Steven C. Sutton

Comparison of the Gravimetric, Phenol Red, and 14C-PEG-3350 Methods to Determine Water Absorption in the Rat Single-Pass Intestinal Perfusion Model

This study was undertaken to determine whether the gravimetric method provided an accurate measure of water flux correction and to compare the gravimetric method with methods that employ nonabsorbed markers (eg, phenol red and 14C-PEG-3350). Phenol red, 14C-PEG-3350, and 4-[2-[[2-(6-amino-3-pyridinyl)-2-hydroxyethyl]amino]ethoxy]-methyl ester, (R)-benzene acetic acid (Compound I) were co-perfused in situ through the jejunum of 9 anesthetized rats (single-pass intestinal perfusion [SPIP]). Water absorption was determined from the phenol red. 14C-PEG-3350, and gravimetric methods. The absorption rate constant (ka) for Compound I was calculated. Both phenol red and 14C-PEG-3350 were appreciably absorbed, underestimating the extent of water flux in the SPIP model. The average ±SD water flux (μg/h/cm) for the 3 methods were 68.9±28.2 (gravimetric), 26.8±49.2 (phenol red), and 34.9±21.9 (14C-PEG-3350). The (average±SD) ka for Compound I (uncorrected for water flux) was 0.024±0.005 min−1. For the corrected, gravimetric method, the average±SD was 0.031±0.001 min−1. The gravimetric method for correcting water flux was as accurate as the 2 “nonabsorbed” marker methods.

Fed and fasted gastric pH and gastric residence time in conscious beagle dogs.

The gastric pH values are controversial in the literature. Some suggest the dog gastric pH is higher than human and dog gastric pH after fed with particular diet is uncertain. Gastric pH in 16 male beagle dogs was measured using Bravo pH telemetry system. For the fed study, the dogs received 10 or 200 g of dog dry food (5L18) 15 min before dosing the Bravo pH capsule, followed by a 50 mL of water to aid in swallowing. It was surprising to find a small, but statistically significantly lower pH in the fed compared to the fasted stomach. The average gastric pH in fasted dogs was 2.05 and 1.08 and 1.26 for 10 and 200 g fed dogs. The average gastric emptying time of the capsule was 1.4, 9.4 and 20 h for fasted, 10 g fed and 200 g fed dogs, respectively. The inter-individual variability was higher in fasted dogs than in fed dogs. The results showed the gastric pH in each colony of dogs can be different from reported values in the literature. It emphasizes that the importance of measuring the pH in each colony when dogs are used to evaluate pharmacokinetics of pH sensitive drugs or formulations.

Relationship Between Drug Absorption Enhancing Activity and Membrane Perturbing Effects of Acylcarnitines

Acylcarnitines with chain lengths of 2 to 18 carbon atoms were tested for their effects on rat intestinal brush border membrane order (S) by fluorescence polarization of 1,6-diphenyl-l ,3,5-hexatriene (DPH). These results were compared to the previously reported effectiveness of the acylcarnitines as absorption enhancers of the poorly absorbed antibiotic cefoxitin. Acylcarnitines with fatty acids less than 12 carbon units in length were ineffective in increasing drug absorption and perturbing brush border membrane order. Long-chain acylcarnitines (12–18 carbons) significantly increased the bioavailability of cefoxitin and decreased the lipid order of brush border membranes. The results suggest that, in order to promote drug absorption, the acylcarnitines must surpass a critical chain length (10 carbon units) to partition effectively into the membrane and, in addition, must perturb the lipid order beyond a threshold value (15–20%). Membrane perturbing capacity may serve as an indicator of the absorption enhancing potential of other aliphatic-type compounds.

In vitro models for selection of development candidates. Permeability studies to define mechanisms of absorption enhancement

Considerable attention has been given to alternative strategies for improving the absorption of peptides and hydrophilic drugs across the intestinal epithelium. One approach to overcoming this restriction is to co-administer drugs with absorption enhancers. Although in many cases, improvements in drug absorption by enhancers have been ascribed to mucosal damage, it now appears that enhancement can be separated from damage by manipulating the concentration and exposure of the epithelium to these agents. There are a number of agents that increase mucosal permeability in a reversible manner and without overt alterations to the intestinal mucosa. Increasing evidence suggests the most promising enhancers alter paracellular rather than transcellular permeability of the mucosal epithelium. Several classes of agents that have been tested as absorption promoters are discussed, including experimental evidence as to their mechanism of action and toxicity. The structural and functional features of cell tight junctions are also addressed, particularly, as they are involved in the regulation of paracellular permeability. In addition, results from studies performed using a variety of in vitro model systems are presented that have helped elucidate the specific cellular mechanisms of absorption enhancement. The future prospects of absorption enhancers is promising. However, a number of safety concerns and formulation design issues must be considered before the application of absorption enhancers to routine oral drug delivery in humans can be realized.

Simultaneous in vitro measurement of intestinal tissue permeability and transepithelial electrical resistance (TEER) using Sweetana-Grass diffusion cells.

A simple modification of the commercially available Sweetana–Grass (S-G) side-by-side diffusion cells, allowing the simultaneous measurement of tissue permeability and transepithelial electrical resistance (TEER), has been described and validated for rat excised, muscle-free intestinal tissue. The TEER-lowering effects of a series of acylcarnitines were shown to be correlated with previously reported in vitro (i.e., membrane perturbation) and in vivo (i.e., absorption enhancement) activity. The TEER-lowering effect of palmitoyl carnitine chloride (PCC) was also shown to be reversible. The effects of PCC on TEER and the permeability of poorly absorbed compounds (cefoxitin and lucifer yellow) were simultaneously determined. Compared to controls (mannitol-treated), PCC immediately produced a rapid drop in colon TEER. By 5 min post-PCC addition, colon TEER was 50% of control; by 10 min post-PCC addition, colon TEER was 17% of control. After a lag of about 5–10 min post-PCC addition, the cefoxitin or lucifer yellow permeability coefficient increased more than 20-fold. The modified S-G cells provide a simple and reproducible method whereby flux and TEER can be simultaneously determined, providing a valuable link between the effect of absorption enhancers on TEER measurements and the increased permeability of poorly absorbed compounds.

A computational model for particle size influence on drug absorption during controlled-release colonic delivery.

The effect of particle size on the percent drug absorbed is computationally modeled for controlled-release dosage forms that deliver drug particles to the colon. The relative benefit of reducing particle size is mapped on a diagram of the drugs absorption rate constant (estimated from rat intestinal perfusion, CACO-2 or human intubation permeation rates) versus the drugs solubility. Some drugs fall into a limit of high percentage absorption even with large particles such that particle size reduction has little impact. Another group of drugs is solubility limited such that even with small particles, absorption is negligible. Between the two regions, only drugs with sufficiently high absorption rates are influenced by the drug dissolution rate and thereby the particle size. The size of this region is a function of dosing rate. Comparisons between calculated particle size effects on colon absorption as a function of colon volume suggest caution when using animal models to predict bioavailability from colonic drug delivery. This volume dependence also suggests that the particle size influence will vary as a function of the digestive cycle.

Solid nanocrystalline dispersions of ziprasidone with enhanced bioavailability in the fasted state.

Reducing the absorption difference between fed and fasted states is an important goal in the development of pharmaceutical dosage forms. The goal of this work was to develop and characterize a solid nanocrystalline dispersion (SNCD) to improve the oral absorption of ziprasidone in the fasted state, thereby reducing the food effect observed for the commercial formulation. A solution of ziprasidone hydrochloride and the polymer hydroxypropyl methylcellulose acetate succinate (HPMCAS) was spray-dried to form a solid amorphous spray-dried dispersion (SDD), which was then exposed to a controlled temperature and relative humidity (RH) to yield the ziprasidone SNCD. The SNCD was characterized using powder X-ray diffraction, thermal analysis, microscopy, and in vitro dissolution testing. These tools indicate the SNCD consists of a high-energy crystalline form of ziprasidone in domains approximately 100 nm in diameter but with crystal grain sizes on the order of 20 nm. The SNCD was dosed orally in capsules to beagle dogs. Pharmacokinetic studies showed complete fasted-state absorption of ziprasidone, achieving the desired improvement in the fed/fasted ratio.

Predicting injection site muscle damage I : Evaluation of immediate release parenteral formulations in animal models

AbstractPurpose. The goal of this study was to find a resource sparing alternative to the rabbit lesion model (RbLV) for assessing injection site toleration in extended release (ER) intramuscular (IM) formulation screening. Methods. ER formulations (danofloxacin oily and aqueous suspensions) were evaluated in RbLV, rat and rabbit plasma creatine phospho-kinase (CK), and rat foot edema (RFE) models as described in the companion article. Results. None of the short term models could consistently predict acute and chronic effects of the. For example, RFE predicted little muscle damage from aqueous vehicle (0.03 ± 0.03 g) and 60 mg/ml (0.08 ± 0.03 g) formulation; while RbLVdays l–3 was marked and greater (p < 0.05) for 60 mg/ml (6.0 ± 3.1) than vehicle (2.2 ± 2.9) formulations. Furthermore, RbLVdays l–3 for vehicle (6.5 ± 7.5) and 60 mg/ml (4.9 ± 4.6) danofloxacin oily formulations were worse (p < 0.05) than oil alone (1.4 ± 2.2); an observation not predicted by CK models, since they apparently reflected only the acute muscle damage of formulation components immediately available to surrounding tissue at the time of injection. Conclusions. The CK models may be useful to screen those ER formulations with unacceptable acute damage due to immediately available components. However, to evaluate potential delayed effects from ER formulations, the long-term model RbLV was still recommended.

Enhanced Bioavailability of Cefoxitin Using Palmitoyl L-Carnitine. I. Enhancer Activity in Different Intestinal Regions

The conditions under which the absorption enhancer palmitoyl L-carnitine chloride (PCC) improved the bioavailability of the poorly absorbed antibiotic cefoxitin throughout the rat intestine has been studied. Cefoxitin alone was appreciably absorbed only in the duodenum (31% vs <7% elsewhere). PCC solutions (3 mg/rat, pH 4.0) enhanced cefoxitin bioavailability (F) by 0-, 22-, 16-, and > 32-fold in the duodenum, jejunum, ileum, and colon regions, respectively. The inability of PCC to improve F in the duodenum could not likely be attributed to enzymatic degradation of the enhancer, since coadmin-istration with protease and esterase inhibitors produced similar results (F = 30%). Coadministration of PCC solution with cefoxitin in the unligated or ligated colon, increased F to 33 and 76%, respectively. Qualitatively similar results were seen with PCC suspensions (3 mg/rat, pH 6.0). Maintaining a high concentration of cefoxitin and PCC in a restricted region (i.e., by ligating a 2- to 3-cm section of the colon) afforded a two- to threefold advantage over an unligated colon section. The difference in cefoxitin bioavailability between ligated and unligated colon was probably due to sample spreading and subsequent/simultaneous dilution.

Enhanced Bioavailability of Cefoxitin Using Palmitoylcarnitine. II. Use of Directly Compressed Tablet Formulations in the Rat and Dog

The performance of tablets containing the absorption enhancer palmitoylcarnitine chloride (PCC) and the antibiotic cefoxitin (CEF) was determined by direct placement of tablets in the rat stomach, small intestine, and colon. While the bioavailability (F) of tablets containing 12 mg CEF without PCC ranged from 0.6 to 3.9%, the addition of 24 mg PCC resulted in an enhanced CEF bioavailability in the rat colon (mean ± SD: F = 57 ± 19%) and rat jejunum (F = 71 ± 16%) but not in the rat stomach. Following oral administration to dogs, tablets of 200 mg CEF without or with 600 mg PCC resulted in the same low bioavailabilities (7.0 ± 10.3 and 7.0 ± 3.6%, respectively). However, when these tablets were enteric coated, PCC improved CEF bioavailability from 2.44 ± 1.84 to 29.0 ± 13.4%. Therefore, the use of enteric-coated direct compressed tablets containing PCC and direct compression excipients improved the peroral bioavailability of a poorly absorbed compound.