels from 15.25 ± 0.32 to 16.04 ± 0.72 IU/ml.
Further, a significant elevation of HbA1c (%) was observed in diabetic rats as compared to control. The excess amount of glucose present in the blood of diabetic patients reacts with hemoglobin to form HbA1c. The increase in the level of HbA1c is directly proportional to the concentration of glucose present in the blood, and the level of HbA1c decreases with improvement in glycemic control mechanisms. In the management and prognosis of diabetes, several investigators have suggested HbA1c be used as the most reliable marker. Our study gave a clear view that the oral administration of purified fraction and glibenclamide significantly decreases HbA1c level possibly due to normoglycemic control mechanisms, which also reflect the decreased protein glycation condensation reactions.
Glycogen level in various tissues reflects the insulin activity as it enhances intracellular deposition of glycogen by stimulating glycogen synthase and suppressing glycogen phosphorylase. Hence, the significant reduction of hepatic glycogen level in diabetic rats as compared to control rats can be attributed to the deficiency of insulin. On oral administration of purified fraction and glibenclamide, remarkable increase in hepatic glycogen content was observed, suggesting the role of purified fraction in stimulating or regeneration of β-cells of islets of Langerhans in the pancreas to secrete insulin. The result signifies the efficiency of purified fraction in controlling diabetes by improving glucose metabolism and reestablishing normal blood glucose level.
Diabetes is often associated with hyperlipidemia and hypertriglyceridemia. Major risk factor in the development of coronary heart disease is dyslipidemia, and its prevalence is 95% in diabetic patients. Under diabetic condition, excess mobilization of fat occurs from the adipose tissue due to underutilization of glucose which results in hyperlipidemia. The rise in blood sugar is accompanied with the increase in TC, TGs, and VLDL and fall of HDL. These possible effects may be due to influence of insulin in control of lipolysis or alteration in the activity of cholesterol biosynthesis enzymes. Similarly, polyphenolic extract of I. frutescens leaves at a dose of 300 mg/kg has shown to exhibit antihyperlipidemic effects while at the same time increasing HDL-c.
Administration of active fraction to diabetic rats had significant effect on serum lipid profile. An increased level of HDL-c and a decrease in elevated TC, TGs, and LDL-c level were observed compared to diabetic control rats, which is explained by the increase in plasma insulin level after treatment. This rise in plasma insulin level activates the lipoprotein lipases and thus hydrolysis of stored TGs into free fatty acids. Lowered TG levels observed in the present study are of significant importance and show the efficiency of active fraction in stimulating lipase bound to endothelium which hydrolyzes the TGs into fatty acids. Further, TC, TGs, VLDL, and HDL are biomarkers of hyperlipidemia and atherosclerosis. Hence, the significant reduction in VLDL and increase in HDL strengthen the hypolipidemic effect of active fraction.
SGOT and SGPT are the biomarkers for liver functioning. As per the results obtained, a significant increase in serum SGOT and SGPT levels was observed in STZ-induced diabetic control rats. This might be due to the leakage of these marker enzymes from the liver cytosol into the blood circulation; it represents the toxicity of STZ on the liver. Since liver is the vital organ which is responsible for glucose and lipid homeostasis in the body, diabetes is attributed to liver dysfunction. Hepatic damage is associated with necrosis of liver cells, lipid peroxidation, and elevation in the serum biochemical parameters such as SGOT and SGPT. Administration of active fraction at 50 mg/kg b.w. to the diabetic rats for a period of 28 days maintained SGOT and SGPT levels in normal range. This indicates the ability of purified fraction to normalize liver cells, causing parenchymal cell regeneration, thus providing fortification against fragility of membrane and preventing the leakage of these marker enzymes. Our study clearly suggested the hepatoprotective action of purified fraction.
Histological studies of vital organs are one of the best methods to understand the significant effect of administered compound. Since pancreas and liver are the major organs affected during diabetes, these vital organs were studies for the morphological and physiological changes. According to the results obtained, minute and reduced number of islet cells was observed in diabetic-induced rats. However, experimental rats administered with purified fraction showed the regeneration of the normal cell count and size of islet cells, especially in the central β-cell region. Quiescent β-cells are stable and formed by neogenesis or by replication of the preexisting differentiated cells. Treatment of diabetic rats with purified fraction and glibenclamide results in the regeneration of the β-cells might be due to the prevention of free radical formation, which is in agreement with the reports of other plant extracts having β-cell regenerative potential.
| » Conclusion|| |
The results of this study showed that active fraction obtained from methanolic extract of I. frutescens possessed antidiabetic properties as shown in its ability to reduce blood glucose level of STZ-induced diabetic rats. This confirmation justifies its use in ethnomedical medicine for the treatment of diabetes. Further studies should be undertaken to identify the active antihyperglycemic compound. Comprehensive chemical and pharmacological investigation should be carried out to isolate the active compound and appropriate elucidation of its mechanism of action. The result suggests that it is worth undertaking further studies on possible usefulness of the I. frutescens in managing diabetes mellitus.
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Conflicts of interest
There are no conflicts of interest.
| » References|| |
Lopez AD, Mathers CD. Measuring the global burden of disease and epidemiological transitions: 2002-2030. Ann Trop Med Parasitol 2006;100:481-99.
Biessels GJ, Staekenborg S, Brunner E, Brayne C, Scheltens P. Risk of dementia in diabetes mellitus: A systematic review. Lancet Neurol 2006;5:64-74.
Ramírez G, Zavala M, Pérez J, Zamilpa A.In vitro
screening of medicinal plants used in Mexico as antidiabetics with glucosidase and lipase inhibitory activities. Evid Based Complement Alternat Med 2012;2012:701261.
Bailey CJ, Day C. Metformin: Its botanical background. Pract Diabet Int 2004;21:115-7.
Rajagopal K, Sasikala K. Antihyperglycaemic and antihyperlipidaemic effects of Nymphaea stellata
in alloxan-induced diabetic rats. Singapore Med J 2008;49:137-41.
Verma RK, Singh N, Gupta MM. Triterpenoids of Ichnocarpus frutescens
. Fitoterapia 1987;L8:271-2.
Savithramma N, Yugandhar P, Suhrulatha D. Traditional medicinal plants used by local people of Kailasakona – A sacred grove of Chittoor district, Andhra Pradesh, India. Int J Pharm Pharm Sci 2015;7:408-11.
Barik R, Jain S, Qwatra D, Joshi A, Tripathi GS, Goyal R, et al.
Antidiabetic activity of aqueous root extract of Ichnocarpus frutescens
in streptozotocin-nicotinamide induced type-II diabetes in rats. Indian J Pharmacol 2008;40:19-22.
] [Full text]
Dash DK, Ghosh T, Yeligar VC, Murugesh K, Nayak SS, Maiti BC, et al
. Effect of Ichnocarpus frutescens
extract on antihyperglycemic, antihyperlipidemic and antioxidants status in streptozotocin-induced diabetic rats. Orient Pharm Exp Med 2007;7:244-53.
Pandurangan A, Khosa RL, Hemalatha S. Anti-inflammatory and analgesic activity of Ichnocarpus frutescens
. Pharmacol Online 2008;1:392-9.
Kumarappan C, Srinivasan R, Jeevathayaparan S, Rajinikanth R, Naveen Kumar HS, Senthilrajan S, et al
. Ichnocarpus frutescens
: A valuable medicinal plant. Pharmacol Online 2015;19:18-37.
Markhan KR. Hydrolysis and analysis of glycosides. In: Techniques of Flavonoid Identification. London, UK: Academic Press; 1982. p. 52-7.
Kavishankar GB, Lakshmidevi N. Antidiabetic effect of a novel N-trisaccharide isolated from Cucumis prophetarum
on streptozotocin-nicotinamide induced type-2 diabetes rats. Phytomedicine 2014;21:624-30.
De Moura LP, Gomes RJ, Leme JA, de Araujo MB, de Mello MA. Hepatic steatosis markers in diabetic rats trained at the aerobic/anaerobic transition. J Liver 2013;2:137.
Sudhakar NS, Pattabhiraman TN. A new colorimetric method for the estimation of glycosylated haemoglobin. Clini Chim Acta 1981;109:267-74.
Muruganandan S, Srinivasan K, Gupta S, Gupta PK, Lal J. Effect of mangiferin on hyperglycemia and atherogenicity in streptozotocin diabetic rats. J Ethnopharmacol 2005;97:497-501.
Bhandary MJ, Chandrashekar KR, Kaveriappa KM. Medical ethanobotany of the Siddis of Uttara Kannada district, Karnataka, India. J Ethanopharmacol 1995;47:149-58.
Anbu J, Nithya S, Kannadhasan R, Kishore S, Anjana A, Suganya S. Antioxidant and protective effect of aqueous extract of Ichncarpus frutescens
and Cyperus rotundus
against cisplatin induced testicular toxicity in rodents. Int J Pharm Pharm Sci 2012;4:437-41.
Arunachalam K, Parimelazhagan T. Antidiabetic activity of aqueous root extract of Merremia tridentata
(L.) Hall. F. In streptozotocin-induced-diabetic rats. Asian Pac J Trop Med 2012;5:175-9.
Dash DK, Yeligar VC, Nayak SS, Ghosh T, Rajalingam D, Sengupta P, et al
. Evaluation of hepatoprotective and antioxidant activity of Ichnocarpus frutescens
(Linn.) R. Br. on paracetamol-induced hepatotoxicity in rats. Trop J Pharma Res 2007;6:755-65.
Antu KA, Riya MP, Mishra A, Anilkumar KS, Chandrakanth CK, Tamrakar AK, et al.
Antidiabetic property of Symplocos cochinchinensis
is mediated by inhibition of alpha glucosidase and enhanced insulin sensitivity. PLoS One 2014;9:e105829.
Pari L, Amarnath Satheesh M. Antidiabetic activity of Boerhaavia diffusa
L.: Effect on hepatic key enzymes in experimental diabetes. J Ethnopharmacol 2004;91:109-13.
Eddouks M, Jouad H, Maghrani M, Lemhadri A, Burcelin R. Inhibition of endogenous glucose production accounts for hypoglycemic effect of Spergularia purpurea
in streptozotocin mice. Phytomedicine 2003;10:594-9.
Rajasekar R, Manokaran K, Rajasekaran N, Duraisamy G, Kanakasabapathi D. Effect of Alpinia calcarata
on glucose uptake in diabetic rats-an in vitro
and in vivo
model. J Diabetes Metab Disord 2014;13:33.
Monnier VK, Cerami A. Non-enzymatic glycosylation and browning in diabetes and aging. Diabetes 1982;31:57-66.
Golden S, Wals PA, Okajima F, Katz J. Glycogen synthesis by hepatocytes from diabetic rats. Biochem J 1979;182:727-34.
Uttra KM, Devrajani BR, Ali Shah SZ, Devrajani T, Das T, Raza S, et al
. Lipid profile of patients with diabetes mellitus (A multidisciplinary study). World Appl Sci J 2011;12:1382-4.
Sharma SB, Nasir A, Prabhu KM, Murthy PS, Dev G. Hypoglycaemic and hypolipidemic effect of ethanolic extract of seeds of eugenia jambolana in alloxan-induced diabetic rabbits. J Ethnopharmacol 2003;85:201-6.
Kumarappan CT, Rao TN, Mandal SC. Polyphenolic extract of Ichnocarpus frutescens
modifies hyperlipidemia status in diabetic rats. J Cell Mol Biol 2007;6:175-87.
Ramachandra Setty S, Quereshi AA, Viswanath Swamy AH, Patil T, Prakash T, Prabhu K, et al.
Hepatoprotective activity of Calotropis procera
flowers against paracetamol-induced hepatic injury in rats. Fitoterapia 2007;78:451-4.
Govan AT, Macfarlane PS, Callander R. Pathology Illustrated. 2nd
ed. Edinburgh, New York: Churchill Livingstone; 1986.
Gupta R, Mathur M, Bajaj VK, Katariya P, Yadav S, Kamal R, et al.
Evaluation of antidiabetic and antioxidant activity of Moringa oleifera
in experimental diabetes. J Diabetes 2012;4:164-71.
[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3]