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Project Title:

Phototrophical production of hydrogen from aromatic compounds

Ref.No.: 20

Project Type and Category:

Hydrogen production

Project Duration:

1.1.1990 - 31.12.1992

Project Participants:

Prof. Dr.Friedrich Giffhorn, Angewandte Mikrobiologie, Universitaet des Saarlandes
Partners:
1. Prof. Dr. H.J. Klemme, Institut für Mikrobiologie der Universität Bonn
2. Dr. H. Cypionka, Prof. Dr. N. Pfennig, Fakultät für Biologie der Universität Konstanz
3. Prof. Dr. J. Oelze, Institut für Biologie II, Mikrobiologie, der Universität Freiburg

Sponsor:

BMFT (BMBF) ""Biological Hydrogen production""

Project Budget and
Funding:

138,000 DM

Project Description and Objectives:

Research of biological hydrogen production at competitive costs. The basic idea is to use industrial waste as a substrate to feed bacteria that produce hydrogen.

Technical Goals:

Find out conditions and material combinations with optimized bacteria grow and hydrogen production.

Project Status

Finalized

Preliminary or Final Results:

Purple nonsulfur bacteria are able to grow phototrophically with aromatic substrates (Evans and Fuchs 1988) and they may evolve hydrogen under nitrogen limited conditions. This production of molecular hydrogen is catalysed by a side reaction of the nitrogenase complex (Willison et. al. 1983). A combination of hydrogen production as a potential energy source and the degradation of aromatic waste compounds derived from industrial processes might become an interesting process. The capability of five strains of the phototrophic bacterium Rhodopseudomonas palustris to produce molecular hydrogen from the aromatic acids benzoate, p-hydroxybenzoate, cinnamate and D- and L-mandelate was investigated. Optimal hydrogen production was achieved when the strains were grown anaerobically in the light at 10000 lux under nitrogen limitation using 1 mM L-glutamate as nitrogen source. In the presence of 2 mM benzoate or L-mandelate as carbon and electron source, strain DSM 131 produced 45% H2 of the maximal theoretical value and strain F2 32%, respectively. Increased hydrogen production correlated with increased nitrogenase activities, but hydrogen formation was not further stimulated by inhibition of the hydrogen uptake hydrogenase with EDTA (Fissler et al. 1994). Enhancement of the yield is described for the hydrogen production from nonaromatic organic substrates with immobilized cells of Rhodobacter capsulatus (Francou and Vignais 1984), Rhodospirillum rubrum (von Felten et Al. 1985) or Rhodobacter sphaeroides (Sasikala et al. 1990). As compared toliquid cultures immobilized cells exhibit in many systems improved production characteristics and stabilities. The entrapment in polymeric matrices anchors the living cells in a network, but allows free diffusion of the substrates and products. Natural polymers like agar or alginate (Von Felten et al. 1985) or immobilization on porous glas are suitable matrices, which avoid losses of activity during the immobilization procedure. Cells of the purple nonsulfur bacterium Rhodopseudomonas palustris DSM 131 were immobilized in agar, agarose, *-carrageenan or Na-alginate gel, respectively. With alginate beads, prepared by an emulsion technique and an optimal cell load of 10 mg dry weight/ml gel, the hydrogen production from aromatic acids was doubled as compared to growing liquid cultures. Hydrogen yields of 60%, 57%, 86% or 88% of the maximal theoretical value were obtained from mandelate, benzoylformate, cinnamate or benzoate, respectively. Benzoate concentrations above 16.5 mM were inhibitory. During a period of 55 days the process of hydrogen evolution with immobilized cells was repeated in five cycles with slowly decreasing efficiency (Fissler et al. 1996).

Related Reference Papers and Other Publications:

Related projects: Ref.No.: 208,21, 23, 107, 59
Literature:
            Evans WC, Fuchs G (1988) Ann Rev Microbiol 42: 289-317. Felten P von,
            Zürrer H, Bachofen R (1985) Appl Microbiol Biotechnol 23: 15-20. Fissler
            J, Schirra C, Kohring G-W, Giffhorn F (1994) Appl Microbiol Biotechnol 41:
Publications:
Kohring, G.-W., J. Fißler, C. Schirra, F. Giffhorn: Hydrogen production from aromatic acids by liquid cultures and immobilized cells of Rhodopseudomonas
palustris. In: Verziroglu, T.N., C.-J. Winter, J.P. Baselt, G. Kreysa (eds.): Hydrogen Energy Progress XI (Proceedings of the 11th World Hydrogen Energy Conference, Stuttgart) (1996), 2743-2748
Fißler, J., G.-W. Kohring, F. Giffhorn: Enhanced hydrogen production from aromatic acids by immobilized cells of Rhodopseudomonas palustris. Appl. Microbiol. Biotechnol.44, 43-46 (1995)
Fißler, J., C. Schirra, G.-W. Kohring, F. Giffhorn: Hydrogen production from aromatic acids by Rhodopseudomonas palustris. Appl. Microbiol. Biotechnol. 41, 395-399 (1994)
Kohring, G.-W., J. Fißler, F. Giffhorn: Degradation of aromatic compounds by purple nonsulfur bacteria. In: D. Behrens, J. Wiesner (eds.): Mikrobiologische Reinigung von Böden, DECHEMA, Frankfurt, 263-268 (1992)
Kohring, G.-W., J. Fißler, B. Hermann, C. Schirra, F. Giffhorn: Anaerobic degradation of substituted aromatic compounds by purple nonsulfur bacteria. In: G. Kreysa, A.J. Driesel (eds.): DECHEMA Biotechnology Conferences 5, DECHEMA, Frankfurt, 873-876 (1992)
Kohring, G.-W., C. Schirra, S. Schmitt, F. Giffhorn: Anaerobic degradation of phenol and chlorinated aromatic compounds by purple nonsulfur bacteria. International Symposium: Soil Decontamination Using Biological Processes, Karlsruhe, Deutschland 485-490 (1992)
Posters und reports:
Kohring, G.-W., J. Fißler, F. Giffhorn: Degradation of aromatic compounds by purple nonsulfur bacteria. Poster, Jahrestagung der VAAM in Freiburg (1991)
Kohring, G.-W., J. Fißler, F. Giffhorn: Abbau aromatischer Verbindungen durch chwefelfreie Purpurbakterien. Poster, DECHEMA Fachgespräch Umweltschutz ""Mikrobiologische Reinigung von Böden"", (1991)
Kohring, G.-W., J. Fißler, F. Giffhorn: Hydrogen Production by puple nonsulfur bacteria during growth on aromatic compounds. Vortrag, VII International Symposium on Photosynthetic Procaryotes in Amherst, Massachusetts, USA (1991)
Hermann, B., G.-W. Kohring, F. Giffhorn: Isolation of Purple Nonsulfur Bacteria, that Degrade Phloroglucinol to H2 and CO2. Poster Jahrestagung der VAAM in Düsseldorf (1992)

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Project Title: Phototrophical production of hydrogen from aromatic compounds		Ref.No.: 20
Project Type and Category:	Hydrogen production
Project Duration:	1.1.1990 - 31.12.1992
Project Participants:	Prof. Dr.Friedrich Giffhorn, Angewandte Mikrobiologie, Universitaet des Saarlandes Partners: 1. Prof. Dr. H.J. Klemme, Institut für Mikrobiologie der Universität Bonn 2. Dr. H. Cypionka, Prof. Dr. N. Pfennig, Fakultät für Biologie der Universität Konstanz 3. Prof. Dr. J. Oelze, Institut für Biologie II, Mikrobiologie, der Universität Freiburg
Sponsor:	BMFT (BMBF) ""Biological Hydrogen production""
Project Budget and Funding:	138,000 DM
Project Description and Objectives:	Research of biological hydrogen production at competitive costs. The basic idea is to use industrial waste as a substrate to feed bacteria that produce hydrogen.
Technical Goals:	Find out conditions and material combinations with optimized bacteria grow and hydrogen production.
Project Status	Finalized
Preliminary or Final Results:	Purple nonsulfur bacteria are able to grow phototrophically with aromatic substrates (Evans and Fuchs 1988) and they may evolve hydrogen under nitrogen limited conditions. This production of molecular hydrogen is catalysed by a side reaction of the nitrogenase complex (Willison et. al. 1983). A combination of hydrogen production as a potential energy source and the degradation of aromatic waste compounds derived from industrial processes might become an interesting process. The capability of five strains of the phototrophic bacterium Rhodopseudomonas palustris to produce molecular hydrogen from the aromatic acids benzoate, p-hydroxybenzoate, cinnamate and D- and L-mandelate was investigated. Optimal hydrogen production was achieved when the strains were grown anaerobically in the light at 10000 lux under nitrogen limitation using 1 mM L-glutamate as nitrogen source. In the presence of 2 mM benzoate or L-mandelate as carbon and electron source, strain DSM 131 produced 45% H2 of the maximal theoretical value and strain F2 32%, respectively. Increased hydrogen production correlated with increased nitrogenase activities, but hydrogen formation was not further stimulated by inhibition of the hydrogen uptake hydrogenase with EDTA (Fissler et al. 1994). Enhancement of the yield is described for the hydrogen production from nonaromatic organic substrates with immobilized cells of Rhodobacter capsulatus (Francou and Vignais 1984), Rhodospirillum rubrum (von Felten et Al. 1985) or Rhodobacter sphaeroides (Sasikala et al. 1990). As compared toliquid cultures immobilized cells exhibit in many systems improved production characteristics and stabilities. The entrapment in polymeric matrices anchors the living cells in a network, but allows free diffusion of the substrates and products. Natural polymers like agar or alginate (Von Felten et al. 1985) or immobilization on porous glas are suitable matrices, which avoid losses of activity during the immobilization procedure. Cells of the purple nonsulfur bacterium Rhodopseudomonas palustris DSM 131 were immobilized in agar, agarose, *-carrageenan or Na-alginate gel, respectively. With alginate beads, prepared by an emulsion technique and an optimal cell load of 10 mg dry weight/ml gel, the hydrogen production from aromatic acids was doubled as compared to growing liquid cultures. Hydrogen yields of 60%, 57%, 86% or 88% of the maximal theoretical value were obtained from mandelate, benzoylformate, cinnamate or benzoate, respectively. Benzoate concentrations above 16.5 mM were inhibitory. During a period of 55 days the process of hydrogen evolution with immobilized cells was repeated in five cycles with slowly decreasing efficiency (Fissler et al. 1996).
Related Reference Papers and Other Publications:	Related projects: Ref.No.: 208,21, 23, 107, 59 Literature: Evans WC, Fuchs G (1988) Ann Rev Microbiol 42: 289-317. Felten P von, Zürrer H, Bachofen R (1985) Appl Microbiol Biotechnol 23: 15-20. Fissler J, Schirra C, Kohring G-W, Giffhorn F (1994) Appl Microbiol Biotechnol 41: Publications: Kohring, G.-W., J. Fißler, C. Schirra, F. Giffhorn: Hydrogen production from aromatic acids by liquid cultures and immobilized cells of Rhodopseudomonas palustris. In: Verziroglu, T.N., C.-J. Winter, J.P. Baselt, G. Kreysa (eds.): Hydrogen Energy Progress XI (Proceedings of the 11th World Hydrogen Energy Conference, Stuttgart) (1996), 2743-2748 Fißler, J., G.-W. Kohring, F. Giffhorn: Enhanced hydrogen production from aromatic acids by immobilized cells of Rhodopseudomonas palustris. Appl. Microbiol. Biotechnol.44, 43-46 (1995) Fißler, J., C. Schirra, G.-W. Kohring, F. Giffhorn: Hydrogen production from aromatic acids by Rhodopseudomonas palustris. Appl. Microbiol. Biotechnol. 41, 395-399 (1994) Kohring, G.-W., J. Fißler, F. Giffhorn: Degradation of aromatic compounds by purple nonsulfur bacteria. In: D. Behrens, J. Wiesner (eds.): Mikrobiologische Reinigung von Böden, DECHEMA, Frankfurt, 263-268 (1992) Kohring, G.-W., J. Fißler, B. Hermann, C. Schirra, F. Giffhorn: Anaerobic degradation of substituted aromatic compounds by purple nonsulfur bacteria. In: G. Kreysa, A.J. Driesel (eds.): DECHEMA Biotechnology Conferences 5, DECHEMA, Frankfurt, 873-876 (1992) Kohring, G.-W., C. Schirra, S. Schmitt, F. Giffhorn: Anaerobic degradation of phenol and chlorinated aromatic compounds by purple nonsulfur bacteria. International Symposium: Soil Decontamination Using Biological Processes, Karlsruhe, Deutschland 485-490 (1992) Posters und reports: Kohring, G.-W., J. Fißler, F. Giffhorn: Degradation of aromatic compounds by purple nonsulfur bacteria. Poster, Jahrestagung der VAAM in Freiburg (1991) Kohring, G.-W., J. Fißler, F. Giffhorn: Abbau aromatischer Verbindungen durch chwefelfreie Purpurbakterien. Poster, DECHEMA Fachgespräch Umweltschutz ""Mikrobiologische Reinigung von Böden"", (1991) Kohring, G.-W., J. Fißler, F. Giffhorn: Hydrogen Production by puple nonsulfur bacteria during growth on aromatic compounds. Vortrag, VII International Symposium on Photosynthetic Procaryotes in Amherst, Massachusetts, USA (1991) Hermann, B., G.-W. Kohring, F. Giffhorn: Isolation of Purple Nonsulfur Bacteria, that Degrade Phloroglucinol to H2 and CO2. Poster Jahrestagung der VAAM in Düsseldorf (1992)