A highly sensitive method for determining urine homogentisic acid (HGA) is required to provide adequate diagnosis and therapy for alkaptonuria in early stages. In this study, we developed a highly sensitive high-performance liquid chromatography with electrochemical detection (HPLC-ECD) for determining HGA in urine. In order to obtain a chromatogram of HGA by HPLC-ECD, an oxidation current was monitored at +0.5 V vs. Ag/AgCl. The peak heights of HGA showed linearity (r = 0.999) ranging from 4.2 ng/mL to 168 ng/mL, and the detection limit was 1.2 ng/mL (signal-to-noise ratio, S/N = 3). In recovery tests using human control urine spiked with an HGA standard, the recoveries of HGA were more than 93.2 %, and the relative standard deviations (n = 6) were less than 1.9 %. As an in vivo application using male Wistar rats, the level of urine HGA, which was metabolized from tyrosine in tyrosine-enriched food, was determined by this HPLC-ECD method. The determination of HGA in urine by this HPLC-ECD method requires only 0.1 mL of a rat urine specimen and simple sample preparation consisting of dilution and filtration.

中文翻译：

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高效液相色谱-电化学检测法测定高纯酸及其在饲喂富含酪氨酸食物的大鼠尿液中的应用。

需要一种高度敏感的测定尿高尿酸（HGA）的方法，以在早期阶段对链球菌尿症提供足够的诊断和治疗。在这项研究中，我们开发了一种具有电化学检测（HPLC-ECD）的高灵敏度高效液相色谱法，用于测定尿液中的HGA。为了通过HPLC-ECD获得HGA的色谱图，在+0.5 V对Ag / AgCl的条件下监测氧化电流。HGA的峰高呈线性（r = 0.999），范围为4.2 ng / mL至168 ng / mL，检测极限为1.2 ng / mL（信噪比，S / N = 3）。在使用掺有HGA标准品的人体对照尿液进行的恢复测试中，HGA的回收率高于93.2％，相对标准偏差（n = 6）小于1.9％。作为使用雄性Wistar大鼠的体内应用，尿液HGA的水平 通过HPLC-ECD方法测定富含酪氨酸的食物中酪氨酸代谢产物的含量。通过这种HPLC-ECD方法测定尿液中的HGA，只需要0.1 mL的大鼠尿液样本和简单的样品制备，包括稀释和过滤。