Large scale production of d-allose from d-psicose using continuous bioreactor and separation system

l-Rhamnose isomerase (l-RhI) from Pseudomonas stutzeri LL172 can convert d-psicose to d-allose. Partially purified recombinant l-RhI from Escherichia coli was immobilized on BCW-2510 Chitopearl beads and utilized to produce d-allose. Total 20,000 units of immobilized enzyme converted d-psicose to d-allose without remarkable decrease in the enzyme activity over 17 days. When 50% d-psicose (w/w) was applied to a column with a flow rate of 0.8 ml/min at 42 °C, approximately 30% d-psicose was isomerized to d-allose for 17 days. However, by reducing the flow rate to 0.4 ml/min after 17 days, d-allose was transformed at the same rate for 13 days. The total of 27 l reaction mixture was separated by Simulated-Moving-Bed Chromatograph system. Approximately 2.2 l/d of 50% (w/w) reaction mixture was separated continuously. After separation, d-allose and d-psicose fractions were 3 l of approximately 10% (w/w) with 95% purity and 10 l of approximately 8% (w/w) with 95% purity per day, respectively. The separated d-allose solution was concentrated up to about 50% and crystallized gradually by being kept at room temperature. Crystals of d-allose were separated from the syrup by filtration and 1.65 kg crystals of 100% purity were obtained. The d-allose crystal yield from the d-psicose substrate was approximately 10%.