TITLE: USDA LTAR Common Experiment measurement: Concentration of phosphorus, potassium, and sulfur in aboveground biomass
AUTHORS: John L. Kovar, Ann Marie Fortuna

[DESCRIPTION]
An adequate supply of plant nutrients is an important part of sustainable crop production and the overall health of a crop. Knowing the amount of each nutrient removed in the harvested biomass (grain, forage, etc.) is the first step in calculating plant nutrients required. When the removed nutrients are not replaced, soil fertility declines over time, leading to a loss of yield potential. Phosphorus (P) is a primary macronutrient, with total P concentration in crops generally varying from 0.1 to 0.5%. Phosphorus can be found in roots, stems, and leaves, but most P is in seeds and fruit. It is a component of phytin, a major storage form of P in seeds and tubers. Potassium (K) is a primary macronutrient stored in the stems and foliage of aboveground biomass. Seeds contain only small amounts of K. Potassium is mobile in plant tissues and can easily leach. Therefore, estimates of plant biomass removed and remaining in the field are valuable for determining soil K fertility. Sulfur (S) is a secondary macronutrient in seeds and plant tissues. Nodule formation in legume crops requires S. Research has shown that S fertilization may improve nitrogen (N)-use efficiency in many crops by increasing N uptake. Determine P, K, and S concentrations in the biomass by first digesting the material. The nutrient concentrations in the digestate can then be determined using inductively coupled plasma-optical emission spectrometry (ICP-OES).

[STEPS]
SECTION: Sample collection
1. In most cases, biomass samples will be collected at each harvest, potentially resulting in multiple samples per year.
SECTION: Sample collection
2. For the common experiment, it is important to collect samples from BAU and ASP treatments when plants are at the same phenological (developmental) stage.
SECTION: Sample collection
3. This process may require multiple samplings if crops significantly differ in developmental stage between or among the treatments.
SECTION: Sample processing
4. Dry the biomass samples in a forced-air oven at 60 °C until a constant weight is reached (48 to 72 hours).
SECTION: Sample processing
5. Grind a subsample to pass a 2-mm screen, using a Wiley or similar mill.

SECTION: Sample processing
6. Grind a 2-mm subsample to pass a 1-mm screen, using a Udy cyclone or similar mill.

SECTION: Sample processing
7. If samples are stored for an extended period, redry before analysis.
SECTION: Sample analysis
8. Send samples to a commercial laboratory or analyze them at an ARS or University laboratory if
equipment and personnel are available. 
SECTION: Sample analysis
9. Before the concentration of nutrients in biomass can be determined, it is first necessary to
destroy the organic matrix and solubilize the nutrients of interest.
SECTION: Sample analysis
10. Several methods are available for this purpose, the most common of which are :
i) addition of concentrated acid to the sample followed by heating
ii) addition of concentrated acid to the sample in a sealed vessel followed by microwave heating (Bryson and Mills, 2014; Havlin and Soltanpour, 1980).

SECTION: (i) Nitric and hydrochloric acid digestion with peroxide, open vessel
11. This method involves the acid dissolution of the organic matrix by digestion via external heating (hot plate, digestion block, or automated digestion block system), with the subsequent quantitative determination of P, K, and S in the digestate.
SECTION: (i) Nitric and hydrochloric acid digestion with peroxide, open vessel
12. Pre-digest a representative sub-sample containing up to 0.5 g of dry matter in nitric acid overnight.

SECTION: (i) Nitric and hydrochloric acid digestion with peroxide, open vessel
13. Digest the sample in heated nitric acid and hydrogen peroxide.
SECTION: (i) Nitric and hydrochloric acid digestion with peroxide, open vessel
14. After cooling, filter the vessel contents, centrifuge or allow to settle, dilute with nitric or hydrochloric acid to volume, and analyze by ICP-OES.
SECTION: (ii) Microwave-assisted digestion, closed vessel
15. This method involves the dissolution of the organic matrix in a closed vessel by nitric acid digestion via microwave heating for the subsequent quantitative determination of P, K, and S.
SECTION: (ii) Microwave-assisted digestion, closed vessel
16. Digest a representative sub-sample in concentrated nitric acid using a suitable laboratory microwave unit.
SECTION: (ii) Microwave-assisted digestion, closed vessel
17. Cap and heat a fluorocarbon (PFA or TFM) microwave vessel containing the sample and acid in the microwave unit.
SECTION: (ii) Microwave-assisted digestion, closed vessel
18. After cooling, filter the vessel contents, centrifuge or allow to settle, dilute to volume, and analyze by ICP-OES.
SECTION: Calculations and sufficiency ranges
19. Report P, K, and S concentrations on a dry-matter basis. 
SECTION: Calculations and sufficiency ranges
20. Calculate the amount of P, K, or S leaving the field by multiplying the biomass concentration by the amount of biomass removed (g nutrient/kg dry matter × kg dry matter/ha).
SECTION: Quality assessment and quality control
22. Recommend a standard operating procedure for your laboratory for measuring P, K, and S, with
the following items included:
SECTION: Quality assessment and quality control
22.1. Perform instrument calibration and maintenance according to the manufacturer’s instructions and log the results.
SECTION: Quality assessment and quality control
22.2. Prepare a standard calibration curve on the day of analysis with a minimum of six (6) concentrations, including a blank. Run calibration check standards throughout the assay every 20 samples to verify calibration accuracy and adjust for any drift. Values should be within 10% of the known value.
SECTION: Quality assessment and quality control
22.3. Run a certified reference standard (e.g., National Institute of Standards and Technology (NIST)) with each analytical sequence to verify calibration and method accuracy. Certified values should fall between 90% and 110% recovery. The results should not vary more than three standard deviations from the accepted mean value.
SECTION: Quality assessment and quality control
22.4. Determine method detection limits (MDL) based on the open-vessel nitric/hydrochloric acid or the closed-vessel nitric acid digestion matrices.
SECTION: Sample analysis
8.1. When choosing a commercial lab, consider the following factors : 
(1) whether the laboratory participates in a proficiency testing program, e.g., NAPT or ALP
(2) whether the laboratory returns samples for archiving purposes. 

 

SECTION: Illustrative information
23.  
 
 
 
 
 
 
SECTION: Recommendations for data collection
24. Table 1. Summary of recommendations for measurement of aboveground biomass P, K, and S concentrations.
 
 

SECTION: Calculations and sufficiency ranges
21. When reviewing P, K, and S data, it may be helpful to review sufficiency ranges for row crops, fruits, and vegetables. The Bryson and Mills (2014) reference contains tables for macro- and micro-nutrients although these will vary by environment. Utilize local publications for greater accuracy.