Détermination du 5-méthyltetrahydrofolate dans le sang total par chromatographie liquide

Given the clinical importance of vitamin B9 and its role in many disorders, its determination and particularly of one of its predominant circulating form: 5-methyltetrahydrofolate is a necessity. The present study describes a liquid chromatography method that allows determination of 5-methyltetrahydrofolate in whole blood. Defrosted whole blood samples were deproteinized in an ascorbic acid solution at + 37°C, then submitted to a liquid-liquid partition extraction. The extract is then analyzed by reversed phase liquid chromatography followed by fluorimetric detection within 9 minutes. Different system suitability parameters and validation procedure tests were applied to evaluate the reliability o the proposed method. System suitability parameters were satisfactory for the intended application. Precision results for 5-methyltetrahydrofolate were better than 2 % for chromatographic analysis and better than 5 % for the entire procedure and interassay. Average recovery results was 92.50 %. Limits of detection and quantification were respectively 8 and 10 nmol/l. The proposed liquid chromatography method determines 5-methyltetrahydrofolate in whole blood with a satisfactory reliability.