This book is freely available for research and educational purposes. XML to PDF by RenderX XEP XSL-FO F ormatter, visit us at š¯—£š¯——š¯—™ | principle of practical biochemistry and experimental part. BIOCHEMISTRY. Book Ā· January with 25, Reads. Publisher: 1st ed. Try this link Practical Textbook of Biochemistry for Medical Students You may also check out this Textbook of Biochemistry for Medical Students.

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We are hopeful that this practical biochemistry book will help the medical students to envisage about the various facts encountered in the reactions in the body. For every medical practitioner and enlighted patients, Biochemistry has been playing a significant role. This book will certainly be of significant importances to . The given manual is designed according to the curriculum on biochemistry for the students It is intended to save the student's time and optimize their practical.

Principles and Techniques of Practical Biochemistry 5th Ed. Wilson, K. The first edition of this multiauthor lab practical book appeared in , and the fact that it is now in its fifth edition attests to its usefulness to the biochemistry teaching community.

Laboratory work has of course moved on a great deal since can you remember what sorts of techniques you were using in ? It is hard to foresee what the next edition might contain in a world of mass spectrometric sequencing and DNA arrays, as well as robotics.

It is quite a problem for those teaching to undergraduates both to keep up with what is going on and also to have suitable equipment on which to train the students. In general, I suppose we must be into teaching the basic principles, the theory behind the methods, and concepts such as accuracy and statistical treatments and then rely on undergraduate projects or summer lab experiences or schemes where students take a year out and work in industry to allow the undergraduates to get experience on current instrumentation.

It is not a lab recipe book either. No part of this publication should be reproduced, stored in a retrieval system, or transmitted in any form or by any means: This book has been published in good faith that the material provided by authors is original.

Every effort is made to ensure accuracy of material, but the publisher, printer and authors will not be held responsible for any inadvertent error s. In case of any dispute, all legal matters are to be settled under Delhi jurisdiction only. First Edition: Clinical biochemistry plays a tremendous impact on the diagnosis and treatment of patients.

Medical students should be aware of the practicals, diagnostic parameters and their estimations. They should acquire sound knowledge about the diagnostic reports and its implications which aids in diagnosis and prognosis of the disease. Biochemistry is the most fast growing subject, extensively applicable to understand the disease at molecular level. Estimations of various biochemical parameters definitely give an insight to understand the normal metabolism and its aberrations leading to diseases, which forms the foundation for medicine.

Biochemistry should be encouraged in relation to health and disease which will make the subject more interesting and fascinating to the students. We are hopeful that this practical biochemistry book will help the medical students to envisage about the various facts encountered in the reactions in the body.

In our teaching biochemistry to undergraduate medical students we have realized practicals is where they have shown some amount of enthusiasm towards biochemistry. Towards achieving this goal, each practical session has been reorganized such that it becomes easy to understand. Wherever possible, the subject is presented in tabular format such that it becomes very concise; and all test results are given in color such that the simple task of reading the book itself realizes one doing the practicals.

Fundamental concepts and principles behind each experiment are explained in a simple way. Our only genuine concern is to help you to understand the subject in an easy and organized way such that this little knowledge comes as a big help not only in your exams but also in your future medical career. We will be glad to accept constructive criticism and fruitful suggestion to make this book a better one.

I take this opportunity to thank computer operators Mrs Veena Jayaram and Mr Sunder for their help in preparing the manuscript.

Laboratory Hazards and First Aid Laboratory Safety Rules Specimen Collection and Processing Glasswares Used in Biochemistry Laboratory Qualitative Analysis of Carbohydrates Qualitative Analysis of Proteins Nonprotein Nitrogenous Substances Qualitative Analysis of Normal Urine Analysis of Abnormal Constituents in Urine Hemoglobin and its Derivatives Spot Tests Principles of Colorimetry Estimation of Blood Sugar Estimation of Blood Urea Estimation of Urine Creatinine Estimation of Serum Inorganic Phosphate Estimation of Serum Total Proteins Glucose Tolerance Test Estimation of Serum Cholesterol Estimation of Plasma Ascorbic Acid Flame Photometer CSF Analysis Estimation of Albumin in Urine Biophysics is a growing enterprise worldwide, driven primarily by the widespread realization of the major contribution that can be made to biological science by a combination of truly state-of-the-art physical measurements with modern molecular biology.

The field occupies a unique and central position at the intersection of the biological, chemical, physical, and computational sciences. Biophysics takes a quantitative, physical, non-phenomenological approach to biology that is firmly rooted in the principles of condensed-phase physics and physical chemistry.

Biophysicists are driven primarily by their curiosity about how biological systems work at the molecular level. While they routinely employ the methods of molecular biology, their primary focus is on development of novel structural and dynamical tools that enable uniquely incisive studies of systems ranging in complexity from single proteins in vitro to the complex interactions of biopolymers in live cells.

Biophysicists as a group most often develop the novel, sophisticated experimental methods that reveal molecular level details with unprecedented clarity. The state of the art in X-ray crystallography, solution phase and solid-state NMR, atomic force microscopy, single-molecule methods, EPR, and fluorescence microscopy continues to evolve in ways that better elucidate biological structure and function.

In parallel, biophysicists are developing powerful new computational tools based on firmly established physical principles that are sufficiently accurate to greatly enhance insights from experiment. Just as the tools of molecular biology gradually become useful to biophysicists, overtime the new tools developed by biophysicists gradually find widespread use among all biological scientists.

Practical, functional knowledge of physical principles that underline nursing procedures and the operation of machinery that nurses use. Technical knowledge from the science of physics that applies specifically to nursing performance and understand certain biomedical phenomenon like how does a suction apparatus operates?

What is the most efficient way to move a heavy object or a patient? How does air get in and out of the lungs? In addition study of biophysics helps a nurse understand following contents of nursing: In this situation, a nurse takes a measurement of physical quantity and compare measured value of physical quantity with a standard to determine its relationship with that standard. The standards of measurement is called a unit. The numerical value of a physical quantity, therefore, refers to the number of standard unit of measurement.

Thus measurement of any quantity has two characteristicsā€”a numerical value and a unit. For example, you measure the birth weight of a baby as 3. Then 3. Although the number of physical quantities that we measure is very large, we do not need a very large number of standards to compare every measurement.

It is so because all the physical quantities are not independent quantities in so far as their measurement is concerned. For example, velocity of a body is measured in units of length meter and time seconds. A few independent standards have been chosen to fix the units of certain physical quantities.

These independent standards are length, mass and time. Such units fixed by independent standards are called fundamental units. For example, ā€” One meter: These units are called derived units. For example, the unit of volume is cubic meter which is derived from the unit of length. The unit of speed is derived from units of length and time.

They differ from each other because different standards of measurement are used for fundamental quantities. Table 1. The two systems of measurement most frequently used in nursing practice are the MKS also called metric and the FPS also called English. You may note from Table 1.

A Course In Practical Biochemistry

The unit of length in English system is the foot. The unit of length in the Metric system is the meter. Similarly, in microscopic work, a very small unitā€” micron is used. The various multiples of units of length are listed in Table 1. The unit of mass in Metric system and SI system is the kilogram kg.

A physical balance ordinarily measures the mass of a body. Some of these units are used in measuring food items for special diets, amount of drugs, weights of patients, etc. Although commonly we use the terms mass and weight in the same sense, the two terms have different meanings in physics.

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In physics, concept of mass and weight are different. Mass of a body is the quantity of matter contained in it. On the other hand, weight is defined as the gravitational force with which a body is pull towards the center of the earth. In the SI system, the unit of weight is Newton. For example, an object at sea level weighs more than it does on a high mountain because the value of the acceleration due to gravity of the earth on the object is greater at sea level.

The mass, however, remains the same everywhere. The mass of an object is measured by a physical balance whereas its weight is measured by a spring balance. Mass is a scalar quantity while weight is a vector quantity because it is directed towards the center of the earth.

Mass of a person is the same on the earth as well as on the moon, but weight of the person is different at these two places because their pulls on the person are different. A person weighs six times more on earth than on the moon. Whereas mass and weight have the same numerical value, it is important in solving problems to indicate the unit specifically, as one of force weight or as one of mass. Units of Time The unit of time is the second and is based on the natural clock.

Let us consider some of the measurements of time you make in the course of your work. However, for studying the heart beat of a patient by electrocardiography, greater accuracy in the measurement of time is required. In this case the beating of the heart must be accurately measured in tenths or hundredths of a second. In nursing practice, you may come across situations when a measurement taken in Metric unit must be changed to the corresponding English unit and vice versa.

For this reason, approximate equivalents commonly used in the hospital are given in Table 1. Discuss the meaning and importance of biophysics in nursing. Discuss the concept of units and fundamental and derived units. Describe the different system of the units. List of the basic units of length, weight, mass and time. San Francisco, Biophysical Chemistry ; Semi-classical method of calculation of rates of chemical reactions proceeding in polar liquids.

J Electroanal Chem ; Eugenie V. Biological Physics.

New York. American Institute of Physic, An Introduction to Physics in Nursing. The CV Mosby Company, Glaser R, Biophysics, Springer, Glaser R.

An Introduction. Springer Verlag, Heidelberg, Glaser, Roland. An Introduction Corrected ed. Springer, ; Fundamental Physics. Paedeep Publishers, Oncise Physics, New Delhi: Selina Publishers, August Intermediate Physics for Medicine and Biology 4th Edition. Springer, International Physics for Medicine and Biology 4th ed. Lal S. Principles of Physics. Meyer B Jackson.

Molecular and cellular biophysics. New York: Cambridge Publication, Nicolas Rashevsky, Mathematical biophysics. Perutz MF.

Proceedings of the Royal Society of London. Series, ; B Proteins and Nucleic Acids, Elsevier, Amsterdam, Proteins and Nucleic Acids: Structure and Function. Elsevier, Philip C, Nelson. Biological Physics Updated Edition. WH Freeman, Physical Biology of the Cell.

Garland Science, Oxford, Rashevsky, N. Physico- Mathematical Foundations of Biology - Vol. Dover Publications, Third Revised Edition, Rodney MJ, Cotterill.

Wiley, Medical physiology and biophysics. Saunders Sneppen K, Zocchi G. Physics in Molecular Biology. Cambridge University Press, Physics in Molecular Biology 1 ed. Cambridge University Press ; Theory of Enzyme Catalysis. Molekuliarnaya Biologia Moscow ; 6: The medical students should have the knowledge of the various tests, diagnostic investigations done in biochemistry laboratory. They should also be aware of all potential hazards and the safety measures.

Damage should be rectified immediately. The mouth of the pipette should be plugged with cotton or piece of rubber while filling. If the corrosive solution swallowed is an acid: If the corrosive solution swallowed is an alkali: Laboratory Hazards and First Aid 7 From strong alkalies: The first aid kit should contain: One should be aware that accidental injuries can occur either from direct contact through skin, by inhaling vapors, powder or swallowing by mistake while pipetting.

Strong acids like sulfuric acid, hydrochloric acid and strong alkalies like potassium hydroxide, sodium hydroxide, etc. Benzene is toxic to bone marrow; carbon tetrachloride and halogenated hydrocarbons are toxic to liver, etc.

Hence, their use should be minimized in assays. Skin contact with them must be strictly avoided and rubber or plastic gloves should be used while handling these substances.

Certain precautionary measures must be followed like: Perchloric acid should be kept in fume cupboard. Picric acid should be stored in a container of water tightly closed with cork or rubber stopper. Ether should be kept in brown or dark bottles away from sunlight since on exposure to sunlight they form peroxides, that when raised to certain sufficient concentration cause violent explosion.

Laboratory Hazards and First Aid 9 4. Cylinder containing inflammable gases like hydrogen, propane, acetylene should be kept outside the laboratory when not in use. Radioactive Waste Expert opinion has to be taken for the disposal of radioactive waste, and their guidelines have to be strictly followed.

Flushing radioactive substances down the sink can be very dangerous as they pollute the underground water table. That is true for the rules used in a biochemistry lab. Conduct yourself in a responsible manner at all times in the laboratory. Follow all written and verbal instructions carefully. Do not touch any equipment, chemicals, or other materials in the laboratory area until you are instructed to do so.

Laboratory Safety Rules 11 4. Be prepared for your work in the laboratory. Read all procedures thoroughly before entering the laboratory. Never fool around in the laboratory.

Always work in a well-ventilated area. Observe good housekeeping practices. Work areas should be kept clean and tidy at all times. Be alert and proceed with caution at all times in the laboratory. Notify the teacher immediately, if you observe any unsafe conditions. Dispose all chemical wastes properly. Never mix chemicals in sink drains. Sinks are to be used only for water. Check with your teacher for disposal of chemicals and solutions. Keep hands away from face, eyes, mouth and body while using chemicals or lab equipment.

Wash your hands with soap after performing all experiments. Any time when chemicals, heat, or glassware are used, students must wear safety goggles. Dress properly during a laboratory activity. Long hair, dangling jewelry, and loose or baggy clothing are hazardous in the laboratory. Long hair must be tied back, and dangling jewelry and baggy clothing must be secured. Shoes must completely cover the foot. No sandals allowed in the laboratory. A lab coat should be worn during laboratory experiments.

Report any accident spill, breakage, etc. Do not panic. If a chemical splashed into your eye s or on your skin, immediately flush with running tab water for at least 20 minutes. All chemicals in the laboratory are to be considered dangerous.

Avoid handling chemicals with fingers.

Do not taste, or smell any chemicals. Check the label on all chemical bottles twice before removing any of the contents. Take only as much chemical as you need.

Never return unused chemicals to their original container.

Introductory Practical Biochemistry

Never remove chemicals or other materials from the laboratory area. Never pipette by mouth. Never handle broken glass with your bare hands. Use a brush and dustpan to clean up broken glass. Place broken glass in the designated glass disposal container.

Examine glassware before each use. Never use chipped, cracked or dirty glassware. Do not immerse hot glassware in cold water. The glassware may shatter.

Heated glassware remains very hot for a long time. They should be set aside in a designated place to cool, and picked up with caution, using tongs.

Never look into a container that is being heated as there are chances of it getting splashed to the face or eyes. Clean all the glassware you have and put them on the shelf in a proper order.

Wipe and clean the table. Put all chemicals back to their respective places. Put off the gas burner. Use a pipette bulb to draw liquid above the calibration mark. Remove the bulb and cover the pipette with your forefinger. Dry the pipette tip with a tissue. Laboratory Safety Rules 15 4. Rotate the pipette using the thumb and the other fingers to let in air so that the liquid drains slowly until the meniscus reaches the calibration mark.

To deliver the liquid, hold the pipette vertically and let the pipette tip touch the wall of the receiving container. When the delivery is completed, touch the tip of the pipette to the wall of the container. Always keep the pipette tip under liquid surface when you draw up liquid. Never use the bulb to blow air inside the pipette, this will introduce dust and make the pipette dirty.

Blood is the most frequently used body fluid for analytical purposes. Ideally all estimations should be performed within hr after collection.

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For extracting serum, allow the blood to clot at room temperature for 30 minutes. Loosen the clot by a stick, and centrifuge for 10 minutes at RPM. Separate the serum and label it. They can be used later for analysis. Special care should be taken to avoid hemolysis. Hemolyzed samples alter the values of many chemical estimations because of the release of RBC contents, which can cause color change.

False high results may be obtained because of hemolysis. Hemolyzed samples affect bilirubin and enzyme estimations giving erroneous results. Venipuncture is more commonly performed for obtaining blood. Disposable needles are used to eliminate the hazards of infections. Whole blood, serum or plasma can be selected depending upon the methods by which the biochemical parameters are to be investigated.

Serum is the fluid portion of clotted blood while plasma is the fluid portion of unclotted blood. Various anticoagu- lants are used depending on the parameters to be analyzed.

Blood is collected and mixed with some chemicals that prevent clotting. These chemicals are called anticoagulants.

Most of the anticoagulants used in the laboratory, act by binding calcium as an insoluble salt. Oxalates, citrates and EDTA chelate calcium ion.

The routinely used anticoagulants are: Potassium oxalate 2. Double oxalate 3. Ammonium oxalate 4. Sodium citrate 5.

EDTA ethylene diamine tetra acetate 7. Sodium fluoride to prevent glycolysis; used for glucose estimation 8. Acid citrate dextrose ACD. Preservation of Urine Samples Several changes like urinary decomposition, precipitation of phosphates, crystallization of uric acid and bacterial action may alter the urinary composition, if it is kept for long periods, especially in the collection of 24 hours urine samples.

Also urine may become alkaline due to precipitation of uric acid and urates. Specimen Collection and Processing 19 Various preservatives are used depending on the analysis of parameters in urine. The common ones are concentrated hydrochloric acid HCl toluene and liquid petroleum. Before carrying out any estimation in urine, the urinary deposits must be mixed well. These glasswares may be volumetric graduated or non-volumetric non-graduated. Volumetric glasswares include flasks, measuring cylinders, pipettes, etc.

Non-volumetric glasswares include beakers, funnels, bottles and test tubes, etc. They are used mainly for the preparation of the solutions and reagents. Flasks These have capacities of ml. Different types of flasks are available. Conical flasks Erlenmeyer type: These are used for performing titration, and for boiling the solutions.

Evaporation is minimum because of the conical shape. Flat- bottomed round flasks: These are used mainly for heating the liquids.

Glasswares Used in Biochemistry Laboratory 21 c. Round bottomed flasks: These can withstand high temperature. So they are used to evaporate samples to dryness, distillation of water, alcohol and other organic compounds. Volumetric flasks: They are flat bottomed, pear-shaped vessels with long narrow necks with a specific volume mark and fitted with a stopper. Graduated Measuring Cylinder Graduated measuring cylinders are narrow, straight side vessels that are used to measure specific volumes.

They are available in sizes ranging from 10 ml to several liters. A high degree of accuracy is not possible because of their wider bore. Burettes Burettes are long, graduated tubes with a stop cork at one end, available in capacities of 10 to ml. These devices are used to deliver known volumes of liquid into a container accurately. By measuring from one graduated line to another graduated line, one can deliver even fractional volumes less than 1 ml of liquid with a high degree of accuracy.

They are used mainly for titrations and also to dispense corrosive reagents. Funnels usually have short or long, thin stems. These funnels are used with filter paper to remove particles from solutions. Funnels with wide mouthed stems that allow solids to pass through easily are used for transferring solids into a container. They are made up of plain white or amber colored glass. Amber colored bottles are useful to store certain light sensitive chemicals like silver nitrate.

Drop Bottles Drop bottles have a narrow neck with a slotted glass stopper, available in ml capacities. They are used for delivery of drops of solutions such as stains and indicator solutions and are made up of white or brown glass. Wash Bottles Wash bottles are usually plastic bottles with a delivery tube at the top. They are of various types differing in their levels of accuracy and precision which includes complex adjustable or automatic pipettes.

Manual Pipettes a.

Introduction to Practical Biochemistry

To deliver type of pipettes TD: These pipettes must be held vertically and the tip must be placed against the side of the accepting vessel to drain liquid by gravity. Common pipettes included under TD type are, graduated and volumetric pipettes.

Glasswares Used in Biochemistry Laboratory 23 b. Graduated pipettes: These pipettes are available from 0. Mohr pipettes and serological pipettes. Mohr pipettes are glass tubes of uniform diameter with a tapered delivery tip, have graduations made at uniform intervals but well above the tapered delivery tip. These are mainly used for pipetting distilled water and reagents. However, 0. The serological pipettes are either of TD or blow- out pipettes.

Volumetric pipettes: These pipettes are not graduated but designed specially to deliver a specific quantity of the specimen. They have an open- ended bulb holding the bulk of the liquid, a long glass tube at one end that has the mark to describe the extent to which the pipette is to be filled and a tapered delivery portion. These pipettes hold and deliver only the specific volumes indicated at the upper end of the pipette.

These are used mainly to pipette specimen and standards, and are very accurate. Pipettes must be refilled or rinsed out with the appropriate solvent after the initial liquid has been drained from the pipettes.

It consists of capillary tubing with a line demarcating a specific volume. These are filled to the line by capillary action. Pasteur pipettes: This pipette has a rubber bulb attached to the top of a glass tubing.

It is tapered at the tip and is especially useful in delivering urine samples.

A mechanical plunger does this work. These are frequently used in the laboratory to repeatedly add a specific volume of a reagent. They are mainly of push button type Eppendorf type and are piston operated devices to dispense liquid. Suggestions are made for further work in more advanced classes.

As well as the practical method the experiments are accompanied by background information, discussion of results, references for further study and illustrations. A volume that has been long awaited The editor is to be congratulate d on his efforts As well as practical details nearly all the procedures ar e accompanied by background information and references for further study. The book will have a ready appeal to teachers who are looking for new ideas at all levels of expertise in practical biochemistry.

Endeavour qu: Teachers of biochemistry will turn to this book again and again to find their inspiration This book can be recommended to all teachers as a valuabe collection of practical biochemistry experiments. We are always looking for ways to improve customer experience on Elsevier.

We would like to ask you for a moment of your time to fill in a short questionnaire, at the end of your visit. If you decide to participate, a new browser tab will open so you can complete the survey after you have completed your visit to this website. Thanks in advance for your time. Skip to content. Search for books, journals or webpages All Webpages Books Journals. View on ScienceDirect. Published Date:In case of reusable ones, the needle point should be kept sharp.

This leads to certain unbalanced forces on the surface. Inorganic constituents present in the given sample of normal urine are chloride, calcium, phosphorus, inorganic sulfates, ammonia, sodium, potassium and magnesium. Since then, he is actively involved in teaching Biochemistry to post-graduate students.

Clinical Approach to Practical Biochemistry

Experiment Observation Inference Moisten a piece of Black spots are Indicates the filter paper with seen on the presence of few drops of filter paper. Cool the solution. Wiley, Specific Urease Test Principle:

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