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ARS Home » Midwest Area » Ames, Iowa » National Laboratory for Agriculture and The Environment » Agroecosystems Management Research » Research » Publications at this Location » Publication #333794
Title: Malachite green method for determining Phosphorus concentration in diverse matrices

Author
item RAHUTOMO, SUROSO - Iowa State University
item Kovar, John
item THOMPSON, MICHAEL - Iowa State University

Submitted to: Communications in Soil Science and Plant Analysis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/19/2019
Publication Date: 6/27/2019
Citation: Rahutomo, S., Kovar, J.L., Thompson, M.L. 2019. Malachite green method for determining Phosphorus concentration in diverse matrices. Communications in Soil Science and Plant Analysis. https://doi.org/10.1080/00103624.2019.1635140.
DOI: https://doi.org/10.1080/00103624.2019.1635140

Interpretive Summary: Phosphorus (P) exists in many forms in soils, sediments, and waters. Various extraction procedures have been developed to assess the distribution of P among the different fractions. Soil is initially extracted with a “weak” extracting solution in the first step followed by “stronger” extracting solutions in subsequent steps. The amounts of P extracted can then be used to predict biological availability or the potential for environmental degradation. Depending on the type of extractant, the traditional ways of quantifying P in extracts are either the molybdate blue-ascorbic acid (AA) colorimetric method or via inductively coupled plasma atomic emission spectrometry (ICP-AES). If several P forms are to be characterized in one sample, the separate analytical methods require additional time and labor. Thus, two comparison studies were conducted to investigate the use of the malachite green (MG) colorimetric method for determining P concentrations in diverse extracting solutions. The first experiment compared the MG method to the AA method to determine P extracted during fractionation analysis, while the second compared the MG method to ICP-AES for determining P in oxalate extracts. For the MG method, a 96-well standard micro-plate was used, and absorbance was read on universal micro plate reader. The results showed that the MG and AA methods agreed for determining P concentration in the extracts of water, hydrochloric acid, and concentrated sulfuric acid. A slight discrepancy between the two methods was found for extracts of sodium bicarbonate and sodium hydroxide. Differences among the sediments likely led to the discrepancy between the two colorimetric methods. In the determination of oxalate-extractable P, the MG method produced significantly higher values for P concentration than those obtained by ICP-AES analysis. This discrepancy may have been related to analytical issues with the ICP-AES method itself, rather than the extraction procedure. Despite the discrepancy for some extracts, the micro-plate system-MG method could be valuable because of its simplicity, sensitivity, and rapidity compared to the AA or ICP-AES methods.

Technical Abstract: Phosphorus (P) fractionation and extraction with ammonium oxalate are two methods commonly used to assess P dynamics in riparian soils and stream sediments. The traditional way of quantifying P in fractionation extracts is the molybdate blue-ascorbic acid (AA) colorimetric method. Ammonium oxalate-extractable P (Pox) often is determined via inductively coupled plasma atomic emission spectrometry (ICP-AES). Separate analytical methods require additional time and labor. Thus, two comparison studies were conducted to investigate the use of the malachite green (MG) colorimetric method for determining P concentrations in diverse matrices. The first experiment compared the MG method to the AA method to assess the sequential extraction of P during fractionation analysis, while the second compared the MG method to ICP-AES for determining Pox. For the MG method, a 96-well standard micro-plate was used, and absorbance was read on universal micro plate reader. The results showed that the MG and AA methods agreed for determining P concentration in the extracts of water, 1 M HCl, and concentrated H2SO4. A slight discrepancy between the two methods was found for extracts of 0.5 M NaHCO3 and 0.1 M NaOH, and concentrated HCl. Differences among the sediment materials subjected to fractionation analysis likely led to the discrepancy between the two colorimetric methods. In the determination of Pox, the MG method resulted in significantly higher values for P concentration than those obtained by ICP-AES analysis. This discrepancy may have been related to analytical issues with the ICP-AES method, including matrix interference, selection of the proper analytical wavelength, and the different densities of the standards and samples. Despite the discrepancy for some extracts in the sequential extraction and greater Pox values, the microplate system-MG method could be valuable because of its simplicity, sensitivity, and rapidity compared to the AA or ICP-AES methods.