核黄素、光黄素、光色素及其他有关物质的高效液相色谱测定

HPLC determination of riboflavin,lumiflavin,lumichrome and other related substances

目的:建立一种反相高效液相色谱法测定核黄素的含量及其有关物质。方法:采用Agilent Zarbox C18(250 mm×4.6mm,5μm)色谱柱,以甲醇(A)-0.1%醋酸(B)为流动相梯度洗脱(0～15 min,30%A;15～25 min,30%A→60%A;25～35min,60%A),流速1.0 mL.min-1,检测波长267 nm,柱温为室温,进样量20μL,外标法定量。结果:在该色谱条件下,核黄素及其有关物质能够得到很好的分离,分离度大于1.5,理论塔板数按核黄素计大于5000;核黄素浓度在1～30μg.mL-1范围内与峰面积呈良好的线性关系(r=0.9999);定量限和检测限分别为1.08 ng(S/N=10)和0.36 ng(S/N=3)。结论:该高效液相色谱法准确、灵敏、可靠,专属性好,可用于核黄素的含量及其有关物质的测定。

Objective：To establish an RP-HPLC method for the determination of the content of riboflavin and its related substances.Methods：The analysis was carried on an Agilent Zarbox C18（250 mm×4.6 mm,5 μm） column;The mobile phase was composed of methanol（A） and 0.1% acetic acid（B） with gradient elution（0-15 min,30% A;15-25 min,30% A→60% A;25-35 min,60% A） at the flow rate of 1.0 mL·min-1;The detection wavelength was 267 nm,and the column temperature was room temperature;Loaded volume was 20 μL.Results：Using this method,riboflavin and its related substances were well separated and the degree of separability was more than 1.5,the number of theoretical plates with riboflavin was more than 5000;The linear curve was obtained in the range of 1-30 μg·mL-1（r=0.9999）;The minimal quantifiable level and the minimal detectable level were 1.08 ng（S/N=10） and 0.36 ng（S/N=3）.Conclusion：The method proposed for determination of the content and its related substances of riboflavin is specific,sensitive,reliable,accurate,and can be used for the determination of the content of riboflavin and its related substances.