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Methodological Features of the Spectrophotometric Determination of Proteins in Biological Fluids Using Reactions with Brompyrogallol Red

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Abstract

Determination of proteins in biological fluids is rather important for diagnostics in current clinical practice. The results of total protein determination depend on the amino acid composition of the proteins present in the biological fluid. In this paper, some aspects of the spectrophotometric determination of proteins in biological fluids are discussed, in particular, the methodological features of the technique based on the reaction of proteins with brompyrogallol red (BPGR). The most important advantage of BPGR in the determination of proteins in biological fluids is rather high and equal sensitivity of the dye to the proteins of albumin and globulin fractions, thus minimizing the errors attributed to the mismatch of the protein composition of the analyzed samples and calibration solutions used. The aim of the work was to study the influence of conditions and shelf life of the BPGR solution on the analytical properties of the solution in the spectrophotometric determination of proteins in biological fluids. Stability of the optical and analytical properties of BPGR solutions was studied using Fisher’s and Student’s criteria under conditions of different storage temperatures and nature of the stabilizer (ethanol or sodium benzoate) in the reagent solutions. Verification of the accuracy of the total protein determination by the proposed method was carried out in spike tests. The introduced additives of standard solutions were prepared from the “Total protein” or “Albumin” calibrators. The developed method of the spectrophotometric determination of the mass concentration of proteins in urine by the reaction with bromopyrogallol red was tested on real objects, metrologically certified, and listed in the Federal Register of certified measurement techniques. Analytical and metrological studies have shown that the developed method of protein determination with a reagent based on BPGR provides equal and high sensitivity of the determination of albumin and globulin protein fractions in human biological fluids. To increase the shelf life of the reagent solution and preserve the analytical properties of the solution, we recommend using ethanol as a stabilizer.

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Notes

  1. Instruction RС No. FSH 2007/01435 on the use of a set of reagents for the quantitative determination of total protein in urine and cerebrospinal fluid with pyrogallol red (TOTAL PROTEIN PGK FS “Diakon-DS”). Approved by the Order of Roszdravnadzor dated July 20, 2010 No. 6830 – Pr/10.

  2. GOST R 53079.4–2008. Clinical laboratory technologies. Quality assurance of clinical laboratory research. Part 4. Rules for conducting the preanalytical stage, Moscow: Standartinform, 2009, p. 65.

  3. RMG 61—2010. Indicators of accuracy, correctness, precision of methods of quantitative chemical analysis. Assessment methods, Moscow: Publishing house of standards, 2004, p. 42.

  4. Mass concentration of proteins in urine. Spectrophotometric measurement technique using bromopyrogallol red. Certified by the Federal State Budgetary Institution “GKHI”. Certificate of attestation of the measurement technique dated 04/27/2018 No. S.MI 02067847.04.RA.RU.311345–2018.

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ACKNOWLEDGMENTS

This work was carried out using the scientific equipment of the Ecological Analytical Core Facility Center of the Kuban State University.

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Authors and Affiliations

  1. Kuban State University, 350040, Krasnodar, Russia

    T. B. Pochinok, P. V. Anisimovich & Z. A. Temerdashev

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  1. T. B. Pochinok
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Correspondence to T. B. Pochinok.

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Translated by P. Kuchina

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Pochinok, T.B., Anisimovich, P.V. & Temerdashev, Z.A. Methodological Features of the Spectrophotometric Determination of Proteins in Biological Fluids Using Reactions with Brompyrogallol Red. Inorg Mater 57, 1367–1373 (2021). https://doi.org/10.1134/S0020168521140077

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