Medical Biochemistry PDF

(with Clinical Concepts & Case Studies) Dr. U. Satyanarayana Dr. U. Chakrapani Co-published with SECTION 1 (with Clinical Concepts & Case Studies) Dr. U. Satyanarayana...

(with Clinical Concepts & Case Studies) Dr. U. Satyanarayana Dr. U. Chakrapani Co-published with SECTION 1 (with Clinical Concepts & Case Studies) Dr. U. Satyanarayana M.Sc., Ph.D., F.I.C., F.A.C.B. Professor of Biochemistry & Director (Research) Dr. Pinnamaneni Siddhartha Institute of Medical Sciences (Dr. NTR University of Health Sciences) Chinaoutpalli, Gannavaram (Mdl) Krishna (Dist), A.P., India Dr. U. Chakrapani M.B.B.S., M.S., D.N.B. Co-published with Since 1960 ELSEVIER A division of Reed Elsevier India Pvt. Ltd. Books & Allied Pvt. Ltd.  Cjpdifnjtusz-!5f Satyanarayana and Chakrapani ELSEVIER A division of Reed Elsevier India Private Limited Mosby, Saunders, Churchill Livingstone, Butterworth-Heinemann and Hanley & Belfus are the Health Science imprints of Elsevier. © 2013 Dr. U. Satyanarayana First Published: March 1999 Revised Reprint: August 2000 Second Revised Edition: June 2002 Revised Reprint: 2004, 2005 Third Revised Edition (multicolour): 2006 Revised Reprint: 2007, 2010 Fourth Revised Edition: 2013 All rights are reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, or otherwise without the prior permission of the publishers. ISBN: 978-81-312-3601-7 Medical knowledge is constantly changing. As new information becomes available, changes in treatment, procedures, equipment and the use of drugs become necessary. The author, editors, contributors and the publisher have, as far as it is possible, taken care to ensure that the information given in this text is accurate and up-to-date. However, readers are strongly advised to confirm that the information, especially with regard to drug dose/usage, complies with current legislation and standards of practice. Please consult full prescribing information before issuing prescriptions for any product mentioned in this publication. This edition of Biochemistry, 4e by Dr. U. Satyanarayana and Dr. U. Chakrapani is co-published by an arrangement with Elsevier, a division of Reed Elsevier India Private Limited and Books and Allied (P) Ltd. ELSEVIER A division of Reed Elsevier India Private Limited. Registered Office: 305, Rohit House, 3 Tolstoy Marg, New Delhi-110 001. Corporate Office: 14th Floor, Building No. 10B, DLF Cyber City, Phase II, Gurgaon–122 002, Haryana, India. BOOKS AND ALLIED (P) Ltd. Registered Office: 8/1 Chintamoni Das Lane, Kolkata 700009. Corporate Office: No. 1-E(1) ‘Shubham Plaza’ (1st Floor), 83/1, Beliaghata Main Road, Kolkata 700 010, West Bengal, India. Cover Design Depicts the universal energy currency of the living world—ATP, predominantly synthesized by the mitochondria of the cell (the functional unit of life), in comparison with the international currencies—$, £, €, `, ¥. Printed and bound at..... Copyright.indd i 6/7/2013 4:31:26 PM  Preface to the Fourth Edition This book ‘Biochemistry’ has undoubtedly become one of the most preferred text books (in India and many other countries) by the students as well as teachers in medical, biological and other allied sciences. It is certainly a book of choice and a true companion to all learning biochemistry, hence appropriately regarded by many as ‘Bible of Biochemistry’. This book has undergone three editions, several reprints, and revised reprints in a span of 13 years. The advances in biochemistry are evergrowing due to exponential growth of the subject. Further, the critical comments, frank opinions and constructive suggestions by teachers and students need to be seriously considered. All this necessitates frequent revision of the book. In this fourth edition, a thorough revision and update of each chapter with latest advances has been done. The main emphasis of this edition is an improved orientation and treatment of human biochemistry in health and disease. A wide variety of case studies with relevant biochemical profiles (along with diagnosis and discussion) are newly added as an appendix. In addition, several newer aspects of biochemistry are covered in this edition, some of them are listed below. l Triacylgylcerol/fatty acid cycle l ω-fatty acid l Metabolic syndrome l Soluble and insoluble fiber l Glucose toxicity l Trans fatty acids l Estimated average glucose l Nutrigenomics l Peptide nucleic acids l Detailed information on antivitamins l Pseudogenes l Dental caries l Recombinant ribozymes l Amino acids as neurotransmitters l Epigenetic regulation of gene expression l Disorders of membrane transport l Metagenomics l Diagnostic importance of various body fluids and tissues l Therapeutic diets l Enzyme patterns in diseases l Atkins diet l Cystatin C l Dietary antioxidants l Pleural fluid l High fructose corn syrups l High sensitive CRP It is a fact that I represent a selected group of individuals authoring books, having some time at disposal, besides hard work, determination and dedication. I consider myself as an eternal reader and a regular student of biochemistry. However, it is beyond my capability to keep track of the evergrowing advances in biochemistry due to exponential growth of the subject. And, this makes me nervous whenever I think of revising the book. I honestly and frankly admit that I have to depend on mature readers for subsequent editions of this book. AN INVITATION TO READERS, WELL WISHERS AND SUBJECT EXPERTS I have to admit that it is not all the time possible for me to meet the readers individually and get their feedback. I sincerely invite the readers, my well wishers and experts in biochemistry subject to feel free and write to me (Email ID: [email protected]) expressing their frank opinions, critical comments and constructive suggestions. And this will help me to further improve the book in subsequent revisions. Dr. U. SATYANARAYANA  Preface to the First Edition Biochemistry is perhaps the most fascinating subject as it deals with the chemical language of life, be it human, animal, plant or microorganism. No other science subject has as much application as biochemistry to the disciplines of medicine, health, veterinary, agriculture bioengineering and technology. This necessitates a totally different outlook for the books on biochemistry subject. There are many biochemistry textbooks on the market. Some of them are purely basic while others are applied, and there are very few books which cover both these aspects together. For this reason, the students learning biochemistry in their undergraduate courses have to depend on multiple books to acquire a sound knowledge of the subject. This book, ‘Biochemistry’ is unique with a simultaneous and equal emphasis on basic and applied aspects of biochemistry. This textbook primarily is an integration of medical and pure sciences, comprehensively written to meet the curriculum requirements of undergraduate courses in medical, dental, pharmacy, life-sciences and other categories (agriculture, veterinary, etc.) where students learn biochemistry as one of the subjects. The tendency among the students (particularly medical) is to regard biochemistry as being mostly concerned with unimportant and complicated metabolic (chemical) pathways. This book gives a new orienta- tion to the subject of biochemistry so that the students appreciate the great importance and significance of the application of biochemistry to medicine. This book is designed to develop in students a sustained interest and enthusiasm to learn and develop the concepts in biochemistry in a logical and stepwise manner. It incorporates a variety of pedagogic aids, besides colour illustrations to help the students understand the subject quickly and to the maximum. The summary and biomedical/clinical concepts are intended for a rapid absorption and assimilation of the facts and concepts in biochemistry. The self-assessment exercises will stimulate the students to think rather than merely learn the subject. In addition, these exercises (essays, short notes, fill in the blanks, multiple choice questions) set at different difficulty levels, will cater to the needs of all the categories of learners. It will not be out of place to mention here how-and when-the book was born. The entire book was written in the early morning hours (between 2 AM-6 AM; when the world around is fast asleep), during which period I carry out my intellectual activities. After a sound sleep, a fresh mind packed with creative ideas and innovative thoughts, has largely helped me to write this book. My wife pleaded with me that I should not write topics like diabetes, cancer, AIDS at home. In deference to her sentiment, I made a serious attempt to write those topics during my leisure time in the Department. But when I went through them in my serene mood of the early morning hours, I had to discard them in disappointment and rewrite them. Truly, each page of this book was conceived in darkness and born at daybreak ! This textbook is a distillation of my knowledge and teaching experience in biochemistry, acquired during the past 25 years. It contains predigested information on biochemistry for good understanding, assimilation and reproducibility. Each page is crafted with a fine eye. The ultimate purpose of this book is to equip the reader with comprehensive knowledge in biochemistry with reference to basic as well as applied aspects. Although I have made every effort to make the book error free, I am under no illusion. I welcome comments, criticism and suggestions from the faculty, students and other readers, and this will help me make improvements in the next edition. Dr. U. SATYANARAYANA [ ii ]  Acknowledgements I owe a deep debt of gratitude to my parents, the late Sri U. Venkata Subbaiah, and Smt. Vajramma, for cultivating in me the habit of early rising. The writing of this book would never have been possible without this healthy habit. I am grateful to Dr. B. S. Narasinga Rao (former Director, National Institute of Nutrition, Hyderabad) for disciplining my professional life, and to my eldest brother Dr. U. Gudaru (former Professor of Power Systems, Walchand College of Engineering, Sangli) for disciplining my personal life. My elder son, U. Chakrapani (MBBS) deserves a special place in this book. He made a significant contribution at every stage of its preparation—writing, verification, proof-reading and what not. I had the rare privilege of teaching my son as he happened to be a student of our college. And a major part of this book was written while he was learning biochemistry. Thus, he was the first person to learn the subject of biochemistry from my handwritten manuscript. The student-teacher relation (rather than the father-son) has helped me in receiving constant feedback from him and restructure the book in a way an undergraduate student would expect a biochemistry textbook to be. Next, I thank Dr. G. Pitcheswara Rao (former Professor of Anatomy, SMC, Vijayawada) for his constructive criticism and advice, and Dr. B. Sivakumar (Director, National Institute of Nutrition, Hyderabad) for his helpful suggestions on the microfigures. Last but not least, I thank my wife Krishna Kumari and my younger son, Amrutpani, without whose cooperation and encouragement this book could never have been written. The manuscript was carefully nurtured like a new born baby and the book has now become a full-pledged member of our family. ACKNOWLEDGEMENTS TO THE FOURTH EDITION I am grateful to a large number of faculty members, students, friends and pen friends who directly or indirectly helped me to revise and improve the content and quality of the book. I have individually and personally thanked all of them (who number a few hundreds!). I once again express my gratitude to them. I thank Dr (Mrs) U.B. Vijaya Lakshmi, MD, Associate Professor of Biochemistry at our college who participated to comprehensively prepare case studies with biochemical correlations, besides improving the biomedical/ clinical aspects in some chapters. My special thanks goes to one student, and an ardent fan of my books, Mr. Y. Nagendra Sastry (Ph.D), who has been studying my books regularly for over 7-8 years. His constant feedback and suggestions have certainly contributed to improve this book. I express my gratitude to Mr. M.S.T. Jagan Mohan (my former colleague), who has helped me with his frequent interactions to revise the book, and make it more student-friendly. I express my sincere thanks to Mr Arunabha Sen, Director, Books & Allied (P) Ltd, Kolkata for his whole hearted support and constant encouragement in revising the book, and taking all pains to bring it out to my satisfaction. I thank Mr. Shyamal Bhattacharya for his excellent page making and graphics-work in the book. I am grateful to Mr. Abhijit Ghosal for his help in the cover design. I thank my wife, Krishna Kumari, my younger son Amrut Pani and my daughter-in law Oohasri for their constant support and encouragement. My special thanks to my grand daughter Maahe (2 years) whose ever smiling face, sweet words and deeds infuse energy into my academic activities. I am grateful to Uppala Author-Publisher interlinks, Vijayawada for sponsoring and supporting me to bring out this edition. Dr. U. SATYANARAYANA [ iii ]  Scope of Biochemistry The term Biochemistry was introduced by Carl Neuberg in 1903. Biochemistry broadly deals with the chemistry of life and living processes. There is no exaggeration in the statement, ‘The scope of biochemistry is as vast as life itself !’ Every aspect of life-birth, growth, reproduction, aging and death, involves biochemistry. For that matter, every movement of life is packed with hundreds of biochemical reactions. Biochemistry is the most rapidly developing and most innovative subject in medicine. This becomes evident from the fact that over the years, the major share of Nobel Prizes earmarked for Medicine and Physiology has gone to researchers engaged in biochemistry. The discipline of biochemistry serves as a torch light to trace the intricate complexicities of biology, besides unravelling the chemical mysteries of life. Biochemical research has amply demonstrated that all living things are closely related at the molecular level. Thus biochemistry is the subject of unity in the diversified living kingdom. Advances in biochemistry have tremendous impact on human welfare, and have largely benefited mankind and their living styles. These include the application of biochemistry in the laboratory for the diagnosis of diseases, the products (insulin, interferon, growth hormone etc.) obtained from genetic engineering, and the possible use of gene therapy in the near future. Organization of the Book This textbook, comprising 43 chapters, is organized into seven sections in the heirarchical order of learning biochemistry. l Section I deals with the chemical constituents of life—carbohydrates, lipids, proteins and amino acids, nucleic acids and enzymes. l Section II physiological chemistry includes digestion and absorption, plasma proteins, hemoglobin and prophyrins, and biological oxidation. l Section III incorporates all the metabolisms (carbohydrates, lipids, amino acids, nucleotides, minerals) l Section IV covers hormones, organ function tests, water, electrolyte and acid-base balance, tissue proteins and body fluids, and nutrition. l Section V is exclusively devoted to molecular biology and biotechnology (DNA-replication, recombination, and repair, transcription and translation, regulation of gene expression, recombinant DNA and biotechnology) l Section VI gives relevant information on current topics such as human genome project, gene therapy, bioinformatics, prostaglandins, diabetes, cancer, AIDS etc. l Section VII deals with the basic aspects for learning and understanding biochemistry (bioorganic chemistry, biophysical chemistry, tools of biochemistry, genetics, immunology). Each chapter in this book is carefully crafted with colour illustrations, headings and subheadings to facilitate quick understanding. The important applications of biochemistry to human health and disease are put together as biomedical/clinical concepts. Icons are used at appropriate places to serve as ‘landmarks’. The origins of biochemical words, confusables in biochemistry, practical biochemistry and clinical biochemistry laboratory, case studies with biochemical correlations, given in the appendix are novel features. The book is so organized as to equip the readers with a comprehensive knowledge of biochemistry. [ iv ]  Contents SECTION ONE SECTION FIVE Chemical Constituents of Life Molecular Biology and Biotechnology 1 ➤ Biomolecules and the cell 3 24 ➤ DNA-replication, recombination and repair 523 25 ➤ Transcription and translation 542 2 ➤ Carbohydrates 9 26 ➤ Regulation of gene expression 566 3 ➤ Lipids 28 27 ➤ Recombinant DNA and biotechnology 578 4 ➤ Proteins and amino acids 43 5 ➤ Nucleic acids and nucleotides 69 SECTION SIX 6 ➤ Enzymes 85 Current Topics 7 ➤ Vitamins 116 28 ➤ Human genome project 619 29 ➤ Gene therapy 625 SECTION TWO 30 ➤ Bioinformatics 634 Physiological Biochemistry 31 ➤ Metabolism of xenobiotics (detoxification) 638 32 ➤ Prostaglandins and related compounds 644 8 ➤ Digestion and absorption 165 33 ➤ Biological membranes and transport 650 9 ➤ Plasma proteins 182 34 ➤ Free radicals and antioxidants 655 10 ➤ Hemoglobin and porphyrins 196 35 ➤ Environmental biochemistry 662 36 ➤ Insulin, glucose homeostasis, 11 ➤ Biological oxidation 221 and diabetes mellitus 669 37 ➤ Cancer 685 SECTION THREE 38 ➤ Acquired immunodeficiency 32 Metabolisms syndrome (AIDS) 695 12 ➤ Introduction to metabolism 241 SECTION SEVEN 33 13 ➤ Metabolism of carbohydrates 244 Basics to Learn Biochemistry 14 ➤ Metabolism of lipids 285 39 ➤ Introduction to bioorganic chemistry 703 15 ➤ Metabolism of amino acids 330 40 ➤ Overview of biophysical chemistry 708 34 16 ➤ Integration of metabolism 380 41 ➤ Tools of biochemistry 719 17 ➤ Metabolism of nucleotides 387 42 ➤ Immunology 732 35 43 ➤ Genetics 737 18 ➤ Mineral metabolism 403 36 APPENDICES SECTION FOUR Answers to Self-assessment Exercises 745 Clinical Biochemistry and Nutrition 37 I Abbreviations used in this book 751 19 ➤ Hormones 427 II Origins of important biochemical words 756 38 20 ➤ Organ function tests 453 III Common confusables in biochemistry 759 21 ➤ Water, electrolyte and IV Practical biochemistry—principles 763 acid-base balance 468 V Clinical biochemistry laboratory 769 22 ➤ Tissue proteins and body fluids 487 VI Case studies with biochemical correlations 772 23 ➤ Nutrition 502 INDEX 779 CHEMICAL CONSTITUENTS OF LIF LIFEE 1 Biomolecules and the Cell 3 2 Carbohydrates 9 3 Lipids 28 4 Proteins and Amino acids 43 5 Nucleic acids and Nucleotides 69 6 Enzymes 85 7 Vitamins 116 Section I “This page intentionally left blank"  Section 1 Chemical Constituents of Life Chapter Biomolecules and the Cell 1 The cell speaks : “I am the unit of biological activity; Organized into subcellular organelles; Assigned to each are specific duties; Thus, I truly represent life!” T he living matter is composed of mainly six elements—carbon, hydrogen, oxygen, nitrogen, phosphorus and sulfur. These elements organic compounds. It is believed that man may contain about 100,000 different types of molecules although only a few of them have together constitute about 90% of the dry weight been characterized. of the human body. Several other functionally important elements are also found in the cells. Complex biomolecules These include Ca, K, Na, Cl, Mg, Fe, Cu, Co, I, The organic compounds such as amino acids, Zn, F, Mo and Se. nucleotides and monosaccharides serve as the monomeric units or building blocks of complex Carbon—a unique element of life biomolecules—proteins, nucleic acids (DNA and RNA) and polysaccharides, respectively. The Carbon is the most predominant and versatile important biomolecules (macromolecules) with element of life. It possesses a unique property to their respective building blocks and major form infinite number of compounds. This is functions are given in Table 1.1. As regards attributed to the ability of carbon to form stable lipids, it may be noted that they are not covalent bonds and C C chains of unlimited biopolymers in a strict sense, but majority of length. It is estimated that about 90% of them contain fatty acids. compounds found in living system invariably contain carbon. Structural heirarchy of an organism The macromolecules (proteins, lipids, nucleic Chemical molecules of life acids and polysaccharides) form supramolecular Life is composed of lifeless chemical assemblies (e.g. membranes) which in turn molecules. A single cell of the bacterium, organize into organelles, cells, tissues, organs Escherichia coli contains about 6,000 different and finally the whole organism. 3 4 BIOCHEMISTRY TABLE 1.1 The major complex biomolecules of cells Biomolecule Building block Major functions (repeating unit) 1. Protein Amino acids Fundamental basis of structure and function of cell (static and dynamic functions). 2. Deoxyribonucleic acid (DNA) Deoxyribonucleotides Repository of hereditary information. 3. Ribonucleic acid (RNA) Ribonucleotides Essentially required for protein biosynthesis. 4. Polysaccharide (glycogen) Monosaccharides (glucose) Storage form of energy to meet short term demands. 5. Lipid Fatty acids, glycerol Storage form of energy to meet long term demands; structural components of membranes. Chemical composition of man Prokaryotic and eukaryotic cells The chemical composition of a normal man, The cells of the living kingdom may be weighing 65 kg, is given in Table 1.2. Water is divided into two categories the solvent of life and contributes to more than 1. Prokaryotes (Greek : pro – before; karyon – 60% of the weight. This is followed by protein nucleus) lack a well defined nucleus and possess (mostly in muscle) and lipid (mostly in adipose relatively simple structure. These include the tissue). The carbohydrate content is rather low various bacteria. which is in the form of glycogen. 2. Eukaryotes (Greek : eu – true; karyon – nucleus) possess a well defined nucleus and are more complex in their structure and function. THE CELL The higher organisms (animals and plants) are composed of eukaryotic cells. The cell is the structural and functional unit of life. It may be also regarded as the basic unit A comparison of the characteristics between of biological activity. prokaryotes and eukaryotes is listed in Table 1.3. The concept of cell originated from the contributions of Schleiden and Schwann (1838). However, it was only after 1940, the EUKARYOTIC CELL complexities of cell structure were exposed. The human body is composed of about 1014 cells. There are about 250 types of specialized TABLE 1.2 Chemical composition of a normal man cells in the human body e.g. erythrocytes, (weight 65 kg) nerve cells, muscle cells, E cells of pancreas. Constituent Percent (%) Weight (kg) An eukaryotic cell is generally 10 to 100 Pm in diameter. A diagrammatic representation Water 61.6 40 of a typical rat liver cell is depicted in Protein 17.0 11 Fig.1.1. Lipid 13.8 9 The plant cell differs from an animal cell by Carbohydrate 1.5 1 possessing a rigid cell wall (mostly composed of cellulose) and chloroplasts. The latter are the Minerals 6.1 4 sites of photosynthesis. Chapter 1 : BIOMOLECULES AND THE CELL 5 TABLE 1.3 Comparison between prokaryotic and eukaryotic cells Characteristic Prokaryotic cell Eukaryotic cell 1. Size Small (generally 1-10 Pm) Large (generally 10-100 Pm) 2. Cell membrane Cell is enveloped by a rigid cell wall Cell is enveloped by a flexible plasma membrane 3. Sub-cellular Absent Distinct organelles are found organelles (e.g. mitochondria, nucleus, lysosomes) 4. Nucleus Not well defined; DNA is found Nucleus is well defined, surrounded by a as nucleoid, histones are absent membrane; DNA is associated with histones 5. Energy metabolism Mitochondria absent, enzymes of Enzymes of energy metabolism are located energy metabolism bound to in mitochondria membrane 6. Cell division Usually fission and no mitosis Mitosis 7. Cytoplasm Organelles and cytoskeleton Contains organelles and cytoskeleton absent (a network of tubules and filaments) The cell consists of well defined subcellular Nucleus organelles, enveloped by a plasma membrane. By differential centrifugation of tissue Nucleus is the largest cellular organelle, homogenate, it is possible to isolate each surrounded by a double membrane nuclear cellular organelle in a relatively pure form envelope. The outer membrane is continuous (Refer Chapter 41). The distribution of major with the membranes of endoplasmic reticulum. enzymes and metabolic pathways in different At certain intervals, the two nuclear membranes cellular organelles is given in the chapter have nuclear pores with a diameter of about 90 on enzymes (Refer Fig.6.6). The subcellular nm. These pores permit the free passage of the organelles are briefly described in the following products synthesized in the nucleus into the pages. surrounding cytoplasm. Mitochondrion Plasma membrane Rough endoplasmic reticulum Vacuole Ribosomes Golgi apparatus Nucleus Nucleolus Smooth endoplasmic reticulum Lysosome Peroxisome Cytoskeleton Cytosol Coated pits Fig. 1.1 : Diagrammatic representation of a rat liver cell. 6 BIOCHEMISTRY Nucleus contains DNA, the repository of carbohydrates, lipids and amino acids (e.g., citric genetic information. Eukaryotic DNA is acid cycle, E-oxidation). The matrix enzymes associated with basic protein (histones) in the also participate in the synthesis of heme and ratio of 1 : 1, to form nucleosomes. An assembly urea. Mitochondria are the principal producers of nucleosomes constitutes chromatin fibres of of ATP in the aerobic cells. ATP, the energy chromosomes (Greek: chroma – colour; soma – currency, generated in mitochondria is exported body). Thus, a single human chromosome is to all parts of the cell to provide energy for the composed of about a million nucleosomes. The cellular work. number of chromosomes is a characteristic The mitochondrial matrix contains a circular feature of the species. Humans have 46 double stranded DNA (mtDNA), RNA and chromosomes, compactly packed in the nucleus. ribosomes. Thus, the mitochondria are equipped The nucleus of the eukaryotic cell contains a with an independent protein synthesizing dense body known as nucleolus. It is rich in machinery. It is estimated that about 10% of the RNA, particularly the ribosomal RNA which mitochondrial proteins are produced in the enters the cytosol through nuclear pores. mitochondria. The ground material of the nucleus is often The structure and functions of mitochondria referred to as nucleoplasm. It is rich in enzymes closely resemble prokaryotic cells. It is such as DNA polymerases and RNA polymerases. hypothesized that mitochondria have evolved Hutchinson-Gilford progeria syndrome from aerobic bacteria. Further, it is believed that (HGPS) is a rare condition of aging beginning at during evolution, the aerobic bacteria developed birth (incidence I in 5 million births). HGPS a symbiotic relationship with primordial occurs as a result of distortion of nuclear anaerobic eukaryotic cells that ultimately led to envelope due to accumulation of abnormal the arrival of aerobic eukaryotes. protein namely lamina A. Endoplasmic reticulum Mitochondria The network of membrane enclosed spaces The mitochondria (Greek: mitos – thread; that extends throughout the cytoplasm chondros – granule) are the centres for the constitutes endoplasmic reticulum (ER). Some of cellular respiration and energy metabolism. They these thread-like structures extend from the are regarded as the power houses of the cell nuclear pores to the plasma membrane. with variable size and shape. Mitochondria are rod-like or filamentous bodies, usually with A large portion of the ER is studded with dimensions of 1.0 u 3 Pm. About 2,000 ribosomes to give a granular appearance which mitochondria, occupying about 1/5th of the total is referred to as rough endoplasmic reticulum. cell volume, are present in a typical cell. Ribosomes are the factories of protein biosynthesis. During the process of cell The mitochondria are composed of a double fractionation, rough ER is disrupted to form small membrane system (Refer Fig.11.5). The outer vesicles known as microsomes. It may be noted membrane is smooth and completely envelops that microsomes as such do not occur in the cell. the organelle. The inner membrane is folded to form cristae (Latin – crests) which occupy a The smooth endoplasmic reticulum does not larger surface area. The internal chamber of contain ribosomes. It is involved in the synthesis mitochondria is referred to as matrix or mitosol. of lipids (triacylglycerols, phospholipids, sterols) The components of electron transport chain and metabolism of drugs, besides supplying Ca2+ and oxidative phosphorylation (flavoprotein, for the cellular functions. cytochromes b, c1, c, a and a3 and coupling Golgi apparatus factors) are buried in the inner mitochondrial membrane. The matrix contains several enzymes Eukaryotic cells contain a unique cluster of concerned with the energy metabolism of membrane vesicles known as dictyosomes Chapter 1 : BIOMOLECULES AND THE CELL 7 which, in turn, constitute Golgi apparatus (or by diffusion, for reutilization by the cell. Golgi complex). The newly synthesized proteins Sometimes, however, certain residual products, are handed over to the Golgi apparatus which rich in lipids and proteins, collectively known as catalyse the addition of carbohydrates, lipids or lipofuscin accumulate in the cell. Lipofuscin is sulfate moieties to the proteins. These chemical the age pigment or wear and tear pigment which modifications are necessary for the transport of has been implicated in ageing process. As the cell proteins across the plasma membrane. dies, the lysosomes rupture and release hydrolytic enzymes that results in post-morteum autolysis. Certain proteins and enzymes are enclosed in membrane vesicles of Golgi apparatus and The digestive enzymes of cellular compounds secreted from the cell after the appropriate are confined to the lysosomes in the best interest signals. The digestive enzymes of pancreas are of the cell. Escape of these enzymes into cytosol produced in this fashion. will destroy the functional macromolecules of the cell and result in many complications. The Golgi apparatus are also involved in the occurrence of several diseases (e.g. arthritis, membrane synthesis, particularly for the muscle diseases, allergic disorders) has been partly formation of intracellular organelles (e.g. attributed to the release of lysosomal enzymes. peroxisomes, lysosomes). Inclusion cell (I-cell) desease is a rare Lysosomes condition due to the absence of certain hydrolases in lysosomes. However, these enzyme are Lysosomes are spherical vesicles enveloped syntherized and found in the circulation. I-cell by a single membrane. Lysosomes are regarded disease is due to a defect in protein targetting, as as the digestive tract of the cell, since they are the enzymes cannot reach lysosomes. actively involved in digestion of cellular substances—namely proteins, lipids, carbo- Peroxisomes hydrates and nucleic acids. Lysosomal enzymes Peroxisomes, also known as microbodies, are are categorized as hydrolases. These include single membrane cellular organelles. They are the enzymes (with substrate in brackets)— spherical or oval in shape and contain the D-glucosidase (glycogen), cathepsins (proteins), enzyme catalase. Catalase protects the cell from lipases (lipids), ribonucleases (RNA). the toxic effects of H2O2 by converting it to H2O The lysosomal enzymes are responsible for and O2. Peroxisomes are also involved in the maintaining the cellular compounds in a dynamic oxidation of long chain fatty acids (> C18), and state, by their degradation and recycling. The synthesis of plasmalogens and glycolipids. Plants degraded products leave the lysosomes, usually contain glyoxysomes, a specialized type of + A living cell is a true representative of life with its own organization and specialized functions. + Accumulation of lipofuscin, a pigment rich in lipids and proteins, in the cell has been implicated in ageing process. + Leakage of lysosomal enzymes into the cell degrades several functional macromolecules and this may lead to certain disorders (e.g. arthritis). + Zellweger syndrome is a rare disease characterized by the absence of functional peroxisomes. 8 BIOCHEMISTRY peroxisomes, which are involved in the three types – microtubules, actin filaments and glyoxylate pathway. intermediate filaments. The filaments which are polymers of proteins are responsible for the Peroxisome biogenesis disorders (PBDs), are structure, shape and organization of the cell. a group of rare diseases involving the enzyme activities of peroxisomes. The biochemical INTEGRATION OF abnormalities associated with PBDs include CELLULAR FUNCTIONS increased levels of very long chain fatty acids (C24 and C26) and decreased concentrations of The eukaryotic cells perform a wide range of plasmalogens. The most severe form of PBDs is complex reactions/functions to maintain tissues, Zellweger syndrome, a condition characterized and for the ultimate well-being of the whole by the absence of functional peroxisomes. The organism. For this purpose, the various victims of this disease may die within one year intracellular processes and biochemical reactions after birth. are tightly controlled and integrated. Division of a cell into two daughter cells is good example of Cytosol and cytoskeleton the orderly occurrence of an integrated series of cellular reactions. The cellular matrix is collectively referred to as cytosol. Cytosol is basically a compartment Apoptosis is the programmed cell death or containing several enzymes, metabolites and cell suicide. This occurs when the cell has salts in an aqueous gel like medium. More recent fulfilled its biological functions. Apoptosis may studies however, indicate that the cytoplasm be regarded as a natural cell death and it differs actually contains a complex network of protein from the cell death caused by injury due to filaments, spread throughout, that constitutes radiation, anoxia etc. Programmed cell death is cytoskeleton. The cytoplasmic filaments are of a highly regulated process. 1. Life is composed of lifeless chemical molecules. The complex biomolecules, proteins, nucleic acids (DNA and RNA), polysaccharides and lipids are formed by the monomeric units amino acids, nucleotides, monosaccharides and fatty acids, respectively. 2. The cell is the structural and functional unit of life. The eukaryotic cell consists of well defined subcellular organelles, enveloped in a plasma membrane. 3. The nucleus contains DNA, the repository of genetic information. DNA, in association with proteins (histones), forms nucleosomes which, in turn, make up the chromosomes. 4. The mitochondria are the centres for energy metabolism. They are the principal producers of ATP which is exported to all parts of the cell to provide energy for cellular work. 5. Endoplasmic reticulum (ER) is the network of membrane enclosed spaces that extends throughout the cytoplasm. ER studded with ribosomes, the factories of protein biosynthesis, is referred to as rough ER. Golgi apparatus are a cluster of membrane vesicles to which the newly synthesized proteins are handed over for further processing and export. 6. Lysosomes are the digestive bodies of the cell, actively involved in the degradation of cellular compounds. Peroxisomes contain the enzyme catalase that protects the cell from the toxic effects of H2O2. The cellular ground matrix is referred to as cytosol which, in fact, is composed of a network of protein filaments, the cytoskeleton. 7. The eukaryotic cells perform a wide range of complex functions in a well coordinated and integrated fashion. Apoptosis is the process of programmed cell death or cell suicide.  Section 1 Chemical Constituents of Life Chapter Carbohydrates 12 The carbohydrates speak : “We are polyhydroxyaldehydes or ketones; Classified into mono-, oligo- and polysaccharides; Held together by glycosidic bonds; Supply energy and serve as structural constituents.” arbohydrates are the most abundant organic 1. They are the most abundant dietary source C molecules in nature. They are primarily composed of the elements carbon, hydrogen and of energy (4 Cal/g) for all organisms. 2. Carbohydrates are precursors for many oxygen. The name carbohydrate literally means organic compounds (fats, amino acids). ‘hydrates of carbon’. Some of the carbohydrates 3. Carbohydrates (as glycoproteins and glyco- possess the empirical formula (C.H2O)n where lipids) participate in the structure of cell n d 3, satisfying that these carbohydrates are in membrane and cellular functions such as cell fact carbon hydrates. However, there are several growth, adhesion and fertilization. non-carbohydrate compounds (e.g. acetic acid, C2H4O2; lactic acid, C3H6O3) which also appear 4. They are structural components of many as hydrates of carbon. Further, some of the organisms. These include the fiber (cellulose) of genuine carbohydrates (e.g. rhamnohexose, plants, exoskeleton of some insects and the cell C6H12O5; deoxyribose, C5H10O4) do not satisfy wall of microorganisms. the general formula. Hence carbohydrates cannot 5. Carbohydrates also serve as the storage be always considered as hydrates of carbon. form of energy (glycogen) to meet the immediate Carbohydrates may be defined as energy demands of the body. polyhydroxyaldehydes or ketones or compounds CLASSIFICATION which produce them on hydrolysis. The term ‘sugar’ is applied to carbohydrates soluble in OF CARBOHYDRATES water and sweet to taste. Carbohydrates are often referred to as saccharides (Greek: sakcharon–sugar). They Functions of carbohydrates are broadly classified into three major groups— Carbohydrates participate in a wide range of monosaccharides, oligosaccharides and poly- functions saccharides. This categorization is based on the 9 10 BIOCHEMISTRY TABLE 2.1 Classification of monosaccharides with selected examples Monosaccharides (empirical formula) Aldose Ketose Trioses (C3H6O3) Glyceraldehyde Dihydroxyacetone Tetroses (C4H8O4) Erythrose Erythrulose Pentoses (C5H10O5) Ribose Ribulose Hexoses (C6H12O6) Glucose Fructose Heptoses (C7H14O7) Glucoheptose Sedoheptulose number of sugar units. Mono- and oligo- liberated on hydrolysis. Based on the number of saccharides are sweet to taste, crystalline in monosaccharide units present, the oligo- character and soluble in water, hence they are saccharides are further subdivided to commonly known as sugars. disaccharides, trisaccharides etc. Monosaccharides Polysaccharides Monosaccharides (Greek : mono-one) are the Polysaccharides (Greek: poly-many) are poly- simplest group of carbohydrates and are often mers of monosaccharide units with high mole- referred to as simple sugars. They have the cular weight (up to a million). They are usually general formula Cn(H2O)n, and they cannot be tasteless (non-sugars) and form colloids with further hydrolysed. The monosaccharides are water. The polysaccharides are of two types – divided into different categories, based on the homopolysaccharides and heteropolysaccharides. functional group and the number of carbon atoms Aldoses : When the functional group in H MONOSACCHARIDES— monosaccharides is an aldehyde C O , they STRUCTURAL ASPECTS are known as aldoses e.g. glyceraldehyde, glucose. Stereoisomerism is an important character of monosaccharides. Stereoisomers are the Ketoses : When the functional group is a keto compounds that have the same structural C O group, they are referred to as ketoses formulae but differ in their spatial configuration. e.g. dihydroxyacetone, fructose. A carbon is said to be asymmetric when it is Based on the number of carbon atoms, the attached to four different atoms or groups. The monosaccharides are regarded as trioses (3C), number of asymmetric carbon atoms (n) tetroses (4C), pentoses (5C), hexoses (6C) and determines the possible isomers of a given heptoses (7C). These terms along with functional compound which is equal to 2n. Glucose groups are used while naming monosaccharides. contains 4 asymmetric carbons, and thus has 16 For instance, glucose is an aldohexose while isomers. fructose is a ketohexose (Table 2.1). Glyceraldehyde The common monosaccharides and disaccha- —the reference carbohydrate rides of biological importance are given in the Table 2.2. Glyceraldehyde (triose) is the simplest mono- saccharide with one asymmetric carbon atom. It Oligosaccharides exists as two stereoisomers and has been chosen Oligosaccharides (Greek: oligo-few) contain as the reference carbohydrate to represent the 2-10 monosaccharide molecules which are structure of all other carbohydrates. Chapter 2 : CARBOHYDRATES 11 TABLE 2.2 Monosaccharides and disaccharides of biological importance Monosaccharides Occurrence Biochemical importance Trioses Glyceraldehyde Found in cells as phosphate Glyceraldehyde 3-phosphate is an intermediate in glycolysis Dihydroxyacetone Found in cells as phosphate Its 1-phosphate is an intermediate in glycolysis Tetroses D-Erythrose Widespread Its 4-phosphate is an intermediate in carbohydrate metabolism Pentoses D-Ribose Widespread as a constituent of For the structure of RNA and nucleotide RNA and nucleotides coenzymes (ATP, NAD+, NADP+) D-Deoxyribose As a constituent of DNA For the structure of DNA D-Ribulose Produced during metabolism It is an important metabolite in hexose monophosphate shunt D-Xylose As a constituent of glycoproteins Involved in the function of glycoproteins and gums L-Xylulose As an intermediate in uronic acid pathway Excreted in urine in essential pentosuria D-Lyxose Heart muscle As a constituent of lyxoflavin of heart muscle Hexoses D-Glucose As a constituent of polysaccharides The ‘sugar fuel’ of life; excreted in urine in (starch, glycogen, cellulose) and diabetes. Structural unit of cellulose in plants disaccharides (maltose, lactose, sucrose). Also found in fruits D-Galactose As a constituent of lactose Converted to glucose, failure leads to (milk sugar) galactosemia D-Mannose Found in plant polysaccharides For the structure of polysaccharides and animal glycoproteins D-Fructose Fruits and honey, as a constituent Its phosphates are intermediates of glycolysis of sucrose and inulin Heptoses D-Sedoheptulose Found in plants Its 7-phosphate is an intermediate in hexose monophosphate shunt, and in photosynthesis Disaccharides Occurrence Biochemical importance Sucrose As a constituent of cane sugar and Most commonly used table sugar supplying beet sugar, pineapple calories Lactose Milk sugar Exclusive carbohydrate source to breast fed infants. Lactase deficiency (lactose intolerance) leads to diarrhea and flatulence Maltose Product of starch hydrolysis, An important intermediate in the digestion of occurs in germinating seeds starch 12 BIOCHEMISTRY H C O H C O relation with glyceraldehyde. It may be noted H C OH HO C H that the D- and L-configurations of sugars are primarily based on the structure of CH2OH CH2OH glyceraldehyde, the optical activities however, D-Glyceraldehyde L-Glyceraldehyde may be different. H C O H C O Racemic mixture : If d- and l-isomers are present in equal concentration, it is known as H C OH HO C H racemic mixture or dl mixture. Racemic mixture HO C H H C OH does not exhibit any optical activity, since the H C OH HO C H dextro- and levorotatory activities cancel each H C OH HO C H other. CH2OH CH2OH In the medical practice, the term dextrose is D-Glucose L-Glucose used for glucose in solution. This is because of the dextrorotatory nature of glucose. Fig. 2.1 : D-and-L- forms of glucose compared with D- and L- glyceraldehydes (the reference carbohydrate). Configuration of D-aldoses The configuration of possible D-aldoses D- and L-isomers starting from D-glyceraldehyde is depicted in The D and L isomers are mirror images of Fig.2.2. This is a representation of Killiani- each other. The spatial orientation of H and Fischer synthesis, by increasing the chain length OH groups on the carbon atom (C5 for of an aldose, by one carbon at a time. Thus, glucose) that is adjacent to the terminal primary starting with an aldotriose (3C), aldotetroses (4C), alcohol carbon determines whether the sugar is aldopentoses (5C) and aldohexoses (6C) are D- or L-isomer. If the OH group is on the right formed. Of the 8 aldohexoses, glucose, mannose side, the sugar is of D-series, and if on the left and galactose are the most familiar. Among side, it belongs to L-series. The structures of these, D-glucose is the only aldose mono- D- and L-glucose based on the reference mono- saccharide that predominantly occurs in saccharide, D- and L-glyceraldehyde (glycerose) nature. are depicted in Fig.2.1. Configuration of D-ketoses It may be noted that the naturally occurring monosaccharides in the mammalian tissues are Starting from dihydroxyacetone (triose), there mostly of D-configuration. The enzyme machinery are five keto-sugars which are physiologically of cells is specific to metabolise D-series of important. Their structures are given in Fig.2.3. monosaccharides. Epimers Optical activity of sugars If two monosaccharides differ from each Optical activity is a characteristic feature of other in their configuration around a single compounds with asymmetric carbon atom. specific carbon (other than anomeric) atom, they When a beam of polarized light is passed are referred to as epimers to each other (Fig.2.4). through a solution of an optical isomer, it will be For instance, glucose and galactose are epimers rotated either to the right or left. The term with regard to carbon 4 (C4-epimers). That is, dextrorotatory (d+) and levorotatory (l–) are they differ in the arrangement of OH group at used to compounds that respectively rotate the C4. Glucose and mannose are epimers with plane of polarized light to the right or to the left. regard to carbon 2 (C2-epimers). An optical isomer may be designated as The interconversion of epimers (e.g. glucose D(+), D(–), L(+) and L(–) based on its structural to galactose and vice versa) is known as Chapter 2 : CARBOHYDRATES 13 CHO HCOH Aldotriose CH2OH (3C) D-Glyceraldehyde CHO CHO HCOH HOCH Aldotetroses HCOH HCOH (4C) CH2OH CH2OH D-Erythrose D-Threose CHO CHO CHO CHO HCOH HOCH HCOH HOCH Aldopentoses HCOH HCOH HOCH HOCH (5C) HCOH HCOH HCOH HCOH CH2OH CH2OH CH2OH CH2OH D-Ribose D-Arabinose D-Xylose D-Lyxose CHO CHO CHO CHO CHO CHO CHO CHO HCOH HOCH HCOH HOCH HCOH HOCH HCOH HOCH Aldo- HCOH HCOH HOCH HOCH HCOH HCOH HOCH HOCH hexoses (6C) HCOH HCOH HCOH HCOH HOCH HOCH HOCH HOCH HCOH HCOH HCOH HCOH HCOH HCOH HCOH HCOH CH2OH CH2OH CH2OH CH2OH CH2OH CH2OH CH2OH CH2OH D-Allose D-Altrose D-Glucose D-Mannose D-Gulose D-Idose D-Galactose D-Talose Fig. 2.2 : The structural relationship between D-aldoses shown in Fischer projection. (The configuration around C2 (red) distinguishes the members of each pair). epimerization, and a group of enzymes— The term diastereomers is used to represent namely—epimerases catalyse this reaction. the stereoisomers that are not mirror images of one another. Enantiomers Enantiomers are a special type of STRUCTURE OF GLUCOSE stereoisomers that are mirror images of each other. The two members are designated as For a better understanding of glucose D- and L-sugars. Enantiomers of glucose are structure, let us consider the formation of depicted in Fig.2.5. hemiacetals and hemiketals, respectively Majority of the sugars in the higher animals produced when an aldehyde or a ketone reacts (including man) are of D-type (Fig.2.5). with alcohol. 14 BIOCHEMISTRY CH2OH CH2OH C O CH2OH CH2OH C O HOCH C O C O HOCH HCOH CH2OH HOCH HCOH HCOH HCOH C O HCOH HCOH HCOH HCOH CH2OH CH2OH CH2OH CH2OH CH2OH Dihydroxyacetone D-Xylulose D-Ribulose D-Fructose D-Sedoheptulose Fig. 2.3 : Structures of ketoses of physiological importance. OR2 Anomers—mutarotation H R1 C + R2 OH R1 C H The D and E cyclic forms of D-glucose are O OH known as anomers. They differ from each other Aldehyde Alcohol Hemiacetal in the configuration only around C1 known as The hydroxyl group of monosaccharides can anomeric carbon (hemiacetal carbon). In case of react with its own aldehyde or keto functional D anomer, the OH group held by anomeric group to form hemiacetal and hemiketal. Thus, carbon is on the opposite side of the group the aldehyde group of glucose at C1 reacts CH2OH of sugar ring. The reverse is true for with alcohol group at C5 to form two types E-anomer. The anomers differ in certain physical of cyclic hemiacetals namely D and E, as depicted and chemical properties. in Fig.2.6. The configuration of glucose is Mutarotation : The D and E anomers of conveniently represented either by Fischer glucose have different optical rotations. The formulae or by Haworth projection formulae. specific optical rotation of a freshly prepared glucose (D anomer) solution in water is +112.2° Pyranose and furanose structures which gradually changes and attains an Haworth projection formulae are depicted by equilibrium with a constant value of +52.7°. In a six-membered ring pyranose (based on pyran) the presence of alkali, the decrease in optical or a five-membered ring furanose (based on rotation is rapid. The optical rotation of furan). The cyclic forms of glucose are known as E-glucose is +18.7°. Mutarotation is defined as D-D-glucopyranose and D-D-glucofuranose the change in the specific optical rotation (Fig.2.7). representing the interconversion of D and E H C O H C O H C O H H 2 2 H C OH H C OH HO C H O C C O HO C H HO C H HO C H HO C H H C OH 4 4 HO C H H C OH H C OH H C OH HO C H H C OH H C OH H C OH HO C H H C OH CH2OH CH2OH CH2OH HO C H H C OH D-Galactose D-Glucose D-Mannose H C H H C H OH HO Fig. 2.4 : Structures of epimers (glucose and galactose L-Glucose D-Glucose are C4-epimers while glucose and mannose are C2-epimers). Fig. 2.5 : Enantiomers (mirror images) of glucose. Chapter 2 : CARBOHYDRATES 15 H 1 OH 1 HO 1 H C H C O C H C OH H C OH H C OH (A) HO C H O HO C H HO C H O H C OH H C OH H C OH 5 5 5 H C H C OH H C CH2OH CH2OH CH2OH D-D-Glucose D-Glucose E-D-Glucose (+ 112.2q) (aldehyde form) (+ 18.7q ) CH2OH CH2OH CH2OH O OH O H H H H OH H H H (B) O C H OH H OH H OH H HO OH HO HO H H OH H OH H OH D-D-Glucopyranose D-Glucose E-D-Glucopyranose (aldehyde form, acyclic) Fig. 2.6 : Mutarotation of glucose representing D and E anomers (A) Fischer projections (B) Haworth projections. forms of D-glucose to an equilibrium mixture. 1% open chain form. In aqueous solution, the E Mutarotation depicted in Fig. 2.6, is summarized form is more predominant due to its stable below. conformation. The D and E forms of glucose are interconvertible which occurs through a linear D-D-Glucose Equilibrium mixture E-D-Glucose form. The latter, as such, is present in an + 112.2° + 52.7° + 18.7° insignificant quantity. (Specific optical rotation [D]20 D ) Mutarotation of fructose : Fructose also The equilibrium mixture contains 63% exhibits mutarotation. In case of fructose, the E-anomer and 36% D-anomer of glucose with pyranose ring (six-membered) is converted to furanose (five-membered) ring, till an equilibrium is attained. And fructose has a specific optical O O rotation of –92° at equilibrium. The conversion of dextrorotatory (+) sucrose to levorotatory fructose is explained under inversion of sucrose (see later in this chapter). Pyran Furan CH2OH CH2OH REACTIONS OF MONOSACCHARIDES O H C OH O Tautomerization or enolization H H H H The process of shifting a hydrogen atom from OH H OH H HO OH OH one carbon atom to another to produce enediols H is known as tautomerization. Sugars possessing H OH H OH anomeric carbon atom undergo tautomerization D-D-Glucopyranose D-D-Glucofuranose in alkaline solutions. Fig. 2.7 : Structure of glucose-pyranose When glucose is kept in alkaline solution for and furanose forms. several hours, it undergoes isomerization to form 16 BIOCHEMISTRY H Sugar H C OH CuSO4 H C O C O H C O Enediol H C OH HO C H HO C H Sugar acid 2+ + HO C H R HO C H Cu Cu D-Fructose R R D-Glucose D-Mannose 2H2O + Cu2O 2Cu(OH) H C OH It may be noted that the reducing property of C OH sugars cannot help for a specific identification of HO C H any one sugar, since it is a general reaction. R Enediol Oxidation (common) Depending on the oxidizing agent used, the terminal aldehyde (or keto) or the terminal Fig. 2.8 : Formation of a common enediol from glucose, fructose and mannose alcohol or both the groups may be oxidized. For (R corresponds to the end 3 carbon common structure). instance, consider glucose : 1. Oxidation of aldehyde group (CHO o COOH) results in the formation of gluconic acid. D-fructose and D-mannose. This reaction— known as the Lobry de Bruyn-von Ekenstein 2. Oxidation of terminal alcohol group transformation—results in the formation of a (CH2OH o COOH) leads to the production of common intermediate—namely enediol—for all glucuronic acid. the three sugars, as depicted in Fig.2.8. Reduction The enediols are highly reactive, hence sugars When treated with reducing agents such as in alkaline solution are powerful reducing sodium amalgam, the aldehyde or keto group of agents. monosaccharide is reduced to corresponding alcohol, as indicated by the general formula : Reducing properties H The sugars are classified as reducing or non- 2H H C O H C OH reducing. The reducing property is attributed to the free aldehyde or keto group of anomeric R R carbon. The important monosaccharides and their In the laboratory, many tests are employed to corresponding alcohols are given below. identify the reducing action of sugars. These D-Glucose o D-Sorbitol include Benedict’s test, Fehling’s test, Barfoed’s D-Galactose o D-Dulcitol test etc. The reduction is much more efficient D-Mannose o D-Mannitol in the alkaline medium than in the acid D-Fructose o D-Mannitol + D-Sorbitol medium. D-Ribose o D-Ribitol The enediol forms (explained above) or sugars Sorbitol and dulcitol when accumulate in reduce cupric ions (Cu2+) of copper sulphate tissues in large amounts cause strong osmotic to cuprous ions (Cu+), which form a yellow effects leading to swelling of cells, and certain precipitate of cuprous hydroxide or a pathological conditions. e.g. cataract, peripheral red precipitate of cuprous oxide as shown neuropathy, nephropathy. Mannitol is useful to next. reduce intracranial tension by forced diuresis. Chapter 2 : CARBOHYDRATES 17 H C O H C O configuration on these two carbons give the H C OH C same type of osazones, since the difference is masked by binding with phenylhydrazine. Thus HO C H Conc. H2SO4 H C O glucose, fructose and mannose give the same H C OH H C type (needle-shaped) osazones. 3H2O H C OH C Reducing disaccharides also give osazones— CH2OH CH2OH maltose sunflower-shaped, and lactose powder- D-Glucose Hydroxymethyl furfural puff shaped. H C O H C O Formation of esters H C OH C The alcoholic groups of monosaccharides H C OH Conc. H2SO4 H C may be esterified by non-enzymatic or O enzymatic reactions. Esterification of carbo- H C OH H C 3H2O hydrate with phosphoric acid is a common CH2OH H C reaction in metabolism. Glucose 6-phosphate D-Ribose Furfural and glucose 1-phosphate are good examples. Fig. 2.9 : Dehydration of monosaccharides ATP donates the phosphate moiety in ester with concentrated H2SO4. formation. GLYCOSIDES Dehydration Glycosides are formed when the hemiacetal When treated with concentrated sulfuric acid, or hemiketal hydroxyl group (of anomeric monosaccharides undergo dehydration with an carbon) of a carbohydrate reacts with a hydroxyl elimination of 3 water molecules. Thus hexoses group of another carbohydrate or a non- give hydroxymethyl furfural while pentoses give carbohydrate (e.g. methyl alcohol, phenol, furfural on dehydration (Fig.2.9). These furfurals glycerol). The bond so formed is known as can condense with phenolic compounds glycosidic bond and the non-carbohydrate (D-naphthol) to form coloured products. This is moiety (when present) is referred to as aglycone. the chemical basis of the popular Molisch test. In case of oligo- and polysaccharides, they are H C O first hydrolysed to monosaccharides by acid, and + H2N NH C6H5 H C OH this is followed by dehydration. R Bial’s test : Pentoses react with strong HCl to Glucose Phenylhydrazine form furfural derivatives which in turn react with orcinol to form green coloured complex. Bial’s H C N NH C6H5 test is useful for detection of xylose in urine in essential pentosuria. H C OH Mucic acid test : Galactose when treated with R Glucohydrazone nitric acid forms insoluble mucic acid crystals. H2N NH C6H5 Osazone formation H C N NH C6H5 Phenylhydrazine in acetic acid, when boiled with reducing sugars, forms osazones in a C N NH C6H5 reaction summarized in Fig.2.10. R Glucosazone As is evident from the reaction, the first two carbons (C1 and C2) are involved in osazone Fig. 2.10 : A summary of osazone formation (R represents C3 to C6 of glucose). formation. The sugars that differ in their 18 BIOCHEMISTRY The monosaccharides are held together by formed are amino sugars e.g. D-glucosamine, glycosidic bonds to result in di-, oligo- or D-galactosamine. They are present as consti- polysaccharides (see later for structures). tuents of heteropolysaccharides. Naming of glycosidic bond : The N-Acetylneuraminic acid (NANA) is a nomenclature of glycosidic bonds is based on derivative of N-acetylmannose and pyruvic acid. the linkages between the carbon atoms and the It is an important constituent of glycoproteins status of the anomeric carbon (D or E). For and glycolipids. The term sialic acid is used to instance, lactose—which is formed by a bond include NANA and its other derivatives. between C1 of E-galactose and C4 of glucose— Certain antibiotics contain amino sugars is named as E(1 o 4) glycosidic bond. The other which may be involved in the antibiotic activity glycosidic bonds are described in the structure e.g. erythromycin. of di- and polysaccharides. 5. Deoxysugars : These are the sugars that Physiologically important glycosides contain one oxygen less than that present in the 1. Glucovanillin (vanillin-D-glucoside) is a parent molecule. The groups CHOH and natural substance that imparts vanilla flavour. CH2OH become CH2 and CH3 due to the 2. Cardiac glycosides (steroidal glycosides) : absence of oxygen. D-2-Deoxyribose is the most Digoxin and digitoxin contain the aglycone important deoxysugar since it is a structural steroid and they stimulate muscle contraction. constituent of DNA (in contrast to D-ribose in 3. Streptomycin, an antibiotic used in the RNA). Feulgen staining can specifically detect treatment of tuberculosis is a glycoside. deoxyribose, and thus DNA in tissues. Fucose is 4. Ouabain inhibits Na+ – K+ ATPase and a deoxy L-galactose found in blood group blocks the active transport of Na+. antigens, and certain glycoproteins. 5. Phlorhizin produces renal damage in 6. L-Ascorbic acid (vitamin C) : This is a experimental animals. water-soluble vitamin, the structure of which closely resembles that of a monosaccharide. DERIVATIVES OF MONOSACCHARIDES There are several derivatives of monosaccha- rides, some of which are physiologically important (Fig.2.11) DISACCHARIDES 1. Sugar acids : Oxidation of aldehyde or primary alcohol group in monosaccharide results Among the oligosaccharides, disaccharides in sugar acids. Gluconic acid is produced from are the most common (Fig.2.12). As is evident glucose by oxidation of aldehyde (C1 group) from the name, a disaccharide consists of two whereas glucuronic acid is formed when primary monosaccharide units (similar or dissimilar) held alcohol group (C6) is oxidized. together by a glycosidic bond. They are 2. Sugar alcohols (polyols) : They are crystalline, water-soluble and sweet to taste. The produced by reduction of aldoses or ketoses. For disaccharides are of two types instance, sorbitol is formed from glucose and 1. Reducing disaccharides with free aldehyde mannitol from mannose. or keto group e.g. maltose, lactose. 3. Alditols : The monosaccharides, on reduction, yield polyhydroxy alcohols, known as 2. Non-reducing disaccharides with no free alditols. Ribitol is a constituent of flavin aldehyde or keto group e.g. sucrose, trehalose. coenzymes; glycerol and myo-inositol are components of lipids. Xylitol is a sweetener used Maltose in sugarless gums and candies. Maltose is composed of two D-D-glucose 4. Amino sugars : When one or more units held together by D (1 o 4) glycosidic bond. hydroxyl groups of the monosaccharides are The free aldehyde group present on C1 of second replaced by amino groups, the products glucose answers the reducing reactions, besides Chapter 2 : CARBOHYDRATES 19 H C O H C OH OH OH CH2OH HO C H H OH H C OH H H H C OH CH2OH OH H H C OH HO H Glycerol COOH H OH D-Glucuronic acid myo -Inositol H CH2OH O O HOCH2 O OH O H3C C HN H OH COO– H H H OH H CH2OH H H OH H H H H OH H H HO OH OH H H NH2 HO H D-2-Deoxyribose D-Glucosamine N-Acetylneuraminic acid Fig. 2.11 : Structures of monosaccharide derivatives (selected examples). the osazone formations (sunflower-shaped). Inversion of sucrose Maltose can be hydrolysed by dilute acid or the Sucrose, as such is dextrorotatory (+66.5°). enzyme maltase to liberate two molecules of But, when hydrolysed, sucrose becomes D-D-glucose. levorotatory (–28.2°). The process of change In isomaltose, the glucose units are held in optical rotation from dextrorotatory (+) together by D (1 o 6) glycosidic linkage. to levorotatory (–) is referred to as inversion. The hydrolysed mixture of sucrose, containing Cellobiose is another disaccharide, identical glucose and fructose, is known as invert sugar. in structure with maltose, except that the former The process of inversion is explained below. has E (1 o 4) glycosidic linkage. Cellobiose is formed during the hydrolysis of cellulose. Hydrolysis of sucrose by the enzyme sucrase (invertase) or dilute acid liberates one molecule Sucrose each of glucose and fructose. It is postulated that sucrose (dextro) is first split into D-D- Sucrose (cane sugar) is the sugar of commerce, glucopyranose (+52.5°) and E-D-fructofuranose, mostly produced by sugar cane and sugar beets. both being dextrorotatory. However, E-D- Sucrose is made up of D-D-glucose and E- fructofuranose is less stable and immediately gets D-fructose. The two monosaccharides are held converted to E-D-fructopyranose which is together by a glycosidic bond (D1 o E2), between strongly levorotatory (–92°). The overall effect is C1 of D-glucose and C2 of E-fructose. The that dextro sucrose (+66.5°) on inversion is reducing groups of glucose and fructose are converted to levo form (–28.2°). involved in glycosidic bond, hence sucrose is a non-reducing sugar, and it cannot form osazones. Lactose Sucrose is an important source of dietary Lactose is more commonly known as milk carbohydrate. It is sweeter than most other sugar since it is the disaccharide found in milk. common sugars (except fructose) namely glucose, Lactose is composed of E-D-galactose and E-D- lactose and maltose. Sucrose is employed as a glucose held together by E (1 o 4) glycosidic sweetening agent in food industry. The intestinal bond. The anomeric carbon of C1 glucose is free, enzyme—sucrase—hydrolyses sucrose to glucose hence lactose exhibits reducing properties and and fructose which are absorbed. forms osazones (powder-puff or hedgehog shape). 20 BIOCHEMISTRY CH2OH CH2OH POLYSACCHARIDES O O H H H H H H Polysaccharides (or simply glycans) consist of 1 4 HO OH H O OH H repeat units of monosaccharides or their OH derivatives, held together by glycosidic bonds. H OH H OH They are primarily concerned with two important Glucose Glucose functions-structural, and storage of energy. Maltose (D-D-glucosyl (1 o 4) D-D-glucose) Polysaccharides are linear as well as branched polymers. This is in contrast to structure of proteins and nucleic acids which are CH2OH only linear polymers. The occurrence of O O branches in polysaccharides is due to the fact H H HOH2C H H that glycosidic linkages can be formed at any 1 2 OH H O H HO one of the hydroxyl groups of a monosaccharide. HO CH2OH Polysaccharides are of two types H OH OH H Glucose 1. Homopolysaccharides on hydrolysis yield Fructose Sucrose only a single type of monosaccharide. They (D-D-glucosyl (1 o 2) E-D-fructose) are named based on the nature of the monosaccharide. Thus, glucans are polymers of glucose whereas fructosans are polymers of