Dehydrogenation reaction at C1(2) positions is typical and representative of industrial production of steroid drugs. Anti-inflammatory activity can be doubled when the nucleus of the anti-inflammatory steroid hormone drug introduces double bonds at the C1(2) positions. Arthrobacter simplex is currently the most widely studied and used strain for C1(2) dehydrogenation. Therefore, breeding Arthrobacter simplex with high-efficiency dehydrogenation ability is of great significance. In order to obtain high-efficiency strains, the research proposed a new screening strategy based on image process technique: firstly, a color reaction between 2,4-dinitrophenylhydrazine (DNPH) and 9α-hydroxyandrost-4-ene-3,17-dione (9α-OH-AD) was established to characterize the dehydrogenation ability of the strain; secondly, the color data of strains mutated by atmospheric and room temperature plasma (ARTP) in the “color reaction” were automated and analyzed for dehydrogenation ability prediction using optimized support vector machine model. Result showed that the prediction accuracy reached as high as 96% in verification experiments. After a series of mutagenesis, including breaking the bottleneck of a single mutation in ARTP, the dominant strain ARLU-146 was finally obtained from 5168 strains. Its initial conversion rate was 0.8059 g/L/h, with a conversion of 94.41% at 24 h, compared to the original strain ASP which increased the transformation rate by more than 10%. By further process optimization, a high conversion (94.34% within 20 h) with high substrate (85 g/L cortisone acetate) was achieved. According to literature research, it is the highest conversion at this substrate concentration. • A high-throughput screening method was developed by using image processing and machine learning technique. • “Mutation bottleneck” of single ARTP mutagenesis was surpassed by complex mutagenesis. • A high substrate (85 g/L CA) and high transformation rate craft (94.34% within 20 h) were built.

Shanxi aged vinegar (SAV), a traditional Chinese cereal vinegar, is produced using solid-state fermentation (SSF) technology. Organic acids are the key flavor compounds of vinegar. However, the metabolic mechanism of organic acids during SSF process is still unclear. In this study, metatranscriptomics was used to explore the metabolic profile of main organic acids in SSF. The results show that carbon metabolism is the dominant pathway during fermentation, among which pyruvate metabolism, glycolysis and starch and sucrose metabolism associated with organic acids were the most abundant. The metabolic pathways of acetic acid and lactic acid shift from acetyl-P and pyruvate pathways at early and middle-early stages of fermentation to acetaldehyde and L-lactaldehyde pathways at later stages, respectively, and Lactobacillus and Acetobacter are the predominant microorganisms contributed to them. Temperature and acetic acid are proven to be the environmental factors that regulate the metabolic activity during SSF. This study sheds new lights on metabolism of flavor substances in the spontaneous ecosystems of traditional fermented food.

Background The industrial vinegar residue (VR) from solid-state fermentation, mainly cereals and their bran, will be a potential feedstock for future biofuels because of their low cost and easy availability. However, utilization of VR for butanol production has not been as much optimized as other sources of lignocellulose, which mainly stem from two key elements: (i) high biomass recalcitrance to enzymatic sugar release; (ii) lacking of suitable industrial biobutanol production strain. Though steam explosion has been proved effective for bio-refinery, few studies report SE for VR pretreatment. Much of the relevant knowledge remains unknown. Meanwhile, recent efforts on rational metabolic engineering approaches to increase butanol production in Clostridium strain are quite limited. In this study, we assessed the impact of SE pretreatment, enzymatic hydrolysis kinetics, overall sugar recovery and applied atmospheric and room temperature plasma (ARTP) mutant method for the Clostridium strain development to solve the long-standing problem. Results SE pretreatment was first performed. At the optimal condition, 29.47% of glucan, 71.62% of xylan and 22.21% of arabinan were depolymerized and obtained in the water extraction. In the sequential enzymatic hydrolysis process, enzymatic hydrolysis rate was increased by 13-fold compared to the VR without pretreatment and 19.60 g glucose, 15.21 g xylose and 5.63 g arabinose can be obtained after the two-step treatment from 100 g VR. Porous properties analysis indicated that steam explosion can effectively generate holes with diameter within 10–20 nm. Statistical analysis proved that enzymatic hydrolysis rate of VR followed the Pseudop-second-order kinetics equation and the relationship between SE severity and enzymatic hydrolysis rate can be well revealed by Boltzmann model. Finally, a superior inhibitor-tolerant strain, Clostridium acetobutylicum Tust-001, was generated with ARTP treatment. The water extraction and enzymolysis liquid gathered were successfully fermented, resulting in butanol titer of 7.98 g/L and 12.59 g/L of ABE. Conclusions SE proved to be quite effective for VR due to high fermentable sugar recovery and enzymatic hydrolysate fermentability. Inverse strategy employing ARTP and repetitive domestication for strain breeding is quite feasible, providing us with a new tool for solving the problem in the biofuel fields.