Organic Anion-Transporting Polypeptide 1B1/1B3-Mediated Hepatic Uptake Determines the Pharmacokinetics of Large Lipophilic Acids: In Vitro-In Vivo Evaluation in Cynomolgus Monkey.

It is generally presumed that uptake transport mechanisms are of limited significance in hepatic clearance for lipophilic or high passive-permeability drugs. In this study, we evaluated the mechanistic role of the hepato-selective organic anion-transporting polypeptides (OATPs) 1B1/1B3 in the pharmacokinetics of compounds representing large lipophilic acid space. Intravenous pharmacokinetics of 16 compounds with molecular mass ∼400-730 Da, logP ∼3.5-8, and acid pKa <6 were obtained in cynomolgus monkey after dosing without and with a single-dose rifampicin-OATP1B1/1B3 probe inhibitor. Rifampicin (30 mg/kg oral) significantly (P < 0.05) reduced monkey clearance and/or steady-state volume of distribution (VDss) for 15 of 16 acids evaluated. Additionally, clearance of danoprevir was reduced by about 35%, although statistical significance was not reached. A significant linear relationship was noted between the clearance ratio (i.e., ratio of control to treatment groups) and VDss ratio, suggesting hepatic uptake contributes to the systemic clearance and distribution simultaneously. In vitro transport studies using primary monkey and human hepatocytes showed uptake inhibition by rifampicin (100 µM) for compounds with logP ≤6.5 but not for the very lipophilic acids (logP > 6.5), which generally showed high nonspecific binding in hepatocyte incubations. In vitro uptake clearance and fraction transported by OATP1B1/1B3 (ft,OATP1B) were found to be similar in monkey and human hepatocytes. Finally, for compounds with logP ≤6.5, good agreement was noted between in vitro ft,OATP1B and clearance ratio (as well as VDss ratio) in cynomolgus monkey. In conclusion, this study provides mechanistic evidence for the pivotal role of OATP1B-mediated hepatic uptake in the pharmacokinetics across a wide, large lipophilic acid space. SIGNIFICANCE STATEMENT: This study provides mechanistic insight into the pharmacokinetics of a broad range of large lipophilic acids. Organic anion-transporting polypeptides 1B1/1B3-mediated hepatic uptake is of key importance in the pharmacokinetics and drug-drug interactions of almost all drugs and new molecular entities in this space. Diligent in vitro and in vivo transport characterization is needed to avoid the false negatives often noted because of general limitations in the in vitro assays while handling compounds with such physicochemical attributes.

Investigation of pH Influence on Skin Permeation Behavior of Weak Acids Using Nonsteroidal Anti-Inflammatory Drugs.

As a continuing effort to understand the skin permeation behavior of weak acids and bases, the objectives of the present study were to evaluate skin permeation of nonsteroidal anti-inflammatory drugs (NSAIDs) under the influence of pH, investigate the mechanism of pH effect, and examine a previous hypothesis that the effective skin pH for drug permeation is different from donor solution pH. In vitro permeability experiments were performed in side-by-side diffusion cells with diclofenac, ibuprofen, flurbiprofen, ketoprofen, and naproxen and human skin. The donor solution pH significantly affected skin permeation of NSAIDs, whereas no effect of the receiver pH was observed. Similar to previous observations, the apparent permeability coefficient versus donor solution pH relationships deviated from the predictions (fractions of unionized NSAIDs) according to the acid/base theory. The influences of the viable epidermis barrier, polar pathway transport, ion permeation across skin, and effective skin pH were investigated. The effective pH values for skin permeation determined using the NSAIDs (weak acids) in this study were different from those obtained previously with a weak base at the same donor solution pH conditions, suggesting that the observed permeability-pH relationships could not be explained solely by possible pH differences between skin and donor solution.

Optimal choice of pH for toxicity and bioaccumulation studies of ionizing organic chemicals.

It is recognized that the pH of exposure solutions can influence the toxicity and bioaccumulation of ionizing compounds. The present study investigates whether it can be considered a general rule that an ionizable compound is more toxic and more bioaccumulative when in the neutral state. Three processes were identified to explain the behavior of ionizing compounds with changing pH: the change in lipophilicity when a neutral compound becomes ionized, electrical attraction, and the ion trap. The literature was screened for bioaccumulation and toxicity tests of ionizing organic compounds performed at multiple pH levels. Toxicity and bioconcentration factors (BCFs) were higher for acids at lower pH values, whereas the opposite was true for bases. The effect of pH was most pronounced when pH - pK(a) was in the range of -1 to 3 for acids, and -3 to 1 for bases. The factor by which toxicity and BCF changed with pH was correlated with the lipophilicity of the compound (log K(OW) of the neutral compound). For both acids and bases, the correlation was positive, but it was significant only for acids. Because experimental data in the literature were limited, results were supplemented with model simulations using a dynamic flux model based on the Fick-Nernst-Planck diffusion equation known as the cell model. The cell model predicts that bases with delocalized charges may in some cases show declining bioaccumulation with increasing pH. Little information is available for amphoteric and zwitterionic compounds; however, based on simulations with the cell model, it is expected that the highest toxicity and bioaccumulation of these compounds will be found where the compounds are most neutral, at the isoelectric point.

Particle size reduction and pharmacokinetic evaluation of a poorly soluble acid and a poorly soluble base during early development.

AIM: The aim of the present study was to find out if nanosuspensions were a better choice compared with microsuspensions, for the present substances with water solubility in the order of 2-3 µM (pH 6.8, small intestinal pH) and no permeability limitations. The ambition was also to understand what the higher solubility in the stomach for BA99 means in terms of absorption properties of the substance. METHOD: The pharmacokinetic parameters of a poorly soluble acid (AC88) and a poorly soluble base (BA99) administered orally as nanosuspensions have been compared with those from microsuspensions using rat as in vivo species. RESULTS: A significant difference was observed between the two suspensions for AC88 already at the lowest dose, 5 µmol/kg (the particle size of the nanosuspensions and the microsuspensions was about 200 nm and 14 µm, respectively). These results were further confirmed at a high dose (500 µmol/kg). However, for BA99, there were no significant differences between the two formulations at any dose investigated (the particle size of the nanosuspensions and the microsuspensions was about 280 nm and 12 µm, respectively). CONCLUSIONS: The study demonstrated a clear correlation between particle size and in vivo exposures for an acidic compound, the nanosuspensions providing the highest exposure. For a basic compound, on the other hand, with the present properties and doses, a microsuspension was sufficient. In the latter case, the higher solubility at gastric pH, because of the basic pK(a), limits the need for particle reduction.

Comparison of acid clearance of noncarbonated and carbonated soft drinks in the mouth.

The aim of this study was to measure differences in oral pH after subjects drank carbonated and noncarbonated acidic soft drinks. Oral pH was measured in 17 subjects at 4 oral sites with beetrode electrodes after each subject drank 50 mL of noncarbonated and carbonated blackcurrant beverages. There were no statistical differences for either beverage regarding the time taken to recover a pH of 5.5, 6.0, and 6.5. Oral pH recovered to 6.5 within 90 seconds with both drinks. The median lowest pH recorded after 30 seconds at the 4 sites for both drinks was > pH 4.9, but there was no statistical difference in the clearance between the drinks. The implication may be that carbonation is not directly responsible for the erosive potential of different beverages.

Ouabain-sensitive bicarbonate secretion and acid absorption by the marine teleost fish intestine play a role in osmoregulation.

The gulf toadfish (Opsanus beta) intestine secretes base mainly in the form of HCO3- via apical anion exchange to serve Cl- and water absorption for osmoregulatory purposes. Luminal HCO3- secretion rates measured by pH-stat techniques in Ussing chambers rely on oxidative energy metabolism and are highly temperature sensitive. At 25 degrees C under in vivo-like conditions, secretion rates averaged 0.45 micromol x cm(-2) x h(-1), of which 0.25 micromol x cm(-2) x h(-1) can be accounted for by hydration of endogenous CO2 partly catalyzed by carbonic anhydrase. Complete polarity of secretion of HCO3- and H+ arising from the CO2 hydration reaction is evident from equal rates of luminal HCO3- secretion via anion exchange and basolateral H+ extrusion. When basolateral H+ extrusion is partly inhibited by reduction of serosal pH, luminal HCO3- secretion is reduced. Basolateral H+ secretion occurs in exchange for Na+ via an ethylisopropylamiloride-insensitive mechanism and is ultimately fueled by the activity of the basolateral Na+-K+-ATPase. Fluid absorption by the toadfish intestine to oppose diffusive water loss to the concentrated marine environment is accompanied by a substantial basolateral H+ extrusion, intimately linking osmoregulation and acid-base balance.

Poly(amidoamine) (PAMAM) dendritic nanostructures for controlled site-specific delivery of acidic anti-inflammatory active ingredient.

The purpose of the investigation was to evaluate the potential of polyamidoamine (PAMAM) dendrimer as nanoscale drug delivery units for controlled release of water insoluble and acidic anti-inflammatory drug. Flurbiprofen (FB) was selected as a model acidic anti-inflammatory drug. The aqueous solutions of 4.0 generation (G) PAMAM dendrimer in different concentrations were prepared and used further for solubilizing FB. Formation of dendrimer complex was characterized by Fourier transform infrared spectroscopy. The effect of pH on the solubility of FB in dendrimer was evaluated. Dendrimer formulations were further evaluated for in vitro release study and hemolytic toxicity. Pharmacokinetic and biodistribution were studied in male albino rats. Efficacy of dendrimer formulation was tested by carrageenan induced paw edema model. It was observed that the loaded drug displayed initial rapid release (more than 40% till 3rd hour) followed by rather slow release. Pharmacodynamic study revealed 75% inhibition at 4th hour that was maintained above 50% till 8th hour. The mean residence time (MRT) and terminal half-life (THF) of the dendritic formulation increased by 2-fold and 3-fold, respectively, compared with free drug. Hence, with dendritic system the drug is retained for longer duration in the biosystem with 5-fold greater distribution. It may be concluded that the drug-loaded dendrimers not only enhanced the solubility but also controlled the delivery of the bioactive with localized action at the site of inflammation.

Polyethylenimine with acid-labile linkages as a biodegradable gene carrier.

Polyethylenimine (PEI) is a gene carrier with high transfection efficiency. However, PEI has high cytotoxicity, which depends on its molecular weight. To reduce the cytotoxicity, degradable PEIs with acid-labile imine linkers were synthesized with low molecular weight PEI1.8K (1.8 kDa) and glutadialdehyde. The molecular weights of the synthesized acid-labile PEIs were 23.7 and 13 kDa, respectively. The half-life of the acid-labile PEI was 1.1 h at pH 4.5 and 118 h at pH 7.4, suggesting that the acid-labile PEI may be rapidly degraded into nontoxic low molecular weight PEI in acidic endosome. In a gel retardation assay, plasmid DNA (pDNA) was completely retarded at a 3:1 N/P (nitrogen of polymer/phosphate of DNA) ratio. The zeta potential of the polyplexes was in the range of 46.1 to 50.9 mV and the particle size was in the range of 131.8 to 164.6 nm. In vitro transfection assay showed that the transfection efficiency of the acid-labile PEIs was comparable to that of PEI25K. In toxicity assay, the acid-labile PEI was much less toxic than PEI25K, due to the degradation of acid-labile linkage. Therefore, the acid-labile PEIs may be useful for the development of a nontoxic polymeric gene carrier.

Altered expression of renal acid-base transporters in rats with lithium-induced NDI.

Prolonged lithium treatment of humans and rodents often results in hyperchloremic metabolic acidosis. This is thought to be caused by diminished net H+ secretion and/or excessive back-diffusion of acid equivalents. To explore whether lithium treatment is associated with changes in the expression of key renal acid-base transporters, semiquantitative immunoblotting and immunocytochemistry were performed using kidneys from lithium-treated (n = 6) and control (n = 6) rats. Rats treated with lithium for 28 days showed decreased urine pH, whereas no significant differences in blood pH and plasma HCO3- levels were observed. Immunoblot analysis revealed that lithium treatment induced a significant increase in the expression of the H+-ATPase (B1-subunit) in cortex (190 +/- 18%) and inner stripe of the outer medulla (190 +/- 9%), and a dramatic increase in inner medulla (900 +/- 104%) in parallel to an increase in the expression of type 1 anion exchanger (400 +/- 40%). This was confirmed by immunocytochemistry and immunoelectron microscopy, which also revealed increased density of intercalated cells. Moreover, immunoblotting and immunocytochemistry revealed a significant increase in the expression of the type 1 electrogenic Na+-HCO3- cotransporter (NBC) in cortex (200 +/- 23%) and of the electroneutral NBCn1 in inner stripe of the outer medulla (250 +/- 54%). In contrast, there were no changes in the expression of Na+/H+ exchanger-3 or of the Cl-/HCO3- exchanger pendrin. These results demonstrate that the expression of specific renal acid-base transporters is markedly altered in response to long-term lithium treatment. This is likely to represent direct or compensatory effects to increase the capacity for HCO3- reabsorption, NH4+ reabsorption, and proton secretion to prevent the development of systemic metabolic acidosis.

Intracellular proton mobility and buffering power in cardiac ventricular myocytes from rat, rabbit, and guinea pig.

Intracellular pH (pHi) is an important modulator of cardiac function. The spatial regulation of pH within the cytoplasm depends, in part, on intracellular H+ (Hi+) mobility. The apparent diffusion coefficient for Hi+, DHapp, was estimated in single ventricular myocytes isolated from the rat, guinea pig, and rabbit. DHapp was derived by best-fitting predictions of a two-dimensional model of H+ diffusion to the local rise of intracellular [H+], recorded confocally (ratiometric seminaphthorhodafluor fluorescence) downstream from an acid-filled, whole cell patch pipette. Under CO2/HCO3--free conditions, DHapp was similar in all three species (mean values: 8-12.5 x 10-7 cm2/s) and was over 200-fold lower than that for H+ in water. In guinea pig myocytes, DHapp was increased 2.5-fold in the presence of CO2/HCO3- buffer, in agreement with previous observations in rabbit myocytes. Hi+ mobility is therefore low in cardiac cells, a feature that may predispose them to the generation of pHi gradients in response to sarcolemmal acid/base transport or local cytoplasmic acid production. Low Hi+ mobility most likely results from H+ shuttling among cytoplasmic mobile and fixed buffers. This hypothesis was explored by comparing the pHi dependence of intrinsic, intracellular buffering capacity, measured for all three species, and subdividing buffering into mobile and fixed fractions. The proportion of buffer that is mobile will be the main determinant of DHapp. At a given pHi, this proportion appeared to be similar in all three species, consistent with a common value for DHapp. Over the pHi range of 6.0-8.0, the proportion is expected to change, predicting that DHapp may display some pHi sensitivity.

Direct measurement of acid permeation into rat oesophagus.

BACKGROUND AND AIMS: The early responses of the oesophageal mucosa to acid perfusion may predict subsequent pathology. Mucosal responses to luminal acid may result either from acid permeating through the mucosa or from other unknown transduction mechanisms. In order to better understand the dynamics of acid permeation into the oesophageal mucosa, we measured interstitial pH (pH(int)) of the oesophageal basal epithelial layer, pre-epithelial layer thickness, and blood flow in rats in vivo during luminal acid challenge. A novel confocal microscopic technique was used in vitro to measure pH(int) from defined cellular sites in response to luminal and basolateral acidification. METHODS: 5-(and-6)-Carboxyfluorescein (CF) and carboxy-seminapthorhodofluor-1 (SNARF-1) fluorescence was used to measure pH(int) by conventional and confocal microscopy, respectively, in urethane anaesthetised rats. Pre-epithelial layer thickness was measured optically with carbon particles as markers. Blood flow was measured with laser Doppler flowmetry. RESULTS: Luminal acidification failed to alter pH(int) in vivo and in vitro, but pH(int) was lowered by modest serosal acidification. Pre-epithelial layer thickness and blood flow increased significantly during luminal surface acid perfusion. Indomethacin had no effect on any acid related response. CONCLUSION: In this first dynamic measurement of oesophageal acid permeation and pre-epithelial layer thickness, pH(int) was preserved in spite of high luminal acidity by two complementary techniques. Despite the apparent permeability barrier to acid permeation, oesophageal blood flow and thickness responded to luminal acid perfusion.

Relevance of ineffective oesophageal motility during oesophageal acid clearance.

BACKGROUND: Oesophageal clearance of acid reflux consists of an initial volume clearance followed by neutralisation of the acidified mucosa by swallowed saliva (chemical clearance). Ineffective oesophageal motility (IOM), a frequent finding in patients with gastro-oesophageal reflux disease (GORD), has been claimed to underlie prolonged acid clearance by affecting oesophageal emptying and saliva transport. Intraluminal impedance allows non-radiological monitoring of movement of oesophageal liquids. AIMS: To evaluate the relevance of IOM during oesophageal volume and chemical clearance using combined pH impedance measurements. SUBJECTS: Impedance was validated with fluoroscopy to study volume clearance in three healthy subjects. Acid clearance tests were performed in 10 healthy subjects in the upright and supine positions, before and after oesophageal peristaltic disruption with sildenafil 50 mg. METHODS: After instillation of an acid bolus, simultaneous manometry, pH, and impedance were used to study oesophageal motility, chemical clearance, and volume clearance, respectively. RESULTS: Impedance allowed assessment of volume clearance accurately, showing a strong correlation with fluoroscopy (r(2)=0.89). Sildenafil provoked a graded impairment in oesophageal motility in healthy subjects without affecting saliva secretion. In the upright position, volume clearance was slightly prolonged only with severe IOM (>80% abnormal peristaltic sequences). In the supine position, severe IOM significantly prolonged chemical and volume clearance. Moderate IOM (30-80% abnormal peristalsis) had no effect. With normal peristalsis and moderate IOM, clearance times were similar in the upright and supine positions. Severe IOM however had a greater impact on clearance in the supine than in the upright position. CONCLUSION: Ineffective oesophageal motility has little effect on oesophageal clearance during upright acid reflux. With supine reflux, only severe IOM is associated with prolonged oesophageal clearance.

Dietary accumulation of perfluorinated acids in juvenile rainbow trout (Oncorhynchus mykiss).

Perfluorinated acids (PFAs) recently have emerged as persistent global contaminants after their detection in wildlife and humans from various geographic locations. The highest concentrations of perfluorooctane sulfonate are characteristically observed in high trophic level organisms, indicating that PFAs may have a significant bioaccumulation potential. To examine this phenomenon quantitatively, we exposed juvenile rainbow trout (Oncorhynchus mykiss) simultaneously to a homologous series of perfluoroalkyl carboxylates and sulfonates for 34 d in the diet, followed by a 41-d depuration period. Carcass and liver concentrations were determined by using liquid chromatography-tandem mass spectrometry, and kinetic rates were calculated to determine compound-specific bioaccumulation parameters. Depuration rate constants ranged from 0.02 to 0.23/d, and decreased as the length of the fluorinated chain increased. Assimilation efficiency was greater than 50% for all test compounds, indicating efficient absorption from food. Bioaccumulation factors (BAFs) ranged from 0.038 to 1.0 and increased with length of the perfluorinated chain; however, BAFs were not statistically greater than 1 for any PFA. Sulfonates bioaccumulated to a greater extent than carboxylates of equivalent perfluoroalkyl chain length, indicating that hydrophobicity is not the sole determinant of PFA accumulation potential and that the acid function must be considered. Dietary exposure will not result in biomagnification of PFAs in juvenile trout, but extrapolation of these bioaccumulation parameters to larger fish and homeothermic organisms should not be performed.

Bioconcentration and tissue distribution of perfluorinated acids in rainbow trout (Oncorhynchus mykiss).

Rainbow trout (Oncorhynchus mykiss) were exposed simultaneously to a homologous series of perfluoroalkyl carboxylates and sulfonates in a flow-through system to determine compound-specific tissue distribution and bioconcentration parameters for perfluorinated acids (PFAs). In general, PFAs accumulated to the greatest extent in blood > kidney > liver > gall bladder. Carboxylates and sulfonates with perfluoroalkyl chain lengths shorter than seven and six carbons, respectively, could not be detected in most tissues and were considered to have insignificant bioconcentration factors (BCFs). For detectable PFAs, carcass BCFs increased with increasing length of the perfluoroalkyl chain, ranging from 4.0 to 23,000, based on wet weight concentrations. Carboxylate carcass BCFs increased by a factor of eight for each additional carbon in the perfluoroalkyl chain between 8 and 12 carbons, but this relationship deviated from linearity for the longest PFA tested, possibly because of decreased gill permeability. In general, half-lives (3.9-28 d) and uptake rates (0.053-1.700 L/kg/d) also increased with increasing length of the perfluoroalkyl chain in all tissues. Sulfonates had greater BCFs, half-lives, and rates of uptake than the corresponding carboxylate of equal perfluoroalkyl chain length, indicating that hydrophobicity, as predicted by the critical micelle concentration, is not the only determinant of PFA bioaccumulation potential and that the acid function must be considered.

Regulation of AE2-mediated Cl- transport by intracellular or by extracellular pH requires highly conserved amino acid residues of the AE2 NH2-terminal cytoplasmic domain.

We reported recently that regulation by intracellular pH (pH(i)) of the murine Cl-/HCO(3)(-) exchanger AE2 requires amino acid residues 310-347 of the polypeptide's NH(2)-terminal cytoplasmic domain. We have now identified individual amino acid residues within this region whose integrity is required for regulation of AE2 by pH. 36Cl- efflux from AE2-expressing Xenopus oocytes was monitored during variation of extracellular pH (pH(o)) with unclamped or clamped pH(i), or during variation of pH(i) at constant pH(o). Wild-type AE2-mediated 36Cl- efflux was profoundly inhibited by acid pH(o), with a value of pH(o50) = 6.87 +/- 0.05, and was stimulated up to 10-fold by the intracellular alkalinization produced by bath removal of the preequilibrated weak acid, butyrate. Systematic hexa-alanine [(A)6]bloc substitutions between aa 312-347 identified the greatest acid shift in pH(o(50)) value, approximately 0.8 pH units in the mutant (A)6 342-347, but only a modest acid-shift in the mutant (A)6 336-341. Two of the six (A)6 mutants retained normal pH(i) sensitivity of 36Cl- efflux, whereas the (A)6 mutants 318-323, 336-341, and 342-347 were not stimulated by intracellular alkalinization. We further evaluated the highly conserved region between aa 336-347 by alanine scan and other mutagenesis of single residues. Significant changes in AE2 sensitivity to pH(o) and to pH(i) were found independently and in concert. The E346A mutation acid-shifted the pH(o(0) value to the same extent whether pH(i) was unclamped or held constant during variation of pH(o). Alanine substitution of the corresponding glutamate residues in the cytoplasmic domains of related AE anion exchanger polypeptides confirmed the general importance of these residues in regulation of anion exchange by pH. Conserved, individual amino acid residues of the AE2 cytoplasmic domain contribute to independent regulation of anion exchange activity by pH(o) as well as pH(i).

Characterization of the efflux transporter(s) responsible for restricting intestinal mucosa permeation of the coumarinic acid-based cyclic prodrug of the opioid peptide DADLE.

PURPOSE: To elucidate the efflux transporter(s) responsible for restricting the permeation of a coumarinic acid-based cyclic prodrug of the opioid peptide DADLE (CD) thorough Caco-2 cell monolayers. METHODS: The cellular permeability characteristics of CD were investigated using Caco-2 cells, Madin-Darby canine kidney-wild type II cells (MDCK-WT). MDCK cells transfected with the human MDR1 gene (MDCK-MDR1), and MDCK cells transfected with human MRP2 gene (MDCK-MRP2). These cells were grown as monolayers onto microporous membranes. The disappearance from the donor side and appearance on the receiver side of CD were monitored by HPLC. The substrate activity of CD for P-gp was determined by using GF120918. a known P-gp specific inhibitor. The substrate activity of CD for MRP2 was determined by using cyclosporin A (CsA), a known MRP2 and P-gp inhibitor. RESULTS: In Caco-2 cells, the ratio of the apparent permeability coefficients (Papp) of CD flux in the basolateral (BL) to apical (AP) direction vs. the flux in the AP-to-BL direction (Papp-BL-to-AP/Papp AP-to-BL) was 71. In the presence of GF120918 (2 microM), the Papp BL-to-AP/Papp AP-to-BL ratio was decreased to 16. In the presence of CsA (25 microM), the ratio was decreased to 5.6. In MDCK-WT. MDCK-MDR1, and MDCK-MRP2 cells, the Papp BL-AP/Papp AP-to-BL ratios of CD were 13, 35, and 22, respectively. CsA (25 microM) greatly decreased the Papp BL-P-AP/Papp AP-to-BL ratios in MDCK-WT and MDCK-MDR1 cells to 1.5 and 3.2, respectively. However, in MDCK-MRP2 cells. CsA (25 microM) decreased the ratio only to 11. A mixture of GF120918 (2 microM) and CsA (25 microM) decreased the Papp BL-to-AP/Papp AP-to-BL ratios of CD in MDCK-WT, MDCK-MDR1, and MDCK-MRP2 cells to 1.4, 2.7, and 5.4. respectively. CONCLUSIONS: These data suggest that CD is a good substrate for both P-gp and MRP2 and that the restricted permeation of this cyclic prodrug in Caco-2 cells and in the intestinal mucosa is probably due to its substrate activities for both of these efflux transporters.

Effect of carbonate salts on the kinetics of acid-catalyzed dimerization of adefovir dipivoxil.

PURPOSE: The chemical stability and product(s) distribution of adefovir dipivoxil (ADV) was examined in the presence of soluble and insoluble carbonate salts. METHODS: Chemical stability of ADV in the solid state at 60 degrees C/30% RH was examined. Stability was also examined in the presence of excess formaldehyde vapor at 23 degrees C/53% RH. ADV and its degradation product(s) were determined by reverse phase HPLC. RESULTS: Addition of aqueous soluble carbonate salts, such as sodium carbonate, compromised the stability of ADV in solid state. However, aqueous insoluble carbonates, such as calcium carbonate and magnesium carbonate, enhanced the stability of ADV as compared to the control formulation. Pivalic acid, a degradation product of ADV, was shown to accelerate the degradation rate of ADV in solid state. The de-stabilizing effect of this acid on ADV stability was diminished in the presence of magnesium carbonate. Pivalic acid also increased the rate at which ADV dimers were formed in the presence of formaldehyde vapor. Addition of insoluble carbonates reduced the rate of formaldehyde-catalyzed dimerization of ADV. CONCLUSIONS: Addition of insoluble carbonate salts decreased the rate of degradation of ADV by minimizing the extent of formaldehyde-catalyzed dimerization in solid state.

In situ acid resistance of in vivo formed white spot lesions.

Thin sections of natural enamel lesions, so-called white spots (WS), and areas of sound enamel (SEn) adjacent to the WS were exposed to an intraoral environment for 2 weeks. Thin sections of WS samples, clamped in a PMMA holder, were microradiographed before and after exposure to intraoral conditions. Acid resistance was evaluated by lesion depth and mineral changes during the cariogenic challenge. The results show that there were statistically significant differences in lesion depth, mineral loss and mineral volume percent at the surface before and after the intraoral cariogenic challenge at least at p<0.05, except for a change in mineral volume percent at the surface of WS samples. This exception indicates that no mineral change occurred in the surface layer of WS. The fact of 2.8 and 1.8 times higher ratios of SEn over WS of mean changes in lesion depth and mineral loss data, respectively, seems to indicate a quantitative difference in acid resistance level of WS lesions compared with the areas of SEn. Regarding the site of mineral changes, a distinctive feature of WS samples is that mineral loss occurs at the bottom of lesions. In contrast, areas of SEn produce a typical subsurface type of lesions. From this in situ study, it can be concluded that the surface of WS samples was apparently much more acid-resistant (at least approximately 2 times) than the areas of SEn that received a similar intraoral acid challenge.

Evaluation of using dog as an animal model to study the fraction of oral dose absorbed of 43 drugs in humans.

PURPOSE: To conduct a retrospective evaluation of using dog as an animal model to study the fraction of oral dose absorbed (F) of 43 drugs in humans and to briefly discuss potential factors that might have contributed to the observed differences in absorption. METHODS: Mean human and dog absorption data obtained under fasted state of 43 drugs with markedly different physicochemical and pharmacological properties and with mean F values ranging from 0.015 to 1.0 were obtained from the literature. Correlation of F values between humans and dogs was studied. Based on the same references, additional F data for humans and rats were also obtained for 18 drugs. RESULTS: Among the 43 drugs studied, 22 drugs were virtually completely absorbed in both dogs and humans. However, the overall correlation was relatively poor (r2 = 0.5123) as compared to the earlier rat vs. human study on 64 drugs (r2 = 0.975). Several drugs showed much better absorption in dogs than in humans. Marked differences in the nonliner absorption profiles between the two species were found for some drugs. Also, some drugs had much longer Tmax values and prolonged absorption in humans than in dogs that might be theoretically predicted. Data on 18 drugs further support great similarity in F between humans and rats reported earlier from our laboratory. CONCLUSIONS: Although dog has been commonly employed as an animal model for studying oral absorption in drug discovery and development, the present study suggests that one may need to exercise caution in the interpretation of data obtained. Exact reasons for the observed interspecies differences in oral absorption remain to be explored.