Höld K M, Hubbard D L, Wilkins D G, Rollins D E
Center for Human Toxicology, Department of Pharmacology and Toxicology, University of Utah, Salt Lake City 84112-9457, USA.
J Anal Toxicol. 1998 Oct;22(6):414-7. doi: 10.1093/jat/22.6.414.
Hair pigmentation is a critical factor in the interpretation of the concentration of certain compounds and their metabolites incorporated into hair. Melanin is responsible for the pigmentation. The color and the melanin content of human hair samples differs over a wide range. Once deposited into hair, drug may remain detectable for a period of months to years. However, if drug disposition into hair is influenced by those properties attributed to hair color, then certain persons may test positive more frequently than other persons. Removal of the melanin from hair digests prior to drug analysis may reduce the effect of melanin on the total drug concentration by excluding the drug bound to the pigment. In this study, the effect of melanin removal by centrifugation of hair digests on cocaine concentrations was investigated. Two sets of hair samples from five cocaine users were analyzed for cocaine and metabolites. A solution consisting of 10 mL of 0.5M Tris buffer (pH 6.4) to which is added 60 mg D,L-dithiothreitol, 200 mg SDS, and 200 U Proteinase K, was used to digest the hair. Two milliliters of this solution was added to 20 mg of hair and incubated at 37 degrees in a shaking water bath (90 oscillations/min) overnight. The samples were removed from the water bath and mixed. One set was centrifuged at 2000 rpm and divided into supernatant and melanin pellet. The other set was not centrifuged. Internal standards were added to all tubes. The samples were further extracted, derivatized, and analyzed by gas chromatography-mass spectrometry. A mean of 8.8% (standard deviation [SD] 7.0%) of the total cocaine concentration (supernatant and pellet) was left behind in the pellet. The same experiment was repeated except that the melanin pellet was redigested with 0.1 N HCl. After redigestion of the melanin pellet, the mean cocaine concentration in the pellet was 3.8% +/- 4.0% (mean +/- SD) of the total cocaine concentration in hair. These data demonstrate that removal of melanin from hair digests by centrifugation does not eliminate hair color bias when interpreting cocaine concentrations.
头发色素沉着是解释某些化合物及其代谢产物在头发中的浓度的关键因素。黑色素负责色素沉着。人类头发样本的颜色和黑色素含量差异很大。一旦药物沉积在头发中，可能在数月至数年的时间内都能被检测到。然而，如果药物在头发中的沉积受头发颜色相关特性的影响，那么某些人检测呈阳性的频率可能比其他人更高。在药物分析之前从头发消化物中去除黑色素，通过排除与色素结合的药物，可能会降低黑色素对总药物浓度的影响。在本研究中，研究了通过离心头发消化物去除黑色素对可卡因浓度的影响。对来自五名可卡因使用者的两组头发样本进行了可卡因及其代谢产物分析。使用一种由10 mL 0.5M Tris缓冲液（pH 6.4）组成的溶液，其中加入了60 mg D,L-二硫苏糖醇、200 mg十二烷基硫酸钠和200 U蛋白酶K来消化头发。将两毫升该溶液加入20 mg头发中，并在37度的振荡水浴（90次振荡/分钟）中孵育过夜。将样本从水浴中取出并混合。一组以2000 rpm离心，分为上清液和黑色素沉淀。另一组未进行离心。向所有试管中加入内标。样本进一步提取、衍生化，并通过气相色谱-质谱联用仪进行分析。黑色素沉淀中残留的可卡因总浓度（上清液和沉淀）平均为8.8%（标准差[SD] 为7.0%）。除了用0.1 N盐酸对黑色素沉淀进行再次消化外，重复相同的实验。在对黑色素沉淀进行再次消化后，沉淀中可卡因的平均浓度为头发中可卡因总浓度的3.8%±4.0%（平均值±标准差）。这些数据表明，在解释可卡因浓度时，通过离心从头发消化物中去除黑色素并不能消除头发颜色偏差。
