New Application of pH-Mediated Acid Stacking Technique for Amphoteric Compounds in Capillary Electrophoresis: Example Assay of Blood Glutathiones

Abstract

Both reduced and oxidized forms of glutathiones were firstly stacked and detected using pH-mediated acid stacking method, in which glutathiones were stacked as cations and separated as anions. Factors, such as injection time, sweeping time, buffer pH, concentration of sodium chloride in sample matrix, that influenced stacking and separation were systematically studied and optimized. Under the optimum condition, the enhancement factors of ~20 times for both reduced and oxidized forms of glutathiones could be easily obtained within 20 min with satisfied sensitivities (limit of detections were 0.12 and 0.06 μmol/L for reduced and oxidized glutathione, respectively, at signal-to-noise ratio, S/N = 3), linearity range (0.3-300.0 and 0.6-300.0 μmol/L for reduced and oxidized glutathione, respectively), recoveries (>98%) and reproducibilities (relative standard deviation <5.1% for peak height). The proposed method provides an alternation way for assaying of glutathiones, as well as amphoteric compounds, in blood sample.

Determination of glutathione in blood via capillary electrophoresis with pH‐mediated stacking

“…Micellar electrokinetic chromatography approaches were the first to be developed, based on the use of alkaline (pH 7-9) background electrolytes (BGEs) containing sodium dodecyl sulfate (SDS) [32,33]. The second group includes capillary zone electrophoresis (CZE) approaches based on the separation of analytes in positive polarity mode using alkaline BGEs (pH ∼7.5-8.5) [34][35][36][37][38][39]. In this case, GSH and GSSG have a negative charge and migrate against the electroosmotic flow (EOF); however, a high level of EOF enables their rapid determination (5-10 min).…”

“…The authors in Ref. [36] used acid stacking to achieve an LOQ (GSSG) of ∼3 µM (in terms of blood), but this required an increase in separation time from ∼11 to 20 min. In addition, as noted in Ref.…”

The Influence of Crenotheraphy with SHS Water on the Concentration of Glutathione in Blood of People with Rheumatoid Arthritis

“…However, HPLC methods are disadvantaged by the long time required to complete each assay, materials consumed and high costs of instrumentation. During the last decade increased use of capillary electrophoresis (CE) occurred while a lot of attempts have been focused on glutathione separations [56][57][58][59]. Main advantages of CE are low consumptions of buffers and solvents; requirement of a small sample volume and fast separation.…”

Sensitivity enhancement in capillary electrophoresis and their applications for analyses of pharmaceutical and related biochemical substances

“…Under the optimum condition, the signal enhancement of 20 times for both reduced and oxidized forms of glutathiones was observed. LODs were 0.12 and 0.06 μmol/L for reduced and oxidized glutathione, respectively [105]. Additionally, acid‐mediated stacking has been employed for the analysis of d , l ‐kynurenine, in human serum and urine samples by cationic‐β‐CD and its synergistic dual chiral selector system with α‐CD in 50 mM borax borate buffer (pH 9.0) [106].…”

“…Sample stacking techniques have been performed on many pharmaceutical substances. Table 1 [15][16][17][21][22][23]66,98,[100][101][102][103][104][105] describes selected applications of sample stacking techniques in CE for pharmaceutical and related biochemical substances.…”