MACROSCOPIC MICROSCOPIC MOLECULAR EVALUATIONS MANGIFERIN CONTENT AND BIOACTIVE POTENTIALS OF MANGIFERA INDICA LEAVES IN THAILAND

Abstract

Mango (Mangifera indica) has been noted that it is one of the most ancient and valuable fruit crop and has been considered to be ‘Ayurveda king of fruit’. It is also found to be the major sources of mangiferin, which has many pharmacological effects, that can be detected in all parts of mango. Mango trees have long been cultivated since the early history of Thailand. Thai people ate mango leaves as vegetables with anti-dysentery and anti-flatulence properties. The leaves, a waste material gained from timming of post-harvest could be used as the good reasonable source of mangiferin. Currently, Thai mangoes have over 174 cultivars have been cultivated. They have confronted with confusions about numerous synonym nomenclatures and needed to be correctly identified. However, neither of previous studies provided any macroscopic, microscopic nor molecular descriptive evidences in combination. There still has little information about mangiferin content in Thai mango leaves as well as their biological activities such as antidiabetic, antimicrobial or anticancer. This study investigated selected seventeen Thai mango cultivars popularly cultivated in Thailand, on macroscopic, microscopic leaf characteristics and their genetic relationships using ISSR markers; in addition, it also evaluated mangiferin content in selected mango leaves and some biological activities such as antidiabetic, antimicrobial and anticancer of mango leaf extract and mangiferin. For selected Thai mango identifications, seventeen Thai mango cultivars, M. caloneura and B. macrophylla were collected throughout Thailand (each of them from three different locations). Macroscopic characters together with their genetic characters had a potential to identify among seventeen Thai mango cultivars as well as microscopic leaf constant number, as a supporting evidence, in combination with macroscopic and molecular characteristics was able to use as a helpful tool for more accurate identification. Fifteen Mangifera indica ‘Okrong’ leaf samples were collected from different locations in Thailand for evaluated mangiferin content. They were determined by TLC-densitometry and TLC-image analysis. TLC quantitation was validated. The TLC plate was developed with a saturated mobile phase; ethyl acetate: methanol: formic acid (3.9 : 6 : 0.1). Mangiferin spots were clearly detected under UV 254 nm. Mangiferin contents were 4.992 ± 1.025 and 4.311 ± 0.987 g / 100 g of dried crude drug, respectively. For antidiabetic activities, yeast α-glucosidase activity (from Saccharomyces cerevisiae) and rat α-glucosidase activity (from intestinal acetone powders from rat) were determined by using 1 mM of p-nitrophenyl-α-D-glucopyranoside (PNPG) as the substrate; while, pancreatic α-amylase activity (from porcine pancreas) using 1 mM of 2-chloro-4 nitrophenol-α-D-maltotroside (CNPG-3) as substrate. The absorbance was measured at 405 nm. Both mango leaf extract and mangiferin possessed a dose response relationship with a great inhibitions, especially yeast α-glucosidase (mango leaf extract; IC50=0.050 mg/ml), rat α-glucosidase activity (mangiferin; IC50 = 0.433 mg/ml) when compared to acarbose (IC50 = 11.926 and 0.449 mg/ml, respectively). For antimicrobial activities, thirteen representatives gram-positive bacteria, gram-negative bacteria and fungi were used to demonstrate zone of inhibitions and MIC, MBC and MFC. For disk diffusion, mango leaf extract showed inhibition zones against some of tested gram-positive bacteria; whereas, mangiferin showed inhibition zones against some of tested both gram-positive and gram-negative bacteria. For broth microdilution, mango leaf extract and mangiferin showed the most potent inhibition against Kocuria rhizophila with MIC values of 15.63 and 62.5 µg/ml and MBC values of 2000 and ≥ 2000 µg/ml, respectively. Anticancer activity was evaluated against five human cancer cell lines compared to two human normal cell lines using MTT assay. For cytotoxicity, mango leaf extract, ≥ 200 µg/ml, showed cytotoxicity against tested cancer cell lines. Both mango leaf extract and mangiferin increased % survival of skin fibroblast.