Co-cultivation of diazotrophic terrestrial cyanobacteria and Arabidopsis thaliana.

Diazotrophic cyanobacteria are able to fix N2 from the atmosphere and release it as bioavailable nitrogen what other organisms can utilize. Thus, they could be used as living nitrogen supplier whereby the use of fertilizer could be reduced in agricultural industry what results in a decrease of laughing gas released during fertilizer production. The diazotroph cyanobacterium Desmonostoc muscorum (D. muscorum) was characterized in shake flasks cultivated in nitrogen-free and nitrogen-containing medium. Similar growth rates were reached in both cultivations and the release of ammonium by D. muscorum was detected under nitrogen depletion. Subsequently, D. muscorum was co-cultivated with Arabidopsis thaliana (A. thaliana) in nitrogen-free medium. Additionally, the plant was cultivated in nitrogen containing and nitrogen-free medium without D. muscorum as reference. A co-cultivation led to higher growth rates of the cyanobacterium and similar growth of A. thaliana with similar maximum photochemical efficiency of photosystem II compared to the growth of nitrogen containing medium. Further, accumulation of cyanobacterial cells around the roots of A. thaliana was detected, indicating a successfully induced artificial symbiosis. Based on these results, D. muscorum could be a promising cyanobacterium as living nitrogen supplier for plants.

Pink bacteria-Production of the pink chromophore phycoerythrobilin with Escherichia coli.

Phycoerythrobilin (PEB) is an open-chain tetrapyrrole derived from heme and plays an important role as light-harvesting pigment in the phycobiliproteins of cyanobacteria and red algae. Furthermore, PEB can also function as an antioxidant with potential use as a natural acid stable food colorant. PEB is not commercially available and large, pure quantities can only be obtained by laborious methanolysis of red algae followed by liquid chromatography. Here we describe an improved method for high yield production and purification of PEB in Escherichia coli via heterologous expression where the two required enzymes heme oxygenase and PEB synthase subsequently convert the substrate heme provided by the host cell. Experiments in shaking flasks resulted in the highest product yield of 680.23 ±â€¯42.75 µg PEB per g cell dry weight, by induction with 0.1 mM IPTG. Scale-up to batch-operated fermentation in a 2 L bioreactor reached product concentrations up to 5.02 mg PEB L-1 by adjustment of aeration, induction time, media composition and supplementation of precursors. A further approach included separation of PEB from developed foam above the culture. This enabled continuous product collection during cultivation and simplified product purification. Produced PEB was validated via UV-vis spectroscopy, high pressure liquid chromatography and mass spectrometry.

Determination of triterpenic acids and screening for valuable secondary metabolites in Salvia sp. suspension cultures.

Plant in vitro cultures are a prospective alternative for biochemicals production, for example the triterpenes oleanolic and ursolic acid present in plants and cell cultures of Salvia sp. Our objective was to develop a suitable analysis protocol for evaluation of triterpenic acid yield in plant raw material and in vitro cultures supporting selection processes. Moreover, valuable bioactive compounds had to be revealed. Thus, different strategies enhancing the separation for a sensitive and effective HPLC-UV method were investigated and the developed method was validated for linearity, precision, accuracy, limits of detection and quantification. A baseline separation of these isomers enabled detection limits of below 0.4 microg/mL and quantification limits of about 1.2 microg/mL. Over the tested concentration range a good linearity was observed (R2 > 0.9999). The variations in the method were below 6% for intra- and inter-day assays of concentration. Recoveries were between 85-98% for both compounds using ethanol as extraction solvent. Additionally, metabolite profiling of cell suspension culture extracts by GC-MS has shown the production variability of different plant metabolites and especially the presence of plant phenols and sterols. These studies provide a method suitable for screening plant and cell culture productivity of triterpenic acids and highlighted interesting co-products of plant cell cultures.