Semi-automatic amino acid sequencing and D/L-configuration determination of peptides with detection of liberated N-terminal phenylthiocarbamoylamino acids

A semi-automatic method for the simultaneous determination of the sequence and the D/L-configuration of amino acids in peptides is reported. The autom ated sequencing program was set up based both on the yield in the cyclizati on/cleavage reaction with 40 mmol dm(-3) boron trifluoride (BF3) etherate a t 48 degrees C for 5 min and on the recovery of anilinothiazolinone- (ATZ-) amino acids extracted with ethyl acetate and 1-chlorobutane. The D/L-confi guration of phenylthiocarbamoylamino acids was determined manually after hy drolysis of ATZ-amino acids by HPLC using a combination of a reversed phase and a chiral stationary phase of beta-cyclodextrin. The hydrolysis conditi ons for ATZ-amino acids were examined with HCl to suppress the amino acid r acemization, and 0.1 mol dm(-3) HCl at room temperature for 5 min was selec ted for hydrolysis. The sequence and D/L-configuration determination of a s ynthetic beta-amyloid 1-40 peptide was carried out and up to 10 residues co uld be successfully analyzed by the semi-automated sequencing procedure wit hout racemization.