The HPLC profile of crude enzyme-treated onion extract was compared with those from non-treated or inactivated enzyme treated extracts. The peaks at tR 21.0 and 25.8 min (open arrows) were remarkably increased after fermentation, while the peaks at tR 19.0 and 22.7 min (closed arrows), which were the major compounds in control groups, were significantly decreased (Fig. 2). By repeated chromatography, the two decreased compounds were
collected and identified as quercetin-3,40-di-O-b-D-glucoside (1) and quercetin-40-O-b-D-glucoside (2) while the two increased compounds were isolated and identified as quercetin-3-O-b-D-glucoside (3) and quercetin (4) based on comparison of their NMR data with those in the literatures (Fernandez et al., 2005; Fossen
et al., 1998). The chemical structures are presented in Fig. 3.
A. kawachii was documented as having strong glucosidase activity
(Do, 2003). Therefore, increased compounds might be converted
from the enzymatic cleavage of corresponding quercetin
glucosides. It was expected that both 1 and 2 should be converted
into quercetin since they have only glucose as a sugar in their
structures and A. kawachii has been known for its strong b-glucosidase
activity. When purified 1 and 2 were separately treated with
crude enzyme, 2 was fully converted into 4, while most of 1 was
changed into 3, which still had one glucose moiety at quercetin
C-3 (Fig. 4). This is presumably due to the glucose binding position,
namely the position at C-3 of 1, which is sterically hindered
between the catechol moiety and carbonyl group.