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| Titel |
Humeomics for a structure-bioactivity relationship of Humic Substances |
| VerfasserIn |
Antonio Nebbioso, Alessandro Piccolo, Riccardo Spaccini |
| Konferenz |
EGU General Assembly 2011
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| Medientyp |
Artikel
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| Sprache |
Englisch
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| Digitales Dokument |
PDF |
| Erschienen |
In: GRA - Volume 13 (2011) |
| Datensatznummer |
250055333
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| Zusammenfassung |
| Humic Substances (HS) represent a class of naturally occurring organic compounds and are
commonly found in soils, sediments and natural bodies of water1. They have an
effect on physical-chemical and biological properties of such environments, and
are fundamental for their quality and fertility2. The heterogeneous and complex
nature of humic components represents the main difficulty against the chemical
characterization and structure elucidation of them. Recent findings have provided significant
experimental evidence supporting the hypothesis that HS are organized in supramolecular
architecture3. Such structure is composed by relatively simple molecules held together
by weak bonds and allows the single compounds to be selectively separated and
quali-quantitatively characterized with appropriate analytical methods, such as NMR4,5 and
mass spectrometry (MS)4,6. A comprehensive description of all components of HS,
defined as “Humeomics” would bring an important achievement for research in
agrarian and environmental chemistry. In order to accomplish this goal it is required
to obtain sub-fractions of HS as homogeneous as possible and characterize their
content.
A humic acid (HA) from a volcanic forestal soil (Allic Fulvudand) was isolated, purified,
and fractionated1,7. The first step of this method consisted in a liquid extraction with a
mixture of dicholormethane and methanol 2:1. This yielded an organic extract not bound to
the residue via covalent bonds. Such residue underwent two trans-esterification stages in
methanol, the first using BF3 12%, and the second using KOH 1M. The extracts thus obtained
were further separated in water/chloroform liquid extraction. Such components were defined
as weakly and strongly ester bound to the residue. This step yielded also a residue, which is
thereafter hydrolyzed in aqueous HI 47% to separate the organic matter bound via
ether bonds. The final product showed minimal chemical reactivity. Such residual
matter was separated with HP-SEC chromatography in order to separate fractions
according to their hydrodynamic volume. The fractions obtained were analyzed with
the following techniques: 1. Elemental analysis 2. NMR mono and bidimensional
spectroscopy, in both solid and solution state (Bruker AV300 e AV400) 3. GC-MS
and pyrolysis-GC-MS (Perkin-Elmer Turbomass Gold) 4. HPSEC coupled to MS
with Elettrospray source and Orbitrap detector (Thermo Finnigan Orbitrap, High
resolution).
The organosoluble fractions show linear aliphatic and, to a lesser extent, aromatic
compounds. In hydrosoluble fractions several compounds with a chemical formula similar to
saccarides are detected. This is suggested by formulas with high content of oxygen and
nitrogen atoms. Aliphatic compounds are also revealed to a lesser extent. In the final residue
quantitative assessment shows aromatic (mostly quaternary)>aliphatic>carbonyl molecules
in order of abundance. From NMR relaxation time measurements, a more rigid architecture is
attributed to this material with respect to the starting HA. Mass spectrometry revealed
formulas with extensive oxygen substitution and unsaturation index, which can be attributed
to the NMR signals in sp2 carbon region. The MS analysis of the final residue showed for
fatty acids, from the first to the tenth HPSEC fraction, an increase in the ratio between 1)
short and long chained homologues and 2) hydrophilic and hydrophobic homologues,
suggesting a correlation between the chemical composition of aggregates and their
hydrodynamic properties: stacking between longer, saturated carbon chains results in larger
aggregates whereas smaller hydroxylated compounds result in weaker aggregation
forces.
The humeomic approach to HS analysis represents a solid basis for a more reliable and
detailed characterization of Natural Organic Matter.
REFERENCES
1) F.J. Stevenson (1994). Humus Chemistry: Genesis, Composition, Reactions. John
Wiley & Sons, New York
2) P.M. Huang, M.K. Wang and C.H. Chiu, (2005) Pedobiologia, 49, 609-635
3) Piccolo A., (2002). Adv. Agron. 75, pp. 57-134
4) Piccolo A., Conte P., Trivellane E., Van Lagen B., Buurman P., (2002). Environ. Sci.
Technol., 36, pp. 76-84
5) Simpson A. J., (2002). Magn. Reson. Chem. 40, pp. S72-S82
6) Grasset L., Amblès A., (1998). J. Anal. Appl. Pyrol. 47, pp. 1-12
7) Kolattukudy P.E., Kronman K., Poulose A.J., 1975. Plant Physiol. 55 pp. 567-573 |
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