Maurer H H, Schmitt C J, Weber A A, Kraemer T
Department of Toxicology, Institute of Pharmacology and Toxicology, University of Saarland, Homburg (Saar), Germany.
J Chromatogr B Biomed Sci Appl. 2000 Oct 1;748(1):125-35. doi: 10.1016/s0378-4347(00)00270-x.
Specific detection of amanitins in body fluids is necessary for an early diagnosis of an intoxication with amanita mushrooms. In this paper, a liquid chromatographic-mass spectrometric assay after immunoaffinity extraction (IAE-LC-MS) is described for the determination of alpha- and beta-amanitin in urine. The method has been validated according to the criteria established by the Journal of Chromatography B. The assay was found to be selective. The calibration curves for alpha- and beta-amanitin were linear from 5 to 75 ng/ml. Intra- and inter-day accuracy and precision were inside the required limits. Amatoxins in frozen urine samples or immunoaffinity extracts were stable for more than 6 months, and the IAE columns could be used more than fifty times without remarkable loss in performance. LOD for alpha- and beta-amanitin was 2.5 ng/ml and LOQ for both was 5.0 ng/ml. The absolute recoveries of alpha- and beta-amanitin were 63% and 58% for the low quality control and 61% and 57% for the high quality control. The absolute recovery for the internal standard gamma-amanitin methyl ether at 25 ng/ml was 60%. The analysis of 5 authentic urine samples from patients intoxicated by amanita mushrooms showed a good correlation between the results measured by radioimmunoassay and the IAE-LC-MS assay. A partial validation showed that the assay was also suitable for plasma analysis.
对体液中的鹅膏毒素进行特异性检测对于早期诊断鹅膏菌中毒至关重要。本文描述了一种免疫亲和萃取后液相色谱 - 质谱分析法（IAE - LC - MS），用于测定尿液中的α - 鹅膏毒素和β - 鹅膏毒素。该方法已根据《色谱杂志B》制定的标准进行了验证。结果表明该分析方法具有选择性。α - 鹅膏毒素和β - 鹅膏毒素的校准曲线在5至75 ng/ml范围内呈线性。日内和日间的准确度和精密度均在规定范围内。冷冻尿液样本或免疫亲和提取物中的鹅膏毒素在6个月以上保持稳定，并且免疫亲和萃取柱可使用五十多次而性能无明显损失。α - 鹅膏毒素和β - 鹅膏毒素的检测限为2.5 ng/ml，两者的定量限均为5.0 ng/ml。低质量控制时α - 鹅膏毒素和β - 鹅膏毒素的绝对回收率分别为63%和58%，高质量控制时分别为61%和57%。25 ng/ml内标γ - 鹅膏毒素甲醚的绝对回收率为60%。对5例鹅膏菌中毒患者的真实尿液样本分析表明，放射免疫分析法与IAE - LC - MS分析法的检测结果具有良好的相关性。部分验证表明该分析方法也适用于血浆分析。
