PHARMACOKINETIC EVALUATION OF AMOXICILLIN/CLAVULANICACID AND ANTACID INTERACTION Anab Fatima B.Pharm, M.Phil.(Pharmaceutics) Thesis submitted for the award of degree of Ph.D In Pharmaceutics DEPARTMENT OF PHARMACEUTICS FACULTY OF PHARMACY UNIVERSITY OF KARACHI. 1 2 In the name of ALLAH The most Beneficient and tha most Merciful………………. ALLAHUMA SALLEY ALA SYEDENA MAULANA MUHAMMEDUN WA ALA AALEHYY WA ASHABHI BARIK WASALLIM WA SALLO ALEHEY SURAT AL-FATIHAH 1. IN THE NAME OF ALLAH,THE MOST GRACIOUS,THE MOST MERCIFUL. 3 2. ALL THE PRAISES AND THANKS BE TO ALLAH,THE LORD OF THE A LAMIN(MANKIND,JINN AND ALL THAT EXIST). 3. THE MOST GRACIOUS,THE MOST MERCIFUL. 4. THE ONLY OWNER(AND THE ONLY RULING JUDGE)OF THE DAY OF RECOMPENSE(THE DAY OF RESURRECTION). 5. YOU(ALONE) WE WORSHIP,AND YOU(ALONE) WE ASK FOR HELP(FOR EACH AND EVERY THING). 6. GUIDE US TO STRAIGHT WAY. 7. THE WAY OF THOSE WHOM YOU HAVE BESTOWED YOUR GRACE,NOT(THE WAY) OF THOSE WHO EARNED YOUR ANGER NOR OF THOSE WHO WENT ASTRAY.(AAMEEN) 4 DEDICATED TO: My beloved Parentral Grand Father(late),Grand Mother(late),Parents and all those persons whose prayers bring me to this stage. ACKNOWLEDGEMENT “If ALLAH assist you, then there is none that can overcome you” (Holy Quran 3:160) All praise to Almighty Allah who gives me strength, confidence and courage to complete the thesis. I am heartiest thankful to my research supervisor Prof.Dr.S.Baqir.S.Naqvi for his valuable guidance, sincere advice, moral support and encouragement throughout the research project. He was really much helpful during research work for managing expensive chemicals and accessories for rapid completion of the project. • I am really so much thankful to Dr.Rabia Ismail Yousuf and Mrs.Rehana Saeed Assistant Professor Department of Pharmaceutics, University of Karachi for their kind and humble cooperation throughout my research project. 5 • I am also thankful to Dr. Mohammed Harris Shoaib, Mrs. Sabahat jabeen,Mr.Iyad Naeem,Dr.Shaista Assistant Professors and Ms.Fauzia Israr lecturer Department of Pharmaceutics,Faculty of Pharmacy,University of Karachi for their sincere suggestions and help whenever required during this research project. • Worthful acknowledgement to Prof.Dr.Dilnawaz Sheikh and Prof.Dr.Rafi Sheikh for their resilient support and reinforcement during the research work. • Special thanks to Prof.Dr. Javeid Iqbal Dean ,Faculty of Pharmacy,Hamdard University for his co-operation and valuable suggestions during my research project. • I am also thankful to each and every teacher including Prof.Dr.Fauzia, Prof.Dr.Nighat and Dr.Shahnaz for providing me knowledge in any way. • I appreciated all non-teaching staff of department of Pharmaceutics,University of Karachi for their co-operation during this work. • Thanks to my friend Miss. Shaheen Parveen who sincerely encouraged me during my research work. • I am also obliged to library incharge Mr.Azhar for providing me books during this research work. • I am thankful to Mr.Amjad and their Lab staff of Liaqat National Hospital for providing me plasma for my research. • I am so much thankful to Mr. Jibran and Mr.Ilyas from Imam Clinic for helping me to provide plasma from Husaini blood bank and for conducting my clinical study at Imam Clinic. • I am also indebted to whole administration of Imam Clinic including Dr.Ali Imam, Dr.Noman Palekar, Dr.Hisala and other HR and nursing staff for their humble Cooperation during my clinical study. • I am also thankful to Dr.Sajjad, Mr.Ghulam Abbas, Mr.Asim of Dr.Tahir Shamsi lab and Mr.Ayaz from Hamdard University for providing their support and help in sampling of blood during my study. • I am really thankful to Dr.Tanveer Abbas, Assistant Professsor department of Microbiology,University of Karachi for telling me useful links for literature survey. • Special thanks for my parents prayers that gave me strength during my work without which this work would not be completed. • Special thanks for my all family members especially my brothers Sheikh Abdul Khaliq,Sheikh Abdul Rehman and Sheikh Mohammed Jiyad for their kind cooperation during my research project. • I sincerely acknowledge and appreciate those volunteers namely Farrukh Najam, Mehdi Raza, Syed Zaad Ali, Mohammed Rafiq, Mohammed Asif Basheer, Syed Mohammed Mohsin Rizvi, Mohammed Uzair Shah, Waqas Ali, Adil Waqas and H.M.Bilal Anjum who participated in this research project. • Words for those who helped me in any way for the completion of this research work. 6 May ALLAH Bless Them All.(Aameeen) TABLE OF CONTENTS ABSTRACT……………………………………………………………………………………. KHULASA……………………………………………………………………………………… 1.INTRODUCTION……………………………………………………………………………..1 1.1 Drug Interaction……………………………………………………………………………...2 1. 2.Drug-Drug Interactions(DDIs)……………………………………………………………...3 1.3. Pharmacokinetic Determination of Drug-Drug Interactions…………..9 1.4. Pharmaceutical analysis of drugs in dosage forms……………………..10 1.5. ANALYSIS OF ANTIBIOTIC USING HPLC…………………….…12 1.6. HISTORY/STORY OF PENICILLIN…………………………………14 1.6.4. PHARMACOKINETIC OF PENICILLIN…………………………………………..17 7 1.6.5. AMOXICILLIN/CLAVULANIC ACID(AUGMENTIN)……………………………18 2. OBJECTIVE OF THE STUDY……………………………………………………26-27 3. LITERATURE REVIEW…………………………………………………………..28-62 4. EXPERIMENTAL……………………………………………………………………...63 4.1.PHYSICO-CHEMICAL TESTS OF DIFFERENT CO-AMOXICLAV BRANDS…….64 4.1.1. WEIGHT VARIATION TEST………………………………………………………..64 4.1.2 THICKNESS VARIATION TEST……………………………………………………65 4.1.3. HARDNESS TEST…………………………………………………………………….65 4.1.4. DISINTEGRATION TEST……………………………………………………………65 4.1.5. DISSOLUTION TEST…………………………………………………………………65 4.1.6. CONTENT ASSAY FOR CO-AMOXICLAV TABLETS BY HPLC………………67 4.2. PHARMACOKINETIC EVALUATION OF CO-AMOXICLAV TABLET……..68 4.3. Validation Parameters………………………………………………………………….74 4.4. System suitability parameters………………………………………………………….74 4.4.1. Specificity ……………………………………………………………………………….74 4.4.2. Linearity…………………………………………………………………………………74 4.4.3. Intra-Day and Inter-Day Accuracy and Precision……………………………………74 4.4.4. Lower Limit of Quantification (LLOQ) and Limit of Detection (LOD)…………….75 4.4.5. Recovery…………………………………………………………………………………75 4.4.6. Stability………………………………………………………………………………….75 4.4.7. Freeze and Thaw stability……………………………………………………………...75 4.4.8. Long Term Stability…………………………………………………………………….76 8 4.5. Pharmacokinetic Analysis……………………………………………………………...76 4.5.1. Compartmental…………………………………………………………………………76 4.5.2. Non-Compartmental……………………………………………………………………76 4.7. SERUM BACTERICIDAL TEST (SBT)…………………………………………….77 5.RESULTS…………………………………………………………………………………80- 139 % RELATIVE BIO-AVAILABILITY………………………………………………………161 7.DISCUSSION……………………………………………………………………………162- 177 8.REFERENCES …………………………………………………………………………230-245 9.APPENDIX…………………………………………………………………………………..246 NATIONAL BIOETHICS COMMITTEE(NBC) PAK ISTAN APPROVAL…………..247 PUBLICATION FROM RESEARCH WORK……………………………………………..248 LIST OF PUBLICATIONS……………………………………………………………..........249 LIST OF TABLES TABLE 1……………………………………………………………………………………6 & 7 THE MOST DANGEROUS DRUG INTERACTION.……………………………………………………………………………6 & 7 TABLE 2………………………………………………………………………………………...83 DETAILS OF DIFFERENT AMOXICILLIN/CLAVULANIC ACID PRODUCTS AVAILABLE IN PAKISTAN………………………………………………………………….83 TABLE 3………………………………………………………………………………………...84 9 WEIGHT VARIATION TEST OF AMOXICILLIN/CLAVULANIC ACID BRANDS AVAILABLE IN PAKISTAN………………………………………………………………….84 TABLE 4…………………………………………………………………………...……………86 THICKNESS VARIATION TEST OF AMOXICILLIN/CLAVULANIC ACID BRANDS AVAILABLE IN PAKISTAN………………………………………………………………….86 TABLE 5………………………………………………………………………………………...87 HARDNESS VARIATION TEST OF AMOXICILLIN/CLAVULANIC ACID BRANDS AVAILABLE IN PAKISTAN…………………………………………………………………87 TABLE 6………………………………………………………………………………………...90 DISSOLUTION OF DIFFERENT BRANDS OF AMOXICILLIN/CLAVULANIC ACID TABLET(250/125 mg) AVAILABLE IN PAKISTAN……………………………………….90 TABLE 7………………………………………………………………………………………91 HPLC ASSAY OF DIFFERENT BRANDS OF AMOXICILLIN/CLAVULANIC ACID TABLET(250/125 mg) AVAILABLE IN PAKISTAN…………………………………….....91 TABLE 8(a)……………………………………………………………………………………..92 CALIBRATION CURVE OF AMOXICILLIN STD in Human Plasma(Linearity,Accuracy and Precision)…………………………………………………92 TABLE 8(b)…………………………………………………………………………………….93 Back Calculated Concentration of Amoxicillin Standard in Human Plasma……………..93 TABLE 9(a)……………………………………………………………………………………..96 CALIBRATION CURVE OF CLAVULANIC ACID STD in Human Plasma (Linearity, Accuracy and Precision)………………………………………………………………………..96 TABLE 9(b)…………………………………………………………………………………….97 Back Calculated Concentration of Clavulanic Acid Standard in Human Plasma………97 TABLE 10(a)…………………………………………………………………………………100 INTRA-DAY PRECISION AND ACCURACY OF AMOXICILLIN IN HUMAN PLASMA(n=3)………………………………………………………………………………...100 TABLE 10(b)…………………………………………………………………………………..100 10 INTRA-DAY PRECISION AND ACCURACY OF CLAVULANIC ACID IN HUMAN PLASMA(n=3)……………………………………………………………………...…………100 TABLE 11(a)……………………………………………………………………………...101-102 INTER-DAY PRECISION AND ACCURACY OF AMOXICILLIN IN HUMAN PLASMA(n=5)……………………………………………………………………………101-102 TABLE 11(b)……………………………………………………………………………...103-104 INTER-DAY PRECISION AND ACCURACY OF CLAVULANIC ACID IN HUMAN PLASMA(n=5)…………………………………...……………………………………….103-104 TABLE 12(a)…………………………………………………………………………………..105 LOWER LIMIT OF QUANTIFICATION (LLOQ) FOR AMOXICILLIN……………..105 TABLE 12(b)…………………………………………………………………………………..106 Back Calculated Concentration of Amoxicillin……………………………………………..106 TABLE 13(a)…………………………………………………………………………………..107 LOWER LIMIT OF QUANTIFICATION (LLOQ) FOR CLAVULANIC ACID………107