KINETICS OF APPEARANCE OF 4 HEMORPHINS FROM BOVINE HEMOGLOBIN PEPTIC HYDROLYSATES BY HPLC COUPLED WITH PHOTODIODE-ARRAY DETECTION

The kinetics of appearance of hemorphins during peptic hydrolysis of b ovine hemoglobin was investigated by reverse-phase high-performance li quid chromatography (RP-HPLC) coupled with a photodiode array detector . The degree of hydrolysis (DH) of hemoglobin by pepsin was determined and different defined DH of hydrolysates were obtained. The analysis of these hydrolysates by HPLC coupled with a photodiode array detector allowed us to identify and quantify the hemorphins in every hydrolysa te and to determine the quantitative evolution of hemorphins as a func tion of DH. It indicated that hemoglobin was a direct precursor of LVV -hemorphin-5 and LVV-hemorphin-7. These peptides were demonstrated to be secondary substrates for pepsin to generate VV-hemorphin-5 and VV-h emorphin-7. Moreover, LVV-hemorphin-7 was more stable towards pepsin t han LVV-hemorphin-5. The affinity of pepsin towards some peptidic bond s was also demonstrated.