Efficient high performance liquid chromatograph/ultraviolet method for determination of diclofenac and 4′-hydroxydiclofenac in rat serum

Kaphalia, Lata; Kaphalia, Bhupendra S.; Kumar, Santosh; Kanz, Mary F.; Treinen-Moslen, Mary

doi:10.1016/j.jchromb.2005.10.045

Cited by 37 publications

(26 citation statements)

References 32 publications

(43 reference statements)

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“…All peaks were isocratically eluted within 14 min at a mobile phase flow rate of 1 ml/min and detected using UV at 280 nm. A single unidentified peak eluted at 6.25 min was assumed to be 5DFN (Kaphalia et al, 2006). No authentic standard for 5DFN was commercially available, but because the chromatophoric difference between 5DFN and 4DFN is presumed negligible, 4DFN was used as the reference.…”

Section: Methodsmentioning

confidence: 99%

“…Analyses of DFN and its metabolites were performed using a modified version of the method described by Kaphalia et al (2006). In brief, samples were separated on a Hypersil Gold C 18 column (4.6 ϫ 250 mm, 5-m particle size; Thermo Fisher Scientific) after a sample injection volume of 100 l. The mobile phase consisted of CH 3 CN and 0.75 mM sodium acetate buffer, pH 5.0 at a ratio of 2:3.…”

Section: Accurate Estimation Of Enzyme Kinetics From Depletion Datamentioning

confidence: 99%

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The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

Sjögren¹,

Lennernäs²,

Andersson³

et al. 2008

Drug Metab Dispos

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ABSTRACT:The use of multiple depletion curves for the estimation of maximum velocity of the metabolic reaction (V max ), the Michaelis constant (K m ), and intrinsic clearance (CL int ) was thoroughly evaluated by means of experimental data and through a series of Monte Carlo simulations. The enzyme kinetics of seven compounds were determined using the multiple depletion curves method (MDCM), the traditional initial formation rate of metabolite method (IFRMM), and the "in vitro t 1 ⁄2" method, and the parameter estimates that were derived from the three methods were compared. The impact of a change in enzyme activity during the incubation period on the parameter estimates and the possibility to correct for this were also investigated. The MDCM was in good overall agreement with the IFRMM. Correction for a change in enzyme activity was possible and resulted in a better concordance in CL int estimates. The robustness of the method in coping with different rates of substrate turnover and variable starting concentrations were also demonstrated through Monte Carlo simulations. Furthermore, the limitations imposed by assumptions inherent in the in vitro t 1 ⁄2 method were demonstrated both experimentally and by simulations. This study demonstrates that the MDCM is a robust and efficient method for estimating enzyme kinetic variables with high accuracy and precision. The method may potentially be used in a wide range of applications, from pure enzyme kinetics to in vitrobased predictions of the pharmacokinetics of compounds with multiple and/or unknown metabolic pathways.Estimation of metabolic intrinsic clearance (CL int ) is currently included in many drug discovery programs. The most frequently used assay is the "in vitro t 1 ⁄2" method (T 1 ⁄2M), in which CL int is derived from the monoexponential slope of a single depletion curve (Obach, 1999). This method is used both for ranking compounds with respect to metabolic stability and for the prediction of metabolic clearance in animals and humans. The data are usually obtained from incubations with microsomes or fresh or cryopreserved hepatocytes (Iwatsubo et al., 1997;Obach et al., 1997;Rodrigues, 1997;Ito and Houston, 2005). CL int , defined as the maximum velocity of the metabolic reaction (V max ) divided by the Michaelis constant (K m ), the substrate concentration that yields half of V max , is by this means used as the link between fundamental enzyme kinetics and in vivo pharmacokinetic variables (Rane et al., 1977). Although the T 1 ⁄2M approach is fast, it is built on the assumption that the initial concentration (C 0 ) is well below K m . This assumption is often valid, but, if not, the method will underestimate CL int and thus underpredict the rate of hepatic clearance in vivo. The basic assumption of C 0 Ͻ ϽK m is usually not confirmed and may be one of several contributing factors to the tendency of systematic underprediction of hepatic clearance seen in the literature (Carlile et al., 1999;Obach, 1999;Ito and Houston, 2005). The ability to predict cleara...

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Section: Methodsmentioning

confidence: 99%

Section: Accurate Estimation Of Enzyme Kinetics From Depletion Datamentioning

confidence: 99%

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

Sjögren¹,

Lennernäs²,

Andersson³

et al. 2008

Drug Metab Dispos

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“…Several HPLC methods have been described in the literature for the determination of diclofenac in raw material, pharmaceutical dosage forms, and biological fluids [7,8]. Also HPLC methods are available in literature for separate determination of metoprolol tartrate or phenol red [2,[8][9][10][11].…”

Section: Introductionmentioning

confidence: 99%

“…Also HPLC methods are available in literature for separate determination of metoprolol tartrate or phenol red [2,[8][9][10][11]. Therefore, the aim of this study was to determine segmental permeability and absorption characteristics of diclofenac using a newly developed and validated reversed-phase liquid chromatographic method for the simultaneous determination of diclofenac, metoprolol tartrate and phenol red in intestinal perfusion samples.…”

Section: Introductionmentioning

confidence: 99%

Determination of Regional Intestinal Permeability of Diclofenac and Metoprolol Using a Newly-Developed and Validated High Performance Liquid Chromatographic Method

Kaynak¹,

Buyutuncel²,

Cagler³

et al. 2015

Trop. J. Pharm Res

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“…2-4) It has been determined by a variety of analytical techniques, such as UV/Vis spectrophotometry, [5][6][7][8][9][10][11] fluorimetry, [12][13][14] reflectometry, 15) liquid chromatography, [16][17][18][19] and flow injection analysis with solid-phase spectroscopic detection. 20) However, as far we know, there is no kinetic-spectrophotometric method for the determination of diclofenac in the literature.…”

mentioning

confidence: 99%

Determination of Diclofenac Sodium in Commercial Pharmaceutical Formulations and Human Control Serum Using a Kinetic-Spectrophotometric Method

Mitić

Miletić

Pavlović

et al. 2007

CHEMICAL & PHARMACEUTICAL BULLETIN

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Diclofenac, derived from benzeneacetic acid, is a nonsteroidal anti-inflammatory drug (NSAID) that is more usually found as a sodium or potassium salt with potent anti-inflammatory, analgesic, and antipyretic properties. Its mechanism of action is associated principally with the inhibition of prostaglandin synthesis (specifically, inhibition of cyclooxygenase).1) Diclofenac is indicated for a variety of conditions such as acute and chronic arthritis, rheumatoid arthritis, osteoarthritis, and ankylosing spondylitis. Diclofenac also has been demonstrated to be an effective migraine treatment in several placebo-controlled studies.2-4) It has been determined by a variety of analytical techniques, such as UV/Vis spectrophotometry, [5][6][7][8][9][10][11] fluorimetry, [12][13][14] reflectometry, 15) liquid chromatography, [16][17][18][19] and flow injection analysis with solid-phase spectroscopic detection. 20) However, as far we know, there is no kinetic-spectrophotometric method for the determination of diclofenac in the literature.The aim of this work is to develop a simple, rapid, reliable, precise and accurate kinetic method for the determination of diclofenac sodium in pharmaceutical preparations. The method is based on a ligand-exchange reaction. Diclofenac shows complexing ability with cobalt(II). The complex agreed with the empirical formula CoD 2 · H 2 O and the cobalt ions are co-ordinated through the carboxylate group of the ligand (diclofenac, D). 21,22) ExperimentalApparatus The reaction rate was monitored spectrophotometrically by measuring the rate of change of absorbance at 376 nm. The readings were performed on a Perkin-Elmer Lambda 15 UV/Vis spectrophotometer, connected to a thermo-circulating bath.pH measurements were carried out using a Hanna Instruments pH meter. In addition, high precision volume micropipettes (Lab Mate ϩ ) of 50, 500 and 1000 ml were used for handling or pipetting the solutions.The solutions were thermostated at 22Ϯ0.1°C before the beginning of the reaction.Potentiometric titrations were conducted using a Titrino 716 DMS in dynamic equivalence-point titration (DET) mode. The evaluation of EPs was based on the zero crossing of the second derivate of the titration curve.Reagents A diclofenac stock solution (1ϫ10 Ϫ3 mol l Ϫ1 as the sodium salt) was prepared from pharmaceutical 99.9% certified products supplied by a pharmaceutical laboratory (Galenika, a.d., Belgrade, Serbia). The solution was stored at 4°C. Working solution was prepared daily from this solution, as required, by dilution with water.Acetic acid solution (HAc, 10 mol l Ϫ1) was prepared from glacial HAc (Merck).1-Nitroso-2-naphthol solution (1ϫ10 Ϫ3 mol l Ϫ1 ) (Merck) was prepared by dissolving a known amount in 5 ml absolute ethanol and diluting it with water (total volume 50 ml).The stock cobalt(II) solution (1.7ϫ10 Ϫ3 mol l Ϫ1 ) was prepared by dissolving CoCl 2 ϫ6H 2 O (Merck) in water.Ionic strength was kept constant at 0.1 by adding an appropriate amount of NaCl solution (mol l Ϫ1). Analytical grade chemicals an...

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Efficient high performance liquid chromatograph/ultraviolet method for determination of diclofenac and 4′-hydroxydiclofenac in rat serum

Cited by 37 publications

References 32 publications

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

Determination of Regional Intestinal Permeability of Diclofenac and Metoprolol Using a Newly-Developed and Validated High Performance Liquid Chromatographic Method

Determination of Diclofenac Sodium in Commercial Pharmaceutical Formulations and Human Control Serum Using a Kinetic-Spectrophotometric Method

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Efficient high performance liquid chromatograph/ultraviolet method for determination of diclofenac and 4′-hydroxydiclofenac in rat serum

Cited by 37 publications

References 32 publications

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CLint), Maximum Velocity of the Metabolic Reaction (Vmax), and Michaelis Constant (Km): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CLint), Maximum Velocity of the Metabolic Reaction (Vmax), and Michaelis Constant (Km): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

Determination of Regional Intestinal Permeability of Diclofenac and Metoprolol Using a Newly-Developed and Validated High Performance Liquid Chromatographic Method

Determination of Diclofenac Sodium in Commercial Pharmaceutical Formulations and Human Control Serum Using a Kinetic-Spectrophotometric Method

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The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations

The Multiple Depletion Curves Method Provides Accurate Estimates of Intrinsic Clearance (CL_int), Maximum Velocity of the Metabolic Reaction (V_max), and Michaelis Constant (K_m): Accuracy and Robustness Evaluated through Experimental Data and Monte Carlo Simulations