摘要
目的优化盐酸氨基葡萄糖的柱前衍生反应条件及其含量测定的高效液相色谱条件。方法以异硫氰酸苯酯(PITC)为衍生化试剂,采用响应面优化PITC浓度、时间及介质pH值等反应条件;采用C18色谱柱(250mm×4.6mm,5 m),流动相为甲醇-水,流速为1.0 ml/min,梯度洗脱,检测波长245 nm。结果最佳衍生化反应条件为:取0.5ml供试品溶液,加入2.5 ml pH 7.14的PBS溶液,室温放置15 min,再加入2.0 ml 0.25%PITC试剂,于80℃水浴41.75min后迅速冷至4℃,用流动相定容至10 ml。在该色谱条件下,盐酸氨基葡萄糖衍生物的保留时间为7.5 min,在50～500 g/ml浓度内呈良好线性(r=0.9983),回收率为99.56%,RSD<2.43%。结论该方法操作简单、结果重复性好,适用于盐酸氨基葡萄糖制剂含量的测定。
Objective To optimize the condition of pre-column derivatization and establish RP-HPLC method for determination of glucosamine hydrochloride （GAH）. Methods The phenylisothiocyanate（PITC） was used as derivatization reagent. The derivatization conditions were optimized with Box-Behnken design response surface. HPLC was adopted with ODS C18 column （250mm 4.6mm, 5gm） . The mobile phase consisted of methanol-water （gradient elution） with the flow rate of 1.0ml/min, detection wavelength of 245 nm. Results The derivatization condition was as follows： 2.0ml of 0.25% PITC methanolic solution was added into the mixture （0.5ml sample solution along with 2.5ml of pH7.14 phosphate buffer solution） after 15min, and then the above solution was placed in water bath at 80℃ for 41.75 min. The sample was cooled at 4℃ at once before volumed to 10ml with HPLC mobile phase. The retention time of the derivative was 7.5 min. The standard curves for GAH showed linearity at the concentration of 50-500 gg/ml. The average recovery was 99.56%（RSD 0.51%）. Conclusion This method was found to be easy and specific. It is applicable for determination of GHA in preparations.
出处
《营养学报》
CAS
CSCD
北大核心
2013年第4期387-390,共4页
Acta Nutrimenta Sinica
基金
宁波市科技局自然科学基金项目(No.2011A610034)
