气相色谱-质谱联用技术在3-羟基-3-甲基戊二酸尿症诊断中的应用  

Application of Gas Chromatography-Mass Spectrometry Quantitative Techniques in Diagnosis of 3-Hydroxy-3-Methylglutaric Aciduria

作　　者：江敏妍[1] 刘丽[1] 李秀珍[1] 程静[1] 蔡燕娜[1] 彭敏芝[1] 

机构地区：[1]广州医学院附属广州市妇女儿童医疗中心内分泌代谢科,广州510623

出　　处：《实用儿科临床杂志》2012年第20期1557-1560,共4页Journal of Applied Clinical Pediatrics

基　　金：国家科技部973项目(2006BAI05A07);广州市科技局支撑项目(2010J-E231-1)

摘　　要：目的建立尿素酶预处理气相色谱-质谱联用技术(GC-MS)定量检测3-羟基-3-甲基戊二酸的方法,用于临床鉴别诊断3-羟基-3-甲基戊二酸尿症(3-HMG)。方法尿3-羟基-3-甲基戊二酸经尿素酶预处理,残余物进行三甲基硅烷化(BSTFA-TMCS)衍生,衍生化后进行GC-MS测定分析,采用总离子扫描模式进行检测。3-羟基-3-甲基戊二酸存在2种衍生化形式,即2个硅烷化(di-TMS)和3个硅烷化(tri-TMS)形式,分别选择特征离子m/z 175(di-TMS)和363(tri-TMS)为定量离子,内标十七烷酸选择特征离子m/z 327为定量离子。分析该法精密度、稳定性、样本回收率。用该法检测10例代谢性酸中毒、低血糖、未伴酮症、高度怀疑3-HMG的患儿尿样。结果 3-羟基-3-甲基戊二酸定量范围为6.17～1 234.56μmol·L-1,线性方程为Y=0.284 3X-0.003,线性相关系数为0.998 6,低、中、高水平批内变异系数为2.17%、7.79%、5.78%;批间变异系数为4.19%、4.73%、9.24%;回收率为88.25%～108.47%。诊断3-HMG 1例,尿3-羟基-3-甲基戊二酸水平为219.86 mmol·mol-1肌酐。结论尿素酶预处理GC-MS测定尿3-羟基-3-甲基戊二酸具有较高的回收率、精密度和准确性,可为临床诊断3-HMG提供参考依据。Abstract： Objective To establish the urine 3 - hydroxy - 3 - methylglutaric acid determination by gas chromatography - mass spec- trometry （ GC - MS） using urease pretreatment for the diagnosis of 3 - hydroxy - 3 - methylglutaric aciduria （ 3 - HMG）. Methods The uri- nary 3 - hydroxy - 3 - methylglutaric acid was pretreated with urease, added the internal standard （ heptadecanoic acid） , removed protein, dried by nitrogen, derivatized with BSTFA/TMCS （ N, O - bistrimethylsilyltrifluoroacetamide/Trimethylchorosilane） and was converted into a trimethylsilyl derivative. The derivatization was measured and analyzed by GC - MS using total ion currency. The monitored ions at m/z 175 in 3 - hydroxy - 3 - methylglutaric acid di - TMS and at m/z 363 in tri - TMS were used for quantification. The monitored ions at nv/z 327 in in- ternal standard were also used for quantification. The precision,accuracy and recovery were analyzed. Urine samples of 10 patients with meta- bolic acidosis, hypoglycemia without ketosis and highly suspected 3 - HMG were determined. Results The limit of 3 - hydroxy - 3 - methyl- glutaric acid was 6.17 - 1 234.56 ptmol ~ L-t. The equation was Y=0. 284 3X -0. 003. The correlation coefficient was 0. 998 6. The intra - assay coefficient of variation（CV） were 2. 17% , 7.79% , 5.78%. Inter - assay CV were 4. 19% , 4.73% , 9.24%. The recovery were 88.25% - 108.47%. One patient with 3 - HMG was diagnosed, and the level of 3 - hydroxy - 3 - methylglutaric acid was 219.86 mmol~ mol - ~ creatinine. Conclusions Urinary levels of 3 - hydroxy - 3 - methylglutaric acid determination by GC - MS using urease pretreatment shows high recovery,precision and accuracy. It can be used in the diagnosis of 3 - HMG.

关 键 词：3-羟基-3-甲基戊二酸 气相色谱 质谱 有机酸尿症 

分 类 号：R725.8[医药卫生—儿科]