Phenolic acid decarboxylase of Aspergillus luchuensis plays a crucial role in 4-vinylguaiacol production during awamori brewing

Aspergillus luchuensis has been used to produce awamori, a distilled liquor, in Okinawa, Japan. Vanillin, derived from ferulic acid (FA) in rice grains, is one of the characteristic flavors in aged and matured awamori, known as kusu. Decarboxylation of FA leads to the production of 4-vinylguaiacol (4-VG), which is converted to vanillin by natural oxidization. However, the mechanism underlying FA conversion to 4-VG has remained unknown in awamori brewing. In our previous studies, we showed that phenolic acid decarboxylase from A. luchuensis (AlPAD) could catalyze the conversion of FA to 4-VG, and that AlPAD is functionally expressed during koji making (Maeda et al., J. Biosci. Bioeng., 126, 162–168, 2018). In this study, to understand the contribution of AlPAD to 4-VG production in awamori brewing, we created an alpad disruptant (Δalpad) and compared its 4-VG productivity to that of the wild-type strain. The amount of 4-VG in the distillate of moromi prepared with the wild-type strain showed a significant increase, proportional to the time required for koji making. In the Δalpad strain, the amount of 4-VG was very small and remained unchanged during the koji making. In an awamori brewing test using koji harvested 42–66 h after inoculation, the contribution of AlPAD to 4-VG production was in the range of 88–94 %. These results indicate that AlPAD plays a key role in 4-VG production during awamori brewing.