A new spectrophotometric method for the determination of methyldopa

A new, simple and low cost spectrophotometric method for the determination of methyldopa in pharmaceutical preparations was developed. The method was based on the coupling of methyldopa with 2,6-dichloroquinone-4-chlorimide (DCQ). The absorbance maximum (λ_max) of the resulted colored product was at 400 nm. Different buffers were used to determine the optimal pH for the reaction. About 1% w/v acetate buffer with pH 8.0 gave the optimal pH required for the reaction. Of the different solvents tried, water and ethanol were found to be the most suitable solvents. Beer’s law was obeyed in concentration range of 4–20 μg/ml methyldopa. The correlation coefficient was found to be (r = 0.9975). The limit of detection and limit of quantification were 1.1 μg/ml and 3.21 μg/ml, respectively. The reaction ratio between methyldopa and DCQ was studied and found to be 1:3. The work included the study of the possible interference of hydrochlorothiazide found in combination with methyldopa tablets. The method was validated and results obtained for the assay of two different brands of methyldopa tablets were compared with the BP method (colorimetric). The repeatability and reproducibility of the developed method were evaluated and the obtained results quoted. The derivative formed as a result of the reaction of methyldopa with DCQ was isolated and its possible mechanistic pathway was suggested.     

Application of HPLC for the simultaneous determination of aceclofenac, paracetamol and tramadol hydrochloride in pharmaceutical 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.     

Development and Validation of Stability-indicating RP-HPLC Method for Estimation of Pamabrom in Tablets

The present study depicts the development of a validated RP-HPLC method for the determination of the pamabrom in presence of degradation products or other pharmaceutical excipients. Stress study was performed on pamabrom and it was found that it degrade sufficiently in acidic, alkali and oxidative condition but less degradation was found in thermal and photolytic condition. The separation was carried out on Enable G 120 A⁰ (250×4.6 mm, 5 μ) column having particle size 5 μ using methanol: water (75:25 v/v) with pH 4.0 adjusted with ortho phosphoric acid as mobile phase at flow rate of 1 ml/min. The wavelength of the detection was 280nm. A retention time (R_t) nearly 3.9 min was observed. The calibration curve for pamabrom was linear (r² = 0.9997) from range of 10-60 μg/ml with limit of detection and limit of quantification of 1.41 μg/ml and 4.28 μg/ml, respectively. Analytical validation parameter such as selectivity, specificity, linearity, accuracy and precision were evaluated and relative standard deviation value for all the key parameters were less than 2.0%. The recovery of the drug after standard addition was found to be 101.35%. Thus, the developed RP-HPLC method was found to be suitable for the determination of pamabrom in bulk as well as stability samples of tablets containing various excipients.     

New Stability-Indicating RP-HPLC Method for Determination of Diclofenac Potassium and Metaxalone from their Combined Dosage Form

A simple, precise and accurate isocratic RP-HPLC stability-indicating assay method has been developed to determine diclofenac potassium and metaxalone in their combined dosage forms. Isocratic separation was achieved on a Hibar-C₁₈, Lichrosphere-100^® (250 mm × 4.6 mm i.d., particle size 5 μm) column at room temperature in isocratic mode, the mobile phase consists of methanol: water (80:20, v/v) at a flow rate of 1.0 ml/min, the injection volume was 20 μl and UV detection was carried out at 280nm. The drug was subjected to acid and alkali hydrolysis, oxidation, photolysis and heat as stress conditions. The method was validated for specificity, linearity, precision, accuracy, robustness and system suitability. The method was linear in the drug concentration range of 2.5–30 μg/ml and 20–240 μg/ml for diclofenac potassium and metaxalone, respectively. The precision (RSD) of six samples was 0.83 and 0.93% for repeatability, and the intermediate precision (RSD) among six-sample preparation was 1.63 and 0.49% for diclofenac potassium and metaxalone, respectively. The mean recoveries were between 100.99–102.58% and 99.97–100.01% for diclofenac potassium and metaxalone, respectively. The proposed method can be used successfully for routine analysis of the drug in bulk and combined pharmaceutical dosage forms.     

Development of Validated Bioanalytical HPLC-UV Method for Simultaneous Estimation of Amlodipine and Atorvastatin in Rat Plasma

A simple, economical and robust analytical high-performance liquid chromatography-ultraviolet method was developed and validated for simultaneous chromatographic elution of two cardiovascular drugs viz. amlodipine and atorvastatin in biological fluid for the first time. Only two liquid chromatography–mass spectrometry/mass spectrometry methods are available in literature for quantitation of selected pair of analytes. The bioanalytical method was developed in rat plasma by using Thermo beta-basic C₁₈ (100×4.6 mm, 5 μm) and mobile phase was composed of dibasic phosphate buffer (pH 3.0):acetonitrile in the ratio of 55:45 at a flow rate of 1 ml/min with ultraviolet detection monitored at 240 nm. The selected chromatographic conditions were found to effectively separate amlodipine (5.1 min) and atorvastatin (12.1 min). The parametric statistics,i.e. correlation coefficient of 0.999, was assessed for both the drugs having linearity over the tested concentration range (0.05 to 10.0 μg/ml) in rat plasma using an unweighted calibration curve. The mean recovery (%) was more than 92.8% for both the drugs using protein precipitation method. The accuracy of samples for six replicate measurements at lower limit of quantitation level was within limit. The method was validated and was successfully applied to the nonclinical pharmacokinetic study of combination tablets containing amlodipine and atorvastatin in six Sprague Dawley rats.     

Development and Validation of a Simultaneous HPLC Method for Estimation of Bisoprolol Fumarate and Amlodipine Besylate from Tablets

A fast, robust and stability indicating RP-HPLC method was developed for simultaneous determination of bisoprolol fumarate and amlodipine besylate in tablets. The mobile phase was mixture of 25 mM ammonium acetate adjusted to pH 5.0 and methanol (65: 35) at 0.8 ml/min. The stationary phase was Luna C18-2 column (3 μ, 50×4.6 mm ID). UV detection was performed at 230 nm. Retention time was 1.45 min and 3.91 min for bisoprolol and amlodipine, respectively. Linearity was established in the range of 8–33 μg/ml. Mean recovery was 99.1% and 98.6% for bisoprolol fumarate and amlodipine besylate, respectively.     

Development and Validation of Stability-indicating High Performance Liquid Chromatographic Method for the Estimation of Everolimus in Tablets

The present study depicts the development of a validated reversed-phase high performance liquid chromatographic method for the determination of the everolimus in presence of degradation products or pharmaceutical excipients. Stress study was performed on everolimus and it was found that it degrade sufficiently in oxidizing and acidic conditions but less degradation was found in alkaline, neutral, thermal and photolytic conditions. The separation was carried out on Hypersil BDS C18 column (100×4.6 mm, 5 μ) column having particle size 5 μ using acetate buffer:acetonitrile (50:50 v/v) with pH 6.5 adjusted with orthophosphoric acid as mobile phase at flow rate of 1 ml/min. The wavelength of the detection was 280 nm. A retention time (R_t) nearly 3.110 min was observed. The calibration curve for everolimus was linear (r²=0.999) from range of 25-150 μg/ml with limit of detection and limit of quantification of 0.036 μg/ml and 0.109 μg/ml, respectively. Analytical validation parameters such as selectivity, specificity, linearity, accuracy and precision were evaluated and relative standard deviation value for all the key parameters were less than 2.0%. The recovery of the drug after standard addition was found to be 100.55%. Thus, the developed RP-HPLC method was found to be suitable for the determination of everolimus in tablets containing various excipients.     

A Stability-Indicating RP-HPLC Assay Method for 5-Fluorouracil

The present study describes the development of a validated RP-HPLC method for the determination of 5-fluorouracil in presence of its degradation products or other pharmaceutical excipients. Stress studies were performed on 5-fluorouracil and it was found that it degrades sufficiently in alkaline conditions, while negligible degradation was observed in acidic, neutral, oxidative and photolytic conditions. The peaks of the degradation products were not observed in the chromatogram due to the nonchromophoric nature of the degradation moiety formed. The separations were carried out on a C-18 reversed phase column (Phenomenex; Prodigy ODS3V, 250×4.6 mm, 5 μ) using 50mM KH₂PO₄ (pH, 5.0) as mobile phase at a flow rate of 1.2 ml/min and temperature of 30°. The wavelength of detection was 254 nm. A retention time of nearly 6 minutes was obtained. Analytical validation parameters such as specificity and selectivity, linearity, accuracy and precision were evaluated. The calibration curve for 5-fluorouracil was linear (r²=0.999±0.0005) from range of 10 μg/ml to 100 μg/ml. Relative standard deviation values for all the key parameters, was less than 2.0 %. The recovery of the drug after standard addition to the degraded sample was found to be 104.69%. Thus, the developed RP-HPLC method was found to be suitable for the determination of 5-fluorouracil in bulk as well as stability samples of the pharmaceutical dosage forms containing various excipients.     

UV and Three Derivative Spectrophotometric Methods for Determination of Ezetimibe in Tablet Formulation

UV, first, second and third derivative spectrophotometric methods have been developed for the determination of ezetimibe in pharmaceutical formulation. The solutions of standard and sample were prepared in methanol. For the first method, UV spectrophotometry, the quantitative determination of the drug was carried at 233 nm and the linearity range was found to be 6-16 μg/ml. For the first, second and third derivative spectrophotometric methods the drug was determined at 259.5 nm, 269 nm and 248 nm with the linearity ranges 4-14 μg/ml, 4-14 μg/ml and 4-16 μg/ml. The calibration graphs constructed at their wavelength of determination were found to be linear for UV and derivative spectrophotometric methods. All the proposed methods have been extensively validated. The described methods can be readily utilized for the analysis of pharmaceutical formulation. There was no significant difference between the performance of the proposed methods regarding the mean values and standard deviations.     

RP-HPLC Estimation of Ramipril and Telmisartan in Tablets

A rapid high performance liquid chromatographic method has been developed and validated for the estimation of ramipril and telmisartan simultaneously in combined dosage form. A Genesis C18 column having dimensions of 4.6×250 mm and particle size of 5 μm in isocratic mode, with mobile phase containing a mixture of 0.01 M potassium dihydrogen phosphate buffer (adjusted to pH 3.4 using orthophosphoric acid): methanol:acetonitrile (15:15:70 v/v/v) was used. The mobile phase was pumped at a flow rate of 1.0 ml/min and the eluents were monitored at 210 nm. The selected chromatographic conditions were found to effectively separate ramipril (R_t: 3.68 min) and telmisartan (R_t: 4.98 min) having a resolution of 3.84. The method was validated in terms of linearity, accuracy, precision, specificity, limit of detection and limit of quantitation. Linearity for ramipril and telmisartan were found in the range of 3.5-6.5 μg/ml and 28.0-52.0 μg/ml, respectively. The percentage recoveries for ramipril and telmisartan ranged from 99.09-101.64% and 99.45-100.99%, respectively. The limit of detection and the limit of quantitation for ramipril was found to be 0.5 μg/ml and 1.5 μg/ml respectively and for telmisartan was found to be 1.5 μg/ml and 3.0 μg/ml, respectively. The method was found to be robust and can be successfully used to determine the drug content of marketed formulations.     

Stability-Indicating RP-HPLC Method for the Simultaneous Determination of Prazosin, Terazosin, and Doxazosin in Pharmaceutical Formulations

The current study was carried out with an attempt to separate similarly structured title drugs by liquid chromatography. Spectrophotometric techniques were generally insufficient under these conditions because of the spectral overlapping of drugs with similar functional groups. The pharmaceutical drugs prazosin, terazosin, and doxazosin contain the same parent quinazoline nucleus, thus making it especially difficult to separate the former two drugs because of their very similar structures. A simple and sensitive method for the routine determination of these drugs in pharmaceutical formulations was attempted. We found that the mobile phase consisting of A: ACN–diethylamine (0.05 ml), B: methanol, and C: 10 mM Ammonium acetate separated these drugs effectively. Separations were carried out on a new Kromasil C18 column (250 × 4.6 mm, 5.0 μm) at 254 nm wavelength. The calibration curve was found to be linear in the range of 2–500 μg/ml. The stated method was then validated in terms of specificity, linearity, precision, and accuracy. Additionally, the proposed method reduced the duration of the analysis.     

Development and Validation of a Reversed-Phase High-performance Liquid Chromatography Method for Determination of Desmopressin in Chitosan Nanoparticles

A simple isocratic reversed-phase high performance liquid chromatographic method was developed for determination of released desmopressin from chitosan nanoparticles in the in vitro media. The chromatographic separation was achieved with acetonitrile/water (25:75, v/v), in which water contained 0.1% v/v trifluoroacetic acid with pH=2.5 as mobile phase, a Chromolith^® Performance RP-18e column (150×4.6 mm; 5 μm) kept at 40° and ultraviolet detection at 220 nm. The compound was eluted isocritically at a constant flow rate of 1.6 ml/min. The method was validated according to the International Conference on Harmonisation guidelines. The validation characteristics included accuracy, precision, linearity rang, selectivity, limit of detection, limit of quantitation and robustness. The calibration curve was linear (r>0.9999) over the concentration rang 0.5-100 μg/ml. The limit of detection and limit of quantitation in the release media were 0.05 and 0.5 μg/ml, respectively. The proposed method had an accuracy of and intra- and inter-day precision <4.2. Furthermore, to evaluate the performance of the proposed method, it was used in the analysis of desmopressin level in real samples containing chitosan nanoparticles in the in vitro media.     

Hantzsch pre-column derivatization for simultaneous determination of alendronate sodium and its pharmacopoeial related impurity: Comparative study with synchronous fluorometry using fluorescamine

High performance liquid chromatographic (HPLC) method with a pre-column derivatization based on Hantzsch condensation reaction was applied for simultaneous determination of alendronate sodium (ALN) and its main related impurity, 4-Aminobutanoic acid (ABA) at its pharmacopeial limit. The separation of colored condensation products of ALN and ABA were achieved on Agilent Zobrax Eclipse SB-C18 analytical column (250 × 4.6 mm, 5 μm) using a mobile phase composed of acetonitrile–0.1 M acetate buffer, pH 5.0 (15:85, v/v). The flow rate was 1 mL min⁻¹. The detection was carried out at 340 nm using photo-diode array detector. Peak areas were used for the linear regression line in the range of 10–500 and 0.2–40 μg mL⁻¹ for ALN and ABA, respectively. Different conditions for the optimization of the derivatization reactions as well as for the HPLC measurement were studied. The proposed method was validated for linearity, precision, accuracy, specificity and robustness. This method was used to check the purity of ALN in the presence of ABA (related impurity) at the pharmacopeial limit (0.5%). For comparison purpose, another method was proposed which involves synchronous fluorescence measurement after ALN reaction with fluorescamine. In this method, the third derivative synchronous spectra were estimated as peak to peak measurement from 339 to 370 nm for ALN determination with LOD and LOQ of 24 and 73 ng mL⁻¹, respectively, showing very high sensitivity. Both methods have been applied for determination of the alendronate sodium (ALN) in bulk and pharmaceutical preparations without interference of additives in tablets or oral solution.     

Effect of centchroman coadministration on the pharmacokinetics of metformin in rats

Objectives:

To study the effect of centchroman, a non-steroidal oral contraceptive, coadministration on the pharmacokinetics of metformin in rats.

Materials and Methods:

The pharmacokinetic interaction of metformin was studied in normal Sprague-Dawley female rats with and without centchroman coadministration. Blood samples were analyzed using a validated high-performance liquid chromatography method to generate the pharmacokinetic profile of metformin. The C_max and t_max were directly read from the concentration–time data. Other pharmacokinetic parameters were estimated using non-compartmental analyses.

Results:

Metformin was monitored up to 10 h, and it exhibited a double-peak phenomenon. The C_{max 1}, 2.62 ± 0.32 μg/ml, and C_{max 2,} 2.96 ± 0.65 μg/ml, occurred after 0.75 and 3 h post-dose, respectively. The mean residence time (MRT), AUC_{0-4 h} and volume of distribution (Vd/F) were 4.20 ± 0.30 h, 8.53 ± 1.89 μg.h/ml and 14.24 ± 5.42 L/kg, respectively. Following centchroman coadministration, metformin showed significantly (P < 0.05) higher C_max (C_{max 1,} 3.96 ± 0.55 μg/ml and C_{max 2,} 5.21 ± 0.59 μg/ml), AUC_{0-4 h} (12.28 ± 0.73 μg.h/ml) and Vd/F (18.29 ± 1.19 L/kg), but lower MRT (3.19 ± 0.36 h) than the values obtained after metformin dosing alone. However, AUC_0-t (17.74 ± 5.58 μg.h/ml) and clearance (3.76 ± 0.80 L/h/kg) remained unchanged.

Conclusions:

The results indicate that centchroman coadministration increases the rate but not the extent of absorption of metformin in rats. However, it does not seem to alter the pharmacokinetics of metformin to clinically significant levels.     

Method Development and Validation of a Stability-Indicating RP-HPLC Method for the Quantitative Analysis of Dronedarone Hydrochloride in Pharmaceutical Tablets

A simple, precise, and accurate HPLC method has been developed and validated for the quantitative analysis of Dronedarone Hydrochloride in tablet form. An isocratic separation was achieved using a Waters Symmetry C₈ (100 × 4.6 mm), 5 μm particle size column with a flow rate of 1 ml/min and UV detector at 290 nm. The mobile phase consisted of buffer: methanol (40:60 v/v) (buffer: 50 mM KH₂PO₄ + 1 ml triethylamine in 1 liter water, pH=2.5 adjusted with ortho-phosphoric acid). The method was validated for specificity, linearity, precision, accuracy, robustness, and solution stability. The specificity of the method was determined by assessing interference from the placebo and by stress testing the drug (forced degradation). The method was linear over the concentration range 20–80 μg/ml (r² = 0.999) with a Limit of Detection (LOD) and Limit of Quantitation (LOQ) of 0.1 and 0.3 μg/ml respectively. The accuracy of the method was between 99.2–100.5%. The method was found to be robust and suitable for the quantitative analysis of Dronedarone Hydrochloride in a tablet formulation. Degradation products resulting from the stress studies did not interfere with the detection of Dronedarone Hydrochloride so the assay is thus stability-indicating.     

Adiponectin serum levels correlate with insulin resistance in type 2 diabetic patients

The adipose tissue is not only an inert storage depot for lipids, but also it secretes a variety of bioactive molecules, known as adipokines, which affect whole-body homeostasis. Adiponectin is the most abundant of these adipocytokines and is known to have a regulatory effect on the metabolism of glucose and lipid. The main objectives of this study were to evaluate the serum levels of adiponectin and to establish a correlation between adiponectin serum levels and the degree of insulin resistance in type 2 diabetic patients. Eighty participants were enrolled in this study; 61 type 2 diabetic patients and 19 apparently healthy subjects. Serum level of adiponectin was measured by enzyme-linked immunosorbent assay (ELISA) for each participant. Data collection sheet was filled with all required information for each participant. Adiponectin level in the diabetic patients (5.05 ± 2.61 μg/ml) was lower than in non-diabetic healthy controls (5.71 ± 2.35 μg/ml). When the results were compared according to gender, diabetic females showed significantly higher adiponectin levels (5.76 ± 2.64 μg/ml) than diabetic males (4.366 ± 2.43 μg/ml, P = 0.035). In addition, female diabetic patients with abdominal obesity (waist circumference (WC) ⩾ 88 cm) had lower adiponectin levels (5.58 ± 2.58 μg/ml) than diabetic females without abdominal obesity (6.96 ± 3.12 μg/ml). The correlation analysis indicated that adiponectin had a significant positive correlation with age (r = −0.450, P < 0.001). In conclusion, female diabetic patients had a statistically significant higher adiponectin level than male diabetic patients which could indicate a gender effect. Adiponectin levels were inversely related to insulin resistance; as patients with abdominal obesity had lower serum levels of adiponectin.     

Development and Validation of RP-HPLC Method for Simultaneous Estimation of Atorvastatin Calcium and Fenofibrate in Tablet Dosage Forms

A reverse phase high performance liquid chromatographic method was developed for the simultaneous estimation of atorvastatin calcium and fenofibrate in tablet formulation. The separation was achieved by Luna C18 column and methanol:acetate buffer pH 3.7 (82:18 v/v) as mobile phase, at a flow rate of 1.5 ml/min. Detection was carried out at 248 nm. Retention time of atorvastatin calcium and fenofibrate was found to be 3.02+0.1 and 9.05+0.2 min, respectively. The method has been validated for linearity, accuracy and precision. Linearity for atorvastatin calcium and Fenofibrate were in the range of 1-5 μg/ml and 16-80 μg/ml, respectively. The mean recoveries obtained for Atorvastatin calcium and fenofibrate were 101.76% and 100.06%, respectively. Developed method was found to be accurate, precise, selective and rapid for simultaneous estimation of atorvastatin calcium and fenofibrate in tablets.     

Gas chromatographic method for the determination of lumefantrine in antimalarial finished pharmaceutical products

A simple method has been developed and validated for quantitative determination of lumefantrine in antimalarial finished pharmaceutical products using gas chromatography coupled to flame ionization detector. Lumefantrine was silylated with N,O–bis(trimethyl-silyl)trifluoro-acetamide at 70°C for 30 minutes, and chromatographic separation was conducted on a fused silica capillary (HP-5, 30 m length × 0.32 mm i.d., 0.25 μm film thickness) column. Evaluation of the method within analytical quality-by-design principles, including a central composite face-centered design for the sample derivatization process and Plackett–Burman robustness verification of the chromatographic conditions, indicated that the method has acceptable specificity toward excipients and degradants, accuracy [mean recovery = 99.5%, relative standard deviation (RSD) = 1.0%], linearity (=0.9986), precision (intraday = 96.1% of the label claim, RSD = 0.9%; interday = 96.3% label claim, RSD = 0.9%), and high sensitivity with detection limits of 0.01 μg/mL. The developed method was successfully applied to analyze the lumefantrine content of marketed fixed-dose combination antimalarial finished pharmaceutical products.     

Development and Validation of a Stability-Indicating LC-Method for the Simultaneous Estimation of Levodropropizine,Chloropheniramine, Methylparaben,Propylparaben, and Levodropropizine Impurities

A simple, fast, and efficient RP-HPLC method has been developed and validated for the simultaneous estimation of Levodropropizine, Chloropheniramine, Methylparaben, Propylparaben, and the quantification of Levodropropizine impurities in the Reswas syrup dosage form. A gradient elution method was used for the separation of all the actives and Levodropropizine impurities by using the X-Bridge C18, 150 mm × 4.6 mm, 3.5 μm column with a flow rate of 1.0 mL/min and detector wavelength at 223 nm. The mobile phase consisted of a potassium dihydrogen orthophosphate buffer and acetonitrile. All the peaks were symmetrical and well-resolved (resolution was greater than 2.5 for any pair of components) with a shorter run time. The limit of detection for Levodropropizine and its Impurity B was 0.07 μg/ml & 0.05 μg/ml, whereas the limit of quantification was 0.19 μg/ml & 0.15 μg/ml respectively. The method was validated in terms of precision, accuracy, linearity, robustness, and specificity. Degradation products resulting from the stress studies were well-resolved and did not interfere with the detection of Levodropropizine, Chloropheniramine, Methylparaben, Propylparaben, and Levodropropizine Impurity B, thus the test method is stability-indicating. Validation of the method was carried out as per International Conference on Harmonization (ICH) guidelines.     

Stability-Indicating RP-HPLC Method for the Simultaneous Estimation of Doxofylline and Terbutalinesulphate in Pharmaceutical Formulations

An isocratic, stability-indicating, reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed for the quantitative determination of doxofylline and terbutaline sulphate, used for the treatment of respiratory problems. The chromatographic separation was achieved on a Zorbax-SB Phenyl 250 × 4.6mm × 5 μm column with the mobile phase consisting of a mixture of 25 mM ammonium acetate (pH 5.0) : acetonitrile (85:15 %v/v) at a flow rate of 1.0 ml/min. The eluate was monitored at 274 nm using a PDA detector. Forced degradation studies were performed on the bulk sample of doxofylline and terbutaline sulphate using acid (0.1N HCl), base (0.1N NaOH), oxidation (10% hydrogen peroxide), photolytic, and thermal degradation conditions. Good resolution was observed between the degradants and analytes. Degradation products resulting from the stress studies did not interfere with the detection of doxofylline and terbutaline sulphate, thus the assay is stability-indicating. The method has the requisite accuracy, selectivity, sensitivity, and precision for the simultaneous estimation of doxofylline and terbutaline sulphate in bulk and pharmaceutical dosage forms. The limit of quantitation and limit of detection were found to be 1.16 μg/ml and 0.38 μg/ml for doxofylline, 2.08 μg/ml and 0.62 μg/ml for terbutaline sulphate, respectively.