This practical part serves as an introduction to the work with microorganisms and the necessary equipment.
- Production of solid and liquid media
- Sterile working under the Cleanbench
- Cultivation of microorganisms in shaking flasks
- Microscopy and cell counting
- Determination of biomass via bio dry mass or by photometer
Block II: Fermentation
In this block the operation and control of a fermenter and the supervision of a cultivation should be practiced. Thereby, operating parameters of the fermenter will be adjusted and a cell culture from pre-culture to cell harvest will be performed.
- Preparation of a cultivation (autoclaving, inoculation etc.)
- Operating parameters for a submerged cultivation (pH-value, T, kla-value)
- Monitoring the growth of microorganisms (bio-dry matter, metabolism)
Block III: Reconditioning
The aim of the experiment is to isolate an enzyme (e.g. alcohol dehydrogenase) from a microorganism (e.g. Saccharomyces cerevisiae) using various purification steps. Starting from the raw extract of the cell disruption, different methods are applied, each successively increasing the purity of the enzyme to be isolated.
- Cell Digestion
- Heat denaturation of proteins
- Salting out proteins
- Chromatographic protein purification
- Affinity chromatography
Block IV: Biotransformation
The aim of biotransformation is the determination of the Michaelis constant and the maximum reaction rate of an enzyme (e.g. of lactate dehydrogenase from rabbit muscle) and the inhibition of enzyme activity by various inhibitors (e.g. oxamate and reduced nicotinamide adenine dinucleotide (NADH+H+)).
- Conversion of substrates by biocatalysts
- Analysis of kinetic data