ORAL BIOAVAILABILITY Basic Principles, Advanced Concepts, and Applications potx

In the 1960s and 1970s, appli-cation of the physical sciences to the problem of oral drug delivery produced the first wave of major advances that shaped the development of the modern com

Thomas J Long School of Pharmacy and Health Sciences

University of the Pacific

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Library of Congress Cataloging-in-Publication Data:

Oral bioavailability : basic principles, advanced concepts, and applications / edited by Ming Hu, Xiaoling Li.

p ; cm – (Wiley series in drug discovery and development)

Includes bibliographical references.

ISBN 978-0-470-26099-9 (cloth)

1 Drugs–Bioavailability 2 Drug development 3 Intestinal absorption I Hu, Ming, Ph D.

II Li, Xiaoling, Ph.D III Series: Wiley series in drug discovery and development.

[DNLM: 1 Biological Availability 2 Drug Delivery Systems 3 Intestinal Absorption QV 38]

RM301.6.O73 2011

615 .19– dc22

2011002983 oBook ISBN: 978-1-118-06759-8

ePDF ISBN: 978-1-118-06752-9

ePub ISBN: 978-1-118-06758-1

10 9 8 7 6 5 4 3 2 1

to my mom Qihua Chang whose constant love and encouragement persists to this date,

to my wife Yanping Wang whose company endears constant push for perfection, and

to my children Vivian and William whose energy and noise are missed now they are in college.

—Ming Hu Dedicated to my grandmother Yunzhi Su,

my parents Bailing Li and Jie Hu,

my wife Xinghang, and

my children Richard and Louis for their unconditional love, encouragement, and understanding.

—Xiaoling Li

Ming Hu and Xiaoling Li

2 Physicochemical Characterization of Pharmaceutical Solids 7

Smita Debnath

Lauren Wiser, Xiaoling Gao, Bhaskara Jasti, and Xiaoling Li

4 In Vitro Dissolution of Pharmaceutical Solids 39

Josephine L P Soh and Paul W S Heng

5 Biological and Physiological Features of the Gastrointestinal

Paul C Ho

6 Absorption of Drugs via Passive Diffusion and Carrier-Mediated

Miki Susanto Park and Jae H Chang

7 In Vitro–In Vivo Correlations of Pharmaceutical Dosage Forms 77

Deliang Zhou and Yihong Qiu

Rashim Singh and Ming Hu

9 Efflux of Drugs via Transporters —The Antiabsorption Pathway 111

Jae H Chang, James A Uchizono, and Miki Susanto Park

Leslie M Tompkins and Hongbing Wang

vii

11 Protein Binding of Drugs 145

Antonia Kotsiou and Christine Tesseromatis

12 Urinary Excretion of Drugs and Drug Reabsorption 167

Pankaj Gupta, Bo Feng, and Jack Cook

13 Pharmacokinetic Behaviors of Orally Administered Drugs 183

Jaime A Y´a˜nez, Dion R Brocks, Laird M Forrest, and Neal M Davies

Venugopal P Marasanapalle, Xiaoling Li, and Bhaskara R Jasti

15 Drug–Drug Interactions and Drug–Dietary Chemical Interactions 233

Ge Lin, Zhong Zuo, Na Li, and Li Zhang

16 Anatomical and Physiological Factors Affecting Oral Drug

Ayman El-Kattan, Susan Hurst, Joanne Brodfuehrer, and Cho-Ming Loi

Zhong Qiu Liu and Ming Hu

18 Drug Transporters and Their Role in Absorption and Disposition

David J Lindley, Stephen M Carl, Dea Herrera-Ruiz, Li F Pan, Lori B Karpes,

Jonathan M E Goole, Olafur S Gudmundsson, and Gregory T Knipp

Takashi Sekine and Hiroyuki Kusuhara

John R Cardinal and Avinash Nangia

21 Lipid-Based and Self-Emulsifying Oral Drug Delivery Systems 343

Sravan Penchala, Anh-Nhan Pham, Ying Huang, and Jeffrey Wang

22 Prodrug Strategies to Enhance Oral Drug Absorption 355

Sai H S Boddu, Deep Kwatra, and Ashim K Mitra

Puchun Liu and Steven Dinh

Marilyn E Morris and Yash A Gandhi

25 Interplay Between Efflux Transporters and Metabolic Enzymes 401

Stephen Wang

26 Regulatory Considerations in Metabolism- and Transporter-Based Drug

Yuanchao (Derek) Zhang, Lei Zhang, John M Strong, and Shiew-Mei Huang

27 Caco-2 Cell Culture Model for Oral Drug Absorption 431

Kaustubh Kulkarni and Ming Hu

CONTENTS ix

28 MDCK Cells and Other Cell-Culture Models of Oral Drug Absorption 443

Deep Kwatra, Sai H S Boddu, and Ashim K Mitra

29 Intestinal Perfusion Methods for Oral Drug Absorptions 461

Wei Zhu and Eun Ju Jeong

30 Liver Perfusion and Primary Hepatocytes for Studying

Cindy Q Xia, Chuang Lu, and Suresh K Balani

31 In vivo Methods for Oral Bioavailability Studies 493

Ana Ruiz-Garcia and Marival Bermejo

32 Determination of Regulation of Drug-Metabolizing Enzymes and

Bin Zhang and Wen Xie

33 Computational and Pharmacoinformatic Approaches to Oral

Miguel ´ Angel Cabrera-P´erez and Isabel Gonz´alez- ´ Alvarez

In Spring of 1983, I took a position at The University

of Michigan There I met my first Chinese student, Ming

Hu, from mainland China, and began a personal and

professional relationship that has lasted for nearly 30 years

He is now a Professor at the University of Houston and

one of the two editors of this book I am very pleased to

have observed his contributions to science and his success

as a scientist over the nearly 30 years I have known him

and followed his career It is a pleasure to write this

foreword for this book coedited by Ming and his former

classmate at Shanghai Medical University, Prof Xiaoling

Li at University of the Pacific

This book has two purposes, to give readers a

contem-porary understanding of the science of oral bioavailability

and to present the state-of-the-art tools that can be used

to advance the science of oral bioavailability and solve

problems in the development of drug products for oral

administration It presents the advances in the science of

oral bioavailability over the last five decades This

mul-tidisciplinary scientific field has steadily progressed from

an emphasis on physical sciences such as solubility and

solid state properties, to incorporating the significant recent

advances in the biological sciences that emphasize

trans-porters, enzymes, and the biological and physiological

pro-cesses that influence their expression and function

I will note some of the evolutionary and perhaps

revo-lutionary steps this field of oral bioavailability has taken

over last five decades In the 1960s and 1970s,

appli-cation of the physical sciences to the problem of oral

drug delivery produced the first wave of major advances

that shaped the development of the modern commercial

oral dosage form and the science of oral bioavailability

Important physicochemical principles and strategies such

as manipulation of dissolution via physical manipulation

of the drug and drug product and chemical modificationusing prodrugs were developed These approaches are rou-tinely considered and applied in the drug product devel-opment process today The principles governing sustainedand controlled release formulations were developed in those

“early” years (e.g., Higuchi equation), and have becomewidely applied in the later decades of the twentieth cen-tury In the 1980s, important progress in the science oforal bioavailability was led by the development of twocritical absorption models, rat intestinal segment perfu-sion model (developed in my laboratory) and Caco-2 cellmono-layer culture model (developed in Dr Ronald T Bor-chardt’s lab) Prof Hu studied in both laboratories, and was

an early contributor to the development of both of thesesystems for the study of oral absorption These methodshave since become widely adapted by the pharmaceuticalindustries This set the basis for predicting oral absorptionand partitioning bioavailability into its component process,dissolution/release, transport/permeation, and metabolism,notability distinguishing absorption and systemic availabil-ity During the 1980s, major advances were also made

in the study of metabolism in the intestine as well as

the liver, particularly the cytochrome P 450s and resultant

potential drug– drug interaction mechanisms In addition topredicting oral absorption, my laboratory also pioneeredthe concept of exploiting the intestinal mucosal cell pep-tide transporter (hPEPT1) to improve the oral absorption ofpolar drugs by making a prodrug, chemically combining thedrug and an amino acid with a peptide-bond like structure.This mechanistic concept is the basis for the absorption

of many polar drugs and prodrugs The development ofseveral approved prodrugs including valacyclovir and val-ganciclovir, while originally empirical, is based on these

xi

xii FOREWORD

transport mechanisms In the 1990s, I established the

con-cept of the Biopharmaceutical Classification System (BCS),

partitioning drugs into classes for drug development and

drug product regulation This BCS approach has found wide

use in drug discovery, development as well as regulation

It has been adapted by regulatory authorities and

govern-ments around the world as a basis for the regulation of drug

product quality

During this same period, the US Food and Drug

Administration began the mandate of requiring studies that

predict drug–drug interactions based on the sciences that

were developed during the past two decades Study of efflux

transporters began in the 1990s and has exploded in the

twenty-first century While efforts to make an inhibitor of

p-glycoprotein for anticancer application have not produced

an approved drug, it is likely that the future will see such

a development The explosion in the study of transporters

is ongoing, with the recent addition of efflux transporters

such as multidrug resistance-related proteins (MRPs), breast

cancer resistant protein (BCRP), and uptake transporters

such as organic anion transporting peptides (OATP), organic

anion transporters (OATs), and carboxylic acid transporter

(CAT) Such advances in our mechanistic understanding

of oral bioavailability will most certainly lead to future

advances in therapy

The advances in the science of oral bioavailability is

driven by the needs to develop orally administered drugs,

which remains the most acceptable patient compliant means

of administering drugs to patients across the globe today

Although the scientific basis was most often the pursuit

of industrial scientists, a lack of rapid advancement in

the science of oral bioavailability became recognized as

a hurdle in the drug development process in the early1990s as many highly potent compounds (high affinity

ligands), for example, HIV in vitro were inactive in humans.

In a timely or even a watershed event, the NationalInstitute of Health in 1994 organized a conference on “OralBioavailability,” where scientists of various backgroundswere organized to address the complex problem facingpotent yet poorly bioavailable drug candidates, particularlyanti-HIV candidates Senior managements in many ofthe major pharmaceutical companies became aware ofand recognized the importance of “bioavailability” as thepharmaceutical industry was working hard to fast track thedevelopment of anti-HIV drugs This led to investment

by the pharmaceutical industries in the technology andscientists to tackle this oral delivery problem While actualnumbers can be hard to obtain and interpret, my impression

is that the attention to bioavailability has led to thedecrease in the percentage of clinical trial failures due tooral bioavailability problems Looking even further intothe future, I believe the science of oral bioavailabilitywill be driven by the needs for personalized medicine,individualized treatment plan tailored to patients, and

by the commercial need to increase the efficiency andefficacy of oral drug product development This bookprovides a comprehensive survey of the modern study

of the science of oral bioavailability in the twenty-firstcentury

GORDON L AMIDON, Ph.D

The University of Michigan, Ann Arbor, MI

Since the concept of bioavailability has been introduced,

significant progress has been made in understanding the

science of oral bioavailability and in improving the oral

delivery of drugs Yet, we also find that there is still

much to be discovered to have a good handle on oral

bioavailability As a subject, bioavailability encompasses

the knowledge and technologies from various disciplines

A pharmaceutical scientist in a specific research area will

benefit from a treatise on the topic Hence, the objective

of this book is to provide the framework for fundamental

concepts and contemporary practice of bioavailability in

pharmaceutical research and drug development

It is our belief that this book provides both the basic

concepts to a novice and the advanced knowledge to

veteran pharmaceutical scientists and graduate students

in related research fields Chapter 1 gives a high level

summary of this book The basic concepts of bioavailability

are covered in Chapter 2–13 From Chapter 14 to 26,

the advanced concepts of bioavailability are discussed

in greater depth Various approaches and methods for

improving and studying bioavailability are highlighted in

Chapter 27 to 33 The comprehensive coverage of topics

on bioavailability in this book offers readers a choice of

logically building their knowledge on bioavailability from

basic concepts to advanced applications or `a la carte based

on their specific needs

A book with such diverse contents requires a ciplinary effort Without the efforts of contributors fromdifferent areas, this book would have not been a reality

multidis-We would like to personally thank all authors for theircontributions and patience during the completion of thisbook project Sincere thanks are gratefully extended to MrJonathan Rose at John Wiley and Sons, Inc and Dr BingheWang (the book series editor) for their patience, under-standing, support, and confidence in us We would alsolike to express our appreciations to Mrs Kathy Kassab forher invaluable secretarial assistance, and to Haseen Khanfor her tireless effort in the book production Finally, wewould like to thank the world renowned scientist and lead-ing expert in bioavailability, Prof Gordon L Amidon forwriting an insightful and inspiring forward for this book

Suresh K Balani, Drug Metabolism and

Pharmacokinet-ics, Millennium Pharmaceuticals, Inc., 35 Landsdowne

Street, Cambridge, MA 02139

Marival Bermejo, Department of Engineering, Pharmacy

and Pharmaceutical Technology Section, School of

Pharmacy, Universidad Miguel Hern´andez de Elche,

Carretera Alicante Valencia km 87, San Juan de Alicante

03550, Alicante, Spain

Sai H.S Boddu, Division of Pharmaceutical Sciences,

University of Toledo, Toledo, OH

Dion R Brocks, Faculty of Pharmacy, University of

Alberta, Alberta, Canada

Joanne Brodfuehrer, Department of Pharmacokinetics,

Dynamics and Metabolism, Pfizer Global Research and

Development, Cambridge, MA

Miguel Angel ´ Cabrera-P´erez, Molecular Simulation

and Drug Design Department, Centro de Bioactivos

Qu´ımicos, Universidad Central “Marta Abreu” de Las

Villas, Carretera a Camajuan´ı, Km 51/2, Santa Clara,

Villa Clara, C.P 54830, Cuba

John R Cardinal, J R Cardinal Consulting LLC,

Wilm-ington, NC

Stephen M Carl, Department Industrial and Physical

Pharmacy, College of Pharmacy, Nursing and Health

Sciences, Purdue University, West Lafayette, IN 47907

Jae H Chang, Department of Drug Metabolism and

Pharmacokinetics, Genentech, South San Francisco, CA

Jack Cook, Clinical Pharmacology, Specialty Care

Busi-ness Unit, Pfizer Inc., Groton, CT

Neal M Davies, Department of Pharmaceutical Sciences,

College of Pharmacy, Washington State University,Pullman, WA

Smita Debnath, Merck Frosst Canada Ltd, Kirkland,

Bo Feng, Pharmacokinetics, Dynamics and Metabolism,

Pfizer Inc., Groton, CT

Laird M Forrest, Department of Pharmaceutical

Chem-istry, University of Kansas, Lawrence, KS

Yash A Gandhi, Department of Pharmaceutical Sciences,

School of Pharmacy and Pharmaceutical Sciences,University at Buffalo, State University of New York,Buffalo, NY

Xiaoling Gao, Department of Pharmaceutics and

Medici-nal Chemistry, Thomas J Long School of Pharmacy andHealth Sciences, University of the Pacific, Stockton, CA95211

Current Affiliation: Institute of Medical Sciences, hai Jiaotong University School of Medicine, Shanghai,

Shang-PR China

Isabel Gonz´alez- ´ Alvarez, Department of Engineering:

Pharmacy and Pharmaceutical Technology section,School of Pharmacy, Universidad Miguel Hern´andez deElche, Carretera Alicante Valencia km 87., San Juan

03550, Alicante, Spain

xv

Jonathan M.E Goole, Laboratory of Pharmaceutics and

Biopharmaceutics, Universite Libre de Bruxelles,

Insti-tute of Pharmacy, 1050 Brussels, Beligum

Olafur S Gudmundsson, Discovery Pharmaceutics,

Pharmaceutical Candidate Optimization, Bristol-Myers

Squibb, Princeton, NJ

Pankaj Gupta, Clinical Pharmacology, Specialty Care

Business Unit, Pfizer Inc., Groton, CT

Paul W.S Heng, Department of Pharmacy, National

University of Singapore, Singapore

Dea Herrera-Ruiz, Universidad Aut´onoma del Estado de

Morelos, Facultad de Farmacia, Cuernavaca, Mexico

Paul C Ho, Department of Pharmacy, National University

of Singapore, Singapore

Ming Hu, Department of Pharmacological and

Pharma-ceutical Sciences, College of Pharmacy, University of

Houston, 1441 Moursund Street, Houston, TX 77030

Shiew-Mei Huang, Offices of Clinical Pharmacology,

Center for Drug Evaluation and Research, Food and

Drug Administration, Building 51, Room 3106 10903

New Hampshire Avenue, Silver Spring, MD 20993

Ying Huang, Department of Pharmaceutical Sciences,

College of Pharmacy, Western University of Health

Sciences, Pomona, CA 91766

Susan Hurst, Department of Pharmacokinetics, Dynamics

and Metabolism, Pfizer Global Research and

Develop-ment, Groton, CT

Bhaskara R Jasti, Department of Pharmaceutics and

Medicinal Chemistry, Thomas J Long School of

Phar-macy and Health Sciences, University of the Pacific,

Stockton, CA 95211

Eun Ju, Korea Institute of Toxicology (KIT), 19

Sin-seongno, Yuseong, Daejeon, 305–343, Republic of

Korea

Gregory T Knipp, Department Industrial and Physical

Pharmacy, College of Pharmacy, Nursing and Health

Sciences, Purdue University, 575 Stadium Mall Dr.,

Room 308A, West Lafayette, IN 47907–2091

Antonia Kotsiou, Department of Pharmacology,

Are-taieion University Hospital, Vas Sophias 76, 11528,

Athens, Greece

Kaustubh Kulkarni, Department of Pharmacological and

Pharmaceutical Sciences, College of Pharmacy,

Univer-sity of Houston, 1441 Moursund Street, Houston, TX

77030

Hiroyuki Kusuhara, Laboratory of Molecular

Pharma-cokinetics, Graduate School of Pharmaceutical Sciences,

The University of Tokyo, Tokyo, Japan

Deep Kwatra, Division of Pharmaceutical Sciences,

School of Pharmacy, University of Missouri-KansasCity, 2464 Charlotte Street, 5005 Rockhill Road, KansasCity, MO 64108-2718

Na Li, Department of Pharmacology, The Chinese

Univer-sity of Hong Kong, Hong Kong

Xiaoling Li, Department of Pharmaceutics and Medicinal

Chemistry, Thomas J Long School of Pharmacy andHealth Sciences, University of the Pacific, Stockton, CA95211

Ge Lin, School of Biomedical Sciences, Faculty of

Medicine, The Chinese University of Hong Kong, HongKong

David J Lindley, Department Industrial and Physical

Pharmacy, College of Pharmacy, Nursing and HealthSciences, Purdue University, West Lafayette, IN 47907

Puchun Liu, Noven Pharmaceuticals, Inc., 11960 SW 144

Street, Miami, FL 33186

Zhong Qiu Liu, Department of Pharmaceutics, School of

Pharmaceutical Sciences, Southern Medical University,Guangzhou 510515, China

Cho-Ming Loi, Department of Pharmacokinetics,

Dynam-ics and Metabolism, Pfizer Global Research and opment, San Diego, CA

Devel-Chuang Lu, Drug Metabolism and Pharmacokinetics,

Mil-lennium Pharmaceuticals, Inc., 35 Landsdowne Street,Cambridge, MA 02139

Venugopal P Marasanapalle, Department of

Pharmaceu-tics and Medicinal Chemistry, Thomas J Long School ofPharmacy and Health Sciences, University of the Pacific,Stockton, CA 95211

Current Affiliation: Forest Research Institute, 220 SeaLane, Farmingdale, NY 11735

Ashim K Mitra, Division of Pharmaceutical Sciences,

School of Pharmacy, University of Missouri-KansasCity, 2464 Charlotte Street, 5005 Rockhill Road, KansasCity, MO 64108-2718

Marilyn E Morris, Department of Pharmaceutical

Sci-ences, School of Pharmacy and Pharmaceutical SciSci-ences,University at Buffalo, State University of New York,Buffalo, New York, NY

Avinash Nangia, Vaunnex Inc., Sharon, Massachusetts

Li F Pan, Department Industrial and Physical Pharmacy,

College of Pharmacy, Nursing and Health Sciences,Purdue University, West Lafayette, IN 47907

CONTRIBUTORS xvii

Miki Susanto Park, Department of Pharmaceutics and

Medicinal Chemistry, Thomas J Long School of

Phar-macy and Health Sciences, University of the Pacific,

Stockton, CA 95211

Sravan Penchala, Department of Pharmaceutical

Sci-ences, College of Pharmacy, Western University of

Health Sciences, Pomona, CA 91766

Anh-Nhan Pham, Department of Pharmaceutical

Sci-ences, College of Pharmacy, Western University of

Health Sciences, Pomona, CA 91766

Yihong Qiu, Global Pharmaceutical Regulatory Affairs,

Abbott Laboratories, 200 Abbott Park Rd, RA71-Bldg

AP-30-1, Abbott Park, IL, 60064–6157

Ana Ruiz-Garcia, Clinical Pharmacology, Oncology

Divi-sion, Pfizer Inc, 10646 Science Center Dr CB-10, San

Diego, CA 92121

Takashi Sekine, Department of Pediatrics, Toho

Univer-sity School of Medicine, Tokyo, Japan

Rashim Singh, Department of Pharmacological and

Phar-maceutical Sciences, College of Pharmacy, University

of Houston, 1441 Moursund Street, Houston, TX

Josephine L.P Soh, Pfizer Global Research and

Develop-ment, UK

John M Strong,∗ Offices of Pharmaceutical Sciences,

Center for Drug Evaluation and Research, Food and

Drug Administration, Building 51, Room 3106 10903

New Hampshire Avenue, Silver Spring, MD 20993

Christine Tesseromatis, Department of Pharmacology,

Medical School, Athens University, M Assias 75,

11527, Athens, Greece

Leslie M Tompkins, Department of Pharmaceutical

Sci-ences, School of Pharmacy, University of Maryland, 20

Penn Street, Baltimore, MD 21201

James A Uchizono, Department of Pharmaceutics and

Medicinal Chemistry, Thomas J Long School of

Phar-macy and Health Sciences, University of the Pacific,

Stockton, CA 95211

Hongbing Wang, Department of Pharmaceutical Sciences,

School of Pharmacy, University of Maryland, 20 Penn

Street, Baltimore, MD 21201

Jeffrey Wang, Department of Pharmaceutical Sciences,

College of Pharmacy, Western University of Health

Sciences, 309 E Second Street, Pomona, CA 91766

∗Deceased.

Stephen Wang, Drug Metabolism and Pharmacokinetics,

Merck Research Laboratories, 2015 Galloping HillRoad, Kenilworth, NJ 07033

Current Affiliation: DMPK/NCDS, Millennium: TheTakeda Oncology Company, 35 Landsdowne Street,Cambridge, MA 02139

Lori B Karpes, Department Industrial and Physical

Phar-macy, College of PharPhar-macy, Nursing and Health ences, Purdue University, West Lafayette, IN 47907

Sci-Lauren Wiser, Department of Pharmaceutics and

Medici-nal Chemistry, Thomas J Long School of Pharmacy andHealth Sciences, University of the Pacific, Stockton, CA95211

Cindy Q Xia, Drug Metabolism and

Pharmacokinet-ics, Millennium Pharmaceuticals, Inc., 35 LandsdowneStreet, Cambridge, MA 02139

Wen Xie, Center for Pharmacogenetics and Department of

Pharmaceutical Sciences, University of Pittsburgh, 633Salk Hall, 3501 Terrace Street, Pittsburgh, PA 15216

Jaime A Y´a ˜nez, Department of Drug Metabolism and

Pharmacokinetics (DMPK), Alcon Laboratories, Inc.,

6201 S Freeway, Fort Worth, TX 76134

Bin Zhang, Center for Pharmacogenetics and Department

of Pharmaceutical Sciences, University of Pittsburgh,Pittsburgh, PA 15216

Lei Zhang, Offices of Clinical Pharmacology, Center

for Drug Evaluation and Research, Food and DrugAdministration, Building 51, Room 3106 10903 NewHampshire Avenue, Silver Spring, MD 20993

Current affiliation: Frontage Laboratories, Inc., Exton,

PA 19341

Li Zhang, School of Pharmacy, Faculty of Medicine, The

Chinese University of Hong Kong, Hong Kong

Yuanchao (Derek) Zhang, Offices of Clinical

Pharmacol-ogy, Center for Drug Evaluation and Research, Food andDrug Administration, Building 51, Room 3106 10903New Hampshire Avenue, Silver Spring, MD 20993Current affiliation: Frontage Laboratories, Inc., Exton,PA

Deliang Zhou, Manufacturing Science and Technology,

Global Pharmaceutical Operations, Abbott Laboratories,North Chicago, IL

Wei Zhu, Department of Pharmaceutical Sciences and

Clinical Supplies, Merck and Co., Inc., 770 SumneytownPike, P.O Box 4, WP 75B-210, West Point, PA 19486

Zhong Zuo, School of Pharmacy, Faculty of Medicine,

The Chinese University of Hong Kong, Hong Kong

Portal vein Blood

Bile

Bypass hepatocytes

NAT FMO

2D6 2E1 3A4(+5)

2B6

2C19 2C9(+8)

2D6 2E1 3A4(+5)

2B6

2C19 2C9(+8)

2D6 2E1 3A4(+5)

2B6

2C19 2C9(+8)

2D6 2E1 3A4(+5)

2B6

2C19 2C9(+8)

2D6 2E1 3A4(+5) (a)

Figure 8.1 (a) Contribution of individual human enzyme systems to metabolism of marketed

drugs; (b) contribution of individual P450s in metabolism of drugs UGT indicates

uridinedinu-cleotide phosphate (UDP) glucuronosyl transferase; FMO, flavin-containing monooxygenase; NAT,

N-acetyltransferase; MAO, monoamine oxidase; P450, cytochrome P450 Source: Adapted from

2%

(b) (a)

CYP1A2 CYP2B6 CYP2C CYP2D6 CYP2E1 CYP3A4 other

researchers identify new substrate drugs Source: Adapted from Wang and Tompkins (2008).

SULT1A1 FMO5

OATP2 Carboxylesterase CYP7A

XRE

CYP1A CYP1B UGT1A1 UGT1A3 UGT1A6 BCRP

Figure 10.6 Target genes of PXR, CAR, and AhR (a) PXR and CAR are shown dimerized totheir common partner, RXR and sitting response elements XREM (xenobiotic response enhancermodule) and PBREM (phenobarbital response enhancer module), respectively Overlapping targetgenes are boxed in the center with CAR-specific targets shown above and PXR-specific targetsshown below (b) AhR is shown bound to partner, ARNT and activating its target genes afterbinding to the XRE (xenobiotic response element; often called the dioxin response element).UGT1A1 represents a common target gene of all three receptors

OAT4

PepT1/2 OCTN2 OAT1

OAT3

MATE-1/2

MRP2/4 PgP

BCR BCRP P-gp MRP

MCT

MRP OCT

POT Intracellular accumulation

POT?

Nucleus

E

TAP-1 TAP-2

R

Passive Transcellular Diffusion

Abluminal

Figure 18.1 A representative depiction of a number of transporters expressed in a human intestinal

cell illustrating the complexity of the system Source: Modified from Carl et al (2007).

r

Efflux pump

Prodrug not recognized

S I D E Biotransformation of prodrug to drug

Enzymes or chemicals

Drug

Prodrug

Transporter r

Transporter ATP

ATP

Efflux pump

Drug not recognized

Figure 22.1 Role of efflux and influx transporters in intestinal absorption

Paracellular transport

Carrier mediated transport

e l l

c y

Tight cell junction

Carrier mediated efflux transporter

y t o p l a s

ATP

Active

transport

Transporter Facilitator protein Efflux pump

Figure 22.3 Intestinal drug transport mechanisms

(a)

(b)

7

1 2 6

7 27

CYP2C8 CYP2C9 CYP2C19 CYP2D6 CYP3A Non-CYP Phase I

41

40 19 3 16 47

13

39

1A1 1A3 1A4 1A6 1A8 (extrahepatic) 1A9

1A10 (extrahepatic) 2B7

2B15 Other

Figure 26.2 (a) Distribution of CYP and non-CYP phase I enzyme pathways for 65 oral drugs(NMEs approved between 2003 and 2008) (3) (b) Distribution of UGT enzyme pathways reported

for 103 drugs from the literature (Kiang et al., 2005) and Drugs@FDA Most of them are expressed

in the liver except for UGT1A8 and UGT 1A10

Figure 28.4 Simplified structure of P-gp structure and function.

CDF-Mrp-2 Substrate

Ca 2+ buffer pretreated for 10 min Ca 2+ buffer pretreated for 10 min

+MK571 (Ca2+buffer pretreated for 10 min) +MK571 (Ca2+ buffer pretreated for 10 min)

MK571: a Mrp-2 inhibitor

BC BC

Figure 30.6 Fluorescence and phase-contrast micrographs of hepatocytes treated with CDFDA

in the presence and absence of MK571 These results demonstrate that depletion of Ca2 + opensthe tight junction and enables compounds to be released from bile canaliculi (BC) MK571, whichdid not disrupt BC, blocked the excretion of CDF into BC via its inhibitory effect on Mrp2

(wild type)

m PXR knockout

h PXR

transgene

(loss-of-function)

(humanized function) (gain-of-function)

Figure 32.2 Strategies to create the loss-of-function knockout, gain-of-function transgenic, and

the combined “humanized” function models Source: Adapted from Gong et al (2005), with the

permission of the publisher

1.1 INTRODUCTION

Oral bioavailability of a drug is a measure of the rate and

extent of the drug reaching the systemic circulation and is

a key parameter that affects its efficacy and adverse effects

Therefore, study of oral bioavailability has received

consid-erable attention in scientific arena Unfortunately, we are

unable to predict bioavailability as a priori to this date,

although we have made significant progress in

understand-ing various components of this complex puzzle, includunderstand-ing

solubility (e.g., aqueous solubility), partition coefficients

(e.g., octanol/water), absorption (e.g., permeability across

the Caco-2 cell membrane), metabolism (e.g.,

microsome-mediated phase I metabolism), and excretion (e.g., efflux

via p-glycoprotein) However, understanding a few of these

components would not allow us to accurately predict a

drug candidate’s bioavailability in humans Therefore, oral

bioavailability remains to be a highly experimental

param-eter that eludes prediction from modern computational

or experimental approaches, although some preliminary

Oral Bioavailability: Basic Principles, Advanced Concepts, and Applications, First Edition Edited by Ming Hu and Xiaoling Li.

© 2011 John Wiley & Sons, Inc Published 2011 by John Wiley & Sons, Inc.

progress has been made in recent years Continued progress

to develop a better and more thorough understanding ofphysicochemical and biochemical profiling of drug or drug-like molecules would be needed to alleviate the problemsassociated with bioavailability, and some progress has beenmade in the last decade (Ho and Chien, 2009) Poor oralbioavailability is also one of the leading causes of fail-ures in clinical trials This is because compounds withlow bioavailability would have a highly variable expo-sure between individuals If a compound has an averagebioavailability of 5%, it would easily vary in the range of0.5–10%, a 20-fold difference This difference makes theselection of an appropriate dose particularly difficult sincetoo little may yield no impact and too much could result intoxicity, which is not acceptable for most drugs that desirechronic administration

The reasons why oral bioavailability is such a lenge for development of drugs or drug-like substances(e.g., nutraceuticals) are several-fold: first, many physic-ochemical and biological factors contribute to the bioavail-ability of a compound; second, many scientific disciplinesare involved but few, if any, scientists are good at morethan one specific area; third, reliable scaling from ani-mal models to humans is often absent; and fourth, oralbioavailability is often seriously affected by diet andpolypharmacy, neither of which can be adequately con-trolled in a standard clinical trial, considering the diversity

chal-of the population— the elderly and seriously ill patients

In addition, we are normally able to gain access only tolimited body fluids such as blood and urine, and fluidssurrounding the target tissues/cells are often not accessi-ble This limitation makes bioavailability, a measure of theextent and rate of absorption and the elimination processes,

1

2 BARRIERS TO ORAL BIOAVAILABILITY—AN OVERVIEW

bile and kidney, so other elimination route (e.g., exhalation) is not shown (See insert for color representation of the figure.)

really representing only systemic blood exposure to drugs

(Fig 1.1) Therefore, it is not surprising that bioavailability

would sometimes not satisfactorily correlate with efficacy

Oral bioavailability remains a major challenge to

the development of nutraceuticals and naturally derived

chemopreventive agents For example, many scientists are

interested in developing plant-derived polyphenols into

chemopreventive agents Polyphenols are derived from

plants and consumed in the form of fruits, vegetables,

spices, and herbs In different regions of the world, a

large percentage of dietary polyphenols are consumed

in the form of flavonoids from various sources of food

intake, although cultural and dietary habit dictates which

forms of polyphenols are consumed (Fletcher, 2003; Slavin,

2003; Aggarwal et al., 2007) On the other hand, a large

percentage of population do not take sufficient quantities

of fruits and vegetables for a variety of reasons (Adhami

and Mukhtar, 2006) Therefore, scientists are interested

in developing a pill that will mimic the effects of

ingesting fruits and vegetables Yet, today their effort

has not produced a single polyphenolic chemopreventive

agent; the unsuccessful attempt may be attributed to the

poor bioavailability of polyphenols (usually <5%) Poor

bioavailability makes the evaluation of a chemopreventive

agent a particular challenge, since the clinical trials for

chemopreventive agents often involve a large population

for a prolonged period and extremely high costs

When all of the above-mentioned challenges are takeninto consideration in the product development of drugs orchemopreventive agents, it is obvious that developing anappropriate oral dosage form for drug candidate or can-didate of chemopreventive agent is not a trivial or straightforward task Although pharmaceutical scientists have greatdifficulty in predicting and enhancing bioavailability, thereward is also immense as the vast majority of top rev-enue and prescription leaders are orally administered drugs.Therefore, we devote this chapter to briefly introduce each

of the factors that influence bioavailability and guide thereaders to the appropriate chapters in this book where theycan obtain in-depth contents of each related topic

As an oral dosage form enters the oral cavity and thenthe gastrointestinal (GI) tract, several barriers must beovercome before it can reach the systemic circulation andthe therapeutic target On its way to the therapeutic target,

a drug in a given dosage form will need to first overcomethe preabsorption barrier formed by the hostile acidic andenzymatic environment in the stomach and intestine Thenthe drug would encounter the primary barrier formed bythe biological membrane, that is, the wall of the GI tract.Once a drug successfully passes the intestinal epitheliumbarrier, the drug will need to overcome another barrierconsisting of transporters and enzymes, which utilize theefflux mechanism to pump the drug back to the intestineand degrade the drug via the first-pass effect There are

many factors that will affect a drug molecule’s ability

to overcome these barriers to reach and remain in the

systemic circulation These factors include the inherent

physicochemical properties of the drug molecules,

biologi-cal characteristics of the GI tract, pathophysiologibiologi-cal state,

drug–drug or drug–food interactions, etc

1.1.1 Physicochemical Factors

Various physicochemical factors will affect the oral

bioavailability of a drug The importance of

physicochemi-cal properties of a drug molecule in drug absorption or

per-meation was illustrated by Lipinski’s “rule of 5” (Lipinski

et al., 2001) Because of the importance of

physicochemi-cal properties, a thorough characterization of drug substance

would provide fundamental information for drug discovery,

as well as for formulation and dosage form development

The characterization of key physicochemical properties of

drug substances is described in Chapter 2 One of the key

physicochemical properties that play a crucial role in the

drug absorption/permeation is solubility Solubility defines

the maximum concentration of a drug available for

absorp-tion or permeaabsorp-tion, while another important

physicochem-ical property, dissolution rate, controls the rate of the drug

available for absorption or permeation Factors that affect

solubility and dissolution rate surely will also influence the

bioavailability of the drug Variation of pH in the GI tract

causes drugs to behave differently in terms of solubility and

dissolution rate along the GI tract For an acidic drug, a low

solubility and slow dissolution rate in the stomach, where

pH is low, can be expected, while for a basic drug, poor

sol-ubility owing to precipitation in the intestinal fluids, where

pH is high, would happen An understanding of the basic

concept of solubility and dissolution rate forms a solid

foun-dation for comprehending bioavailability Physicochemical

factors also dictate the permeability of drug molecules

Solubility and permeability of a drug are such important

factors for drug absorption or bioavailability The combined

effect of these two factors would determine the

developa-bility and bioavailadevelopa-bility of a compound to a certain extent

Chapters 3 and 4 discuss the two important factors related

to drug absorption, namely, solubility and dissolution rate

Chapter 6 provides the fundamentals for drug permeation

or absorption Chapter 7 correlates the physicochemical

parameters in vitro and in vivo.

1.1.2 Biological Factors

Oral delivery is a preferred route for the administration

of small molecule drugs, because the intestine has a very

large surface area, in excess of 200 m2, which is the

size of a tennis court Since oral absorption is limited by

the drugs with molecular weight <600 Da and effective

absorption window in the GI tract, permeability of drug

through intestinal membrane, physiology of GI tract, andmetabolism of drugs in absorption and transport havebecome important factors with respect to bioavailability

GI tract is not always a hospitable place for drugabsorption Enzymes are secreted in the GI tract at arate of about 45 g per day in adult humans Althoughthe primary functions of these enzymes are to digestnutrients such as protein, carbohydrates, and nucleotides,their presence is one of the primary reasons why proteinand genetic materials (for gene therapy) cannot be deliveredorally, unless special formulation approaches are used

In addition to surviving in the hostile environment, adrug needs to overcome the barriers posted by theintestinal epithelium Intestinal epithelium is a complextissue with advanced cellular structures and metabolicfunctionality The presence of cellular junctions, especiallytight junction, severely impedes the passage of molecules

with molecular weight >200 Da via the paracellular route.

Therefore, the vast majority of the drug molecules mustuse the transcellular route Transcellular route is affected

by a myriad of interrelated but sometimes competingbiological factors Although it was always believed thatlipophilic molecules have an easy access to the transcellularroute, the presence of various efflux transporters thatpreferentially bind with lipophilic molecules could seriouslylimit the absorption of lipophilic molecules In addition,

if a molecule is too lipophilic (e.g., log P > 5), it may

be retained in the cellular membrane Because intestinalepithelial cells have a functional existence of only three

to four days (near or at the tip of the intestinal villus),molecules that bind too tightly will be eventually lostwhen the epithelial cells slough off Hydrophilic drug

molecules with molecular weight >200 Da cannot penetrate

the intestinal epithelium by passive diffusion; they musthave special structural motifs that make them attractive forthe nutrient transporters such as amino acid transporters(Chapter 17), the small peptide transporter 1 (or PepT1)(Chapter 18), organic ion transporters (Chapter 19), andnucleobases transporters Assuming drug molecules getinto the epithelial cells, there are intestinal first-passmetabolisms capable of further degrading their chance

to reach the systemic circulation These metabolisms areprimary phase II metabolism although CYP3A4 is thought

to be decently active in the enterocytes In Chapters 5, 6,

8, and 10, the barriers to oral bioavailability have beendescribed in greater details, with emphasis on GI biology(Chapters 5 and 16), drug absorption (Chapter 6) andmetabolism pathways (Chapter 8), and drug excretion bythe enterocytes (Chapter 9)

The last major barrier to oral bioavailability, perhapsthe most well-known one, is the first-pass metabolism inthe liver Since all drugs absorbed via the GI tract (exceptthe last few centimeters of the rectum) have to enter theportal vein and encounter hepatocytes (each of which can

4 BARRIERS TO ORAL BIOAVAILABILITY—AN OVERVIEW

be called metabolic superstar ), escaping liver metabolism

is the last step in the oral absorption process

In addition to these important factors, protein binding,

which affects drug distribution and free drug available for

metabolism, has also featured in this book (Chapter 11)

Lastly, another general factor that affects the systemic

exposure, elimination via urine, is discussed in Chapter 12

Taken together, substantial information is provided on the

pharmacokinetic behaviors of drugs following oral

admin-istration (Chapter 13), many of which can be explained

using the information learned from previous chapters

1.1.3 Diet and Food Effects

Development of drugs is becoming more global and

multi-dimensional The day where a standard diet is appropriate

for clinical trials across the globe is probably over

Tradi-tionally, diet and food effects have focused on the protein

content, caloric intake, and fat amounts, and few if any have

carefully examined the effects of other more exotic dietary

components such as spices More recently, consumers are

taking ever large quantities of dietary supplements with

increasing frequency and variety Although we are unable

to completely address how these changes in the diet will

impact drug bioavailability, various attempts have been

made Chapter 14 has shed some light on this topic

1.1.4 Drug Interactions

Drug interactions remain a serious concern for the

develop-ment of new drugs On the basis of the target patient

popula-tion, certain types of drug interactions are not acceptable to

the manufacturer, FDA (Food and Drug Administration of

the United States of America), or both Traditionally, drug

interactions are classified into pharmacodynamic

interac-tions and pharmacokinetic interacinterac-tions and this book mainly

deals with the latter in Chapter 15, since it is a book focused

on oral bioavailability

Classical pharmacokinetic drug interactions typically

involve phase I metabolic enzymes, and clinical examples

of this type of interactions are well documented in the

literature From a pharmacokinetic point of view, drug

interaction may cause a rise or a fall in body exposure of

drug, that is, change in Cmax(maximal drug concentration)

and/or AUC (area under the curve) values From a

mechanistic point of view, a rise in exposure is typically

related to inhibition of enzyme activities or down regulation

of relevant metabolic enzymes, whereas a fall in exposure

is typically related to activation of dormant enzymes or

induction of relevant metabolic enzymes

More recently, FDA is contemplating the inclusion of

efflux transporters into the drug interaction universe, and

provisional guidance has been issued This could further

complicate the drug development process and increase

the complexity and cost of development The reasons areseveral-fold First, many drugs undergo efflux and phase

I metabolism simultaneously and therefore it is difficult

to sort out the precise mechanisms of drug interactions.Second, there are few demonstrated clinical cases whereinteractions with efflux transporters have been confirmed

as the sole source of drug interactions Third, metabolicenzymes may develop significant interplay with the effluxtransporters such that it would be necessary to interactwith both components of the disposition in order to displayclinically significant effects Many of these are discussed

in Chapter 26

1.1.5 Formulation Factors

Based on the physicochemical and biological factors thataffect the bioavailability, we can use different strategies toovercome the barriers for bioavailability (Chaubal, 2004).One can design a dosage form that can avoid the harshenvironment in the stomach or optimally utilize the absorp-tion window For example, an enteric-coated dosage formwill not dissolve until it reaches the intestine while a gas-troretentive drug delivery system can prolong the residenttime of dosage forms in the GI tract Oral dosage forms can

be coated with rate-limiting membranes that can controlthe rate of drug release from the dosage form Increasedsolubility and dissolution rate are effective ways to improvebioavailability One can create an effective dissolution ratethat supplies the proper amount of soluble drug for absorp-tion Nanoparticles are increasingly becoming an importantpart of modern drug dosage form design as incorporation ofnanoparticles can often alleviate challenges associated withpoor solubility Varieties of pharmaceutical technologiesand drug delivery approaches have been used to improvethe physicochemical properties of the drugs Approachesfor various dosage form design and solubility/dissolutionenhancement can be found in Chapters 20 and 21.Enhancement of solubility and dissolution rate allowsformulation scientists to manipulate the factors related todrug substances for improving bioavailability To improvebioavailability, one can also modulate the permeability ofdrug across the intestinal epithelium Chapters 22 and 23represent some of the attempts in this direction

1.2 SCIENTIFIC DISCIPLINES INVOLVED

It was once thought that the intestinal tract is a very modating organ for drug absorption because this organ isbuilt to absorb nutrients If drugs are good for us, shouldthe intestinal tract be there to do what benefits us? It wasnot until 1990s that the myth—if medicinal chemists can

accom-develop active compounds in vitro, formulation scientists can make a finished product to deliver them in vivo —was

found to be untrue Development of two classes of

compounds, renin inhibitors and HIV protease inhibitors,

convinced drug development scientists and senior

manage-ment in pharmaceutical companies that oral bioavailability

matters because the intestine is not just an absorption organ

Scientists with various training and education

back-grounds are involved in the development of orally

administered drugs Aside from classical biologists and

medicinal chemists that are involved in the drug

discov-ery phase, more preclinical ADME (absorption,

distribu-tion, metabolism, and excretion) works are now integrated

into the drug discovery area Once a candidate is selected,

additional ADME work plus toxicology will be needed

to further advance the candidate into clinical trials Then,

physician researchers, nurse practitioners, biostatisticians,

marketing professionals, and pharmaceutical economists

become involved in the clinical studies At the same time, a

different group of scientists, many with engineering

back-ground, are making decisions on the manufacturing and

processing parameters Therefore, it is not entirely

unex-pected that scientists in different groups do not always

have the proper background to fully understand each other

One of the purposes of this book is to provide an

easy-to-understand section for the scientists in different areas to

understand ADME and their terminologies

One of the important goals of this book is to give a

prac-tical guide to the use of several state-of-the-art technologies

and methodologies to readers who may or may not be

famil-iar with these techniques in order to gain a basic

understand-ing and knowledge for practical approaches and/or methods

The readers can dive into the contents ranging from

advanced reviews on various important efflux transporters

that affect drug absorption and excretion (Chapter 24),

to the coupling between efflux transporters and enzymes

(Chapter 25), to computational methods and approaches to

predict bioavailability (Chapter 33) For the technology and

methodology, the readers can find a detailed description on

the Caco-2 cell culture model (Chapter 27), MDCK (Madin

Darby canine kidney and other related cell culture models

(Chapter 28), intestinal perfusion (Chapter 29), liver

perfu-sion (Chapter 30), primary hepatocytes (Chapter 30), in vivo

pharmacokinetics (Chapter 31), and methods to determine

regulation of enzymes and transporters (Chapter 32)

Rapid advances in the human genomics and proteomics

promise to better predict factors determining human

responses to drugs Although the price of sequencing the

whole human genome remains out of the practical range at

present, rapid advances in this area are expected to make

the practice an economic reality in the not so distant future

Recent passage of a law by the Congress of the United

States of America to ban discrimination based on genetic

information should provide the legal framework to protect

an individual’s right to utilize his/her genetic information

for better health care This law and progress in the

economics of human genome sequencing will mean that,

in the near future, we could develop criteria that will dosepatients according to his/her genetic makeup—a radicalprogress in the field of individualized pharmacy We willall welcome the day that geneticists become active partic-ipants in the drug development process, instead of limitingtheir participation only in the drug discovery process

Oral bioavailability remains a big challenge for smallmolecules, and an even bigger challenge for macromolec-ular drugs such as protein Despite decades of effort, there

is no product for oral insulin This book devotes itself tothe study of various biological and physicochemical prin-ciples and methodologies that can be used to understandthe oral bioavailability problems and to devise strategiesthat can be used to overcome these problems Although

we still cannot predict bioavailability as a priori at this

time, it is getting closer to the moment when we would beable to do so for the small molecular drugs Efforts under-taken by various drug delivery companies are on the brink

of achieving oral delivery of active insulin Therefore, thescience of oral bioavailability is closer than ever in the his-tory of drug development to become an enabler of drugdevelopment, instead of an obstacle to drug development.Together with the advent of individualized genomic infor-mation, we are heading to a day when each of the patientscould receive drug according to his/her conditions We areall very hopeful that this day is within our grasp in the nearfuture

REFERENCES

Adhami VM and Mukhtar H (2006) Polyphenols from green tea

and pomegranate for prevention of prostate cancer Free Radic

Res 40:1095–1104.

Aggarwal BB, Sundaram C, Malani N, and Ichikawa H (2007)

Curcumin: the Indian solid gold Adv Exp Med Biol 595:1–75.

Chaubal MV (2004) Application of drug delivery technologies in

lead candidate selection and optimization Drug Discov Today

9:603–609.

Fletcher RJ (2003) Food sources of phyto-oestrogens and their

precursors in Europe Br J Nutr 89(Suppl 1):S39–43.

Ho RJ and Chien JY (2009) Drug delivery trends in clinical trialsand translational medicine: Updated analysis of ClinicalTri-

als.gov database J Pharm Sci 98:1928–1934.

Lipinski CA, Lombardo F, Dominy BW, and Feeney PJ (2001)Experimental and computational approaches to estimate sol-ubility and permeability in drug discovery and development

settings Adv Drug Deliv Rev 46:3–26.

Slavin J (2003) Why whole grains are protective: biological

mechanisms Proc Nutr Soc 62:129–134.

2.2.1 Ultraviolet/visible (UV/vis) diffuse

2.3.3 X-ray powder diffractometry (XRPD) 12

2.3.4 Thermal methods of analysis 13

2.3.5 Thermogravimetric analysis (TGA) 13

2.3.6 Differential thermal analysis (DTA) 13

2.3.7 Differential scanning calorimetry (DSC) 13

Oral Bioavailability: Basic Principles, Advanced Concepts, and Applications, First Edition Edited by Ming Hu and Xiaoling Li.

© 2011 John Wiley & Sons, Inc Published 2011 by John Wiley & Sons, Inc.

2.1 INTRODUCTION

Solid dosage forms, such as tablets and capsules, are a mon means of administration of pharmaceutically active

com-ingredient (API) in humans (Gennaro, 1985; Brittain et al.,

1991; Brittain, 1995) They are manufactured by ing a number of powdered solids together, most com-monly, blending or mixing of multiple components, milling

process-or size reduction, granulation which may be done eitherusing a granulating fluid or in the dry state (roller com-paction), compression into tablets and coating (Lachman

et al., 1986) All these processes may be influenced by the

physical properties of the solids, and, thus, their tance is being increasingly recognized A number of testshave been included in the United States Pharmacopeia(USP) to characterize these physical properties of pow-dered solids (The United States Pharmacopeia and NationalFormulary, 2002) It is important to have a comprehensiveunderstanding of the physical characteristics earlier duringproduct development to prevent future problems such aswas observed with ritonavir In this case, a new, more ther-modynamically stable, less soluble polymorph was being

impor-formed during bulk drug manufacturing after the product

was launched in the market As a consequence, stability

and bioavailability of the product were at risk (Bauer et al.,

2001)

The API and excipients used in formulations can exist

in various physical phases that can impact processability,

7

stability, and performance of the formulation, which have

been briefly summarized below

2.1.1 Crystalline and Amorphous Phases

Crystalline materials are characterized by a regular,

well-defined, and long-range (>20 A) periodicity in the˚

arrangement of the constituent atoms, ions, or molecules

They exhibit sharp melting points and characteristic

X-ray powder patterns Amorphous materials lack long-range

order in their molecular arrangement They do not have

melting points and their X-ray patterns show a broad halo

The most common route of obtaining amorphous solids is

by rapid cooling of a melt below its melting point where

the structural characteristics of the liquid are maintained

but the viscosity is much higher This is considered as

a “supercooled liquid.” On further cooling, a

character-istic temperature known as the glass transition

tempera-ture (Tg )is observed below which the solid is “kinetically

frozen” into an unstable glassy state with properties

dif-ferent from both the supercooled liquid and crystalline

form Amorphous solids possess higher free energy than

their crystalline counterparts due to which they have higher

apparent solubilities, dissolution rates, and enhanced

chem-ical reactivities (Hancock and Zografi, 1997) For example,

the amorphous form of sulfapyridine was found to have a

higher apparent solubility and dissolution rate than its

crys-talline counterpart (Gouda et al., 1977) When the same

dose was administered to dogs, therapeutically adequate

concentrations were obtained with amorphous novobiocin

while the crystalline form was not absorbed at all This

difference in bioavailability was attributed to the

differ-ences in apparent solubility and dissolution rates between

the amorphous and crystalline phases (Mullins and Macek,

1960)

2.1.2 Polymorphic Forms

Polymorphism is the ability of a compound to crystallize

as two or more phases having different arrangements

and/or molecular conformations in the crystal lattice

(Brittain, 1999) Polymorphs of a given compound are

chemically identical but, in general, different in structure

and properties including dissolution rates, melting point,

density, hardness, and crystal shape The choice of the

polymorphic form may determine the physical and chemical

stability, compressibility, and bioavailability of the drug

substance (Haleblian et al., 1971) A compound can exist

in a number of polymorphic forms but only one form

is thermodynamically stable at a given temperature and

pressure, while the others are metastable The metastable

polymorphs have higher free energies, apparent solubilities,

and dissolution rates than their stable counterparts The

larger the free energy difference between the stable

and metastable polymorphs, the higher is the expecteddifference in solubility (Brittain, 1999) For example, whendifferent polymorphs of tolbutamide (forms I, II, III, andIV) were administered to dogs, forms II and IV showedhigher bioavailabilities than forms I and III (Kimura

et al., 1999) While it is generally preferred to formulate

the most thermodynamically stable form, under somecircumstances, it may be desirable to use the metastableforms in formulations because of their higher dissolutionrates However, owing to higher reactivity of metastablephases, their physical, as well as chemical, stability needs

to be carefully monitored during processing and storage

2.1.3 Solvates

Solvates are adducts or molecular complexes containingsolvent molecules within the crystal structure in either sto-ichiometric or nonstoichiometric proportions If the incor-porated solvent molecule is water, the solvate is referred

to as a hydrate (Vippagunta et al., 2001) The

incorpora-tion of the solvent molecule in the crystal lattice results

in differences in physical and pharmaceutical properties.Differences in solubility of the hydrated and anhydrousphases may result in a difference in bioavailability Whenanhydrous ampicillin and ampicillin trihydrate were admin-istered to dogs, the peak serum levels following adminis-tration of the anhydrate were higher and occurred earlier

(Poole et al., 1968) Similar results were observed after

their oral administration to humans (Ali and Farouk, 1981).Similarly, differences in the solubility of the anhydrousand the dihydrate phases of carbamazepine resulted indifferences in bioavailability when administered to dogs.The bioavailability of carbamazepine dihydrate was lowerthan that of the corresponding anhydrate forms (Kobayashi

et al., 2000).

The discussion above indicates that it is important tocharacterize materials to select and control the desirable

form in formulations Brittain et al (1991) have defined

a systematic approach for physical characterization ofpharmaceutical solids where the material properties wereclassified as molecular, particulate, and bulk properties Theobjective of this chapter is to summarize the techniquesused for physicochemical characterization of solids based

on this classification This chapter provides only a briefoutline and the reader is encouraged to go through thereferences for a deeper understanding of the subject

2.2 MOLECULAR LEVEL PROPERTIES

These are defined as those characteristics that could bemeasured at a molecular level They include spectroscopictechniques and are based on properties such as molecularinteractions and molecular bond energies These studies

MOLECULAR LEVEL PROPERTIES 9

can be performed at a very early stage with the advantage

of using minimum amount of material and providing

information regarding polymorphic form, solvate phase,

and crystallinity

2.2.1 Ultraviolet/Visible (UV/vis) Diffuse Reflectance

Spectroscopy

Although UV/vis techniques are widely used for the

anal-ysis of solutions, it can be adapted for the

character-ization of solids This is performed by using diffuse

reflectance techniques instead of in the transmission mode

It is associated with the fraction of radiation that

pene-trates into the bulk and then emerges (Brittain, 1995) The

instrumentation consists of a light source, a

monochro-mator, an integrating sphere, and a detector The

instru-ment and sample preparation may be optimized to

mini-mize undesirable specular reflectance (surface reflectance

of the incident beam) Several diffuse reflectance

theo-ries have been proposed but the Kubelka –Munk theory

is the most generally accepted Diffuse reflectance can be

k and s = the molar absorption and the scattering

coefficients, respectively, and

R∞= the reflectivity of an infinitely thick sample

The equation is valid in weakly absorbing systems

with a small particle size (∼1 μm) without

signif-icant contribution from specular reflectance (Kubelka,

1948)

UV/vis diffuse reflectance spectroscopy has been used

to evaluate solid–solid interactions in formulations The

effect of formulation composition and processing variables

on the microenvironment in solid dosage forms was

eval-uated using indicator probes providing a measure of the

physicochemical nature of the formulation in the solid state

(Govindarajan et al., 2006a) The technique has also been

used to correlate pH of the solution before freeze-drying and

chemical reactivity of the freeze-dried material

(Govindara-jan et al., 2006b), and evaluation of the Maillard reaction

between a primary amine and lactose (Wu et al., 1970).

2.2.2 Vibrational Spectroscopy

Infrared (IR) and Raman spectroscopies are complementary

techniques widely used for the characterization of

phar-maceutical solids The IR region in the electromagnetic

spectrum can be divided into three regions; the near-IR

(4000–14,000 cm−1), mid-IR (400– 4000 cm−1), and

far-IR (100– 400 cm−1) with the near- and mid-IR generally

being used for analysis When a sample is irradiated,absorption of IR energy results in transitions betweenmolecular vibrational and rotational energy levels (either ofsingle pairs of atoms or groups of atoms) The molecularvibrations depend on the structure of the analyte and thuscan be used for the identification of a molecular identity

IR and Raman spectroscopies complement each other byevaluating various functional groups While polar groupssuch as C O and NH are likely to be IR active, bondssuch as C C and SS are more likely to be Raman active(Brittain, 1995)

Spectrometers usually consist of an electromagneticsource, a sample chamber, and a detector For IR anal-ysis, sample preparation can be carried out using differ-ent methods including (i) alkali halide pellet, where theanalyte is pulverized with either KBr or KCl and com-pressed into discs; (ii) mull preparation, where the analyte

is mixed with ∼1 mg of mineral oil; (iii) use of a neatpowdered sample in diffuse reflectance infrared Fouriertransform spectroscopic technique (DRIFTS), it is nonin-vasive but is particle size dependent; (iv) attenuated totalreflectance (ATR), where the sample is placed in contactwith an IR transmitting crystal with a high refractive indexthrough which the IR beam is directed and then penetrates

a few micrometers into the sample; and (v) tic spectroscopy—modulated IR radiation is selectivelyabsorbed by the sample Similar to DRIFTS, no samplepreparation is required for Raman spectroscopy where anal-ysis can be carried out on small samples of the neat mate-rial Both IR and Raman spectroscopes can be combinedwith an optical microscope enabling analysis of a few crys-tals (Bugay, 2001)

photoacous-Since IR and Raman techniques are based on themolecular structure of materials, they can be used forthe analysis of polymorphs and solvates Five forms

of tranilast (three polymorphs and two solvates) werecharacterized by using IR and Raman spectroscopes Whileform II was determined to be a conformationally distinctivepolymorph, form III had a different packing with weakerintermolecular hydrogen bonds Owing to its ease andrapidity, IR spectroscopy was used to determine polymorph

contamination during process development (Vogt et al.,

2005) Some authors have reported the use of thesetechniques for the analysis of materials with differentdegrees of crystallinity (Okumura and Otsuka, 2005)and solid dispersions (Konno and Taylor, 2006) Ramanspectroscopy was used to detect amorphous indomethacin

at a level of 2% w/w using a calibration curve ofindomethacin with various degrees of crystallinity Agrowing application of NIR spectroscopy is in the field

of process analytical technologies (PATs) such as real-timemonitoring of blending processes (El-Hagrasy and Drennen,2005)

2.2.3 Solid-State Nuclear Magnetic Resonance

(SSNMR)

Nuclear magnetic resonance (NMR) spectroscopy has been

extensively used to analyze molecules in the solution phase

The use of NMR for the study of solids is now being widely

used for the characterization of pharmaceutical solids

There are several interactions that a nucleus with a magnetic

moment undergoes when placed in a magnetic field Some

of these include Zeeman interactions (interaction between

magnetic moment of the nucleus and the external magnetic

field), dipole –dipole interaction (magnetic coupling of two

nuclei through space), chemical shift (magnetic shielding by

surrounding electrons), and spin–spin couplings (indirect

coupling between two spins) A thorough discussion of

all these interactions is outside the scope of this chapter;

therefore, references are provided (Brittain, 1995; Tishmack

et al., 2003).

There are, however, some differences in the analysis of

solutions and solids The strong dipolar interactions and

chemical shift anisotropy (CSA) in solids lead to broad

peaks in the spectrum These are not observed in solutions

because of the fast random motions of the samples CSA

occurs in solids since the orientation of the molecules

is fixed with respect to the magnetic field These issues

can be overcome by probes that can be subjected to high

decoupling power and by rapidly spinning the sample

Magic angle spinning (MAS) can also be used wherein

the line width can be minimized by spinning the sample

at an angle of 54.7◦with the magnetic field (Brittain, 1995;

Tishmack et al., 2003).

Solid-state nuclear magnetic resonance (SSNMR) has

been used for the qualitative and quantitative

character-ization of solids that can exist as polymorphs, solvates,

or in the amorphous state MAS NMR was used for the

evaluation of several polymorphs and hydrates of

olanza-pine (Fig 2.1) (Reutzel-Edens et al., 2003) The spectra

were characterized by highly resolved, sharp resonances

The unique chemical shifts reflected the presence of the

carbon nuclei in different polymorphic forms and states of

hydration Presence of form II in form III as an impurity

was observed using X-ray diffraction (XRD) and this was

confirmed by using SSNMR (Reutzel-Edens et al., 2003).

In a review by Shah et al (2006), several examples are

discussed where SSNMR was used for the evaluation of

amorphous forms of drugs

2.2.4 Solubility

Solubility is defined as the concentration of the dissolved

solid in a solvent medium in a saturated solution in

equilibrium with the solid at a defined temperature and

pressure (Martin et al., 1983) The time dependence of this

solubilization process is often measured, which is referred

Chemical Society Copyright© 2003

to as dissolution testing (Grant and Higuchi, 1990) (see

Chapter 4 for details) Various dissolution testing methods

of dosage forms have been reported in the USP Some ofthe factors affecting solubility include the physical form

of the solid, the nature and composition of the solvent,temperature, and pressure (The United States Pharmacopeiaand National Formulary, 2002) Solubility is the topic

of discussion in Chapter 3, and therefore has not beendiscussed in detail here

PARTICULATE LEVEL PROPERTIES 11

However, it is important to recognize the influence of the

physicochemical characteristics on the solubilization and

dissolution rate of API from a dosage form At a given

temperature, the metastable phases such as the amorphous

form or metastable polymorphic form will theoretically

have a higher dissolution rate than the corresponding

stable form attributed to the free energy differences (Grant

and Higuchi, 1990) The anhydrate form of theophylline

has an apparent solubility twice that of its hydrate in

aqueous media due to its higher thermodynamic activity

However, at equilibrium, its solubility was determined

to be the same as that of the hydrate due to its

conversion to the hydrate form (Shefter and Higuchi, 1963)

Phadnis and Suryanarayanan (1997) have discussed the

impact of incorporating the metastable form of anhydrous

theophylline in tablets Owing to recrystallization of the

stable form on storage, the dissolution of these tablets failed

the USP dissolution test In a literature review by Pudipeddi

and Serajuddin (2005), solubility ratios of polymorphs

and anhydrates/hydrates of a number of APIs have been

reported Typically, it was determined that the solubility

ratio of polymorphs was less than 2 Some examples, where

the solubility differences have resulted in differences in

bioavailability, have been discussed in Section 2.1

2.3 PARTICULATE LEVEL PROPERTIES

These are properties that are associated with individual solid

particles that can be characterized by evaluating a small

amount of powder

2.3.1 Particle Morphology

A crystal is bounded by a number of planar faces and

the arrangement of these faces is termed habit (Brittain,

1995) Particulates can be classified into the following

based on their different crystal habits (The United States

Pharmacopeia and National Formulary, 2002):

1 acicular—needle-like particles, width and thickness

are similar;

2 columnar—rod-like particles, larger and thicker than

acicular particles;

3 flake —thin flat particles;

4 plate —flat particles with similar length and width,

thicker than flakes;

5 lath—thin blade-like particles;

6 equant—particles of similar length, width, and

thick-ness

These different particle morphologies can impact the

flow of materials, dissolution characteristics, and

compres-sion properties An evaluation of various habits of celecoxib

showed that the lath-shaped crystals had better flow and

compressibility characteristics (Banga et al., 2007)

Swami-nathan and Kildsig (2000) demonstrated that the surfaceroughness of particle carriers had a significant effect on thestability of powder mixtures Mixtures containing highlytextured surfaces segregated to a lesser extent than thosewith a smoother texture probably because of a higher num-ber of sites of adhesion In the case of dipyridamol [crystalmodification of dipyridamol using different solvents andcrystallization conditions (Adhiyaman and Basu, 2006)],crystals with different morphology were produced by crys-tallizing from various solvents These crystals exhibiteddifferent dissolution rates Similar observations have beenreported for phenytoin where the dissolution rate increasedbecause of inclusion of dopants improving wettability by ahabit-related increase in the polar crystal face areas (Chow

et al., 1995).

Particle morphology can be evaluated by optical andelectron microscopic techniques Widely used opticalmicroscopes include the light and polarizing microscopes.The light microscope simply consists of a magnifying lenswith a stage to mount the sample A polarizing microscope

is a light microscope with two polarizers The technique

is based on the effect the analyte has on the transmittedlight For further details, the reader is encouraged to review

the work of Kuhnert-Brandst¨atter (1971), McCrone et al.

(1978), and Brittain (1995)

Scanning electron microscopy is widely used for uating particle morphology of pharmaceuticals It has anelectron gun generating an electron beam, a column withlenses for beam focusing, the sample chamber, and detector.When the electron beam interacts with an inner electron ofthe sample it is ejected with a discrete amount of energy

eval-This is referred to as the secondary electron These

sec-ondary electrons escaping from the surface of the sampleare attracted to the detector, accelerated, and visible light isproduced The emitted light is detected by a photomultipliertube (Brittain, 1995)

2.3.2 Particle Size Distribution

Particle size distribution of the API and the excipientsplays an important role during the process of mixing orblending The magnitude of the gravitational and inertialforces is largely determined by the particle size of thecomponents being blended Most powders having meanparticle size below 100 μm are not free flowing due

to high interparticulate forces (Lachman et al., 1986).

Owing to these forces, blending is more uniform when thecomponents have similar particle sizes Some techniquesthat can be used for the determination of particle sizedistribution include sieve analysis, microscopy, and lightscattering techniques The USP has described the methodfor powders where at least 80% of the particles are larger

than 75 μm Sieves are stacked on top of one another in

ascending degrees of coarseness, and the test powder is

placed on the top sieve The nest of sieves is subjected

to a standardized period of agitation, and the weight of

the material retained on each sieve is determined, which

provides the weight percentage of powder in each sieve

size range The size parameter involved in determining

particle size distribution by analytical sieving is the length

of the side of the minimum square aperture through which

the particle passes (The United States Pharmacopeia and

National Formulary, 2002)

Particle size of the powders can influence several

properties of formulations such as content uniformity

where it was observed that the narrower the particle

size distribution and smaller the particle size, the better

is the content uniformity (Zhang and Johnson, 1997)

Rohrs et al (2006) developed a method for determining

the particle size distribution width and the dose required

to conform with the content uniformity requirements of

the USP Literature reports also discuss the influence of

particle size on powder flow (Podczeck and Newton, 1999;

Kachrimanis et al., 2005) and granulation and tabletting

properties (Sun and Grant, 2001; Herting and Kleinebudde,

2007) Generally, powders with smaller particle sizes yield

tablets with a higher tensile strength In the study by

Sun and Grant (2001), different size fractions of l-lysine

monohydrochloride dihydrate were compressed Crystals

with smaller particle size resulted in compacts with higher

tensile strength at a given pressure attributable to the larger

number of contact points with the smaller sized particles

Dissolution rate is inversely proportional to the particle size,

that is, the smaller the particle size of the API, the higher

is the dissolution rate (Florence and Salole, 1976; Elamin

et al., 1994).

In a recent publication, Shekunov et al (2006) have

reviewed the various methods of measuring particle size

distributions Laser diffraction is being widely used in the

pharmaceutical industry due to its robustness, short

mea-surement time, high precision, and flexibility Generally,

such instruments use a standard He –Ne laser light source

and the diffracted light is detected by a position-sensitive

detector The measurement is strongly influenced by the

shape of the particles Some other techniques used for the

measurement of particle size distribution include dynamic

light scattering (used for emulsions and colloids), the

coul-ter councoul-ter (used for nonagglomerated systems), and the

Anderson Cascade Impactor (for inhalations)

2.3.3 X-ray Powder Diffractometry (XRPD)

X-ray powder diffractometry (XRPD) is an excellent

tech-nique for the characterization of crystalline phases Every

crystalline solid phase has a unique X-ray pattern which can

form the basis for its identification In a powder mixture,

each crystalline phase produces its pattern independently ofthe other constituents in the mixture Thus, XRPD enablesnot only the identification of one API in the presence ofexcipients, but also the simultaneous identification of morethan one API in formulations (Brittain, 1995)

An X-ray diffractometer exposes the sample to

electro-magnetic radiation lying between the ultraviolet and γ -rays

in the electromagnetic spectrum When the X-rays are dent on the crystalline solids, diffraction occurs, that is,the X-rays are scattered in all directions A peak in theX-ray pattern is observed when these scattered beams are

inci-in phase and reinci-inforce each other, and this is definci-ined byBragg’s law Thus, when a monochromatic beam of wave-

length λ is incident on a sample at an angle θ diffraction

occurs, if

nλ = 2d sin θ

d is the distance between the planes in the crystal (angstrom

units) and n is the order of reflection The USP also

provides an introduction to X-ray diffractometry (Brittain,1995; The United States Pharmacopeia and NationalFormulary, 2002)

XRPD can be used to identify different crystalline formssuch as hydrates and polymorphs Various crystalline solidforms of fluprednisolone have been characterized by usingXRPD In this study, XRPD was used for the identification

of (i) anhydrous polymorphic forms; (ii) solvates; and (iii)anhydrous form existing in both crystalline and amorphous

states (Haleblian et al., 1971) Phadnis and Suryanarayanan

(1997) have evaluated the phase transformations occurringduring processing and storage of anhydrous theophyllinetablets and its implication of tablet dissolution using XRPD

40

Figure 2.2 XRD patterns of (a) stable anhydrous theophylline I,(b) stable theophylline monohydrate II, and (c) metastable anhy-

drous theophylline I* (Phadnis et al., 1997) Source: Reprinted

with permission from Wiley-Liss, Inc, a subsidiary of John Wileyand Sons, Inc Copyright© 1997

PARTICULATE LEVEL PROPERTIES 13

Quantitative analysis of various components in a

formulation can be carried out using XRPD When

appro-priate absorption corrections are made, the peak intensities

of each component are proportional to the weight fraction

of that component in the mixture This method was used

for determining the amounts of anhydrous carbamazepine

and carbamazepine dihydrate in a mixture (Suryanarayanan,

1989) Low levels of contamination of α-carbamazepine

(<5% w/w) in β-carbamazepine could also be detected and

quantified (Phadnis et al., 1997) XRPD can also be used

for the quantification of crystalline material in the

pres-ence of the amorphous form as was reported by Surana and

Suryanarayanan (2000) Crystalline sucrose was detected

and quantified in a mixture with amorphous sucrose up to

levels of 1–5% w/w of the crystalline form

XRPD can also be carried out under a controlled

envi-ronment enabling the monitoring of phase transformations

as they occur The temperature (variable temperature PXRD

(powder X-ray diffraction)) and the water vapor pressure

of the chamber can be changed to evaluate phase

trans-formations Solute crystallization in a ternary system

dur-ing all stages of freeze-drydur-ing was monitored by X-ray

while simulating the process in situ in the chamber of

the diffractometer (Pyne et al., 2003) The influence of

both temperature and water vapor pressure on the

dehy-dration of carbamazepine dihydrate was evaluated by using

variable temperature XRPD While X-ray amorphous

car-bamazepine was formed at water vapor pressures of ≤5.1

Torr, crystalline anhydrous γ -carbamazepine was formed

at≥12 Torr Thus, XRPD in these controlled environments

helped elucidate the anhydrous phases obtained under

dif-ferent dehydration conditions (Han et al., 1998).

2.3.4 Thermal Methods of Analysis

Thermal analysis is a group of techniques in which

a property of the sample is measured against time or

temperature while the temperature of the sample, in a

specified environment, is changed This temperature change

may be linear, or the sample property may be measured

when the temperature is held constant These techniquesare very useful in preformulation studies where the APIproperties such as polymorphism, state of hydration, degree

of crystallinity, purity, and degradation characteristics can

be measured (Brittain, 1995)

2.3.5 Thermogravimetric Analysis (TGA)

It is a measure of the thermally induced weight loss

of a material as a function of applied temperature It

is a useful method for the quantitative analysis of totalvolatile content of a solid Therefore, it can be used tocharacterize desolvation and degradation processes In arecent study, the dehydration of calcium benzoate hydrateswas evaluated using thermogravimetric analysis (TGA).The weight loss observed during TGA corresponded tothe stoichiometric water content of the hydrates (Fig 2.3;Terakita and Byrn, 2006) The influence of grinding onthe loss of acetonitrile from quinapril hydrochloride wascharacterized using TGA (Han and Suryanarayanan, 1997;

Guo et al., 2000) It is a complementary technique to

differential scanning calorimetry (DSC) and differentialthermal analysis (DTA)

2.3.6 Differential Thermal Analysis (DTA)

The difference in temperature between a sample and

a reference as a function of temperature is monitored.Therefore, if the sample undergoes an endothermic reaction,its temperature will lag behind that of the reference.However, in the case of an exothermic reaction itstemperature will exceed that of the reference DTA has beenmost commonly used to determine temperatures at whichthermal events occur In the above study by Terakita andByrn (2006), DTA was used as a complementary techniquefor the characterization of dehydration (Fig 2.3)

2.3.7 Differential Scanning Calorimetry (DSC)

DSC is widely used for the characterization of ceuticals, and the USP also has a general chapter on this

pharma-30

60 90 120 150 Temperature (°C)

0 2 4 6 8 10 12

–20

–15 –10 –5 0 5 10 15 20

Figure 2.3 TGA/DTA scans of (a) calcium benzoate trihydrate and (b) monohydrate using an openpan at 5◦C/min (Terakita and Byrn, 2006) Source: Reprinted with permission from Wiley-Liss,Inc., a subsidiary of John Wiley and Sons, Inc Copyright© 2006

Exotherm corresponding to degradation Glass transition with enthalpic recovery

Figure 2.4 DSC curves (heating rate of 10◦C) of TGME samples milled for 30, 60, and 300 s

(Shalaev et al., 2002) Source: Reprinted with permission from Wiley-Liss, Inc., a subsidiary of

John Wiley and Sons, Inc Copyright© 2002

technique (The United States Pharmacopeia and National

Formulary, 2002) This method of analysis is similar to

DTA There are two types of measurement—power

com-pensated and heat flux While in power-comcom-pensated DSC,

the sample and the reference are kept at the same

temper-ature (heat flow required to maintain the same tempertemper-ature

is monitored), in heat flux DSC the temperature

differen-tial between the sample and the reference is monitored

DSC can be used to monitor thermal events, including

melting point, crystallization, desolvation, and glass

tran-sitions The impact of milling on the thermal properties

of tetraglycine methyl ester (TGME) was characterized by

using DSC (Fig 2.4) Change in the melting endotherm

due to milling was observed indicating creation of a

dis-order An endotherm associated with enthalpic recovery at

the glass transition temperature was also observed (Shalaev

et al., 2002) Thus, the study exhibits the utility of the

tech-nique for characterizing several phase transformations, that

is, melting, glass transition, and degradation For further

understanding of the technique, the reader is referred to the

review by Clas et al (1993).

2.3.8 Modulated DSC

In this technique, a controlled temperature modulation is

overlaid on the conventional linear heating or cooling

rate to produce a continuously changing nonlinear sample

temperature This helps in separating out overlapping

thermal events such as enthalpic recovery and glass

transition (Royall et al., 1998).

2.3.9 Pressure DSC

The sample is subjected to different pressures, and thus

used to separate overlapping endotherms, for example,

dehydration and vaporization of water The use of this

technique to characterize the dehydration of carbamazepinedihydrate and ampicillin trihydrate has been demonstrated

by Han and Suryanarayanan (1997, 1998), respectively

2.4 PROPERTIES ASSOCIATED WITH THE BULK LEVEL

These include all the bulk material properties includingparticle shape, size distribution, powder flow, and density.The science and technology of small particles is referred to

as micromeritics as proposed by DallaValle (1948) These

particle properties can strongly influence various processesduring manufacturing, and hence knowledge about theseproperties and control over them are important Severaltechniques are available for characterizing these properties

2.4.1 Surface Area

The surface area per unit weight or volume can influencedissolution, chemical reactivity, and bioavailability of theAPI In the case of excipients, flowability is a major factorthat is influenced by the surface area; for example, grades

of lactose and Avicel with larger particles, thus smallersurface area have better flow properties (DallaValle, 1948;

Brittain et al., 1991) The surface area can be calculated

from the particle size distribution described earlier Someother techniques available for its direct determination aredescribed below

Adsorption Method

For the determination of the surface area of an adsorbent,the volume of gas (in cubic centimeters) can beplotted against the pressure of the gas adsorbed pergram of adsorbent The BET equation can be used to

PROPERTIES ASSOCIATED WITH THE BULK LEVEL 15

more accurately determine the volume of nitrogen gas

V m in cubic centimeters that 1 g of the powder can

adsorb when a monolayer is complete (Martin et al.,

1983)

Air Permeability Method

The resistance to the flow of air through a plug of

compacted powder is proportional to the surface

area of the powder Therefore, the permeability

across a given pressure drop in the plug is inversely

proportional to the specific surface area The plug

of powder can be regarded as a series of capillaries

and the internal surface depends on the particle

surface area The flow rate depends on the surface

area of the particles, the degree to which the

particles are compressed, and the irregularity of

the capillaries The Kozeny–Carman equation can

be used to determine the surface area using this

A= the cross-sectional area of the plug,

K = a constant,

ε= the porosity,

V = volume of air flowing through the

capillary of length l in t seconds, and

η = the viscosity of air (Martin et al., 1983).

2.4.2 Porosity

The porosity of a solid is defined as the ratio of the

void volume to the bulk volume (total sample volume)

and is frequently represented as a percentage Porosity

can influence the dissolution of both powders and tablets

For instance, an insoluble drug may dissolve more or less

rapidly in aqueous medium depending on their adsorption

of moisture or air (Martin et al., 1983) Properties such

as powder flow (Ohta et al., 2003) and dissolution rate of

tablets are also influenced by its pore structure and porosity

(Otsuka et al., 2007) Pore radii and volume measurements

can be carried out by using either gas adsorption or mercury

porosimetry (Martin et al., 1983) For mercury porosimetry,

the sample is placed in a holder with a tapered calibrated

stem The sample holder and stem are then filled with

mercury and pressure is applied to force the mercury into

the pores The amount of mercury that penetrates the sample

can be determined from the decrease in volume in the

calibrated stem, which is indicative of sample porosity

(Martin et al., 1983).

2.4.3 Density

Density is defined as the weight per unit volume.Pharmaceutical processes, including material transfer,blending, and flow (for example, through a hopper or afeeder), can be significantly influenced by the density of thecomponents Reports in the literature discuss the influence

of the difference in densities on segregation, and thus

con-tent uniformity issues during manufacturing (Rippie et al.,

1964; Venables and Wells, 2001)

Three types of densities can be defined:

is calculated from the weight of the liquid that thesample displaces In flotation, a few crystals of thepowder are suspended in the liquid whose density isclose to that of the material and the temperature isvaried till the crystals float It is at this temperature,that the density of the powder and the liquid arethe same and thus the density of the powder can

be determined All these techniques are discussed indetail by Duncan-Hewitt and Grant (1986)

Granule or Particle Density

It is the weight of the particles per unit volume asdetermined by mercury intrusion (considering poreslarger than 10 μm) It is determined by a methodsimilar to liquid displacement Mercury is used as theliquid since it can penetrate the intraparticle space but

not the internal pores of the particles (Martin et al.,

1983)

Bulk Density

The volume occupied by a powder is referred to as

the bulk volume Density determined from the bulk

volume and weight of the powder is referred to

as bulk density The bulk density depends on the

size, shape, porosity, and cohesiveness of powders

For smaller particles (<10 μm), flow may be

restricted because the cohesive forces are equal to

the gravitational forces Flat particles such as needles

and flakes tend to pack loosely to give powders

with high porosity Poor flow may also arise due to

high moisture content and surface roughness (which

increases friction and cohesiveness) One of the

most widely used methods to determine bulk density

is to pour the powder into a graduated cylinder

and measure the bulk volume and the weight of

the powder Tap density is also often measured by

subjecting the powder in the graduated cylinder to a

number of measured taps, for example, 500 or 1000

Another variation of this measurement involves the

tapping of the cylinder for specified number of taps,

until a constant volume is obtained (Martin et al.,

1983)

These powder characteristics such as the density,

particle size distribution, and shape can significantly

influence the flow of powders or granules, as discussed

so far Pharmaceutical processability such as blending,

encapsulation, and compression depend on the flowability

of the granules Several methods can be used to measure

the flowability of powders

2.4.4 Angles of Repose

This is the angle formed between cone of a powder and

the horizontal plane Powders with lower angles of repose

exhibit better flow The rougher and more irregular the

surface of the particles, the higher will be the angle of

repose Particles with more spherical shape have a lower

angle of repose (Martin et al., 1983).

2.4.5 Compressibility or Carr’s Index

This can be calculated using the following equation:

v = the tapped volume (after being subjected to a

specified number of taps) and

v0 = the bulk volume (Lachman et al., 1986).

Generally, values lower than 15% indicate good flow,

whereas values above 25% indicate poor flowability

Several examples exist in literature that illustrate the

utility of Carr’s Index to evaluate the flow characteristics

(Podczeck and Newton, 1999; Nagel and Peck, 2003)

2.4.6 Water Sorption

Sorption of water by pharmaceutical solids can have

an impact on both their chemical and physical stability.Water sorption/desorption isotherms can be generatedeither gravimetrically or volumetrically For gravimetricmeasurements, a specific weight of the material is placed on

a balance in a chamber or environment whose temperatureand relative humidity are controlled When the solid doesnot sorb any more moisture (i.e., it has a constant weight),the relative humidity and/or the temperature of the samplemay be changed Thus, the moisture sorption at eachhumidity condition can be monitored This is referred

to as the continuous measurement of moisture sorption.

Figure 2.5 presents the continuous water vapor sorptionprofiles of amorphous trehalose prepared by differentmethods (freeze-dried, spray dried, melt quenched, and

dehydrated) (Surana et al., 2004) Alternatively, different

samples of the solid can be placed in chambers wherethe humidity is controlled by saturated salt solutions Thesamples are removed periodically and weighed, until thesample is at equilibrium (i.e., its weight does not change

between weighings) This is referred to as the discontinuous

method (Brittain, 1995).

Solids can come in contact with water due to contactwith hygroscopic excipients, either during processing (such

as wet granulation) or during storage (from moisture

in the head space) At low relative humidities, whilecrystalline solids absorb moisture on the surface, watervapor is absorbed into the bulk of amorphous material

As the humidity increases, multilayer sorption may occurand at a characteristic RH the solid may dissolve in thesorbed film As discussed earlier, the amorphous form of

0 0.0

2.0 4.0

Dehydrated I

Dehydrated II

Spray-dried 8.0

RH (%)

Figure 2.5 Water vapor sorption profiles of amorphous trehalose

prepared by different methods (Surana et al., 2004) Source:

Reprinted with permission from SpringerLink Copyright© 2004

REFERENCES 17

solids tend to sorb more moisture than their crystalline

counterparts Certain pharmaceutical processes, including

milling, compaction, and spray drying, may induce a change

in the physical form of the API creating a local disorder

(Vadas et al., 1991) These high-energy spots may sorb

moisture to a greater extent than the crystalline state These

regions can undergo considerable change and affect the bulk

properties of the material

While this chapter provides a brief introduction to some

of the widely used techniques for the physicochemical

characterization of pharmaceuticals, references are provided

for a deeper understanding Several examples are provided

to understand the implication of these properties during

manufacturing It should be emphasized that an early

systematic characterization of material properties helps in

avoiding issues during formulation development

ACKNOWLEDGMENT

The author would like to thank Dr Raj Suryanarayanan,

Dr Sophie-Dorothee Clas, Dr Hubert Dumont, Dr Raghu

Cavatur, Ms Cecilia Madamba, and Ms Mary-Lynn Gaal

for their contributions to this chapter

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