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Hari Kishan Reddy Ganthi et al., IJSID, 2012, 2 (4), 471-490INTRODUCTIONParacetamol (acetaminophen) is one of the most popular over-the-counter (OTC) analgesic and antipyretic drugs.Paracetamol (1-8) is available in different dosage forms: tablet, capsules, drops, elixirs, suspensions and suppositories.Aceclofenac is a non-steroidal anti-inflammatory drug (NSAID). It is used for the relief of pain and inflammation inrheumatoid arthritis, osteoarthritis and ankylosing spondylitis. The dose is 100 mg twice daily. It should not be given topeople with porphyria or breast-feeding mothers, and is not recommended for children.Chlorzoxazone is used to relieve pain and stiffness caused by muscle strains and sprains. It is used in combination withphysical therapy, analgesics (such as aspirin or acetaminophen), and rest. The side effects are upset stomach, drowsiness,dizziness, lightheadedness, weakness, skin rash or itching, yellowing of the skin or eyes and stomach pain.Chemical structures of all ingredients were represented in figure-1.All three ingredients are available in liquid pharmaceuticaldosage forms.Paracetamol Chlorozoxazone AceclofenacFigure-1: Chemical structure of all ingredientsAll ingredients have reported methods for individual and other combination products, but the objective of the presentstudy is to develop a single RP-HPLC method for the estimation of Paracetamol, Aceclofenac and Chlorozoxazone informulations and the developed and validated method is simple, novel, rugged and linear.MATERIALS AND METHODSSelection of mobile phase:Various buffer salts, pH values were tried with different organic solvents (acetonitrile or methanol) for the optimization ofmobile phase. Finally well shaped and high resolution was achieved with pH 6.0 phosphate buffer and acetonitrile at 67:33 v/vratio.Chemicals and reagents:Ortho phosphoric acid, triethyl amine (AR Grade) was procured from S.D fine chemicals. High pure (NLT 98.5%) standards(Aceclofenac, Paracetamol and Chlorozoxazone) were used for this study. HPLC grade acetonitrile were procured fromSpectrochem Pvt. Ltd. Water is prepared by mili Q system (Milli-pore).Buffer preparation: Diluted 2.0mL of ortho phosphoric acid to 1000 mL of water. Adjusted to pH 6.0 with triethyl amineFiltered through 0.45µm membrane filter and degassed.Mobile Phase: Mixed the buffer and acetonitrile in the ration 67:33 v/v.Diluent: Mobile phase.HPLC conditions:Column : Inertsil ODS 3V (150X4.6mm, 5μm)Wavelength : 275nmInjection volume:20 μLInternational Journal of Science Innovations and Discoveries, Volume 2, Issue 4, July-August 2012472
Hari Kishan Reddy Ganthi et al., IJSID, 2012, 2 (4), 471-490Flow rate : 1.0 mL/minTemperature : 30°CRun time : 30minMobile phase : Buffer and Acetonitrile 67:33 v/vPreparation of standard solution: For 100/ 500 / 500 mg Tablets:Accurately weigh and transfer about 100 mg of Aceclofenac working standard into 50 mL volumetric flask, add to it about30 mL of acetonitrile and sonicate to dissolve, dilute up to the mark with acetonitrile and mix well. (Concentration ofAceclofenac is about 2000µg/mL)Accurately weigh and transfer 100 mg of Paracetamol working standard, and 100 mg of Chlorzoxazone working standardin to 200 mL volumetric flask, add to it about 10 mL of above Aceclofenac stock solution and sonicate to dissolve, dilute upto the mark with mobile phase and mix well. (Concentration of Aceclofenac is about 100 µg/mL, concentration ofParacetamol is about 500µg/mL and concentration of Chlorzoxazone is about 500µg/mL ).Preparation of standard solution: For 100/ 325/ 250 mg Tablets:Accurately weigh and transfer about 100 mg of Aceclofenac working standard into 50 mL volumetric flask, add to it about30 mL of acetonitrile and sonicate to dissolve, dilute up to the mark with acetonitrile and mix well. (Concentration ofAceclofenac is about 2000µg/mL)Accurately weigh and transfer 65 mg of Paracetamol working standard, and 50 mg of Chlorzoxazone working standard into 200 mL volumetric flask, add to it about 10 mL of above Aceclofenac stock solution and sonicate to dissolve, dilute up tothe mark with mobile phase and mix well. (Concentration of Aceclofenac is about 100 µg/mL, concentration ofParacetamol is about 325µg/mL and concentration of Chlorzoxazone is about 250µg/mL).Preparation of standard solution: For 100/ 500 mg Tablets:Accurately weigh and transfer about 100 mg of Aceclofenac working standard into 50 mL volumetric flask, add to it about30 mL of acetonitrile and sonicate to dissolve, dilute up to the mark with acetonitrile and mix well. (Concentration ofAceclofenac is about 2000µg/mL)Accurately weigh and transfer 100 mg of Paracetamol working standard, in to 200 mL volumetric flask, add to it about 10mL of above Aceclofenac stock solution and sonicate to dissolve, dilute up to the mark with mobile phase and mix well.(Concentration of Aceclofenac is about 100 µg/mL, concentration of Paracetamol is about 500µg/mL).Preparation of standard solution: For 100/ 500/15 mg Tablets:Accurately weigh and transfer about 100 mg of Aceclofenac working standard into 50 mL volumetric flask, add to it about30 mL of acetonitrile and sonicate to dissolve, dilute up to the mark with acetonitrile and mix well. (Concentration ofAceclofenac is about 2000µg/mL)Accurately weigh and transfer 100 mg of Paracetamol working standard, in to 200 mL volumetric flask, add to it about 10mL of above Aceclofenac stock solution and sonicate to dissolve, dilute up to the mark with mobile phase and mix well.(Concentration of Aceclofenac is about 100 µg/mL, concentration of Paracetamol is about 500µg/mL).Preparation of standard solution: For 100mg Tablets:Accurately weigh and transfer about 100 mg of Aceclofenac working standard into 50.0 mL volumetric flask, add to itabout 30 mL of acetonitrile and sonicate to dissolve, dilute up to the mark with acetonitrile and mix well. (Concentrationof Aceclofenac is about 2000µg/mL)International Journal of Science Innovations and Discoveries, Volume 2, Issue 4, July-August 2012473
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Page 7 and 8: Linearity of Test Method (Inferred

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