Reflections of fermentation

Reflect on this. As white light passes through the back of a glass slide, light beams are reflected and superimposed to form a characteristic interference pattern. A shift in this pattern signals a change in the thickness of the chip. (Adapted from Anal. Chem. 2001, 73, 4313–4318.)

Large-scale fermentation processes are frequently used to produce vital therapeutic compounds, including a range of antibiotics. Generally, these reactions are monitored with expensive HPLC-MS techniques, including somewhat laborious extraction steps and the significant use of organic chemicals. However, according to Günther Jung and colleagues, the simple detection of subtle changes in light interference patterns may facilitate compound-specific on-line monitoring of important industrial fermentation.

Using a technique called reflectometric interference spectroscopy (RIfS), the German scientists directly followed the production of the glycopeptide antibiotic balhimycin in real time (Anal. Chem. 2001, 73, 4313–4318). They accomplished this by taking advantage of the glycopeptide antimicrobial’s high binding specificity to D-Ala-D-peptide termini.

Such peptide structures were immobilized onto a silanized glass slide that was coated with inert material (to prevent nonspecific binding), forming a plate of selective balhimycin targets. When white light is passed through the underside of the plate, it is reflected at each of the different layers—glass, coating, and peptides. By detecting the overall reflected light intensity at each wavelength, a single interference pattern can be plotted. When the fermentation broth is brought into contact with the plate, binding of the antibiotic forms a new layer, shifting the interference plot by a degree proportional to the amount of product that is present, that is, the thickness of the new layer.

Using this optical biosensor approach, the scientists monitored a fermentation reaction for balhimycin over time for product concentration, and the results corresponded almost exactly to those determined from HPLC-MS analysis. The only preparation needed between sampling and analysis in the biosensor method was a short centrifugation step to remove floating particles. The researchers are currently working to replace this step with a filter–dilution system that would be even more amenable to on-line monitoring in industrial fermenters. The RIfS technique should be applicable to a range of other important processes in which distinctive ligand–receptor interactions can be exploited.

—DAVID FILMORE

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