Determination of free and bound phenolic acids in beer

The aim of this study was the qualitative and quantitative determination of free and bound phenolic compounds, mainly phenolic acids, in beer. In spite of the wide literature describing the content of free phenolic acids in beer, data concerning its content of bound forms are scarce or missing. Moreover, the experimental conditions commonly reported in the literature to detect bound phenolic acids by alkaline hydrolysis result in loss of several phenolic acids, particularly dihydroxy-derivatives. Recently, we described that the addition of ascorbic acid, a strong antioxidant, and ethylenediaminetetraacetic acid, a metal chelator, totally prevents the loss of phenolic acids during alkaline hydrolysis. On this basis, we developed a hydrolytic procedure based on alkaline hydrolysis with 2 N NaOH containing ascorbic acid (1% w/v) and ethylenediaminetetraacetic acid (10 mM). In these conditions, a complete recovery of caffeic acid (98.7±4.3% of expected value) following hydrolysis of chlorogenic acid (5′-caffeoylquinic acid, an ester of caffeic acid with quinic acid) was obtained. In the present study we took advantage of this hydrolytic procedure to quantitatively measure free and total (free plus bound) phenolic acids in beer. After alkaline hydrolysis, which released bound phenolic acids, a remarkable increase in the content of 4-hydroxyphenylacetic acid, vanillic acid, caffeic acid, syringic acid, p-coumaric acid, ferulic acid and sinapic acid was observed in beer from three different brands. Our results show that the most of phenolic acids in beer are present as bound forms and only a small portion can be detected as free compounds.