Zaopei is the direct source of Chinese liquor (Baijiu). Adding functional strains to Zaopei is a potential strategy to regulate Baijiu brewing, mainly including the two ways of solid-state fortified Jiuqu (SFJ) and liquid-state fortified agent (LFA). Here, to explore their regulated details, the response patterns of Zaopei microecosystem and the changes in the product features were comprehensively investigated. The results showed that SFJ more positively changed the physicochemical properties of Zaopei and improved its ester content, from 978.57 mg/kg to 1078.63 mg/kg over the fermentation of 30 days, while LFA decreased the content of esters, alcohols, and acids. Microbial analysis revealed that SFJ significantly increased Saccharomycopsis and Aspergillus from the start of fermentation and induced a positive interaction cluster driven by the added functional Paenibacillus, while LFA exhibited a community structure near that of the original microecosystem and led to a simpler network with the reduced microbial nodes and correlations. Metabolism analysis found that both SFJ and LFA weakened the flavor-producing metabolism by suppressing some key enzyme pathways, such as EC 3.2.1.51, EC 4.2.1.47, EC 1.1.1.27, EC 1.1.1.22, EC 1.5.1.10, EC 1.14.11.12. As a result, SFJ improved the raw liquor yield by 28.5 % and endowed the final product with a more fragrant aroma, mainly through ethyl (E)-cinnamate, ethyl isovalerate, ethyl phenacetate with the higher odor activity values, while LFA promoted the yield by 13.2 % and resulted in a purer and less intense aroma through the aroma-active β-damascenone, ethyl heptoate, ethyl phenacetate. These results facilitated the regulated mechanism of SFJ and LFA on Baijiu brewing and indicated that the used functional strains in this study could be applicated in SFJ way for the further industrial-scale application.

As a rare disease leading to male infertility, idiopathic hypogonadotropic hypogonadism (IHH) has strong heterogeneity of clinical phenotype and gene mutation. At present, there is no effective diagnosis and treatment method for this disease. This study is to explore the possible new pathogenic gene of idiopathic hypogonadotrophic hypogonadism and the pathological mechanism affecting its occurrence. We performed a whole-exome sequencing on 9 patients with normosmic idiopathic hypogonadotropic hypogonadism (nIHH), 19 varicocele patients with asthenospermia, oligospermia, or azoospermia, 5 patients with simple nonobstructive azoospermia, and 13 normal healthy adult males and carried out comparative analysis, channel analysis, etc. After preliminary sequencing screening, 309–431 genes harbouring variants, including SNPs and indels, were predicted to be harmful per single patient in each group. In genetic variations of nIHH patients’ analysis, variants were detected in 10 loci and nine genes in nine patients. And in co-analysis of the three patient groups, nine nIHH patients, 19 VC patients, and five SN patients shared 116 variants, with 28 variant-harbouring genes detected in five or more patients. We found that the NEFH, CCDC177, and PCLO genes and the Gene Ontology pathways GO:0051301: cell division and GO:0090066: regulation of anatomical structure size may be key factors in the pathogenic mechanism of IHH. Our results suggest that the pathogenic mechanism of IHH is not limited to the central nervous system effects of GnRH but may involve other heterogeneous pathogenic genetic variants that affect peripheral organs.

The active ester-synthesis microorganisms in medium-high temperature Daqu (MHT-Daqu) largely impact the strong-flavor Baijiu quality, while their actual composition and metabolic mechanism remain unclear. Here, to explore how the active microbiota contributes to MHT-Daqu ester biosynthesis, metatranscriptomic and metaproteomic analyses coupled with experimental verification were performed. The results showed that the MHT-Daqu microbiota with the higher ester-forming ability exhibited a more active dynamic alteration from transcription to translation. The genera Aspergillus, Bacillus, Leuconostoc, and Pediococcus could transcribe and translate obviously more ester-forming enzymes. In the ester-synthesis metabolic network, the synergetic microbiota confirmed by interaction analysis, containing Eurotiales, Bacillales, and Saccharomycetales, played an essential role, in which the Eurotiales and its representative genus Aspergillus contributed the highest transcript and protein abundance in almost every metabolic process, respectively. The recombined fermentation verified that their corresponding genera could produce the ester and precursor profiles very close to that of the original MHT-Daqu active microbiota, while the microbiota without Aspergillus caused a polar separation. These results indicated that the synergetic microbiota with Aspergillus as the core dominated the metabolic network of ester synthesis in MHT-Daqu. Our study provides a detailed framework of the association between the active synergetic microbiota and ester synthesis in MHT-Daqu.

BACKGROUNDThe ester-synthesis enzymes influenced by environmental factors during Daqu-making process largely determine the flavor of Chinese liquor, but the main ester-synthesis enzyme and its key influencer remain unclear. Here, the volatile ester profiles over the whole Daqu-making process, under different treatments, for at least 90 days, were carefully analyzed, and the potential ester-synthesis enzymes, as well as their dependently environmental factors, were explored. RESULTSIn the detected 46 volatile esters, only the short-chain (C4-C8) and medium-chain (C9-C13) ester content obviously changed, as the primary contributor discriminating different samples. Their trends were both consistent with that of the alcohols and the primary metabolism, which included alcohol acyltransferases (AATs) reaction with alcohols and acyl-CoAs as the substrates. Among the potential ester-synthesis enzymes, the typical AAT activity also exhibited the highest correlation with the short- and medium-chain esters (r > 0.78, P < 0.05). The Mantel test between environmental factors and ester production showed that temperature of Daqu was directly correlated with the short-chain esters (r = 0.58, P < 0.01) and AAT activity (r = 0.56, P < 0.01). Further, the short- and medium-chain ester content in Daqu under the treatment nearer to the reported optimal temperature of 40-50 degrees C of AATs reaction was overall higher than that of the other treatment Daqu. CONCLUSIONThis study revealed that the temperature-dependent AATs reaction was the main enzymatic method producing the short- and medium-chain esters over the whole Daqu-making process. The results could contribute to the flavor improvement of Baijiu. (c) 2022 Society of Chemical Industry.

BACKGROUND:The saccharification function of daqu is usually characterized by two indicators: saccharification power and liquefaction power. Daqu provides diverse microbial saccharifying enzymes for hydrolyzing carbohydrate in Baijiu fermenting grain. Obviously, the composition of microbial communities and enzymatic genes in different types of daqu cultured at varied temperatures is different. However, these differences in saccharification function are not fully understood.RESULTS:The findings suggested that the saccharification power and liquefaction power of jiangxiangxing daqu were lower than those of nongxiangxing daqu throughout the production process. We employed metagenomics to find evidence that a mode of multiple saccharifying enzymes involving amylase, cellulase and hemicellulase originating from various microbes exists in daqu. Moreover, a totality of 541 related differential genes were obtained, some of which, annotated to genera of Aspergillus, Lactobacillus and Weissella, were significantly enriched (P < 0.05) in nongxiangxing daqu, while others, annotated to thermophilic genera of Virgibacillus, Bacillus, Kroppenstedtia and Saccharopolyspora, showed a higher relative abundance in jiangxiangxing daqu (P < 0.05).CONCLUSION:Various microbial communities of daqu showed diverse saccharification capacity during cultivation of different parameters. These findings are helpful in comprehending the saccharification functional genes of daqu. © 2022 Society of Chemical Industry.

The use of probiotics to promote the release of bioactive peptides during Cheddar cheese ripening was investigated. The proteolytic and angiotensin-converting enzyme inhibitory (ACE-I) activities and ACE-I peptide profile of Cheddar cheese with different probiotics were analysed and compared. Molecular docking was used to analyse the binding site of peptide to ACE molecule. Compared with other cheeses, ACE-I activity of Lactobacillus helveticus cheese was the highest (79.71%), and the degree of proteolysis was higher and the molecular mass was lower. Most of the peptides were less than 1.6 kDa. In addition, 43.31e51.59% of the peptides in cheese originated from b-casein, mainly from the degradation of starter and probiotics. SKVLPQ and YQEPVLGP, which were uniquely identified in L. helveticus cheese, could interact with the S1 and S2 active pocket of ACE, respectively. Especially, YQEPVLGP could bind to ACE at multiple sites and was an efficient ACE inhibitory peptide.(c) 2022 Elsevier Ltd. All rights reserved.

A general method for the direct synthesis of highly homogeneous and dense polymerized carbon nitride (PCN) nanosheet films on F: SnO2 (FTO) is developed. Detailed photoelectrochemical (PEC) water-splitting studies reveal that the as-synthesized PCN films exhibit outstanding performance as photoanode for PEC water-splitting. The optimal PCN photoanode exhibits excellent photocurrent density of 650 µA cm-2 , and monochromatic incident photon-to-electron conversion efficiency (IPCE) value up to 30.55% (λ = 400 nm) and 25.97% (λ = 420 nm) at 1.23 VRHE in 0.1 m KOH electrolyte. More importantly, the PCN photoanode has an excellent hole extraction efficiency of up to 70 ± 3% due to the abundance of active sites provided by the PCN photoanode nanosheet, which promotes the transport rates of OER-relevant species. These PCN films provide a new benchmark for PCN photoanode materials.

Summary Daqu provides hydrolytic enzymes that saccharifying the macromolecular carbohydrate in fermenting grains. The process of saccharification is the premise of Baijiu production. But little is known about the protein composition associated with the activity of saccharifying enzymes in daqu. This study investigated related proteins between nongxiangxing daqu and jiangxiangxing daqu by metaproteomics. We present that the amylase system (α‐amylase, glucoamylase and α‐glucosidase) and cellulase system (cellobiohydrolases, endoglucanase and β‐glucosidase) are regarded as the main saccharifying enzymes. Combined with protein‐based taxonomic annotation, the key proteins involving in amylase system were mainly from Kroppenstedtia , Lichtheimia , Byssochlamys and Thermomyces , while Thermoascu contributed the most to cellulase system. Moreover, it was found that the up‐regulated amylase proteins at thermophilic fermentation period in jiangxiangxing daqu in comparison with nongxiangxing daqu were affiliated to Aspergillus , Rhizomucor , Byssochlamys and Thermomyces, along with Thermoascu contributing to cellulase system. These findings will provide clues for improving saccharification efficiency in Baijiu production.

采用宏基因组学技术对8种中日酒曲中微生物群落结构进行解析.研究表明,邛崃米曲MB和清香型小曲QX中主要有根霉属(Rhizopus);2种黄酒麦曲TH和FHC物种组成相对丰富,主要为糖多孢菌属(Sacchar-opolyspora)和横梗霉属(Lichtheimia)等;4种日本酒曲(NP_MS、NP_SJR、NP_SJW和NP_SY)均主要为曲霉属(As-pergillus,均＞91％),其中主要为米曲霉(Aspergillus oryzae)、黄曲霉(Aspergillus flavus)、白曲霉(Aspergillus kawachii)和黑曲霉(Aspergillus niger).8种中日酒曲理化指标检测结果显示,日本酒曲的平均糖化力、液化力和还原糖含量均高于中国酒曲,中国酒曲的平均发酵力略高于日本酒曲.8种中日酒曲理化指标与其微生物种群结构具有明显相关性.此外,根据8种中日酒曲KEGG(Kyoto Encyclopedia of Genes and Genomes)功能注释发现,中日酒曲内部微生物代谢活动均较为活跃,其中有关碳水化合物代谢和氨基酸代谢的相关基因表达量占比最高,且碳水化合物代谢在8种中日酒曲之间具有显著性差异.研究结果揭示了8种中日酒曲之间微生物种群结构的差异,为后续解析中日酒曲中微生物代谢机理,并筛选特定功能微生物,开发功能型强化酒曲提供了理论基础.

Luzhou-flavor baijiu (LFB) is brewed by the combined action of various microorganisms, and its flavor is affected by the microbial community and the genes they express, but which genes are the key ones during LFB brewing is less clear. Based on our previous studies the genes ME2 and adhE were identified as key genes, but which role they play was also unknown. In this study functional microorganisms were screened based on the key genes ME2 and adhE, and they were identified to be Rummeliibacillus suwonensis, Clostridium tyrobutyricum and Lactobacillus buchneri. Then simulated fermentation experiments were carried out with the functional microorganisms, and during the fermentation process expression of the key genes and the amounts of the main flavors were detected to analyze the role of the key genes. The results showed that the key gene ME2 was significantly positively correlated with the contents of the main acids, however the key gene adhE and the formation of the main esters in the LFB brewing process was a significant positive correlation. This study verified the two key genes ME2 and adhE complement each other in the LFB brewing process, playing an important role in promoting the formation of flavor substances, and are very beneficial to improve the quality of LFB.

Chinese Jiang-flavor Baijiu is the most widely consumed liquor. Jiang-flavor Daqu, a fermentation starter, is important sources of key flavors of Jiang-flavor Baijiu. Some microbes play significant roles in flavor formation of Daqu. In order to clarify the microbial population that promotes the formation of Daqu flavor, we use high throughput sequencing technology combined with headspace solid-phase microextraction gas chromatography-mass spectrometry to investigate microbial population and volatile compounds in Jiang-flavor Daqu. In addition, the dynamic changes of physicochemical factors and enzyme activities in Jiang-flavor Daqu were investigated. Correlations between microbial population, volatile compounds, physicochemical factors, and enzyme activities of Jiang-flavor Daqu were disclosed by redundancy analysis and Spearman correlation analysis. A total of 66 volatile compounds were identified and 14 primary microorganisms were selected. Results showed that high temperature environment could promote the formation of acids, aldehydes and ketones, phenols, furans by affecting the growth of Monascus, Trichomonascus, Cutaneotrichosporon, Wallemia, Millerozyma, Nigrospora, Cladosporium, Bacillus, and Pediococcus in the early fermentation stage. While high nitrogen environment was more suitable for the growth of Virgibacillus and Kroppenstedtia, who could promote the formation of pyrazines in the late fermentation stage. Practical applications This study has provided a scientific basis for the directed regulation of Daqu fermentation through physicochemical factors, developed scientific basis for artificially constructing Daqu microbial population and obtaining an easy-to-operate, reproducible fermentation system for Daqu production.

Chinese Baijiu, the national liquor of China, is also one of the world-renowned distilled spirits. Jiuqu, a fermentation starter, is the masterpiece of traditional food biotechnology with a long history in China. Compared to western brewing technology of using malt and yeast in stages, the mode of using jiuqu (or combined with pit mud, a activated sludge) for Baijiu brewing is recognized in China. So far, culture-dependent, molecular biology technology and omics approaches have provided deeper insights into study the jiuqu microbiome. Jiuqu is considered to be a microecosystem with diverse and unique microorganisms, in which fascinating features of saccharification, alcohol fermentation, and flavors generation appear. However, knowledge about microbial diversity of jiuqu and their roles involved in Baijiu fermentation is still fragmentary and not systematic enough. Therefore, this review introduced and compared three basic types of jiuqu starter: daqu, xiaoqu and fuqu. We described in detail the microbial succession during preparation of jiuqu, aiming to emphasize that succession process promotes to formation of its stable distribution of microbial community. We also summarized the main species originated from jiuqu and their various properties. Furthermore, reported results about fermentation features of saccharification, alcohol fermentation, and flavors generation in jiuqu microbiome were generalized, and these features were emphasized to play the potential role in Baijiu brewing. Although the current work is limited, promising clues can be traced for future investigation on comprehensive metabolic processes in jiuqu. • The various microorganisms present in jiuqu, and each has its specific property. • Microbial succession in preparation promotes to distribution of microbial community. • Jiuqu contributes the potential of saccharification, fermentation and flavors generation to Baijiu brewing.