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Sensitivity and Quantitation using a Finnigan LCQ Deca XP Plus Ion Trap Mass Spectrometer

ctron).
Injection volume: 5 μL
Flow Rate: 400 μL/min
Mobile phase A: water containing 10 mM ammonium acetate B: acetonitrile containing 0.1 % formic acid
Gradient: 45% B for 3 minutes

Mass Spectrometer
Ionization mode: Positive electrospray ionization (ESI)
Capillary temperature: 350C
Spray voltage: 4.5 kV
Sheath gas: 45 units Sweep gas: 10 units
Maximum inject time: 400 ms
Isolation width: 1.0
Collision energy: 40%
Number of segments: 2
Segment 1: Full-scan MS/MS m/z 301.2 → m/z [100-305]
Segment 2: Full-scan MS/MS Scan event 1 m/z 309.2 → m/z [100-311] Scan event 2 m/z 314.2 → m/z [100-320]

Analysis
The standard samples were analyzed with five replicate injections at each concentration level and a blank (MeOH) injection between each level.

Results and Discussion
Eleven calibration standards over a concentration range of four orders were used to evaluate the quantitative performance of the LCQ Deca XP Plus mass spectrometer. Figure 1 shows the extracted ion chromatograms from full-scan MS/MS of alprazolam (500 fg on-column) and alprazolam-d5 internal standard (50 pg on-column) in protein precipitated bovine plasma. These chromatograms were generated by summing the two most intense product ions observed in the full-scan MS/MS spectra (Figure 2) acquired for the analyte (m/z 274 + 281) and the internal standard (m/z 279 + 286). The LOD was determined to be 500 fg for alprazolam, with a signal-tonoise of 5 to 1.

Figure 3 shows the extracted ion chromatograms from full-scan MS/MS of temazepam and alprazolam spiked in protein-precipitated bovine plasma at 0.2 pg/μL (1 pg oncol
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