Soil Testing

Certificate Course in Agricultural Biotechnology

BIOFERTILIZER TECHNOLOGY

Institute team:

Dr. B.K.Datta
Dr. R.Datta
Dr. S.K.Das
Dr. S.K.Si
Mr. S. Sahoo
Mr. D. Biswas
Mr. S. Giri
Mr. T. Nayek

Expert Consultants:

Dr. P.K. Singh, IRAI, New Delhi
Dr. O.P. Rupela, ICRISAT, Hyderabad
Dr. R.K.Basak, BCKV, Mohanpur, W.B
Dr. D.J. Bagyaraj, Univ. of Agric. Sciences, Bangalore
Dr. R. Kale, Univ. of Agric. Sciences, Bangalore
Dr. Sunil Pabbi, IARI, New Delhi
Dr. Aloke Adholeya, TERI, New Delhi

Vivekananda Institute of Biotechnology

First published in July, 2004

by Vivekananda Institute of Biotechnology
Nimpith, South 24 Parganas, West Bengal, India

Designed by Tellywallah, Calcutta
Printed by Swapna Printing Works Private Limited, Calcutta

All rights reserved.

c Vivekananda Institute of Biotechnology, 2004

This manual is sold subject to the condition that it shall not, by way of trade or otherwise,
be lent, resold, hired out or otherwise circulated without the publisher's prior consent
in any form of binding or cover other than that in which it is published.

Foreword

Fifty years back a monk, Swami Buddhanandji, was deeply inspired by Swami
Vivekananda’s ideals. He started a different kind of journey of life and a seed
of development was sown at Nimpith. Late Swami Buddhanandaji established
this Ashram.

Vivekananda Institute of Biotechnology is a branch of that tree. The
Institute initiated its activities in 1991. Its aim is to develop an advanced,
functional, research as well as a resource center for the people of the Sunderbans.
One of the chosen fields is Agricultural Biotechnology. For the last few years the
Institute has been implementing programmes on entrepreneurship development,
in this field, in rural areas. The present manual is one of such activity to support
this programme, which is the need of the hour.

The Department of Science and Technology, Govt of India and United
Nations Development Programme have come forward benevolently in bringing
out this manual. It is a result of the combined efforts of scientists at our Institute
and of other National Institutes and Universities. Senior scientists like Dr.
P.K.Singh, Dr. O.P.Rupela, Dr. Radha Kale, Dr. Ranjan Basak, Dr. Sunil Pabbi,
Dr. D.J. Bhagyaraj and Dr. Alok Adholeya have enriched this effort by their
valuable guidance and by sharing their experience . Shri Gautam Bose, Shri Amit
Kumar and Shri Pradip Nair of Tellywallah have worked hard and extensively to
make it in this present form.

We hope that this great effort will be used by the rural agro-biotechnologists,
whose services, we believe, will bring a new dawn to rural India.

Nimpith Swami Sadananda
July 2004 Chairman

Preface

In today’s world, technology is moving very fast, in certain sectors at a faster
pace. Biotechnology is such an area. The rural India which is depending on
agriculture for its day to day life provides an immense market for new technologies.
The only condition is the proper training and marketing.

The present manual is the outcome of the project ‘Technical Human
Resource Development – Vocational Training For Employment Generation’
supported by UNDP & DST, Govt. of India. The objective of the programme is
to develop human resources through competency based training in innovative
areas for the production and service sectors in new, high technology areas based
on market needs.

Agricultural biotechnology is the area in which we have worked on under
this project. This manual is first of its kind in this series. It deals with Biofertilizer
Technology which has six modules - Soil testing and fertilizer recommendation,
Production and Application of Blue-green algae, Azolla, Microbial inoculants,
Vesicular Arbuscular Mycorrhiza and Vermicompost.

The target are the rural youth, who have passed their 10th std. It is
designed and presented in such a way that a complicated subject like soil testing
or microbial inoculant production becomes an easily adaptable skill, a demystification
of technology indeed. All the techniques mentioned here are of world standard
but no doubt many other options can also be opted for, for instance, in the
section of microbial inoculant production only the use of vessel is mentioned in
this manual though the use of other types of fermentors or shakers are also
possible.

This effort is the result of hard work of a team of our Institute, the same
was complemented by experts of other organizations, which are of world repute.
Mr. Khudiram Sardar, Mr. Diwakar Haldar, , Mr.Tarun Das, Mr. Deepankar Haldar,
Mr.Tapan Haldar and Mr. Shubankar Malik have helped while filming the experiments
for this manual.

Shri Gautam Bose , Shri Amit Kumar, Shri Pradip Nair and Shri Partha
Bhattacharya of Tellywallah gave their best to make this dream a reality by
filming, designing, adding inputs and finally printing the manual.

Nimpith Dr. B.K. Datta
July 2004 Principal Scientist, VIB

Using this manual

This manual has been written to complement classroom lectures pertaining to
the Biofertilizer Technology Section of the “Certificate Course in Agricultural
Biotechnology” taught at the Vivekananda Institute of Biotechnology — VIB.

However, it may also be read by anyone with school level knowledge of
Science who is interested in setting up a soil testing lab or a microbial inoculant
production facility.

We have departed from traditional styles of writing Course material.
Instead of dry and forbidding lists of procedures and equations, the manual tries
to expose the student to the various aspects of the three disciplines that this
course straddles — Microbiology, Agricultural Science and Chemistry. Of course,
one cannot entirely avoid equations and procedures! But we felt it is possible
to present these topics in a friendlier manner.

Our approach to making the subject “friendlier” and also maintaining
sufficient scientific detail was two-pronged.

We’ve split the material being discussed into two parts — quite literally.
The odd-numbered pages in this manual contain descriptions of experiments
and processes in a step-by-step manner, devoid of detailed explanations — rather
like a traditional lab manual which expects students to follow the steps described
bothering to evoke an interest in the topic at hand.

The objective of the VIB course was the opposite. As scientists we are amazed
and intrigued by our respective fields of study. We wanted the reader to feel this
amazement as well; to undestand that science is an exciting subject!

To help us out, we enlisted the help of Shubham, an inquisitive,
imaginary friend, who can’t stop asking questions. Shubham can
be found loitering around on the even-numbered pages — asking
questions about the text on the facing page. The even-numbered
pages also contain supplementary information about the topic at
hand. Definitions, little bits of trivia, brief forays into the history
of science, suggestions for further reading, tips on how to simplify
a process and so on may also be found on even numbered pages.

The course material was meant to enable an interested student set up
his own Soil Testing Centre and an Inoculant Production Centre. Which is why
the material in this manual is presented in a “modular” fashion — typical of
industrial processes. Therefore, operations like autoclaving, working in a Laminar
Flow Cabinet and using a Fermentor have been dealt with in a separate section.
The step-by-step description concentrates on following the process being discussed
segment-wise.

The line at the bottom of each odd-numbered page is a “station map”.
This helps the students to see the “whole railway line” and on which station he
stands — this is typical of any industrial process where one goes from
A (A test tube of bacteria) to B (application of the inoculant in the field) and the
various stages that must be crossed to do so.

Using this manual

In sections where extra explanation on the left hand pages was unnecessary,
the step-by-step description continues.

Addressing the reader directly (”You could then....” “We might look at
this from another angle....”) is something most textbooks never do but we felt
there was no reason why the manual should not attempt to make the reader
feel as if he was in a classroom while reading it.

The manual is accompanied by an interactive CD-ROM. This contains all
the course material viewable in a non-linear fashion. This enables a student
to quickly refer to topics without having to flip through the manual. Further, all
the “modules” that are part of the production process, were filmed during the
making of this manual and the footage, with narration, is available on the CD.
This could be used in the classroom too, during, say, the first time the topic
is brought up for discussion by the instructor.

The glossary in this manual is available in a searchable format on the
CD-ROM.

The CD-ROM contains two computer programmes written specifically for Soil
Testing Centres. Chemicalc and X-Base are a scientific formula calculator and
a Soil Database, respectively. X-Base allows Soil Testing Centres to share their
data over the Internet with no additional software or hardware requirements
apart from a very rudimentary computer system and a telephone line.

A few of the pages also contain material that is not immediately relevant
to the course (eg., a brief description of the gene cascade in R.Meliloti that leads
to the production of nitrogenase, may be found in the pages that describe
Rhizobia). These portions are italicised. These sections are rare and are only
found, if at all, in the introductory part of the section. These could, if nothing
else, be food for thought for an inquisitive reader.

Biotechnology is the fastest growing scientific field around the world. As
scientists around the world learn more about the amazing internal workings of
living things, literature — this manual included — needs to be updated with a
regularity, which, to a scientist, is more exciting than monotonous. The authors
would greatly appreciate any feedback about this manual.

VIB hopes that this initiative will help fuel the next agricultural revolution
in our country — one powered not just by fertilizers and technology but also by
a more aware and knowledgable farmer who understands the science behind
the word 'biotechnology'...

VIB team

Nimpith
July 2004

Soil Testing - Collection and preparation of soil samples

Composite soil samples, packed for lab. analysis 1-1

Introduction..

A bit about the Soil Collection Process

The process of soil testing begins in the field — where collection of samples
is done. This needs to be done methodically — to ensure that the samples
taken from the field represent the soil characteristics of the entire field.
The process stated here is simple and easy to follow — though it does seem
elaborate when you first read about it!
Locating areas to take samples...

Start at the bottom -left corner of the field and walk
along the path indicated by the arrows. Along the
way, mark areas (with a piece of wood — or anything
easy to locate) from where you will be taking soil
samples. Avoid marking prohibited areas (see the
next page).

The marked areas are shown in Red in the figure.

Notice, that the red dots along the path are not
necessarily on the path itself.
That’s perfect — because this is a random sample.

Apparatus required

Plastic bucket, spade and wooden stakes or markers.

Why is it necessary to go through such an elaborate process?

It’s quite simple — ideally you’d want to test all the soil in your field
— but that would take a lot of time. So, it becomes necessary to take
samples from different places in the field.

But how do you ensure that the samples taken to the testing centre
represent, more or less, the soil in the field? The answer is the reason
behind the elaborate process.

By taking samples from randomly selected spots, you ensure that
your cumulative soil sample represents the soil in most areas of the field. The
zig zag walk is just to establish a technique of selecting random spots.

1-2 Collection and Preparation of soil samples

Collecting soil samples

The locations from where soil samples
are taken may be marked by wooden
stakes as shown. The stakes need not
be numbered.

The locations are chosen by walking
along a zig-zag path starting at any
corner of the field. These are marked by
white spots in the picture.

A typical location is shown in the next
picture.

Each collection location needs to be
cleared of vegetation and dust. This is
done by scraping away a very thin layer
of soil.

Collection Grinding Partitioning Storage

Drying Mixing Sieving

Collection and Preparation of soil samples 1-3

Prohibited samples...

Prohibited Samples?

It’s just another thing to do with the statistics (see page
2.1). Your soil samples need to represent the average soil
characteristic of the field. However, every field has areas
in it that tend to distort the average characteristic because
the soil there has properties very different from the rest
of the soil.

(It’s called deviation from the mean, by the way)

So, you need to ignore these areas when collecting samples.

These include

— Areas near gates, farmways, buildings etc. and areas on crop hills and in rows.
— Areas where organic/chemical manure is or was kept.
— Areas which are permanently in the shade.

Why must the pit be 6” (15 cm) deep?

6” is the depth of tillage i.e., the depth up to which the root system of the crop
penetrates. Since soil testing is carried out to determine the availability of
nutrients to crops, samples i.e., the furrow slices, are 6” long.

Why should a PVC bucket be used?

An iron bucket may have rust, which might contaminate the sample. Presence
of iron would skew the results of the organic carbon test as we shall see later.
Jute or nylon bags which may have been used to store fertilizer etc. must not
be used too.

PVC (or any other plastic) would not contaminate the soil. Besides, they are easy
to clean.



Remove a wedge-shaped lump of soil
from the cleared sampling location and
discard it.

The resulting pit should be about 6 inches
(15 cm) deep.

From the two larger surfaces of the pit,
remove a half inch thick slice of soil —
called the furrow-slice. Thus, from each
sampling location on the field, two furrow-
slices are obtained.

Carry these in a PVC bucket.





After collection — what now?

You now have a bucketful of furrow-slices. However, it would be time-consuming
to test all the soil in each of the furrow-slices separately and then average the
results. Ideally, you would want to test a relatively small sample of soil which,
nevertheless, represented the soil present in the entire field.

This is achieved by thoroughly mixing the soil. Further, to simplify the lab
experiments, the samples are ground and sieved... We’ll discuss this issue in the
next section...



After all the samples are collected, they
could be taken directly to the laboratory
— if one is close enough. Else, the
samples may be prepared on location
itself, and a composite soil sample may
be sent for testing.

The samples are now air-dried in the
shade.




Drying the soil samples

Why are the samples dried?

In dried soil, any reversible chemical reactions that usually take place in it are
in equilibrium.

However, drying does change the chemical constituents of soils. Ferrous iron
is oxidised to ferric iron, exchangeable potassium content increases or decreases
depending upon the soil and hydrogen ion activity changes to some extent.

Therefore, concentration of ferrous iron (if required) should be determined with
a field-moist soil sample. Also, concentration of exchangeable potassium and
soil pH may also be determined without drying the sample.

Dried soil is easier to grind.

For reasons mentioned earlier, metallic apparatus must not be used for grinding
as particles might break off and contaminate the soil.

Therefore, a wooden mortar and pestle must be used.


Grinding the soil samples

Spread a clean polythene sheet on the
ground and place a wooden mortar on
it. Transfer the dried soil samples to the
mortar.

With a wooden pestle, grind the soil to
break down any aggregates.

After grinding, transfer the soil to the
polythene sheet and spread evenly across
the surface.




Mixing

Why is mixing important? After all, we just ground the samples — that
should have mixed everything quite well.

Mixing is necessary to ensure that the composite soil sample (all the soil samples
taken from different areas in the field) represents the field’s soil composition as
closely as possible even in small quantities.

For example, to determine the amount of phosphorus, as little as 2.5 g of soil
is used in the experiment. You would have collected nearly 3 kg of soil from the
field of which only about 500g of soil is sent to the soil-testing centre.

Therefore, unless properly mixed, it is likely that soil from some parts of the
field might not reach the testing centre.

Yes, during grinding, mixing does take place — but it is random and might not
be enough. Besides, like the collection stage, the mixing stage described here
is to establish a process that minimises chances of statistical error.


Mixing

Lift one end of the sheet and fold it till
the soil collects in the centre. Repeat the
process with the diagonally opposite
corner as shown in the picture... and
then with the other two corners...

This will cause the soil to collect in the
centre of the sheet. This process is called
Mixing and must be repeated 5 times.

Cone the soil and flatten the top.




Partitioning the soil sample

Partitioning?

At this stage, you have about 3kg of a composite soil sample.
This would be a rather unwieldy package to transport to the
testing labs. Besides, the lab will need only about 300g of soil
for all the tests.

Here is where the purpose behind the monotonous mixing process
becomes apparent. Because the sample is mixed, statistically,
the average chemical composition of the field is represented by
surprisingly small amounts of soil — as little as a few grams!

So, you don’t need to send in all the soil you’ve collected painstakingly — you
could send in 500g which is relatively easier to transport and store.

There is a catch though — selecting the final soil sample must also be done at
random . This is fulfilled by the next Stage — Partitioning.

This involves halving the mass of the soil sample in successive stages till it is
about 500g. At each stage, a random portion of soil is selected.

The method described here is one of many that may be used to quarter the
soil sample. There are others, such as : The Riffle Technique and The Paper
Quartering Technique.

However, the method described needs no extra apparatus and is very easy.
Hence its use in this manual.



Divide the soil into 4 equal portions, as
shown.

Discard any two diagonally opposite
portions.

This process is called Partitioning.





Continue partitioning the sample till about
500g of soil remains.

The amount of soil retained depends
upon the number of experiments that
the Soil Chemist intends to conduct. 500
g is enough if all tests are to be carried
out.

Transfer all the soil to a sieve.




Sieving the soil sample

What mesh size is to be used?

A fine sieve (80 mesh) is used for determination of oxidisable
organic carbon and the elements i.e., Nitrogen, Phosphorus and
Potassium.
A coarse sieve (20 mesh) is used for determination of soil pH
and salinity.

The entire volume of the partitioned sample should pass through
the sieve. Soil aggregates that are too large to be sieved should
be ground in a mortar and sieved again.


Sieving the soil sample

Sieve the soil.

Preparation is complete. The soil particles
have been ground, mixed, partitioned
and sieved.

Before the soil is transported to the lab,
it must be stored in polythene bags.




Packing the soil sample


Packing the soil sample

Seal the open end of the bag with thread.

Label the bag. The information required
by the laboratory for testing is shown in
the Information Sheet (see Page 9-28)

The samples are now ready for transport.
The picture shows 3 polythene bags —
these are samples from adjoining fields
all headed to the testing centre.




Soil testing - Determination of soil pH

pH electrodes dipped into a soil-water suspension for pH measurement 1-21

Introduction

A bit about pH...

pH is the quantitative measure of acidity or alkalinity of liquid
solutions. A solution with a pH value less than 7 is considered
acidic and a solution with a pH more than 7 is considered alkaline.
pH 7 is considered neutral.
Soil pH between 6 and 8 is safe for most crops. If the tested
sample has a pH value outside this “safe range”, steps must be
taken to artificially correct the problem.

The acidity of a solution is directly proportional to its hydrogen
ion concentration.

The term pH is derived from p representing the German word potenz, ‘power’,
+ H, the symbol for hydrogen.

pH meters are extremely sensitive instruments. They consist of one (or two)
glass electrodes connected to a digital display. The pH of a solution is displayed
when the electrodes of the meter are dipped in it.

Soil water suspension : A suspension, as opposed to a solution, is a
heterogeneous mixture, i.e., its constituents may be separated by physical
means. The mixture of soil in water is therefore a suspension, not a solution.

Apparatus and reagents required
Buffer tablets of pH 4.0 and 7.0, a top loading balance, a 100mL beaker, a
wash bottle, a glass rod and a pH meter

Determination of Lime Requirement : An acidic soil is treated with Lime to
increase its pH. Recommendation of liming is also done after a pH test. The
difference being the addition of an extra ingredient to the soil.

Take 5g of soil instead of 20g as shown here. Add 5mL of distilled water and
then add 10mL of SMP Extractant Buffer.

Proceed with the pH exactly as shown in the following pages. When you obtain
the pH value, refer to the Lime Recommendation Table on page 9-28.

1-22 Determination of soil pH

Preparing a soil suspension

Weigh out 20g of soil.

Transfer the soil to a 100mL beaker.

Add 50ml of distilled water. This creates
a soil-water suspension.

The soil : water ratio for conducting this
test should be 1 : 1.25

Stir the suspension occasionally for about
half an hour or shake in a shaker for 5
minutes.

Preparing the soil suspension Measuring the pH of the sample

Calibrating the pH meter

Determination of soil pH 1-23

Using the pH meter...

Calibration? Why is it necessary?

Think about this — how does the meter know a solution’s pH? It doesn’t. It’s
just programmed to display different pH values depending upon the voltage
across its electrodes.

The electrodes, though, are sensitive to a whole lot of other things — like
temperature for instance. So, even though a change in room temperature will
not change the pH of a solution, it will cause the electrodes to report the pH
incorrectly.

And this is true of any measuring instrument. We calibrate by measuring a known
amount and then re-programming the meter to display that amount — this is
sometimes as simple as pushing a switch.

In this experiment, we calibrate the meter with 2
buffer solutions — with pH values of 7.0 and 4.0.
See page 1-26 for a description of buffer solutions.

First, we dip the
electrodes in the pH
7.0 buffer. While we
were conducting the
experiment, the
meter read 7.2. This
was because it was set to measure correctly at a
slightly lower room temperature. So, the adjustment
knob was turned till the display read 7.0.

Between readings, wash the electrodes with distilled
water and wipe them dry with a piece of clean
tissue paper.

Repeat the process with a buffer solution of pH 4.0. The volumetric flask in the
third picture contains the pH buffer solution.

The meter might need a few minutes to “warm
up”. The time varies from model to model and you
should check the literature that came with your
meter. Generally, “warming up” takes a few minutes.
Also, it takes a few seconds for the display to
stabilise after you’ve dipped the electrodes in a
solution. So, wait a while before noting down a
pH reading.


Calibration

Calibrate the pH meter with any two
known buffer solutions.

Just prior to taking any readings, stir the
soil-water suspension with a glass rod.

Dip the electrodes of the pH meter into
the suspension and take a reading.




A buffer story

A bit about Buffer Solutions

We know about pH and how it describes the acidity or alkalinity of a solution.
Now, why is a solution acidic? Or alkaline? Modern definitions of acidity refer to
the ability of the compounds in a solution to accept or release electrons — the
Lewis Concept.

But historically, an acid was a compound that, in solution, could release hydrogen
ions into the solution, and a base was a compound which could accept hydrogen
ions.

Since HCl dissociates into H+ and Cl- ions, in solution, it is an acid.

There are situations when we want the pH of a solution to remain constant —
irrespective of change in the concentrations of its acidic or alkaline constituents.
This is done by adding an acid-base pair to the solution that acts as a reservoir,
or buffer. What this reservoir does is suppress or increase the dissociation of
other compounds depending upon the pH of the solution. A commonly used
buffer solution is the NH4Cl - NH4OH pair. These are readily soluble chemicals
and keep each other’s dissociated concentrations in check — in line with the
solubility product principle.

The NH4Cl - NH4OH pair is an alkaline buffer and maintains the pH of the solution
at around 8.5.

You can read more about Buffer Solutions in books on Physical Chemistry.

A thin “chemical film” is deposited on the electrodes each time a pH measurement
is taken. Unless removed, this film causes the electrodes to report inaccurate
values. Therefore, between readings, wash the electrodes of the pH meter with
a stream of distilled water and then wipe them dry with tissue paper.

When not in use keep the electrodes dipped in distilled water.


Meter readings...

The pH of the sample tested is 7.79. The
display on most pH meters takes about
a minute to stabilise.




Soil Testing - Determination of salinity

Conductivity readings are taken from the supernatant liquid.. 1-29

Introduction

A bit about salinity...

The determination of the quantity of water-soluble salts is
of special importance for arid, semi-arid as well as coastal
areas. It helps in taking reclamation measures as well as in
the selection of crops which differ in their tolerance to salts.

While we could measure the concentrations of the salts by
chemical analysis, it would be time-consuming, expensive
— and entirely unnecessary. That’s because, we don’t need
to identify all the salts lurking about — we’re only interested
in the water-soluble ones. The concentration of all of these salts taken together
is what matters to plants.

So, we take a more practical approach to measure soil salinity — we add distilled
water into the soil and stir it till the soluble salts get dissolved. Then, we measure
the electrical conductivity of the water.

And how does that tell us anything about salinity? Indirectly, it does — because
in solution, ions are the carriers of electric charge and therefore, the electrical
conductivity of a solution is directly proportional to its soluble salt concentration.

The conductivity of a soil sample is measured with the help of a conductivity
meter and is expressed in mmhos/cm. or, in SI units, in dS/m.

You don’t even need to calculate the concentrations for the purposes of
recommending fertilizers. The recommendation is based upon the conductivity
measurement itself. ( See the recommendation tables). Most soil testing labs
mention “Electrical Conductivity” or just “E.C.” in their reports.


0.01N KCl solution, a 100mL beaker and a conductivity meter.

Using a conductivity meter...

An electrical conductivity meter is very similar to a pH meter. It also consists
of an electrode connected to a digital display. Measurements are made by dipping
the electrode in the solution being tested.

The precautions to be observed while using this instrument are the same as
those with a pH meter (see page 1-24 ).

1-30 Determination of salinity

Calibrating the conductivity meter

This is a typical Conductivity Meter. Like
the pH meter, its electrode must be kept
immersed in distilled water when not in
use.

Calibrate the meter. This is done with
distilled water and a 0.1N Potassium
Chloride solution.

Distilled water should display 100 in the
digital panel.

Then, dip the electrode in a 0.1N KCl
solution.

Calibration of the Conductivity meter Measuring conductivity

Supernatant Liquid

Determination of salinity 1-31

A bit about supernatant liquid

What is Supernatant liquid?

The liquid that floats above a suspension after it
has been allowed to stand for a while is termed
“Supernatant”. When measuring conductivity, the
conductivity cell should remain in the Supernatant
liquid and not touch the soil below as shown.

So, why are we measuring the conductivity of the
supernatant liquid? After all, during the pH
experiment we had specified that the soil should
be in suspension while the reading was being taken.

The supernatant liquid is a solution. The salts present in the soil dissolve in water
and dissociate into ions, which are charged particles. The concentration of soluble
salts in the soil may, therefore, be calculated from the conductivity of the
supernatant liquid.

Soil - water ratio? Why is that important?

Soil to water ratio should be 1:2. The ratio influences the amount of salts in the
extract. Some laboratories use different ratios while conducting this test. Either
way, the ratio must (and is) always mentioned in a soil analysis report.


Conductivity readings...

Adjust the cell constant knob till the
meter displays 14.1 m.mhos/cm.

The soil water suspension from the pH
experiment is allowed to stand till a clear
supernatant liquid is obtained.
After setting the range switch to
maximum, the electrode is dipped in the
supernatant liquid.

Reduce the range setting on the meter
one at a time till the most appropriate
setting is found.

In this case, the conductance of the soil
sample is 1.20 m.mhos/cm.

Calibration of the Conductivity meter Measuring conductivity

Supernatant Liquid

Determination of salinity 1-33

Soil Testing - Determination of available organic Carbon

Diphenylamine indicator being added drop by drop 1-35

Introduction

A bit about Oxidisable organic carbon...

Decomposed plants and microbial residues are the constituents of
organic matter. The percentage of oxidisable organic matter can
be determined by multiplying its percentage of organic carbon by
1.724.

Oxidisable organic carbon consists of partly decomposed residues
of plants, animals and microorganisms. This constitutes most of
the usable carbon present in the soil.

The other forms of carbon which are present, but not useful as a source of
nutrients, include inorganic carbon (such as carbonates), elemental carbon (such
as coal and graphite) and completely decomposed organic carbon.

For areas known to have very low organic matter content take 2g of soil in the
conical flask, for peat soils, take 0.05g and for areas known to have about
1-2% of organic carbon content, take 0.5g of soil.


1N potassium dichromate solution, 0.5N ferrous ammonium sulphate solution,
diphenylamine indicator, concentrated sulphuric acid and 85% orthophosphoric
acid solution.

500mL conical flask, titration setup (50mL burette, chromyl chloride solution
to clean the burette and titration stand) 10mL bulb-type pipette, chemical
balance, 1000mL volumetric flask and two watch glasses.

Why are two conical flasks used?

This experiment is based upon the Walkley and Black method according
to which soil is digested with chromic acid resulting in the oxidation of
its organic content.

The excess chromic acid is determined by titration with a standard ferrous
ammonium sulphate solution.

After titration, in the case of the soil sample, the amount of titrant consumed
is obtained. The amount of titrant consumed in a blank titration (without soil)
could be calculated stoichiometrically. But this would require accurate weighing
of all the reagents involved in the reaction.

Therefore, it is much simpler to perform a blank titration to obtain the required
figures i.e. the volume of ferrous iron solution consumed.

1-36 Determination of oxidisable organic carbon

Oxidising the carbon in the soil sample

Take two 500mL conical flasks.

Add 1g of soil to one of the flasks.

Add 10mL of K2Cr2O7 to each of the
flasks with a pipette.

Oxidizing the carbon in the soil Titration of the soil sample suspension

Blank Titration Calculations

Determination of oxidisable organic carbon 1-37

Precautions...

Concentrated sulphuric acid is a very corrosive chemical. It fumes in contact
with moisture. Observe the following precautions when using sulphuric acid :

The acid must be poured into the beaker along a glass rod or along its inner
walls.

DO NOT use a pipette to measure out the acid - if any of the acid gets into
your mouth, there might not be enough of it left to talk about the experience!

This step should ideally be carried out in an Exhaust Cabinet because the fumes
are extremely corrosive as well. Don’t try to smell the fumes however tempting
it might seem!

What happens during the half hour?

The oxidisable organic carbon in soil is oxidised by potassium dichromate

3C + 2K2Cr2O7 + 8H2SO4 = 3CO2 + 8H2O + 2K2SO4 + 2Cr2 (SO4)3

Potassium dichromate is converted to potassium sulphate and chromium sulphate.
Cr6+ is reduced to Cr3+. The colour of the oxidised form of chromium(Cr6+) is
yellow (or orange) and that of it’s reduced form (Cr 3+ ) is green.

The volume of K2Cr2O7 solution added to the soil should be large enough so that
only a small fraction of it is reduced - which is indicated by yellow (or orange)
c o l o u r o f t h e r e a c t i o n m e d i u m a f t e r c o m p l e t i o n o f ox i d a t i o n .

This occurs over a period of 30 minutes.

Because sulphuric acid fumes, reagents
might get deposited on the watch glass.
Therefore, when adding water, if you see
flecks of chemicals deposited on the watch
glass, rinse them and allow the water to
drip into the conical flasks.


Oxidising the carbon in the soil sample

With a measuring cylinder, add 20 mL.
of concentrated sulphuric acid to each
of the flasks.

Cover the flasks with watch-glasses and
allow them to stand for about half an
hour.

Then, add about 200 mL of distilled
water to each of the flasks.




Titration

A bit about titration...

Titration is a process by which the amount of an oxidisable or reducible substance
in solution is determined by measuring the volume of a standard reagent required
to react with it.

The burette used must be cleaned with chromyl chloride prior to titration. Dirty
burettes are the most common cause of errors.

Carry out the blank titration first. This will give you a general idea about the
volume of titrant, i.e., Ferrous ammonium sulphate that will be consumed. With
this value in mind, the titration of the soil sample usually takes less time.

Observe the colours that the solution assumes during the process. In the first
phase, the solution is a dark burgundy. After a while, it turns violet. This indicates,
approximately, the mid point of the titration. Local action is also observed at
this point. The remainder of the titration needs to be carried out carefully, i.e.,
by agitating the contents of the flask after every 2 drops.

The end-point is indicated by a sudden change of colour of the solution to viridian,
or dark green.

During the titration of the soil sample, all the indicative colours are more cloudy
than those observed during the blank titration. This is due to suspended soil
particles.

Why is Orthophosphoric acid used?

Orthophosphoric acid, H3PO4, is added so that the colour change at
end point is clearly defined.

Diphenylamine should be added just prior to titration. This is to avoid the
potassium dichromate from oxidising the indicator instead of the organic
content of the soil sample being tested.
During titration, a small amount of diphenylamine is oxidised, however, the
error is negligible.


Blank Titration

Add 10mL of orthophosphoric acid to
each of the flasks.

Just prior to titration, add about 10 drops
of Diphenylamine indicator to the flask.
The solution turns a dark burgundy.

The blank titration is done first.

Titration is done with a 0.5N Ferrous
ammonium sulphate solution.




Local action... And a few calculations

What is local action?

Local action is a phenomenon observed midway during titration. At
this stage, even though the titration is not complete, a faint, localised
“end point” may be observed in the solution where the titrant drops
fall.
To observe local action during this experiment allow a drop of titrant,
i.e., Ferrous ammonium sulphate, to drop on the solution without
agitating the flask as is normally done during titration.The solution
in the immediate vicinity of the drop turns green momentarily.

A few Calculations...

The CD-ROM has a Chemical calculator that does all the work for you but
since we’ve set out to understand the science behind Agricultural Biotechnology,
let’s dive headfirst into yet another bout with theory — and learn a bit about
Stoichiometry.

The Appendix contains an article that explains why you need to add and divide
all these numbers...

The percentage of oxidisable organic carbon (%OC) in the soil sample is given
by

% O.C. = [VK x (1– VS/VB) x SK x 0.3] / W

where

Vk = Volume of Potassium dichromate solution

VS = Volume of Ferrous iron solution consumed in titration with soil

VB = Volume of Ferrous iron solution consumed in blank titration

Sk = Strength of Potassium dichromate solution

W = Weight of soil


Indicative colours during titration...

Midway through the titration, the colour
of the solution turns to clear purple.
At this stage, local action may be
observed.

End point is indicated by a sudden change
of colour to viridian, or dark green.

Titrate the soil sample as well.

Notice that all the indicative colours with
the soil sample are cloudy.

The picture shows the titrated soil sample
suspension at end point.




Soil Testing - Determination of available Nitrogen

Ammonia bubbling up the neck of a Kjeldahl flask 1-45

A bit about the Experiment

A bit about the Experiment

Plants generally take up nitrogen as nitrate under aerobic conditions.
In anaerobic situations, some crops, such as rice, can take up
nitrogen as ammonium ions. Most of the nitrogen present in soil
is present in complex compounds. This is considered as a potential
reserve source and, as such, it may be measured to assess the
nitrogen-supplying capacity of the soil.

Soil testing centres do not usually conduct a separate test for
determining the quantity of available nitrogen in a soil sample brought to them
for testing. Instead, they calculate this quantity directly from the quantity of
oxidisable organic carbon.

And how exactly is that possible? The ratio of the amount of oxidisable organic
carbon is proportional to the amount of nitrogen in a given area. The ratio is
unique to each region. These ratios have been tabulated. In Nimpith, where VIB
is located, the ratio is 1 : 5. With this value, we need only perform the organic
carbon test to determine the quatities of both nitrogen and oxidisable organic
carbon.


Boric acid solution, Mixed indicator, 0.32% potassium permanganate solution,
2.5% sodium hydroxide solution, liquid paraffin.

Kjeldahl flask(s), distillation setup, titration setup, 250mL conical flask and
a few glass beads.

Glass beads and liquid paraffin

These are used to reduce frothing and the formation of bubbles in the solution
when the flask is heated.

The bubbles may carry soil into the delivery tube and deposit them in the conical
flask connected to the other end of the tube. The presence of soil makes it hard
to detect the end point when we titrate the contents of the conical flask. More
on this topic later...

1-46 Determination of Available Nitrogen

Extracting the Nitrogen as Ammonia

Take 20g of soil in a Kjeldahl Flask.

Add 20mL of distilled water.

Coat a few glass beads in liquid paraffin
and put them in the flask.

Extracting the Nitrogen as Ammonia Calculations

Titration of the condensate

Determination of Available Nitrogen 1-47


A bit about Kjeldahl and the flask he invented...

A Danish chemist called J.G.C.T. Kjeldahl came up with the brilliant idea of
estimating nitrogen concentrations in organic substances by distilling it out as
ammonia — which can be easily assayed. For boiling the organic substance, he
made a round-bottomed glass flask with a long neck. A special heat-resistant
glass is used which does not crack when heated to high temperatures and is
expposed to relatively cooler liquids at the same time.

The entire assembly is called a Kjeldahl setup or unit and the flask also bears
its inventor’s name.



Then add 100mL each of 0.32%
potassium permanganate and 2.5%
sodium hydroxide solutions.

Heat the flask to about 80oC on an electric
heater.

Ammonia is evolved. The gas escapes
into the delivery tube attached to the
Kjeldahl flask.





Why do we use mixed indicator?

In this test the pH changes at two distinct points. The first is when
the ammonia is absorbed by the boric acid and the solution changes
from bright pink to green. The second occurs during the titration
of the solution with sulphuric acid. The solution then changes back
to pink.

These changes occur at different pH values and a single indicator
is not sufficient since indicators exhibit a colour shift only in a
small pH range. Thus, we need two indicators which will show us both these
changes. Hence a mixed indicator — which is a mixture of Methyl Red, Bromocresol
green and Ethanol — is used in this experiment.



The evolved gases condense and are
collected in a conical flask containing
Boric Acid solution and Mixed Indicator.

The Ammonia is absorbed by the acid —
indicated by a change in colour of the
solution to green. Continue boiling the
contents of the Kjeldahl flask till about
100mL of distillate is collected.

Titrate the distillate with 0.02N sulphuric
acid.





My end point is brown!

You did not pour in enough paraffin. Or, perhaps, you didn't use
enough glass beads. These ingredients are added to reduce the
surface tension of the solution in the Kjeldahl flasks. This greatly
reduces bubble formation...

The bubbles often carry small amounts of soil and deposit it in
the conical flask. This “muddies” the indicative colours during
titration and hence the end point appears brownish...

A few calculations

Substitute the observed volume, V, of sulphuric acid consumed in the following
equation to calculate the amount of available nitrogen (in kg per hectare) of
the soil sample —

V X 31.36 kg/Ha


Titration of the condensate... and Calculations

Local action is observed distinctly during
titration.

End-point is indicated by a change in
colour from green to a brownish-pink.

Note the value of sulphuric acid
consumed.

Carry out a blank titration — with the
contents of the conical flask
corresponding to the Kjeldahl flask
without soil.




Titration of the condensate... and Calculations


Soil Testing - Determination of available Potassium

Flame view - the orange-red colour indicates the presence of potassium 1-55

A bit about Potassium

A bit about potassium

In soil, potassium may be found in four compound forms - Water
soluble, Exchangeable, Fixed and Lattice-bound. Of these, plants
are interested only in the first two since they cannot assimilate
potassium when it is present in the last two types of compounds.
Potassium is the most abundant meta-cation in plant cells. Oddly
though, soil humus furnishes very little potassium during
decomposition. Also, it occurs in plants only as a mobile, soluble
ion, K+, rather than as an integral part of any specific compound -
but, it is known to affect important aspects of a plants life such as
cell division, formation of carbohydrates, translocation of sugars and resistance
of the plant to certain diseases. Over 60 enzyme actions are known to require
potassium for activation.

Which is why it forms the “K” part of the NPK trio - the three major important
elements that plants require for proper growth. Incidentally, the “K” comes from
“Kalium” which is what potassium used to be called.

And a bit about the experiment

In the next experiment, when testing for phosphorus, we will learn about a
technique called curve fitting. This experiment also uses the same principle but
the curve fitting itself is done electronically by the machine itself.

So what machine are we talking about? It’s called a Flame Photometer. It consists
of two parts — the gas compressor (that’s the first picture on the right) and the
Aspirator/Measurement unit (the second picture). The principle on which this
gizmo operates is that every element, when burnt in a flame, emits energy in
a set series of wavelengths. Simply put, each element burns with a different
colour. Further, the intensity of colour is directly proportional to the concentration
of the element. So, by measuring the intensity of colour of flame aspirated with
the sample, and comparing it with a known set of colour intensities, the photometer
c a n d e t e r m i n e t h e c o n c e n t ra t i o n o f p o t a s s i u m i n t h e s a m p l e .

The gas compressor regulates the flow of LPG to the Photometer. The gas burns
with a nearly colourless (or faint blue) flame. The soil sample extract is then
sucked into the flame in minute quantites. This causes the water to vapourise
instantly and the compounds in it burn in the flame. The colour of the flame
changes depending upon the elements present in the extract. This is detected
by electronic sensors which calculate the intensity of the colour.

Potassium burns with an orange-yellow flame.

Apparatus and reagents list is on page 1-60

1-56 Determination of available Potassium

Calibrating the photometer

Set the Compressor to supply gas at a
pressure of 0.45kg/cm2.

Ignite the flame and then calibrate the
photometer with solutions whose
potassium concentrations are known.

The calibration must be done with 4
standard solutions.

Calibrating the Flame Photometer Measuring the concentration

Extracting the Potassium from the sample Calculations

Determination of available Potassium 1-57

Calibrating a Flame Photometer

Enter “Calibration” mode We’re working with pretty The smart meter now
using the control panel. high concentrations of realises that it needs to
The buttons you have to Potassium here. plot a Standard Curve. So
press will vary for different you need to tell it how
models. Some meters are designed many standard samples
for micro-analysis — like you’re going to use to plot
Usually, you’ll see a the one here. In our case, the curve. We’ll use 4
numbered list of options. we need to tell it to expect samples. Most meters are
In this case, we press “5” Potassium in high good enough to accurately
on the numeric keypad to concentrations. “fit a curve” with this
enter Calibration mode. number of samples.

It now wants to know the Sampling time! Aspirate The flame colour changes
concentrations, in ppm, of each of the stock solutions immediately to a bright
the standard solutions one by one — in the order orange-yellow. The
we’re going to use. Here, in which you keyed them change in intensity of the
we use solutions with 100, into the meter. We typed colour will be barely
75, 50 and 25 ppm in the concentrations in noticable to the naked
concentrations. descending order (100,75, eye. The meter however,
50,25), so, we’ll have to can distinguish each
a s p i ra t e t h e 1 0 0 p p m colour precisely.
solution first.


Calibrating a Flame Photometer

After each sample is The flame becomes After aspirating distilled
aspirated, the meter colourless when distilled water, repeat with the next
demands a “washing” with water is aspirated. It might Standard sample. Note
distilled water — just like also be a faint blue colour. that the display in the
the E.C. meter and the pH The picture here has been picture reads STD4, or
m e t e r. N o t i c e h o w deliberately modified to Standard Sample No. 4.
different measuring exaggerate the colour of
apparatus all have the the flame — so that you This was taken when we’d
same operating principles. can easily compare the already aspirated the first
It’s really quite simple — flame colours with and three samples. Now, we
no magic, just science! without the sample. aspirate the 4th sample.

Between each sample, the Calibration is over. Now, Shubham, it seems, has
meter will ask you to the meter is ready to test no question to ask on this
aspirate distilled water. the soil sample extract - topic and wants to get on
After all the 4 samples are we don’t know the with the experiment!
aspirated, the meter concentration of Potassium
sounds a satisfied beep in this. The meter will
and tells you happily that analyse the colour of the
calibration is over. flame, plot its density on
the standard curve that it’s
drawn for itself and tell us
t h e c o n c e n t ra t i o n o f
Potassium.




Extracting the potassium from the soil sample

Ammonium Acetate? Why is this used?

Like in the Phosphorus experiment, we need to find a way to
extract the Exchangeable Potassium from the soil sample. That’s
what Ammonium acetate is used for.

The ratio of soil : Ammonium acetate should be 1:5. That is, if
you used 5.0g of soil, take 25mL of ammonium acetate.

Ammonium acetate dissociates to yield ammonium ions

CH3COONH4- CH3COO- + NH4+
The NH4+ ions replace the K+ ions, held on exchange sites of soil colloids. As a
result, K+ ions are released into solution. Perfect for our purpose!

The chemical equation above has two arrows pointing in both directions. This
indicates a reversible reaction — i.e. one that occurs simultaneously in both
directions. However, each reversible reaction has an equilibrium point at which
the rates of both the forward and backward reactions remain constant.

Chemistry can be a really exciting subject! You can find out more about reversible
equations in any textbook on Physical Chemistry. See the Appendix for a list.


1N ammonium acetate solution of pH 7.0, 1000ppm potassium solution of
pH 7.0.

10mL pipette, 150mL conical flask with a rubber stopper, 50mL volumetric
flask, 100mL measuring cylinder, a funnel, Whatman no. 42 filter paper and
a flame photometer. This experiment is shown using a direct read-out electronic
flame photmeter.



Take 5g of soil in a 150mL conical flask.

Add 25mL of 1N Ammonium Acetate.
The pH of the solution should be 7.0.

Cork the flask and agitate its contents
for about 30 minutes. This can be done
with a mechanical shaker.





Whatman No. 42... Whatman No. 1... Who is this man called
What?

It’s a brand name. “Whatman”, the company, makes filter paper
— and a lot of other paper products used in Chemical analysis.
The paper is graded according to its relative porosity. Hence,
No. 1, No. 42 etc.

An interesting feature of these papers is that they are “ashless”.
This means that you can burn them and they do not leave behind
a residue. This property is useful in a lot of experiments — such
as gravimetric analysis, in which the filter paper is burned after it is used for
filtration thereby leaving all the precipitate behind for weighing. Neat!

Also, whoever started up the company was probably called “Whatman”... If that
helps at all....



Filter the suspension through Whatman
No. 42 paper.

The filtrate is used for determining
Potassium concentration. Transfer the
filtrate to beakers for use with the Flame
Photometer.

Aspirate the filtrate (the soil sample
extract).




Calculations...

The results and some calculations....

Easy as pie! The smart Flame Photometer tells you the concentration of
Potassium in the extract after politely asking you to wait for a while. In our
case, the concentration of Potassium was 22.3.

The K 2 O content of the soil is calculated using this formula -
K20 (in kg/hectare)= [ 2 X CK ppm X Ve ] / Ws
where

CK ppm = Concentration of the Potassium in ppm obtained from the
Photometer
Ve = Volume of Ammonium Acetate used

Ws = Weight of soil taken, in grams

You can use Chemi-Calc, the calculator on the CD-ROM, to do the calculations.
Or, if you want to show off a bit as well (like the Photometer did), use this trick

Multiply the CK ppm amount by 10!
Remember that the ratio of soil to extractant used should be 1 : 5. Which means
that if you followed the steps, you would have taken 5g of soil and 25mL of
Ammonium acetate. Those values then cancel out to give 5 in the numerator
part of the equation. Multiply that by 2(also in the numerator) and you get
[ CK ppm X 10]

But beware, the trick works ONLY if you measured out the Ammonium acetate
and the soil carefully to at least a couple of decimal places. So 25.01mL and
5.02g of soil is fine. But 25.5 mL and 5.2g of soil means your experiment
will be approximately OK but you cannot show off with the calculation trick!

The formula gives you the amount of K2O present in the soil (in kg per hectare).
To find the amount of Potassium multiply the result from the calculation above
by 0.83.

However, this is not necessary for recommendation since we are interested in
the K2O amount — Why? Because that’s the compound that Fertilizers Companies
refer to! They aren’t very smart, are they?


Measuring the concentration of Potassium in the soil sample

Presence of potassium is indicated if the
flame changes to a yellow-orange colour.

The minute colour differences between
the colours emitted by different elements
are not distinguishable by the naked eye.
The flame view is provided primarily for
adjusting the stability of the flame and
verifying that the nozzles of the aspirator
are not contaminated by residues from
previous experiments.

The concentration of Potassium, in ppm,
is displayed on the screen after a few
seconds.

Most soil testing labs are not equipped with direct-display Photometers like the
one used here. Older equipment requires the user to plot a Standard Curve
manually on Graph Paper. If such equipment is used, the process described in
Section 7 (the procedure used to determine the amount of Phosphorus) is
applicable here as well.




Soil Testing - Determination of available Phosphorus

The blue colour indicates presence of phosphorus 1-67

Preparing a Standard Curve

A bit about available Phosphorus...

Phosphorus occurs in soil in both organic and inorganic forms, most of which is
not easily available to plants. A portion of the total Phosphorus is absorbed by
plants during their growth in the form of H2PO4= . This is what we refer to as
available phosphorus.
What is a Standard Curve and what is it’s use?

Simply put, it is a graphical means of determining unknowns that are
variables of a linear equation. The catch is, that this is an equation of the
form y = n x, where n might vary randomly as x varies. So how is that
linear, you may ask. It is, approximately, because if we define n as n + e, then
we find that e is a very small positive or negative number.

The easier method to solve the problem, is to plot, on graph paper, a few (x,y)
pairs and draw ONE line — the Standard Curve — connecting as many points
as possible. If e was a relatively large number then we would have no option but
to resort to esoteric mathematical tools like regression analysis because then
the equation would cease to be approximately linear. But because e is a small
number, we would find that most points are either on — or very close to — the
straight line drawn and points are scattered almost equally on either side of the
line.

Then, to find the value of y for any given x (or vice versa) all you need to do is
find the corresponding point on the line. Thus, we’ve found the solution to a
linear equation by graphical means. Let’s call it the Graph Technique. Sounds
difficult? It isn’t. As an exercise, try plotting the following value pairs on a sheet
of graph paper

(x,y) = (0 , 0) , (1.1 , 1) , (4.9 , 5) , ( 6.2 , 6) , (7.3 , 7) and (11 , 11).

Draw a line that joins the points - you will find that the points are scattered to
either side of the straight line joining (0 , 0) and (11 , 11).

Using this line, find out the value of y when x is 8. You get y = 8. Now, the
actual value of y might not be 8 exactly, but the graph shows that it would be
pretty close to 8, if not exactly 8. In most cases, as in our present experiment,
the small error is negligible.

Olsen's extractant or Bray and Kurtz No. 1 extractant, Standard 100ppm
phosphate solution, ammonium molybdate reagent (containing antimony
potassium tartrate and ascorbic acid), 2,4-dinitrophenol, P-free charcoal.
Eight 25mL volumetric flasks, 10mL graduated pipette, 10mL measuring
cylinder, a funnel, graph paper, Whatman No. 42 filter paper and a colorimeter.

1-68 Determination of available Phosphorus


Take eight 25 mL volumetric flasks and
add 1mL, 2mL, 3mL, 4mL, 5mL and
10mL of 2 ppm Phosphate solution to
six of them.

Leave two flasks blank.

Add 5mL of a 2ppm Standard Phosphate
solution to one of the empty flasks.

Add 5mL of Olsen's Reagent to the same
flask.

Preparing a Standard Curve Measuring the concentration of Phosphorus

Extracting Phosphorus from the soil sample Calculations

Determination of available Phosphorus 1-69


A step-by-step description of the process...
First we need to plot a Standard Curve. In this test, it is plotted to determine,
approximately, the relationship between the intensity of colour and the needle
deflection of the Colorimeter (the reading on the scale) — the assumption being
that the relationship would be linear. It is linear, by the way... Which makes this
an ideal candidate for using the Graph Technique described on page no. 1-68.

Now, we need a few stock solutions of Phosphorus to plot the Standard Curve.
We take 6 such solutions. Then, we need to find out how much the Colorimeter
would read when the solutions are placed in it.

At this stage two problems crop up —

(1) The Phosphorus in the soil is present primarily as a Phosphate of Calcium,
Iron and Aluminium — which are all colourless. So the Phosphorus needs to be
extracted AND

(2) the new compound, in solution, must have a colour whose intensity varies
linearly with respect to its concentration.

The extraction part is done by adding either Olsen’s extractant which is a 0.5M
NaHCO3 solution or by Bray and Kurtz No.1 Extractant.
Ammonium molybdate reagent or "Reagent B" (which is a cocktail of Ammonium
Molybdate, Tartarate and ascorbic acid) is used to obtain a heteropoly complex
called phosphomolybdic acid, which is reduced, partially, by the ascorbic to give
a blue coloured solution. The amount of the complex produced is directly
proportional to the Phosphorus concentration. Problem solved!

The two flasks to which we did NOT add any stock solution are used to simplify
a technical hitch (see page 1-78) when Olsen's extractant is used.

The Flask with Olsen's reagent described in the facing page will be used to carry
out a mini-titration to determine the amount of acid required to lower the
solution's pH to 3.

The other flask will be used as a "blank". This lets us know if any of the chemicals
we are using contain Phosphorus as an impurity. Shubham will be lurking on
these pages to elaborate as we discuss the steps of the experiment!



Add two drops of 2,4-dinitrophenol
indicator to the flask.

The solution turns yellow.

Then, with a graduated pipette or
dropper, add 2.5M Sulphuric acid to the
flask till the yellow colour is discharged.

This indicates that the pH of the solution
is 3.

Note down the volume of acid consumed.
In our test, 0.4mL of acid was required
to lower the pH when 5mL of Olsen's
reagent was added.

This flask is not needed any more and
may be removed from the work area.

This leaves 7 flasks — 6 with standard
phosphate solutions and one empty flask
for the blank test.





More on the pH value

The value obtained in the previous step — 0.4mL of acid —
pertained to 5mL of Olsen's extractant. So, if we use, say, 50mL
of the reagent, we need to add 4mL of 2.5M sulphuric acid to
adjust the pH of the solution.

Since we've standardised the procedure (always using 5mL of
filtrate and so on) the value obtained earlier — 0.4mL — is used
throughout.



We now have seven flasks left.

To each of the flasks, add 5mL of Olsen's
reagent.

Add exactly 0.4mL of 2.5M Sulphuric
Acid to each of the flasks to adjust the
pH of the solution to 3.

Add approximately 10mL of distilled
water to each of the flasks.

Then, add 4mL of Reagent B — the
Ammonium Molybdate reagent mixture
- to each of the flasks.

The picture shows the reagent being
pipetted into the flask containing 10mL
of the Standard Phosphate solution.




Using a colorimeter

Using the Colorimeter

Colorimeters are usually analogue — measurements are indicated by the deflection
of a needle over a semicircular scale — and they look very ancient in the
laboratory, surrounded by digital equipment. However, they are very precise
instruments as well!

Colorimeters have a light-proof slot where the sample to be tested is inserted.
If the sample is opaque, a full-scale deflection is observed, whereas a transparent
sample does not cause any deflection of the needle.

Calibration

The zero and full scale deflections are set by
“measuring” a sample of distilled water and
a black, opaque cylinder (that comes as a
standard accessory with the meter and is
designed specifically for this purpose.)

Turn the adjustment knob to set the needle
to the zero when calibrating with distilled
water. With the opaque cylinder in the metering
slot, turn the adjustment knob so that the
needle deflects all the way and stops in front
of the “infinity” mark.

More on the Standard Curve...

After you’ve measured the optical densities of all the six solutions,
plot them on a sheet of graph paper.

The concentrations of Phosphorus in each of the flasks, in parts
per million, go on the x-axis, while the corresponding optical
densities go on the y-axis.

Draw a straight line that connects most of the dots. That’s your
Standard Curve! By the way, you might be wondering why it’s called the Standard
Curve when it is a straight line. Well, it would be a curve if the equation were
not linear. For instance, if the relationship between y and x were quadratic
( y = nx2 + c) then the graph would be a curve. Besides, (and this might sound
silly) mathematically, a straight line is also a curve!



Add distilled water to each of the flasks
till the volume of solution in them is
exactly 25mL

The 25mL value is to be maintained for
all solutions. The value determines the
concentration of the phosphate solution.

Remember that ppm is a measure of
concentration.

Allow the flasks to stand for 10 minutes.
The solutions assume a blue tinge.

Measure the optical densities of all the
solutions using a colorimeter.

The blank sample should register a "zero"
deflection while the sample with 10mL
of the standard phosphate solution should
register the highest optical density.




Phosphorus free or not...

On to the extraction
We have the Standard Curve and now we proceed to extracting
the Phosphorus from the soil sample.

This is the part that you will be doing more often since a Standard
Curve only needs to be prepared once a day.

Remember to note down the volume of acid used to adjust the
pH of Olsen's Reagent. In our case, the volume is 0.4mL.

Olsen’s Method

This method is used to extract Phosphorus in soils with pH above 6.

The Bray and Kurtz Method is employed to extract Phosphorus from soils with
pH below 6. The reagent used for extraction, in this case is a solution of 0.03
N Ammonium fluoride in 0.025 N HCl.

Olsen's reagent is added at 1 : 20 ratio to the soil. Thus, when 2.5g of soil are
taken for extraction, 50mL of Olsen's Reagent is to be used.

Bray and Kurtz Reagent is added at a 1 : 10 ratio.

Usually you will find yourself using Olsen’s Method more often. The steps followed
for employing both the methods are identical except for the ratio mentioned
above.

Preparation of the Standard Curve should also be done with the same reagent
used for extraction.


Extracting Phosphorus from the sample

Take 2.5g of soil in a 150mL conical flask.

Add about 0.5g of Phosphorus-free
charcoal to the conical flask.

Prior pH tests on the soil sample indicate
that its pH is 7.2. Hence, Olsen's Reagent
is used as an extractant.

Add 50mL of Olsen's Reagent to the
flask.





2,4-dinitrophenol and ascorbic acid

2,4-dinitrophenol is used as an indicator. The test is to be carried out at a pH
of 3.0. Therefore, we add 2.5M sulphuric acid, drop by drop, till the yellow colour
of the 2,4-dinitrophenol is discharged indicating that the pH of the solution is
exactly 3.0.

This causes a conflict with Reagent B — the ammonium molybdate mixed with
antimony potassium tartarate and ascorbic acid. Ascorbic acid cannot be used
if we use 2,4-dinitrophenol.

Some laboratories use stannous chloride in conjunction with pure ammonium
molybdate. The problem with this method is that the blue complex formed is
very unstable and the colour disappears in a few minutes. In a soil testing centre,
where you could be testing 100 samples everyday, you cannot use Stannous
Chloride.

So we have a bit of problem! The solution is to carry out a kind of “mini-titration”
to determine exactly how much 2.5M sulphuric acid is required to bring the pH
of the solution down to 3.0. We do this by adding 5mL of Olsen’s Extractant to
a 10mL volumetric flask and then adding two drops of 2,4-dinitrophenol. The
solution assumes a yellow colour. Then, we add 2.5M Sulphuric acid to the flask
drop by drop through a graduated pipette, till the yellow colour is discharged.
This gives us the amount of acid required to correct the pH for 5mL of extractant.

Note that one does not need to lower the pH when using Bray and Kurtz Extractant.
After addition of this reagent, we directly add 4mL of Reagent B and then top
up with distilled water to 20mL.



Agitate the contents of the flasks for half
an hour in a mechanical shaker.

No. 42 filter paper.

Then, transfer exactly 5mL of the filtrate
to a 25mL volumetric flask.

Add 0.4 mL of 2.5M Sulphuric acid.




The Standard Curve

0.16

0.14

0.12

0.10
Optical Density

0.08

0.06

0.04

0.02

0
0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8
Concentration of Phosphorus (in ppm)
The Standard Curve...again

The relation between concentration and optical density is linear. And like we
know, it is plotted by measuring the optical densities of solutions with known
strengths — marked red.

After you’ve completed the experiment, you have an optical density reading of
a solution with an unknown concentration of phosphorus - marked blue. The
x-axis value gives you the concentration, in ppm, of phosphorus in the soil
sample.
A Standard Curve is usually prepared daily in the soil testing lab.

Calculations

Substitute the ppm concentration of phosphorus obtained into the following
equation to obtain the availibility per hectare.

P2O5 (kg/hectare) = Vc x Ve x cppm x 2 x 2.2 /Vf x W
where
cppm = ppm concentration obtained from the standard curve
Vc = Volume of the coloured solution in mL
Ve = Volume of extractant taken for extraction of P from soil in mL
Vf = Volume of filtrate taken for colour development in mL
W = Weight of soil taken in grams


Measuring the concentration of Phosphorus

Add 4mL of ammonium molybdate
reagent.

The solution turns blue within a few
minutes.

Add distilled water to the flask till the
volume of solution in it is exactly 25mL.

The solution turns blue.

After 10 minutes, measure the optical
density of the solution in a Colorimeter.

Plot the value on the standard curve you
obtained earlier. This is marked in blue
on the Standard Curve on the facing
page.




Soil Testing - Determination of Gypsum requirement

At end-point, the solution changes from a wine-red to a greenish-blue 1-83

A bit about the experiment

A bit about the experiment
Alkali soils contain large amounts of sodium. This element has huge
ions. Soil aeration and permeability are two conditions that are adversely
affected by the monstrous Sodium atoms hogging space.

To improve the situation, Sodium cations (Cations are atoms that have lost an
electron and, hence, are positively charged) are displaced by Calcium cations.
The Sodium is then leached away by percolating water.

Gypsum — CaSO4.2H20 — is used as a source of Calcium. The following pages
describe the test to determine how much Gypsum needs to be applied to a field.


Two 250mL conical flasks, a 150mL conical flask, a funnel, Whatman No. 42
filter paper, titration setup.

Saturated gypsum solution, ammonium hydroxide-ammonium chloride buffer
solution, EBT indicator, EDTA solution.

1-84 Determination of Gypsum requirement

Adding Gypsum to the soil sample

Ta ke t w o 2 5 0 m L c o n i c a l f l a s k s

To one of the flasks, add 5g of soil. Leave
the other flask empty. It will be used to
carry out a blank test.

To the flask containing soil, add 100mL
of a saturated gypsum solution.

Determination of Gypsum requirement 1-85


Remember this...

Add 5mL of the SAME gypsum solution that you added to the soil sample.
The experiment is to determine how much Gypsum in the 100mL solution is
adsorbed by the soil sample. The amount of gypsum adsorbed directly affects
the strength of the solution.

We compare the strengths of the two solutions by titrating equal volumes —
5mL of filtrate and 5mL of the original solution.

The difference in the volume of EDTA consumed in both the titrations is used to
determine the Gypsum requirement of the field where the sample was taken.



To the other flask, add 5mL of the same
saturated gypsum solution.

Agitate the soil suspension in a
mechanical shaker for 5 minutes.

No. 42 filter paper.


Why add a buffer?

Why do we add the buffer solution?

If iron is present in large amounts in the soil sample, it will interfere
with the titration and skew the results.
The ammonium hydroxide-ammonium chloride buffer stabilises the
pH of the solution at 10. This is highly alkaline and causes any
Iron present in the solution to precipitate as a hydroxide — thereby
ensuring that the titration gives us the required results.

Other elements such as copper and nickel are also present in most
soils but not in amounts enough to cause significant errors in the experiment.


Titration of the filtrate

Transfer 5mL of the filtrate to a 150mL
conical flask.

Then, add 1mL of ammonium chloride-
ammonium hydroxide buffer solution and
25mL of distilled water to both the flasks
- the flask for the blank test and the
flask with 5mL of filtrate.

Add 2-3 drops of EBT indicator. The
solutions assume a wine-red colour.


A few calculations

A sight for sore eyes...

One of the most interesting aspects of Analytical Chemistry are the colours one
sees. Fiery reds, moody blues and shocking pinks... They're all on display.
Titration is a case in point. Even veteran chemists are sometimes distracted by
the beautiful display of a solution magically changing colour.

This particular titration is perhaps the most interesting of the ones described in
this book. Wine Red to blue... The introductory page to this section contains a
full page picture of the solution at end-point.

A few calculations...

The amount of Gypsum that the soil sample requires is calculated by substituting
values into the following formula -

Gypsum requirement (tons/Ha) = SEDTA(Vb-Vs) X 688.68
where

Vs = Volume of the EDTA required for titration of the soil filtrate
Vb = Volume of the EDTA required for blank titration
SEDTA = Strength (in N) of the EDTA solution


Titrating the filtrate and the blank

Titrate with EDTA.

End Point is indicated by a change in
colour to blue-green.

Note down the value of EDTA consumed.

Titrate the blank next.

Note down the volume of EDTA
consumed.


Soil Testing - Reagent preparation

Extraction of phosphorus using Olsen’s Reagent 1-93

Take all the usual precautions while using chemicals —
The bibliograpy at the end of this manual lists quite a few books
on lab-practice. Read one of them. Or, ask a lab technician to show
you the precautions. Observing a technician at work is not
recommended — lab technicians sometimes skip safety measures
while they work. Ask them to explain the procedures to you while
they're not at work.

1-94 Reagent preparation

We’ve followed a graphical method of explaining the procedure for preparing lab
reagents. Most of the reagents are prepared simply by adding a weighed amount
of solid (crystalline) reagent to distilled water and mixing the two. A 1L volumetric
flask is commonly used.

In cases where the reagent may not be soluble in distilled water at room
temperature, a little acid is also added to the mixture.

The preparation procedure is described in a
step-by-step format.

The capacity of the container is mentioned to its
left. Make sure you use a flask/beaker of
the right capacity.

1L The colour that the solution assumes at the END
is shown in a circle below the container capacity.

A white circle indicates a colourless solution.

If the flask needs to be heated, it is indicated
by an orange “flame” at the bottom.

A step that involves corrosive chemicals is
indicated by a red circle.

A step that involves transfer of the solution to
another container midway through the preparation
(for, say, filtration) is indicated by an arrow
p o i n t i n g o u t wa r d f r o m t h e c o n t a i n e r.

Step 1

Step 2
Add 20mL of sulphuric acid

Filter the solution and store in a reagent
bottle

Reagent preparation 1-95

1N Potassium Dichromate

1L

Take 49.04g of potassium dichromate and
put it in the flask

Add 800mL of distilled water

Agitate the contents by shaking the flask
gently till the salt dissolves

After the salt is dissolved, add more distilled
water till the volume of the solution is 1L


0.5N Ferrous Ammonium Sulphate

1L

Take 196.1g of ferrous ammonium sulphate
in the flask


Add about 20mL of concentrated sulphuric
acid. DO NOT use a pipette — use a
measuring cylinder

gently till the salt dissolves.



SMP Buffer solution (pH 7.5)

1L

1.8g of p-nitrophenol

2g of Calcium acetate

2.5mL of triethanolamine

3g of Potassium Chromate

53g Calcium Chloride


gently till the salts dissolve


Adjust the pH of the solution to 7.5 with
dilute Sodium Hydroxide.


Diphenylamine Indicator

250mL

Take 500mg of Diphenylamine indicator
powder in the beaker


Add about 100mL of concentrated
sulphuric acid. DO NOT use a pipette
— use a measuring cylinder

gently till the indicator dissolves

After the salt is dissolved, store it in a
brown reagent bottle


0.32% Potassium Dichromate

1L

Take 3.2g of potassium dichromate in the
flask





2.5% Sodium Hydroxide

1-74

1L

Take 25g of sodium hydroxide in the flask





[Methyl Red + Bromocresol Green + Ethanol] MixedIndicator

250mL

Take 0.07g of Methyl Red indicator in
the beaker

Add 0.1g of Bromocresol Green

Add 100mL of Ethanol

Swirl the contents of the beaker to mix
them well

After the contents are mixed thoroughly,
store the indicator in a brown reagent
bottle


Boric Acid + Mixed Indicator

1L

Take 20g of boric acid in the flask


Warm up to 80oC
Add 200mL of Ethanol

Add 20mL Mixed Indicator

Add 0.05N Sodium Hydroxide till the solution
turns reddish-purple



Olsen’s Extractant

1L

Take 42g of sodium bicarbonate in the flask


gently till the salt dissolves.

Adjust the pH to 8.5 by adding either dilute
HCl or dilute NaOH solution



2,4-dinitrophenol Indicator

250mL

Take 500mg of 2,4-dinitrophenol
indicator powder in the beaker

Add distilled water while stirring the
solution to make a supersaturated
solution

Filter or decant the solution

Store the filtrate in a reagent bottle


Standard 100ppm Phosphate solution

1L

Take 0.4392g of potassium orthophosphate in
the flask


Agitate the contents by shaking the flask gently
till the salt dissolves



2.5M Sulphuric acid

1L

Take 800mL of distilled water

Add 140mL of sulphuric acid

gently

After mixing the acid, add more distilled


[Ammonium Molybdate + Tartarate + Ascorbic acid] ReagentB

500mL
Take 12g ammonium
molybdate (AR grade)
in the beaker

Add 250mL of distilled
water

2L
Ta k e 0 . 2 9 0 8 g o f
antimony potassium
tartarate reagent in
the beaker
Add 100mL of
distilled water.

Take 1L of 2.5M sulphuric acid in the flask. Mix
throughly and add water till the volume of the solution
is 2L.

250mL

-200mL

Take 1.056g ascorbic
acid in the beaker


1N Ammonium Acetate of pH 7.0

1L

Take 77.08g of ammonium acetate in the flask




Adjust the pH of the solution to 7 by adding a
little dilute acid/alkali


Standard Potassium Solution (1000ppm)

1L

Take 1.908g of potassium chloride in the flask





Abiotic — Anaerobic

Abiotic Factor in the Environment : Nonliving forms (physical and chemical) in
the environment that influence the environmental processes

Absorptive : A substance that can absorb and hold water

Aseptic Condition : A situation where there is no microorganism

Acetylene : An organic gas with the chemical formula C2H2

Acetylene Reduction Assay (ARA) : The nitrogen fixing enzyme, Nitrogenase, in
addition to the reduction of nitrogen to form ammonia, can also reduce acetylene
to form ethylene. The rate of this reaction is measured and extrapolated as
nitrogenase activity. This assay is called as ARA.

Acid Soil : A soil with a pH value less than 7. Usually applied to surface layer
or root zone, but may be used to characterize any horizon or sample.

Adaptability : Power of an organism to make itself fit in altered environmental
condition

Adhesion : Physical attachment between two different bodies.

Adventitious Root : The roots of a mature embryophyta developed from parts
other than primary root initial

Aeration Device : A device by which air can pass through a system

Aerobic : An organism, a system or a reaction that requires oxygen for its
operation

Agar-agar : A polysaccharide derived from seaweeds (Red algae) used as
solidifying agent in culture medium

Air-dry : The state of dryness (of soil) at equilibrium with the moisture content
in the surrounding atmosphere.The actual moisture content will depend upon
the relative humidity and the temperature of the surrounding atmosphere.

Alkaline Soil : Any soil that has pH greater than 7. Usually applied to surface
layer or root zone but may be used to characterize any horizon or a sample.

Ambient Temperature : Temperature of the surrounding environment.

Amylase : An enzyme that breaks starch into glucose

Anaerobic : An organism, a system or a reaction that does not require oxygen
for its operation

7-2 Glossary of Terms

Annuli — Biofertilizer

Annuli : Plural of annulus: ring-like segments of the cylindrical body of annelids,
like earthworm

Anterior Region : Proximal part of an animal body

Antibacterial : Any chemical or physical agent or any phenomenon that inhibit
bacterial activity

Antifungal Compound : Compound that inhibits the growth of fungus or destroy
it.

Aquatic Environment : Combination of all the physical, chemical and biological
factors in an water body, which influence living forms

Arid Region : Dry and bare region where the sunlight is of high intensity

Ashby's Medium : A medium for the culture of Azotobacter, the constituents of
which was developed by Ashby

Autoclave : A device for wet sterilization

Auxins : A group of bioactive compounds having promoting effect on plant growth
through cell division, elongation and differentiation

Available Forms of Plant Nutrients : The form of plant nutrients that are soluble
in water and can be absorbed by the plant root

Available Nutrient : That portion of any essential element or compound in the
soil that can be absorbed readily and assimilated by growing plants.
Available Forms Of Plant Nutrients : Simple, inorganic and water-soluble forms
of plant nutrients that can readily be absorbed by the plant root system

Bacteria : Prokaryotic (without true nucleus) unicellular microorganism having
cell wall and spore producing capability; present in all parts of biosphere

Beneficial Microorganism : The microorganisms that improve the soil quality by
their biological action

Biochemical Process : Chemical reactions taking place within the cell or external
to the cell but always with the help of enzymes produced by the cells

Biodegradable : The substances that can be decomposed by the microbial activity

Biodegradation : Decomposition of complex biological macromolecules of organic
bodies into simpler forms by bacterial activity

Biofertilizer : Preparation containing living or latent cells of microorganisms,

Glossary of Terms 7-3

Biological — CFU

which, when applied to soil, increase their number in soil and improves soil
fertility through their biological action

Biologically Degradable Organic Waste : Organic waste that can be decomposed
by microbial activity

Biomass : Total amount of living and nonliving organic content

Biotic Factor in Environment : Living forms that influence environmental processes

Blue Green Algae (BGA) : The BGA are prokaryotic microorganisms living on
water or moist soil. They are photosynthetic and some of them are nitrogen-
fixers.

Broth : Liquid culture containing bacterial cells

Buffer : It is a solution containing an acid and a base, or a salt that tends to
maintain a constant H+ concentration. A buffer tablet with a specific value (as
4.0, 7.0, 9.2) when dissolved in 100 ml distilled water, gives a buffer solution
of that specific pH value.

C-4 plants : Normally, the first stable product in photosynthesis is phosphoglyceric
aldehyde, a three-carbon compound (C-3) plant). In some plants, the first stable
compound is a 4-carbon compound — malic acid. These plants are called C-4
plants. The photosynthetic efficiency of these plants is comparatively high

Calcareous Soil : Soil containing sufficient calcium carbonate (often with magnesium
carbonate) to effervesce visibly when treated with cold 0.1N HCl.

Carbofuran : A pesticide
Carbohydrate : Organic compounds that are composed of carbon, hydrogen and
oxygen, where the ratio of hydrogen and oxygen is 2:1, as in water

Carbohydrate : A class of biological macromolecule constituted of carbon, hydrogen
and oxygen, where the ration of hydrogen to oxygen is 2:1 as in water molecule

Carrier : A finely granular solid phase that can hold and carry microbial cells in
sufficient quantity giving suitable environment to them

Cellulase : The enzyme that breaks cellulose, the major component of plant cell
wall

CFU : Colony forming unit. Spreading of bacterial suspension on plate distribute
the individual cells separately on the surface. On incubation, each cell develops
one colony. But a few cells may remain in clumps. Such clumps, comprising of
two or few cells, will develop individual colonies. Thus enumeration of microbial
culture is made in terms of the number of CFU rather than the number of cells.


Chemical Energy — Culture

Chemical Energy : Energy remaining latent within high-energy chemical compounds
( as in carbohydrate ); to be derived after breakdown by chemical means
(respiration)

Chemical Environment : Chemical factors like concentrations of different chemical
compounds in a system
Chemical nitrogen Fertilizer : Nitrogenous compounds that are applied in field
as nitrogen source of crop plant e.g. Urea, Di-ammonium phosphate

Chitinase : An enzyme that degradess chitin, a component of plant cell wall

Circulatory System : Combination of organs in animal body responsible for the
circulation of metabolites, excretory products etc.

Cocoon : The fertilized eggs of earthworms

Coelom : The body cavity of lower animals

Colonization : Increase in number of individuals in a particular area through
propagation

Colonization of Microorganism : Development of a microbial community in a
particular area through growth (cell division)

Colour : A land may show two different colours, viz., light colour and dark colour.
Soil samples from these two-coloured lands should be collected separately and
analysed as they vary in their properties.
Combined Nitrogen : Compounds containing nitrogen ( ammonia, nitrite, nitrate)

Composite Soil Sample : Soil samples collected from a number of sites of a soil
unit are thoroughly mixed to represent the properties of the soil unit. The mixed
sample is termed composite soil sample.

Compost : Organic residues, or a mixture of organic residues and soil that have
been piled, moistened and allowed to undergo biological decomposition.

Contaminants : Any undesired living form in microbial culture

Contamination : Appearance of any undesired living form in a pure living population
(as in microbial culture)

Cross-fertilisation : Union of male and female sex cells of two different organisms

Culture : Growing living cells or tissue in artificial or semi-natural nutrient medium


Culture Medium — Endogeic Earthworms

Culture Medium : A solution consisting of all the required food substances to
culture living cells or tissue

Cumulative Effect : Combined effects of different factors

Curling : A disease of plant where the leaves or fronds are curled inwardly or
outwardly, caused by bacteria, fungus or viruses.

Cytokinin : A group of bioactive compounds having promoting effect on plant
cell division, specially division of cytoplasm.

Decomposing Microorganisms : The microorganisms that are involved in the
degradation of complex biological macromolecules into the simpler forms

Decomposition : Degradation of complex biological macromolecules present in
organic bodies into smaller and simpler forms by bacterial activity.

Defence Mechanism : A measure to protect an organism from pathogenic agents.

Digestive System : A system in animal body responsible for intake, digestion
and absorption of food

Dilution Plating Method : A method of serial dilution of microbial culture, soil,
food etc., plating the dilutes and consequent incubation to enumerate the number
of viable cells or colony forming units of microorganisms, general or specific.

Diptera : An insect group comprising of insects with single pair of wings

Dominance : Prevalence

Dorsal Side : The backside

Doubling Time : The time required to double its content through multiplication

Dull Colour : Colour not so bright

EC : Electric Conductivity

Effluent : Liquid industrial waste of sewage

Electrical Conductivity of Soil (EC) : The measurement of electrical conductivity
is based on the principle that ions are the carriers of electricity. The electrical
conductivity of a solution increases with increase in soluble salt concentration.
The electrical conductivity of a soil is measured with the help of conductivity
meter and is expressed in mmhos/cm. (now in dS/m). Thus the measurement
of EC helps us to know the concentration of water-soluble salts in the soil.

Endogeic Earthworms : The earthworms that obtain their food from the deep


Endomycorrhiza — Essential Plant Nutrients

layer of substrate / soil

Endomycorrhiza : Mycorrhiza where the fungus remain within the root tissue

Enumeration : Counting of a particular object in relation to space of a particular
event in relation to time

Environment : The combination of all the physical, chemical and biological agents
surrounding an organism that has significant effect on the organism.

Environmental Factors : All the physical, chemical and biological components of
the environment, which influence the ecosystem including the living forms.

Enzyme : A protenacious macromolecule of biological origin and of special
structural and functional organisation that regulate a specific biological reaction

Enzyme, Chitinase : See chitinase

Enzyme, Nitrogenase : See nitrogenase

Enzyme, Amylase : See amylase

Enzyme, Cellulase : See cellulase

Enzyme, Lipase : See lipase

Enzyme, Protease : See protease

Epigeic Earthworms : The earthworms that obtain their food from the surface
of substrate /soil

Oesophagus : The proximal part of the digestive canal beyond the pharynx

Essential Plant Nutrients : Nutrient means something that serves as food or
nourishment. There are 16 chemical elements known to be essential for the
growth of the higher plants. These are C, H, O (absorbed from air and water),
N, P, K, Ca, Mg, S (macronutrients absorbed from the soil), Mn, Zn, Cu, Cl, B,
Mo (micronutrients absorbed from the soil). In the absence of these elements
plants develop deficiency symptoms character tic of the deficient element. Hence
the name essential plant nutrients.


Ethylene — Gibberrellic Acid

Ethylene : An organic gas with the chemical formula C2H4, can be produced by
the reduction of acetylene.

Evaporation : Loss of water in form of vapour

Excretory System : The system in animal body responsible for the release of
harmful products produced as by-products during metabolism

Fermenter : A system within which optimum condition for the growth and
metabolism of microorganism is established artificially

Fertility Status of Soil : It is the nutrient status of the soil. Soil fertility means
the ability or the capacity of the soil to provide the essential plant nutrients in
forms readily available to the plant.

Fertilization : Union of gametes of two opposite sexes

Fertilizer Gap : Difference between the amount of fertilizer applied in field and
the amount that is really absorbed by the crop

Fertilizer Requirement : The quantity of certain plant nutrient elements needed,
in addition to the amount supplied by the soil, to increase plant growth to a
designated optimum.

Fertilizers : Natural or artificial substance containing the chemical elements that
improve the growth and productiveness of plants. Fertilizers enhance the natural
fertility or replace the chemical elements taken from the soil by previous crops.
Modern chemical fertilizers include one or more of the three elements most
important in plant nutrition — N, P, and K.

Flora : The total set of plant community in a given ecosystem

Fossil : Any article that proves the presence of particular living forms in remote
past

Fossil Fuel : The fuel derived from plants and animals of remote past (coal and
petroleum)

Free-living microorganism : The microorganisms that can live freely in environment
without any biological relationship with others

Fungi : Eukaryotic (having true nucleus) microorganism having a plant-like cell
wall but no chlorophyll; capable of spore production, heterotropic

Fungicide : A chemical agent that inhibit or eliminate fungal infection in plants
or seeds.

GA : Gibberellic acid, a plant hormone


Genital pore — Host specificity

Genital Pore : A pore through which sex cells are released in males

Gibberellins : A group of compounds having promoting effect on plant growth,
especially during germination and differentiation of roots.

Gizzard : A strong muscular organ in digestive canal responsible for the maceration
of ingested food

Gregarious : Living in a flock or company

Green Manure : Some highly nitrogen-containing plants that are cultivated and
mixed with the soil by tilling. The plant nutrients are released after decomposition

Grinding : It is the process of breaking or crushing the soil samples by wooden
mortar, roller, motorized grinder into soil aggregates taking care that primary
sand particles are not crushed.

Growth Promoting Substances : Natural or synthetic substances that promote
the growth and development of plants

Gypsum Requirement : The quantity of gypsum or its equivalent required to
reduce the exchangeable sodium percentage of a given increment of soil to an
acceptable level.

Hard And Sticky Soil : The soil where the particles remain tightly adhered to
give the soil hard nature when dry and sticky nature when wet

Hardpan : A hardened soil layer, in the lower A or in the B-horizon, caused by
cementation of soil particles with organic matter or with materials such as silica,
sesquioxides, or calcium carbonate. The hardness does not change appreciably
with changes in moisture content and pieces of the hard layer do not slake in
water. 'A' horizon is the surface horizon of a mineral soil having maximum organic
matter accumulation, maximum biological activity and / or eluviations of materials
such as iron and aluminium oxides and silicate clays. 'B' horizon is the one
beneath the 'A’ that is characterised by one or more of the following (i) concentration
of silicate clays, iron and aluminium oxides and humus alone or in combination
(ii) a blocky structure and (iii) coatings of iron and aluminium oxides that give
darker, stronger or redder colour.

Heavy Soil : Soil with high proportion of clay

Heterotropic : The living forms that cannot synthesise their own food and depend
upon other autotropic organisms for their food requirement.

Host : An organism that harbours other parasitic or symbiotic organism

Host Specificity : Specificity of pathogenic of symbiotic microorganism to recognise
and infect a suitable host


Hyaline— Light soil

Hyaline : Transparent

IBA : Indole Butyric Acid — a plant hormone

Incubation : Keeping any biological system in its optimum condition of operation.

Incubator : A device where the optimum condition is maintained for the operation
of a biological system

Ingestion of Food : Uptake of food in the digestive canal

Inoculation : Addition of starter culture (inoculum) to the medium or soil to
initiate the growth

Inoculum : A little amount of microorganism that is added to the culture media
or soil to initiate the growth

Inorganic Phosphate : Inorganic compounds containing phosphate (viz. ammonium
phosphate, potassium dihydrogen phosphate etc.)

Intestine : The distal part of the digestive canal

Invertebrate : An animal having no backbone

Jensen's Medium : A medium for the culture of Azotobacter, the constituents of
which was developed by Jensen.

Laminar Flow Cabinet (LFC) : A device where microbiological operations are done
in aseptic condition

Leaf Litter : Leaves lying about

Leghaemoglobulin : A proteinacious iron compound present in the root nodules
of leguminous plants

Lepidoptera : An insect group comprising of insect with two pair of wings, wings
fold upwards when at rest

Light Soil : Sandy soil


Lime (Calcium) Requirement — Microbial Inoculants

Lime (Calcium) Requirement : The amount of agricultural limestone ( a sedimentary
rock composed of over 50% calcium carbonate) required per acre to a soil depth
of 6 inches (15cm) or on about 910,000kg of soil to raise the pH of the soil to
a desired value under field conditions.

Limiting Factor : The critical factor that limits the activity of a system in spite
of abundance of other regulating factors

Lipase : An enzyme that breaks fats into fatty acid and glycerol

Loamy Soil : Soil with equal parts of sand and clay

Log-phase : Rapid growth phase of living forms

Maceration : Softening and grinding of the ingested food

Macronutrient : A chemical element necessary in large amounts (usually 50 ppm
in the plant) for the growth of plants. See also Essential Plant Nutrients.

Management Unit : It is the unit of soil representing a distinct management
practice that the previous crop has received, e.g., if a portion of the previously
cropped land was irrigated and the other portion was not irrigated, the land has
two soil units viz., irrigated and non-irrigated land. Separate soil samples are
to be collected and analysed to represent each distinct soil unit.

Medium, Ashby's : See Ashby's Medium

Medium, Culture : See Culture Medium

Medium, Jensen's : See Jensen's Medium

Medium, P.K. : See P.K. Medium

Medium, Slant : See Slant

Medium, YEM : See YEM Medium

Mineralisation : A process of breakdown of complex biological macromolecules
into simplest chemical forms that are available to the plant

Metabolism : All the biochemical reactions occurring within the living cell

Metabolites : All the intermediate compounds and end products of metabolism
Microbial Inoculants : Liquid or carrier based preparation of living or latent cells
of microorganisms


Micro-habitat — Nitrogen Fixing Micro-organisms

Micro-habitat : Isolated habitat (a region where an organism dwells) with very
small area where all the environmental conditions suitable for the growth and
activity of the organism is established

Micronutrient : A chemical element necessary in only extremely small amounts
(less than 50 ppm in the plant ) for the growth of the plants. See also Essential
Plant Nutrients.

Microorganism : Living forms that can be identified under microscope, very
minute in size, visible by naked eye when in groups or colony and cannot be
identified

Microorganisms : The living beings that are too small to be identified by naked
eye; can be observed clearly and identified by microscope

Mineralisation : Conversion of complex organic compounds to simple inorganic
forms

Mixing (soil) : The methodical process of mixing soil for lab analysis. This process
is part of a set of techniques to obtain a composite soil sample that represents
the entire plot of land.

Moist Soil : Soil with water near 100% of its water holding capacity

Monotelic : The organism breeding only once in whole life

Mycorrhiza : A symbiotic association between fungus and root system of higher
plants where both are benefited

NAA : Naphthalene Acetic Acid — a plant hormone

Nematode : Soil born, very minute, cylindrical, invertebrate animal

Niche : Position of an organism in relation to habitat, food habit, role in the
ecosystem, etc.

Nitrogen-Fixation : Conversion of atmospheric nitrogen into ammonia

Nitrogenase : The enzyme that catalyses reduction of nitrogen into ammonia in
nitrogen fixing microorganisms

Nitrogen-fixing Microorganisms : The microorganisms that can convert atmospheric
nitrogen into ammonia


Nocturnal — PK Medium

Nocturnal : An animal that feeds at night

Nodulation Efficiency : Efficiency of rhizobial culture to infect the root of specific
leguminous plant and to develop nodule.

Nodule : Tumorous growth in a part of root of leguminous plant caused by nodule
forming, nitrogen-fixing bacteria — Rhizobium

Non-palatable : Not tasteful and non-desirable

Nutrients : Elements required for the nutrient of an organism

Optimum Condition : The condition most suitable for the operation of a particular
activity

Organic Carbon : The carbon in the form of organic compound

Organic Carbon (%) : The organic carbon that is estimated involves mostly the
partly decomposed organic carbon and rarely the completely decomposed ones.
It excludes 90 to 95% of less active organic matters that are not beneficial for
plant growth (charcoal, graphite).

Organic Humus : Decomposed organic matter in soil

Organic Matter : Materials derived from the dead bodies of living forms

Organic Phosphate : Organic compounds containing phosphate (viz. nucleic acids,
phospholipids etc.)

Organic Waste : Waste material derived from plant or animal source, e.g. straw,
paper, cow dung, fruit and vegetable waste etc.

Ova : Plural of ovum — the female sex cell

Ovary : The female reproductive gland where female sex cells, ova, are produced

Oviduct : A tubular system through which the ovum is transported from ovary
to female pore

Oxygenic Photosynthesis : The photosynthetic process where the hydrogen
required for the reduction of carbon dioxide is derived from water molecule and
oxygen is released as a by-product

P.K. Medium : Pikovskia's medium for the culture of phosphate solubilising
microorganisms


Partitioning — Photosynthesis

Partitioning : The process of dividing the thoroughly mixed soil samples into four
parts (quarters), also called quartering technique. IN this multi-step process,
the soil sample is divided into 4 equal portions and two portions are selected
for analysis. The selected portions are mixed and partitioned repeatedly till about
500g of soil is left.

Pesticide : Chemical agent that inhibit or eliminate pest manifestation in plants
or seeds.

pH : pH is the quantitative measure of acidity or alkalinity of liquid solutions. A
solution with a pH value less than 7 is considered acidic and a solution with a
pH more than 7 is considered alkaline. The solution with a pH 7 is considered
neutral. There are certain important points to be kept in mind while measuring
soil pH, such as: 1. Soil particles should be kept suspended by stirring just before
dipping the electrode ( electrode is either of the two points by which an electric
current enters or leaves a battery or any other electrical device). 2. Drying
changes the soil pH. In the soil report it is thus essential to mention whether
the dried or field moist samples were tested. 3. The ratio of soil: water should
also be mentioned in the report as H ion concentration in soil-water suspension
decreases with increasing dilution and so the pH increases. 4.Washing the
electrode with distilled water from a wash bottle, after every pH measurement
has to be done compulsorily. 5.When not in use, electrodes should be kept
immersed in distilled water.

pH Buffer : A substance that has the property to keep the pH constant in spite
of minute change of the system

Pharynx : The cavity forming the upper part of the digestive canal

Phosphate, Insoluble : The forms of phosphate that are not soluble in water and,
hence, are not available to the plants

Phosphate Solubilisation : Conversion of insoluble forms of phosphate ( as tri-
calcium phosphate) into its soluble form ( as mono-calcium phosphate)

Phosphate Solubilising Microorganisms (psm's) : The microorganisms that convert
the insoluble forms of phosphate (as tri-calcium phosphate) into soluble form

Phosphate, Soluble : The forms of phosphate that are soluble in water and hence
available to the crop plants (e.g. mono-calcium phosphate)

Photosynthesis : A process by which green plants (including algae) can synthesise
carbohydrates by reducing carbon dioxide by water and sunlight with the help
of chlorophyll pigment
Photosynthesis, Oxygenic : see Oxygenic photosynthesis


Photosynthesis, Oxygenic — Recycling Process

Physical Environment : Physical factors like pH, temperature, light etc. in a
system

Physical Factors : Light, temperature, humidity etc.

Pigment : Coloured organic compound that gives colour to the living body, as
a whole or in part (e.g. Chlorophyll in plants, Haemoglobin in blood)

Plant Growth Promoting Substances : Natural or synthetic compounds that
promote plant growth

Plant Growth Regulator : The natural or synthetic compounds that regulate plant
growth and development

Plant Hormones : Natural or synthetic compounds that regulates germination,
growth, development and other biological activities in plants, e.g. auxins

Plant Nutrients : Elements required for the growth and development of plant

Plant Pathogen : Microbial agents which infect plants and cause disease

Plates : Solid medium in petridishes

Pollution : A change in the physical, chemical or biological components of the
environment that affect the living forms

Polytelic : Organism breeding several times in whole life

Posterior Region : Distal part of an animal body

Propagules : Modified plant parts (including seeds) that are used as agents of
propagation

Protease : The enzyme that breaks protein

Protozoa : An animal group comprising of unicellular organisms

Recycling : Alteration of different forms of elements in nature in cyclic manner
in relation to time

Recycling of Elements : Sequential alteration of different forms of elements in
nature in a cyclic manner

Recycling Process : A process by which the elements in the nature are converted
in cyclic manner from one form to other in relation to time and space


Reduction — Slant

Reduction : A chemical reaction where electron or hydrogen atom is added to
a molecule.

Reproductive Potential : Power of an organism to reproduce and hence increase
in number in relation to time

Reproductive System : The system in living organism responsible for its propagation

Residual Effect : The effect of any fertilizer on the next crop

Resistance : Capability to strive against

Respiration : A process of breakdown (oxidation) of food substances to release
energy

Restoration Of Soil Fertility : A phenomenon in which all the events involving
increasing soil fertility occur simultaneously in a balanced manner so the soil
fertility remain constant in relation to time.

Rhizosphere : Volume of the soil surrounding the root system where the influence
of the root is governed

Saline Soil : A non-sodic soil containing sufficient soluble salts to impair its
productivity but not containing excessive exchangeable sodium. The conductivity
of the saturation paste extract is greater than 4dS/m.

Sandy Soil : Soil with high proportion of sand and very little proportion of clay

Seed Pelleting : Covering of individual seed with some protective and inert
substances to form small ball-like structure

Self-Fertilization : Union of male and female sex cells of same organism

Seminal Vesicle : A vesicular structure in the male reproductive system where
semen, the fluid containing spermatozoa, is stored

Semi-natural Condition : A condition where natural condition is established by
assembling the natural components

Sewage : Refuse carried-off by roadside drain in domestic region

Slant : Solid medium in the test tube where the tube is kept in slanting position


Soil Amendment — Solubilisation

Soil Amendment : Any substance such as lime, sulphur, gypsum, and sawdust,
which is used to alter the properties of a soil, generally to make it more productive.
Strictly speaking, fertilizers are soil amendments, but the term is used most
commonly for materials other than fertilizers.

Soil Fertility : The ability of a soil to supply all the essential nutrients in an
optimum amount and balance and in a form readily available to the plants
concerned

Soil Microorganisms : The group of microorganisms (bacteria, actinomycetes,
fungi, algae etc.), which live in soil

Soil Moisture : Water content of soil

Soil Productivity : The capacity of a soil for producing a specified plant or sequence
of plants under a specified system of management i.e. the capacity of the soil
to produce crops per unit area

Soil Reaction : It is the degree of acidity or alkalinity expressed as pH; it has
tremendous indirect effects on plant growth. Abnormally high or alkaline pH
(above 9) or low pH (below 4) is toxic to plants. Between these extremes, the
effect is usually on the nutrient availability. Soil reaction can be modified. Soils
can be made more alkaline (pH increased) by adding calcium, magnesium,
sodium or potassium. Soils can be made more acid by adding substances that
produce strong acids in the soil like fertilizers containing sulphur, ammonium
sulphate or superphosphate.

Soil Salinity : The amount of soluble salts in a soil, expressed in terms of
percentage, parts per million (ppm), or other convenient ratios.

Soil Strata : Plural of stratum; soil layer

Soil Testing : It is the rapid chemical analysis of a soil to estimate its available
nutrient status, reaction and salinity.

Soil Texture : It describes the size of the soil particles. The texture determines
drainage rate and the total amount of stored water in the soil. On the basis of
soil texture, soils can be clay, loam or sandy. The greater the quantity of smaller
particles (clay), the less the drainage rate and the more water held in storage.
On the other hand sandy soil store little water and have a high drainage rate.
Loam (having 20% or less clay, 30-50% silt and 30-50% sand particles) is the
best for cultivation as it is neither too dry nor too wet.

Solar Energy : Energy derived from sunlight
Solitary : Occurring singly

Solubilisation : Conversion of insoluble compounds into their soluble forms


Spermathecial Pore — Tolerant Limit

Spermathecial Pore : A pore in temporarily female earthworm where spermatozoa
are released during sexual copulation

Spermatozoa : The male sex cell

Sprouted Seeds : Seeds where tender seedlings have been developed at the
early stage of germination.

Starter Culture : Little amount of microbial culture that is introduced to the
sterilised medium to initiate the growth

Starter Culture : Little amount of culture added to the production system for the
initiation of growth

Sterilisation : The process of elimination of any living form from an object

Sterilisation, Dry : Making of an object free from any living form by means of
dry heat

Sterilisation, Moist : Making of an object free from any living form by means of
moist heat

Stereoscopic Microscope : A modified version of compound microscope where
living cells are examined

Stunted Growth : Growth that has been checked by a disease

Sustainable Effect : And effect which can be kept up over years

Symbiosis : An association between two organisms where both are benefited

Symbiont : Individual partner in symbiosis

Symbiosis : An association between two organisms where both are benefited

Symbiotic Bacteria : Bacteria that live in symbiotic association with other organism

Symbiotic Microorganism : A microorganism that lives in a mutually beneficial
association with another organism

Synergistic Interaction : Combined effect of two or more components on the
same thing or phenomenon

Testis : The male reproductive gland where spermatozoa, the male sex cells, are
produced
Tolerant Limit : Critical point of stress beyond which the organism cannot tolerate
the stress


Top Dressing — Volatilisation

Top Dressing : An application of fertilizer to a soil after the crop stand has been
established.

Topography : The physical features of an area of land are called the topography,
e.g., an undulating land is divided into three categories, viz., lowland, medium
land, and highland. Separate soil samples should be taken and tested to represent
each category as their properties would vary.

Toxic : Harmful to living form

Transplant Shock : Temporal loss of vitality of seedlings when they are transplanted
from the nursery bed to the field

Unfavourable : Not suitable

Unfavourable Environmental Condition : The physical, chemical and biological
status of the environment that is not suitable for the growth, development or
other biological activity of an organism.

Utilisation Efficiency of Chemical Fertilizers : Capability of soil condition to use
the applied chemical fertilizer for crop growth and productivity

Vas Deferens : The tubular system through which semen are transported from
seminal vesicle to genital pore

Vesicular Arbuscular Mycorrhyzae (VAM) : A symbiotic association between fungi
and root system of higher plants where special structures, vesicles and arbuscles,
are developed by the fungi within the root
Ventral Side : The front side

Versatile : Turning freely from one place to another

Vesicle and Arbuscle : Structures developed by the fungi within the vascular
system of root in Vesicular Arbuscular Mycorrhyza (VAM)

Vestigial Organ : Underdeveloped and non-functional organ, which is well
developed and functional in individuals of other groups

Viable Cells : Microbial cells that retain the property to divide and show all the
microbial activities.

Volatilisation : Very quick evaporation

Water-Holding Capacity : Capacity of a substance to retain water content against
gravitational force


YEM Medium — Notes

YEM Medium : Yeast-Extract Mannitol medium for culture of Rhizobium

NOTES - You can add words to the glossary here. You could also use the Glossary
programme on the CD-ROM to modify this glossary or create your own glossary.


Bibliography

Root nodules on a clover plant 8-1

Bibliography

The following online sources were consulted while writing this manual

Online museums on microbiology -

http://www.bacteriamuseum.org
http://www.microbe.org
http://www.ucmp.berkeley.edu

The Tree of Life Web project

http://www.tolweb.org

University and college (*.edu) sites

http://commtechlab.mse.edu
http://www.cme.msu.edu/sites/dlc-me
http://helios.bto.ed.ac.uk/bto/
http://soils1.cses.vt.edu/
http://hcs.osu.edu/hcs200/Intro.html
http://web.reed.edu/academic/departments/biology/nitrogen/
http://www.lifesci.ucla.edu/mcdbio/html/ri4.htm
http://www.ma.psu.edu/~lkh1/iss/
http://www.wsu.edu:8080/~hurlbert/pages/101hmpg.html
http://www.ulst.ac.uk/faculty/science/bms/
http://webcd.usal.es/web/psm/abstracts/Mariano.htm
http://www.asahi-net.or.jp/~it6i-wtnb/azolla~E.html
http://www.safs.bangor.ac.uk/dj/lectures/nit-fix/lecture2.html

Indiana Bioloab

http://www.disknet.com/indiana-biolab

Misc. Sources, Research organisations etc.

http://www.socgenmicrobiol.org.uk
http://www.asmusa.org
http://www.sanger.ac.uk/Projects/Microbes
http://www.erin.utoronto.ca/~w3msa
http://www.indiaagronet.com
http://fukuokafarmingol.net
http://www.microbiologyonline.org.uk

More information on the Internet may be found by running a search

http://www.google.com
http://www.dmoz.org

8-2

Bibliography

Copyright information for photographs sourced online -

Electron micrograph - Azotobacter (c) 1995, Stu Pankratz

Electron micrograph - Rhizobia on a clover root hair tip - (c) 1995, Frank Dazzo

Books

The article on Rhizobium has been reproduced with almost no changes from
“Rhizobium, Root Nodules and Nitrogen Fixation”- Society for General Microbiology,
January 2002, Edited by Janet Hurst.

Basak,R.K. 2000, Fertilizers. Kalyani Publishers Ludhiana, New Delhi.

Basak, R.K. 2000, Soil Testing and Fertiliser Recommendation, Kalyani Publishers.
New Delhi.

Bear, P.E. 1953, Soil and Fertilizers .4th ed. John Wiley and Son, Inc. NewYork.
Black.C.A. 1965, Method of Soil Analysis, Part-2 Am. Soc. Agron. Inc. Madison,
Wisconsin, USA.

Chopra, S.L. and Kanwar, J.S. 1982, Analytical Agricultaral Chemistry. Kalyani
Publishers, New Delhi.

Guzhov, Yu, 1989, Genetics and Plant breeding for Agriculture, Mir Publishers,
Moscow.

Graham, P.H, Harris S.C., (Eds) Biological nitrogen fixation technology for tropical
agriculture

Hesse, P.R. 1994, A Textbook of Soil Chemical Analysis CBS Publishers and
Distributors, Delhi

Jackson, M.L. 1973, Soil Chemical Analysis, Pentice Hall of India Pvt. Ltd. New
Delhi.

Kale, R. D. 1998, Earthworm — Cinderella of Organic Farming, Prism Books Pvt.
Ltd., Calcutta.

Metson, A.J. 1956, Methods of Chemical Analysis for Soil Samples. New Zealand
Dept. Sci. and Ind, Res. R.E. Owen . Govt. Printer, Wellington, New Zealand.

Motsara, M.R., Bhattyacharya, P. and Srivastava, B. 1995. Biofertilizer-Technology,
Marketing and Usases. Fertilizer Development and Consultation Organisation,
New Delhi.

8-3

Bibliography

Natesh, S., Chopra V.L., Ramchandran S., 1987, Biotechnology in Agriculture,
Oxford Publishers / IBH .

Piper,C.S. 1942, Soil and Plant Analysis: a laboratory manual of methods for the
examination of soil and the determination of the inorganic constituents of plants.
Univ. of Adelaide, Australia.

Peech, M., L.T. Alexander, L.A. Dean and J.F. Reed 1974, Methods of Soil Analysis
for soil fertility investigations U.S.Dept. Agro. Cir. 757. 25P.

Tandon, HLS 1993, Method of Analysis of Soils, Plants, Water and Fertlizers.
Fertilizer Development and Consultation Organization, NewDelhi.110048 ( India)

Yagodin, B.A. (Ed.) 1984, Agricultural Chemistry, Mir Publishers, Moscow.

8-4

Tables

Seedlings being prepared at the VIB nursery 9-1

USING THE TABLES
N P2O5 K2O

1) Rice H 30 20 20 Basal-Full P and K
Prekharif Topdressing of N
Duration M 40 20 20 1/2 after 1st weeding
(100 days)
Direct Seeded (15-20 DAS)
Aus,HYV,Heera L 60 30 30 1/2 at 30-35 DAS
Aditya,
Prasanta,
Kalyani-2,
Khanika

USING THE TABLES - Sample Table 1

This table type shows recommendation of N, P2O5 and K2O on the basis
of soil test results.

H, M and L refer to High, Medium and Low results from the lab.

The blue background line shows Nitrogen recommendations, the orange -
Phosphate and the green backgound line shows Potash recommendation.

Time of application and other remarks are noted is mentioned in the last column.

The crops for which recommendations are available on that page are shown in
green at the top left or right of the page.

The tabulated values are in kg per hectare.

For example, say, a farmer wishes to grow HYV rice on his field (which is 2
hectares in area) in the pre-kharif season. The soil tests indicate that his field
is low in N, Medium in P and Low in K.

The recommendation (highlighted in red in the sample table) would be
(60 x 2 = 120)kg/ha of N, (20 x 2= 40)kg/ha of P2O5 and (30 x 2=60)kg/ha
of K2O.

9-2 RECOMMENDATION TABLES etc.

USING THE TABLES
HIGH MEDIUM LOW
N P2O5 K2O N P2O5 K2O N P2O5 K2O

I. Hill Zone
A.Higher elevation (above 1500 m)

Rainfed

Potato 100 75 75 125 100 100 150 100 100
Cabbage 100 50 50 120 50 50 150 60 60
Fallow
Ginger 120 60 60 120 60 60 120 60 60
Fallow

Irrigated

Potato 100 75 75 125 100 100 150 100 100
Cabbage 100 50 50 120 50 50 150 60 60
Vegetables 80 40 40 100 50 50 120 60 60

USING THE TABLES - Sample Table 2

This table type is more comprehensive than the previous one. It shows
recommendation of N, P2O5 and K2O on the basis of both agroclimatic
factors in West Bengal and soil test results.

The blue background lines shows Nitrogen recommendations, the orange -
Phosphate and the green backgound lines shows Potash recommendation.

The agroclimatic region for which recommendations are available on that page
are shown in green at the top left or right of the page and also in bold large
type at the beginning of each section.

The tabulated values are in kg per hectare.

For example, say, a farmer wishes to grow cabbages on his irrigated field (which
is 1 hectare in area) in the pre-kharif season. His farm is located in a hilly region
of Bengal. The soil tests indicate that his field is low in N, Medium in P and Low
in K.

The recommendation (highlighted in red) would be 150kg of N, 50kg of P2O5
and 60kg of K2O.

Further, the crops are listed in recommended cropping sequences. Each
season is marked by a yellow square The recommended crops for that season
are listed beside and below it. The seasons are in the order Summer/Kharif/Pre-
Kharif and Rabi.

Thus, a valid sequence for the farmer (with a rainfed plot) would be, say, cabbage
in the first season and ginger in the next — but he may not grow potatoes and
cabbages because they are listed as alternatives for the same season.
RECOMMENDATION TABLES etc. 9-3

RICE FERTILIZER RECOMMENDATION USING SOIL TEST RESULTS
N P2O5 K2O

1) Rice H 30 20 20 Basal-Full P and K
Duration M 40 20 20 1/2 after 1st weeding
(100 days)
Direct Seeded (15-20 DAS)
Aus,HYV,Heera L 60 30 30 1/2 at 30-35 DAS
Aditya,
Prasanta,
Kalyani-2,
Khanika

2) Rice H 30 20 20 Basal-1/4 N, full P and K
Duration M 50 25 25 1/2 at 15 DAT
(100-115days)
Transplanted
Aus,HYV,Rasi, L 60 30 30 1/4 at 30-35 DAT
Tulsi
IET-2333,
Annada

3) Rice H 20 20 20 Basal-1/2 full P and K
Kharif, Topdressing of N
Transplanted,
Traditional M 40 20 20 1/2 at P.I. Stage
and improved In case topdressing of N is not
Rupsail, Roghusail, possible due to stagnation of
Bhasamanlk, the entire quantity of N upto
Patnai-23 30 kg/ ha should be applied
SR-26B,Nagra L 50 25 25 as basal.
Tilakkachari,
water depth
upto 50 cm.

4) Rice H 40 20 20 Basal-1/4 N,full P and K
Kharif,short Topdressing of N
duration
(115-125days) M 50 25 25 1/2 at 15 DAT
HYV,IR-64,
IR-36,
Ratna,Khitish
Vikash L 60 30 30 1/4 at 30-35 DAT
IET-4786,Later

Kharif,Medium Topdressing of N
duration
(125-135days) M 60 30 30 1/2 at 15 DAT
HYV,Jaya,Ajaya


RICE
N P2O5 K2O

Kunti,
Shasyasree,
Vikarmacharya,
Prakash,
Pratap L 80 40 40 1/4 at 40-45 DAT
IR-20

6) Rice
Kharif,Medium
duration
(140-150days)
(a)Water depth H 50 25 25 Basal-1/4 N,full P and K
(15-30cm.) Topdressing of N
HYV IR-42, 1/2 at 21 DAT
Shalibahan
Pankaj M 60 30 30 1/4 at 55-60 DAT
Swarnadhan,
Mansarobar,
Swarna,
Bipasha,
Sabitri,
Gayitri L 80 40 40
(b)Water depth H 50 25 25 Basal-1/4 N,full P and K
Suresh,Biraj
Jogen M 60 30 30 1/2 at tillering
Tulashi,
Rajashree L 80 40 40 1/4 at P.I
(c)Water depth H 20 20 20 Basal-1/4 N,full P and K
Sabita,Nalini M 40 20 20 1/2 at tillring, 1/4 at P.I.
Amulya, if split application of N is not
Matangini possible due to stagnation of
Purendu, water, entire fertiliser upto the
Jitenda L 50 25 25 level of 30 kg/ha along with
full P and K need be applied
as basal.
(d)Water depth Fertiliser dose as
(above 100cm) above
Dinesh,
Prunendu,
Jitendra

Boro,HYV,Tulsi Topdressing of N
IR-64,Khitish M 100 50 50 1/2 at tillering
IR-36, 1/4 at P.I
Shasyasree,
IET-4786 L 120 60 60


WHEAT, POTATO,
SUGARCANE and JUTE
N P2O5 K2O

8) Wheat
HYV
Earlysown
irrigated H 80 40 40 Basal-1/4 N,full P and K
K-9107,HP-1731 Topdressing of N
Late sown 1/4 at 21 DAS
HD-2643,
HP-1633 M 100 50 50 1/4 at 40-50 DAS
Sonalika
Normal sown
HUM-468,
UP-262, L 120 60 60
Sonalika

9) Potato H 150 100 100 Basal-3/4 N, full P and K
K.Jyoti, M 200 125 125 Topdressing of N
K.Chandramukhi 1/4 At 1st earthing up
K.Badsha L 250 150 150

10) Sugarcane H 100 50 50 Basal-1/3 N,full P and K
Early upland Topdressing of N
Co J-64
Co 7218 1/3 at 40-45 DAP
Co 87263,
Co S-687 M 150 75 75 1/3 at 80-90 DAP
Medium
duration
Bo-91,
Co 62033
CoS-776 L 200 100 100

11) Jute H 30 20 30 Basal-full P and K
Topdressing of N
a)Olitorius M 40 20 40 1/2 after 1st weeding
JRO-632, (15 DAS)
JRO-524
JRO-7835,
JRO-878 L 50 25 50 1/2 after 35-42 DAS
b)Capsularies H 40 20 20 Basal-full P and K
JRC-7447 M 50 25 25 Topdressing of N
1/2 after 1st weeding
JRC-212 L 60 30 30 (15 DAS)
1/2 at 30-42 DAS


OIL SEEDS
N P2O5 K2O

12) Oilseeds H 60 30 30 Basal-1/4 N,full P and K
Sarsan and Topdressing of N
Toria 1/2 at 30-35 DAS
a)Irrigated M 80 40 40 Basal-full NPK
Benoy,Subinoy L 100 50 50
b)Unirrigated H 30 20 20
Benoy,Subinoy M 40 20 20 In case of rains,20 kg N
L 50 30 30 as topdressing
13) Oil seeds Raj
a)Irrigated H 80 40 40 Basal-1/2 N,full P and K
Sita,Sarama, M 100 50 50 Topdressing of N
Bhagirathi L 120 60 60 1/2 at 40-45 DAS
b)Unirrigated
Sita,Sanjukta 40 20 20 Basal-full NPK
Asech

14) Oil seeds
Til
a)Irrigated

Tilottama 50 25 25 Basal-1/2 N,full P and K
Rama Topdressing of N
80 40 40 1/2 at Flower initiation
b)Unirrigated 25 25 25 Basal-Full NPK for oilseed
Tilottama crops, SSP may be chosen as
Rama phosphate sources to meet
the requirement of S alongwith
P. In case of other sources of
P, calcium sulphate should be
applied.
15) Oil seeds
Linseed
a) Irrigated 40 20 20 Basal-2/3 N,full P and K
Garima,Neela Topdressing of N
Mukta 1/3 at 30 DAS
b)Unirrigated 20 20 20 Basal-full NPK
Neela

16) Oil Seeds
Groundnut
a)Rainfed,
Kharif
AK 12-24,
JL-24 20 30 45 Basal-full NPK
ICGS-44,
Somnath,
ICGS-II,


OIL SEEDS and PULSES
N P2O5 K2O

Girnar-I
b)Irrigated H 20 40 60 Basal-full NPK
Rabi Summer M 20 60 60 SSP is preferred as phosphate
Som nath sources as SSP contains S and
Girnar-1 L 20 60 60 P. For further requirement of
S,application of 200-250 kg
gypsum/ha may be applied
before pegging. Liming should
be done in low pH soils for
correction of acidity and supply
of Ca.

17) Oil Seeds H 30 30 30 Basal-1/2 N,full P and K
Sunflower M 40 40 40 Topdressing of N
Modern L 60 40 40 1/2 at 30 DAS

18) Pulses
Pea,Dhusar,
GF-68 20 40 20 Basal-full NPK
Garden pea
Bonavilla
Arkel

19) Pulses
Arhar 20 50 20 Basal-full NPK
TAT-10
(120-125
days) Liming must be done in acid
Sweta, Chuni soil having pH below 5.5.
(180 days)
Rahi Seed treatment with
(160 days) Rhizobium culture is
recommended.

20) Pulses
Gram 20 50 20 Basal-full NPK
Mahamaya-1 Seed treatment with
Mahamaya-2 Rhizobium culture and
Anuradha liming in acid soil are
necessary.
21) Pulses
Kalai 20 40 20 Basal-full NPK
Kalindi Seed treatment with
Goutam Rhizobium culture and
Sarada liming in acid soil are
necessary.


PULSES and MAIZE
N P2O5 K2O

22) Pulses
Mug 20 40 20 Basal-full NPK
Sonali, Seed treatment with
Panna, Rhizobium culture and
Pusa, liming in acid soil are
Baisakhi necessary.
K-850

23) Pulses
Lentil 20 50 20 Basal-full NPK
Asha,Subrata, Seed treatment with
Ranjan Rhizobium culture and liming
in acid soil are necessary.
24) Pulses
Khesari 1 or 2 DAP Spray (2%)
Nirmal,
BIOL

25) Pulses
Soyabean H 20 30 20 Basal-full NPK
JS-2,Pusa-16 M 30 60 40 SSP is preferred as phosphate
Soyamax L 40 60 40 sources

26) Maize

a) Kharif
Hybrid &
Composite H 40 20 20 Basal-1/2 N,full P and K
Kishan
Composite M 60 30 30 Topdressing of N
Azad Uttam 1/4 at 30 DAS
Composite
Megha L 80 40 40 1/4 at tasselling
b) Rabi
Early-Diara
Arun H 60 30 30 Basal-1/2 N,full P and K
Tarun,Probha M 90 45 45 Topdressing of N Medium-
Agoti-76 1/4 at 30 DAS
Late-Vijay,
Ganga L 120 60 60 1/4 at tasselling
Safed-2,Kishan
Composite
Vikram,
Deccan-101


VEGETABLES
N P2O5 K2O

27) Vegetables H 60 40 40 Basal-1/2 N,full P and K
Topdressing of N
Summer Bhindi M 80 50 50 1/4 at 21 DAS
Parvani
Krani L 100 60 60 1/4 at 35 DAS
Pusa Sawani

28) Vegetable H 80 40 40 Basal-1/2 N,full P and K
Topdressing of N
a) Summer
Brinjal M 100 50 50 1/4 at 21 DAS
Rajpur
Selection L 120 60 60 1/4 at 42 DAS
b) Hybrid
Brinjal 180 100 100
H 100 60 60 Basal-1/2 N,full P and K
Topdressing of N
29) Vegetable M 120 75 75 K Topdressing of N
Arum L 150 80 80 1/4 at 21 DAS
1/4 at 42 DAS

Summer Topdressing of N
Bottle gourd, M 40 20 20 1/4 at 21-28 DAS
Sweet gourd,
Bitter gourd
Pumpkim,
Cucumber L 60 30 30

Summer M 100 60 50 Topdressing of N
Pointed gourd L 120 60 50 1/4 at 21 DAP
1/4 at 42 DAP

Winter Brinjal M 80 40 40 Topdressing of N
L 100 50 50 1/4 at 21 DAT
1/4 at 42 DAT

Winter Cabbage M 150 60 80 Topdressing of N
L 200 60 90 1/4 at 21 DAT
1/4 at 42 DAT


VEGETABLES
N P2O5 K2O

Winter
Cauliflower M 120 60 80 Topdressing of N
L 150 80 80 1/4 at 15 DAT
1/4 at 35 DAT

Winter Onion Topdressing of N
Sukh Sagar M 125 100 100 1/2 at 30 DAT
Pusa Ratna SSP is preferred as source of
Pusa Red L 140 100 100 phosphate as SSP contains S
Red Globe besides P and Onion
Patnai White requires Sulphur.

Winter Beet Topdressing of N
Carrot M 75 50 50 1/4 at 21 DAS
1/4 at 42A DAS
Turnip For Garlic, SSP is preferred
Garlic L 100 60 60 as source of P as Garlic needs
S and SSP contains S besides
P.

Winter Tomato M 100 50 50 Topdressing of N
L 120 60 60 1/4 at 21 DAT
1/4 at 42 DAT
38) Vegetable 180 90 90 Basal-1/2 N,full P and K
Hybrid Tomato Topdressing of N
1/4 at 21 DAT
1/4 at 42 DAT
Radish M 50 60 60 Topdressing of N
L 50 60 60 1/4 at 21 DAT
1/4 at 42 DAT

Knolkhol M 80 80 80 Topdressing of N
L 80 80 80 1/2 at 21 DAT

Chilli M 80 50 60 Topdressing of N
L 100 60 80 1/4 at 30 DAT
1/4 at 60 DAT
42)Vegetable H 150 80 100 Basal-1/2 N,full P and K
Elephant’s
foot M 175 100 120 Topdressing of N
(Kavur) L 200 120 140 1/2 at 90 DAS


VEGETABLES
N P2O5 K2O

Winter Tomato M 100 75 75 Topdressing of N
L 120 75 75 1/4 at 21 DAT
1/4 at 42 DAT

Frenchbean M 50 60 75 Topdressing of N
Clusterbean L 60 80 90 1/4 at flower initiation stage
Pea

Water melon M 80 50 50 Topdressing of N
Sugar Baby, L 100 60 60 1/4 at 21 DAS
Adhary 1/4 at 42 DAS
Ashahi Yamao

Kakrol Topdressing of N
1/2 at 30 DAP

Palak Topdressing of N
1/2 at 21 DAS
1/2 at 42 DAS

Sweet, Topdressing of N
Trumeric 1/2 at 30 DAS

Ginger, Topdressing of N
Turmeric 1/4 at 21 DAS
1/4 at 42 DAS

Katwa Danta Topdressing of N
Puin 1/4 at 21 DAS
1/4 at 42 DAS


WEST BENGAL SPECIFIC RECOMMENDATIONS HILL ZONE
HIGH MEDIUM LOW

I. Hill Zone
A.Higher elevation (above 1500 m)

Rainfed

Potato 100 75 75 125 100 100 150 100 100
Cabbage 100 50 50 120 50 50 150 60 60
Fallow
Ginger 120 60 60 120 60 60 120 60 60
Fallow

Irrigated

Potato 100 75 75 125 100 100 150 100 100
Cabbage 100 50 50 120 50 50 150 60 60
Vegetables 80 40 40 100 50 50 120 60 60

B.Lower elevation (above 1500 m)

Rainfed

Maize 40 20 20 50 25 25 60 30 30
Rice 50 25 25 60 30 30 80 40 40
Mustard 60 30 30 60 30 30 60 30 30
Soyabean 20 30 20 20 60 40 20 60 40
Vegatables 80 40 40 100 50 50 120 60 60
Ragi 40 20 20 50 25 25 50 25 25
Rice 50 25 25 60 30 30 80 40 40
Vegetables 80 40 40 100 50 50 120 60 60
Ginger 120 60 60 120 60 60 120 60 60
Fallow

Irrigated

Maize 30 20 20 50 25 25 60 30 30
Rice 50 25 25 60 30 30 80 40 40
Potato 100 75 75 125 100 100 150 100 100
Maize 40 20 20 60 30 30 80 40 40
Rice 50 25 25 60 30 30 80 40 40
Vegetables 80 40 40 100 50 50 120 60 60
Vegetables 80 40 40 100 50 50 120 60 60
Rice 40 20 20 50 25 25 60 30 30
Potato 100 75 75 125 100 100 150 100 100


TERAI ZONE
HIGH MEDIUM LOW
II.Terai Zone
A.Upland

Rainfed

Jute 30 20 20 40 20 30 50 25 40
Vegetables 80 40 40 100 50 50 120 60 60
Jute 30 20 30 40 20 40 50 25 50
Niger/Toria 30 20 20 40 20 20 50 30 30
Rice 30 20 20 40 20 20 50 25 25
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 60 30 30
Pulses 20 40 20 20 40 20 20 40 20

Irrigated

Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Potato 100 75 75 125 100 100 150 100 100
Rice 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Potato 100 75 75 125 100 100 150 100 100
Rice 30 20 20 30 20 20 30 20 20
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 100 50 50 120 60 60 150 80 80

B.Medium Land

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 60 30 30
Wheat 60 30 30 70 35 35 80 40 40
Jute 30 20 20 40 20 40 50 25 50
Wheat 60 30 30 70 35 35 80 40 40
Rice 30 20 20 40 20 20 50 25 25
Vegetables 80 40 40 100 50 50 120 60 60

Irrigated

Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Potato 100 75 75 125 100 100 150 100 100
Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80

TERAI ZONE and GANGETIC ZONE
HIGH MEDIUM LOW

Vegetables 100 50 50 120 60 60 150 80 80
Rice 30 20 20 40 20 20 50 25 25
Wheat 80 40 40 100 50 50 120 60 60
Jute 30 20 30 40 20 40 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Tobacco 40 20 40 50 25 50 60 30 60

C.Low Land

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Pulse 2% DAP 2% DAP 2% DAP
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 60 30 30

Irrigated

Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Vegetables 80 40 40 80 40 40 100 50 50
Rice 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Vegetables 80 40 40 80 40 40 100 50 50
GM 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Rice 80 40 40 100 50 50 120 60 60

III. Gangetic (New Alluvial Zone)

A.Upland

Rainfed

Jute 30 20 30 40 20 30 50 25 40
Pulse(Kalai) 20 40 20 20 40 20 20 40 20
Jute 30 20 30 40 20 30 50 25 40
Mustard 30 20 20 40 20 30 50 30 30
Rice 30 20 20 40 20 20 50 25 25
Pulse(Kalai) 20 40 20 20 40 20 20 40 20
Rice 30 20 20 40 20 20 50 25 25
Mustard 30 20 20 40 20 30 50 30 30


GANGETIC ZONE
HIGH MEDIUM LOW
Irrigated

Jute 30 0 20 40 0 20 50 20 25
Rice 30 20 20 40 20 20 50 25 25
Potato 150 100 100 200 125 125 250 150 150
Jute 30 20 20 40 20 40 50 25 50
Rice 30 20 20 40 20 20 50 25 25
Wheat 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 25
Vegetables 100 50 50 120 60 60 150 80 80
Sesame
Rice 30 20 20 40 20 20 50 25 25
Potato 150 100 100 200 125 125 250 150 150
Rice 30 20 20 40 20 20 50 25 25
Mustard 80 40 40 100 50 50 110 60 60
Vegetables 100 50 50 120 60 60 150 80 80

B.Medium Land

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 50 25 25
Jute 30 20 20 40 20 20 50 25 50
Rai,Mustard 30 20 20 40 20 20 50 30 30
Rice 30 20 20 40 20 20 50 25 25
Pulses 20 40 20 20 40 20 20 40 20

Irrigated

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150
Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Wheat 80 40 40 100 50 50 120 60 60
Sesame
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Rice 30 20 20 40 20 20 60 30 30
Mustard 60 30 30 80 40 40 100 50 50
Vegetables 100 50 50 120 60 60 150 80 80


GANGETIC and VINDHYA ALLUVIAL ZONE
HIGH MEDIUM LOW

C.Low Land

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
(Poyra)
G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
(Poyra)
Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Oilseed 20 20 20 20 20 20 20 20 20
(Linseed
Poyra)

Irrigated

G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Rice 80 40 40 100 50 50 120 60 60
Sesame 50 25 25 50 25 25 50 25 25
Rice 30 20 20 40 20 20 60 30 30

IV. Old (Vindhya Alluvial Zone)
A. Upland

Rainfed

Rice 30 20 20 40 20 20 50 25 25
Mustard, 30 20 20 40 20 20 50 30 30
Toria
Jute 30 20 30 40 20 30 50 25 40
Mustard 30 20 20 40 20 20 50 30 30
Jute 30 20 30 40 20 30 50 25 40
Pulse 20 40 20 20 40 20 20 40 20
Rice 30 20 20 40 20 20 50 25 25
Pulse 20 40 20 20 40 20 20 40 20
(Kalai)

Irrigated

Jute 30 0 20 40 0 20 50 20 25
Rice 30 20 20 40 20 20 50 25 25
Potato 150 100 100 200 125 125 250 150 150


VINDHYA ALLUVIAL ZONE
HIGH MEDIUM LOW

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 50 25 25
Wheat 80 40 40 100 50 50 120 60 60
Jute 30 20 20 40 20 20 50 25 25
Rice 30 20 20 40 20 20 50 25 25
Vegetables 100 50 50 120 60 60 150 80 80
Jute 30 0 20 40 0 20 50 20 25
Vegetables 80 40 40 100 50 50 120 60 60
Vegetables 100 50 50 120 60 60 150 80 80
Rice 30 20 20 40 20 20 50 25 25
Rice 30 0 20 40 20 20 50 20 20
Potato 150 100 100 200 125 125 250 150 150
Sesame
Rice 30 20 20 40 20 20 50 25 25
Potato 150 100 100 200 125 125 250 150 150

B.Medium Land

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 60 30 30
Pulses 20 40 20 20 40 20 20 40 20

Irrigated

Jute 30 0 20 40 0 20 50 20 25
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150
Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Wheat 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 50 25 25
Rice 30 0 20 40 0 20 60 20 20
Potato 150 100 100 200 125 125 250 150 150
Rice 30 20 20 40 20 20 60 30 30
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Sesame
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 25
Vegetables 100 50 50 120 60 60 150 80 80


VINDHYA ALLUVIAL and RED-LATERITE ZONE
HIGH MEDIUM LOW
C. Lowland

Rainfed

Jute 30 20 30 40 20 40 50 25 50
Rice 30 20 20 40 20 20 60 30 30
Pulse/ 2% DAP 2% DAP 2% DAP
Oilseeds
Poyra
Khesari
Linseed

Mung 20 40 20 20 40 20 20 40 20
Rice 30 20 20 40 20 20 60 30 30

Irrigated

G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Rice 80 40 40 100 50 50 120 60 60
Sesame 50 25 25 50 25 25 50 25 25
Rice 30 20 20 40 20 20 60 30 30

V.Red & Laterite Zone
A.Upland

Rainfed
G.Nut 20 30 45 20 30 45 20 30 45
Kulthi 20 40 0 20 40 0 20 40 0
Soyabean 20 30 20 30 40 30 40 30 30
Niger 30 20 20 40 20 20 50 30 30
Rice 30 20 20 40 20 20 60 30 30
Kulthi 20 40 0 20 40 0 20 40 0
Maize 40 20 20 60 30 30 80 40 40
Oilseed 30 20 20 40 20 20 50 30 30
(Toria/
Niger/
Safflower
Millet 30 20 20 40 20 20 40 20 20
(Jowar/
Ragi)
Kulthi 20 40 0 20 40 0 20 40 0
Arhar 20 50 20 20 50 20 20 50 20
Fallow


RED-LATERITE ZONE
HIGH MEDIUM LOW
N P2O 5 K2O N P2O 5 K2O N P2O 5 K2O

Irrigated

Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 20
Potato 150 100 100 200 125 125 250 150 150
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 50 25 25
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 80 40 40 100 50 50 120 60 60
Vegetables 80 20 40 100 25 50 120 30 60
Vegetables 100 50 50 120 60 60 150 80 80
Maize 40 20 20 60 30 30 80 40 40
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150

B.Medium Land

Rainfed

Rice 30 20 20 40 20 20 60 30 30
Sesame 20 20 20 30 20 20 40 20 20
Rice 30 20 20 40 20 20 60 30 30
Pulse 20 40 20 20 40 20 20 40 20
Rice 30 20 20 40 20 20 60 30 30
Mustard 30 20 20 40 20 20 50 30 30

Irrigated

Rice 30 20 20 40 20 20 50 25 25
Rice 30 0 20 40 0 20 50 20 20
Potato 150 100 100 200 125 125 250 150 150
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 20
Wheat 80 40 40 100 50 50 120 60 60
Sesame
Rice 30 20 20 40 20 20 60 30 30
Potato 150 100 100 200 125 125 250 150 150
Maize 40 20 20 60 30 30 80 40 40
Rice 30 20 20 40 20 20 60 30 30
Mustard 80 40 40 100 50 50 120 60 60
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 20
Potato 150 100 100 200 125 125 250 150 150


RED-LATERITE ZONE and COASTAL ZONE
HIGH MEDIUM LOW

C. Low Land

Rainfed

G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
(Poyra)

Irrigated

G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Rice 80 40 40 100 50 50 120 60 60
Rice 50 25 25 50 25 25 50 25 25
Sesame 30 20 20 40 20 20 60 30 30

VI.Coastal Zone Saline
A. Upland

Rainfed

Fallow
Vegetables 80 40 40 100 50 50 120 60 60
Chilli 40 20 20 50 25 25 60 30 30
Rice 30 20 20 40 20 20 60 30 30
Fallow

Irrigated

Fallow
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 60 30 30
Watermelon 80 40 40 100 50 50 120 60 60
Vegetables 80 40 40 100 50 50 120 60 60
Vegetables 50 25 25 60 30 30 100 50 50
Vegetables 100 50 50 120 60 60 150 80 80


COASTAL ZONE - SALINE
HIGH MEDIUM LOW

B. Medium Land

Rainfed

Fallow
Rice 30 20 20 40 20 20 60 30 30
(Poyra
Khesari)

Irrigated

Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 0 40 0 0 50 20 20
Chilli 80 50 60 80 50 60 100 60 80
Fallow
Rice 30 20 20 40 20 20 60 30 30
Watermelon 80 40 40 100 50 50 120 60 60
Vegetables 80 40 40 100 60 60 150 80 80
Rice 30 0 0 40 0 0 50 20 20
Vegetables 100 50 50 120 60 60 150 80 80
Fallow
Rice 30 20 20 40 20 20 60 30 30
Cotton 40 20 20 40 20 20 40 20 20

C.Low Land

Rainfed

Fallow
Rice 30 20 20 40 20 20 40 20 20
(Khesari
Poyra)

Irrigated

G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Sunflower 30 30 30 40 40 40 60 40 40
G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Chilli 40 20 20 60 30 30 80 40 40
Fallow
Rice 30 20 20 40 20 20 60 30 30
Cotton 40 20 20 40 20 20 40 20 20


COASTAL ZONE - LESS SALINE
HIGH MEDIUM LOW

Less Saline

A. Upland

Rainfed

Fallow
Vegetables 80 40 40 100 50 50 120 60 60
Toria 30 20 20 40 20 20 50 30 30
Fallow
Rice 30 20 20 40 20 20 60 30 30
Kalai 20 40 20 20 40 20 20 40 20
Khesari 2% DAP 2% DAP 2% DAP

Irrigated

Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 20
Wheat 80 40 40 100 50 50 120 60 60
G.Nut 20 30 45 20 30 45 20 30 45
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 80 40 40 100 50 50 120 60 60
Vegetables 50 25 25 60 30 30 100 50 50
Vegetables 100 50 50 120 60 60 150 80 80
Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 50 25 25
Mustard 60 30 30 80 40 40 100 50 50
Vegetables 80 40 40 100 50 50 120 60 60
Rice 0 20 20 40 20 20 60 30 30
Chilli 40 20 20 60 30 30 80 40 40

B.Medium Land

Rainfed

Fallow
Rice 30 20 20 40 20 20 60 30 30
Fallow
Rice 30 20 20 40 20 20 60 30 30
Barley 20 20 20 30 25 25 40 30 30
Fallow
Rice 30 20 20 40 20 20 60 30 30
Sunflower/ 20 20 20 20 20 20 30 30 30
Safflower


COASTAL ZONE - LESS SALINE
HIGH MEDIUM LOW

Irrigated

Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 0 40 0 0 50 20 20
Wheat 80 40 40 100 50 50 120 60 60
Rice 30 20 20 40 20 20 60 30 30
Chilli 40 20 20 60 30 30 80 40 40
G.Nut 20 30 45 20 30 45 20 30 45
Rice 30 20 20 40 20 20 60 30 30
Vegetables 100 50 50 120 60 60 150 80 80

C.Low Land

Rainfed

Fallow
Rice 30 20 20 40 20 20 60 30 30
Cotton 40 20 20 40 20 20 40 20 20
Fallow
Rice 30 20 20 40 20 20 60 30 30
Sunflower 20 20 20 20 20 20 30 30 30
Rice 30 0 20 40 0 20 60 20 30
Pulses 2% DAP
(Poyra)

Irrigated

Vegetables 80 40 40 100 50 50 120 60 60
Rice 30 0 20 40 0 20 50 20 20
Sugarbeet 80 40 40 100 50 50 120 60 60
G.M. 0 25 0 0 25 0 0 25 0
Rice 30 0 20 40 0 20 60 0 30
Rice 80 40 40 100 50 50 120 60 60
Mung 20 40 20 20 40 20 20 40 20
Rice 30 20 20 40 20 20 50 25 25
Rice 80 40 40 100 50 50 120 60 60


Late monsoon Pre-monsoon Post-monsoon

Fruit FYM N P2O5 K2O FYM N P2O5 K2O FYM N P2O5 K2O
kg g g g kg g g g kg g g g
Mango 50 1 kg 50 750 750
Litchi 40 250 375 375 40 250 375 375
Jackfruit 40 250 375 375 40 250 375 375
Sapota 40 250 375 375 40 250 375 375
Cashewnut 15 1 kg 1 kg 330

RECOMMENDATION TABLES etc.
Custard Apple 20 125 150 150 20 125 150 150
Pomegranate 20 125 125 125 20 125 125 125
Coconut 40 250 125 500 40 250 125 500
Guava 25 200 200 150 25 200 150 150
Pineapple 0.5 5 2.5 5 5 2.5 5
Citrus 20 130 130 130 20 130 130 130 20 130 130 130

Papaya 3 months 3 months 3 months
5 50 50 50 5 50 50 50 5 50 50 50

Banana 1st 30 day period 2nd 45 day period 3rd 45 day period
10 25 10 20 - 25 10 20 10 25 10 20
4th 45 day period 5th 45 day period 6th 45 day period
- 25 10 20 - 25 10 20 - 25 10 20
7th 45 day period 8th 45 day period
- 25 10 20 - 25 10 20

All figures are per plant per year

9-25
NPK requirement of FRUIT CROPS

RHIZOBIUM NODULATION - MOST PROBABLE NUMBERS

n=4 n=2
s=10

40 20 >7 x 108
39
38 19 6.9 x 108
37 3.4 x 108
36 18 1.8 x 108
35 1.0 x 108
34 17 5.9 x 107 s=8
33 3.1 x 107
32 16 1.7 x 107 >7 x 106
31 1.0 x 107
30 15 5.8 x 106 6.9
29 3.1 x 106 3.4
28 14 1.7 x 106 1.8
27 1.0 x 106 1.0
26 13 5.8 x 105 5.8 x 105 s=6
25 3.1 x 105 3.1
24 12 1.7 x 105 1.7 >7 x 104
23 1.0 x 105 1.0
22 11 5.8 x 104 5.8 x 104 6.9
21 3.1 x 104 3.1 3.4
20 10 1.7 x 104 1.7 1.8
19 1.0 x 104 1.0 1.0
18 9 5.8 x 103 5.8 x 103 5.8 x 103
17 3.1 x 103 3.1 3.1
16 8 1.7 x 103 1.7 1.7
15 1.0 x 103 1.0 1.0
14 7 5.8 x 102 5.8 x 102 5.8 x 102
13 3.1 x 102 3.1 3.1
12 6 1.7 x 102 1.7 1.7
11 1.0 x 102 1.0 1.0
10 5 5.8 x 101 5.8 x 101 5.8 x 101
9 3.1 x 101 3.1 3.1
8 4 1.7 x 101 1.7 1.7
7 1.0 x 101 1.0 1.0
6 3 5.8 x 1 5.8 x 1 5.8 x 1
5 3.1 x 1 3.1 3.1
4 2 1.7 x 1 1.7 1.7
3 1.0 x 1 1.0 1.0
2 1 0.6 x 1 0.6 0.6
1 <0.6 <0.6 <0.6
0 0

Approx range 109 107 105


RHIZOBIUM NODULATION - MOST PROBABLE NUMBERS

Using the table
This table is used to find the number of 'active' rhizobia in your culture or carrier
bags. 'Active' rhizobia refers to the Rhizobia capable of forming nodules. This
number can be calculated by comparing your test results with results obtained
in tests with large numbers of replicates.

The large number gives statistically accurate results. Comparing the results
allows you to deduce the most probable number of active rhizobia in your carrier.

After performing a nodulation efficiency test with dlutions of your carrier, you
would end up with a table that looks like the one on page 2-82.

On the table on the opposite page, n is the number of replicates - the number
of tubes of each dilution - in the set. s is the number of dilutions.

To find the most probable number of nodulating rhizobia, look for the number
of nodulated units you found under the cooresponding replicates column. The
cooresponding entry in the appropriate dilutions column (labelled s=10 etc.).

For example, in the sample table on page 2-82, a total of 21 nodules were
formed with 4 replicates. Thus, we look for the number ‘21’ under the ‘n=4’
column. Then, we move along the row to the corresponding entry under the
‘s=10’ column since we tested with 10 dilutions.

The most probable number of rhizobium nodulating units is 3.1x104


LIME RECOMMENDATION USING SMP TEST VALUES

Target pH Using the table
Sample pH 6.4 6.8 This table is used to determine the
amount of pure CaCO3 required
to increase the pH of one acre of
4.8 10.6 12.4 land.
4.9 10.1 11.8 Pure CaCO3 required, in tons per acre The ‘Sample pH’ column lists pH
values obtained from a pH test
5.0 9.6 11.2 using the SMP buffer solution.
5.1 9.1 10.6 The ‘Target pH’ columns list the
amount of pure CaCO3 required
5.2 8.6 10.0 to raise the pH of one acre of land
to 6.4 and 6.8.
5.3 8.2 9.4
This table lists values for pure
5.4 7.7 8.9 CaCO3 and not commercial lime.
5.5 7.2 8.3 For conversion, use the following
values —
5.6 6.7 7.1
100kg pure CaCO3
5.7 6.2 7.1
= 56kg pure CaO
5.8 5.7 6.5 = 74kg pure Ca(OH)2
= 84kg pure MgCO3
5.9 5.2 6.0 = 92kg pure CaCO3.MgCO3
= 116kg pure CaSiO3
6.0 4.7 5.4
1 bigha = 0.33 acre = 0.13 hectare
6.1 4.2 4.8

6.2 3.7 4.2

6.3 3.2 3.7

6.4 2.7 3.1

6.5 2.2 2.5

6.6 1.7 1.9

6.7 1.2 1.4


A few tips on Lime recommendation —

1) Fine textured acidic soils with high organic carbon content require more lime
than coarse soils that are low in organic carbon. Liming coarse textured soil is
best done in split doses as overliming is a risk in these soils.

2) Lime is corrosive and is hazardous to seedlings. Liming should be done atleast
a couple of weeks before transplanting the crop. Also, advise the farmer to wear
gloves and a mask while liming his or her field.

3) Root penetration of most crops is 6 inches (15cm). The recommendation
tables assume this to be the case. Some crops have roots that penetrate 9 inches
(23 cm) into the soil. If the farmer intends to grow such a crop, increase the
recommendation by 50%.

4) Fine liming materials such as quicklime (CaO) react with soil much faster than
coarse liming materials. Therefore, fine liming materials are to be applied more
frequently and in less quantities than coarse materials.

5) Liming a neutral or alkaline soil is a waste of time and money.

6) If, for whatever reason, liming materials are unavailable, recommend a
minimum dose of 300-500kg per hectare. This will not significantly neutralise
the soil in the plot but will act as a Ca fertilizer because acidic soils are often
deficient in Calcium.

7) Prolonged use of calcitic limestone causes deficiency of magnesium in the
field. To offset this, recommend application of a magnesium supplying mineral.
A few such minerals are — MgSO4 (Epsom salts) MgO (Magnesia) and Mg(NO3)2.
Dolomitic limestone (CaCO3.MgCO3) may also be recommended.

8) The final acidity of the soil is critical for a good yield. Crops that require a
high dosage of lime amendment — crops that require a neutral or nearly neutral
soil — are: wheat, tobacco and sweet potatoes. Crops that require less lime
are: potatoes, rice, rye and watermelon. Crop such as tea, coffee (arabica) and
pineapples can survive in relatively acidic soils and require very little lime
amendment. These estimates, again, depend on a whole lot of other factors such
as terrain, soil texture and irrigation types.

9) Talk to the farmer! Recommendation of soil amendments (fertilizer included)
is not really an exact science. Socio-economic factors play an important role in
the farmer-field-crop relationship. For instance, a farmer may not have funds
to comply with an optimal recommendation. In such cases, the ‘optimal’
recommendation must take into account his available funds...

9-29

LIST OF ABBREVIATIONS

Most abbreviations are expanded in the main body of the manual or in the
Glossary.

ARA - Acetylene Reduction Assay
CFU - Colony forming unit
EBT (indicator) - Erichrome Black T
EDTA - Ethylene Diamine Tetraacetic Acid
FYM - Farm yard manure
SMP (extractant buffer) - Shoemaker, MacLean and Pratt (the scientists who
devised the pH determination test using this chemical.

9-30

INFORMATION SHEET
Sample No. X-base ID

Date of Collection

Farmer’s name

Address

G.P

Block

Mouza No.

Plot No.

Land area.

Land type : Upland - medium land - low land

Soil texture : Sandy - Loam - Clay

Irrigation facility : Irrigated area - Non-irrigated area.

Source of water : River - Pond - Canal - Tube well - Well

Drainage condition : Good - Medium - Bad.

Slope of the land : No slope - slight slope - middle slope - steep slope.

Height of water level in chary rice :

Information on previous crop :

Name and variety of the crop :

Dose of organic manure, if applied :

Dose of fertilizer, if applied :

Yield :

Information of the crop that will be grown:

Name and variety of the crop :

Season (Pre kharif / kharif / rabi ) :

SIGNATURE
9-31

Soil Testing

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