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Determination of potato carboxypeptidase inhibitor in African Green Monkey plasma using 96-well SPE and LC–MS/MS

Journal of Pharmaceutical and Biomedical Analysis
Publication Date
DOI: 10.1016/j.jpba.2004.09.059
  • Potato Carboxypeptidase Inhibitor
  • Cpi
  • Peptide Analysis
  • Protein Analysis
  • Lc–Ms/Ms


Abstract Potato carboxypeptidase inhibitor (CPI), a peptide with multiple isoforms (MW > 4000 Da) was determined from African Green Monkey plasma using a PE Sciex API-3000 LC–MS/MS in the positive ionization mode with the turbo ionspray interface (450 °C). Samples were prepared using an Oasis MCX 96-well solid phase extraction plate and chromatographed on an Allure C 18 HPLC Column (50 mm × 1.0 mm, 5 μm) using gradient elution. Upon analysis of the extracts using LC–MS/MS, the concentration of CPI was calculated using a single MS/MS transition ( m/z 830.5 → 221.0) that was reflective of the mass concentration (μg/mL) of main the CPI isoforms present in plasma from monkeys after they were given an intravenous dose of CPI. The assay was linear for CPI over concentrations of 0.05–10 μg/mL when extracting 200-μL aliquots of African Green Monkey plasma. The assay was applied to the determination of CPI in African Green Monkey plasma samples in two separate analytical runs (correlation of standard curves, r 1 = 0.9991 and r 2 = 0.9953). Quality control (QC) samples were run at 0.05, 0.1, 0.2, 0.5, 1.0, 2.0 and 5.0 μg/mL for each assay. Average ranges ( n = 12) for accuracy and precision for all concentrations of QCs during the two runs were 92.0–102.0% of expected potency and 10.4–21.8% (coefficient of variations), respectively.

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