Chen Yulong, Yao Qin, Zeng Xuan, Hao Cui, Li Xiulian, Zhang Lijuan, Zeng Pengjiao
Department of Gynaecology, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China.
Medical Research Center, The Affiliated Hospital of Qingdao University, Qingdao, Shandong, China.
Front Oncol. 2022 Nov 3;12:1014159. doi: 10.3389/fonc.2022.1014159. eCollection 2022.
Altered glycan levels in serum have been associated with increased risk of cancer. In this study, we have developed and validated a HPLC-based method to analyze monosaccharide composition (D-mannose, Glucosamine, Galactosamine, Glucuronic acid, D-glucose, D-galactose, D-xylose, L-fucose) in human serum, with L-rhamnose, being used as internal standard. Monosaccharides obtained from hydrolyzed serum samples were derivatized by 1-Phenyl-3-methyl-5-pyrazolone. A ZORBAX XDB-C18 column(150×4.6mm) was used for chromatographic separation with 100 mM ammonium acetate buffer (NH4Ac-HAc, PH=5.5, solvent A), acetonitrile (ACN, solvent B) as a mobile phase. The calibration standard curves for the eight monosaccharides showed good linearity over the range of 2.5-500μg/mL with R > 0.995. The relative standard deviation values for intra-day and inter-day precision were ≤ 5.49%. Recovery was 69.01-108.96%. We observed that this column exhibited high specificity and selectivity to separate monosaccharides from serum. This method was then applied to quantitatively analyze the serum monosaccharide levels in 30 patients with endometrial cancer and 30 matched healthy controls. Statistical analysis indicated that the serum monosaccharide levels were significantly higher in patients compared with healthy controls (P value< 0.0001). Overall, we report here a simple, reliable, low-cost, and reproducible HPLC method for the separation and quantification monosaccharides in the human serum, which has potential value to serve as a screening marker for endometrial cancer.
血清中聚糖水平的改变与癌症风险增加有关。在本研究中，我们开发并验证了一种基于高效液相色谱法（HPLC）的方法，用于分析人血清中的单糖成分（D-甘露糖、氨基葡萄糖、半乳糖胺、葡萄糖醛酸、D-葡萄糖、D-半乳糖、D-木糖、L-岩藻糖），以L-鼠李糖作为内标。从水解血清样品中获得的单糖用1-苯基-3-甲基-5-吡唑啉酮进行衍生化。使用ZORBAX XDB-C18柱（150×4.6mm）进行色谱分离，以100 mM醋酸铵缓冲液（NH4Ac-HAc，PH=5.5，溶剂A）、乙腈（ACN，溶剂B）作为流动相。八种单糖的校准标准曲线在2.5-500μg/mL范围内显示出良好的线性，R>0.995。日内和日间精密度的相对标准偏差值≤5.49%。回收率为69.01-108.96%。我们观察到该柱对从血清中分离单糖具有高特异性和选择性。然后将该方法应用于定量分析30例子宫内膜癌患者和30例匹配的健康对照者的血清单糖水平。统计分析表明，患者的血清单糖水平显著高于健康对照者（P值<0.0001）。总体而言，我们在此报告了一种简单、可靠、低成本且可重复的HPLC方法，用于分离和定量人血清中的单糖，该方法具有作为子宫内膜癌筛查标志物的潜在价值。
