Valorization of starch-rich traditional Chinese Herbal residues into 5-Keto-d-gluconic acid via multi-enzymes cascade

Multi-enzyme cascade valorizes starch-rich TCHRs into 5KGA; • NADP + regeneration built via Ga5DH-SBFA-CAT coupling; • Reverse activity suppressed, enabling efficient GA-to-5KGA conversion; • Fed-batch production of 5KGA reached 203 g/L with TTN of 9.5 × 10 4 . Herbal residues (HRs) are emerging as an important and sustainable biomass resource, driven by the diversification of global health concepts and the rapid growth in related consumption. Herein, we established a valorization route from starch-rich traditional Chinese herb residues (TCHRs) to high-value 5‑keto‑D‑gluconic acid (5KGA) via a multi-enzymatic cascade. Starch-rich TCHRs were first converted to gluconic acid (GA) through enzymatic hydrolysis and oxidation (>80% conversion). Subsequently, an oxidized cofactor regeneration system was established to efficiently convert GA into 5KGA by coupling gluconate-5-dehydrogenase (Ga5DH) with a synthetic bridged-flavin analogue (SBFA), thereby mitigating constraints arising from Ga5DH’s high reductive activity. This system catalyzed the irreversible dehydrogenation of GA to 5KGA, achieving complete conversion of 100 mM GA within 4 h (total turnover number, TTN = 1.0 × 10 4 ). Furthermore, upon scaling up in a 1-L bioreactor with continuous operation for 48 h, 203.3 g/L (950 mM) of 5KGA was produced with a TTN of 9.5 × 10 4 . This value-upgrading strategy provides an efficient and economically viable biotransformation pathway for starch-based non-edible biomass feedstocks. Download: Download high-res image (112KB) Download: Download full-size image