Solvent effect on kinetics of appearance of haemorphins in the course of peptic hydrolysis of bovine haemoglobin

Abstract

The effect of the composition of the solvent on the peptic hydrolysis of bovine haemoglobin was studied to improve the preparation of two opioid peptides. Peptic hydrolysis was performed for 24 h at 23 °C in 0.1 M sodium acetate buffer, pH 4.5. A HPLC method was reported for isolating LVV‐haemorphin‐7 and VV‐haemorphin‐7 in one step. The kinetics of appearance of haemorphins was investigated in the presence of 20% (v/v) ethanol, a stabilizing solvent of haemoglobin or urea, a denaturant agent. Ethanol improved the yield of VV‐haemorphin‐7, whereas urea improved the yield of the two haemorphins. Because the amounts of haemorphins observed in urea were greater than in ethanol, the denatured state of haemoglobin is more favourable to obtaining LVV‐haemorphin‐7 and W‐haemorphin‐7. The peptide bonds that give rise to haemorphins would be more accessible to the pepsin.