title: Characterization and quantitative amino acids analysis of analgesic peptides in cinobufacini injection by size exclusion chromatography, matrix-assisted laser desorption/ionization time of flight mass spectrometry and gas chromatography mass spectrometry.
abstract: Cinobufacini injection that comes from the water extract of Bufo bufo gargarizans Cantor skin is widely used for cancer treatment in China. Peptide is one of its major types of constituents, however the biological effects and content of this injection are little reported. In present study, the analgesic effect of peptides was determined and evaluated by in-vivo models. To characterize and quantitatively analyze these peptides, a reliable and efficient method combining size exclusion chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry with amino acid analysis was developed. The peptides presented as a series of analogs with similar molecular weights mostly ranging from 2 to 8 kDa. The amino acid analysis by gas chromatography mass spectrometry (GC-MS) was developed to determine both free and combined amino acids (FAA and CAA) in cinobufacini injection. This method achieved good linearity (R(2) , 0.9909-0.9999) and low limit of detection and quantification. FAA and CAA samples were efficiently analyzed by modified Phenomenex EZ: faast procedure. For the sample analysis, the method showed good repeatability (relative standard deviation, RSD ≤ 10%). For most FAA and CAA the mean recoveries were >80% with RSD <10%. The GC-MS based method is useful for quality assurance of both FAA and CAA in cinobufacini injection.
authors.affiliation: Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China.
authors.forename: Xu
authors.initials: X
authors.lastname: Wu
authors.forename: Nan
authors.initials: N
authors.lastname: Si
authors.forename: Gao
authors.initials: G
authors.lastname: Bo
authors.forename: Hao
authors.initials: H
authors.lastname: Hu
authors.forename: Jian
authors.initials: J
authors.lastname: Yang
authors.forename: Baolin
authors.initials: B
authors.lastname: Bian
authors.forename: Hai Yu
authors.initials: HY
authors.lastname: Zhao
authors.forename: Hongjie
authors.initials: H
authors.lastname: Wang
mesh_terms: D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry; D000664:Amphibian Venoms / Q000737:chemistry* / Q000494:pharmacology; D000700:Analgesics / Q000737:chemistry* / Q000494:pharmacology; D000818:Animals; D001522:Behavior, Animal / Q000187:drug effects; D002850:Chromatography, Gel / Q000379:methods*; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D057230:Limit of Detection; D016014:Linear Models; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D010455:Peptides / Q000737:chemistry* / Q000494:pharmacology; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
chemical_list: D000596:Amino Acids; D000664:Amphibian Venoms; D000700:Analgesics; D010455:Peptides; C545813:huachansu
keywords: GC-MS; amino acid; analgesic effect; cinobufacini injection; peptide