The racemization of anN-methylamino-acid residue during peptide-bond formation and mixed-anhydride activation has been investigated using Ala-MeLeu-Gly and Ala-MeLeu as model peptides. The results were compared with those for Ala-Leu-Gly, Ala-Leu, and Ala-Pro. The extents of racemization were determined by analysis of the diastereomeric products of the reactions after deprotection, using an amino-acid analyzer. Extensive racemization was detected after the hydrolysis of the mixed anhydrides of Bz-MeLeu, Z-Ala-MeLeu, and Z-Ala-Leu, but not of Boc-Ala-Pro and Z-MeIle. Significant racemization (2.8–39%) was observed when Z-Ala-MeLeu was coupled with Gly-OBzl by various methods in the presence of salts such as triethylamine hydrochloride orp-toluenesulfonate. Only coupling through theN-hydroxysuccinimide (HONSu) ester gave stereochemically pure product. In the absence of salt, less racemization was observed, but only couplings usingN-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline andN,N′-dicyclohexylcarbodiimide–HONSu gave essentially pure products. Polar solvents promoted racemization, but excess base did not. Chemical evidence that the racemization intermediate is an oxazolium-5-oxide has been obtained by trapping the intermediate as an addition product (a pyrrole) in 85% yield.The yields obtained by various coupling methods have been determined for several model peptides. Couplings at the carboxyl group of anN-methylamino acid gave high yields only in the absence of salt, except for coupling by the HONSu ester method. Couplings to anN-methylamino group gave high yields, except for coupling by thep-nitrophenyl ester method. Couplings to Ala-MeLeu-OBu′ gave higher yields than couplings to Ala-MeLeu-OBzl, presumably due to piperazine-dione formation by the latter.