Direct determination of stability of protease inhibitors in plasma by HPLC with automated column-switching

• Published in: Journal of pharmaceutical and biomedical analysis Vol. 19; no. 3; pp. 343 - 349
• Main Authors: Peng, Sean X, Strojnowski, Michael J, Bornes, David M
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.03.1999; Elsevier Science
• Subjects: Analysis; Animals; Biological and medical sciences; Chromatography, High Pressure Liquid - instrumentation; Chromatography, High Pressure Liquid - methods; Column-switching; Drug Stability; Evaluation Studies as Topic; General pharmacology; Hydroxamic Acids - blood; Matrix Metalloproteinase Inhibitors; Medical sciences; MMP inhibitors; Pharmacology. Drug treatments; Protease Inhibitors - blood; Protease Inhibitors - chemistry; RAM column; Rats; Reproducibility of Results; Sensitivity and Specificity; Time Factors; MMP inhibitors; Column-switching; Drug stability; RAM column; Automation; Column switching; Rat; Enzyme; Chemical stability; Rodentia; HPLC chromatography; Enzyme inhibitor; Metabolism; In vitro; Metalloproteins; Blood plasma; Direct method; Peptidases; Vertebrata; Mammalia; Animal; Detection limit; Hydrolases; Comparative study; Quantitative analysis
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/S0731-7085(98)00127-7

Automated column-switching HPLC methods were developed and utilized for the direct analyses of three hydroxamic acid based metalloprotease inhibitors in rat plasma. These column-switching methods involved the use of a restricted-access media (RAM) precolumn and a column-switching valve, allowing the complete automation of sample preparation and HPLC. The plasma samples were directly injected onto a precolumn packed with SPS/ODS stationary phase and then backflushed onto an ODS analytical column using a 6-port column-switching device. The drug stability in rat plasma was determined using both the automated and traditional HPLC methods. The results obtained from the automated column-switching methods were in good agreement with those from traditional methods that involve sequential protein precipitation, liquid extraction, solvent evaporation, and sample reconstitution. In addition to the elimination of labor-intensive and time-consuming sample preparation procedures, the column-switching methods allowed on-line analyte enrichment and accurate determination of drug stability in plasma with detection limits in the range of 10–20 ng ml −1. This work represents, for the first time, a drug stability study in plasma by automated column-switching HPLC technique with the use of a RAM column. Our column-switching methods can be readily adapted to any existing HPLC system with minimal hardware modification.