Determination of Gossypol in Feeds by HPLC

Abstract

An HPLC method for determination of goosypol in feed was developed. Gossypol in food was extracted with acetic acid-water-phosphoric acid (85 : 15 : 1) for 20 min in a water bath at 100°C. The extract was diluted with acetone-water (1 : 1), and injected into the HPLC. HPLC was performed with a Shodex C18M4E (4.6 mm i.d.×250 mm) column at a flow-rate of 1.0 mL/min, using a mobile phase of methanol-water (9 : 1) adjusted to pH 2.6 with phosphoric acid, and gossypol was detected with a UV detector (254 nm). A recovery test was conducted with cottonseed spiked with gossypol at 1,000 and 5,000 mg/kg, and with 2 kinds of formula feed spiked with gossypol at 58 and 580 mg/kg. The mean recoveries of gossypol were 90.8-105.0% and the relative standard deviations (RSD) were within 3.0%. A collaborative study was conducted with cottonseed and formula feed spiked with gossypol at 305 mg/kg in 8 laboratories. The average content of gossypol in cottonseed was 6,090 mg/kg, and the repeatability and reproducibility as the relative standard deviation (RSD_r and RSD_R) were 3.3% and 4.4%, while the mean recovery, RSD_r and RSD_R of gossypol in formula feed were 87.0%, 2.7% and 5.5%, respectively.