The dilution rate varies with the population density of the culture and maintains the density within a narrow range. Organisms grow at the maximum rate characteristic of the organism and the conditions. The growth rate can be changed by modifying the nutrient medium, gas concentration, or incubation temperature. A disadvantage of the turbidostat is that all nutrient concentrations in the culture chamber are necessarily higher than the minimum, resulting in inefficient utilization of nutrients.
The turbidostat system for continuous culture of Hydrogenomonas bacteria, developed by Battelle Memorial Institute ([ref.194]), includes electrolysis of water in a separate unit. Hydrogen and oxygen are fed separately up to the point of injection into the culture vessel, and the mixed volume is kept very small to minimize am possibility of explosion. However, the two gases may be injected simultaneously if there is a demand for both.
In the chemostat, growth of the organisms is limited by maintaining one essential nutrient concentration below optimum. A constant feed of medium, with one nutrient in limiting concentration and with constant removal of culture at the same rate, is used to achieve the steady state. The dilution rate is set at an arbitrary value, and the microbial population is allowed to find its own level. By appropriate setting of the dilution rate, the growth rate may be held at any desired value from slightly below the maximum possible to nearly zero. This constitutes a self-regulating system and allows selection of a desired growth rate.
A combined electrolysis-chemostat method, developed by Magna Corp., maintained the hydrogen-producing electrode of an electrolysis cell in the bacterial culture. Resting cells of Hydrogenomonas eutropha consumed hydrogen produced at the cathode of an electrolysis cell built into a specially constructed Warburg flask. Attempts to immobilize Hydrogenomonas cells on a porous conductor were partially successful. This system could lower the volume requirements compared with those for the isolated subsystems. Disadvantages of this integrated system include electrolysis of the bacterial medium, possibly resulting in toxic breakdown products, and the possible effects of electric power and the KOH electrolyte on the bacteria. The main disadvantage of an integrated system would be the disparity between optimal conditions for efficient electrolysis and efficient bacterial conversion, particularly temperature and pH, with the combination possibly resulting in considerably higher power and weight demands.
Both continuous-culture approaches are being studied with NASA support. The turbidostat offers the greatest potential efficiency in weight and volume, but uses nutrient materials less efficiently and is more complex. The chemostat is less efficient in weight and volume, but has greater simplicity and reliability.
Hydrogenomonas eutropha has been grown in 15-liter batch cultures and in 2.1-liter continuous cultures. A 20-liter continuous culture, sufficient to balance the requirements of a man, is under development.
The potential problem areas in large-scale continuous production of the bacteria include assuring genetic stability, preventing or controlling bacteriophage and foreign bacterial contamination, and preventing heterotrophic growth caused by exposure to organic material from the urine. Genetics of hydrogen bacteria and phage infection have been studied by DeCicco. Research on these problems indicates that they are not of major importance, but cause significant effects and must be eliminated or controlled.
Bacterial Composition and Nutrition
Hydrogenomonas bacteria can be used for at least part of the astronauts' diet. The washed bacteria have a mild taste and are being studied for their total energy content, protein and lipid digestibility, and vitamin content. Carbon and nitrogen balances, and respiratory quotient are to be determined in animals fed the bacteria as their sole food source. No toxic constituents have been discovered. Sonicated and cooked bacteria, when fed to white rats as 12 percent of the solids of a nutritionally balanced diet, were eaten readily and produced no ill effects. Net utilization of the protein appears to be somewhat lower than casein and about the same as legume proteins.
The composition of Hydrogenomonas eutropha is shown in [table XI]. The composition of the bacteria varies with the age and growth phase of the cells and with the medium and gas available. It is possible to modify the growth conditions to grow the type of bacteria desired for nutritive purposes.