Luận văn In vitro release of ketoprofen from proprietary and extemporaneously manufactured gels

i IN VITRO RELEASE OF KETOPROFEN FROM PROPRIETARY AND EXTEMPORANEOUSLY MANUFACTURED GELS A Thesis Submitted to Rhodes University in Fulfilment of the Requirements for the Degree of MASTER OF SCIENCE (PHARMACY) by Ralph Nii Okai Tettey-Amlalo December 2005 Faculty of Pharmacy Rhodes University Grahamstown ii ABSTRACT Ketoprofen is a potent non-steroidal anti-inflammatory drug which is used for the treatment of rheumatoid arthritis. The oral administration of ketoprofen can cause gastric irritation and adverse renal effects. Transdermal delivery of the drug can bypass gastrointestinal disturbances and provide relatively consistent drug concentrations at the site of administration. The release of ketoprofen from proprietary gel products from three different countries was evaluated by comparing the in vitro release profiles. Twenty extemporaneously prepared ketoprofen gel formulations using Carbopol® polymers were manufactured. The effect of polymer, drug concentration, pH and solvent systems on the in vitro release of ketoprofen from these formulations were investigated. The gels were evaluated for drug content and pH. The release of the drug from all the formulations obeyed the Higuchi principle. Two static FDA approved diffusion cells, namely the modified Franz diffusion cell and the European Pharmacopoeia diffusion cell, were compared by measuring the in vitro release rate of ketoprofen from all the gel formulations through a synthetic silicone membrane. High-performance liquid chromatography and ultraviolet spectrophotometric analytical techniques were both used for the analysis of ketoprofen. The validated methods were employed for the determination of ketoprofen in the sample solutions taken from the receptor fluid. Two of the three proprietary products registered under the same manufacturing license exhibited similar results whereas the third product differed significantly. Among the variables investigated, the vehicle pH and solvent composition were found have the most significant effect on the in vitro release of ketoprofen from Carbopol® polymers. The different grades of Carbopol® polymers showed statistically significantly different release kinetics with respect to lag time. When evaluating the proprietary products, both the modified Franz diffusion cell and the European Pharmacopoeia diffusion cell were deemed adequate although higher profiles were generally obtained from the European Pharmacopoeia diffusion cells. iii Smoother diffusion profiles were obtained from samples analysed by high-performance liquid chromatography than by ultraviolet spectrophotometry in both diffusion cells. Sample solutions taken from Franz diffusion cells and analysed by ultraviolet spectrophotometry also produced smooth diffusion profiles. Erratic and higher diffusion profiles were observed with samples taken from the European Pharmacopoeia diffusion cell and analysed by ultraviolet spectrophotometry. The choice of diffusion cells and analytical procedure in product development must be weighed against the relatively poor reproducibility as observed with the European Pharmacopoeia diffusion cell. iv ACKNOWLEDGEMENT I would like to sincerely thank the following people: My supervisor, Professor John Michael Haigh, for giving me this opportunity to undertake my research study with him and for his guidance, encouragement and financial support. I would also like to thank his wife, Mrs Lil Haigh for the keen interest and trust she has in me. Mr Dave Morley, Mr Leon Purdon and Mr Tichaona Samkange for their advice, assistance and technical expertise in the laboratory and to Mrs Prudence Mzangwa for ensuring the tidiness of the laboratory. The Dean and Head, Professor Isadore Kanfer and the staff of the Faculty of Pharmacy for the use of departmental facilities. Professor Roger Verbeeck for giving me his book on Dermatological and Transdermal Formulations and GraphPad PRISM® statistical software. My colleagues in the Faculty for their friendship and support. Sigma-Aldrich (Atlasville, South Africa) for the donation of ketoprofen which was facilitated by Professor Rod Bryan Walker. Aspen Pharmacare (Port Elizabeth, South Africa), Gattefossé (Saint-Priest, France), Noveon Inc. (Cleveland, USA) for the donation of excipients. My parents, especially my Father, who has always believed in me and taught me the true meaning of hard work. Finally I would like to give thanks to the Almighty God for strength, determination and power to succeed no matter what storms or challenges life brings my way. I would also like to thank him for the life of my son. v STUDY OBJECTIVES Ketoprofen is a non-steroidal anti-inflammatory, analgesic and antipyretic drug used for the treatment of rheumatoid osteoarthritis, ankylosing spondylitis and gout. It is more potent than the other non-steroidal anti-inflammatory drugs (NSAIDs) with respect to some effects such as anti-inflammatory and analgesic activities. Although ketoprofen is rapidly absorbed, metabolized and excreted, it causes some gastrointestinal complaints such as nausea, dyspepsia, diarrhoea, constipation and some renal side effects like other NSAIDs. Therefore, there is a great interest in developing topical dosage forms of these NSAIDs to avoid the oral side effects and provide relatively consistent drug concentrations at the application site for prolonged periods. The objectives of this study were: 1. To develop and validate a suitable high-performance liquid chromatographic method for the determination of ketoprofen from topical gel formulations. 2. To develop and validate a suitable ultraviolet spectrophotometric method for the determination of ketoprofen from topical gel formulations. 3. To extemporaneously manufacture topical gel formulations using Carbopol® polymers and study the effect of polymer type, pH, loading concentration and solvent composition on the in vitro release of ketoprofen. 4. To compare and contrast the in vitro release rates of ketoprofen from proprietary gel products and extemporaneously prepared topical gel formulations using the Franz diffusion cell and the European Pharmacopoeia diffusion cell. 5. To compare and contrast the in vitro release rates of different proprietary gel products and extemporaneously prepared topical gel formulations using ultraviolet spectrophotometry and high-performance liquid chromatography utilizing both the Franz diffusion cell and the European Pharmacopoeia diffusion cell. vi TABLE OF CONTENTS ABSTRACT................................................................................................................................... ii ACKNOWLEDGEMENT........................................................................................................... iv STUDY OBJECTIVES................................................................................................................. v TABLE OF CONTENTS ............................................................................................................ vi LIST OF TABLES ...................................................................................................................... xii LIST OF FIGURES ................................................................................................................... xiii CHAPTER ONE ........................................................................................................................... 1 TRANSDERMAL DRUG DELIVERY ...................................................................................... 1 1.1 PAST PROGRESS, CURRENT STATUS AND FUTURE PROSPECTS OF TRANSDERMAL DRUG DELIVERY ...................................................................... 1 1.1.1 Introduction........................................................................................................1 1.1.2 Rationale for transdermal drug delivery ............................................................2 1.1.3 Advantages and drawbacks of transdermal drug delivery .................................2 1.1.4 Innovations in transdermal drug delivery ..........................................................5 1.2 PERCUTANEOUS ABSORPTION............................................................................ 6 1.2.1 Introduction........................................................................................................6 1.2.2 Human skin ........................................................................................................7 1.2.2.1 Structure and functions of skin ..........................................................................7 1.2.2.2 The epidermis.....................................................................................................8 1.2.2.3 The viable epidermis........................................................................................10 1.2.2.4 The dermis .......................................................................................................10 1.2.3 Routes of drug permeation across the skin ......................................................10 1.2.3.1 Transcellular pathway......................................................................................11 1.2.3.2 Intercellular pathway .......................................................................................11 1.2.3.3 Appendageal pathway......................................................................................12 1.2.4 Barrier function of the skin ..............................................................................13 1.2.5 Enhancing transdermal drug delivery ..............................................................14 1.2.5.1 Chemical approach...........................................................................................14 1.2.5.1.1 Chemical penetration enhancers ..................................................................16 1.2.5.2 Physical approach ............................................................................................17 1.2.5.2.1 Iontophoresis................................................................................................17 1.2.5.2.2 Electroporation.............................................................................................19 1.2.5.2.3 Phonophoresis ..............................................................................................20 1.2.5.2.4 Microneedle .................................................................................................22 1.2.5.2.5 Pressure waves .............................................................................................23 1.2.5.2.6 Other approaches .........................................................................................23 1.2.5.2.7 Synergistic effect of enhancers ....................................................................24 vii 1.2.6 Selection of drug candidates for transdermal drug delivery ............................24 1.2.6.1 Biological properties of the drug .....................................................................25 1.2.6.1.1 Potency.........................................................................................................25 1.2.6.1.2 Half-life........................................................................................................25 1.2.6.1.3 Toxicity ........................................................................................................25 1.2.7 Physicochemical properties of the drug...........................................................25 1.2.7.1 Oil-water partition co-efficient ........................................................................25 1.2.7.2 Solubility and molecular dimensions...............................................................26 1.2.7.3 Polarity and charge ..........................................................................................26 1.3 MATHEMATICAL PRINCIPLES IN TRANSMEMBRANE DIFFUSION........ 27 1.3.1 Introduction......................................................................................................27 1.3.2 Fickian model...................................................................................................27 1.3.2.1 Fick’s first law of diffusion..............................................................................27 1.3.2.2 Fick’s second law of diffusion.........................................................................28 1.3.3 Higuchi model..................................................................................................30 1.4 METHODS FOR STUDYING PERCUTANEOUS ABSORPTION..................... 32 1.4.1 Introduction......................................................................................................32 1.4.2 Diffusion cell design ........................................................................................32 1.4.2.1 Franz and modified Franz diffusion cell..........................................................33 1.4.2.2 European Pharmacopoeia diffusion cell ..........................................................34 CHAPTER TWO ........................................................................................................................ 36 KETOPROFEN MONOGRAPH .............................................................................................. 36 2.1 PHYSICOCHEMICAL PROPERTIES OF KETOPROFEN................................ 36 2.1.1 Introduction......................................................................................................36 2.1.2 Description.......................................................................................................36 2.1.3 Stereochemistry................................................................................................37 2.1.4 Melting point....................................................................................................37 2.1.5 Solubility..........................................................................................................37 2.1.6 Dissociation constant .......................................................................................38 2.1.7 Maximum flux .................................................................................................38 2.1.8 Partition co-efficient and permeability co-efficient .........................................38 2.1.9 Optical rotation ................................................................................................38 2.1.10 Synthesis ..........................................................................................................39 2.1.11 Stability ............................................................................................................42 2.1.12 Ultraviolet absorption ......................................................................................43 2.1.13 Infrared spectrum.............................................................................................43 2.1.14 Nuclear magnetic resonance spectrum.............................................................43 viii 2.2 CLINICAL PHARMACOLOGY OF KETOPROFEN .......................................... 45 2.2.1 Anti-inflammatory effects................................................................................45 2.2.2 Analgesic and antipyretic effects .....................................................................45 2.2.3 Mechanism of action........................................................................................45 2.2.4 Therapeutic use ................................................................................................47 2.2.4.1 Indications........................................................................................................47 2.2.4.2 Contraindications .............................................................................................47 2.2.5 Adverse reactions.............................................................................................48 2.2.6 Toxicology .......................................................................................................49 2.2.7 Drug interactions..............................................................................................50 2.2.8 Pharmaceutics ..................................................................................................51 2.3 PHARMACOKINETICS OF TOPICAL KETOPROFEN .................................... 52 CHAPTER THREE.................................................................................................................... 56 IN VITRO ANALYSIS OF KETOPROFEN ............................................................................ 56 3.1 DEVELOPMENT AND VALIDATION OF AN HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC METHOD FOR THE DETERMINATION OF KETOPROFEN.................................................................................................... 56 3.1.1 Method development .......................................................................................56 3.1.1.1 Introduction......................................................................................................56 3.1.1.2 Experimental ....................................................................................................57 3.1.1.2.1 Reagents.......................................................................................................57 3.1.1.2.2 Instrumentation ............................................................................................57 3.1.1.2.3 Ultraviolet detection.....................................................................................59 3.1.1.2.4 Column selection .........................................................................................59 3.1.1.2.5 Mobile phase selection.................................................................................61 3.1.1.2.6 Preparation of selected mobile phase...........................................................62 3.1.1.2.7 Preparation of stock solutions......................................................................63 3.1.1.3 Optimisation of the chromatographic conditions.............................................63 3.1.1.3.1 Detector wavelength ....................................................................................63 3.1.1.3.2 Choice of column.........................................................................................63 3.1.1.3.3 Mobile phase composition ...........................................................................64 3.1.1.4 Chromatographic conditions............................................................................65 3.1.1.5 Conclusion .......................................................................................................65 3.1.2 Method validation ............................................................................................66 3.1.2.1 Introduction......................................................................................................66 3.1.2.2 Accuracy and bias ............................................................................................66 3.1.2.3 Precision...........................................................................................................67 3.1.2.3.1 Repeatability ................................................................................................67 3.1.2.3.2 Intermediate precision..................................................................................68 3.1.2.3.3 Reproducibility ..........