Tank fermentation is a novel fermentation technology; however, the role of microorganisms in Liupao tea quality is unknown. Using high-throughput sequencing, untargeted metabolomics, and targeted absolute quantification, the microbial community, chemical profile, metabolites, and metabolic pathway of Liupao tea were discovered. Sphingomonas, Aquabacterium, Wolbachia, Blastobotrys, Aspergillus, Rasamsonia, and Candida were found to be the most common microorganisms, and throughout fermentation, Blastobotrys was the dominant fungal genus. A total of 40 compounds were identified as differentially changed metabolites (p < 0.05, VIP >1) (VIP: variable important in projection) and significantly contributing to tank fermentation, flavonoids and amino acid derivatives decreased, while nucleotides significantly increased. Gallic acid, EGCG, CG, EGC, EC, etc. could be identified as markers to assess the degree or completion of Liupao tea fermentation. There were significant (p < 0.01) correlations between microbial populations and metabolites, with Sphingomonas being significantly (p < 0.01) associated with 27 metabolites and Blastobotrys being significantly (p < 0.01) associated with 12 metabolites. The metabolic pathways of catechins, flavonoids, and nucleotides elucidate the factors influencing tea quality formation. This research improved our understanding of how Liupao tea processing affects its chemical composition and quality formation.

Tank fermentation is a novel approach to fermenting teas; however, the species of microorganisms present remain unclear. The microbial community composition of Liupao tea at various stages of tank fermentation was analyzed using high-throughput sequencing. Sphingomonas, Aquabacterium, Pelomonas, Acinetobacter, Blastobotrys, Aspergillus, Debaryomyces, and Aureobasidium were the predominant genera, which is different from pile fermentation. Fifteen genera (including Lactobacillus, Debaryomyces, Candida, Allobaculum, Flavobacterium, Caulobacter, Blastobotrys, Aspergillus, and Rasamsonia) were identified as biomarkers. PICRUSt analysis predicted that the most abundant functional genes were related to metabolism of carbohydrates, amino acids, cofactors, vitamins, and other secondary metabolites. Using the pure culture method, 283 strains were isolated at various stages of fermentation, representing 20 genera and 29 species of bacteria, and 11 genera and 18 species of fungi. Most of the dominant Sphingomonas, Staphylococcus, Aspergillus, and Blastobotrys identified by sequencing were also isolated. Of these, Sphingomonas olei, Aspergillus luchuensis, Aspergillus niger, Aspergillus aculeatus, Aspergillus amstelodami, Blastobotrys adeninivorans, Candida metapsilosis, and Candida blankii were beneficial strains that might be used to ferment Liupao tea. This study provides a basis for the development of processing technologies and utilization of microbial strains in the production of dark teas. PRACTICAL APPLICATION: Microbial diversity in tank-fermented Liupao tea was reported for the first time. 8 microorganisms were the predominant genera. The species, functions and potential risks of microorganisms was revealed. We clarified the differences between tank and pile fermentation.

为探索六堡茶发酵过程中多酚氧化酶(polyphenol oxidase,PPO)的来源及其酶学活性,采用不同pH值缓冲液、保护剂和液料比对发酵前后六堡茶PPO进行提取,通过(NH4)2SO4沉淀、透析和Sephadex?G-100凝胶过滤层析纯化PPO.结果表明:从六堡茶中分离纯化得到2个PPO同工酶,分子质量分别为59?kDa(PPO1)和70?kDa(PPO2),其中PPO1为茶树PPO,PPO2由发酵过程中微生物产生.底物邻苯二酚对PPO1和PPO2的最适浓度为0.5?mol/L,PPO1和PPO2的最适pH值分别为6.5和6.0,最适温度分别为40?℃和35?℃,PPO1的热稳定性优于PPO2,两者在90?℃条件下处理5?min几乎完全灭酶.研究表明,六堡茶发酵后会产生新的同工酶,二者共同提高黑茶品质,为黑茶加工条件的控制和黑茶PPO的研究提供一定的理论基础.

探究六堡茶渥堆发酵过程中乳酸菌多样性并筛选具有降胆固醇活性菌株,旨在评价其作为潜在益生菌的体外降胆固醇能力,预防人体因饮食不均衡导致的高血脂症.采用高通量测序、纯培养法分析不同渥堆发酵时期六堡茶中乳酸菌多样性,并对其体外降胆固醇能力、胃肠耐受性进行研究.结果表明,从25个样品中共检测到乳酸菌4个属(乳杆菌属(Lactobacillus)、魏斯氏菌属(Weissella)、乳球菌属(Lactococcus)和肠球菌属(Enterococcus))5个种,其中乳杆菌属为优势菌.共分离得到76株乳酸菌,包括55株戊糖乳杆菌(Lactobacillus pentosus)、3株铅黄肠球菌(Enterococcus casseliflavus)、5株类肠膜魏斯氏菌(Weissella paramesenteroides)、8株希腊魏斯氏菌(Weissella hellenica)、5株台湾乳球菌(Lactococcus taiwanensis).通过乳酸菌体外降胆固醇实验,发现胆固醇去除率为0％～30％,其中戊糖乳杆菌L131的去除胆固醇率最高,为(29.56±0.37)％;在pH值分别约为3.0、8.0的人工胃液、人工胰液中,L131有最高存活率,分别为(92.70±0.71)％和(77.54±4.81)％,显著高于其他菌株(P<0.05).本研究探明六堡茶渥堆发酵过程中乳酸菌的多样性,且发现具有降胆固醇特性的优良乳酸菌株,为健康黑茶产品和茶源益生菌的开发提供理论基础.