摘要：

AIM: To establish a new sensitive microplate-based method to determine alpha -glucosidase inhibiting activity and provide a reliable high-throughput way for screening alpha-glucosidase inhibitors in vitro. METHODS: The fitting combination of enzyme and substrate in a certain reaction was tested. Acarbose, the most popular alpha-glucosidase inhibitor in clinical use was used to validate the established method. Calibration curve, wavelength fidelity and kinetic analysis, together with the effect of altered incubation time, temperature and pH were then studied. RESULTS: The details of assay procedure and evaluation of factors affecting the measurement are described. As little as 160μl assay system with the combination of 2.5 mmol·L~(-1) PNPG and 0.2 U·ml~(-1) alpha -glucosidase was performed in a 96-well plate. The optimal action was finally achieved by incubated at 37℃, pH 7.0 for 15 min and measured at 400 nm. Results from the validation exercises by Acarbose strongly demonstrated the accuracy and reliability of the proposed approach. CONCLUSION: This method reported in the current paper makes it possible to rapidly examine large numbers of samples for the presence of alpha-glucosidase inhibitors in very small sample volumes. Such action may help to pace the development of potential oral medications from natural products protecting patients against postprandial hyperglycemic toxicity and therefore treating diabetes mellitus and the related complications.

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