PURPOSE: To establish the method of determining C-terminal amino acid which can deal with increasingly slighter quantities of protein, etc., by treating denatured protein or denatured peptide to convert a terminal carboxyl group to an acetyl group, hydrolyzing the resultant product, converting the liberated amino ketone body to a deriv. and separating and identifying the same.

CONSTITUTION: The protein, etc., are subjected to the denaturation treatment by performic acid oxidation, etc., to form the denatured protein, etc., in which the three-dimensional structure is broken down and the C-terminal is exposed. After the protein, etc., are dried, the protein, etc., are treated with acetic anhydride in the presence of an org. ternary base in an anhydrous aprotic solvent to convert the carboxyl group at the terminal to the acetyl group. The amino ketone body corresponding to the terminal amino acid part having the C- terminal is liberated as the mixture with the other amino acid if the product is hydrolyzed by using hydrochloric acid, etc. The desired amino ketone body is separated and identified by a hydrazone deriv., etc.