F.J. González-Vila

13C- and 15N-NMR spectroscopic examination of the transformation of organic nitrogen in plant biomass during thermal treatment

Abstract Structural changes in lignocellulosic biomass heated under conditions comparable to those encountered in several types of natural or planned burnings have been studied by solid-state 13C- and 15N-CPMAS NMR spectroscopy of 15N-enriched ryegrass (Lolium rigidum) after being subjected to progressive thermal treatment. The solid-state 15N-NMR spectra of biomass subjected to severe heating revealed amide-N in forms which are resistant to the thermal treatment. Progressive burning was found to occur in two well-defined stages: In the early stage the free amino acid and some NH2 groups were removed, but no substantial disruption of the peptide structure was observed. In the final stage of burning the amide-N was converted to heterocyclic structures such as pyrroles, imidazoles and indoles. Some evidence for the presence of pyridines and phenazines was also found at this stage. These findings suggest that a major portion of the N is in forms that may survive most natural fires and that their stability towards further microbial degradation is increased by the heating. The solid-state 13C-NMR spectra revealed that the carbohydrate fraction is converted into condensed dehydrated material producing intense signals in the aromatic region.

13C NMR assessment of decomposition patterns during composting of forest and shrub biomass

A laboratory experiment was designed to investigate the degradation patterns of leaves from 12 forest and shrub species typical of Mediterranean ecosystems by solid-state 13 C NMR. The spectral data have been compared with those for the major organic fractions, and elementary composition in three transformation stages (zero time, intermediated and advanced (168 d)). The plant material in general showed a selective depletion of lipid and water-soluble products and a concentration in acidinsoluble residue (Klason lignin fraction), but the increasing percentage of total alkyl carbons (not observed in pine leaves) suggests that recalcitrant aliphatic material accumulates in the course of the 168 d incubation, when the total weight losses were up to 660 g kg ˇ1 . This contrasts with the fact that the concentration of extractable alkyl C (i.e. the lipid fraction) decreased in all cases. The results for the diAerent plants suggested some general transformation trends simultaneous to specific biodegradation patterns. The non-ameliorant, soil acidifying species (i.e. those a priori considered to favor the accumulation of humus with low biological activity) have high initial concentrations of extractives, alkyl structures and comparatively lower percentages of O-alkyl structures. The decay process in these species is not associated to the increase of the alkyl-to-O-alkyl ratio, which is shown by the ameliorant species. Superimposed on these major trends, the biomass of the diAerent plants underwent divergent paths in the course of composting, leading to, for example, (i) accumulation of recalcitrant, nonhydrolyzable alkyl and aromatic structures (Retama, Genista); (ii) enrichment of resistant O-alkyl structures such are stable fractions of carbohydrate and tannins (Pinus, Calluna); and (iii) accumulation of aliphatic extractives with the lowest stabilization of protein in resistant forms (Arctostaphylos, Ilex). In particular, in the acidifying species, the spectral patterns suggest that the apparent stability of the aromatic domain is compatible with selective preservation of tannins together with aliphatic structures. Such specific tendencies are also illustrated by the diAerence spectra (0 vs 168 d) which suggest that early humification processes are highly heterogeneous and distinct rather than the selective degradation of lipid and water-soluble fractions and carbohydrates, and they may include stabilization of tannins and aliphatic (cutin- and protein-like) macromolecules. 7 2000 Elsevier Science Ltd. All rights reserved.

Pyrolysis derivatization of humic substances 1. Pyrolysis of fulvic acids in the presence of tetramethylammonium hydroxide

Abstract Humic Acids (HAs) extracted from soils of very different origin have been analyzed by pyrolysis in the presence of tetramethylammonium hydroxide. The thermal degradation products consisted mainly of aliphatic series such as fatty acids methyl esters and α,ω-methoxy fatty acid methyl esters. Triterpenoid compounds with ursane, oleanane and hopane skeletons were also detected in several samples. Aromatic units derived from lignin moieties were also detected although in minor amounts and corresponded mainly to guaiacyl and syringyl units. This procedure seems to release the more labile, aliphatic moieties attached to the aromatic nuclei of the HA structure.

Effects of fire on humic and lipid fractions in a Dystric Xerochrept in Spain

Fire-induced changes in humic and lipid soil fractions were assessed by two approaches. Comparisons were made of the nature and amounts of the two fractions in samples of a Dystric Xerochrept under two pine stands, one that had been burned two years earlier and one that had not. Further comparisons were made of humic and lipid fractions in a soil sample before and after heating in the laboratory. Separation and identification of the compounds in these soil fractions were carried out by preparative liquid chromatography followed by GC-MS in the case of lipids; the humic acids were previously degraded by successive treatments with sodium persulphate and potassium permanganate. The greatest differences were in the distribution patterns of soil lipids (alkanes, fatty acids): accumulation of the homologues of lower molecular weight (< C20) was dominant in the post-fire samples. For soil resin acids, these samples had higher values for dehydroabietic and secodehydroabietic acids and lower ones for pimaric acid. The degradative studies on the humic acids showed that the relative yields of aromatic products were comparatively higher in the samples after burning. Alkanoic diacids were found to be the least resistant to heating, whereas greater amounts of alkanes in the less condensed polymer structures of the humic acids were observed in the samples after burning. The relative amounts and distribution patterns of the fatty acids released showed no great differences in the humic acids.

Preservation of aliphatic macromolecules in soil humins

Abstract Two series of alkali-insoluble preparations of soil organic matter (humins) were isolated from three representative types of terrestrial humus by two methods, respectively based on (i) ultrasonic disaggregation followed by flotation in a bromoform-ethanol mixture and (ii) partition in water-methylisobutylketone. The humins amounted to between 7–42% of the total carbon in the soil heavy fraction and were characterized by pyrolysis followed by gas chromatography-mass spectrometry. The pyrolysis patterns of these particulate soil organic fractions differed quantitatively and qualitatively from those typical for terrestrial humic acid or lignocellulosic materials. The humins isolated by both methods yielded major series of alkyl products (56–81% of the total chromatographic area) including very variable amounts of alkanes, fatty acids and olefins. The aromatic pyrolysis products consisted mainly of alkyl-substituted mono and di-phenols, naphthalenes, phenanthrenes, fluorenes and methoxyphenols. The pyrograms of the humins isolated by partition were much more varied and produced a greater yield of alkanes than those isolated by flotation. The differences between humins from the three types of humus were small and tended to coincide in both series of preparations. Both the 13 C NMR spectra and the atomic ratios suggested the presence of stable carbohydrate-like material, which was not evident in the pyrograms or in the total yield of hydrolyzable sugars. The results suggest that humins represent the high-stability aggregate compartments of active terrestrial soils where structural rearrangement of aliphatic biogenic macromolecules may take place, simultaneously to the condensation of recalcitrant macromolecular material tightly bonded on reactive mineral surfaces.

Thermally assisted hydrolysis and alkylation of lignins in the presence of tetra-alkylammonium hydroxides

Three different milled lignins isolated from bamboo, pine and beech, corresponding to the three different structural groups, have been subjected to pyrolysis in the presence of tetramethylammonium hydroxide (TMAH) and tetrabutylammonium hydroxide (TBAH). Pyrolysis of the lignins in the presence of TMAH releases the methyl esters and methyl ethers of the different lignin monomers. Several moieties bearing carboxylic acid groups, not previously detected using pyrolytic techniques, could be released from the three lignins, suggesting that the presence of these moieties in the lignin structure have been underestimated.

Solid state NMR studies of fire-induced changes in the structure of humic substances

Abstract Several structural changes were recognized by CPMAS 13 C-NMR and resolution-enhanced IR spectroscopy after humic acids (HA) and fulvic acids (FA) were subjected to laboratory heating at 350°C. The NMR results suggested that the HA underwent decarboxylation, selective transformation of aliphatic material, and increase in the number of aromatic carbons when subjected to heating for 120 s, whereas after subsequent heating no predominant aromatic neoformations were observed, and the elimination of alkyl structures occurred preferentially at this stage. The infrared spectroscopy revealed a well-defined lignin pattern in the original HA, that tended to disappear with heating. The results obtained in the laboratory experiments agreed with several of the differences found between the NMR patterns of the humic substances from the unburnt soil area and from that affected by forest fire. It is suggested that the changes observed play important roles in the colloidal properties and the water-repellency observed in the heated humic substances and in the post-fire soil area.

Chemical composition of lipophilic extractives from Eucalyptus globulus Labill. wood

Summary The composition of lipophilic extractives in the chloroform soluble fraction of the acetone extract from Eucalyptus globulus wood has been examined. The lipid extract was fractionated by solid-phase extraction on aminopropyl-phase cartridges into four different fractions of increasing polarity. The total lipid extract and the resulting fractions were analyzed by gas chromatography and gas chromatography-mass spectrometry, using high temperature capillary columns. The main compounds identified included sterols, sterol esters, fatty acids, steroid ketones, hydrocarbons and triglycerides. Minor compounds such as fatty alcohols, mono- and diglycerides, waxes and tocopherols were also identified among the lipids from E. globulus wood.

Molecular alterations of organic fractions from urban waste in the course of composting and their further transformation in amended soil

Abstract The evolution of various analytical characteristics (including the concentrations of water-soluble fractions, free and esterified lipids, and humic-like substances) of solid urban waste in the course of a 7-week composting process have been monitored in two independent piles composted during different seasons (Spring/Autumn). Whereas the concentration of water-soluble and mineral fractions tended to increase during composting, the opposite was observed for total organic matter and free lipid. Unmatured compost showed comparatively high amounts of esterified lipid. The absence of monotonic trends shown by this fraction, and by total humic-like substances indicate two successive stages of compost microbial reworking. Gas chromatography-mass spectrometry (GC-MS) analysis of the lipid fractions progressively removed by step-wise chemolysis allows the appraisal of different lipid species, which differ in their resistance to biodegradation. Extended composting does not contribute to the accumulation of progressively aromatised humic-type materials similar to those present in soil. Both pyrolysis (Py)-GC-MS and spectroscopic techniques (Fourier transform infrared spectroscopy, FT-IR, and 13 C nuclear magnetic resonance spectroscopy, NMR) confirm that compost-derived humic acid-like substances are not structurally comparable to soil humic acids. Finally, the fate of the compost organic fractions has been investigated in two soils amended with different doses of the final compost. In general, the most conspicuous compost-induced effects were reflected by the differences in the qualitative and quantitative composition of the soil lipid. The results indicate that compost application cannot be considered to contribute to the mid-term accumulation of stable forms of organic matter in soil.

Thermally assisted hydrolysis and alkylation as a novel pyrolytic approach for the structural characterization of natural biopolymers and geomacromolecules

Abstract The high-temperature reaction of macromolecules with tetra-alkylammonium hydroxides has significantly improved the use of the conventional pyrolytic procedures which are widely used for the structural characterization of natural biopolymers and geomacromolecules. This new pyrolytic approach has overcome some of the analytical limitations of conventional pyrolysis allowing the detection of polar compounds as their alkyl derivatives. We report recent developments of the high-temperature hydrolysis and alkylation procedure, carried out in the presence of tetra-alkylammonium hydroxides, for the structural characterization of natural biopolymers and geomacromolecules such as lignins, humic substances, coals, asphaltenes and kerogens.