3 years ago

The mechanism of interactions between tea polyphenols and porcine pancreatic alpha-amylase: Analysis by inhibition kinetics, fluorescence quenching, differential scanning calorimetry and isothermal titration calorimetry

The mechanism of interactions between tea polyphenols and porcine pancreatic alpha-amylase: Analysis by inhibition kinetics, fluorescence quenching, differential scanning calorimetry and isothermal titration calorimetry
Lijun Sun, Frederick J. Warren, Michael J. Gidley
Scope This study aims to use a combination of biochemical and biophysical methods to derive greater mechanistic understanding of the interactions between tea polyphenols and porcine pancreatic α-amylase (PPA). Methods and results The interaction mechanism was studied through fluorescence quenching (FQ), differential scanning calorimetry (DSC) and isothermal titration calorimetry (ITC) and compared with inhibition kinetics. The results showed that a higher quenching effect of polyphenols corresponded to a stronger inhibitory activity against PPA. The red-shift of maximum emission wavelength of PPA bound with some polyphenols indicated a potential structural unfolding of PPA. This was also suggested by the decreased thermostability of PPA with these polyphenols in DSC thermograms. Through thermodynamic binding analysis of ITC and inhibition kinetics, the equilibrium of competitive inhibition was shown to result from the binding of particularly galloylated polyphenols with specific sites on PPA. There were positive linear correlations between the reciprocal of competitive inhibition constant (1/Kic), quenching constant (KFQ) and binding constant (Kitc). Conclusion The combination of inhibition kinetics, FQ, DSC and ITC can reasonably characterize the interactions between tea polyphenols and PPA. The galloyl moiety is an important group in catechins and theaflavins in terms of binding with and inhibiting the activity of PPA. One of the leading causes of type II diabetes mellitus is excess fluctuations in post-prandial blood sugar. It is believed that certain compounds present in tea (called polyphenols) may act to slow the digestion of starch, reducing fluctuations in post-prandial blood glucose. In this study, detailed experimental information is provided regarding the interaction between polyphenols and amylase, the enzyme that digests starch, that will allow us to identify which of these compounds in tea are best at reducing starch digestion

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/mnfr.201700324

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