## Medium Optimization

At this stage it is important to consider the optimization of a medium such that it meets as many as possible of the seven criteria given in the introduction to this chapter. The meaning of optimization in this context does need careful consideration (Winkler, 1991). When considering the biomass growth phase in isolation it must be recognized that efficiently grown biomass produced by an 'optimized' high productivity growth phase is not necessarily best suited for its ultimate purpose, such as synthesizing the desired product. Different combinations and sequences of process conditions need to be investigated to determine the growth conditions which produce the biomass with the physiological state best constituted for product formation. There may be a sequence of phases each with a specific set of optimal conditions.

Medium optimization by the classical method of changing one independent variable (nutrient, antifoam, pH, temperature, etc.) while fixing all the others at a certain level can be extremely time consuming and expensive for a large number of variables. To make a full factorial search which would examine each possible combination of independent variable at appropriate levels could require a large number of experiments, x", where * is the number of levels and n is the number of variables. This may be quite appropriate for three nutrients at two concentrations (23 trials) but not for six nutrients at three concentrations. In this instance 3C (729) trials would be needed. Industrially the aim is to perform the minimum number of experiments to determine optimal conditions. Other alternative strategies must therefore be considered which allow more than one variable to be changed at a time. These methods have been discussed by Stowe and Mayer (1966), McDaniel et al. (1976), Hendrix (1980), Nelson (1982), Greasham and Inamine (1986), Bull et al. (1990) and Hicks (1993).

When more than five independent variables are to be investigated, the PlackettBurman design may be used to find the most important variables in a system, which are then optimized in further studies (Plackett and Burman, 1946). These authors give a series of designs for up to one hundred experiments using an experimental rationale known as balanced incomplete blocks. This technique allows for the evaluation of X — 1 variables by X experiments. X must be a multiple of 4, e.g. 8, 12, 16, 20, 24, etc. Normally one determines how many experimental variables need to be included in an investigation and then selects the Plackett Burman design which meets that requirement most closely in multiples of 4. Any factors not assigned to a variable can be designated as a dummy variable. Alternatively, factors known to not have any effect may be included and designated as dummy variables. As will be shown shortly in a worked example (Table 4.16), the incorporation of dummy variables into an experiment makes it possible to estimate the variance of an effect (experimental error).

Table 4.16 shows a Plackett Burman design for seven variables (A-G) at high and low levels in which two factors, E and G, are designated as 'dummy' variables. These can then be used in the design to obtain an estimate of error. Normally three dummy variables will provide an adequate estimate of the error. However, more can be used if fewer real variables need to be studied in an investigation (Stowe and Mayer, 1966). Each horizontal row represents a trial and each vertical

Trial |

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