Determination of B2 and B6 vitamers in serum by capillary electrophoresis-molecular fluorescence-charge coupled detector

Abstract

A method for the separation by capillary zone electrophoresis and determination of vitamers of two important water-soluble vitamins, i.e., B2 and B6, is proposed here. The working conditions for optimal separation were obtained by a multivariate methodology in order to succeed in the best resolution in the shortest analysis time. The optimization of the buffer composition together with other variables, such as analysis and injection voltage, temperature of the capillary cassette, and injection time, resulted in a solution of 30 mM KH2PO4 adjusted to pH 8.5 with formic acid. Concerning the detection step, the target analytes were quantified by molecular fluorescence, for which two different detectors, a photomultiplier tube (PMT) and a charge-coupled detector (CCD), were compared in terms of resolution, sensitivity, and precision. A Xe-Hg lamp was used as an irradiation source in the two cases. The best option was the CCD, which provides three-dimensional electropherograms and enables to solve the overlapped peaks. Besides, the sensitivity of the CCD was similar to that of the PMT, due to the treatment data, obtaining limits of detection and quantification from 1.16 to 27.1 ng/g and from 3.83 to 89.4 ng/g, respectively. The method was applied to the serum samples for which a prior liquid-liquid extraction using ethanol in an acid medium was mandatory for eliminating the interferences and concentrating the analytes by a factor of 5. The rapidity of the analysis (13 min for the electrophoretic separation) and the excellent precision (repeatability and within-laboratory reproducibility between 2.86 and 4.11% and 7.03 and 8.45%, respectively, both expressed as relative standard deviation) demonstrated the capability of the proposed method for a clinical routine analysis.

Supercritical fluid extraction as an on‐line clean‐up technique for determination of riboflavin vitamins in food samples by capillary electrophoresis with fluorimetric detection

“…Three different aqueous buffers were tested: phosphate, carbonate and borate buffers. In agreement with previous works [31,32], the phosphate buffer solution (PBS) exhibit a higher EOF and the most favourable S/ Ns for all RF vitamins, in comparison to the use of both borate and carbonate buffer solutions. The ionic strength of the PBS was tested, at a constant pH (8.5), in the 10 and 50 mM concentration range.…”

“…There is a growing interest in knowing not only total RF content, but also flavin composition of food samples. The common separation techniques used for the determination of RF and its metabolites in different foods are LC [26,27], TLC [28,29] and CE [30,31]. Pérez-Ruiz et al [32] have reported the determination of RF, FMN and FAD in biological tissues by CZE with LIF detection.…”

All‐Metal Clusters that Mimic the Chemistry of Halogens

“…These substances may participate in enzymatic processes as co‐enzymes or their precursors, act as antioxidants, and affect genetic regulation processes. Therefore, absence of vitamins will cause serious physiological problems 1, 2. Among all kinds of vitamins, the B group of vitamins is the most important constituent, endowed with a great variety of chemical structures.…”

Determination of residual cell culture media components by MEKC

“…While some CE methods have been developed for the determination of water‐soluble vitamins as main components in food or nutritional supplements, very little has been focused on the trace level analysis. Priego‐Capota and Castro used CZE with a molecular fluorescence charge coupled detector to measure vitamin B 2 (riboflavin) and B 6 (pyridoxine) 19. Although the detection limit was obtained at part per billion levels, extraction was required for eliminating interferences and off‐line sample concentration was also needed when the method was applied to serum samples.…”