Population PKPD of voclosporin in renal allograft patients.

The aims of this population-pharmacokinetic/pharmacodynamic (POP-PKPD) analysis of voclosporin in renal allograft patients were to build a POP-PKPD model for voclosporin and calcineurin activity (CNa) and identify clinically relevant covariates that could assist dosing of the drug. POP-PKPD modeling was performed using a stochastic approximation of the standard expectation maximization (SAEM) algorithm for nonlinear mixed-effects as implemented in Monolix™ 3.2. Voclosporin whole blood concentrations were obtained from de novo renal allograft patients and assayed using a validated LC/MS/MS assay. CNa was measured using a (32)P-radiolabeled assay. A two-compartment model with simultaneous sigmoid inhibitory Emax model was used to describe the PKPD relationship between voclosporin concentration and CNa. The POP-PKPD model was then utilized to simulate an optimal initial dosing strategy. Eighty-seven patients were included in the POP-PKPD study. Population mean estimates (relative standard error, rse) for oral clearance (CL/F) and first compartment volume of distribution (V1), were 717 mL min(-1) (35%) and 2010 mL (17%), respectively. Maximum CNa Inhibition (Imax), effective concentration (C50), and baseline immunosuppression (S0) were 0.87 pmol/min/mg (8.0%), 123 ng/mL (10%), and 1.15 pmol/min/mg (4.0%), respectively. Covariate analyses demonstrated that age and body surface area significantly influenced CL/F: CLi=717(Agei/48.8)-0.57(BSAi/1.99)1.1, while serum triglycerides significantly altered S0: S0i=1.15(TRIGi/1.97)0.15.

Pharmacokinetics of voclosporin in renal impairment and hepatic impairment.

Voclosporin is a novel calcineurin inhibitor intended for prevention of organ graft rejection and treatment of lupus nephritis. These studies evaluated the effect of renal or hepatic impairment on pharmacokinetics of voclosporin. Thirty-three subjects were enrolled into 1 of 4 groups based on renal function as defined by creatinine clearance and 18 subjects were enrolled into 1 of 3 groups based on hepatic function defined by Child-Pugh classes. Voclosporin 0.4 mg/kg was administered orally. Geometric mean ratios (renal/hepatic impairment-to-normal) and 90% confidence intervals for Cmax and AUC were calculated. A default no-effect interval of 80-125% was set. Although 90% confidence intervals exceeded the no-effect intervals for both parameters, individual Cmax and AUC plots indicate almost complete overlapping range of values for mild and moderate renal impairment and normal subjects. Severe renal impairment resulted in a 1.5-fold increase in AUC without an increase in Cmax . Mild to moderate hepatic impairment resulted in a 1.5- to 2-fold increase in voclosporin exposure. Voclosporin can be administered safely to patients with mild to moderate renal impairment without dose modification. Appropriate safety monitoring with concentration-based adjustments in transplantation are recommended for patients with severe renal impairment, and for patients with hepatic impairment.

Changes in the temporal and spatial expression of H beta 58 during formation and maturation of the chorioallantoic placenta in the Rat.

Cloning and sequencing of a cDNA amplified by RNA fingerprinting at the implantation site of pregnant rats revealed 80% similarity with H beta 58, previously shown to be essential for formation of the chorioallantoic placenta in the mouse. H beta 58 mRNA was detected in the endometrium of hormonally sensitized rats stimulated to undergo decidualization and in the contralateral uterine horns lacking a decidual stimulus, indicating that uterine expression of H beta 58 mRNA did not require decidualization or the presence of a blastocyst. Immunodetection in the early postimplantation uterus (Days 6-8 of pregnancy) showed H beta 58 localized in the luminal and glandular epithelia and some stromal cells. Decidual cells at Day 6 of pregnancy expressed H beta 58, and by Day 9 of pregnancy, the protein localized throughout the maternal decidua. The temporal and spatial distribution of H beta 58 in the developing chorioallantoic placenta was assessed at Days 10, 12, and 14 of pregnancy. Immunoreactive H beta 58 localized to erythroid cells within the developing fetal vasculature of the chorioallantoic primordia at Day 10 of pregnancy. By Day 12, the fetal vasculature extended into the placental labyrinth, and the erythroid stem cells continued to strongly express H beta 58. At Day 14 of pregnancy, immunoreactivity became evident in the trophoblast giant cells and syncytiotrophoblast of the fetal placenta. As the chorioallantoic placenta matured (Day 18), H beta 58 mRNA was 3.6-fold higher in the labyrinth compared with the junctional region. Stable cell lines (HRP/LRP) isolated from the rat labyrinthine placenta expressed H beta 58 mRNA and protein. The expression pattern of H beta maternal and fetal placental tissues and its early expression in fetal erythroid stem cells during formation and maturation of the chorioallantoic placenta suggest that H beta 58 plays key roles in the regulatory networks that control hematopoietic development and placentation.

Pharmacokinetics of metoprolol enantiomers following single and multiple administration of racemate in rat.

The chiral beta-adrenergic blocking agent metoprolol (MET), which is marketed as a racemate, is a highly extracted drug with rapid absorption. The enantiomeric disposition of MET is reported following racemic administration as a single and as multiple oral dosing four times per day for four days in male Sprague-Dawley rats (n=6 in each group). Plasma was collected and enantiomeric concentrations of MET were determined using a stereospecific HPLC assay. The R/S ratio for AUC is not statistically different from unity either after single or after multiple administration of racemate. The oral clearance after single dose was 1.99+/-0.87 and 2. 26+/-0.85 ml min(-1) kg(-1) for R- and S-MET, respectively. These values were decreased to 0.59+/-0.21 and 0.64+/-0.26 ml min(-1) kg(-1) after multiple administration of racemate. The corresponding values for the elimination half-lives were approximately 35 and 33 min after single and multiple dose administration for both enantiomers, respectively. These results may suggest a saturable first pass metabolism of MET as its enantiomers are accumulated in plasma following multiple dosing in the rat model.

Effects of hyperlipidemia on the pharmacokinetics of nifedipine in the rat.

PURPOSE: The effect of hyperlipidemia on nifedipine pharmacokinetics was studied. The mechanisms by which hyperlipidemia affects pharmacokinetics of drugs are mainly undetermined. Hyperlipidemia may decrease the fraction of unbound drug in plasma and/or decrease intrinsic ability of the cytochrome P-450 systems due to excess membrane cholesterol. Hyperlipidemia is a primary risk factor for coronary artery disease leading to hypertension and ischemic heart disease, for which nifedipine, a calcium channel blocker, is used. METHODS: Poloxamer 407 (P407)-induced hyperlipidemic rat model was used to study the effects of hyperlipidemia on the pharmacokinetics of nifedipine (6 mg kg-1 given i.v., i.p. and p.o.). Total plasma cholesterol levels increased from 0.82-2.02 to 5.27-11.05 mmol L-1 48 h post P407 administration (1 g kg-1, i.p.). Protein binding studies were conducted by an ultrafiltration method. RESULTS: Hyperlipidemia significantly decreased CLTB by 38% and CLTB/F by 45 and 42% following po and i.p. doses, respectively, thereby increasing AUC0-infinity, Cmax and half-life. Absolute bioavailability and Vdss remained unchanged. AUC0-infinity was affected to the same extent in each route of administration, therefore, the effect was mainly systemic rather than presystemic. Hyperlipidemia significantly lowered the fraction unbound in plasma by approximately 31%. CONCLUSIONS: The altered pharmacokinetics of nifedipine by P407-induced HYPERLIPIDEMIA may be, at least in part, due to the decrease in fraction unbound in plasma. A decrease in intrinsic clearance, however, cannot be ruled out.

Pharmacokinetics of single oral and multiple intravenous and oral administration of acebutolol enantiomers in a rat model.

Acebutolol (AC), is a chiral, beta-adrenergic blocking agent which possesses partial agonist activity and is metabolized to an equipotent chiral metabolite, diacetolol (DC). The enantiomeric disposition of AC is reported following racemic administration as a single oral (p.o., 50 mg kg(-1)) or as a multiple thrice daily intravenous (i.v.) or p.o. dosing for four days in male Sprague-Dawley rats (n = 6). Enantiomeric concentrations of AC and DC in plasma and urine were determined using a stereospecific HPLC assay. The bioavailabilities of R- and S-enantiomer were 0.40 and 0.39 after single dose administration of AC respectively. These values were increased to 0.51 and 0.53 after multiple dosing. Although no significant differences were found in AUC0-infinity after single i.v. as compared with AUC0-tau after multiple i.v. dosing of AC, the 39 and 45% increase in mean AUC0-tau were found after multiple p.o. dosing over the corresponding AUC0-infinity, for the single p.o. dose of AC for R- and S-enantiomer, respectively. The disposition of DC as well as the urinary excretion of metabolite was stereoselective in favor of R-enantiomer after oral administration of AC. These results indicate that AC enantiomers have low availability and moderate extraction through the first-pass metabolism in a rat model. The higher AUC values after p.o. multiple dosing may suggest a saturable first-pass metabolism of AC.

Gender differences in autoimmune diseases: estrogen increases calcineurin expression in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) predominantly affects women (9:1 compared to men) of childbearing age and often decreases its intensity in postmenopausal women, suggesting that sex hormones play a role in its pathogenesis. Comparison of steady-state levels of calcineurin mRNA using RNase protection assays revealed increased calcineurin expression in response to estradiol in cultured T cells from nine female lupus patients. Calcineurin mRNA levels did not increase significantly in T cells from eight age-matched normal control female volunteers. Estrogen-dependent calcineurin mRNA increased in a dose-dependent fashion, while progesterone and dexamethasone did not increase calcineurin mRNA in patient cells. Lupus T cell calcineurin mRNA increased in response to estradiol at 6 h but not at 3 h. Calcineurin phosphatase activity increased in lupus T cell extracts after incubation of cells with estradiol, while phosphatase activity in normal T cells was unaffected by estrogen. Calcineurin expression in T cells from patients with vasculitis and rheumatoid arthritis taking medications similar to those taken by the lupus patients was unaffected by estradiol. This study provides the first evidence for a molecular marker of estrogen action in lupus patients and suggests that estrogen-dependent changes in lupus T cell calcineurin could alter proinflammatory cytokine gene regulation and T-B cell interactions.

Grapefruit juice and orange juice effects on the bioavailability of nifedipine in the rat.

Previous studies with rats indicate that nifedipine undergoes both hepatic and extrahepatic presystemic metabolism after peroral (po) administration, and that its bioavailability is increased and absorption delayed by concomitant administration of grapefruit juice concentrate (GJC). Hence, the effects of GJC could be to delay stomach emptying and inhibit nifedipine metabolism in the small-intestinal wall and liver or, alternatively, to impede nifedipine absorption until reaching the large intestine where gut wall presystemic metabolism is not a factor. The mechanism(s) of action of GJC might be partially resolved by comparison with orange juice concentrate (OJC), which has a similar consistency but lacks inhibitory effects on nifedipine presystemic metabolism, and also by giving regular-strength solutions of the two juices, both on which should not significantly affect stomach emptying. This study compared the po bioavailability of nifedipine (6 mg kg-1) in male Sprague-Dawley rats coadministered GJC, OJC, grapefruit juice regular strength (GJRS), orange juice regular strength (OJRS), or (tap) water. Nifedipine plasma concentration-time profiles in the GJRS, OJRS, and (tap) water groups displayed a single peak. Both GJC and OJC groups have double-peak profiles (indicating delayed gastric emptying); however, the majority of the nifedipine dose in both cases was absorbed during the interval of the second peak, which occurred several hours postdosing. GJC significantly increased nifedipine bioavailability (relative bioavailability 2.02, compared with (tap) water), indicating that GJC may affect both extrahepatic and hepatic first-pass metabolism, although a reduction in systemic nifedipine clearance cannot be ruled out. Surprisingly, GJRS had no significant effect on nifedipine bioavailability. OJC did not increase nifedipine bioavailability, further suggesting that the delay in nifedipine absorption by GJC or OJC results from delayed gastric emptying.

Extrahepatic first-pass metabolism of nifedipine in the rat.

The peroral (p.o.) bioavailability of nifedipine is reported to range from about 45 to 58% in the rat; this compares favourably to human beings. The metabolism of nifedipine is similar in rats and humans (oxidation of the dihydropyridine ring), with the liver believed to be solely responsible for the systemic clearance of the drug and the observed first-pass effect after p.o. dosing. The purpose of this study was to determine whether intestinal metabolism also contributes to the first-pass elimination of nifedipine in the rat. The systemic availabilities of nifedipine doses given by po, intracolonic (i.c.), and intraperitoneal (i.p.) routes of administration were compared to that for an intravenous (i.v.) dose (in each case a dose of 6 mg kg-1 was given) using adult male Sprague-Dawley rats (249-311 g, n = 6 or 7/group). The geometric mean of systemic nifedipine plasma clearance after i.v. dosing was 10.3 mL min-1 kg-1. The nifedipine blood-to-plasma ratio was found to be about 0.59. Therefore, the systemic blood clearance of nifedipine was about 17.5 mL min-1 kg-1; which, compared to the hepatic blood flow of rats (55 to 80 mL min-1 kg-1) showed that nifedipine is poorly extracted by the liver (0.22 < or = EH < = 0.32). The mean absolute bioavailabilities of the p.o., i.p., and i.c. doses were 61, 90, and 100%, respectively. Assuming complete absorption of the extravascular nifedipine doses these results indicate that, in addition to hepatic extraction, substantial first-pass elimination of nifedipine occurs within the wall of the small intestine but not the colon of the rat.

Pharmacokinetics of the enantiomers of verapamil after intravenous and oral administration of racemic verapamil in a rat model.

Verapamil is a chiral calcium channel blocking drug which is useful clinically as the racemate in treating hypertension and arrhythmia. The published pharmacokinetic data for verapamil enantiomers in the rat model are limited. Utilizing a stereospecific high-performance liquid chromatographic (HPLC) assay, the enantiomeric disposition of verapamil is reported after intravenous (1.0 mg kg-1) and oral (10 mg kg-1) administration of racemic verapamil to the rat model. After intravenous administration the systemic clearance of R-verapamil was significantly greater than that of S-verapamil; 34.9 +/- 7 against 23.7 +/- 3.7 mL min-1 kg-1 (mean +/- SD), respectively. After oral administration, the clearance of R-verapamil was significantly greater than that of S-verapamil, 889 +/- 294 against 351 +/- 109 mL min-1 kg-1, respectively. The apparent oral bioavailability of S-verapamil was greater than that of R-verapamil, 0.074 +/- 0.031 against 0.041 +/- 0.011, respectively. These data suggest that the disposition of verapamil in the rat is stereoselective; verapamil undergoes extensive stereoselective first-pass clearance after oral administration and the direction of stereoselectivity in plasma is opposite to that observed in the human.

Pharmacokinetics of acebutolol enantiomers after intravenous administration of racemate in a rat model: a dosing range comparison.

Acebutolol (AC) is a chiral beta-adrenergic blocking drug, possessing intrinsic sympathomimetic activity (ISA), and is useful clinically as the racemate in treating hypertension. Utilizing a stereospecific high-performance liquid chromatographic (HPLC) assay, the enantiomeric disposition of AC and its major metabolite diacetolol (DC) are reported after intravenous administration of single 5, 15, 30, and 50 mg kg-1 doses of racemate to male Sprague-Dawley rats. The mean area under the plasma concentration versus time curve (AUC) values display a linear relationship with respect to the administered dose. No statistical differences are observed in apparent volume of distribution (Vd), terminal elimination half-life (t1/2), total body clearance (Clt), or renal clearance (Clr) with respect to dose administered. Generally, R-S ratios for AUC following AC administration are statistically different from unity (p < 0.05). However, for the 50 mg kg-1 doses the R-S ratio for AUC is not statistically different from one. For DC, the plasma disposition is nonstereoselective in plasma. The amount of R-DC recovered in urine, however, was greater than that of the antipode (R:S = 1.92 +/- 0.29). This study suggests that the enantiomeric disposition of intravenous AC is linear within the investigated range of 5-50 mg kg-1 racemate in rats.

Oestrogen and progesterone control basic fibroblast growth factor mRNA in the rat uterus.

Cell proliferation and differentiation in the rodent uterus are probably controlled by the interaction of female sex steroids with polypeptide growth factors. Uterine basic fibroblast growth factor (bFGF) mRNA was measured by RNase protection during the time (days 2-4) of endometrial cell proliferation in the pregnant rat. bFGF transcripts were detected at each of the 3 days of pregnancy examined. To investigate the influence of oestrogen and progesterone on bFGF mRNA accumulation, ovariectomized rats were treated with oestradiol for 48 h followed by a single injection of oestradiol, progesterone, the two steroids co-injected or oil vehicle alone. Uterine RNA was collected 6 h after the last hormone injection. Steroid treatments increased steady-state uterine bFGF mRNA compared with vehicle control animals as measured by RNase protection. Northern blot analysis of c-fos and c-jun mRNAs from these same treatment groups revealed increased protooncogene expression in the uterus of hormone treated rats compared with the control animals. Temporal analysis of bFGF mRNA in ovariectomized rats at 1, 3 and 6 h after acute oestrogen and oestrogen-progesterone co-administration showed a dual pattern of transcript accumulation. Both hormone treatments increased bFGF mRNA within 1 h compared with vehicle injected rats. Co-administration of the two hormones, however, repressed bFGF mRNA accumulation relative to oestrogen at 3 and 6 h. Together, these studies provide evidence that bFGF control of uterine cell proliferation in pregnant rats can occur from newly synthesized bFGF. Moreover, the results suggest that progesterone is a potent stimulator of bFGF expression in the uterus.

Stereoselective metabolism of rac-mexiletine by the fungus Cunninghamella echinulata yields the major human metabolites hydroxymethylmexiletine and p-hydroxymexiletine.

rac-Mexiletine is an orally effective class 1b antiarrhythmic agent used to treat ventricular arrhythmias. In vivo experiments have demonstrated. It is predominantly metabolized by the liver with < 10% excreted as unchanged drug. The major mammalian metabolites have been identified as p-hydroxymexiletine (PHM) and hydroxymethylmexiletine (HMM). The purpose of our study was to determine whether the fungus Cunninghamella echinulata, which possesses a cytochrome P450 system analogous to that found in humans, could be used as a suitable in vitro model for studying the oxidative metabolism of rac-mexiletine. To accomplish this, a high performance liquid chromatographic assay was used that was capable of simultaneously quantifying the enantiomers of mexiletine, HMM, and PHM. Utilizing this procedure, it was demonstrated that C. echinulata stereoselectively converted rac-mexiletine into HMM (4% of added drug) and PHM (32% of added drug) after an incubation period of 50 hr. In addition, metabolite biosynthesis could be optimized by altering fermentation media components. Seven media values and seven pH values were evaluated. It was determined that a medium at pH 7.0 containing yeast extract and sucrose yielded optimal amounts of metabolites.

Renal, biliary and intestinal clearance of sotalol enantiomers in rat model: evidence of intestinal exsorption.

Biliary clearance (Clb) of sotalol (STL) enantiomers was assessed in anaesthetized Sprague-Dawley rats (419 +/- 9 g, mean +/- SEM, n = 4) following administration of a 10 mg kg-1 i.v. dose of the racemate. Clb for S- and R-STL (0.0675 +/- 0.0090 and 0.0662 +/- 0.0089 mL min-1 kg-1, respectively) represented approximately 0.3% of systemic clearance (Cls) values for S- and R-STL (20.4 +/- 2.2 and 20.7 +/- 2.0 mL min-1 kg-1, respectively). Bile:plasma concentration ratios at 1, 2, and 3 h post-dose were approximately 1.4, 1.3, and 1.2 for both STL enantiomers. Renal clearance (Clr) and intestinal clearance (Cli) of STL enantiomers were assessed in conscious Sprague-Dawley rats (325 g, n = 4) following administration of a 10 mg kg-1 i.v. dose of the racemate. STL enantiomers were predominantly eliminated intact in the urine: Clr for S- and R-STL (26.3 +/- 3.2 and 28.7 +/- 4.2 mL min-1 kg-1 respectively) accounted for approximately 96% of Cls for S- and R-STL (27.5 +/- 3.3 and 29.9 +/- 4.2 mL min-1 kg-1, respectively). Approximately 4% of the dose was recovered in the faeces, corresponding to Cli values of 1.16 +/- 0.17 and 1.26 +/- 0.19 mL min-1 kg-1 for S- and R-STL, respectively. Total recovery of the administered dose in urine and faeces was 99.7 +/- 0.2 and 99.8 +/- 0.5% for S- and R-STL, respectively. It is concluded from these results in the rat model that (i) STL enantiomers are predominantly eliminated intact in urine; (ii) STL enantiomers are excreted intact in bile, and to a much larger extent in the faeces, thus suggesting the presence of intestinal exsorption of STL; (iii) STL does not appear to be metabolized; and (iv) Cls, Clr, Clb, and Cli are negligibly stereoselective.

The nifedipine gastrointestinal therapeutic system (GITS). Evaluation of pharmaceutical, pharmacokinetic and pharmacological properties.

Nifedipine is a short-acting calcium antagonist formulated into several different oral preparations, each of which offers a distinct drug release profile. Of these, the nifedipine gastrointestinal therapeutic system (GITS) affords (rate)-controlled-release (CR) and once-daily administration. Although it is recognised that CR drug formulations may enhance the treatment compliance of patients by reducing the number of daily doses, there are several pharmaceutical, pharmacokinetic and pharmacological considerations which may influence the ultimate selection of a particular dosage form. The formulation design of the nifedipine GITS involves an osmotic pump process which provides approximately zero-order delivery of the drug. This mechanism serves to prevent the possibility of dose dumping, but more importantly allows for maintenance of the relatively constant plasma drug concentrations assumed necessary to maintain smooth control of blood pressure. The pharmacokinetic characteristics of the nifedipine GITS have been evaluated in both single- and multiple-dose studies. The GITS formulation provides drug concentrations which reach a plateau within 6 hours after administration of a single dose, and continue at that concentration until at least 24 hours after administration. In this way large fluctuations in plasma drug concentrations are avoided, which may improve the efficacy and tolerability of the drug. Although a trend showing a small increase in the 24-hour plasma nifedipine concentrations has been observed by our group from some single-dose studies, it does not appear to be clinically relevant. One potentially important disadvantage of the GITS compared with 'naturally' long-acting agents is that the 'intrinsic' pharmacokinetic properties of nifedipine may be exposed in poorly compliant patients, leading to extended periods of subtherapeutic drug concentrations. Drug delivery by the nifedipine GITS is unaffected by changes in pH and gastrointestinal (GI) motility, but the rate of drug release can increase slightly with food intake (although absolute bioavailability remains unchanged). No studies have been conducted to determine the average GI transit time of this particular dosage form, but it is possible that inadequate retention may occur in some patients, perhaps leading to less optimal clinical outcomes. For example, the median GI transit time for both oxprenolol and metoprolol Oros drug delivery systems has been reported as 27.4 hours, with individual times ranging from 5.1 to 58.3 hours. The possibility of inadequate GI retention of the nifedipine GITS is perhaps more likely in patients who have pre-existing GI motility disorders or who are taking other medications that enhance GI motility. The interaction between grapefruit juice and nifedipine is interesting, considering that the exact mechanism involved has yet to be determined. Nonetheless, inhibition of presystemic metabolic processes (probably involving liver enzymes but possibly also enzymes contained within the wall of the small intestine) is likely to be a factor in the increased bioavailability of nifedipine observed in individuals coingesting grapefruit juice. Thus, potential nifedipine formulation differences with respect to the degree of interaction with grapefruit juice may occur if a significant degree of extrahepatic metabolism is involved. The majority of clinical trials with the nifedipine GITS have assessed its efficacy in patients with mild-to-moderate essential hypertension, and have found it to be at least equivalent to other dosage forms of the drug. Since there is limited information available directly comparing the efficacy and adverse effects of the different types of nifedipine formulation, little attention has been focused on this subject. However, modifying the rate and duration of nifedipine release may profoundly affect the clinical performance of this drug. A slower rate of intravenous nifedipine infusion has been shown to reduce the incide

Influence of cimetidine coadministration on the pharmacokinetics of sotalol enantiomers in an anaesthetized rat model: evidence supporting active renal excretion of sotalol.

Sotalol (STL) is an amphoteric, chiral beta-adrenergic blocking drug useful in the treatment of both hypertension and ventricular arrhythmias. In the human and rat, STL enantiomers are predominantly cleared from the body by the kidney as intact drug. The renal clearance (Clr) of STL enantiomers substantially exceeds the glomerular filtration rate (GFR) in the human and rat. In this report, the hypothesis that STL enantiomers are excreted by an active renal transport system was investigated in the rat by coadministering racemic STL (10 mg kg-1) with cimetidine, an inhibitor of renal tubular secretion of organic cations. To compare the effects of short-term and sustained cimetidine exposure on STL enantiomer disposition, cimetidine was administered either as a single bolus (30 mg kg-1, n = 7) immediately prior to the STL dose, or as a 30 mg kg-1 bolus plus a 50 mg kg-1 infusion over the 6 h study period (n = 7). Blood and urine samples were collected over 6 h, during which time anaesthesia was maintained via intraperitoneal administration of pentobarbital. Cimetidine bolus and cimetidine infusion reduced STL enantiomer Clr by 43 and 59%, respectively, compared with respective saline controls. Significant stereoselectivity was observed in the cimetidine infusion group: systemic clearance, Clr (R > S), and AUC (S > R), although the magnitude of stereoselectivity was less than 5%. This study supports the hypothesis that STL enantiomers are predominantly cleared from the rat via a renal cationic transport mechanism and that this system can be competitively inhibited by the presence of cimetidine.

Protein binding of sotalol enantiomers in young and elderly human and rat serum using ultrafiltration.

The protein binding of sotalol (STL) enantiomers was evaluated using an ultrafiltration technique with serum from young (32 +/- 2 years, n = 5) and elderly (73 +/- 6 years, n = 5) male and female humans, and young (8 weeks, n = 4) and elderly (60 weeks, n = 3) male Sprague-Dawley rats. Serum samples were collected and immediately frozen at -20 degrees C. Within 1 week, the serum samples were thawed at room temperature, and adjusted to pH 7.4 using 0.05 M phosphate buffer, pH 5.0. Aliquots were spiked with 250 ng mL-1 and 500 ng mL-1 of each STL enantiomer, placed in ultrafiltration sets (Microsep, 30K molecular weight cut-off), capped, equilibrated to 37 degrees C, and centrifuged at 1850g for 1.5 h at 37 degrees C. Aliquots of ultrafiltrate and unspun serum were analysed for STL enantiomer concentration using a stereospecific HPLC assay. In all groups, bound fraction was less than 7% for both STL enantiomers. There were no significant differences in bound fraction between groups, or between enantiomers. Adsorption of STL enantiomers to the ultrafiltration device and membrane, evaporative loss of serum samples during centrifugation, and protein concentration in each ultrafiltrate sample were all negligible. It is concluded that the binding of STL in human and rat serum at therapeutic concentrations and physiological temperature and pH is negligible and non-stereoselective.

Pharmacokinetics of verapamil and norverapamil enantiomers after administration of immediate and controlled-release formulations to humans:evidence suggesting input-rate determined stereoselectivity.

Verapamil is a racemic calcium channel-blocking drug that undergoes extensive hepatic first-pass metabolism to an active metabolite, norverapamil. The enantiomers of verapamil and norverapamil have differing negative inotropic, chronotropic, and dromotropic activities and differing effects on vascular smooth muscles; the S-enantiomers having greater activity. It is hypothesized that the R/S concentration ratio of verapamil enantiomers may be input-rate dependent. The pharmacokinetics of verapamil and norverapamil enantiomers were studied in 11 young, healthy male and female volunteers after oral administration of 80 mg immediate-release (IR) verapamil every 8 hours, and a 240 mg dose once daily of controlled-release (CR) formulation on two separate occasions. Both dosage regimens were continued for 1 week with a minimum 1-week period between the two drug treatments. After the last dose of each regimen, plasma samples were collected over the period corresponding to the dosing interval. Enantiomer concentrations were determined using a microwave-facilitated precolumn derivatization with high performance liquid chromatographic quantification. Stereospecific assay revealed that: (1) stereoselective R- and S-enantiomer disposition occurred regardless of formulation administered; (2) a trend of R:S concentration ratios of verapamil differed between the two formulations; and (3) fluctuations between Cmax and Cmin values of the two formulations were statistically different over respective dosing intervals (greater fluctuation after CR administration). Using nonstereospecific data analyses, however, the pharmacokinetic parameters for verapamil and norverapamil were similar for both formulations over a 24-hour period. We suggest that kinetic differences can be attributed to differences in release rates of drug from the tablet matrices. The relative bioavailabilities of verapamil and norverapamil from the two products may, therefore, be subject to input rate-dependent processes.

Stereospecific high-performance liquid chromatographic assay of lomefloxacin in human plasma.

This report describes an HPLC assay developed for the quantification of the enantiomers of lomefloxacin (LFLX), a quinolone antibiotic, in plasma. Following addition of racemic acebutolol (internal standard, IS), plasma samples were extracted at pH 7 with a mixture of chloroform-isopentyl alcohol-diethyl ether (71.25:3.75:25, v/v/v). The organic layer was evaporated, and LFLX and IS enantiomers in the resulting residue were derivatized with chloroform solutions of 1% triethylamine and 1% (S)-(+)-(1-naphthyl)ethyl isocyanate, followed by 2% ethyl chloroformate (ECF) 1 min later. Ethanolamine was added 30 s after the addition of ECF. The enantiomers were separated as diastereomers on an 8 x 100 mm Radial Pak normal phase column using a mobile phase of hexane-chloroform-methanol (64.5:33:2.5, v/v/v) pumped at 2.0 ml min-1. The IS was detected by fluorescence at 245 and 420 nm (excitation and emission, respectively) during the first 12 min, after which time the wavelengths were 280 and 470 nm for detection of LFLX. The method: (1) was sensitive and showed excellent linearity (10-1000 ng ml-1, r2 > 0.99) between added enantiomer concentrations and peak-area-ratio (LFLX/IS); and (2) separated LFLX and IS enantiomers within 25 min. The assay is suitable for the quantification of LFLX enantiomers in plasma samples.