Sample processing and HPLC/tandem mass spectrometry conditions used for the 33 CNS drugs examined
Caffeine, chlorpromazine, meprobamate, and phenytoin were analyzed on a PE-Sciex API 4000 mass spectrometer. All other drugs were analyzed on a PE-Sciex API 3000. Mobile phase A consisted of 100% methanol. Mobile phase B consisted of 10 mM ammonium acetate containing 1% 2-propanol, pH 6.8. Mobile phase C consisted of 100% acetonitrile. Gradients were performed as detailed under Materials and Methods. Samples were injected onto either a Phenomenex Synergi Max-RP 2.0 × 30 mm, 4-μm column (column 1) or a Primesphere 5-μm HC-C18 2.0 × 30 mm column (column 2) maintained at 50†C.
Drug | Protein Aliquot | Buffer Aliquot | Injection Volume | MS Polarity | MRM Transition | Retention Time | Initial Condition | Intermediate Condition | Column |
|---|---|---|---|---|---|---|---|---|---|
| μl | μl | μl | min | ||||||
| Buspirone | 10 | 50 | 10 | + | 386.3:122.1 | 1.45 | 90% B:10% C | 10% B:90% C | 1 |
| Caffeine | 50 | 50 | 3 | + | 195.0:137.9 | 0.70 | 90% B:10% C | 10% B:90% C | 2 |
| Carbamazepine | 10 | 50 | 10 | + | 237.2:194.2 | 1.33 | 90% B:10% C | 10% B:90% C | 1 |
| Carisoprodol | 10 | 50 | 10 | + | 261.4:176.3 | 1.42 | 90% B:10% C | 10% B:90% C | 1 |
| Chlorpromazine | 10 | 100 | 10 | + | 319.2:245.7 | 0.75 | 95% B:5% C | 15% B:85% C | 1 |
| Citalopram | 10 | 50 | 10 | + | 324.5:262.2 | 1.40 | 90% B:10% C | 10% B:90% C | 1 |
| Clozapine | 10 | 50 | 3 | + | 327.1:269.8 | 0.80 | 20% B:80% A | 100% B | 1 |
| Cyclobenzaprine | 10 | 50 | 10 | + | 276.5:231.3 | 1.51 | 90% B:10% C | 10% B:90% C | 1 |
| Diazepam | 10 | 50 | 10 | + | 285.2:193.0 | 1.48 | 80% B:20% C | 15% B:85% C | 1 |
| Fluoxetine | 10 | 50 | 10 | + | 310.3:148.3 | 1.58 | 90% B:10% C | 10% B:90% C | 1 |
| Fluvoxamine | 10 | 50 | 10 | + | 319.2:200.1 | 1.44 | 90% B:10% C | 10% B:90% C | 1 |
| Haloperidol | 10 | 50 | 5 | + | 376.1:165.2 | 1.39 | 90% B:10% C | 10% B:90% C | 1 |
| Hydrocodone | 50 | 50 | 10 | + | 300.4:199.0 | 1.56 | 15% A:85% B | 95% A:5% B | 1 |
| Hydroxyzine | 10 | 50 | 5 | + | 375.2:201.0 | 1.58 | 90% B:10% C | 10% B:90% C | 1 |
| Lamotrigine | 50 | 50 | 10 | + | 256.0:211.0 | 1.50 | 5% A:90% B:5% C | 47.5% A:5% B:47.5% C | 2 |
| Meprobamate | 50 | 50 | 5 | + | 219.0:158.0 | 0.71 | 90% B:10% C | 50% B:50% C | 1 |
| Methylphenidate | 50 | 50 | 10 | + | 234.0:174.3 | 1.51 | 90% B:10% C | 10% B:90% C | 1 |
| Metocloperamide | 50 | 50 | 3 | + | 300.4:227.2 | 1.44 | 90% B:10% C | 50% B:50% C | 1 |
| Midazolam | 10 | 50 | 10 | + | 326.3:291.3 | 1.69 | 30% A:70% B | 90% A:10% B | 1 |
| Morphine | 10 | 50 | 10 | + | 779.3:649.5 | 1.26 | 10% A:90% B | 90% A:10% B | 1 |
| Nortriptyline | 10 | 50 | 10 | + | 264.4:233.3 | 1.82 | 10% A:90% B | 90% A:10% B | 1 |
| Paroxetine | 10 | 50 | 10 | + | 330.3:192.2 | 1.47 | 80% B:20% C | 15% B:85% C | 1 |
| Phenytoin | 10 | 50 | 9 | - | 251.0:102.0 | 0.70 | 10% A:90% B | 95% A:5% B | 2 |
| Propoxyphene | 10 | 50 | 10 | + | 340.3:266.3 | 1.61 | 90% B:10% C | 10% B:90% C | 1 |
| Risperidone | 10 | 50 | 10 | + | 411.3:191.3 | 1.39 | 95% B:5% C | 15% B:85% C | 1 |
| 9-OH-Risperidone | 10 | 50 | 3 | + | 427.0:207.0 | 0.70 | 10% A:90% B | 80% A:20% B | 1 |
| Selegiline | 10 | 50 | 10 | + | 188.4:119.2 | 1.63 | 90% B:10% C | 10% B:90% C | 1 |
| Sertraline | 10 | 50 | 10 | + | 306.2:159.1 | 1.55 | 5% A:90% B:5% C | 45% A:5% B:50% C | 1 |
| Sulpiride | 10 | 50 | 10 | + | 342.3:112.2 | 1.45 | 10% A:90% B | 90% A:10% B | 1 |
| Thiopental | 10 | 50 | 10 | - | 241.1:57.5 | 1.77 | 10% A:90% B | 90% A:10% B | 1 |
| Trazodone | 10 | 50 | 10 | + | 372.2:176.1 | 1.56 | 90% B:10% C | 10% B:90% C | 1 |
| Venlafaxine | 50 | 50 | 10 | + | 278.4:215.3 | 1.32 | 95% B:5% C | 5% B:95% C | 2 |
| Zolpidem | 10 | 50 | 9 | + | 308.4:235.0 | 1.72 | 20% A:60% B:20% C | 80% A:20% C | 1 |
MS, mass spectrometry; MRM, multiple reaction monitoring.