An automated approach for rapid on-line monitoring of the solid and liquid phases present in bioprocesses based on mid-IR Fourier transform spectrometry is introduced. The principles of this new approach are presented using the example of the quantitative and qualitative analysis of poly(beta-hydroxybutyric acid) (PHB) accumulated in living bacterial cells as well as dissolved glucose during two 27-h fermentation processes. FT-IR spectra were recorded on-line using a diamond-attenuated total reflection (ATR) cell connected to the fermentation broth by means of a computer-controlled flow system. For calibration of the FT-IR method, standard reference analysis procedures for PHB (gas chromatography) and glucose (HPLC) were used. While pumping the fermentation broth through the flow cell, the recorded spectra corresponded to the fermentation solution, which allowed the determination of glucose in a range from 3.8 to -10.3 g/L. Upon stopping the flow, the cells settled on the ATR surface, and the thus recorded spectra enabled the determination of the intracellular PHB content of Escherichia coli in a range from 0.005 to 0.766 g/L. Errors of cross-validation of 0.264 g/L for glucose and 0.037 g/L for PHB were obtained. Application of one PLS calibration model to another fermentation was possible with prediction errors of 0.493 g/L for glucose and 0.105 g/L for PHB. Furthermore, from the position and shape of the PHB carbonyl band, it could be concluded that the PHB granules inside the E. coli are predominantly amorphous.

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In this chapter we report on the molecular biology of crystalline surface layers of different bacterial groups. The limited information indicates that there are many variations on a common theme. Sequence variety, antigenic diversity, gene expression, rearrangements, influence of environmental factors and applied aspects are addressed. There is considerable variety in the S-layer composition, which was elucidated by sequence analysis of the corresponding genes. In Corynebacterium glutamicum one major cell wall protein is responsible for the formation of a highly ordered, hexagonal array. In contrast, two abundant surface proteins from the S-layer of Bacillus anthracis. Each protein possesses three S-layer homology motifs and one protein could be a virulence factor. The antigenic diversity and ABC transporters are important features, which have been studied in methanogenic archaea. The expression of the S-layer components is controlled by three genes in the case of Thermus thermophilus. One has repressor activity on the S-layer gene promoter, the second codes for the S-layer protein. The rearrangement by reciprocal recombination was investigated in Campylobacter fetus. 7-8 S-layer proteins with a high degree of homology at the 5' and 3' ends were found. Environmental changes influence the surface properties of Bacillus stearothermophilus. Depending on oxygen supply, this species produces different S-layer proteins. Finally, the molecular bases for some applications are discussed. Recombinant S-layer fusion proteins have been designed for biotechnology.

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