Fukuyama Yuko, Nakajima Chihiro, Furuichi Keiko, Taniguchi Kenichi, Kawabata Shin-Ichirou, Izumi Shunsuke, Tanaka Koichi
Koichi Tanaka Laboratory of Advanced Science and Technology, Shimadzu Corporation , 1, Nishinokyo-Kuwabaracho, Nakagyo-ku, Kyoto 604-8511, Japan.
Anal Chem. 2013 Oct 15;85(20):9444-8. doi: 10.1021/ac4018378. Epub 2013 Sep 24.
Hydrophobic peptides are difficult to detect in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), because of the hydrophilic properties of conventional matrices and the low affinity for hydrophobic peptides. Recently, we reported on alkylated dihydroxybenzoic acid (ADHB) as a matrix additive for hydrophobic peptides; however, the peptides were detected in the rim of the matrix-analyte dried spot. Here, we report on a novel matrix, alkylated trihydroxyacetophenone (ATHAP), which is a 2,4,6-trihydroxyacetophenone derivative incorporating a hydrophobic alkyl chain on the acetyl group and thus is expected to have an affinity for hydrophobic peptides. ATHAP increased the sensitivity of hydrophobic peptides 10-fold compared with α-cyano-4-hydroxycinnamic acid (CHCA), in which the detection of hydrophilic peptides was suppressed. The peptides were detected throughout the entire matrix-analyte dried spot using ATHAP, overcoming the difficulty of finding a "sweet spot" when using ADHB. In addition, ATHAP functioned alone as a matrix, unlike ADHB as an additive. In phosphorylase b digests analysis, hydrophobic peptides, which were not detected with CHCA for 1 pmol, were detected with this matrix, confirming that ATHAP led to increased sequence coverage and may extend the range of target analytes in MALDI-MS.
由于传统基质的亲水性以及对疏水肽的低亲和力，疏水肽在基质辅助激光解吸/电离质谱（MALDI-MS）中难以检测。最近，我们报道了烷基化二羟基苯甲酸（ADHB）作为疏水肽的基质添加剂；然而，肽是在基质-分析物干斑的边缘被检测到的。在此，我们报道了一种新型基质，烷基化三羟基苯乙酮（ATHAP），它是一种2,4,6-三羟基苯乙酮衍生物，在乙酰基上引入了疏水烷基链，因此有望对疏水肽具有亲和力。与α-氰基-4-羟基肉桂酸（CHCA）相比，ATHAP将疏水肽的灵敏度提高了10倍，而CHCA对亲水肽的检测有抑制作用。使用ATHAP时，肽在整个基质-分析物干斑中都能被检测到，克服了使用ADHB时寻找“最佳点”的困难。此外，与作为添加剂的ADHB不同，ATHAP单独作为基质发挥作用。在磷酸化酶b消化物分析中，对于1皮摩尔的样品，用CHCA无法检测到的疏水肽，用这种基质能够检测到，这证实了ATHAP导致序列覆盖率增加，并可能扩大MALDI-MS中目标分析物的范围。
