Koichi Tanaka Laboratory of Advanced Science and Technology (KTLAST), Shimadzu Corporation, Nishinokyo-Kuwabaracho Nakagyo-ku, Kyoto, 604-8511, Japan.
J Mass Spectrom. 2013 Nov;48(11):1217-23. doi: 10.1002/jms.3283.
We describe here an optimization study of the sample preparation conditions for sensitive detection of peptides by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Among many factors in the conditions, we varied the percent acetonitrile in the peptide solution, the percent acetonitrile in the matrix solution and the α-cyano-4-hydroxycinnamic acid (CHCA) concentration in the matrix solution. CHCA was chosen because it is the most frequently used matrix for analyzing peptides. The well-established dried-droplet method was employed for sample deposition. The examined range of the concentration of CHCA was from 0.01 to 10 mg/ml, and the MeCN content of the solvent for matrix/analyte was 10% to 50%. The indicator for the detection sensitivity was the S/N ratio of the peaks of peptides used. Highly increased sensitivity (100- to 1000-fold) was observed for the optimal CHCA concentration of 0.1 mg/ml in 20% MeCN/0.1% aq. trifluoroacetic acid (TFA), as compared with the conventional concentration (10 mg/ml) in 50% MeCN/0.1% aq. TFA. For example, the limit of detection of human ACTH 18-39 was 10 amol/well for the optimal condition but 10 fmol/well for the conventional condition. The optimal condition (0.1 mg/ml CHCA in 20% MeCN/0.1% aq. TFA) was verified with five model peptides and provided significant improvement in sensitivity (by two to three orders of magnitude) compared with the conventional conditions. Optimizing the CHCA concentration and solvent composition significantly improved the detection sensitivity in the analysis of peptides by MALDI-MS.
我们在这里描述了一种优化方案，用于优化肽的基质辅助激光解吸/电离质谱（MALDI-MS）分析中的样品制备条件。在这些条件中，我们改变了肽溶液中的乙腈百分比、基质溶液中的乙腈百分比和基质溶液中的α-氰基-4-羟基肉桂酸（CHCA）浓度。选择 CHCA 是因为它是最常用于分析肽的基质。采用成熟的干液滴法进行样品沉积。所考察的 CHCA 浓度范围为 0.01 至 10mg/ml，基质/分析物溶剂中的乙腈含量为 10%至 50%。检测灵敏度的指标是所用肽峰的 S/N 比值。与传统浓度（50%乙腈/0.1%三氟乙酸（TFA）中 10mg/ml）相比，在 20%乙腈/0.1%水三氟乙酸（TFA）中，最佳 CHCA 浓度（0.1mg/ml）可使灵敏度提高 100 至 1000 倍。例如，对于最佳条件，人 ACTH 18-39 的检测限为 10amol/孔，但对于传统条件，检测限为 10fmol/孔。最佳条件（0.1mg/ml CHCA 在 20%乙腈/0.1%水 TFA 中）用五种模型肽进行了验证，与传统条件相比，灵敏度提高了两个数量级。优化 CHCA 浓度和溶剂组成可显著提高 MALDI-MS 分析中肽的检测灵敏度。
