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We tested the influence of two light intensities [40 and 300 μmol PAR / (m2s)] on the fatty acid composition of three distinct lipid classes in four freshwater phytoplankton species. We chose species of different taxonomic classes in order to detect potentially similar reaction characteristics that might also be present in natural phytoplankton communities. From samples of the bacillariophyte Asterionella formosa, the chrysophyte Chromulina sp., the cryptophyte Cryptomonas ovata and the zygnematophyte Cosmarium botrytis we first separated glycolipids (monogalactosyldiacylglycerol, digalactosyldiacylglycerol, and sulfoquinovosyldiacylglycerol), phospholipids (phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, and phosphatidylserine) as well as non-polar lipids (triacylglycerols), before analyzing the fatty acid composition of each lipid class. High variation in the fatty acid composition existed among different species. Individual fatty acid compositions differed in their reaction to changing light intensities in the four species. Although no generalizations could be made for species across taxonomic classes, individual species showed clear but small responses in their ecologically-relevant omega-3 and omega-6 polyunsaturated fatty acids (PUFA) in terms of proportions and of per tissue carbon quotas. Knowledge on how lipids like fatty acids change with environmental or culture conditions is of great interest in ecological food web studies, aquaculture, and biotechnology, since algal lipids are the most important sources of omega-3 long-chain PUFA for aquatic and terrestrial consumers, including humans.
We tested the influence of two light intensities [40 and 300 μmol PAR / (m2s)] on the fatty acid composition of three distinct lipid classes in four freshwater phytoplankton species. We chose species of different taxonomic classes in order to detect potentially similar reaction characteristics that might also be present in natural phytoplankton communities. From samples of the bacillariophyte Asterionella formosa, the chrysophyte Chromulina sp., the cryptophyte Cryptomonas ovata and the zygnematophyte Cosmarium botrytis we first separated glycolipids (monogalactosyldiacylglycerol, digalactosyldiacylglycerol, and sulfoquinovosyldiacylglycerol), phospholipids (phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, and phosphatidylserine) as well as non-polar lipids (triacylglycerols), before analyzing the fatty acid composition of each lipid class. High variation in the fatty acid composition existed among different species. Individual fatty acid compositions differed in their reaction to changing light intensities in the four species. Although no generalizations could be made for species across taxonomic classes, individual species showed clear but small responses in their ecologically-relevant omega-3 and omega-6 polyunsaturated fatty acids (PUFA) in terms of proportions and of per tissue carbon quotas. Knowledge on how lipids like fatty acids change with environmental or culture conditions is of great interest in ecological food web studies, aquaculture, and biotechnology, since algal lipids are the most important sources of omega-3 long-chain PUFA for aquatic and terrestrial consumers, including humans.
In this dissertation, an electric field-assisted method was developed and applied to achieve immobilization and alignment of biomolecules on metal electrodes in a simple one-step experiment. Neither modifications of the biomolecule nor of the electrodes were needed. The two major electrokinetic effects that lead to molecule motion in the chosen electrode configurations used were identified as dielectrophoresis and AC electroosmotic flow. To minimize AC electroosmotic flow, a new 3D electrode configuration was designed. Thus, the influence of experimental parameters on the dielectrophoretic force and the associated molecule movement could be studied. Permanent immobilization of proteins was examined and quantified absolutely using an atomic force microscope. By measuring the volumes of the immobilized protein deposits, a maximal number of proteins contained therein was calculated. This was possible since the proteins adhered to the tungsten electrodes even after switching off the electric field. The permanent immobilization of functional proteins on surfaces or electrodes is one crucial prerequisite for the fabrication of biosensors.
Furthermore, the biofunctionality of the proteins must be retained after immobilization. Due to the chemical or physical modifications on the proteins caused by immobilization, their biofunctionality is sometimes hampered. The activity of dielectrophoretically immobilized proteins, however, was proven here for an enzyme for the first time. The enzyme horseradish peroxidase was used exemplarily, and its activity was demonstrated with the oxidation of dihydrorhodamine 123, a non-fluorescent precursor of the fluorescence dye rhodamine 123.
Molecular alignment and immobilization - reversible and permanent - was achieved under the influence of inhomogeneous AC electric fields. For orientational investigations, a fluorescence microscope setup, a reliable experimental procedure and an evaluation protocol were developed and validated using self-made control samples of aligned acridine orange molecules in a liquid crystal.
Lambda-DNA strands were stretched and aligned temporarily between adjacent interdigitated electrodes, and the orientation of PicoGreen molecules, which intercalate into the DNA strands, was determined. Similarly, the aligned immobilization of enhanced Green Fluorescent Protein was demonstrated exploiting the protein's fluorescence and structural properties. For this protein, the angle of the chromophore with respect to the protein's geometrical axis was determined in good agreement with X-ray crystallographic data. Permanent immobilization with simultaneous alignment of the proteins was achieved along the edges, tips and on the surface of interdigitated electrodes. This was the first demonstration of aligned immobilization of proteins by electric fields.
Thus, the presented electric field-assisted immobilization method is promising with regard to enhanced antibody binding capacities and enzymatic activities, which is a requirement for industrial biosensor production, as well as for general interaction studies of proteins.