Studies on the Impact of Malting and Mashing on the Free, Soluble Ester-Bound, and Insoluble Ester-Bound Forms of Desired and Undesired Phenolic Acids Aiming at Styrene Mitigation during Wheat Beer Brewing

Abstract

Mitigation studies on styrene in wheat beer found no correlation between the free phenolic acid contents in the processing steps and the final concentrations of the toxicologically relevant styrene and the desired aroma-active vinyl aromatics in beer, which can be explained by the presence of phenolic acid releasing enzymes that are still active after kiln-drying and by the yeast′s own feruloyl esterase activity. The present study contributed to a better understanding of the coherence between the free, soluble ester-bound, and insoluble ester-bound forms of cinnamic, p-coumaric, and ferulic acid during malting and mashing of barley and wheat varieties. Concentration differences in malt by factors of up to 1700 were found between the total cinnamic acid contents (an undesired precursor of the toxicologically relevant styrene) and the total contents of p-coumaric and ferulic acid (both desired precursors of the aroma-active compounds 4-vinylphenol and 2-methoxy-4-vinylphenol). In grain and malt, cinnamic acid occurred predominantly in a soluble form, whereas the desired precursors were mainly insoluble ester-bound. This had a direct effect on the transfer rates from malt into wort, which were found to be >100% for cinnamic acid, revealing that a complete transfer was accompanied by an additional biosynthesis, but only <8% for the desired phenolic acids. Interestingly, in the wort, cinnamic and pcoumaric acid contents were dominated by the free form, while ferulic acid was mostly still soluble ester-bound. Overall, the use of barley malts led to an introduction of cinnamic, p-coumaric, and ferulic acid into the wort in a ratio of 2:14:84, and the use of wheat malt in a ratio of 1:2:97.

The malting parameters: steeping, germination, withering, and kilning temperature and aeration rate as possibilities for styrene mitigation in wheat beer

“…To study the impact of the kilning temperature within experiment 4, a set of dark malts was additionally produced to the pale malts with a withering temperature of 50 °C and a kilning temperature of 200 °C (2 min). Further, from all these malts, the corresponding wort was produced according to the standard mashing procedure of congress mash (50 g of malt, 400 mL of water) (MEBAK ® 4.1.4.2 [30], EBC 4.5.1 [31]), since a former study [35] revealed that changes within the malts can differ from the impact that varying malting parameters have on the final state in wort.…”

“…Total, soluble, and free phenolic acids were isolated and their contents in malts were determined as previously described in detail [35].…”

“…The determination of soluble and free phenolic acids in wort was performed as recently described [35].…”

“…Positive atmospheric pressure chemical ionization (APCI + ) was applied using the following conditions: vaporizer temperature, 250 °C; discharge current, 4.0 µA; sheath gas pressure, 30 arbitrary units (au); auxiliary gas pressure, 10 au; declustering voltage, -10 V; and capillary temperature, 300 °C. The mass spectrometer was operated in the multiple reaction monitoring (MRM) mode, and the most intensive fragment ion was chosen as the quantifier ion, together with two qualifier ions [35].…”

“…Calibration lines were determined from mixtures of known contents of the unlabeled analyte and the corresponding stable isotopically labeled internal standard in seven different concentration ratios (10:1, 5:1, 3:1, 1:1, 1:3, 1:5, and 1:10) with good linearity (R 2 > 0.99) for all analytes in the applied range [35].…”