METHAMPHETAMINE . . . on Wipes by SPE: METHOD 9109, Issue 1, dated 17 October 2011 - Page 4 of 33
d. Cap securely and mix contents by inverting the tubes end over end on a rotary mixer or equivalent at 10-30 rpm for at least one hour. e. Check the pH which should be about ≤ 4. If needed, adjust the pH with diluted (2.5 to 3 M) sulfuric acid drop-wise, mixing the contents by shaking or inversion a few times after each addition of acid before checking the pH. f. After mixing, transfer 10 mL of supernatant to a 25-mL glass centrifuge tube. NOTE: If extraction is to be performed on a subsequent day, store samples in a refrigerator. Analytes are stable in the desorption solution for at least one week refrigerated. 8. Solid phase extraction procedure: a. Column selection: Select one of the SPE columns listed in the EQUIPMENT section. Each brand of column has a slightly different conditioning procedure and resistance to flow. Other brands of SPE columns may also work. Elution profiles of drugs to be analyzed need to be determined before use of columns other than those specified. b. Setting up columns: Attach SPE columns to vacuum ports on the manifold. Attach vacuum line to vacuum pump capable of 25-30 psi vacuum. c. Conditioning: Condition each column with 1 column volume (3 mL) of methanol followed by 1 column volume of Type II deionized water. For some brands (e.g., Speedisk®) the conditioning volume is 1/3 column volume. Check product literature. d. Loading: Load each SPE column with 5 mL of the sample acid desorbate solution. Adjust vacuum so that the flow rate is about 1-2 mL/minute. The vacuum required to obtain that flow rate varies with brand of SPE column. e. First wash: Wash each column with 1 column volume (3 mL) of 0.1 M aqueous hydrochloric acid. For some brands (e.g., Oasis® or Speedisk®) this volume may be decreased to 2 or 1 mL, respectively. f. Second wash: Wash each column with 1 column volume of methanol. Add the methanol in 2 or 3 separate aliquots to ensure that the aqueous acid is flushed through. Discard all effluents. g. Drying: Remove last traces of water in the SPE columns by pulling air through the columns under increased vacuum (e.g., 25 psi) for 5 minutes. Silica-based SPE columns or columns with high resistance to flow may require a longer time to reach dryness. h. Elution: Position 13 x 100 mm collection tubes under each column. Elute analytes with 3 mL of elution solution (80:20:2 methylene chloride:isopropanol:concentrated ammonium hydroxide v/v, freshly prepared). Adjust vacuum so that the flow rate is 1 mL/minute or less. For some brands (e.g., Speedisk®) this flow rate may occur without applied vacuum. Most of the analytes (e.g., amphetamine, ephedrine, methamphetamine, etc.) are eluted in the first milliliter. 9. Evaporation: To each collection tube containing eluate, add about 5 μL crystal violet solution and 100 μL of 0.3 M hydrochloric acid in methanol. The samples are evaporated to dryness under gently blowing nitrogen at 25-35 °C. The samples should be removed from the evaporation bath within a few minutes after dryness. A mixed whitish and purple residue will remain. The purple color of the crystal violet helps to make the residue more visible when dried. The color of the crystal violet remains a constant blue to blue-violet during concentration and drying. 10. Derivatization: (Perform under the hood.) Add 100 μL of acetonitrile containing the optional dibromooctafluorobiphenyl secondary internal standard. Add 25 μL MSTFA and 25 μL MBHFBA in that order. Cap tubes between additions to prevent atmospheric humidity from affecting the reagents. (See note below. Have no more than 5 or 6 tubes uncapped at a time.) Vortex each tube about 4-5 seconds. Using Pasteur pipettes, transfer each mixture to low-volume (300-500 μL) amber autosampler vials and cap vials. NOTE 1: Some derivatization takes place at room temperature, especially trimethylsilylation. Derivatization is completed on-column after injection. No prior heating is required or recommended. NOTE 2: The color of the reconstituted solution should be deep blue to violet. If the color turns light blue or turquoise upon standing, moisture may be present (the vials may not have Method rev. 1.1.1
NIOSH Manual of Analytical Methods (NMAM), Fifth Edition
