本试验旨在建立一种测定棉籽粕中游离棉酚含量的高效液相色谱(HPLC)法的优化方法。试验采用丙酮提取棉籽粕样品,经超声、离心、抽滤、旋转蒸发处理,用乙腈-0.2%磷酸(体积比为85: 15)溶液溶解;色谱条件为:色谱柱Agilent TC-C₁₈ (250 mm×4.6 mm,5 μm),流动相为乙腈-0.2%磷酸(体积比为85: 15)溶液,流速1.0 mL/min,紫外检测波长235 nm,进样量20 μL,柱温25 ℃。结果得出,游离棉酚的回归标准曲线为y=189 347x+24 969(r=0.999 99),在游离棉酚含量为0.122 4~78.336 μg/mL范围内线性良好,最低定量限为0.52 mg/kg;棉籽粕样品36 h内4 ℃稳定性的相对标准偏差在1.51%~3.43%;平均回收率在94.72%~99.49%,相对标准偏差在1.72%~3.13%。可知,本方法稳定性好,准确度、灵敏度和回收率高。

An optimized method based high-performance liquid chromatographic (HPLC) was developed for the determination of free gossypol content in cottonseed meal. The gossypol in cottonseed meal was extracted with acetone by ultrasound, centrifugation, suction filter, rotating evaporation, and then dissolved with the mixture of acetonitrile and 0.2% phosphoric acid (85: 15, V/V). The optimal conditions of HPLC were as follows: the column was Agilent TC-C₁₈ (250 mm×4.6 mm, 5 μm), and mobile phase was the mixture of acetonitrile and 0.2% phosphoric acid (85: 15, V/V) with a flow rate of 1.0 mL/min. UV detected wavelength was set at 235 nm, the injected sample volume was 20 μL and the column temperature was 25 ℃. The results showed that the equation of the standard curve was y=189 347x+24 969 (r=0.999 99), which had a highly linear relationship when gossypol content ranged from 0.122 4 μg/mL to 78.336 μg/mL, and the minimum detection limit of gossypol was 0.52 mg/kg. The relative standard deviation of gossypol extracted from cottonseed meal stability at 4 ℃ within 36 h was 1.51% to 3.43%. The average recovery rate of the method was 94.72% to 99.49% with relative standard deviation of 1.72% to 3.13%. In conclusion, the results show that the proposed HPLC method is of good stability, accuracy, sensitivity, and high efficiency.