Search

Reactive oxygen species (ROS) are thought to play an important, although poorly understood, role in modifying the composition of terrigenous-derived dissolved organic matter (tDOM) upon export to aquatic systems. Of the dominant ROS in natural systems, OH has been shown to substantially modify lignin under laboratory conditions, simulating processes responsible for transformation of DOM in natural waters. Utilizing high performance liquid chromatography (HPLC) in conjunction with ultrahigh resolution mass spectrometry, we examined HPLC fractionated lignin extracts in an effort to establish which fractionated components of the lignin extract were (i) most susceptible to attack by OH and (ii) most likely to be responsible for compounds observed in the DOM. The results indicate that the most polar components of the lignin extract, representing tDOM, could be responsible for a large portion of newly generated formulas observed within DOM in natural waters. This suggests that partially oxidized lignin may be readily altered to compounds observed in DOM which are not typically associated with terrigenous OM, such as alicyclic and condensed aromatic-like compounds.

It has been well accepted that humic acids in soils comprise both aromatic and aliphatic structural units that harbor mainly oxygenated functional groups such as carboxylic acids, aldehydes, ketones, and hydroxyl groups. Recent mass spectral data obtained from electrospray ionization-Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICRMS) suggest that the main functional groups in humic acids are carboxylic acids bound to both alicyclic and condensed aromatic molecules. It was suggested that these structural entities originate from oxidation of lignin. To provide additional support, detailed structural characteristics were obtained here using several multi-dimensional nuclear magnetic resonance (NMR) techniques to observe the molecular connectivity of humic acid components, with a focus on carboxylic acids in particular. The humic acid sample examined had been shown to exhibit structural characteristics similar to a larger set of soil humic acids. In this study, many carboxylic acid resonances were found to correlate with protons that could be assigned to aliphatic and alicyclic chemical shift regions. Functional groups with heteroatoms such as alcohols, some olefins and an assortment of methyl groups were found to correlate within 2–3 bonds of carboxylic acid groups. The collection of structural components and their proximity to a variety of carboxylic groups supports the occurrence of carboxylated alicyclic molecules as important elements of the aliphatic portion of soil humic acids and a pathway from lignin is consistent with recent findings.

Pyrogenic organic matter (Py-OM), generated via the incomplete combustion of biomass, is well studied due to the presence of slow-cycling, condensed aromatic compounds (ConAC) known to sequester in soils and sediments. Recently, dissolved Py-OM (Py-DOM) has received interest, due to its higher mobility and potential to be transferred through watersheds to aquatic systems. Py-DOM quantities, molecular identities and importance to global carbon budgets and cycles are only beginning to be understood. Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) analyses were performed on Py-DOM isolated from oak and grass biochars produced over a range of temperatures (250–650 °C), and the data are compared to complementary 1H NMR spectroscopic and benzenepolycarboxylic acid biomarker (BPCAs) analyses. Py-DOM is revealed to be a heterogeneous mixture of compounds ranging in relative aromaticity and oxygenation. FTICR-MS analyses demonstrate a decrease in Py-DOM relative oxygen content and a concurrent increase in aliphatic character and heteroatomic (N, Cl) content. 1H NMR and BPCAs analyses detail low O/C dissolved ConAC not observed by FTICR-MS, demonstrating the necessity for a multiple proxy approach to Py-DOM characterization. Heterogeneous Py-DOM is explained as resulting from pyrolysis-initiated and radical-mediated functional group cleavage, aromatic condensation and aromatic ring-opening reactions. Oak biomass progresses faster along a char maturity continuum for a given pyrolysis temperature, perhaps due to its greater lignin content or radical quenching by grass cuticular material. Biomass species and pyrolysis temperature likely result in different Py-DOM compositions and fates and must be considered when evaluating the impacts of wildfires and biochar applications.

Long-term stabilization of soil organic matter (SOM) plays an important role in the carbon cycle. Hence, understanding biogeochemical mechanisms of soil C sequestration is crucial to control its release to the atmosphere. This study aims at investigating the biogeochemical mechanisms of soil C sequestration. An exploratory assessment was carried out on the information about the soil C levels provided by the molecular composition of humic acids (HAs) analyzed by electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). Significant PLS forecasting model for total soil C was obtained using as descriptors the 131 compounds in common in all the HAs detected by FTICR-MS, and its variable importance for projection (VIP) was plotted in the space defined by their atomic ratios using van Krevelen diagrams. The results indicated that significant relationship exists between the HAs molecular composition and the soil organic C levels. The VIP values for the different groups of compounds illustrate how HA contains information about the amounts of C stored in the soil: the HAs in the soils with high levels of organic C have significantly (P < 0.1) higher proportions of unsaturated lipid and lignin-derived compounds; on the other side, low soil organic C levels are associated to HAs with comparatively high proportions of saturated lipid compounds., Financial support by Spanish CICyT (grant CGL2013-43845-P) is gratefully acknowledged. Marco A. Jiménez-González thanks the Spanish Ministry of Economy and Competitiveness (MINECO) for funding his pre-doctoral fellowship (BES-2014-069238).

Algae are regarded as the form of biomass most likely to provide sufficient quantities of fuels without impacting our food supplies. Studies investigating the potential of hydrothermal treatment of algae to produce biofuels show that, in many instances, the produced oils do not resemble crude oils and have a high heteroatom content. In this study, Scenedesmus spp. algae and isolated algaenan, a type of biopolymeric cell wall in certain algae and an important precursor to some kerogens, are subjected to hydrous pyrolysis in efforts to mimic the thermal maturation occurring in sediments as a proxy for biofuels production. Our study shows that algaenan can be subjected to hydrous pyrolysis to yield a hydrocarbon rich mixture that resembles many fossil fuel crude oils. More importantly, separation of the algaenan prior to the hydrothermal treatment can yield a paraffin rich crude requiring little additional processing to attempt to reproduce the geological process that gave us crude oils from ancient Type I kerogen. Although it requires algaenan isolation as a prerequisite, this could be a first step in the direction of producing oils without need for further upgrading. Whole algae, however, yield additional oxygenated products derived from oxygenated biopolymers even though the paraffins derived from algaenan dominate.

Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) can begin to tease apart the molecular character of sedimentary organic matter (SOM). We therefore tested five different solvents (aqueous base, CHCl3, MeOH, pyridine and water) for their ability to extract a representative fraction from two organic rich lacustrine sediments, Mangrove Lake, Bermuda (MLB) and Mud Lake, Florida (MLF). Following comparison using liquid state nuclear magnetic resonance spectroscopy (NMR) and negative ion mode electrospray ionization mass spectrometry (FTICRMS) we found that pyridine was the optimal solvent, extracting a more diverse (10–100× greater integration for carbonyl, amide and amine groups) and a larger number of peaks on average (1375–1450 vs. 380–1450). Comparison of the pyridine extracts between MLB, MLF and two organic poor sediments from the Mississippi River Delta and Bayou Grande (Pensacola, FL) showed that only 4.9% of the molecular formulae were common to all four and that unique formulae made up the highest proportion of the assignments. The use of pyridine for extracting immature (Holocene) SOM for FTICRMS analysis can therefore be widely applied to immature sediments and produce representative spectra.

Exposure of lignin-derived organic matter (OM) to hydroxyl radicals originating from Fenton type reactions generates condensed aromatic and alicyclic aliphatic compounds, as shown using ultrahigh resolution mass spectrometry. Although condensed aromatic compounds are common in soil and dissolved OM, their presence has been attributed largely to combustion. A non-pyrogenic route for the formation of condensed aromatic compounds from lignin is suggested here, specifically that hydroxyl radical-initiated oxidation of lignin is capable of producing black carbon-like condensed aromatic compounds. Alicyclic aliphatic compounds are also produced, likely as part of a concerted process involving ring opening, polymerization and/or cyclization and hydrogen abstraction. Hydroxyl radicals associated with lignin degradation are produced through photochemistry in aqueous systems and enzymatic microbial processes in soil.

The isolation of algaenan from its parent algae is the first step employed in the effort to characterize it. Three different protocols are the most followed in the isolation of algaenan. Because different structures have been proposed for algaenan from the same parent algae but isolated using different isolation protocols, we examined the possible effects the isolation procedure has on the algaenan chemistry. Our results indicate that the three different commonly used procedures have an impact on the structure of the recovered algaenan, because they involve different chemical treatments of varying strengths for the removal of non-algaenan components. The modifications to the overall similar dominant aliphatic composition include the formation of esters through transesterification, inclusion of proteins, destruction of crystallinity and hydrolysis of esters.

Although rates and mechanisms of early diagenesis have been well studied, the effects of microbial metabolism on the molecular composition of the sedimentary organic matter (SOM) over long periods of time need more investigation. In this study, we characterize the early diagenesis of marine SOM from organic rich sediments of the Ocean Drilling Program site 1082 located off Namibia, in the vicinity of the Benguela coastal upwelling system. We used both Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance spectroscopy ( 13 C NMR) to assess the quantitative partitioning of the organic carbon into major compound classes (aliphatic, aromatic, ester, carboxylic, amide and carbons from carbohydrates). Then, we calculate the SOM composition in the main biomolecules (proteins, carbohydrates, lipids and lignin) on the basis of previous 13 C NMR based estimates of the molecular composition of the organic mixtures. Results show that the SOM is still labile at 7 m below the seafloor (mbsf) and composed of about 25% proteins and 15% carbohydrates. With increasing depth, the protein content exponentially decreases to 13% at 367 mbsf, whereas the carbohydrate content decreases linearly to 11%. The lignin and lipid content consistently represent around 10% and 40% of the SOM, respectively, and show an increase with depth, due mostly to selective enrichment as the more labile components are lost by degradation. Thus, these components of the SOM are considered refractory at the depth scale considered. The calculated remineralization rates are extremely slow ranging from 5.6 mol C m −3 ky −1 at the top of the core to 0.2 mol C m −3 ky −1 according to the organic carbon flux to the seafloor. Knowing the labile carbon losses, we propose a method to calculate the initial TOC before the diagenesis took place.

Suberan is a biopolymer located in the periderm (cork) of above ground parts of plants or the endoderm of roots. It acts as a protective barrier between plants and the environment and is commonly associated with another suberinic biopolymer, suberin. The structure of suberan has not been suggested, but is believed to have structural components similar to suberin. With the use of destructive and non-destructive techniques, including Fourier transform infrared (FTIR) spectroscopy, flash pyrolysis–gas chromatography–mass spectrometry (py–GC–MS), one and two-dimensional high-resolution magic angle spinning (HRMAS) nuclear magnetic resonance (NMR) spectroscopy, the structure of suberan from the Betula nigra bark, is now described. Based on data obtained from B. nigra bark, the non-saponifiable residue, considered to be suberan, is a polymethylenic biopolymer with alkyl chain length consisting mainly of 22 carbons, which is consistent with previous degradative studies. Furthermore, esters, acids and primary alcohols were also observed in suberan, which contribute to a glyceryl structural entity. The long-chain polymethylenic structures demonstrate a high degree of crystallinity, providing a primary structural difference between suberan and suberin.

Gilsonite, a naturally occurring asphaltite bitumen, consists of a complex mixture of organic compounds. In the present study, advanced one and two dimensional solid state and solution 1 H, 13 C and 15 N nuclear magnetic resonance (NMR) and electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FT-ICR-MS) were employed to investigate its composition and structure. 13 C NMR yielded a carbon aromaticity of 27%. Aromatic moieties in gilsonite were primarily single rings or small clusters of fused rings. Half of the aromatic carbons of gilsonite can be accounted for by pyrroles. 15 N and 13 C cross polarization-magic angle spinning (CP-MAS) NMR showed that most nitrogen in gilsonite was pyrrolic. The aromatic rings were heavily substituted with alkyl chains, as evidenced by 1 H 13 C correlation spectra. Advanced solid state NMR spectral editing techniques clearly identified specific functional groups such as CCH 3 , CCH 2 , and C CH 2 (exomethylene). 1 H 13 C wideline separation (WISE) NMR helped identify mobile and non-protonated alkyl carbons. FT-ICR-MS indicated that ∼64% of calculated formulae generated by ESI were aliphatic, while only about 0.8–2.5% of formulae contained possible aromatic rings. All of the assigned formulae contained at least one heteroatom (N, O or S), indicating that ionization by ESI was selective for the polar fraction of gilsonite and potentially less reflective of the overall chemical character of gilsonite than NMR spectroscopy. By combining the information obtained from advanced NMR and ultrahigh resolution MS we propose a structural model for gilsonite as a mixture of many pyrrolic and a few fused aromatic rings highly substituted with and connected by mobile aliphatic chains.

Molecular level characterization of complex biopolymers in nature is a key element to understanding the composition of natural organic matter (NOM) and fossil organic matter formation, such as kerogen and coal. Characterization of such geopolymers is difficult due to their heterogeneity and insolubility in common aqueous or organic solvents. Here, a strategy for extracting a representative sample is presented using the combined techniques of nuclear magnetic resonance (NMR) and Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) for chemical analysis and characterization of NOM. A variety of NOM samples (wood, kerogen, bitumen, whole sediments) are shown as examples for implementing the strategy, which include solvent extractions using pyridine. For most samples, the extracts are confirmed to be chemically representative of the insoluble solid, by comparing the liquids NMR spectrum of the extract to that of the whole, unfractionated NOM utilizing high resolution magic angle spinning (HRMAS) NMR. To assist in unambiguous peak assignment, a technique for post acquisition spectral denoising, using wavelet transformation (WT), is also employed on the 1D and 2D NMR spectra. The findings from the NMR data lay a foundation for the subsequent electrospray ionization FTICR-MS analysis of the pyridine extracts, since this instrument has the limitation of only analyzing liquid samples. The FTICR-MS analyses can, therefore, be used to molecularly represent the structural components of the NOM. Various compositional insights have been obtained on these samples, indicating the efficacy of the analytical techniques used.

This study describes a new approach for characterizing high molecular weight compounds in Type I kerogen, involving both nuclear magnetic resonance (NMR) spectroscopy and Fourier transform ion cyclotron mass spectrometry (FTICR-MS). Kerogen isolated from the Mahogany zone of the Green River Formation was examined directly using high resolution magic angle spinning (HRMAS) NMR to obtain liquid-like multidimensional spectra. It was then successively extracted with n-pentane, dichloromethane and pyridine. Pyridine extraction was also performed for comparison with the successive extractions. Using solid-state NMR, we show that the sum of the successive extracts and the single pyridine extract are quantitatively representative of the unextracted kerogen. This suggests that a non-invasive characterization of Green River kerogen can be obtained by examining the soluble extracts, all of which were subjected to ESI-FTICR-MS to identify a wide series of compounds. Series of polar CHO, CHOS and CHON compounds between C12 and C50+ were found. In all the extracts the two homologous series of acids (CnH2nO2 and CnH2n−2O4) dominated. Collision-induced dissociation was also employed to identify the different functional groups comprising the different series. The CHO series contained carboxylic acid and alkoxyl groups, whereas the CHOS and CHON series contained sulfoxide groups and nitrile-type compounds. The results also show that pyridine extraction can be used either for screening a large series of samples or for the specific study of CHO compounds. However for a detailed and complete study of the different homologous series we recommend using the successive extraction protocol.

An algal culture of Botryococcus braunii race A and its residue obtained after 201 days of oxic biodegradation are characterized using high resolution magic angle spinning (HRMAS) NMR spectroscopy, a hybrid technique which provides molecular details non-invasively. The high resolution of two dimensional NMR using HRMAS gives precise structural assignments of different classes of compounds contained in the initial algae and biodegraded algae. The data indicate that some intracellular lipids and proteinaceous materials are more environmentally labile while other long chain polymethylenic structures with ether functional groups, likely from algaenan, are observed to survive the degradation process. The identification of similar structural entities in both fresh and degraded material clearly points to selective preservation as the main process responsible for the early diagenetic transformation of this type of algae.

The thermal reactivity of organic matter in source rocks is usually kinetically represented by a set of parallel and independent first order reactions. This approach assumes that only defunctionalization reactions take place upon thermal decomposition, regardless of the chemical nature of kerogen. We have developed a new method for evaluating maturation pathways for an important kerogen-forming geopolymer, algaenan, involving molecular dynamic reactive modeling based on quantum mechanics to reproduce maturation. To achieve this, a structural model is first constructed based on models from the literature and analytical characterization of our samples using modern 1D and 2D nuclear magnetic resonance spectroscopy (NMR), fourier transform infrared spectroscopy (FTIR) and elemental analysis. Then, thermal decomposition of the algaenan is performed at low conversion in order to describe the initial transformations analytically. In an additional step, the observed chemical changes are quantitatively and qualitatively compared to simulated maturation from the molecular models. From this simulated maturation detailed reaction schemes are extracted for primary cracking mechanisms

A major obstacle for characterizing dissolved organic matter (DOM) with ultrahigh resolution mass spectrometry has been its low concentrations in natural waters. Many previous mass spectrometric studies of both terrestrial and marine DOM typically have isolated and concentrated the DOM using solid phase C18 extraction disks, ultrafiltration, or XAD resins, all of which are known to discriminate against many different classes of compounds. We have, for the first time, developed an approach to directly analyze natural water samples with less than 6 mg/l DOC (dissolved organic carbon), using electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS). We demonstrate the sensitivity and ability of sequential selective ion accumulation (SSIA) to detect the thousands of components in a single freshwater DOM sample without any significant pretreatment. By utilizing SSIA, the baseline noise decreases while signal to noise ratios of the peaks increase, allowing for approximately 40% more formulas to be assigned to peaks in the mass spectra.

Covalent and non-covalent interaction of proteinaceous materials in soils and sediments has been suggested as an important mechanism for immobilizing nitrogen in numerous types of environments. In a previous study (Hsu P.-H., Hatcher, P.G., 2005. New evidence for covalent coupling of peptides to humic acids based on 2D NMR spectroscopy: A means for preservation. Geochimica et Cosmochimica Acta 69, 4521–4533), we provided molecular evidence for covalent, as well as non-covalent, bonding between 15N-labeled peptides and humic acid molecules using the 2D HSQC (heteronuclear single quantum coherence) NMR technique. In this report, we examine the influence of aromaticity and aliphaticity of peptides and humic materials on these covalent and non-covalent interactions. We use 2D NMR techniques to evaluate bonding interactions of 15N labeled peptides, having different aromatic and aliphatic properties, with three humic acids that vary in degree of aromaticity. The peptide containing primarily aromatic amino acid residues is observed to form covalent and non-covalent bonds with mainly aromatic-rich humic acids. The peptide composed of aliphatic amino acid residues shows, on the other hand, only bonding interactions with aliphatic-rich humic acids. These observations provide the first direct molecular evidence that aliphatic functional groups are involved in bonding with proteinaceous materials. The process may play an important role in sequestration of proteinaceous materials in sedimentary systems such as marine systems where the humic materials are mainly aliphatic in nature.

The molecular composition of water-soluble products generated by the natural degradation of charcoal particles over a period of 100 years in a temperate forest soil has been investigated by ultrahigh resolution mass spectrometry with electrospray ionization. The detectable products are condensed aromatic ring structures extensively substituted with oxygen-containing functional groups, indicating that oxidation and dissolution of charcoal black carbon occurs on a centennial timescale. Many of the same species are also detected within the dissolved organic matter (DOM) of the forest’s soil pore waters. We introduce the calculation of carbon normalized double bond equivalents (DBE/C) as a structural determinant for the empirical formulas obtained by mass spectral analysis. A threshold DBE/C value of 0.7 serves as a criterion for identifying species with condensed aromatic ring structures (CARS). A comparison with ultrahigh resolution mass spectra from previous studies shows that many of the CARS extracted directly from soil BC have the same mass (within 1 ppm) and empirical formulas as CARS detected in volcanic ash soil humic acid (HA) from Japan, and Amazonian Rio Negro DOM. The similarity of water-soluble condensed aromatics present within, and exported from fire-impacted soils of geographically and climatically disparate ecosystems indicates that the CARS reported herein are the molecular fingerprint of black carbon degradation in soils. Understanding the production mechanisms, reactivity, and fate of these molecular species should provide new insight to BC degradation and cycling. The soil charcoal particles at this site are infiltrated by filamentous microorganisms, suggesting that saprophytic fungi may be important to soil BC degradation processes.

Charcoal of Eucalyptus grandis and Eucalyptus saligna was produced with different final charring temperatures. Ground charcoal samples (120 mesh) were treated with HNO3 25% (w/w) during 4 h under reflux conditions. Humic (HA) and fulvic acids (FA) were extracted from the reaction products utilizing the standard method recommended by the International Humic Substances Society (IHSS). The HA and FA yields were obtained by quantifying carbon in each fraction. The highest HA yields were obtained at 450 °C final charring temperature (430 mg C-HA g−1charcoal), and the highest FA yields at 300–350 °C (310–304 mg C-FA g−1 charcoal). Elemental analysis, thermogravimetry, FTIR and 13C NMR were used for HA characterization. Significant chemical modifications were observed after acid oxidation, such as increments in nitrogen and oxygen contents, and formation of carboxylic groups. Based on the characteristics of the obtained products, we conclude that charcoal synthetic HAs and natural pyrogenic derived HAs have similar characteristics.

The structure of cutan, and a cutin/cutan mixture from the Agave americana leaf cuticle is herein described by use of the technique of one and two-dimensional high-resolution magic angle spinning (HRMAS) NMR spectroscopy, with added information from solid-state 13 C cross-polarization magic angle spinning (CPMAS) and Bloch decay NMR spectroscopy. Cutin in the cutin/cutan mixture is found to contain ester functionalities, in line with previous degradative approaches. Furthermore, epoxy groups, free primary alcohols and carboxylic acid groups are also observed in cutin. A significant new finding is the evidence for α-branched fatty acids/esters in cutin. Cutan has mainly free primary hydroxyls, as well as long-chain carboxylic acids, which have been previously predicted to form ester linkages with trihydroxylated benzene units. Another significant finding in cutan is the evidence for benzenecarboxylic acids that are ester linked with fatty alcohols. The long-chain polymethylenic groups in cutan are shown to express a high degree of crystallinity similar to that observed in soil organic matter.

Black carbon is an important component of natural organic matter that may play a significant role in global climate change. However, the complexity of natural organic matter has resulted in a milieu of methods and accompanying black carbon values that make it difficult to delineate the proportion of black carbon in soil and sedimentary total carbon budgets. Here we report on a chemical oxidation method that removes lignin and then facilitates black carbon to be estimated from solid-state 13C Nuclear Magnetic Resonance (NMR) spectroscopy. The resulting black carbon values are lower in comparison to those reported by other researchers but may be a more accurate method for estimating black carbon in a range of natural organic matter samples from different environments.

We propose and demonstrate an approach involving use of C18 solid phase disk extraction coupled with high resolution mass spectrometry for obtaining non-invasive molecular level information on dissolved organic matter (DOM) from river water. With this approach, DOM extraction from acidified natural water can be achieved rapidly with a simple filtration setup at a remote field site. From total organic carbon and UV-Vis absorbance measurements, we show that a large portion (over 60%) of the original DOM in water is recovered without the interference of salts. NMR analysis indicates that the C18-isolated material has a similar distribution of functional groups as the original DOM but 2-D NMR details are greatly enhanced. Electrospray ionization mass spectrometry and high resolution mass spectrometry were employed to study DOM at molecular level. Highly resolved mass spectra of DOM (resolving power δ m m 50% > 80,000 at m/z

Botryococcus braunii cells were grown until the late-stationary phase of growth and subsequently decomposed under oxic conditions for 201 days using a microbial consortium obtained from a freshwater lake.Degradation exhibited multi-G model kinetics, with a ‘labile’ fraction lost at a rate two to three times slower than those observed for the degradation of other previously studied phytoplankton, and a ‘refractory’ fraction lost even more slowly.Scanning electron microscopy of the 201-day detritus, as previously seen for the kerogen, indicates the preservation of cell wall material with loss of intracellular contents.Detrital samples analyzed by pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS) and solid-state ramp-CPMAS 13 C NMR, however, indicates the preservation of highly aliphatic material, algaenan, as well as ‘intrinsically labile’ proteinaceous components.These results further support the encapsulation hypothesis that proteins may be sterically protected from enzymatic attack via intimate associations with refractory, macromolecular organic matter. # 2003 Elsevier Science Ltd.All rights reserved.

Solid-state CP–MAS 13C and 15N nuclear magnetic resonance (NMR) spectroscopy and flash pyrolysis–gas chromatography–mass spectrometry (Py-GC–MS) techniques have been used to characterize the organic-rich sedimentary deposit from Mangrove Lake, Bermuda. The chemical nature of organic nitrogen in sediment samples from different depths was studied as it relates to the early diagenesis of proteinaceous material in the reducing marine environment. Although the majority of proteins were lost during early diagenesis, part of the proteinaceous material became resistant to proteolysis and chemical hydrolysis. The proteinaceous material is proposed to have survived early diagenesis via encapsulation by refractory, aliphatic organic matter.

A molecular and isotopic study of sediment cores from a sinkhole lake, Mud Lake, Florida, USA, was performed in order to relate documented changes in the regional terrestrial vegetation and water table over the last ∼5500 years to molecular and isotopic proxies for biological sources of organic matter to the lake sediments. Temporal shifts in the source of organic matter to the sediment, as determined by the stable carbon-isotope composition of bulk organic matter and biomarkers, correspond with previously defined regional scale transitions in forest ecosystems ( Quercus to Pinus at ∼5500 14 C yr BP and Taxodium expansion at 2500 14 C yr BP) and coincident increases in the Floridan water table. The δ 13 C values for total sedimentary organic carbon showed a shift from terrestrial and aquatic macrophyte sources (−27.8‰, in sediments dated at ∼5400 14 C yr BP) within the basal peat, to a cyanobacterial-dominated sapropel (−18.1‰) at the surface. A comparison of the δ 13 C values of bulk sediment and biomarkers representative of cyanobacterial and algal input (e.g. 7- and 8-methylheptadecane and the n -alkane C 17 ) indicates that the present shallow lake was fully developed by ∼2400 14 C yr BP. Differences among the δ 13 C values of specific biomarkers derived from vascular plants (C 29 and C 31 n -alkanes and CuO lignin oxidation products) and from cyanobacteria and algae are nearly equivalent in magnitude to the shift recorded in δ 13 C of TOC, indicating their effective use as source proxies in this system.

12 pages, 5 figures, 4 tables, 45 references., Humic isolates from different layers of an organic-matter-rich algal sapropel from Mangrove Lake, Bermuda, were subjected to solid-state 15N nuclear magnetic resonance (NMR) spectroscopy in order to reveal chemical structural interrelationships that allow delineation of diagenetic pathways. Amide-N, most likely from peptide-like material, was found to represent the main organic nitrogen form in all humic fractions at all depths of the sapropel. Amides were also identified in the residues from 6 M HCl hydrolysis of the alkaline insoluble extract (formerly called humin). These results indicate that some peptide-like material resists both alkaline and acid hydrolysis. We suggest that the chemical, and also the biological recalcitrance of peptide-like material, is related to encapsulation into the paraffinic network of algaenans. Being hydrophobic in nature, algaenan biomacromolecules may represent an effective means of protection against the attack of hydrophilic enzymes and aqueous extraction solutions.

Thermochemolysis with tetramethylammonium hydroxide (TMAH) was utilized to analyze peptide-like material in insoluble residues of geochemical samples. Product identification was performed by gas chromatography/mass spectrometry using bovine albumin and di-, tri- and oligo-peptides. Most of the identified amino acid derivatives originate from the cleavage of the peptide bonds and subsequent methylation of the carboxylic and the amino groups. Some products are explained by deamination of aromatic amino acids. Only two products were identified that experienced chemical rearrangement after methylation. TMAH/thermochemolysis of algal material resulted in products comparable to those obtained from albumin. Amino acid derivatives were also identified among the TMAH/thermochemolysis products of refractory biopolymer of the alga Scenedesmus communis and the HCl-hydrolysis residue of the humin from an algal sapropel. These results strongly indicate that for those samples, hydrolysis fails to extract all proteinaceous and peptide-like components, possibly because these approaches rely on extraction of peptides and amino acids into solution and some of these are entrapped within a non-extractable hydrophobic network. However, alternative explanations for this behavior are possible. For example, at the higher temperatures and pressures and strongly basic conditions used in TMAH themochemolysis, penetration of this hydrophobic barrier takes place thus allowing breakdown and methylation of protein remnants.

Molecularly-uncharacterized organic matter comprises most reduced carbon in soils, sediments and natural waters. The origins, reactions and fates of these ubiquitous materials are relatively obscure, in large part because the rich vein of geochemical information that typically derives from detailed structural and stereochemical analysis is yet to be tapped. This discussion highlights current knowledge about the origins and characteristics of molecularly uncharacterized organic matter in the environment and outlines possible means by which this structurally uncharted frontier might best be explored.

The Ems-Dollart estuary (on the border of the Netherlands and Germany) was chosen for a pilot study to characterize ultrafiltered dissolved organic matter (UDOM) in estuarine systems. UDOM samples were taken from four locations with salinities varying from 0.43 to 20 parts per thousand. The UDOM in these samples was concentrated using cross-flow ultrafiltration and represents the size fraction of DOM between 1000 Dalton and 0.2 mu m. The samples were analyzed by analytical pyrolysis, stable carbon isotope and radiocarbon analysis, and solid state C-13 NMR, All UDOM samples throughout the estuary showed relatively similar pyrolyzates and C-14 activities (c. 87%). Solid-state C-13 NMR spectra were also similar, except for the sample with the highest salinity. UDOM delta(13)C values ranged from -27.78 to -25.40 parts per thousand with increasing salinity. This suggests mixing of river DOM with seawater DOM of comparable ages. The absence of pyrolysis products of unaltered polysaccharides, lignins, proteins and lipids indicates that the Ems-Dollart estuary UDOM consists of a large fraction of refractory organic matter and a very small fraction of fresh organic matter. (C) 2000 Elsevier Science Ltd. All rights reserved.

Laser micropyrolysis GC–MS was used for in situ analysis of the coal and lignin components of a synthetic mixture. Designed to mimic environmental matrices such as soils and sediments, the mix comprised several possible soil precursors and was also amended with [4,9- 13 C]pyrene as part of concurrent research on the interaction of PAH pollutants and sedimentary organic matter. The labeled spike was consistently detected as the major pyrolysate in the in situ analyses of both lignin and coal components of the synthetic mix, indicating its effective sorption by these moieties of the mix. The remaining hydrocarbon distribution detected from the lignin was dominated by guiaicyl (i.e. methoxyphenol) compounds, whereas high abundances of aromatic (e.g. benzene, naphthalene, phenol and alkyl derivatives thereof) and aliphatic (e.g. n- alkene/alkane, prist-1-ene, hopanes) products were detected in the coal. Apart from the high concentrations of the 13 C-spike, these data were very similar to molecular data obtained from the respective pyroprobe pyrolysis GC–MS analysis of pure lignin and coal samples. The untainted (i.e. apart from the 13 C-spike) molecular signatures detected from the in situ analysis of the coal and lignin constituents indicates minimal organic contamination from the other constituents of the synthetic mix, successfully demonstrating the capability of the laser micropyrolysis GC–MS technique to selectively analyse the discrete organic entities within complex and heterogeneous mixtures.

Recent work suggests that nitrogen in humic acids exists primarily as amide functional groups that mirror those in protein. However, the mode for the existence of such labile materials as protein still remains unclear. With the combined applications of NMR spectroscopy, tetramethylammonium hydroxide (TMAH) thermochemolysis, pyrolysis/GC/MS, and elemental analysis, we propose that the survival of proteins in humic acids is carried out by encapsulation into hydrophobic domains of humic acids. To test this hypothesis, we simulated encapsulation of 15N-labeled protein extracts into humic acids and demonstrated that complete hydrolysis of the protein is prevented by the encapsulating humic acid. Results from this study constitute evidence to support the encapsulation mechanism involved in the formation of refractory organic nitrogen during sediment diagenesis.

This paper presents the results from an assessment of the application of a new molecular analytical procedure, 13 CTMAH thermochemolysis, to study the chemical modification of lignin by white-rot and brown-rot fungi. This technique diAers from other molecular chemolysis procedures (e.g. TMAH thermochemolysis and CuO alkaline oxidation) as it enables one to determine the amount of hydroxylated aromatic components in degraded lignin residues through a selective lignin depolymerization and 13 C-labeled methylation reaction. Major diAerences were observed in the chemical composition and yield of lignin monomers released from a limited sample set of field and laboratory inoculation brown-rot and white-rot degraded residues when analyzed by 13 C-TMAH thermochemolysis. The brown-rot residues were characterized by high yields of 3,4-dihydroxy phenyl compounds, presumably due to fungal demethylation of methoxyl groups on guaiacyl lignin, and relatively low yields of aromatic acids that result from microbial side chain oxidation. The white-rot residues were characterized by low yields of demethylated lignin monomers but relatively high yields of monomers exhibiting side chain oxidation. If generally applicable, this distinct chemical functionality has important implications for the chemical reactivity and solubility of degraded wood residues and consequently the cycling of terrestrial carbon in the geosphere. The 13 C-TMAH thermochemolysis procedure provides a rapid and sensitive tool for tracking microbial modifications of lignin in terrestrial environments including coastal sediments, forest soils and waters receiving terrestrial organic matter. # 2000 Elsevier Science Ltd. All rights reserved.

Soluble polylabdanoids were isolated from five Tertiary Class I resinites by sequential extraction and were characterized by 13 C and 1 H NMR spectroscopy and Py–GC–MS. The structure and maturation characteristics of soluble extracts were strikingly similar to those observed for polylabdanoids in the native resinite. Soluble and insoluble materials undergo parallel geotransformation processes similar to those observed previously, including (i) exomethylene and total olefinic carbon depletion, (ii) double bond redistribution as indicated by ∑C14/∑C15 pyrolysis product ratios and (iii) A-ring defunctionalization. Resonances at 138 and 127 ppm in 13 C NMR spectra were shown to be those of aromatic and olefinic structures in mature samples, indicating that double bond content had been overestimated in the past. Proton NMR analyses have revealed several structural features previously unobserved. Resonances at 5.3 ppm of mature polymers were assigned to trisubstituted olefins in cyclized/cross-linked polylabdanoids. Further evidence in support of this pathway is derived from 13 C NMR spectra, from which an increase in aliphatic carbon content paralleling the loss of olefinic structures was observed.

The mechanism by which heated tetramethylammonium hydroxide (TMAH) degrades the lignin biopolymer was investigated by the novel application of 13C-labeled TMAH (13C-TMAH) in the thermochemolysis of a synthetic model guaiacyl lignin dimer. GC-MS analysis of the products showed labeling patterns consistent with a base-catalyzed intramolecular displacement of the β-phenoxy group and the formation of two intermediate guaiacyl propane epoxides, a γ-hydroxy-α,β-epoxide and an α-hydroxy-β,γ-epoxide. Methoxide then functions as a nucleophile to open the epoxide ring. These results substantiate the base-catalyzed reactions previously postulated by Gierer (1970) to explain alkali wood pulping and also explain the facile formation and distribution of lignin derivatives obtained in the TMAH thermochemolysis of natural samples. The absence of substantial numbers of bonds involving propyl-aryl ether linkages with adjacent hydroxyl groups is the limiting factor in the complete decomposition of lignin by TMAH thermochemolysis, as propyl-aryl ether linkages without adjacent hydroxyl groups cannot react via this mechanism. This helps to explain why materials such as highly degraded lignin residues, with significant side chain alteration and type III kerogens above the rank of lignite, where aliphatic-aryl ether linkages are thought to be insignificant, are reported to give low yields of TMAH thermochemolysis products.

Selected aliphatic biopolyesters (cutins, cuticles and a suberin) isolated from different plants have been analyzed using thermochemolysis with tetramethylammonium hydroxide (TMAH). This method consists of a high-temperature saponification/transesterification, and yields methyl esters of fatty acids and the methyl ethers of alcohols, which are subsequently analyzed by gas chromatography and gas chromatography–mass spectrometry. The main compounds produced from the analyzed samples correspond to the methyl derivatives of long-chain fatty acids, hydroxy fatty acids and α,ω-alkanedioic acids. The composition of the released compounds are similar to those reported in the literature using different depolymerization methods. The main advantage of the procedure is that it is easily performed in glass tubes with very low amounts of sample and without additional derivatization steps prior to gas chromatographic analysis because the products are methylated in situ. The method also avoids the laborious and time consuming sample preparation of extractive methods and the use of large amounts of solvents.

Select ion monitoring/gas chromatography/mass spectrometry (SIM/GC/MS) in conjunction with 13C-labeling has been utilized in order to monitor the thermal degradation of 13C-labeled n-C25 mixed with a typical marine derived crude oil from the Arabian peninsula and an oil derived from terrestrial source rocks in the Ardjuna Basin. Pyrolyses were carried out in anhydrous closed gold vessels at times ranging from 1–360 h under isothermal conditions (350–425°C) at a pressure of 120 bar and the degradation of the labeled n-C25 followed first order kinetics in the two oil systems. For each temperature regime, the rate of 13C-labeled n-C25 decomposition was suppressed in comparison to the neat system with the Arabian Light marine oil exerting a greater influence on the n-C25 thermal stability. It was determined that the activation energies for the n-C25 compared favorably to the neat pyrolysis experiments [Ea(marine oil) = 67.5 kcal/mol, Ea(terrestrial oil) = 67.9 kcal/mol and Ea(neat) = 68.2 kcal/mol]. However, the pre-exponential factor obtained for the n-C25 decomposition varied by as much as a factor of five in the two oils in comparison to the neat pyrolysis experiments. An extrapolation of these kinetic parameters to geologic conditions shows that the n-C25 will be more thermally stable in high temperature reservoirs than previously anticipated. Using a similar procedure as was used on the quantification of the 13C-labeled n-C25, n-alkanes, the major products stemming from the degradation of the 13C-labeled n-C25, containing enrichments in 13C were quantified by SIM/GC/MS, as well. In monitoring and quantifying the overall enrichments of individual n-alkanes in the oils, it was shown that the mechanism for the thermal degradation of 13C-labeled n-C25 is similar between the neat system and the Ardjuna oil, but differs somewhat in the Arabian Light oil.

The cell walls of the fresh water green microalgae Tetraedron minimum, Scenedesmus communis and Pediastrum boryanum are composed of highly resistant, non-hydrolyzable aliphatic biopolymers as revealed by 13C-NMR, FTIR and thermal and chemical degradations. The biopolymers are composed of long-chain even-carbon-numbered ω9-unsaturated ω-hydroxy fatty acid monomers varying in chain length from 30 to 34 carbon atoms. These monomers are intermolecularly ester linked to form linear chains in which the unsaturations act as the starting position of ether cross-linking. S. communis biosynthesises a more densely cross-linked algaenan than T. minimum and P. boryanum. The monomers of T. minimum have, on average, larger chain lengths than those of P. boryanum and S. communis. The polyether nature of these algaenans makes them highly resistant against degradation, so that they are selectively preserved in the sedimentary record. Therefore, these algaenans probably are important precursors for Type I kerogens.

Soluble polylabdanoids isolated by sequential solvent extraction have been characterized by liquid-state {sup 13}C- and {sup 1}H NMR and {sup 13}C-{sup 1}H HMQC (heteronuclear correlation) NMR spectroscopy in addition to solid-state NMR and Py-GC-MS techniques. Two Holocene resins originating from Santander, Colombia and Mombasa, Kenya were analyzed. Soluble polymers were isolated by extraction with a 1:1 (v/v) methylene chloride-methanol mixture following sequential extractions with methylene chloride and methanol. The molecular weight of polymer extracts was shown by GPC analyses to exceed that of non-polymeric occluded terpenoids. Py-GC-MS, solid-state {sup 13}C CP/MAS and {sup 13}C cross-polarization/depolarization NMR spectroscopy results indicated that chemical compositions of soluble polymers isolated from immature resins are highly representative of the structure of corresponding insoluble polymers, i.e. polylabdatrienes. These data provide evidence for cross-linking or cyclization of side-chain olefinic carbons during or shortly after polymerization. Generally, the characterization of soluble resin polymers by liquid-state NMR spectroscopy has proven to be an excellent means for investigating the maturation mechanism of polylabdanoid resinites, and has potential for furthering the application of Class I resinites as geothermal indicators.

Coalification is a process that transforms plant remains under the influence of time, temperature, and possibly pressure to a black, generally lustrous solid organic fossil fuel having a very complex chemistry. Although much has been learned in the past two centuries on coal chemistry, we still have little knowledge of the way plant materials undergo these transformations. The problem has been the existence of adequate characterization tools. The advent of some new tools for structural elucidation of macromolecules has led to some revised thinking on coalification. Originally thought to be a process involving the full degradation of plant remains and subsequent reconstitution of these remains, the general consensus today is that the process involves a selective preservation of certain resistant plant components followed by some minor reorganization of the biopolymers that survive. The major components of plants do degrade, and the resistant ones often constitute only a small fraction of the original mass of the plant materials. This paper examines the most recent literature concerning coalification and focuses on the chemistry associated with coalification of certain recognizable plant remains that have been at the center of research activity for the past two decades. Thus, we examine the recent knowledge of the chemistry of resistant biopolymers in extant and fossil wood, cuticles, resins, spores, and algae, knowledge that has been assembled to establish the reaction pathways towards coal macromolecules.

6 pages, 2 figures, 2 tables, 23 references., Ultrasonic dispersion of geochemical samples suspended in water and subsequent homogenization by ball milling is widely used for fractionation of organic matter. The effect of these treatments on organic matter is investigated with lignin as a model compound. Structural alterations detectable by solid-state 13C nuclear magnetic resonance (NMR) spectroscopy were examined. Comparison of the solid-state 13C NMR spectra of untreated lignin and lignin mixed with quartz or soil did not reveal evidence for structural changes in the organic matter composition after ultrasonic dispersion and subsequent ball-milling. The chemical structure of organic matter in geochemical samples is not affected by these treatments as far as such structural alterations can be detected by solid-state 13C NMR spectroscopy., This work was financially supported by the Deutsche Forschungsgemeinschaft (Ko 1035/6-land 2) and the Deutscher Akademischer Austauschdienst (Ref. 315, D/94/16993).

The influence of brown coal emissions from a briquette factory on the organic matter of a Mollisol has been investigated. Non-contaminated, heavily contaminated soils and airborne particles are compared by using chemical and spectroscopic methods (13C CPMAS-NMR, Py-GC-MS, 14C dating). Bulk soils as well as their particle-size fractions are investigated. Organic carbon content of the bulk contaminated soil (138.6 g C/kg) is higher by a factor of six than the non-contaminated soil. For particle-size fractions, the highest content of organic carbon is found in the fine sand fraction (347.2 g C/kg). Furthermore, organic matter of the contaminated site shows a trend for relatively higher abundance of aliphatic compounds in the fraction ranging from 6 to 2000 μm, whereas for particles

13C and 15N NMR spectroscopy was applied to modern (a mixed algal culture, its algaenan and its compost), ancient (algal derived sediments from Mangrove Lake, Bermuda) and fossilized algal residues (Torbanite, Green River Shale) for the purpose of establishing the forms of nitrogen algal remains and evaluating their long-term stabilities. The results indicate that proteinaceous material can resist microbial degradation in sediments as old as 4000 yr, possibly in refractory biopolymers, by encapsulation into their macromolecular network. The 15N NMR spectra of the Torbanite and the Green River Shale show a relative enrichment of heterocyclic-N, which may derive from selective preservation of biogenic heterocyclic compounds or by rearrangement of peptide chains during diagenesis.

Tetramethylammonium hydroxide (TMAH) thermochemolysis has been applied to the diagenetically resistant aliphatic biopolymer cutan derived from the leaf cuticles of Agave americana. Data obtained from the cutan sample using this new analytical method are vastly different in comparison with conventional flash pyrolysis data of the same sample. Under conventional flash pyrolysis conditions, the dominant products were alkanes, alkenes, and α, ω-alkadienes. Under thermochemolysis conditions, however, the cutan biopolymer yielded fatty acid methyl esters (FAMEs) of varying carbon chain length, along with a large number of benzenecarboxylic acid methyl esters that point to the possibility of a chemical structure containing functionalized benzene rings. This type of structure is absent from the current structural model of cutan.

When analyzed by the TMAH thermochemolysis procedure and solid-state 13C NMR, DHP guaiacyl lignin is found to yield products which are somewhat similar to those observed in fresh or degraded gymnosperm lignin. The relative amounts of the various TMAH products, particularly the stereoisomeric pairs of 3,4-dimethoxyphenyl-trimethoxypropane, however, are not identical, perhaps indicating that disordered and randomly polymerized DHP lignin is not similar to a possibly more ordered network in natural lignin.

The presence of benzenecarboxylic acid methyl esters in thermochemolysis or pyrolysis products of humic acids, produced in the presence of tetramethylammonium hydroxide (TMAH), has recently been attributed to the existence of benzenecarboxylic acid moieties in the original materials being examined by this new technique for structural elucidation. Our studies of wood and model compounds, materials free of carboxylic acid-containing structures, by this method indicate that certain benzenecarboxylic acids are produced by the TMAH reagent in the course of reaction at elevated temperatures. This indicates that one should exercise due caution in interpreting the presence of these benzenecarboxylic acid methyl esters as pre-existing structural humic acids.

Comparison of the new TMAH thermochemolysis procedure with the traditional CuO oxidation procedure for lignin characterization in gymnosperm wood and soil formed from degraded gymnosperm wood indicates that an excellent agreement exists between the two methods, especially with regard to the yield of lignin monomers. Moreover, the TMAH procedure appears to be more sensitive for calculation of benzenecarboxylic acid/benzaldehyde ( Ad Al ) ratios, displaying a larger dynamic range than that observed by CuO oxidation. If one considers the simplicity with which the TMAH procedure is carried out, this technique is much more suited to routine analyses of large numbers of samples.

Three polylabdanoid resinites ranging in age from Eocene to Miocene were analyzed by pyrolysis-gas chromatography-mass spectrometry and solid-state nuclear magnetic resonance spectroscopy. The goal of the study was to determine effects on the chemical composition of polylabdanoid resinites following maturation processes. The samples studied included two Victorian Brown Coal resinites from the Yallourn and Morwell coal seams in Victoria, Australia, and a resinite from the Brunner coal measure in Nelson, New Zealand. Results of CPMAS 13 C NMR experiments have determined a progressive decrease in total olefinic carbon content (per diterpenoid) with increasing maturity. Olefin reduction occurs primarily via a diminution in exocyclic methylene carbons of the labdatriene precursors (i.e. communic acid, communol, and/or biformene and their analogs). Analysis of Soxhlet extracted samples by py-GC-MS has shown a transformation from pyrolysis products indicative of communic acid polymers (hydronaphthenic acids) to a pyrolyzate containing a significant proportion of alkyl(hydro)naphthalenes. Condensation of the resinite polymer occurring in conjunction with defunctionalization reactions has been proposed.

Flash pyrolysis in the presence of tetramethylammonium hydroxide (TMAH) has been utilized for the characterization of lignin in a gymnosperm (pine), angiosperm (alder), and nonwoody ( Juncus effusus L.) vascular plant tissue. The gymnosperm tissue yielded primarily methylated guaiacyl derivatives resulting from cleavage of lignin β-O-4 bonds and subsequent methylation of acidic hydroxy groups. The angiosperm tissue also yielded guaiacyl-type compounds in addition to an analogous suite of methylated syringyl lignin derivatives. The nonwoody angiosperm yielded methylated cinnamyl lignin derivatives in addition to methylated guaiacyl and syringyl compounds. Analysis of alder wood with this technique at a low temperature of 310°C has demonstrated that the dominant reaction with TMAH is mainly a thermally assisted chemolytic degradation rather than degradation induced by pyrolytic bond cleavage. Accordingly, we now call the technique TMAH thermochemolysis.

Information on the type and amount of refratoryy biopolymers produced by plants is still insufficient. The aim of the present work was to determine the chemical (structural) composition of spruce cuticles as a source material for humification. Intact cuticles were isolated by conventional techniques from fully developed needles of Norway spruce (Picea abies (L.) Karst.). The cuticles were subjected to a series of selective treatments to remove different types of polymers. Extraction with organic solvents, to remove lipids and waxes, was followed by saponification to remove the cutin polyester. Finally, the cuticle residues were hydrolyzed to remove polysaccharides. Through investigations combining CP/MAS 13C-NMR spectroscopy, analytical pyrolysis, and wet chemical methods (CuO oxidation), the chemical composition of the polymer was determined in the cuticle, and in the residues obtained by the selective chemical treatments. These data show that the isolated spruce cuticles consist of extractable lipids, polysaccharides, and cutin, biopolymers commonly found in plant cuticles. In addition, a lignin-type polymer was identified, which was selectively isolated after the treatments described above, from the final residue. In conjunction with results from a microscopic survey of the cuticles, these investigations provide evidence for the presence of a lignin-like polymer as a component of intact cuticles of Norway spruce. The consequences of this finding for the humification process of spruce litter are discussed.

In this study 1 Eocene wood sample and 25 fossil Eocene seed coats of water plants from 6 different localities on the south coast of England are analyzed using scanning electron and light microscopy, in addition to Curie-point pyrolysis-GC(-MS) and solid state 13 C NMR, in order to characterize their morphological and chemical alteration in relation to different lithologies. Results from the translucent inner seed coat layers, tegmens, reveal that these tissues consist of aliphatic type biomacromolecules similar to cutan. The tegmens remain morphologically and chemically unaltered. By contrast, the results from the outer seed coat layers, testae, show that these are composed of polyphenolic macromolecules derived from lignin-celluloses. The testae are, on average, morphologically best preserved in fine-grained sediments, whereas best chemical preservation is observed in testae from coarse-grained sediments. Testae associated with pyrite in blue or green fine-grained sediments show poor quality chemical preservation, whereas those from dark brown and grey clays with pyrite show chemical preservation more similar to that of testae from coarse-grained sediments. These results show that diagenetic alteration of organic matter is not only determined by burial history but can also be greatly influenced by differences in lithology.