Nakahara Y, Kikura R
National Institute of Health Sciences, Tokyo, Japan.
J Chromatogr B Biomed Sci Appl. 1997 Oct 24;700(1-2):83-91. doi: 10.1016/s0378-4347(97)00332-0.
A sensitive GC-MS method was developed for the quantitative analysis of ephedrine (EP), phenylpropanolamine (PPA) and methylephedrine (ME) in animal and human hair. After washing with 0.1% sodium dodecyl sulfate, hair samples (10 mg) were added with deuterated internal standards, extracted by 1-h sonication and over night soaking in 2 ml of 5 M HCl-methanol (1:20) at room temperature. Following evaporation of the liquid phase, the residue was dissolved in phosphate buffer solution (pH 6.0) and purified using a solid-phase extraction procedure with Bond Elut Certify columns. Two types of derivatization were compared - using trifluoroacetic anhydride (TFAA) and pentafluoropropionic anhydride (PFPA) - for discrimination of EP and methamphetamine (MA). Derivatized extracts were analyzed by GC-MS in the EI mode using a capillary column (OV-1 equivalent). From the results comparing three GC-MS conditions, PFP-derivatives separated with a temperature gradient of 20 degrees C/min from 60 degrees C to 280 degrees C gave the best resolution between EP and MA. ME was analyzed as a trimethylsilyl derivative using N,O-bis-trimethylsilyl acetamide at the above GC condition. The assay was linear from 0.5 to 50 ng/mg (r=0.998) and capable of detecting less than 50 pg of derivatized EP, PPA and ME on-column. Intra-assay precision was characterized by C.V. values from 5 to 16% in the concentration range of 1-10 ng/mg hair. The method was used for the quantitative determination of EP, PPA and ME in the hair obtained from three rats with dark brown hair after ten intraperitoneal injections (5 mg/kg/day) of the three drugs and from three male and one female volunteers with black hair after an oral dose of 50 mg/day of EP-HCl for three days. Hair samples were collected by shaving from the back of rats and cutting from the scalp of humans 28 days after the first dose. The incorporation rates of EP, PPA and ME into hair (the ratios of [hair concentration] to [AUC]) obtained from the animal experiment were 0.10, 0.07 and 0.03, respectively, which are a little lower than those (0.14, 0.10 and 0.04) of their desoxy-compounds, MA, amphetamine and dimethylamphetamine. EP was detected at an average of 2.25 ng/mg (n=4) in human scalp hair and at a range of 1-29 ng/mg (n=3) in human beard hair until day 14, but its metabolite (PPA) was at a trace level in the hair of the four subjects. The method was successfully used for detection of ME and EP in the hair of a neonate and its mother who was abusing Bron syrup containing ME during the pregnancy.
建立了一种灵敏的气相色谱 - 质谱联用（GC - MS）方法，用于定量分析动物毛发和人发中的麻黄碱（EP）、苯丙醇胺（PPA）和甲基麻黄碱（ME）。用0.1%十二烷基硫酸钠洗涤后，向毛发样品（10 mg）中加入氘代内标，于室温下用2 ml 5 M盐酸 - 甲醇（1:20）超声处理1小时并浸泡过夜进行提取。液相蒸发后，残渣溶解于磷酸盐缓冲溶液（pH 6.0）中，并用Bond Elut Certify柱通过固相萃取程序进行纯化。比较了两种衍生化方法——使用三氟乙酸酐（TFAA）和五氟丙酸酐（PFPA）——以区分EP和甲基苯丙胺（MA）。衍生化提取物在电子轰击（EI）模式下使用毛细管柱（相当于OV - 1）通过GC - MS进行分析。通过比较三种GC - MS条件的结果，以20℃/min的温度梯度从60℃升至280℃分离得到的PFP衍生物在EP和MA之间具有最佳分离度。在上述GC条件下，ME作为三甲基硅烷基衍生物使用N,O - 双（三甲基硅基）乙酰胺进行分析。该测定法在0.5至50 ng/mg范围内呈线性（r = 0.998），并且能够在柱上检测到低于50 pg的衍生化EP、PPA和ME。批内精密度在毛发浓度范围为1 - 10 ng/mg时，变异系数（C.V.）值为5%至16%。该方法用于定量测定三只深棕色毛发大鼠在腹腔注射（5 mg/kg/天）这三种药物十次后毛发中的EP、PPA和ME，以及三名男性和一名女性志愿者在口服50 mg/天盐酸麻黄碱三天后黑发中的上述物质。首次给药28天后，通过剃除大鼠背部毛发和剪取人类头皮毛发收集毛发样品。从动物实验获得的EP、PPA和ME在毛发中的掺入率（[毛发浓度]与[AUC]的比值）分别为0.10、0.07和0.03，略低于它们的脱氧化合物MA、苯丙胺和二甲基苯丙胺的掺入率（0.14、0.10和0.04）。在人类头皮毛发中，平均检测到EP为2.25 ng/mg（n = 4），在人类胡须毛发中，直到第14天检测到的范围为1 - 29 ng/mg（n = 3），但其代谢物（PPA）在四名受试者的毛发中处于痕量水平。该方法成功用于检测一名新生儿及其母亲毛发中的ME和EP，其母亲在怀孕期间滥用含ME的布朗糖浆。
