Diosgenin

Diosgenin is a key precursor for steroidal pharmaceuticals; however, traditional acid hydrolysis produces significant environmental pollution, whereas enzymatic conversion presents a more environmentally sustainable alternative. Herein, four saponinases were identified from Aspergillus tubingensis HG57: the β-glucosidases ATGH I and ATGH II, which hydrolyzed residual β-d-glucosyl moieties, and the rhamnosidases ATRH I and ATRH II, which cleaved terminal α-L-(1 → 2) rhamnosyl linkages in steroidal saponins. ATGH I (7.46 U mg^-1) and ATRH I (285.31 U mg^-1) demonstrated some of the highest catalytic efficiencies documented to date. Both enzymes displayed broad pH tolerance and thermostability (≤50 °C), with ATRH I maintaining over 90 % activity in 2 M rhamnose. Structure-guided engineering of ATGH I led to the development of an improved variant, ATGH M, which displayed a two-fold improvement in catalytic efficacy (14.99 U mg^-1). High-density fermentation produced enzyme-rich supernatants (ATGH M: 98.52 U mL^-1; ATRH I: 2564.36 U mL^-1) that facilitated gram-scale, one-pot enzymatic hydrolysis to produce diosgenin. This bioprocess achieved 138 % of the diosgenin yield obtained by traditional acid hydrolysis while markedly reducing wastewater pollutants, lowering chemical oxygen demand by 76 %, sulfate by 99 %, and acidity by nearly 100 %. The integration of efficient saponinase discovery, protein engineering, and high-density fermentation established a green and scalable enzymatic platform for diosgenin production, offering a sustainable alternative to conventional acid hydrolysis.

This study employed an integrated approach combining in vivo metabolism analysis, network pharmacology, experimental validation and quantitative analysis to investigate the anti-inflammatory material basis and mechanism of action of Paris polyphylla var. yunnanensis (PPY). Firstly, rats were orally administered extracts from the roots and leaves of PPY, and plasma, urine, and feces samples were collected at different time points post-administration. Using UHPLC-Q/TOF-HRMS technology, a total of 26 xenobiotics (18 prototypes and 8 metabolites) were identified from the rat samples. Subsequently, network pharmacology analysis predicted 19 core targets and preliminarily screened 10 key bioavailable components with potential anti-inflammatory activity. Further in vitro cell experiments evaluated the bioactivity of these key components, revealing that polyphyllin II exhibited the most significant anti-inflammatory effect, while pennogenin and diosgenin also demonstrated anti-inflammatory activity within certain concentration ranges. Finally, HPLC-DAD analysis was used to compare the content differences of major active components in PPY from different geographical origins. The results showed that samples from Hanzhong (Shaanxi) and Chengdu (Sichuan) contained higher levels of pennogenin-type saponins, whereas those from Honghe (Yunnan) and Yuxi (Yunnan) were richer in diosgenin-type saponins. In summary, this comprehensive strategy significantly contributes to elucidating the anti-inflammatory material basis and mechanism of PPY, while also providing a reference for the systematic discovery of active components in the quality evaluation of traditional Chinese medicines.