荧光高效液相色谱法测定血清中总唾液酸含量的分析  

Analysis of the total sialic acid content in serum determinated with fluorometric high-performance liquid chromatography

作　　者：柏丹丹[1] 李红卫[2] 潘莉莉[2] 王冰[3] 古桂雄[1] 

机构地区：[1]苏州大学儿科研究所,江苏苏州215003 [2]厦门大学公共卫生学院,福建厦门361005 [3]厦门大学医学院,福建厦门361005

出　　处：《中国儿童保健杂志》2012年第11期1022-1025,共4页Chinese Journal of Child Health Care

基　　金：江苏社会发展(BE2011673);苏州市科技局(SZS201108;SYS201145)

摘　　要：【目的】探讨荧光高效液相色谱法测定血清中总唾液酸含量的最佳条件。【方法】按优化实验的原则,将人体血清分别用三氟乙酸、硫酸、盐酸80℃水解,酸化浓度分别为0.05、0.10、0.15、0.20mol/L,酸解时间分别为30、60、90、120min,经50℃避光衍生150min后进行色谱分析。【结果】用0.10mol/L TFA溶液80℃水解60min测定的唾液酸浓度最佳,唾液酸的最适宜浓度在0.625～80μmol/L范围内,其平均回收率为94.0%,精密度、稳定性、重复性试验的相对标准差分别为1.3%、1.1%和1.0%。Neu5Ac的最低检出限为0.02μmol/L,16～19岁健康青少年血清中唾液酸(Neu5Ac)含量的平均值为(2.12±0.15)mmol/L。【结论】荧光高效液相色谱法测定唾液酸简单、重复性好、灵敏度高,可广泛用于人体血清唾液酸含量测定。[Objective] To determine the optimum conditions for total sialic acid analysis in serum using the fluoro- metric high-performance liquid chromatography （HPLC）. [Methods] The sialic acid standard operating fluid contained N-acetylneuraminic acid （Neu5Ac）,N-glyeolylneuraminic acids （Neu5Gc） and ketodeoxynonulosonic acid （KDN）. The hu- man plasma would be hydrolyzed with the trifluoroacetic acid （TFA）,sulphuric acid and hydrochloric acid at different con- centration of 0. 05,0.10,0.15 mol/L and 0.20 mol/L at 80℃ and at varying time of 30,60,90 rain and 120 min respectively. The hydrolyzed sample was treated with 4,5-Methylenedioxy-l,2- phenylenediamine dihydrochloride （DMB,Sigma） derivative reagent at 50 ℃ for 150 min away from light. [Results] The maximum release of sialic acid was obtained with an equal volume of 0.1 mol/L TFA for 60 min at 80℃. The optimum detection limit of sialic acid was in 0. 625-80 μmol/L, the average recovery rate was 94.0 % and the relative standard deviation （RSD） for the precision,stability and repeatability test were 1.3 %, 1.1% and 1.0 %, respectively. The minimum detection limit of NeuSAc was 0.02μmol/L and the average content of sialic acid in human serum was （2.12±0.15） mmol/L in 16-19 years old healthy adolescents. [Conclusion] The total sialic acid content determinated with fluorometric HPLC method is a simple,high sensitivity and good repeatability,which can be widely used in determination of sialic acid concentration in serum.

关 键 词：血清 唾液酸 荧光高效液相色谱 

分 类 号：R722[医药卫生—儿科]