Description

       Disaccharides such as sucrose are hydrolyzed by α-glucosidase in the small intestine. The resulting glucose is rapidly absorbed into the blood vessels, causing a rise in blood glucose. Taking an agent to reduce α-glucosidase activity is very useful as a preprandial treatment for diabetics and for people worried about their blood sugar levels. It has been reported that some natural products show this activity. However, it is not easy to detect the inhibitory activity of α-glucosidase, since natural products often have a dark color that interferes with the conventional measurement method that uses synthetic substrates for α-glucosidase. In addition, an IC50 value is normally used to describe inhibitory activity, but an IC50 value is not effective for expressing intensity when the control value varies between measurements. The test also requires special skills on the part of technicians. This situation prompted us to develop a new measurement process that can be applied to colored samples. Douchi, a traditional Chinese soybean-based fermented food that has a reputation for being healthenhancing, shows many local variations. Some types have angiotensin-converting enzyme inhibitory activity, as reported in the 2004 Annual Report. This new method was applied to douchi to search for highly active products. Rat small intestine acetone powder was used as an enzyme source of α-glucosidase. This enzyme was placed in a 96-well microplate. Diluted samples were made and added to each well, followed by warming at 37℃ for 40 min with the synthetic substrate 4-nitrophenyl-β-D-glucopyranoside, after which the reaction was stopped by addition of alkali. The absorbance at 405 nm was measured after color development at the same time. A blank without enzyme was measured simultaneously. Mulberry leaf extract was used as the positive contrast. The slope value of the absorbance-dilution straight line was -18.23, indicating that the sample showed strong suppressive activity (Fig. 1A). Coffee was used as the negative sample. The slope value of the line was 0.176, which we interpret as being close to zero (Fig. 1B). The variation index in this assay was 3.39% (n = 4) for intra-assay and 2.68% (n = 3) for inter-assay. Water-extracted samples (extraction concentration 40% (w/v)) of various douchi chosen at random in China were measured using this method (Fig. 2). BM (Jiangxi), KKHBS (Hunan), and MK (Sichuan) showed conspicuously higher activity than the other samples. In summary, a simple and sensitive α-glucosidase assay method was developed for colored samples, and douchi, a famous Chinese traditional food, was measured using this method. Some douchi samples showed strong α- glucosidase-suppressive activity.

Figure, table

1.  

   

   Fig. 1. Absorbance-dilution curves of mulberry leaf extract (A) and coffee (B)

2.  

   

   Fig. 2. α-glucosidase-inhibitory activity of various water-extracted samples.
   Douchi samples from China are shown as open columns, extracts of Japanese miso and natto as the red columns, voglibose (anti-diabetes) as the blue column, and guava tea as the green column.