Science

Laborato
KHARADAR GENERAL HOSPITAL
SCHOOL OF NURSING
AFFILIATED WITH DOW UNIVERSITY
OF HEALTH SCIENCES

ry
(BIOCHEMISTRY)

Acknowledgment: Compiled By: Supervised by: Edited by:

Dr. Fauzia Imtiaz Rehan Ahmad Samundar Khan Talat Parveen Shah
Shehla Naeem Zafar Faculty Senior Faculty Principal
Rasheed Ahmed Khan Muhammed Shahid
M. Adeel Coordinator
 PREFACE

The main goal of this scientific manual is to enhance the knowledge of undergraduate
nursing students to understand the biochemistry of life specially structure and function of
macromolecules and their application in health and illness.

With the use of these standard laboratory practicum which are included in the under
graduate nursing curricula will be more beneficial in the clinical area for the patients and
have greater impact in the outcome related to health sciences. understanding of the
essential macromolecules e.g. carbohydrates, lipids and proteins are crucial for the
nursing undergraduates to cope the challenges in medical sciences.

I am very grateful to Ms.Talat Parveen Shah, Principal Kharadar General Hospital School
of Nursing for her encouragement that enables me to write this manual and I also
appreciate the support from the Mohummad Shahid BSN program coordinator. Mr.Rehan
Ahmed, Faculty of KGH-SON, also has graeter contribution in the development of this
Scientific Manual for Biochemitery.

Further suggestions towards improvement of this manual will be highly appreciated.

Samundar Khan
Senior Faculty
Khararad General Hospital
School of Nursing

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 Kharadar General Hospital School of Nursing

CERTIFICATE
This is certify that:

Mr. / Ms. _______________________________________Roll No._____

Of _______________________class has carried out the necessary practical
work as prescribed by Kharadar General Hospital School of Nursing for the
Year________________

_______________ __________________ __________________

Faculty Program Coordinator Head of
Department

Date: - _________________

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 TABLE OF CONTENTS

TITLE PAGE------------------------------------------------------------------------1

PREFACE-------------------------------------------------------------------------- --------2

CERTIFICATE---------------------------------------------------------------------------- 3

TABLE OF CONTENTS---------------------------------------------------------------- 4

INDEX---------------------------------------------------------------------------------------- 5

PRACTICAL-1 HAZARDS & THEIR PRECAUTIONARY

FIRST AID MEASURES-------------------------------------- 6

PRACTICAL-2 SOLUTIONS----------------------------------------------------12

PRACTICAL-3 PROPERTIES OF ORGANIC COMPOUNDS------------19

PRACTICAL-4 CARBOHYDRATES------------------------------------------24

PRACTICAL-5 PROTEINS------------------------------------------------------28

PRACTICAL-6 LIPIDS-----------------------------------------------------------32

PRACTICAL-7 ENZYME KINETICS-----------------------------------------34

PRACTICAL-8 INTRODUCTION OF BODY MASS INDEX (BMI) ---37

PRACTICAL-9 BASAL METABOLIC RATE (BMR) ---------------------41

APPENDIX –I (Some Basic Conversions and Calculations) ----------------44

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 INDEX

S. Date OBJECT OF THE Page Initials Remarks

No. PRACTICAL No.

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PRACTICAL NO.1

HAZARDS & THEIR PRECAUTIONARY FIRST AID MEASURES

1.1 OBJECTIVES:

At the end of this lab students will be able to:

1 List the lab instruction to be followed.

2 List various hazards and accidents can occur during work in lab.

3 Give precautionary measures which can prevent such situations.

4 Give first aid in general.

5 Discuss safety measures we can follow for managing accidental

conditions.

6 Solve the given case.

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1.2 INSTRUCTIONS

TO BE FOLLOWED WHILE WORKING IN SCIENCE

LABORATORY

1. Always wear a lab. Coat while working in laboratory.

2. Do not forget to bring your practical manual in practical class

3. Use equipment and laboratory ware correctly and with care.

4. Always use neat and clean dry test tube.

5. Always keep the test tube in tilting position away from you and others
while heating.

6. Heat only contents of the test tube, not whole.

7. Keep on moving the test tube on the flame while heating.

8. Never move about in the laboratory with a filled pipette in hand. The
falling of reagent may be dangerous for you and other persons in the
laboratory.

9. Breakable items should be kept in proper racks and never at the edge
of the working table.

10.Do not place eatable son the working table.

11.Check that nothing except the required electrical appliance is on.

12.At the end of the work clean all working tables and glass wares.

13.After throwing chemical in the sink, flush with plenty of tap water to
prevent corrosions of drainage pipes.

14.In case of any emergency report to supervising teacher immediately.

15.Read label on the reagent bottle carefully before using the reagents

16.After using the reagent, close the bottle properly.

17.Keep finger nails short

18.No smoking in the working zone of the laboratory.

19.Switch off your mobile phones while working in the lab do not use
hands free or any cell accessories.

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1.3 SAFETY MEASURES FOR SCIENCE LABORATORY

1.3.1 A. General Safety Precautions/Safety Rules

1. Work quietly and carefully, with full attention.

2. Wear appropriate clothing, according to laboratory procedure.

3. Always work under supervision of lab incharge.

4. Inform your supervisor about health-related problems or allergies, if

you have any.

5. Never eat or drink in the Lab area.

6. Do not attempt Lab activities at home unless told to do so by your

teacher, and only under the direct supervision of an experienced
person.

7. Avoid unnecessary touching of substances in the lab.

8. Smell substances using the proper technique (wafting fumes toward

you).

9. Pour substances properly and safely.

10.Clean up all spills immediately

11.Dispose of harmful substances by following teacher's directions.

12.Wash hands after handling substances and before leaving the Lab.

13.Rinse off substances immediately that come into contact with skin
or clothing.

14.Do not perform unauthorized experiments.

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 15.Report all accidents immediately to your teacher/Lab Incharge.

16.If vapors generated are toxic, use a fume hood.

17.Wear chemical splash goggles.

18.Wear a chemical resistant apron.

19.Wear chemical resistant gloves.

20.Tie back long hair.

21.Do not wear loose and short sleeves.

22.Do not wear sandals.

23.Do not wear contact lenses.

24.No food or beverages.

25.No gum chewing.

26.Know the location of all of the science lab safety equipment, exits
and telephone. (Safety showers, eye wash, fire blankets, fire
extinguishers).

1.3.2 B. Handling a Heat Source

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 1. In test tubes, Use heat-resistant glass (Pyrex or Kimax) - never use
cracked glass.

2. Always keep the open end of the test tube pointed away from
everyone.

3. Never allow any container to boil dry. Use tongs or gloves to pick up
hot objects.

4. Turn off hot plate when not in use. Unplug cords by pulling on the
plug, not the cord.

5. Report and replace equipment that has frayed or damaged cords.

6. Make sure electrical cords are placed properly where no one will trip
over them.

7. Treat burns using cold water or ice.

8.

1.3.3 C. Handling an Open Flame

1. Locate fire safety equipment before using any open flame (fire
blanket, fire extinguishers, fire alarm, first-aid kit).

2. Know the proper procedures for using a Bunsen burner In the Lab.

3. Remove all flammable substances from the room before lighting a

flame.

4. Use a test tube holder if the test tube is being heated in an open flame.

5. Point the open end away from yourself and others.

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 6. Gently move the test tube back and forth over the flame so that it is
heated evenly.

1.3.4 D. Recommendations

1. Dispose of broken glassware as instructed by your teacher.

2. Report broken or damaged equipment immediately (DO NOT USE

IT).

3. Clean up work area completely when you are finished.

4. Wash all glassware thoroughly and place in drying racks.

5. Report all accidents to the teacher immediately (no matter how minor)

1.4 HAZARDS SYMBOLS

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1.4.1 DISCUSSION:
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PRACTICAL NO.2

SOLUTIONS

2.1 OBJECTIVES:
At the completion of this lab students will be able to:

1. Define stock solution standard.

2. Define serial dilution.

3. Prepare stock solution of 1gm glucose/100ml and 10gm of

NaCl/100ml.

4. Prepare serial dilutions of 50mg, 100mg, 150mg and 200mg/100ml

from the stock solution of glucose.

5. Prepare 0.9%, 3% and 5% saline solution.

6. Convert 70mg%, 160mg%and 200mg% of glucose in mmol/L.

7. Convert 14.2mg% of Na and 0.14mg% of K into meq/L.

8. Discuss the significance of:

a. 5%, 10% and 25% dextrose solution.

b. 0.9, 3% and 5%of saline solution in clinical practice.

2.2 OBJECT: To study concept of solution

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2.3 SOLUTION:
Solution is a homologenous mixture of two or more Substances.
A solution consists of two parts (I) solute (ii) solvent

2.3.1 SOLUTE:
Solute is smaller and dispersed part of solution, which is usually a solid but
may be liquid.
.
2.3.2 SOLVENT:
Solvent is larger part and it is the dissolving material of Solution, usually it is
liquid but may be solid.

2.4 STANDARD SOLUTION:

It is a solution when we know the exact concentration, % or quantity of
solute and solvent.

2.5 TYPES OF SOLUTIONS:

From the biochemistrical point of view, the solutions are the following four
types.

(1) PERCENT SOLUTION (%)

(2) MOLAR SOLUTION (M)
(3) MOLAL SOLUTION (m)
(4) NORMAL SOLUTION (N)

2.5.1 PERCENT SOLUTION:

1. W / W % = (Mass of Solute /Mass of Solution) X 100

2. W / V % = (Mass Of Solute/Volume Of Solution) ) X 100

3. V / V % = (Volume of Solute/Volume Solution) X100

2.5.2 MOLARITY:
Number of Moles of Solute per Unit Volume (L=dm 3)

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 M = (Number of Mole of Solute / Volume of Solution) Or

M =MASS OF SOLUTE (IN GRAMS) / (MOLECULER MASS X VOLUME OF

SOLUTION IN L).

2.5.3 MOLALITY:
Number of Moles of Solute Mixed Per Kg of Solvent.

m = Number of Moles of Solute /Mass of Solvent (IN Kg) OR

m =Mass of Solute (In Grams) / (Molecular Mass of Solute Mass of

Solvent (In Kg).

2.5.4 NORMALITY:
Gram equivalent weight of solute dissolved per liter of solution. N = gram
equivalent weight of solute /volume of solution in L.

2.5.5 ACID:
A compound / element / ion that tend to donate H + ion or accept a pair of
electron in an aqueous solution.
2.5.6 BASE:
A compound / element / ion that tends to donate OH - ion or a pair of electron
or accepts H+ Ion in Aqueous Solution.
2.5.7 PH:
Power H+ ions in solution (negative log of H+ ion molar concentration).
PH = - Log [H+] where [H+] =Molar concentration of H+
2.5.8 BUFFER:
A solution that tends to resist change in pH upon slight addition of acid or
base it is composite of strong acid weak base or strong base and weak acid.

2.5.9 ISOTONIC SOLUTION:

A solution with an osmotic pressure or concentration equal to that of a
specified other solution, usually taken to be within a cell. It therefore neither
gains nor losses water by osmosis.

2.5.10HYPOTONIC SOLUTION:
A solution with an osmotic pressures less than that of a specified other
solution. 2.5.11 HYPERTONIC
SOLUTION:
A solution with an osmotic pressure greater than that of a specified other
solution.

2.5.12 OSMOLALITY:

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The number of particles of solute per unit weight of water irrespective of the
size and nature of particles. Units are mmol/kg of water.

2.5.13 OSMOLARITY:
The number of particles of solute per liter of solution. Units are mmol/L

What are the clinical importance’s of the solution?

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2.6 METHODS FOR CONVERSION OF UNITS

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Conversion of mg/dl to mmol/L:

To convert mg/100ml to mg/L

- Multiply by 10
- Divide mg/L volume by it molecular weight
mg per 100ml x10
mmol/L =
mol wt

Conversion of mg % to meq/L

- multiply by 10

- Divide by equivalent weight

mg % x 10
meq/L =
eq.wt

Conversion of mmol/L to meq/L or vice versa.

mmol/L
meq/L = valency

mmol = meq/L X valency

If valency of the solute is 1, then

meq/L = mmol

Molarity:

Molarity: no. of moles of solute/1000ml solution

Normality: gm eq.wt of solute/1000ml of solution

Molality: no. of moles of solute/1000gm solvent

OBJECTIVE: To prepare 0.95 % NaCl solution (Normal Saline).

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REQUIREMENT:

1. Measuring cylinder
2. Beakers, Glass rod
3. Weighing Scale
4. Colorless bottle,
5. NaCl crystal & Water.

PROCEDURE:
1. Weigh 0.95g NaCl and transfer in the beaker.
2. Add water to mix it & make up to mark 100ml.

USES:
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OBJECTIVE: To prepare 5% Dextrose Solution.

REQUIREMENT:
1. Measuring cylinder

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 2. Beakers, Glass rod
3. Weighing Scale
4. Colorless bottle
5. Dextrose (glucose) & Water.

PROCEDURE:

1. Weigh 5g the Dextrose and transfer in the beaker.

2. Add water to mix it & make up to mark 100ml.

USES:

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PRACTI
CAL NO.3

PROPERTIES OF ORGANIC COMPOUNDS

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OBJECTIVES:

At the completion of this lab students will be able to:

1. Define organic compound.

2. List the types of organic compound.

3. Describe the uses of organic compound.

4. Describe the characteristic of organic compounds.

5. Differentiate between organic and inorganic compound.

3.1 INTRODUCTION OF ORGANIC COMPOUNDS

Organic Compounds:

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Organic compound is the study of the structure, properties, composition,
reactions, and preparation (by synthesis or by other means) of chemical
compounds that contain carbon. These compounds may contain any number
of other elements, including hydrogen, nitrogen, oxygen, the halogens as well
as phosphorus, silicon and sulfur. Because of their unique properties, multi-
carbon compounds exhibit extremely large variety and the range of
application of organic compounds is enormous. They form the basis of, or are
important constituents of many products (paints, plastics, food, explosives,
drugs, petrochemicals, to name but a few) and (apart from a very few
exceptions) they form the basis of all earthly life processes.

3.2 Types of Organic Compounds:

You can find details of the physical and chemical properties for each of these
types of compound:

A. Aliphatic compounds

These are compounds where the functional group is not attached directly to a
benzene ring.

 Alkanes . . .

 Alkenes . . .

 Halogenoalkanes (haloalkanes or alkyl halides) . . .

 Alcohols . . .

 Aldehydes and ketones . . .

 Carboxylic acids . . .

 Acyl chlorides (acid chlorides) . . .

 Acid anhydrides . . .

 Esters . . .

 Amides . . .

 Nitriles . . .

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  Amines . . .

 Amino acids and other biochemistry . . .

B. Aromatic compounds

These are compounds based on benzene rings. This isn't a complete list, but
shows only those compounds where there are important differences between
them and their aliphatic equivalents.

 Arenes (benzene and methylbenzene) . . .

 Halides (e.g. chlorobenzene) . . .

 Phenol . . .

 Phenyl amine (aniline) and diazonium compounds . . .

3.3 Uses of Organic Compounds:

There are several organic compounds that have different uses in chemical,
medical and industrial fields

A. Chemical Uses:
In chemical sector they are used as a subject of research so that most type of
compounds or elements can be discovered. They are also used as supportive
to other studies because of having a specific type of nature and property.

B. Medical Uses:

When it comes to medicinal uses then stable halocarbon organic compounds

are used for the purpose of anesthesia.

C. Industrial Uses:

They are also used in industries in the form of heal tolerant, metalworking
lubricants and inert synthetic oils.

3.4 Characteristic of Organic Compound:

1. Organic compounds are generally covalently bonded.

2. It allows for unique structures such as long carbon chains and rings.

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3. Organic compounds typically melt, boil, sublimate, or decompose
below 300 °C.

4. Neutral organic compounds tend to be less soluble in water compared

to many inorganic salts, with the exception of certain compounds such
as ionic organic compounds and low molecular weight alcohols and
carboxylic acids where hydrogen bonding occurs.

5. Organic compounds tend to dissolve in organic solvents which are

either pure substances like ether or ethyl alcohol, or mixtures, such as
the paraffinic solvents such as the various petroleum ethers and white
spirits.

6. A unique property of carbon in organic compounds is that its valency

does not always have to be taken up by atoms of other elements, and
when it is not, a condition termed unsaturation results.

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PRACTICAL NO 4

CARBOHYDRATES

OBJECTIVE

At the completion of this lab students will be able to:

1. Discuss the structure of mono & disaccharides with example.

2. Define & differentiate between reducing & non reducing sugar.

3. Discuss the significance of Benedicts test & Molish test.

4. Discuss the principle of Benedicts & Molish test.

5. Perform the Benedicts & Molish test.

6. Discuss the Nutritional values of Monosaccharide & Disaccharides.

7. Discuss the sources & availability of mono & disaccharides.

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4.1 Structure of Monosaccharides and Disaccharides

a. Monosaccharides

Monosaccharides are the simplest units of carbohydrates and the simplest

form of sugar. They are the building blocks of more complex carbohydrates
such as disaccharides and polysaccharides. Physically, they are usually
colorless, can dissolve in water, and have the appearance of a crystal-like
substance.

Structure of Monosaccharides

Monosaccharides are the simplest units of carbohydrates, they are made up

of carbon, hydrogen, and oxygen atoms. The following are some example of
monosaccharides

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26
 b. Disaccharides:

Disaccharides, the simplest polysaccharides, are the products of a

condensation reaction between two monosaccharides. Disaccharide is one of
four groups of Carbohydrates (monosaccharide, disaccharide, polysaccharide,
and oligosaccharide).
Example and structure of disaccharides:
The most common disaccharides are Sucrose, Lactose, and Maltose.

1. Sucrose is the sugar often found in the grocery store and is produced
by plants. It is a sugar derived from fructose and glucose. It is obtained
from cane as a transport form of carbohydrates.

Sucrose

2. Maltose is created by condensation reaction of the two glucoses,

forming a α-1,4-O-glycosidic linkage. It is the second member of an
important biochemical series of glucose chains. Maltose can be broken
down into two glucose molecules by hydrolysis.

Maltose
3. Lactulose is a synthetic (man-made) sugar that is not absorbed by the
body but is broken down in the colon into products that absorb water
into the colon, thus softening stools. Its primary use is to treat
constipation.

Lactulose

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4.2 Reducing and Non-reducing sugars
Any carbohydrate which is capable of being oxidized and causes the
reduction of other substances without having to be hydrolysed first is known
as reducing sugar, but those which are unable to be oxidised and do not
reduce other substances are known as non-reducing sugars. Generally; all the
free monosaccharides having free aldehyde or hydroxyl ketonic group are
capable of being oxidised. After being oxidised they cause the reduction of
the other substances and so known as reducing sugars.

Difference between Reducing and Non-reducing sugars

No Reducing Sugars Non-reducing Sugars

1 Such sugar bear a free aldehyde These sugars do not have such
(-CHO) or ketonic (-CO) group groups

2 Reducing sugars have the Non reducing sugar fail to reduce

capacity to reduce cupric ions of the cupric ions of Benedict's
Benedict's or Fehling solution to
cuprous ions. solution to cuprous ions.
3 Examples: Maltose, Examples: Sucrose, Trehalose
Lactose,Melibiose, Gellobiose,
Gentiobiose

4.3 Significance and principle of Benedicts test & Molish test:

Presence of carbohydrates can be confirmed qualitatively by several tests.

Due to the presence of different numbers of sugar units, specific
carbohydrates exhibit typical colour reactions that form the basis for their
identification. In this kit the following tests will be performed for the
qualitative analysis of carbohydrates:

1. Molisch’s Test:
All carbohydrates (monosaccharides, disaccharides and polysaccharides) give
a positive reaction for Molisch test. It is based on the dehydration of the
carbohydrate by Sulphuric acid to produce an aldehyde, which condenses
with two molecules of α-naphthol, resulting in appearance of a purple ring at
the interface.

2. Benedict’s test:
Benedict’s test is based upon the participation of the aldehyde and ketone
groups in a chemical reaction. In this procedure, Benedict’s reagent, which
contains a basic solution of cupric ions, is mixed with a sugar. The copper
ions will attract the electrons from the aldehyde or ketone group changing the
charged copper ions to a neutral metal. Copper ions are blue in color whereas

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copper metal is orange. Disaccharides will react with the Benedict’s reagent
only if it contains an exposed aldehyde or ketone group. This type of sugar is
known as a reducing sugar.

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4.4 Identification of Carbohydrates present in a given sample
MOLISCH’S TEST

POSITIVE NEGATIVE
Carbohydrate present carbohydrate absent

IODINE TEST

POSITIVE NEGATIVE
Polysaccharide Monosaccharide /
Blue - starch Disaccharide
Purple - Dextrin
Reddish- glycogen

BENEDICTS TEST

POSITIVE NEGATIVE
Reducing sugar non reducing disaccharides
Monosaccharide –e.g glucose e.g. sucrose.
Fructose C.T
Disaccharide – e.g. Maltose
Lactose Thymol blue test

BARFOED TEST

POSITIVE NEGATIVE
With in3minutes more than 3minutes
Monosaccharide present reducing disaccharide present

SELIWANOFF’S TEST

POSITIVE NEGATIVE
With in 30 sec after 30 sec
Ketosugar (fructose) aldo sugar(glucose)

C.T
OSAZONE TEST

Glucose Fructose Maltose lactose

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MOLISCH’S TEST

4.5 GENERAL TEST FOR CARBOHYDRATES

OBJECT:

To detect presence of carbohydrate in a given solution.

PRINCIPLE:
When carbohydrates are treated with strong acid like H2SO4 they undergo
dehydration and form hydroxy methyl furfurals with hexoses.When these
furfurals are treated with alpha-naphthol give purple colored compound.

REAGENTS:
1- Alpha naphthol solution
2- Concentrated H2 SO4

Procedure Observation Result

1. Take 5ml of given Purple colored ring

solution in a test tube appear
2. Add 2 drops of at junction of two
naphthol solution & solutions
Mix thoroughly
3. Add 3ml concentrated
H2 SO4 along the
side of test tube till
purple colored
complex ring appear.

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4.6 BENEDICT’S TEST FOR REDUCING SUGARS

Objective:
To detect the presence of reducing sugar in a given solution.

Principle:
Reducing sugar in strong alkaline medium form enediols. Enediols are
powerful reducing agents, reduces the metallic ions like cupric ions to
cuprous ions
Heating
Reducing Sugar Enediols form of reducing
Alkaline medium
sugar

CuSO4 Cu++ + So4--

Na2 CO3 + H2O NaOH
NaOH + Cu SO4 Cu (OH) 2
Cu (OH) 2 + Na citrate boilBoil cupric hydroxide- Na citrate
complex
Cu (OH) 2 + Sugar (enediols) CuO2 + Sugar acid

Reagents:
Benedict reagent (CuSO4, Na2CO3 and Na citrate)

Procedure Observation Result

1. Take 5ml of Red precipitates of

benedicts cuprous oxide appear.
qualitative reagent
in test tube & boil
it
2. Then add 8 drops
of given solution
3. Again boil for 2
minutes.

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PRACTICAL NO 5

PROTEINS

At the completion of this lab students will be able to:

1. Define Protein.

2. Classify Proteins on the basis of structure, nutrition & functions with

example.

3. Briefly describe the structure of proteins in terms of primary,

secondary, tertiary & quaternary structures.

4. Give the principle & significance of Biuret test & the coagulation test.

5. Perform the Biuret test & Heat coagulation test.

6. Give the nutritional values of protein in terms of their function &

proportion in diet.

7. List some important sources of dietary protein & specify their

nutritional class.

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5.1 Protein
The human body uses proteins for many things, including repairing and
building tissues, acting as enzymes, aiding the immune system, and serving
as hormones. Each of these important functions requires a slightly different
form of protein. In spite of their differences in structure, all proteins contain
the same basic sub-components.

Proteins are long chains of amino acids. Amino acids are the building blocks
of protein. In other words, amino acids are like the links in a chain. The chain
itself represents the protein molecule. Protein chains are then twisted and
folded together in specific ways to create certain molecules.

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This example shows our primary protein chain as it's twisted into a helical
shape, folded into a sheet, and then twisted all over again into an intricate
globular shape. In this case, the final product is a protein molecule known as
hemoglobin, which can be found in your blood.

5.2 Classification of Proteins

Proteins are important macromolecules of the cells, formed by the
polymerization of amino acids according to the sequence of genetic code in
the mRNA. Proteins are the mode of expression of the genetic information.
They perform a variety of duties in the cells such as they act as the structural
components of cells, enzymes, hormones, pigments, storage proteins and
some toxins in the cells. The proteins are classified into many categories
based on different criterions.

Proteins are classified based on the following three criterions:

1. Classification based on STRUCTURE of Protein
2. Classification based on COMPOSITION of Protein
3. Classification based on FUNCTIONS of Proteins

1. Classification of Proteins based on the Structure of Proteins:

Based on the structure, proteins are classified into 3 groups.

(A). Fibrous Proteins
(B). Globular Proteins
(C). Intermediate Proteins

(A). Fibrous Proteins:

 They are linear (long fibrous) in shape.
 Secondary structure is the most important functional structure of
fibrous proteins.
 Usually, these proteins do not have tertiary structures.
 Physically fibrous proteins are very tough and strong.
 They are insoluble in the water.
 Long parallel polypeptide chains cross linked at regular intervals.
 Fibrous proteins form long fibres or sheaths.
 Functions of fibrous proteins: perform the structural functions in the
cells.
 Examples of fibrous proteins: Collagen, Myosin, Silk and Keratin.

(B). Globular Proteins:

 Globular proteins are spherical or globular in shape.
 The polypeptide chain is tightly folded into spherical shapes.

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  Tertiary structure is the most important functional structure in globular
proteins.
 Physically they are soft than fibrous proteins.
 They are readily soluble in water.
 Most of the proteins in the cells belong to the category of globular
proteins.
 Functions: Form enzymes, antibodies and some hormones.
 Example: Insulin, Haemoglobin, DNA Polymerase and RNA
Polymerase

(C). Intermediate Proteins:

 Their structure is intermediate to linear and globular structures.
 They are short and more or less linear shaped proteins
 Unlike fibrous proteins, they are soluble in water.
 Function: blood clotting proteins
 Example: Fibrinogen

2. Classification of Proteins based on Composition:

Two broad categories of proteins according to its composition, they are:

(A). Simple Proteins
(B). Conjugated Proteins

(A). Simple Proteins

 Simple proteins composed of ONLY amino acids.
 Proteins may be fibrous or globular.
 They possess relatively simple structural organization.
 Example: Collagen, Myosin, Insulin, Keratin

(B). Conjugated Proteins

 Conjugated proteins are complex proteins.
 They contain one or more non-amino acid components.
 Here the protein part is tightly or loosely bound to one or more non-
protein part.
 The non-protein parts of these proteins are called prosthetic groups.
 The prosthetic group may be metal ions, carbohydrates, lipids,
phosphoric acids and nucleic acids.
 The prosthetic group is essential for the biological functions of these
proteins.
 Conjugated proteins are usually globular in shape and are soluble in
water.

36
  Most of the enzymes are conjugated proteins.

3. Classification of Protein based on Functions:

(A). Structural Proteins:

 Form the component of the connective tissue, bone, tendons, cartilage,
skin, feathers, nail, hairs and horn.
 Most of them are fibrous proteins and are insoluble in water.
 Example: Collagen, Keratin and Elastin.

(B). Enzymes:
 They are the biological catalysts.
 Enzymes reduce the activation energy of reactants and speed-up the
metabolic reactions in the cells.
 Most of them are globular conjugated proteins
 Example: DNA Polymerase, Nitrogenase, Lipase

(c). Hormones:
 They include the proteinaceous hormones in the cells.
 Example: Insulin, Glucagon, ACH

(D). Respiratory Pigments

 They are coloured proteins
 All of them are conjugated proteins and they contain pigments
(chrome) as their prosthetic group.
 Example: Haemoglobin, Myoglobin

(E). Transport Proteins

 They transport the materials in the cells
 They form channels in the plasma membrane
 They also form one of the components of blood and lymph in animals.
 Example: Serum albumin

5.3 structure of proteins in terms of primary, secondary, tertiary &

quaternary structures

37
A. Primary Structure
The amino acid sequence of a protein is
encoded in DNA. Proteins are
synthesized by a series of steps called
transcription and translation. Often, post-
translational modifications, such as
glycosylation or phosphorylation, occur
which are necessary for the biological
function of the protein. While the amino
acid sequence makes up the primary
structure of the protein

B. Secondary Structure
Stretches or strands of proteins or
peptides have distinct characteristic local
structural conformations or secondary
structure, dependent on hydrogen
bonding. The two main types of
secondary structure are the α-helix and
the ß-sheet.

C. Tertiary Structure
The overall three-dimensional shape of an entire protein molecule is
the tertiary structure. The protein molecule will bend and twist in such a way
as to achieve maximum stability or lowest energy state. Although the three-
dimensional shape of a protein may seem irregular and random, it is
fashioned by many stabilizing forces due to bonding interactions between the
side-chain groups of the amino acids.

D. Quaternary Structure
Many proteins are made up of multiple polypeptide chains, often referred to
as protein subunits. These subunits may be the same (as in a homodimer) or
different (as in a heterodimer). The quaternary structure refers to how these
protein subunits interact with each other and arrange themselves to form a
larger aggregate protein complex. 

38
5.4 SCHEME FOR THE DETECTION OF PROTEINS

A. GENERAL TEST FOR DETECTION OF PROTEIN

Biuret test Lead acetate test Sulphosalicyclic acid test

B.TEST FOR AMINO ACID

Xanthoproteic test Hopkins’s Cole test Millon’s test Sulphur test

C. IDENTIFICATION OF INDIVIDUAL PROTEINS

Acetic acid test

White precipitate
Positive (Casein) No precipitate
Negative (Albumin, globulin,
C.T.
Gelatin & peptones) C.T.

Heat coagulation test

Neumann test

Salt saturation test Coagulation formed No Coagulation

(Albumin & globulin) May be Gelatin &
peptones
½ saturation test (with NH4SO4 salt) C.T.

Test Positive for Globulin Color Test

Negative

C.T.
Test for Albumin
Full saturation test (withNH4SO4 salt)
C.T.

Negative Biuret test Negative Biuret test

(With filtrate) (With filtrate)

39
 5.5 GENERAL TEST FOR DETECTION OF PROTEINS

1. BIURET TEST

Objective:
To detect the presence of protein in given sample
Principle:
Peptide linkages present in protein molecule react with cupric ion in alkaline
medium provided by (NaOH) give purple color complex.
Reagents:
1- Sodium hydroxide (5% solution) 2- Copper sulphate (1%
solution)

Procedure Observation Inference

1. Take a test tube. Add Purple color It shows

3ml of NaOH in the appears in the
presence of
test tube. test tube.
2. Add 0.5ml of protein.
copper sulphate
solution. Mix the
content
thoroughly.
3. Add 3.5ml of the
given solution.
4. Note down the color of
the content in the test
tube.

40
5.6 HEAT COAGULATION TEST (FOR ALBUMIN & GLOBULIN)

Objective:

To detect the presence of albumin and globulin in given solution.

Principle:

When heat coagulable proteins are heated at their isoelectric PH they undergo
coagulation.

Procedure:

- Take 5ml of given solution in a clean test tube.

- Incline the test tube slightly and heat upper portion of solution (lower
portion will act as on control for comparison).

Observation:

A dense clot will be formed in upper part of solution.

Inference:

Albumin & Globulin are present.

5.7 Sources of protein

Some sources of dietary protein include:

 Lean meat, poultry and fish
 Eggs
 Dairy products like milk, yoghurt and cheese
 Seeds and nuts
 Beans and legumes (such as lentils and chickpeas)
 Soy products like tofu
 Some grain and cereal-based products are also sources of protein, but
are generally not as high in protein as meat and meat alternative
products.

41
PRACTICAL NO 6

LIPIDS

OBJECTIVE

At the completion of this lab students will be able to:

1. Define lipids.

2. Give classification of lipids.

3. Discuss the factor which affects the solubility of lipids.

4. Detect the relative solubility of lipids in various polar solvents.

5. Define emulsification.

6. Give the principle of emulsification test.

7. Perform emulsification test.

42
6.1 Lipid
Lipids are any of a large group of organic compounds that are oily to the
touch and insoluble in water. Lipids include fatty acids, oils, waxes, sterols,
and triglycerides. They are a source of stored energy and are a component of
cell membranes.

6.2 Classification of Lipids

Lipids can be classified into two major classes:

 Nonsaponifiable lipids, and

 Saponifiable lipids.

a. Nonsaponifiable Lipids
A nonsaponifiable lipid cannot be broken up into smaller molecules by
hydrolysis, which includes triglycerides, waxes, phospholipids, and
sphingolipids.

b. Saponifiable Lipids
A saponifiable lipid contains one or more ester groups allowing it to undergo
hydrolysis in the presence of an acid, base, or enzymes. Nonsaponifiable
lipids include steroids, prostaglandins, and terpenes.
Each of these categories can be further broken down into non-polar and polar
lipids.
Nonpolar lipids, such as triglycerides, are used for energy storage and fuel.
Polar lipids, which can form a barrier with an external water environment, are
used in membranes. Polar lipids include glycerophospholipids and
sphingolipids.
Fatty acids are important components of all of these lipids.

6.3 Types of Lipids

Within these two major classes of lipids, there are several specific types of
lipids important to live, including fatty acids, triglycerides,
glycerophospholipids, sphingolipids, and steroids. These are broadly
classified as simple lipids and complex lipids.

1. Simple Lipids
Esters of fatty acids with various alcohols.

1. Fats: Esters of fatty acids with glycerol. Oils are fats in the liquid
state.

43
 2. Waxes: Esters of fatty acids with higher molecular weight monohydric
alcohols

2. Complex Lipids
Esters of fatty acids containing groups in addition to alcohol and a fatty acid.

1. Phospholipids: Lipids containing, in addition to fatty acids and

alcohol, a phosphoric acid residue. They frequently have nitrogen-
containing bases and other substituents, eg, in glycerophospholipids
the alcohol is glycerol and in sphingophospholipids the alcohol is
sphingosine.

2. Glycolipids (glycosphingolipids): Lipids containing a fatty acid,

sphingosine, and carbohydrate.

3. Other complex lipids: Lipids such as sulfolipids and amino lipids.

Lipoproteins may also be placed in this category

3. Precursor and derived lipids

These include fatty acids, glycerol, steroids, other alcohols, fatty aldehydes,
and ketone bodies, hydrocarbons, lipid soluble vitamins, and hormones.
Because they are uncharged, acylglycerols (glycerides), cholesterol, and
cholesteryl esters are termed neutral lipids.
Some of the different types of lipids are described below:

a. Fatty Acids
Fatty acids are carboxylic acids (or organic acid), often with long aliphatic
tails (long chains), either saturated or unsaturated.

i. Saturated fatty acids

When a fatty acid is saturated it is an indication that there are no carbon-
carbon double bonds. The saturated fatty acids have higher melting points
than unsaturated acids of the corresponding size due to their ability to pack
their molecules together thus leading to a straight rod-like shape.

ii. Unsaturated fatty acids

If a fatty acid has more than one double bond then this is an indication that it
is an unsaturated fatty acid.
“Most naturally occurring fatty acids contain an even number of carbon
atoms and are unbranched.”
Unsaturated fatty acids, on the other hand, have a cis-double bond(s) that
create a kink in their structure which doesn’t allow them to group their
molecules in straight rod-like shape.

44
Role of Fats
Fats play several major roles in our body. Some of the important roles of fats
are mentioned below:

 Fats in correct amounts are necessary for the proper functioning of our
body.
 Many fat-soluble vitamins need to be associated with fats in order to
be effectively absorbed by the body.
 They also provide insulation to the body.
 They are an efficient way to store energy for longer periods.

b. Waxes
Waxes are “esters” (an organic compound made by replacing the hydrogen
with acid by an alkyl or another organic group) formed from long-chain
carboxylic acids and long-alcohols.
Waxes are seen all over in nature. The leaves and fruits of many plants have
waxy coatings, which may protect them from dehydration and small
predators.
The feathers of birds and the fur of some animals have similar coatings which
serve as a water repellent.
Carnauba wax is valued for its toughness and water resistance(great for car
wax).

c. Phospholipids
Membranes are chiefly made of phospholipids which are
Phosphoacylglycerols.
Triacylglycerols and phosphoacylglycerols are similar however the terminal
OH group of the phosphoacylglycerol is esterified with phosphoric acid
instead of fatty acid which leads to the formation of phosphatidic acid.
The name phospholipid comes from the fact that phosphoacylglycerols are
lipids that contain a phosphate group.

d. Steroids
The chemical messengers in our bodies are known as hormones which are
organic compounds synthesized in glands and delivered by the bloodstream
to certain tissues in order to stimulate or inhibit the desired process.
Steroids are a type of hormone which is usually recognized by their
tetracyclic skeleton, consisting of three fused six-membered and one five-
membered ring, as shown in the diagram above. The four rings are designated

45
as A, B, C & D as noted in blue, and the numbers in red represent the
carbons.

e. Cholesterol
Cholesterol is an important lipid found in the cell membrane. It is a sterol,
which means that cholesterol is a combination of steroid and alcohol.
It is an important component of cell membranes and is also the basis for the
synthesis of other steroids, including the sex hormones estradiol and
testosterone, as well as other steroids such as cortisone and vitamin D.

6.4 FACTORS AFFECTING LIPID SOLUBILITY 

i) Chemical nature of the molecule

If a molecule is a lipid, it is necessarily lipid soluble and will readily pass

through the lipid component of membranes.  That means that, if these
substances get into the body, they are not restricted to any one location... they
can diffuse to locations anywhere in the body.

Some examples of biological lipids include all of the steroid hormones and
all of the lipid soluble vitamins.  Note that all of these molecules are built
around a cholesterol "nucleus," so they are all structurally similar.

ii) Atomic or molecular formula weight

The smaller a molecule is, the more likely it is to fit through a space in the
membrane.  Thus, the smaller the molecular weight of a substance, the more
lipid soluble it is Neither of the following substances are lipids and neither
will diffuse very quickly through a membrane... Nonetheless, molecule B is
smaller and will diffuse through a membrane before molecule A will.

iii) Valence or charge and sphere of hydration

Any substance which is charged (eg. any ion) will attract water molecules. 
Water molecules carry no charge but act as if they do.  The H+ containing
end of H2O acts as if it is positively charged and the O2-containing end acts
as if it is negatively charged.  Thus, the H+ bearing end of H2O is attracted to
anions and the O2-bearing end is attracted to cations.  Molecules of H2O will
literally form a "ball of water" or "sphere of hydration" that will diffuse along
with the ion they surround.  This ball of water makes the ion physically
bigger and, therefore, less lipid soluble.  Typically, the greater the valence,
the greater the size of the sphere of hydration.

46
iv) Charge density and sphere of hydration

Charge density is a difficult concept to grasp.  Let's suppose we have 2 ions

with the same charge, for example, Li+ and K+.  Note that the atomic mass of
Li+ (7) is much less than that of K+ (39).  That means that Li+ is smaller
than K+.  Nonetheless, K+ and Li+ have the same charge.  Charges move. 
As charges move around, sometimes they are close to the outside of the ion
and sometimes they are closer to the center of the ion.  Whenever a charge is
close to the outside of the atom, it will attract more H2O molecules into its
sphere of hydration.  Note (below) tha Li+ is smaller than  K+.  Thus, on
average, the charge on Li+ is almost always closer to the outside of the
Li+ ion than is the charge on K+.  Because the charge on Li+ is closer to the
outside of Li+ more of the time, Li+ ends up having a bigger sphere of
hydration than K+.  That means that, even though K+ is bigger than Li+,
Li+ is less lipid soluble than K+ because Li+ has a bigger sphere of hydration
around itself. 

v) Prevailing concentration gradient (slope or steepness of the gradient)

If it is possible for a substance to get across a membrane, the speed at which

it diffuses through the membrane will be directly related to the concentration
gradient.  Solutes diffuse from an area of high solute concentration to an area
of low solute concentration.  If glucose was higher outside cells than inside
cells, which direction would the glucose concentration gradient be driving the
glucose?  That's right, into the cell.  Suppose we had a child with diabetes
mellitus I and we wanted to reduce their plasma glucose level.  We ask them
to exercise more frequently.  When they exercise, what happens to the
glucose inside their cells.  That's right, they burn it!  If they burn glucose
inside their cells, what are they doing to the glucose concentration gradient? 
That's right, by reducing the amount of glucose inside their cells, they are
making the glucose gradient steeper so that glucose will enter their cells more
easily, in turn reducing their plasma glucose level.

vii) route of administration

The route of administration of a drug does not really affect the lipid solubility
of a drug.  The route of administration does affect how long it will be until
the drug appears in the bloodstream.  A drug or nutrient is not considered to
have entered the body until it has been absorbed into the bloodstream.  If a
drug is injected directly into the bloodstream, it has instantly entered the
body.  If a drug is injected into a muscle, taken orally or applied topically to
the skin, it may take longer to be absorbed into the bloodstream.

47
 6.5 EMULSIFICATION TEST

Objective:
To demonstrate the process of emulsification of lipids in given sample

Principle: -
In the presence of emulsifying agent like bile salt, Na2CO3 and soap the
surface tension of H2O is lowered due to which lipid molecule are broken
down into small droplets.

Reagents: -
Bile salt solution 5%

Procedure: -
- Take 2 test tubes and mark them A and B
In test tubes A– take 1ml of given sample of oil and add 1 ml water.
In test tubes B–take 1ml of given sample of oil and add 1ml of bile
salt solution.
- Shake them well and allow to standing.

Observation: -
In test tube A- two separate layers are seen.
In test tube B- oil is broken down into small particle.

Inference: -
Test tube A- no emulsification is seen.
Test tube B- emulsification of oil in presence of bile salt.

48
PRACTICAL NO 7.

ENZYME KINETICS

OBJECTIVE

At the completion of this lab students will be able to:

1. Define enzymes and enlist Its classification According to IUB

2. Enumerate the factors regulating the activity of an enzyme.

3. Perform the test to determine the effect pH on the activity of salivary

amylase.

4. Perform the test to determine the effect temperature on the activity of

salivary amylase.

49
7.1.A. Definition of Enzymes
Enzymes are Proteins that speeds up the rate of a chemical reaction in a
living organism. An enzyme acts as catalyst for specific chemical reactions,
converting a specific set of reactants called substrates into specific products.
7.1.B. Classification of Enzymes
Enzymes are broadly of two types:
1.) Intracellular or Endoenzymes:-
They are functional within cells where they are synthesized.
2.)Extracellular or Exoenzymes:
These enzymes are active outside the cells.
Example: Digestive enzymes like Pepsin, Trypsin, Amylase.
Classification of Enzymes by IUB System,
Enzymes are classified by complex system, suggested by commission on
enzymes of International Union of Biochemistry (IUB). Based on their action
they are divided into 6 major classes. Each enzyme is assigned a 4 Digit code
number.
(1) Oxido-Reductases:
Enzymes in this class are involved in Oxidation-Reduction reactions.
Example: Alcohol Dehydrogenase.
(2)Transferases:
Enzymes that catalyze transfer of Functional groups are called as
Transferases.
Example: Phosphorylases
(3)Hydrolases:
These are enzymes that bring about hydrolysis of various compounds.
Example: Lipase
(4)Lyases:
Enzymes specialized in addition or removal of water.
Example: Aldolase
(5)Isomerases:
Enzymes involved in all isomerization reactions.
Example: Phosphotriose Isomerase.
(6)Ligases:
Enzymes catalyzing synthetic reactions where two molecules are joined
together and ATP are used.
Example: Succinate thiokinase

7.2 Factors effecting regulation of enzyme function

50
 1. Enzyme concentration
2. Substrate concentration
3. Temperature
4. PH
5. Salinity
6. Activators
7. Inhibitors

1. Enzyme concentration
To understand the effect of increasing the enzyme concentration upon the
reaction rate, the substrate must be present in an excess amount; i.e., the
reaction must be independent of the substrate concentration. Any change in
the amount of product formed over a specified period of time will be
dependent upon the level of enzyme present.

2. Substrate concentration
Substrate concentration is the amount of substratepresent that can be turned
into product and is most commonly measured in molarity. Increasing Substrat
Concentration increases the rate of reaction. This is because
more substrate molecules will be colliding with enzyme molecules, so more
product will be formed.

3. Temperature
Like most chemical reactions, the rate of an enzyme-catalyzed reaction
increases as the temperature is raised. A ten degree Centigrade rise in
temperature will increase the activity of most enzymes by 50 to 100%.
Variations in reaction temperature as small as 1 or 2 degrees may introduce
changes of 10 to 20% in the results. In the case of enzymatic reactions, this is
complicated by the fact that many enzymes are adversely affected by high
temperatures.

4. PH

Enzymes are affected by changes in pH. The most favorable pH value - the
point where the enzyme is most active - is known as the optimum pH. The
optimum pH value is vary greatly from one enzyme to another.

5. Salinity

51
High concentrations of any salt decrease water activity, and also that may
reduceenzyme activity. If the salt concentration is close to zero, the charged
amino acid side chains of the enzyme molecules will attract to each other.
The enzyme will denature and form an inactive precipitate.

6. Activators

Enzyme activators are molecules that bind to enzymes and increase

their activity. ... In some cases, when a substrate binds to one catalytic
subunit of an enzyme, this can trigger an increase in the substrate affinity as
well as catalytic activity in theenzyme's other subunits, and thus the substrate
acts as an activator.

7. Inhibitors

Enzyme inhibitors are substances which alter the catalytic action of the
enzyme and consequently slow down, or in some cases, stop catalysis.

52
 Objective:

To determine the effect of temperature & pH on activity of salivary amylase.

Procedure:

(1) Take a mouthful of water. Keep rolling in mouth as long as you can.
Then spit out in a beaker, add 10ml of distilled water to it stock solution
of amylase enzyme.

(2) Take 6ml of stock solution in two test tubes & add 3ml of 1% starch
solution in each test tube. Perform Iodine test in one & Benedicts test
in other test tube.

(3) For pH: - Take 2 set of test tubes as A& B. In test tubes A1 & A2 Add
0.1% HCl &3ml of stock solution & 3ml of 1% of starch solution,
perform Iodine &Benedicts test. In test tube B1 & B2take 3ml 0.1%
NaOH & 3ml of stock solution & 3ml of starch solution, perform Iodine
& Benedicts test.

(4) For temperature: - Take 3sets of test tubes C.D.E. In test tubes C1&
C2 add 3ml of stock solution &3ml of starch solution. Keep it at room
temp. For 5minutes then perform Iodine & Benedicts test. In test tubes
D1&D2add 3ml of stock solution & 3mlof starch solution & keep it in
water bath at 4oc̊for 15minutes then perform Iodine & Benedicts test. In
test tubes E1&E2 add 3mlof stock solution & 3mlof starch solution &
boil it for 5minuets then perform iodine &Benedicts test.

7.3 OBSERVATION FOR THE EFFECT OF PH:

Test
Time Substrat pH Temp. Iodine Benedi Observation Inference
Tubes e(1%Sta Test cts
(Stock rch) Test
sol.)
3ml
A1 3ml 3ml.1 Room 2-5drops --------
%HCl temp.
A2 3ml 3ml.1 Room ---------- 5ml
%HCl temp.
B1 3ml 3ml.1 Room 2-5drops ---------
%NaO temp.
H
B2 3ml 3ml.1 Room ---------- 5ml
%NaO temp.
H

53
 7.4 OBSERVATION FOR THE EFFECT OF TEMPERATURE:

Test
Time Substrat pH Temp. Iodine Benedicts Observation Inference
Tubes e(1%Sta Test Test
(Stock rch)
sol.)
3ml
C1 3ml -------- Room 2-5drops ---------
---- temp
C2 3ml -------- Room 5ml
---- temp
D1 3ml -------- 37c 2-5drops ----------
----
D2 3ml -------- 37c --------- 5ml
----
E1 3ml -------- Boiling 2-5drops
- temp.
E2 3ml -------- Boiling 5ml
- temp

RESULT:-

______________________________________________________________
______________________________________________________________
______________________________________________________________
__________________________________________________________
_____________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________
______________________________________________________________

54
Practical No.8

INTRODUCTION OF BODY MASS INDEX (BMI)

OBJECTIVE

At the completion of this lab students will be able to:

1. Define BMI.

2. What are the measure causes of obesity?

3. Give the principle of BMI.

4. What are the factors that minimize the risk of obesity?

5. What are the other methods for estimating body composition or body

fats?

6. Briefly describe the consequences of obesity.

55
8.1 INTRODUCTION OF BODY MASS INDEX (BMI)

Body weight is the most frequently used method of an assessing nutritional

status it should be used in combination with measurement of body height to
establish whether a person is under weight or over weight.
The body mass index uses height and weight to determine healthy weight. It
is calculated by:

BMI (kg/m²) = Weight (in kg)

Height (in m²)

Body weight reflects both lean body mass and adipose tissues and cannot be
used as a method for describing body composition or the percentage of fat
tissue present. During physical training, body fat usually decreases and lean
body mass increases.
Obesity is defined as a condition characterized by excess body fat. Clinically,
obesity and overweight have been defined in terms of BMI. Over weight and
obesity have become national health problems, increasing the risk of
hypertension, hyperlipidemia. Type II diabetes, coronary heart disease and
other health problems.
However the nutritional status can be assessed by evaluating the persons
dietary intake, taking anthropometric measurement (example BMI)
performing a physical examination and conducting laboratory test.

8.2 CLASSIFICATION OF OVER WEIGHT AND OBESITY BY BMI

BMI (kg/m²) OBESITY CLASS

Under weight < 18.5
Normal 18.5-24.9
Over weight 25.0-29.9
Obese 30.0-34.9 I
35.0-39.9 II
Extremely obese > 40 III

56
8.3 Most common causes of obesity
 Physical inactivity
 Overeating
 Genetics
 A diet high in simple carbohydrates
 Frequency of eating
 Medications
 Psychological factors
 Diseases such as hypothyroidism, insulin resistance, polycystic ovary
syndrome, and Cushing's syndrome are also contributors to obesity.

8.4 Factors that minimize the risk of obesity

Many of the strategies that produce successful weight loss and maintenance
will help prevent obesity. Improving your eating habits and increasing
physical activity play a vital role in preventing obesity. Things you can do
include:
 Eat five to six servings of fruits and vegetables daily. A vegetable
serving is one cup of raw vegetables or one-half cup of cooked
vegetables or vegetable juice. A fruit serving is one piece of small to
medium fresh fruit, one-half cup of canned or fresh fruit or fruit juice,
or one-fourth cup of dried fruit.
 Choose whole grain foods such as brown rice and whole wheat bread.
Avoid highly processed foods made with refined white sugar, flour and
saturated fat.
 Weigh and measure food to gain an understanding of portion sizes.
Balance the food "checkbook." Eating more calories than you burn for
energy will lead to weight gain.
 Weigh yourself regularly.
 Avoid foods that are high in "energy density" or that have a lot of
calories in a small amount of food.
 Crack a sweat: accumulate at least 30 minutes or more of moderate-
intensity activity on most, or preferably, all days of the week.
Examples include walking a 15-minute mile, or weeding and hoeing
the garden.
 Make opportunities during the day for even just 10 or 15 minutes of
some calorie-burning activity, such as walking around the block or up
and down a few flights of stairs at work. Again, every little bit helps. 

57
8.5 Consequences of obesity:

If you are obese, severely obese, or morbidly obese, you may have:
Major health risks
 Shorter Life Expectancy
 Compared to people of normal weight, obese people have a 50% to
100% increased risk of dying prematurely
 Obese people have more risk for:
 Diabetes (type 2)
 Joint problems (e.g., arthritis)
 High blood pressure
 Heart disease
 Gallbladder problems
 Certain types of cancer (breast, uterine, colon)
 Digestive disorders (e.g., gastroesophageal reflux disease)
 Breathing difficulties (e.g., sleep apnea, asthma)
 Psychological problems such as depression
 Problems with fertility and pregnancy
 Urinary Incontinence
Difficulties with day-to-day living
 Normal tasks become harder when you are obese, as movement is
more difficult
 You tend to tire more quickly and you find yourself short of breath
 Public transport seats, telephone booths, and cars may be too small for
you
 You may find it difficult to maintain personal hygiene

58
8.6 OBJECTIVE:
To calculate BMI (body mass index) and correlate it to ideal body weight.

A. REQUIREMENTS:
Weighing scale, measuring tape.

B. PRINCIPLE:

The Body Mass Index (BMI) is a measurement tool that compares your
height to your weight and gives you an indication of whether you are
overweight, underweight or at a healthy weight for your height.

C. PROCEDURE:

1) Collect a few household materials to find your Body Mass Index

(BMI): your bathroom scale; a yardstick; a pencil and paper and
calculator.

2) Weigh yourself on your bathroom scale. (The following formula is

calculated in pounds.)

3) Measure your height in inches using the yardstick.

4) Use the pencil, paper and calculator to perform the following BMI
calculation.

5) Take your height in inches and square the number (i.e. multiply the
number of inches by the same number of inches).

6) Divide your weight in pounds by the second figure (your height in

inches squared).

7) Multiply that answer by 703. The answer is your Body Mass Index.

8) Judge your personal BMI result using the above chart.

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RESULT:
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PRACTICAL NO. 9

BASAL METABOLIC RATE (BMR)

OBJECTIVES:

At the completion of this lab students will be able to:

1. Define BMR.

2. Why it’s important to know your BMR?

3. Calculate your BMR.

4. Which equation is used to calculate the BMR?

5. List the factors which affect the BMR.

6. Discuss result.

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 INTRODUCTION OF BASAL METABOLIC RATE

9.1 BASAL METABOLIC RATE (BMR)

Basal metabolic rate (BMR) or resting metabolic rate (RMR) is the amount of
energy expended while at rest in a neutrally temperate environment, in the
post-absorptive state (meaning that the digestive system is inactive, which
requires about twelve hours of fasting in humans). The release of energy in
this state is sufficient only for the functioning of the vital organs, such as the
heart, lungs, brain and the rest of the nervous system, liver, kidneys, sex
organs, muscles and skin. BMR decreases with age and with the loss of lean
body mass. Increasing muscle mass will also increases BMR.

BMR is measured under very restrictive circumstances when a person is

awake. An accurate BMR measurement requires that the person's sympathetic
nervous system not be stimulated, such a condition whose prerequisite is that
of complete rest. A more common and closely related measurement, used
under less strict conditions, is resting metabolic rate (RMR).

The most well-known way to calculate your BMR is the Harris Benedict
equation. This formula accounts for the factors of height, weight, age, and
sex to calculate your basal metabolic rate (BMR). This calculation is more
accurate than determining calorie needs based only on total bodyweight.
However, the Harris Benedict equation does not take lean body mass into
consideration. Calculations will be accurate in all but the extremely muscular
and the extremely obese.

Nutrition and dietary considerations

Basal metabolic rate is usually by far the largest component of total caloric
expenditure. However, the Harris-Benedict equations are only approximate
and variation in BMR (reflecting varying body composition), in physical
activity levels, and in energy expended in thermogenesis make it difficult to
estimate the dietary consumption any particular individual needs in order to
maintain body weight. 2000 calories is often quoted but is no more than a
guideline.

9.2 Factors Affecting BMR:

BMR is the largest factor in determining overall metabolic rate and how
many calories you need to maintain, lose or gain weight. BMR is determined
by a combination of genetic and environmental factors, as follows:

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 Genetics. Some people are born with faster metabolisms; some with
slower metabolisms.

 Gender. Men have a greater muscle mass and a lower body fat
percentage. This means they have a higher basal metabolic rate.

 Age. BMR reduces with age. After 20 years, it drops about 2 per cent,
per decade.

 Weight. The heavier your weight, the higher your BMR. Example: the
metabolic rate of obese women is 25 percent higher than the metabolic
rate of thin women.

 Body Surface Area. This is a reflection of your height and weight.

The greater your Body Surface Area factor, the higher your BMR. Tall,
thin people have higher BMRs. If you compare a tall person with a
short person of equal weight, then if they both follow a diet calorie-
controlled to maintain the weight of the taller person, the shorter
person may gain up to 15 pounds in a year.

 Body Fat Percentage. The lower your body fat percentage, the higher
your BMR. The lower body fat percentage in the male body is one
reason why men generally have a 10-15% faster BMR than women.

 Diet. Starvation or serious abrupt calorie-reduction can dramatically

reduce BMR by up to 30 percent. Restrictive low-calorie weight loss
diets may cause your BMR to drop as much as 20%.

 Body Temperature/Health. For every increase of 0.5C in internal

temperature of the body, the BMR increases by about 7 percent. The
chemical reactions in the body actually occur more quickly at higher
temperatures. So a patient with a fever of 42C (about 4C above
normal) would have an increase of about 50 percent in BMR.

 External temperature. Temperature outside the body also affects

basal metabolic rate. Exposure to cold temperature causes an increase
in the BMR, so as to create the extra heat needed to maintain the
body's internal temperature. A short exposure to hot temperature has
little effect on the body's metabolism as it is compensated mainly by
increased heat loss. But prolonged exposure to heat can raise BMR.

 Glands. Thyroxin (produced by the thyroid gland) is a key BMR-

regulator which speeds up the metabolic activity of the body. The more
thyroxin produced, the higher the BMR. If too much thyroxin is
produced (a condition known as thyrotoxicosis) BMR can actually
double. If too little thyroxin is produced (myxoedema) BMR may
shrink to 30-40 percent of normal. Like thyroxin, adrenaline also
increases the BMR but to a lesser extent.

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  Exercise. Physical exercise not only influences body weight by
burning calories, it also helps raise your BMR by building extra lean
tissue. (Lean tissue is more metabolically demanding than fat tissue.)
So you burn more calories even when sleeping.

9.3 OBJECTIVE:

To calculate the Basal Metabolic Rate (BMR).

A. REQUIREMENTS:

Measuring scale, weighing scale.

B. FORMULA:

For men: BMR = 66 + (13.7 X wt in kg) + (5 X ht in cm) – (6.8 X age in

years)

For women: BMR = 655 + (9.6 X wt in kg) + (1.8 X ht in cm) – (4.7 X age in
years)

Note: Please keep in mind when calculating your BMR that 1 inch is equal to
2.54 cm and 1kg is equal to 2.2lbs. Using the metric equivalents are
necessary to arrive at the right total calculation.

C. PROCEDURE:

1. Step 1
Weigh yourself. Begin with your current body weight. You may be
tempted to
cheat on this number, but don't. Using accurate information benefits
you in the
long run.
2. Step 2
Measure how tall you are in your stocking feet.
3. Step 3
Determine your activity level. On a scale from sedentary to very
active, determine where it lies.
4. Step 4
Know your age. If you have been lying about your age for years, now
is the time to get real.
5. Step 5
Calculate your BMR. You can either use the formula of adding your weight
4.4 times to your height in inches times 4.7 and taking the sum of that
number. Thenmultiply it by your age in years times 4.7, then subtract that

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answer from your weight and age, and then add 655. Or, search for a basil
metabolic rate calculator on the Internet, and plug in your numbers.

RESULT:
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APPENDIX -I
(Some Basic Conversions and Calculations)

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1. Conversions

o 1 gram (g) = 1000 milligrams (mg)

o 1 milligram (mg) = 1000 microgram (ug)
o 1 microgram (ug) = 1000 nanogram (ng)
o 1 nanogram (ng) = 1000 picogram (pg)
o 1 gram = 10e12 picograms
o 1 mole = 1000 millimoles (mmole)
o 1 millimole (mmole ) = 1000 micromoles (umole)
o 1 micromole (umole ) = 1000 nanomoles (nmole)
o 1 nanomole (nmole ) = 1000 picomoles (pmole)
o 1 mole = 10e12 picomoles
o A 1 micromolar solution (1uM) = 1 picomoles per microliter
(1pmoles/ul):
o 1uM = 10e-6 moles/liter x 10[-6] liters/ul = 10e-12 moles/ul = 1
pmole/ul

2. Percent solutions

A 1% solution is defined as 1 gram per 100 ml volume. To find the

weight (in grams) needed for a particular volume (in ml) of solution,
convert the desired percentage to a decimal (divide by 100) and
multiply by the ml final volume desired.

Example 1: To make 4 liters of 10% SDS:

10%

------ X 4000 ml = 400 g SDS

100

In practice, dissolve 400 g SDS in about 3500 ml dH2O. After the

powder is in solution, the volume of the solution is adjusted with
dH2O to a final volume of 4 liters.

Example 2: To make 250 ml of 0.8% TA agarose:

0.8%

------ X 250 ml = 2 g agarose required.

100

In practice (when the % solution is very low) we do not worry about

the volume displacement and add 250 ml TA buffer to 2 g agarose.

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3. Concentration calculations

Concentration required x total volume

Volume of stock required = ---------------------------------------------------
Stock concentration

Example 1:

To make 10 ml of a 15 mM solution from a 500 mM stock:

15 mM x 10 ml
Volume of stock required = ------------------------ = 0.3 ml or 300 ul
500 mM

Example 2:

To make 10 ml of a 0.1% solution from a 20% stock:

0.1% x 10 ml
Volume of stock required = ---------------------- = 0.05 ml or 50 ul
20%

Length

The standard unit of length in the metric system is the meter. Other units of
length and their equivalents in meters are as follows:
1 millimeter = 0.001 meter
1 centimeter = 0.01 meter
1 decimeter = 0.1 meter
1 kilometer = 1000 meters

We abbreviate these lengths as follows:

1 millimeter = 1 mm
1 centimeter = 1 cm
1 meter = 1 m
1 decimeter = 1 dm
1 kilometer = 1 km

For reference, 1 meter is a little longer than 1 yard or 3 feet. It is about half
the height of a very tall adult. A centimeter is nearly the diameter of a dime, a
little less than half an inch. A millimeter is about the thickness of a dime.

Volume

The standard unit of volume in the metric system is the liter. One liter is
equal to 1000 cubic centimeters in volume. Other units of volume and their
equivalents in liters are as follows:
1 milliliter = 0.001 liter

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1 centiliter = 0.01 liter
1 deciliter = 0.1 liter
1 kiloliter = 1000 liters

From these units, we see that 1000 milliliters equal 1 liter; so 1 milliliter
equals 1 cubic centimeter in volume. We abbreviate these volumes as
follows:
1 milliliter = 1 ml
1 centiliter = 1 cl
1 deciliter = 1 dl
1 liter = 1 l
1 kiloliter = 1 kl

For reference, 1 liter is a little more than 1 quart. One teaspoon equals about
5 milliliters.

Mass

The standard unit of mass in the metric system is the gram. Other units of
mass and their equivalents in grams are as follows:
1 milligram = 0.001 gram
1 centigram = 0.01 gram
1 decigram = 0.1 gram
1 kilogram = 1000 grams

We abbreviate these masses as follows:

1 milligram = 1 mg
1 centigram = 1 cg
1 decigram = 1 dg
1 gram = 1 g
1 kilogram = 1 kg

For reference, 1 gram is about the mass of a paper clip. One kilogram is
about the mass of a liter of water.

Time

The following conversions are useful when working with time:

1 minute = 60 seconds
1 hour = 60 minutes = 3600 seconds
1 day = 24 hours
1 week = 7 days
1 year = 365 1/4 days (for the Earth to travel once around the sun)

In practice, every three calendar years will have 365 days, and every fourth
year is a "leap year", which has 366 days, to make up for the extra quarter
day over four years. The years 1992, 1996, 2000, and 2004 are all leap years.

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This gives us a total of 52 complete 7 day weeks in each calendar year, with
1 day left over (or 2 in a leap year).

The year is divided into 12 months, each of which has 30 or 31 days, except
for February, which has 28 days (or 29 days in a leap year).

Decimals in measurement

We use decimals to specify units of measurement when we need more

precision about length, volume, mass, or time. For example, when specifying
the height of a person 1.63 meters tall, to say that person is 1 or 2 meters tall
doesn't give us a very good idea of how tall that person really is.
The prefixes for the different units of length, volume, and mass in the metric
system obey the following rules:

Prefix Multiply by
milli- 0.001
centi- 0.01
deci- 0.1
deka- 10
hecto- 100
kilo- 1000

So for example:
1 hectometer = 100 meters
1 centigram = 0.01 gram
3 milliliters = 3 × (0.001 liters) = 0.003 liters
0.9 kilometers = 0.9 × (1000 meters) = 900 meters

Examples:
metre (common base unit)
kilometre = 1000 metres
hectometre = 100 metres (not commonly used)
decametre = 10 metres (a measure used in naval artillery)
1
decimetre = ⁄10 of a metre
centimetre = 1⁄100 of a metre
= 1⁄1000 of a
millimetre
metre
litre (common base unit)

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kilolitre = 1000 litres (not commonly used)
(used for beer kegs, 1 keg is approx. 1⁄2 of a
hectolitre = 100 litres
hectolitre)
decalitre = 10 litres (not a commonly used measure)
decilitre = 1⁄10 of a litre
centilitre = 1⁄100 of a litre
millilitre = 1⁄1000 of a litre

71