title: Novel chiral derivatization reagents possessing a pyridylthiourea structure for enantiospecific determination of amines and carboxylic acids in high-throughput liquid chromatography and electrospray-ionization mass spectrometry for chiral metabolomics identification.
abstract: This paper reports the synthesis and the application of novel derivatization reagents possessing a pyridylthiourea structure for the enantiospecific determination of chiral amines and carboxylic acids in high-throughput LC-ESI-MS/MS. The novel reagents, i.e., (R)-N-(3-pyridylthiocarbamoyl)pyrrolidine-2-carboxylic acid (PyT-C) and (S)-3-amino-1-(3-pyridylthiocarbamoyl)pyrrolidine (PyT-N), were evaluated as chiral derivatization reagents for the enantiomeric determination of chiral amines and carboxylic acids, respectively, in terms of separation efficiency by reversed-phase chromatography and detection sensitivity by ESI-MS/MS. The chiral amines and carboxylic acids were easily labeled with PyT-C and PyT-N, respectively, at 60°C in 60min in the presence of 2,2'-dipyridyl disulfide (DPDS) and triphenylphosphine (TPP) as the activation reagents. The resulting diastereomers were completely separated by reversed-phase chromatography using a small particle (1.7μm) ODS column (Rs=3.54-6.00 for carboxylic acids and Rs=3.07-4.75 for amines). A highly sensitive detection at the sub-fmol level was also obtained from the SRM chromatograms at a single monitoring ion, m/z 137.0 (0.72-1.46fmol for carboxylic acids and 0.55-1.89fmol for amines). The proposed procedure using PyT-C and PyT-N was applied to the determination of chiral amines and carboxylic acids spiked into human saliva, as a model study of chiral metabonomics identification. dl-Amino acid methyl esters and N-acetyl dl-amino acids, which are the representatives as the chiral amines and carboxylic acids, in the saliva were clearly identified by the present method. Because the same product ion at m/z 137.0 was obtained from collision-induced dissociation (CID) of protonated molecular ions of all the derivatives, the proposed procedure using both reagents (i.e., PyT-C and PyT-N) seems to be useful for chiral metabolomics identification having selected functional groups (i.e., amines and carboxylic acids).
authors.affiliation: Laboratory of Analytical and Bio-Analytical Chemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan.
authors.forename: Ryuji
authors.initials: R
authors.lastname: Nagao
authors.forename: Haruhito
authors.initials: H
authors.lastname: Tsutsui
authors.forename: Toshiki
authors.initials: T
authors.lastname: Mochizuki
authors.forename: Takahiro
authors.initials: T
authors.lastname: Takayama
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Kuwabara
authors.forename: Jun Zhe
authors.initials: JZ
authors.lastname: Min
authors.forename: Koichi
authors.initials: K
authors.lastname: Inoue
authors.forename: Kenichiro
authors.initials: K
authors.lastname: Todoroki
authors.forename: Toshimasa
authors.initials: T
authors.lastname: Toyo'oka
mesh_terms: D015082:2,2'-Dipyridyl / Q000031:analogs & derivatives* / Q000737:chemistry; D000588:Amines / Q000032:analysis / Q000737:chemistry*; D002264:Carboxylic Acids / Q000032:analysis / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000379:methods*; D004220:Disulfides / Q000737:chemistry*; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D008297:Male; D055432:Metabolomics / Q000379:methods; D012463:Saliva / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013237:Stereoisomerism
chemical_list: D000588:Amines; D002264:Carboxylic Acids; D004220:Disulfides; C001105:2,2'-dipyridyl disulfide; D015082:2,2'-Dipyridyl