Application of microdialysis for the determination of muscle and subcutaneous tissue concentrations after oral and topical ibuprofen administration.

BACKGROUND: The topical administration of non-steroidal antiinflammatory drugs (NSAIDs) is widely used for the treatment of soft tissue pain. However, it is not known whether effective tissue concentrations are reached with the topical route. OBJECTIVE: To evaluate and compare unbound muscle and subcutaneous tissue ibuprofen concentrations with use of microdialysis after topical and oral administration. METHODS: In a 2-way crossover design, 11 healthy volunteers received either 800 mg oral ibuprofen or 16 g of 5% ibuprofen gel applied onto the skin of the thigh (defined area, 17 x 19 cm). Microdialysis catheters were inserted into the medial vastus muscle (25 to 30 mm) and into the subcutaneous adipose layer of the thigh (4 to 5 mm). Dialysate was collected in 20-minute intervals up to 5 hours. RESULTS: Essentially all of the orally administered dose was recovered in urine as ibuprofen or metabolites during 24 hours, but only about 0.55% of the topically administered dose was recovered. The relative systemic bioavailability of ibuprofen gel, based on urine recovery data, was (mean +/- SD) 0.57%+/-0.30%. Mean values of the dialysate areas under the drug concentration-time curves after topical and oral administration were 731.2+/-605.0 and 176.6+/-122.9 ng x h x mL(-1) for subcutaneous tissue and 63.5+/-90.3 and 213.4+/-117.2 ng x h x mL(-1) for muscle, respectively. Muscle dialysate concentrations after topical administration varied considerably among the subjects. CONCLUSION: These results suggest that, if target tissue concentrations correlate directly with the degree of pain relief, patients with pain caused by dermal or subcutaneous tissue damage will have greater pain relief after topical administration of ibuprofen accompanied with less systemic side effects. In addition, a proportion of patients with muscle pain may also experience pain relief from topical ibuprofen.

Synthesis and biological evaluation of substituted flavones as gastroprotective agents.

Flavone (1) was found to protect against ethanol-induced gastric damage in rats; however, it is known that certain compounds in the flavone class, including flavone itself, are inducers of hepatic drug metabolizing enzymes. With the hope of identifying gastroprotective flavones that have minimal effects on drug metabolizing enzymes, we have synthesized and evaluated selected flavone analogs. Gastroprotective potency in the ethanol model was retained by methoxy substitution in the 5-position (4) and by methoxy (12) or methyl (14) substitution in the 7-position. A number of substituted analogs of the potent molecule 5-methoxyflavone (4) were also synthesized, and in many cases, these substitutions provided gastroprotective molecules. In order to assess liver enzyme induction potential, two of the gastroprotective flavones, 7-methoxyflavone (12) and 5-methoxy-4'-fluoroflavone (26), were examined for their effect on liver microsomal cytochrome P450 and 7-ethoxyresorufin O-dealkylase (CYP1A) activity. These two compounds caused minimal changes in the cytochrome P450 concentration and were considerably less potent than beta-naphthoflavone as inducers of CYP1A enzyme activity. Furthermore, following oral administration to rats, 5-methoxy-4'-fluoroflavone (26) was found to protect against indomethacin-induced gastric damage. These results indicate that, through appropriate substitution, flavones can be obtained that are gastroprotective but have minimal effects on drug-metabolizing enzymes.

Celiac vagotomy reduces suppression of feeding by jejunal fatty acid infusions.

We investigated the role of the celiac branch of the vagus nerve in suppression of food intake produced by jejunal fatty acids infusions. Following selective celiac vagotomy or sham surgery, adult, male Sprague-Dawley rats received 7 h infusions of linoleic acid or saline through indwelling jejunal catheters on four consecutive days. Although linoleic acid still produced significant suppression of intake in rats with celiac vagotomy, it was less effective in these animals than in controls. The temporal pattern of results suggested that celiac afferent fibers are involved in mediating both pre- and postabsorptive effects of infused fatty acids.

Jejunal or portal vein infusions of lipids increase hepatic vagal afferent activity.

Jejunal infusions of linoleic acid, corn oil, or caprylic acid significantly increased hepatic vagal afferent activity, whereas saline infusions were ineffective. The magnitude of response was greatest with either linoleic acid or corn oil. Hepatic portal infusions of linoleic acid, Liposyn II, or caprylic acid significantly increased hepatic vagal afferent activity, whereas 5% albumin/phosphate buffer vehicle was ineffective. The magnitude of response was greatest with either linoleic acid or Liposyn II. These data show that either jejunal or portal infusions of lipids increase activity of hepatic vagal afferents and could potentially serve as a complementary and/or alternative substrate to celiac vagal afferents in mediating the effects of jejunal infusions of lipids in suppressing food intake.

Comparison of tebufelone distribution in rat blood plasma and inflamed and noninflamed tissues following peroral and intravenous administration.

It is recognized that some acidic nonsteroidal antiinflammatory drugs (NSAIDs) accumulate in the synovial fluids of inflamed joints. The distribution of tebufelone, a member of the di-tert-butylphenol class of NSAIDs, between rat plasma and paw tissues (inflamed and noninflamed) was determined. Tebufelone was administered (doses providing 80% maximal effectiveness) perorally (po, 10 mg/kg) or intravenously (i.v., 0.5 mg/kg) to Sprague-Dawley rats just prior to injection of an inflammatory adjuvant (carrageenan) into one of the hind paws. Levels of the drug were measured in plasma, inflamed paws, and noninflamed paws at prescribed times postdosing by capillary gas chromatography/stable isotope dilution mass spectrometry. Tebufelone levels, as determined from areas under the curves of concentration versus time were sevenfold (po) and 11-fold (i.v.) higher in paw tissue than in plasma. Inflamed and noninflamed tissues have equal affinity for the drug. Calculated terminal-phase half-lives of tebufelone in paw tissue were similar following i.v. and po dosing (approximately 14 h) and equivalent to the plasma half-life of the drug after po dosing (approximately 10 h). The plasma half-life determined following i.v. dosing was considerably lower (2.8 h). The anti-inflammatory activity of tebufelone appears to correlate more closely with tissue distribution profiles than plasma drug levels at the dose levels examined.

Evaluation of ketorolac concentrations in plasma and gingival crevicular fluid following topical treatment with oral rinses and dentifrices.

Two clinical studies were conducted to determine the relative amounts of ketorolac detectable locally in the gingival crevicular fluid (GCF) and systemically in plasma after oral, topical drug administration. The rinse study compared topical administration of three concentrations of ketorolac tromethamine (0.1%, 0.5%, and 0.01%) in oral rinse formulations administered topically and a perorally administered capsule (10 mg), and the dentifrice study compared two concentrations of ketorolac in dentifrice formulations (0.15% and 1.0%) with a 0.1% oral rinse, all treatments administered topically. The dose-corrected systemic availability of the three oral rinses evaluated in the rinse study relative to the peroral capsule was about 15%. However, the ratios of the observed maximum GCF ketorolac concentration to maximum plasma ketorolac concentration ranged from 22 to 49, compared to less than 1 for the peroral ketorolac capsule. Using this ratio as an estimate of the ability of a treatment to target the drug to the gingival tissue, these data indicate that the ketorolac oral rinses achieved greater delivery of drug to the gingival tissue (presumed site of action for periodontitis) with a lower systemic drug load than peroral administration of a ketorolac capsule. The dose-corrected relative systemic bioavailabilities for the dentifrice treatments with respect to the 0.1% rinse in the dentifrice study were 59.2% and 86.4% for the 1.0% and 0.15% dentifrices, respectively, indicating that significantly less ketorolac was systemically available from the two dentifrices relative to the oral rinse. The relative bioavailabilities of ketorolac in the GCF after dosing with the dentifrice formulations with respect to the rinse were 89.1% for the 1.0% dentifrice and 19.7% for the 0.15% dentifrice. Thus, the 1.0% dentifrice appears to provide statistically equivalent levels of ketorolac to the gingival tissue as the 0.1% oral rinse with significantly less systemic exposure. The T1/2 of ketorolac in the GCF was about 0.5 h for all three treatments, which is significantly less than the plasma half-life of about 5.3 h. These data suggest that GCF levels of ketorolac should remain above the IC50 for PGE2-stimulated IL-1 bone resorption for about 7 h following treatment, assuming continuation of the first-order elimination observed over the first two postdosing hours. We conclude that oral rinses and dentifrices are effective and preferred vehicles for administration of ketorolac for use in treatment of periodontitis.

Absorption, bioavailability, and pharmacokinetics of tebufelone in the rat.

Tebufelone (NE-11740) is a member of the new di-tert-butylphenol class of anti-inflammatory agents. It exhibits good inhibitory activity against cyclooxygenase and 5-lipoxygenase in vitro. It also shows excellent anti-inflammatory activity and inhibits bone resorption in vivo in the rat adjuvant arthritis model at an oral dose level of 1 to 2 mg/kg. The absorption, bioavailability, and pharmacokinetics of tebufelone were investigated in male Sprague-Dawley rats. Tebufelone labeled with carbon-14 was administered intravenously at doses of 0.5 and 2 mg/kg and perorally at doses of 2 and 10 mg/kg to fasted rats. Plasma samples taken from the rats at timed intervals were analyzed for total radiolabel by scintillation counting and for tebufelone by a mass spectrometric method. Comparison of the total radiolabel and tebufelone areas under the curves (AUCs) of concentration of tebufelone versus time from the 2-mg/kg intravenous and 2-mg/kg oral doses indicates that tebufelone is completely absorbed and 100% bioavailable at this dose level in the rat. The AUCs are a linear function of dose at the 0.5- and 2-mg/kg dose levels, but the AUC of the 10-mg/kg dose exhibits a nonproportional increase, suggesting saturation of elimination processes at this higher dose.

Automated gas chromatography/tandem mass spectrometry assay for tebufelone and a 13C,(18)O-labeled analog in plasma: applicabilityto absolute bioavailability determination.

An automated capillary gas chromatography/tandem mass spectrometry (GC/MS/MS) assay for the simultaneous quantitation of tebufelone (TE) and 13C, (18)O-labeled TE (TE-CO) in plasma was developed. This method permits the use of stable isotope coadministration (TE and TE-CO dosed concurrently via peroral and intravenous routes, respectively) for the determination of TE absolute bioavailability. The selectivity of MS/MS conducted on a triple-quadrupole instrument allowed minimal sample preparation and rapid analysis. Electron ionization produced molecular ions (M+) for TE, TE-CO, and the 3-methyl-TE internal standard which were selected in Q1 to undergo collisionally activated dissociation in Q2. Quantitation was achieved through monitoring product ions at m/z 248, 251 and 248, respectively, in Q3. A 2-1000 ng per sample (40 pg to 20 ng injected) quantitation range provided access to an effective 1-5000 p.p.b. plasma concentration range (0.2-2 g samples) for both TE and TE-CO. The assay showed no bias and less than 8% relative standard deviation over the entire range. The method was used to determine plasma levels of TE and TE-CO in four dogs receiving 2.5:2.5 mg/kg TE:TE-CO, intravenously. The pharmacokinetics of both isotopomers proved to be identical, indicating no isotope effect and verifying the chemical stability of the (18)O-carbonyl label under these dosing conditions. In addition, the applicability of this analytical approach to the determination of TE peroral bioavailability was initially tested in dogs.

Interspecies scaling of tebufelone pharmacokinetic data and application to preclinical toxicology.

PURPOSE: Retrospective application of allometric scaling techniques to tebufelone, a nonsteroidal antiinflammatory agent, in order to better understand the systemic exposure relationships between the doses administered to the species used in toxicology studies and the doses given to human subjects and patients in clinical studies. METHODS: Non-compartmental estimates of tebufelone's total body volume of distribution during the terminal phase (Vz) and clearance (CL) obtained from intravenous dosing to rat, monkey, dog, and human were allometrically scaled to body weight, and body weight and brain weight, respectively. AUCs determined from single or multiple dose pharmacokinetic studies and from preclinical toxicology studies were plotted versus dose adjusted for bioavailability and divided by allometrically scaled clearance to produce an allometric relationship suggesting a non-linear increase in AUC with dose across the four species. RESULTS: Segmental linear regression analysis of this relationship indicates a change point associated with an AUC of approximately 2,300 ng-hr/mL. Elevations in serum levels of various liver enzymes or associated signs of hepatic toxicity occur in some, but not all of the animals exposed for more than three weeks in repeat dosing studies at the actual dose that this represents. CONCLUSIONS: The analysis suggests that doses producing tebufelone plasma levels above a certain threshold AUC and duration of exposure to parent tebufelone are associated with increased risks of hepatic effects. Whether this is because metabolic shifts occur at these doses cannot be determined from these data.

Determination of two mebeverine metabolites, mebeverine alcohol and desmethylmebeverine alcohol, in human plasma by a dual stable isotope-based gas chromatographic-mass spectrometric method.

A dual stable isotope-based GC-MS method was developed for the simultaneous determination of two metabolites of mebeverine, mebeverine alcohol and desmethylmebeverine alcohol, in human plasma. Plasma samples were treated with beta-glucuronidase to cleave the glucuronide conjugates of both compounds prior to analysis. The treated plasma was prepared for analysis by solid-phase extraction using octadecylsilane cartridges. The isolated metabolites were derivatized and analyzed by GC-MS using selected-ion monitoring. Plots of peak-area ratio were linear with metabolite concentration from 2 to 200 ng/ml and the limit of detection for both metabolites was 0.5 ng/ml. The GC-MS methodology was applied to the analysis of plasma from human subjects following peroral administration of mebeverine. Pharmacokinetic parameters for both metabolites were determined and suggest that relative systemic mebeverine exposure may potentially be assessed using metabolite kinetics, if the latter subsequently are demonstrated to be linear with mebeverine dose.

Responses of celiac and cervical vagal afferents to infusions of lipids in the jejunum or ileum of the rat.

Multiunit celiac and single-unit cervical recordings of vagal afferents were performed before and during infusions of fatty acids, triglycerides, or saline into either the ileum or jejunum of the rat. In multiunit recordings, lipids increased activity of vagal afferents to a greater extent than saline. The greatest increases in vagal afferent activity resulted from infusions of linoleic acid, conjugated linoleic acid, or oleic acid. The triglycerides, corn oil or Intralipid, were less effective than the fatty acids in affecting vagal afferent activity. Ileal pretreatment with the hydrophobic surfactant Pluronic L-81 significantly attenuated the response of celiac vagal afferents to ileal infusion of linoleic acid. Single-unit recordings of cervical vagal afferents supported the multiunit data in showing lipid-induced increased vagal afferent activity in approximately 50% of ileal units sampled and 100% of a limited number of jejunal units sampled. These data demonstrate that free fatty acids can activate ileal and jejunal vagal afferents in the rat, and this effect can be attenuated by pretreatment with a chylomicron inhibitor. These data are consistent with the view that lipid-induced activation of vagal afferents could be a potential substrate for the inhibitory effects of intestinal lipids on gastrointestinal function, food intake, and body weight gain.

Comparison of tissue concentrations after intramuscular and topical administration of ketoprofen.

PURPOSE: To assess whether topical ketoprofen, which has been reported to provide analgesic effects in clinical studies, reaches predictable tissue concentrations high enough to account for the reported analgesia. Intramuscular ketoprofen was used as positive control. METHODS: Muscle and subcutaneous tissue concentrations were assessed by microdialysis. Plasma and tissue concentrations after intramuscular injection were described using a three-compartment population pharmacokinetic model. The prediction performance of the model was assessed by superimposing tissue concentrations of 12 subjects that did not participate in the present study. RESULTS: Most dialysate concentrations after topical dosing of ketoprofen (100 mg) were below the quantification limit of 0.47 ng/ml. Plasma concentrations increased slowly and reached an apparent plateau of 7-40 ng/ml at 10-12h. No decline was observed up to 16 h. Tissue concentrations after intramuscular injection (100 mg) were about 10 times higher than those after topical dosing. Tissue concentrations measured in the majority of the 12 subjects that did not participate in the present study were found within the range of two-thirds of the predicted concentrations. CONCLUSION: Predictable and cyclooxygenase-inhibiting concentrations of ketoprofen were achieved in subcutaneous and muscle tissue after intramuscular but not after topical dosing. Thus, the tissue concentrations of ketoprofen after topical administration can hardly explain the reported clinical efficacy of topical ketoprofen.

Suppression of food intake, body weight, and body fat by jejunal fatty acid infusions.

Three experiments investigated effects of jejunal lipid infusions given on 4 or 21 consecutive days in adult, male Sprague-Dawley rats. In experiment 1, 7-h infusions of linoleic or oleic acid (0.2 ml/h for 7 h; total load = 11.5 kcal) on 4 consecutive days reduced total intake (ad libitum consumption of the liquid diet Boost, Mead Johnson, plus load) by approximately 15% and decreased weight gain compared with 4-day tests with saline administration. In experiment 2, linoleic acid at 0.1 ml/h for 7 h (5.7 kcal) was ineffective, whereas the same load delivered in 3.5 h produced effects similar in magnitude to those in the first experiment. In experiment 3, jejunal infusions of linoleic acid (0.2 ml/h for 7 h) on 21 consecutive days reduced mean total intake by 16%, body weight by 10%, and carcass fat by 48% compared with controls receiving saline. The net decrease in caloric intake may reflect the combined activation of pre- and postabsorptive mechanisms, and it suggests a possible treatment for obesity.

The antitussive effect of dextromethorphan in relation to CYP2D6 activity.

AIMS: To test the hypothesis that inhibition of cytochrome P450 2D6 (CYP2D6) by quinidine increases the antitussive effect of dextromethorphan (DEX) in an induced cough model. METHODS: Twenty-two healthy extensive metaboliser phenotypes for CYP2D6 were studied according to a double-blind, randomised cross-over design after administration of: (1) Placebo antitussive preceded at 1 h by placebo inhibitor; (2) 30 mg oral DEX preceded at 1 h by placebo inhibitor (DEX30); (3) 60 mg oral DEX preceded at 1 h by placebo inhibitor (DEX60); (4) 30 mg oral DEX preceded at 1 h by 50 mg oral quinidine sulphate (QDEX30). Cough frequency following inhalation of 10% citric acid was measured at baseline and at intervals up to 12 h. Plasma concentrations of DEX and its metabolites were measured up to 96 h by h.p.l.c. RESULTS: Inhibition of CYP2D6 by quinidine caused a significant increase in the mean ratio of DEX to dextrorphan (DEX:DOR) plasma AUC(96) (0.04 vs 1.81, P<0.001). The mean (+/-s.d.) decrements in cough frequency below baseline over 12 h (AUEC) were: 8% (11), 17% (14.5), 25% (16.2) and 25% (16.9) for placebo, DEX30, DEX60 and QDEX30 treatments, respectively. Statistically significant differences in antitussive effect were detected for the contrasts between DEX60/placebo (P<0.001; 95% CI of difference +80, +327) and QDEX30/placebo (P<0.001, +88, +336), but not for DEX30/placebo, DEX30/DEX60 or DEX30/QDEX30 (P=0.071, -7, +241; P=0.254, -37, +211; P=0.187, -29, +219, respectively). CONCLUSIONS: A significant antitussive effect was demonstrated after 60 mg dextromethorphan and 30 mg dextromethorphan preceded by 50 mg quinidine using an induced cough model. However, although the study was powered to detect a 10% difference in cough response, the observed differences for other contrasts were less than 10%, such that it was possible only to imply a dose effect (30 vs 60 mg) in the antitussive activity of DEX and enhancement of this effect by CYP2D6 inhibition.