Optimization of the quenching method for metabolomics analysis of Lactobacillus bulgaricus.
This study proposed a quenching protocol for metabolite analysis of Lactobacillus delbrueckii subsp. bulgaricus. Microbial cells were quenched with 60% methanol/water, 80% methanol/glycerol, or 80% methanol/water. The effect of the quenching process was assessed by the optical density (OD)-based method, flow cytometry, and gas chromatography-mass spectrometry (GC-MS). The principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were employed for metabolite identification. The results indicated that quenching with 80% methanol/water solution led to less damage to the L. bulgaricus cells, characterized by the lower relative fraction of prodium iodide (PI)-labeled cells and the higher OD recovery ratio. Through GC-MS analysis, higher levels of intracellular metabolites (including focal glutamic acid, aspartic acid, alanine, and AMP) and a lower leakage rate were detected in the sample quenched with 80% methanol/water compared with the others. In conclusion, we suggested a higher concentration of cold methanol quenching for L. bulgaricus metabolomics due to its decreasing metabolite leakage.
MOE Key Laboratory of Dairy Science, College of Food Science, Northeast Agricultural University, Harbin 150030, China; School of Public Health, Jilin Medical College, Jilin 132013, China.
Ming-ming
MM
Chen
Ai-li
AL
Li
Mao-cheng
MC
Sun
Zhen
Z
Feng
Xiang-chen
XC
Meng
Ying
Y
Wang
D005285:Fermentation; D005434:Flow Cytometry; D005516:Food Microbiology; D008401:Gas Chromatography-Mass Spectrometry; D048668:Lactobacillus delbrueckii / Q000378:metabolism*; D016018:Least-Squares Analysis; D055442:Metabolome; D055432:Metabolomics / Q000379:methods* / Q000706:statistics & numerical data; D000432:Methanol; D025341:Principal Component Analysis
D000432:Methanol