Chemostat

A chemostat (from chemical environment is static) is a bioreactor to which fresh medium is continuously added, while culture liquid containing left over nutrients, metabolic end products and microorganisms are continuously removed at the same rate to keep the culture volume constant. By changing the rate with which medium is added to the bioreactor the specific growth rate of the microorganism can be easily controlled within limits. A chemostat (from chemical environment is static) is a bioreactor to which fresh medium is continuously added, while culture liquid containing left over nutrients, metabolic end products and microorganisms are continuously removed at the same rate to keep the culture volume constant. By changing the rate with which medium is added to the bioreactor the specific growth rate of the microorganism can be easily controlled within limits. One of the most important features of chemostats is that microorganisms can be grown in a physiological steady state under constant environmental conditions. In this steady state, growth occurs at a constant specific growth rate and all culture parameters remain constant (culture volume, dissolved oxygen concentration, nutrient and product concentrations, pH, cell density, etc.). In addition, environmental conditions can be controlled by the experimenter. Microorganisms growing in chemostats usually reach a steady state because of a negative feedback between growth rate and nutrient consumption: if a low number of cells are present in the bioreactor, the cells can grow at growth rates higher than the dilution rate as they consume little nutrient so growth is less limited by the addition of limiting nutrient with the inflowing fresh medium. The limiting nutrient is a nutrient essential for growth, present in the medium at a limiting concentration (all other nutrients are usually supplied in surplus). However, the higher the number of cells becomes, the more nutrient is consumed, lowering the concentration of the limiting nutrient. In turn, this will reduce the specific growth rate of the cells which will lead to a decline in the number of cells as they keep being removed from the system with the outflow. This results in a steady state. Due to the self-regulation, the steady state is stable. This enables the experimenter to control the specific growth rate of the microorganisms by changing the speed of the pump feeding fresh medium into the vessel. Another important feature of chemostats and other continuous culture systems is that they are well-mixed so that environmental conditions are homogenous or uniform and microorganisms are randomly dispersed and encounter each other randomly. Therefore, competition and other interactions in the chemostat are global, in contrast to biofilms. The rate of nutrient exchange is expressed as the dilution rate D. At steady state, the specific growth rate μ of the micro-organism is equal to the dilution rate D. The dilution rate is defined as the flow of medium per unit of time, F, over the volume V of culture in the bioreactor Specific growth rate μ is inversely related to the time it takes the biomass to double, called doubling time td, by: Therefore, the doubling time td becomes a function of dilution rate D in steady state: Each microorganism growing on a particular substrate has a maximal specific growth rate μmax (the rate of growth observed if growth is limited by internal constraints rather than external nutrients). If a dilution rate is chosen that is higher than μmax, the cells cannot grow at a rate as fast as the rate with which they are being removed so the culture will not be able to sustain itself in the bioreactor, and will wash out. However, since the concentration of the limiting nutrient in the chemostat cannot exceed the concentration in the feed, the specific growth rate that the cells can reach in the chemostat is usually slightly lower than the maximal specific growth rate because specific growth rate usually increases with nutrient concentration as described by the kinetics of the Monod equation. The highest specific growth' rates (μmax) cells can attain is equal to the critical dilution rate (D'c): where S is the substrate or nutrient concentration in the chemostat and KS is the half-saturation constant (this equation assumes Monod kinetics).