1999 Progress Report: Dependence of Metal Ion Bioavailability on Biogenic Ligands and Soil Humic Substances

EPA Grant Number: R825960
Title: Dependence of Metal Ion Bioavailability on Biogenic Ligands and Soil Humic Substances
Investigators: Higashi, Richard M. , Fan, Teresa W-M. , Lane, Andrew N.
Institution: University of California - Davis
EPA Project Officer: Lasat, Mitch
Project Period: January 1, 1998 through December 31, 2001
Project Period Covered by this Report: January 1, 1998 through December 31, 1999
Project Amount: $345,816
RFA: EPA/DOE/NSF/ONR - Joint Program On Bioremediation (1997) RFA Text |  Recipients Lists
Research Category: Hazardous Waste/Remediation , Land and Waste Management


Organic matter can strongly affect metal ion binding to soil and sediment. In fact, production of a major form of organic matter-low-molecular weight organic ligands-is the principal mechanism by which plants and microbes acquire metal ions, so that the chemistry of biogenic organic matter is the key to understanding mechanisms of bioavailability for bioremediation purposes.

Thus, the complex interaction between metal ions, biogenic ligands, and humic substances must be understood to engineer the proper organisms and conditions for bioremediation of metal ion contamination. We propose to investigate this critically lacking area of knowledge through the following objectives: (1) determine the sorption behavior of metal ions on isolated humic substances in the presence of biogenic and synthetic ligands; (2) conduct a subset of experiments from Objective 1 as longer term aging experiments; (3) investigate the properties of isolated humic substances that are involved in Objectives 1 and 2; (4) assess the relationship of Objectives 1 and 2 to metal ion bioavailability to vascular plants, including evaluation of soils from a Federal demonstration site (McClellan AFB); and (5) use the findings from Objectives 1 through 4 to identify key rhizospheric processes that regulate metal bioavailability, additionally incorporating findings from a complementary project on the biochemistry of metal ion ligands in plants/mycorrhizal systems.

Progress Summary:

Studies of bioavailability of soil pollutants such as Cd(II) to plants necessarily involves a complex mixture of other metals (nutrients), inorganic anions, and a suite of biogenic ligands such as organic acids. A major organic phase involved in both organic and metal pollutant retention is humic substances (HS). Understanding the chemistry imposed by these complex mixtures is crucial, and therefore is a major focus of our effort.

Using pyrolysis-GC/MS and 2-D NMR (NOESY), we have uncovered kinetic and structural interactions of certain biogenic ligands, Cd(II), and a soil HS that limit the formation of ligand-Cd complexes that might affect bioavailability of metal ions to plants (Higashi, et al., 1998). Results from further studies eventually led to our use of photon-correlation spectroscopy (PCS) and excitation-emission matrix fluorescence (EEM fluorescence) techniques. The PCS revealed the formation, as well as shifts in size ranges, of surprisingly large colloidal aggregates (up to 1,000 nm in size) upon addition of Cd(II), which can drastically alter the availability of metal chelating groups (Higashi, et al., 1999). This dynamic association/aggregation was not unexpected, but the strong response of aggregation, large size ranges, and the number of size ranges of aggregate colloids could not have been predicted prior to the measurement. Concomitant with the size changes were dramatic changes in major peaks, revealed through EEM fluorescence. Some of these peaks changed over 25-fold in intensity upon the addition of Cd(II), some going up, and others going down. We are currently pursuing the identity of these peaks.

Generally, the key to these phenomena lies in the chemical structures of HS, and how they are organized three-dimensionally. We recently published in Environmental Science and Technology (Fan, et al., 2000) a comprehensive study of HS structure from forest soil used for bioavailability studies, which is summarized here. The soil was extracted for humic substances (HS) that was then demetallated with 4,5-dihydroxy-1,3-benzene disulphonate ("Tiron"). This yielded HS that was readily soluble in water at neutral pH, with a persistent inorganic composition of at about 25 percent, as determined by X-ray fluorescence and consistent with FTIR analysis. The high solubility at neutral pH greatly facilitated studies by solution-state 1-D and 2-D 1H , 13C, 31P, and 13C-1H NMR, which was supported by FTIR and pyrolysis-GC/MS analyses. Carbohydrates containing pyranoses, hydroxyphenylpropyl structures, phosphate mono/diesters, plus phosphatidic acid esters were evident. The amino acids Gly, Ala, Leu, Ile, Val, Asp, Ser, Thr, Glu, and Pro were identified in this HS as peptidic from scalar coupling in one 2-D NMR technique (TOCSY) and dipolar interactions in another technique (NOESY), then confirmed by acid digestion of HS followed by 2-D 1H NMR and GC-MS analysis. The above techniques, together with a third 2-D NMR method (HSQC), sketched a mobile nature for the peptidic side groups, but relatively rigid for the aromatic groups. This means that the peptidic moeities may be more accessable to water-borne contaminants than the aromatic groups.

Thus, both lines of studies - the dynamic experiments and the structural investiga- tions?are now poised to address the accessability of reactive functional groups, using the different approaches as appropriate.

Future Activities:

The two-pronged program will now be bumped up to three simultaneous approaches. On the first front, we will continue the ternary and higher order interaction experiments with HS, metal ions, and organic chelators, now focusing on the dynamics of the HS colloids, particularly aggregation/disaggreagation. The colloid dynamics evidence from PCS analysis demonstrates how three components can actually have quarternary (or perhaps higher order) interactions. This aspect is important in many contexts (e.g., facilitated transport of metals by colloids). On the second front, structural characterization of HS from the other experimental soils to be used in the experiments will be completed in a similar manner as described by Fan, et al. On the third front, we are now beginning to study multimetal ion bioavailability to plants (Higashi, et al., 1999), comparing the hydroponic (with HS) with whole soil systems.

Journal Articles on this Report : 2 Displayed | Download in RIS Format

Other project views: All 28 publications 8 publications in selected types All 4 journal articles
Type Citation Project Document Sources
Journal Article Fan TW-M, Higashi RM, Lane AN. Chemical characterization of a chelator-treated soil humate by solution-state multinuclear two-dimensional NMR with FTIR and pyrolysis-GCMS. Environmental Science & Technology 2000;34(9):1636-1646. R825960 (1999)
R825960 (2000)
R825960 (Final)
R825433 (Final)
R825433C007 (Final)
  • Abstract: ACS-Abstract
  • Journal Article Higashi RM, Fan TW-M, Lane AN. Association of desferrioxamine with humic substances and their interaction with cadmium(II) as studied by pyrolysis-gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy. Analyst 1998;123(5):911-918. R825960 (1999)
    R825960 (Final)
    R825433 (Final)
    R825433C007 (Final)
  • Abstract: RSC-Abstract
  • Supplemental Keywords:

    soil, sediments, adsorption, chemical transport, heavy metals, bioremediation, environmental chemistry, Scientific Discipline, Geographic Area, Waste, Water, Ecosystem Protection/Environmental Exposure & Risk, Bioavailability, Contaminated Sediments, Remediation, Environmental Chemistry, State, Fate & Transport, Bioremediation, West Coast, fate, fate and transport, sorption, biogenic ligands, humic substances, NMR spectroscopy, pyrolysis GCMS, rhizospheric, contaminated sediment, soils, adsorption, chemical transport, vascular plants, metal ion bioavailability, FTIR microspectroscopy, wheat, California, phytoremediation, sediments, soil humic substances, biogenic organic matter, heavy metals, metal compounds, metals, fluoroescence spectrophotometry, duckweed

    Progress and Final Reports:

    Original Abstract
  • 1998
  • 2000 Progress Report
  • Final Report