Comparative pharmacokinetic and bioequivalence evaluation of two formulations of morniflumate 350-mg tablets in healthy male subjects
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BACKGROUND: Morniflumate is a nonsteroid anti-inflammatory drug (NSAID) that inhibits cyclooxygenase-1, 2 (COX-1, 2). OBJECTIVE: This study aimed to compare the pharmacokinetics (PKs) and assess the bioequivalence of two different formulations of morniflumate 350-mg tablets in healthy Korean male subjects. METHODS: A randomized, single-dose, two-period, two-sequence crossover study was conducted with 38 subjects. Subjects received a single dose of two tablets of either a test or a reference formulation and the alternated formulation in the next period. Serial blood samples for the PK analysis were collected over 12 hours. PK parameters were determined by a noncompartment analysis. PK parameters, including the maximum concentration (Cmax) and the area under-the-concentration-time curve from time zero to the time of the last quantifiable concentration (AUClast) were compared in bioequivalence tests. RESULTS: The Cmax of the test and reference formulations were 985.72 ± 6.80 mg/L and 947.09 ± 6.73 mg/L, respectively, while the AUClast values were 2675.92 ± 7.84 mg×h/L and 2653.06 ± 7.78 mg×h/L, respectively. The geometric mean ratios (90% confidence interval) of the test formulation to the reference formulation for Cmax and AUClast were 1.0715 (0.9469 - 1.2124) and 1.0592 (0.9592 - 1.1695), respectively. CONCLUSIONS: The new formulation of morniflumate 350-mg tablet showed a PK profile similar to that of the marketed formulation, and the results of this study fell within in the conventional criteria of bioequivalence.
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Simultaneous determination of morniflumate and its major active metabolite, niflumic acid, in human plasma by high-performance liquid chromatography in stability and pharmacokinetic studies.

A rapid, sensitive and stable high-performance liquid chromatography (HPLC) method was developed and validated for the simultaneous determination of morniflumate and its major active metabolite, niflumic acid, in human plasma. HPLC analysis was carried out using a 5 µm particle size, C18 -bonded silica column with a mixture of acetonitrile and 0.005 m potassium phosphate monobasic in water (60:40, v/v) as the mobile phase and UV detection at 287 nm. The method involved the treatment with 50 µL of 0.4 m hydrochloric acid for the stability of morniflumate, extraction with diethylether and evaporation to dryness under a nitrogen stream. The lower limit of quantitation for morniflumate and niflumic acid was 50 and 500 ng/mL, respectively. The calibration curves for morniflumate and niflumic acid were linear over the concentration range of 50-20,000 ng/mL and 500-50,000 ng/mL, respectively, with correlation coefficients greater than 0.9995 and inter- or intra-batch coefficients of variation not exceeding 13.79%. The variability (percentage difference) of incurred sample re-analysis did not exceed 11.72% and all of the repeat samples fell within 20% of the mean value. This assay procedure was applied successfully to an examination of the pharmacokinetics of morniflumate and its metabolite, niflumic acid, in human subjects.

Contribution of pH to systemic exposure of niflumic acid following oral administration of talniflumate.

PURPOSE: Systemic exposure to niflumic acid was significantly increased when talniflumate was given orally together with a meal. To clarify the underlying mechanism, an in vitro dissolution study of talniflumateonducted at different pH values, and magnesium hydroxide was co-administered in healthy volunteers. METHODS: In vitro dissolution tests of talniflumate tablets were performed in a USP Paddle apparatus at pH 1.2, 4.0, and 6.8, respectively, in the presence and absence of Tween 80 (2%). Serial samples of the talniflumate solution were taken and analyzed on a high-performance liquid chromatography (HPLC)/ultraviolet system. Healthy volunteers were divided randomly into two groups, and each volunteer received a single 740-mg dose of talniflumate, with or without 1 g of magnesium hydroxide, following an overnight fast. The plasma concentrations of niflumic acid were measured using HPLC coupled with tandem mass spectrometry. RESULTS: Talniflumate was completely insoluble at each of the tested pHs in the absence of Tween 80. The drug was slowly and steadily dissolved (54%) at pH 4 in the presence of the surfactant, but the extent of dissolution was only 15 and 0.5% at pH 1.2 and 6.0, respectively. Magnesium hydroxide co-administered with talniflumate significantly increased systemic exposure to niflumic acid: the mean maximum plasma concentration (C (max)) and area under the concentration-time curve (AUC (inf)) were augmented by 2.0- and 1.9-fold, respectively, compared with those in the absence of the antacid. Magnesium hydroxide significantly accelerated the appearance of niflumic acid in plasma by 2.8-fold. CONCLUSIONS: Magnesium hydroxide increases the rate and extent of systemic exposure to niflumic acid owing to the enhanced solubility of talniflumate and absorption of niflumic acid. The possible combination of talniflumate and an antacid should be considered in the development of pharmaceutical formulations.

Determination of talniflumate and niflumic acid in human plasma by liquid chromatography-tandem mass spectrometry.

A simple and rapid quantification method was developed for determining both talniflumate and niflumic acid in human plasma. After simple protein precipitation with acetonitrile, the analytes were chromatographed on a reversed-phase C(18) column and detected by LC/MS/MS with electrospray ionization. The assay accuracy and precision were within the FDA guidance for the analytical method validation. This method was used to measure the plasma concentrations of both compounds from healthy subjects after a single oral dose of talniflumate, 740 mg.

Liquid chromatography-mass spectrometric method for the sensitive determination of niflumic acid in human plasma and its application to pharmacokinetic study of talniflumate tablet.

A sensitive LC-MS method was developed and validated for the determination of niflumic acid (NFA), the active metabolite of the talniflumate formulation, in human plasma. The analyses were performed on C(18) column using acetonitrile-ammonium acetate buffer (pH 5.7, 40:60) as a mobile phase with quadrupole MS detection of NFA at m/z 281 in a negative ion-monitoring mode. Calibration curve was linear in the concentration range of 1-1000ng/mL in human plasma. The higher sensitivity of LC-MS allowed low concentrations of NFA to be determined at initial drug absorption and terminal elimination phases following oral administration of talniflumate tablet.

Effect of food on systemic exposure to niflumic acid following postprandial administration of talniflumate.

PURPOSE: Talniflumate was designed as a prodrug of niflumic acid, a potent analgesic and anti-inflammatory drug, which is widely prescribed for treating rheumatoid diseases. The prandial effect on talniflumate absorption remains unclear; therefore, this study investigated the effect of food on the systemic exposure to niflumic acid in healthy volunteers. METHODS: Volunteers received a single 740-mg dose of talniflumate 30 min after consuming a high-fat breakfast, a low-fat breakfast, or no food (fasting condition). Plasma concentrations of both talniflumate and niflumic acid were measured using validated high-performance liquid chromatography coupled to tandem mass spectrometry. RESULTS: The maximum concentration of niflumic acid was 224 +/- 193 ng/ml at approximately 2.7 h in the fasted condition compared with 886 +/- 417 ng/ml (p < 0.05) at 1.8 h and 1,159 +/- 508 ng/ml (p < 0.01) at 2.2 h with the low- and high-fat meals, respectively. The mean area under the curve from zero to infinity (AUC(inf)) values after the low- and high-fat meals were four- and fivefold, respectively, the value while fasting (p < 0.05). CONCLUSIONS: It is strongly recommended that talniflumate be taken after a meal to increase systemic exposure to its active metabolite. Our results suggest a reduction in the daily dosage of talniflumate when taken with food.

(19)F NMR spectroscopic characterization of the interaction of niflumic acid with human serum albumin.

The interaction of a non-steroidal anti-inflammatory drug, niflumic acid (NFA), with human serum albumin (HSA) has been investigated by (19)F nuclear magnetic resonance (NMR) spectroscopy. A (19)F NMR spectrum of NFA in a buffered (pH 7.4) solution of NaCl (0.1 mol L(-1)) contained a single sharp signal of its CF(3) group 14.33 ppm from the internal reference 2,2,2-trifluoroethanol. Addition of 0.6 mmol L(-1) HSA to the NFA buffer solution caused splitting of the CF(3) signal into two broadened signals, shifted to the lower fields of 14.56 and 15.06 ppm, with an approximate intensity ratio of 1:3. Denaturation of HSA by addition of 3.0 mol L(-1) guanidine hydrochloride (GU) restored a single sharp signal of CF(3) at 14.38 ppm, indicating complete liberation of NFA from HSA as a result of its denaturation. These results suggest that the binding is reversible and occurs in at least two HSA regions. Competitive (19)F NMR experiments using warfarin, dansyl-L: -asparagine, and benzocaine (site I ligands), and L: -tryptophan and ibuprofen (site II ligands) revealed that NFA binds to site I at two different regions, Ia and Ib, in the ratio 1:3. By use of (19)F NMR with NFA as an (19)F NMR probe the nonfluorinated site I-binding drugs sulfobromophthalein and iophenoxic acid were also found to bind sites Ia and Ib, respectively. These results illustrate the usefulness and convenience of (19)F NMR for investigation of the HSA binding of both fluorinated and nonfluorinated drugs.

Stability studies of some glycolamide ester prodrugs of niflumic acid in aqueous buffers and human plasma by HPLC with UV detection.

Glycolamide esters (compounds 1-17) of 2-(3-trifluoromethyl-phenylamino)nicotinic acid (niflumic acid, CAS 4394-00-7) have been synthesized and evaluated as possible prodrugs. In-vitro hydrolysis studies were conducted at selected pH values (1.2, 3.5, 4.8, 7.4 and 7.8) and in human plasma at 37 +/- 0.5 degree C using HPLC with UV detection. The aqueous (pH 7.4 and 7.8) and enzymatic rates of hydrolysis were substantially affected by the nature of promoieties in this series. The compounds showed good chemical stability in the buffers of low pH values (1.2, 3.5 and 4.8) and appreciable hydrolysis under alkaline conditions and in human plasma. They exhibited long hydrolytic half-lives of 7-46 h in aqueous buffer solutions (pH 7.4 and 7.8) and 14-21 min in human plasma, respectively. It was observed that N,N-disubstituted and cyclic glycolamide derivatives showed 2 fold more hydrolysis in the alkaline pH than monosubstituted derivatives, whereas the piperidino and thiomorpholino derivatives did not undergo chemical hydrolysis. The compounds contain two possible sites for hydrolysis with an increased hydrolytic susceptibility at the terminal aliphatic carbonyl site in aqueous buffers and human plasma solutions. They were found to be cleaved at two hydrolytic carbonyls, namely the nicotinyl (2-5 % in enzymatic hydrolysis) and the aliphatic site (7-55 % and 70-85 % in buffer and plasma hydrolysis, respectively) as revealed by HPLC analysis. The glycolamide ester prodrugs of niflumic acid underwent chemical and enzymatic hydrolysis to release mainly the metabolite 2-(3-trifluoromethyl-phenylamino) nicotinic acid carboxymethyl ester (III) and not the parent drug 2-(3-trifluoromethyl-phenylamino)nicotinic acid. The structure of the metabolite was confirmed by liquid chromatography-mass spectroscopy (LCMS).

Quantitation of niflumic acid in human plasma by high-performance liquid chromatography with ultraviolet absorbance detection and its application to a bioequivalence study of talniflumate tablets.

A rapid and simple HPLC method with UV detection (288 nm) was developed and validated for quantitation of niflumic acid in human plasma, the active metabolite of talniflumate. After precipitation with 100% methanol containing the internal standard, indomethacin, the analysis of the niflumic acid level in the plasma samples was carried out using a reverse phase C18 CAPCELL PAK (5 microm, 4.6 mm x 250 mm) column. The chromatographic separation was accomplished with an isocratic mobile phase consisting of a mixture of 0.1M sodium acetate in water and acetonitrile (37:63, v/v), adjusted to pH 6.4. This HPLC method was validated by examining its precision and accuracy for inter- and intra-day runs in a linear concentration range of 0.02-5.00 microg/mL. Stability of niflumic acid in plasma was excellent, with no evidence of degradation during sample processing (autosampler) and 30 days storage in a freezer. This validated method was successfully applied to the bioequivalence study of talniflunate in healthy volunteers.

Simultaneous determination of niflumic acid and its prodrug, talniflumate in human plasma by high performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method has been developed for the simultaneous determination of niflumic acid and its prodrug, talniflumate, in human plasma. Niflumic acid and talniflumate were eluted isocratically with methanol-water (73:27, v/v, adjusted to pH 3.5 by acetic acid) at a fl ow rate of 1 mL/min. Indomethacin was used as an internal standard. Signals were monitored by an UV detector at 288 nm. Retention times of indomethacin, niflumic acid and talniflumate were 5.9, 7.2 and 13.5 min, respectively. Calibration plots were linear over the range 50-5000 ng/mL for niflumic acid and 100-5000 ng/mL for talniflumate. The limits of quantitation were 50 ng/mL for niflumic acid and 100 ng/mL for talniflumate. The intra- and inter-day relative standard deviations (RSD) of niflumic acid and talniflumate were less than 10% and the accuracies were higher than 90%. This method is rapid, sensitive and reproducible for the determination of niflumic acid and talniflumate in human plasma.

Detection of the non-steroidal anti-inflammatory drug niflumic acid in humans: a combined 19F-MRS in vivo and in vitro study.

This study describes for the first time results of a (19)F-MRS study on humans exposed to the fluorinated non-steroidal anti-inflammatory drug niflumic acid. The accumulation and elimination of this commercially available selective prostaglandin synthase inhibitor is studied after an oral bolus in the human liver, in blood plasma and in urine samples. The in vivo spectra of the liver display two resonances with a similar increase in signal intensity during the investigation period of 240 min. One resonance refers to the parent compound niflumic acid (P), whereas the second resonance corresponds to a metabolite (M1) formed by the biotransformation by liver enzymes. The spectroscopic comparison with model compounds suggests 4'-hydroxyniflumic acid as the metabolite. During the entire experiment the concentration ratios of these resonances (P/M1) ranged between 0.7 and 0.9, indicating a high metabolite concentration most probably due to an efficient first pass metabolism. Both resonances (P, M1) were observed in the in vitro study of the blood plasma samples after plasma protein denaturation. However, in comparison to the liver spectra, the amount of the metabolite M1 is very small with a P/M1-ratio of 36.6 after 90 min and 16.1 after the end of measurement. This finding suggests an efficient biliary excretion of the metabolite M1, which bypasses the blood circulation system. Both resonances are also identified in the native urine samples. The signal intensity of the parent compound dominates the spectra of all urine samples, whereas the signal intensity of M1 increases slowly reaching a similar value to the parent compound P at the end of the measurement. This observation demonstrates an effective renal elimination of niflumic acid and suggests the existence of an enterohepatic circuit with a re-entry mechanism for the biliary excreted metabolite M1. In the urine spectra, an additional metabolite M2 is found. This resonance exhibits a low but constant signal intensity. The chemical origin of this metabolite is unclear.

Evaluation of several solid phase extraction liquid chromatography/tandem mass spectrometry on-line configurations for high-throughput analysis of acidic and basic drugs in rat plasma.

Several configurations using 6- and 10-port switching valves were studied for high flow, on-line extraction of rat plasma coupled to an electrospray triple quadrupole mass spectrometer. Each plasma sample was diluted 1:1 with an aqueous internal standard solution. The sample was injected into a 2.1 x 20 mm cartridge column packed with 25 microm divinylbenzene/N-vinylpyrrolidone packing using 100% aqueous mobile phase at 4 mL/min. After sample loading and sample cleanup, the analytes were eluted from the extraction column with a 1.0-min gradient at 0.4 mL/min. The samples were either analyzed directly after elution from the extraction column or after additional separation using a short high performance liquid chromatography (HPLC) column. The different configurations were tested using an acidic drug (diflunisal) and a basic drug (clemastine) in rat plasma. On-line analysis was performed by injecting 200 microL of diluted plasma. The mass spectrometer was operated in the multiple reaction monitoring (MRM) mode. All calibration standards gave relative standard deviations (RSDs) below 5%. The total time per sample was 3 min.

Pharmacokinetic parameters and killing rates in serum of volunteers receiving amoxicillin, cefadroxil or cefixime alone or associated with niflumic acid or paracetamol.

Pharmacokinetic parameters and killing rates in serum of volunteers receiving amoxicillin, cefadroxil or cefixime alone or associated with niflumic acid or paracetamol were studied. Niflumic acid (250 mg) or analgesic and antipyretic drugs such as paracetamol (500 mg) are often combined with antibiotics to avoid inflammation and pain in acute ear, nose and throat diseases. Pharmacokinetic interactions between these two classes of drugs have been described in experimental models, and exceptionally in humans. The aim of the present investigation was to study the interactions of these two drugs with three antibiotics (amoxicillin 500 mg x 2, cefadroxil 500 mg x 2, cefixime 200 mg and one placebo capsule) on pharmacodynamic parameters and on rate of killing in the serum of six healthy volunteers receiving the antibiotic associated or not with the product in a randomized cross-over double-blind trial. The bacteria most often involved in sinusitis, bronchitis and otitis media (Haemophilus influenzae, Streptococcus pneumoniae, Staphylococcus aureus) three target diseases for oral cephalosporins and amoxicillin, were chosen for bacteriological study. Blood samples were obtained at 0.25, 0.50, 1, 1.5, 2, 4, 6 and 12 h after oral administration of antibiotics alone or associated with the drugs. There was a wash-out period of at least 1 week between the eleven sequences. Antibiotics were measured by two methods: bioassay and high performance liquid chromatography (HPLC). All serum samples obtained at peak level, 4 and 6 h were tested for killing rate. Area under the time kill curve was calculated by the trapezoidal rule method and relative bioactivity in percent was defined as follows: (AUC control - AUC test)/AUC control x 100. No pharmacokinetic interaction was found in the AUC and T1/2 of the plasma concentrations of the antibiotics or associated with the drugs, regardless of dose, as determined by HPLC or microbiological assay. For these beta-lactam antibiotics killing rate was found to be time-dependent. Bactericidal activity was improved on H. influenzae when cefixime was associated with niflumic acid and became concentration-dependent. A significant concentration relation was also found with niflumic acid or paracetamol associated with cefixime on Strep. pneumoniae.

[Non-steroidal anti-inflammatory agents and pregnancy. A study of renal and digestive toxicity of niflumic acid in the perinatal period]. / Anti-inflammatoires non stéroïdiens et grossesse. Etude sur la toxicité rénale et digestive en période périnatale de l'acide niflumique.

We report a case of intrauterine exposure to niflumic acid in a preterm neonate (35 weeks of gestation). The mother received niflumic acid (750 mg daily) during the last four days of gestation. Severe oligohydramnios was present. A transient neonatal renal failure, but also abdominal complications were observed for several days. Maternal and neonate niflumic acid levels were studied and showed an important placental transfer. Niflumic acid is a prostaglandin synthetase inhibitor. It is important that the nephrotoxicity of this vasoactive drug be known. Maternal administration of niflumic acid during the last days of gestation can induce fetal and neonatal adverse effects, especially renal failure. Prescription of all nonsteroidal anti-inflammatory drugs must be very cautious during pregnancy.

Determinaçäo dos efeitos das drogas antiinflamatórias näo esteróides (indometacina, butazona, clinoril, naprosin, benflogin e inflaril) nos leucogramas de ratos portadores de processo inflamatório crônico / Assessment of the effects of non-steroidal anti-inflammatory drugs (indomethacin, phenylbutazone, sulindac, naproxen, benzydamine and neflumic acid) on rat leucograms with chronic inflammatory process

Neste trabalho procurou-se avaliar os efeitos dos antiinflamatórios näo esteróides: Indometacina (indocid), Butazona (Fenilbutazona), Clinoril (Sulindac), Naprosin (Naproxen), Benflogin (Cloridrato de Benzidamina) e Inflaril (Acido neflúmico) nos leucogramas de ratos portadores de um processo inflamatório crônico provocado pela introduçäo intradérmica de lamínulas de vidro nos períodos de 3, 12 e 18 dias. O sangue para a contagem total dos leucócitos, eletrônica e diferencial, esfregaço, foi obtido por punçäo intracardíaca (Burhoe). A Indometacina, o Clinoril e a Butazona indicaram diminuiçäo de linfócitos e eosinófilos e aumento de monócitos e neutrófilos em todos os períodos e observaçäo em todos os períodos; o Inflaril reduziu o número de linfócitos, neutrofilia e eosinopenia em todos os períodos; o Inflaril reduziu o número de linfócitos e eosinófilos de aumentou os monócitos, com exceçäo de 3§ período, e os neutrófilos nos três períodos; e o Benflogin elevou os linfócitos na 1ª e 3ª fases, e os monócitos nos três períodos, e reduziu os neutrófilos nos dois primeiros, e os eosinfófilos nos dois últimos períodos. Todas as drogas usadas provocaram reduçäo de leucócitos em todos os períodos de tratamento, exceçäo feita ao Naprosin no 3§, ao Benflogin no 2§ e ao Inflaril no 1§ e 2§ períodos

[Determination of the concentrations of niflumic acid in the plasma and synovial tissue following repeated administration of capsules]. / Détermination des concentrations d'acide niflumique dans le plasma et le tissu synovial après administration répétée de gélules.

Niflumic acid was assayed in the plasma and synovial tissue of 12 adults about 10 hours following the end of a 1 g niflumic acid daily administration for 2 days; an arthroscopy of the knee had been carried out in these patients with various degenerative infections. The ratio of niflumic acid synovial concentrations to plasmatic concentrations was 1.27, showing the affinity of this drug for the synovial tissue.

The pharmacokinetics and availability of niflumic acid in humans.

The pharmacokinetic parameters and relative availability of niflumic acid in two different pharmaceutical preparations were studied in 12 subjects after a single oral administration. Total plasma clearance averaged 45 ml/min, and the half-life of elimination approximately 2 h, giving a distribution volume of 0.12 l/kg on the average. The values of these pharmacokinetic parameters were in agreement with the general characteristics of this type of substance, a weak acid strongly bound to plasma proteins. Comparison of the systemic availability of the two oral forms showed no difference; they were probably close to 100%.

Quantitative determination of niflumic acid and its beta-morpholinoethyl ester in human plasma by gas-liquid chromatography.

Niflumic acid and its beta-morpholinoethyl ester are extracted from plasma with diethyl ether. After methylation with diazomethane the solution is evaporated to dryness and the residue dissolved in methanol before injection in the chromatographic column. Using a nitrogen-sensitive detector the method permits the determination of 100 ng of each compound in 1 ml of plasma. The coefficient of variation is 5.3% and 4.8% for the acid and the ester, respectively, at the 2-microgram level.