Operational Comparisons of Size Exclusion Chromatography in On-line Recovery of Large Molecules

초록

Cyclodextrin homologues are oligosaccharides, synthesized from starch through cyclization by cyclodextrin glycosyltransferase (CGTase, EC 2.4.1.19). There are many kinds of CD derivatives, but -, - and -CD, composed of 6, 7 and 8 glucopyranose units, respectively, are the major products. Recently the usages of CDs expanded not only as an additive for medicinal products, foodstuffs and cosmetics, but also as a separating media in separation of enantiomers and other processes. The number of applications of CD derivatives are also increasing. CDs production in aqueous solution is affected by many factors, such as the product inhibition on CGTase or product degradation by CGTase and so on. Thus, modified processes to overcome these disadvantages are needed, and ATPS may become a solution. Cyclodextrin homologues (CDs), produced by cyclodextrin glycosyltrans- ferase (CGTase), were simultaneously partitioned in aqueous two-phase system (ATPS). Partition coefficients of CDs were measured in PEG/salt and PEG/dextran systems. Phosphate, citrate, sulfate were tested as salts. ATPS of PEG/salt and PEG/dextran had the partition coefficients of the CDs, larger than unity. However, PEG/dextran system was observed better than PEG/salt as CGTase activity decreased sharply with salt concentration. Enzymatic reaction occurred mainly in PEG-rich bottom phase because of the low partition coefficient of CGTase. The resulting CDs transferred to the PEG-rich top phase, obeying the diffusional partition. In the ATPS of 7% PEG (M.W. 20,000) and 9% dextran (M.W. 40,000), 7mg/ml of CDs were obtained in top phase at 4.5 hours. The interconversion and disproportiona-tion among the CDs and linear oligosaccharides were also studied.