Determination of hesperetin and its conjugate metabolites in serum and urine

Hesperidin and hesperetin are flavanones in many Citrus fruits with antiulcer, antioxidation and anticancer activities. Hesperetin is the aglycone and a metabolite of hesperidin. HPLC analysis was developed for the determination of hesperetin and its conjugate metabolites in serum and urine prior to and after hydrolysis following oral administrations of hesperetin, hesperidin, or decoction of Pericarpium Citri Reticulatae (Chenpi). The hydrolysis was conducted away from light by using glucuronidase/sulfatase in the presence of ascorbic acid at 37°C. The HPLC methods employed a LiChrospher 100 RP-18e column, using acetonitrile/1% acetic acid (36/65) and methanol/acetonitrile/1% acetic acid (17/26/57) as mobile phases for serum and for urine, respectively. For internal standard, 5,7-dimethoxycoumarin was used. The present methods are rapid, sensitive, reproducibility, precise, and accurate with good recoveries. They are applicable for the pharmacokinetic studies of hesperidin and hesperetin as well as Citrus foods or herbs containing the them.