Continuous preparation of two opioid peptides and recycling of organic solvent using liquid/liquid extraction coupled with aluminium oxide column during haemoglobin hydrolysis by immobilized pepsin

Previously, success in producing and extracting continuously LVV-heamorphin-7 and VV-haemorphin-7 in an aqueous/butan-2-ol-octan-1-ol biphasic medium in the course of hydrolysis of bovine haemoglobin by immobilized pepsin was achieved. In this paper, the coupling of the continuous hydrolysis of haemoglobin by pepsin immobilized on a duolite with concomitant extraction of the two haemorphins and haem by a butan-2-ol-octan-1-ol mixture, their adsorption on an aluminium oxide column and the recycling of the solvent mixture in the reactor are described. A steady-state for haemorphin concentrations in the regenerated organic phase at the outlet of the continuous-stirred-tank-reactor (CSTR) was achieved and maintained for more than 10 bed volumes. Finally, the two haemorphins were selectively eluted from the aluminium oxide column tank to a volatile ethanolamine solution. (C) 2008 Elsevier Ltd. All rights reserved.