Abstract
Syringic acid (SA) is a natural derivative of syringaldehyde (SD), derived from lignin depolymerization. Its application in the food industry focuses on the properties of natural functional ingredients; it is mainly used as a food antioxidant and food preservative, but can also be used as an ingredient to enhance food flavor and functional foods. This compound exhibits a remarkable spectrum of biological activities, including potent antioxidant, anti-inflammatory, neuroprotective, hypoglycemic, detoxifying, and anti-cancer effects, positioning it as a highly promising candidate for pharmaceutical and nutraceutical applications. In this study, suitable sites were first screened through homologous sequence alignment, and a variant of aryl-alcohol oxidase (CgAAO) with high efficiency in catalyzing the conversion of SD to SA was obtained via site-directed mutagenesis. A deep eutectic solvent (DES) system based on choline chloride/urea (ChCl/UR) in water was developed to enhance SA production. Additionally, key parameters of the biological reaction were optimized, including temperature, pH, metal ions, as well as the type and dosage of DES. The optimal performance was achieved using recombinant E. coli pRSFDuet-CgAAO-Y335F whole-cell biocatalysts, yielding 75% and producing 0.75 g/L SA in 100 mM KPB buffer (pH 7.0) containing 5 wt% ChCl/UR and 1 mM Fe3+. This study established a novel biosynthetic pathway for SA that was efficient, mild, green, and environmentally friendly.