Phytochemical investigation of the stems of Tetracera indica Merr. and in-vitro antidiabetic evaluation of isolated compounds

Abstract

Tetracera indica Merr. (Family: Dilleniaceae) is a large, woody, rain forest climber species. It is found throughout the Malaysia and is locally and commonly known as Mempelas paya or sand paper plant. Aerial parts of T. indica have been traditionally used to cure different disorders including diabetes and its related infirmities in Malaysia. The traditional claims of this plant as an antidiabetic agent have not been properly scientifically evaluated. Hence, the aim of this study was to explore in vitro antidiabetic potential of the stems of T. indica as well as to isolate the compounds responsible for its in vitro antidiabetic property. The in vitro antidiabetic activity investigation of the stems ethanol extract, sub-fractions and isolated compounds from the bioactive fraction was carried out on 3T3-L1 pre-adipocytes and adipocytes. Upon thorough investigations, four flavonoids (MHQ1-Wogonin, MHQ-2-Norwogonin, MHQ-3-Quercetin, MHQ-4-Techtochrysin) and two terpenoids (MHQ-5-Stigmasterol, MHQ-6-Betulinic acid) were isolated from the stems ethanol extract of T. indica and further subjected to cytotoxicity test against 3T3-L1 adipocytes with regard to evaluate their antidiabetic potential. All the compounds were isolated and purified through silica gel and sephadex LH-20 repeated column chromatography and recrystallization with different solvents. Their structures were elucidated through 1H-and 13C-NMR spectroscopy. Cytotoxicity test was performed through MTT assay on 3T3-L1 pre-adipocytes to determine the safe dose of the extract, sub-fractions and all the isolated compounds for further in vitro antidiabetic evaluation on 3T3-L1 pre-adipocytes. The stems ethanol extract, sub-fractions and isolated pure compounds i.e. MHQ1, MHQ2, MHQ3 and MHQ4 were further subjected to adipogenesis to investigate insulin like activity or insulin sensitizing activity for the purpose of evaluating antidiabetic potential. All compounds were introduced to the cells in different safe concentrations as well as in different adipogenic cocktails. The adipogenic cocktails were modified by the addition of compounds to be investigated and rosiglitazone in the presence or absence of insulin. Results vividly showed that MHQ-1, MHQ-2 and MHQ-4 induced adipogenesis like insulin and enhanced adipogenesis like rosiglitazone significantly. Furthermore, MHQ1, MHQ2 and MHQ-4 as well as rosiglitazone as positive control were subjected to fluorescence glucose uptake test by 2-NBDG (fluorescent glucose analogue) on mature adipocytes. Results suggested significant glucose uptake activity by MHQ1, MHQ2 and MHQ-4. It is concluded and suggested that the further in-depth research study on the isolated compounds might help to discover a new safe compound with strong antidiabetic activity. Results further confirm the traditional use of T. indica in the management of diabetes in Malaysia.