Imai K, Matsunaga H, Santa T, Homma H
Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
Biomed Chromatogr. 1995 Jul-Aug;9(4):195-6. doi: 10.1002/bmc.1130090410.
A potential use for phenylisothiocyanate (PITC), the most popular Edman reagent, is presented for analysis of the amino acid sequence and configuration in peptides containing D/L-amino acids. After derivatization with PITC of the N-terminal amino acid (L-Tyr) of an enkephalin analogue, [D-Ala2, D-Leu5]-enkephalin, followed by cleavage/cyclization with trifluoroacetic acid at 50 degrees C for 5 min, the liberated 2-anilino-5-thiazolinone-L-Tyr (ATZ-L-Tyr) was further treated with 20% aqueous trifluoroacetic acid at 50 degrees C for 10 min. The resultant phenylthiohydantoin (PTH)-L-Tyr was then separated on a chiral stationary phase (a penylcarbamoylated cyclodextrin column), retaining its configuration. The residual sequence and configurations of the peptide (D-Ala-Gly-L-Phe-D-Leu) were also determined by separating the corresponding PTH-D- or L-amino acids on chiral columns. This method may be applicable to an automatic Edman sequence analyzer for the configuration determination of peptides containing D/L-amino acids.
介绍了最常用的埃德曼试剂异硫氰酸苯酯（PITC）在分析含有D/L-氨基酸的肽段氨基酸序列和构型方面的潜在用途。用PITC对脑啡肽类似物[D-Ala2，D-Leu5]-脑啡肽的N端氨基酸（L-Tyr）进行衍生化，然后在50℃下用三氟乙酸裂解/环化5分钟，释放出的2-苯胺基-5-噻唑啉酮-L-Tyr（ATZ-L-Tyr）再在50℃下用20%的三氟乙酸水溶液处理10分钟。然后在手性固定相（苯基氨基甲酰化环糊精柱）上分离得到的苯乙内酰硫脲（PTH）-L-Tyr，其构型得以保留。通过在手性柱上分离相应的PTH-D-或L-氨基酸，也确定了肽段（D-Ala-Gly-L-Phe-D-Leu）的剩余序列和构型。该方法可能适用于自动埃德曼序列分析仪，用于确定含有D/L-氨基酸的肽段的构型。
