The use of capillary high performance liquid chromatography with electrospray mass spectrometry for the analysis of small volume blood samples from serially bled mice to determine the pharmacokinetics of early discovery compounds.
The methodology described demonstrates, for the first time, the feasibility of performing pharmacokinetic studies in the serially bled mouse model to support the early development of discovery compounds. Sample analysis, using capillary high performance liquid chromatography combined with tandem mass spectrometry, has facilitated the achievement of this milestone and has successfully been applied to determine pharmacokinetic information following both intravenous and oral administration of a single discovery compound. The methodologies described demonstrate potential for a reduction in the amount of new chemical entity required to undertake pharmacokinetic studies. Typically, such studies are performed in larger rodents with a significantly increased body mass (ten times in the case of the rat) and therefore it follows that to undertake the same experiment in the mouse would require ten times less compound to effect an equivalent dose. Conventionally, pharmacokinetic studies to obtain both intravenous and oral information, e.g. clearance and half-life, and the resultant bioavailability have been performed using two parallel groups of rodents, collecting blood by exsanguination, separating off the plasma and analysing this using conventional liquid chromatography/tandem mass spectrometry. The use of capillary high performance liquid chromatography (HPLC) has facilitated the analysis of small volume blood samples by increasing the effective sensitivity of the analytical method. Consequently, we have established a protocol for serially bleeding mice thus reducing the number of animals and so further reducing the amount of compound required for such experiments. This paper reports data obtained from collected and processed blood volumes of <20 microL with the subsequent injection of only 1 microL of precipitated extract onto a capillary column.
Fraser
I J
IJ
Department of International Development Support, BioMet, GlaxoWellcome Research and Development, Park Road, Ware, Herts. SG12 0DP, UK.
Dear
G J
GJ
Plumb
R
R
L'Affineur
M
M
Fraser
D
D
Skippen
A J
AJ
D000284:Administration, Oral; D000818:Animals; D001572:Benzofurans / Q000008:administration & dosage / Q000097:blood / Q000493:pharmacokinetics*; D001774:Blood Chemical Analysis / Q000379:methods*; D001800:Blood Specimen Collection / Q000379:methods; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods; D007275:Injections, Intravenous; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D010599:Pharmacokinetics; D051381:Rats; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation / Q000379:methods
D001572:Benzofurans; C403054:GW 290569