AN ABSTRACT OF THESIS OF Ning-Ning Yang, for the degree of Master of Science in Pharmacy presented on June 11, 1997. Title: Product Formulations and In Vitro-In Vivo Evaluation of a Novel "Tablet-in-a-Bottle" Suspension Formulation of Amoxicil lin and Clavulanic Acid. Abstract approved: Redacted for Privacy This thesis describes a novel "Tablet-in-a-Bottle" oral suspension formulation. Ingredients with unstable physical or chemical characteristics can be placed in a core tablet, and then dry compression coated with an outer layer which provides separation from other components. The new suspension formulation comprises fast disintegrating clavulanic acid (KCA) tablets with a powder mixture containing amoxicillin. Hardness, friability, flow properties and weight uniformity of tablets for three different formulations were investigated and were all improved in a third formulation. Stability tests under different humidities were conducted. Amoxicillin and clavulanic acid in the new formulations showed the same stabilities when compared with the marketed product Augmentin®. Preliminary pharmacokinetics and bioavailability of one new formulation were evaluated by comparing in vitro release rates and in vivo urinary excretion rates. In vitro dissolution studies were carried out according to the USP )OCIII paddle method. The new formulation showed faster release rates during the first hour when stirring speed was 25 rpm. However, when 75 rpm stirring speed was applied, the dissolution profiles for the new formulation and the reference marketed product were identical. A randomized two-way crossover bioequivalence study was designed to evaluate the bioavailabilities. Cmax, Tmax and AUC0_9t of amoxicillin were within ±20% of the reference pharmacokinetic values. However, Cmax and Tmax of clavulanate were not within ±20%. Bioeqivalence between this new suspension formulation and the marketed product (Augmentin®) were evaluated using a two one-sided t-test. There is not sufficient statistical support with this test to conclude that the two products are bioequivalent. However, this is most likely due to small sample size and high intersubject variation and statistical support for bioequivalence is expected in a larger study group. ©Copyright by Ning-Ning Yang June 11, 1997 All Rights Reserved Product Formulations and In Vitro-In Vivo Evaluation of a Novel "Tablet-in-a-Bottle" Suspension Formulation of Amoxicillin and Clavulanic Acid. by Ning-Ning Yang A THESIS submitted to Oregon State University in partial fulfillment of the requirements of the degree of Master of Science Completed June 11, 1997 Commencement June 1998 Master of Science thesis of Ning-Ning Yang presented on June 11, 1997 APPROVED: Redacted for Privacy ilMajor Pro sor, representing Pharmacy Redacted for Privacy Dean of College of Pharmacy Redacted for Privacy Dean of Gradu School I understand that my thesis will become part of the permanent collection of Oregon State University libraries. My signature below authorizes release of my thesis of any reader upon request. Redacted for Privacy Nin: mg Yang, Author ACKNOWLEDGMENT Without the unconditional love and support from my grate parents, everything will be impossible for me. Thanks to my loving sister and brother, they are not only my family but also my best friends. Thanks to my major professor, Dr. Ayres, who brought me into a very interesting field and showed me his creativity, patience and kindness. He can always help me out of my difficulties during my graduate study. Thanks to Dr. Coll for her advice, support and friendship. She was there with me in the lab when I went through all kinds of problems about my experiments. Thanks to all faculty and staff of School of Pharmacy for their help. Also I am grateful to the technical support of Lisa Kam, Jennifer Pelster and Christine Eskander. I will remember all the graduate students in the department who made my school life interesting. Especially thanks to my friends. I will never forget the time we spend together, the fun we had and the encouragement you gave. Life here will always be part of my precious memories. Finally, my LORD, who not only looked after me but also guided my every step. He is always there when I need Him. CONTRIBUTION OF AUTHORS Dr. James Ayres was involved in the formulation design, data interpretation, and writing of each manuscript. Dr. Jacqueline Coll assisted in methodology development and data collection for the study. TABLE OF CONTENTS Page INTRODUCTION 1 SUSPENSION FORMULATION DEVELOPMENT AND STABILITY STUDIES 2 ABSTRACT 3 INTRODUCTION 4 MATERIALS AND METHODS 5 Materials 5 HPLC System. 5 Derivatization Procedure for Clavulanic Acid Assays 6 Imidazole Reagent 6 Mobile Phase and Internal-Standard Solutions 7 Standards for Calibration Curve 7 Aqueous Stability of Derivative of Clavulanic Acid 9 Formulation Design 9 Initial Formulation (Formulation A) 10 Tablet Hardness and Friability for Formulation A 12 Tablet Weight Uniformity 14 Flow Properties for Formulation A 14 Formulation B 15 Tablet Hardness and Friability for Formulation B 17 TABLE OF CONTENTS (Continued) Page Tablet Weight Uniformity 18 Flow Properties for Formulation B 19 Formulation C 20 Tablet Hardness and Friability for Formulation C 22 Tablet Weight Uniformity 23 Formulation Stability Study 25 High Humidity Study 25 Low Humidity Study I 26 Low Humidity Study II 26 RESULTS AND DISCUSSION 27 Aqueous Stability of Imidazole Derivative of Clavulanic Acid 27 Formulation Stability Study 34 Humidity Studies 45 CONCLUSIONS 57 REFERENCES 58 PRELIMINARY PHARMACOKINETICS AND BIOAVAILABILITY EVALUATION 59 ABSTRACT 60 INTRODUCTION 61 MATERIALS AND METHODS 62 TABLE OF CONTENTS (Continued) Page UV Analysis 62 Dissolution Studies 62 Subjects 63 Study Design 63 Collection of Urine Samples 63 Buffer Solution 64 Data Analysis 64 RESULTS AND DISCUSSION 65 Dissolution Studies 65 Biostudies 65 CONCLUSIONS 79 REFERENCES 80 CONCLUSIONS 82 BIBLIOGRAPHY 83 APPENDIX Individual Biostudy Data Plots 86 LIST OF FIGURES Figure Page 1.1 Typical calibration curve for clavulanic acid analyzed using HPLC 8 1.2 Aqueous stability of imidazole dervative of clavulanic acid 29 1.3 Degradation of clavulanic acid dervative over 16 hours 30 1.4 Degradation of clavulanic acid dervative after 1.5 days 31 1.5 Degradation of clavulanic acid dervative after 6 days 32 1.6 Aqueous stability of imidazole dervative of clavulanic acid 33 1.7 pH profile of suspension formulation A vs. Augmentin® 35 1.8 Amoxicillin stability of formulation A vs. Augmentin® 36 1.9 Amoxicillin stability of formulation A vs. Augmentin® (corrected) 37 1.10 Clavulanic acid stability of formulation A vs. Augmentin® 38 1.11 pH profile of suspension formulation B vs. Augmentin® 40 1.12 Amoxicillin stability of formulation B vs. Augmentin® 41 1.13 Clavulanic acid stability of formulation B vs. Augmentin® 42 1.14.a Amoxicillin/clavulanate suspension appearance 43 1.14.b Amoxicillin/clavulanate suspension appearance 44 1.15 Amoxicillin profile in high humidity study 47 1.16 Amoxicillin profile in low humidity study I (corrected) 48 1.17 Clavulanic acid profile in high humidity study 49 1.18 Amoxicillin profile in low humidity study I 50 1.19 Amoxicillin profile in low humidity study I (corrected) 51 LIST OF FIGURES (Continued) Figure Page 1.20 Clavulanic acid profile in low humidity study I 52 1.21 Amoxicillin profile in low humidity study II 53 1.22 Clavulanic acid profile in low humidity study II 54 1.23 Amoxicillin profile for effect of stearic acid on tablet 55 1.24 Clavulanic acid profile for effect of stearic acid on tablet 56 2.1 Dissolution profiles of amoxicillin from new formulation and Augmentin at 25 rpm and 75 rpm. 66 2.2 Dissolution profiles of clavulanic acid from new formulation and Augmentin® at 25 rpm and 75 rpm. 67 2.3 Cumulative mean urinary data of amoxicillin for new formulation and Augmentin® 69 2.4 Cumulative mean urinary data of clavulanic acid for new formulation and Augmentin® 70 2.5 Mean excretion rates of amoxicillin for new formulation and Augmentin® 71 2.6 Mean excretion rates of amoxicillin for new formulation and Augmentin® 72 LIST OF TABLES Table Page 1.1 Ingredients and amount used for KCA tablet of formulation A 11 1.2 Ingredients and amount used for powder mix of formulation A .. 12 1.3 Results of tablet hardness test for formulation A 13 1.4 Results of friability for formulation A 13 1.5 Weight of core tablet of formulation A 14 1.6 Weight of finished tablet of formulation A 14 1.7 Flow properties for formulation A 15 1.8 Ingredients and amount used for KCA tablet of formulation B 16 1.9 Ingredients and amount used for powder mix of formulation B .. 17 1.10 Results of tablet hardness test for formulation B 18 1.11 Results of friability for formulation B 18 1.12 Weight of core tablet of formulation B 19 1.13 Weight of finished tablet of formulation B 19 1.14 Flow properties for formulation B 19 1.15 Ingredients and amount used for KCA tablet of formulation C 21 1.16 Ingredients and amount used for powder mix of formulation C 22 1.17 Results of tablet hardness test for formulation C 23 1.18 Results of friability for formulation C 23 1.19 Weight of core tablet of formulation C 23 1.20 Weight of fmished tablet of formulation C 24 LIST OF TABLES (Continued) Table Page 1.21 Flow properties for formulation C 24 1.22 Individual flow properties for ingredients used in formulation C 24 2.1 Mean amoxicillin pharmacokinetic parameters 65 2.2 Mean clavulanic acid pharmacokinetic parameters 65 2.3 Amoxicillin pharmacokinetic parameters