a b s t r a c tNine kinds of chlorogenic acids (CGAs) account for 80% of the total CGA content in green coffee beans. They consist of three subgroups of caffeoylquinic acids (CQAs), feruloylquinic acids (FQAs), and dicaffeoylquinic acids (diCQAs). We previously reported the inhibitory effects of 5-CQA on porcine pancreas a-amylase (PPA) isozymes, PPA-I and PPA-II. In this paper, we investigated the PPA-I inhibition by eight kinds of CGAs. The IC 50 values of CQAs, FQAs, and diCQAs against the PPA-I-catalysed hydrolysis of pnitrophenyl-a-D-maltoside were 0.08-0.23 mM, 1.09-2.55 mM, and 0.02-0.03 mM, respectively. All CQAs and FQAs and 3,5-diCQA showed mixed-type inhibition with binding to the enzyme-substrate complex (ES) being stronger than to the enzyme (E). 3,4-DiCQA and 4,5-diCQA showed mixed-type inhibition, but, conversely are suggested to bind to E stronger than ES.

Coffee silverskin (CS) is a thin tegument of the outer layer of green coffee beans and a major by-product of the roasting process to produce roasted coffee beans. CS extracts obtained by the treatment of CS with subcritical water at 25-270°C were investigated for their antioxidant activity using hydrophilic oxygen radical absorption capacity (H-ORAC) and DPPH radical scavenging capacity assays. The antioxidant activity increased with increasing the extraction temperature and the highest activity was observed with the extracts obtained at 270°C. The H-ORAC and DPPH values of the extracts were 2629±193 and 379±36μmol TE/g of CS extract, respectively. High correlation (R=0.999) was observed between H-ORAC and DPPH values for the CS extracts. High correlation of the antioxidant activity was also observed with protein and phenolic contents in the extracts. The CS extracts could be useful as a good source of antioxidative materials.

Chlorogenic acid (5-caffeoylquinic acid, 5-CQA) is a kind of polyphenol and is richly included in green coffee beans. The inhibitory effects of 5-CQA and its components, caffeic acid (CA) and quinic acid (QA), on the two porcine pancreas alpha-amylase (PPA) isozymes, PPA-I and PPA-II, were investigated using p-nitrophenyl-alpha-D-maltoside as substrate at pH 6.9 and 30 degrees C. The inhibition potencies of the respective inhibitors against both PPA isozymes were almost the same and in the order of 5-CQA > CA >> QA. Their IC(50) values were 0.07-0.08 mM, 0.37-0.40 mM, and 25.3-26.5 mM, respectively. The inhibition mechanisms of 5-CQA and CA were investigated by kinetic analyses, and the inhibitor constants K(i) and K(i)' (for the free enzyme and enzyme-substrate complex, respectively) were determined. It was indicated that 5-CQA and CA showed mixed-type inhibition with K(i) > K(i)' against both PPA-I and PPA-II. The binding of PPA-I or PPA-II with 5-CQA or CA was all exothermic and enthalpy-driven. QA is a poor inhibitor, and its inhibitory mode was unique and hardly analyzed by a simple Michaelis-Menten-type interaction between the enzyme and inhibitor. However, it was shown that the inhibitory activity of CA was enhanced 5 times by ester-bond formation with QA in the form of 5-CQA. These results provide us with significant hints for the development of alpha-amylase inhibitors useful for the prevention of diabetes and obesity.

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5-Caffeoylquinic acid (5-CQA) is generally referred to as chlorogenic acid and exhibits various biological activities such as antioxidant activity and porcine pancreas α-amylase inhibitory activities. 5-CQA may be useful as an antioxidant for food and to prevent diabetes and obesity. The degradation of 5-CQA and caffeic acid (CA) in an aqueous solution at 37 °C and pH 5.0-9.0 was studied. The degradation of 5-CQA and CA, demonstrating time and pH dependence (i.e., the rate constant, k, was higher at higher pH), was satisfactorily described by the Weibull equation. The stability of 5-CQA at pH 7.4 and 9.0 was improved by adding (-)-epigallocatechin gallate (EGCG) and ascorbic acid (AA). Moreover, the degradation of 5-CQA in the presence of EGCG or AA could be described by the Weibull equation. The k value in the presence of EGCG or AA was dependent on their concentration.