Further studies on the MEKC and MEEKC of UV-absorbing compounds in human urine

Micellar electrokinetic capillary chromatography (MEKC), a mode of capillar y electrophores is, can separate cations, anions and uncharged molecules si multaneously and with high resolution. So ii is ideal for separating endoge nous and exogenous analytes in complex biofluids, such as urine. Our previo us system used beta -CD-enhanced SDS MEKC to aid resolution and could resol ve a large number of peaks from human urine in about 12 minutes. in this study we have investigated a number of alternative additives to enh ance resolution even further. Various compounds have been claimed, in the l iterature to improve MEKC e.g. mixed micelles, organic solvents, and urea. We investigated a number of such compounds as additives in our standard uri ne system but without much success. However some modified cyclodextrins wer e found to increase both resolution and the migration window. In particular sulphated beta -cyclodextrin gave excellent improvements in both resolutio n and selectivity. The optimised conditions (6.2 mM S-beta -CD in 25 mM sod ium tetraborate, pH 10, 75 mM SDS) resolved 70 components in a normal urine pool in < 15 minutes using < 10 nl of urine. Faster separations but with s ome loss of resolution were obtained by injection from the detector end of the capillary (short-end injection). in addition we have investigated the use of microemulsion electrokinetic ch romatography MEEKC but found that it offered no real advantages over MEKC f or urine analysis. This optimised MEKC separation is simpler, faster and gives higher resoluti on than gradient RP-IC.