Department of Biology, University of Modena and Reggio Emilia, Modena, Italy.
Clin Chem Lab Med. 2010 Aug;48(8):1133-9. doi: 10.1515/CCLM.2010.217.
Glycosaminoglycans (GAGs) are natural complex polysaccharides that are important in several pathological processes. Urinary GAGs have long been investigated for their possible modifications in many pathological conditions. In some cases, they have been found to have diagnostic utility. As a result, the measurement of GAGs in urine is gradually gaining importance. Cetylpyridinium chloride (CPC) and cetyltrimethylammonium bromide (CETAB) are generally used to extract urinary GAGs prior to analysis. In this study, we evaluated the extraction of human urinary GAGs using CPC in comparison with CETAB.
Extracted urinary GAGs were qualitatively and quantitatively analyzed by agarose-gel electrophoresis in the presence of sequential staining and densitometric scanning. This procedure was able to give more reproducible and reliable results for urinary GAGs, and high performance liquid chromatography (HPLC) was used for the evaluation of chondroitin sulfate (CS) disaccharides.
Differences were observed between CPC and CETAB extract protocols. The absolute amount of CS evaluated by electrophoresis was found to be similar for the two protocols. However, the heparan sulfate (HS) concentration was calculated to be approximately 3.3 times greater for CPC than CETAB. When calculated in relative percentage, 33.6% HS was determined for CPC and 10.0% for CETAB. These results show a quantitative expression for greater recovery of HS by using CPC protocol than CETAB. No significant differences were found between CS quantified by agarose-gel and HPLC. In addition, no differences were observed for the CS disaccharide composition purified by using CPC or CETAB, and quite similar results were observed for 4s/6s disaccharide ratios and charge density values.
Extract procedures for urinary GAGs using CPC or CETAB are able to recover similar amounts of CS quantified by agarose-gel electrophoresis and HPLC. However, CPC yields greater recovery of HS than the CETAB protocol; an increase of approximately 3.3 times as evaluated by electrophoresis. This different capacity of HS extraction between CPC and CETAB should be considered when urinary GAGs of subjects affected by various diseases and related pharmacological treatments are considered, or meta-analysis is performed comparing various studies and trials performed under different experimental conditions.
糖胺聚糖（GAGs）是天然的复杂多糖，在多种病理过程中具有重要作用。长期以来，人们一直在研究尿液中的 GAGs，以了解其在许多病理情况下的可能变化。在某些情况下，它们已被发现具有诊断效用。因此，尿液中 GAG 的测量逐渐变得重要。氯化十六烷基吡啶（CPC）和溴化十六烷基三甲铵（CETAB）通常用于在分析前提取尿液中的 GAG。在这项研究中，我们评估了 CPC 与 CETAB 提取人尿液 GAG 的效果。
使用琼脂糖凝胶电泳在连续染色和密度扫描的情况下对提取的尿液 GAG 进行定性和定量分析。该程序可提供更具重现性和可靠性的尿液 GAG 结果，并用高效液相色谱法（HPLC）评估硫酸软骨素（CS）二糖。
观察到 CPC 和 CETAB 提取方案之间存在差异。电泳评估的 CS 绝对量两种方案相似。然而，用 CPC 计算的肝素硫酸盐（HS）浓度比 CETAB 高约 3.3 倍。按相对百分比计算，CPC 为 33.6% HS，CETAB 为 10.0% HS。这些结果表明，与 CETAB 相比，使用 CPC 方案可更有效地回收 HS。用琼脂糖凝胶和 HPLC 定量的 CS 无显著差异。此外，用 CPC 或 CETAB 纯化的 CS 二糖组成无差异，4s/6s 二糖比和电荷密度值也有非常相似的结果。
使用 CPC 或 CETAB 提取尿液 GAG 的程序能够回收与琼脂糖凝胶电泳和 HPLC 定量的 CS 相似的量。然而，与 CETAB 方案相比，CPC 方案回收的 HS 量更大，电泳评估增加约 3.3 倍。当考虑受各种疾病和相关药物治疗影响的受试者的尿液 GAG 时，或在不同实验条件下进行的各种研究和试验的荟萃分析时，应考虑 CPC 和 CETAB 之间提取 HS 的这种不同能力。
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