0 10 30 50 70 90 Duration of sterilization (min)
Fig. 5.5. The effect of the time of sterilization on the yield of a subsequent fermentation (Richards, 1966).
The thermal destruction of essential media components conforms approximately with first order reaction kinetics and, therefore, may be described by equations similar to those derived for the destruction of bacteria:
where xt is the concentration of nutrient after a heat treatment period, t, x0 is the original concentration of nutrient at the onset of sterilization, k is the reaction rate constant. It is important to appreciate that we are considering the decline in the concentration of the nutrient component, whereas we consider the decline in the number of contaminants. The effect of temperature on the reaction rate constant may be expressed by the Arrhenius equation:
Therefore, a plot of the natural logarithm of the reaction rate against 1/T will give a straight line, slope — (E/R). As the value of R, the gas constant, is fixed
Table 5.1. The effect of sterilization time on glucose concentration and product accretion rate in an antibiotic fermentation (Corbett, 1985)
Time at Amount of added Relative
121° (min) glucose remaining (%) accretion rate
60 35 90
40 46 92
30 64 100
the slope of the graph is determined by the value of the activation energy (E). The activation energy for the thermal destruction of B. stearothermophilus spores has been cited as 67.7 kcal mole™1, whereas that for thermal destruction of nutrients is 10 to 30 kcal mole"' (Richards, 1968). Figure 5.6 is an Arrhenius plot for two reactions — one with a lower activation energy than the other. From this plot it may be seen that as temperature is increased, the reaction rate rises more rapidly for the reaction with the higher activation energy. Thus, considering the difference between activation energies for spore destruction and nutrient degradation, an increase in temperature would accelerate spore destruction more than medium denaturation.
In the consideration of Del factors it was evident that the same Del factor could be achieved over a range of temperature/time regimes. Thus, it would appear to be advantageous to employ a high temperature for a short time to achieve the desired probability of sterility, yet causing minimum nutrient degradation. Thus, the ideal technique would be to heat the fermentation medium to a high temperature, at which it is held for a short period, before being cooled rapidly to the fermentation temperature. However, it is obviously impossible to heat a batch of many thousands of litres of broth in a tank to a high temperature, hold for a short period and cool without the heating and cooling periods contributing considerably to the total sterilization time. The only practical method of materializing the objective of a short-time, high-temperature treatment is to sterilize the medium in a continuous stream. In the past the fermentation industry was reluctant to adopt continuous sterilization due to a number of disadvantages outweighing the advantage of nutrient
quality. The relative merits of batch and continuous sterilization may be summarized as follows-
Advantages of continuous sterilization over batch sterilization
(i) Superior maintenance of medium quality.
(iii) Easier automatic control.
(iv) The reduction of surge capacity for steam.
(v) The reduction of sterilization cycle time.
(vi) Under certain circumstances, the reduction of fermenter corrosion.
Advantages of batch sterilization over continuous sterilization
(i) Lower capital equipment costs.
(ii) Lower risk of contamination — continuous processes require the aseptic transfer of the sterile broth to the sterile vessel.
(iii) Easier manual control.
(iv) Easier to use with media containing a high proportion of solid matter.
The early continuous sterilizers were constructed as plate heat exchangers and these were unsuitable on two accounts:
(i) Failure of the gaskets between the plates resulted in the mixing of sterile and unsterile streams.
(ii) Particulate components in the media would block the heat exchangers.
However, modern continuous sterilizers use double spiral heat exchangers in which the two streams are separated by a continuous steel division. Also, the spiral exchangers are far less susceptible to blockage. However, a major limitation to the adoption of continuous sterilization was the precision of control necessary for its success. This precision has been achieved with the development of sophisticated computerized monitoring and control systems resulting in continuous sterilization being very widely used and it is now the method of choice. However, batch sterilization is still used in many fermentation plants and, thus, it will be considered here before continuous sterilization is discussed in detail.
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