Exercise 1 Date : ____________

GENERAL PRECAUTIONS WHILE WORKING IN NUTRITION LABORATORY

Laboratory is a place where a person can experiment, observe and acquire knowledge. To
perform such experiments, different apparatuses, equipments and reagents are used. Animal
Nutrition Laboratory requires the use of clean glassware, preparation of samples, weighing of
samples, solution preparation, reagents making, filteration, drying, condensation, extractions,
estimation and calculations.
Following precautions are to be taken while working in a Nutrition Laboratory:
1. Laboratory safety:
(i) Laboratory should be well ventilated and fitted with exhaust fans for effective
removal of fumes.
(ii) Use apron and other devices like gloves, goggles etc. depending upon the
material to be handled.
(iii) DO NOT ADD WATER TO ACIDS. Keep acids off skin and protect eyes
from spattering. If acids are spilled on the skin, wash off immediately with tap
water. Gaseous nitrogen oxides from HNO3 can cause severe lung damage.
Copious flow of fumes is there when both concentrated HNO3 and HCl are
mixed together.
(iv) If acid falls on clothes, neutralize the same with few drops of dilute ammonia
solution or some weak alkali solution.
(v) If acid spills on the floor or the table, neutralize it with some weak alkali and
wipe off with duster.
(vi) If you happen to suck acid into your mouth during pipetting, wash your mouth
quickly with water and then rinse with a weak solution of washing soda.
(vii) Use fume hood to protect against any type of fumes.
(viii) Avoid use of equipments for purpose other than intended.
2. Cleaning of laboratory wares:
Glass and porcelain wares should be thoroughly washed with some detergent, then
extensively rinsed with tap water followed by rinsing with distilled water. If grease film
remains after cleaning with detergent, a cleaning solution containingsodium or potassium
dichromate in concentrated H2SO4 may be used. After this, rinsing is necessary to remove the
last traces of dichromate.

1
Preparation of cleaning solution:
Mix 15-20 g potassium dichromate to 50 ml H2O. Dissolve it by warming not by
heating. Then add 600 ml of concentrated H2SO4. This solution is only effective when it is
heated upto 70°C. If solution turns green by sometime, then discard it.
Some other cleaning solutions:
- Alcoholic solution of potassium hydroxide
- Mixture of sulphuric acid and nitric acid.
- Equal volumes of approximately 0.1N potassium permanganate and conc. H2SO4.
PRECAUTIONS IN THE USE OF PIPETTES
1. Pipettes must be clean. No drops should adhere to the walls after the pipette has
been drained.
2. Before use, the pipette should be dry or rinsed three times with small volumes of
the solution to be measured.
3. While filling a pipette, be sure that its tip is well below the surface of the liquid.
This is extremely important when strong acids, bases and other corrosive or
poisonous solutions are handled.
4. Draw the liquid a little above the mark, then carefully remove all drops adhering
to the outside of the pipette stem by wiping with a clean piece of filter paper.
5. Carefully adjust the meniscus of the liquid to the mark while the pipette is held
vertically.
6. Hold the pipette vertically while draining it when the liquid has ceased to run,
touch the tip of the pipette to the wall of the receiving vessel. A small drop of
liquid will remain in the tip of the ordinary transfer pipette. Do not try to remove
this drop. However, when using an Ostwald pipette, this last drop is removed by
blowing the pipette.
7. Use care to prevent contamination of the pipette tip. Support in such a way that tip
does not rest on the bench top.
Drainage time of various
pipettes
Capacity (ml) 2 10 20 25 50
Approximate drainage time (sec) 10 20 28 30 35

2
PRECAUTIONS IN THE USE OF BURETTES AND MICRO-BURETTES
1. The burette must be clean, when a liquid is delivered from a burette, no adhering
drops should be left on the walls. If drops are formed, the burette must be thoroughly
cleaned before it can be used.
2. Before use, the burette should be rinsed three times with small volumes of the
standard solution with which it is to be filled.
3. Fill a micro burette by attaching a rubber tube to the top. Suck up the solution from a
beaker held beneath the tip. Remove the rubber tube and wipe off the burette tip with
a clean filter paper before adjusting the meniscus to the zero mark.
4. After filling a burette, it is essential to remove all air bubbles from the delivery tip
before measurements are made.
5. Be sure that the burette delivery tip or stopcock does not leak.
6. During titrations, do not empty the burette too rapidly. Accurate measurements
depend largely upon uniform drainage.
7. Make careful readings. Have your eyes level with the bottom of the meniscus.
8. Observe the proper use of significant figures.
PRECAUTIONS IN USING FLASKS AND BEAKERS
1. They should not be heated directly on a flame but should be placed on asbestos
wire gauge. This ensures the equal distribution of heat across the bottom.
2. They should be constantly shaken to reduce bumping. If it is conical flask, hold it
by a paper collar and swirl the centrists.
3. In a beaker, if liquid is to be heated, stir the contents with glass rod.
Funnels: They are used for filteration i.e. separating a liquid from solid particles. So, keep
them clean and free from dust particles.
MEASURING CYLINDERS:
1. While reading volume of a liquid in a measuring cylinder, hold it vertically
and place the eye level with liquid surface. The reading in line with lowest
part of curved surface surface gives the volume of liquid.
2. While pouring a liquid (measured volume) use a glass rod to prevent
splashing.
TEST TUBES:
1. Must be placed in a rack (wooden or plastic).
2. While heating, hold the test tube with a test tube holder.

3
 3. On a flame, it should be shaken to prevent overheating.
4. Hold the open end away from you and your classmates to prevent mishap
while heating a test tube.
HANDLING OF GLASS APPARATUSES
- Glass apparatus used is delicate and thin walled so should be handled with care.
- Do not place a hot apparatus on the cold surface.
- Do not heat thick walled glass apparatus. Never pour hot liquids into them.
- They are liable to break.
- Measuring cylinder should not be heated.
- Always fill a container (glass) ¾ of its capacity with liquid to be heated, as it may
overflow.
- Do not use crackedor broken glassware, it may cause injury or break during the
experiment.
Other things to take care:
- Mortar and pestle should be washed and cleaned.
- Tongs should be used to hold crucibles.
- Spatula should not be heated.
For laboratory accidents/mishaps
First aid box-should contain
- Bandages, gauge, cotton wool, leucoplasts
- Scissors, forceps, safety glass, droppers
- Vaseline, boric acid powder, sodium bicarbonate powder
- Glycerin, sarson oil, picric acid and tannic acid solution
- Methylated spirit, rectified spirit
- Burnol for burns-antiseptic ointment
Study Questions

4
 Instructor’s Signature
Date:
Exercise 2 Date : ____________

FAMILIARIZATION WITH VARIOUS FEEDSTUFFS

Food is a material which after ingestion is capable of being digested, absorbed and
utilized. Any naturally occurring ingredient or material fed to animals for the purpose of
sustaining growth and development is called feed. Livestock feeds are generally classified
according to the amount of specific nutrient they furnish in the ration. These can be divided
into the following major categories:
1. Roughages 2. Concentrates 3. Supplements 4. Additives
Feeds

(A) (B) (C) (D)

Roughages Concentrates Supplements Additives
(A)
Roughages

Dry Green

Crop residue Hay Industry Natural Cultivated

Straws Tree waste
Stovers leaves Bagasse
Husk
Grass land Leguminous Non-
Pastures Leguminous
Tree fodder Berseem Maize
Subabul Lucerne Bajra
Jamun Shaftal Sorghum
Sewani Senji Guinea grass
Aquatic Cowpea Napier grass
water
Weed Guara Sudax
marine
algea Oats
Barley

5
 Rye grass

The roughages are fibrous and bulky feeds of whole plant or agricultural crops residue, and
they are relatively inferior source of available energy because of relatively higher amount of
crude fibre. Almost all feeds of plant origin containing higher than 18 per cent crude fibre on
dry matter basis are known as roughages. However, some exceptions in concentrates
containing more than 18 per cent crude fibre are cotton seed, sunflower seed, gram chuni etc.
The self explanatory description of feedstuffs and other feeds like ingredients has been
described in a chart form
(B)
> 60% TDN Concentrates <18% CF
<35% NDF

Energy Feeds Protein feeds

Grains By products Animal protein Vegetable protein

- Maize -Wheat brain -Meat meal - Mustard cake
-Wheat -Rice bran -Bone meal -Soybean meal
-Rice -Rice polish -Blood meal -Cotton seed cake
-Sorghum -Molasses -Fish meal -Linseed cake
-Bajra -Feather meal -Til cake
-Skim milk powder -Sunflower cake
-Groundnut cake
-Maize cake
-Arharchuni
-Gram chuni
-Maize gluten meal
-Moongchuni

6
 (C)
Supplements

Vitamins Mineral mixture

Salt (NaCl)
DCP (Di Calcium
Phosphate)
LSP
(D)
Additives

Antibiotic Hormones Enzymes Antifungal Acidifiers Flavours

Probiotics

Date ___________________
Observations:
S. No. Name of the ingredient/sample Description

7
 Instructor’s Signature
Date:

8
Exercise-3 Date______
PREPARATION AND PROCESSING OF SAMPLES FOR CHEMICAL ANALYSIS
(Herbage, faeces, urine and silages)
The quality of the ingredient is the foundation upon which an animal ration is built.
Therefore, proper smapling and evaluation of incoming feed ingredient is a critical step in
manufacturing quality feed. The objective is to check the quality of feedstuffs at the time of
purchase, processing for manufacturing of feed / pelleted feed / compounded feed, safe
storage and of processed feed. Sampling also required during digestion and metabolism trial
for the determination of digestibility and to know the balance of nutrients or minerals.
Sample: Small amount of representative feed or an ingredient from the bulk.
Sampling effects
 The selection of a ingredient for a feed formula
 The selection of a supplier for an ingredient
 The acceptance or rejection of a shipment of an ingredient
 Economics of the enterprise
The best time to take sample from bags is when the lot is being moved out or moved
in. The scale of sampling will be:

Number of bags in the lot Number of bags to be sampled

Up to 30 bags All bags
31 to 300 bags 30 bags
301 to 1000 bags 50 bags
1001 to 2000 bags 100 bags
2001 and above 120 bags

Selection of bags for sampling, according to the above scale shall be done at random
and shall cover bags from various positions of the stock and in which bag is to be sampled is
directed by the formula: N/n
Where,N = total number of bags
n = total number of bags to be sampled
The sample from a bag is drawn with the help of a 'Parkhi' or slotted tube sampler by
probing diagonally from one corner of the bag to the opposite corner, with the probe inserted
as nearly horizontal as possible. This is known as Primary sample.

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Sampling from bulk: Select the spots for sampling at various depths. Samples should
preferably be drawn when the feed is being moved out or moved in. The scale of sample is as
under:

Quality of consignment Number of spots from where samples to

be drawn
Up to 30 tones 30
31 to 100 tones 50
101 tones and 300 50 spots + 10% of excess

Sampling of bulk is done either with a thermo sampler or a deep pin probe. These
samples are inserted in a close position and when the required depth is achieved, the tube is
then opened. Then, allowed to collect. This is also called a Primary sample.
Composite sample: All the primary samples are mixed thoroughly and blended to constitute
a homogenous mixture.
Check sample: It is the one, which is carefully subdivided and portions sent to a number of
laboratories for analysis and used as check on laboratory assay procedures.
Duplicate samples: It is representative portion of a existing sample, provided to additional
laboratory to resolve differences if, any.
Official sample: It is one, taken by government officials for regulatory purposes.
Purchasing sample: It is the portion of the sample submitted by the supplier to the
purchaser.
Referee sample: It is often taken by impartial sample and sent to a referee laboratory for the
purpose of arriving at settlement between buyer and seller.
Reference sample: A sample of known characteristics, kept as guide or comparison check
for inspection or analysis. It is useful in comparing incoming ingredients against
specifications. It is also used in comparing finished products for colour, texture or other
visual specification.
Working sample: It is the portion or portions of a sample used for analysis.

Tools for sampling

The mandatory requirement of any sampling equipment is that it will accept all
particle sizes that occur in the material being sampled, without discrimination and that the
materials used in the equipment will not contaminate the sample, e.g. trace minerals.
Sampling of green fodder:

10
  Select randomly 10 points representing whole of the field and grab 3-5kg of the plant
material from three points, without making any choice.
 Spread these on the floor evenly and make another sampling from it weighing 1-2 kg.
 Chop it into small piece (about 1 cm) with the help of a hand chaff cutter.
 Spread chaffed fodder on the floor evenly and make spot sampling from different
place weighing ½ kg.
Sampling of silage
Collect the sample from different parts of silo pit. Cut into pieces of 3 to 5 cm length
with the help of a spade on clean surface.
Sampling of hay and straw
Take about one kg of straw from various places in the heap. Chaff the sample, dry it
and grind in a mill through 1 mm sieve and preserve in plastic bags or bottles.
Sampling of concentrate feeds
Take representative samples from different parts of each bag. Mix the sample
thoroughly and determine its dry matter content. Grind the sample and preserve for further
analysis.
Sampling of liquid feeds
The major problem in sampling is due to segregation or setting. The liquid feed
should be mixed and agitated by circulating or stirring before it is sampled. Liquid samples
are taken by dippers or draw off pipes.
Faeces sampling
a) Large animals: Mix daily collection on a clean surface manually. Take 1/100th part of
dry matter estimation and 1/500th part for nitrogen estimation. For nitrogen estimation
preserve the sample in bottle with 40% (w/v) H2SO4 after through mixing. Dried
faeces are ground and stored in bottles for further analysis.
b) Small animals: Grind daily collection and mix thoroughly. Take 1/5th part for dry
matter and 1/50th part for nitrogen determination as described above.
c) Poultry faeces: Collect droppings on polythene sheets. Separate contaminations like
feathers and feed and mix thoroughly. Take 1/20 th and 1/100th part in glass bottles for
dry matter and nitrogen determination, respectively. For nitrogen estimation, preserve
the sample with 5 per cent (w/v) H2SO4.
Urine sampling

11
 The samples of urine (5-20% depending upon the size of the animal) are collected
daily in bottles and evaporation of ammonia-N is prevented by adding 5% (w/v) H2SO4.
Milk sampling
Just after milking, milk is thoroughly mixed with plunger (agitator) and sample is
taken in bottles. Potassium dichromate or mercury chloride is added as a preservative agent.
The samples are stored in refrigerator until analyzed.
Storage of samples
The nature of storage depends upon the kind of analytical report required. The
purpose of storage is to preserve the material for a period as long as required without
affecting significant change in its components.
The following processes are commonly used:
1. Freezing
2. Cold storing
3. Freeze drying
4. Oven drying
5. Use of chemical / preservatives
6. Irradiation
Store the samples in cool, dark and dry place with proper labeling showing the following:
 Name of the ingredient
 Code number
 Date
 Batch number
 Signature
Study Questions:

Instructor’s Signature
Date:
Exercise -4 Date______
PREPARATION OF SOLUTIONS
(A) Principles of volumetric analysis
Solution:

12
 A solution is a homogenous liquid mixture, which consists of a solid and liquid where
the solid is dispersed in the liquid at the molecular level. The solid is referred to as a solute
and the liquid is referred to as the solvent. When both the components are liquid, the
component in excess is called as solvent and other as solute.
The amount of solute dissolved in an unit volume of solvent is turned as
concentration. A standard solution is the solution of known concentration.
Expression of a solution concentration
Most chemical reactions occur in a liquid solvent / solute (i.e. solution environment).
Typically the solute (s) in a solution will be the reactants. Concentration is a way to express
the amount of solute per unit amount of solution / solvent. There are several ways that
concentration can be expressed.
a) Mass per cent: It means the number of grams of solute per 100 g of solution.
For example, 10 g sodium chloride in 90 g water is a 10% by mass solution.
Mass per cent = (mass of solute / mass of solution)  100
= 10 g / (10 g + 90 g)  100
= 10%
b) Volume per cent: it means the number of milliliters of solute per 100 ml of solution.
The volume per cent of a solution cannot be calculated directly from the volumes of
its components because the final volume may not equal the sum of the components
volumes. To prepare volume per cent solution, first determine the final volume and
concentration of the desired solution and then determine the amount of solute. Dilute
the solute in sufficient solvent to produce the final volume of desired solution. For
example, to prepare 100 ml of a 10% by volume solution of acetic acid, dilute 10 ml
acetic acid with distilled or deionized water to make 100 ml of solution.
Note: Solutions of concentrated reagents, such as 37% hydrochloride acid and 85%
phosphoric acid, are per cent solution by mass.
c) Molar solution: Molar solution is one that contains one mole or one molecular
weight in grams of a substance in each litre of the solution, whether the substance is
in the form of molecules, ions or any other species. it is designated as M.
Molar methodof expressing concentration is useful due to fact that equal volumes of
equimolar solutions contain equal number of molecules.
To make a 1 M solution of sodium hydroxide, slowly add 40 g sodium hydroxide to
500 distilled or deionized water in a 1000 ml volumetric flask. When all solid is dissolved

13
and the solution is at room temperature, dilute to the mark and invert the flask several times
to mix.
d) Molarity (M)
Gram moles of solute
Molarity (M) = 
Volume of solution in litres

Mass of solution in grams

Molarity (M) = 
Gram molecular mass of solute  volume of solution
e) Molality (m): Molality of a solution may be defined as the number of gram moles of
the solute dissolved in 1000 g of the solvent. It is represented by small letter 'm'.
Gram moles of solute
Molarity (M) = 
Mass of solvent (kg)

Mass of solute (g)

= 
Gram molecular mass of solute  mass of solvent (kg)
A solution containing one mole of solute per 1000 g of solvent has molality equal to
one and is called molal solution and expressed as moles per kilogram (mol kg-1). Molality of
a solution does not change with temperature.
f) Normality (N): A normal solution of a regent is one that contains 1 g equivalent
weight per litre of solution.
Number of gram equivalent
Normality (M) = 
Number of litres

Number of milligram equivalents

= 
Number of milliliters (ml)

Hence, number of mg equivalent = number of ml  normality. If the volumes of

solution of two different substances A and B which react with one another are V A ml and
VBml respectively, then these volumes severally contain the same-number of g equivalents or
mg equivalent of A and B. Thus
VA normality A = VB normality B
Neutralization reactions:

14
 Equivalent weight of an acid is that weight of it which contains one gram atom of
replaceable hydrogen i.e. 1.1008 (more accurately 1.0078) g of hydrogen. The equivalent
weight of a monobasic acid such as hydrochloride acid will therefore contain 1 g molecular
weight (or 1 mol) in a litre of solution. The equivalent weight of a diboric acid (e.g. sulphuric
acid) is likewise ½ and respectively of its molecular weight.
Molecular weight of an acid
Equivalent weight of an acid = 
Basicity
Basicity of an acid is equal to the number of replaceable hydrogen atom present in
one molecule of the acid.
36.46
 Equivalent weight of HCl =  = 36.46
1

98
Equivalent weight of H2SO4 =  = .49
2

106
 Equivalent weight of (COOH)2 2H2O =  = 53
2
Equivalent weight of a base: It is that weight of it which contains one replaceable
hydrogen group, 7.008 g of ionizable hydroxyl; which is equivalent to 1.008 g of hydrogen.
The equivalent weights of sodium hydroxide and potassium hydroxide are 1 mol of calcium
hydroxide and barium hydroxide ½ mol.
Molecular weight of alkali (g)
Equivalent weight of an alkali = 
Acidity
Acidity of an alkali is equal to the number of replaceable hydroxyl groups present in
one molecular of the alkali.
40
Equivalent weight of KOH =  = 40
1

56
Equivalent weight of KOH =  = 56
1
106
Equivalent weight of Na2CO3 =  = 53
2
Oxidation – reduction reactions

15
 The equivalent weight of an oxidizing or reducing agent is most defined as that
weight of the reagent which reacts with or contains 1.008 g of available hydrogen or 8.000 g
of available oxygen. By 'available' is meant capable of being utilized in oxidation or
reduction. The amount of available oxygen may be indicated by writing the hypothetical
equation e.g.
2KMnO4 = K2O + 2 MnO + 50
i.e. in acid solution 2 KMnO4 gives up 5 atoms of available oxygen, which is taken up by the
reducing agent, hence its equivalent weight is 2 KMnO 4 / 10. For potassium dichromate in
acid solution, the hypothetical equation is:
K2Cr2 O7 = K2O + Cr2 O3 + 30
The equivalent weight is K2O + Cr2 O3 + 30
The equivalent weight is K2Cr2 O7 / 6
Equivalent weight of oxidizing substances: Gram equivalent weight of an oxidizing
substance is that weight of the substance in grams which is equivalent to 8 grams of available
oxygen.
a) Equivalent weight of KMnO4 in acidic medium can be determined as follow:
2 KMnO4 + 3 H2SO4 = K2SO4 + 2 MnSO4 + 2 H2O + 50
2 (39+55+64) = 316 = 80 (5  16)
This shows that:
316 g of KMnO4 can give 80 grams of oxygen or 5 atoms of oxygen for oxidation.
So, 2 KMnO4 5 (0) = 10H
Molecular weight 158
Equivalent weight of KMnO4 =  =  = 31.6
5 5
b) Equivalent weight of KMnO4 in alkaline medium
2 KMnO4 + H2O  2 MnO2 + 2 KOH + 30
KMnO4 3 (0)  6 H
Molecular weight 158
Equivalent weight of KMnO4 =  =  52.67
3 3

Titrations:
In titration a known volume of a reactant with known concentration, is slowly
added to a vessel with another reactant until the reaction is complete. The point at which

16
the reaction is complete is known as the 'endpoint'. Typically the end point is determined
visually through either a colour change or formation of a precipitate.
Table : Change in colour of various indicators at different pH range

Indicator Acid colour Base colour pH range

Cresol red Red Yellow 0.2-1.8
Thymol blue (acid range) Red Yellow 1.2-3.0
Red cabbage extract Red Yellow 2.4-4.5
Bromphenol blue Yellow Blue 3.0-4.6
Bromcresol green Yellow Blue 3.8-5.4
Methyl red Red Yellow 4.2-6.2
Bromcresol purple Yellow Purple 5.2-6.8
Bromthymol blue Yellow Blue 6.0-7.6
Phenol red Yellow Red 6.8-8.4
Cresol red Yellow Red 7.2-8.8
Thymol blue (base range) Yellow Blue 8.0-9.6
Phenolphthalein Colourless Red 9.4-10.5
Thymolphthalein Colourless Blue 10.1-12.1
Alizarin yellow R yellow Lilac 10.1-12.1
Trinitrobenzene Colourless Orange 12.0-14.0

Important titrations and choice of indicators

Titration Indicator
1. Strong acid and strong base Methyl orange as phenolphthalein
2. Weak acid and strong base Phenolphthalein
3. Strong acid and weak base Methyl red / methyl orange
4. Weak acid and weak base Methyl orange

Study Questions :

17
 Instructor’s signature
Date:

18
 Exercise-5 Date______
PREPARATION OF STANDARD SOLUTION
(B) Preparation of Standard Solution
Solution of approximate normal (one normal of 1N) strength can be prepared by
dissolving a little more than the gram equivalent weight of the substance per litre of the
solution and then exact standardization is done with some primary standard solution.
(i) Primary Standards:
A primary standard solution is one which can be prepared directly by weighing. Other
solutions of approximate strength can then be titrated and standardized with the help of this
primary solution. Pot. hydrogen iodate, sodium carbonate, borax, succinic acid, oxalic acid
and sodium oxalate may be employed as primary standards.
Sodium carbonate (Na2CO3)
Anhydrous sodium carbonate of A.R. (analytical reagent) quality should be selected.
It reacts with the acid according to the following equation:
Na2CO3 + 2HCl = H2O + 2NaCl + CO2
Eq. wt of Na2CO3 = 53 (106/2=53)
Sodium carbonate is hydroscopic and therefore it must be made perfectly anhydrous
before it is weighed.
Procedure for N/10 Na2CO3:
Take about 6-7 g of Na2CO3 (A.R.) in a nickel crucible and heat it in a hot air oven at
about 100°C for an hour or so to drive out the moisture. Now accurately weight 5.3g of the
salt on an analytical balance and dissolve it in a little freshly boiled distilled water. Transfer it
to a one litre measuring flask and make volume up to the mark. Shake well and label it as.
0.1N or N/10 Na2CO3
Preparation of 0.1N hydrochloric acid
The concentrated acid is 10-12 N. Therefore, to prepare a standard solution, say
decinormal of the acid, it is diluted roughly one hundred times. Take about 10 ml of strong
hydrochloric acid from a burette in a one litre measuring flask and make volume upto the
mark with distilled water and shake well. Put this solution in the burette and pipette out 10 ml
of 0.1 N Na2CO3 in a conical flask. Add a drop of methyl orange as indicator. Allow the acid
to run in, drop by drop till the yellow colour in the flask changes to red. Repeat the
experiment to get at least three concordant readings.

19
Calculations:
Suppose 10 ml of 0.1N Na2CO3 = 8ml of HCl
N1V1 = N2V2
0.1N 10 = N28
N2 = 0.125
To prepare 1 litre of decinormalHCl, the amount of 0.125N acid required is
10000.1000.125 = 800 ml. Take 800 ml of 0.125N acid and dilute to 1 litre. Label it as
0.1N HCl and check its strength again by titrating 10 ml of its which must neutralize exactly
10 ml of 0.1N Na2CO3.
Preparation of 0.1N sulphuric acid
Concentrated sulphuric acid is 34-36 normal. Add 3.5 ml of sulphuric acid in a one
litre beaker which has already been half filled with distilled water. Now, transfer it to a one
litre measuring flask; make volume upto the mark and shake well. Titrate this solution against
0.1N Na2CO3 using methyl orange as indicator. To prepare 0.1N acid, follow the steps as in
case of preparation of 0.1N hydrochloric acid.
Preparation of 0.1N potassium permanganate (KMnO4)
Potassium permanganate and potassium dichromate are used as oxidizing agents in
the volumetric analysis, in the presence of dilute sulphuric acid.
Great care should be taken to prepare a standard solution of KMnO 4 as it is not
available in a very pure form.
N/10 KMnO4
Its standard solution can be prepared by standardization of against a standard solution
of sodium oxalate or oxalic acid. The equivalent weight of sodium oxalate
COONa
 = 67 and that of KMnO4 = 31.6 and of oxalic acid = 63
COONa

To prepare decinormal solution of sodium oxalate, weigh 6.7 g of it and dissolve in water.
Make volume to one litre and shake well. For 0.1 N KMnO 4solution, weigh 3.2 g of it and
dissolve in H2O to make the volume to one litre. Titrate against standard sodium oxalate
solution after acidification and dilute accordingly to make exactly 0.1N KMnO 4. Remember
that KMnO4 solution acts as a self indicator and therefore no indicator is to be added from
outside.

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Note: Standarization of solution can be done directly also by taking 50-100 mg (exact
quantity) of the solid primary standard and adding about 100 ml of distilled water to it and
titrating with the test solution.

Study Questions:

Instructor’s signature
Date:

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Exercise 6 Date : ____________

WEENDE’S SYSTEM OF ANALYSIS (PROXIMATE ANALYSIS)

The scheme of analysis of feedstuffs, also known as 'Weende System' or
'Proximate Analysis' was proposed by two German workers (Henneberg and Stohmann) at
the Weende Experiment Station in Germany. It is a system for approximating the nutritive
value of feedstuffs without conducting feeding trial. The scheme of analysis separates the
feed components into six fractions such as:
1. Moisture and dry matter
2. Mineral matter/ash
3. Crude protein
4. Crude fat/ether extract
5. Crude fibre
6. Nitrogen-free extract
Proximate analysis scheme of feedstuffs consists of organic and inorganic components.
Organic component consists of carbohydrates, fats and oils, proteins and other
nitrogenous compounds along with a number of other organic compounds. The inorganic
component consists of minerals like calcium, sodium, potassium, magnesium, iron,
molybdenum, copper, zinc, chloride, sulphate, silicate, carbonate etc. First five fractions
are determined in lab, while sixth i.e. NFE is calculated by subtracting from hundred.
NFE=100-(CP+CF+EE+ASH+H2O)

Summary of fractions of proximate analysis

Fractions Major components Procedure/Method

Moisture Water and any other volatile Drying at approximately
compounds 100°C to a constant weight
Ash Mineral elements Ignition at 500°±10°C for
4-5 hours
Crude protein Proteins, amino acids, non protein N by Kjeldahl digestion
N
Crude fibre Cellulose, some hemi-cellulose Residue left after boiling
and lignin with acid and alkali
Ether extract Fats, oils, waxes, sterols etc. Extraction with petroleum
ether
N-free extract Starch, sugars, some cellulose, Remainder, i.e. 100 minus

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hemicellulose and some lignin or sum of the above fractions
artifacts

Instructor’s Signature
Date:

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Exercise–7 Date _______

TO DETERMINE MOISTURE CONTENT OF FEEDS AND FODDERS

(OVEN DRYING METHODS-A COMMON PROCEDURE)
Equipment:
 Oven
 Desiccators
 Aluminium moisture dishes with lids
For fodders:
 Electronic balance with platform
 Aluminium tray 200 g
Theory:
Feedstuffs are dried in an oven at 100 ± 1˚C to a constant weight. The loss of weight
of sample is reported as moisture content. In this procedure, volatile substances present in
feedstuffs are also accounted as water.
Procedure:
Weigh in duplicate 2-5 ± 0.1 g sample into aluminium dishes which previously have
been dried, cooled in desiccator and weighed. Use covered aluminium dishes. Place the
dishes with lids removed in the oven and dry for 12-18 hours at 100 ± 1˚C. After this period,
replace the cover while still in oven and transfer the dishes to desiccator. Allow sufficient
time to attain room temperature and weigh it. Report loss in weight or moisture. Percent dry
matter is calculated by subtracting percent moisture from 100.
Observations:
Weigh of empty dish (W)
Weigh of dish plus sample before drying (W1)
Weigh of dish plus sample after drying (W2)
Calculations:
W 1−W 2
Moisture content of feedstuffs (%) = ×100
W 1−W
Loss∈weight
= ×100
g , sample
Weight of sample after drying
Or ×100=% DM
Weight of sample before drying
100 - % DM = % Water / moisture

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25
Study questions:
1. Enlist different methods of drying samples.
2. Name the method used for estimation of moisture in fermented feeds like silage.

Instructor’s Signature
Date:

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Exercise – 8 Date _______
(A) TO DETERMINE THE TOTAL ASH CONTENT OF FEEDS AND FODDERS
Equipment:
 Desiccators
 Muffle furnace
 Large crucibles
 Tong
Theory:
Ash is the inorganic residue remaining after a feedstuff is ignited to carbon free, usually at a
temperature not exceeding red heat (500-600 ˚ C).
Procedure:
Weigh in duplicate 2 ± 0.1 g samples into crucibles. Burn the organic matter by placing the
sample on a hot plate till fumes stops and then keep the crucible in a muffle furnace raising
the temperature up to 600˚C slowly by setting the regulator at low speed. Ignite the sample
for four hours at 600˚C. Transfer crucibles directly to desiccator (not above 100˚C), cool and
weigh. Report residue as percentage ash to first decimal place on an air dry basis.
Observations:
1) Samples 2) Duplicate
Weight of empty crucible (W0) =
Weight of crucible plus sample (W1) =
Weight of sample (W1-W0) =
Weight of crucible + Sample after ignition (W2) =
Weight of residue (W2-W0) =
Calculations:
W 2−W 0
% ash = × 100
W 1−W 0
(B) TO DETERMINE THE ACID-INSOLUBLE ASH CONTENTS OF FEEDS AND
FODDERS
The total ash (content) obtained after muffling the feed materials at 600˚C. Dissolve
the ash in 25 ml HCl(1+1) and transfer to 250 ml beaker keep the beakers on hot plate and
evaporate the acid till half of it remains in the beakers. Now, add 5 ml conc. HCl and heat to
dryness and continue heating for another 30 minutes on a water bath to dehydrate silica. Now
add 5 ml conc. HCl and 5 ml water to the residue. Heat for a few minutes on water bath.
Filter through whatman filter paper. The insoluble material left on the filter paper is called the

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acid insoluble ash. The residue along with the filter paper are dried and again muffled at
600˚C to get the weight of the acid insoluble ash.
Calculations:
Weight of crucible + sample after ignition = W3
W 3 −W 0
x 100
% AIA = W 1−W 0
Study questions:

Instructor’s Signature

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 Date:
Exercise – 9 Date _________

TO DETERMINE THE CRUDE PROTEIN CONTENT OF FEEDS AND FODDERS

Equipment:
 Kjeldhal flask
 250 ml conical flasks
 50 ml burette
 50 ml cylinder
 Digestion and distillation apparatus
Reagents:
Chemically pure H2SO4
NaOH – 40%
Digestion mixture (9.0 g K2SO4 + 1.0 g CuSO4)
Standard solution H2SO4(0.1 N)
Boric acid solution – 4 %
Mixed indicator:
0.1 g methyl red plus 0.5 g bromocresol green in 100 ml of 95 % ethanol. Adjust the colour
of the solution with drops of dilute HCl or NaOH to bluish purple mid colour. Add 5 ml
mixed indicator per litre of boric acid solution.
Theory:
The organic nitrogen present in feedstuffs is converted into ammonium sulphate by boiling
with concentrate sulphuric acid. The reaction proceeds instantaneously in two steps as follow:
Protein + H2SO4→ CO2 + NH3 + SO2 + H2O
2NH3 + H2SO4 → (NH4)2SO4
The digestion mixture acts as a catalyst. Copper sulphate lower the temperature at which
the reaction takes place while potassium sulphate raises the boiling point of the mixture.
The ammonium sulphate derived from feed nitrogen is converted into ammonia by
adding excess 40 % alkali.
(NH4)2 SO4 + NaOH → 2NH3 + Na2SO4 + 2 H2O
The ammonia is received in boric acid solution by steam distillation. The ammonia
boric acid complex (ammonium borate) is titrated with standard sulphuric acid.
NH4OH + H3BO3 → NH4B(OH)4
2 NH4B(OH)4 + H2SO4 → (NH4)2SO4 + H3BO3 + 2 H2O

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Procedure:
Weigh 1-2 g sample on filter paper and transfer wrapped sample into kjedhal flask.
Add approximately 5 g digestion mixture and 30 ml conc. H2SO4. Digest the sample until
the solution turns clear green, boil briskly for 30 minutes. Digestion should be carried out in
fume cup board and the exhaust fan should be turned on before starting the digestion. The
flask is rotated occasionally during digestion until all the black specks disappear. If any
material sticks to the neck of the flask, cool, then wash down with a small amount of water
from wash bottle and redigest. Allow the flask to cool in a closed fume cupboard. A reagent
blank be run simultaneously. Now transfer it into a 250 ml flask and make the final volume to
250 ml by adding distilled water. Now take 50 ml out of 250 ml flask in one litre conical
flask, then add 40 % NaOH and then do the steam distillation.
Prepare distillate receivers in duplicate by taking 25- 30 ml boric acid indicator
solution in each of 250 ml conical flask. Add 2 drops of mixed indicator, in case it was not
added previously in stock boric acid solution.
Place the flask beneath the condenser tube such that the tip of the condenser dips in
boric acid solution. Turn on condenser water. The hot plate should be switched on well in
advance prior to connecting 1000 ml conical flask. Otherwise, the boric acid solution may be
sucked back through the condenser, if this happens, the condenser and connecting bulb must
be washed thoroughly with distilled water and stream. The determination should be repeated.
Add 300 ml water in cool contents of the flask and a few grains of zinc to prevent bumping
and reduce frothing. Keep the flask in tilted position away from your body while adding the
water. Then add 100 ml 40% NaOH solution in the flask. Immediately connect the flask to
the condenser and distil until all the ammonia has passed over into the receiver.
The first 125 to 150 ml distillate will generally contain all the ammonia. When the
distillation is practically completed lower the receiving flask before turning off the heater and
allow about 5 ml more to distil through in order to wash the condenser tip.
Titrate with standard(0.1 N) sulphuric acid to a pink/red (same as boric acid). Correct
the blank determination on reagents.

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Observations:
(1) (2)
Strength of the acid used =
Final burette reading =
Initial burette reading =
Acid used =
Reagent blank reading =
Actual acid used =
Calculations:
Two atoms of nitrogen (in the form of ammonia) have neutralized one molecule of H 2SO4
with the formation of (NH4)2SO4. Hence each ml of 1 N acid neutralized by the ammonia is
equivalent to 14 mg of nitrogen.

N × amount of acid used (ml) ×14 × 100 250

× 50 = K
% Nitrogen = 1000× g , sample

N = Normality of acid
% crude protein = % K × 6.25
Study questions:

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 Instructor’s signature
Date:

32
Exercise-10 Date _______

TO DETERMINE THE ETHER EXTRACT CONTENT OF FEEDS AND FODDERS

(Soxhlet Extraction Method) AND TO CALCULATE NITROGENFREE EXTRACT
(NFE)
Equipment:
 Soxhlet extraction unit
 Heating mantles
 Extraction thimbles
 50 ml beakers
 Steam bath
Reagents:
Petroleum ether (boiling range, 40 to 60o C)
Theory:
The sample is extracted continuously with freshly distilled portions of petroleum ether. The
vapours of ether, boiling in the flask pass into the condensers where they condense and drop back on
the sample, thus dissolving the ether soluble material. The soluble substances along with ether are
siphoned back into the extraction flask. This process is continuous and eventually all the ether soluble
material is removed from the sample. The extract is termed “Crude fat” or Ether extract. The latter
term is more accurate since in addition to true fats, the extract also contains a number of other fat like
substances. In some plant materials, the true fat may constitute less than 50% of the total extract.
Procedure
Weigh 3-5gram sample in the extraction thimble. Place the thimble in a soxhlet extractor and
connect it to the flask. Pour sufficient quantity of petroleum ether to run the siphon. Place the
extractor along with the flask on the heating mantle and connect the condenser with the water tap.
Turn on the water. Switch on the heating element and as soon as ether begins to boil, adjust the heat
so that the ether boils smoothly. Extract for 8 hours. After this, remove the thimble and collect the
ether in the extractor so that only a small quantity is left in the flask. Stop boiling and collect the ether
from the extractor in the bottle. Transfer the residual ether containing fat into tared beaker and wash
the flask with redistilled ether in order to transfer last traces of the fat. Evaporate on steam bath in
fume cupboard. Place beaker in oven at 105 o C for 30 minutes. Cool in desiccators and weigh. Report
ether extract content. Save the fat free residue in the thimble for the determination of crude fibre.

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Observations:
Duplicate samples
(1) (2)
Wt. of sample (W) =
Wt. of beaker + ether extract (W2) =
Wt. of empty beaker (W1) =
Wt. of ether extract (W2-W1) =
Calculations
(W2-W1)
(%) Ether extract = ×100
(W)
Determination of Nitrogen Free Extract (NFE)
It is commonly referred to as NFE.
NFE represents the sum of the remaining undetermined portions of the feed. It includes
mostly sugars, starches and also some of the soluble hemicelluloses. It is determined by substracting
the sum of % of moisture, %CP, % CF, % EE and % ash content from 100.
NFE = 100 - (% moisture + % CP + % CF + % EE + % ash)
Study Questions:
1. What is the importance of the ether extract estimation?
2. 3 gms of moisture free groundnut cake was extracted with petroleum ether. The weight of the
receiver flask before and after extraction were 62.1850 and 62.4360 gms respectively.
Calculate the % fat content of the groundnut cake.
3. 5 gms of moisture free chick mash was extracted with petroleum ether. The weights of the
receiver flask before and after extraction were 52.9920 and 53.2540 gms respectively.
Calculate the percentage fat content of the chick mash.
4. Explain the limitations of the method of Proximate analysis.

Instructor’s Signature
Date :

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Exercise-11 Date _______
TO DETERMINE THE CRUDE FIBRE CONTENT OF FEEDS AND FODDERS
Equipment:
 Hot plate
 Desiccator
 600 ml tall spoutlesspyrex corning beaker
 Filtering cloth- about 45 thread per inch
 Filtration/suction flask 500ml with gasket
 Funnels
 Large crucibles
 Wash bottles
 Horn spatula
Reagents:
 1.25% H2SO4 solution (0.255 N)
 1.25% NaOH solution (0.312 N) use carbonate-free water
 The strength of these solutions must be checked by titration
 Ethyl alcohol/rectified spirit (colourless)
Theory:
In livestock feeding, the information on the crude fibre content of a feed is essential because
feed that are high in fibre are less digestible than those that are lower in fibre. This group of substance
include cellulose and other carbohydrates which are not dissolved by weak acid and alkali. Crude
fibre is lost on ignition of dried residue remaining after digestion of sample with 1.25% H 2SO4 and
1.25% NaOH solutions under specific conditions.
Procedure:
Weigh 2-5±0.1gm samplesof fat free residue into 600 ml beaker avoiding fibre contamination
from paper or brush. Add 175ml distilled water and heat to boil. Then add 25 ml 10% H 2SO4 and boil
for 30 minutes. Rotate the beaker every 5 minutes. Maintain the volume constant by adding distilled
water, if necessary. At the end of 30 minutes, remove the beaker and filter through the cheese cloth
placed on the funnel. Wash with boiling water until the washings are no longer acidic. Check with
blue litmus paper.
Transfer the residual material into the original beaker with the help of a wash bottle and make
the volume to 175 ml. Heat and bring to boil. Add 25ml 10% NaOH and reflux for 30 minutes. After
this remove the beaker and filter the contents to the cheese cloth. Wash with boiling water until
washings are no longer basic to red litmus. Wash into the large crucible with a minimum amount of
water.

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 Dry the contents in crucible at 100 oC for overnight. Cool in a dessicator and weigh. Ash the
contents of the crucible in an electrical muffle furnace at about 600 oC for 2 hours, cool and weigh.
Report the loss in weight on ignition as crude fibre.
Observations: Duplicate samples
(1) (2)
Wt. of sample (W) =
Wt. of crucible + residual material =
(a) After drying (W2) =
(b) After ignition (W1) =
Loss in Weight (W2-W1) =
(W2-W1)
% Crude fibre= ×100
Gm sample
Study Questions:
1. 2 gms of a moisture and fat free cattle feed was digested alternately with 1.25% H 2SO4 for 30
minutes and 1.25% NaOH for 30 minutes. Calculate the crude fibre content with the
following data.
(a) Weight of the silica crucible plus crude fibre residue = 33.5513 gms
(b) Weight of the silica crucible + ash = 33.3200 gms
2. 2 gms of a moisture and fat free chich mash was taken for crude fibre estimation. Calculate
the fibre content with the following data.
(a) Weight of the silica crucible plus crude fibre residue = 32.4194 gms
(b) Weight of the silica crucible plus ash = 32.1706 gms

Instructor’s Signature
Date:

36
Exercise-12 Date _______
TO DETERMINE THE CALCIUM CONTENT OF FEEDS AND FODDERS
Equipment:
 Small crucibles
 Muffle furnace
 250 ml volumetric flask
 50 ml pipette
 Funnel
 Filter paper Whatsman No. 42
 One mark 50 ml pipette
Reagents:
0.05 N standard KMnO 4 solution, saturated ammonium oxalate solution, methyl red
indicator, dilute HCl (1+4)
Theory:
Calcium is precipitated as calcium oxalate. The precipitate is dissolved in sulphuric acid and
the solution is titrated with standard potassium permanganate (0.05N). The equation can be written as
follows:

Ca + 
Oxalate Calcium oxalate

+ H2SO4 + CaSO4
2 KMnO4 + 3H2SO4 K2SO4 + 2MnSO4 + 3H2O + 5O

+ O2 2CO2 + H2O

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Procedure
Char 2-5 samples on a hot plate and ash in muffle furnace for 4 hours at 600 o C. Dissolve the
ash in 25 ml HCl (1+1) and transfer to 100 ml beaker. Add 5 ml conc. HCl and heat to dryness and
continue heating for another 30 minutes on a water bath to dehydrate silica.
Now add 5 ml conc. HCl and 5 ml water to the residue. Heat for a few minutes on water bath.
Filter through Whatman filter paper and dilute to 250 ml with distilled water. Keep a part of this
solution for the estimation of phosphorus. The residue remain on the filter paper represents acid
insoluble ash.
Pipette 50 ml of solution in a beaker. Add 10 ml of saturated ammonium oxalate solution and
heat the solution to boiling. Add 2 drops methyl red and mix the content thoroughly. Neutralize the
content with ammonia and boil again until the precipitate is coarsely crystalline. Add a few drops of
dilute HCl until the colour is adjusted to faint pink. Allow the mixture to stand for 4 hours. Filter the
precipitate (calcium oxalate) through Whatman filter paper No. 42. Wash the precipitate with hot
water or with 0.5% ammonium hydroxide solution until the precipitate is free from chloride and
oxalate. Dissolve the precipitate along with filter paper in dilute sulphuric acid in the same beaker in
which calcium was precipitated. Heat the content to about 60 oC and titrate with 0.05 N KMnO4 to a
pink end point. Correct for blank and calculate % calcium.
Observations:
Duplicate samples
(1) (2)
Wt. of feed sample (W) =
Final volume of the test sol. = 250 250
Aliquot taken for dilution = 50 50
Burette final reading (b) =
Initial reading (a) =
Actual KMnO4 used = (b-a)
1 ml 0.05N KMnO4 = 0.001 g Ca++
1 ml 0.1N KMnO4 = 0.002 g Ca++
Calculations
0.001 x ml KMnO4 x 250
% calcium in feed = ×100
50 x gm sample

ml KMnO4 x 0.5
= ×100
gm sample
Study Questions:
1. What are the precautions to be taken while estimating calcium in a feed sample?

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2. 10 gms of sesbania leaves were ashed and the ash solution was made up to 100 ml. 10 ml of the
ash solution was precipitated as calcium oxalate. The acidified precipitate required 44 ml of N/10
KMnO4. Calculate the calcium content of the sample.
3. 10 gms of fish meal was ashed and the ash solution was made up to 100 ml of the solution were
precipitated for calcium as calcium oxalate. The precipitate required 35.0 ml of N/10 KMnO 4 for
titration. Calculate the calcium content of the sample.

Instructor’s Signature
Date:

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Exercise-13 Date_________
TO DETERMINE THE PHOSPHOROUS CONTENT OF FEEDS AND FODDERS
Equipment:
 Water bath
 250 ml beaker
 Funnel
 Whatman filter paper No. 42
 25 ml pipette
Reagents:
 HNO3
 0.1N Sodium hydroxide
 0.1N H2SO4
 2% Sodium nitrate
 1% Phenolphthalein indicator
 Molybdate solution
Dissolve 100 g of HNO3 in a mixture of 144 ml of ammonia and 271 ml of water.
Add this solution with constant stirring into a mixture of 400 ml of nitric acid and
1148 ml of water.
Decant the solution and add 100 ml of nitric acid. Filter before use.
Theory:
In acid solution the phosphate is precipitated as ammonium phosphomolybdate. The
washed precipitate is dissolved in a measured excess of standard alkali and the excess
of alkali is titrated against acid.
(NH4)3 Po4 12HoO3 + 23 NaOH = 11 Na2HoO4+(NH4)2HoO4+NaNH4HPO4+11H2O
Procedure:
Pipette 25 ml of solution in a beaker and add 10 ml of nitric acid and a piece of red
litmus paper. Add ammonia drop wise till the litmus paper changes colour. Dilute to
75-100 cc. Heat on a water bath at 45-50°C.
Add 25 ml of molybdate solution. The mixture is gently stirred and allowed to stand
for 2 hours. The yellow precipitate of ammonium phosphomolybdate is collected by
filtration over Whatman filter paper No. 42. Wash the precipitate with 2% sodium
nitrate until the filtrate is no longer acidic to blue litmus.

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Observations:
Wt. of sample =
Volume of acidic extract =
Aliquot taken for titration =
Amount of NaOH (0.1N) used for dissolving ppt. = (X)
Amount of acid (0.1N) for titration = (Y)
(a) Final burette reading =
(b) Initial burette reading =
Acid used = (Y)
Actual amount of NaOH (0.1N) used for dissolving out
Calculations:
1 ml 0.1N NaOH = 0.309 mg P2O5
Study questions:

Instructor’s signature
Date:
Exercise – 14 Date_________

41
 DEMONSTRATION OF THE DETERGENT METHOD OF FORAGE ANALYSIS
(VAN SOEST SCHEME OF ANALYSIS)
The Weende method of crude fibre(CF) and NFE analysis possess some limitations.
As per this method, the CF is assumed to be the indigestible part and should contain
cellulose, hemicellulose and lignin. Similarly, the NFE fraction is assumed to contain the
easily digestible parts like protein, starch, soluble sugars etc. however, it is not exactly true.
During the acid and alkali digestion, part of cellulose, hemicellulose and lignin comes out of
the CF fraction, as lignin is soluble in alkali and cellulose and hemicellulose both in acid and
alkali. Consequently, some part of cellulose, hemicellulose and lignin appear in NFE fraction.
Further, ruminants are able to utilize CF fraction efficiently.
P. J. Van Soest in 1965 developed a method which is efficient to take care of the
defects in the estimation of CF and NFE by proximate analysis. He partitioned the
carbohydrates into various fractions by a system of analysis using detergents. This method
makes use of the concept that the dry matter of plant origin consists of two principal parts.
(i) Cell wall: This includes cellulose, hemicellulose, fibre bound nitrogen, lignin and
acid insoluble ash (silica). This fraction is insoluble in neutral detergent solution but
soluble in acid detergent solution.
(ii) Cell contents: Consists of soluble carbohydrates, starch, pectin, soluble nitrogen,
organic acids and minerals. This fraction is soluble in neutral detergent solution.
Estimation of Neutral Detergent Fibre (NDF)
Principle:
The feed sample is treated with neutral detergent solution, which dissolves the cell
contents and the residue left is known as neutral detergent fibre.
Apparatus needed:
(i) Refluxing apparatus
(a) Spoutless beaker
(b) Round bottom flask as condenser
(ii) Sintered glass crucible (grade-I)
(iii) Electronic balance
(iv) Vacuum pump
(v) Hot plate
(vi) Wash bottle
(vii) Hot air oven
(viii) Muffle furnace

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Reagents needed:
(i) Neutral detergent solution for 1 litre solution
(a) Sodium lauryl sulphate :30 g
(b) Disodium ethylene diamino tetra acetate (EDTA) dehydrate :18.61 g
(c) Sodium borate decahydrate (Na2B4O7.10 H2O) :6.81 g
(d) Disodium hydrogen phosphate (anhydrous) (Na2HPO4) :4.56 g
(e) 2-ethoxyethanol (ethylene glycol mono ethyl ether) :10 ml
(f) Distilled water :upto 1 L
(ii) Acetone
(iii) Decalin
Preparation of Neutral Detergent Solution (NDS):
(i) Put EDTA and Na2B4O7.10H2O together in a large beaker. Add around 500 ml
distilled water and heat until dissolved.
(ii) Add disodium lauryl sulphate and 2-ethoxyethanol in the same beaker and mix
thoroughly.
(iii) In another beaker, put Na2HPO4and add around 200 ml distilled water. Heat it
until dissolved.
(iv) Then mix both the solutions and make the volume 1 L in a volumetric flask.
(v) Finally check the pH range 6.9 to 7.1. If the solution is properly made, pH
adjustment is not required.
Procedure:
(i) Take 0.5 g air dry sample (1mm) in a spoutless beaker.
(ii) Add 100 ml of ND solution and 2 ml decalin to it.
(iii) Reflux for 60 minutes from the unset of boiling.
(iv) Filter the reagent through a pre-weighed sintered crucible, wash thrice with
hot distilled water and finally twice with acetone under vacuum.
(v) Dry crucible at 100 °C for overnight and weigh it after cooling in a desiccator.
(vi) Express the recovered NDF on percent basis.
Observations:
(i) Weight of feed sample =Wg
(ii) Weight of empty crucible =W1 g
(iii) Weight of crucible + NDF =W2 g

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Calculation:
(W 2 – W 1 )
NDF %= [ W ] ×100

Estimation of Acid Detergent Fibre (ADF)

Principle:
The feed sample is treated with acid detergent solution, which dissolves the cell
content and hemicelluloses and the residue left is known as acid detergent fibre (ADF). It
contains cellulose, lignin and silica.
Apparatus:
(i) Same as required for NDF estimation.
Reagents needed:
(i) Acid detergent solution
(ii) Acetone
(iii) Decalin
For 1 litre solution:
a) Cetyltrimethyl ammonium bromide (CTAB) : 20g
b) 1N H2SO4: Upto 1 litre
Preparation of Acid Detergent Solution:
a) Analytical grade H2SO4 (specific gravity 1.84 and purity 97%) is standardized to 1N
H2SO4. (Take 27.42 ml of H2SO4 and make the volume upto 1 litre.)
b) Put CTAB in a beaker and add around 500 ml of 1N H2SO4 in it. Thoroughly mix it
and make the volume 1 litre by the 1N H2SO4.
Procedure:
i. Take 0.5 g feed sample (1mm) in the spoutless beaker.
ii. Add 100 ml of acid detergent solution and 2 ml of decalin to it.
iii. Reflux it for 60 minutes from the unset of boiling.
iv. Filter on a previously weighed crucible with washing with hot distilled water thrice
and finally with acetone twice with the help of a suction pump.
v. Keep the crucible in hot air oven at 1000C for overnight and take the weight of the
crucible after cooling in desiccator.
Observations:
i. Weight of sample =Wg
ii. Weight of empty crucible = W1 g
iii. Weight of crucible + ADF = W2 g

44
Calculations:
( W 2 – W 1)
ADF %= [ W ] × 100

Estimation of Acid Detergent Lignin (ADL)

Principle:
The acid detergent fibre (lingo cellulose + acid insoluble ash) is treatedwith 72%
(W/W) H2SO4, which dissolves the cellulose. The ADL can then be determined by the loss in
weight due to ignition.
Apparatus:
(i) Glass tray (ii) Same required for DNF except refluxing apparatus
Reagents needed:
(i) 72% (W/W) H2SO4 (ii) Acetone
Preparation of 72% (W/W) H2SO4:
Distilled water : 280 ml
Conc. H2SO4 : 400 ml
Take 280 ml of distilled water in a large flask and then add 400 ml of conc. H2SO4
slowly with stirring. The flask must be cooled in water bath(sink).
Procedure:
(i) Place the crucible containing ADF on the glass tray.
(ii) Fill the crucible (about half) with cooled H 2SO4 and stir it with a small rod
breaking all lumps.
(iii) As acid drain away, refill the crucible with 72% H2SO4 and stir it occasionally.
(iv) After 3 hours filter the crucible with washings with hot distilled water thrice
andfinally with acetone twice with the help of suction pump.
(v) Keep the crucible in hot air oven at 100°C for overnight and take the weight
after cooling in desiccator.
(vi) Ignite the crucible in muffle furnace at 500to 550°C for 3 hours andtake the
weight after cooling in desiccator.
Observations:
(i) Weight of sample =Wg
(ii) Weight of crucible + ADL = W1 g
(iii) Weight of crucible + ash = W2 g

Calculation:

45
 (W 1 – W 2 )
ADL %= [ W ] ×100

Calculation of hemicelluloses and cellulose:

Hemicellulose = NDF – ADF
Cellulose = ADF- ADL
Study Questions:

Instructor’s Signature
Date:

46
Exercise – 15 Date________
QUALITATIVE DETECTION OF UNDESIRABLE CONSTITUENTS AND
COMMON ADULTRENTS OF FEEDS (ANTI-NUTRITIONAL FACTORS)
Anti-nutritional Factors:
Anti-nutritional factors are defined as those generated in natural feedstuffs by the
normal metabolism of the species from which the material originates and by different
mechanisms exerting effects contrary to optimum nutrition. These anti-nutritional factors are
often referred as toxic factors as they produce deleterious effects when eaten by animals.
1. Nitrates and Nitrites
Nitrate rather nitrite is toxic to animals. In ruminants, nitrate is readily reduced to nitrite.
It oxidises the ferrous ion of haemoglobin to ferric state, producing brown pigment
“Meth-haemoglobin”, which is incapable of transferring oxygen to the body tissues and
toxic symptoms are seen when 30% of haemoglobin is converted.
Source:
Nitrate is present usually in the form of potassium nitrate. The total nitrate
content in feeds fluctuates between 0.04 to 4.5%. The common plants known
to accumulate nitrate to dangerous levels are sweet clover, Johnson grass, oats,
rape, alfalfa, rye grass, sudan grass, wheat and corn. Acute poisoning occurs
when the fodder contains more than 1 % nitrate (DM basis) or the water with
1500 ppm.
Principle:
When nitrate-nitrite present in the plant sample is allowed to react with diphenylamine
under highly acidic conditions, the intensity of the colour developed is directly proportional
to the presence of nitrate/nitrite in the feedstuff.
Material needed:
(i) Fresh fodder crop
(ii) 2% diphenylamine solution prepared in conc. sulphuric acid
Procedure:
(i) Prepare extract of the plants by macerating 2-3 g of fresh fodder crop with water
in the ratio of 1:5.
(ii) Place 5-6 drops of 2% diphenylamine solution on a glass plate.
(iii) Put 2-3 drops of the plant extract on the above solution and mix it.
(iv) Simultaneously, run a control and compare the colour development (conversion of
green extract to blue colour) with the control.

47
2. Tannins:
Tannins are polyphenolic compounds which precipitate proteins from aqueous
solution.
Source:
The main sources of tannins are tree leaves, beans, salseed meal, mahua cake.
Principle:
When tannin is allowed to react with vanillin, it produces a typical pink colour. The
intensity of the colour is directly proportional to the tannin content of the feedstuff.
Materials needed:
(i) Fresh leaves/feedstuffs
(ii) Vanillin prepared in methanol (4 g vanillin in 100 ml methanol)
(iii) 1N HCl
(iv) Filter paper
Procedure:
(i) Crush the fresh leaves or prepare the extract of the feedstuffs.
(ii) Dip the filter paper in the extract or moisten it.
(iii) Then dip the filter paper in vanillin for 15 seconds and then for 5 seconds in
1N HCl.
(iv) The development of pink colour within 15 minutes to one hour indicates
presence of tannins, which may be compared with a control.
3. Hydrocyanic acid:
The plant generally contains cyanogenicglucosides like amygdalin (glucoside of
benzaldehyde cyanohydrin), dhurin (glucoside of β-hydroxybenzaldehyde
cyanohydrin) and linamarin (glucoside of acetone cyanohydrin) which can be
hydrolysed to prussic acid (HCN) by the same plant when the plants are damaged or
digested by the animals.
Source:
Hydrocyanic acid in trace amounts is fairly wide spread in the form of glucosides but
relatively high levels are found in jowar (sorghum), sudan grass, maize, linseed and
cassava root.
Materials needed:
(i) Put one gram of crushed feed material in the test tube and add 5-8 drops of
chloroform or toluene.
(ii) Dip the filter paper in picric acid solution, dry it and again dip it in sodium

48
 carbonate solution and again dry it.
(iii) Suspend the filter paper in side test tube with the help of rubber cork and pin.
(iv) Incubate it at about 38-40°C overnight.
(v) The intensity of colour changing from yellow to reddish brown indicates the
amount of cyanogens present in the sample, while comparing it with a control
sample.
4. Saponins:
Saponins are glycosides and occur in wide variety of plants. They can be
isolated by extraction from plant material with hot water or ethanol. Upon complete
hydrolysis with acids, strong bases or enzymes, they yield a lipid soluble aglycone,
which are either steroids or triterpenoids and sugars [hexoses, pentoses and saccharin
(uronic) acids].
They are bitter to taste, form foam in aqueous solution and haemolyse red
blood cells. They tend to alter the permeability of the cell wall and exert toxicity
effects. Saponins are highly toxic to cold blooded animals.
Source:
The common plants which contain saponins include lucerne, soybean, tea
leaves, mahua cake, beet root and clovers.
Principle:
The lipid soluble aglycone and water soluble glycone parts of the saponin jointly
confer lower surface tension and form stable foam in aqueous solution.
Materials needed:
(i) Feedstuffs/fodder
(ii) Test tube
Procedure:
(i) Prepare aqueous extract of the sample in the test tube and shake it.
(ii) Formation of persistent foam in the liquid surface indicates the
presence of saponins, which Ca be compared with a control.
Adulterant:
These are the substances which are mixed with the pure feed ingredients to earn more
profit through increase in weight, increase in CP content etc.
1. Salt in fish meal:
Salt is generally added to fish meal to increase the weight. More than 4% salt in fish
meal causes many abnormalities in birds.

49
 Principle:
When the sample containing salt (chloride ions of NaCl/Kcl) is allowed to react with
silver nitrate in presence of potassium chromate solution, the degree of brick red
precipitates formed indicates the absence of salt in the feed sample.
Materials needed:
(i) Fish meal
(ii) 0.1N silver nitrate
(iii) Saturated potassium chromate
(iv) Beaker
(v) Heater
(vi) Muslin cloth
Procedure:
(i) Prepare extract of the sample by boiling 2g of the ground sample with
90 ml water and filtering it.
(ii) Take 10 ml extract in a beaker.
(iii) Add 3-4 drops of saturated potassium chromate solution in it.
(iv) Add 0.1N AgNO3 solution drop wise.
(v) The intensity of brick red precipitate indicates the absence of salt
(NaCl/KCl) in the sample.
2. Urea in feed sample:
Urea solution is generally sprayed on poor quality cakes to increase the nitrogen
content. The BIS specification allows only 1% urea in compounded cattle feed while
in poultry feed, the urea should not be added.
Principle:
When the sample containing urea is allowed to react with dimethyl amino
benzaldehyde reagents in the presence of Carrez solutions (zinc acetate in glacial acetic
acid) and 2 (potassium ferrocyanide) solution, the intensity of yellow colour indicates the
amount of urea present in the feed.
Procedure:
(i) Prepare extract of the feed sample by mixing 1g sample, 5 ml each of Carrez-
1 and 2 solution, distilled water upto 500 ml in a volumetric flask and filtering
it after 30 minutes.
(ii) Take 5 ml filtrate in a test tube.

50
 (iii) Ad 5 ml DMAB reagent in it and shake well.
(iv) The degree of yellow colour indicates the amount of urea present in the
sample.
(v) Compare it with a control sample.
Common adulterants of different feed ingredients
Sr. No. Feed ingredient Adulterant
1. Groundnut cake Ground husk
Urea
Non-edible oil cakes
2. Mustard cake Argemone (Argemonemoxicom) seeds
Fibrous feed ingredients
Urea
3. Soybean meal Urea
4. Wheat bran Ground rice hulls
Saw dust
5. Fish meal Common salt
Urea
6. Mineral mixture Common salt
Marble powder
Sand
Limestone
7. Molasses Water

Study questions:

Instructor’s Signature
Date:
Exercise-16 Date:____________
CALCULATION OF NUTRITIVE VALUE OF DIFFERENT FEEDSTUFFS

51
A. Evaluation of a sole ration:
Chemical composition of a ration though is a good index of its nutritive value but its
nutritional worth for particular specie of animal is based on its digestion and metabolism in
the body. Nutritional value of a ration is determined from digestibility, digestible crude
protein, total digestible nutrients, nutritive ratio and starch equivalent values for livestock and
poultry.
Objective:
 To determine the nutritive value viz, digestibility coefficients, digestible crude protein
(DCP), total digestible nutrients (TDN), nutritive ratio (NR) and starch equivalent
(SE) of feeds and fodders.
Procedure:
For estimating the nutritive value of feeds and fodders first step is to conduct a
digestion/metabolic trial. Record the average daily feed intake and average daily faecal
material voided. Analyse the feeds and dung voided for dry matter, crude protein, crude fibre,
ether extract, ash contents and indicator (if added). The digestibility can be calculated directly
by using total collection method and indirectly by using an indicator.
Digestibility coefficients (DC) of nutrients:
A. Direct method
Feed int ake−faeces voided
×100
Digestibility coefficients of DM = Feed int ake
When there is feed reside them
Feed offered − Feed refused faces voided
×100
Digestibility coefficients of DM = Feed offered −Feed refused
Digestibility coefficients of Nutrient =
Feed int ake×Nutrient in feed−faeces voided×Nutrient in feaces
×100
Feed int ake×Nutrient in feed
B. Indirect method (indicator method)

% indicator in feed % nutrient in faeces

% DC of a nutrient =
(
100− 100× ×
% indicator in faeces % nutrient in feed )

Determining nutritive value:

1. Digestible nutrients:

52
% Digestible nutrient in a diet/ration =

% nutrient in the diet /ration×% DC of nutrient

100
2. Total digestible nutrients (TDN):
TDN = % digestible crude protein (DCP) + % digestible nitrogen free extract +
% digestible crude fibre (DCF) + % digestible ether extract (DEE)  2.25
3. Nutritive ratio (NR):
% DNFE+% DCF +DEE×2.25 % TDN−% DCP
NR = % DCP or % DCP
4. Starch equivalent (SE): It is the per cent ratio of fat deposited per unit of feed to the fat
deposited per unit of starch.
Weight of fat deposited /weight of feed
×100
SE = Weight of fat deposited /starch
Based on the fat producing power of various nutrients, Kellener calculated the starch
equivalents for each nutrient as follows:
Digestible nutrient Factor for starch
equivalent
Crude protein 0.95
Starch (CF + NFE) 1.00
Ether extract (roughages) 1.91
Ether extract (gains/brans/gain by products) 2.21
Ether extract (oil seeds, oil cakes and animal products) 2.41
Correction factor for crude fibre
Type of roughage % of CF to be multiplied
by factor
Dry roughages (straw, hay etc) unchopped 0.58
Dry roughages finally chopped 0.29
Green fodders (on fresh basis) % CF
4 0.29
5 0.31
6 0.34
7 0.36
8 0.38
9 0.40
10 0.43
11 0.45
12 0.48
13 0.50
14 0.53

53
 15 0.55
16 or more 0.58
Under practical conditions SE can be calculated as follows:
SE = % DCP  0.95 + % DEE  (1.91/2.21/2.41) + % DNFE  1.0 + % DCF  1
Example: Find the nutritive value of the berseem from the following table:
Per cent chemical composition
DM CP CF* EE Ash NFE
Berseem intake (kg) 50 20 16 24 3 5 52
Faeces voided (kg) 20 20 12 28 3 5 50
Solution
I. Calculations for digestibility coefficients
Daily average DM CP CF EE NFE
Berseem (kg) (A) 50×20 10×16 10×24 10×3 10×52
100 = 100 100 100 100
10.0 = 1.6 = 2.4 = 0.3 = 5.2
Faeces (B) 20×20 4×12 4×28 4×3 4×50
100 = 100 = 100 100 = 100
4.0 0.48 = 1.12 0.12 =2
Nutrients 10-4 = 6.0 1.6-0.48 = 2.40-1.12 0.30-0.12 5.2-2.0 =
digested (kg) 1.12 = 1.28 = 0.18 3.2
% digestibility 6×100 1.12×100 53.3 60.0 61.54
coefficient 10 = 1.6
60 = 70

II. Calculation for Total Digestible Nutrients and Starch Equivalent

% composition % digestibility % digestible Starch

of feed coefficient nutrients equivalent
Crude protein 16 70.0 16×70 11.2  0.95 =
10.64
100 =
11.20
Crude fibre 24 53.30 24×53.3 12.79  1 =
100 12.79
=
12.79
NFE 52 61.54 52×61.54 32  1 = 32
100 =

54
 32.0
EE* 3 60.00 3×60 1.80  1.91 =
3.438
100 = 1.80
DM 20 60.00 60×20 –
100 =
12.00
TDN* – – 60.04 –
SE – – – 56.83
Study Questions:
Q.1: Why we get the apparent digestibility coefficient rather than the true digestibility?
Q.2: Why in chicken, the term dry matter metabolizability rather than dry matter
digestibility is used?
Q.3: Why the correction factor for crude fibre in calculating the starch equivalent is used?
Exercise 4. A bullock was fed 4.0 kg hay / day and voided 5.8 kg faeces. Find the
digestibility coefficients, DCP, TDN, NR and SE from the following data:
Percent
H2O CP CF EE Ash NFE
Hay 16.00 13.00 28.00 2.50 6.00
Faeces 75.00 3.50 10.00 0.54 2.00

Exercise 5. Find the digestibility coefficients of various proximate components of feed

fed to growing pigs from the data given below:
Intake/ Outgo DM CP EE CF Ash NFE
Feed 763 96.0 20.0 6.0 7.5 7.0
Faeces 475 30.0 24.0 8.0 21.0 22.0
Exercise 6. Determine the metabolizability of the nutrients and digestibility of crude
protein in chicken from the following data:
Intake/ DM CP EE CF Ash NFE
Outgo
Feed 110 90 16 7.0 4.0 8 0.4
Dropping 62 45 21 20.0 2.0 25 2
Intestinal contents – – 20 – – – 2.5
(5th segment)

55
 Instructor’s Signature
Date:

Exercise – 17 Date:_____________
CALCULATION OF DE, ME & NE FOR BALANCE OF NUTRIENTS
Energy is defined as the capacity to do work. Energy is available in chemical,
thermal, electrical and radiation forms. In the feeds, it is present in the chemical form which
on consumption by the animal is broken down in the living system to release energy for doing
mechanical work, maintaining the integrity of cell membranes and maintaining the body
temperature. Energy released in the body is used with different efficiencies in various animal
species for different physiological functions. Energy value of a feed for different species of
the livestock and poultry are different. In the body, energy is partitioned as given below:

56
Objective:
 To determine the digestible, metabolisable and net energy of feeds for different animals.
Procedure:
Energy value of different feeds is determined by conducting a metabolic trial. From
the exact intake of feed, faeces and urine excreted, the DE and ME values are calculated.
Gross energy (calorific value) of feed, faeces and urine are obtained by completely burning
the sample in a bomb calorimeter. Calculations for livestock are made as given below:
GE i−GE f
DE / kg diet = Kg feed int ake
DE −(GE u +GPD )
ME / kg diet = Kg feed int ake
Where
GEi = Gross energy intake (through feed) Kcal / g
GEf= Gross energy of faeces (Kcal/g)
GEub = Gross energy of urine (Kcal/ml)
GPD = Gaseous product of digestion

57
 In poultry birds, there is common opening for faeces and urine thus the ME is
calculated as:
GE i−GE f
ME / kg diet = Kg feed int ake
In pigs, GPD is very small thus it is negligible therefore the ME is the difference of
DE and urinal energy. GPD for different animals is different and is calculated for cattle and
sheep from the following equations:
E = 4.012X + 17.68 g (cattle)
E = 3.41X + 9.89 g (sheep)
Where
E is methane production (1 g methane produces 13.34 Kcal)
X is digestible carbohydrates in 100 g i.e. sum total of digestible CF and Digestible
NFE.
NE = ME – Heat Increment (HI)
The heat increment of digestion is approx. 10% of GEi
Example-1: Calculate the DE, ME and NE for cattle from the following data:
Feed /Faeces Intake / GE (Kcal/g or Digestible CF Digestible NFE
Voided ml)
Berseem 10 4.0 1.30 2.56
Faeces 4 4.2 – –
Urine 15 litres 0.14 – –

DE (Kcal) = 10  1000  4.0 – 4  1000  4.2

= 40,000 – 16,800 = 23,200 Kcal = 23.2 MCal
ME = DE – (UE + GPD)
GPD = [4.102  (Dig. CF + Dig. NFE) + 17.68  Calorific value of
methane/g (13.34)
GPD = [4.012  (13 + 25.6) + 17.68]  13.34
= [4.012  38.6 + 17.68]  13.34
= (154.86 + 17.68)  13.34
= 172.54  13.34 = 2301.68 = 2.3 MCal
ME = DE – (UE + GPD)
= 23.2 – 2.3 = 17.4 MCal
NE = ME – HI

58
 = 17.4 – 10% of GEi
= 17.4 – 10X / 100  40.0
= 17.4 – 4 = 13.4 MCal
Example-2: Determination of ME for poultry and pigs
Poultry Pigs
a b ab a b ab
Feed/ Energy Total Feed/ Energy Total
Faeces/ Kcal/ g or Faeces/ Kcal/ g
Urine ml Urine or ml
Intake (g) 88.0 3.60 316.8 1600 4.5 7200
Faeces (g) 33.0 3.40 112.2 450 3.6 1620
Urine (ml) – – – 1050 0.18 189
Total – – – – – 7200 – 1620
digestible = 5580
energy
Total ME – – 316.8 – – – 5580 – 189 =
112.2 = 5391
204.6
DE (Kcal/ – – – – – 5580 1.6 =
kg diet) 3487.5
ME (Kcal/ – – 204.6  – – 5391  1.6 =
kg diet) 0.088 = 3369
23.25
DE
×100
DE as per cent of GEi =
GE i

ME
×100
ME as per cent of GEi =
GE i
ME i
×100
ME as per cent of DEi = DE i

Study Questions:
1. Find out the ME as % of DE intake using the date of Practical No.3, Problem 1.
2. Determine the ME Kcal / kg for broilers from following data:
DM intake 125 g/day, droppings voided 47 g with GE of 3.80 and 2.10 for feed and
droppings, respectively.
3. Calculate the ME as % of DE in pigs from the following data:

59
 DM intake 1650 g, faeces 475 g and urine 1200 ml per day. The GE of feed, faeces and
Urine are 4.75, 3.5 and 0.15 Kcal per g or ml.

Instructor’s Signature
Date:
Exercise-18 Date:______________
CALCULATING THE NUTRIENT REQUIREMENTS OF AN ANIMAL
The base of scientific feeding is the knowledge of nutrient requirements of an animal /
bird. The animal / bird needs nutrients for two major components in their bodies i.e.
1. Maintenance
2. Production
The maintenance requirements vary with the animal body weight while production
includes the growth in growing animals, milk, meat, egg and wool production and
reproduction. In monogastric animals (pigs, poultry birds etc.) the requirements for
maintenance and growth are not partitioned and are calculated together as a single
component. While in ruminants the requirements are considered separately. The requirement
for growth varies with the rate of growth which is proposed to be achieved. For ruminants the
requirements for maintenance, growth, milk production as per the fat content of milk and
wool production (sheep) for different species are given in tables in Annexure. It may be noted

60
here that there is a growing trend away for using the total digestible nutrient system for
expressing the useful energy for feeds and the energy needs of the animal. Modern feeding
standards state digestible energy (DE) or metabolizable energy (ME) needs in term of
kilocalories. DE and/or ME figures can be computed from the commonly accepted average
equivalent:-
1 lb TDN = 2,000 Kcal DE = 1640 Kcal ME
(0.4536 kg)
or 1 kg TDN= 4.409 McaI DE = 3.62 Mcal ME
Objective:
 To determine the actual daily nutrients needed for maintenance, growth (growing
animals), milk production (lactating animals), wool production (sheep) and
reproduction of the animals.
Procedure:
 Record the weight of the animal.
 Note the physiological status of the animal viz. lactating, gestating.
 Note down the, daily requirement of the animal for DCP, TDN &.ME as per its body
weight.
 Record the rate of growth if the animal is growing.
 Note down the DCP and TDN requirements from the tables for DCP and TDN (The
requirements for growth and maintenance for growing animals are given together).
 Check the milk yield and fat per cent of the milk (in case of lactating animals).
 Note down the requirements from the table.
 Check the stage of gestation and record requirements. The animals in the advance
stage of pregnancy need extra allowance for fetus development.
 Add all the requirements of all the nutrients separately for maintenance, growth,
lactation and gestation.
Example-I
Calculate the requirements for the following growing animals:
1. Cow weighing 260 kg with a daily growth rate of 550 g.
2. Buffalo weighing 260 kg with a daily growth rate of 550 g.
3. Growing lamb 20 kg with growth rate of 100 g.
4. Growing kids 15 kg with growth rate of 150 g.
Solution:

61
 See the tables of requirements of each type of growing animal. The values for
different animals are tabulated as under:
Animal Body Growth Requirement for
weight rate (g) DM DCP TDN Calcium Phosphorus
(kg) (kg) (g) (kg) (g) (g)
Cow 260 550 6.70 350 3.70 20.0 16.0
Buffalo 260 550 6.50 420 3.60 20.0 16.0
Lamb 20 100 0.68 63 0.41 3.4 2.3
Kids 15 150 0.785 53 0.51 4.2 2.8

Example-II
Determine the requirements for a buffalo / cow weighing 450 kg yielding 10 litres of
milk with 6% fat.
Phase of animal Requirement of
DM DCP (g) TDN (kg) Calcium (g) Phosphorus
(kg) (g)
Maintenance 6.0 280 3.4 18 14
Milk production – 57  10 = 570 0.41  10 = 3.1  10 = 31 2.1  10 = 21
10 litres 4.10
Total requirement 6.0 850 7.8 49 35

Study Questions:
1. Determine the requirements of a buffalo weighing 550 kg and producing 9 litres of milk
with 7.5% fat.
2. Determine the nutritional requirements of a lamb/kid weighing 25 kg and gaining 150 g
per day.

62
 Instructor’s Signature
Date:
ANNEXURE-I

COMPOSITION AND NUTRITIVE VALUE OF COMMON INDIAN FEEDING

STUFFS ON DRY MATTER BASIS

Ingredients C.P. E.E. C.F. Ash N.F.E D.C.P. T.D.N. M.E.

% % % % . % % (Mcal/kg)
%
Green fodders
Berseem 17.3 1.9 25.9 14.2 40.7 12.8 62.3 2.26
Cowpea 28.1 3.0 26.7 9.2 33.0 20.3 62.2 2.24
Lucerne 20.2 2.3 30.1 10.7 36.7 16.2 60.2 2.17
Guinea grass 7.9 1.2 38.2 15.5 37.0 5.8 65.1 2.34
Jowar 12.8 2.6 26.5 12.7 45.4 7.8 52.0 1.87
Jowar (flowering) 7.6 1.9 30.7 9.4 50.4 3.4 56.3 2.02
Maize (flowering) 6.4 0.9 29.9 11.6 51.2 4.7 71.2 2.56
Maize (dough stage) 5.1 1.5 26.9 7.3 59.2 1.6 65.2 2.34
Oats (milk stage) 6.4 2.3 28.7 9.3 53.3 2.8 55.1 1.98
Napier grass hybrid 11.5 2.2 25.9 15.9 44.5 7.6 60.0 2.16
Paragrass 9.4 0.0 29.3 10.4 50.1 7.9 56.2 2.02
Bajra 8.8 1.9 24.9 8.2 56.2 5.3 55.4 1.99
Guar 18.1 1.9 31.9 10.4 37.7 12.0 60.0 2.16
Silages
Jowar 5.9 1.8 37.3 10.6 44.4 2.4 51.1 1.84
Maize 7.9 1.1 24.6 11.3 55.1 3.4 61.1 2.20
Oats 8.5 2.0 30.7 13.7 45.1 4.1 62.6 2.23

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Hays
Berseem 14.7 1.6 30.6 12.1 41.0 10.3 65.8 2.36
Lucerne 21.3 1.4 29.4 12.7 35.2 16.4 55.9 2.01
Cowpea 11.5 1.4 32.8 8.6 45.8 7.6 55.6 2.00
Straws/stovers
Bajra 3.1 1.2 40.4 6.8 48.5 0.9 53.4 1.92
Barley 2.2 0.9 47.4 8.1 41.4 0.5 44.5 1.60
Ragi 3.7 0.5 36.5 8.6 50.7 0.2 52.6 1.89
Paddy 3.8 1.6 30.6 20.4 43.6 0.0 44.5 1.60
Wheat 2.4 1.0 39.6 10.1 46.9 0.0 48.9 1.76
Grains
Arhar (chuni) 16.2 4.3 16.7 5.7 57.1 14.4 74.1 2.67
Cotton seed 19.5 18.3 19.8 5.6 36.8 12.9 88.8 3.20
Gram (chuni) 19.6 4.8 7.5 2.6 65.5 13.6 87.5 3.15
Bajra 11.6 5.0 1.4 2.9 79.1 5.2 60.6 2.18
Jowar 12.5 3.4 2.2 2.0 79.9 6.1 85.7 3.08
Maize 10.6 3.3 2.2 1.8 82.1 7.0 87.1 3.13
Oats 8.8 1.9 18.3 8.5 62.5 4.4 71.7 2.58
Rice (broken) 7.0 0.4 0.6 4.6 87.4 5.6 90.6 3.26
Wheat 10.5 1.9 1.9 1.9 83.8 6.3 92.3 3.32
Ragi 10.3 1.2 3.7 3.9 80.9 6.2 78.6 2.82
Oilseed cakes
Cotton seed (decorticated) 38.5 9.2 6.3 8.0 38.0 31.6 86.0 3.09
Groundnut (solvent ext.) 38.3 1.2 4.2 7.5 31.8 49.1 77.0 2.78
Rape (solvent ext.) 53.3 0.8 12.6 9.2 42.1 32.3 86.8 3.12
Sesame 42.5 4.7 5.2 12.2 35.4 34.0 80.0 2.52
Soybean 48.0 5.2 7.4 6.3 33.8 41.4 85.0 3.07
By-products
Gram husk 3.7 0.9 50.4 6.0 39.0 0.0 61.3 2.21
Maize gluten 48.2 8.6 0.5 2.3 40.4 39.1 84.1 3.70
Molasses 4.3 0.1 - 10.8 84.8 2.4 60.0 3.00
Guar-meal 46.7 5.1 6.0 7.2 53.0 42.5 83.5 3.86
Rice bran 14.0 14.4 14.1 15.8 41.7 9.1 76.1 2.74
Wheat bran 11.5 2.9 12.7 10.4 62.5 8.7 70.4 2.53
Tapioca chips 2.9 0.4 10.9 9.5 76.3 1.5 83.3 3.68

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 MANUAL OF
PRACTICALS IN ANIMAL NUTRITION
For B.V.Sc. & A.H.
As per VCI minimum standards of veterinary education Degree
course regulation 2016

BY
DR. JASMINE KAUR
DR. J.S. LAMBA
DR. R.S. GREWAL
DR. J.S. HUNDAL
DR. UDEYBIR SINGH
DR. A.P.S. SETHI
DR. M. WADHWA

DEPARTMENT OF ANIMAL NUTRITION

COLLEGE OF VETERINARY SCIENCES
GURU ANAND DEV VETERINARY AND ANIMAL SCIENCES
LUDHIANA-141004

2017

65
 INDEX

Sr. Title Page Date of Date of Instructor's

No. Practical Submission Signature
1. GENERAL PRECAUTIONS WHILE 1-4
WORKING IN NUTRITION
LABORATORY

2. FAMILIARIZATION WITH VARIOUS 5-8

FEEDSTUFFS

3. PREPARATION AND PROCESSING 9-12

OF SAMPLES FOR CHEMICAL
ANALYSIS
(Herbage, faeces, urine and silages)

4. PREPARATION OF SOLUTIONS 13-18

( A) Principles of volumetric analysis

5. PREPARATION OF STANDARD 19-21

SOLUTION
(B) Preparation of Standard Solution

6. WEENDE’S SYSTEM OF ANALYSIS 22-23

(PROXIMATE ANALYSIS)

7. TO DETERMINE MOISTURE 24-25

CONTENT OF FEEDS AND
FODDERS
(OVEN DRYING METHODS-A
COMMON PROCEDURE)

8. (A) TO DETERMINE THE TOTAL 26-27

ASH CONTENT OF FEEDS AND
FODDERS
(B) TO DETERMINE THE ACID-
INSOLUBLE ASH CONTENTS OF
FEEDS AND FODDERS

9. TO DETERMINE THE CRUDE 28-30

PROTEIN CONTENT OF FEEDS AND
FODDERS

66
10. TO DETERMINE THE ETHER EXTRACT 31-32
CONTENT OF FEEDS AND FODDERS
(SOXHLET EXTRACTION METHOD)
AND TO CALCULATE NITROGEN FREE
EXTRACT (NFE)

11. TO DETERMINE THE CRUDE FIBRE 33-34

CONTENT OF FEEDS AND FODDERS

12. TO DETERMINE THE CALCIUM 35-37

CONTENT OF FEEDS AND FODDERS

13. TO DETERMINE THE 38-39

PHOSPHOROUS CONTENT OF
FEEDS AND FODDERS

14. DEMONSTRATION OF THE 40-44

DETERGENT METHOD OF FORAGE
ANALYSIS (VAN SOEST SCHEME
OF ANALYSIS)

15. QUALITATIVE DETECTION OF 45-49

UNDESIRABLE CONSTITUENTS
AND COMMON ADULTRENTS OF
FEEDS (ANTI-NUTRITIONAL
FACTORS)

16. CALCULATION OF NUTRITIVE 50-54

VALUE OF DIFFERENT
FEEDSTUFFS

17. CALCULATION OF DE, ME & NE 55-58

FOR BALANCE OF NUTRIENTS

18. CALCULATING THE NUTRIENT 59-61

REQUIREMENTS OF AN ANIMAL

19. ANNEXURE 62-63

67
 FORWARD

The advancement in the livestock sector is only possible by scientific feeding of

balanced ration to various categories of livestock and poultry by providing the desired
nutritional environment for exploiting their genetic potential to the maximum, as feed cost
represent 70-75% of the total cost of production. It has direct effect on profitability from the
animal enterprises. To maintain the standard of feeding, the feed ingredients to be used in
formulating the rations/diets for the different categories of livestock and poultry should be
evaluated nutritionally. Analytical techniques are powerful tools in the pursuit of scientific
teaching and research. Animal Nutrition a course of undergraduate students is the basic
teaching programme which includes familiarization with various feed stuffs, preparation and
processing of samples for chemical analysis, proximate analysis for the feeds and fodders,
detection of anti-nutritional factors and adulterants, etc. All these topics have well taken care
of by the authors in the preparation of this manuscript. Complete knowledge of all these
aspects will make the students of B.V.Sc. & A.H. competent enough. I greatly appreciate the
efforts made by the authors bringing Animal Nutrition Manual for under graduate course in
complete and concise manner. I am sure this manual shall be of immense use to the student,
teachers and research workers. I feel pleasure in recommending this manual for the use in the
teaching of B.V.Sc. & A.H. as per Veterinary Council of India-minimum standard of
veterinary education - Degree course (B.V.Sc. & A.H.) regulation 2016.

Dean
College of Veterinary Science
GADVASU, Ludhiana.

68
 PREFACE

The purpose of this manual is to fulfill the needs of the students, teachers and research
workers for the practical portion of the undergraduate course of Animal Nutrition as per the
Veterinary Council of India-minimum standards of veterinary education-Degree course
(B.V.Sc. & A.H.) regulation 2016.

The manual is divided into different aspects mentioned in VCI regulation 2016.
Starting with information on Animal Nutrition Laboratory, familiarization with various
feedstuffs, preparation and processing of samples for chemical analysis, Weende's system of
analysis for feeds and fodders, Demonstration of Van Soest scheme of analysis, Qualitative
detection of undesirable constituents and common adulterants. An attempt has been made to
provide detailed procedures along with material required observation and calculation.
Authors hope that the manual will provide the desired information to undergraduate students,
teachers and research workers.

Authors are highly thankful to Dr. Manju Wadhwa, Senior Nutritionist cum Head,
Department of Animal Nutrition for providing her valuable guidance and moral support in
preparing the manuscript. Authors admit that the manuscript may have some discrepancies
and there is always scope for improvement so suggestions are always welcome.

All authors

69