Development of a reliable method based on ultra-performance liquid chromatography coupled to tandem mass spectrometry to measure thiol-associated oxidative stress in whole blood samples

Escobar, Javier; Sánchez-Illana, Ángel; Kuligowski, Julia; Torres-Cuevas, Isabel; Solberg, Rønnaug; Garberg, Håvard Tetlie; Huun, Marianne Ullestad; Saugstad, Ola Didrik; Vento, Máximo; Cháfer-Pericás, Consuelo

doi:10.1016/j.jpba.2016.02.007

Cited by 40 publications

(21 citation statements)

References 32 publications

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“…The chromatographic method was adapted from Escobar et al (2016) and optimized empirically based on MS response, peak shape and retention time. Acetonitrile and 0.1% ( v /v) formic acid in ultrapure water resulted in a good MS response with a narrow cystine peak.…”

Section: Resultsmentioning

confidence: 99%

“…The present chromatographic method was adapted from the UPLC‐MS/MS assay described by Escobar et al (2016). By comparing our HPLC‐MS/MS method with the UPLC‐MS/MS method, we have further optimized the method in terms of sensitivity (the volume of sample injected is, 1.0 vs 5.0 μL), and limit of detection for cystine (0.1 vs 0.34 μmol/L).…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, disulfide exchange reactions of cystine with other sulfhydryl groups can lead to the loss of cystine in the cells. To avoid these artifactual oxidation–reduction reactions and in order to have an adequate measurement of cystine in the cells, derivatization with N ‐ethylmaleimide (NEM) is required immediately during sample preparation (de Graaf‐Hess et al, 1999; Escobar et al, 2016). NEM, an alkylating agent, forms stable bonds with sulfhydryl containing molecules, enabling them to be permanently blocked and prevent disulfide bond formation.…”

Section: Introductionmentioning

confidence: 99%

“…Graaf‐Hess et al (1999) have applied such method for measuring cystine in leukocytes and fibroblasts of cystinotic patients. On the other hand, Escobar et al (2016) have recently reported a rapid and reliable bioanalytical assay for detection of cystine and other related thiols in whole blood using ultra‐performance liquid chromatography coupled to tandem mass spectrometry (UPLC‐MS/MS). Unfortunately, these methods are hampered in use because of their high limit of detection and poor cystine recovery.…”

Section: Introductionmentioning

confidence: 99%

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Quantification of cystine in human renal proximal tubule cells using liquid chromatography–tandem mass spectrometry

Jamalpoor

Sparidans

Casellas

et al. 2018

Biomedical Chromatography

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Nephropathic cystinosis is characterized by abnormal intralysosomal accumulation of cystine throughout the body, causing irreversible damage to various organs, particularly the kidneys. Cysteamine, the currently available treatment, can reduce lysosomal cystine and postpone disease progression. However, cysteamine poses serious side effects and does not address all of the symptoms of cystinosis. To screen for new treatment options, a rapid and reliable high‐performance liquid chromatography–tandem mass spectrometry (HPLC‐MS/MS) method was developed to quantify cystine in conditionally immortalized human proximal tubular epithelial cells (ciPTEC). The ciPTEC were treated with N‐ethylmaleimide, lysed and deproteinized with 15% (w/v) sulfosalicylic acid. Subsequently, cystine was measured using deuterium‐labeled cystine‐D4, as the internal standard. The assay developed demonstrated linearity to at least 20 μmol/L with a good precision. Accuracies were between 97.3 and 102.9% for both cell extracts and whole cell samples. Cystine was sufficiently stable under all relevant analytical conditions. The assay was successfully applied to determine cystine levels in both healthy and cystinotic ciPTEC. Control cells showed clearly distinguishable cystine levels compared with cystinotic cells treated with or without cysteamine. The method developed provides a fast and reliable quantification of cystine, and is applicable to screen for potential drugs that could reverse cystinotic symptoms in human kidney cells.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Quantification of cystine in human renal proximal tubule cells using liquid chromatography–tandem mass spectrometry

Jamalpoor

Sparidans

Casellas

et al. 2018

Biomedical Chromatography

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“…The reduced/oxidised glutathione (GHS/GSSG), and its precursors γ-l-glutamyl-l-cysteine (γ-Glu-Cys) and S-adenosyl-L-homocysteine (SAH), were analysed by UPLC-MS/MS according to Escobar et al [18].…”

Section: Introductionmentioning

confidence: 99%

Pterostilbene Prevents Early Diabetic Retinopathy Alterations in a Rabbit Experimental Model

Millán

Torres-Cuevas

et al. 2019

Nutrients

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Oxidative stress generated by diabetes plays a key role in the development of diabetic retinopathy (DR), a common diabetic complication. DR remains asymptomatic until it reaches advanced stages, which complicate its treatment. Although it is known that good metabolic control is essential for preventing DR, knowledge of the disease is incomplete and an effective treatment with no side effects is lacking. Pterostilbene (Pter), a natural stilbene with good antioxidant activity, has proved to beneficially affect different pathologies, including diabetes. Therefore, our study aimed to analyse the protective and/or therapeutic capacity of Pter against oxidant damage by characterising early retinal alterations induced by hyperglycaemia, and its possible mechanism of action in a rabbit model of type 1 diabetes mellitus. Pter reduced lipid and protein oxidative damage, and recovered redox status and the main activities of antioxidant enzymes. Moreover, the redox regulation by Pter was associated with activation of the PI3K/AKT/GSK3β/NRF2 pathway. Our results show that Pter is a powerful protective agent that may delay early DR development.

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Determination of proflavine in rat whole blood without sample pretreatment by laser desorption postionization mass spectrometry

Chen

et al. 2017

Anal Bioanal Chem

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A novel pretreatment-free method involving laser desorption postionization (LDPI) coupled with time-of-flight mass spectrometry (MS) was developed for the monitoring of proflavine level in rat whole blood. It comprises a protocol for dosing via intravenous administration and collection of whole blood, followed by direct LDPI-MS analysis without any sample pretreatment. An intense ion signal at m/z 209 was observed from whole blood without any interference signals, except some background signals below m/z 100. The calibration curve was established with use of 9-phenylacridine as the internal standard for proflavine determination from the plotting of the peak ratios of proflavine to the internal standard, with a correlation coefficient (R ) greater than 0.99. The limit of detection was estimated to be 0.48 pmol/mm and the quantification range was 0.5-16.5 μg/mL for proflavine. In addition, only a minimal matrix effect was observed, as expected from considerations of the desorption and ionization mechanism. Interday and intraday accuracy and precision were calculated to be within 13% and 82-114%, respectively. Estimated concentrations of proflavine residue in whole blood were also successfully obtained at selected time points after dosing. The proposed method is simple, low cost, and sensitive, and should be seen as a complementary method for monitoring drug levels in blood. Graphical Abstract Monitoring proflavine levels in rat whole blood at different time points using laser desorption postionization mass spectrometry (LDPI-MS).

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Development of a reliable method based on ultra-performance liquid chromatography coupled to tandem mass spectrometry to measure thiol-associated oxidative stress in whole blood samples

Cited by 40 publications

References 32 publications

Quantification of cystine in human renal proximal tubule cells using liquid chromatography–tandem mass spectrometry

Quantification of cystine in human renal proximal tubule cells using liquid chromatography–tandem mass spectrometry

Pterostilbene Prevents Early Diabetic Retinopathy Alterations in a Rabbit Experimental Model

Determination of proflavine in rat whole blood without sample pretreatment by laser desorption postionization mass spectrometry

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