Developmental changes in the levels and redox potentials of main hemolymph thiols/disulfides in the Jamaican field cricket Gryllus assimilis.

Main thiols and disulfides were determined in the hemolymph of the Jamaican field cricket Gryllus assimilis at various developmental stages. On the basis of these data, redox potentials of the glutathione, cysteine and homocysteine redox systems were calculated. The concentrations of all thiols studied decreased during development (at a stage of 6 molts) with respect to young crickets, and increased again in adult insects. Redox potentials of the glutathione and cysteine systems increased from values of -131.0±5.6 mV and -86.9±17.1 mV, respectively in young crickets to -58.0±3.6 mV and -36.1±4.2 mV, respectively, at the stage of 6 molts and decreased to values of -110.4±24.8 mV and -66.3±12.2 mV, respectively, in adult insects. Redox potentials of the glutathione and cysteine systems in the hemolymph of young and adult insects were similar to those reported for human plasma.

Determination of Total Apigenin in Herbs by Micellar Electrokinetic Chromatography with UV Detection.

Apigenin is a naturally occurring plant flavone that exhibits strong antioxidant, anti-inflammatory, and antitumor properties. A MEKC-UV based method was developed for the determination of total apigenin in selected herbs. Application of pseudostationary phase in the form of SDS micelles resulted in great repeatability of retention times and peak areas. A buffer solution consisting of 30 mmol/L sodium borate (pH 10.2), 10% acetonitrile, and 10 mmol/L sodium dodecyl sulfate was found to be the most suitable BGE for the separation. The method was validated and calibrated for total apigenin in the range of 1.0-100 µmol/L (R (2) = 0.9994). The limits of detection and quantification were 0.48 µmol/L and 0.92 µmol/L, respectively. This precise and robust method was successfully applied to the analysis of plant samples for total apigenin content.

Novel MEKC method for determination of sodium 2-mercaptoethanesulfonate in human plasma with in-capillary derivatization and UV detection.

Sensitive electrophoretic method for determination of total sodium 2-mercaptoethanesulfonate (mesna) in human plasma, based on the stacking with high salt concentration in MEKC and in-capillary derivatization with 2-chloro-1-methyllepidinium tetrafluoroborate followed by UV detection was developed. In the method 0.03molL(-1)pH 7 phosphate buffer with the addition of 0.01molL(-1) SDS, and 10% ACN was used as a BGE. The limit of quantification (LOQ) of the method was 0.5µmolL(-1). Linearity in detector response was observed over the range of 0.5-10µmolL(-1) with the correlation coefficient 0.9971. The intra- and inter-day accuracy (three concentration levels, 5 days, n=3) of the method ranged from 97.2 to 110.0% and from 94.0 to 101.2%, respectively. The novel MEKC method with UV detection proved to be suitable for determination of total mesna in human plasma.

A versatile method for analysis of saliva, plasma and urine for total thiols using HPLC with UV detection.

A simple and rapid HPLC method using 2-chloro-1-methyllepidinium tetrafluoroborate (CMLT) as a derivatization reagent was developed for simultaneous determination of homocysteine (Hcy), glutathione (GSH), γ-glutamylcysteine (γ-GluCys), cysteinylglycine (CysGly), N-acetylcysteine (NACys) and cysteine (Cys) in human saliva, plasma and urine. Separation of the analytes was achieved in just 7min using an HPLC, followed by UV detection at 355nm. Chromatographic separation was accomplished on Aeris PEPTIDE XB-C18 (150mm×4.6mm, 3.6µm) column from Phenomenex with a gradient elution: 0-4.0min, 7-30% B; 4.0-5.5min, 30-7% B; 5.5-7.5min, 7% B; (A: B, v/v); (A) 0.5% CH3COOH and (B) EtOH. Mobile phase was delivered at a flow rate 1.0mLmin(-1). Linearity in detector response for total thiols was observed over the range of 0.1-20µmolL(-1) for Hcy, GSH and γ-GluCys, 0.25-50µmolL(-1) for NACys and CysGly and 5-300 for Cys. The LOQ values for Hcy, GSH, γ-GluCys, NACys, CysGly and Cys were 0.05, 0.05, 0.10, 0.06, 0.12 and 0.08µmolL(-1), respectively. The method was successfully implemented to analysis of the samples donated by 15 apparently healthy volunteers and 10 patients.

Determination of homocysteine thiolactone in urine by field amplified sample injection and sweeping MEKC method with UV detection.

Homocysteine thiolactone (Hcy-thiolactone), an intramolecular thioester, easily acylates free-amino groups in proteins, which impairs or alters the protein's biological function. Here, we describe new capillary electrophoresis assay for the determination of Hcy-thiolactone in human urine based on a field amplified sample injection and sweeping MEKC with UV detection. The two steps procedure relies on sample liquid-liquid extraction followed by CE separation and UV detection at 240nm. The Hcy-thiolactone standard added to the urine before the extraction step shows that the response of the detector is linear within the range studied, from 0.1 to 1µmolL(-1) urine. The intra- and interday precision and recovery were 3.2-14.4% (average 5.1% and 9.3%) and 92.5-112.6% (average 99.8% and 99.1%), respectively. The lower limit of quantification was 0.09nmol Hcy-thiolactone in 1mL of urine. The proposed method was applied for the analysis of 15 urine samples donated by apparently healthy volunteers. The average concentration of the analyte was 0.170±0.029µmolL(-1).