Machnik M, Geyer H, Horning S, Breidbach A, Delahaut P, Schänzer W
Institut für Biochemie, Deutsche Sporthochschule Köln, Cologne, Germany.
J Chromatogr B Biomed Sci Appl. 1999 Feb 19;723(1-2):147-55. doi: 10.1016/s0378-4347(98)00527-1.
A method for the detection of clenbuterol in human scalp hair by gas chromatography-high-resolution mass spectrometry (GC-HRMS) is described. The sample preparation involved chemical digestion of the protein structure, which was achieved by incubating the hair with 1 M KOH at 70 degrees C. A single extraction step with tert.-butyl methyl ether provided approximately 90% of the analyte, which was dried and derivatized with N-methyl-N-trimethylsilyltrifluoroacetamide (MSTFA) to yield clenbuterol N,O-bis-trimethylsilyl (TMS). Hair was collected from four pregnant women who were therapeutically treated with Spiropent (clenbuterol-HCl) and from the infant of one female patient. Hair samples were taken during the application time and two to six months after completion of clenbuterol administration. The detection limit of the method was approximately 4 ng clenbuterol/g hair when 25 mg hair material were processed and 2 ng/g for 50 mg hair samples (corresponds to 4 pg per injection). The method allows clenbuterol to be measured retrospectively for up to at least six months. The levels of clenbuterol determined in hair ranged from 2 to 236 ng/g. No clenbuterol was found in the hair of the infant, which was taken five and a half months after delivery. To improve sample preparation, an additional purification step via immuno affinity chromatography (IAC) was integrated. The IAC purified extracts showed reduced biological background interference and an improved limit of detection (0.8 ng/g).
描述了一种通过气相色谱-高分辨率质谱法（GC-HRMS）检测人头皮毛发中克伦特罗的方法。样品制备包括对蛋白质结构进行化学消解，这通过在70℃下用1M氢氧化钾孵育毛发来实现。用叔丁基甲醚进行单次萃取步骤可提供约90%的分析物，将其干燥并用N-甲基-N-三甲基硅烷基三氟乙酰胺（MSTFA）衍生化以生成克伦特罗N,O-双三甲基硅烷基（TMS）。从四名接受 Spiropent（盐酸克伦特罗）治疗的孕妇以及一名女性患者的婴儿处采集毛发。在用药期间以及克伦特罗给药结束后两到六个月采集毛发样本。当处理25mg毛发材料时，该方法的检测限约为4ng克伦特罗/克毛发，对于50mg毛发样本则为2ng/g（相当于每次进样4pg）。该方法允许对克伦特罗进行至少长达六个月的回顾性测量。毛发中测定的克伦特罗水平范围为2至236ng/g。在分娩后五个半月采集的婴儿毛发中未发现克伦特罗。为了改进样品制备，引入了通过免疫亲和色谱（IAC）的额外纯化步骤。IAC纯化的提取物显示出降低的生物背景干扰和改进的检测限（0.8ng/g）。
