Simultaneous determination of total plasma glutathione, homocysteine, cysteinylglycine, and methionine by high‐performance liquid chromatography with electrochemical detection

Houzé, Pascal; Gamra, Stéphanie; Madelaine, Isabelle; Bousquet, Bernard; Gourmel, Bernard

doi:10.1002/jcla.1018

Cited by 68 publications

(48 citation statements)

References 26 publications

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“…After centrifugation at 14 000 g for 5 min, the supernatant was diluted 1:2 with the mobile phase and 50 ml were injected into the HPLC system (HP 1100, Hewlett-Packard, Waldbronn, Germany). Mobile phase consisted of 50 mM NAH 2 PO 4 , 1.0 mM ion-pairing reagent 1-octanesulfonic acid, and 5% acetonitrile (v/v), adjusted to pH 2.7 with 85% phosphoric acid (Houze et al 2001).…”

Section: Determination Of the Folate Content By A Microbiological Tesmentioning

confidence: 99%

Placental markers of folate-related metabolism in preeclampsia

Mislanova¹,

Martsenyuk²,

Huppertz³

et al. 2011

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The etiology and degree of clinical symptoms of preeclampsia depend on genotypic and phenotypic maternal and trophoblast factors, and elevated levels of plasma homocysteine (Hcy) are one of the pathogenetic factors of preeclampsia. To assess the impact of the folate-related metabolism, we characterized the indices of this metabolism in 40 samples from uncomplicated term placentas and 28 samples from preeclamptic pregnancies by quantifying the total content of folate, methionine (Met), Hcy and related cysteine, and glutathione (GSH) in compliance with the 677 C/T genotype of methylene tetrahydrofolate reductase (MTHFR). The prevalence of MTHFR genotypes was not significantly different between the two groups. The polymorphism of MTHFR was not unambiguously connected with the content of total placental Met, Hcy and related cysteine, and GSH either in uncomplicated or in complicated pregnancies. By contrast, the combination of the heterozygous MTHFR genotype with folate deficiency in the samples from preeclamptic pregnancies was characterized by a statistically significant decrease in the Met content, a trend toward increased Hcy levels and a tight association between metabolically directly and indirectly related compounds, e.g. positive relation between Hcy versus cysteine and folate versus GSH and negative relation between folate versus Hcy and both Hcy and cysteine versus GSH. We demonstrated the expression of cystathionine-b-synthase (CBS) in human placenta at term by RT-PCR and western blot analysis, for the first time, and confirmed its catalytic activity and the accumulation of cysteine and CBS in placental explants cultivated in the presence of elevated Hcy concentrations. We suggest that disturbance in placental folate-related metabolism may be one of the pathogenetic factors in preeclampsia.

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Section: Determination Of the Folate Content By A Microbiological Tesmentioning

confidence: 99%

Placental markers of folate-related metabolism in preeclampsia

Mislanova¹,

Martsenyuk²,

Huppertz³

et al. 2011

Reproduction

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“…4A). Besides these compounds, we identified and quantified cysteinylglycine according to [57]. The concentrations of cysteine (8.3 l 0.2 lM), cystine (10.2 l 0.5 lM), homocysteine (9.6 l 0.1 lM), GSH (11.6 l 0.3 lM), GSSG (4.5 l 0.2 lM), and cysteinylglycine (35 l 0.4 lM) were determined.…”

Section: Determination Of the Thiols In Different Biological Matricesmentioning

confidence: 99%

Simultaneous determination of eight biologically active thiol compounds using gradient elution‐liquid chromatography with Coul‐Array detection

Petrlová¹,

Mikelová²,

Stejskal³

et al. 2006

J of Separation Science

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The most active form of sulfur in biomolecules is the thiol group, present in a number of biologically active compounds. Here we present a comprehensive study of thiol analysis using flow injection analysis/HPLC with electrochemical detection. The effect of different potentials of working electrodes, of organic solvent contents in the mobile phase, and of isocratic and gradient elution on simultaneous determination of thiol compounds (cysteine, cystine, N-acetylcysteine, homocysteine, reduced and oxidised glutathione, desglycinephytochelatin, and phytochelatins) are described and discussed. These thiol compounds were well separated and detected under optimised HPLC-electrochemical detection conditions (mobile phase: 80 mM trifluoroacetic acid and methanol with a gradient profile starting at 97:3 (TFA:methanol), kept constant for the first 8 min, then decreasing to 85:15 during one minute, kept constant for 8 min, and finally increasing linearly up to 97:3 from 17 to 18 min; the flow rate was 0.8 mL/min, column and detector temperature 25 degrees C, and the electrode potential 900 mV). We were able to determine tens of femtomoles (3 S/N) of the thiols per injection (5 microL), except for phytochelatin5 whose detection limit was 2.1 pmole. This technique was consequently used for simultaneous determination of compounds of interest in biological samples (maize tissue and human blood serum).

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“…However, many electroanalyses of biological samples require the prior use of a separation technique as samples may contain many reactive species that may impede the electrochemical detection. [1][2]4,8,15,17,[20][21][22][23][24] This becomes a particular problem when continual and high throughput monitoring is needed.…”

Section: Introductionmentioning

confidence: 99%

“…[7][8][9]36 Consequently, reports that focus on electrochemical detection in human plasma are usually coupled with a separation technique, such as HPLC. [1][2]4,8,15,17,[20][21][22][23] In our experiments, human plasma was diluted to 25% with 0.15 M PBS (pH 7.2), and an optimal catechol concentration of 1.0 mM was used, because higher concentrations resulted in coagulation. Lastly, all three electrodes on the disposable screen-printed electrode, itself, were fully used due to the small quantity of available human plasma.…”

Section: Detection In Human Plasmamentioning

confidence: 99%

Selective Thiol Detection in Authentic Biological Samples with the Use of Screen-printed Electrodes

Lee

Compton

2015

ANAL. SCI.

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A selective voltammetric determination of homocysteine and glutathione was applied to cell tissue culture media and human plasma via a single two-step method. The two-step method relies on the 1,4-Michael addition reaction between electro-oxidized catechol and the target thiol. Furthermore, the procedure relies on the differing reaction kinetics of the ortho-quinone with various thiol species giving different responses as a function of the scan rate. At faster scan rates homocysteine is only detected, while at slower scan rates the adduct signal reflects both homocysteine and glutathione. As a result, the quantification of both homocysteine and glutathione can be determined with a combination of both sets of data. The previous proof-of-concept (P. T. Lee, D. Lowinsohn, and R. G. Compton, Sensors, 2014, 14, 10395), is applied to the quantification of thiols in both tissue culture media and human plasma alone. Analytical parameters were determined for both homocysteine and glutathione in the respective media and the linear range. The sensitivities in tissue culture media are ca. 3 nA μM -1 and ca. 1 nA μM -1 and the limits of detections are ca. 2 μM and ca. 1 μM for homocysteine and glutathione, respectively. In human plasma, the sensitivities were determined to be 94 and 39 nA μM -1 , and the limit of detections are ca. 0.8 μM and ca. 0.8 μM for homocysteine and glutathione, respectively.

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Simultaneous determination of total plasma glutathione, homocysteine, cysteinylglycine, and methionine by high‐performance liquid chromatography with electrochemical detection

Cited by 68 publications

References 26 publications

Placental markers of folate-related metabolism in preeclampsia

Placental markers of folate-related metabolism in preeclampsia

Simultaneous determination of eight biologically active thiol compounds using gradient elution‐liquid chromatography with Coul‐Array detection

Selective Thiol Detection in Authentic Biological Samples with the Use of Screen-printed Electrodes

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