Floating matrix dosage form for dextromethorphan hydrobromide based on gas forming technique: In vitro and in vivo evaluation in healthy volunteers

The objective of this study was to develop the dextromethorphan hydrobromide sustained-release (DMB-SR) tablets using floating technique to prolong the gastric residence time and compared their pharmacokinetic behavior with conventional sustained release tablets. DMB-SR floating tablets were prepared employing hydroxypropyl methylcellulose (HPMC) as hydrophilic gel material, sodium bicarbonate as gas-generating agent and hexadecanol as floating assistant agent. An orthogonal experiment design method was used to select the optimized formulation. The floating tablets were evaluated for uniformity of weight, hardness, friability, drug content, floating characteristics, in vitro release and in vivo bioavailability. The optimized tablets were prepared with HPMC K4M 25 mg, sodium bicarbonate 20 mg and hexadecanol 18 mg. The prepared tablets could float within 3 min and maintain for more than 24 h. The data of physical parameters were all lie within the limits. Drug release at 12 h was more than 85%. The comparative pharmacokinetic study was performed by administration of the DMB-SR floating tablets and conventional DMB-SR tablets. The area under curve of plasma concentration–time (AUC) of floating tablets was slightly higher than that of reference tablets, T_max was prolonged apparently. The results showed the floating tablets are a feasible approach for the sustained-release preparation of drugs, which have limited absorption sites in the stomach.     

Preformulation studies and optimization of sodium alginate based floating drug delivery system for eradication of Helicobacter pylori

The aim of this study was to design a local, floating, mucoadhesive drug delivery system containing metronidazole for Helicobacter pylori eradication. Face-centered central composite design (with three factors, in three levels) was used for evaluation and optimization of in vitro floating and dissolution studies. Sodium alginate (X₁), low substituted hydroxypropyl cellulose (L-HPC B1, X₂) and sodium bicarbonate (X₃) concentrations were the independent variables in the development of effervescent floating tablets. All tablets showed acceptable physicochemical properties. Statistical analysis revealed that tablets with 5.00% sodium alginate, 38.63% L-HPC B1 and 8.45% sodium bicarbonate content showed promising in vitro floating and dissolution properties for further examinations. Optimized floating tablets expressed remarkable floating force. Their in vitro dissolution studies were compared with two commercially available non-floating metronidazole products and then microbiologically detected dissolution, ex vivo detachment force, rheological mucoadhesion studies and compatibility studies were carried out. Remarkable similarity (f₁, f₂) between in vitro spectrophotometrically and microbiologically detected dissolutions was found. Studies revealed significant ex vivo mucoadhesion of optimized tablets, which was considerably increased by L-HPC. In vivo X-ray CT studies of optimized tablets showed 8 h gastroretention in rats represented by an animation prepared by special CT technique.     

CYP2B6 haplotype and biological factors responsible for hepatotoxicity in HIV-infected patients receiving efavirenz-based antiretroviral therapy

Data on the pharmacogenetic markers of CYP2B6 and biological factors associated with hepatotoxicity in HIV-infected patients receiving an efavirenz-based antiretroviral therapy (ART) regimen are very limited. A total of 134 HIV-infected Thai adults were prospectively enrolled to receive a once-daily regimen of efavirenz 600 mg/tenofovir/lamivudine. Seven single nucleotide polymorphisms (SNPs) within CYP2B6 were genotyped using real-time PCR. At 12 weeks after ART, plasma efavirenz concentrations at 12 h after dosing were measured. The mean ± standard deviation patient age was 37 ± 8 years, and 77.6% were male. The median (IQR) CD4 count was 43 cells/mm³ (17–105 cells/mm³). Eighteen patients (13.4%) had positive anti-HCV and 5 patients (3.7%) had positive HBsAg. The frequencies of heterozygous/homozygous mutants of each SNP were 64C>T (11%), 499C>G (0%), 516G>T (55%), 785A>G (63%), 1375A>G (0%), 1459C>T (3%) and 21563C>T (62%). The three most frequent haplotypes identified included *1/*6 (40.3%), *1/*1 (34.3%) and *6/*6 (8.2%). The median (IQR) plasma efavirenz concentration was 2.3 mg/L (1.4–3.7 mg/L). At 24 weeks, median (IQR) serum ALP was 98 mg/dL (73–133 mg/dL) and direct bilirubin was 0.11 mg/dL (0.10–0.19 mg/dL). The proportion of grade 1 and grade 2 elevated serum ALP was 12.7% and 1.5%, respectively. By multivariate analysis, factors associated with high ALP, total bilirubin and direct bilirubin included CYP2B6 haplotype *6/*6, high serum ALP at Week 0 and positive anti-HCV (all P < 0.05). In summary, HIV-infected patients with the pharmacogenetic marker ‘CYP2B6 haplotype *6/*6’ may have increased susceptibility to hepatotoxicity with efavirenz-based ART.     

The influence of CYP2A6 polymorphisms and cadmium on nicotine metabolism in Thai population

We investigated the influence of genetic, cadmium exposure and smoking status, on cytochrome P450-mediated nicotine metabolism (CYP2A6) in 182 Thai subjects after receiving 2 mg of nicotine gum chewing for 30 min. The urinary excretion of cotinine was normally distributed over a 2 h period (logarithmically transformed). Individuals with urinary cotinine levels in the ranges of 0.01–0.21, and 0.52–94.99 μg/2 h were categorized as poor metabolizes (PMs: 6.5%), and extensive metabolizers (EMs: 93.5%), respectively. The majority of EMs (45%) carried homozygous wild-type genotypes (CYP2A6*1A/*1A, CYP2A6*1A/*1B and CYP2A6*1B/*1B), whereas only 1% of PMs carried these genotypes. Markedly higher frequencies of EMs were also observed in all heterozygous defective genotypes including the null genotype (*4C/*4C; 1 subject).A weak but significant positive correlation was observed between total amounts of urinary cadmium excretion and total cotinine excretion over 2 h. Our study shows generally good agreement between CYP2A6 genotypes and phenotypes. Smokers accumulated about 3–4-fold higher mean total amounts of 2-h urinary cadmium excretion (127.5 ± 218.2 ng/2 h) than that of non-smokers (40.5 ± 78.4 ng/2 h). Among the smokers (n = 16), homologous wild-type genotype *1/*1 was significantly the predominant genotype (6/16) compared with other defective allele including *4C/*4C. In addition, 2 h urinary excretion of cotinine in smokers of all genotypes was significantly higher than non-smokers. The proportion of smokers who smoked more than 5 cigarettes/day was significantly higher in EMs in all CYP2A6 genotypes (n = 14) than in PMs (n = 0).     

CYP2C9 and VKORC1 in therapeutic dosing and safety of acenocoumarol treatment: implication for clinical practice in Hungary

The purpose of this work was to investigate the contribution of CYP2C9 and VKORC1 to acenocoumarol (AC) dose variability, bleeding events in Hungary. The study recruited 117 patients on long-term AC therapy (INR 2–3), and 510 healthy individuals to model the findings. Patients were genotyped for alleles proved to affect lower AC overdose CYP2C9*2, CYP2C9*3, VKORC1*2. Additionally, we tested VKORC1*3, VKORC1*4 to examine their effect in patients with higher AC requirements. Most impact on dose reduction is accountable for CYP2C9*2/*3 (59%) and for VKORC1*2/*2 (45.5%), and on dose increase for newly evaluated VKORC1*3/*4 (22.5%) diplotypes. VKORC1*3 and *4 alleles seem to balance the dose-reducing effect of VKORC1*2 allele. Being a carrier of combination of VKORC1*2 and CYP2C9*2,*3 polymorphisms, rather than of one of these SNPs, is associated with higher risk of over-anticoagulation (up to 34.3%) in long-term AC treatment. The pharmacogenetic dosing algorithm involving VKORC1, CYP2C9 diplotypes and age explains 30.4% of AC dosing variability (p < 6.10 × 10⁻⁹). Correlation between the studied diplotypes and bleeding events could not be revealed.     

The algal hepatoxoxin okadaic acid is a substrate for human cytochromes CYP3A4 and CYP3A5

The hepatotoxin okadaic acid (OA) was incubated with nine human recombinant cytochrome P450s (1A1, 1A2, 2C8, 2C9, 2C19, 2D6, 2E1, 3A4 and 3A5). Both CYP3A4 and CYP3A5 converted OA to a mixture of the same four metabolites, but incubation with CYP3A4 resulted in higher levels of conversion. Michaelis–Menten parameters, K_m (73.4 μM) and V_max (7.23 nmol of metabolites nmol^?1 min^?1) for CYP3A4 were calculated by analyzing double-reciprocal plots. LC–MSⁿ analysis and chemical interconversion indicate that metabolites 2 and 3 are the 11S-hydroxy and 11R-hydroxy okadaic acid respectively, while metabolite 4 is 11-oxo okadaic acid. LC–MSⁿ analysis of metabolite 1 shows a molecular ion which corresponds to an addition of 16 amu to OA, also suggesting hydroxylation, but the specific site has not been identified. The same four metabolites were produced upon incubation of okadaic acid with pooled human liver microsomes. This transformation could be completely inhibited with ketokonazole, and inhibitor of the CYP3A family of enzymes. The metabolites were determined to be only slightly less potent inhibitors of serine threonine protein phosphatase 2A (PP2A) when compared to OA. As PP2A is the principle molecular target for OA, these oxidative transformations may not effectively detoxify OA.     

Delivery of Nerve Growth Factor to Brain Via Intranasal Administration and Enhancement of Brain Uptake

The main objective of the study was to investigate the efficacy of chitosan to facilitate brain bioavailability of intranasally administered nerve growth factor (NGF). In vitro permeability studies and electrical resistance studies were carried out across the bovine olfactory epithelium using Franz diffusion cells. The bioavailability of intranasally administered NGF in rat hippocampus was determined by carrying out brain microdialysis in Sprague–Dawley rats. The in vitro permeation flux across the olfactory epithelium of NGF solution without chitosan (control) was found to be 0.37 ± 0.06 ng/cm²/h. In presence of increasing concentration of chitosan (0.1%, 0.25%, and 0.5%, w/v) the permeation flux of NGF was found to be 2.01 ± 0.12, 3.88 ± 0.19, and 4.12 ± 0.21 ng/cm²/h respectively. Trans-olfactory epithelial electrical resistance decreased ～34.50 ± 4.06% in presence of 0.25% (w/v) chitosan. The C_max in rats administered with 0.25% (w/v) chitosan and NGF was 1008.62 ± 130.02 pg/mL, which was significantly higher than that for rats administered with NGF only 97.38 ± 10.66 pg/mL. There was ～14-fold increase in the bioavailability of intranasally administered NGF with chitosan than without chitosan. Chitosan can enhance the brain bioavailability of intranasally administered NGF. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:3640–3646, 2009     

A novel coating concept for ileo-colonic drug targeting: Proof of concept in humans using scintigraphy

The in vivo proof of concept of a novel double-coating system, based on enteric polymers, which accelerated drug release in the ileo-colonic region, was investigated in humans. Prednisolone tablets were coated with a double-coating formulation by applying an inner layer composed of EUDRAGIT® S neutralised to pH 8.0 and a buffer salt (10% KH₂PO₄), which was overcoated with layer of standard EUDRAGIT® S organic solution. For comparison, a single coating system was produced by applying the same amount of EUDRAGIT® S organic solution on the tablet cores. Dissolution tests on the tablets were carried out using USP II apparatus in 0.1 N HCl for 2 h and subsequently in pH 7.4 Krebs bicarbonate buffer. For comparison, tablets were also tested under the USP method established for modified release mesalamine formulations. Ten fasted volunteers received the double-coated and single-coated tablets in a two-way crossover study. The formulations were radiolabelled and followed by gamma scintigraphy; the disintegration times and positions were recorded. There was no drug release from the single-coated or double-coated tablets in 0.1 N HCl for 2 h. The single-coated tablets showed slow release in subsequent Krebs bicarbonate buffer with a lag time of 120 min, while in contrast drug release from the double-coated tablets was initiated at 60 min. In contrast, using the USP dissolution method, normally employed for modified release mesalamine products, no discrimination was attained. The in vivo disintegration of the single-coated EUDRAGIT® S tablets in the large intestine was erratic. Furthermore, in 2 volunteers, the single-coated tablet was voided intact. Double-coated tablets disintegrated in a more consistent way, mainly in the ileo-caecal junction or terminal ileum. The accelerated in vivo disintegration of the double-coating EUDRAGIT® S system can overcome the limitations of conventional enteric coatings targeting the colon and avoid the pass-through of intact tablets. Moreover, Krebs bicarbonate buffer has the ability to discriminate between formulations designed to target the ileo-colonic region.     

Formulation Optimization and Bioavailability After Oral and Nasal Administration in Rabbits of Puerarin-Loaded Microemulsion

The main objective of the study was to investigate the efficacy of microemulsion (ME) to facilitate bioavailability of puerarin (PUE) after oral and nasal administration. The pseudo-ternary phase diagrams were constructed to screen the ME components and optimize the ME formulation. The optimized formulation for bioavailability assessment consisted of 20% Tween 80, 20% glycerin, and 1.6% ethyl oleate. The solubility (27.8 mg/mL) of PUE in ME was significantly improved compared to that (4.58 mg/mL) of crude PUE in water. The ME droplets were spherical with a mean particle diameter of 23.4 nm. After nasal (5 mg/kg) and oral (20 mg/kg) administration of a single dose of PUE as ME to fasted rabbits, the absolute bioavailability was 34.58% and 13.54%, respectively. It showed a shorter T_max (0.75 h) for nasal administration than that (1.0 h) for oral administration of PUE-loaded ME. The C_max of PUE-loaded ME was 0.55 μg/mL after nasal administration and 0.64 μg/mL after oral administration, respectively. The results showed that nasal administration might be a promising route to enhance the absorption of PUE in the form of ME.     

Influence of ABCB1, CYP3A4*18B and CYP3A5*3 polymorphisms on cyclosporine A pharmacokinetics in bone marrow transplant recipients

The aim of this study was to retrospectively evaluate the effect of polymorphisms in the CYP3A4, CYP3A5 and ABCB1 genes on the dose-adjusted concentration and dose requirement of cyclosporine A (CsA) in Chinese recipients during the early period after bone marrow or hematopoietic stem cell transplantation. Ninety-one bone marrow transplant recipients were genotyped by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay or by direct sequencing for the C1236T, G2677T/A and C3435T polymorphisms in CYP3A4*18B, CYP3A5*3, and ABCB1, respectively. The concentration at zero before administration (C₀) and concentration at 2 h after administration (C₂) of whole blood CsA were measured by fluorescence polarization immunoassay. Dose-adjusted C₀ and C₂ were determined and compared among groups with different genotypes. Compared with CYP3A5*3/*3 individuals, CYP3A5*1/*1 subjects have a significantly lower dose-adjusted C₀ and C₂ at days 1–10 and a higher dose requirement for CsA at days 16–30 (p < 0.05). In addition, homozygotes for the ABCB1 3435T mutant have a significantly higher dose-adjusted C₀ and C₂ and a lower dose requirement compared with wildtype (p < 0.05). Similar results were also derived for carriers of the T-G-C haplotype in CYP3A5 producers compared with non-carriers (p < 0.05 and p < 0.01, respectively). In summary, the ABCB1 3435T SNP, T-G-C haplotype in CYP3A5 producers, and CYP3A5*3 SNP are all associated with differences in CsA pharmacokinetics and dose requirements during the first month after bone marrow or hematopoietic stem cell transplantation. Genetic testing can therefore help to determine initial dosage and individualize immunosuppressive therapy.     

Pilot toxicokinetic study and absolute oral bioavailability of the Fusarium mycotoxin enniatin B1 in pigs

The aim of present study was to reveal the toxicokinetic properties and absolute oral bioavailability of enniatin B1 in pigs. Five pigs were administered this Fusarium mycotoxin per os and intravenously in a two-way cross-over design. The toxicokinetic profile fitted a two-compartmental model. Enniatin B1 is rapidly absorbed after oral administration (T_1/2a = 0.15 h, T_max = 0.24 h) and rapidly distributed and eliminated as well (T_1/2elα = 0.15 h; T_1/2elβ = 1.57 h). The absolute oral bioavailability is high (90.9%), indicating a clear systemic exposure. After intravenous administration, the mycotoxin is distributed and eliminated rapidly (T_1/2elα = 0.15 h; T_1/2elβ = 1.13 h), in accordance with oral administration.     

Human urotensin II in internal mammary and radial arteries of patients undergoing coronary surgery

AimsInternal mammary (IMA) and radial artery (RA) have different incidence of vasospasm and long-term patency rates in arterial grafting. We compared the vasoreactivity of human urotensin II (hU-II) and its receptor with mechanism investigations in IMA and RA.MethodsIMA and RA taken from patients undergoing coronary bypass surgery were studied in organ baths. Urotensin receptor expression was determined by RT-PCR.ResultshU-II contracted IMA with pD₂ of 8.57 ± 0.41 and 45.4 ± 9.1% E_max of contraction to 100 mM KCl, whereas caused less contractile responses in RA (pD₂:8.30 ± 0.79, E_max:20.4 ± 4.8%, p < 0.05). Nifedipine inhibited hU-II-contraction in IMA. In U₄₆₆₁₉-precontraction, hU-II elicited comparable relaxation in IMA (pD₂:8.39 ± 0.43, E_max:56.1 ± 4.0%) and RA (pD₂:9.03 ± 0.46, E_max:65.2 ± 7.1%). The relaxation was abolished by endothelium denudation and by indomethacin, oxadiazoloquinoxalinone or N^ω-nitro-l-arginine, oxyhemoglobin, and Ca²⁺-activated K⁺ channel (K_Ca) blockers. Urotensin receptor mRNA was detected in both arteries.ConclusionshU-II is an important spasmogen in arterial grafts with receptors expressed in IMA and RA. hU-II elicits stronger contraction in IMA than in RA and a moderate endothelium-dependent relaxation attributable to nitric oxide, prostacyclin, and endothelium-derived hyperpolarizing factor with involvement of K_Ca activation. The relaxant response of endothelium-intact IMA and RA to hU-II demonstrates the importance of preservation of endothelium in these grafts.     

A new and simple HPLC method for determination of etamsylate in human plasma and its application to pharmacokinetic study in healthy adult male volunteers

A new and simple HPLC assay method was developed and validated for the determination of etamsylate in human plasma. After protein precipitation with 6% perchloric acid, satisfactory separation was achieved on a HyPURITY C₁₈ column (250 mm × 4.6 mm, 5 μm) using a mobile phase comprising 20 mM sodium dihydrogen phosphate-2 hydrate (pH was adjusted to 3.5 by phosphoric acid) and acetonitrile at a ratio of 95:5 v/v. The elution was isocratic at ambient temperature with a flow rate of 0.75 ml/min. Allopurinol was used as internal standard. The calibration curve was linear over the range from 0.25 to 20 μg/ml (r² = 0.999). The limit of quantification for etamsylate in plasma was 0.25 μg/ml. The within day coefficient of variance (%CV) ranged from 3.9% to 10.2%, whereas the between-day %CV ranged from 3.1% to 8.7%. The assay method has been successfully used to estimate the pharmacokinetics of etamsylate after oral administration of a 500 mg tablet under fasting conditions to 24 healthy Egyptian human male volunteers. Various pharmacokinetic parameters including AUC_0–t, AUC_0–∞, C_max, T_max, t_1/2, MRT, Cl/F, and V_d/F were determined from plasma concentration–time profile of etamsylate.     

Pharmacogenetic relevance of the CYP2C9*3 allele in a tenoxicam bioequivalence study performed on Spaniards

We performed a study to quantify CYP2C9 and CYP2C8 alleles influence on the variability observed in tenoxicam pharmacokinetic (PK) and implication in a bioequivalence study design performed on Spaniards. Eighteen healthy volunteers were included in an open, randomized, crossover, phase I bioequivalence study. Significant increases were found in CYP2C9*3 alleles vs. *1 and *2 in AUC_0–∞ (median (min–max)): 256 (230–516) vs. 150 (100–268) and 169 (124–197) μg h/mL (p < 0.01) and half-life time (t1/2) 102 (79–36) vs. 56 (45–94) and 64 (60–80) h (p < 0.01). Non-significant differences were observed in C_max 1.9 (1.8–2.9) vs. 2.4 (1.7–3.4), 2.5 (1.6–2.9) μg/mL or in according to CYP2C8 alleles presence. CYP2C9*3 allele is associated to a longer elimination time of tenoxicam. PK parameters calculated in bioequivalence studies (AUC_0–∞, t1/2) may be influenced by the presence of CYP2C9*3 allele resulting in a high variability. Thus, bioequivalence studies of tenoxicam formulations should be designed considering genotype profile.     

Alteration of pharmacokinetics of proguanil in healthy volunteers following concurrent administration of efavirenz

Efavirenz and proguanil are likely to be administered concurrently for the treatment of patients with HIV and malaria. The metabolism of proguanil is mediated principally by CYP2C19 while efavirenz is known to inhibit this enzyme. This study therefore investigated the effect of efavirenz on proguanil disposition. Fifteen healthy volunteers were each given 300 mg single oral doses of proguanil alone or with the 9th dose of efavirenz (400 mg daily for 11 days) in a crossover fashion. Blood samples were collected at pre-determined time intervals and analyzed for proguanil and its major metabolite, cycloguanil, using a validated HPLC method. Co-administration of proguanil and efavirenz resulted in significant increases (p < 0.05) in C_max, T_max, AUC_T and elimination half-life (T_1/2β) of proguanil compared with values for proguanil alone [C_max: 2.55 ± 0.24 mg/l vs 3.75 ± 0.48 mg/l; T_max: 2.80 ± 0.99 h vs 4.80 ± 0.99 h; AUC_T: 45.58 ± 12.75 mg h/l vs 97.00 ± 23.33 mg h/l; T_1/2β: 16.50 ± 4.55 h vs 23.24 ± 4.08 h]. Also, efavirenz caused a pronounced decrease in the AUC(metabolite)/AUC(unchanged drug) ratio of proguanil along with a significant decrease (p < 0.05) in C_max and AUC of the metabolite.These results indicate that efavirenz significantly alters the pharmacokinetics of proguanil. These suggest that the protection against malaria by proguanil may be decreased when the drug is co-administered with efavirenz and the antimalarial efficacy is dependent on cycloguanil plasma levels.     

Preparation of estradiol chitosan nanoparticles for improving nasal absorption and brain targeting

The estradiol(E₂)-loaded chitosan nanoparticles (CS-NPs) were prepared by ionic gelation of chitosan with tripolyphosphate anions (TPP). The CS-NPs had a mean size of (269.3 ± 31.6) nm, a zeta potential of +25.4 mV, and loading capacity of E₂ CS-NPs suspension was 1.9 mg ml⁻¹, entrapment efficiency was 64.7% on average. Subsequently, this paper investigated the levels of E₂ in blood and the cerebrospinal fluid (CSF) in rats following intranasal administration of E₂ CS-NPs. E₂-loaded CS-NPs were administered to male Wister rats either intranasally or intravenously at the dose of 0.48 mg kg⁻¹. The plasma levels achieved following intranasal administration (32.7 ± 10.1 ng ml⁻¹; t_max 28 ± 4.5 min) were significantly lower than those after intravenous administration (151.4 ± 28.2 ng ml⁻¹), while CSF concentrations achieved after intranasal administration (76.4 ± 14.0 ng ml⁻¹; t_max 28 ± 17.9 min) were significantly higher than those after intravenous administration (29.5 ± 7.4 ng ml⁻¹ t_max 60 min). The drug targeting index (DTI) of nasal route was 3.2, percent of drug targeting (DTP%) was 68.4%. These results showed that the E₂ must be directly transported from the nasal cavity into the CSF in rats. Finally, compared with E₂ inclusion complex, CS-NPs improved significantly E₂ being transported into central nervous system (CNS).     

Food–drug interactions: Effect of capsaicin on the pharmacokinetics of simvastatin and its active metabolite in rats

Capsaicin (trans-8-methy-N-vanilly-6-nonenamide, CAP), the main ingredient responsible for the hot pungent taste of chilli peppers. However, little is known about the metabolic interactions between CAP and clinically used drugs. This study attempted to investigate the effect of CAP on the pharmacokinetics of simvastatin (SV), a cytochrome P450 (CYP) 3A substrate and an important cholesterol-lowering agent. CAP (3, 8 or 25 mg/kg), ketoconazole, dexamethasone or 5% CMC-Na was given to rats for seven consecutive days and on the seventh day SV (80 mg/kg) was administered orally. The results showed that when a single dose of SV was administered to rats fed with CAP over one week, AUC_0→∞, C_max of SV and its acid metabolite was significantly decreased in comparison to the control treatment. Pretreatment of rats with CAP resulted in an decrease in the AUC_0–∞ of SV of about 67.06% (CAP 3 mg/kg, P < 0.05), 73.21% (CAP 8 mg/kg, P < 0.01) and 77.49% (CAP 25 mg/kg, P < 0.01) compared with the control group. The results demonstrate that chronic ingestion of high doses of CAP will decrease the bioavailability of SV to a significant extent in rats.     

Transport of 5,5-diphenylbarbituric acid and its precursors and their effect on P-gp,MRP2 and CYP3A4 in Caco-2 and LS180 cells

AimTo examine the transport of 5,5-diphenylbarbituric acid sodium (T2007) and its mono- (MMMDPB) and di- (T2000) methoxymethylated precursors and their inducibility potential in Caco-2 and LS180 cells.MethodsTransport studies of T2000, MMMDPB and T2007 in Caco-2 cells were performed in Transwells. P-gp and CYP3A4 activities were assayed by [³H]digoxin and rhodamine 123 cellular retention and testosterone 6β-hydroxylation, respectively. Expressions of PXR, VDR and CAR mRNA and CYP3A4, MDR1/P-gp and MRP2 mRNA and protein were determined by qPCR and Western blotting, respectively. PXR siRNA was used to assess the involvement of PXR.ResultsThe P_app(A→B)s and P_app(B→A)s of T2000, MMMDPB and T2007 were similar (30–35 × 10^?6 cm/s) in Caco-2 cells. Treatment for 3 days with T2000 (15 μM), MMMDPB (70 μM) and T2007 (300 μM) generally furnished a greater induction in LS180 cells over the Caco-2 cells due to the higher, natural abundance of PXR. Changes in expression were confined mostly to MDR1 and CYP3A4: in LS180 cells, treatment for 3 days increased MDR1 and CYP3A4 but not MRP2 mRNA, and elevated P-gp and CYP3A4 protein expression that led to decreased cellular accumulation of [³H]digoxin and rhodamine 123, and enhanced testosterone 6β-hydroxylase activity towards T2007, respectively. The silencing of PXR by PXR siRNA in LS180 cells significantly attenuated the induction of MDR1 and CYP3A4.ConclusionsT2000, MMMDPB, and T2007 exhibited high permeabilities but are not P-gp substrates. T2007 and its analogs upregulated CYP3A4 and MDR1 modestly via the PXR.     

Improving sublingual delivery of weak base compounds using pHmax concept: Application to propranolol

The purpose of the present work was to provide theoretical and experimental support in generating an optimal pH (pH_max) for a representative weak base compound (propranolol), that can lead to enhanced sublingual absorption. Initially equations for pH-solubility and pH-permeability profiles were derived and compared to the profiles obtained experimentally. Excellent correlation (R² = 0.999) of solubility profiles was obtained using non-linear regression, and the permeability profiles further predicted that at certain pH (pH_max), optimal mucosal permeation could be achieved. Subsequently, in a pharmacokinetics study, a buffered sublingual propranolol tablet, designed to achieve its pH_max (when dissolved in saliva), were compared to that from a marketed product (Inderal^® which could not achieve pH_max) in 8 healthy subjects. Each subject received the products sublingually for 15 min followed by swallowing the remaining drug–saliva. The plasma propranolol concentrations of AUC during first 30 min from the buffered tablet were significantly higher than that from the Inderal^® tablet (p < 0.05), and no significant differences in the remaining AUC were observed. These in vitro and in vivo results on propranolol provided experimental confirmation of the pH_max concept as well as its utility in sublingual drug delivery. Such an approach may be applicable to other similar compounds to improve sublingual drug delivery.     

The prototypic pharmacogenetic drug debrisoquine is a substrate of the genetically polymorphic organic cation transporter OCT1

Debrisoquine is a probe drug for in vivo phenotyping of human CYP2D6 metabolic activity. However, debrisoquine is positively charged under physiological conditions and it is unclear how it enters the hepatocytes to undergo CYP2D6 metabolism. We analysed whether debrisoquine is a substrate of the hepatic organic cation transporter OCT1 and whether drug–drug interactions at OCT1, or polymorphisms in OCT1 gene, affect debrisoquine uptake.Debrisoquine showed low carrier-independent membrane permeability (P_e of 0.01 × 10^?6 cm/s in artificial PAMPA membranes) and strongly inhibited the uptake of the model OCT1 substrate MPP⁺ (IC₅₀ of 6.2 ± 0.8 μM). Debrisoquine uptake was significantly increased in HEK293 cells overexpressing OCT1 compared to control cells. The OCT1-mediated uptake of debrisoquine followed Michaelis–Menten kinetics (K_M of 5.9 ± 1.5 μM and V_max of 41.9 ± 4.5 pmol/min/mg protein) and was inhibited by known OCT1 inhibitors and by commonly used drugs. OCT1-mediated debrisoquine uptake was reduced or missing in cells expressing loss-of-function OCT1 isoforms. Deletion of Met420 or substitution of Arg61Cys or Gly401Ser reduced V_max by 48, 63 and 91%, respectively, but did not affect the K_M. The OCT1 isoforms carrying Cys88Arg or Gly465Arg substitutions completely lacked OCT1-mediated debrisoquine uptake.In conclusion, debrisoquine is a substrate of OCT1 and has the potential to be used as a phenotyping marker for OCT1 activity. Moreover, variations in debrisoquine metabolic phenotypes and their associations with diseases may be due not only to genetic variations CYP2D6, but also in OCT1.