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The composition of dissolved organic matter (DOM) in extremely deep groundwaters has rarely been investigated. Focusing on deep fracture waters in billion year old rocks in South Africa, we hypothesized the DOM would be predominantly derived from in situ microbial sources, consistent with H2-driven lithoautotrophic ecosystems previously documented in these fluids. We collected groundwater from 0.6 to 3.4 km depths via boreholes in mines, and characterized the DOM by a variety of analytical methods. Dissolved organic carbon concentrations ranged from 0.25 to 4.9 mg C/L. Low molecular weight organic acids (formate, acetate, lactate, propanoate) and amino acids (after hydrolysis) accounted for varying fractions of the total dissolved organic carbon. UV spectrophotometry, scanning fluorimetry, and nuclear magnetic resonance (NMR) spectra indicated lower concentrations of aromatic compounds than are commonly found in surface waters and shallow groundwaters. Molecular weights of aromatic compounds measured by size-exclusion chromatography were dominantly 500–1000 Da. Scanning fluorimetry revealed humic substances in two shallow (578 and 1300 m) samples; deeper samples showed peaks indicating aromatic amino acids. Intensive study of one sample from a previously sampled borehole in Beatrix mine resulted in large NMR peaks for aliphatics and carboxyls with lesser aromatics peaks; negative electrospray ion Fourier transform ion cyclotron resonance-mass spectrometry detected protein-, lipid-, and lignin-like compounds. These results are consistent with DOM in the form of microbial metabolites, and modified cell components (membrane lipids and proteins). These findings support previous geomicrobiological reports of chemolithoautotrophic microbial ecosystems functioning in isolation from photosynthetic primary production.

Deep continental subsurface fracture water systems, ranging from 1.1 to 3.3 km below land surface (kmbls), were investigated to characterize the indigenous microorganisms and elucidate microbial carbon sources and their cycling. Analysis of phospholipid fatty acid (PLFA) abundances and direct cell counts detected varying biomass that was not correlated with depth. Compound-specific carbon isotope analyses (δ13C and Δ14C) of the phospholipid fatty acids (PLFAs) and carbon substrates combined with genomic analyses did identify, however, distinct carbon sources and cycles between the two depth ranges studied. In the shallower boreholes at circa 1 kmbls, isotopic evidence indicated microbial incorporation of biogenic CH4 by the in situ microbial community. At the shallowest site, 1.05 kmbls in Driefontein mine, this process clearly dominated the isotopic signal. At slightly deeper depths, 1.34 kmbls in Beatrix mine, the isotopic data indicated the incorporation of both biogenic CH4 and dissolved inorganic carbon (DIC) derived from CH4 oxidation. In both of these cases, molecular genetic analysis indicated that methanogenic and methanotrophic organisms together comprised a small component ( In the samples collected from >3 kmbls in Tau Tona mine (TT107, TT109 Bh2), the CH4 had an isotopic signature suggesting a predominantly abiogenic origin with minor inputs from microbial methanogenesis. In these samples, the isotopic enrichments (δ13C and Δ14C) of the PLFAs relative to CH4 were consistent with little incorporation of CH4 into the biomass. The most 13C-enriched PLFAs were observed in TT107 where the dominant CO2-fixation pathway was the acetyl-CoA pathway by non-acetogenic bacteria. The differences in the δ13C of the PLFAs and the DIC and DOC for TT109 Bh2 were ∼−24‰ and 0‰, respectively. The dominant CO2-fixation pathways were 3-HP/4-HB cycle > acetyl-CoA pathway > reductive pentose phosphate cycle.

This study investigated the effects of iron on natural organic matter (NOM) absorbance, fluorescence, and molecular weight. Addition of iron(III) to 5 and 10 mg C/L XAD-8 isolated NOM samples at pH 5.5 caused absorbance of visible light to increase, fluorescence intensity of peaks at excitation/emission wavelengths 230/435 nm and 320/450 nm to decrease, and a small shift from intermediate to higher molecular weight components. Iron(II) had little or no effect on XAD-8 isolate properties. Addition of iron(III) to XAD-4 isolated NOM solutions also caused absorbance of visible light to increase. Iron(III) addition to XAD-4 isolate solutions caused a greater decrease in fluorescence intensity than was observed for the XAD-8 isolate, but little or no change in the molecular weight distribution. Overall, our results suggest that the effects of iron on NOM molecular weight, absorbance, and fluorescence properties depend upon the redox state of iron and the composition of the NOM. These results also suggest that ...

This study examines the importance of several possible mechanisms causing sunlight-mediated changes in the amounts of bacterial utilization and biomass growth on dissolved organic matter (DOM) from allochthonous sources. Our results demonstrate that, while hydroxyl radical reactions with DOM can be an important process increasing its bioavailability, other photoreactions will cause most of the sunlight-induced increases unless hydroxyl production rates are high (.;7 mmol L 21 d 21 ). Low molecular weight carboxylic acids could not account for most of the observed sunlight and hydroxyl-induced increases in DOM bioavailability. Both sunlight and hydroxyl-mediated reactions significantly decreased the bacterial growth efficiency of DOM, indicating that photochemical reactions affect not only the fraction of the total DOM pool available to bacteria on ecologically relevant timescales but also the substrate quality and ultimately the environmental fate of this material. Extrapolation of these results to field conditions suggests that photochemical and biochemical mineralization could be an important sink of DOC and source of bioavailable carbon in the Plum Island estuary during the summer months. Dissolved organic matter (DOM) is a heterogeneous mixture of natural organic compounds that is present in all natural waters. The sources of this carbon in freshwater systems include both in situ biological production (autochthonous sources) and detrital carbon from the surrounding terrestrial watershed (allochthonous sources). At one time, relatively labile autochthonous material was thought to be the dominant source of substrates for bacterial growth in these systems (Cole et al. 1982). More recently, it has become clear that relatively recalcitrant allochthonous (or humic) organic matter can also be a major source of energy and carbon for bacterial growth in many freshwaters (Tranvik 1988; Moran and Hodson 1990). One important environmental factor that may change the utilization of DOM by bacteria is exposure to light. A variety of studies have shown that irradiation of DOM by natural and/or simulated sunlight can increase both the amount of DOM that is susceptible to bacterial utilization and the abil1

Flow injection analysis was used to study the effect of a fulvic acid on the kinetics of iron(II) oxidation and iron colloid formation under conditions approximating fresh natural waters. While iron(II) oxidation in high-carbonate inorganic solutions is predicted well by a recently proposed homogeneous model, it overestimates the oxidation rate in low-carbonate solutions, possibly due to the formation of an intermediate iron(II) colloid or surface species. Results in fulvic acid solutions are consistent with the formation of an iron(II)–fulvic acid complex at both pH 6.0 and 8.0 which accelerates the overall oxidation rate relative to inorganic solutions. However, iron(III) complexation by fulvic acid greatly slows the formation of iron colloids, stabilizing dissolved iron(III). Decreased pH and increased ionic strength slow and decrease iron colloid formation. Evidence of a kinetic control on the distribution of iron(III) between organically complexed and colloidal forms is presented.

A combination of flow-injection analysis and kinetic analysis was used to examine the speciation of iron(II) and iron(III) in fulvic acid solutions as a function of pH, ionic strength, and time. This methodology was used to follow a shift in iron speciation from faster to slower reacting species over a timescale of several days. This speciation data shows that both iron(II) and iron(III)–fulvic acid complexes are important iron species in humic-containing natural waters and that their amounts and their rates of transformation to colloidal iron are controlled primarily by the kinetics of thermal (dark) reduction and iron(II) oxidation. The kinetic analysis methodology also yielded the rate constants for the thermal reduction of iron by the fulvic acid. These rate constants decrease with increasing pH and are independent of ionic strength. While thermal reduction was found to be too slow to produce large amounts of steady state iron(II) at circumneutral pH, it does provide a mechanism for iron redox cycling in the absence of photochemical or biochemical processes.

In this study, we examine the role of the hydroxyl (OH*) radical as a mechanism for the photodecomposition of chromophoric dissolved organic matter (CDOM) in sunlit surface waters. Using gamma-radiolysis of water, OH* was generated in solutions of standard humic substances in quantities comparable to those produced on time scales of days in sunlit surface waters. The second-order rate coefficients of OH* reaction with Suwannee River fulvic (SRFA; 2.7 x 10(4) s(-1) (mg of C/L)(-1)) and humic acids (SRHA; 1.9 x 10(4) s(-1) (mg of C/L)(-1)) are comparable to those observed for DOM in natural water samples and DOM isolates from other sources but decrease slightly with increasing OH* doses. OH* reactions with humic substances produced dissolved inorganic carbon (DIC) with a high efficiency of approximately 0.3 mol of CO2/mol of OH*. This efficiency stayed approximately constant from early phases of oxidation until complete mineralization of the DOM. Production rates of low molecular weight (LMW) acids including acetic, formic, malonic, and oxalic acids by reaction of SRFA and SRHA with OH* were measured using HPLC. Ratios of production rates of these acids to rates of DIC production for SRHA and for SRFA were similar to those observed upon photolysis of natural water samples. Bioassays indicated that OH* reactions with humic substances do not result in measurable formation of bioavailable carbon substrates other than the LMW acids. Bleaching of humic chromophores by OH* was relatively slow. Our results indicate that OH* reactions with humic substances are not likely to contribute significantly to observed rates of DOM photomineralization and LMW acid production in sunlit waters. They are also not likely to be a significant mechanism of photobleaching except in waters with very high OH* photoformation rates.

Molecular weight (MW) of humic materials is a key factor controlling proton and metal binding and organic pollutant partitioning. Several studies have suggested preferential adsorption of higher MW, more aromatic moieties to mineral surfaces; quantification of such processes is fundamental to develo

The analysis of carbon type distribution and chemical structure of natural organic matter (NOM) by 13C NMR spectroscopy is important for understanding its origins and reactivity. While prior work has used solution-state NMR, solid-state NMR has the potential to provide this information with less instrument time and sample processing, while providing an array of advanced filtering techniques. Quantitative and qualitative analyses with 13C solid-state magic angle spinning (MAS) NMR techniques are described for three commercially available samples and one fulvic acid sample isolated from the Rio Grande in New Mexico. This study demonstrates the utility of solid-state 13C NMR for aquatic NOM structural characterization by determining the % carbon type for three standard natural organic materials and comparing these results to the existing solution-state 13C NMR determinations. The solid-state 13C MAS NMR results are used to determine % carbon distribution, estimates of elemental composition (%C, %H, %(O+N)), aromatic fraction (fa), nonprotonated aromatic fraction (faN), an estimate of aromatic cluster size, and ratio of sp2 to sp3 carbons. Additionally, Gaussian deconvolution is used for a more detailed analysis of carbon type than frequency band integration techniques. The solid-state 13C NMR results of these analyses indicate the chemical composition of Rio Grande fulvic acid has higher aromatic fraction and nonprotonated aromatic fraction (fa =0.32, faN = 0.21) and a lower sp2/sp3 fraction and cluster size (sp2/sp3 = 0.66, C = 6) compared to Suwannee River fulvic acid (fa = 0.18, faN = 0.10, sp2/sp3 = 0.82, C = 8).

Synchronous fluorescence (SF) spectra of dissolved organic matter (DOp/I) show promise for mixing studies and can resolve multiple river inputs to large water masses in a single fast measurement. This study examines the effects of physicochemical variables on DOM SF independent of mixing. Our results show that changes in the pH and metal ion concentration do not affect SF in the alkaline Cuyahoga-Erie sy:;tem. The effect of temperature is small and correctable. Model predictions suggest that photobleaching of DOM SF can limit the use of this method to short periods (3-7 d, depending on conditions). SF spectra are sensitive enough to detect 1% river water mixed in Lake Erie water. The intrinsic fluorescence of natural waters that arises from dissolved organic matter (DOM) provides a potentially useful method of tracing the mixing of rivers into large water masses like the oceans or the Great Lakes. Although fluorescence spectroscopy is fast, sensitive, and adaptable to shipboard or remote sensing (Mackey et al. 1989), application of this method to circulation studies is hampered by a number of potential interferences. In particular, laboratory studies indicate that DOM fluorescence spectra are a function of physical and chemical variables in the environment. On the other hand, results of several field studies demonstrate conservative behavior of DOM fluorescence. Conservative behavior has been observed in the mixing of the Rhine into the North and Wadden Seas (Zimmerman and Rommets 1974) and the mixing of smaller rivers in the southeastern United States in the Atlantic (Dorsch and Bidleman 1982; Willey and Atkinson 1982). In each case, the maximum fluorescence was found to correlate with a known conservative tracer (salinity) as freshwater was diluted with seawater. While conservative behavior is desired for mixing studies, it is somewhat surprising in light of laboratory studies of DOM fluorescence. The maximum fluorescence intensity of DOM is known to be a function of pH (Laane 1982; Ferrari 199 1), metal ion concentration (Saar and Weber 1980; Ferrari 1991; Cabaniss 1992; Cook and Langford 1995), and exposure to sunlight (Kramer 1979; Kieber et al. 1990; Kouassi and Zika 1990; Chen and Bada 1992). Furthermore, fluorescence quantum efficiency is a function of temperature, and the in situ generation of new fluorophores (e.g. algal

Synchronous fluorescence spectra of standard humic substances (HS) at variable pH can only be adequately explained by positing large (≥7) numbers of fluorescent factors. Although most of the variation (99.9%) can be explained using only two or three major factors, careful representation of the experimental error shows numerous additional minor factors that have been overlooked in previous work. The experimental error in these spectra was measured by repeated spectral acquisition and estimated by a linear regression equation relating the standard deviation, σ , to fluorescence intensity. Each of the six HS examined gave a unique synchronous spectral shape at pH 7.0, although most showed similar patterns of pH dependence. These results indicate that these synchronous fluorescence measurements have a high information content, suitable for tracer studies of HS.

Ethnotraditional use of plant-derived natural products plays a significant role in the discovery and development of potential medicinal agents. Plants of the genus Taraxacum, commonly known as dandelions, have a history of use in Chinese, Arabian and Native American traditional medicine, to treat a variety of diseases including cancer. To date, however, very few studies have been reported on the anti-carcinogenic activity of Taraxacum officinale (TO). In the present study, three aqueous extracts were prepared from the mature leaves, flowers and roots, and investigated on tumor progression related processes such as proliferation and invasion. Our results show that the crude extract of dandelion leaf (DLE) decreased the growth of MCF-7/AZ breast cancer cells in an ERK-dependent manner, whereas the aqueous extracts of dandelion flower (DFE) and root (DRE) had no effect on the growth of either cell line. Furthermore, DRE was found to block invasion of MCF-7/AZ breast cancer cells while DLE blocked the invasion of LNCaP prostate cancer cells, into collagen type I. Inhibition of invasion was further evidenced by decreased phosphorylation levels of FAK and src as well as reduced activities of matrix metalloproteinases, MMP-2 and MMP-9. This study provides new scientific data on TO and suggests that TO extracts or individual components present in the extracts may be of value as novel anti-cancer agents.

Ethnotraditional use of plant-derived natural products plays a significant role in the discovery and development of potential medicinal agents. Plants of the genus Taraxacum, commonly known as dandelions, have a history of use in Chinese, Arabian and Native American traditional medicine, to treat a variety of diseases including cancer. To date, however, very few studies have been reported on the anti-carcinogenic activity of Taraxacum officinale (TO). In the present study, three aqueous extracts were prepared from the mature leaves, flowers and roots, and investigated on tumor progression related processes such as proliferation and invasion. Our results show that the crude extract of dandelion leaf (DLE) decreased the growth of MCF-7/AZ breast cancer cells in an ERK-dependent manner, whereas the aqueous extracts of dandelion flower (DFE) and root (DRE) had no effect on the growth of either cell line. Furthermore, DRE was found to block invasion of MCF-7/AZ breast cancer cells while DLE blocked the invasion of LNCaP prostate cancer cells, into collagen type I. Inhibition of invasion was further evidenced by decreased phosphorylation levels of FAK and src as well as reduced activities of matrix metalloproteinases, MMP-2 and MMP-9. This study provides new scientific data on TO and suggests that TO extracts or individual components present in the extracts may be of value as novel anti-cancer agents.

Plants used in folklore medicine continue to be an important source of discovery and development of novel therapeutic agents. In the present study, we determined the effects of crude aqueous extracts of a panel of medicinal plants on the growth and invasion of cancer cells. Our results showed that extracts of L. tridentata (Creosote Bush) and J. communis L. (Juniper Berry) significantly decreased the growth of MCF-7/AZ breast cancer cells. The latter as well as A. californica (Yerba Mansa) inhibited invasion into the collagen type I gel layer. Furthermore, the phosphorylation levels of extracellular signal-regulated kinase 1 and 2 (ERK1/2) decreased when the cells were exposed to aqueous extracts of L. tridentata, J. communis L. and A. californica. This study provides original scientific data on the anticancer activity of selected aqueous medicinal plant extracts used in traditional medicine.

Plants used in folklore medicine continue to be an important source of discovery and development of novel therapeutic agents. In the present study, we determined the effects of crude aqueous extracts of a panel of medicinal plants on the growth and invasion of cancer cells. Our results showed that extracts of L. tridentata (Creosote Bush) and J. communis L. (Juniper Berry) significantly decreased the growth of MCF-7/AZ breast cancer cells. The latter as well as A. californica (Yerba Mansa) inhibited invasion into the collagen type I gel layer. Furthermore, the phosphorylation levels of extracellular signal-regulated kinase 1 and 2 (ERK1/2) decreased when the cells were exposed to aqueous extracts of L. tridentata, J. communis L. and A. californica. This study provides original scientific data on the anticancer activity of selected aqueous medicinal plant extracts used in traditional medicine.

Native American medicinal plants are traditionally used to prevent and treat a variety of diseases, including cancer. These herbal preparations are alleged to have many biological activities, such as stimulation or suppression of immune responses and antiproliferative effects on cancer cells. In the present study, we investigated the effects of aqueous and ethanol extracts from two Native American plants, Ligusticum porteri (Osha) and Anemopsis californica (Yerba Manza), on the growth of human MCF-7/AZ breast and HCT8/E11 colon cancer cells. The aqueous and ethanol extracts from A. californica potently inhibited growth of MCF-7/AZ in a concentration-dependent manner, whereas the growth of HCT8/E11 was unaltered. Extracts from L. porteri showed no activity on either cell line. In addition, we observed that the extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) activities were markedly decreased when exposed to both extracts from A. californica. These results suggest that the growth inhibitory effect of A. californica in breast cancer cells is ERK-mediated.

Native American medicinal plants are traditionally used to prevent and treat a variety of diseases, including cancer. These herbal preparations are alleged to have many biological activities, such as stimulation or suppression of immune responses and antiproliferative effects on cancer cells. In the present study, we investigated the effects of aqueous and ethanol extracts from two Native American plants, Ligusticum porteri (Osha) and Anemopsis californica (Yerba Manza), on the growth of human MCF-7/AZ breast and HCT8/E11 colon cancer cells. The aqueous and ethanol extracts from A. californica potently inhibited growth of MCF-7/AZ in a concentration-dependent manner, whereas the growth of HCT8/E11 was unaltered. Extracts from L. porteri showed no activity on either cell line. In addition, we observed that the extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) activities were markedly decreased when exposed to both extracts from A. californica. These results suggest that the growth inhibitory effect of A. californica in breast cancer cells is ERK-mediated.

This research compared raw filtered waters (RFWs), XAD resin isolates (XAD-8 and XAD-4), and reverse osmosis (RO) isolates of several surface water samples from McDonalds Branch, a small freshwater fen in the New Jersey Pine Barrens (USA). RO and XAD-8 are two of the most common techniques used to isolate natural organic matter (NOM) for studies of composition and reactivity; therefore, it is important to understand how the isolates differ from bulk (unisolated) samples and from one another. Although, any comparison between the isolation methods needs to consider that XAD-8 is specifically designed to isolate the humic fraction, whereas RO concentrates a broad range of organic matter and is not specific to humics. The comparison included for all samples: weight average molecular weight (Mw), number average molecular weight (Mn), polydispersity (rho), absorbance at 280 nm normalized to moles C (epsilon280) (RFW and isolates); and for isolates only: elemental analysis, % carbon distribution by 13C NMR, and aqueous FTIR spectra. As expected, RO isolation gave higher yield of NOM than XAD-8, but also higher ash content, especially Si and S. Mw decreased in the order: RO > XAD-8 > RFW > XAD-4. The Mw differences of isolates compared with RFW may be due to selective isolation (fractionation), or possibly in the case of RO to condensation or coagulation during isolation. 13C NMR results were roughly similar for the two methods, but the XAD-8 isolate was slightly higher in 'aromatic' C and the RO isolate was slightly higher in heteroaliphatic and carbonyl C. Infrared spectra indicated a higher carboxyl content for the XAD-8 isolates and a higher ester:carboxyl ratio for the RO isolates. The spectroscopic data thus are consistent with selective isolation of more hydrophobic compounds by XAD-8, and also with potential ester hydrolysis during that process, although further study is needed to determine whether ester hydrolysis does indeed occur. Researchers choosing between XAD and RO isolation methods for NOM need to consider first the purpose of the isolation; i.e., whether humic fractionation is desirable. Beyond that, they should consider the C yield and ash content, as well as the potential for alteration of NOM by ester hydrolysis (XAD) or condensation/coagulation (RO). Furthermore, the RO and XAD methods produce different fractions or isolates so that researchers should be careful when comparing the compositions and reactivities of NOM samples isolated by these two different techniques.

An iron flow-injection analysis system has been optimized for the analysis of iron in waters high in dissolved organic carbon. The method detects either dissolved iron(II) or total dissolved iron with a detection limit of 10 nM, precision of 0.65% at 1 microM, and a dynamic range of four orders of magnitude. There are minimal interferences (1%) from other metals at environmental concentrations. The iron(II) method measures iron(II) in the presence of excess iron(III) with less than 1% interference. When used with pre-acidified samples, the total dissolved iron method agrees well with electrothermal atomic absorption spectrometry for a variety of natural waters with a range of dissolved organic carbon (3-36 mg C/L) and iron (1-28 microM) concentrations. When used with samples at their ambient pH, the total dissolved iron method detects dissolved iron, but not colloidal iron (size fraction 0.05-0.45 micron).