[Direkt zum Inhalt](https://www.sigmaaldrich.com#main-content) [![Merck](https://www.sigmaaldrich.com/static/logos/purple/merck.svg)](https://www.sigmaaldrich.com/AT/de) Produkte Warenkorb0 ATDE Produkte [Anmelden / Registrieren](https://www.sigmaaldrich.com/oidc-sign-in) [Bestellungssuche](https://www.sigmaaldrich.com/AT/de/order-lookup) [Schnelleinkauf](https://www.sigmaaldrich.com/AT/de/quick-order) Warenkorb0 [Home](https://www.sigmaaldrich.com/AT/en)[Solid Phase Extraction (SPE)](https://www.sigmaaldrich.com/AT/en/applications/analytical-chemistry/sample-preparation/solid-phase-extraction)The Extraction of Amphetamine and Related Drugs using Molecularly Imprinted Polymer SPE # The Extraction of Amphetamine and Related Drugs using Molecularly Imprinted Polymer SPE __Christine Widstrand, Staffan Bergström, Anna-Karin Wihlborg1, An Trinh2__ 1MIP Technologies AB, Scheelevägen 22, 220 07, Lund, Sweden, 2Supelco, 595 North Harrison Road, Bellefonte, PA, 16823, USA *Reporter US Volume 26.1* [[email protected]](mailto:[email protected]) __Section Overview__ - [Introduction](https://www.sigmaaldrich.com#introduction) - [Molecularly Imprinted Polymers](https://www.sigmaaldrich.com#molecularly-imprinted-polymers) - [Extraction and Analysis of Amphetamine and Amphetamine Related Drugs](https://www.sigmaaldrich.com#extraction-analysis) - [Lower Limits of Quantitation Achieved using SupelMIP SPE](https://www.sigmaaldrich.com#lower-limits-of-quantitation) - [Improved Recovery & Reduced Ion-Suppression Achieved using SupelMIP SPE](https://www.sigmaaldrich.com#improved-rocovery) - [Conclusion](https://www.sigmaaldrich.com#conclusion) - [Related Materials](https://www.sigmaaldrich.com#related-materials) ## [](https://www.sigmaaldrich.com)Introduction Amphetamine and amphetamine related drugs consist of a class of stimulants and hallucinogens used by students, athletes, and recreation drug users. Over the last 15 years, abuse of these drugs has become a global problem. In 2004, there were over 17,000 methamphetamine lab seizures in the US; and as a result, The Combat Methamphetamine Act of 2005 was signed into on law in 2006 to regulate over-the-counter sales of ephedrine, pseudoephedrine, and phenylpropanolamine products which are precursors used for the illicit manufacture of amphetamine and methamphetamine (1). In a 2005 report conducted by the European Monitoring Centre for Drugs and Drug Addiction, use of amphetamine and ecstacy has been growing throughout Europe (2). In recent years, a variety of screening/detection techniques have been commercially available such as ELISA and other immuno-based colorimetric and point of collection (POC) kits/devices. Although such assays are fast and efficient, many of which are typically not class-selective and will only detect some amphetamines and not others (3). Or, some immunoassays are not selective enough and recognize such a wide range of sympathomimetic amines, the positive predictive value (PPV) can range from 0-90% (4). In either case, an alternative and highly selective class-specific confirmation assay is required to verify immuno-based screening results. In this report we describe a simple, fast and class selective method for trace extraction of amphetamine related drugs from urine samples using a molecularly imprinted polymer SPE (SupelMIP™) specifically designed for selective extraction of amphetamine related drugs (__Figure 1__). The SupelMIP technique is also compared against a recently published extraction technique using a conventional hydrophilic polymer SPE phase (5). ![ Chemical Structures of Amphetamines and Related Drugs Investigated](https://www.sigmaaldrich.com/content/dam/cms-commons/sigmaaldrich/marketing/global/images/technical-documents/articles/analytical-chemistry/solid-phase-extraction/the-extraction-of-a.png "the-extraction-of-a") __Figure 1.__ Chemical Structures of Amphetamines and Related Drugs Investigated ## [](https://www.sigmaaldrich.com)Molecularly Imprinted Polymers Molecularly imprinted polymers (MIPs) are a class of highly cross-linked polymer-based molecular recognition elements engineered to bind one target compound or a class of structurally related compounds with high selectivity. Selectivity is introduced during MIP synthesis in which a template molecule, designed to mimic the analyte, guides the formation of specific cavities or imprints that are sterically and chemically complementary to the target analyte(s). As a result, multiple interactions (e.g., hydrogen bonding, ionic, Van der Waals, hydrophobic) can take place between the MIP cavity and analyte functional groups. The strong retention offered between a MIP phase and its target analyte(s) allows for the use of exhaustive wash procedures during solid phase extraction that results in superior sample cleanup prior to analysis. This leads to cleaner extracts, lower detection limits and a more efficient sample cleanup process. An illustration of the selective cavity is shown in __Figure 2__. ![Illustration of a Selective MIP Cavity](https://www.sigmaaldrich.com/content/dam/cms-commons/sigmaaldrich/marketing/global/images/technical-documents/articles/analytical-chemistry/solid-phase-extraction/the-extraction-of-b.png "the-extraction-of-b") __Figure 2.__ Illustration of a Selective MIP Cavity ## [](https://www.sigmaaldrich.com)Extraction and Analysis of Amphetamine and Amphetamine Related Drugs Amphetamine and the related drugs methamphetamine, phentermine, MDA, MDMA and MDEA were extracted from urine using both the SupelMIP SPE and a conventional hydrophilic polymer SPE via the procedures described in __Table 1__. Extracts were further analyzed via LC-MS-MS (__Table 2__). SupelMIP SPE- Amphetamines MethodPublished Amphetamine Method Using Conventional Hydrophilic Polymer SPE Phase (5) __Sample Pre-Treatment____Sample Pre-Treatment__ Human urine samples were spiked with internal standard (methamphetamine-d3 and MDM A-d5) and diluted 1:1 (v/v) with 10mM ammonium acetate buffer, pH 8. Adjust to pH 7.5-8.5 using NH, or HAc __SPE Procedure__Human urine samples were spiked with internal standard. Spiked and blank urine acidified with 100 µl 5 M HCI per 10 mL  urine. __SPE Procedure__ SupelMIP SPE -  Amphetamines, 25 mg/3 mL  (53228-U) 1.Condition and equilibrate MIP phase with 1 mL methanol, and 1 mL 10 mM ammonium acetate buffer, pH 8 2\. Load 1 mL pre-treated sample on to the cartridge. 3\. Wash (elute interferences) using the following wash scheme: • 2 x 1 mL DI water (Do not let column dry!) • 1 mL  60/40 MeCN/DI water followed by 5-10 minute vacuum (-1 bar, -20 inHg, or -70 kPA) to dry the column •  1 mL  1% HAc in MeCN 4\. Elute the amphetamine drugs with 2 x 1 mL 1% formic acid in methanol. Apply - 0.4 bar (-12 in Hg) between each fraction. 5\. Evaporate under nitrogen to dryness and reconstitute with 150µl  LC mobile phase (90% A and 10% B) prior to LC-MS-MS analysis Conventional Hydrophilic Polymer SPE Phase, 30  mg/1 mL 1.  Condition and equilibrate SPE phase with 1 mL methanol and 1 mL DI water 2\. Load 1 mL  pre-treated sample on to the cartridge. 3\. Wash (elute interferences) with 1 mL 5% methanol containing 2% ammonium hydroxide and with 1 mL 20% methanol containing 2% ammunium hydroxide 4\. Elute the amphetamine drugs with 0.5 mL  20% methanol with 2% acetic acid 5\. Evaporate under nitrogen and reconsittute with 150 µl  LC mobile phase prior to LC-MS-MS analysis Table 1. Comparison of SupelMIP SPE – Amphetamines Method and Conventional Hydrophilic Polymer SPE Method (53228-U) Column:Ascentis C18, 15 cm x 2.1 mm, 5µm particles, ([581304-U](https://www.sigmaaldrich.com/AT/en/product/supelco/581304U)) Instrument:Shimadzu LC-20/Applied Biosystems/MDS SCIEX APl3200 Mobile Phase:Solvent A - DI Water + 0.05% TFA Solvent B - Acetonitrile + 0.05% TFA Gradient:Initial: 90% A - 10% B 7 min 70% A - 30% B 10-11 min 10% A - 90% B 11.2 min 90% A - 10% B Temp.:Ambient Flow Rate:0.2 mL/min Ion Mode:Positive Ion Source:Turbospray Ion Spray Voltage:5500V Source Temp.:600 °C Collision Gas:6 psi Inj.:20 µl Det.:MS-MS MRM transition and retention times:Compound Amphetamine Methamphetamnie Methamphetamine D8 Phentermine MDA MDMA MDMA D 6 MDEARt (min) 7.8 8.3 8.3 8.7 8.0 8.5 8.5 9.2Q1/Q3 136/119and 136/91 1S0/119and 1S0/91 1S8/124 and 1S8/93 1S0/133 and 1S0/91 180/163and 180/10S 194/163and 194/10S 199/16S and 199/136 208/163 and108/105 Table 2. LC-MS-MS Conditions ([581304-U](https://www.sigmaaldrich.com/product/supelco/581304U)) ## [](https://www.sigmaaldrich.com)Lower Limits of Quantitation Achieved using SupelMIP SPE The amphetamine drugs listed in __Figure 1__ were spiked into urine at the levels of 15 and 50 ng/mL, and extracted/ analyzed using the procedures described in __Table 1__ and __Table 2__. The lower limit of quantitation (LLOQ) for each analyte was estimated for both the SupelMIP SPE and conventional hydrophilic SPE methods (signal to noise ratio of 10:1) and summarized in __Table 3__. Using the SupelMIP method, LLOQs in the range of 2.5 – 43.0 pg/mL (an order of magnitude lower than the conventional hydrophilic polymer SPE method) were achieved. To further illustrate the high selectivity achieved using the SupelMIP SPE extraction method, urine was spiked with 15 pg/mL amphetamine and extracted/analyzed as above. An amphetamine peak was easily detected using the SupelMIP SPE extraction method. In contrast, no amphetamine response was observed using the conventional hydrophilic polymer SPE method (__Figure 3__).  SupelMIP (pg/mL)Conventional Hydrophilic Polymer SPE (pg/mL) Methamphetamine6.652 Amphetamine7.3138 Phentermine1.5141 MDA4.3261 MDMA3.056 MDEA2.552 Table 3. LLOQ for SupelMIP SPE vs. Conventional Hydrophilic Polymer SPE ![Amphetamine Spiked Urine Samples (15 pg/mL) cleaned up with SupelMIP SPE vs. Conventional Hydrophilic Polymer SPE](https://www.sigmaaldrich.com/content/dam/cms-commons/sigmaaldrich/marketing/global/images/technical-documents/articles/analytical-chemistry/solid-phase-extraction/the-extraction-of-f.png "the-extraction-of-f") __Figure 3.__ Amphetamine Spiked Urine Samples (15 pg/mL) cleaned up with SupelMIP SPE vs. Conventional Hydrophilic Polymer SPE ## [](https://www.sigmaaldrich.com)Improved Recovery & Reduced Ion-Suppression Achieved using SupelMIP SPE Urine samples were spiked with the amphetamine drugs listed in __Figure 1__ at the levels of 0.010, 0.015, 0.50, 1.0, and 5.0 ng/mL and extracted/analyzed using the methods described in __Table 1__ and __Table 2__. Relative recovery for each spike concentration was determined against deuterated internal standards. Greater than 80% relative recovery was achieved for all the spike levels tested (with the exception of MDA) using the SupelMIP SPE method. In contrast, 0% recovery was observed for all the amphetamine drugs at spike levels 0.010 and 0.015 ng/mL, and at spike levels 0.5, 1.0 and 5.0 ng/mL, recoveries above 80% were only observed for methamphetamine and MDMA using the conventional hydrophilic polymer method (data not shown). Blank urine samples were cleaned up according to both SPE procedures. The SPE extracts were spiked with a mixture of the amphetamine drugs post extraction. Standards for the calibration curve were prepared in the reconstitution solvent. The standard calibration curves were compared to the matrix-matched samples for both methods, without using internal standard corrections. __Table 4__ summarizes the percentage of ion suppression at 10 and 100 ng/mL for the SupelMIP SPE and the hydrophilic polymer SPE method. From the table, ion suppression is significantly less for the SupelMIP SPE.  SupelMIP (ng/mL)Conventional Hydrophilic Polymer SPE (ng/mL)  __10____100____10____100__ Methamphetamine0%3%31%12% Amphetamine5%10%33%28% Phentermine3%2%36%22% MDA30%23%51%48% MDMA5%13%45%36% MDEA10%4%65%52% Table 4. % Ion-Suppression of SupelMIP SPE vs. Hydrophilic Polymer SPE ## [](https://www.sigmaaldrich.com)Conclusion[](https://www.sigmaaldrich.com) In this report, we demonstrated the utility of a molecularly imprinted polymer SPE (SupelMIP) method designed for the class-selective extraction of amphetamine and related drugs from urine. When compared to a recently published conventional hydrophilic polymer SPE method, the SupelMIP method offers reduced ion-suppression and achieved lower LOQs/LODs by an order of magnitude. The highly selective SupelMIP method also offered increased recovery and reproducibility for better sensitivity, precision and accuracy. ## Related Materials Bitte entschuldigen Sie, es ist ein unerwarteter Fehler aufgetreten Response not successful: Received status code 500 ### References 1\. The Combat Meth Act of 2005. Available at: . [http://www.deadiversion.usdoj. gov/meth/q\_a.html](http://www.deadiversion.usdoj.%20gov/meth/q_a.html) 2\. 2005\. European Monitoring Centre for Drugs and Drug Addiction, Available at: .. Annual Report . . [http://www.emcdda.eu.int/](http://www.emcdda.eu.int/) 3\. Walsh, Forensic Science International, 2007. In Press . 4\. Woodworth A, Saunders AN, Koenig JW, Moyer TP, Turk J, Dietzen DJ. 2006. Differentiation of Amphetamine/Methamphetamine and Other Cross-Immunoreactive Sympathomimetic Amines in Urine Samples by Serial Dilution Testing. 52(4):743-746. [https://doi.org/10.1373/clinchem.2005.060616](https://doi.org/10.1373/clinchem.2005.060616) 5\. Fuh M, Wu T, Lin T. 2006. Determination of amphetamine and methamphetamine in urine by solid phase extraction and ion-pair liquid chromatography?electrospray?tandem mass spectrometry. Talanta. 68(3):987-991. [https://doi.org/10.1016/j.talanta.2005.06.057](https://doi.org/10.1016/j.talanta.2005.06.057) __Related Articles__ - [Determination of Formaldehyde and Acetaldehyde in Air Using DNPH Cartridges and Automated On-Line Desorption Followed by HPLC](https://www.sigmaaldrich.com/AT/en/technical-documents/protocol/environmental-testing-and-industrial-hygiene/air-testing/aldehydes-air-dnph) - [Effect of Container Materials on Milk Off-Flavors](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/gas-chromatography/container-materials) - [Pesticide Residue Analysis in Ginger Powder with QuEChERS](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/quechers/analysis-of-pesticide-residues-in-ginger-powder) - [Analysis of Pesticide Residues in Pistachios Using QuEChERS Extraction and Cleanup with Supel™ QuE Z-Sep+](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/quechers/analysis-of-pesticide-residues-in-pistachios) - [HybridSPE-Phospholipid Technology](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/small-molecule-hplc/hybridspe-precipitation-technology) - [Impact of Mobile Phase Additives on LC-MS Sensitivity, Demonstrated using Spice Cannabinoids](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/small-molecule-hplc/lcms-spice-cannabinoids) - [Solvent Analysis for 40 Per and Polyfluoroalkyl Substances](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/small-molecule-hplc/solvent-analysis-for-40-per-and-polyfluoroalkyl-substances) - [Analysis of Drugs of Abuse in Urine](https://www.sigmaaldrich.com/AT/en/technical-documents/technical-article/analytical-chemistry/solid-phase-extraction/analysis-of-drugs-of-abuse-in-urine-after-cleanup-with-hlb) - [View More](https://www.sigmaaldrich.com/AT/en/search/facet-search?focus=sitecontent&term=facet-search) __Related Product Categories__ - [Solid Phase Extraction (SPE) and QuEChERS](https://www.sigmaaldrich.com/AT/en/products/analytical-chemistry/analytical-sample-preparation/spe-and-quechers) - [HPLC Columns](https://www.sigmaaldrich.com/AT/en/products/analytical-chemistry/analytical-chromatography/hplc-columns) Oben __Melden Sie sich an, um fortzufahren.__ Um weiterzulesen, melden Sie sich bitte an oder erstellen ein Konto. Anmelden__Sie haben kein Konto?__Registrieren An unknown error has occured. - Deutsch - DE - English - EN [Weitere Informationen](https://www.sigmaaldrich.com)