Miniaturized sample preparation method for determination of amphetamines in urine

A simple and miniaturized sample preparation method for determination of amphetamines in urine was developed using on-column derivatization and gas chromatography-mass spectrometry (GC-MS). Urine was directly applied to the extraction column that was pre-packed with Extrelut and sodium carbonate. Amphetamine (AP) and methamphetamine (MA) in urine were adsorbed on the surface of Extrelut. AP and MA were then converted to a free base and derivatized to N-propoxycarbonyl derivatives using propylchloroformate on the column. Pentadeuterated MA was used as an internal standard. The recoveries of AP and MA from urine were 100 and 102%, respectively. The calibration curves showed linearity in the range of 0.50-50 microg/mL for AP and MA in urine. When urine samples containing two different concentrations (0.50 and 5.0 microg/mL) of AP and MA were determined, the intra-day and inter-day coefficients of variation were 1.4-7.7%. This method was applied to 14 medico-legal cases of MA intoxication. The results were compared and a good agreement was obtained with a HPLC method.     

Simultaneous analysis of six amphetamines and analogues in hair, blood and urine by LC-ESI-MS/MS. Application to the determination of MDMA after low ecstasy intake

A rapid and sensitive method using LC-MS/MS triple stage quadrupole for the determination of traces of amphetamine (AP), methamphetamine (MA), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA, "ecstasy"), 3,4-methylenedioxyethamphetamine (MDEA), and N-methyl-1-(3,4-methylenedioxyphenyl)-2-butanamine (MBDB) in hair, blood and urine has been developed and validated. Chromatography was carried out on an Uptisphere ODB C(18) 5 microm, 2.1 mm x 150 mm column (Interchim, France) with a gradient of acetonitrile and formate 2 mM pH 3.0 buffer. Urine and blood were extracted with Toxitube A (Varian, France). Segmented scalp hair was treated by incubation 15 min at 80 degrees C in NaOH 1M before liquid-liquid extraction with hexane/ethyl acetate (2/1, v/v). The limits of quantification (LOQ) in blood and urine were at 0.1 ng/mL for all analytes. In hair, LOQ was <5 pg/mg for MA, MDMA, MDEA and MBDB, at 14.7 pg/mg for AP and 15.7 pg/mg for MDA. Calibration curves were linear in the range 0.1-50 ng/mL in blood and urine; in the range 5-500 pg/mg for MA, MDMA, MDEA and MBDB, and 20-500 pg/mg for AP and MDA. Inter-day precisions were <13% for all analytes in all matrices. Accuracy was <20% in blood and urine at 1 and 50 ng/mL and <10% in hair at 20 and 250 pg/mg. This method was applied to the determination of MDMA in a forensic case of single administration of ecstasy to a 16-year-old female without her knowledge during a party. She suffered from hyperactivity, sweating and agitation. A first sample of urine was collected a few hours after (T+12h) and tested positive to amphetamines by immunoassay by a clinical laboratory. Blood and urine were sampled for forensic purposes at day 8 (D+8) and scalp hair at day 60 (D+60). No MDMA was detected in blood, but urine and hair were tested positive, respectively at 0.42 ng/mL and at 22 pg/mg in hair only in the segment corresponding to the period of the offence, while no MDA was detectable. This method allows the detection of MDMA up to 8 days in urine after single intake.     

Rapid analysis of amphetamines in blood using head space-solid phase microextraction and selected ion monitoring

A simple and rapid method for analysis of methamphetamine (MA) and amphetamine (AP) in blood was developed using head space-solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry/electron impact ionization-selected ion monitoring (GC-MS/EI-SIM). A vial containing a blood sample, sodium hydroxide, and pentadeuterated methamphetamine as an internal standard, was heated at 80 °C for 20 min. The extraction fiber of the SPME was exposed for 5 min in the head space of the vial. First, heptafluorobutyric anhydride solution was injected into the injection port of the GC-MS to make heptafluorobutyramide (HFB) derivatives of amphetamines, and compounds absorbed on the fiber were detached by exposing the fiber in the injection port. Straight calibration curves of MA and AP were obtained from 0.01 to 2 μg/g in blood, respectively. No interfering substances were found, and the time for analysis was 30 min for one sample.     

The study of metabolite-to-parent drug ratios of methamphetamine and methylenedioxymethamphetamine in hair

The metabolite-to-parent drug ratios were determined in the hair of 2444 methamphetamine (MA) abusers who had produced MA-positive hair results from 2001 to May 2005 and in the hair of 53 ecstasy abusers who had produced positive methylenedioxymethamphetamine (MDMA) hair results from 2002 to May 2005. For the hair analyses, hair strands were washed, cut into small pieces and extracted for 20 h in 1 mL methanol containing 1% HCl. Drugs in the extract were determined by gas chromatography-mass spectrometry (GC-MS) using selective ion monitoring after derivatization with trifluoroacetic anhydride. The six range groups were divided as follows on the basis of MA concentrations in hair (n = 2389): 0.5-5 ng/mg (n = 950), 5-10 ng/mg (n = 582), 10-20 ng/mg (n = 503), 20-30 ng/mg (n = 160), 30-40 ng/mg (n = 80), more than 40 ng/mg (n = 114) to assess the correlations between MA concentrations and metabolite-to-parent drug ratios. In groups of higher MA concentrations, lower ratios of AP/MA were found, and there was a statistically significant difference among six range groups. Comparisons of age groups (tens, twenties, thirties, forties, fifties, and sixties) and male and female subjects for the ratios of AP/MA showed a statistically significant difference. The detection of metabolites and the parent drug with reasonable ratios was found to be a useful indicator for distinguishing internal drug incorporation from external contamination. In our study, MA users can produce 0.4-116% (mean = 9%) of amphetamine (AP) concentrations in hair, and ecstasy users 1-110% (mean = 12%) of methylenedioxyamphetamine (MDA) in appropriately washed hair samples.     

Preparation and application of a fortified hair reference material for the determination of methamphetamine and amphetamine

Quality assurance is one of the major issues in forensic analytical laboratories, where the need for a reference material (RM) has rapidly increased. RMs are very useful for method development and validation, internal quality control or proficiency tests. In the present study, we prepared a RM using drug-free hair for the determination of methamphetamine (MA) and its main metabolite, amphetamine (AP) according to the recommendations of ISO Guide 35. The concentrations of MA and AP were determined using two extraction methods, agitation with 1% HCl in methanol at 38 degrees C and ultrasonication with methanol/5M HCl (20:1), followed by gas chromatography/mass spectrometry (GC/MS) after derivatization with trifluoroacetic anhydride (TFAA). The assignment of values was conducted through the homogeneity study and characterization of the material. Furthermore, an internal proficiency test was performed with the prepared RM, of which the results were compared with those of the authentic hair RM prepared in our previous study. As a result, a hair RM containing MA and AP was prepared at the level of 4.86+/-0.69 ng/mg and 4.63+/-0.44 ng/mg, respectively. Most participants showed satisfactory performances in the internal proficiency test with the both RMs. The hair RM prepared in this study demonstrated its suitability for quality assurance in forensic laboratories.     

芬氟拉明和苯丙胺类兴奋剂的固相微萃取

用固相微萃取技术从血中提取芬氟拉明、苯丙胺和甲基苯丙胺。在 70℃条件下用 10 0 μm聚二甲基硅氧烷萃取头吸附 15min。重氢甲基苯丙胺作内标 ,采用柱前衍生化的进样方式 ,气质联用仪测定。选择离子m /z2 6 8(芬氟拉明 )、m/z2 40 (苯丙胺 )、m /z2 5 4(甲基苯丙胺 )和m/z2 5 8(重氢甲基苯丙胺 ,内标 )的峰面积比定量。血中检测浓度可达 0 0 1～ 0 0 3μg/g。通过解剖例中芬氟拉明的实际测定 ,证明这是一个从血液中提取分析苯丙胺类衍生物的快速准确的方法     

Highly sensitive analysis of methamphetamine and amphetamine in human whole blood using headspace solid-phase microextraction and gas chromatography-mass spectrometry

A simple and highly sensitive method for analysis of derivatized methamphetamine (MA) and amphetamine (AM) in whole blood was developed using headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry electron impact ionization selected ion monitoring (GC-MS-EI-SIM). A whole blood sample, deuterated-MA (d(5)-MA), as an internal standard (IS), tri-n-propylamine and pentafluorobenzyl bromide were placed in a vial. The vial was heated and stirred at 90 degrees C for 30min. Then the extraction fiber of the SPME was exposed at 90 degrees C for 30min in the headspace of the vial while being stirred. The derivatives adsorbed on the fiber were desorbed by exposing the fiber in the injection port of a GC-MS. The calibration curves showed linearity in the range of 0.5-1000ng/g for both MA and AM. The time for analysis was about 80min per sample. In addition, this proposed method was applied to two autopsy cases where MA ingestion was suspected. In one case, MA and AM concentrations in the mixed left and right heart blood were 165 and 36.9ng/g, respectively. In the other case, MA and AM concentrations were 1.79 and 0.119 microg/g in the left heart blood, and 1.27 and 0.074 microg/g in the right heart blood, respectively.     

Simultaneous analysis of six amphetamines and analogues in hair, blood and urine by LC-ESI-MS/MS: Application to the determination of MDMA after low Ecstasy intake

A rapid and sensitive method using LC-MS/MS triple stage quadrupole for the determination of traces of amphetamine (AP), methamphetamine (MA), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA, “ecstasy”), 3,4-methylenedioxyethamphetamine (MDEA), and N-methyl-1-(3,4-methylenedioxyphenyl)-2-butanamine (MBDB) in hair, blood and urine has been developed and validated. Chromatography was carried out on an Uptisphere ODB C₁₈ 5 μm, 2.1 mm × 150 mm column (Interchim, France) with a gradient of acetonitrile and formate 2 mM pH 3.0 buffer. Urine and blood were extracted with Toxitube A^® (Varian, France). Segmented scalp hair was treated by incubation 15 min at 80 °C in NaOH 1 M before liquid–liquid extraction with hexane/ethyl acetate (2/1, v/v). The limits of quantification (LOQ) in blood and urine were at 0.1 ng/mL for all analytes. In hair, LOQ was <5 pg/mg for MA, MDMA, MDEA and MBDB, at 14.7 pg/mg for AP and 15.7 pg/mg for MDA. Calibration curves were linear in the range 0.1–50 ng/mL in blood and urine; in the range 5–500 pg/mg for MA, MDMA, MDEA and MBDB, and 20–500 pg/mg for AP and MDA. Inter-day precisions were <13% for all analytes in all matrices. Accuracy was <20% in blood and urine at 1 and 50 ng/mL and <10% in hair at 20 and 250 pg/mg. This method was applied to the determination of MDMA in a forensic case of single administration of ecstasy to a 16-year-old female without her knowledge during a party. She suffered from hyperactivity, sweating and agitation. A first sample of urine was collected a few hours after (T + 12 h) and tested positive to amphetamines by immunoassay by a clinical laboratory. Blood and urine were sampled for forensic purposes at day 8 (D + 8) and scalp hair at day 60 (D + 60). No MDMA was detected in blood, but urine and hair were tested positive, respectively at 0.42 ng/mL and at 22 pg/mg in hair only in the segment corresponding to the period of the offence, while no MDA was detectable. This method allows the detection of MDMA up to 8 days in urine after single intake.     

Correlation of methamphetamine results and concentrations between head, axillary, and pubic hair

This study was designed to compare the qualitative results and concentrations of methamphetamine (MA) and its metabolite amphetamine (AP) in head hair and hair collected from different parts of the body (axillae and pubis). Hair from subjects (N = 14) suspected MA users was simultaneously collected. Hair preparation involved washing step, fine cutting, overnight extraction, derivatization by the trifluoroacetic anhydride, and gas chromatography/mass spectrometry (GC/MS) using selective ion monitoring. In this study, we found a good correlation of the qualitative results for MA between head hair and hair on other parts of the body, but there were some differences in concentrations of MA and AP. Namely, the concentrations of MA and AP were higher in axillary and pubic hair than in head hair.     

Distribution of methamphetamine and amphetamine in drug abusers’ head hair

In order to aid the interpretation of hair results from methamphetamine (MA) abusers the MA and amphetamine (AP) concentrations in 2070 hair samples were statistically evaluated. The MA and AP concentrations in hair were put into three groups arbitrarily representing low, medium and high ranges and the metabolite-to-parent drug ratios of each group were examined. The concentration ranges proposed here were also applied to the interpretation of five authentic cases. The low, medium and high ranges of MA were 0.5–4.2, 4.2–24.5 and 24.5–608.9 ng/mg and those of AP were 0.1–0.4, 0.4–1.7 and 1.7–41.4 ng/mg. The AP-to-MA ratios showed large variation but a tendency that it decreased as the MA ranges increased. This evaluation was very useful to presume the severity of individuals’ MA abuse and to provide law enforcement agencies more understandable information. It could also facilitate the court's decision regarding specific circumstances surrounding the drug-related crimes.     

The prevalence of drugs in injured drivers

In mid 2009 Victoria introduced compulsory drug testing of blood taken from all injured drivers taken to hospital. Δ(9)-Tetrahydrocannabinol (THC), methylamphetamine (MA) and 3,4-methylenedioxy-methylamphetamine (MDMA) are prohibited and if drivers are positive to any amount an automatic penalty is enforced. Laboratory screens were conducted on preserved blood using ELISA testing for cannabis metabolite and methylamphetamines and a fully validated LC-MS/MS method for 105 drugs including THC, amphetamines, opioids, benzodiazepines, antidepressants and antipsychotics and a number of other psychoactive substances using a minimum of two transitions per drug. Conventional GC-testing for ethanol was used to screen and quantify the presence of alcohol. 1714 drivers were tested and showed alcohol in 29% (≥ 0.01 g/100mL) and drugs in 35%. The positive rate for the three drugs prohibited by legislation was 12.5%. The prevalence of THC, MA and MDMA was 9.8%, 3.1%, and 0.8%, respectively. The range of THC concentrations in blood was 2-42 ng/mL (median 7) of which 70% had a concentration of 10 ng/mL or higher. The range of concentrations for MA and MDMA was 0.02-0.4 and 0.03-0.3mg/L (median for both drugs was 0.05 mg/L). Drugs of any type were detected in 35% of cases. The other drugs were largely prescribed drugs such as the antidepressants (9.3%) and benzodiazepines (8.9%). Neither 6-acetylmorphine nor cocaine (or benzoylecgonine) was detected in these cases.     

Detection of phentermine in hair samples from drug suspects

Phentermine (PT) has been widely used as an anti-obesity drug. This drug has to be used with caution due to its close resemblance with amphetamines in its structure and toxicity profile. Recently, PT is in distribution by illegal modes and is found to be available through sources such as the internet, thus their misuse and/or abuse is threatening to be a serious social issue. In the present study, 32 cases of drug suspects were observed for PT abuse, detected using hair samples for drug analysis. PT and other amphetamines, such as methamphetamine (MA), amphetamine (AP), 3,4-methylenedioxyamphetamine (MDMA) and 3,4-methylenedioxyamphetamine (MDA), were extracted using 1% HCl in methanol for 20 h at 38°C. The extracts were derivatized with trifluoroacetic anhydride (TFAA) and analyzed using gas chromatography/mass spectrometry (GC/MS). Among the 32 cases of PT abuse, MA and its main metabolite, AP were identified in seven cases and MDMA and its main metabolite, MDA were detected in two other cases.     

A rapid and convenient LC/MS method for routine identification of methamphetamine/dimethylamphetamine and their metabolites in urine

A rapid and sensitive LC/MS method was developed for the simultaneous analysis of N,N-dimethylamphetamine (DMA), N,N-dimethylamphetamine N-oxide (DMANO), methylamphetamine (MA) and amphetamine (A) in urine samples. Employing an Alltech C18 column for solid phase extraction followed by LC/MS analysis using an Alltech Platinum EPS C18 column with a mixture of ammonium formate (0.01 M, pH 3) and acetonitrile (77:23, v/v) as mobile phase at a flow rate of 0.2 mL/min, simultaneous identification and quantitation of A, MA, DMA and DMANO in urine can be achieved using a 5-min chromatographic run. The calibration ranges were 0.10-3.0 micro g/mL for DMANO, 0.05-3.0 micro g/mL for DMA and 0.05-5.0 micro g/mL for both MA and A. The intra-, inter-day precision and accuracy for all analytes, spiked at three different concentrations in quality control samples, were in the ranges of 1.7-8.6, 4.1-10.0, -11.6 to 12.9%, respectively. The newly developed method was applied to the analysis of urine samples obtained from 118 suspected MA/DMA abusers, with the presence of MA confirmed in their urine samples under the drug-use surveillance program. Of these 118 samples, 43 were found to contain DMANO and 11 with both DMANO and DMA.     

Long-term stability of various drugs and metabolites in urine, and preventive measures against their decomposition with special attention to filtration sterilization

The long-term stability of drugs and metabolites of forensic interest in urine, and preventive measures against their decomposition have been investigated, with special attention to filtration sterilization. An aseptic urine collection kit, which was recently developed based on filtration sterilization, was utilized for the aseptic collection and storage of urine samples. For evaluating preservation measures, methamphetamine (MA), amphetamine (AP), nitrazepam (NZ), estazolam (EZ), 7-aminoflunitrazepam (7AF), cocaine (COC), and 6-acetylmorphine (6AM) were spiked into urine at 500 ng/mL each, and were monitored for 6 months at 25, 4, and -20 degrees C, after the addition of NaN(3) and/or filtration sterilization using the aseptic collection kit. In severely contaminated urine with bacteria, there were significant losses of 7AF and NZ, and slight decomposition of MA and AP at 25 degrees C. However, such degradation was successfully suppressed by the use of the kit, though the use of the kit and NaN(3) were preferred for 7AF. The kit was also effective in preventing the hydrolyses of COC and 6AM, while it was suggested that the common preservative NaN(3) can accelerate the hydrolysis of such ester-type drugs and metabolites.     

Methamphetamine body packer: acute poisoning death due to massive leaking of methamphetamine

We encountered three methamphetamine (MA) body packers presenting simultaneously, one of whom died. Three Nigerian men (39, 35, and 37 years old) who attempted to smuggle were found to contain 35 (498 g), 21 (292 g), and 5 packages (73 g) of methamphetamine hydrochloride (MA-HCl) in their stomachs, respectively. Packages were wrapped with plastic film and Scotch tape. The 39-year-old man died with acute poisoning from c. 20 g of MA-HCl that had leaked from the packages into the stomach. His plasma MA concentration was 8.6 microg/mL when he was hospitalized (17 h before his death). Autopsy findings showed extreme pulmonary congestion and edema as well as moderate hepatic edema and several petechiae. Quantitative analysis was performed by gas chromatography/mass spectrometry. Extremely high concentrations of MA and its metabolite amphetamine (AP) were found in cardiac blood (63.5 microg/mL and 1.2 microg/mL), urine (4,518 microg/mL and 72.4 microg/mL), gastric contents (8,490 microg/mL and 16.9 microg/mL), and in all other autopsy samples. These high concentrations confirmed that the cause of death was acute MA poisoning. Furthermore, impurity-profiling analysis of the seized MA revealed that the MA smuggled by the three suspects originated from the same batch.     

SPE/UPLC法检测血中吗啡、苯丙胺类及氯胺酮

目的建立SPE/UPLC方法在同一条件下同时检测血中吗啡、苯丙胺类及氯胺酮。方法采用SCX 3cc（60mg）固相萃取柱萃取血中吗啡、MA、MDMA、MDA及氯胺酮,用超高效液相色谱（UPLC）-二极管阵列检测器（PDA）检测,结合保留时间和紫外光谱进行定性、定量分析,对实验各环节进行优化,并进行实际案例检测。结果吗啡、MA、MDMA、MDA、氯胺酮的固相萃取提取回收率分别为81.4%±2.51%、88.2%±2.48%、91.8%±2.03%、93.8%±1.46%、74.8%±2.27%,峰面积和质量浓度的线性关系良好（r〉0.999）,线性范围分别为0.08～100μg/mL、0.4～100μg/mL、0.2～75μg/mL、0.3～75μg/mL、0.4～100μg/mL,检出限分别为30pg、200pg、80pg、100pg、200pg。结论本文所建方法适用于血中吗啡、苯丙胺类、氯胺酮常见毒品的筛选及定量分析。     

荧光免疫标记抗体法测定血痕ABO血型

目的探讨采用荧光免疫标记抗体法测定血痕血型的可行性和优点。方法根据抗原抗体特异性结合的原理,首先对抗A、抗B单克隆抗体进行荧光标记,然后使荧光标记抗体与相应抗原(血痕)在最佳条件下结合,最后荧光显微镜镜检,判定血痕的血型。结果采用该方法测定血痕的血型结果准确、灵敏度高、操作简单、检验时间短。结论该方法可以作为一种测定血痕血型的检验方法使用。     

甲基苯丙胺及其代谢产物在急性中毒豚鼠体内的分布

目的研究甲基苯丙胺 (MAP)及其代谢产物苯丙胺 (AP)在急性中毒豚鼠体内的含量分布。方法应用GC/NPD技术 ,以 4 苯基丁胺 (4 PBA)为内标 ,样品经水解后碱化或直接碱化至pH >11,环已烷混旋提取 ,三氟乙酸酐 (TFA)微波衍生化 ,测定MAP急性中毒豚鼠体液和组织中MAP及AP的含量。结果急性中毒死亡豚鼠体内各器官和体液中MAP及AP含量最高为肺 ;其次为肝、脑、肾、脾、肠、心、血 ;再次为胃、胆汁 ;最少是尿。结论MAP在动物体内代谢迅速 ,组织或体液中MAP和AP浓度的比值与豚鼠给药后存活时间有关     

Determination of amphetamine-type stimulants,ketamine and metabolites in fingernails by gas chromatography–mass spectrometry

A gas chromatography–mass spectrometry (GC–MS) method was developed and validated for the simultaneous qualification and quantification of methamphetamine (MA), amphetamine (AP), 3,4-methylenedioxy-N-methylamphetamine (MDMA), 3,4-methylenedioxy-N-amphetamine (MDA), ketamine (KET) and norketamine (NKT) in fingernails. Fingernail samples (20 mg) were washed with distilled water and methanol, digested with 1.0 M sodium hydroxide at 95 °C for 30 min, and then extracted with ethyl acetate. Extract solutions were evaporated to dryness, derivatized using heptafluorobutyric anhydride (HFBA) at 60 °C for 30 min, and analyzed by GC–MS. The linear ranges were 0.1–20.0 ng/mg for AP, MDMA and NKT, 0.2–20.0 ng/mg for MA and MDA, and 0.4–20.0 ng/mg for KET, with the coefficients of determination (r² ≥ 0.9989). The intra- and inter-day precisions were within 7.1% and 10.6%, respectively. The intra- and inter-day accuracies were ?10.9% to 0.8% and ?4.3% to 4.5%, respectively. The limits of detections (LODs) and the limits of quantifications (LOQs) for each analyte were lower than 0.094 ng/mg and 0.314 ng/mg, respectively. The recoveries were in the range of 72.3–94.9%. The average fingernail growth rates of two subjects for three years and six subjects for two months were 3.12 mm/month and 3.16 mm/month, respectively. The method proved to be suitable also for the simultaneous detection and quantification of MA, MDMA, KET and their metabolites in fingernails.     

Stereoselective analyses of selegiline metabolites: possible urinary markers for selegiline therapy

The stereoselective analysis of selegiline metabolites in human urine and plasma by gas chromatography using the chiral column with the non-chiral reagent was investigated for the differentiation of selegiline therapy from the methamphetamine (MA) abuse. This method gave clear separations of MA and amphetamine (AM) isomers without any artifactual optical-opposite peaks due to the reagent. After the administration of selegiline tablets, desmethylselegiline (DMS), MA and AM were observed as (−)-isomers in the urine and plasma. Within the first 48 h after dosing, approximately 40% of selegiline administered was excreted in urine as these three metabolites. The parent drug, selegiline, was not detected in any urine or plasma samples. On the other hand, MA and AM were observed only as (+)-isomers in the urine of MA abusers. For the distinction of selegiline users from street MA abusers in urinalysis, (−)-DMS, a specific metabolite of selegiline, was not a suitable marker. (−)-DMS rapidly disappeared from urine and was excreted only 1% of the given dose. By the moment analysis with the trapezoidal integration, the mean residence times of (−)-DMS in plasma and urine were 2.7 and 3.8 h, respectively, which were 5–20 times shorter than those of (−)-MA or (−)-AM. The values of AM/MA in the urine increased from 0.24 to 0.67 (r=0.857) along with time after the selegiline administration. This ratio was not a sufficient marker to differentiate selegiline users from MA abusers, although the values of AM/MA in 74% of MA abusers were less than 0.24. The present GC technique improved the chiral analyses of MA and AM. This chiral analysis is the most useful technique to avoid the misinterpretation in the discrimination between clinical selegiline therapy and illicit MA use.