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S. Makhsatova

National Center for Biotechnology, 13/5 Korgalzhyn Road, Astana, 010000, Kazakhstan

A. Kulyyassov

National Center for Biotechnology, 13/5 Korgalzhyn Road, Astana, 010000, Kazakhstan


Anionic detergents like SDC are commonly used in biochemical and molecular biology research for solubilizing and separating proteins and nucleic acids. The detection method using methylene blue (MB) can be employed to determine the concentration of anionic detergents in these experimental preparations. The quantification protocol for surface active substances was optimized, with the consideration of factors such as pH, salts and the spectrophotometer wavelength. Several extraction methods were employed to remove the detergent from the sample, namely, ethyl acetate extraction, mineral oil and acid precipitation. All of the methods have about the same efficiency. The MB method was used as a standard technique for the precise quantification of detergent amounts which allowed as achieving an approximate concentration range between the lower limit of ca. 0.02% and 0.1% of the upper limit. Consequently, we weren’t able to detect the concentration of the detergent lower than 0.02%. In the presence of anionic detergents, the electrostatic interaction between positively charged MB and the anions forms a complex that can be extracted into chloroform, allowing for quantification. Without the formation of this complex, water-soluble MB remains immiscible in chloroform.


Calibration curves, Liquid Chromatography tandem Mass Spectrometry (LC-MS/MS), methylene blue (MB), methylene blue active substances (MBAS), sodium deoxycholate (SDC), sodium dodecyl sulfate (SDS), spectrophotometry, serial dilution

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Badmus S. O., Amusa H. K., Oyehan T. A., Saleh T. A. Environmental risks and toxicity of surfactants: overview of analysis, assessment, and remediation techniques // Environ Sci Pollut Res Int. ‒ 2021. ‒ Vol. 28, № 44. ‒ P. 62085-62104. Crossref

Abbott D. C. The colorimetric determination of anionic surface-active materials in water // Analyst. ‒ 1962. ‒ Vol. 87, № 1033. ‒ P. 286-293. Crossref

Ghorbani F., Ekhtiari M., Moeini Chaghervand B., Moradi L., Mohammadi B., Kajbafzadeh A. M. Detection of the residual concentration of sodium dodecyl sulfate in the decellularized whole rabbit kidney extracellular matrix // Cell Tissue Bank. ‒ 2022. ‒ Vol. 23, № 1. ‒ P. 119-128. Crossref

Zvarova B., Uhl F. E., Uriarte J. J., Borg Z. D., Coffey A. L., Bonenfant N. R., Weiss D. J., Wagner D. E. Residual Detergent Detection Method for Nondestructive Cytocompatibility Evaluation of Decellularized Whole Lung Scaffolds // Tissue Eng Part C Methods. ‒ 2016. ‒ Vol. 22, № 5. ‒ P. 418-28. Crossref

Laemmli U. K. Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 // Nature. ‒ 1970. ‒ Vol. 227, № 5259. ‒ P. 680-685. Crossref

Botelho D., Wall M. J., Vieira D. B., Fitzsimmons S., Liu F., Doucette A. Top-Down and Bottom-Up Proteomics of SDS-Containing Solutions Following Mass-Based Separation // Journal of Proteome Research. ‒ 2010. ‒ Vol. 9, № 6. ‒ P. 2863-2870. Crossref

Ikonomou M. G., Blades A. T., Kebarle P. Investigations of the electrospray interface for liquid chromatography/mass spectrometry // Analytical Chemistry. ‒ 1990. ‒ Vol. 62, № 9. ‒ P. 957-967. Crossref

Rundlett K. L., Armstrong D. W. Mechanism of Signal Suppression by Anionic Surfactants in Capillary Electrophoresis−Electrospray Ionization Mass Spectrometry // Analytical Chemistry. ‒ 1996. ‒ Vol. 68, № 19. ‒ P. 3493-3497. Crossref

Epton S. R. A Rapid Method of Analysis for Certain Surface-Active Agents // Nature. ‒ 1947. ‒ Vol. 160, № 4075. ‒ P. 795-796. Crossref

Mukerjee P. Use of Ionic Dyes in Analysis of Ionic Surfactants and Other Ionic Organic Compounds // Analytical Chemistry. ‒ 1956. ‒ Vol. 28, № 5. ‒ P. 870-873. Crossref

Hayashi K. A rapid determination of sodium dodecyl sulfate with methylene blue // Analytical Biochemistry. ‒ 1975. ‒ Vol. 67, № 2. ‒ P. 503-506. Crossref

George A. L., White G. F. Optimization of the methylene blue assay for anionic surfactants added to estuarine and marine water // Environmental Toxicology and Chemistry. ‒ 1999. ‒ Vol. 18, № 10. ‒ P. 2232-2236. Crossref

Jurado E., Fernandez-Serrano M., Nunez-Olea J., Luzon G., Lechuga M. Simplified spectrophotometric method using methylene blue for determining anionic surfactants: applications to the study of primary biodegradation in aerobic screening tests // Chemosphere. ‒ 2006. ‒ Vol. 65, № 2. ‒ P. 278-85. Crossref

Masuda T., Tomita M., Ishihama Y. Phase transfer surfactant-aided trypsin digestion for membrane proteome analysis // J Proteome Res. ‒ 2008. ‒ Vol. 7, № 2. ‒ P. 731-40. Crossref

Golz E. K., Vander Griend D. A. Modeling methylene blue aggregation in acidic solution to the limits of factor analysis // Anal Chem. ‒ 2013. ‒ Vol. 85, № 2. ‒ P. 1240-6. Crossref

Ghosh A. K. Study of the self-association of methylene blue from protonation equilibriums // Journal of the American Chemical Society. ‒ 1970. ‒ Vol. 92, № 22. ‒ P. 6415-6418. Crossref

Saputra E., Saputra R., Nugraha M. W., Irianty R. S., Utama P. S. Removal of Methylene Blue from aqueous solution using spent bleaching earth // IOP Conference Series: Materials Science and Engineering. ‒ 2018. ‒ Vol. 345. Crossref

Lundell N., Schreitmuller T. Sample preparation for peptide mapping--A pharmaceutical quality-control perspective // Anal Biochem. ‒ 1999. ‒ Vol. 266, № 1. ‒ P. 31-47. Crossref

Xing G., Zhang J. M., Chen Y., Zhao Y. M. Identification of four novel types of in vitro protein modifications // Journal of Proteome Research. ‒ 2008. ‒ Vol. 7, № 10. ‒ P. 4603-4608. Crossref