HYPOGLYCEMIC AND HEPATOPROTECTIVE EFFECT OF MERREMIA VITIFOLIA IN MICE
HTML Full TextHYPOGLYCEMIC AND HEPATOPROTECTIVE EFFECT OF MERREMIA VITIFOLIA IN MICE
Kawser Ahmed, Mohammad Hasanuzzaman and Prodip Kumar Baral *
Department of Pharmacy, Noakhali Science and Technology University, Noakhali, Bangladesh.
ABSTRACT: Traditionally, Merremia vitifolia meets multipurpose medicinal uses in tribal areas. Therefore, the study worked on hypoglycemic and hepatoprotective effects on animal models to meet the research gap on this plant. Methanolic stem extract at 200 mg/kg and 400mg/kg were administrated orally to determine the effects on blood glucose and hepatic enzymes. The highest dose showed a significant (p < 0.01) reduction in blood glucose. Maximum glucose level reduction (52.1%) was observed at 400 mg/kg extract with 3.5±0.6 mmol/L after 3 h, while reference glibenclamide (10mg/kg) reduced 25.2% with 4.4±0.8 mmol/L. The plasma levels of alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase of CCl4 intoxicated mice were 108.8, 43.4, and 268.8 IU/L at 400 mg/kg, where the values for reference silymarin (100 mg/kg) were 103.4, 54.3, and 237.7 IU/L, respectively. The reduction of alkaline phosphate and alanine aminotransferase levels were moderately (p<0.01) and minimally (p<0.05) significant at concentration 400 mg/kg, respectively. Therefore, the study explored the plant's potential hypoglycemic and hepatoprotective effects, where the hypoglycemic effect was strongly significant.
Keywords: Merremia, Hypoglycemic, Hepatoprotective, α-glucosidase, Alanine aminotransferase
INTRODUCTION: Traditional herbal medicine has a long history to use for hypoglycemic effects and hepatoprotective effects all over the world 1, 2. A study describes the uses of 800 and 150 plants in Mexico and India for hypoglycemic effects, respectively. It is considered that the plant metabolites contain active compounds that reduce blood glucose levels by the following mechanisms, such as stimulating endogenous insulin release or drop of sugar absorption or improving insulin sensitivity 3.
On the other hand, traditional natural products convey any probable modes of hepatoprotection, including immunomodulation, stimulation of reductive enzymes, modulation of hepatic DNA synthesis, lipid accumulation Suppression, and reduction of hepatic fibrosis 4, 5.
A study included 509 species belonging to 140 genera that exhibit significant anti-diabetic activity 6. Ficus benghalensis 7, Momordica charantia 8, Ficus hispida 9, Ficus elastic 10 and Pterocarpus marsupium 11 are some prominent traditional medicines used for the treatment of diabetes. Another study enlisted 100 medicinal plants clinically proven for hepatoprotective effects 12 and some of the significant plants are Phoenix dactylifera 13, Aquilaria agallocha 14 and Salix caprea 15.
The genus Merremia of over 100 plants belongs to family Convolvulaceae and is an abundant source of structurally varied phytochemicals with medicinal potential 16. Additionally, its species meet nutrition values up to a safety level and are prohibited from chronic and prolonged administration to humans as well as animals 17. Some therapeutically valuable Merremia plants are M. borneensis, M. emarginata, M. mammosa, M. peltate, M. tridentate, and M. vitifolia 18–20. The plants provide a wide spectrum of activities, including antimicrobial, analgesic, anticancer, antidiabetic, hepatoprotective activities. Specifically, M. mammosa and M. tridentate for antidiabetic activity 21, 22, M. emarginata for hepatoprotective action 23. Merremia vitifolia is a perennial and climbing plant widely distributed in South and East Asia. Its leaf and rhizome are mainly used in tribal areas to treat fever, inflammation, rheumatism, jaundice, dysentery, and urinary diseases 24–26. Nevertheless, only a few studies on the plant are available. A recent study revealed the substantial antioxidant, thrombolytic, and anti-nociceptive potential of the methanolic leaf extract 27. However, no work on hypoglycemic effects was determined on any animal model. Moreover, no evidence of hepatotoxicity or hepatoprotective was presented. As chemical constituents metabolized in the liver and are an essential mark for drug safety, our present aim was to investigate the level of hypoglycemic and hepatoprotective effect of value addition on traditional medicine.
MATERIALS AND METHODS:
Chemicals and Drugs: Analytical grade Methanol (Merck, Germany); Standard Drugs: Glibenclamide and Silymarin (Square Pharmaceuticals Limited, Bangladesh); Dextrose (Glaxo SmithKline Bangladesh).
Experimental Animals: Six-seven weeks old Swiss albino mice of both sexes with a mean body weight of 25 ± 5.0 g were procured from Animal House at the Department of Pharmacy, Jahangirnagar University, Savar, Bangladesh. The animals were fed with a commercial rat pellet diet ad libitum during the entire experimental period.
Collection and Identification of the Plant: M. vitifolia was collected from Noakhali Science and Technology University, Sonapur, Noakhali and identified by Bangladesh National Herbarium, Mirpur-1, Dhaka. The identification code was DACB46503.
Preparation of Plant Sample: The collected plant stem was dried in the dark for ten days and ground into a coarse powder. Roughly 500g of powder was produced and stored in an airtight container in at cool, dark, and dry place until extraction commenced.
Cold extraction for M. vitifolia: The powder was soaked in 1500 ml of 80% methanol at room temperature for two weeks, accompanied by occasional shaking and stirring. Then the solution was filtered using a filter cloth and Whitman filter paper every seven days. The rotary evaporation (Yamato Scientific, Japan) at 45⁰C and evaporation at ambient conditions gave nearly 65 g of gummy, concentrate and greenish-black extract. Then, the extract was stored in a refrigerator in a beaker covering it with aluminum foil.
Hypoglycemic Test: A total of four groups of mice (n=10) and equally designed by sex were taken to perform hypoglycemic tests according to the described method with a little bit of modification 28. For Group I (Control), mice were given only distilled water (10 mL/kg of mouse body weight), while Group II was used as the standard Group and treated with glibenclamide (10mg/kg). Group III and IV were treated with 200 mg/kg of stem extract and 400mg/kg body weight, respectively. Fasting blood glucose measurement followed by immediate administration (p.o.) of test samples. After 1 hour, dextrose (2 g/kg) solution was administered to all groups, and blood was collected from the mouse tail vein after 1 h, 2 h, and 3 h of administration. Glucose concentration was measured by Accu-Check electronic glucometer (Roche, Germany).
Hepatoprotective Effect on CCl4-induced Mice: The mice were divided into five groups (n = 10) and followed the method described by Li et al. 29. Group I (negative control) animals were administered a single dose of water (25 ml/kg, p.o.) daily for 7 days and received olive oil (8 ml/kg, i.p.) on day 7. Group II (Toxicant control) received water (25 ml/kg, p.o.) once daily for 7 days and received 0.2% CCl4 in olive oil (8 ml/kg, i.p.) on day 7. Group III (positive control) received standard drug silymarin (100 mg/kg, p.o.) once daily for 7 days and received 0.2% CCl4 in olive oil (8 ml/kg, i.p.) on day 7. Groups IV–V were administered orally 200 & 400 mg/kg of the extract once daily for 7 days, respectively. The Groups III–V animals were simultaneously administered 0.1% CCl4 in olive oil (8 ml/kg, i.p.) on day 7 after 2 h of the silymarin and extract administration. Animals were sacrificed after 24 h of treatment, and blood was collected through cardiac puncture, and the serum was separated by centrifugation at 6000 × g for 15 min. Liver function biochemical markers alkalinephosphatase (ALP) levels, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were evaluated according to the supplier's specifications from the standard kits.
Statistical Analysis: Data are expressed as mean ± standard error of the mean (SEM) and analyzed using the Statistical Package for the Social Sciences (SPSS), version 20.0 software. The difference between group means was analyzed with a one-way analysis of variance (ANOVA) followed by Dunnet ‘t’ test. P <0.05 was considered as statistically significant.
RESULTS:
Hypoglycemic Effects: The blood glucose levels of all groups were nearly 7 mmol/L before 1 hr of administration. In contrast, several changes were observed in all administrations except control Table 1. After 1 hr of administration, both standard and extracts showed a minute decrease of glucose level, and the most minor reduction was exhibited by the standard at 6.5±0.5 mmol/L. In 2nd hour, the standard also exhibited the lowest glucose level, although MV 400 had a substantial fall of that level with 5.6±0.8 mmol/L. Dramatically, in 3rd hour, MV 400 showed a significant decrease (p<0.01) in glucose level, where it surpassed the hypoglycemic effect of glibenclamide (10mg/kg) and reached the maximum reducing figure at 3.5±0.6 mmol/L. At that end point, MV 400 exhibited 52.1 hypoglycemic effect compared to the blood glucose level before administration.
TABLE 1: EFFECT OF METHANOLIC STEM EXTRACT OF M. VITIFOLIA ON BLOOD GLUCOSE LEVEL (MEAN ± SEM)
Treatment | Blood Glucose Concentration (mmol/L) | |||
-1 Hour | +1 Hour | +2 Hour | +3 Hour | |
Control | 7.0±0.5 | 7.1±0.5 | 7.4±0.2 | 7.2±0.3 |
Standard | 7.0±0.4 | 6.5±0.5 | 5.1±0.6 | 4.4±0.8* |
MV200 | 7.3±0.4 | 7.1±0.5 | 7.0±0.4 | 5.7±0.5 |
MV400 | 7.2±0.4 | 7.1±0.6 | 5.6±0.8* | 3.5±0.6** |
Here, MV stands for methanol extract of M. vitifolia; *p<0.05; **p<0.01
Hepatoprotective Test: The results of the hepatoprotective effect of extracts on CCl4- intoxicated rats are shown in Table 2. As indicated from the results, CCl4-intoxicated animals showed an increase in the alkalinephosphatase, alanine aminotransferase, and aspartate aminotransferase levels compared to the negative control group. Treatment of animals with M. vitifolia at 200 and 400 mg/kg, p.o., or silymarin 100 mg/kg, p.o., significantly decreased the level of serum marker enzymes.
TABLE 2: PROTECTIVE EFFECT OF M. VITIFOLIA AND SILYMARIN ON CARBON TETRACHLORIDE (CCL4) INDUCED ELEVATION IN ALKALINEPHOSPHATASE, ALANINE AMINOTRANSFERASE, AND ASPARTATE AMINOTRANSFERASE LEVELS (MEAN ± SEM)
Group | Alkaline phosphatase (IU/L) | Alanine aminotransferase (IU/L) | Aspartate aminotransferase (IU/L) |
Negative Control | 188.2±15.6 | 55.9±10.0 | 274.0±66.8 |
Toxicant Control | 204.2±22.9 | 63.7±14.2 | 327.0±36.1 |
Positive Control | 103.4±21.7** | 54.3±8.3 | 237.7±45.6* |
MV200 | 188.6±27.0 | 54.3±12.4 | 282.7±30.7 |
MV400 | 108.8±15.3** | 43.4±4.0* | 268.8±59.9 |
Here, MV stands for methanol extract of M. vitifolia; *p<0.05;**p<0.01
M. vitifolia at 400 mg/kg exhibited 108.81±15.3 IU/L of ALP concentration, almost a 50% decrease from the toxicant control group. Alanine aminotransferase concentration showed surprising results;400 mg/kg dosehad a value of 43.4±4.0 U/L, whereas positive control provided a higher value of 54.3±8.3 IU/L. Regarding aspartate aminotransferase level, 400 mg dose also showed the least value between the two concentrations, and positive control exhibited the least among all groups. The reduction of alkaline phosphate and alanine aminotransferase levels were moderately (p<0.01) and minimally (p<0.05) significant at concentration 400 mg/kg, respectively.
DISCUSSION: The study determined the hypoglycemic and hepatoprotective effects of the methanolic extract of the stem of M. vitifolia on mice models. Upon administration of the extract with two concentrations, the results showed a significant lowering of blood glucose and liver enzyme levels that marked potential phytoconstituents at the plant metabolite.
Empirically, the people in Luwu, Southern Sulawesi, use M. vitifolia to cure diabetes mellitus (Hasanah et al., 2019). However, the 400 mg/kg dose surpasses the blood glucose level for 10 mg/kg glibenclamide in this study. Although no in vivo activity on hypoglycemic activity is available, one study on n-hexane extract of the plant leaves showed the inhibition of α-glucosidase, which is associated with glucose absorption inhibition by the intestinal epithelium membrane. Phytol, neophytadiene, β-caryophyllene, stigmasterol, γ-sitosterol, and lup-20(29)-en-3-one probably contribute to α-glucosidase inhibitory activity 31. Some plants under the genus Merremia also exhibited hypoglycemic activity. A study confirmed the potent antidiabetic activity of M. emarginata in streptozotocin-induced diabetic rats. The enzymatic activities for carbohydrate metabolism were significantly increased; however, glucose-6-phosphatase, fructose-1, 6-bisphosphatase were significantly decreased, and interestingly it showed the pancreatic β-cells regeneration in this study 32. Another study described that flavonoid-rich fraction of M. tridentata revealed strong hypoglycemic effects in alloxan-induced diabetic mice compared with the references, glibenclamide, and metformin. At the same time, the plant exhibited good α-amylase and α-glucosidase inhibitory activity compared with acarbose 33. The root extract of M. tridentate also possessed significant antidiabetic activity in streptozotocin (STZ)-induced diabetic rats 34. Another Merremia species M.mammosa exhibited wound healing in a diabetic rat model 35.
In the case of hepatoprotective effects against CCl4 intoxicated mice, alkalinephosphatase, alanine aminotransferase, and aspartate aminotransferase levels were decreased significantly. Additionally, the ingestion of the plant extracts reduced comparatively enlarged intoxicated liver size. However, a recent study on a genus of the plant explored that M. tridenta ameliorated the liver damage caused by the CCl4 and reduced serum aspartate aminotransferase, alanine aminotransferase, alkalinephosphatase, acyl carrier protein and total Bilirubin content to normal levels 36. Another species showed a hepatoprotective effect, where four out of ten elevated enzymes level by CCl4 were significantly reduced upon administration of M. borneensis. Moreover, pretreatment with M. borneensis against rats treated with CCl4 showed hepatic enzymatic and non-enzymatic antioxidant molecules in increased level, and histopathological improvement, substantially 37.
One recent study confirmed the isolation of 27 phytochemicals in this plant and some proposed compounds are azedarachin C, stigmastan-3,6-dione, pheophorbide A, and isoquinaldonitrile, 3-methyl-, 2-oxide, however, comprehensive bioactivity testing had not been done on these compounds 25. The present finding must keep a query for extensive investigation on the plant's phytochemicals that exactly impart in these actions.
Limitation of this Study: The study evaluated the pharmacological insight on hypoglycemic and hepatoprotective effects only that is not enough for the plant's therapeutic profiling. It should take more than two doses to determine the dose-response relationship meaningful. Moreover, no hint can be conveyed about the molecules responsible for the two pharmacological aspects.
CONCLUSION: The study is the scientific evidence of hypoglycemic and hepatoprotective activities of M. vitifolia that justify the ethnopharmacological diversity of the plant. Although plant extract exhibited less significant hepatoprotection, it may be highlighted for antidiabetic activity as an alternative medicine.
Declaration:
Ethical Issue: The ethical committee, Faculty of Science of Noakhali Science and Technology University, Bangladesh, had approved the research with the code: NSTU/SCI/EC/2020/132.
Consent for Publication: Not Applicable
Availability of Data and Materials: There has no restriction to sharing any data.
CONFLICTS OF INTEREST: There is no conflict of interest
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How to cite this article:
Ahmed K, Hasanuzzaman M and Baral PK: Hypoglycemic and hepatoprotective effect of Merremia vitifolia in mice. Int J Pharm Sci & Res 2024; 15(8): 2314-19. doi: 10.13040/IJPSR.0975-8232.15(8).2314-19.
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Article Information
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2314-2319
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English
IJPSR
Kawser Ahmed, Mohammad Hasanuzzaman and Prodip Kumar Baral *
Department of Pharmacy, Noakhali Science and Technology University, Noakhali, Bangladesh.
pkb.phar@nstu.edu.bd
12 February 2024
25 June 2024
06 July 2024
10.13040/IJPSR.0975-8232.15(8).2314-19
01 August 2024