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.

A study of the chemiluminescence behavior of cephalosporins with luminol and its analytical application.

The chemiluminescence intensity of luminol-dissolved oxygen was decreased when cephalosporins were mixed with luminol. The decrease chemiluminescence intensity was linear with the logarithm of cephalosporins concentration over the range from nanogram to microgramme level, with the limits of detection at nanogram level. The sensitivities of determination for cephalosporins were in the order of cefoperazone > ceftriaxone > cefuroxime > cefaclor > cefalexin > cefradine. The proposed method was applied to monitor the excretion of cefradine in human urine after taken cefradine capsules. The possible chemiluminescence mechanism and relationship between the determination sensitivities and generations of cephalosporins were also discussed.

[Analysis of multiple cephalosporins in blood and urine by HPLC].

OBJECTIVE: To establish a new high performance liquid chromatography (HPLC) method for determining the concentration of cefazolin, cefradine, cefoperazone and cefotaxime in blood and urine, as well as to investigate its applicability. METHODS: Protein in blood and urine was precipitated directly by acetonitrile with acetanilide was used as the internal standard using Agilent Zorbax SB-Aq column (250 mm x 4.6 mm, 5 microm). The mixed solvents of water (triethylamine 0.12%, acetic acid 0.12%) and acetonitrile were used as the mobile phase to separate cephalosporins using gradient elution method at 1 mL/min (flow rate) and 254 nm (detection wavelength). RESULTS: The working curve of four cephalosporins showed a good correlation (r = 0.9993), with the detection limit up to 0.01 microg/mL. The recovery rate was more than 81.2%. CONCLUSION: This method is fast, easy and accurate. It is suitable for biological analysis of the 4 cephalosporins of the blood and urine in practical cases.

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.

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.

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.

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.

Initial study of using a laminar fluid diffusion interface for sample preparation in high-performance liquid chromatography.

This report describes a new microfluidic device called the H Filter for sample preparation prior to HPLC. The H Filters make possible a diffusional transfer of an analyte from a sample stream into a stream of a "receiver" fluid. Existing mathematical models can be used for optimizing experimental conditions. The authors have selected the extraction of the antibiotic cephradine from blood to demonstrate the utility of the new device. The extracts of blood samples spiked with cephradine levels between 0.2 and 100 microg/ml were analyzed using a C8 reversed-phase column and UV detection at 260 nm. The HPLC results were in good agreement with theory. The recovery of 32.2+/-2.8% was uniform over the entire range of cephradine concentrations. The new method completely avoids the use of centrifuges, that is otherwise typical for most current methodologies for the preparation of blood samples prior to HPLC analysis.

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.

An alternative approach for assessment of rate of absorption in bioequivalence studies.

The partial area method was investigated for evaluation of equivalency in the rate of absorption of immediate release formulations. The applicability of the method was demonstrated with four drugs with different pharmacokinetic/pharmacodynamic characteristics. The confidence interval approach currently employed for bioequivalence determinations was applied to the relevant absorption parameters, including Cmax and partial AUCs. The method was found to be more discriminating than Cmax and/or Tmax in the evaluation of the absorption rate of drugs. The cutoff time or point for partial AUC calculation may vary with the type of drug under study, depending on its clinical use and onset of action. The method was shown to be useful in the assessment of rate of absorption in bioequivalence studies.

Peroral absorption of cefroxadine in patients within the first day after severe trauma: comparison to cefroxadine pharmacokinetics in fasted, healthy volunteers.

Peroral absorption of cefroxadine given to 7 24-h fasted trauma patients by nasogastric tube within the first day of admission was compared to that obtained in fasted healthy volunteers. The trauma patients exhibited significantly lower Cmax and reduced AUC. Even though rate and extent of bioavailability cannot be determined from these two different population groups since the total clearance must be assumed to be different in patients and healthy subjects, a reduced bioavailability is assumed based on pathophysiologic reflections.

Pharmacokinetics of cephradine administered intravenously and orally to young and elderly subjects.

The pharmacokinetics of IV and oral cephradine in healthy young male and female volunteers (ages 19 to 25, n = 10) were compared to those of older individuals (ages 65 to 81, n = 9). Subjects received 1 gram of cephradine by a 5-minute intravenous (IV) infusion followed the next day by a 1-gram oral dose. Serial serum and urine samples collected over a period of 12 hours after the dose were analyzed for cephradine concentration by a microbiologic assay. After IV administration, mean serum cephradine concentrations in the elderly group were significantly higher at both 6 hours (1.52 +/- 0.41 mcg/mL) and 8 hours (0.73 +/- 0.22 mcg/mL) than in the young group at 6 hours (0.43 +/- 0.11 mcg/mL). Total systemic clearance was significantly lower (2.64 +/- 0.34 vs. 4.81 +/- 0.59 ml/min/kg) and the elimination half-life was significantly longer (1.71 +/- 0.20 vs 1.12 +/- 0.13 hours) in the elderly group (P = .0001). Systemic cephradine clearance correlated positively with creatinine clearance (r2 = 0.34, P = .0110) and negatively with age (r2 = 0.79, P = .0052). The mean volume of distribution was not significantly different between the two groups. Mean renal clearance was significantly lower in the elderly group (P = .0001), but more than 80% of the dose was excreted in the urine within 6 hours in both groups. After oral administration, the mean peak concentration and time to peak concentration did not differ between groups. The relative oral bioavailability was approximately 94% in both groups. The mean serum concentrations in the elderly were higher at both 6 and 8 hours than in the young group at 6 hours. There were no differences in pharmacokinetic parameters between male and female subjects. Because of reduced cephradine clearance secondary to an age-related decline in renal function, administration of cephradine every 8 hours, rather than every 6 hours, may be sufficient in elderly patients.

Pharmacokinetics of cefroxadine after infusion to healthy volunteers.

The pharmacokinetics of cefroxadine in healthy human volunteers was studied in plasma and urine, after an infusion administration of 1 g of this drug for 0.5 h. Blood and urine samples were microbiologically quantified by diffusion on solid gelose using Bacillus Subtilis ATCC 6633 as the test organism. Plasma and urine profiles were fitted to a reduced two-compartment model not having inactivating biotransformation as a route of elimination. The parameters of distribution for this model show a rapid disposition (t1/2 alpha = 0.28 h) and an average volume of the central compartment of 0.15 +/- 0.04 l/kg (range 0.20-0.09). The dominant terminal half-life (beta-phase) was 1.17 +/- 0.26 h (range 0.90-1.67). The total body volume 0.41 +/- 0.09 l/kg (range 0.30-0.52) indicates that there is no diffusion of the antibiotic into intracellular fluids of poorly perfused tissues due to its high elimination rate.

Cephradine (Velosef) penetration of mandibular bone.

The concentration of cephradine in serum and mandibular bone was assayed in 28 patients undergoing 3rd molar surgery following a single 1 g intravenous injection. Serum and cortical bone samples taken simultaneously, contained mean cephradine concentrations of 42.11 micrograms/ml and 2.61 micrograms/g respectively. These results, when compared with those reported for other bony sites including the femoral head and knee, show a reduced bone penetration with a bone-to-serum ratio of approximately 0.06:1.

Cephradine penetration of mandibular bone.

The concentration of cephradine in serum and mandibular bone was assayed in 22 patients undergoing third molar surgery following a single 1-g intramuscular injection. Serum and cortical bone samples taken simultaneously contained mean cephradine concentrations of 12.41 micrograms/mL and 1.25 micrograms/g, respectively. These results, when compared with those reported for other bony sites, including femoral head and knee, show a reduced bone penetration with a bone-to-serum ratio of approximately 0.10:1.

Determination of serum and bone concentrations of cephradine and cefuroxime by HPLC in patients undergoing hip and knee joint replacement surgery.

Bone and serum concentrations of cephradine and cefuroxime were measured by HPLC in 21 patients undergoing hip and knee joint replacement surgery. An intravenous dose of 750 mg of each cephalosporin was given to patients at induction of anaesthesia. The serum and bone concentrations of both compounds were similar in individual patients although there was considerable interpatient variation. The mean concentrations in the femoral head bone supernatant in hip replacements were 7.2 mg/kg of cephradine and 8.0 mg/kg of cefuroxime. In knee replacement the femoral condyle bone concentrations were 5.1 mg/kg of cephradine and 4.2 mg/kg of cefuroxime, and for tibial bone 5.6 and 4.6 mg/kg respectively. Both cephradine and cefuroxime diffuse well into bone giving bone concentrations effective against common pathogens such as Staphylococcus aureus and S. epidermidis.

Microbore liquid chromatographic determination of cadralazine and cephalexin in plasma with large-volume injection.

The application of microbore systems (15 cm X 1 mm I.D. columns filled with Nucleosil C18, 5 microns particle size) to the determination of cephalexin and cadralazine in plasma was investigated. Factors such as mobile phase flow-rate, detector flow-cell volume and injection volume were examined with regard to the needs of routine drug analysis. Mobile phase flow-rates of 50-60 microliters/min were used. A flow cell with an optical path length of 6 mm and an intermediary volume (2.4 microliters) was selected for UV detection in order to obtain sufficient sensitivity. Large volumes of non-eluting solvent containing the drug were injected on the column. The addition of an ion-pairing reagent to samples containing cephalexin and cefroxadin prior to the injection was found to improve the chromatographic performance. The blood sample size required for analysis with microbore columns was smaller than that with conventional columns. The analysis time was similar and the limit of quantitation was also similar, provided that large sample volumes were injected on the microbore column.