Soil Science/Agronomy/Horticulture 326 Manual... · final soil depth is approximately equal to that in the greenhouse pot. Water is then added quickly to the soil surface to get complete

Laboratory Manual Soil Science/Agronomy/Horticulture 326

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CONTENTS

Introduction 1

Exercise 1 Plant Response to N, P, and K 2

2 Nitrogen Requirement of Different Plant Species 7

3 Plant Response to Nutrient Sources and Soil Placement 12

4 Soil pH, pH Buffering Capacity, and Organic Matter Content 18

5 Soil Potassium Buffer Power 23

6 Mineralization of Organic Nitrogen 28

7 Tissue Testing 31

8 Total P and K concentrations in Plant Tissue 37

9 Total N in Plant Tissue 40

10 Determination of Available P and K in Soil 43

11 Determination of Soil pH, Lime Requirement and Soluble Salts 47

12 Development of Nutrient Deficiency Symptoms in Plants 53 Growing in Solution Culture


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INTRODUCTION

During the semester, you will complete 11 of the 12 greenhouse and laboratory exercises containedin this manual. The final exercise will be conducted as a demonstration in the greenhouse. In mostinstances, you will be assigned to work jointly with another student in your laboratory section. Thisprovides the opportunity to exchange ideas and discuss results as you observe. However, everystudent is expected to turn in individual laboratory reports and data sheets.

The 11 exercises are organized into four units with the following objectives:

Unit I: To demonstrate plant responses to soil applications of essential nutrients undergreenhouse conditions. You will study the response of various crops to applications ofnitrogen, phosphorus and potassium and compare crop responses to different sourcesand methods of application of these nutrients. Exercises 1, 2 and 3 make up this unit.

Unit II: To become familiar with analytical methods for determining some of the soilproperties and processes that affect plant growth. Exercises 4, 5 and 6 make up thisunit.

Unit III: To examine plant analysis as a means of identifying nutrient disorders, verifyingthe adequacy of soil fertilization, and gaining a more detailed understanding of howplants respond to soil treatments. Exercises 7, 8 and 9 make up this unit .

Unit IV: To introduce soil analysis as a tool for assessing the fertility status of soils and forserving as a basis for fertilizer and lime recommendation. Exercises 10 and 11 makeup this unit.

For each laboratory exercise, results obtained by each student or student pair are tabulated anddistributed to the whole class so that all of the students can see how their results fit into the “bigpicture”. For several of the exercises, each student will do additional analysis of the group data. Theeducational value of these exercises depends on the reliability of the greenhouse and laboratoryresults of each student. Sloppy work by just a few students can destroy much of the learning valueof many of the exercises.

Grading

Your performance in the laboratory accounts for 30% of your course grade. The following factorsare taken into consideration in determining your laboratory grade:

1. Careful attention to detail in setting up and carrying out each exercise.2. Turning in data sheets and assigned reports for all exercises on time.3. Errors in data entry and calculations.4. Neatness of your work area in the laboratory and in the greenhouse.5. Regular watering of pots for each greenhouse exercise.

Subject matter covered in the laboratory will be included as a separate laboratory examination at thetime the final lecture examination is given.


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EXERCISE 1

PLANT RESPONSE TO N, P, AND K

Nitrogen, phosphorus, and potassium are justifiably classified as primary nutrients, not only becauseplants require them in relatively large quantities, but also because these are the three nutrients thatmost commonly limit plant growth and crop production. Thus, unless a soil has been heavilyfertilized in recent times, it is generally possible to observe responses to N, P, and K under the veryintensive cropping that occurs in the greenhouse. This is less likely for secondary and micronutrients.

There are distinct advantages, but also distinct limitations, for studying plant response to nutrientapplications made under greenhouse conditions:

Table 1-1: Advantages and Limitations of Using the Greenhousefor Studying Plant Nutrient Responses.

Advantages

1. Through careful control of the environment,conditions can be such that only the factorbeing studied is growth-limiting.

2. Root volume is restricted to the soil in thepot in which the plants are growing. Aside fromgaseous nutrients absorbed through thestomata (e.g., S as SO2) or falling on the leavesvia atmospheric deposition (dust), all of thenutrients taken up by the plant must come fromthe soil in the pot. In the field, plants take upnutrients from an unknown volume of surfacesoil and also from the subsoil below thefertilized zone.

3. Large numbers of treatments can be testedat relatively low cost and low expenditure oftime. For the same cost in money and time,greenhouse experiments enable one to study amuch wider range of soils and soilamendments.

Limitations

1. The environment is artificial. Plants mayreact differently in the field because ofdifferences in factors such as temperature,humidity or radiation level.

2. The ratio of crop dry matter to soil volumeexplored by the roots is much higher ingreenhouse pots than in the field. Because ofthis, nutrient deficiencies occur at a higher levelof available nutrients than in the field. The ratioof transpiring surface to water storage in thesoil is also much higher in the greenhouse, sofrequent watering is required.

3. Results obtained in the greenhouse cannotbe applied directly to field conditions. Forexample, a soil testing method for a specificnutrient may correlate well with uptake of thatnutrient in the greenhouse, but the critical levelfor yield response must be determined fromfield calibration studies.

Because of these differences between field and greenhouse conditions, greenhouse studies arerestricted largely to studies such as the relative plant availability of various forms of a nutrient orto screening experiments that serve to develop technologies such as soil testing methods.

The full advantage of greenhouse studies cannot be realized unless special precautions aretaken to minimize experimental error. The soil must be mixed thoroughly so as to behomogeneous, nutrients must be applied as uniformly as possible, and efforts must be made toobtain a uniform stand of the test crop. But all of these precautions are a waste of time unlesssoil moisture is controlled properly. Most potted plants grow in artificial potting medium – often a


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Exercise 1 (Continued)

mixture containing coarse ingredients such as sand, peat, compost, vermiculite, perlite, etc. –intended to promote rapid drainage of excess water. Leaching of nutrients is extreme underthese conditions so this practice is not suitable for studying plant response to nutrient addition.

Growing potted plants in soil (other than sand) is generally so tricky due to problems of over-watering that it is usually not recommended to the public. We will grow potted plants in soil bycarefully controlling soil water content over a narrow range that is favorable to the plant and notconducive to leaching. The proper soil moisture is that corresponding to the amount of water thepot of soil can retain against drainage due to the force of gravity. This is commonly referred to asthe field moisture capacity (FMC) of the soil.

The FMC of the soil used in greenhouse studies generally cannot be guessed accurately. It mustbe measured. The apparatus commonly employed is shown in Figure 1.

To measure the FMC, you need a beaker that will hold a depth of soil equal to the depth of soil inthe greenhouse pots that will be used, a glass tube, and glass wool or cotton batting. Thepurpose of the glass wool or cotton batting and the glass tube is to allow air to escape when thesoil surface is flooded with water. The beaker is initially filled to approximately 1/3 the depth thatthe soil will have in the pot in the greenhouse and gently tapped on a table top or in the palm ofyour hand two to three times to pack the soil. The process is repeated twice more so that thefinal soil depth is approximately equal to that in the greenhouse pot. Water is then added quicklyto the soil surface to get complete coverage of the soil surface. The amount of water addedshould not wet the soil to more than about 1/3 of its total depth in about 5 minutes. The beaker isthen covered with plastic sheeting to prevent water evaporation.

After 24 to 48 hours, the soil is examined. If it is wet to its entire depth, too much water has beenadded and the whole procedure needs to be repeated. If wet to approximately 80% of its depth,a soil sample weighing 20 to 50 grams is removed from the middle portion of the wetted zone. The sample is weighed to +/- 0.01 g and dried for 24 hours at 105 oC. The dried soil weight isthen determined and the soil’s FMC is calculated.

Calculation of FMC

Soil moisture is always expressed on a dry-weight basis. The formula used is:

% H2O = (weight H2O) x 100% (weight dry soil)

= (weight wet soil) ! (weight dry soil) x 100% (weight dry soil)

(weight wet soil) = % H2O x (weight dry soil) + (weight dry soil) 100%

These formulas are used to calculate the soil weights needed in steps 2 and 8 of this exercise.


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Figure 1-1: Apparatus for estimating field moisture capacity of soilsfor greenhouse studies.


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MaterialsThe materials needed are: soil passed through a 2-mm sieve, air-dried, and mixed; pots; pot liners;balances; plastic sheets; nutrient sources (nutrient treatments are shown in a separate hand-out),seed; labels; marking pens; deionized water .

Procedure

1. Select three pots and line each with a plastic bag.

2. Compute the weight of air-dry soil that isequivalent to 1500 g of oven-dry soil. Thisweight is ________ g.

3. Weigh the amount of air-dry soil calculatedin step (2) into each pot.

4. Spread the soil from the first pot onto thesheet of plastic provided and add the nutrientsfor the treatment assigned to you. Thoroughlymix the treated soil and return the soil to thefirst pot.

5. Label the container with your name(s), labsection, and amount of the variable nutrient(N, P, or K) in mg/kg.

6. Repeat steps 4 and 5 for the remaining pots.

7. Randomly label the pots A, B, or C.

8. Compute the weight of the container whenthe soil is adjusted to its field moisturepercentage (FMC) of %. Include thistotal weight on your pot label.

9. Remove about a cup of soil from the sur-face of pot A and level the remaining soil.

10. Add approximately 3/4 of the water that willbe needed to bring the soil to FMC. (A specificvolume of water is not required at this point aslong as FMC is not exceeded.)

Remarks

1. Individual pots should weigh within 10 g ofeach other.

2. The air-dry soil contains _____ % water(get this value from the instructor).

3. Weigh the soil to +/! 10 g.

4. The added nutrients should be uniformlydistributed throughout the pot.

5-8. Pot label

The total weight is comprised of:

9. Save the soil for step 11.

10. If all of the water were added to the soilsurface (step 15), the soil at the surface couldbecome disturbed and some of the seedsuncovered.

Pot, plastic bag, and label ________ g

Oven dry soil ________ g

Water at FMC ________ g

Total weight ________ g

Required Information Example

Name(s) ______________ Jane D. & John Q.

Replicate (A, B or C)_ ___ Pot A

Lab Section ___________ Section 301 Nutrient added_____mg/kg 150 mg K/kg


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Procedure

11. Place ____ seeds of corn on the soil andcover with the soil just removed.

12. Place the pot on the balance and adddeionized water to adjust the container to thetotal weight computed in step (8). Use agraduated beaker.

13. Fold the plastic liner over the soil surfaceto minimize water loss by evaporation until thecorn germinates and emerges.

14. Thin to 4 seedlings per pot (or other numberas directed by your lab instructor).

15. Water the pots to the computed weightthree times weekly during the first two weeksand daily thereafter.

Continue watering until harvest time!

16. At the designated time, cut the plants atthe soil surface and place in the paper bagprovided. Label the bag and place in crop drier.Dry at 55 oC.

17. After the plants have dried to constantweight, record the dry weight of the plants plusbag to +/! 0.01 g.

18. Grind the samples and put the groundtissue into labeled plastic bags. You willanalyze the tissue for N, P and K (Ex. 8 & 9).

19. Weigh each empty paper bag and calcu-late the dry weight of the tissue.

20. Weigh 150 to 200 mg of each groundsample into a 50 ml beaker for P & K analysisin Exercise (8).

21. Weigh 100 to 150 mg of ground plant tissueand transfer quantitatively to a dry digestiontube for nitrogen analysis in Exercise (9).

Remarks

11. Wait until all of the water has infiltratedthe soil. The instructor will tell you how manyseeds to plant.

12. Hereafter, all watering will be done byweight using deionized water. Tap watercontains Ca, Mg, Fe, N and other unknowns.

13. As soon as plants emerge, uncover theplastic bag from the soil surface and fold downover the outside of the pot.

14. Shake the soil from roots of the seedlingsremoved back into the pot; discard plantsremoved. The remaining 4 plants should bespaced uniformly in the pot.

15. Water loss by evapotranspiration will below the first two weeks until there is significantleaf surface area. Notice changes in water usebetween cloudy days and bright, sunny days.

Harvest after completing Exercise (7).

16. Save all plant parts including desiccatedleaves that may have fallen off during harvest.Label the bag with the same information as onthe pot label.

17. For research, the samples would be keptin the drier until weighed to avoid absorption ofmoisture from the atmosphere.

18. Grind to pass a 20-mm screen. Sampleswill be taken for Ex. (8) in step (20) and for Ex.(9) in step (21).

19. Subtract the weight of each empty bagfrom the weight of tissue + bag. Fill out theData Sheet and hand it in.

20. Record the weights on the Data Sheet forExercise (8). These samples will be ashedprior to analyzing for P & K.

21. Record the weight to +/! 1 mg. Thedigestion tube should be dry so that the tissuewill not adhere to the neck.


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EXERCISE 2

NITROGEN REQUIREMENT OF DIFFERENT PLANT SPECIES

Nitrogen was established as an essential element in plant nutrition in the 19th century. Plantresponse to nitrogen is manifested in the production of vigorous plant growth with dark green leafcolor. Nitrogen is an important constituent of the chlorophyll molecule as well as amino acids,proteins, nucleotides, nucleic acids, amines, and amides.

The plow layer of most soils contains nitrogen mainly in the organic form, ranging from 0.08 to 0.4%(1,600 to 8,000 lbs per acre plow layer). Over the growing season, only 2 to 3% of this organicnitrogen is made available to crops under Wisconsin climatic conditions. Soils low in organic matterwill supply very little nitrogen. Continuous cropping without replacement of nitrogen reduces a soil'sability to supply nitrogen; thus, the need for nitrogen fertilizers to supplement natural supplies.

Nitrogen is taken up by plants as nitrate (NO3-) or ammonium (NH4

+) ions. Most plants can utilizeboth forms of nitrogen in their growth processes. An imbalance of nitrogen or an excess of thisnutrient in relation to P, K, and S prolongs the growing period and delays maturity. Too muchnitrogen produces succulent plants, which makes them more susceptible to disease. Some plantsshow weakening of stems causing lodging.

Nitrogen requirements vary among plant species. In this exercise, you will determine the optimumnitrogen rate for biomass production under greenhouse conditions by different plant species. In thefield, the nitrogen concentration varies with stage of maturity and portion of the plant sampled. Thenitrogen concentration of most plant parts decreases as the plant matures. When nitrogen is theyield-limiting factor, chlorophyll in the lower leaves breaks down and nitrogen is translocated to theupper leaves. Thus, deficiency symptoms for this element show up first on the older leaves. Therange in nitrogen concentration in the leaves of several crops is shown in the accompanying table.

Table 2-1. Nitrogen Concentration in the Leaves of Various Crops

Crop Nitrogen range(%, dry wt. basis)

Vegetable Crops (Geraldson and Tyler, 1990)Celery 2.5 - 4.0Kale and collards 4.0 - 5.0Lettuce 2.5 - 4.0Onion 1.5 - 2.5Pea 3.1 - 3.6Pepper 3.0 - 4.5

Potato 3.0 - 5.0Spinach 4.0 - 6.0

Sweet corn 2.6 - 3.5Sweet potato 3.2 - 4.2Tomato 2.5 - 6.0Turnip 3.5 - 4.5Watermelon 2.0 - 3.0

Cotton (Sabbe and Zelinski, 1990) 3.0 - 4.3Peanut 2.7 - 3.8Soybean (Small and Ohlrogge, 1973) 4.3 - 5.5Sugar cane blades (Bowen, 1990) 1.5 - 2.7


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Table 2-1. Nitrogen Concentrations in the Leaves of Various Crops (continued)

Nitrogen rangeCrop (%, dry wt. basis)

Small grains : Oats, wheat, barley (Westfall et al., 1990) 1.7 - 3.5Sugar cane blades (Bowen, 1990) 1.5 - 2.7Rice (Westfall et al., 1990) 2.5 - 4.5Sorghum (Whitney, 1970) 3.0 - 5.0Corn (Jones, 1990) 2.7 - 3.5Forage crops:

Alfalfa (Kelling and Matocha, 1990) 2.5 - 5.5Bromegrass (Krueger and Scholl, 1970) 2.5 - 3.6Orchardgrass (Kresge and Younts, 1963) 3.2 - 3.5Tall fescue (Hallock et al., 1966) 3.4 - 3.8Kentucky bluegrass (Butler and Hodges, 1967) 2.6 - 3.2Bermudagrass (Adams et al., 1967) 2.6 - 3.2Johnsongrass (Spooner et al., 1971) 1.6 - 1.8Millet (Clapp and Chambles, 1970) 2.5 - 3.5Pangola grass (Harris et al., 1968) 1.7 - 2.0Sorghum-sudan and sudangrass (Dotzenko, et al., 1966) 2.0 - 3.0

Annual and perennial ryegrass (Thomas et al., 1952) 3.8 - 4.2Creeping bentgrass 4.5 - 5.5

References Cited

Adams, W.E., A.W. White, R.D. McCreery, and R.N. Dawson. 1967. Agron. J. 59:247-250.

Bowen, J.E. 1990. In R.L. Westerman (ed.) Soil Testing and Plant Analysis, p.454. No. 3, Soil Sci. Soc. Am. Book Series. Madison.

Butler, J.D., and T.K. Hodges. 1967. J. Hortic. Sci. 2:62-63.

Clapp, J.G., Jr., and D.S. Chambles. 1970. Crop Sci. 10:345-349.

Dotzenko, A.D., N.E. Hamburg, G.O. Hinze, and W.H. Leonard. 1966. Colorado Agric. Exp. Sta. Tech. Bull. 87.

Geraldson, C.M., and K.B. Tyler. 1990. In R.L. Westerman (ed.) Soil Testing and Plant Analysis, pp.549-562. No. 3, Soil Sci. Soc.Am. Book Series. Madison.

Hallock, D.L., R.H. Brown, and R.E. Blaser. 1966. Virginia Agric. Exper. Stn. Agron. Res. Rep. 112.

Harris, H.D., V.N. Schroder, and R.L. Silman. 1968. Fla. Agric. Exp. STa. Tech. Bull. 725.

Jones. J.B. 1990. Plant analysis as an aid in fertilizing corn and sorghum. In R.L. Westerman (ed.) Soil Testing and Plant Analysis,pp.603-643. No. 3, Soil Sci. Soc. Am. Book Series. Madison.

Kelling, K.A., and J.E. Matocha. 1990. In R.L. Westerman (ed.) Soil Testing and Plant Analysis, pp.549-562. No. 3, Soil Sci. Soc. Am.Book Series. Madison.

Kresge, C.B., and S.E. Younts. 1963. Agron. J. 55:161-164.

Krueger, C.R., and J.M. Scholl. 1970. Wis. Agric. Exp. Sta. Res. Rep. 69.

Sabbe, W.E., and L.J. Zelinski. 1990. In R.L. Westerman (ed.) Soil Testing and Plant Analysis, pp.469-493. No. 3, Soil Sci. Soc. Am.Book Series. Madison.

Small,H.G., Jr., and A.J. Ohlrogge. 1973. In L.M. Walsh and J.D. Beaton (eds.), Soil Testing and Plant Analysis, pp. 315-327. SoilSci. Soc. Am., Inc. Madison.

Spooner, A.E., W.R. Jeffrey, and H.J. Hunneycutt. 1971. pp. 12-15. In Ark. Agric. Exp. Sta. Bull.769.

Thomas, B.A., A. Thompson, V.A. Oyanuga, and R.H. Armstrong. 1952. Exp. Agric. 22:10-22.

Westfall, D.G., D.A. Whitney, and D.M. Brandon. 1990. In R.L. Westerman (ed.) Soil Testing and Plant Analysis, pp.495-519. No. 3,Soil Sci. Soc. Am. Book Series. Madison.

Whitney, D.A. 1970. Kansas State Univ. mimeo 3a-162-1-300.


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Materials

Soil passed through a 2-mm sieve and air-dried; pots with a capacity of about 2 liters, plastic bagpot liners; balances with a capacity of 2 kg or more; plastic sheets for mixing soils and amendments;ammonium nitrate; triple super phosphate; potassium chloride; gypsum (CaSO4 * 2H2O) and zincsulfate; labels, marking pens, corn seeds; deionized water.

Procedure

1. Select four pots and line each with a plasticbag.

2. Compute the weight of air-dry soil that isequivalent to 1500 g of oven-dry soil.

3. Weigh g of soil of air-dry soilinto each pot.

4. Compute the weights of NH4NO3 required togive the following concentrations of N. Pot N rate, mg/kg A 0 B 75 C 150 D 300

5. Spread the soil from each pot(sequentially) onto the plastic sheet provided,and add the appropriate amount of NH4NO3 togive the desired concentration of N.

6. Also add to each pot, 200 mg/kg P astriple super phosphate, 300 mg/kg K as KCl, 18mg/kg S as CaSO4

* 2H2O, and 2 mg/kg Zn as

ZnSO4.

7. Mix the treated soil thoroughly; return thesoil to the pot.

8. Remove up to one cup of soil from thesurface of each pot and level the remaining soil.

Remarks

1. The pot weights should not vary by morethan +/- 10 g.

2. The air-dry soil contains % moisture.Use the moisture formula from Exercise 1.

3. This is the amount of air-dry soil calculatedin step (2). Check your calculations with the labinstructor before proceeding.

4. Show your calculations and results to the labinstructor before proceeding.

5. Begin with the control treatment (no N) andproceed to the highest N rate to minimizecontamination of the plastic sheet with N.

6. Use the balance provided.

7. Label the N treatments: A. 0 N; B. 75 N; C. 150 N; D. 300 N.

8. This will be used later to cover the seed.Small seeds will require less than one cup tocover them. Check with the instructor.


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Pete P. & Robin H.Corn75 mg/kg Lab 301Planted 2/5/02


Procedure

9. Add approximately 3/4 of the amount ofwater needed to bring the soil to FMC.

10. After the water has infiltrated, plant thenumber of seeds suggested by your labinstructor; cover with soil removed in step 8.

11. Label the pot as in Exercise 1. Includethe rate of N applied and the crop planted.

12. Place the pot on the balance and adjustthe soil to its FMC (_______ %) by carefullyadding enough water to attain the calculatedtotal weight.

13. Close the plastic liner over the soil surface.As soon as the plants emerge, uncover thesoil and fold the plastic bag over the sides ofthe pot.

14. Record the number of plants emergeddaily until the number is constant. Then thinplants to the number designated by your labinstructor.

15. Water to FMC as often as needed toassure adequate moisture. Water lossbetween waterings should not exceed 30% ofthe total water in the soil at FMC.

Remarks

9. Use a graduated beaker. Exact volume isnot needed at this point. See step 12 for thetotal amount of water to add, and apply about3/4 of that amount.

10. The number of seeds depends on expectedpercent germination and the eventual size of theplants.

11. Sample label:

12. Calculation of the total weight at FMC:

Pot + plastic bag + label ________ g

Soil (oven-dry basis) ________ g

Water at FMC ________ g

Total weight ________ g

13. This will minimize water loss by evaporationand prevent the surface soil from dryingexcessively and delaying germination.

14. Discard the thinned plants.

15. The permanent wilting point is about 1/2FMC, but photosynthesis is slowed at about 3/4FMC.


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Procedure

16. At the designated time, cut the plants ineach pot at the soil surface and place inseparate paper bags. Label the bags the sameas the labels on the pots.

17. Weigh the dried plant tissue plus the bag.

18. Grind the tissue and put the ground tissuein a labeled plastic bag. The tissue will beanalyzed later.

19. Weigh the paper bag after the tissue hasbeen removed for grinding.

20. Record the dry matter yield on the datasheet.

Remarks

16. Place the samples in the box provided. Thelab instructor will put them in the drier and drythem to constant weight at 60o to 70o C.

17. Do not remove the tissue from the bag atthis point. Record the weight of the tissue plusthe bag.

18. Do not discard the bag after grinding.

19. Subtract the bag weight from the weightrecorded in step 18.

20. Leave your data sheet with the instructor


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EXERCISE 3

PLANT RESPONSE TO NUTRIENT SOURCE AND PLACEMENT

When fertilizing plants, the primary consideration is the appropriate rate of application. The chemicalforms of the nutrients in the fertilizer are generally of secondary importance. There are, however,some instances where the chemical form of the nutrient is of concern. For example, cranberry usesnitrogen only in the ammonium form, and the rate of nutrient release to plants from fertilizer isimportant when fertilizing container-grown plants or turfgrass.

Fertilizer is any material that contains one or more essential plant nutrients, is applied primarily forits nutrient content, and is known to promote plant growth through an increase in nutrient supply. Inthe fertilizer industry, fertilizers that contain one or two essential plant nutrients are known asfertilizer materials. They may be applied to the soil as such but are often mixed with other fertilizermaterials to provide a complete fertilizer -- one that contains all three of the primary plant nutrients,N, P, and K.

A given nutrient may be obtained from several different fertilizer materials. These materials differ innutrient concentration and, very often, in the chemical form in which the nutrient exists. In thisexercise, the class will observe plant responses to different fertilizer materials. Examples of fertilizermaterials and their distinguishing characteristics are given in Table 3-1.The guaranteed nutrientcontent of fertilizers is expressed as their "grade". (See Table 3-1.) Grade designations consist ofthree numbers separated by hyphens. An example is 10-8-16. These numbers signify that thefertilizer contains a minimum of 10% total N, 8% citrate-soluble phosphate (expressed as P2O5), and16% water-soluble potash (expressed as K2O). The numbers are always given in the samesequence: N-P2O5-K2O. A fertilizer such as ammonium nitrate contains no P or K, and its grade is33.5-0-0. Note that the theoretical N concentration of NH4NO3 is 35%.

Fertilizers are not pure chemicals. The cost of purifying them is prohibitive, and there is no need toremove ordinary impurities from a product that will be applied to the soil. Also, there is no P, P2O5,K, or K2O as such in fertilizer. Phosphorus is always accompanied by a cation (usually Ca2+ or NH4

+)and potassium by an anion (usually Cl- or SO4

2-). Use of the terms phosphate (P2O5) and potash(K2O) is a carry-over from the early days of agricultural chemistry when elements analyzed in a totalelemental analysis of soils were reported as oxides. To convert % P2O5 to the elemental form (P),multiply % P2O5 by 0.44; to convert K2O to the elemental form (K), multiply by 0.83.

Inorganic Fertilizer Materials

Raw materials for inorganic fertilizers come directly from nature. Atmospheric nitrogen is the principalsource of nitrogen fertilizers. Air is about 79% N2 by volume and contains about 36,000 tons overevery acre of the earth's surface. Combining hydrogen with atmospheric nitrogen under pressure,heat, and a suitable catalyst produces anhydrous ammonia (NH3):

N2 + 3 H2 =====> 2 NH3

The H2 is obtained from methane (CH4) or hydrolysis of water. When methane is used, urea is oftenmade in the same fertilizer plant by converting the C from methane to CO2 and reacting the CO2 withNH3:

2 NH3 + CO2 =====> (NH2)2CO + H2O


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Other nitrogen fertilizers are made by reacting NH3 with various acids such as H3PO4, HNO3, andH2SO4.

Fertilizer phosphorus comes from "phosphate rock," a calcium phosphate ore deposit that may beof either igneous or sedimentary origin and that contains the phosphate mineral apatite. Becauseof the low solubility of apatite, treatment of the ore with strong acids (H3PO4, H2SO4, HNO3) isnecessary to produce soluble phosphate products.

Fertilizer potassium is obtained by mining deposits that were created by the evaporation of ancientseas under arid conditions. In contrast to rock phosphate, potash ore can be used directly aspotassium chloride (often called "muriate of potash"), potassium sulfate, or as potassium magnesiumsulfate. Processing is usually necessary, however, to remove impurities such as common salt (NaCl).

Organic materials

Several organic materials, including manure and crop residues, can be added to soil to increasenutrient supply, especially nitrogen. Activated sewage sludge, dried blood, and fish contain a higherpercentage of nitrogen than do most manures and crop residues.

Most of the nitrogen from organic sources becomes available within the first three to four weeksfollowing application. Thereafter, the amount of nitrogen released is very small.

Some organic materials contain significant amounts of phosphorus but are typically low in potassium(Table 3-1). Applications of animal manure that are heavy enough to meet the nitrogen requirementsof a crop usually provide more than enough phosphorus for that crop. Such rates of application overan extended period of time can increase available phosphorus to excessive levels.

Slow Release Nitrogen Materials

When water-soluble nitrogen fertilizers are applied to soil, significant amounts of nitrogen may belost from the soil by leaching, denitrification, or ammonia volatilization. Use of slow-release nitrogenfertilizers generally reduces these losses. However, slow-release nitrogen fertilizers are tooexpensive for economical use in feed, forage, and fiber production.

Slow-release nitrogen fertilizers are produced by altering the solubility of materials or by includingcompounds that require microbial activity for transforming organic N to available forms. A commonmethod used is to coat water-soluble compounds with materials that are water-insoluble but containcracks and/or pores. Water can enter by diffusion through these openings creating a saturatedsolution which is then forced out through the same openings or which builds up pressure sufficientto disrupt the coating material. The most common example of a coated nitrogen fertilizer is sulfur-coated urea (SCU). This material is urea with a coating of elemental sulfur, a binding agent, asealant, and a microbiocide. Nitrogen release rates can be varied by controlling the thickness of thesulfur coating.


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Table 3-1: Composition of some common fertilizers.

Fertilizer Formula Grade Nutrient Conc. Nutrient Conc.

% %

Inorganic N itrogen Materials

Anhydrous am monia NH3 82-0-0 N 82 -- --

Am monium nitrate NH4NO3 33.5-0-0 N 33.5 -- --

Am monium sulfate (NH2)2SO4 21-0-0 N 21 S 26

Calcium nitrate Ca(NO3)2 15.5-0-0 N 15.5 Ca 24

Monoammonium phosphate NH4H2PO4 11-48-0 N 11 P 21

Diam monium phosphate (NH4)2HPO4 18-46-0 N 18 P 20

Potassium nitrate KNO3 13.5-0-44.5 N 13.5 K 37

Urea CO(NH2)2 46-0-0 N 46 -- --

Urea am monium nitrate soln. NH4NO3 28-0-0 N 28 -- --

+ CO(NH2)2

Inorganic Phosphorus Materials

Normal superphosphate Ca(H 2PO4)2 0-20-0 P 8.7 Ca 20

+ CaSO4

Triple superphosphate Ca(H 2PO4)2 0-46-0 P 20 Ca 14

Monoammonium phosphate NH4H2PO4 11-48-0 P 21 N 11

Diam monium phosphate (NH4)2HPO4 18-46-0 P 20 N 18

Inorganic Potassium Materials

Potassium chloride KCl 0-0-60 K 50 Cl 46

Potassium sulfate K2SO4 0-0-50 K 42 S 17

Potassium nitrate KNO3 13.5-0-44.5 K 37 N 13.5

Potassium m agnesium sulfate K2SO4 + 0-0-22 K 18 Mg 11

2 MgSO4 S 22

Organic Fertilizer Materials

Material Nutrient Amount Nutrient Amount Nutrient Amount

lb/ton lb/ton lb/ton

Beef m anure1 N 14 P 3.9 K 9

Chicken manure1 N 25 P 10.9 K 10

Dairy manure1 N 10 P 2.2 K 8

Sheep manure1 N 28 P 4.2 K 20

Swine m anure1 N 10 P 2.8 K 8

% % %

Sewage sludge (dry) N 5.4 P 2.5 K 0.4

Activated sewage sludge (dry) N 6.0 P 0.9 K 0.5

Turkey dropping compost (dry) N 5.0 P 0.9 K 3.3

_________________________________

1 As excreted, without bedding. Most of the inorganic N is urea or NH3, which can be lost upon drying.

Slow-Release Materials

Material N conc. Material N conc.

% %

Sulfur-coated urea 32-37 Methylene-coated urea 32

Resin-coated urea 34 Isobutylidenediurea 31

Urea formaldehyde 35-42


15


There are three types of coating that can be applied to nitrogen materials:

1. Impermeable coatings with small pores through which solutions of the nutrients diffuse.

2. Impermeable coatings that must be broken by abrasive, chemical, or biological actionbefore nitrogen can be released.

3. Semi-permeable coatings through which water diffuses and creates internal pressuressufficient to disrupt the coating.

Uncoated organic fertilizer compounds of low water solubility are used as nitrogen sources for high-value crops, turf, and ornamentals. These compounds are produced by the reaction of urea and anumber of aldehydes to form compounds that are sparingly soluble in water. Two of the most commonare the ureaforms (UF) and isobutylidenediurea (IBDU). Urea reacts with formaldehyde in thepresence of a catalyst to form a mixture of compounds under the generic name ureaforms -- alsoknown as methylene ureas.

There are numerous potential processes for the dispersal of soluble fertilizer salts containing nitrogen,phosphorus, and potassium, etc. into asphalt, water, paraffins, oils, gels, polymers, and resins, whichare referred to as matrixes. Nitrogen compounds of limited water solubility, such as urea-formaldehyde, have also been incorporated into a matrix or have been embodied in expandedvermiculite, perlite, clay, glass frits, and similar materials. Matrix materials have been found to beeffective in rice production, pastures, and vegetable crops but too costly for field crop production.

Influences of Nutrient Source and Method of Application

Plant response to fertilizer is conditioned by the amount of nutrient applied, the method of application,and soil characteristics. Exercise 1 demonstrates nutrient rate effects. The present exercise examinesthe influence of nutrient source and method of application on plant response to nitrogen, phosphorus,and potassium.

Variation in plant response to nitrogen in different fertilizer materials relates primarily to losses of thenutrient from soil-plant systems. The mechanisms for nitrogen loss are leaching of nitrate (NO3

-),denitrification, and volatilization of ammonia (NH3) from the soil surface. Leaching cannot occur inclosed pots. However, nitrogen loss through denitrification and volatilization can occur. The extent ofdenitrification varies with the level of NO3

- in the soil solution and the aeration status of the soil.Volatilization of nitrogen is restricted to situations wherein solutions on or near the soil surface containhigh concentrations of NH4

+ and the pH approaches or surpasses 7.3.

Plant response to different phosphatic fertilizers varies with the solubility of the phosphate, the ionsassociated with the phosphate, and conditions prevailing for diffusion transport of phosphate ions toplant root surfaces. Plant response to row applications of phosphatic fertilizers is significantly reducedwhen the water solubility of the phosphate is less than about 50%. Plant response to phosphate is alsoreduced when the fertilizer is placed in a soil zone that is subject to periodic drying. Plant recoveryof fertilizer phosphate can often be enhanced by including ammoniacal nitrogen in the fertilizer.


16


All common potassium fertilizers are totally soluble in water. Potassium moves in soil much morereadily than phosphorus though not nearly as fast as NO3

-. Consequently, when variation in plantresponse to different potassium sources is observed, the most common reason is a nutritive effect ofthe anion associated with K in the fertilizer.

Materials The materials needed are: soil passed through a 2-mm sieve, air-dried, and mixed; pots; pot liners;

labels; plastic sheets, balances; fertilizer materials; marking pens; sorghum seeds; deionized water.

Procedure

1. Select four pots and line each with aplastic bag.

2. Compute the weight of moist soil that isequivalent to 1.5 kg of oven-dry soil.

3. Weigh _______ kg of air-dry soil (equi-valent to 1.5 kg of oven-dry soil) into each pot.

4. Spread the soil from the Pot A onto aplastic sheet, and add N, P, K, S, and Zn asinstructed. Mix thoroughly, and return themixture to the pot.

5. Repeat for Pot B. Label pots A and B"incorporated".

6. Repeat step 4 with Pots C and D, but donot apply the nutrient that will be topdressed atthis time.

7. Remove 1 cup of soil from the surface andlevel the remaining soil.

8. Add approximately 3/4 of the amount ofwater needed to bring the soil to FMC.

9. After the water has infiltrated, plant 15sorghum seeds or other number suggested byyour lab instructor; cover with the soil removedin step 7.

Remarks

1. The pot weights should not vary by morethan +/- 10 g.

2. The air-dry soil contains _______ %moisture. Use the moisture formula fromExercise 1.

3. Check your calculations with the labinstructor before proceeding.

4. This is the “incorporated” treatment.Fertilizer materials and amounts to be usedare given on a separated handout.

5. The incorporated and topdressed treat-ments are done in duplicate.

6. These pots receive the “topdressed” treat-ment. The topdressed nutrient will be appliedafter planting and final watering.

7. This will be used later to cover the seed.

8. Use a graduated beaker. Exact volume isnot needed at this point. See step 11 for totalamount of water to add and apply about 3/4 ofthat amount.

9. The number of seeds to plant depends onexpected germination percentage.


17


Procedure

10. Label the pot, including fertilizer sourceand placement.

11. Place the pot on the balance and carefullyadd water to adjust the soil to its total weightwhen at a field moisture percentage of _____ %.

12. Weigh the the fertilizer for the nutrient thatwill be topdressed, and spread uniformly overthe soil surface of Pot C. Repeat for Pot D.

13. Place the four pots on the bench spaceassigned for your section, but do not cover thesoil surface with the plastic liner.

14. After 7 days, thin to 5 uniform plants perpot. Water pots as needed.

15. When you water, notice any differences inplant height or color associated with fertilizerplacement.

16. At the designated time, cut the plants ineach pot at the soil surface and place inseparate paper bags labeled with the sameinformation as on the pot label. Dry to constantweight at 55o C.

17. Weigh the dry samples plus bags to +/-0.01 g.

18. Grind the samples and put the groundtissue in labeled plastic bags for later chemicalanalysis.

19. Weigh the empty bags and calculate the netweight of the dry tissue.

Remarks

10. Sample LabelIrma B. & J.B. QuickLab 301MilorganiteTopdressed - Pot CPlanted 2/12/02

11. The total weight of the pot at FMC is:

Pot, plastic bag, and label _______ g

Soil (oven-dry basis) _______ g

Water at FMC _______ g

Total _______ g

12. The topdressed nutrient is applied afterwatering so that it is not incorporated bywatering.

13. Moisture accumulates on the undersurface of the liner and drops onto the soilsurface. This tends to incorporate the moresoluble topdressed materials in a non-uniformpattern.

14. Thin any plants that emerge after initialthinning.

15. Also, notice any inhibition of germination asinfluenced by fertilizer placement.

16. Be sure to include all plant parts. Cut allplants at a uniform height.

17. Don't forget to weigh the empty bags aftergrinding!

18. The instructor will demonstrate the use ofthe tissue grinder.

19. Record your data on the Data Sheet andhand it to the instructor.


18

EXERCISE 4

SOIL pH, pH BUFFERING CAPACITY AND ORGANIC MATTER CONTENT

Soil pH Buffering Capacity

Soil pH buffering capacity is the ability of soil to resist a change in its pH when acid-forming or base-forming materials are added. The capacity to resist pH change is an extremely important characteristicof soil. Without this property, the pH of the soil would fluctuate widely whenever water passed throughthe profile or fertilizers or other materials were applied as a means of enhancing crop growth. In humidregions, soils without pH buffering ability would quickly become very acid and mineral weatheringwould accelerate drastically. Thus, agricultural sustainability of a soil is very strongly related to its pHbuffering capacity.

Soil pH buffering capacity is directly related to the number of pH-dependent cation exchange sites inthe soil. Sources of pH-dependent charge include the weak acid functional groups on organic matter(primarily carboxylic and phenolic functional groups) and broken oxygen bonds at the edges of layersilicates and oxides of Fe and Al (which can exhibit partial positive or negative charges depending onwhether they are protonated or not). Under acid conditions, most of the pH-dependent cationexchange sites are occupied by H+ or Al3+, which must be neutralized in order to raise the soil pH. [Asthe pH rises above 5, Al3+ is precipitated as Al(OH)3]. Consequently, there is a direct relationshipbetween the pH buffering capacity of an acid soil and its pH-dependent cation exchange capacity.Neutralization of hydrogen on a pH-dependent site creates a negative charge on that site whichincreases the cation exchange capacity.

The pH buffering capacity of Wisconsin soils is largely determined by organic matter content and toa lesser extent by clay content. Fine textured soils generally have higher organic matter and claycontents than well-drained sandy soils and thus have higher buffering capacities.

In humid tropical regions where intensive weathering has taken place and soils are frequently verystrongly acid (pH < 5), pH buffering capacity derives to a large extent from the protonation and de-protonation of amorphous sesquioxides and from the neutralization of exchangeable aluminum. It isnot until the pH of these soils is adjusted to above pH 5.5 that organic functional groups begin to playa significant role in buffering soil pH. These soils may have a net positive surface charge (exhibitanion exchange capacity) when very acid and convert to negative surface change (exhibit cationexchange capacity) when the pH is raised.

The pH buffering capacity of a soil is a direct indication of the amount of acidity or alkalinity that needsto be neutralized in order to bring about a specified change in pH. Thus, the higher the pH bufferingcapacity of an acid soil, the greater the amount of liming material that must be applied in order to raisethe pH a specified amount. Organic soils have very high buffering capacities. It is generally noteconomical to lime them to as high a pH as is the practice for mineral soils.

In this exercise, class members will measure pH, pH buffering capacity, and organic matter indifferent soils. Compilation of the results obtained by all students will permit examination of how thepH buffering capacities of Wisconsin soils differ and relate to organic matter content.


19


Soil Organic Matter

Soil organic matter can be measured several different ways. In the method used here, the soil sampleis reacted with an excess of a strong oxidizing agent (chromic acid in H2SO4), and the excess istitrated with a standard ferrous solution. Heat to speed the reaction is supplied by the heat of dilutionof concentrated H2SO4. The procedure is not specific for carbon but determines any easily oxidizedsubstance. However, experience has shown that the amount of material oxidized by this treatment isequivalent to oxidation of approximately 77% of the total carbon in soil organic matter. This value doesnot necessarily apply to soil organic matter from other climatic zones. Since soil organic mattercontains approximately 58% carbon, a good approximation of the organic matter content can beobtained. Presence of reducing substances (chlorides, Feo ,Fe2+, Mn2+) in the soil leads to high results.

To avoid the use of the heavy metal, Cr, in chromic acid and subsequent disposal problems and tospeed operations, the UW Soil & Plant Analysis Lab estimates soil organic matter by weight loss whena sample is heated to 360 oC. The sample is first heated at 105 oC to remove moisture, then at 360oC to burn off organic matter. At higher temperatures, there is the danger of weight loss from structuralwater in some soil clays and from decomposition of carbonates. Presence of CaSO4 *2H2O (gypsum)or NaHCO3 in sub-humid soils leads to high estimates of organic matter determined by weight losson ignition.

The most modern methods of soil carbon analysis employ combustion of a soil sample at 600 to 1000oC in a stream of O2 gas, with infrared detection of carbon as CO2. Heat is applied either using aninduction or resistance furnace. Above 900 oC, carbon from carbonate minerals (if present) is releasedso that organic carbon is not identical to total carbon. With these high temperature dry combustionmethods, instrumentation costs are higher but no assumptions regarding completeness of combustion,loss on ignition of inorganic soil constituents, or average oxidation state of organic carbon arerequired.

Reactions and Equations

In this reaction, organic C, which has an average oxidation state of (0), is oxidized with therelease of 4 e- per C atom:

C(0) + 2 H2O ===> C(IV)O2 + 4 H+ + 4 e-

The Cr(VI) in the dichromate ion, Cr2O72-, as chromic acid, undergoes reduction to Cr(III):

Cr(VI)2O72- + 14 H+ + 6 e- ===> 2 Cr(III)3+ + 7 H2O

And, overall, on a 12 e- basis:

3 C(0) + 2 Cr(VI)2O72- + 16 H+ ===> 3 C(IV)O2 + 4 Cr(III)3+ + 8 H2O

The unit of interest for our purposes is mmol of electrons undergoing this oxidation/reductionreaction, i.e., mmol(e-). [Remember: one mole of electrons contains 6.02x1023 electrons(Avogadro’s number).] This unit, mmol(e-), is more relevant to the purpose than the molarconcentration of the dichromate. After the chromic acid has reacted with organic carbon, theexcess electron acceptors remaining as unreacted Cr(VI) are determined by electron titration witha standardized ferrous iron, Fe(II), solution, which is oxidized to Fe(III):

6 Fe2+ + Cr2O72- + 14 H+ ===> 2 Cr3+ + 6 Fe3+ + 7 H2O


20


Apparatus

Soil samples of different texture, pH and organic matter; scoop calibrated for 10 g of a "light coloredsilt loam" (2 million lbs/acre plow-layer); 20-mL pipets; plastic vials; pH meter; 500-mL conical flasks;25-mL dispensers; 25- or 50-mL burettes; magnetic stirrer.

Reagents

KOH solutions: Dissolve 11.2 g KOH in about 800 mL of deionized water and dilute to one liter to give0.2 M KOH. Dilute 25, 50, 100, and 200 mL of the 0.2 M KOH to one liter. The resulting concentrationswill be 0, 0.005, 0.01, 0.02, and 0.04 M. Addition of 25 mL of these solutions to 10 g of soil will give the equivalent of 0, 12.5, 25, 50, and 100 mmol KOH per kg of soil, respectively.

Solid NaF.

Concentrated H2SO4.

Standard 0.1667 M K2Cr2O7: Dissolve 49.04 g of K2Cr2O7 in water and dilute to 1 liter. Theconcentration with respect to electrons transferred in reducing Cr(VI) to Cr(III) is 1.000 mole/L or 1.000mmol/mL.

Ferroin indicator: Dissolve 3.7 g of o-phenanthroline monohydrate and 1.74 g of FeSO4 * 7H2O in 250mL of water.

Ferrous solution, 0.5 M: Dissolve 196.1 g of Fe(NH4)2(SO4)2 * 6 H2O in 800 mL of water containing 20mL of concentrated H2SO4 and dilute to 1 liter. The Fe2+ in this solution slowly oxidizes on exposureto air so it must be standardized against the dichromate solution daily.

Procedure for pH and pH Buffering Capacity

Procedure

1. Measure 10 g of the soil assigned to youinto each of five plastic vials.

2. Add 25 mL of the solutions as shownbelow, using the appropriate dispenser:

mmol KOH addedVial no. Solution per 10 g soil 1 Deionized water 0

2 0.005 M KOH 0.125

3 0.01 M KOH 0.250

4 0.02 M KOH 0.500

5 0.04 M KOH 1.000

Remarks

1. Use the calibrated scoop technique, whichthe lab instructor will demonstrate.

2. Be sure to keep a record of what solutionwas added to which vial.


21


Procedure

3. Shake the soil suspension intermittently forone hour.

4. Measure the pH of each suspension, usinga glass electrode pH meter.

5. On graph paper, plot [mmol KOHadded]/[kg(soil)] on the y-axis vs [soil pH] on thex-axis for each KOH addition.

6. From the resulting graph, find the buffercapacity (BC) from the slope of the lineconnecting the 0 KOH addition and the pointrepresenting each KOH addition.

7. Turn in the completed data sheet and graphto your lab instructor.

Remarks

3. Cap the vials and shake vigorously for 15seconds every 10 minutes.

4. The instructor will standardize anddemonstrate the use of the pH meter.

5. A computer generated graph is acceptable.

6. Slope = [y2 ! y1]/[x2 ! x1]

= mmol KOH/kg(soil) ! 0 pH in treated vial ! pH in vial 1

7. Next week, you will be asked to use theclass data to plot pH buffering capacity vs soilorganic matter.

Procedure for Determining Soil Organic Matter

Procedure1

1. Weigh 1 to 2 g of oven-dried, medium-textured soil to +/! 0.01 g and transfer to a 500-mL conical flask.

2. Add 20 mL of 0.1667 M K2Cr2O7 and swirl tomix.

3. In a fume hood and under instructorsupervision, add 20 mL conc. H2SO4; swirl theflask gently for 1 minute.

4. Allow to stand for 30 minutes.

5. Dilute the suspension with about 200 mL ofwater.

6. Add 0.2 g of NaF and 10 drops of o-phenanthroline indicator.

Remarks

1. Weigh out 0.1 to 0.2 g for organic soils, 2 to3 g for light-colored sandy soils.

2. Use a pipette and pipetting bulb. Molaritybased on electrons transferred is 1.000 M or1.000 mmol(e-)/mL.

3. Use gloves and eye protection! Do not getany H2SO4 on your clothing. SO2 fumes will begenerated from heat of dilution.

4. The reaction proceeds slowly anddiminishes as the flask cools.

5. Exact volume is not important; dilutionproduces a clearer endpoint in a turbid solution.

6. The NaF complexes Fe3+ produced whenFe2+ is oxidized. Fe3+ interferes with theendpoint. The o-phenanthroline is a redoxindicator, not a pH indicator.


22


Procedure1

7. Place a magnet in the flask and the flask onthe magnetic stirrer. Titrate with 0.5 M ferroussolution to a burgundy endpoint. The color of thesolution at the beginning may be anywhere fromorange-yellow to dark green, depending on theamount of organic matter in the sample. Astitration proceeds, the color of the solution shiftsto green, then turbid gray near the endpoint. Thecolor changes abruptly to wine-red at theendpoint.

8. Run a reagent blank (same procedure exceptno soil used).

9. Calculate:Oxidized soil organic matter, mmol(e-)/g.2

Percent carbon.3

Percent soil organic matter.4

10. Rinse all glassware and place in the dishpanprovided.

Remarks

7. If less than 4 mL of ferrous solution is used,the procedure should be repeated with a smallersample as there is danger that the oxidation ofthe organic matter was not complete.

8. This determines any oxidizable materialintroduced in the reagents and glassware.

9. See calculations below.

10. Turn in the data sheet to your instructor.

1 Based on a variation of the method of Walkley and Black. Soil Sci. 37:29 (1934) as reported byD.W. Nelson and L.E. Sommers, 1996. Methods of Soil Analysis, Part 3. Chemical Methods. SSSA/ASA. Madison, WI.

2 Oxidized soil organic matter, mmol(e-)/g :

mmol(e-)/g = (mL Fe2+ for blank !mL Fe2+ for sample) x (Conc. of Fe2+ solution, mmol(e-) /mL)Soil sample weight, g

3 Percent carbon:

% C = (Oxidized soil organic matter, mmol(e-)/g ) x [0.012 g C / 4 mmol(e-)] x (1/0.77) x 100%

where it is assumed that 4 mmol of e- are required to oxidize 1 mmol (12 mg or 0.012 g) of soil organic carbon and that the oxidation using the heat of dilution of concentrated sulfuricacid is 77% complete.

4 Percent soil organic matter (% SOM)

% SOM = % C x (1/0.58)

where it is assumed that the soil organic matter is 58% C by weight.


23

EXERCISE 5

SOIL POTASSIUM BUFFER POWER

The plant availability of nutrients in soil directly relates to the quantities of the nutrients that come incontact with root surfaces. The quantities of nutrients transported to root surfaces depend on theconcentrations of the nutrient ions in the soil solution and the ability of the soil to replenish the ionswhen plant absorption occurs.

The labile forms of a nutrient include those in the soil solution and those in equilibrium with thedissolved forms. These forms vary with the nutrient and soil being considered. Labile forms ofpotassium that are potentially available to plants include dissolved, exchangeable, and somenonexchangeable. The K immediately available for plant nutrition is that present in the soil solutionat the root surface. Replenishment of solution K from the solid phase is of great importance as theconcentration of K in the soil solution is low. Of the solid-phase forms, exchangeable K is the mostreadily available because it is in rapid equilibrium with the soil solution. Soils that contain littleexchangeable K depend on transformations from nonexchangeable forms to replenish theexchangeable and solution phases upon depletion. This nonexchangeable labile K pool is the Kassociated with the soil micaceous mineral, illite, or the K fixed in vermiculite. The equilibria existingbetween solution, exchangeable, and nonexchangeable K forms is reversible, but attainment ofequilibrium with nonexchangeable K forms is relatively slow. In terms of plant nutrition, the rate oftransfer between the various labile phases is of prime importance.

The importance of replenishing dissolved K is illustrated by the following example: The concentration of K in the soil solution varies over an approximate range of 1 to 10 mg/L. At fieldmoisture capacity (assuming FMC = 25% by weight), a silt loam soil weighing 2,000,000 lbs/acre plowlayer contains 500,000 lbs of water per acre plow layer. This is equal to about 227,000 kg (or liters)of water. If the concentration of K is 10 mg/L, the soil solution in an acre plow layer will contain2,270,000 mg of K or 2.27 kg of K. This is equal to 5 lbs of K. Since a 5 ton/acre alfalfa cropremoves about 300 lbs of K per acre, it is apparent the amount of K in solution at any given time isless than 2% of the crop’s requirement. The rest must be released from solid-phase forms over thegrowing season.

Mathematical models of plant nutrient uptake require equations describing the diffusion of nutrientsfrom the bulk soil to the plant root. These equations include a term called the buffer power for a givennutrient. Nutrient buffer power is defined as the change in concentration of total labile nutrient per unitchange in concentration of the nutrient in solution. Concentrations are expressed on a unit volume ofsoil basis (e.g., g/m3 soil or mg/cm3 soil). A high buffer power indicates a high ratio of labile solid-phase nutrient to that in the soil solution whereas a buffer power of 1 indicates that all of the labilenutrient is in the soil solution (e.g., nitrate-N). The buffer power varies with the nutrient considered,the adsorption capacity of the soil and the degree of saturation of the nutrient adsorbing sites. For agiven soil, phosphate will usually show the highest buffer power, calcium, magnesium, potassium,ammonium and sodium intermediate, and sulfate, chloride and nitrate low buffer powers.

The purpose of this exercise is to obtain estimates of K buffer power of a soil that has received variousK additions. You will be using the K concentration in a 4 mM Sr(NO3)2 extract as an estimate of theK concentration in the soil solution (extracting the actual soil solution is difficult). The K extracted with1 M NH4OAc or Bray P-1 solution will be used as estimates of total labile K. Later, you will plot classdata of [labile K/cm3 (soil)]/[dissolved K/cm3 (soil)] for soil samples with different K additions to seehow buffer power varies with varying concentrations of total labile K.


24


Materials

Soil extraction flasks (50-mL conical flasks); filter tubes; filter paper; soil samples with different Kadditions; flame photometer; balance; scoop calibrated for 10 g of light-colored silt loam; dispenserbottles; and the following reagents:

4 mM Sr(NO3)2 solution: Dissolve 0.846 g Sr(NO3)2 in about 800 mL of deionized water and dilute to1 liter.

Bray P-1 extracting solution (0.03 M NH4F+ 0.025 M HCl): Dissolve 1.11 g of NH4F in about 900 mLof water; add 2.1 mL of concentrated HCl and dilute to 1 liter.

NH4OAc extracting solution (1 M): Add 57 mL glacial acetic acid to about 600 mL of deionized water.Add slowly, with mixing, 267 mL of concentrated NH4OH. Cool. Adjust to pH 7.7 +/! 0.2 with HOAcor NH4OH, and dilute to 1 liter.

Standard K stock solution (315 mg/L K): Dissolve 1.0895 g of oven-dried (105 oC) KH2PO4 in about900 mL of deionized water. Add 5 mL of concentrated H2SO4 as a preservative and dilute to 1 liter.This solution contains 250 mg/L P and is also used for the standard stock P solution in Exercise 11.

Working K standards: Dilute the volumes of 315 mg/L K shown in the table below to 100 mL with 4mM Sr(NO3)2, water, or 1 M NH4OAc for Sr(NO3)2-extractable K, Bray P-1 extractable K, or NH4OAc-extractable K, respectively.

Sr(NO3)2-K Bray P-1 and NH4OAc-K K stock solution Conc. of K K stock solution Conc. of Kper 100 mL standard in dilute standard per 100 mL standard in dilute standard

mL mg/L mL mg/L

0 0 0 0 1 3.15 2 6.3 2 6.3 4 12.6 4 12.6 8 25.2 6 18.9 12 37.8 8 25.2 16 50.4


25


Procedure for Determining Potassium Buffer Power

Procedure

1. Label six 50-mL soil extraction flasks withnumbers 1 to 6.

2. Weigh 10 g of the assigned soil andtransfer to soil extraction flask [1].

3. Weigh 2 g of the assigned soil (twice) andtransfer to soil extraction flasks [2] & [3].

4. To the 10-g sample in flask [1] and toempty flask [4], add 25 mL of 4 mM Sr(NO3)2

solution.

5. Add 20 mL of the Bray P-1 extractant tothe 2-g sample in soil extraction flask [2] and toempty soil extraction flask [5].

6. Add 20 mL of 1 M NH4OAc extractingsolution to the 2-g sample in soil extraction flask[3] and to empty soil extraction flask [6].

7. Place the soil extraction flasks on the orbitalshaker and shake for 30 minutes.

8. Filter each suspension and reagent blankthrough Whatman No. 2 filter paper.

9. Set the flame photometer to read 0 "% T"with the extracting solution. (Use water to set 0% T with the Bray-1 extracts.)

10. Set the flame photometer to read 100% Twith 25.2 mg/L K in the Sr(NO3)2 solution. Itmay be necessary to repeat steps 9 and 10.

.

Remarks

1. Keep track of what sample went in whatflask.

2-4. Weigh to +/! 0.01 g. Use a funnel totransfer the soil to the extraction flask. Include areagent blank (no soil) to account for Kcontamination in the reagents, glassware, filterpaper, your finger tips (especially smokers), etc.Use the appropriate dispensers for the differentextractants..

5. This solution is used for available K inWisconsin and Michigan soil testing labs.

6. This solution is used to measure available Kin many other states.

7. Exchange reactions proceed quickly, butthis amount of time is needed to ensure thatequilibrium is reached.

8. Use the funnel tubes provided. Collect 7 to10 mL of filtrate.

9. The lab instructor will demonstrate how touse the flame photometer.

10. Standards must be made up in the samematrix as the samples. Different matrices havedifferent viscosities and electrolyte compositionsthat affect K flame analysis.


26


Procedure

11. Aspirate filtrates from the Sr(NO3)2 flasks(1) and (4) and record % T to +/- 0.5.

12. Repeat steps 8-10 using K standards inwater and your Bray P-1 extracts (2) and (5).

13. Repeat steps 8-10 using K standards inNH4OAc and your NH4OAc extracts (3) and (6).

14. Using the appropriate standard curve,convert % T to mg/L K in solution.

15. Record your data on the data sheet andcalculate K buffer power estimates1 for your soilsample using K extracted by NH4OAc and BrayP-1 as estimates of labile K and the Kconcentration in Sr(NO3)2 as the estimate of Kconcentration in the soil solution.

16. Rinse all glassware and place in thedishpan provided.

Remarks

11-13. Aspirate both the sample and the reagentblank. If the sample reading exceeds 100 % T,dilute the sample with the appropriate extractionsolution and re-run. Take the dilution factor intoaccount in your calculations.

14. Note that the x and y axes may not be atthe same scale on the three standard curves.

15. Equations for estimating the buffer powerbased on this one sample are given below andon the Data Sheet. Later you will plot class datato see how buffer power varies with K additionsfor the soil that you used.

16. The lab assistants will wash the glasswarelater.

1 This approximation assumes that the plot of Labile K vs Dissolved K is linear and passes throughthe origin. The class data for this soil that you will plot later will show how the buffer power (slopeof the plotted line at a given point) varies with the amount of labile K in the soil.

Calculations are on the next page.


27


Calculations

The units for K buffer power are: weight of labile K per unit volume of soil weight of dissolved K per unit volume of soil

Labile nutrient per unit volume of soil = labile K extracted volume of soil used

Dissolved K per unit volume of soil = weight of K per unit volume of Sr(NO3)2 x volume of solution volume of soil

The volume of solution per unit volume of soil is the volumetric fraction of water in the soil. If wewant to determine the buffer power at FMC, we must convert FMC from % by weight, FMCw, to %by volume, FMCv. This requires multiplying FMCw by soil bulk specific gravity, DB(soil)/D(water).

FMCv = FMCw x DB(soil) = weight of water x weight of soil/volume of soil D(water) weight of soil weight of water/volume of water

where the numerical value for D(water) is assumed to be 1 for metric units of g/cm3 or Mg/m3.

Therefore:

Dissolved K per unit volume of soil at FMCv = weight of K per unit volume of Sr(NO3)2 x FMCv.

Calculation of Dissolved K, g/m3 at FMCv

KSr(NO3)2 in sample, mg/L = Measured Ksr(NO3)2 in sample, mg/L x DF

Corrected dissolved K, mg/L = KSr(NO3)2 in sample, mg/L ! KSr(NO3)2 in blank, mg/L

Corrected dissolved K, mg/1000 cm3 = Corrected dissolved K, mg/L x 1 L/1000 cm3

Dissolved K, g/m3 (solution) = Corrected dissolved K, mg/1000 cm3 x 106 cm3/m3 x 1 g/1000 mg(Note that dissolved K, g/m3(solution) is numerically equal to corrected dissolved K, mg/L)

Dissolved K, g/m3(soil) = Dissolved K, g/m3(solution) x FMCv% 100%

Calculation of Labile K, g/m3

The following equations apply to Labile K determined for either NH4OAc or Bray extractants:

Corrected Labile K, mg/L = (K concentration in sample, mg/L ! K concentration in blank) x DF

Labile K, g/m3(soil) = Corrected Labile K, mg/L x 20 cm3(solution) x DB(soil), Mg/m3(soil) 2 g(soil) x 1000 mg/g x 1000 cm3/L x 1 Mg/106g

Using the numerical values without including all of the units:

Labile K, g/m3(soil) = Corrected Labile K, mg/L x DB(soil) x10

Buffer Power Approximation at FMCv

Buffer power approximation = [Labile K, g/m3(soil)] [Dissolved K, g/m3(soil)]


28

EXERCISE 6

MINERALIZATION OF ORGANIC NITROGEN

The rate of mineralization of organic N is controlled by the nature of the organic material and by thesuitability of the soil environment for microbiological growth. Finely divided organic material with highenergy content and favorable ratios of carbon to nitrogen, phosphorus, and sulfur is decomposedmuch more readily than soil humus or coarse residues with high lignin content and high ratios ofcarbon to nitrogen, phosphorus, or sulfur. As for growth conditions, the requirements for optimumgrowth of microorganisms are similar to those of higher plants: soil pH near neutrality, soil moistureslightly below field capacity, and an adequate supply of all essential mineral nutrients.

In this exercise, each pair of students will be assigned the same silt loam soil but the organic N sourcewill vary. After two weeks of incubation, the soils will be analyzed for ammonium and nitrate nitrogen.Examination of the class results will reveal some of the factors regulating mineralization of organic N.

Materials

Silt loam soil; dried and ground organic materials with different C:N ratios; quartz sand; glass vials;polyethylene sheets, rubber bands; 5-g calibrated scoop; balances; wash bottles; dispensers; burette,Kjeldahl flasks, Kjeldahl steam distillation apparatus; and the following reagents:

2 M KCl: Add 148 g KCl to about 800 mL of deionized water in a 1-liter volumetric flask. Dissolve thesalt and dilute to volume.

Devarda's alloy (50% Cu, 45% Al, 5% Zn): Ball-mill the alloy to pass a 100-mesh screen and at least75% through a 300-mesh screen. Store in a stoppered bottle.

MgO: Heat "heavy" MgO in a muffle furnace at 600 to 700 oC for 2 hr. Cool in a desiccator over KOHpellets, and store in a tightly stoppered bottle.

Boric acid indicator solution: Dissolve 40 g H3BO3 in about 900 mL of deionized water. Add 25 mL ofmixed indicator consisting of 0.33 g bromcresol green and 0.165 g methyl red dissolved in 500 mL ofethanol. Mix and dilute to volume.

THAM, 0.050 M: Dissolve 6.057 g of tris-hydroxyamino methane (THAM), oven-dried at 105 °C, indeionized water and dilute to 1 liter.

Standard H2SO4 (0.0357 M): Dilute 2 mL of concentrated H2SO4 to 1 liter with deionized water. Mixwell and standardize against 0.050 M tris-hydroxyamino methane (THAM). Calculate the amount ofwater that needs to be added to dilute the H2SO4 to exactly 0.0357 M. Add the required amount ofwater and check the resulting concentration by titrating with the THAM solution. One mL of 0.0357 MH2SO4 contains 0.0174 mmol of H+ which will neutralize 1 mg of NH3-N.


29


Procedure

1. Measure one 5-g scoop of the assigned soiland two 5-g scoops of quartz sand into each oftwo glass vials.

2. Add 0.100 g of the assigned organicmaterial to one of the vials and mix.

3. Add 3 mL of deionized water to each vial.

4. Cover the vials with squares of polyethylenesheeting and fasten in place with rubber bands.

5. Label the vials and place in the incubatorset at 35 +/! 2 oC.

6. After two weeks, remove the samples fromthe incubator.

7. Scoop 5 g of the original soil into a thirdvial.

8. Add 10 to 15 mL of 2 M KCl to each vial;shake the vial until all of the soil and sand issuspended; then decant the solution into a 100-mL Kjeldahl distillation flask.

9. Repeat step 8 twice more.

10. Add one scoop of Devarda’s alloy (0.2 g), adrop of surfactant and, just before connectingthe flask to the distillation unit, add one scoop ofMgO (0.2 g).

Remarks

1. The sand helps to provide better aerationpromoting faster mineralization anddiscouraging denitrification of NO3

-.

2. Weigh to +/! 0.001 g. This application isequivalent to 20 tons per acre (assuming 2millions lbs of soil per acre).

3. Use a pipette.

4. Polyethylene is permeable to gases but notto water.

5. Mineralization will be relatively rapid at thistemperature.

6. You can find them in the incubator with allof the others if you labeled them properly.

7. This is a check on the amount of inorganicN in the unincubated soil. The incubated samplewith no amendment applied is used to measurethe amount of inorganic N mineralized from soilorganic matter.

8. Mineralized N will be present as NH4+ and

NO3-. The high concentration of K+ in the KCl will

displace exchangeable NH4+ into solution.

9. All of the inorganic N should be in solution.Do not try to transfer the soil to the distillationflask. Sand causes "bumping" during distillation.

10. Devarda's alloy is a reducing agent,converting NO3

- to NH4+; MgO raises the pH

high enough to convert NH4+ to NH3 without

hydrolyzing organic N. If the MgO is allowed todissolve in the solution before connecting theflask, some NH3 may be lost.


30


Procedure

11. Place 5 mL of boric acid indicator solutioninto a 50-mL conical flask and position the flaskunder the condenser of the steam distillationapparatus.

12. Carefully connect the distillation flask to theapparatus and introduce steam.

13. Collect about 15 mL of distillate; then shutoff the steam.

14. Remove the distillation flask.

15. Titrate the distillate to a faint pink color withthe standard H2SO4 solution.

16. Calculate the amount of N in each soilsample and the amount of N mineralized. Givethe data to the instructor.

17. Rinse all glassware and place it in the dishpan provided.

Remarks

11. Boric acid traps NH3: NH3 + H3BO3 ----> NH4B(OH)4.

12. This is a delicate glass apparatus. The labinstructor will demonstrate how to use it.

13. Most of the NH3 comes over in the first 5mL.

14. Place the distillation flask in the sink to cool.

15. Be sure to read the burette before and aftertitrating.

16. See calculations below and on the datasheet.

17. Use tap water. Glassware will be cleanedfurther by a lab assistant.

Chemical Reactions Involved

NH3 + H3BO3 + H2O ===> NH4B(OH)4

NH4B(OH)4 + H+ ===> NH4+ + H3BO3 + H2O

Calculations

1 mmol of H2SO4 contains 2 mmol of H+, therefore: (0.0357 mmol H2SO4/mL) x (2 mmol H+/mL H2SO4) = 0.0714 mmol H+/mL H2SO4

0.0714 mmol H+/mL H2SO4 reacts with 0.0714 mmol of NH4B(OH)4 containing 0.0714 mmol of N

(14 mg N/mmol N) x 0.0714 mmol N/mL H2SO4 = 1.00 mg N/mL H2SO4

N mineralized, mg = (N extracted from incubated soil, mg) ! (N extracted from original soil, mg)

N mineralized from added org. N = N mineralized (org. N added) ! N mineralized (not added)

N mineralized, mg/kg(soil) = N mineralized, mg x 1000 g(soil)/kg(soil) Soil weight, g(soil)


31

EXERCISE 7

PLANT TISSUE TESTING

Tissue testing refers to a rapid, semi-quantitative analysis of plant tissue in the field. It is usedprimarily for trouble-shooting purposes but can also be used to:

• Call attention to the need for laboratory tests.• Supplement soil testing to determine whether the fertilizer

recommendation was adequate -- or excessive.• Verify deficiency symptoms.• Survey large areas quickly.• Follow the uptake of nutrients in field research plots.

Plant analysis, on the other hand, is a quantitative analysis of one or more elements in plant tissue.It is carried out in a laboratory and is used where more precise results are required.

Experience is required for reliable results with most tissue testing kits. This is so because thecomposition of plant tissue varies with age and portion of plant sampled. It will be affected also byweather and other factors. The N, P, and K contents of plant sap are higher early in the growingseason than toward maturity. With inadequate nutrition, plants go through a period of stress about thetime of flowering or early seed formation. Nutrient demands are high at this time, and the soil may notbe able to supply them as fast as required. The use of starter fertilizer may provide a supply of readilyavailable nutrients early in the season but may be inadequate later. Also, early in spring plants utilizenutrients accumulated in the soil over winter. Testing in the spring may not reflect accurately asufficient supply of nutrients later.

In the tissue testing kit used in this exercise, plant sap is analyzed for NO3--N, H2PO4

-, and K+. Evenplants growing under N stress will have some NO3

- accumulating overnight. This is assimilated readilyinto organic N in daylight. Consequently, tissue testing should not be done early in the morning.

In this exercise, test strips will be used to estimate the concentrations of nitrate-N and potassium incell sap. “Cardy” nitrate-nitrogen and potassium meters will also be used. The “Cardy” meters utilizeion-specific electrodes sensitive to nitrate and potassium ions (similar to the glass electrode used tomeasure hydrogen ion activity). Results from the two methods will be compared. Phosphorus in plantsap will be measured as H2PO4

- with freshly-prepared reagents and filter paper strips.

Reagents for Tissue Test Kits

A complete list of the reagents employed in an early tissue testing kit and the procedures for preparingthe test papers is given by Morgan and Wickstrom (1956)1:

Nitrate powder: Mix 10 g MnSO4 * H2O, 2 g finely-powdered Zn, 4 g sulfanilic acid, and 2 g alpha-naphthylamine with 25 g BaSO4. Add 75 g citric acid and another 75 g of BaSO4. Thoroughly mix allingredients. Grind any coarse materials to a fine powder before adding to the mixture. (Test stripscontaining these or similar ingredients are now available.2)

P-K reagent no. 1, concentrated: (0.0032 M (NH4)6Mo7O24 * 4 H2O in 3.75 M HCl): Dissolve 4 g ofammonium molybdate in 137 mL of deionized water. Slowly, with stirring, add 63 mL conc. HCl.


32


P-K reagent no. 1, diluted: Dilute 10 mL of concentrated P-K reagent no. 1 with 40 mL of deionizedwater.

P reagent no. 2: Add approximately 2 g of stannous oxalate powder to 30 mL of deionized water.Shake before using.

Check the chemicals. Last year's kit is useless unless the chemicals have been stored in arefrigerator. There are simple tests to check for deterioration:

• Nitrate strips: In good condition, the strips should be white. They turn gray as thechemicals deteriorate.

• Phosphate chemicals: Since saliva contains phosphate enzymes, wet an area onthe test paper with your tongue. Run the P test on that area and another areawhere no saliva has been placed. This should give the contrast between dark bluereaction to no reaction if the chemicals are good.

• Potassium strips: Tips should be a bright orange color. When washed with theacid P-K reagent, the orange tip should be a pale yellow. The tips are brownishorange and brownish yellow, respectively, if the strips are too old.

The nitrate and potassium test strips are generally good for one season. Unopened test containersstored in a refrigerator and opened containers kept in a cool place away from moisture condensationmay be good for more than one season. The test strips should be kept in a box separate from theliquid chemicals. The phosphorus solutions can deteriorate rapidly depending on temperature andcontamination.

Procedure for Using the Plant Tissue Test Kits

Procedure

Nitrate1. Cut one corn plant just above soil level.

2. With a pair of pliers, squeeze plant sap fromthe cut end of the stem onto the white tip of atest strip.

3. After ten seconds, observe the color at thetip of the strip and compare it with the colorscale on the kit.

4. Use the nitrate table in the following sectionto interpret the results.

Remarks

Nitrate1. Nitrate concentration is usually highest atthe base of the stem because that which is notassimilated in the roots is converted to organicforms in the upper portion of the plant.

2. Be sure to use a clean pair of pliers andsqueeze out enough sap to wet the tip of thetest strip.

3. Nitrate reacts with the chemicals coating thetip of the test strip to give a pink to purple color.

4. Record the results on your data sheet.


33


Procedure

Phosphorus

1. With a pair of pliers, squeeze plant sap fromthe cut end of the stem onto the test papersupplied for the P test.

2. Apply a single drop of P-K reagent no. 1 tothe wetted area.

3. Next apply one drop of stannous oxalate.

4. From the relative intensity of the blue colorthat develops, estimate the relative level of P inthe plant from the phosphorus table on the nextpage.

Potassium

1. With a pair of pliers, squeeze sap from thecut end of the stem onto the orange tip of thepotassium test strip.

2. After 60 seconds, dip the tip of the test stripinto a small test tube containing P-K reagent no.1 or 0.7% HNO3.

3. After soaking for 60 seconds, remove thestrip and compare the color of the test zone withthe color scale on the kit.

4. Interpret the reading using the potassiumtable on the next page.

Remarks

Phosphorus

1. Squeeze out enough sap to wet an areaequivalent at least to the size of a dime.

2. The molybdate in this reagent forms acomplex with the phosphate.

3. Stannous oxalate reduces the phospho-molybdate complex to a blue colored compound.

4. Record the results on your data sheet.

Potassium

1. Be sure to thoroughly wet the orange tip ofthe test strip. The K in the sap reacts with thesodium cobalti-nitrite in the test strip to form anorange compound of low solubility.

2. The acid solution dissolves the sodiumcobalti-nitrite but not the orange potassiumcobalti-nitrite.

3. The intensity of the orange color reflects theconcentration of K in the sap.

4. Record the results on your Data Sheet.


34


Interpretation of Tissue Test Kit Results

NitrateEstimated concentrations of nitrate associated with the color of the tip of the nitrate test strip towhich plant sap has been applied are given in the table below:

Approximate concentration Color Interpretation N NO3

- - - - - mg/kg - - - -White Very low 0 - 1 0 - 5Light pink/purple Low 2 - 7 10 - 30Pinkish purple Medium 15 - 25 60 - 100Deep purple High 55 - 110 250 - 500

PhosphorusThe sufficiency of P in the test plant is estimated from the color developed on the test paper towhich plant sap was applied. The interpretation is given in the table below:

Color InterpretationColorless or very light blue Very lowLight blue LowFair blue MediumDeep blue High

PotassiumThe sufficiency of K in the test plant is estimated from the color of the tip of the test strip to whichplant sap was applied. The interpretation is given in the table below:

Color Interpretation Approx. K Conc. mg/kg

Pale yellow Very low 0 - 250Yellowish orange Low 250 - 450Medium orange Medium 700 -1000Deep orange High > 1000


35


“Cardy” Nitrate and Potassium Meters

Calibration (To be done by instructor): Calibrate the nitrate meter with standard solutions containing 150and 2000 mg/kg NO3

- (34 and 452 mg/kg N). Follow the instructions provided with the meter. Convertvalues displayed as NO3

- to NO3- - N by multiplying displayed value by 0.226. Calibrate the potassium

meter using standard solutions containing 150 and 2000 mg/kg K. Follow the instructions provided withthe meter.

Nitrate Determination with the “Cardy” Nitrate Meter

Procedure

1. Cut one corn plant just above the soil.

2. Place the sampling strip provided on thesensor of the Cardy Nitrate Meter.

3. Squeeze sap from the cut end with pliersonto the sampling strip.

4. Close the cover and wait for the reading tostabilize.

5. Convert the mg/kg NO3- reading from the

meter to mg/kg NO3--N.

6. Rinse the sensor with deionized water andblot dry with a paper towel.

7. Repeat steps 2 -5 twice.

8. Calculate the average of the three readings.

Remarks

1. Save all plant parts for dry matter analysis.all plants must be cut at the same height.

2. Use tweezers to handle the strips so as notcontaminate them with perspiration from yourfingertips.

3. Saturate an area approximately 1 cm x 1cm with sap.

4. The meter should stabilize in 30 to 45seconds.

5. mg/kg NO3--N = mg/kg NO3

- x 0.226

6. Blot gently so as not to scratch the sensor.

7. The procedure should be repeated to obtaina representative average measurement. In thefield, the procedure would be repeated on 10 to20 plants from different parts of the field.

8. Record the average readings on your datasheet.


36


Potassium Determination with the “Cardy” Potassium Meter

Procedure

1. Cut one corn plant just above the soil.

2. Place the sampling strip provided on thesensor of the “Cardy” Potassium Meter.

3. Squeeze sap from the cut end with pliersonto the sampling strip.

4. Close the cover and wait for the reading tostabilize.

5. Rinse the sensor with deionized water andblot dry with a paper towel.

6. Repeat steps 2 -5 twice.

8. Calculate the average of the three readings.

Remarks

1. Save all plant parts for dry matter analysis. allplants must be cut at the same height.

2. Use tweezers to handle the strips so as notcontaminate them with perspiration from yourfingertips.

3. Saturate an area approximately 1 cm x 1 cmwith sap.

4. The meter should stabilize in 30 to 45seconds.

5. Blot gently so as not to scratch the sensor.

6. The procedure should be repeated to obtaina representative average measurement. In thefield, the procedure would be repeated on 10 to20 plants from different parts of the field.

8. Record the average readings on your datasheet.

1 Morgan, N.D., and G.A. Wickstrom. 1956. Give your plants a blood test: Guide to quick tissue tests.Better Crops with Plant Food Magazine. American Potash Inst.

2 EM Science, Gibbstown, NJ 08027.


37

EXERCISE 8

TOTAL P AND K CONCENTRATIONS IN PLANT TISSUE

Dry combustion of plant tissue in a muffle furnace drives off the organic C, H, and O as CO2 and H2O,leaving behind carbonates, oxides, phosphates, borates, and sulfates of the cations present in thetissue. Dry combustion also volatilizes N, Cl, and some Mo and cannot be used for their analysis. SomeP, B, and S will volatilize as well if precaution is not taken before combustion to ensure that there is anexcess of cations over anions in the combustion vessel. If the tissue is deficient in cations, strontiumacetate can be added to ensure an excess of cations without interfering with the elements normallytested for in the tissue.

The inorganic compounds formed during dry combustion, with the exception of SiO2, are readilydissolved in acid solutions. Once dissolved, the concentrations of the constituent elements can bedetermined by appropriate methods. The results of the analyses are expressed as percentage or mg/kgof each element in the tissue. These concentrations, when multiplied by the dry weight of the plants, givethe amount of the element absorbed (total uptake). By comparing uptake from fertilized and unfertilizedsoils, it is possible to estimate the amount of added nutrient recovered by the crop.

Materials

Muffle furnace, 50-mL beakers, acid-washed filter paper, funnels, 3- and 25-mL pipettes,spectrophotometer tubes, spectrophotometer, flame photometer.

2 M HCl: Dilute 170 mL concentrated HCl to 1 liter.

HNO3 - vanadomolybdate reagent: Dissolve 0.62 g of NH4VO3 in 300 mL of hot water; cool and add 125mL of concentrated HNO3. Dissolve 12 g of (NH4)6Mo4O24 @4 H2O in 400 mL of water, add to the vanadatesolution, and dilute to 1 liter.

Standard P solution (250 mg/L): Dissolve 1.0984 g of oven-dried (105 °C) KH2PO4 in about 900 mL ofdeionized water. Add 5 mL of concentrated HNO3 as a preservative and dilute to 1 liter. (This is thesame as the 315 mg/L K solution used in exercise 5.)

Dilute standard P solutions: Pipette 0, 10, 20, 40, 60, and 100 mL of the 250 mg/L P standard solutioninto 500-mL volumetric flasks. Add 50 mL of 2 M HCl and dilute to 500 mL. The P concentrations ofthese standard solutions are 0, 5, 10, 20, 30, and 50 mg/L.

NH4OAc, 4 M: Add 228 mL glacial acetic acid (HOAc) to about 400 mL of deionized water. Slowly, withmixing, add 267 mL of concentrated NH4OH. Cool. Adjust the pH to 7.0 +/! 0.2 with HOAc or NH4OH,and dilute to 1 liter.

NH4OAc, 1 M: Transfer 250 mL of 4 M NH4OAc to a 1-liter volumetric flask, and dilute to volume withwater.

Standard K solution, 5,000 mg/L: Dissolve 9.535 g of oven-dried (105 °C) KCl in water and dilute to 1liter.


38


Standard Na solution, 2,000 mg/L: Dissolve 5.084 g of oven-dried (105 °C) NaCl in water and dilute to1 liter.

Concentrated K, Na standard solution; 2,000 mg/L K + 40 mg/L Na: Pipette 100 mL of 5,000 mg/L Ksolution and 5 mL of 2,000 mg/L Na solution into a 500-mL volumetric flask. Dilute to volume, and adda crystal of thymol as a preservative. Store in a refrigerator.

Dilute K and Na standard solutions: Pipette 0, 5, 10, 15, 20, and 25 mL of 2,000 mg/L K + 40 mg/L Nasolution into 500-mL volumetric flasks. Add 42 mL of 4 M NH4OAc and 18 mL of 2 M HCl to each flask,and dilute to 500 mL.

Procedure

1. Weigh 150 to 200 mg of ground plant tissueand transfer to a 50-ml beaker.

2. Ash at 500 °C for two hours.

3. Cool the samples to room temperature.

4. Add 3 ml of 2 M HCl to dissolve the ash.

5. Add 25 ml of deionized water, mix and filter.

6. Pipette 3 ml of the filtrate into a spectro-photometer tube. Add 3 ml of water and 3 ml ofthe vanadomolybdate reagent, and mix.

7. Let stand for at least 10 minutes.

8. Wipe the bottom half of the spectro-photometer tube with lens tissue.

Remarks

1. Record weights to the nearest mg.

2. Ashing at a higher temperature could result involatilization of some K, P, S, B, Mo, Cl, and Na,(or even melt the beakers).

3. The beakers could crack if removed from thefurnace while hot, and convection currents coulddisturb the ash.

4. Some effervescence may be seen ascarbonates in the ash react with the acid.

5. The sample is diluted to get the concen-trations of P and K into the desired concentrationranges for analysis..

6. A yellow P-vanadomolybdate complex isformed. The intensity of the color is proportionalto the concentration of P.

7. The color takes 10 minutes to develop fullyand is stable for 2 to 3 days.

8. Fingerprints or other smudges in the opticalpath result in false, high P readings.


39


Procedure

9. Read the absorbance (red scale on spectro-photometer) of the sample at 440 nm. Convertthe absorbance reading to mg/L P in solutionusing the standard P curve.

10. Calculate % P in the tissue.

11. Pipette 3 ml of the filtrate (step 5) into a 50-ml beaker or flask. Add 3 ml of deionized waterand 3 ml of 1M NH4OAc, and mix.

12. Aspirate this solution into the flame photo-meter, and read the % Transmission on theappropriate scale.

13. Calculate % K in the sample..

Remarks

9. Absorbance equals [2 ! log (% T)]. It is usedin preference to % T (% of incident lighttransmitted through the sample) for plotting thestandard curve because it gives a near linearrelationship with concentration.

10. See the Calculations section below.

11. The NH4OAc serves as a "radiation buffer" inthe flame photometric determination of K, helpsto prevent salt crystals from clogging theatomizer, and eliminates differences in matrixproperties which might affect the flow ratethrough the atomizer.

12. Convert this reading to mg/L K in solutionusing the standard K curve.

13. See the Calculations section below. Leaveyour data sheet with the instructor.

Calculations

%P in tissue:= [P(extract ! blank), mg/L] x 28 mL x [9mL/3mL] x [1 L/1000 mL] x 100%

[tissue wt, mg]

= 0.084 L x [P(extract ! blank), mg/L] x 100% [tissue wt, mg]

% K in tissue:= [K(extract ! blank), mg/L] x 28mL x [9mL/3mL] x [1 L/1000 mL] x 100%

[tissue wt, mg]

= 0.084 L x [K(extract ! blank), mg/L x 100% [tissue wt, mg]


40

EXERCISE 9

TOTAL NITROGEN IN PLANT TISSUE

The procedure described below measures the organic nitrogen in plant tissue. It does not measure NO3--

N quantitatively. However, in most instances NO3--N is an insignificant fraction of the total N in plant

tissue. When NO3- is expected to be significant, addition of salicylic acid during tissue digestion results

in conversion of NO3- to NH4

+, which is measured.

The majority of the N detected by this method is found in proteins. Proteins contain about 16% N.Hence, the so-called crude protein content of plant material is commonly estimated by multiplying % Nby 6.25 (1 ) 0.16).

In the following procedure, hot, concentrated H2SO4 is used to oxidize the plant tissue and release theorganic N in the form of NH4

+. Selenium and copper are added as catalysts to Na2SO4 in a "digestionmix". The Na2SO4 raises the boiling point of the H2SO4. The net result is relatively rapid and completedegradation of the plant tissue. Even so, the digestion process requires 2 to 3 hours and is, therefore,completed by lab assistants prior to lab time.

Materials

Micro-Kjeldahl digestion apparatus, 100-mL digestion flasks, analytical balance, steam distillationapparatus, 50-mL conical flasks, burette, and the following reagents:

Digestion mix: Mix thoroughly 5 g Se metal (Toxic!) and 32 g anhydrous CuSO4 with 1,000 g ofanhydrous Na2SO4.

Mixed indicator: Dissolve 0.130 g of bromcresol green and 0.065 g of methyl red in 100 mL of ethanol.

Boric acid, 4%: Dissolve 40 g of H3BO3 in about 900 mL of deionized water, add 25 mL of mixedindicator solution containing 0.33 g bromcresol green and 0.165 g methyl red dissolved in 500 mL ofethanol, and dilute to 1 liter with deionized water.

Sulfuric acid, concentrated: If nitrates are to be included with "total N," dissolve 75 g of salicylic acid inone 2-liter bottle of H2SO4.

NaOH, 15 M: Dissolve 600 g of NaOH in deionized water and dilute, when cool, to 1 liter. (Technicalgrade NaOH is satisfactory.)

THAM, 0.050 M: Dissolve 6.057 g of tris-hydroxyamino methane (THAM), oven-dried at 105 °C, indeionized water and dilute to 1 liter.

Standard H2SO4 (0.0357 M): Dilute 2 mL of concentrated H2SO4 to 1 liter with deionized water. Mix welland standardize against 0.050 M tris-hydroxyamino methane (THAM). Calculate the amount of waterthat must be added to dilute the H2SO4 to exactly 0.0357 M. Add the required amount of water, andcheck the resulting concentration by titrating with the THAM solution. One mL of 0.0357 M H2SO4

contains 0.0174 mmol of H+ which will neutralize 1 mg of NH3-N.


41


Procedure for Determining Total Nitrogen in Plant Tissue

Procedure

1. Weigh 100 to 150 mg of ground plant tissueand transfer quantitatively to a dry digestion tube.

2. Add 2 g of digestion mix.

3. Add 5.0 mL of concentrated H2SO4 and allowto react for 30 minutes at room temperature.

4. Place on the digestion apparatus, and applylow heat until the initial reaction has subsided.

5. Increase the temperature to just below theboiling point of the H2SO4. Digest for 30 minutesafter the solution becomes clear.

6. Disconnect the heaters, and allow the flasksto cool until they can be held in the hand.

7. Slowly and carefully add 15 to 25 mL ofdeionized water with a wash bottle and allow tocool.

8. Add 5 mL of boric acid indicator solution to a50-mL conical flask, and place the flask under thecondenser of the steam distillation apparatus.

9. Slowly add 15 mL of 15 M NaOH down theside (inside) of the Kjeldahl flask, andimmediately connect the flask to the distillationapparatus.

Remarks 1. Record the weight to +/! 1 mg. The digestiontube should be dry so that the tissue will notadhere to the neck.

2. Use the scoop provided.

3. Do this in a fume hood and use gloves andeye protection! Do not get any H2SO4 on yourclothing.

4. This will take 30 to 60 minutes.

5. The solution will be colored because of theCu in the digestion mix, but it should not beturbid.

6. If the flasks cool down too far, the Na2SO4

will solidify, and getting it to re-dissolve may be aproblem.

7. In Chem 103 you were told "Never add waterto H2SO4; always add H2SO4 to water!" In theKjeldahl procedure, there is no suitablealternative to adding water to the H2SO4, so becareful! Point the neck of the flask into the hood.

8. The boric acid traps the NH3 in the distillateas NH4B(OH)4.

9. The object is to get the NaOH to sink to thebottom of the flask without mixing with the H2SO4

before the flask is connected to the still. The15 M NaOH is denser than the diluted acid.


42


Procedure

10. Turn on the steam and distill until 15 mL ofdistillate have been collected.

11. Titrate the distillate with 0.0357 M H2SO4 toa pink endpoint.

12. Repeat steps 2 through 11 without soil added(a blank).

13. Calculate % N in the tissue sample, and turnin your data sheet to the instructor.

Remarks

10. Heat will be generated when the steam mixesthe NaOH and H2SO4. If too much H2SO4 wasadded in step 3 and insufficient NaOH in step 9to neutralize the acid, the N will remain as NH4

+

instead of distilling over as NH3.

11. One mL of this acid = 1 mg of N because theH+ concentration is 1/14 M, and the millimolarweight of N is 14 mg/mmol.

(mmol x mg/mmol = mg)

12. This determines any contribution to the Nmeasurement from the reagents used.

13. See Calculations below.

Chemical Reactions Involved

NH3 + H3BO3 + H2O ===> NH4B(OH)4

NH4B(OH)4 + H+ ===> NH4+ + H3BO3 + H2O

Calculations

1 mmol of H2SO4 contains 2 mmol of H+, therefore:

(0.0357 mmol H2SO4/mL) x (2 mmol H+/mL H2SO4) = 0.0714 mmol H+/mL H2SO4

H+ (sample ! blank), mmol = mL H2SO4 (sample ! blank) x 0.0714 mmol H+/mL H2SO4

1 mmol H+ reacts with 1 mmol of NH4B(OH)4 which contains 1 mmol of N, therefore:

N in sample, mmol = H+ (sample ! blank), mmol

N in sample, mg = N in sample, mmol x 14 mg N/mmol N

(The H+ concentration of 0.0714 mmol/mL H2SO4 is used because 1 mL will react with0.0714 mmol of N; therefore, 1 mL of H2SO4 is the equivalent of 1 mg of N.)

0.0714 mmol N/mL H2SO4 x 14 mg N/mmol N = 1.000 mg N/mL H2SO4 .

% N in tissue = [mg N in sample] x 100% [mg of tissue]


43

EXERCISE 10

DETERMINATION OF AVAILABLE P AND K IN SOIL

Available phosphorus exists in soils as a constituent of both organic and inorganic compounds. Asorganic P undergoes mineralization, the released P begins to equilibrate with the inorganic fraction.Consequently, organic P has not been found to correlate very well with plant uptake of P and is largelyignored in testing for plant-available P in soil.

The rate of P uptake by plants appears to be a function of the solution P concentration at the rootsurface. Initially, this concentration will be that of the bulk soil solution. As uptake progresses, theconcentration near the root drops, and P will have to diffuse from increasingly greater distances. Ifuptake of P is rapid, as in greenhouse cropping experiments and under certain optimum field conditions,the release of P from the solid phase to solution will likely become the rate-limiting step.

Since the solution P concentration is a function of surface P, a test for surface P should also correlatewell with plant uptake. Although a direct determination of surface P by equilibration with radioactive Pis not feasible for routine analysis, various chemical extractants have been found to give highcorrelations with surface P. Most notable among these have been the Bray P-1 extractant (0.03 M NH4Fin 0.025 M HCl), the Mehlich extractant (0.125 M H2SO4 + 0.05 M HCl) and the Olsen extractant (0.5 MNaHCO3). With the Bray extractant, the F- displaces P bound to Al and some Fe surfaces, but it doesnot displace Ca-bound P to any great extent. The weak acidity of the extractant may dissolve some Ca-bound P; but if the soil is highly calcareous, the acid will be neutralized rapidly, liberated Ca2+ willprecipitate the F- as CaF2 and little P will be extracted.

With the NaHCO3 extractant, the HCO3- ion maintains a pH of about 8.5 and, apparently, OH- displaces

Fe- and Al-bound surface P and CO32- displaces Ca-bound P. This extractant is used by many western

states that have soils containing high amounts of free CaCO3. High concentrations of CaCO3 neutralizethe HCl in the Bray extractant and P extracted initially is reprecipitated as calcium phosphate.

Various acid extractants such as 0.3 M HCl, have also been used with success on acid soils. However,any calcium phosphates present will be attacked by these reagents so tests on soils containing calciumphosphate will indicate erroneously high P availabilities. The acid extractants will not give satisfactoryresults on alkaline soils (which usually contain calcium phosphates) nor on acid soils to which rockphosphate has been applied.

Because of the many complex factors involved in P availability to plants, any soil test for P must becalibrated differently for different soils. In the field, the problem is made even more difficult by variationsin the available P supply in the subsoil.

The extractant used in this exercise will be Bray P-1, the most widely-used P test in the U.S.


44


Available potassium. The extractant used most commonly for measuring available soil K is 1 MNH4OAc, which determines exchangeable plus water-soluble K. Other neutral salts may be used, butthe NH4

+ ion, being the same size as K+, can displace some K at the edges of weathered micas that isinaccessible to larger hydrated ions but is available to plants. This extractant does not measuremoderately available or "reserve" K, which in some soils may contribute appreciably to the available Ksupply to the crop. Attempts have been made to measure non-exchangeable available K but none hasbeen generally accepted for use in soil testing laboratories.

The NH4OAc procedure extracts some K that is not immediately available to plants. That is, when plantscan no longer grow in a soil depleted of K, there will still be some K extractable with NH4OAc. The BrayP-1 extractant has a total cation charge concentration of 0.055 M compared with 1 M for NH4OAc, butthe amount of K extracted with this reagent is about 85 to 90% of that extracted with NH4OAc. Moreimportant, however, is the fact that the amounts of K measured by the two extractants are very closelycorrelated.

Interpretation of soil tests for available K is complicated by the fact that drying the soil sample tends torelease K to exchangeable form in low-K soils and convert exchangeable K to a non-exchangeable formin high-K soils. Potassium tests on undried soils usually correlate better with plant uptake than tests ondried soils but handling undried soils in a routine testing laboratory is very difficult and is rarely used.

Reagents

Extracting solution (P-A) (0.03 M NH4F in 0.025 M HCl): Dissolve 1.11 g of NH4F in about 900 mL ofwater. Add 2.1 mL of concentrated HCl and dilute to 1 liter.

Ammonium molybdate soln. (P-B) [0.87 M HCl, 0.0033 M (NH4)6Mo7O24 * 4 H2O, 0.08 M H3BO3]:Dissolve 3.8 g of (NH4)6Mo7O24 * 4 H2O in 300 mL of water at 60 °C; cool. Dissolve 5.0 g H3BO3 in 500mL of water. Mix the two solutions, add 75 mL concentrated HCl (11.6 M), and dilute to 1 liter.

Reductant solution (P-C): Prepare a stock supply of reductant powder by mixing thoroughly andgrinding to a fine powder 2.5 g of 1-amino-napthol-4-sulfonic acid, 5.0 g of Na2SO3 (sodium sulfite),and 146 g of Na2S3O5 (sodium metabisulfite). Dissolve 8 g of the dry powder in 50 mL of warm water.Let stand overnight if possible. A fresh reagent should be made every three weeks. (Some materialmay crystallize upon standing, but this does not affect the performance of the reagent.)

Standard P and K solutions (250 mg/L P and 315 mg/L K): Dissolve 1.0985 g of oven-dried (105°C)KH2PO4 in about 900 mL of deionized water. Add 5 mL of concentrated HNO3 as a preservative, anddilute to 1 liter. Dilute 0, 1, 2, 4, 6, and 8 mL of this standard solution to 100 mL with water. Theresulting P concentrations will be 0, 2.5, 5, 10, 15, and 20 mg/L, respectively; the K concentrations willbe 0, 3.15, 6.3, 12.6, 18.9, and 25.2 mg/L.

Materials

1.5-g soil scoops, 50-mL extraction flasks, 3-mL pipettes, 15-mL dispenser, Whatman no. 2 filterpaper, funnel tubes, spectrophotometer tubes, oscillating shaker, spectrophotometer, flamephotometer.


45


Procedure for Available Phosphorus

Procedure

1. Weigh out 1.50 g of soil from each pot inExercise 1 and transfer each soil sample to a50-mL extraction flask labeled with the letter ofthe replicate from which the sample was taken.

2. Add 15 mL of Bray P-1 solution to thesample and to a "blank" flask.

3. Shake for 5 minutes.

4. Filter the suspension through Whatman no.2 or equivalent filter paper into a funnel tube.

5. Pipette a 3-mL aliquot of the filtrate into aclean spectrophotometer tube.

6. Add 3 mL of ammonium molybdate solution(P-B) into the same spectrophotometer tube withthe pipette provided.

7. Add 5 drops of reductant solution (P-C).

8. Mix the solutions.

9. Allow the solution to stand for 15 minutes,but read the color intensity before 45 min.

10. Read absorbance on the red scale of thespectrophotometer set a wavelength of 660 nm.Set a water blank at 0 absorbance.

11. Calculate P extracted in lbs/acre.

Remarks

1. Use a funnel to transfer the sample to theflask. Recall that you had three replicates inExercise 1. Make sure you know which samplecorresponds to each replicate because the soilanalysis results will be related to the yield ofcorn tissue from each pot.

2. The blank is used to account forcontamination in the procedure.

3. Equilibration is nearly complete in 5 minutes.

4. Collect about 10 mL of filtrate.

5. Save the remaining filtrate for K analysis.

6. Ammonium molybdate forms a complex withH2PO4

-.

7. A blue phosphomolybdate complex isformed in the presence of a reducing agent.

8. Shake the tube by hand without spilling thecontents.

9. The color does not develop fully for 15minutes and fades after 45.

10. Read absorbance for both soil and blanksamples.

11. See Calculations at the end of this exerciseand on the Data Sheet.


46


Procedure for Available Potassium

Procedure

1. Use the solution left in the funnel tube fordetermination of K after the P aliquot has beentaken.

2. Set the flame photometer to read 0 % T withwater.

3. Set the flame photometer to read 100 % Twith the 25.2 mg/L K solution.

4. Aspirate your filtrate from both soil andblank.

5. Use the appropriate standard curve todetermine the concentration of K in the extract.

6. Calculate available K in kg/ha and inlbs/acre. Give the Data Sheet to your instructorwhen completed.

Remarks

1. Available P and available K are determinedin the same extract.

2. The instructor will demonstrate operation ofthe flame photometer.

3. Be sure to use the appropriate K standardsolutions.

4. Read % T to +/! 0.5 % T.

5. The standard curve is plotted as % T vs Kconcentration in mg/L.

6. See Calculations below and on the DataSheet.

Calculations

The calculations are done in metric units and then converted to lbs/acre assum ing the depth of the plow layer to

be 0.2 m (= 20 cm or 7.9 inches).

Calculation of m illigrams of available P and K per kilogram of soil

Avail. P in soil, m g/kg soil = [P conc. (extract ! blank), mg/L] x 15 mL extract x 6 mL x 1 L x 1000 g

1.5 g soil 3 mL 1000 mL 1 kg

Avail. K in soil, mg/kg = [K conc. (extract ! blank ), mg/L] x 15mL extract x 1 L x 1000 g

1.5 g soil 1000 mL 1 kg

Calculation of kilograms of available P or K per hectare plow layer

To convert the available P or K determ ined in the laboratory from units of weight per unit weight (mg/kg) to weight

per unit volume (kg/ha plow layer), the bulk density of the soil is needed. Bulk density, DB, has units of weight per

unit volume. W hen calculating kg/ha, it is convenient to use megagram s per cubic meter, Mg/m 3 (which is

numerically equivalent to g/cm 3), as the units for DB. (1 Mg = 1000 kg; it is also called a tonne).

1 hectare = 10,000 m2. Therefore, a hectare plow layer = 0.2 m (depth) x 10,000 m2 (area) = 2000 m3.

The weight of this hectare plow layer, Mg/ha plow layer = DB, Mg/m3 x 2000 m3/ha plow layer.

Available P or K in soil, kg/ha plow layer:

= (Avail. P or K in soil, mg/kg) x (1kg/106 mg) x (DB, Mg/m3) x (1000 kg/Mg) x (2000 m3/ha plow layer)

Calculation of pounds of available P or K per acre plow layer

Since 1 kilogram = 2.205 pounds and 1 acre = 0.407 hectares, you can convert kilograms per hectare plow

layer to pounds per acre plow layer (assuming the same plow layer depth) as follows:

Available P or K in soil, lbs/acre plow layer:

= (Available P or K in soil, kg/ha plow layer) x (2.205 lbs/kg) x (0.407 ha plow layer/acre plow layer)


47

EXERCISE 11

DETERMINATION OF SOIL pH, LIME REQUIREMENT AND SOLUBLE SALTS

SOIL PH AND LIME REQUIREMENT

The pH of a soil affects many chemical and biological properties of that soil. It affects the availabilityof most of the elements essential for plant growth, the activity of microorganisms, and the cationexchange capacity. Lime recommendations are designed to raise the pH of an acid soil to the optimumpH for the particular crop to be grown1. Alfalfa, for example, grows best at pH 6.8 or above, whereascranberries do better at a pH around 4.5.

In Wisconsin, the lime requirement is calculated from the pH of the soil in water, the pH desired, thepH of the soil after reaction with SMP pH buffer2, weight loss on ignition3 (LOI), and depth of plowing4.Soil organic matter is estimated from LOI and, along with the SMP buffer test, estimates reserveacidity. Water pH represents active acidity. In most North-Central states, the SMP buffer test aloneis used to determine the lime requirement, but Wisconsin research has given better results byincluding organic matter. Empirical equations used to calculate the lime requirement are shown belowfor pH 6.0 and pH 6.8. Equations for other pH levels are given by Kelling et al.1

LR6.0 = [1.92 x (6.0 ! pHwater) x (LOI) + 0.077(pHSMP)] x [plow depth factor]

LR6.8 = [2.92 x (6.8 ! pHwater) x (LOI)] ! 0.092(pHSMP)] x [plow depth factor]

The calculated lime requirements, in tons per acre, are for aglime with a neutralizing index of 60-69.The neutralizing index depends on the neutralizing value and the particle size distribution of the limingmaterial .

Soil pH measurements are affected by the soluble salt content of the soil because the addition of acation to the soil solution will displace a small amount of H+ from the permanent charge sites (but notfrom pH-dependent sites):

HX + K+ <=====> KX + H+,

where X represents a negative charge site on a soil mineral. Consequently, the pH of a recentlyfertilized soil may be artificially depressed by as much as 0.5 pH unit. To circumvent this problem,some labs routinely measure soil pH in 0.01 M CaCl2 or 1 M KCl. Since a salt is added to the soil,previous fertilization will have little additional effect in lowering pH. Wisconsin soil pH values measuredin 0.01 M CaCl2 or 1 M KCl typically are about 0.4 and 1.0 pH unit lower, respectively, than whenmeasured in water. This relationship does not hold true for all soils. An increase in pH is evenpossible in some oxisols having a high anion exchange capacity.____________________1 Kelling, K.A., E.E. Schulte, L.G. Bundy, S.C. Combs, and J.B. Peters. 1991. UWEX Publ. A2809.

2 Shoemaker, R.K., E.O. McLean, and P.F. Pratt. 1961. Soil Sci. Soc. Am. Proc. 25:274-277.

3 Estimated organic m atter, % = 0.07 + [0.89 x LOI, %]

4 Plow depth, inches Plow depth factor

0 - 7.0 1.00

7.1 - 8.0 1.15

8.1 - 9.0 1.31

> 9.0 1.46


48


Procedure for Determining Soil pH and Lime Requirement

Materials

7.5-g soil scoop, plastic vials, 100-mL beakers, dispensers, stirring rods, glass electrode pH meter,oscillating shaker.

Reagents

0.01 M CaCl2: Dissolve 1.48 g of CaCl2 * 2 H2O in water and dilute to 1 liter.

SMP buffer: Dissolve 1.8 g p-nitrophenol and 3.0 g K2CrO4 in about 900 mL of deionized water. Add2.0 g of Ca(OAc)2 and 53.1 g of CaCl2 *

2 H2O. Stir until dissolved. Add 2.5 mL of triethanolamine andmix thoroughly. After all materials have dissolved, let stand overnight; then adjust the pH to 7.7 +/!0.02 with NaOH or HCl, using a glass electrode pH meter. Dilute to 1 liter with deionized water.

Procedure

1. Measure 7.5 g of designated soil samplesinto each of two 40-mL plastic vails.

2. Add 10 mL of water to the first vial.

3. Add 10 mL of 0.01 M CaCl2 to the secondvial.

4. Stir the samples with a glass rod; let stand15 min.

5. Read the pH of the first two samples with aglass electrode pH meter, stirring the samplejust before reading the pH.

6. Add 15 mL of SMP buffer to the first vialcontaining water.

7. Cap the vial and place in a horizontalposition on the oscillating shaker.

8. Shake the sample for 30 minutes; then readthe pH with a glass electrode pH meter.

9. Calculate the lime requirements for peas(pH 6.0) and alfalfa (pH 6.8).

Remarks

1. Use the calibrated scoops.

2. Use the dispenser. This vial is used for pH inwater and SMP pH.

3. Use the dispenser. This vial is used for pH in0.01 M CaCl2.

4. Some time is needed for the soil pH to reachequilibrium.

5. The instructor will standardize the pH meter.

6. Use the dispenser.

7. Shaking is much more efficient in thehorizontal position.

8. This is the SMP pH. Alternatively, shake thesample for 5 min, let stand 1 hr, then shakeanother 5 min.

9. Fill out the first part of the Data Sheet. Theequations are shown on the first page of thisexercise and on the Data Sheet.


49


SOLUBLE SALTS

All soil solutions and surface waters contain some soluble constituents as a result of geologicalweathering of rocks and minerals. In humid regions, these soluble constituents are carried by runoffand percolating waters to streams and oceans. In regions of high evapotranspiration and low rainfallor poor drainage, however, the soluble weathering products tend to accumulate. This accumulationis especially common on the 45% of the earth's land surface receiving less than 20 inches (500 mm)of rainfall annually.

The use of irrigation water containing soluble salts further increases the salt content of soils. In theabsence of leaching, water loss by evapotranspiration results in a net increase in soluble salts.Increasingly high rates of fertilizer application can lead to local salt accumulation problems, even inhumid areas. Salting of highways for ice removal sometimes results in saline soils adjacent to thesehighways.

The measurement of soluble salts has become as widely used in arid and semi-arid regions as hasthe pH measurement in humid areas. It serves as a rapid method for detection of toxic accumulationsof salt and is a valuable aid in soil classification. Frequent analysis of soluble salts aids in determiningwhen leaching is necessary and what the leaching requirement will be.

Ground and surface water vary considerably in soluble salt content. The suitability of water forirrigation is largely dependent on its salt content.

Methods for Measuring Soluble Salts

Gravimetric method. The total dissolved salt content of soil extracts and waters has been determinedfor many years. The procedure consists of evaporating a known volume of solution and weighing theresidue. The results are reported in terms of percent or ppm of salt. This method is useful but time-consuming and dependent on the method of extraction. It may over-estimate the soluble salt contentbecause the presence of moderately-soluble salts such as gypsum.

Resistance-conductance methods. Because pure water contains few ions in solution, it is a very poorconductor of electricity. The addition of ions, however, makes water a better conductor. Materials thatwill not transport an electric current are said to have a high resistance. The unit of resistance is ohm,and ordinary deionized water has a resistance of about 200,000 ohms. As the salt content of a solutionincreases, the resistance decreases.

Resistance methods were once used widely on saturated soil pastes, extracts, and solutions. Muchinformation in the literature prior to 1950 is given in units of resistance. More recently, the results ofsoluble salt measurements have been expressed in units of electrical conductance (EC). Theestimation of salt concentration by electrical conductance utilizes the same theory as resistancemethods, since one is the reciprocal of the other:

Conductance = 1/Resistance

One advantage of using conductance is that it is directly proportional to salt concentration rather thaninversely proportional, as are resistance units.


50


The currently accepted unit of electrical conductance is the Siemen, S, and conductivitymeasurements in soils are expressed as deci-siemens per meter, dS m-1. Accepted units for electricalconductivity have changed in recent years from metric to SI measurement systems. Consequently,all but the most modern tables and instruments are marked in the older system that use the unit,mmho cm,-1 instead of the numerically equivalent SI unit, dS m-1. (If not for the desire to maintainnumerical equivalency, the preferred SI unit would have been siemens per meter, S m-1.)

The conductance of a solution is determined by measuring the current flow between two platinumelectrodes connected to an alternating current voltage source when these electrodes are placed in thetest solution a given fixed distance apart. Hence, the units are given as conductance per unit length(dS m-1).

Extraction Methods

Many soil:water ratios have been used for the extraction of soluble salts. However, none have beenas widely accepted as the saturation extract. In most medium to fine textured soils, the moisturecontent of a saturated soil paste is about four times the amount present at the wilting point and twicethat at field moisture capacity. By using extracts obtained from saturated soils, conductance valuesof soils of different texture may be compared. Obtaining a sample of the saturation extract requiresa relatively large sample and vacuum extraction equipment. In the absence of vacuum extractionequipment, suspensions with extraction ratios of 1:1, 1:2, or 1:5 soil:water have been used. These areeasier to work with for routine analysis than the saturation extract.

The relationship between EC and crop growth is given in Table 11-1. The USDA Soil Salinity Labclassifies a soil as saline if the EC of a saturation extract exceeds 4 dS m-1 .

Table 11-1. Relationship between crop growth and salinity as measured by electrical conductivity.

Electrical conductivity of the saturation extract, dS m-1 at 25 oC

Non-saline Saline 0 2 4 8 16 32

Salinity Yields of very Yields of Only tolerant Only a feweffects mostly sensitive crops many crops crops yield very tolerantnegligible restricted restricted satisfactorily crops yield satisfactorily

Crops differ in their ability to withstand high salt concentrations. Table 11-2 groups crops accordingto their salt tolerance. Within any group, salt tolerance decreases from top to bottom. Spinach, forexample, is less salt-tolerant than garden beet.


51


Table 11-2. Salt-tolerance of crops grouped according to ranges in the electrical conductivityof the saturation extract of saline soil corresponding to a 50% decrement of yield on saline soilfrom that on non-saline soil.1

High salt tolerance Med. salt tolerance Low salt tolerance

Vegetable Crops

EC2 = 10 to 12 EC = 4 to 10 EC = 3 to 4Garden beet Tomato RadishKale Cabbage CeleryAsparagus Lettuce Green beansSpinach Potato

Cucumber

Forage Crops

EC = 12 to 18 EC = 4 to 12 EC = 2 to 4Salt grass White sweet clover White Dutch cloverBermuda grass Perennial ryegrass Meadow foxtailWestern wheat grass Sudan grass Alsike cloverBirdsfoot trefoil Alfalfa Red clover

Orchard grass Ladino cloverBrome grass

Field Crops

EC = 10 to 16 EC = 6 to 10 EC = 4Barley Wheat Field beansSugar beet OatsCanola RiceCotton Corn

_____________________________

1 from U.S.D.A. Handbook Nol 60. 1954. Diagnosis and Improvement of Saline and Alkali Soils.

2 EC = dS m-1.


52


Materials

10-g scoops, 100 mL beakers, dispensers, stirring rods, conductivity meter.

Reagents

0.010 M KCl: Dissolve 0.7456 g of oven-dried (105 °C) KCl in water and dilute to 1 liter. The electricalconductivity (EC) of this solution is 1.412 dS m-1 at 25 °C. It is used to determine the cell constant ofthe conductivity meter.

Procedure

1. Add two 10-g scoops (20 g) of designatedsoil to each of two 100-mL beakers.

2. Add 40 mL of water to the soil in each of thetwo beakers.

3. Stir the samples with a glass rod; let stand15 min.

4. Place the conductivity cell into the beaker of0.010 M KCl. Read the conductivity andcalculate the cell constant1.

5. Place the conductivity cell into the beakercontaining the soil suspension.

6. Record the conductance

7. Calculate the adjusted conductivity of thesample.2

8. Calculate the estimated soil water content atsaturation 3.

9. Calculate the estimated conductivity of thesaturation extract 4.

Remarks

1. One sample of the soil added to the 100-mLbeakers will have had salt added; the other willnot.

2. Use the burette provided. This sample is usedto measure EC.

3. Some time is needed for salts to dissolve.

4. The 0.010 M KCl has a known conductivity of1.412 dS m-1 at 25 °C. The cell constant equalsthe instrument reading divided by1.41.

5. Alternatively, the suspension can be pouredinto the conductivity cell if the cell is held uprightand the air vents are closed.

6. This is the measured conductivity.

7. Multiply the measured conductivity by thecell constant, kc.

8. In medium textured soils, saturation isapproximately 2 x FMC.

9. Fill out the Data Sheet and hand it to theinstructor.

1 kc = [Conductivity of 0.010 M KCl, dS m-1] ) [Measured conductivity of 0.010 M KCl, dS m-1] = [1.41 dS m-1] ) [Measured conductivity of 0.010 M KCl, dS m-1]

2 Adjusted conductivity of sample, dS m-1 = [Measured conductivity of sample, dS m-1] x kc

3 Estimated soil water content at saturation, % = 2 x FMC %

4 Estimated conductivity of saturation extract, dS m-1 = (Adj. cond., dS m-1) x 200 % ) (2 x FMC %) where 200% is the % water (dry soil basis) in a suspension with a 2:1 water:soil ratio.


53

EXERCISE 12

DEVELOPING NUTRIENT DEFICIENCY SYMPTOMS IN PLANTS GROWING IN SOLUTION CULTURE

Applications of the techniques of culturing plants in nutrient solutions range from commercialproduction to investigation of the mechanisms of active ion accumulation by plants and plant tissue.On the commercial scale, solution culture is frequently referred to as soil-less culture or hydroponics,and growers have developed numerous variations of the basic technique. Because of the expenseinvolved, commercial application is normally restricted to high value crops and environments in whichcrops cannot be grown by conventional methods.

On the research side, applications of solution culture techniques are numerous. Nutrient solutions canbe rendered extremely pure (< 10 mg/kg of contaminants) and are, therefore, ideally suited forstudying the essentiality of nutrients and the concentrations of elements in plants as a function of drymatter yield. By employing large solution volumes and providing for a continuous agitation of thesolution, the researcher is able to maintain virtually constant ion concentrations and eliminate diffusionto root surfaces as the rate limiting step in absorption processes. Under these conditions, observedvariations in ion accumulation by plants and their tissues can be ascribed to the mechanisms of ionabsorption and transport by and within plant cells and tissues.

Although solution techniques can be employed productively in many types of research, we must keepin mind the artificiality of the system. Results of solution culture studies cannot be extrapolated tosituations where soil serves as the plant growth medium and little or no control is exerted on the plant'senvironment.

The solution culture outlined here is based on Hoagland's solution, which over the years hasundergone several modifications. The nutrient solution consists of essential major and minor elementsin concentrations and ratios that prevent toxicity, maintain normal growth, and induce clear deficiencysymptoms. Inclusion of Fe in chelate form proved essential for preventing Fe chlorosis, which is verydifficult to control in the early stages of plant growth in solution culture. Aeration is critical, especiallyafter the plants are two to three weeks old because oxygen diffusion through the water is too slow tosupport optimum root respiration in a large root system. Maintaining a well aerated system, however,tends to precipitate Fe(OH)3, making if difficult to maintain adequate Fe in solution.

The purpose of this exercise is to introduce the technique of solution culture of plants and to allow youto view first-hand the deficiency symptoms of the essential mineral elements, N, P, K, Ca, Mg, S, Fe,Cu, Zn, B, Mo and Mn.

Materials

Darkened 5-liter containers, aeration system, deionized water, stock nutrient solutions (Table 12-1),50-ml graduated cylinders , pot labels, seedlings. (The seedlings may be started in sand culture orby using germination paper. This must be planned well in advance of the time needed.)


54


Procedure

1. Select a darkened 5-liter polyethylenecontainer. If a polyethylene container is notavailable, line a 5-liter pot with a clean plasticbag.

2. Add approximately 3 liters of deionizedwater. (Ordinary deionized water is satisfactoryfor !N, !P, !K, !Ca, !Mg, and !Fe treatments.)

3. Add the stock solutions listed in the columnunder the treatment for the element of interestas shown in Table 12-2.

4. Add deionized water to the 5-liter mark onthe container and place the lid on the container.

5. Carefully remove one corn seedling fromthe sand culture or germination paper, and insertit through a hole in the lid.

6. Gently wrap the stem with sufficient cottonbatting to hold the seedling securely.

7. Repeat steps 5 and 6 with the other plantsto be used.

8. Position the lower end of the aeration tubeso that it rests in the center of the bottom of thecontainer. (See figure below.)

9. Label the container with the symbol of theomitted nutrient.

10. Add water twice weekly to bring the level ofthe nutrient solution to the 5-liter mark on thecontainer.

11. Replace the nutrient solution after twoweeks.

12. Note the deficiency symptoms each weekas they develop.

Remarks

1. Light must be excluded to prevent algalgrowth. The plastic bag excludes contaminantswhich are present in most containers.

2. In this manner the stock solutions are dilutedas they are added, thereby avoiding precipitationof relatively insoluble compounds such asCaHPO4.

3. Take the pot to the stock solutions, not thesolutions to the pot. The graduated cylindersmust remain with their respective stocksolutions; otherwise, cross-contamination willoccur.

4. The lid should fit over the top of thecontainer completely and have three 1-inchholes for seedlings and one 3/8-inch hole for theaeration tube.

5. Be careful not to damage the seedling'sleaves or root system.

6. The roots should extend at least two inchesbelow the lid and into the solution.

7. The use of more than one species willprovide some indication of inter-species variationin nutrient deficiency symptoms.

8. This ensures adequate aeration for all plantsin the pot. When all treatments are completed,adjust the air flow rate.

9. E.g., “! N”.

10. Add water more frequently if necessary.

11. The nutrients in solution become depletedwith time, and there is no solid phase (mineral ororganic) to replenish them.

12. No report is required; however, you may beasked to describe some of the deficiencysymptoms on the final examination.


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Table 12-1. Stock nutrient solutions.

Formula Nutrient Stock1 Nutrient Nutrient Nutrient Salt weight content solution concentratio solution concentration g % g/L g/L mg/L mL stock/L mg/L

NH4NO3 80.04 35.0 N 35.7 12,500 N 5 62.5 N

KCl 74.55 52.4 K 28.6 15,000 K 5 75.0 K47.6 Cl 13,700 Cl 16.5 Cl

Ca(NO3)2 * 4 H2O 236.16 17.0 Ca 59.0 10,000 Ca 5 50.0 Ca11.9 N 7,000 N 35.0 N

MgSO4 * 7 H2O 246.50 9.9 Mg 49.3 4,900 Mg 5 24.5 Mg 13.0 S 6,400 S 32.0 S

KH2PO4 136.09 28.7 K 13.6 3,900 K 5 19.5 K 22.8 P 3,100 P 15.5 P

MgCl2 * 6 H2O 203.30 12.0 Mg 41.0 4,900 Mg 5 24.5 Mg34.9 Cl 14,300 Cl 71.5 Cl

Ca(H2PO4)2 * 2 H2O 252.07 15.9 Ca 12.5 2,000 Ca 5 10.0 Ca 24.6 P 3,100 P 15.5 P

Na2SO4 * 10 H2O 142.04 32.4 Na 28.4 9,200 Na 5 46.0 Na 22.6 S 6,400 S 32.0 S

CaCl2 * 2 H2O 147.02 27.3 Ca 36.8 10,000 Ca 5 50.0 Ca48.2 Cl 17,800 Cl 89.0 Cl

MnCl2 * 4 H2O 197.91 27.8 Mn 0.2715 75 Mn 5 0.38 Mn35.8 Cl 97 Cl 0.49 Cl

CuCl2 * 2 H2O 170.48 37.3 Cu 0.0161 6 Cu 5 0.03 Cu52.0 Cl 8 Cl 0.04 Cl

ZnCl2 (95 %) 136.29 45.6 Zn 0.066 30 Zn 5 0.15 Zn49.4 Cl 32 Cl 0.16 Cl

H3BO3 61.83 17.5 B 0.169 30 B 5 0.15 B

H2MoO4 * H2O 179.97 53.3 Mo 0.0019 1 Mo 5 0.005 Mo

__________________________

1 The stock iron solution consists of 5 mM chelating agent in acid form (HEDTA, DTPA, EDDHA, orHBED), 1.35 g FeCl3*6H2O (5 mM), and 0.60 g NaOH (15 mM), prepared by adding the NaOH to thechelating agent in 500 mL deionized water and stirring until dissolved and then slowing adding whilestirring the FeCl3 dissolved in 250 mL water. Dilute to 1 liter and store in an opaque container. (Useof EDTA or citric acid as a chelating agent for hydroponics is discouraged because of the instabilityof the iron chelate.)

.


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Table 12-2. Preparation of dilute nutrient solutions from stock solutions

Stock Com- !N !P !K !Ca !Mg !S !Fe !Mn !Cu !Zn !B !Mo Solution plete ------------------mL stock solution per 5 liters of dilute nutrient solutions-------------------

NH4NO3 25 10* 25 25 37.5 25 25 25 25 25 25 25 25

KCl 25 25 32.5 10* 25 25 25 25 25 25 25 25 25

Ca(NO3)2 25 10* 25 25 10* 25 25 25 25 25 25 25 25

MgSO4 25 25 25 25 25 5* 10* 25 25 25 25 25 25

KH2PO4 10 10 5* ---- 10 10 10 10 10 10 10 10 10

MgCl2 ---- ---- ---- ---- ---- ---- 25 ---- ---- ---- ---- ---- ----

Ca(H2PO4)2 ---- ---- ---- 10 ---- ---- ---- ---- ---- ---- ---- ---- ----

Na2SO4 ---- ---- ---- ---- ---- 25 ---- ---- ---- ---- ---- ---- ----

CaCl2 ---- 25 ---- ---- ---- ---- ---- ---- ---- ---- ---- ---- ----

MnCl2 25 25 25 25 25 25 25 25 ---- 25 25 25 25

CuCl2 25 25 25 25 25 25 25 25 25 ---- 25 25 25

ZnCl2 25 25 25 25 25 25 25 25 25 25 ---- 25 25

H3BO3 25 25 25 25 25 25 25 25 25 25 25 ---- 25

H2MoO4 25 25 25 25 25 25 25 25 25 25 25 25 ----

Fe Chelate 25 25 25 25 25 25 25 5* 25 25 25 25 25

* Omit when nutrient solution is changed. If these elements are omitted in the beginning, severedeficiency symptoms develop very quickly and they are not like those encountered more commonlyin the field.

Figure 12-1. Aeration Device for Solution Cultures.

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Soil Science/Agronomy/Horticulture 326 Manual... · final soil depth is approximately equal to that in the greenhouse pot. Water is then added quickly to the soil surface to get complete