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Isolation and purification of Cu-free methanobactin from Methylosinus trichosporium OB3b.


ABSTRACT:

Background

The isolation of highly pure copper-free methanobactin is a prerequisite for the investigation of the biogeochemical functions of this chalkophore molecule produced by methane oxidizing bacteria. Here, we report a purification method for methanobactin from Methylosinus trichosporium OB3b cultures based on reversed-phase HPLC fractionation used in combination with a previously reported resin extraction. HPLC eluent fractions of the resin extracted product were collected and characterized with UV-vis, FT-IR, and C-1s NEXAFS spectroscopy, as well as with elemental analysis and ESI-MS.

Results

The results showed that numerous compounds other than methanobactin were present in the isolate obtained with resin extraction. Molar C/N ratios, mass spectrometry measurements, and UV-vis spectra indicated that methanobactin was only present in one of the HPLC fractions. On a mass basis, methanobactin carbon contributed only 32% to the total organic carbon isolated with resin extraction. Our spectroscopic results implied that besides methanobactin, the organic compounds in the resin extract comprised breakdown products of methanobactin as well as polysaccharide-like substances.

Conclusion

Our results demonstrate that a purification step is indispensable in addition to resin extraction in order to obtain pure methanobactin. The proposed HPLC purification procedure is suitable for semi-preparative work and provides copper-free methanobactin.

SUBMITTER: Pesch ML 

PROVIDER: S-EPMC3044663 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Isolation and purification of Cu-free methanobactin from Methylosinus trichosporium OB3b.

Pesch Marie-Laure ML   Christl Iso I   Barmettler Kurt K   Kraemer Stephan M SM   Kretzschmar Ruben R  

Geochemical transactions 20110207


<h4>Background</h4>The isolation of highly pure copper-free methanobactin is a prerequisite for the investigation of the biogeochemical functions of this chalkophore molecule produced by methane oxidizing bacteria. Here, we report a purification method for methanobactin from Methylosinus trichosporium OB3b cultures based on reversed-phase HPLC fractionation used in combination with a previously reported resin extraction. HPLC eluent fractions of the resin extracted product were collected and cha  ...[more]

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