Use for humic substance analysis

The determination of molecular sizes and weights for humic substances is a task complicated by the nature of these materials. A brief overview of the methods most commonly used to determine molecular sizes and weights of humic substances has been presented in this chapter. Many of the methods discussed are powerful techniques that can yield information about the molecular size, shape, and weight of humic substances. No single method alone, however, is sufficient to provide a complete understanding of these molecular characteristics. Meaningful and accurate conclusions can only be made by using the data provided by different methods of analysis. 

The development of methods of analysis of tria2ines and thek hydroxy metabohtes in humic soil samples with combined chromatographic and ms techniques has been described (78). A two-way approach was used for separating interfering humic substances and for performing stmctural elucidation of the herbicide traces. Humic samples were extracted by supercritical fluid extraction and analy2ed by both hplc/particle beam ms and a new ms/ms method. The new ms /ms unit was of the tandem sector field-time-of-flight/ms type. 

Supercritical fluid extraction (SFE) is generally used for the extraction of selected analytes from solid sample matrices, but applications have been reported for aqueous samples. In one study, recoveries of 87-100% were obtained for simazine, propazine, and trietazine at the 0.05 ug mL concentration level using methanol-modified CO2 (10%, v/v) to extract the analytes, previously preconcentrated on a C-18 Empore extraction disk. The analysis was performed using LC/UV detection. Freeze-dried water samples were subjected to SFE for atrazine and simazine, and the optimum recoveries were obtained using the mildest conditions studied (50 °C, 20 MPa, and 30 mL of CO2). In some cases when using LEE and LC analysis, co-extracted humic substances created interference for the more polar metabolites when compared with SFE for the preparation of the same water sample. ... 

A problem for both humic- and yellow substances is that for these groups of experimentally defined components of different sources, each analysis will be ambiguous in terms of relative composition and molecular weight distribution. Additionally it appears that almost every scientist working in this field has developed his own extraction procedure (Weber and Wilson, 1975 Mantoura and Riley, 1975 a Schnitzer, 1976 Stuermer and Harvey, 1977). Different extraction times and -procedures result in different compositions of the organic constituents (Laane and Kramer, 1984). Soil humic-and fulvic acids, often used for studies on the interaction with trace elements, and those derived from water have certainly not the same composition and contain not the same distribution of functional groups. Therefore, results should be compared with care (Buffle, 1980 Buffle et al., 1984). 

Note Humic substances (HS) are operationally defined as DOC that is retained on XAD-2 or XAD-8 resins. UDOM is operationally defined as DOC that is retained by a membrane with a 1 nm pore size and 1000 Da molecular weight cutoff. 13C-NMR spectroscopy was used for analysis of carbon functional groups, which are presented as a percentage of the total organic carbon in the sample. THNS, total hydrolyzable neutral sugars THAA, total hydrolyzable amino acids. 

Various separation methods have been used to isolate, fractionate, and characterize humic materials. Originally it was fractionation, based on solubility differences of humic components in diluted alkalis and acids, which laid the ground work for the first classifications of humic substances (HS) in the 19th century (Mulder, 1861 Sprengel, 1837) and provided for operational definition of HS (Kononova, 1966). And now, alkali extraction is the method of choice for isolating HS from solid humus-containing substrates like soil, peat, coal, and so on (Swift, 1996), while hydrophobic resins (e.g., Amberlite XAD resins) are typically used to extract HS dissolved in natural waters (Aiken, 1985). Initial research on HS began with the used simple separation methods to prove, examine, and define characteristics of components of humic matter (Oden, 1919).Today, however, advances in HS research require ever more sophisticated techniques of separation combined with structural analysis (Orlov, 1990 Stevenson, 1994). 

The same methods can be used for determining the chemical composition of the liquid phase as those used for the composition of the solid after digestion or extraction (absorption and emission spectroscopy, electroanalytical methods, ion-selective electrodes, neutron activation analysis, mass spectrometry, etc.). The humic substances of natural waters can also be analyzed. 

The molecules of both HSl and HS2 have a high colloid stability due to their very low collision efficiency and small size which prevent them from settling under normal gravitational acceleration. Their separation in a centrifuge with a g value of some thousands is still practically inefficient. This fact is used for sedimentation analysis to investigate the destabilization and aggregation rate of the coagulation process of humic substances. 

Other studies of humic substances using pyrolysis were done for evaluating the seasonal variations in humic materials [10] in different soils, for the analysis of marine deposits [11], the analysis of composts [12] or of degraded lignins [3]. 

The first application of NMR for the study of humic substances in soil was made by Neyroud and Schnitzer(7) using continuous wave NMR spectroscopy. Later, Gonzalez-Vila et al.(2) were the first to apply Fourier transform NMR for the analysis of dissolved humic substances extracted from soil. The development of... 

Size exclusion chromatography (SEC) has been used to measure molecular weight (MW) distribution of humic substances (3, 6-9). Coupled with detection methods such as molecular fluorescence spectroscopy and dissolved organic carbon analysis (7), electrochemical detection (9), and atomic emission spectroscopy (5), SEC has been used extensively to study humic-metal complexes. A major disadvantage of SEC is that it does not provide adequate resolution for separating humic materials as they do not appear to be made up of distinct fractions with large differences in MW. 

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