Ex-vivo perfusion bioassay: an excellent technique to measure the bioactivity of inhalable insulin coated microcrystals

Alistair Ross, Howard Stevens, Johann Partridge, Barry Moore, Maria Flores, Marie-Claire Parker, Amanda J. Brown, Chris Hillier, Jamie Coleman

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Abstract

Purpose: To measure the bioactivity of inhalable insulin coated microcrystals using a perfusion bioassay that measures its vasodilatory effect on smooth muscle arterial tissue.
Methods: The bioactivity of an insulin protein coated microcrystal (PCMC), a potential candidate for pulmonary drug delivery and commercial insulin was determined on a Danish Myo Tech P110 pressure myograph system. 12 week old Mesenteric resistance arteries from Male Wistar rats were isolated and
immersed in a physiological salt solution (PSS) and attached to 2 opposing hollow glass micro-cannula (outer diameter 80 microns). The PSS was gradually warmed to 37°C (at a pressure less than 5mm Hg) for 1hr. Subsequently the pressure was
increased up to 40mm Hg over a period 15 minutes and equilibrated for a further 15 minutes after gassing with 95%O2 / 5%CO2 to achieve a pH of 7.4 at 37°C. After normalisation by two washes of 123mM KCl and exposure to 1-10mM
noradrenaline the arteries were exposed intraluminally to each insulin preparation by gradual infusion directly into the lumen via a fetal microcannulae inserted to the tip of the glass mounting cannula, at a constant pressure.
Results: The preliminary results (full cummulative response curve yet to be determined) demonstrate insulin mediated relaxation to noradrenaline
preconstriction. The level of constriction drops from 100% to 42% as the concentration of insulin increases from -11 to -9 Log M for the PCMC compared with a drop from 100 % to 65% for the commercial insulin preparation. However the more potent vasodilatory effect found for the insulin PCMC is more likely to be a result of variance introduced in each dilution step than a real increase in potency.
Conclusion: The perfusion bioassay technique provides an excellent method of
measuring insulin bioactivity and indicates the insulin loaded on the microcrystal support is fully active.

Original languageEnglish
PagesM1064
Publication statusPublished - 8 Nov 2002
Event2002 AAPS Annual Meeting and Exposition - Toronto, Canada
Duration: 8 Nov 200214 Nov 2002

Conference

Conference2002 AAPS Annual Meeting and Exposition
CountryCanada
CityToronto
Period8/11/0214/11/02

Keywords

  • perfusion bioassay
  • insulin coated microcrystals

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    Ross, A., Stevens, H., Partridge, J., Moore, B., Flores, M., Parker, M-C., ... Coleman, J. (2002). Ex-vivo perfusion bioassay: an excellent technique to measure the bioactivity of inhalable insulin coated microcrystals. M1064. Poster session presented at 2002 AAPS Annual Meeting and Exposition, Toronto, Canada.