HOBt

HOBt is widely used to suppress racemization in carbodiimide-mediated peptide couplings. Used as an additive, it enhances coupling when HBTU, TBTU, or 1Hbenzotriazole-1-yl-oxy-tris-pyrrolidinophosphonium hexafluorophosphate are used as the coupling reagent. HOBt is supplied as the monohydrate. It does not require any additional drying before use, the water of hydration does not interfere in the coupling reaction. 

Coupling in Solid Phase Peptide Synthesis 

1. Remove the N-protecting group using standard procedures. 

2. Suspend the resin in DCM or DMF (approximately 10 mL/gram resin). 

3. Dissolve 2.5 equivalents (based on resin substitution) of the protected amino acid and 2.5 equivalents of HOBt in a minimum amount of DMF. Add this solution to the resin. 

4. Add 2.5 equivalents of diisopropylcarbodiimide (DIC). Shake the mixture under an inert atmosphere for 3-6 hours. The reaction can be monitored with the Kaiser test. 

5. After the reaction is complete, wash the resin with DMF and DCM.