Metabolic Response of <i>Escherichia coli</i> upon Treatment with Hypochlorite at Sub-Lethal Concentrations

Hypochlorite is a reactive oxygen species that is worldwide as an antibacterial disinfectant. Hypochlorite exposure is known to cause oxidative damage to DNA and proteins. As a response to these effects, the metabolite profiles of organisms treated with sub-lethal doses of hypochlorite are assumed to be severely modified; however, the nature of these changes is hardly understood. Therefore, using nuclear magnetic resonance spectroscopy and gas chromatography-coupled mass spectrometry, we analyzed the time-dependent impact of hypochlorite exposure with a sub-lethal concentration (50 µM) on the metabolite profile of the Escherichia coli strain MG1655. Principle component analysis clearly distinguished between the metabolite profiles of bacteria treated for 0, 5,10, 20, 40, or 60 min. Major changes in the relative amounts of fatty acids, acetic acid, and formic acid occurred within the first 5 min. Comparative gas chromatography-coupled mass spectrometry analyses revealed that the amounts of free methionine and alanine were significantly decreased in the treated cells, demonstrating their susceptibility to hypochlorite exposure. The concentrations of succinate, urea, orotic acid, 2-aminobutyric acid, and 2-hydroxybutyric acid were also severely affected, indicating general changes in the metabolic network by hypochlorite. However, most metabolite levels relaxed to the reference values of untreated cells after 40–60 min, reflecting the capability of E. coli to rapidly adapt to environmental stress factors such as the presence of sub-lethal oxidant levels.次氯酸盐（Hypochlorite）是一类在全球范围内用作抗菌消毒剂的活性氧物种（reactive oxygen species）。已知次氯酸盐暴露会引发DNA与蛋白质的氧化损伤。针对此类损伤，生物体经亚致死剂量次氯酸盐处理后的代谢物谱被认为会发生显著改变，但此类变化的具体机制尚不明晰。因此，本研究采用核磁共振波谱法（nuclear magnetic resonance spectroscopy）与气相色谱-质谱联用法（gas chromatography-coupled mass spectrometry），分析了亚致死浓度（50 µM）次氯酸盐暴露对大肠杆菌MG1655菌株（Escherichia coli strain MG1655）代谢物谱的时间依赖性影响。主成分分析（Principle component analysis）可清晰区分经0、5、10、20、40或60分钟处理的细菌的代谢物谱。脂肪酸、乙酸与甲酸的相对含量在最初5分钟内即发生显著变化。对比气相色谱-质谱联用法分析结果显示，处理组细胞内游离甲硫氨酸与丙氨酸的含量显著降低，表明这两种氨基酸对次氯酸盐暴露具有易感性。琥珀酸盐、尿素、乳清酸、2-氨基丁酸及2-羟基丁酸的浓度也受到显著影响，提示次氯酸盐可引发代谢网络的全局性改变。然而，在40至60分钟后，大多数代谢物水平恢复至未处理细胞的参考值，这反映出大肠杆菌能够快速适应环境胁迫因子（如亚致死浓度的氧化剂暴露）。