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Bioanalysis and Pharmacokinetics

Bioanalysis: The Preclinical Pharmacology Core makes use of three combination triple quadrupole/ion trap mass spectrometers from AB Sciex: a 3200 QTRAP® System, a 4000 QTRAP® System, and a QTRAP® 6500+ System.

All three mass spectrometers are coupled to Shimadzu Prominence liquid chromatography instruments with 80 to 100-place autosamplers. Development of an analytical method requires several steps: optimization of an MS/MS method to detect the compound with optimal sensitivity and accuracy; development of an appropriate chromatographic method; and development of a method for sample preparation and extraction of the compound from the desired biological matrices.

Plasma drug levels graph

Standard conditions, which include use of a reverse phase C18 column with methanol or acetonitrile and water containing 0.1% formic acid as the running buffers, and simple protein precipitation for sample preparation, are attempted first. If any of these approaches prove ineffective, core staff will utilize specialized columns (e.g. anion exchange, polar endcapped columns), alternate buffers or chromatographic gradients, or more elaborate sample preparation (solid phase extraction, liquid-liquid extraction, phospholipid removal). While work is not done under GLP conditions, the Core attempts to follow the FDA Guidance for Industry - Bioanalytical Method Validation (https://www.fda.gov/downloads/Drugs/Guidances/ucm070107.pdf) published in May 2018. 

Sample Requirements: 10 mM DMSO stock

Pharmacokinetics: The Preclinical Pharmacology Core has experience in conducting pharmacokinetic analysis by various routes (IP, IV, PO). The Core will conduct these studies for experimental compounds after evaluation of in vitro metabolic stability or for more established compounds with guidance from the literature. 

Typical Assay Conditions

  • Eight time points.
  • CD-1 and Sprague Dawley strains are typically utilized but other strains are available upon request.
  • Drug levels in plasma and requested tissues can be evaluated. 
  • Pharmacokinetic parameters are typically calculated using the noncompartmental analysis tool from the industry standard PK modeling software, Phoenix WinNonlin (Pharsight Corp.). Compartmental analysis can be conducted upon demand.     

Sample Requirements: 10-20 mg of powder

Prerequisite: evaluation of in vitro metabolic stability and formulation development for new chemical entities (NCEs) or literature references for previously evaluated compounds

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