No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
RP-HPLC method development and validation for the estimation of diclofenac sodium, tramadol hydrochloride and chlorzoxazone from their combined tablet dosage form
Abstract
Objective: The objective of the current study was to develop and validate the RP-HPLC method for the simultaneous estimation of Tramadol Hydrochloride, Chlorzoxazone and Diclofenac sodium from their combined tablet dosage form Methods: The current method describes RP-HPLC method for the estimation of Tramadol Hydrochloride, Chlorzoxazone and Diclofenac sodium from their combined tablet dosage form. The mobile phase used was Acetonitrile: 0.05M Disodium Hydrogen Phosphate buffer pH 3.5 adjusted with 10% v/v Ortho Phosphoric acid(50:50 v/v) and Hypersil ODS C18(250 mm × 4.6 mm, 5.0 μ particle sizes) was used as a stationary phase with detection wavelength of 220 nm. Result: Linearity was obtained in the concentration range of 15-75 μg/ml, 100-500 μg/ml and 20-100 μg/ml for TRM, CHL and DIC respectively. The % recovery was found to be 99.41 -99.84%, 99.30-99.74 % and 99.6-99.97 % for DIC, TRM and CHL respectively. The LOQ was found to be 3.33, 3.95 and 36.71 μg/ml for DIC, TRM and CHL respectively. The proposed method has been validated as per ICH Q2R1 guidelines and successfully applied to the estimation of TRM, DIC and CHL from their combined Tablet dosage form. Conclusion: The method was found to be simple, accurate, precise, and suitable for the estimation of Tramadol Hydrochloride, Chlorzoxazone and Dicofenac sodium from their combined tablet dosage form.
... Additionally, CZ, DIC and paracetamol were analyzed as ternary mixture by HPTLC [8], HPLC [9][10][11], packed column supercritical fluid chromatographic method [12], gas chromatography (GC) [13], spectrophotometry [14] and chemometric and artificial neural network techniques [15]. On the other hand, the ternary mixture of CZ, DIC and TRA was determined in a single analytical report using RP-HPLC [16]. ...
... Based on what has been stated in literature, simultaneous determination of CZ, DIC and TRA was addressed in only one HPLC report [16], taking into consideration that this method was done without presence of related substances and potential impurities of any of three drugs previously mentioned. Moreover, there is no available HPTLC method for simultaneous analysis of this ternary drug combination. ...
... In the present work, an efficient, simple, specific and reliable gradient elution HPLC-DAD and HPTLC procedures for the assay of TRA, CZ and DIC in their combined pharmaceutical dosage form were described. The suggested methods are by far superior to the only reported method for assay of this drug combination [16]. A significant advantage in the study is the separation of three analytes from three of their related substances and potential impurities. ...
In recent years, a lot of single-pill combinations (SPC) are manufactured and are used as a promising choice in treatment of signs and symptoms of osteoarthritis and rheumatoid arthritis. However, this trend made a serious challenge to drug analysts because of the difficulty in the analysis of two or more drugs in presence of each other. In this study, two chromatographic methods were developed for the simultaneous analysis of chlorzoxazone (CZ), diclofenac sodium (DIC) and tramadol hydrochloride (TRA) in presence of three of their related substances and potential impurities. Method I involved application of HPLC with diode array detection where Waters Symmetry C8 column was used as stationary phase. Mixture of ortho-phosphoric acid (0.03 M, pH 3) and acetonitrile was used as a mobile system with gradient elution and flow rate 1 mL/min. Peak areas were measured at the wavelengths 218 nm for TRA and 280 nm for CZ and DIC. Peaks eluted at retention times 2.30, 5.75 and 9.74 min for TRA, CZ and DIC respectively. Method II based on HPTLC separation. In this method, HPTLC silica gel 60 F254 plates were used with mobile phase of hexane: ethyl acetate: methanol: acetic acid (12: 6: 2: 0.1, by volume). Densitometry scanning was performed at 280 nm for all drugs. The retardation factor (Rf) values were 0.14, 0.40 and 0.58 for TRA, DIC and CZ respectively. Both methods were validated according to International Conference on Harmonization (ICH) Guidelines with respect to linearity, ranges, accuracy, precision, specificity, robustness and limits of detection and quantitation. Linearity ranges were 15–100 μg/mL for the three drugs in method I. In method II, the obtained data were linear in the ranges 300–1800 ng/spot for TRA and 25–150 ng/spot for DIC and CZ. Both methods showed good specificity by resolution of the three drugs from the related substances and potential impurities 2-amino-4-chlorophenol, 2.6-dichloroaniline and impurity A of DIC. Finally, both methods were successfully applied to the analysis of a real sample of combined tablets containing the three drugs. The suggested methods could be applied for the studied drugs in QC-lab.
... Various techniques have been explored for the simultaneous detection of multiple drugs, including the concurrent identification of multiple drugs using HPTLC (Kulkarni et al., 2012;Mohite et al., 2013), the LC-ESI-MS/MS-based identification of chlorzoxazone in rat plasma (Wang et al., 2010), simultaneous detection of multiple drugs using HPTLC (Chakraborty et al., 2012;Chawla et al., 1996;Joshi & Rajesh, 2008;Karthikeyan et al., 2009;Konduru et al., 2020;Patel & Patel, 2014;Shaikh & Devkhile, 2008;Vidya et al., 2017), and a swift and sensitive assay for 4-aminophenol in paracetamol drug and tablet formulations through flow injection analysis with spectrophotometric detection (Bloomfield, 2002;Sodhi et al., 1996). ...
This study validates a stability‐indicating LC method for detecting organic impurities in the chlorzoxazone dosage form. Using a Waters X‐Select R HSS T3 analytical column, mobile phase of it was made by mixing of water, methanol, and glacial acetic acid in the ratio of 700:300:10 (v/v/v). The drug product and drug substance were subjected to the stress conditions such as acid, base, oxidation, heat, and photolysis as per the recommendations of the International Conference on Harmonization (Q2) methodology. The study revealed the susceptibility of 4‐chloro‐2‐aminophenol to alkaline environments, emphasizing peak homogeneity and stability. The method verification, per ICH guidelines and USP<1225>, established precision, specificity, linearity, accuracy, and robustness for quality control. The mean impurity recovery ranged from 95.5% to 105.2%, the correlation coefficient ( r ) was greater than 1.000, and the RSD values ( n = 6) ranged from 0.6% to 5.1% across the LOQ–150% ranges. Full‐factorial design tested final method conditions, evaluating multiple parameters concurrently. Graphical optimization within the design space defined strong method requirements, ensuring consistent and reliable outcomes. The study develops and validates chlorzoxazone stability‐indicating methods, employing advanced statistical approaches like design of experiments and factorial design, with resilient conditions established through graphical optimization of the design space.
... Thus, the DLC is one of the few NSAIDs that considered as the "first choice" in treating acute, chronic pain and inflammatory conditions [2]. Several analytical techniques have been used to quantify diclofenac in pharmaceutical preparations including spectrophotometry [5][6][7][8][9][10][11][12], HPLC [13][14][15][16] voltammetry [17][18] and potentiometry [19]. This work focused on the development of a simple, sensitive, selective, and interference-free spectrophotometric procedure depended on the coupling of DLC with diazotized 4-Aminoacetophenone. ...
A recommended, simple and sensitive method for the spectrophotometric determination of diclofenac sodium drug
(DLC) in pharmaceutical formulations was represented in this work. The method is based on the coupling of DLC
with diazotized 4-Aminoacetophenone in basic medium to form an intense yellowish-brown colored diazocoupling
product that showed maximum absorbance at 362 nm. The effect of various factors such as amount of the reagent,
time of the reaction and order of addition were investigated. The experimental results indicated a good linearity (r2
= 0.9954) over a concentration range of (2- 40) µg.mL-1 with a detection limit of 0.447 µg.mL-1 diclofenac sodium
under the optimized conditions. The pharmaceutical excipients did not exhibit any interferences. The low value of
the standard deviation alongside good correlation coefficient confirms the applicability of the method. Accordingly,
the proposed method has been successfully applied to the determination of diclofenac sodium in pharmaceutical
formulations. The results were statistically compared with standard methods by means of t-test and f-test at 95%
confidence level with no significant differences observed.
... The flow rate was adjusted to 0.5 mL/min and UV detection at 270 nm was selected for analysis after the preliminary tests. The retention time of PARA, ACF and CHZ were found to be 0.94, 3.84 and 1.92 min respectively [6]. The blank chromatogram and standard Chromatogram of PARA, ACF and CHZ was shown in Figures 2 and 3. ...
... Chlorzoxazone is assayed by liquid chromatography as per USP. However, several chromatographic methods have been reported for the determination of Chlorzoxazone, in pharmaceutical formulations and/ or in biological fluids, including HPLC [5][6][7][8], stability indicating HPLC [9], LC-MS [10] and HPTLC [11,12] for simultaneous determination of Chlorzoxazone with other drugs. Study for presence of related substances in marketed formulations of Chlorzoxazone is not reported. ...
Chlorzoxazone is one of the most frequently prescribed drugs in the treatment of muscle spasm in combination with NSAIDs. It is reported to undergo hydrolysis in alkaline medium to form 2-amino-4-chlorophenol, which is considered a significant related substance as per USP. Various regulatory authorities like ICH, USFDA, Canadian Drug and Health Agency are emphasizing on the purity requirements and the identification of impurities in active pharmaceutical ingredient (API) as well as in pharmaceutical formulations. To this aim, various marketed formulations were assessed with special attention to identification and quantification of 2-amino-4-chlorophenol by using compendial reversed phase high performance liquid chromatography method. The use of a 250 × 4.6 mm, 5 µm particle size, C 18 column with 70:30:1 %, v/v/v water: acetonitrile: acetic acid as isocratic mobile phase at flow rate 1.5 ml/min enabled separation of the drug from its related substance. UV detection was performed at 280 nm. The method was verified for specificity, linearity, precision and accuracy. The related substance peak was well resolved from drug peak. The linearity of the method was satisfactory over the range 400-2000 ng (correlation coefficient 0.9993). The limits of detection for 2-amino-4-chlorophenol and Chlorzoxazone were 5 and 10.94 ng respectively. The limits of quantitation for 2-amino-4-chlorophenol and Chlorzoxazone were 20 and 33.14 ng respectively. Recovery of Chlorzoxazone ranged from 99.90-100.67%. The method was successfully applied to marketed formulations of Chlorzoxazone for quantitative analysis of Chlorzoxazone and 2-amino-4-chlorophenol.
... These methods include spectrophotometry and multivariate [4][5][6], fluorimetry [7,8], voltammetry [9,10], high-performance liquid chromatography-ultraviolet (HPLC-UV) [11][12][13][14][15], liquid chromatographytandem mass spectrometry (LC-MS/MS) [16][17][18][19], capillary zone electrophoresis [20], and densitometry [21,22]. Other methods were developed for the simultaneous determination of DIC in binary combination with PAR or CHZ that include spectrophotometry [23][24][25], densitometry [26], voltammetry [27], and HPLC-UV [28,29]. Some methods were reported for the simultaneous determination of ternary combination of the three drugs based on densitometry [30,31] and HPLC-UV [32,33]. ...
Objective: The aim of this study is to develop and validate simple, accurate, and precise spectrophotometric methods for the simultaneous determination of diclofenac sodium (DIC), paracetamol (PAR), and chlorzoxazone (CHZ) in ternary mixture using chemometric and artificial neural networks (ANN) techniques.Methods: Three chemometric techniques include classical least squares (CLS), principal component regression (PCR), and partial least squares (PLS) in addition to cascade-forward backpropagation ANN (CFBP-ANN) were prepared using the synthetic mixtures containing the three drugs in methanol. In CLS, PCR, and PLS, the absorbances of the synthetic mixtures in the range 267-295 nm with the intervals Δλ=0.2 nm in their zero-order spectra were selected. Then, calibration or regression was obtained using the absorbance data matrix and concentration data matrix for the prediction of the unknown concentrations of DIC, PAR, and CHZ in their mixtures. In CFBP-ANN, two layers, sigmoid layer with 10 neurons and linear layer were found appropriate for the simultaneous determination of the three drugs in their ternary mixture.Results: The four proposed methods were successfully applied to the analysis of the three drugs in laboratory prepared mixtures and tablets with good percentage recoveries in the range of 98-102%. Relative standard deviation for the precision study was found <1%.Conclusion: The four proposed methods showed simplicity, accuracy, precision, and rapidity making them suitable for quality control and routine analysis of the cited drugs in ternary mixtures and pharmaceutical formulation containing them.
... This study is to develop a reversed-phase high-performance liquid chromatography (RP-HPLC) method for KET and TDL. A literature survey reveals several analytical methods for estimation of KET and TDL individually and in combination with them and other drugs based on ultraviolet (UV) [9][10][11], HPLC [12][13][14][15][16][17][18][19][20][21][22][23], liquid chromatography-mass spectrometry [24], and ion-pair chromatography [25] were reported. However, there are few methods reported for the method development of TDL and KET; the present aim is to develop a more precise, accurate and simple RP-HPLC method for the estimation of TDL and KET. ...
Objective: This study was embarked upon to develop a new, simple, rapid, validated reversed-phase high-performance liquid chromatography (HPLC)method for the estimation of ketorolac tromethamine (KET) and tramadol hydrochloride (TDL) in pharmaceutical dosage forms.Methods: The HPLC method was developed on Shishiedo C18 column (250 mm × 4.6 mm i.d, 5 μ) using methanol: 50 mM phosphate buffer (pH 6.0) in the ratio of 52:48 at 282 nm.Results: Retention time for the drugs was found to be 5.1 and 6.9 minutes for tramadol and ketorolac, respectively. The limit of detection for tramadoland ketorolac were found to be 1.0 and 0.1 μg/ml, limit of quantitation for tramadol and ketorolac were found to be 5.0 and 0.5 μg/ml, respectively. Linearity was established in the range of 20.0-30.0 μg/ml and 8.0-12.0 μg/ml for TDL and KET, respectively. The method was precise with % relative standard deviation <2 for both intra- and interday precision. The accuracy of the method was performed over three levels of concentration, and the recovery was in the range of 98-102%.Conclusion: From the found experimental data, it can be concluded that the developed method is accurate, precise, and selective and can be employedsuccessfully for the estimation of KET and TDL in Pharmaceutical dosage forms.Keywords: Reversed-phase high-performance liquid chromatography, Ketorolac tromethamine, Tramadol hydrochloride.
Objective: A simple, accurate, precise method was developed for the simultaneous estimation of the Chlorzoxazone, Pantoprazole, and Diclofenac in solid dosage form. Method: Chromatogram was run through BDS C18 150 × 4.6 mm, 5 m. Mobile phase containing 0.01% KH 2 PO 4 and Acetonitrile in the ratio of 52:48 v/v was pumped through column at a flow rate of 1.0 ml/min. The buffer used in this method was 0.01%. KH 2 PO 4 , Temperature was maintained at 30 °C. The optimized wavelength for Chlorzoxazone, Pantoprazole, and Diclofenac was 229.0 nm. Results: Retention time of Chlorzoxazone, Pantoprazole, and Diclofenac were found to be 2.229 min, 2.958 min, and 3.568 min. % RSD of system precision for Chlorzoxazone, Pantoprazole and Diclofenac were and found to be 0.9, 0.6 and 0.6, respectively. % RSD of method precision for Chlorzoxazone, Pantoprazole and Diclofenac were and found to be 0.6, 1.0 and 0.7 respectively. % recovery was obtained as 99.86%, 100.07% and 99.70% for Chlorzoxazone, Pantoprazole, and Diclofenac respectively. LOD values are obtained from regression equations of Chlorzoxazone, Pantoprazole and Diclofenac were 0.14 ppm, 0.24 ppm, 1.83 pm, and LOQ values are obtained from regression equations of Chlorzoxazone, Pantoprazole and Diclofenac were 0.42 ppm, 0.72 pm, 5.53 ppm respectively. The regression equation of Chlorzoxazone was y = 8321.9.x + 1397.8Pantoprazolee was y = 9806.1x + 6071.7 and of Diclofenac was y = 2575x + 4338.4. Conclusion: Retention times are decreased so the method developed was simple and economical that can be adopted in regular Quality control test in Industries.
Three green smart spectrophotometric methods namely; absorbance subtraction (AS), amplitude modulation (AM) and amplitude summation (A-Sum) were developed and validated for the determination of a binary mixture of chlorzoxazone (CHZ) and ibuprofen (IBU) without prior separation, using unified regression equation. The proposed spectrophotometric procedures were found to be linear in the range of (2.5-27.5 μg/ml) and (2-22 μg/ml) for CHZ and IBU, respectively. The methods were validated as per ICH guidelines and the results were found to be within the acceptable limits. The specificity was assessed by analyzing synthetic mixtures of both drugs which showed the superiority of AM and A-Sum methods over AS method in the analysis of low concentrations. The proposed methods were applied to pharmaceutical formulations and the results were statistically compared to that of a reported spectrophotometric method. The statistical comparison showed that there is no significant difference among the methods regarding both accuracy and precision.
A new, economic, eco friendly and rapid high-performance thin-layer chromatographic (HPTLC) method was developed and validated for quantitative determination of Tramadol HCl. The HPTLC separation was achieved on an aluminium-backed layer of silica gel 60F 254 using ethyl acetate: methanol: ammonia(25%) (9.0 : 1.0 :0.5 ml(v/v/v)) as mobile phase. Quantitation was achieved by densitometric analysis at 271 nm over the concentration range of 1000-6000 ng/spot. The method was found to give compact spot for the drug (R f 0.76 ± 0.011). The linear regression analysis data for the calibration plots showed good linear relationship with r 2 = 0.9933. The method was validated for precision, recovery, repeatability, and robustness as per the International Conference on Harmonization guidelines. The minimum detectable amount was found to be 116.38 ng/spot, whereas the limit of quantitation was found to be 352.67 ng/spot. Statistical analysis of the data revealed that the method is precise, accurate, reproducible, sensitive and selective for the analysis of Tramadol HCl. The method was successfully employed for the estimation of Tramadol HCl as a bulk drug and in capsule dosage form. 2. INTRODUCTION: Tramadol HCl is a synthetic codeine analogue that is a weak μ-opioid receptor agonist. It is used as an oral non-steroidal anti-inflammatory drug with good analgesic and tolerability profile in various painful conditions. Chemically it is (+/-)-cis-2-[(dimethylamino)methyl]-1-(3-methoxy-phenyl) cyclohexanol [1] . It is official in British Pharmacopoeia [2] , European pharmacopoeia 5.0 and Indian Pharmacopoeia 2010 where potentiometric titration is the official method of assay. Several analytical methods such as spectrophotometric [3] , HPLC [4] and bioanalytical methods [5-7] have beed developed. The present study describes a simple, sensitive and precise HPTLC method for the estimation of Tramadol hydrochloride in bulk and in capsule dosage form.
A simple, precise and accurate reversed-phase liquid chromatographic method has been developed for the simultaneous estimation of aceclofenac (ACF), paracetamol (PCM) and tramadol hydrochloride (TRM) in pharmaceutical dosage form. The chromatographic separation was achieved on a HiQ-Sil™ HS C18 column (250×4.6 mm i.d., 5 μm particle size), kromatek analytical column at ambient temperature. The mobile phase consisted of 40: 60 (v/v); phosphate buffer (pH 6.0): methanol. The flow rate was set to 1.0 mL min(-1) and UV detection was carried out at 270 nm. The retention time (t(R)) for ACF, PCM and TRM were found to be 14.567 ± 0.02, 3.133 ± 0.01 and 7.858 ± 0.02 min, respectively. The validation of the proposed method was carried out for linearity, precision, robustness, limit of detection, limit of quantitation, speci city, accuracy and system suitability. The linear dynamic ranges were from 40-160 μg mL(-1) for ACF, 130-520 μg mL(-1) for PCM and 15-60 μg mL(-1) for TRM. The developed method can be used for routine quality control analysis of titled drugs in pharmaceutical dosage form.
A simple, reverse phase high performance liquid chromatographic method was developed for the simultaneous determination of Paracetamol, Diclofenac Sodium and Chlorzoxazone from tablets. The method described is precise, accurate reproducible and rapid.
A simple, precise, accurate and reproducible spectrophotometric method has been developed for simultaneous estimation of Tramadol Hydrochloride (TRA) and Diclofenac Sodium(DIC) by employing first order derivative zero crossing method in methanol. The first order derivative absorption at 272.45 nm (zero cross point of TRA) was used for quantification of DIC and 282.81 nm (zero cross point of DIC) for quantification of TRA. The linearity was established over the concentration range of 2-18 μg/ml and 5-25 μg/ml for TRA and DIC with correlation coefficient r2 0.9941 and 0.9968, respectively. The mean % recoveries were found to be in the range of 99.2 - 99.73 % and 99.33 - 99.64 % for TRA and DIC, respectively. The proposed method has been validated as per ICH guidelines and successfully applied to the estimation of TRA and DIC in their combined Tablet dosage form.
A rapid and sensitive liquid chromatography-mass spectrometry (LC-MS) method has been developed and validated for quantification of diclofenac potassium in human plasma. The analyte was extracted from human plasma by simple precipitation technique. Bupropion was used as the internal standard. A kromasil CI 8 column provided chromatographic separation of the analyte which was followed by detection with mass spectrometry. The mass transition ion-pair was followed as m/z 296.10 and 214.10 for diclofenac 240.1 and 184.10 m/z for bupropion. The method involves precipitation of diclofenac from plasma, simple isocratic chromatography conditions with mobile phase acetonitrile: water and mass spectrometric detection that enables detection at nanogram levels. The retention times were 1.58 and 1.21 min for diclofenac and bupropion, respectively. The proposed method has been validated with linear range of 150-3181 ng mL for diclofenac. The precision and accuracy values are within ± 10%. The overall recovery of diclofenac was 41.9%.
Two simple, accurate, and precise methods for simultaneous estimation of tramadol hydrochloride and chlorzoxazone in combined dosage form have been described. The first method employs formation and solving of simultaneous equations using 272.20 and 248.30 nm as two analytical wavelengths. The second method is absorption ratio method, which uses 272.20 and 257.50 nm as two analytical wavelengths. Both the methods allow the simultaneous determination of tramadol hydrochloride and chlorzoxazone in concentration ranges employed for this purpose with the standard deviation of < 1.0% in the assay of tablet.