Peptide behaviour and analysis on a chemically stable C18-bonded vinyl alcohol copolymer column with alkaline and acidic eluents.
C18-bonded vinyl alcohol copolymer (ODP) gel showed no weight loss or decrease in column efficiency for alkyl alcohols after being immersed in aqueous solutions of pH 2 and 10 at 50 degrees C for 48 h, and only a 1% weight loss and a slight decrease in alkyl alcohol retention volumes after similar immersion at pH 13. The chemical stability of the ODP gel was further demonstrated in analyses of acidic, neutral and basic peptides on an ODP column with eluents of pH 3-10, which showed that the peptides differ considerably in the sensitivity of their retention behaviour to eluent pH, even though hydrophobic interaction invariably appeared to be the main retention mechanism. The ODP column was therefore applied to the analysis and alignment of lysilendopeptidase (LEP) peptides derived from reduced and S-carboxymethylated carboxyl proteinase (Rcm-P-CP) of Pseudomonas sp. 101. One acid-soluble and seven alkaline-soluble LEP peaks were found in analyses on the ODP column using acidic and alkaline eluents, respectively. The chymotryptic peptides of Rcm-P-CP were first separated on an ODS column with an acidic eluent, and the eight eluates which contained lysine residue, as determined by amino acid analysis, were then analysed on the ODP column with an alkaline eluent, resulting in a further separation of each into several peaks and thus in the recovery of fractions of pure peptides. The LEP peptide alignment was then determined by overlapping the sequences of the chymotryptic peptides with the C- and N-terminal regions of the LEP peptides.
Uchida
T
T
Gel Separation Development Department, Asahi Chemical Industry Co., Ltd., Kawasaki-shi, Japan.
Ohtani
T
T
Kasai
M
M
Yanagihara
Y
Y
Noguchi
K
K
Izu
H
H
Hara
S
S
D002845:Chromatography / Q000379:methods; D004355:Drug Stability; D005782:Gels; D006863:Hydrogen-Ion Concentration; D010455:Peptides / Q000032:analysis*; D011145:Polyvinyls
D005782:Gels; D010455:Peptides; D011145:Polyvinyls