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Development of a Solid‐Phase Extraction–Enzyme‐Linked Immunosorbent Assay for the Determination of 17β‐19‐Nortestosterone Levels in Antifatigue Functional Foods
Author(s) -
Zhang Yan,
Pan Tao,
Fang Guozhen,
Ma Dongyue,
Wang Shuo
Publication year - 2009
Publication title -
journal of food science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.772
H-Index - 150
eISSN - 1750-3841
pISSN - 0022-1147
DOI - 10.1111/j.1750-3841.2009.01311.x
Subject(s) - chromatography , detection limit , solid phase extraction , polyclonal antibodies , chemistry , extraction (chemistry) , matrix (chemical analysis) , conjugate , bovine serum albumin , antibody , biology , immunology , mathematical analysis , mathematics
17β‐19‐nortestosterone (17β‐NT) has been illegally used in antifatigue functional foods to promote muscle growth and improve endurance. A rapid and sensitive solid‐phase extraction–enzyme‐linked immunosorbent assay (SPE‐ELISA) method was developed and successfully applied to analyze the levels of 17β‐NT in antifatigue functional foods. A polyclonal antibody against 17β‐NT was produced from rabbits immunized with the 17β‐NT‐BSA conjugate, and a competitive direct enzyme‐linked immunosorbent assay was developed for the rapid detection of 17β‐NT. The concentration causing 50% inhibition ( IC 50 ) and the limit of detection (LOD) were found to be 0.08 and 0.0055 ng/mL, respectively; this was better than methods previously reported that had a LOD of 2.4 ng/mL. C 18 cartridges were investigated for use in removing the effects of matrix in foods, and the sample purification protocol was optimized. Using the developed SPE‐ELISA method, recoveries of functional food samples were obtained in the range of 71% to 91.5%. Moreover, 2 kinds of antifatigue functional foods were analyzed using the established ELISA and HPLC methods. The correlation coefficient of the results obtained using the 2 methods was greater than 0.98. Thus, the preliminary evaluation of the SPE‐ELISA method proved that it is a specific, sensitive, and precise tool that can be used for the practical detection of 17β‐NT in various antifatigue functional food samples.