同时测定软骨酶解物中硫酸软骨素及透明质酸含量的高效液相色谱法的建立

Establishment of an HPLC Method for Simultaneous Determination of Chondroitin Sulfate and Hyaluronic Acid in Cartilage Hydrolysate

摘要:

为了准确快速同时测定软骨酶解物中硫酸软骨素和透明质酸含量，以半乳糖胺和葡萄糖胺为测定目标物，通过优化色谱条件和方法性试验，建立了一种高效液相色谱的测定方法。使用ZORBAX C18柱分离，用乙腈和乙酸铵溶液为流动相进行梯度洗脱，流速为1.00 mL/min，在波长250 nm下测定。方法性试验表明，硫酸软骨素和透明质酸的质量浓度在100~500 mg/L范围与峰面积线性关系良好，其中硫酸软骨素的检出限为0.948 mg/L，重复性偏差0.089%，平均回收率99.71%，相对偏差0.35%；透明质酸的检出限为0.027 mg/L，重复性偏差0.045%，平均回收率97.74%，相对偏差0.92%。与传统方法相比，该法具有受杂质干扰影响小，准确度高，重复性优异，回收率以及精确度高的优势，可以实现准确快速同时测定软骨酶解物中硫酸软骨素和透明质酸的含量。

Abstract:

To accurately and rapidly determine the contents of chondroitin sulfate and hyaluronic acid in cartilage hydrolysate, galactosamine and glucosamine were used as the target substances. A determination method based on high-performance liquid chromatography was developed by optimizing the chromatographic conditions and methodological tests. The ZORBAX C18 column was used for separation, with acetonitrile and ammonium acetate solution as the mobile phase for gradient elution. The flow rate was 1.00 mL/min, and the measurement wavelength was 250 nm. Methodological tests showed that the mass concentration of chondroitin sulfate and hyaluronic acid followed a good linear relationship with the peak area in the range of 100~500 mg/L. The limit of detection of chondroitin sulfate was 0.948 mg/L, its repeatability deviation was 0.089%, average recovery rate was 99.71%, and relative deviation was 0.35%. Meanwhile, the limit of detection of hyaluronic acid was 0.027 mg/L, its repeatability deviation was 0.045%, average recovery rate was 97.74%, and relative deviation was 0.92%. Compared with the conventional method, the proposed method offers the advantages of being less affected by interference from impurities and having high accuracy, excellent repeatability, high recovery rate, and high precision. Using this method, the contents of chondroitin sulfate and hyaluronic acid in cartilage hydrolysate can be detected rapidly and accurately.