Sucrose → (2R,3S,4R,5R)-2,3,4,5,6-pentahydroxyhexanal

Each of the proteolytic enzyme-treated azuki bean prepared in Example 2 and an enzyme-treated defatted soybean prepared using the same method as Example 2 was dissolved in 0.25 M phosphate buffer (pH 6.8) used as assay buffer to a concentration of 16.7 mg/mL to give a sample. Measurement of α-glucosidase activity was carried out in the manner described below. Specifically, 60 μL of the sample was added to wells of a 96-well plate while 60 μL of assay buffer was added to wells for control. Subsequently, 20 μL of yeast-derived α-glucosidase (Oriental Yeast Co., Ltd.) adjusted to a concentration of 1.25 U/mL (10 μg protein/mL) was added followed by incubating for 10 minutes at 37° C. Next, 20 μL of a 250 mM solution of sucrose in assay buffer was added followed by incubating for 20 minutes at 37° C. (the concentration of each enzyme-treated product during the reaction was 10 mg/mL). Following completion of incubation, 100 μL of 2 M Tris-HCl buffer (pH 7.0) was added to stop the reaction. After stopping the reaction, 50 μL of the reaction liquid in each well was transferred to a separate 96-well plate followed by the addition of 200 μL of a Glucose CII Test Wako reagent (Wako Pure Chemical Industries, Ltd.) and incubating for 5 minutes at 37° C. Following completion of incubation, the absorbance at 505 nm was measured, and the amount of glucose formed was determined from a calibration curve prepared using glucose standards. Furthermore, the α-glucosidase activity was calculated as indicated below based on a value of 100% for the value of α-glucosidase activity of the control. α-Glucosidase activity(%)=(A/B)×100

1. customEach of the proteolytic enzyme-treated azuki bean prepared in Example 2
2. customprepared
3. customto give a sample
4. additionSpecifically, 60 μL of the sample was added to wells of a 96-well plate while 60 μL of assay
5. additionbuffer was added to wells for control
6. additionwas added
7. waitby incubating for 20 minutes at 37° C.
8. additionwas added
9. customthe reaction
10. customthe reaction, 50 μL of the reaction liquid in each well
11. customa separate 96-well plate
12. additionfollowed by the addition of 200 μL of a Glucose CII Test Wako reagent (Wako Pure Chemical Industries, Ltd.)
13. waitincubating for 5 minutes at 37° C

ORD record

ord-25cc405735c24ec7ab0c82363ff47251

Source dataset

uspto-grants-2016_07

DOI

10.6084/m9.figshare.5104873.v1

Patent number

US09399051B2