Table 1.

Racemization of Cys during synthesis of the model peptide, H–Gly–Cys–Phe–NH₂, as a function of the coupling conditions and the Cys protecting group

Coupling conditionsa				Racemization (%)b
Reagent (4 eq)	Additive (4 eq)	Base (8 eq)	Solvent	Cys(Trt)	Cys(Acm)
HCTU	6-Cl-HOBt	DIEA	DMF	8.0	2.2
HCTU	6-Cl-HOBt	PS	DMF	2.0	0.6
HCTU	6-Cl-HOBt	DBU	DMF	2.1	0.7
HCTU	6-Cl-HOBt	TMP	DMF	1.3	0.3
HCTU	6-Cl-HOBt	DIEA	DCM/DMF (v/v, 1/1)	3.3	1.0
HCTU	6-Cl-HOBt	PS	DCM/DMF (v/v, 1/1)	1.2	0.3
HCTU	6-Cl-HOBt	DBU	DCM/DMF (v/v, 1/1)	1.5	0.4
HCTU	6-Cl-HOBt	TMP	DCM/DMF (v/v, 1/1)	0.6	0.1
DIC	HOBt	—	DMF	0.1	< 0.1
EDC · HCl	HOBt	—	DMF	0.9	0.2

^aThe HCTU-mediated reactions were performed using a 1 min preactivation procedure of coupling with Fmoc amino acid/HCTU/6-Cl-HOBt (4/4/4 equiv) in the presence of the bases (8 equiv) listed in Table 1. The carbodiimide-mediated reactions were performed by a 5 min preactivation procedure of coupling with Fmoc amino acid/DIC or EDC · HCl/HOBt (4/4/4 equiv). The coupling time was 30 min and concentrations of the active species were 0.20 M.

^bDefined as (H–Gly–d-Cys–Phe–NH₂)/(H–Gly–l-Cys–Phe–NH₂) × 100.