On-line concentration of peptides and proteins with the hyphenation of polymer monolithic immobilized metal affinity chromatography and capillary electrophoresis

An iminodiacetic acid (IDA)-type adsorbent is prepared at the one end of a capillary by covalently bonding IDA to the monolithic rods of macroporous poly(glycidyl methacrylate-co-ethylene dimethacrylate). Cu(II) is later introduced to the support via the interaction with IDA. By this means, polymer monolithic immobilized metal affinity chromatography (IMAC) materials are prepared. With such a column, IMAC for on-line concentration and capillary electrophoresis (CE) for the subsequent analysis are hyphenated for the analysis of peptides and proteins. The reproducibility of such a column has been proved good with relative standard deviations (RSDs) of dead time of less than 5% for injection-to-injection and 12% for column-to-column (n = 3). Through application on the analysis of standard peptides and real protein samples, such a technique has shown promising in proteome study.

Separation of compounds interacting with liposome membrane in combined prescription of traditional Chinese medicines with immobilized liposome chromatography

Immobilized liposome chromatography (ILC), the stationary phase of which has been regarded as a mimic biomembranes system was used to separate and analyze compounds interacting with liposome membrane in Danggui Buxue decoction, a combined prescription of traditional Chinese medicines (CPTCMs), and its compositions Radix Astragli and Radix Angelica Sinensis. More than 10 main peaks in the extract of Danggui Buxue decoction were resolved on the ILC column, suggesting that more than 10 components in the prescription have significant retention on ILC column. Ligustilide, astragaloside, TV and formononetin, three main bioactive ingredients in Danggui Buxue decoction, were found to have relatively significant, while ferulic acid, another bioactive ingredient in the prescription, relatively weak retention on ILC column. Effects of the eluent pH and amount of immobilized phosphatidylcholine (PC) on separation of interactional compounds in the extract of Danggui Buxue decoction were also investigated. It was found that these two factors strongly affected the retention of some interactional compounds. In addition, the fractions partitioned with different solvents from water extract of this combined prescription were evaluated with this ILC column system. (c) 2005 Elsevier B.V. All rights reserved.