Starter cultures sterilization

The upstream processing element of the manufacture of a batch of biopharmaceutical product begins with the removal of a single ampoule of the working cell bank. This vial is used to inoculate a small volume of sterile media, with subsequent incubation under appropriate conditions. This describes the growth of laboratory-scale starter cultures of the producer cell line. This starter culture is, in turn, used to inoculate a production-scale starter culture that is used to inoculate the production-scale bioreactor (Figure 5.7). The media composition and fermentation conditions required to... 

Reiter et al. (1964) showed that growth of S. aureus in raw, steamed, and pasteurized milk was inhibited by a lactic starter culture. When they neutralized the lactic acid as it was produced, inhibition of the staphylococcus was still evident. Jezeski et al. (1967) also observed that growth of S. aureus in steamed or sterile reconstituted nonfat dry milk was inhibited by an actively growing S. lactis culture. Enter-otoxin was detected in S. aureus-S. lactis mixed cultures when S. lactis was inactivated by bacteriophage but not when the lactic streptococcus grew normally. Further information on S. aureus has been summarized by Minor and Marth (1976). 

Next, grow starter cultures by picking a colony from the above plates into 2 mL LB plus 50 pg/mL kanamycin in sterile 24-well blocks. Cover the blocks with air-pore tape and grow overnight at 37°C, shaking at 180-200 rpm. 

Fermentation can be spontaneous or be induced by specifically added microorganisms. An everyday example of such an induced fermentation is the addition of baking yeast to flour to make bread or cakes. As with bread, fermentation can be done in a normal environment where many different microorganisms are present. A more sophisticated way is to exclude unwanted microorganisms by sterilization of the materials before adding a starter culture. 

The fermentation is usually continuous it proceeds under sterile conditions, at constant temperature, and is started with a defined starter culture to avoid side products as far as possible. Several processes were developed Shell had originally introduced a process that used methane (natural gas) as the feedstock for SCP production. The microorganisms are cultured in an aqueous medium at temperatures of 42 to 45°C and at a pH value of 6.8 under semisterile conditions. The final fermentation broth contains protein at a concentration of 25 g/L. The biomass is concentrated in large sedimentation tanks and then spray-dried. The mass balance equation (Eq. 9.3) shows that large volumes of oxygen are needed and that carbon dioxide and heat must be removed from the reactor. 

Besides the application of a well defined starter culture which has to be tested every time before use, the preconditioning of the food raw material is of critical significance. This preconditioning ranges from simple washing to complete sterilization. Besides, the supplementation of nutrients and the adjustment of a defined pH-value is frequently decisive for the result of a fermentation. 

Successful cultivation of microorganisms for growth and identification requires use of various types of media, either liquid (referred to as hroths) or solid by inclusion of agar. For winemakers, a growth medium may he as simple as diluted sterile grape juice for the activation and expansion of yeast starter cultures or as complicated as that necessary to grow lactic acid bacteria. 

The ability to transfer microorganisms from one container to another without contamination is crucial to success in the microbiology laboratory. These techniques serve as the basis for subsequent work such as starter culture preparation or maintaining viable cultures in long-term storage. Transfer loops are normally used to transfer to the surface of agar (Petri plates and slants), whereas transfer needles are used to prepare stab cultures. Both implements are sterilized by heating in an open flame until red hot (Fig. 13.1). 

Starter cultures are continuously exposed to phage attack in food fermentations. Dairy fermentations are particularly susceptible to phage attack due to the rather limited starter culture repertoire and the use of non-sterile substrates such as milk. Moreover, in many dairy plants starter cultures are cultivated in large volumes and in rich media, which are ideal conditions for phage propagation. Therefore, strategies to avoid the presence and survival of phages in dairy factories have always been a matter of concern. 

Koji starter, also known as seed koji, koji seeds, or tane-koji, provides spores of microorganisms to make koji. Preparation of koji starter is essentially the same as making regular koji for soy paste and soy sauce except that in making koji starter, pure culture and different raw materials are used and longer fermentation is needed to produce abundant spores. In addition, a sterile condition is needed to avoid contamination. 

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