RESEARCH ARTICLESA Systematic Examination of the In Vitro Ussing Chamber and the In Situ Single-Pass Perfusion Model Systems in Rat Ileum Permeation of Model Solutes
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INTRODUCTION
The in vitro Ussing chamber system using an animal mucosal membrane mounted in the diffusion cells to evaluate the absorption characteristics of new drugs at the membrane level has been gaining in popularity in recent years.1., 2., 3., 4. In situ methods in animal models have also been used to determine absorption mechanisms.5., 6., 7. It has been reported that the in vitro permeability of drugs is lower than that in vivo. Lennernas et al.1 suggested that the differences in quantitative
Chemicals and Reagents
For the in vitro Ussing chamber system, a trace amount of radiolabeled permeants was added to pH 7.4 Krebs-bicarbonate Ringer's solution (KBR). The KBR had the following composition (mM): 108.0 NaCl, 4.7 KCl, 1.8 Na2HPO4, 0.4 NaH2PO4, 15.0 NaHCO3, 1.2 MgSO4, 1.25 CaCl2, 11.5 D-glucose. For the in situ system, modified Krebs-Ringer's solution (MKBR) and pH 7.4 phosphate buffer solution were used as the medium. The phosphate buffer solution had the following composition (mM): 57 NaH2PO4, 79 Na2SO4.
Absorption of Steroid Series
In situ absorption studies were systematically performed with a series of steroids varying in lipophilicity, but similar in molecular size. The isoosmotic perfusion solution contained [3H]steroid, phenol red, and MKBR at pH 7.4. Figure 2 shows the fraction of [3H]ED remaining in the perfused intestinal segment, C(ℓ)/C(0), with time and concurrent appearance in blood with time. The condition of steady-state was achieved in 20 min. Thereafter, C(ℓ)/C(0) reached a plateau and the absorption rate in
SIGNIFICANCE
A systematic and mechanistic approach has been applied to understand and quantify the rate-determining factors and barriers in the in situ and in vitro intestinal absorption of a common series of steroids in the rat ileum. Experimental strategies were gainfully employed to define the mass transfer resistances of the ABL, mucosal epithelium, and the submucosal tissue. TC absorption in the ileum had a key role in establishing the permeability of the ABL. Whereas both the in situ and in vitro
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