Development of a more rapid, reduced serum culture system for Caco-2 monolayers and application to the biopharmaceutics classification system.

The objectives were: (1) to develop a more rapid, reduced serum culture system for Caco-2 monolayers, relative to the traditional 21-day, 10% fetal bovine serum (FBS) system; and (2) to determine the biopharmaceutical drug classification of an oral therapeutic agent using this new system. Caco-2 cells were grown in the six well format on polycarbonate filters, in medium containing 2% iron supplemented calf serum (sCS) and a combination of growth factors and hormones. After 4 days in culture, permeabilities of three marker compounds (metoprolol, mannitol, and taurocholate) across monolayers were determined, and compared to permeabilities from the traditional 21-day, 10% FBS system, using cells at similar passage number. Cell morphology, degree of cell differentiation, and the presence of two efflux pumps were assessed. The 2% sCS model was also used to classify the permeability of an oral therapeutic agent as high or low. No difference in permeability was observed for metoprolol transport (P = 0.38) between the two culture methods, and the values obtained were independent of passage number of the cells. Mannitol permeability was about 2-fold higher from the 2% sCS system, as compared to the 10% FBS system. Taurocholate permeability was low indicating the 2% sCS culture at 4 days lacked this particular active transporter capability. Electron micrographs of cells grown in the 2% sCS system at 4 days revealed the presence of microvilli and tight junctions. P-glycoprotein and an efflux pump for furosemide were functionally present. The 2% sCS system indicated the oral therapeutic agent as highly permeable, which agreed with the 10% FBS system. This new system provides a rapid, accurate, and economical option for passive permeability determination, and appears to be applicable to the proposed Biopharmaceutics Classification System (BCS).

Use of stable isotopes for evaluation of drug delivery systems: comparison of ibuprofen release in vivo and in vitro from two biphasic release formulations utilizing different rate-controlling polymers.

Certain delivery systems are intended to release the active ingredient in different phases to obtain the desired therapeutic effect. For these formulations, such as a bilayer tablet, it is desirable to distinguish and measure the release of drug from the different phases simultaneously. Mass spectrometric methods were developed to measure three ibuprofen isotopomers in serum and two in dissolution fluid. The analytical methods were linear (r > or = 0.992) over the concentration range of interest and recovery was greater than 99.2% for all isotopomers. Coadministration of [2H0]ibuprofen, [2H4]ibuprofen, and [2H7]ibuprofen to male beagles demonstrated that the isotopomers were bioequivalent and verified the absence of any kinetic isotope effect due to deuterium incorporation (p = 0.286). These methods were then used to evaluate a bilayer tablet formulation composed of an immediate release layer of 100 mg [2H4]ibuprofen and a sustained release layer with a drug load of 300 mg [2H0]ibuprofen. Two different rate-controlling polymer matrices that provided similar in vitro dissolution profiles were compared in the sustained release phase, while the immediate release formulation remained the same. In male beagles, the HPMC matrix delivered a significantly greater amount of ibuprofen (p < 0.05). The AUC was threefold greater for HPMC (1067 +/- 437 nmole*h/ml) versus EUDRAGIT (320 +/- 51), and Cmax was nearly four times greater (145 +/- 62.1 nmole/ml for HPMC versus 37.9 +/- 14.4 for EUDRAGIT).(ABSTRACT TRUNCATED AT 250 WORDS)

Investigation into substrate cracking of a film-coated bilayered tablet.

Substrate cracking occurred during film coating of a bilayered tablet. The cause was traced to differences in the expansion characteristics of the two layers upon exposure to heat. Thermal mechanical analysis was used to determine the coefficients of thermal expansion of the respective layers.