Bolten Christoph J, Wittmann Christoph
Biotechnology Department, Biochemistry Institute, Westfalian Wilhelms-University, Munster, Germany.
Biotechnol Lett. 2008 Nov;30(11):1993-2000. doi: 10.1007/s10529-008-9789-z. Epub 2008 Jul 5.
Methanol quenching and fast filtration, the two most common sampling protocols in microbial metabolome analysis, were validated for intracellular amino acid analysis in phylogenetically different yeast strains comprising Saccharomyces cerevisiae, Kluyveromyces marxianus, Pichia pastoris, Schizosaccharomyces pombe and Zygosaccharomyces bailii. With only few exceptions for selected amino acids, all yeasts exhibited negligible metabolite leakage during quenching with 60% cold buffered methanol. Slightly higher leakage was observed with increasing methanol content in the quenching solution. Fast filtration resulted in identical levels for intracellular amino acids in all strains tested. The results clearly demonstrate the validity of both approaches for leakage-free sampling of amino acids in yeast.
甲醇淬灭和快速过滤是微生物代谢组分析中两种最常见的采样方法，我们在包括酿酒酵母、马克斯克鲁维酵母、巴斯德毕赤酵母、裂殖酵母和拜耳接合酵母在内的系统发育不同的酵母菌株中，对其用于细胞内氨基酸分析进行了验证。除了少数选定氨基酸有例外情况外，所有酵母在用60%冷缓冲甲醇淬灭过程中，代谢物泄漏都可忽略不计。随着淬灭溶液中甲醇含量增加，观察到泄漏略有增加。快速过滤使所有测试菌株的细胞内氨基酸水平相同。结果清楚地证明了这两种方法用于酵母中无泄漏氨基酸采样的有效性。
