Factors influencing the reaction yields and formation of the side products during coupling of N-carbobenzoxyamino acids with dialkyl or diphenyl aminoalkylphosphonates, by means of mixed anhydride procedure, were evaluated. The recommended procedure, arising from these studies, involves the use of chloroform/ethyl chloroformate/triethylamine system and delicate warming of the reaction mixture during the coupling step.
Factors influencing the reaction yields and formation of the side products during mixed
The factors which affect yield and racemization in custom peptide synthesis by the mixed carbonic anhydride method have been investigated. Good results in test cases were obtained when the tertiary amine necessary for formation of the mixed anhydride contained at least one N-methyl group, and it was not used in excess, indicating the importance of steric factors. Reduction of "basicity" of the amine was found to be favorable; complete racemization was found with an excess of trimethylamine as the tertiary base, and none with an excess of N-methyl-morpholine in the test Z-Gly-Phe-Gly-OEt synthesis in ethyl acetate or tetrahydrofuran.
The more sensitive Bz-Leu-Gly-OEt synthesis gave no racemate with 1 equiv of N-methylmorpholine but some racemate with 2 equiv. The effects on racemization of change of solvent, temperature, time allowed for mixed anhydride formation, and several anions are reported. Evidence for racemization via oxazolones is given. Best yields were obtained with isobutyl chloroformate as the reagent. The principles disclosed here should be useful in improving other acylations.