Automatic and accurate sampling is both convenient and sometimes necessary to obtain detailed information about cell cultures. We developed an autoclavable sampling system in which culture broth was pumped through an ultrafiltration cross-flow module with a novel filtrate collecting principle and a novel regulation of filter back pressure. Filtrate was collected from equal membrane filter areas through holes connected to channels with an even length to the collecting point, resulting in a near plug flow of filtrate and a reduction in the response time to 1 min (98% of full signal of the tracer molecule glucose). Constant pressure difference (0.3 bar) across the membrane filter (30 kDa cutoff value) and prevention of leakage was obtained by squeezing the tubing with culture broth between two flexible spring steel plates fixed at one corner (filtrate flow 1 ml min¿1). The large contact area allowed the tubing to open the passage more when pressure increased. Using this design of sampling system, the metabolite profiles of Aspergillus niger wild type and a phosphofructokinase overexpressing strain (three times wild type) were concluded to be indistinguishable by detailed monitoring of fast transients of substrates and products in batch culture and glucose pulse experiments. The combination of the fast response filter module and prevention of high pressure peaks with the flexible resistance to flow enables long-term (>5 days) and automatic monitoring of cultures of filamentous fungi or other microorganisms with fast changes in extracellular concentrations.
- column liquid-chromatography