Exercise 15: Determination of Available Zinc in Soils

Deficiencies of many of the micronutrients have been recognized in the Midwest. Although these deficiencies affect only a small percentage of the total cropped area, significant yield reductions can result when any one of the micronutrients is present in inadequate amounts. “Minor elements,” as the micronutrients are sometimes called, is not a good term because the term suggests minor importance, which certainly is incorrect. These nutrients are just as important as the macronutrients for proper plant function. Some of the micronutrients are abundant in soils but used by plants in only small amounts. The micronutrients include boron, chlorine, copper, iron, manganese, molybdenum, nickel, and zinc, and tests have been developed to indicate the soil sufficiency of each of these elements.

Most of the work with micronutrient testing has been conducted since the 1950s. The reasons for the apparent increase in micronutrient testing include: 1) use of higher analysis fertilizers containing fewer micronutrients, 2) less widespread use of animal wastes, 3) increased crop yields requiring larger amounts of the elements, 4) aging and erosion of soils with long-term cultivation, 5) changes in air-purity standards, and 6) application of high rates of sewage sludge (municipal biosolids) as fertilizing materials. Furthermore, more attention is being paid to crop quality and the nutritional value of micronutrients in crop quality.

Many of the micronutrients are important components of enzyme systems that serve to catalyze biological reactions. Copper and iron play dominant roles in the energy transformations of plants. Iron exerts a strong influence on chlorophyll production, without which photosynthesis would not be possible. Zinc is necessary for the production of the amino acid tryptophan and the eventual formation of growth regulators. Molybdenum is essential for the utilization of nitrogen and the incorporation of nitrogen into protein. Boron aids in sugar transformations and affects water retention by certain plant parts. Chlorine has an important role in the chemical processes of photosynthesis. Nickel, the most recently recognized micronutrient, has an important role in the enzyme urease.

Many factors can serve to identify micronutrient deficiencies. A good way to confirm a suspected nutrient deficiency is to obtain separate samples of both plant and soil from areas of adjacent normal and abnormal plant growth and have the samples tested. Remember that micronutrient requirements vary from crop to crop. Some crops are sensitive to a given nutrient and will show visual deficiency symptoms when growing in a certain soil, whereas other crops growing on the same soil will not. For example, soybeans grown on calcareous soils in north-central Iowa may show iron chlorosis but corn growing on the same soil likely will not show iron deficiency symptoms.

Many extraction procedures have been tried for the various micronutrients. In some cases, interpretation of the results must be supplemented with other information such as pH, texture, and free-lime content. Two common extractants are used in the Midwest for zinc: 0.1 N HCl and a chelating agent, diethylenetriaminepentaacetic acid (DTPA).

Often interpretation data for micronutrients is much more limited than for macronutrients. Iowa State University recommends only zinc (Zn) for corn and sorghum based on soil testing. Soil test procedures for the other micronutrients have not been calibrated because of either limited occurrence of deficiencies or lack of consistency in the occurrence of deficiencies. The one exception is iron deficiency on soybeans, which has been well documented on high pH soils. Adding soluble forms of iron to these soils gives only short-term benefit. Development of soybean varieties tolerant to low iron availability has been an acceptable solution to the problem.

The extraction of zinc with 0.1 N HCl assumes that all or part of the zinc in soil is acid soluble and will become available to growing plants during a season. This then serves as the index for determining zinc availability. This method is used primarily in neutral and acid soils and is not recommended for calcareous soils.

The test used in Iowa is the extraction of zinc with 0.005 M DTPA in 0.01 M CaCl₂ and 0.1 M triethanolamine (TEA) solution. It has gained wide acceptance because of good results on calcareous soils and has the potential for use to extract iron, manganese, copper, and nickel. This test also can be used to monitor the heavy metals cadmium and lead in soils. The DTPA chelates or binds the metal cation and an equilibrium is established between the extracting solution and the amount of micronutrient in the soil. The pH of the extractant is set at pH7.3 to minimize the dissolution of CaCO₃ from calcareous soils.

With either extracting procedure, the concentration of zinc in solution is most commonly measured by using the atomic absorption spectrophotometer. The extract is passed into a flame in the burning apparatus. A characteristic wavelength of light from a source lamp is passed through the flame and, as the ion, in this case zinc, is excited and the outer electron jumps into a higher orbital, the characteristic wavelength of light is absorbed. The more ions in solution, the greater the absorption of light. The instrument measures the amount of absorption, and can be programmed to convert absorbance units to concentration in parts per million in solution on the output display.

 

*Recommendation for amount to apply in band is based on other states’ information.

 

Procedure

1. Weigh 10.00 ± 0.20 g of assigned soil (< 2.0 mm) into a dry 50-mL centrifuge tube in duplicate. Record the exact weight of soil to be extracted.

Glass or plastic wares should be cleaned by immersing in a 1 N HCl bath, and thoroughly rinsed with deionized water. Soil testing for the micronutrients is particularly difficult because plant requirements for these elements are small, posing the problem of easy contamination of samples unless adequate precautions are taken.

2. Add 20.0 ± 0.1 mL of DTPA extracting solution to each tube.

This gives a 1:2 soil-to-solution ratio. The DTPA is a chelating agent that establishes a chemical equilibrium of the metal ion in the soil, forming a soluble complex.

3. Cap the tube tightly, and set in a shaker horizontally for a shaking period of 1 hr.

Standard laboratory procedures require continuously shaking for 2 hr. The shaking time of 1 hr should give relative but not absolute Zn concentrations. Therefore, we will modify the shaking time to fit the laboratory time period.

4. Centrifuge @ 4000 rpm for 15 min and then pour off the supernatant through a plastic funnel lined with filter paper (Whatman No. 42 or equivalent) into a dry acid-washed 50-mL Erlenmeyer flask.

Refilter if the extract is turbid. The solution must be free of soil particles before it is introduced to the atomic absorption spectrophotometer. Those samples high in extractable Fe will have a yellow color that should not interfere with the Zn determination.

5. Analyze the sample filtrate by atomic absorption spectroscopy (AAS) together with standards ranging from 0.05 to 4.0 µg Zn mL^-1 of DTPA solution. The instructor will provide the specifics on standard preparation and will set the AAS to operate at 213.9 nm, a slit width of 0.7 nm, and an amp setting of 25 (may vary with different lamps). If the sample filtrate is > 4 ppm Zn, dilute the sample with guidance from the instructor.

Atomic absorption relies on the fact that the intensity of a specific wavelength of light from a standard source decreases as the amount of Zn in solution increases during burning. (Have the instructor show you the source lamp.)

6. Calculate the amount of available Zn in the soil; use Table 6 to determine soil test category.

Subtract the absorbance obtained in the blank solution from the absorbance of the soil solution. Determine the concentration of Zn in solution (μg/mL) using the regression line obtained from the standard calibration. Then multiply the μg/mL by the solution/soil factor (mL extractant/g soil analyzed) to obtain Zn concentration in μg/g soil:

Zn concentration (μg/g soil) = Zn in soln (μg/mL) x dilution factor (if any) x (20.0 mL/weight soil extracted in g). Note: micrograms per gram (μg/g) equals part per million (ppm) on a wt:wt basis.

 

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