Characterization of Natural Organic Matter by Nuclear Magnetic Resonance Spectroscopy

Natural organic matter (NOM) is a major intermediate in the global carbon, nitrogen, sulfur, and phosphorus cycles. NOM is also the environmental matrix that frequently controls binding, transport, degradation, and toxicity of many organic and inorganic contaminants. Despite its importance, NOM is poorly understood at the structural chemistry level because of its molecular complexity and heterogeniety. Nuclear magnetic resonance (NMR) spectroscopy is one of the most useful spectrometric methods used to investigate NOM structure because qualitative and quantitative organic structure information for certain organic elements can be generated by NMR for NOM in both the solution and solid states under nondegradative conditions. However, NMR spectroscopy is not as sensitive as infrared or ultraviolet-visible spectroscopy; it is not at present applicable to organic oxygen and sulfur, and quantification of NMR spectra is difficult under certain conditions. The purpose of this overview is to present briefly the “state of the art” of NMR characterization of NOM, and to suggest future directions for NMR research into NOM. More comprehensive texts concerning the practice of NMR spectroscopy and its application to NOM in various environments have been produced by Wilson and by Wershaw and Mikita. Carbon, hydrogen, and oxygen are the major elements of NOM; together they comprise about 90% of the mass. The minor elements that constitute the remainder are nitrogen, sulfur, phosphorus, and trace amounts of the various halogen elements. With the exception of coal, in which carbon is the most abundant element, the order of relative abundance in NOM on an atomic basis is H > C > O > N > S > P = halogens. The optimum NMR-active nuclei for these elements are 1H, 13C, 17O, 15N, 33S, 31P, and 19F. The natural abundances and receptivities of these nuclei relative to 1H are given in Table 12.1. Quadrupolar effects for 17O, 33S, and halogen elements other than 19F lead to line broadening that greatly limits resolution in NMR studies of these elements in NOM.

NMR Studies of the Reaction of Amino Acids with Aqueous Chlorine

Over the past 20 years, gas chromatography/mass spectroscopy (GC/MS) has been widely used to identify trace organic environmental contaminants and to study the mechanisms of the formation or transformation of organic compounds either by natural or man-made processes. In the area of water and wastewater disinfection, GC/MS has been highly successful in identifying numerous volatile organic chlorination by-products, some of which may pose undesirable health risks to humans and aquatic organisms at concentrations found in some waters. However, despite a considerable amount of research in this area much of the chemistry continues to be poorly understood. Analysis of trace organics by GC/MS relies on the assumption that the compounds to be analyzed are (1) volatile and (2) thermally stable to GC temperatures as high as 300 °C. Because nuclear magnetic resonance spectroscopy (NMR) is a mild and nondestructive method of analysis, it can reveal reactions that occur in water that cannot be observed by GC/MS. Until recently the reactions of amino acids with two or more equivalents of aqueous chlorine were believed to produce aldehydes and nitriles according to equation (1). LeCloirec and Martin have reported that the formation of nitriles in such situations may come in part from the reaction of monochloramine with aldehydes (equation (2)). Because reaction (2) may affect the distribution of products in reaction (1), it was important to determine the relationship between these two reactions. This chapter will review the applications of NMR we have used in studies of the products formed upon chlorination of amino acids.

Acquisition and Interpretation of Liquid-state 1H NMR Spectra of Humic and Fulvic Acids

Fourier Transform nuclear magnetic resonance (NMR) spectrometers have become available to many researchers studying humic substances over the last decade. As a result, liquid-state proton (1H) NMR spectrometry has been commonly used to determine the nonexchangeable proton distribution in humic and fulvic acids. The high sensitivity of the 1H nucleus to NMR spectrometry allows spectra to be obtained on a relatively small quantity of sample (10-100 mg) in a short time (10-30 min). 1H NMR spectrometric profiles of humic substances are useful to environmental scientists in determining the source, properties, and degree of transformation (humification) of organic matter that is operationally classified as humic substances. These 1H NMR spectrometric profiles, which provide information about hydrogen distributions in humic substances, are more useful for structural and biogeochemical studies when combined with 13C NMR spectra, which provide information on carbon distributions, and infrared spectra, which provide information on oxygen distributions. These three spectra, in conjunction with elemental composition, molecular weight, and titrimetric data, can then be synthesized to provide average structural characteristics that can be related to source, properties, and degree of humification of the organic material being studied. Special challenges, that are not met when obtaining the spectra of pure compounds, are encountered in obtaining 'H NMR spectra of natural humic substances. These challenges include (1) lack of complete dissolution of macromolecular humic substances at the high concentrations required for NMR studies; (2) significant concentrations of exchangeable protons giving broad peaks that obscure portions of the spectrum; (3) broad peaks of non-exchangeable protons over the entire spectrum that cause difficulties in correct phasing; (4) unstable structures that oxidize, hydrolyze, and structurally rearrange at the high pH conditions under which humic substances are the most soluble; and (5) the presence of unusual structures that complicate straightforward assignment of structure from handbook data. The purposes of this chapter are to describe methods of sample preparation and to provide generally applicable structural assignments whereby 1H NMR spectra suitable for quantitative studies of humic substance structure may be obtained and interpreted.

Cation and Water Interactions in the Interlamellae of a Smectite Clay

Advances in NMR instrumentation and availability have led to increased application to mineral systems and to environmental problems. The sensitivity of high-field NMR systems is nearly sufficient to work at real environmental concentrations. Even with limited sensitivity, the amount of chemical information obtained through NMR spectroscopy makes it a very valuable technique in many model systems. The application of NMR spectroscopy in mineral systems has been primarily limited to studies of the structural metals aluminum and silicon. However, in recent years there have been several publications on mobile cations in minerals, including work on the exchangeable cations in clays. Our interests lie in understanding the sorption of cations in clays, the structural sites available for that sorption, and the role of water in cation–clay interactions. Our goal is to eventually understand the molecular interactions that determine the adsorption and diffusion of cations in clays and, thus, the role of clays in determining cation transport through the geosphere. This fundamental understanding has applications in the fate of heavy metals, radionuclides, and even the mobility of nutrients for plants. It is well known that there are very strong interactions between metals and humic materials and these are also strong contributors to cation mobility. However, for simplicity, we have chosen to focus on the interactions of mobile metal ions with well-characterized clays. An NMR-based approach to this problem can take two complementary directions: first, studies of the structural components of clays such as 29Si and 27Al NMR as a function of cation or hydration; second, NMR studies of probe molecules—which in this case are the cations themselves. High magnetic field, multinuclear NMR spectrometers make it quite possible to study various “uncommon” nuclei with relative ease. It is our experience that using the cations as probe nuclei for studying sorption phenomena yields more information than studies of structural nuclei. This chapter is basically a report of work in progress on several systems that are starting to yield interesting results, which it is hoped will lead to a general understanding of these complex systems.

27Al NMR Study of the Hydrolysis and Condensation of Organically Complexed Aluminum

Aluminum is the most abundant metal of the Earth’s crust, of which it represents approximately 8%, ranking after oxygen and silicon. It exists mainly as oxides. In terrestrial environments, aluminum commonly exists as secondary (authigenic) hydroxide or aluminosilicate minerals, mainly clays. These minerals are highly insoluble at neutral pH. However, aluminum occurs in detectable amounts in natural waters, due to leaching of the soil minerals in acidic conditions. Soil acidity may have a natural origin, such as an acidic (silicic) mother rock, melted snow, dissolved carbonic acid, or biologically generated organic acids. During the past two decades, it has been demonstrated that one of the major origins of increased aluminum mobilization and transport in forested soils is introduction of strong acid through atmospheric sulfur and nitrogen deposition. It has also been shown that aqueous aluminum is the biogeochemical link between atmospheric pollution and damage caused to tree roots and aquatic organisms such as plankton, crustaceans, insects, and fish. Biological studies have shown that the different aluminum species exhibit various toxicities: the most toxic are the monomeric and the polynuclear species; complexation with organic acids results in low toxicity. The significance of aluminum to human health has long been regarded as negligible. There is a possible link between high-level aluminum contamination by renal dialysis or hemodialysis, and neurodegenerative health disorders such as Parkinson’s or Alzheimer’s diseases, but the part played by aluminum is not clear. However, since aluminum salts are used on an industrial level as coagulants and flocculants in water treatment, the aluminum concentration and speciation in drinking water deserve careful monitoring. Because of the specific toxicity of the aluminum species, there has been considerable concern in the past two decades over the speciation of aqueous aluminum present in soils and aquatic systems. To this end, several techniques have been developed in order to partition the aluminum species. The most common among them are chromatographic separation and categorization methods such as timed ferron reaction, and computational methods derived from thermodynamic equilibrium constants. However, significant discrepancies between the results have been noticed, and attributed to the dramatic interference of organic and inorganic anions in the Al fractionation.

Comparative Results of 27Al NMR Spectrometric and Ferron Colorimetric Analyses of Hydroxyaluminum Hydrolysis Products in Aged, Mildly Acidic, Aqueous Systems

The abundant presence of aluminum (Al) in the environment, its commercial importance, and the potential toxic effects of dissolved forms of Al on living organisms have brought about extensive studies of Al hydrolysis products that originate from natural processes and industrial activities. A recent review of past studies of hydroxyaluminum (hydroxy-Al) solutions illustrates the complexity of aluminum hydrolysis. Because of the need to identify and quantify individual Al ion species, initial investigations of concentrated solutions of hydroly/ed aluminum salts using 27A1 nuclear magnetic resonance (NMR) spectrometry began more than 20 years ago in the United Kingdom with continuous-wave instrumentation. Since then, groups in Switzerland, France, and the United States as well as in the UK, involved with geochemical and pharmacological investigations, have applied pulse NMR instruments to conduct research mostly with concentrated Al hydrolysates. In NMR spectrometry, shapes and positions of spectral peaks indicate aluminum nuclei in different chemical environments, and the peak areas are proportional to the number of nuclei in each given symmetry. Figures 8.1 (a) and (b) and 8.2(a) and (b) illustrate liquid-state 27A1 NMR spectra of aluminum hydrolysis products. The well-defined peak at 62.5ppm, downfield from the Oppm reference position for Al(H2O)3+6, has been attributed to the tetrahedrally coordinated aluminum ion within the highly symmetrical configuration of the [AlO4Al12(OH)24(H2O)12]7+ species or “Al13” polyion in aqueous solution. This geometrical arrangement, also termed the “Keggin” structure, was originally deduced from X-ray diffraction studies of aluminum hydroxysulfate and hydroxyselenate salts precipitated from heated concentrated aluminum solutions after being partly neutralized with base. The Al13 structural unit that was postulated has 12 Al ions octahedrally coordinated with OH ions enclosing the central tetrahedral Al ion which is bonded to four of the octahedral ions through bridging by O2− ions. For a given sample specimen, broad NMR spectral peaks indicate the existence of (1) chemical-exchange processes and (2) species having less symmetry and larger electric field gradients than those which give rise to narrower peaks. Mildly acidic solutions of aluminum hydrolysis products exhibit a broad peak downfield from 0ppm centered at ~1 to ~2ppm.

Proton and 19F NMR Spectroscopy of Pesticide Intermolecular Interactions

Sorption of organic pollutants by soils and sediments is one of the main chemical processes that controls pollutant migration in the environment. Information about the molecular mechanisms by which an organic pollutant interacts with other solution-phase constituents and with solid-phase sorbents would be invaluable for more accurate prediction of pollutant fate and transport and for optimal design and application of remediation procedures. Many current models and remediation strategies are based upon the “partition theory” of organic compound sorption, which predicts sorption coefficients from properties such as water solubility or octanol-water partition coefficients. Partition theory is well suited for nonpolar hydrocarbons but may not be appropriate for pesticides with electrophilic or weakly acidic or basic substituents, which may interact with soils or organic matter through specific interactions such as hydrogen bonding or charge-transfer complexes. If a pesticide can form hydrogen bonds or a charge-transfer complex with a sorbent, sorption may be greater than in the absence of specific interactions. Nuclear magnetic resonance (NMR) spectroscopy is well suited for the study of pesticide-solution or pesticide-sorbent interactions because NMR is an element-specific method that is extremely sensitive to the electron density (shielding) near the nucleus of interest. Consequently, solution-state NMR can distinguish between closely related functional groups and can provide information about intermolecular interactions. All nuclei with nonzero nuclear spin quantum number can be studied by NMR spectroscopy. Of the more than 100 NMR-active nuclei, 1H and 19F are the easiest to study because both have natural abundances near 100% and greater NMR sensitivity than any other nuclei. In addition, both 1H and 19F have zero quadrupolar moments, which means that sharp, well resolved NMR peaks can be obtained, at least in homogeneous solutions. Proton NMR is well suited for elucidating molecular interactions in solution but cannot be used to study interactions between pesticides and heterogeneous sorbents such as soils, humic acid, or even cell extracts, since protons in the sorbent generally produce broad peaks that mask the NMR peaks from the solute or sorbate of interest. In contrast, 19F NMR can be used to study interactions between fluorine-containing molecules and heterogeneous sorbents because the fluorine concentration in most natural sorbents is negligible.

Characterization of Nitrogen in Plant Composts and Native Humic Material by Natural-Abundance 15N CPMAS and Solution NMR Spectra

Soil organic matter (SOM) provides one of the major deposits for carbon and nitrogen on the surface of the Earth. It is continuously produced, mainly from dead plant material, by composting and humification processes. During these processes microorganisms thoroughly convert the starting material, which consists mostly of insoluble lignocelluloses. The end products of these processes in average middle-European soils, that contain typically 1 to 5% w/w of organic material, are clay–SOM complexes which are insoluble in all the usual organic and inorganic solvents. The standard aqueous sodium hydroxide extraction procedure dissolves at most 40% of the total organic carbon in all the soils tested by our group. The insoluble majority, the humin fraction, remains as poorly defined aluminosilicate- SOM complexes. During the decomposition and conversion processes the carbon to nitrogen ratio–decreases. Compared to the starting material, SOM is enriched in nitrogen. Under natural conditions, i.e., without the artificial addition of nitrogen in the form of manure or fertilizer, SOM provides the major part of the nitrogen available for plant growth. The chemical characterization of this ubiquitous but ill-defined material has only been partly successful until now. For characterization of the organic carbon in complete soils and extracts, nuclear magnetic resonance (NMR) methods appear to be most promising, especially since the application of high-resolution solid-state methods has become almost a laboratory routine. The combination of proton–carbon cross polarization with high-speed magic angle rotation (the CPMAS technique) permits the study of complete native soils, and thus provides detailed information about the gross chemical structure of the total SOM, without introducing any of the chemical modifications that could result from aggressive chemical extraction procedures. It has been shown by 13C CPMAS and high-resolution (HR) solution 13C NMR studies of a series of typical European soils, in which the concentration of paramagnetic metal ions was fairly low and which contained humic material with an aromatic carbon content ≤20%, that the carbon could be quantitatively assigned. The measurement of the 13C CPMAS spectra of complete native soils with a carbon content in the region of 1 % w/w is rather instrument-time consuming, and appeared to be at the limit of sensitivity.

31P FT-NMR of Concentrated Lake Water Samples

The use of phosphorus-31 Fourier Transform nuclear magnetic resonance (31P FT-NMR) spectroscopy for the study of dissolved organic phosphorus (DOP) in fresh water has been recently established by Nanny and Minear. The fact that NMR is an element-specific technique, is nondestructive, and has the ability to differentiate between similar phosphorus compounds makes it invaluable for the identification and characterization of DOP. Such information regarding DOP is required in order to understand aquatic nutrient cycling. The difficulty with using 31P FT-NMR spectroscopy for such studies is the extremely low DOP concentration; usually ranging from < 1 μg P/L in oligotrophic lakes to approximately 100 μg P/L for eutrophic systems. Nanny and Minear raised the DOP concentration into the NMR detection range, which is on the order of milligrams of phosphorus/liter, by concentrating large volumes of lake water with ultrafiltration (UF) and reverse osmosis (RO) membranes. Volume concentration factors of several ten thousand fold provided DOP concentrations of up to 60 mg P/L. Other DOP concentration methods such as anion exchange, lanthanum hydroxide precipitation, and lyophilization require severe chemical and/or physical transformations of the sample and/or they need long processing times, all of which increase the risk of DOP hydrolysis. Sample concentration with UF and RO membranes does not require the sample to undergo these major changes and is also a relatively rapid concentration method. In addition to these concentration capabilities, the use of ultrafiltration and reverse osmosis membranes permitted fractionation of the DOP samples according to molecular size. Nanny and Minear used three membranes in series with decreasing pore size: 30kDa (kilodaltons), 1 kDa, and RO (95% NaCl rejection) to separate the high-molecular-weight, intermediate-molecular-weight, and low-molecular-weight DOP species. In the intermediate-molecular-weight fraction, Nanny and Minear observed the presence of monoester and diester phosphates. Spectra from ten samples collected over a year typically consisted of a large broad signal in the monoester phosphate region spanning from a chemical shift of 2.00 ppm to −0.50 ppm. The maximum of this signal was usually in the range of 1.00 to 1.50 ppm. This broad signal had a shoulder in the diester phosphate region which sometimes was intense enough to appear as an individual signal.

Solution and Condensed Phase Characterization

Improvements in nuclear magnetic resonance (NMR) instrumentation, magnetic field strength, pulse sequences, and computer technology and software have increased the range of applications and specific elements available for study by NMR. The five chapters in this Part clearly indicate the benefits of these advances, especially regarding studies of aquatic, environmental significance. Each of the studies focuses on environmentally significant issues. For example, chlorination is widely used to disinfect drinking waters, a method that can produce undesirable disinfection by-products. This was first recognized in 1974 with the discovery of trihalomethanes in most finished drinking waters where hypochlorite was used for disinfection. Chapter 7 examines the chlorination of alanine and relates it to the chlorination reactions of acetaldehyde and ammonia, a topic of importance with respect to drinking water disinfection. Aluminum is also widely used in drinking water treatment, and understanding its hydrolysis chemistry and complexation behavior is of great importance to aquatic chemistry. In addition, the aquatic chemistry of aluminum is important because acid rain can release soluble aluminum ions from clay into soil water, possibly damaging terrestrial plant life. Aluminum may eventually reach and accumulate in hydrological systems where it can be toxic to aquatic life. Chapters 8 and 9 focus on 27Al NMR in defining aqueous aluminum speciation in a mildly acidic solution or in the presence of complexing organic compounds. Furthermore, aluminum is of environmental and geochemical significance since it is an integral component of clays, another ubiquitous constituent of natural waters (surface and ground). Interaction between clays, cations, and internal water molecules can be significant in understanding the fate and transport of chemicals through the environment. Since colloidal suspensions of clay materials frequently represent challenges to water and wastewater treatment, understanding of physical and chemical processes are of tantamount importance to the environmental scientist and engineer. Chapter 10 explores cation behavior in clay matrices by using “uncommon” nuclei such as 7Li, 23Na, and 133Cs as probes. This is unique in that many NMR studies of complexation in clay have focused primarily upon the nuclei 27A1 and 29Si.