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(1979). It should be appreciated by this stage that the Del factor does not include a volume term, i.e. absolute numbers of contaminants and survivors are considered, not their concentration. Thus, if the size of a fermenter is increased the initial number of spores in the medium will also be increased, but if the same probability of achieving sterility is required the final spore number should remain the same, resulting in an increase in the Del factor. For example, if a pilot sterilization were carried out in a 1000-dm3 vessel with a medium containing 106 organisms cm-3 requiring a probability of contamination of 1 in 1000, the Del factor would be:

= In 1015 = 34.5. If the same probability of contamination were required in a 10,000-dm3 vessel using the same medium the Del factor would be:

V = In {(106 X 103 X 104)/10-3} = ln(10l3/10-3) = In 1016 = 36.8. Thus, the Del factor increases with an increase in the size of the fermenter volume. The holding time in the large vessel may be calculated by the graphical integration method or by the rapid method of Richards (1968), as discussed earlier, based on the temperature-time profile of the sterilization cycle in the large vessel. However, it must be appreciated that extending the holding time on the larger scale (to achieve the increased V factor) will result in increased nutrient degradation. Also, the contribution of the heating-up and cooling-down periods to nutrient destruction will be greater as scale increases. Maintaining the same nutrient quality on a small and a large scale can be achieved in batch sterilization only by compromising the sterility of the vessel, which is totally unacceptable. Thus, the decrease in the yield of a fermentation when it is scaled up is often due to problems of nutrient degradation during batch sterilization and the only way to eradicate the problem is to sterilize the medium continuously.

Methods of batch sterilization

The batch sterilization of the medium for a fermentation may be achieved either in the fermentation vessel or in a separate mash cooker. Richards (1966) considered the relative merits of in situ medium sterilization and the use of a special vessel. The major advantages of a separate medium sterilization vessel may be summarized as:

(i) One cooker may be used to serve several fermenters and the medium may be sterilized as the fermenters are being cleaned and prepared for the next fermentation, thus saving time between fermentations.

(ii) The medium may be sterilized in a cooker in a more concentrated form than would be used in the fermentation and then diluted in the fermenter with sterile water prior to inoculation. This would allow the construction of smaller cookers.

(iii) In some fermentations, the medium is at its most viscous during sterilization and the power requirement for agitation is not alleviated by aeration as it would be during the fermentation proper. Thus, if the requirement for agitation during in situ sterilization were removed, the fermenter could be equipped with a less powerful motor. Obviously, the sterilization kettle would have to be equipped with a powerful motor, but this would provide sterile medium for several fermenters.

(iv) The fermenter would be spared the corrosion which may occur with medium at high temperature.

The major disadvantages of a separate medium sterilization vessel may be summarized as:

(i) The cost of constructing a batch medium sterilizer is much the same as that for the fermenter.

(ii) If a cooker serves a large number of fermenters complex pipework would be necessary to transport the sterile medium, with the inherent dangers of contamination.

(iii) Mechanical failure in a cooker supplying medium to several fermenters would render all the fermenters temporarily redundant. The provision of contingency equipment may be prohibitively costly.

Overall, the pressure to decrease the 'down time' between fermentations has tended to outweigh the perceived disadvantages of using separate sterilization vessels. Thus, sterilization in dedicated vessels is the method of choice for batch sterilization. However, as pointed out by Corbett (1985), the fact that separate batch sterilizers are used lends further weight to the argument that continuous sterilization should be adopted in preference to batch. The capital cost of a separate batch sterilizer is similar to that of a continuous one and the problems of transfer of sterile media are then the same for both batch and continuous sterilization. Thus, two of the major objections to continuous systems (capital cost and aseptic transfer) may be considered as no longer relevant.

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