癌症患者血清代谢组学的HPLC和LC-MS研究

Study of serum metabolomics in cancer patients based on HPLC and LC-MS

目的本实验采用HPLC法和LC-MS技术检测了健康人和肿瘤患者人血清样品中的代谢产物,鉴定了与肿瘤生长相关的代谢物,以阐明癌症患者血清代谢组学规律。方法 HPLC条件:色谱柱为Phenomenex Luna(150mm×4.6mm,5μm),流动相为水-75%甲醇(0.1%醋酸铵水溶液);检测波长为215nm,进样量为10μL;质谱条件为电喷雾离子检测器(ESI)负离子模式下检测;毛细管电压为2.74kV;锥孔电压为25.52V;离子源温度为100℃;得到保留时间和峰面积,数据经SIMCA-P+软件进行正交-偏最小二乘投影判别分析(OPLS-DA),表达出与肿瘤相关的差异代谢物;利用LC-MS对相关代谢物进行鉴定。结果检测精密度RSD在0.04%~0.61%之间,健康人血清和肿瘤人血清中代谢规律得到明显区分;5种相关代谢组分得到提取和鉴定,分别为苏氨酸,色氨酸,甘油二酯,磷酰胆碱和硬脂酸。结论肿瘤患者和健康人血清代谢规律有明显差异,鉴定出的化合物与肿瘤生长代谢密切相关,可为肿瘤疾病生物标记物的研究提供相关基础依据。

Objective HPLC and LC-MS have been applied in the study of human serum in both health people and cancer patient. Tumor related metabolites have been identified to explain the regulation in serum metabolomics. Method HPLC conditions： Column： Phenomenex Luna （150mm^4.6mm, 5μm）, mobile phase： 0.1% ammonium acetate solution-75%Methol, gradient elution; detect wavelength： 215nm; injection volumn：10~tL. Mass spectrum condition： ESI detector; capillary： 2.74kV; cone：25.52V; source temperature：100℃. The orthogonal partial least squares-discriminant analysis （OPLS-DA） mode was constructed by the serum metabolic profiles, which indicates the relative metabolites of cancer. Result The RSD of retention time of the component is ranging from 0.04% to 0.61%. Five relative metabolites are extracted and identified： threonine, tryptophan, diacylglycerol, phosphorylcholine and stearic acid. Conclusion Differences are shown between health group and cancer group. The five extracted compounds are related to cancer which provide foundation of the study ofbiomarkers in cancer.