Separation of porphyrins by capillary electrophoresis in fused-silica and ethylene vinyl acetate copolymer capillaries with visible absorbance detection

Abstract

A mixture of 5 porphyrins were separated by capillary electrophoresis. A UV‐VIS detector was used to detect the separated components. Separation was performed in 2 types of capillaries, i.e. fused‐silica and ethylene vinyl acetate copolymer (EVA) plastic capillaries. The concentration limit of detection (CLOD), mass limit of detection (MLOD), and relative standard deviations (RSD) of migration time and area were investigated. LOD was comparable to that of epi‐fluorescence detection. LOD was lower when separation was performed in EVA capillary compared to fused‐silica capillary. RSD of migration times of the porphyrins when separated in fused‐silica capillary ranges between 0.5 to 1.6% and in EVA capillary ranges between 0.3 to 1.2%. Area RSD in fused‐silica capillary ranges between 7 to 20% and in EVA capillary ranges between 4.7 to 12.3%. Urine spiked with porphyrins was also analysed by CE using fused‐silica and EVA capillaries. Analysis of urine sample spiked with porphyrins showed that stacking effect of porphyrins was observable only in fused‐silica and not in EVA capillary. This led to similar LOD in fused‐silica capillary to those for EVA capillary. © 1997 John Wiley & Sons, Ltd.

Liquid chromatography and mass spectrometry of haem biosynthetic intermediates: a review

“…Porphyrins have been proposed as a good candidate for CE as they contain two to eight carboxylic groups and therefore should be anionic in an alkaline buffer. The majority of methods used MECC in conjunction with fluorescence or visible absorbance detection for the analysis of porphyrin methyl esters (Li et al, 2005) and free acids (Weinberger et al, 1990;Chiang and Li, 1997;Lim, 2002;Melanson and Lucy, 2002). Chiang and Li (1997) demonstrated that fused-silica capillaries suffered from sample adsorption, while ethylene vinyl acetate capillaries reduced sample adsorption and improved resolution; however, they required longer analysis times compared with fused-silica capillaries.…”

“…The majority of methods used MECC in conjunction with fluorescence or visible absorbance detection for the analysis of porphyrin methyl esters (Li et al ., ) and free acids (Weinberger et al ., ; Chiang and Li, ; Lim, ; Melanson and Lucy, ). Chiang and Li () demonstrated that fused‐silica capillaries suffered from sample adsorption, while ethylene vinyl acetate capillaries reduced sample adsorption and improved resolution; however, they required longer analysis times compared with fused‐silica capillaries. The resolution of type I–IV isomers of coproporphyrin was achieved using MECC with bile salt micellar solution and triton micelles (Wu et al ., ).…”

Micellar electrokinetic chromatographic and capillary zone electrophoretic methods for screening urinary biomarkers of human disorders: A critical review of the state‐of‐the‐art

“…Systematic investigations carried out through the evaluation of different surfactants and the addition of organic solvents to the separation buffers, allowed several authors (see [137][138][139][140][141][142][143]) to achieve also the determination of these neutral and lipophilic analytes. As a further contribution to the exploitation of the full potential of MEKC, the reports dealing with the analysis of urinary porphyrins (see [144][145][146][147]) should be mentioned. Given the structural similarity and the highly varying hydrophobicity of these molecules, this technique appeared again the only one ideally suited for their separation providing also significant advantages in terms of speed of analysis over established procedures.…”

“…Luo et al [144] described a MEKC method employing a mixture of 20 mM sodium phosphate and 20 mM sodium borate pH 9.5 containing 50 mM SDS, for the screening of porphobilinogen) in the urine of patients suspected of having acute intermittent porphyria. Chiang and Li [145] separated a mixture of five porphyrins using either fused-silica or ethylene vinyl acetate copolymer (EVA) plastic capillaries, with UV-Vis detection, comparing the LOD obtained with these different columns. So et al [146] were able to separate positional isomers of porphyrins with biological or clinical significance, such as endogenous coproporphyrin isomers I and III, present in normal urine samples, applying an unusual MEKC method.…”

Separation of Chlorins and Carotenoids in Capillary Electrophoresis

“…Some important interactions which cannot take place or are too weak to be measured in aqueous media, such as solvophobic, electrostatic, donor-acceptor, and analyte-additive interactions were discussed in detail by Bowser et al (12). In the literature, there are some reports on the analysis of pigments by CE (14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26), but in most of them, MEKC technique has been applied to water soluble pigment analyses using uncoated fused silica capillary columns (14)(15)(16)(17)(18)(19). Even with mixed surfactant system in microemulsion electrokinetic chromatography, an efficient separation could not be obtained for mesoporphyrines (19).…”