## Processes

Although a batch sterilization process is less successful in avoiding the destruction of nutrients than a continuous one, the objective in designing a batch process is still to achieve the required probability of obtaining sterility with the minimum loss of nutritive quality. The highest temperature which appears to be feasible for batch sterilization is 121°C so the procedure should be designed such that exposure of the medium to this temperature is kept to a minimum. This is achieved by taking into account the contribution made to the sterilization by the heating and cooling periods of the batch treatment. Deindoerfer and Humphrey (1959) presented a method to assess the contribution made by the heating and cooling periods. The following information must be available for the design of a batch sterilization process:

(i) A profile of the increase and decrease in the temperature of the fermentation medium during the heating and cooling periods of the sterilization cycle.

(ii) The number of micro-organisms originally present in the medium.

(iii) The thermal death characteristics of the 'design' organism. As explained earlier this may be Bacillus stearothermophilus or an alternative organism relevant to the particular fermentation.

Knowing the original number of organisms present in the fermenter and the risk of contamination considered acceptable, the required Del factor may be calculated. A frequently adopted risk of contamination is 1 in 1000, which indicates that Nt should equal 10 3 of a viable cell. It is worth reinforcing at this stage that we are considering the total number of organisms present in the medium and not the concentration. If a specific case is considered where the unsterile broth was shown to contain 1011 viable organisms, then the Del factor may be calculated, thus:

Therefore, the overall Del factor required is 32.2. However, the destruction of cells occurs during the heating and cooling of the broth as well as during the period at 121°C, thus, the overall Del factor may be represented as:

Knowing the temperature-time profile for the heating and cooling of the broth (prescribed by the characteristics of the available equipment) it is possible to determine the contribution made to the overall Del factor by these periods. Thus, knowing the Del factors contributed by heating and cooling, the holding time may be calculated to give the required overall Del factor.

Calculation of the Del factor during heating and cooling

The relationship between Del factor, the temperature and time is given by equation (5.8):

V = A-1- e-(E/RT\ However, during the heating and cooling periods the temperature is not constant and, therefore, the calculation of V would require the integration of equation (5.8) for the time-temperature regime observed. Deindoerfer and Humphrey (1959) produced integrated forms of the equation for a variety of temperature-time profiles, including linear, exponential and hyperbolic. However, the regime observed in practice is frequently difficult to classify, making the application of these complex equations problematical. Richards (1968) demonstrated the use of a graphical method of integration and this is illustrated in Fig. 5.7. The time axis is divided into a number of equal increments, tu t2, i3, etc., Richards suggesting 30 as a reasonable number.

Time

Fig. 5.7. The graphical integration method applied to the increase in temperature over a time period. t2, etc. represent equal time intervals (Richards, 1968).

Time

Fig. 5.7. The graphical integration method applied to the increase in temperature over a time period. t2, etc. represent equal time intervals (Richards, 1968).

For each increment, the temperature corresponding to the mid-point time is recorded. It may now be approximated that the total Del factor of the heating-up period is equivalent to the sum of the Del factors of the mid-point temperatures for each time increment. The value of the specific death rate of B. stearother-mophilus spores at each mid-point temperature may be deduced from the Arrhenius equation using the thermal death characteristic published by Deindoerfer and Humphrey (1959). The value of the Del factor corresponding to each time increment may then be calculated from the equations:

The sum of the Del factors for all the increments will then equal the Del factor for the heating-up period. The Del factor for the cooling-down period may be calculated in a similar fashion.

Calculation of the holding time at constant temperature

From the previous calculations the overall Del factor, as well as the Del factors of the heating and cooling parts of the cycle, have been determined. Therefore, the Del factor to be achieved during the holding time may be calculated by difference: V = V — V — V

vholding overall v heating v cooling

Using our example where the overall Del factor is 32.2 and if it is taken that the heating Del factor was 9.8 and the cooling Del factor 10.1, the holding Del factor may be calculated:

But V = kt, and from the data of Deindoerfer and Humphrey (1961) the specific death rate of B. stearothermophilus spores at 121°C is 2.54 min '.

If the contribution made by the heating and cooling parts of the cycle were ignored then the holding time would be given by the equation:

Thus, by considering the contribution made to the sterilization process by the heating and cooling parts of the cycle a considerable reduction in exposure time is achieved

Richards' rapid method for the design of sterilization cycles

Richards (1968) proposed a rapid method for the design of sterilization cycles avoiding the time-consuming graphical integrations. The method assumes that all spore destruction occurs at temperatures above 100°C and that those parts of the heating and cooling cycle above 100° are linear. Both these assumptions appear reasonably valid and the technique loses very little in accuracy and gains considerably in simplicity. Furthermore, based on these assumptions, Richards has presented a table of Del factors for B. stearothermophilus spores which would be obtained in heating and cooling a broth up to (and down from) holding temperatures of 101-130°C, based on a temperature change of 1°C per minute. This information is presented in Table 5.2, together with the specific death rates for B. stearothermophilus spores over the temperature range. If the rate of temperature change is 1° per minute, the Del factors for heating and cooling may be read directly from the table; if the temperature change deviates from 1° per minute, the Del factors may be altered by simple proportion. For example, if a fermentation broth were heated from 100° to 121°C in 30 minutes and cooled from 121° to 100° in 17 minutes, the Del factors for the heating and cooling cycles may be determined as follows:

From Table 5.2, if the change in temperature had been 1° per minute, the Del factor for both the heating and cooling cycles would be 12.549. But the temperature change in the heating cycle was 21° in 30 minutes; therefore,

and the temperature change in the cooling cycle was 21° in 17 minutes, therefore,

Having calculated the Del factors for the heating and cooling periods the holding time at the constant temperature may be calculated as before.

The scale up of batch sterilization processes

The use of the Del factor in the scale up of batch sterilization processes has been discussed by Banks

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