Determination of ethoxidol in human plasma by microemulsion liquid chromatography

Abstract

One of the most important directions of the development of pharmacological science is the development and introduction into clinical practice of medicines of natural and synthetic origin. A striking representative of domestic cytoprotectors is malic acid salt.

3-hydroxy-6-methyl-2-ethylpyridine, patented under the trade name "Ethoxidol". It is not inferior, and sometimes surpasses similar drugs in its pharmacological activity. Only one work has been found in the literature devoted to the determination of ethoxidol in medicinal preparations by voltammetry. Sensitivity is not sufficient for its determination in blood plasma. It was necessary to develop a more sensitive and selective method.

Microemulsions are often used in variants of capillary electrokinetic chromatography or sample preparation, but the method of microemulsion liquid chromatography (MLC) has not been so widely used. The method has certain advantages over HPLC. In this work, the determination of ethoxidol in blood plasma was carried out by high-performance microemulsion chromatography with fluorimetric detection. 300 mm³ microemulsions were added to 300 mm³ of plasma and thoroughly mixed to precipitate plasma proteins. The addition of microemulsion prevents the co-deposition of ethoxidol together with proteins. The filler fluid was taken and injected into a chromatograph. The degree of extraction was controlled chromatographically by adding a known amount of ethoxidol to the extract from the blank plasma.

In a micellar medium, substances can be solvated differently than in aqueous methanol/acetonitrile solutions. Changes in the spectral characteristics of substances are caused by the presence of oil in the composition of the microemulsion, the introduction of substances into a drop of which leads to a spectral shift. When determining ethoxidol, the excitation and detection wavelengths of λ_ex 287 nm and λ_em 399 nm, respectively, were selected. It should be emphasized separately that the microemulsion additionally allows you to stabilize the components being determined. Thus, in the case of determination of ethoxidol in blood plasma, standard methods of sample preparation (liquid and solid-phase extraction, precipitation of proteins with organic solvents, acids, metal ions) did not lead to the desired result. In the case of dilution of the sample with microemulsion, the extraction was quantitative.

The calibration dependence was linear in the concentration range 0.1-10 mg/dm³. The graph is described by the linear equation S=23.97C-1.257. The correlation coefficient R²=0.998. The detection limit is 50 micrograms/dm³. The relative standard deviation of S(r) is 0.08 (n=3). Thus, a new chromatographic method for determining ethoxidol in human blood plasma is proposed. It is shown that the use of microemulsion as an eluent leads to the stabilization of the drug in solutions and a significant increase in the extraction of ethoxidol from real samples.