Ethyl carbamate (EC), a potential carcinogen, is spontaneously formed from urea and ethanol during Huangjiu fermentation, and urea is derived from arginine (Arg) degradation by Saccharomyces cerevisiae . Previous studies have shown that BTN2 influences Arg uptake and metabolism. In this study, we conducted a comprehensive analysis of the variation of EC, major metabolites, enzyme activities, volatile flavor compounds, amino acids, and general quality within the Arg metabolism pathway during Huangjiu fermentation by wild type, BTN2 Δ, and BTN2 ↑ strains. The results indicated that BTN2 knock out significantly reduced EC, urea, and citrulline by 12.28, 15.6, and 20.18%, respectively. Furthermore, flavors and amino acids among the three groups exhibited minimal variation, resulting in insignificant differences in the sensory evaluation. The primary flavors identified were ethyl lactate and isoamyl alcohol. Cysteine, Arg, and glutamine contributed significantly to the overall flavor profile. These findings suggest that Btn2p is a potential target for the reduction of EC.
Ethanol produced during the Huangjiu fermentation can adversely affect the growth and metabolism of Saccharomyces cerevisiae . It has been established that arginine, one of the major amino acids found in Huangjiu , can protect yeast cells against ethanol stress. Previous studies have indicated that BTN2 , which influences arginine metabolism, is associated with ethanol tolerance in yeast. In this study, we evaluated the response of BTN2 to ethanol stress based on analyses of cell growth, morphology, membrane permeability, membrane integrity, ROS and how BTN2 influenced intracellular metabolites, enzymes, and transcriptional genes in the arginine metabolic pathway. Our findings revealed that BTN2 promoted significantly increases in the growth and ethanol tolerance of S. cerevisiae g. Under 10% ethanol stress, RT-qPCR showed that BTN2 Δ ( BTN2 knock out strain) up-regulated CAR1 , CAR2 , GAP1 , DUR1,2 and DUR3 to increase arginine and urea degradation, BTN2 ↑ ( BTN2 over-expressing strain) up-regulated CAR1 , CAN1 and DUR3 to enhance arginine uptake, catabolism and urea reabsorption, while TPS1 was up-regulated to improve ethanol tolerance ability. Yeast up-regulated both CAR1 (Arginase) and DUR3 (urea transporter) genes in BTN2 knockout and over-expression strains to counteract the ethanol effects. Our findings revealed a novel arginine metabolism under ethanol tolerance of a core functional yeast during Huangjiu fermentation. These findings could contribute to the development of an approach for inhibiting toxic ethyl carbamate (EC) production during Huangjiu fermentation.
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