My Ph.D. thesis is published as paperback:
Maretzek, António F.:
Quantitative in vivo 31P-NMR spektroskopische Untersuchungen zum zellulären Energiehaushalt an den photoautotrophen Mikroorganismen Synechococcus leopoliensis und Chlorella fusca
Aachen: Shaker, 1992
ISBN 3-86111-210-8
Language: German

Below is an abstract, that is quite close to the one in the thesis.

The energy metabolism of two photoautotrophic microorganisms, the procaryotic cyanobacterium Synechococcus leopoliensis and the eucaryotic green alga Chlorella fusca was investigated using 31P-NMR spectroscopy. Two subjects were focused:

Improvements of experimental prerequisites were achieved by the design of an appropriate tube support to meet the physiological and the NMR demands. The NMR tube can be removed without disarranging the other elements as the perfusion tubes, standard capillary, etc., in this way guaranteeing good mechanical precision. The accuracy of the estimation of concentration was tested in in vitro experiments and was found to be +/-4%. Furthermore the optical shutter of the illumination system was automatized, obtaining an illumination control as fast as 1 s for the organisms in the NMR tube.

Improvement of the evaluation of signal intensity is based on a non automatic working deconvolution routine, considering contributions of overlapping signals and avoiding and avoiding misestimation of the baseline.

The efficacy of the specified improvements was verified in in vivo NMR experiments. The deviation of the NTP concentration between independent experiments was in the order of +/-10%. Comparing precision and accuracy of the concentration as determined by extraction methods to the values presented, it has to be stated, that determination of phosphate metabolites in photoautotrophic microorganisms using in vivo 31P-NMR spectroscopy is as reliable as conventional methods.

Concentrations and intracellular pH were determined during steady states of anaerobic, aerobic and photosynthesis conditions. Transient metabolism was studied with anaerobic dark/photosynthesis and anaerobic/aerobic transitions.

NTP concentrations, cytoplasmic pH, phosphorylation potential and AEC during photosynthesis or dark aerobic conditions were higher, cytoplasmic inorganic phosphate concentration lower than those during anaerobic dark conditions, corresponding to literature data. There was no stoichiometry between the increase of the NTP concentration and the decrease of the cytoplasmic inorganic phosphate. The overall concentration of phosphate decreased during photosynthesis or aerobic conditions revealing the phosphorylation of not detectable metabolites (e.g. proteins). During anaerobic conditions in C. fusca an alkalisation whereas during photosynthesis an acidification of the vacuole was observed. An increasing concentration of inorganic phosphate in the vacuole (C. fusca) during anaerobic dark conditions was followed by a decrease during subsequent photosynthesis. Consequently there is no simple relation between inorganic phosphate concentration and pH in the vacuole. Unlike C. fusca S. leopolienses exhibits a 15% higher NTP concentration during photosynthesis than under dark aerobic conditions.

After the onset of photosynthesis or aerobic conditions a transient surplus of the NTP concentration was observed while the reverse transition resulted in an undershooting. Both phenomena are the consequence of the regulation of energy demanding processes. Even after NTP concentration had past its maximum the concentration of the cytoplasmic inorganic phosphate continued decreasing. As a consequence the phosphorylation potential reflects the cellular energetic status better than the AEC since the latter doesn't consider inorganic phosphate. After transition from photosynthesis to anaerobic conditions changes of the inorganic phosphate concentration in C. fusca were almost completed within 4 min contrary to S. leopolienses. Apart from that, transient changes in metabolism were similar for both organism.

The equivalent behavior of S. leopolienses and C. fusca concerning the cytoplasmic inorganic phosphate concentration and their low value during photosynthesis reflects a strong coupling of chloroplast and cytoplasm concerning inorganic phosphate, since there is no chloroplast present in S. leopolienses. This work shows that the concentration of inorganic phosphate must be determined, if cellular energy status is to be evaluated.