为了在普洱茶渥堆发酵中筛选出可降解咖啡碱的菌株,采用咖啡碱液体筛选培养基从不同阶段普洱茶发酵样中筛选目标菌株,结合菌落特征、分生孢子结构以及18βS rDNA序列对目标菌株进行鉴定。并将目标菌株接种至茶叶内进行单菌种发酵,通过高效液相色谱法（HPLC）测定咖啡碱、茶碱、可可碱、3-甲基黄嘌呤等嘌呤碱含量。结果表明,在普洱茶发酵样中筛选出一株有效降解咖啡碱的菌株,此菌株在48βh内对咖啡碱的降解率为87.7%。经鉴定为聚多曲霉（Aspergillus sydowii）,序列相似性为99.8%。在聚多曲霉茶叶单菌种发酵中,咖啡碱含量急剧下降,在发酵结束时仅占干物质重的(0.414±0.077)%;茶碱质量分数大幅度上升,由最初的不足0.1%上升至(2.129±0.246)%,成为主要的嘌呤碱;可可碱含量无显著变化（P<0.05）;3-甲基黄嘌呤有大幅度增加,增幅达130%。本文首次从发酵茶叶中筛选出可降解咖啡碱的菌株,探究了微生物发酵中咖啡碱的转化产物,为低咖啡碱普洱茶的开发提供了菌种。
Abstract
To identify fungi strain involved in caffeine degradation in Pu-erh tea, caffeine liquid medium was used to screen target strain in Pu-erh tea of different stages during solid-state fermentation. The candidate strains were then analyzed for colony morphology, morphological characteristics and 18βS rDNA. The target strain was inoculated into tea for fermentation. Caffeine, theophylline, theobromine and 3-methylxanthine were determined by HPLC during the fermentation process. Results showed that the most effective strain was isolated from Pu-erh tea during solid-state fermentation, which could degrade 87.70% caffeine within 48 hours and was identified as Aspergillus sydowii with similarity of 99.8%. During the inoculated fermentation with Aspergillus sydowii, caffeine declined sharply and only 0.414%±0.077% (w/w) was reserved. Theophylline increased significantly from less than 0.1% (w/w) to 2.129% ±0.246% (w/w) and became the major purine alkaloid. Theobromine was relatively stable without significant change (P<0.05). The 3-Methylxanthine content had an obvious increase by 130%. The fungi stain was firstly isolated and identified to be involved in caffeine degradation, which confirmed fungus will facilitate the production of decaffeinated Pu-erh tea. In addition, caffeine biotransformation products were developed.
关键词
茶碱 /
聚多曲霉 /
咖啡碱 /
普洱茶 /
渥堆发酵
Key words
Aspergillus sydowii /
caffeine /
Pu-erh tea /
solid-state fermentation /
theophylline
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基金
紫鹃茶树调控花色苷生物合成的MBW转录因子复合体研究（C161104）、中国现代农业产业技术体系（CARS-23）
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