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Using a concerted approach of biochemical standard preparation, analytical access via LC-MS/MS, glucose pulse, metabolic profiling, and statistical data analysis, the metabolism dynamics in the aromatic amino acid pathway has been stimulated, monitored, and analyzed in different tyrosine-auxotrophic L-phenylalanine-producing Escherichia coli strains. During the observation window from -4 s (before) up to 27 s after the glucose pulse, the dynamics of the first five enzymatic reactions in the aromatic amino acid pathway was observed by measuring intracellular concentrations of 3-deoxy-d-arabino-heptulosonate 7-phosphate DAH(P), 3-dehydroquinate (3-DHQ), 3-dehydroshikimate (3-DHS), shikimate 3-phosphate (S3P), and shikimate (SHI), together with the pathway precursors phosphoenolpyruvate (PEP) and P5P, the lumped pentose phosphate pool as an alternative to the nondetectable erythrose 4-phosphate (E4P). Provided that a sufficient fortification of the carbon flux into the pathway of interest is ensured, respective metabolism dynamics can be observed. On the basis of the intracellular pool measurements, the standardized pool velocities were calculated, and a simple, data-driven criterion--called "pool efflux capacity" (PEC)--is derived. Despite its simplifying system description, the criterion managed to identify the well-known AroB limitation in the E. coli strain A (genotype delta(pheA tyrA aroF)/pJF119EH aroF(fbr) pheA(fbr) amp) and it also succeeded to identify AroL and AroA (in strain B, genotype delta(pheA tyrA aroF)/pJF119EH aroF(fbr) pheA(fbr) aroB amp) as promising metabolic engineering targets to alleviate respective flux control in subsequent L-Phe producing strains. Furthermore, using of a simple correlation analysis, the reconstruction of the metabolite sequence of the observed pathway was enabled. The results underline the necessity to extend the focus of glucose pulse experiments by studying not only the central metabolism but also anabolic pathways.

Abstract. Diverse modeling and simulation methods are being applied in the area of Systems Biology. Most models in Systems Biology can easily be located within the space that is spanned by three dimensions of modeling: continuous and discrete; quantitative and qualitative; stochastic and deterministic. These dimensions are not entirely independent nor are they exclusive. Many modeling approaches are hybrid as they combine continuous and discrete, quantitative and qualitative, stochastic and deterministic aspects. Another important aspect for the distinction of modeling approaches is at which level a model describes a system: is it at the "macro" level, at the "micro" level, or at multiple levels of organization. Although multi-level models can be located anywhere in the space spanned by the three dimensions of modeling and simulation, clustering tendencies can be observed whose implications are discussed and illustrated by moving from a continuous, deterministic quantitative macro model to a stochastic discrete-event semi-quantitative multi-level model.

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Oxidative racemic resolution of 1,2-diols is a method for the synthesis of enantiopure diols not easily accessed by reduction. The constraints generally found for oxidation to hydroxy ketones can be overcome by coupling various techniques. To circumvent product inhibition, a membrane reactor with solvent extraction of the lipophilic product was chosen. For the oxidative regeneration of NAD+ from NADH anodic oxidation mediated by ABTS was used. The kinetic characteristics of the system were determined independently for each significant system step. However, it proved difficult to simulate the coupled process completely with the kinetic data obtained independently, as true reaction conditions are not covered by kinetic experiments. A mixed approach using a system of ordinary differential equations corrected with data from the process (e.g. enzyme activity) leads to a satisfactory description. This model was applied as a starting point for identifying the relevant process parameters.