Experimental Planning

Once the experimental system has been selected, it is necessary to decide how to carry out the experiment, namely, how many samples will be removed, at what times, and what will be analyzed. The flexibility to decide how many samples will be removed during the fermentation might be limited by the resources available, especially in the case of the Raimbault columns. In terms of the later mathematical analysis, it would be desirable for the data points to be evenly spaced over the whole of the active growth phase, including the acceleration and deceleration phases. It may be necessary to undertake the first fermentation simply to see when things happen, before undertaking a second experiment with better planning of the times at which samples are to be removed (Fig. 15.4).

It is advisable to record sufficient information for the biomass profile to be plotted in terms of both grams of biomass per gram of initial dry substrate (g-biomass g-IDS-1) (i.e., on an absolute basis) and grams of biomass per gram of dry solids (i.e., on a relative basis). See Sect. 14.3 for an explanation of these terms. The following procedure, shown schematically in Fig. 15.5, explains how this can be done. Figure 15.5 also shows the meaning of the various symbols used in the equations presented below. It talks in terms of "flasks", but the same procedure applies if Raimbault columns are used. It assumes that:

• two samples are removed from the same flask (after mixing of the contents): one for moisture content determination and the other for analysis of the amount of biomass or a component of the biomass. Another strategy would be to sacrifice two flasks at each sampling time, using the entire contents of one for biomass determination and the entire contents of the other for determination of the moisture content.

• the biomass determination is done on a moist sample and not a dried sample.

The mass of moist substrate added initially to each flask should be measured, and a sample of this substrate should also be dried in order to determine the initial water content (IWC, expressed as a percentage and on a wet basis). The initial water content should be determined after inoculation, especially if the inoculum brings a significant amount of water into the system. It can be calculated from the moist and dry weights of the removed sample (Fig. 15.5):

Since the original amount of moist substrate added to each flask is known (Moi g added to the "ith" flask), it is simple to calculate the amount of "initial dry substrate" (IDS) that the ith flask holds:

At the time of sampling of the ith flask, after mixing of the substrate in the flask, a sample should be removed, weighed, dried, and reweighed, allowing the moisture content at the time of sampling (WC) to be calculated:

Fig. 15.4. Planning of sampling times. (a) A preliminary experiment should be done to identify at approximately what time the rapid growth period occurs; (b) This information can be used to plan for several samples to be removed during the period of most rapid growth, in order to characterize the growth curve well; (c) The profile that would be obtained if evenly-spaced samples were planned without first identifying the period of rapid growth. Insufficient samples would be removed during the rapid growth period; (d) The underlying fermentation profile assumed in this example. Of course, this profile is an unknown when the initial experimental planning is being undertaken

Fig. 15.4. Planning of sampling times. (a) A preliminary experiment should be done to identify at approximately what time the rapid growth period occurs; (b) This information can be used to plan for several samples to be removed during the period of most rapid growth, in order to characterize the growth curve well; (c) The profile that would be obtained if evenly-spaced samples were planned without first identifying the period of rapid growth. Insufficient samples would be removed during the rapid growth period; (d) The underlying fermentation profile assumed in this example. Of course, this profile is an unknown when the initial experimental planning is being undertaken dry and weigh reweigh a m0-dn inoculated prepared substrate fill pre-weighed flasks

initial moist substrate weights

reweigh

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