An enzyme‐linked immunosorbent assay for the measurement of plasma flavonoids in mice fed apigenin‐ C ‐glycoside

BACKGROUND In the Chenopodiaceae family, the apigenin flavonoids vitexin‐2‐ O ‐xyloside ( VOX ) and vitexin‐2‐ O ‐rhamnoside ( VOR ) are important chemopreventive components. To investigate their bioavailability in in vivo animal studies an enzyme‐linked immunosorbent assay ( ELISA ) method has been developed . RESULTS The ELISA was based on polyclonal antibodies elicited in mice by injecting, as an immunogen, 4′,6″‐ O ‐biapigenin (hinokiflavone, HF ) conjugated to bovine serum albumin ( BSA–HF ). A second immunogen was synthesised by coupling an equimolar mixture of VOX and VOR to BSA ( BSA–F1 ). The BSA–HF elicited a significant antibody response, due to 17 HF hapten groups, coupled to each BSA molecule, whereas BSA–F1 provided a very low antigenicity in respect to control animals. Antiserum raised against BSA–HF showed an antibody titre of 1:1600. Antibodies were found to be specific for the flavonols. Our results show that VOX and its metabolic products reached the concentration of 3.42 ± 0.72 µg  mL −1 in plasma of VOX fed animals, at the net of the control value . CONCLUSIONS By using the ELISA , the concentration of apigenin flavonoids and their metabolites can be detected in VOX ‐ or VOR ‐supplemented animals. The assay represents a useful tool for rapid screening to compare bioavailability of apigenin flavonoids in respect to control animals. © 2013 Society of Chemical Industry