Lentibacillus daqui sp. nov., isolated from high-temperature Daqu, a starter for production of Chinese Jiang-flavour Baijiu.

A Gram-stain-positive, rod-shaped, endospore-forming, aerobic bacterial strain, designated ZS110521T, was isolated from high-temperature Daqu, a starter for production of Chinese Jiang-flavour Baijiu and was characterised by polyphasic taxonomy. This novel isolate grew in the presence of 0-20 % (w/v) NaCl, at pH 6.0-9.0 and 20-50 °C; optimum growth was observed with 8-10 % (w/v) NaCl, at pH 7.0 and 37 °C. A comparative analysis of the 16S rRNA gene sequence (1460 bp) of ZS110521T revealed that it displayed the highest similarity to Lentibacillus populi WD4L-1T (95.5 %), followed by Lentibacillus garicola SL-MJ1T (95.4 %) and Lentibacillus lacisalsi BH260T (95.2 %). ANI and dDDH values between ZS110521T and other strains of species of the genus Lentibacillus were less than 78 and 28 %, respectively. The predominant cellular fatty acids (> 10 %) of ZS110521T were anteiso-C17 : 0 (37.8 %), anteiso-C15 : 0 (28.1 %) and iso-C16 : 0 (15.5 %). The respiratory quinone was identified as menaquinone-7 (MK-7) and the major polar lipids were diphosphatidylglycerol and phosphatidylglycerol. The polyphasic taxonomic data and the results of chemotaxonomic analysis confirmed that ZS110521T represents a novel species, for which the name Lentibacillus daqui sp. nov. is proposed. The type strain of this proposed species is ZS110521T (=CGMCC 1.19456T =JCM 35213T).

Solibacillus daqui sp. nov., isolated from high-temperature Daqu.

A Gram-stain-positive, rod-shaped, endospore-forming, aerobic bacterial strain, designated ZS111008T, was isolated from high-temperature Daqu, a starter for production of Chinese Jiang-flavour Baijiu, and was characterized by polyphasic taxonomy. This novel isolate grew in the presence of 0-5 % (w/v) NaCl, at pH 6.0-9.0 and 25-45 °C; optimum growth was observed with 1 % (w/v) NaCl, at pH 8.0 and 30 °C. A comparative analysis of the 16S rRNA gene sequence (1461 bp) of strain ZS111008T showed highest similarity to Solibacillus silvestris DSM12223T (96.7%), followed by Solibacillus cecembensis PN5T (96.6%) and Solibacillus isronensis AMCK01000046 (96.5%). The DNA G+C content of strain ZS111008T was 37.21 mol%. The respiratory quinone was identified as menaquinone-7 and the major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine and one unknown phospholipid. Lys was detected as the diagnostic diamino acid in the cell wall. Based on morphological characteristics, chemotaxonomic characteristics and physiological properties, strain ZS111008T represents a novel species of the genus Solibacillus, for which the name Solibacillus daqui sp. nov. is proposed. The type strain for this proposed species is ZS111008T (=CGMCC 1.19455T=JCM 35214T).

Metabolite-Based Mutualistic Interaction between Two Novel Clostridial Species from Pit Mud Enhances Butyrate and Caproate Production.

Pit mud microbial consortia play crucial roles in the formation of Chinese strong-flavor baijiu's key flavor-active compounds, especially butyric and caproic acids. Clostridia, one of the abundant bacterial groups in pit mud, were recognized as important butyric and caproic acid producers. Research on the interactions of the pit mud microbial community mainly depends on correlation analysis at present. Interaction between Clostridium and other microorganisms and its involvement in short/medium-chain fatty acid (S/MCFA) metabolism are still unclear. We previously found coculture of two clostridial strains isolated from pit mud, Clostridium fermenticellae JN500901 (C.901) and Novisyntrophococcus fermenticellae JN500902 (N.902), could enhance S/MCFA accumulation. Here, we investigated their underlying interaction mechanism through the combined analysis of phenotype, genome, and transcriptome. Compared to monocultures, coculture of C.901 and N.902 obviously promoted their growth, including shortening the growth lag phase and increasing biomass, and the accumulation of butyric acid and caproic acid. The slight effects of inoculation ratio and continuous passage on the growth and metabolism of coculture indicated the relative stability of their interaction. Transwell coculture and transcriptome analysis showed the interaction between C.901 and N.902 was accomplished by metabolite exchange, i.e., formic acid produced by C.901 activated the Wood-Ljungdahl pathway of N.902, thereby enhancing its production of acetic acid, which was further converted to butyric acid and caproic acid by C.901 through reverse ß-oxidation. This work demonstrates the potential roles of mutually beneficial interspecies interactions in the accumulation of key flavor compounds in pit mud. IMPORTANCE Microbial interactions played crucial roles in influencing the assembly, stability, and function of the microbial community. The metabolites of pit mud microbiota are the key to flavor formation of Chinese strong-flavor baijiu. So far, researches on the interactions of the pit mud microbial community have been mainly based on the correlation analysis of sequencing data, and more work needs to be performed to unveil the complicated interaction patterns. Here, we identified a material exchange-based mutualistic interaction system involving two fatty acid-producing clostridial strains (Clostridium fermenticellae JN500901 and Novisyntrophococcus fermenticellae JN500902) isolated from pit mud and systematically elucidated their interaction mechanism for promoting the production of butyric acid and caproic acid, the key flavor-active compounds of baijiu. Our findings provide a new perspective for understanding the complicated interactions of pit mud microorganisms.

Mining the Factors Driving the Evolution of the Pit Mud Microbiome under the Impact of Long-Term Production of Strong-Flavor Baijiu.

The mud cellar creates a unique microenvironment for the fermentation of strong-flavor baijiu (SFB). Recent research and long-term practice have highlighted the key roles of microbes inhabiting pit mud in the formation of SFB's characteristic flavor. A positive correlation between the quality of SFB and cellar age was extracted from practice; however, the evolutionary patterns of pit mud microbiome and driving factors remain unclear. Here, based on the variation regularity analysis of microbial community structure and metabolites of samples from cellars of different ages (∼30/100/300 years), we further investigated the effects of lactate and acetate (main microbial metabolites in fermented grains) on modulating the pit mud microbiome. Esters (50.3% to 64.5%) dominated the volatile compounds identified in pit mud, and contents of the four typical acids (lactate, hexanoate, acetate, and butyrate) increased with cellar age. Bacteria (9.5 to 10.4 log10 [lg] copies/g) and archaea (8.3 to 9.1 lg copies/g) mainly constituted pit mud microbiota, respectively dominated by Clostridia (39.7% to 81.2%) and Methanomicrobia (32.8% to 92.9%). An upward trend with cellar age characterized the relative and absolute abundance of the most predominant bacterial and archaeal genera, Caproiciproducens and Methanosarcina. Correlation analysis revealed significantly (P < 0.05) positive relationships between the two genera and major metabolites. Anaerobic fermentation with acetate and lactate as carbon sources enhanced the enrichment of Clostridia, and furthermore, the relative abundance of Caproiciproducens (40.9%) significantly increased after 15-day fed-batch fermentation with lactate compared with the initial pit mud (0.22%). This work presents a directional evolutionary pattern of pit mud microbial consortia and provides an alternative way to accelerate the enrichment of functional microbes. IMPORTANCE The solid-state anaerobic fermentation in a mud cellar is the most typical feature of strong-flavor baijiu (SFB). Metabolites produced by microbes inhabiting pit mud are crucial to create the unique flavor of SFB. Accordingly, craftspeople have always highlighted the importance of the pit mud microbiome and concluded by centuries of practice that the production rate of high-quality baijiu increases with cellar age. To deepen the understanding of the pit mud microbiome, we determined the microbial community and metabolites of different-aged pit mud, inferred the main functional groups, and explored the forces driving the microbial community evolution through metagenomic, metabolomic, and multivariate statistical analyses. The results showed that the microbial consortia of pit mud presented a regular and directional evolutionary pattern under the impact of continuous batch-to-batch brewing activities. This work provides insight into the key roles of the pit mud microbiome in SFB production and supports the production optimization of high-quality pit mud.

Novisyntrophococcus fermenticellae gen. nov., sp. nov., isolated from an anaerobic fermentation cellar of Chinese strong-flavour baijiu.

A Gram-stain-negative, coccus-shaped, obligately anaerobic, non-motile and non-spore-forming bacterium, designated strain JN500902T, was isolated from the mud in a fermentation cellar used continuously over 30 years for Chinese strong-flavour baijiu production. Colonies were white, circular, convex and smooth-edged. Growth was observed at 20-40 °C (optimum, 37 °C), at pH 5.0-10 (optimum, pH 7.5), with 0-2 % (w/v) NaCl and with 0-4 % (v/v) ethanol. The Biolog assay demonstrated positive reactions of strain JN500902T in the metabolism of l-fucose and pyruvate. The predominant cellular fatty acids (>10 %) consisted of C16 : 0 and C14 : 0. The major end metabolites of strain JN500902T were acetic acid and ethanol when incubated anaerobically in liquid reinforced clostridial medium. Acetate was the major organic acid end product. The complete genome size of strain JN500902T was 3 420 321 bp with 3327 identified genes. The G+C content was 43.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences affiliated strain JN500902T with the family Lachnospiraceae, having low sequence similarity (92.8 %) to the nearest type strain, Syntrophococcus sucromutans DSM 3224T and forming a clearly distinct branch. Core genome phylogenetic analysis of the isolate and 134 strains belonging to the family Lachnospiraceae also revealed that strain JN500902T was well-separated from other genera of this family as a monophyletic clade. The average nucleotide identity and amino acid identity values between strain JN500902T and 134 Lachnospiraceae strains were less than 74 and 65 %, respectively. Considering its polyphasic characteristics, strain JN500902T represents a novel genus and species within the family Lachnospiraceae, for which the name Novisyntrophococcus fermenticellae gen. nov., sp. nov. is proposed. The type strain is JN500902T (=CICC 24502T=JCM 33939T).

Reclassification of Olsenella gallinarum as Thermophilibacter gallinarum comb. nov. and description of Thermophilibacter immobilis sp. nov., isolated from the mud in a fermentation cellar used for the production of Chinese Luzhou-flavour Baijiu.

A novel Gram-stain-positive, strictly anaerobic, elliptical, non-motile and non-flagellated bacterium, designed LZLJ-2T, was isolated from the mud in a fermentation cellar used for the production of Chinese Luzhou-flavour Baijiu. Growth occurred at 28-45 °C (optimum, 37 °C), at pH 6.0-7.0 (optimum, pH 6.0) and with concentrations of NaCl up to 2 % (w/v; optimum, 0 %). On the basis of 16S rRNA gene sequence similarity, strain LZLJ-2T belonged to the genus Thermophilibacter and was most closely related to Thermophilibacter mediterraneus Marseille-P3256T (similarity 96.9 %), Olsenella gallinarum ClaCZ62T (similarity 96.6 %) and Thermophilibacter provencensis Marseille-P2912T (similarity 96.4 %). In addition, strain LZLJ-2T had high similarity to the genus Olsenella, including Olsenella profusa DSM 13989T (similarity 94.9 %), Olsenella umbonata DSM 22620T (similarity 94.9 %), Olsenella uli ATCC 49627T (similarity 94.22 %), Tractidigestivibacter scatoligenes DSM 28304T (similarity 93.9 %) and Paratractidigestivibacter faecalis KCTC 15699T (similarity 93.25 %). Comparative genome analysis showed that orthoANI values between strain LZLJ-2T and Thermophilibacter mediterraneus Marseille-P3256T, Olsenella gallinarum ClaCZ62T, Thermophilibacter provencensis Marseille-P2912T, Olsenella profusa DSM 13989T, Olsenella umbonata DSM 22620T, Olsenella uli ATCC 49627T, Tractidigestivibacter scatoligenes DSM 28304T and Paratractidigestivibacter faecalis KCTC 15699T were 78.68, 78.99, 78.29, 73.40, 74.00, 74.30, 75.08 and 77.23 %, and the genome-to-genome distance values were respectively 22.3, 22.5, 22.4, 19.6, 20.5, 19.7, 20.5 and 21.5 %. The genomic DNA G+C content of strain LZLJ-2T was 65.21 mol%. The predominant cellular fatty acids (>10 %) of strain LZLJ-2T were C18 : 1 cis 9 (33.7 %), C14 : 0 (22.0 %) and C18 : 1 cis 9 DMA (13.5 %). d-Glucose, sucrose, mannose, maltose, lactose (weak), salicin, glycerol (weak), cellobiose and trehalose (weak) could be used by strain LZLJ-2T as sole carbon sources. Enzyme activity results showed positive reactions with valine arylamidase, leucine arylamidase, crystine arylamidase, acid phosphatase, alkaline phosphatase, esterase (C4) (weakly positive), naphthol-AS-BI-phosphohydrolase, α-glucosidase and ß-glucosidase. The major end products of glucose fermentation were lactic acid and acetic acid. It produced skatole from indole acetic acid, and produced p-cresol from modified peptone-yeast extract medium with glucose. Based on the 16S rRNA gene trees as well as the genome core gene tree, it is suggested that Olsenella gallinarum are transferred to genus Thermophilibacter as Thermophilibacter gallinarum comb. nov. Based on phenotypic, genotypic and phylogenetic data, strain LZLJ-2T is considered to represent a novel species of the genus Thermophilibacter, for which the name Thermophilibacter immobilis sp. nov. is proposed. The type strain is LZLJ-2T (=KCTC 25162T=JCM 34224T).

Daqu microbiota exhibits species-specific and periodic succession features in Chinese baijiu fermentation process.

Daqu, a brick-shaped product spontaneously fermented under an open environment, has been regarded as the starter of fermentation, raw enzyme preparation and raw materials for baijiu production. However, its contribution in baijiu fermentation has not been fully elaborated yet. Here, the effects of daqu microbiota on baijiu fermentation were investigated under both field-scale and lab-scale conditions. In field-scale baijiu fermentation, the dominant daqu microbes (average relative abundance>10.0%), including unclassified_Leuconostocaceae, Thermoascus, and Thermomyces, tended to dominate the early stage (0-7 d). However, the rare daqu microbes (average relative abundance <0.1%, e.g., Kazachstania) tended to dominate the middle and late stages (11-40 d). In addition, some genera showed differences in species diversity between daqu and fermented grains. The average relative abundance of Lactobacillus was over 75% during baijiu fermentation, and most of them were affiliated with Lactobacillus acetotolerans, while Lactobacillus crustorum dominated the Lactobacillus OTUs in daqu. The similar patterns were also observed during lab-scale baijiu fermentation. The results of function prediction showed the enriched metabolic pathways were associated with glycolysis and long-chain fatty acid esters in baijiu fermentation. These results improved the understanding of daqu microbiota function during baijiu fermentation and provided a basic theory to support the regulation of baijiu production.

Microbiota stratification and succession of amylase-producing Bacillus in traditional Chinese Jiuqu (fermentation starters).

BACKGROUND: Jiuqu are vital saccharifying and fermenting agents for Chinese fermented foods. Natural ventilation during Jiuqu fermentation causes changes in temperature, oxygen and moisture content, resulting in mass and heat gradients from the outer to inner areas of Jiuqu blocks. In the present study, microbiota stratification in Jiuqu was investigated by single molecule real-time sequencing and culture isolation. The contributors of Bacillus to amylase activity of Jiuqu and the dynamics of their biomass during Jiuqu fermentation were also analyzed. RESULTS: The dominant orders, genera and species between the inner and outer layers of Huangjiu qu (HJQ) were similar, although they displayed greater variance in two layers of Baijiu qu (BJQ). Bacillus possessed the highest diversity (including 27 species) in Jiuqu. Bacillus licheniformis, Bacillus altitudinis, Bacillus subtilis, Bacillus amyloliquefaciens and Bacillus megaterium were most prevalent in HJQ, whereas B. licheniformis, B. amyloliquefaciens and Bacillus cereus were dominant in BJQ. Isolates of B. amyloliquefaciens, B. subtilis and B. cereus exhibited high activities of amylase and glucoamylase. Quantification of Bacillus members possessing genes of α-amylase revealed that B. cereus and B. licheniformis were the most dominant microbes to secret α-amylase in Jiuqu and their biomass were increasing during Jiuqu fermentation. CONCLUSION: The present study demonstrates the microbial distribution in different layers of Jiuqu and clarifies the Bacillus species processing the activity of α-amylase. These results will help industries control the quality of Jiuqu by rationally selecting starters and optimizing their microbiota. © 2020 Society of Chemical Industry.

Clostridium fermenticellae sp. nov., isolated from the mud in a fermentation cellar for the production of the Chinese liquor, baijiu.

A novel Gram-stain-positive, rod-shaped, obligately anaerobic, non-motile, spore-forming and binary fission encapsulated bacterium, designated strain JN500901T, was isolated from a mud cellar which has been continuously used for the fermentation of Chinese strong-flavour baijiu for over 100 years. Growth of JN500901Toccurred at pH 4.5-8.0 (optimum, pH 5.0), 20-40 °C (37 °C), 0-2 % (w/v) NaCl and 0-10 % (v/v) ethanol. The Biolog assay revealed that strain JN500901T metabolized d-fructose, l-fucose, isomaltulose and l-rhamnose among the 95 studied carbon sources. p-Cresol was the predominant volatile metabolite in the fermentation broth of strain JN500901T incubated in liquid reinforced clostridial medium under anaerobic conditions. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain JN500901T belongs to Clostridiumsensu stricto, and shared the highest sequence similarity to Clostridiumcarboxidivorans DSM 15243T (94.2 %), followed by Clostridiumscatologenes DSM 757T (94.1 %). The dominant cellular fatty acids (>10 %) were C16 : 0 FAME (36.6 %), C19 : 0 cyc 9,10 DMA (19.8 %) and C16 : 1 cis 9 DMA (11.8 %). The complete genome of strain JN500901T contained a circular chromosome of 2.812 Mb with 2611 genes and 31.0 mol% G+C content. Comparative genome analysis of the strain JN500901T, Clostridiumcarboxidivorans DSM 15243T and Clostridiumscatologenes DSM 757T revealed 74.5 and 74.8 % average nucleotide identity, respectively. Based on the phenotypic, biochemical and phylogenetic analyses presented here, strain JN500901T is considered to be a novel species of the genus Clostridiumsensustricto, for which the name Clostridium fermenticellae sp. nov. is proposed. The type strain is JN500901T (=CICC 24501T=JCM 32827T).

Bio-Heat Is a Key Environmental Driver Shaping the Microbial Community of Medium-Temperature Daqu.

"Daqu" is a saccharifying and fermenting agent commonly used in the traditional solid-state fermentation industry (e.g., baijiu and vinegar). The patterns of microbial community succession and flavor formation are highly similar among batches, yet the mechanisms promoting temporal succession in the Daqu microbial ecology remain unclear. Here, we first correlated temporal profiles of microbial community succession with environmental variables (temperature, moisture, and titratable acidity) in medium temperature Daqu (MT-Daqu) throughout fermentation. Temperature dynamics significantly correlated (P < 0.05) with the quick succession of MT-Daqu microbiota in the first 12 d of fermentation, while the community structure was relatively stable after 12 d. Then, we explored the effect of temperature on the MT-Daqu community assembly. In the first 4 d of fermentation, the rapid propagation of most bacterial taxa and several fungal taxa, including Candida, Wickerhamomyces, and unclassified Dipodascaceae and Saccharomycetales species, significantly increased MT-Daqu temperature to 55°C. Subsequently, sustained bio-heat generated by microbial metabolism (53 to 56°C) within MT-Daqu inhibited the growth of most microbes from day 4 to day 12, while thermotolerant taxa, including Bacillus, unclassified Streptophyta, Weissella, Thermoactinomyces, Thermoascus, and Thermomyces survived or kept on growing. Furthermore, temperature as a major driving force on the shaping of MT-Daqu microbiota was validated. Lowering the fermentation temperature by placing the MT-Daqu in a 37°C incubator resulted in decreased relative abundances of thermotolerant taxa, including Bacillus, Thermoactinomyces, and Thermoascus, in the MT-Daqu microbiota. This study revealed that bio-heat functioned as a primary endogenous driver promoting the formation of functional MT-Daqu microbiota.IMPORTANCE Humans have mastered the Daqu preparation technique of cultivating functional microbiota on starchy grains over thousands of years, and it is well known that the metabolic activity of these microbes is key to the flavor production of Chinese baijiu. The pattern of microbial community succession and flavor formation remains highly similar between batches, yet mechanistic insight into these patterns and into microbial population fidelity to specific environmental conditions remains unclear. Our study revealed that bio-heat was generated within Daqu bricks in the first 4 d of fermentation, concomitant with rapid microbial propagation and metabolism. The sustained bio-heat may then function as a major endogenous driving force promoting the formation of the MT-Daqu microbiota from day 4 to day 12. The bio-heat-driven growth of thermotolerant microorganisms might contribute to the formation of flavor metabolites. This study provides useful information for the temperature-based modulation of microbiota function during the fermentation of Daqu.

[Effect of dihydroartemisinin combined irradiation on the apoptosis of human lung cancer GLC-82 cells and its mechanism study].

OBJECTIVE: To study the effect of dihydroartemisinin (DHA) combined irradiation on the apoptosis of human lung cancer GLC-82 cells and to study its mechanism. METHODS: The growth inhibition rate of GLC-82 cells acted by different concentrations DHA was detected using MTT assay at 24, 48, and 72 h, respectively. Clone forming test was used. With multi-target single-hit model, the radiosensitization effect was assessed by calculating sensitizing enhancement ratio (SER).The effect of DHA combined irradiation on the apoptosis of GLC-82 cell cycle distribution and apoptosis were measured by flow cytometry. The protein expression of p53, p21, Bcl-2, and Bax were detected by Western blot. RESULTS: Different concentrations DHA (4, 8, 16, 32, 64, and 128 µg/mL) had cytotoxicity on GLC-82 cells. The IC50 for 24, 48, and 72 h was 38.25,20.58, and 10.36 µg/mL, respectively, in obvious dose- and time-dependent manner. The growth inhibition rate was more significantly increased than that of the blank control group (P < 0.01, P<0.05). DHA had sensitization enhancement effect on GLC-82 cells, with SER of 1.4. DHA combined irradiation could obviously change the structure of GLC-82 cells cell cycle and induce apoptosis (with the apoptosis rate of 21.5%), which was significantly different from that of the blank control group (P < 0.05). Western blot showed the expression of p53 and p21 protein could be increased by DHA combined irradiation, and the expression of Bcl-2 protein down-regulated (P <0.01, P <0. 05). CONCLUSIONS: DHA had stronger cytotoxicity and radiosensitization on GLC-82 cells. Its mechanisms might lie in making the arrest of GLC-82 cells' growth at G0/G1 phase, decreasing the ratio of cells at S phase, restoring the function of p53, decreasing the expression of Bcl-2 protein, and inducing apoptosis in GLC-82 cells.

[Studies on new co-processed excipient consisting of lactose and gelatinized starch].

Co-processed excipients withgelatinized or non-gelatinized starch were prepared by spray drying. Powder and tablet properties of corocessed excipients prepared were compared with those of physical mixtures and spray-dried lactose. Their applicability in traditional Chinese medicine (TCM) powder tableting was tested on two TCM extracts, i.e., the gardenia extract and the Herba Sedi extract. It was shown that gelatinizing starch before co-spray drying with lactose could improve the performance and efficiency of starch as a binder, resulting in remarkable improvement in physicomechanical properties of co-processed excipients prepared. Conpared to self-made and commercially available spray-dried lactose, co-processed excipients achieved better compactability and higher drug loading for TCM extracts. In conclusion, the lactose-gelatinized starch co-processed excipient, with excellent physicomechanical properties, is promising to be explored as a new excipient for direct tableting.

Decoding the Qu-aroma of medium-temperature Daqu starter by volatilomics, aroma recombination, omission studies and sensory analysis.

Qu-aroma is of great significance for evaluation the quality of Daqu starter. This study aimed to decode the Qu-aroma of medium-temperature Daqu (MT-Daqu) via "top-down" and "bottom-up" approaches. Firstly, 52 aroma descriptors were defined to describe the MT-Daqu aroma by quantitative descriptive analysis. Secondly, 193 volatile organic compounds (VOCs) were identified from 42 MT-Daqu samples by HS-SPME-GC-MS, and 43 dominant VOCs were screened out by frequence of occurrence or abundance. By Thin Film (TF)-SPME-GC-O-MS, 27 odors and 90 VOCs were detected in MT-Daqu mixture, and 14 odor-active VOCs were screened out by odor intensity. Thirdly, a five-level MT-Daqu aroma wheel was constructed by matching 52 aroma descriptors and 37 aroma-active VOCs. Finally, Qu-aroma of MT-Daqu was reconstructed with 37 aroma-active VOCs and evaluated by omission experiments. Hereinto, 26 key aroma-active VOCs were determined by OAV value ≥1, including isovaleric acid, 1-hexanol, isovaleraldehyde, 2-octanone, trimethylpyrazine, γ-nonalactone, 4-vinylguaiacol, etc.

[Effects of gender on treatment strategies for elderly patients with acute coronary syndrome].

OBJECTIVE: To explore the effects of gender on treatment strategies for elderly patients with acute coronary syndrome (ACS). METHODS: March 2009 to March 2012, consecutive 619 aged ACS patients undergoing coronary angiography (CA) were screened at our hospital. There were 273 females and 346 males. Risk factors, ACS diagnosis, CA results, treatments and prognosis were compared between female and male groups. RESULTS: The risk factors of body mass index, stroke history, smoking history, hemoglobin (Hb), serum cholesterol (TC), low density lipoprotein (LDL-C) and blood uric acid (UA) levels were significantly lower in female group than those in male group (P < 0.05). The morbidity of diabetes in female group was obviously higher than that in male group (27.8% vs 18.5%, P < 0.05). The prevalence of myocardial infarction history, percutaneous coronary intervention (PCI) and coronary artery bypass grafting (CABG) history in male group were significantly greater than that in female group (48.0% vs 39.9%, P < 0.05; 30.6% vs 22.3%, P < 0.05; 19.9% vs 10.3%, P < 0.01). The rate of combined multiple risk factors (3 or higher) increased significantly in female group (41.8% vs 29.8%, P < 0.05). The incidence of unstable angina pectoris (UAP) and non-ST segment elevation myocardial infarction (NSTEMI) in female group was greater, but there was no statistical significance. The rate of 3-vessel and calcification lesions in female group was significantly elevated compared with male group (36.26% vs 28.61%, P < 0.05). Regarding the choice of treatment strategy, conservative treatment was common in females, but there was no statistical significance between them. PCI, emergency PCI and selective CABG operation were performed more frequently in female group compared with male group (26.0% vs 14.2%, P < 0.01; 14.7% vs 6.6%, P < 0.01; 19.1% vs 7.7%, P < 0.01). The prognosis had no statistical significance between two groups. CONCLUSION: The treatment strategies have certain limitations for female ACS patients. And an more aggressive treatment should be offered to improve the prognosis.

Investigation on fresh bamboo replacing rice husks for brewing light-aroma Baijiu.

Husks are the main source of bran and furfural flavor in traditional Chinese light-aroma Baijiu, but they negatively affect its smell and taste. Here, bran husks were replaced with fresh bamboo to brew light-aroma Baijiu. Flavor components in Jiupei and Baijiu were detected through headspace solid-phase microextraction with gas chromatography-mass spectrometry, and physicochemical properties were assessed; flavor results were obtained from correlation, principal component, and cluster analyses. Starch and reducing sugar content in Jiupei negatively correlated with moisture, alcohol content, and acidity. Fresh bamboo reduced furfural from bran husks in Jiupei by 88.5% and increased alcohol distillation by 51%; it also improved starch efficiency (5%). Surprisingly, isovanillin was found to be present in Baijiu. Total Baijiu yield (57% ± 2.01%) was attained when crushed bamboo size was 1.5 cm × 0.3 cm × 0.3 cm. This study supports the use of fresh bamboo (an eco-friendly alternative for husks) in brewing light-aroma Baijiu. PRACTICAL APPLICATION: The use of fresh bamboo as a replacement for rice husks in brewing light-aroma Baijiu was investigated. It attenuated the chaff taste in light-aroma Baijiu and increased the liquor yield. Surprisingly, isovanillin was also present in the base Baijiu, and it added to the fragrance. This study not only supports the use of bamboo as an auxiliary material for brewing light-aroma Baijiu but also provides a reference for brewing light-aroma Baijiu with alternative auxiliary materials.

Machine learning based age-authentication assisted by chemo-kinetics: Case study of strong-flavor Chinese Baijiu.

The aged Chinese liquor, Baijiu, is highly valued for its superior organoleptic qualities. However, since age-authentication method and aging-mechanism elucidation of Baijiu is still in the exploratory stage, high-quality aged Baijiu is often replaced by lower-quality, less-aged product with fraudulent mislabeling. Authentic high-quality strong-flavor Baijiu was analyzed by gas chromatography-mass spectrometry. Total esters decreased with aging, while acids, alcohols, aldehydes, ketones, terpenes, pyrazines increased. Although concentrations of partial compounds showed non-monotonic profiling during aging, a close positive linear correlation (R2 = 0.7012) of Baijiu Evenness index (0.55-0.59) with aging time was observed, indicating a more balanced composition in aged Baijiu. The reaction quotient (Qc) of each esterification, calculated by the corresponding reactant and product concentration, approached to the corresponding thermodynamic equilibrium constant Kc. This result demonstrated that the spontaneous transformation driven by thermodynamics explained part of the aging compositional profiling. Furthermore, an aging-related feature selection and an age-authentication method were established based on three models combined with five ranking algorithms. Forty-one key features, including thirty-six compound concentrations, four esterification Qc values and the Evenness index were selected out. The age-authentication based on neural network using forty-one input features accurately predicted the age group of Baijiu samples (F1 = 100 %). These findings have deepened understanding of the Baijiu aging mechanism and provided a novel, effective approach for age-authentication of Baijiu and other liquors.

Bacteria and filamentous fungi running a relay race in Daqu fermentation enable macromolecular degradation and flavor substance formation.

As the saccharifying and fermentative agent, medium-temperature Daqu (MT-Daqu) plays an irreplaceable role in the production of strong-flavor Baijiu. Numerous studies have focused on the microbial community structure and potential functional microorganisms, however, little is known about the succession of active microbial community and the formation mechanism of community function during MT-Daqu fermentation. In this study, we presented an integrated analysis of metagenomics, metatranscriptomics, and metabonomics covering the whole fermentation process of MT-Daqu to reveal the active microorganisms and their participations in metabolic networks. The results showed that dynamic of metabolites were time-specific, and the metabolites and co-expressed active unigenes were further classified into four clusters according to their accumulation patterns, with members within each cluster displaying a uniform and clear pattern of abundance across fermentation. Based on KEGG enrichment analysis in co-expression clusters and succession of active microbial community, we revealed that Limosilactobacillus, Staphylococcus, Pichia, Rhizopus, and Lichtheimia were metabolically active members at the early stage, and their metabolic activities were conducive to releasing abundant energy to drive multiple basal metabolisms such as carbohydrates and amino acids. Thereafter, during the high temperature period and at the end of fermentation, multiple heat-resistant filamentous fungi were transcriptionally active populations, and they acted as both the saccharifying agents and flavor compound producers, especially aromatic compounds, suggesting their crucial contribution to enzymatic activity and aroma of mature MT-Daqu. Our findings revealed the succession and metabolic functions of the active microbial community, providing a deeper understanding of their contribution to MT-Daqu ecosystem.

Microbial biogeography of pit mud from an artificial brewing ecosystem on a large time scale: all roads lead to Rome.

IMPORTANCE: Baijiu is a typical example of how humans employ microorganisms to convert grains into new flavors. Mud cellars are used as the fermentation vessel for strong-flavor Baijiu (SFB) to complete the decomposition process of grains. The typical flavor of SFB is mainly attributed to the metabolites of the pit mud microbiome. China has a large number of SFB-producing regions. Previous research revealed the temporal profiles of the pit mud microbiome in different geographical regions. However, each single independent study rarely yields a thorough understanding of the pit mud ecosystem. Will the pit mud microbial communities in different production regions exhibit similar succession patterns and structures under the impact of the brewing environment? Hence, we conducted research in pit mud microbial biogeography to uncover the impact of specific environment on the microbial community over a long time scale.

[Characterization of flowability of pharmaceutical powders based on multivariate analysis method].

The main methods of characterizing the flowability of pharmaceutical powders include repose angle method, HR method, Carr's index method, Jenike flow function method, fractal dimension method, and mass flow rate method, etc. Regarding powders with different flowabilities as the research subject, comprehensive features of pharmaceutical materials were investigated and characterized. The multivariate analysis method was employed to evaluate and analyze flowability values of the tested pharmaceutical materials. Comparing with the method of the mass flow rate, it was feasible to use multivariate analysis method to evaluate the flowability of powders. Simultaneously, the flowability of pharmaceutical materials could be ranked and definitely quantified, and critical values be determined according to the actual production, which has promoted the previous methods dependent only on the single parameter, i.e. repose angle and compression degree methods. A relatively objective standard method of evaluating flowability of powders is formed.