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

Ross, Alistair and Stevens, Howard and Partridge, Johann and Moore, Barry and Flores, Maria and Parker, Marie-Claire and Brown, Amanda J. and Hillier, Chris and Coleman, Jamie (2002) Ex-vivo perfusion bioassay : an excellent technique to measure the bioactivity of inhalable insulin coated microcrystals. In: 2002 AAPS Annual Meeting and Exposition, 2002-11-08 - 2002-11-14.

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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.

    ORCID iDs

    Ross, Alistair, Stevens, Howard, Partridge, Johann, Moore, Barry ORCID logoORCID: https://orcid.org/0000-0002-4943-1632, Flores, Maria, Parker, Marie-Claire, Brown, Amanda J., Hillier, Chris and Coleman, Jamie;