In vitro investigation of the anti-hyperglycemic and chemical composition of Heteromorpha arborescens (Spreng.) Cham leaf extracts used in the management of diabetes mellitus

Diabetes mellitus is currently a major threat all over the world. It is a chronic disorder characterized by elevated blood glucose levels and disturbance in carbohydrate, fat and protein metabolism which often times leads to several complications such as, atherosclerosis, diabetic nephropathy, retinopathy and neuropathy. Due to the side effects associated with synthetic oral hypoglycemic drugs, herbal remedies have become the alternative in the management of diabetes mellitus. Heteromorpha arborescens is one of the many plants used as herbal remedies for the management of diabetes mellitus in South Africa. In addition, the roots, barks and leaves are used in other parts of Southern Africa for the treatment of mental problems, asthma, cough, dysentery and tuberculosis. The roots are fed to malnourished children in Botswana and Swaziland. However, despite the high medicinal importance, there is little or no scientificinformation to prove this claim. Therefore, the basis of this study is to determine the chemical composition as well as perform and invitro investigation of the antihyperglycemic potential of H. arborescens leaf extracts to provide information that could validate the ethno-medicinal claims for the use of this plant in the management of diabetes mellitus. Phytochemical contents and antioxidant activity of the leaf extracts were determined. Phytochemical analysis of the acetone, ethanol, aqueous and blanched extracts of H. arborescens leaves indicated that the total phenol content of the extracts ranged between 15.10 mg GAE/g- 42.50 mg QAE/g, proanthocyanidin, 459-8402.1 mg QE/g and the flavonoid content, 109.24- 235.79 mg QE/g. In addition, alkaloids (7.65percent) and saponin (25.33percent) were present in significantamounts. Based on the IC50 values, the ethanol extract exhibited the highest total antioxidant activity (0.0125 mg/mL) with highest inhibition against DPPH and ABTS radicals (0.06 and 0.049 mg/mL respectively). Relatively high antioxidant activity may be attributed to the polyphenolic contents which possess hypoglycemic potentials. Minimum Inhibitory Concentrations (MIC) of H. arborescens leaf extracts against Bacillus pumilus, Staphylococcus epidermidis, Staphylococcus aureus, Escherichia coli and Klebselia pneumoniae was also determined and results indicated considerable antibacterial activity in the acetone, ethanol and blanched extracts with MIC values ranging from 1.563-12.5 mg/mL; however, the aqueous extract was inactive against all the bacteria strains. The antibacterial activity of the leaf extracts could be advantageous against diabetic related infections. The cytotoxicity, anti-obesity and antidiabetic potentials of blanched, aqueous and ethanol extracts of Heteromorpha arborescens (Spreng.) Cham leaves were also investigated. The results revealed that both ethanol and aqueous extracts indicated considerable inhibition against α-glucosidase (IC50 of 627.29 ± 4.33 μg/mL and 576.46 ± 3.21 μg/mL respectively) while the blanched extract showed weak α- glucosidase inhibition (IC50; 855.38 ± 4.29 μg/mL). The aqueous extract showed the best α- amylase inhibition (IC50; 583.74 ± 5.87 μg/mL) among the assessed extracts. However, weak α- amylase inhibition was observed in the ethanol (IC50; 724.60 ± 4.33 μg/mL) and blanched extracts (IC50; 791.63 ± 3.76 μg/mL) of H. arborescens leaves. Some level of glucose utilization in both C3A and L6 cells was also observed for the aqueous and ethanol extracts which may be attributed to the relatively lower toxicity levels present in them, however, glucose utilization was very weak for the blanched extract, which may be due to higher level of cytotoxicity it possessed. Relatively weak lipase inhibition was observed for the ethanol (IC50; 699.3 ± 1.33 μg/mL), aqueous (IC50; 811.52 ± 3.52 μg/mL) and blanched extract (IC50; 1152.7 ± 4.61 μg/mL) as compared to orlistat (IC50; 56.88 ± 0.11 μg/mL). However, there was no reasonable reduction in lipid accumulation observed in all the extract treated cells. The results obtained suggest that H.arborescens leaf extracts can serve as a potential tool for the development of new strategies for the treatment of diabetes and obesity. However, further analysis is required to ascertain its anti- obesity potential. Also, caution should be taken in the use of the plant at high concentrations in order to ensure safety and efficacy. Analyses were also carried out to determine the nutritional and antinutritional constituents of the plant for possible inclusion in the diet of diabetic patients. Proximate analysis revealed the presence of 8.5 percent total ash, 4.92 percent crude fat, 8.41 percent moisture, 15.74 percent crude protein, 21.48 percent crude fiber, 40.95 percent carbohydrates and 271.04 kcal/100g energy value. Mineral analysis showed that H. arborescens leaves are very rich in K, Ca, and Fe. Considerable amounts of Mg, Mn, Na, P, Cu and Zn were also present. Vitamin analysis showed that the plant has a high content of vitamins A, C and E. The anti-nutrients evaluated were phytate, oxalate, saponin, and alkaloids, all of which were below toxic levels except for saponin which was observed in moderately high level. This study revealed that H. arborescens leaves are a good source of nutrients and mineral elements, (with low anti-nutrient content) that are highly beneficial to human health especially in diabetic individuals, therefore, encouraging its possible inclusion as a vegetable. Essential oil composition of fresh Heteromorphaarborescens leaves were also determined by Solvent-Free-Microwave-Extraction (SFME) and Hydrodistillation (HD) methods and the compositions of both mehods were compared in terms of their chemical compositions, yield, CO2 emission and energy consumption. Solvent Free Microwave extraction method indicated higher oil yield of 0.7 mL/200 g (0.35 percent) as compared to 0.59 mL/200 g (0.295 percent), lower energy consumption and CO2 emission as compared to the hydrodistillation method. In conclusion, H. arborescens leaves indicated considerable potential efficacy in the management for diabetes mellitus and may require further structural elucidation and characterization in order to identify the bioactive constituents.Thesis (PhD) -- Faculty of Science and Agriculture, 202