Joint determination of nine uremic toxins and choline in blood serum using high-performance liquid chromatography with tandem mass spectrometric detection

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Abstract

This paper presents a method for the simultaneous determination of nine uremic toxins and choline in blood serum. The target substances were selected based on literature data as potential biomarkers for assessing the severity and progression of immunoglobulin A nephropathy, a kidney disease that leads to disability, and, without timely treatment, death in young and middle-aged individuals. Ultrafiltration is used to separately determine free and protein-bound indolic uremic toxins in the blood. The use of high-performance liquid chromatography combined with high-resolution tandem mass spectrometry ensures satisfactory accuracy of the analysis without complete chromatographic separation of analytes under standard reversed-phase HPLC conditions. For calibration purposes, an albumin solution in phosphate buffer was used as a surrogate for blood serum. The protein concentration of 45 mg/mL and pH 7.4 match the characteristics of native blood serum. A pilot experiment demonstrated the feasibility of determining key indicators of the gut microbiome’s state—choline and trimethylamine N-oxide (TMAO)—in dried blood spots.

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About the authors

T. I. Alyushina

Research Institute of Hygiene, Occupational Pathology, and Human Ecology of the Federal Medical-Biological Agency of Russia

Email: saveleva@rihophe.site
Russian Federation, Kuzmolovsky Settlement, Leningrad Region

E. I. Savelyeva

Research Institute of Hygiene, Occupational Pathology, and Human Ecology of the Federal Medical-Biological Agency of Russia

Author for correspondence.
Email: saveleva@rihophe.site
Russian Federation, Kuzmolovsky Settlement, Leningrad Region

V. A. Dobronravov

Research Institute of Nephrology, I.P. Pavlov First St. Petersburg State Medical University

Email: saveleva@rihophe.site
Russian Federation, St. Petersburg

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Supplementary files

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2. Рис. 1. Схема определения свободных и связанных с белками уремических токсинов в сыворотке крови.

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3. Рис. 2. Схема определения холина и триметиламин-N-оксида (ТМАО) в сухих пятнах крови.

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4. Рис. 3. Результаты сравнения картриджей Whatman 903 и Биохран на примере определения холина и триметил амин-N-оксида (ТМАО) в технике сухих пятен крови.

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