{"id":43389,"date":"2025-10-03T21:03:32","date_gmt":"2025-10-03T21:03:32","guid":{"rendered":"https:\/\/discover.restek.com\/uncategorized\/analysis-of-d-and-l-amphetamine-and-methamphetamine-enantiomers-for-high-throughput-labs\/"},"modified":"2026-01-03T17:52:22","modified_gmt":"2026-01-03T17:52:22","slug":"analysis-of-d-and-l-amphetamine-and-methamphetamine-enantiomers-for-high-throughput-labs","status":"publish","type":"post","link":"https:\/\/discover.restek.com\/ja\/artikel\/cfss2974\/analysis-of-d-and-l-amphetamine-and-methamphetamine-enantiomers-for-high-throughput-labs","title":{"rendered":"Analysis of d- and l-Amphetamine and Methamphetamine Enantiomers for High-Throughput Labs"},"content":{"rendered":"\n<ul class=\"wp-block-list\">\n<li>Separate d- and l- enantiomers without chiral columns and dedicated instruments.<\/li>\n\n\n\n<li>Simple precolumn derivatization and dilution sample preparation.<\/li>\n\n\n\n<li>Accurately distinguish and quantify licit vs. illicit methamphetamine.<\/li>\n<\/ul>\n\n\n\n<p>Amphetamine and methamphetamine are psychostimulant drugs and occur as two enantiomers, dextrorotary and levorotary, as a result of their chiral center. The dextro-methamphetamine (d-isomer) form is highly abused and is typically found in illicit preparations. However, detection of abuse is complicated because consumption of over-the-counter and prescription medications may yield positive results if the analytical method used cannot distinguish between the enantiomers. Chiral separation of d- and l-methamphetamine and their metabolites d- and l-amphetamine can help determine whether the source was licit or illicit, but chiral columns can be expensive, may necessitate a dedicated instrument, and are not as broadly useful as ubiquitous C18 columns.<\/p>\n\n\n\n<p>In order to provide labs with a high-throughput assay that effectively separates d- and l-amphetamine and methamphetamine enantiomers in urine without the use of a costly and specialized chiral column, we developed the LC-MS\/MS method shown here on a standard reversed-phase Raptor C18 column. Our method employs a simple precolumn derivatization followed by dilution and results in a selective, specific analysis of d- and l-amphetamine and methamphetamine enantiomers that is free from sample matrix interferences. Separation was achieved within a total run time of seven minutes, and quantitation in urine was performed across a linear range of 50-5000 ng\/mL. Validation across this range demonstrated that this method provides reliable analysis of d- and l-amphetamine and methamphetamine enantiomers in a workflow and time frame suitable for high-throughput clinical and forensic toxicology labs.<\/p>\n\n\n<div class=\"wp-block-custom-chromatogram-article-top\"><div class=\"chromatogram-article-placeholder\"><div class=\"figure-heading\"><\/div><div class='chromatogram-article-inner-full'><div class=\"chromatogram-article-inner\">\n<div class=\"wp-block-custom-chromatogram-article\"><div class=\"wp-block-custom-chromatogram-article\"><div class=\"chromatogram-image wide-image\"><img decoding=\"async\" src=\"https:\/\/ez.restek.com\/images\/cgram\/lc_cf0737.png\" alt=\"d &amp; l Amphetamines (DNPA Derivatives) in Urine by LC-MS\/MS\" title=\"-\"><\/div><p class=\"article-id\" style=\"text-align:center\"> LC_CF0737<\/p><div class=\"chromatogram-peaks\"><h4>Peaks<\/h4><table class=\"peaks col-lg-6 col-12\">\n<thead><tr><th><\/th><th style=\"text-align: left;width: 75px\">Peaks<\/th><th style=\"text-align: center;width: 75px\">t<sub>R<\/sub> (min)<\/th><th style=\"text-align: center;width: 75px\">Precursor Ion<\/th><th style=\"text-align: center;width: 75px\">Product Ion 1<\/th><th style=\"text-align: center;width: 75px\">Product Ion 2<\/th><\/tr><\/thead>\n<tbody><tr><td class=\"num\">1.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for l-Methamphetamine*\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/537-46-2\/l-Methamphetamine\" target=\"_blank\" rel=\"noopener\">l-Methamphetamine*<\/a><\/td><td class=\"oth\">3.11<\/td><td class=\"oth\">400.3<\/td><td class=\"oth\">339.0<\/td><td class=\"oth\">323.8<\/td><\/tr>\n<tr><td class=\"num\">2.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for d-Methamphetamine*\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/537-46-2\/d-Methamphetamine\" target=\"_blank\" rel=\"noopener\">d-Methamphetamine*<\/a><\/td><td class=\"oth\">3.38<\/td><td class=\"oth\">400.3<\/td><td class=\"oth\">339.0<\/td><td class=\"oth\">323.8<\/td><\/tr>\n<tr><td class=\"num\">3.<\/td><td class=\"cmpd\">l-Amphetamine*<\/td><td class=\"oth\">4.16<\/td><td class=\"oth\">386.1<\/td><td class=\"oth\">325.0<\/td><td class=\"oth\">308.0<\/td><\/tr>\n<tr><td class=\"num\">4.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for d-Amphetamine*\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/51-64-9\/d-Amphetamine\" target=\"_blank\" rel=\"noopener\">d-Amphetamine*<\/a><\/td><td class=\"oth\">4.55<\/td><td class=\"oth\">386.1<\/td><td class=\"oth\">325.0<\/td><td class=\"oth\">308.0<\/td><\/tr>\n<\/tbody><\/table><div class=\"peaknotes\">*Analytes are DNPA derivatives.<\/div><\/div><div class=\"chromatogram-conditions\"><h4>Conditions<\/h4><div class=\"conditions-container container-fluid\"><div class=\"row\"><table class=\"conditions col-lg-6 col-12\"><tr><th class=\"conditions_header\" scope=\"row\">Column<\/th><td>Raptor C18  (<a target=\"_blank\" href=\"https:\/\/www.restek.com\/p\/9304A12?utm_source=chromatograms&amp;utm_medium=link&amp;utm_campaign=LC_CF0737\" rel=\"noopener\">cat.# 9304A12<\/a>)<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Dimensions:<\/th><td>100 mm x 2.1 mm ID<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Particle Size:<\/th><td>2.7 \u00b5m<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Pore Size:<\/th><td>90 \u00c5<\/td><\/tr><tr><td><\/td><tr><th class=\"sub conditions_header\" scope=\"row\">Guard Column:<\/th><td>Raptor C18 EXP guard column cartridge 5 mm, 2.1 mm ID, 2.7 \u00b5m (<a target=\"_blank\" href=\"https:\/\/www.restek.com\/p\/9304A0252?utm_source=chromatograms&amp;utm_medium=link&amp;utm_campaign=LC_CF0737\" rel=\"noopener\">cat.# 9304A0252<\/a>)<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Temp.:<\/th><td>35 \u00b0C<\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Standard\/Sample<\/th><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Conc.:<\/th><td>500 ng\/mL in urine<\/td><\/tr><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Inj. Vol.:<\/th><td>10 \u00b5L <\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Mobile Phase<\/th><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">A:<\/th><td>0.1% Formic acid in water <\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">B:<\/th><td>0.1% Formic acid in methanol <\/td><\/tr><tr><td><\/td><td><table class=\"cgram_ramp\"><thead><tr><th>Time (min)<\/th><th>Flow (mL\/min)<\/th><th>%A<\/th><th>%B<\/th><\/tr><\/thead><tbody><tr><td>0.00<\/td><td>0.5<\/td><td>40<\/td><td>60<\/td><\/tr><tr><td>5.00<\/td><td>0.5<\/td><td>40<\/td><td>60<\/td><\/tr><tr><td>5.01<\/td><td>0.5<\/td><td>10<\/td><td>90<\/td><\/tr><tr><td>5.50<\/td><td>0.5<\/td><td>10<\/td><td>90<\/td><\/tr><tr><td>5.51<\/td><td>0.5<\/td><td>40<\/td><td>60<\/td><\/tr><tr><td>7.00<\/td><td>0.5<\/td><td>40<\/td><td>60<\/td><\/tr><\/tbody><\/table><\/td><\/tr><\/table><table class=\"conditions col-lg-6 col-12\"><tr><th class=\"conditions_header\" scope=\"row\">Detector<\/th><td>MS\/MS<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Ion Mode:<\/th><td>ESI- <\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Mode:<\/th><td>MRM <\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Instrument<\/th><td>HPLC<\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Sample Preparation<\/th><td>A 500 ng\/mL standard (d- and l-amphetamines and methamphetamines) was prepared in pooled urine. Fifty microliters of the standard was aliquoted into a microcentrifuge tube. Ten microliters of a working internal standard (20 \u00b5g\/mL (\u00b1)-amphetamine-D11 and (\u00b1)-methamphetamine-D11 in water) and 20 \u00b5L of 1M NaHCO<sub>3<\/sub> was added and vortexed at 3000 rpm for 10 seconds. After vortexing, 100 \u00b5L of 0.1% (w\/v) Marfey&#8217;s reagent (1-fluoro-2-4-dinitrophenyl-5-L-alanine amide) in acetone was added, vortexed, and heated at 45 \u00b0C for 1 hour. Samples were allowed to cool to room temperature before the addition of 40 \u00b5L of 1M HCl in water. The sample was then vortexed and evaporated to dryness under nitrogen at 45 \u00b0C. Samples were reconstituted in 1 mL of 40:60 water:methanol (v\/v) and filtered using Thomson SINGLE StEP standard filter vials cat.# 25893 prior to analysis.<\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Notes<\/th><td>Thomson SINGLE StEP standard filter vials cat.# 25893 were used to produce this chromatogram, but have since been discontinued. For assistance choosing a replacement for this application, contact Restek Technical Service or your local Restek representative.<br \/><\/td><\/tr><\/table><\/div><\/div><\/div><div class=\"chromatogram-pdf-link\"><a href=\"https:\/\/ez.restek.com\/images\/cgram\/lc_cf0737.pdf\" target=\"_blank\" rel=\"noopener noreferrer\"><svg xmlns=\"http:\/\/www.w3.org\/2000\/svg\" width=\"18\" height=\"18\" viewBox=\"0 0 18 18\"><g data-name=\"Group 2996\"><path data-name=\"Rectangle 1246\" d=\"M0 0h18v18H0z\" style=\"fill: none;\"><\/path><\/g><g data-name=\"Group 2997\"><path data-name=\"Path 729\" d=\"M13.412 11.4v2.017H5.345V11.4H4v2.017a1.349 1.349 0 0 0 1.345 1.345h8.068a1.349 1.349 0 0 0 1.345-1.345V11.4zm-.672-2.694-.948-.948-1.741 1.735V4H8.706v5.493L6.965 7.758l-.948.948 3.361 3.361z\" transform=\"translate(-.437 -.414)\" style=\"fill: rgb(13, 123, 196);\"><\/path><\/g><\/svg>Download PDF<\/a><\/div><\/div><\/div>\n<\/div><\/div><\/div>\n\n\n<div style=\"height:30px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n        <div class=\"cpb\">\n            <h3 class=\"cpb-heading\">Products Mentioned<\/h3>\n            <hr class=\"cpb-heading-underline\" \/>\n            <div class=\"cpb-list\">\n                                    <div class=\"cpb-item\">\n                        <div class=\"cpb-col cpb-col--left\">\n                            <a class=\"cpb-catalog\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/9304A12\">                                Catalog No. 9304A12                            <\/a>                        <\/div>\n                        <div class=\"cpb-col cpb-col--middle\">\n                            <div class=\"cpb-title\">Raptor C18, 2.7 \u00b5m, 100 x 2.1 mm HPLC\u30ab\u30e9\u30e0<\/div>\n                        <\/div>\n                        <div class=\"cpb-col cpb-col--right\">\n                                                            <a class=\"cpb-view-btn\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/9304A12\">\u88fd\u54c1\u60c5\u5831\u3092\u898b\u308b<\/a>\n                                                    <\/div>\n                    <\/div>\n                                    <div class=\"cpb-item\">\n                        <div class=\"cpb-col cpb-col--left\">\n                            <a class=\"cpb-catalog\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/9304A0252\">                                Catalog No. 9304A0252                            <\/a>                        <\/div>\n                        <div class=\"cpb-col cpb-col--middle\">\n                            <div class=\"cpb-title\">Raptor C18 EXP Guard Column Cartridge, 2.7 \u00b5m, 5 x 2.1 mm, 3-pk.<\/div>\n                        <\/div>\n                        <div class=\"cpb-col cpb-col--right\">\n                                                            <a class=\"cpb-view-btn\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/9304A0252\">\u88fd\u54c1\u60c5\u5831\u3092\u898b\u308b<\/a>\n                                                    <\/div>\n                    <\/div>\n                                    <div class=\"cpb-item\">\n                        <div class=\"cpb-col cpb-col--left\">\n                            <a class=\"cpb-catalog\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/25808\">                                Catalog No. 25808                            <\/a>                        <\/div>\n                        <div class=\"cpb-col cpb-col--middle\">\n                            <div class=\"cpb-title\">EXP Direct Connect Holder for EXP Guard Cartridges, Includes Fitting &amp; Ferrules<\/div>\n                        <\/div>\n                        <div class=\"cpb-col cpb-col--right\">\n                                                            <a class=\"cpb-view-btn\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/25808\">\u88fd\u54c1\u60c5\u5831\u3092\u898b\u308b<\/a>\n                                                    <\/div>\n                    <\/div>\n                                    <div class=\"cpb-item\">\n                        <div class=\"cpb-col cpb-col--left\">\n                            <a class=\"cpb-catalog\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/28307\">                                Catalog No. 28307                            <\/a>                        <\/div>\n                        <div class=\"cpb-col cpb-col--middle\">\n                            <div class=\"cpb-title\">Thomson SINGLE StEP Standard Filter Vials with Screw-Top Caps, 0.2 \u00b5m, PTFE w\/preslit cap, Green Cap, 100-pk.<\/div>\n                        <\/div>\n                        <div class=\"cpb-col cpb-col--right\">\n                                                            <a class=\"cpb-view-btn\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/www.restek.com\/p\/28307\">\u88fd\u54c1\u60c5\u5831\u3092\u898b\u308b<\/a>\n                                                    <\/div>\n                    <\/div>\n                            <\/div>\n        <\/div>\n        \n\n\n<div style=\"height:100px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n","protected":false},"excerpt":{"rendered":"<p>Accurate analysis of amphetamine and methamphetamine isomers can be achieved using chiral columns, but the method presented here provides a simpler, more cost-effective approach for high-throughput labs. Excellent separation of all isomers in urine was obtained in a fast, 7-min LC-MS\/MS quantitative analysis on a Raptor C18 column with no interference from matrix components.<\/p>\n","protected":false},"author":11,"featured_media":6511,"comment_status":"closed","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"_acf_changed":false,"_kad_blocks_custom_css":"","_kad_blocks_head_custom_js":"","_kad_blocks_body_custom_js":"","_kad_blocks_footer_custom_js":"","_kadence_starter_templates_imported_post":false,"_kad_post_transparent":"","_kad_post_title":"","_kad_post_layout":"","_kad_post_sidebar_id":"","_kad_post_content_style":"","_kad_post_vertical_padding":"","_kad_post_feature":"","_kad_post_feature_position":"","_kad_post_header":false,"_kad_post_footer":false,"footnotes":""},"categories":[787],"tags":[],"industries-application":[2223,2226,2239,2240],"post-badge":[],"resource-type":[2623],"product-library":[2499,2481,2483,2513],"resource-technique":[2326,2363],"hf_cat_post":[650,2624],"ppma_author":[414],"class_list":["post-43389","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-artikel","industries-application-klinisch","industries-application-klinische-toxikologie","industries-application-forensik","industries-application-forensische-toxikologie","resource-type-featured-application","product-library-lc-saulen","product-library-flussigchromatographie-produkte","product-library-produkte-zur-probenvorbereitung","product-library-vials-zubehor","resource-technique-flussigchromatografie","resource-technique-ms-ms-de"],"acf":[],"taxonomy_info":{"category":[{"value":787,"label":"Artikel"}],"industries-application":[{"value":2223,"label":"Klinisch"},{"value":2226,"label":"Klinische Toxikologie"},{"value":2239,"label":"Forensik"},{"value":2240,"label":"Forensische Toxikologie"}],"resource-type":[{"value":2623,"label":"Featured Application"}],"product-library":[{"value":2499,"label":"LC S\u00e4ulen"},{"value":2481,"label":"Fl\u00fcssigchromatographie Produkte"},{"value":2483,"label":"Produkte zur Probenvorbereitung"},{"value":2513,"label":"Vials &amp; 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