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Rapid Screening of Amphetamine Drugs in Urine by Positive Ion Electrospray LC/MS/MS

05) and Collection Rack for 13 x 75 mm test tubes (Varian Part No. 12234507).

Sample Preparation

A 100 μL aliquot of a 500 ng/mL deuterated internal standards solution was transferred into individually labeled tubes (double blank tube was urine only). To each tube, a 1 mL of urine sample followed by 0.1 mL of 0.1 N KOH solution was added and mixed by vortex.

The mixture was loaded onto the sorbent bed of an activated 3 mL Focus cartridge pretreated with 1 mL of methanol followed by a 1 mL deionized water wash under gentle vacuum of1 to 2 in. Hg. Next, the sorbent bed was washed with 2 x 1 mL acetonitrile/water (10:90, v/v) under gentle vacuum.

The analyte was collected in a 2 mL autosampler vial by eluting with 2 x 100 μL elution solvent (acetonitrile/methanol/ water/formic acid (22:68:9:1, v/v) under gentle vacuum. The sorbent bed was then flushed with 600 μL of water under vacuum to wash off the elution solvent and dilute the sample for injection. A 10 μL aliquot was injected directly for analysis.

Results and Discussion

The LC method used a six-minute run cycle time with the first peak at 1.65 minutes and the last peak at 4.12 minutes (Figure 1). The two product ions for each analyte can be quantitatively analyzed at the level of 5 ng/mL in urine (Figure 2, about 50 pg on-column). This level is 50 times below the proposed drug cutoff levels published by the Substance Abuse and Mental Health Services Administration (SAMHSA).1 Eight concentration levels were used to generate the calibration curves for the standard. The linearity of the detector response and the response factor-Relative Standard Deviation (rf-RSD) are excellent (Table 1, Figure 3).

The recovery of the drugs from urine was > 85%. The eluent from the Focus cartridge can be injected dir
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