同位素稀释质谱法测定多肽含量

QUANTIFICATION OF PEPTIDE BY ISOTOPE DILUTION MASS SPECTROMETRY

利用Leu-Phe作为模型肽,建立了水解后同位素稀释质谱测定多肽含量的方法。在实验中优化了酸水解的时间,采用高效液相色谱法和质谱法证明模型肽已完全水解。水解后的氨基酸经过HPLC分离,质谱检测采用选择离子监测模式,分别检测苯丙氨酸(m/z=166)和标记苯丙氨酸(m/z=174)的离子。根据苯丙氨酸的含量计算模型肽的含量,并评定了测定结果的不确定度。

An approach for quantification of peptide by hydrolysis followed HPLC isotope dilution method with Leu - Phe as an example was presented . Hydrolysis time was optimized, and HPLC analysis as well as mass spectrometry was used to confirm the 100% successful hydrolysis. Amino acids were separated by liquid chromatography （LC） and then detected by mass spectrometry with selected ion monitor mode of phenylalanine （ m/z = 166 ） and Ds - labelled analogue （ m/z = 174 ）. The original concentration of the peptide was calculated from the phenylalanine concentration. The uncertainty of the result was also evaluated.