Methylxanthines, including caffeine, theobromine and theophylline, are natural and synthetic compounds in tea, which could be metabolized by certain kinds of bacteria and fungi. Previous studies confirmed that several microbial isolates from Pu-erh tea could degrade and convert caffeine and theophylline. We speculated that these candidate isolates also could degrade and convert theobromine through N-demethylation and oxidation. In this study, seven tea-derived fungal strains were inoculated into various theobromine agar medias and theobromine liquid mediums to assess their capacity in theobromine utilization. Related metabolites with theobromine degradation were detected by using HPLC in the liquid culture to investigate their potential application in the production of 3-methylxanthine. Based on theobromine utilization capacity, Aspergillus niger PT-1, Aspergillus sydowii PT-2, Aspergillus ustus PT-6 and Aspergillus tamarii PT-7 have demonstrated the potential for theobromine biodegradation. Particularly, A. sydowii PT-2 and A. tamarii PT-7 could degrade theobromine significantly (p < 0.05) in all given liquid mediums. 3,7-Dimethyluric acid, 3-methylxanthine, 7-methylxanthine, 3-methyluric acid, xanthine, and uric acid were detected in A. sydowii PT-2 and A. tamarii PT-7 culture, respectively, which confirmed the existence of N-demethylation and oxidation in theobromine catabolism. 3-Methylxanthine was common and main demethylated metabolite of theobromine in the liquid culture. 3-Methylxanthine in A. sydowii PT-2 culture showed a linear relation with initial theobromine concentrations that 177.12 ± 14.06 mg/L 3-methylxanthine was accumulated in TLM-S with 300 mg/L theobromine. Additionally, pH at 5 and metal ion of Fe2+ promoted 3-methylxanthine production significantly (p < 0.05). This study is the first to confirm that A. sydowii PT-2 and A. tamarii PT-7 degrade theobromine through N-demethylation and oxidation, respectively. A. sydowii PT-2 showed the potential application in 3-methylxanthine production with theobromine as feedstock through the N-demethylation at N-7 position.

中文翻译：

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Aspergillus sydowii PT-2 通过生物降解可可碱生产 3-甲基黄嘌呤。

甲基黄嘌呤，包括咖啡因、可可碱和茶碱，是茶叶中的天然和合成化合物，可被某些细菌和真菌代谢。先前的研究证实，从普洱茶中分离出的几种微生物可以降解和转化咖啡因和茶碱。我们推测这些候选分离株也可以通过 N-去甲基化和氧化降解和转化可可碱。在这项研究中，将七种茶源真菌菌株接种到各种可可碱琼脂培养基和可可碱液体培养基中，以评估它们在可可碱利用方面的能力。通过在液体培养物中使用 HPLC 检测可可碱降解的相关代谢物，以研究它们在 3-甲基黄嘌呤生产中的潜在应用。基于可可碱利用能力，Aspergillus niger PT-1、Aspergillus sydowii PT-2、Aspergillus ustus PT-6 和 Aspergillus tamarii PT-7 已证明可可碱生物降解的潜力。特别是，A. sydowii PT-2 和 A. tamarii PT-7 可以在所有给定的液体培养基中显着降解可可碱 (p < 0.05)。在 A. sydowii PT-2 和 A. tamarii PT-7 培养物中分别检测到 3,7-二甲基尿酸、3-甲基黄嘌呤、7-甲基黄嘌呤、3-甲基尿酸、黄嘌呤和尿酸，证实存在可可碱分解代谢中的 N-去甲基化和氧化。3-甲基黄嘌呤是液体培养物中常见的可可碱去甲基化代谢产物。A. sydowii PT-2 培养物中的 3-甲基黄嘌呤与初始可可碱浓度呈线性关系，为 177.12 ± 14。06 mg/L 3-甲基黄嘌呤在含有 300 mg/L 可可碱的 TLM-​​S 中累积。此外，5 的 pH 值和 Fe2+ 的金属离子显着促进了 3-甲基黄嘌呤的产生（p < 0.05）。本研究首次证实 A. sydowii PT-2 和 A. tamarii PT-7 分别通过 N-去甲基化和氧化降解可可碱。A. sydowii PT-2 在以可可碱为原料通过 N-7 位的 N-去甲基化显示出在 3-甲基黄嘌呤生产中的潜在应用。