Physiologically Based Pharmacokinetic Modeling of Renally Cleared Drugs in Pregnant Women.

BACKGROUND: Since pregnant women are considerably underrepresented in clinical trials, information on optimal dosing in pregnancy is widely lacking. Physiologically based pharmacokinetic (PBPK) modeling may provide a method for predicting pharmacokinetic changes in pregnancy to guide subsequent in vivo pharmacokinetic trials in pregnant women, minimizing associated risks. OBJECTIVES: The goal of this study was to build and verify a population PBPK model that predicts the maternal pharmacokinetics of three predominantly renally cleared drugs (namely cefazolin, cefuroxime, and cefradine) at different stages of pregnancy. It was further evaluated whether the fraction unbound (f u) could be estimated in pregnant women using a proposed scaling approach. METHODS: Based on a recent literature review on anatomical and physiological changes during pregnancy, a pregnancy population PBPK model was built using the software PK-Sim®/MoBi®. This model comprised 27 compartments, including nine pregnancy-specific compartments. The PBPK model was verified by comparing the predicted maternal pharmacokinetics of cefazolin, cefuroxime, and cefradine with observed in vivo data taken from the literature. The proposed scaling approach for estimating the f u in pregnancy was evaluated by comparing the predicted f u with experimentally observed f u values of 32 drugs taken from the literature. RESULTS: The pregnancy population PBPK model successfully predicted the pharmacokinetics of cefazolin, cefuroxime, and cefradine at all tested stages of pregnancy. All predicted plasma concentrations fell within a 2-fold error range and 85% of the predicted concentrations within a 1.25-fold error range. The f u in pregnancy could be adequately predicted using the proposed scaling approach, although a slight underestimation was evident in case of drugs bound to α1-acidic glycoprotein. CONCLUSION: Pregnancy population PBPK models can provide a valuable tool to predict a priori the pharmacokinetics of predominantly renally cleared drugs in pregnant women. These models can ultimately support informed decision making regarding optimal dosing regimens in this vulnerable special population.

Rapid and simple method for determination of cephradine in human plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS): application to the bioequivalence study.

A rapid and simple procedure was developed for the determination of cephradine in human plasma using liquid chromatography coupled with electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). After trichloroacetic acid (TCA) precipitation of proteins from plasma samples, cephradine and cefaclor (the internal standard; IS) were eluted on a CN column. The isocratic mobile phase used consisted of acetonitrile-water-formic acid (25:75:0.1, v/v/v). Cephradine and the IS were both detected in multiple reaction monitoring (MRM) mode at the transitions: m/z 350.0 --> 90.8 for cephradine and m/z 368.1 --> 106.0 for the IS, respectively. The calibration curve was linear over the concentration range from 0.05 to 50 microg/ml, and correlation coefficients were greater than 0.996. The coefficient of variation of assay precision was less than 9.36%, and its accuracy ranged from 87.92% to 111.16%. The chromatographic run time for each plasma sample was less than 3 min. The developed method was successfully applied to a bioequivalence study of cephradine in healthy male volunteers.

Pharmacokinetic study of cephradine in Pakistani healthy male volunteers.

To observe and discuss the difference in the pharmacokinetics of cephradine in Pakistani population with the reported data of other ethnic origins. A Single group pharmacokinetic study was conducted having six healthy male volunteers of 20-24 years of age. Blood samples were collected at appropriate times up to 7 hours. Plasma concentrations of cephradine was determined by HPLC technique and pharmacokinetic parameters were determined by both compartmental and noncompartmental methods using Kinetica ver 4.4.1 and Winnonlin ver 5.01. Peak plasma concentration was 11.49+/-1.73 microg/ ml achieved at 0.76+/-0.12 hr, after the administration of 250 mg cephradine to fasting volunteers. Area under the serum concentration-time curve was found to be 16.4+/-1.71 g.hr/ ml. Absorption, distribution, disposition and elimination half lives were calculated as 0.183 +/- 0.038 hr, 0.248 +/- 0.143 hr, 2.126 +/- 0.341 hr and 0.441+/-0.193 hr respectively where as the volume of central compartment and total body clearance were found to be 9.65+/-3.78 L and 15.4+/-1.89 L/hr. The plasma concentration time curves showed the absorption rate constant was 3.968 +/- 0.05 hr(-1), disposition rate constant was 0.333+/-0.05 hr(-1), distribution rate constant was 3.64+/-2.18 hr(-1) and elimination rate constant was 1.738+/-0.468 hr(-1). The value of micro-constants i.e. K(12) (central to peripheral compartment) and K(21) (peripheral to central compartment) were found to be 1.529+/- 1.499 hr(-1) and 0.704 +/- 0.44 hr(-1) respectively, where as MRT and AUMC were calculated as 2.04+/-0.09 hr and 35.92+/-1.86 hr(2) microg/ ml. The findings showed that the results of Pakistani subjects are slightly different when compared with the reported data of other ethnic origin.

Quantitative prediction of oral absorption of PEPT1 substrates based on in vitro uptake into Caco-2 cells.

The method for predicting the fraction absorbed (Fa) of the PEPT1 substrates was established based on the in vitro uptake into Caco-2 cells. Uptake of a drug into Caco-2 cells was measured, and the carrier-mediated initial uptake clearance (DeltaCL uptake) was calculated as the difference between the uptake clearance in the absence of glycyl-sarcosine (Gly-Sar) and that in the presence of 30 mM Gly-Sar. The DeltaCL uptake of each drug was then divided by that of cephradine to obtain DeltaCL*uptake, which was a normalized parameter to correct for inter-day and/or inter-cell variability. Then, cephradine (CED), cefixime (CFIX), and cefotiam (CTM) were selected as marker compounds having excellent, medium and poor absorption, respectively. The DeltaCL*uptake and Fa values for CED, CFIX and CTM were fitted to the equation derived from the complete radial mixing (CRM) model, and the scaling factor (A') was obtained. Using the A' value, Fa was predicted from the DeltaCL*uptake value of each drug. Good correlation was observed between the predicted and reported Fa values, which demonstrated that Fa of PEPT1 substrates can be predicted based on the in vitro uptake in Caco-2 cells.

Properties and active substance release kinetics from gelatin-alginate matrices.

The aim of the study was to evaluate the effect of composition and technological processing on pharmaceutical availability of active substance as well as on the properties of porous gelatin-alginate matrices. The active substance carrier included glycerol or peanut oil apart from gelatin and sodium alginate, and some matrices were additionally modified with calcium lactate. The obtained matrices were characterized by good sorption properties and high resistance to proteolytic enzymes. The release of the model antibiotic followed the pattern of first order kinetics, while half-release time in vitro (in the experimental conditions) was 1.5 to 3 hrs.

A specific and rapid HPLC assay for the determination of cefroxadine in human plasma and its application to pharmacokinetic study in Korean.

A specific and rapid high performance liquid chromatographic (HPLC) method with UV detection (254 nm) was developed for the determination of cefroxadine in human plasma. The sample extraction was performed by a simple procedure, vortexing and centrifugation of sample following addition of 60% trichloroacetic acid. Cephalexin was used as an internal standard (I.S.). The HPLC analysis was carried out on a Capcell Pak C18 analytical column with a mobile phase of 50 mM ammonium formate buffer/pH 3.5 and acetonitrile (90:10, v/v). No interference was observed near the peaks of cefroxadine and I.S. The calibration curve was linear over the range of 0.5-40 microg/mL and the lower limit of quantification (LLOQ) was 0.5 microg/mL. The method was validated with excellent sensitivity, accuracy, precision and stability. This assay was successfully applied to determine the pharmacokinetic parameters of cefroxadine in Korean healthy volunteers after an oral administration of two 250 mg cefroxadine capsules. As a result, the plasma half-life was 1.00+/-0.26 h and the mean AUC(0-6 h) was 46.25+/-6.41microgh/mL. The maximum plasma concentration (C(max)) of 17.62+/-4.87 microg/mL reached 1.44+/-0.39 h after administration.

Inhibitory effect of zinc on PEPT1-mediated transport of glycylsarcosine and beta-lactam antibiotics in human intestinal cell line Caco-2.

PURPOSE: The aim of this study was to examine the effects of zinc on the intestinal peptide transporters (PEPT1 and basolateral peptide transporter) and to elucidate the mechanism of the interactions. METHODS: Caco-2 cells were pretreated with zinc, and the uptake studies were carried out. RESULTS: Zinc treatment resulted in the inhibition of [14C]glycylsarcosine (Gly-Sar) uptake via PEPT1 in a concentration-dependent manner, whereas it showed moderate inhibitory effect on the basolateral peptide transporter. Zinc also inhibited the uptake of oral beta-lactam antibiotics such as ceftibuten and cephradine by PEPT1. Kinetic analysis showed that zinc treatment increased Km values without affecting Vmax values of the [14C]Gly-Sar uptake. The inhibition of [14C]Gly-Sar uptake induced by zinc was observed in the presence of an H+ gradient but not in the absence of an H+ gradient. CONCLUSIONS: These results indicate that zinc is a competitive inhibitor of PEPT1. Zinc inhibited the PEPT1 function, possibly by interacting with histidine residues of PEPT1 that are part of an H+-binding site. These findings would provide important information for clinical, physiologic, and biochemical aspects of peptide transporters.

A recirculatory model with enterohepatic circulation by measuring portal and systemic blood concentration difference.

The present study describes a recirculatory model for the evaluation of pharmacokinetic characteristics of drugs possessing enterohepatic circulation (EHC). The advantage of the model is to separately define the extent and rate of absorption for the dosage and EHC after oral administration. Cephradine was used as a model drug and was intravenously or orally administered to rats. Portal and systemic bloods were simultaneously collected in order to estimate various local moments after defining the global moments obtained by non-compartment analysis. For the zero-order moments, bioavailability (BA), the hepatic recovery ratio (Fh), the sum of the local absorption ratio for the dosage and recirculatory local absorption ratio for EHC (F(a)po), and the recirculatory local absorption ratio for EHC (F(a)ehc) after oral administration were estimated to be 95.6, 77.9, 172, and 71.5%, respectively. These data indicate that a complete absorption and substantial EHC contribute high oral exposure of cephradine. For the first-order moments, the sum of the mean local absorption times for the dosage and EHC (t(a)po) and the mean transit time for a single pass of EHC (tc) were 2.50 and 0.117 hr, suggesting a rapid EHC of cephradine compared with the absorption from the dosage. With this model, the absorption rate-time profiles for the dosage and EHC were separately simulated by using a program of nonlinear least squares (MULTI) with fast inverse Laplace transform (FILT). The cumulative biliary excretion ratio (Fbile) calculated by the model was in good agreement with the experimental value obtained in the bile ductcannulated rats. These results suggest that the model proposed in this study would be useful for evaluating the extent and rate of ECH along with absorption from the dosage after oral administration of drugs.

Effect of composition on the physicochemical properties and active substance release from gelatin-alginate sponge.

The aim the study was physicochemically characterize and develop ability of the active substance (cefradine) from the implantable porous carriers. The drug delivery systems consisting of the gelatine and alginic acid sodium salt and glycerol (GL) or peanut oil (AO). Gelatin-alginate sponge was prepared by foamed components and next freeze-dried this foam. The composition of the sponges affected on the sorption ability and on the stability to proteolytic enzymes. Owing to porous structure obtained specific profile of active substance release.

Prolonged intestinal absorption of cephradine with chitosan-coated ethylcellulose microparticles in rats.

Cephradine-containing ethylcellulose microparticles (MPC) were prepared by the solvent evaporation method. Chitosan-coated MPC (Chi-MPC) were prepared by doping MPC with viscous chitosan solution and subsequently drying. When fluorescein isothiocyanate (FITC)-labeled chitosan-coated ethylcellulose microparticles without drug were administered intraduodenally, they moved slowly in the intestine, that is, most of them were retained at the upper and middle parts of the small intestine for more than 8 h, which is considered due to mucoadhesive properties of coated chitosan. When MPC and Chi-MPC was incubated at 37 degrees C in the JP 14 second fluid, pH 6.8, both released the drug slowly with similar release rates. Cephradine solution and suspension, MPC and Chi-MPC were administered intraduodenally to investigate intestinal drug absorption. Only Chi-MPC suppressed the initial plasma level and maintained the plasma concentration for a long time up to 24 h, suggesting Chi-MPC would be useful for prolonged intestinal absorption of cephradine.

Concentration-dependent preferences of absorptive and excretive transport cause atypical intestinal absorption of cyclic phenylalanylserine: small intestine acts as an interface between the body and ingested compounds.

Intestinal absorption of cyclic phenylalanylserine (cyclo(Phe-Ser)), a precursor of gliotoxin, was studied in isolated rat small intestine as a model cyclic dipeptide. Absorption clearance (CLabs) decreased in the presence of glycylsarcosine, cephalexin or cephradine, substrates for H+/oligopeptide cotransporter (PEPT1). CLabs of cyclo(Phe-Ser) also decreased at 4 degrees C. These indicate that cyclo(Phe-Ser) is in part transported by PEPT1. However, Eadie-Hofstee plot of absorption revealed an atypical profile at lower concentrations of cyclo(Phe-Ser) (around 0.1 mM). Moreover, comparative experiments of absorptive and excretive transport showed that excretive transport from the serosal to mucosal side of isolated intestinal tissue at a 0.1 mM cyclo(Phe-Ser) was superior to absorptive transport from the mucosal side to the serosal side, and vise versa at a 1 mM cyclo(Phe-Ser). These results as well as the results of kinetic analysis indicate that intestinal absorption consists of passive transport, carrier-mediated absorptive transport by PEPT1 and carrier-mediated excretive transport, resulting in atypical absorption. Although cyclic dipeptides have potentials for drug, their intestinal absorption may be complex. The results of this study lead us conclude that absorptive and excretive transport by the small intestine acts as an interface between the body and ingested compounds.

Determination of the cephalosporin antibiotic cephradine in human plasma by high-performance liquid chromatography with ultraviolet detection.

A simple and sensitive high-performance liquid chromatographic method, for the determination of cephradine in human plasma samples has been developed and validated. Cephradine and cephaloridine (internal standard) were extracted from human plasma by perchloric acid protein precipitation followed by centrifugation. Aliquots of the extracts were analysed by reversed-phase high-performance liquid chromatography (HPLC) utilising a polymeric reversed-phase PLRP-S column, followed by ultraviolet detection at 260 nm. The method has a working dynamic range from 0.2 to 30.0 microg/ml from 200 microl human plasma. The precision of the method at 0.2 microg/ml was 4.9% (intra-assay) and negligible (inter-assay) as calculated by one-way analysis of variance and the accuracy of the method at 0.2 microg/ml was -4.1% in terms of percentage bias. This method has been successfully applied to clinical studies including an oral bioequivalence study comparing the pharmacokinetics of 500 mg tablets of Kefdrin with 500 mg tablets of Velosef in healthy human volunteers.

pH-dependent inhibitory effects of angiotensin-converting enzyme inhibitors on cefroxadine uptake by rabbit small intestinal brush-border membrane vesicles and their relationship with hydrophobicity and the ratio of zwitterionic species.

The inhibitory effects of five angiotensin-converting enzyme (ACE) inhibitors on the uptake of an aminocephalosporin antibiotic, cefroxadine, by rabbit small intestinal brush-border membrane vesicles were examined in the presence of an inward H+ gradient. Dixon plot analysis showed that all these ACE inhibitors inhibited the uptake of cefroxadine, which is transported by a H+/oligopeptide transporter in the membrane, in the order of enalapril

Cephradin-plaga microspheres for sustained delivery to cattle.

In the field of controlled drug delivery, most of the reported work is aimed at introducing new systems, or at providing basic information on the critical parameters which affect release profiles in vitro and occasionally in vivo. The situation is totally different when one wants to fulfil the specific requirements imposed by the marketing of a sustained release device to be used in humans or in animals eaten by human beings. The control of the release characteristics is then a difficult challenge. In this work, attempts were made to combine cephradin, a hydrophilic beta-lactam antibiotic, and bioresorbable polymeric matrices of a poly(alpha-hydroxy acid) in the form of microspheres with the aim of delivering the antibiotic to cattle at a dose rate of 4-5 mg/kg/day over a 3-4 days period after i.m. injection. PLAGA aliphatic polyesters were selected because they are already FDA approved as matrices. The solvent evaporation technique using PVA as the emulsion stabilizer was selected because it is efficient and can be extended to an industrial scale. Various experimental conditions were used in order to obtain the highest encapsulation yields compatible with the desired specifications. Decreasing the volume of the aqueous phase and adding a water-miscible organic solvent/non-solvent of cephradin failed. In contrast, microspheres containing up to 30% cephradin were prepared after addition of sodium chloride to the aqueous dispersing phase. The amount of entrapped drug was raised to 40% by decreasing the temperature and the pressure. Preliminary investigations using dogs showed that 20% cephradin microspheres prepared under these conditions extended the presence of cephradin in the blood circulation up to 48 h. Increasing the load led to higher blood concentrations but shorter sustained release. The fact that the microspheres were for cattle limited the volume of the injection and thus the amount of microspheres to be administered. The other limiting factors were related to microsphere morphology.

Analysis of intestinal perfusion data for highly permeable drugs using a numerical aqueous resistance--nonlinear regression method.

PURPOSE: To develop, validate and apply a method for analyzing the intestinal perfusion data of highly permeable compounds using the Numerical Aqueous Resistance (NAR) theory and nonlinear regression (NAR-NLR) and to compare the results with the well-established Modified Boundary Layer (MBL) Analysis. METHODS: The NAR-NLR method was validated and the results were compared to the MBL analysis results using previously reported cephradine jejunal perfusion data. Using the Single Pass Intestinal Perfusion (SPIP) method, the concentration dependence of intestinal permeability was investigated for formycin B, proline, and thymidine, three compounds reported to be absorbed by carrier-mediated transport processes. The MBL and NAR-NLR analyses were then applied to the three sets of SPIP data. RESULTS: The results demonstrate that the intrinsic MBL transport parameters were highly variable and, in one case, the analyses failed to give a statistically significant Michaelis constant. The MBL mean dimensionless wall permeabilities (P*w) were greater than the NAR-NLR P*w and were also highly variable. In all cases, the NAR-NLR variability was significantly lower than the MBL variability. The extreme variability in the MBL-calculated P*w is due to the sensitivity of P*w when the fraction of unabsorbed drug (Cm/Co) is low or, alternatively, when P*w approached the aqueous permeability, P*aq. CONCLUSIONS: The NAR-NLR method facilitates the analysis of intestinal perfusion data for highly permeable compounds such as those absorbed by carrier-mediated processes at concentrations below their Km. The method also allows for the use of a wider range of flow conditions than the MBL analysis resulting in more reliable and less variable estimates of intestinal transport parameters as well as intestinal wall permeabilities.

In vitro and in vivo studies on microcapsules and tabletted microcapsules of cephradine.

Cephradine was microencapsulated by coacervation. Ethyl cellulose was used as the polymer and a core/wall ratio of 1:1 was selected. The repose angle, apparent and tapped density, particle size distribution of cephradine microcapsules (CM) and of cephradine powder were examined. Then flat-surfaced tablets of CM were prepared using Avicel PH 101 and magnesium stearate. In vitro and in vivo properties of CM and tabletted CM (both equivalent to 150 mg cephradine) were compared with commercial capsules (equivalent to 250 mg cephradine). The dissolution studies were carried out by the rotating basket method and the agar diffusion method was applied for quantitative determinations. Among the investigated kinetic models for the release of cephradine from CM and tabletted CM the best fit was found with the Higuchi model. In vivo studies were made in rabbits. Bioavailabilities of CM and their tabletted form were higher than that of the commercial capsules. In vitro/in vivo correlations between mean residence time (MRT) and mean dissolution time (MDT) for CM and tabletted CM were calculated. A good correlation was found between the in vitro and in vivo results.

Characteristics of uptake of cefroxadine by rabbit small intestinal brush border membrane vesicles.

Characteristics of transport of an oral aminocephalosporin, cefroxadine, in rabbit small intestinal brush border membrane vesicles were examined. Uptake rate of cefroxadine was saturable in the presence of an inward H+ gradient, and kinetic parameters were similar to those of cephradine. However, the uptake rate was almost linear with the concentration in the absence of an inward H+ gradient up to 5 mM. Overshoot phenomenon was observed in the presence of an inward H+ gradient at 37 degrees C, but it disappeared with decrease of temperature. The Arrhenius plot of uptake rate constant showed a break point at approximately 30 degrees C. Cefroxadine uptake was optimum in the vicinity of pH 5.5 at 37 degrees C, but the dependence on extravesicular pH disappeared at 15 degrees C. The uptake of cefroxadine in the presence of an inward H+ gradient was markedly inhibited by other aminocephalosporins such as cephalexin, but the inhibition was only slight in the absence of an inward H+ gradient. Alkyl alcohols such as n-hexyl alcohol also inhibited H(+)-coupled uptake of cefroxadine at the concentration range at which the alcohols increased the membrane fluidity, and overshoot phenomenon diminished, suggesting that H(+)-coupled transport of cefroxadine is sensitive to the alcohol-induced increase in membrane fluidity. On the other hand, the alcohols rather stimulated its uptake in the absence of an H+ gradient.

Noncompetitive inhibition of cephradine uptake by enalapril in rabbit intestinal brush-border membrane vesicles: an enalapril specific inhibitory binding site on the peptide carrier.

ACE inhibitors, as well as aminocephalosporins with peptide-like structures, are transported by the intestinal peptide carrier. We investigated the transport mechanism using intestinal brush-border membrane vesicles from rabbits and observed that enalapril, an angiotensin converting enzyme inhibitor and substrate of the peptide carrier, noncompetitively inhibited the uptake of cephradine, an aminocephalosporin and substrate of the peptide carrier, with an inhibition constant (Ki) of 2.6 mM when it was present on the cis side (outside) of the vesicles. By contrast, enalaprilat, cefadroxil and GlyPro competitively inhibited cephradine transport with Ki values of 5.4, 3.8 and 5.1, respectively. These results suggest the presence of an enalapril-specific inhibitory binding site on the peptide carrier. In addition, enalapril on the trans side (inside) of the vesicles inhibited the uptake of cephradine, suggesting an apparent reduction of carrier availability by a trapping mechanism. On the other hand, cefadroxil stimulated the uptake of cephradine in the trans experiment, consistent with the concept of countertransport. These findings reveal the uniqueness of enalapril regarding its mode of interaction with the peptide carrier(s) which has been of increasing interest regarding its role in the intestinal absorption of peptide-type drugs.

Disposition kinetics of cephradine in normal and Escherichia coli infected goats.

The pharmacokinetics of cephradine was studied following single and repeated intramuscular injections in normal and Escherichia coli infected goats. Bioavailability of cephradine was determined in normal goats after a single intramuscular dose. The serum concentrations of cephradine following a single and repeated intramuscular administration of 10 mg/kg b.wt. twice daily for five consecutive days, peaked 2 hours after each intramuscular dose with a lower significant value recorded in E. coli infected goats than in normal goats. The absorption half-lives (t0.5(ab)) following a single intramuscular injection of cephradine was significantly higher in E. coli infected goats (1.18 h) than in normal goats (0.64 h). The elimination half-lives (t0.5(beta)) of cephradine were significantly higher in E. coli infected goats than in normal goats following the administration of fifth and ninth doses. The urine and milk concentrations of cephradine were significantly lower in E. coli infected goats than in normal goats. The mean systemic bioavailability of cephradine following a single intramuscular injection in normal goats was 73.9%.

The intestinal uptake of "enzymatically-stable" peptide drugs in rats as influenced by D-glucose in situ.

In previous in situ and in vivo rat perfusion studies, the intestinal absorption of several low molecular weight drugs was increased by the presence of luminal D-glucose. The intent of this study was to determine the potential of this fed-state effect to improve the intestinal uptake of poorly permeable, small peptide and peptide-like drugs. Jejunal wall permeabilities (Pw*) of di-(D-kyotorphin), tri-(cephradine), hexa-(growth hormone releasing peptide, GHRP-6) and octa-(octreotide, a somatostatin analogue) peptides and corresponding net water fluxes were determined in rats using an in situ single-pass perfusion technique. Glucose was shown to enhance the uptake of the smaller (di- and tri-) peptides but not the larger peptides despite the fact that glucose elicited a significant net water absorption with each of the four peptide drugs. It is concluded that glucose enhances jejunal permeabilities of smaller peptides by solvent drag and the enhancement is limited in situ by peptide molecular size. The studies with nonmetabolizable 3-O-methylglucose suggest that the augmentation of the proton gradient across the transmucosal membrane by glucose contributes to the carrier-mediated transport observed with the smaller peptides.