Method of continuously preparing a sterile culture medium

Details Method of continuously preparing a sterile culture medium Method of continuously preparing a sterile culture medium

1994-07-11

1996-04-16

Czaja, Donald E.

Chemistry: molecular biology and microbiology

Apparatus

Bioreactor

210650, 210651, 210653, 210654, 210655, 210788, 422 93, 4352971, C12M 310, C12M 112, B01D 3300

Patent

active

055081967

DESCRIPTION:

BRIEF SUMMARY
The invention concerns a method of continuously preparing a sterile culture medium, wherein various components are combined into a single mainstream appropriate for charging a bioengineering reactor.
The processing-technology stages in a continuous-operation bioengineering reactor essentially comprise culture-medium sterilization, fermentation, and product finishing or product analysis. Successful continuous operation of such a reactor depends on the quality of the incoming sterile medium among other factors. Three methods of preparing it are known, specifically high-temperature short residence time (HTSRT), filtering it sterile, and the use of contaminant-insensitive media.
In HTSRT the flowing non-sterile medium is heated in a heat exchanger to approximately 135.degree. and maintained at that temperature for a few minutes. Any microorganisms are exterminated, and the result is a sterile culture medium. Although it exterminates germs very effectively, there are drawbacks to the method. Various components of the media tend to interact, sugar molecules and amino acids into melanoids for example. Again, some components, vitamins for example, are destroyed by heat.
In filtering sterile, the filter is interposed in the line leading to the reactor and the medium arrives sterile. Since this approach does not involve heat, the quality of the resulting medium is high. When several media are sterilized with the same filter, however, the device will clog up rapidly and becomes unreliable. This approach is accordingly employed fairly infrequently.
Contamination-insensitive components automatically result in a sterile culture medium because they by their very nature do not allow contaminants to proliferate. The only appropriate applications, however, are fermentation processes that occur in a highly acidic milieu or include antibiotics in the culture medium. The use of such procedures is limited.
The object of the present application is a method of continuously preparing a sterile culture medium that can be universally employed, will allow long-term continuous operation, and ensures careful treatment of the culture media.
This object is attained in accordance with the present invention as will now be described. The components arrive in tributaries. The still unsterilized components are combined into a mainstream. The mainstream is forwarded to a transverse-flow filtration module. The module accommodates a membrane that separates the mainstream into a permeate and a retentate. The permeate is forwarded to the reactor. The retentate is diverted to a centrifuge that precipitates any germs. The centrifuged retentate is returned to the mainstream flowing toward the filtration module.
The various components can accordingly be combined in accordance with the present invention even before they are sterilized, which simplifies management and increases flexibility with respect to combining various components. The centrifuge continuously centrifuges microorganisms out of the retentate lines and prevents them from proliferating on or in the membrane. The centrifuge also automatically maintains a high rate of recirculation, which also inhibits the growth of microorganisms on the membrane. Finally, the centrifuge ensures adequate pressure on the membrane.
The mainstream is maintained at a temperature of at least 70.degree. C. by a heat exchanger once the retentate has been returned in one advantageous embodiment of the method. This procedure exterminates many germs and inhibits the growth of heat-resistent types, preventing overload of the centrifuge and filtration module. This temperature, however, is not high enough to damage the components of the media.
The diameters of the pores in the filter membrane in another advantageous embodiment of the method are 0.2 .mu.m or less. This dimension has been demonstrated to allow rapid permeate flow while maintaining the sterilizing capability of the module. The best results have been obtained with pores 0.14 .mu.m in diameter.
To prolong the life of the filtration module, the retentate sho

REFERENCES:
patent: 4832851 (1989-05-01), Bowers et al.
patent: 5174900 (1992-12-01), Nichols
Lavsen et al. Biotechnology and Bioengineering vol. XVIII pp. 1433-1443 (1976).

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