Department of Clinical Laboratory, Second Xiangya Hospital, Central South University, Changsha, PR China.
Clin Chem Lab Med. 2010 Apr;48(4):513-7. doi: 10.1515/CCLM.2010.105.
To provide a more comprehensive clinic marker of tryptophan (TRP) catabolism in patients with systemic lupus erythematosus (SLE), we developed a simple and efficient method that simultaneously measured serum TRP, kynurenine (KYN), and kynurenic acid (KYNA) using high performance liquid chromatography with fluorescence detection (HPLC-FD).
A simple and specific high performance liquid chromatography (HPLC) method was developed for simultaneously quantitative determination of TRP, KYN and KYNA with fluorescence detection (FD) using programmed wavelength and on-column fluorescence derivatization. Thirty patients with SLE and 80 healthy control subjects were analyzed for serum TRP metabolites using the assay we developed. The tryptophan breakdown index (TBI) and neuroprotective ratio (NPR) were calculated.
The retention time of KYN, KYNA and TRP were 8.5 min, 13.7 min and 17.6 min, respectively. The linear range for TRP was 0.245-196 micromol/L, the limit of detection (LOD) was 0.001 micromol/L and average recovery was 103.71%. The linear range for KYN was 0.049-98 v/L, the LOD was 0.0245 micromol/L, and average recovery was 97.45%. The linear range for KYNA was 1.05-2093 nmol/L, the LOD was 0.05 nmol/L, and average recovery was 100.60%. Inter-day and intra-day relative standard deviations (SDs) were <5%. Phenylalanine, tyrosine, 5-hydroxytryptamine and creatinine did not interfere with the method. The results showed great differences in TRP, KYN and KYNA contents and TBI between patients with SLE and healthy controls, but little difference in NPR.
The method is simple, fast, accurate, and meets the requirements for simultaneous determination of TRP, KYN and KYNA in serum.
为了提供系统性红斑狼疮（SLE）患者色氨酸（TRP）分解代谢的更全面临床标志物，我们开发了一种简单有效的方法，使用高效液相色谱-荧光检测（HPLC-FD）同时测量血清 TRP、犬尿氨酸（KYN）和犬尿喹啉酸（KYNA）。
开发了一种简单而特异的高效液相色谱（HPLC）法，使用程序波长和柱上荧光衍生化，荧光检测（FD）同时定量测定 TRP、KYN 和 KYNA。我们开发的测定法分析了 30 例 SLE 患者和 80 例健康对照者的血清 TRP 代谢物。计算色氨酸分解指数（TBI）和神经保护比（NPR）。
KYN、KYNA 和 TRP 的保留时间分别为 8.5、13.7 和 17.6 min。TRP 的线性范围为 0.245-196 μmol/L，检测限（LOD）为 0.001 μmol/L，平均回收率为 103.71%。KYN 的线性范围为 0.049-98 μmol/L，LOD 为 0.0245 μmol/L，平均回收率为 97.45%。KYNA 的线性范围为 1.05-2093 nmol/L，LOD 为 0.05 nmol/L，平均回收率为 100.60%。日内和日间相对标准偏差（SD）均<5%。苯丙氨酸、酪氨酸、5-羟色胺和肌酐不干扰该方法。SLE 患者与健康对照者的 TRP、KYN 和 KYNA 含量及 TBI 差异较大，而 NPR 差异较小。
该方法简单、快速、准确，满足血清中 TRP、KYN 和 KYNA 同时测定的要求。
