Evaluation of quantitative variation of secondary metabolites in Bergenia ciliata (Haw.) using high performance thin layer chromatography

Dear Editor: Therapeutically active metabolite contents in a medicinal plant vary in nature, which may impact on its therapeutic efficacy. Bergenia (Saxifragaceae) is an evergreen perennial herb widely distributed in Central and East Asia with about 30 species reported worldwide. It grows at a range of altitudes from the Khasia hills at 400 feet to the temperate Himalayas from Kashmir to Bhutan at 7,000-10,000 feet [1] . Its distribution over a wide range of altitudinal zones makes it a good candidate for studying variations in its metabolic profiles under different climatic conditions. Bergenin (C-glycoside of 4-O-methyl gallic acid) has been identified as a potent active secondary metabolite in Bergenia and other therapeutically active constituents including, among others, gallic acid (3,4,5 trihydroxybenzoic acid), (+) catechin, and gallicin (Fig. 1). Here, we analyzed three accessions of Bergenia ciliata collected from different altitudes of Uttrakhand, India, to study the quantitative variation in bergenin, gallic acid, (+) catechin, and gallicin by high performance thin layer chromatography (HPTLC). For accurate quantification, different hydrolysis conditions were employed and their effects on individual compound were assessed. We found significant variations in the content of these compounds in the collected rhizomes of Bergenia ciliata ranging from 0.756¡0.3 to 0.85¡0.2 for bergenin, 0.145¡0.2 to 0.27¡0.2 for (+) catechin, 0.027¡0.2 to 0.166¡0.3 for gallic acid and 0.135¡0.3 to 0.186¡0.1 for gallicin. Such variability is expected due to variation in environmental conditions which varies on varying altitudes. Fig. 2 illustrates the effect of climatic and altitudinal variations on the distribution of these four compounds in Bergenia rhizomes, showing that even a small altitudinal variation is capable of inducing quantitative variation in the contents of these compounds. It has been established that biosynthesis of polyphenolic compounds increases on higher altitude in response to lower environmental stress. But the present study revealed that it was not the case for every phenolic compound. Bergenin content showed no significant positive correlation with altitude while gallic acid showed significant correlation with altitude. However, it is difficult to judge whether this is due to a significant shift in the biosynthetic pathways or whether it is merely a mathematical artifact resulting from the fact that one variable varies with altitude, whereas another does not. Phenolic compounds exist in glycoside and aglycone forms [2] . In most cases, the aglycone form is absorbed at a greater rate and has higher antioxidant activity than the glycoside form [3-6] . Hydrolysis of flavonol glycosides to their corresponding aglycones offer a practical method for the quantification of flavonoids in foods [7] . Hydrolysis is also used for quantification of phenolic acids in fruits or other plant part [8] . We exposed Bergenia extract to different hydrolysis conditions. Fig. 1 Chemical structure of A: Bergenin; B: (+)Catechin; C: Gallic