de la Cruz Amorós V, Cortés Castell E, Moya M
Departamento de Pediatría, Universidad Miguel Hernández.
An Esp Pediatr. 1999 Apr;50(4):361-6.
Lysosomal enzyme deficiency results in an accumulation of undegraded material producing structural and functional disorders. Mucopolysaccharidosis is a group of disorders caused by an increase in glycosaminoglycans (GAG) (mucopolysaccharides). Mucopolysaccharidosis, while showing a low incidence, has some clinical manifestations that warrant early diagnosis in order to establish immediate therapeutical action. Although definitive diagnosis is based on quantification of the involved enzymatic activity, it is necessary to have easy to use analytical methods available when there is clinical suspicion of the disease.
This study was made on a population of 632 clinically normal adolescents and children (one month to 18 years of age). In all cases a partial sample of urine to quantify glycosaminoglycans and creatinine expressed as the index GAG/creatinine (mg/mmol) was collected. In a reduced group of 27 people a 24-hour urine sample was collected. Determination of glycosaminoglycan levels was performed with a colorimetric reaction with 1,9-dimethylmethylene blue (DMB). Optimal condition of pH, wavelength and reaction time were established. The values found to be optimal for quantification were defined as pH 4.0, wavelength 528 nm and a reaction time of five minutes. Intra- and inter-assay coefficients of variation were less than 5%.
Validity of the partial urine sample was established by obtaining a linear correlation between the 24 hour urine sample and the partial urine sample with a correlation coefficient of r = 0.89 (p < 0.0001). The results obtained have allowed the establishment of normal values ranging from an index GAG/creatinine of 8.70 mg/mmol in the group aged 1 to 2 years to 1.34 mg/mmol in the group 17 to 18 years of age, with a distribution of intermediate ages adjusted to a logarithmic function. There was a clear difference in the values obtained in urine of patients with clinical suspicion of mucopolysaccharidosis.
Modifications in the spectrophotometric method using DMB as a colorant for quantification of glycosaminoglycans are presented. The feasibility, easy application and sensibility of this method, condition required for its widespread use, are confirmed. Normal reference values from a healthy population have been obtained and shown to be age dependent.
溶酶体酶缺乏会导致未降解物质积累，从而产生结构和功能紊乱。黏多糖贮积症是一组由糖胺聚糖（GAG，即黏多糖）增加引起的疾病。黏多糖贮积症发病率较低，但其某些临床表现需要早期诊断以便立即采取治疗措施。虽然确诊基于对相关酶活性的定量分析，但当临床上怀疑该病时，需要有易于使用的分析方法。
本研究针对632名临床正常的青少年和儿童（年龄从1个月至18岁）展开。所有病例均采集部分尿液样本，以定量糖胺聚糖和肌酐，并将其表示为GAG/肌酐指数（mg/mmol）。在27人的一个缩减组中采集了24小时尿液样本。采用与1,9 - 二甲基亚甲基蓝（DMB）的比色反应测定糖胺聚糖水平。确定了pH、波长和反应时间的最佳条件。确定的用于定量的最佳值为pH 4.0、波长528 nm和反应时间5分钟。批内和批间变异系数均小于5%。
通过获得24小时尿液样本与部分尿液样本之间的线性相关性（相关系数r = 0.89，p < 0.0001），确定了部分尿液样本的有效性。所获得的结果确定了正常参考值范围，1至2岁组的GAG/肌酐指数为8.70 mg/mmol，17至18岁组为1.34 mg/mmol，中间年龄组的分布符合对数函数。临床怀疑患有黏多糖贮积症的患者尿液中所获得的值存在明显差异。
介绍了使用DMB作为糖胺聚糖定量显色剂的分光光度法的改进。证实了该方法的可行性、易于应用和敏感性，这些都是其广泛应用所需的条件。已获得健康人群的正常参考值，并表明其与年龄相关。
