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Plant Anal. Ref. Proc. for S. US (SCSB # 368)
PLANT ANALYSIS REFERENCE PROCEDURES
FOR THE SOUTHERN REGION
OF THE UNITED STATES
SOUTHERN COOPERATIVE SERIES
BULLETIN #368
MAY ‘92
URL: http://www.cropsoil.uga.edu/~oplank/sera368.pdf
Dr. C. Owen Plank
The University of Georgia
Crop & Soil Science Dept.
Athens, GA 30602-7272

http://www.cropsoil.uga.edu/~oplank

E-mail: oplank@arches.uga.edu
ISBN: 1-58161-368-7

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Plant Anal. Ref. Proc. for S. US (SCSB # 368)
May 1992 SCSB #368
PLANT ANALYSIS REFERENCE PROCEDURES FOR
THE SOUTHERN REGION OF THE UNITED STATES
Editor
C.O. Plank
ABSTRACT
This bulletin is the instrument used by the Southern Extension Research Activity-Information Exchange Group-6 (SERAIEG-
6) to document in summary form procedures used by state university plant analysis programs. This document,
records detailed analytical methodologies that are used by the various laboratories throughout the Southern Region. Other
procedures are available for many of the analyses presented in this publication. For information related to similar
procedures, each state university maintains laboratory manuals that may be of further assistance to the reader. The intent
of this document is to provide a reference for current and most widely used plant analysis methods.

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NOTE: Commercial companies are mentioned in this publication solely for the purpose of providing
specific information. Mention of a company does not constitute a guarantee or warranty of its products by
the Agricultural Experiment Stations or an endorsement over products of other companies not mentioned.

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Plant Analysis Reference Procedures for the
Southern Region of the United States
Contents
Sample Preparation ………………………………………………………………………………………………………………1
C. R. Campbell and C. O. Plank
A. Decontamination…………………………………………………………………………………………………..1
B. Drying……………………………………………………………………………………………………………….2
C. Particle-Size Reduction …………………………………………………………………………………………..3
D. Storage………………………………………………………………………………………………………………4
E. Organic Matter Destruction-Dry Ashing ………………………………………………………………………5
F. Organic Matter Destruction-Wet Ashing ………………………………………………………………………7
G. Organic Matter Destruction-Accelerated Wet Digestion ……………………………………………………9
Determination of Total Nitrogen in Plant Samples by Kjeldahl ………………………………………………………. 13
W.H. Baker and T. L. Thompson
Determination of Nitrogen in Plant Tissue Using Continuous Flow, Segmented
Stream AutoAn alyzer …………………………………………………………………………………………………………. 17
R. A. Isaac and W. C. Johnson, Jr.
Determination of Total Nitrogen in Plant Tissue by Combustion…………………………………………………….. 20
C. R. Campbell
Determination of Nitrate Nitrogen in Plant Samples by Selective Ion Electrode ………………………………….. 23
W. H. Baker and T. L. Thompson
Determination of Phosphorus in Plant Tissue by Colorimetry ………………………………………………………… 27
K. P. Moore
Determination of Potassium, Calcium, and Magnesium in Plants by Atomic Absorption
Techniques …………………………………………………………………………………………………………………….. 30
E. A. Hanlon
Determination of P, K, Ca, Mg, Mn, Fe, Al, B, Cu, and Zn in Plant Tissue by Inductively
Coupled Plasma (ICP) Emission Spectroscopy ………………………………………………………………………….. 34
S. J. Donohue and D. W. Aho
Determination of P, K, Ca, Mg, Mn, Fe, Al, B, Cu, and Zn in Plant Tissue by Emission
Spectroscopy…………………………………………………………………………………………………………………… 38
R. A. Isaac and W. C. Johnson, Jr.
Determination of Sulfur in Plant Tissue by Combustion……………………………………………………………….. 42
R. A. Isaac and W. C. Johnson, Jr.
Determination of Sulfur in Plant Tissue by Turbidimetry ……………………………………………………………… 45
C. C. Mitchell

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Determination of Total Manganese, Iron, Copper and Zinc in Plants by Atomic Absorption
Techniques …………………………………………………………………………………………………………………….. 48
E. A. Hanlon
Determination of Boron in Plants by Azomethine-H Method …………………………………………………………. 51
W. E. Sabbe
Determination of Total Boron in Plants by Curcumin Method………………………………………………………… 54
J. W. Odom
Determination of Molybdenum in Plants………………………………………………………………………………….. 57
J. L. Sims and J. D. Crutchfield
Determination of Molybdenum in Plant Tissue Using the Graphite Furnace ………………………………………. 64
D. O. Wilson
Conversion Table ………………………………………………………………………………………………………………. 68

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This bulletin is one in a series of the Southern Cooperative Series and, as such, is in effect a separate publication by each
of the cooperating agencies listed below. Thus, it may be mailed under the frank and indicia of each. Requests for copies
from outside the cooperating states may be addressed to the Georgia Agricultural Experiment Station, Room 2375,
Coliseum, University of Georgia, Athens, Georgia 30602.
Stations and agencies directly participating are as follows:
AGRICULTURAL EXPERIMENT STATIONS
Alabama Agricultural Experiment Station
Auburn University
Auburn, AL 36849-5403
L. T. Frobish, Director
Arkansas Agricultural Experiment Station
University of Arkansas
Fayetteville, AR 72701
G. J. Musick, Director
Florida Institute of Food and Agricultural Sciences
University of Florida
Gainesville, FL 32611
J. M. Davidson, Director
Georgia Agricultural Experiment Station
University of Georgia
Athens, GA 30602
C. W. Donoho, Jr.
Kentucky Agricultural Experiment Station
University of Kentucky
Lexington, KY 40546-0091
C. O. Little, Director
Louisiana Agricultural Experiment Station
Louisiana State University and A&M College
Baton Rouge, LA 70894
K. W. Tipton, Director
Mississippi Agricultural and Forestry
Experiment Station
Mississippi State University
Mississippi State, MS 39762
V. G. Hurt, Director
North Carolina Agricultural Research Service
North Carolina State University
Raleigh, NC 27695-7643
R. J. Kuhr, Director
Oklahoma Agricultural Experiment Station
Oklahoma State University
Stillwater, OK 74078-0500
C. B. Browning, Director
Puerto Rico Agricultural Experiment Station
University of Puerto Rico
Mayaguez, PR 00708
J. A. Quinones, Acting Director
South Carolina Agricultural Experiment Station
Clemson University
Clemson, SC 29634-0351
J. R. Fischer, Director
Tennessee Agricultural Experiment Station
University of Tennessee
Knoxville, TN 37901
D. O. Richardson, Director
Texas Agricultural Experiment Station
Texas A&M University System
University Station
College, TX 77843-2147
R. G. Merrifield, Director
Virginia Agricultural Experiment Station
Virginia Polytechnic Institute and State University
Blacksburg, VA 24061-0402
J. R. Nichols, Director
Virgin Islands Agricultural Experiment Station
College of the Virgin Islands
St. Croix, USVI 00850
D. S. Padda, Director

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STATE EXTENSION SERVICES
Alabama Cooperative Extension Service
Auburn University
Auburn, AL 36849-5403
A. E. Thompson, Director
Arkansas Cooperative Extension Service
University of Arkansas
Little Rock, AR 72203
D. F. Foster, Director
Florida Cooperative Extension Service
University of Florida
Gainesville, FL 32611
J. T. Woeste, Director
Georgia Cooperative Extension Service
The University of Georgia
Athens, GA 30602
C. W. Jordan, Director
Kentucky Cooperative Extension Service
University of Kentucky
Lexington, KY 40546
C. O. Little, Director
Louisiana Cooperative Extension Service
Louisiana State University
Baton Rouge, LA 70803-1900
D. T. Loupe, Director
Mississippi Cooperative Extension Service
Mississippi State University
Mississippi State, MS 39762
H. D. Palmertree, Director
North Carolina Agricultural Extension Service
North Carolina State University
Raleigh, NC 27695-7602
B. K. Webb, Director
Oklahoma Cooperative Extension Service
Oklahoma State University
Stillwater, OK 74078
C. B. Browning, Director
Puerto Rico Cooperative Extension Service
University of Puerto Rico
Mayaguez, PR 00708
J. A. Quinones, Acting Director
South Carolina Cooperative Extension Service
Clemson University
Clemson, SC 29634
B. K. Webb, Director
Tennessee Agricultural Extension Service
University of Tennessee
Knoxville, TN 37901
B. G. Hicks, Director
Texas Agricultural Extension Service
Texas A&M University
College Station, TX 77843
Z. L. Carpenter, Director
Virginia Cooperative Extension Service
Virginia Polytechnic Institute and State University
Blacksburg, VA 24061
J. F. Johnson, Director
Virgin Islands Cooperative Extension Service
University of the Virgin Islands
St. Croix, USVI 00850
D. S. Padda, Director

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STATE DEPARTMENTS OF AGRICULTURE
North Carolina Department of Agriculture
Agronomic Division
Raleigh, NC 27611
D. W. Eaddy, Director

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Preface
Plant analysis has evolved into one of the important tools in crop production. It is a process in which
plant samples are collected from a plant at a specific time during the growing season and analyzed in a
laboratory for various essential nutrients. Nutrients of primary interest are nitrogen (N), phosphorus (P),
potassium (K), calcium (Ca), magnesium (Mg), sulfur (S), manganese (Mn), molybdenum (Mo), iron (Fe),
boron (B), copper (Cu) and zinc (Zn). The plant analysis process also includes an evaluation of the analytical
data to determine whether an element is low, sufficient, or high and finally the formulation of a
recommendation.
Each step in the plant analysis process is equally important. In the laboratory phase many different
procedures are involved which include decontamination, drying, grinding, weighing, ashing, and analysis for
11 to 12 essential nutrients. Consequently, performing a plant analysis involves the use of a variety of
laboratory instruments.
Several laboratories in the Southern Region of the United States offer plant analysis services to
researchers and growers. This bulletin contains reference procedures commonly used by laboratories in the
region. Procedures were selected on the basis of their accuracy and precision as well as their popularity and
acceptance by workers in the area of plant analysis. These procedures also provide a reference for laboratories
to exchange samples to evaluate current plant analysis procedures or to implement new ones.
C. Owen Plank
Editor
Acknowledgements
Appreciation is expressed to all the Southern Extension and Research Activities members who
contributed to this bulletin and to each experiment station representative who submitted procedures.
Appreciation is also expressed to Dr. George Kriz, Administrative Advisor, for his leadership and sincere
support of this work group.
Editorial Committee
C. O. Plank, Editor
C. R. Campbell
F. R. Cox
V. W. Case
E. A. Hanlon
C. C. Mitchell
H. J. Savoy, Jr.
State agricultural experiment stations and extension services are equal opportunity/affirmative action agencies.

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Members of the Southern Extension and
Research Activities Information Exchange
Group-6-Soil Test and Plant Analysis
1991
Administrative Advisor – G. J. Kriz, Associate Director, Agricultural Experiment Station, North
Carolina State University
Alabama C. E. Evans (Rep), C. C. Mitchell
Arkansas W. E. Sabbe (Rep), W. H. Baker, N. Miller, C. Snyder
Florida E. A. Hanlon (Rep), G. Kidder
Georgia C. O. Plank (Rep), R. A. Isaac, M. E. Sumner
Kentucky W. O. Thom (Rep), V. Case, D. Kirkland
Louisiana J. Kovar (Rep), J. Holder
Mississippi W. Houston (Rep), K. Crouse
North Carolina F. R. Cox (Rep), M. R. Tucker, C. R. Campbell
Oklahoma E. Allen (Rep), G. V. Johnson
Puerto Rico O. Muniz-Torres (Rep)
South Carolina R. M. Lippert (Rep), C. L. Parks
Tennessee G. Lessman (Rep), J. J. Jared, H. J. Savoy
Texas L. Unruh (Rep), H. D. Pennington
Virginia S. J. Donohue (Rep)
Virgin Islands E. Craft (Rep)

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Sample Preparation
C. R. Campbell and C. O. Plank*
Sample preparation is critical in obtaining accurate analytical data and reliable interpretation of plant
analysis results. Proven procedures must be followed during decontamination, drying, particle-size reduction,
storage, and organic matter destruction. Each of these preparatory procedures provide opportunities to
enhance the accuracy and reliability of the analytical results.
A. Decontamination
1. Principle
1.1 Plant materials must be clean and free of extraneous substances including soil and dust
particles, and foliar spray residues that may influence analytical results. Generally, the
elements most affected by soil and dust particles are Fe, Al, Si, and Mn. Foliar nutrient spray
and fungicide residues can affect several elements and should be taken into account in the
decontamination process and when evaluating the analytical results. The decontamination
process must be thorough while still preserving sample integrity. Therefore,
decontamination procedures involving washing and rinsing should only be used for fresh,
fully-turgid plant samples.
2. Reagents and Apparatus
2.1 Deionized water.
2.2 0.1 to 0.3% detergent solution (non-phosphate).
2.3 Medium-stiff nylon bristle brush.
2.4 Plastic containers suitable for washing and rinsing tissue samples.
3. Procedure
3.1 Examine fresh plant tissue samples to determine physical condition and extent of
contamination. Unless leaf tissue is visibly coated with foreign substances, decontamination
is usually not required except when Fe (Wallace et al., 1982) Al, Si, or Mn are to be
determined (Jones and Case, 1990).
1Chief-Plant/Waste/Solution Advisory Section, Agronomic Division, North Carolina Department of Agriculture,
Raleigh, NC 27611 and Extension Agronomist, Soil Testing and Plant Analysis, Cooperative Extension Service, The
University of Georgia, Athens, GA 30602.

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3.2 When Al, Si, Mn, and Fe are not of primary interest, plant leaves should be brushed briskly to
remove visible soil and dust particles.
3.3 When plant samples show visible residues from spray applications and when Al, Si, Fe
(Wallace et al., 1982), and Mn are elements of interest, leaves should be washed in a 0.1 to
0.3% detergent solution (Ashby, 1969 and Wallace et al., 1980) followed by rinsing in
deionized water. The wash and rinse periods should be as short as possible (Sonneveld and
Van Dijk, 1982) to avoid danger of N, B, K, and Cl leaching from the tissue (Bhan et al.,
1959).
3.4 After decontamination, samples should be dried immediately to stabilize the tissue and stop
enzymatic reactions.
4. Remarks
4.1 When proper sampling techniques have been utilized, decontamination should be minimized.
4.2 Decontamination is generally not necessary where tissue has been exposed to frequent rainfall
and/or not exposed to nutrient or fungicidal sprays (Jones et al., 1991). Small plants that have
been splattered with soil are the exception to this rule.
4.3 Excessive washing is likely worse than no decontamination since soluble elements, including
B, K, and N, are likely to leach from the tissue.
4.4 Samples should be dipped quickly in the wash and rinse solutions. Sonneveld and van Dijk
(1982) recommended a time of 15 seconds.
4.5 Relatively high concentrations of Al (>100 mg kg-1), Fe (>100 mg kg-1), and Si (>1.0%) are
strong indicators of contamination (Jones et al., 1991). Titanium (Ti) has also been suggested
as an indicator of soil or dust contamination (Cherney and Robinson, 1982).
B. Drying
1. Principle
1.1 Water is removed from plant tisssue to stop enzymatic reactions and to stabilize the sample.
Removal of combined water also facilitates particle size reduction, homogenization, and
weighing.
2. Apparatus
2.1 Forced-air oven equivalent to Blue M Model POM-166E.

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3. Procedure
3.1 Separate or loosen tissue samples and place in paper containers.
3.2 Place containers in forced-air oven and dry at 80oC for 12 to 24 hours. NOTE: The original
condition and sample size will affect drying time.
4. Remarks
4.1 Drying times longer than 24 hours may be required depending on the type and number of
plant samples in the dryer.
4.2 Drying at temperatures under 80oC may not remove all combined water (Jones et al., 1991)
and may result in poor homogenization and incorrect analytical results.
4.3 Drying temperatures above 80oC may result in thermal decomposition and reduction in dry
weight (Jones et al., 1991).
4.4 Enzymes present in plant tissue are rendered inactive at temperatures above 60oC (Tauber,
1949). As a result, air drying may not stabilize samples and prevent enzymatic
decomposition. Samples should, therefore, be properly dried as soon after taking the sample
as possible.
4.5 Quick drying of a limited number of samples can be accomplished using a microwave oven
provided the samples are turned often and the drying process is closely monitored (Carlier
and van Hee, 1971; Shuman and Rauzi, 1981; and Jones et al., 1991).
4.6 If samples absorb significant amounts of moisture during grinding, additional drying may be
required prior to weighing for analysis. Drying time required will vary. Dry to constant
weight by making periodic weighings.
C. Particle-Size Reduction
1. Principle
1.1 Plant tissue samples are reduced to 0.5- to 1.0-mm particle size to ensure homogeneity and to
facilitate organic matter destruction.
2. Apparatus
2.1 Standard and intermediate Wiley-type mills equipped with 20-, 40-, and 60-mesh screens and
stainless steel contact points.
2.2 Cyclotec or equivalent high-speed grinder.
2.3 Medium bristle brush.

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2.4 Vacuum system.
3. Procedure
3.1 After drying, samples should be ground to pass a 1.0-mm screen (20 mesh) using the
appropriate Wiley Mill. A 20-mesh sieve is adequate if the sample aliquot to be assayed is
>0.5 g. However, if the sample aliquot to be assayed is <0.5 g, a 40-mesh screen should be
utilized (Jones and Case, 1990).
3.2 After grinding, the sample should be thoroughly mixed and a 5- to 8-g aliquot withdrawn for
analyses and storage.
3.3 Using a brush or vacuum system, clean the grinding apparatus after grinding each sample.
4. Remarks
4.1 Uniform grinding and mixing are critical in obtaining accurate analytical results.
4.2 Exercise care when grinding very small samples or plant material that is pubescent,
deliquescent, or that has a fibrous texture. These samples are difficult to grind in Wiley mills
and the operator should allow sufficient time for the sample to pass through the screen to
ensure homogeneity. In these instances, experience has shown that Cyclotec or equivalent
high-speed grinders are preferable.
4.3 Most mechanical mills contribute some contamination of the sample with one or more
elements (Hood et al., 1944). The extent of contamination depends on condition of the mill
and exposure time (Jones and Case, 1990). Grier (1966) recommended use of stainless steel
for cutting and sieving surfaces to minimize contamination.
4.4 Routine maintenance should be performed on mills to ensure optimum operating conditions.
Cutting knives or blades should be maintained in sharp condition and in adjustment.
D. Storage
1. Principle
1.1 After particle size reduction and homogenization, samples should be stored in conditions that
will minimize deterioration and maintain sample integrity for weighing and follow-up
analytical work.
2. Apparatus
2.1 Airtight plastic storage containers.
2.2 Storage cabinet located in cool, dark, moisture-free environment

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2.3 Refrigerator.
3. Procedure
3.1 After grinding and homogenization, a representative subsample is taken from the ground
plant material for analyses and storage. The sample should be placed in a container that can
be securely sealed.
3.2 Containers should then be placed in a cool, dry place for storage.
3.3 For long term storage, ground samples should be thoroughly dried, sealed, and placed under
refrigerated conditions (4oC) until the required analysis can be completed.
4. Remarks
4.1 If samples are placed in a cool (4oC), dark, dry environment, storage life is indefinite (Jones
et al., 1991).
4.2 Coin envelopes can also be used for sample storage, however, somewhat greater care must be
exercised in handling to prevent absorption of moisture. Collecting the ground sample in the
envelope and immediately placing into a desiccator cabinet or desiccator will minimize
moisture absorption.
E. Organic Matter Destruction – Dry Ashing
Plant tissue samples previously dried, ground, and weighed are prepared for elemental analysis
through decomposition or destruction of organic matter. Extensive work has been done to evaluate published
methods and to develop new and improved procedures. The best overviews on organic matter destruction are
found in books by Gorsuch (1970) and Bock (1978) and in the review articles by Tolg (1974) and Gorsuch
(1976). Two commonly used methods of organic matter destruction include dry ashing (high temperature
combustion) and wet ashing (acid digestion) (Jones et al., 1991). Both methods are based on the oxidation of
organic matter through the use of heat and/or acids.
1. Principle
1.1 Dry ashing is conducted in a muffle furnace at temperatures of 500 to 550oC for 4 to 8 hours.
For tissue high in carbohydrates and oils, ashing aids (Horwitz, 1980) may be required to
achieve complete decomposition of organic matter. After ashing, the vessel is removed,
cooled, and the ash is dissolved in nitric (HNO3) and/or hydrochloric (HCl) acid. The vessel
is filled to volume and diluted as needed to meet range requirements of the analytical
instrument.
2. Reagents and Apparatus