Starter Culture Preparation

Techniques for growing/expanding LAB starters vary somewhat but can generally be divided into either pure culture or coculture (LAB and wine yeast) methods. In the latter case, most utilize the same yeast strain that is used to carry out alcoholic fermentation. In either case, a properly prepared and expanded LAB starter should yield a stationary-phase inoculum of 10 CFU/mL. 

Inoculum levels generally parallel those of yeast, in the range of 1-5 x 10 cells/mL, corresponding to 1-5% (vol/vol). Where inoculation to red wines is planned during the course of alcoholic fermentation, the volume of starter required should reflect the larger volume of must and not press or finished wine volumes (van der Water, 1993, personal communication). In the case of white wine fermentations or additions to recently fermented wine, the volume of solids is not an issue. 

With one recent exception (Vinflora oenos), rehydration of freeze-dried cultures is usually recommended in sterile distilled or deionized water followed by transfer to expansion media. Most LAB cultures prepared from either laboratory cultures, concentrates, or rehydrated from freeze-dried preparations must be propagated and expanded under winelike conditions prior to inoculation into the total volume of fermentation or wine. This acclimation period is necessary to prevent the significant population crashes observed upon direct inoculation into wine. 

Numerous growth and/or expansion media have been proposed over the years. These have included, at least in the early stages of propagation, 

Ethanol, at 2% (vol/vol), was also be found to stimulatory to LAB starters. Hayman and Monk (1982) reported that inclusion of 1 volume of sterile-filtered (unsulfited) wine to 5 volumes of juice yielded the best growth. It is likely that the addition of wine during expansion enhances acclimation (to both alcohol and wine pH) and, hence, increased numbers of survivors upon final addition to wine. 

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Henick-Kling, T. (1995). Control of malo-lactic fermentation in wine Energetics, flavour modification and methods of starter culture preparation. J. Appl. Bacteriol. 79, 29S-37S. Imazio, S., Labra, M., Grassi, F., Scienza, A., and Failla, O. (2006). Chloroplast microsatellites to investigate the origin of grapevine. Genet. Resour. Crop Evol. 53,1003-1011. 

Henick-Kling, T. 1995. Control of malo-lactic fermentation in wine Energetics, flavour modification and methods of starter culture preparation. J. Appl. Bacterial Symp. Supp. 79, 29S-37S. 

The dried cell concentrate is typically produced by specialized suppliers and distributed worldwide. As the producers of fermented foods want to build up some stock, the cultures should be stable for at least one year. As the activity of the cultures after drying depends on the previous fermentation step (see Section 11.3), and the storage stability depends on the drying process as we will show in this section, the starter culture preparation process must be looked at in a holistic approach. This... 

Hayman, D.C. and P.R. Monk. 1982. Starter culture preparation for the induction of malolactic fermentations in wine. Food Tech. Aust. 34 14-18. 

Lactic acid bacteria. The number of species within lactic acid bacteria used for sausage fermentation is modest. According to Hammes et al. (1985) the following five species can be found in starter culture preparations Lactobacillus plantarum, Lactobacillus sake, Lactobacillus curvatus, Pediococcus pentosaceus and Pediococcus acidilactici. In addition, Lactobacillus pentosus is also used. 

Other microorganisms. In mixed starter culture preparations, Streptomyces and Debaryomyces can be found. Streptomyces griseus subsp. Hiitter is used for introduction of a cellar-ripened sausage aroma and a better colour is obtained due to the enzymic activities of nitrate reductase and catalase (Eilberg and Liepe, 1977). The yeast Debaryomyces hansenii is also applied for aroma enhancement. 

Starter culture prepared using diluted grape juice medium. 

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). 

SokoEek SJ, Hammes WP (1997) Description of a starter culture preparation for vinegar fermentation. Syst Appl Microbiol 20 481-491... 

Starter culture bacteria dominate over naturally occurring microflora. The microorganisms perform the required metabolic activities. Starter culture preparations are free of bacteriophage and microorganisms that may inhibit or reduce starter culture activity. 

Starter cultures of L. oenos ML 34 for inoculating wine are obtained by inoculating the grape juice medium with 1 vol % of a subculture (or another starter culture). The subculture is prepared by inoculating 5 ml of the grape juice medium from a stab culture. The cultures are incubated at room temperature until turbidity is seen,... 

Park and Marth (1972B) prepared a series of cultured milks which contained Salmonella typhimurium. Survival of salmonellae in the products stored at 11 °C ranged from less than three days to more than nine days, depending on species of starter culture, strain of a given species, level of inoculum used to prepare the cultured product, temperature at which the product was cultured, and amount and speed of acid production. In other studies, Park et al (1970) noted that S. typhimurium survived for up to seven to ten months in Cheddar cheese made with a slow acid-producing starter culture and stored at 13° or 7°C, respectively. In contrast, Goepfert et al. (1968) and Hargrove et al. (1969) found that S. typhimurium survived for three to seven months... 

Preparation of starter culture of E. coli 71/18/JM101/JM103... 

These approaches have been reviewed extensively (Jameson, 1990 Ardo, 1997 Fenelon and Guinee, 1997 Fenelon, 2000). Various recommendations for the manufacture of reduced-fat cheeses with improved sensory and textural properties (Mistry et al., 1996 Johnson et al., 1998), (e.g., half-fat Cheddar prepared by homogenization of cream used to standardize the cheese milk) (Nair et al., 2000) the combined effects of increases in milk pasteurization temperature and pH at curd milling, and the use of selected starters and starter culture adjuncts (Guinee et al., 1999 Fenelon et al., 2002) ... 

Both proteolysis and lipolysis are involved in the cheese ripening process. The rate and extent of their interactions are influenced by the rennet preparation used, characteristics of the starter culture, pH, moisture range, salting practices, temperature, and the activity of adventitious microorganisms present in or on the cheese. 

The effect of complex commercial fermentation activators on biogenic amine production was tested by Marques et al. (2008). On the whole, it does not seem that complex nutrient preparations produced for the use with fermentation starter cultures pose a serious threat to biogenic amine production in wines. 

Starter cultures are mainly applied as liquid cultures with about 10 to 10 ° microorganisms per ml. They are also available in a freeze-dried or deep frozen preparation with the advantage of very simple application. 

The primary glycolytic event, the conversion of lactose to lactate, is normally mediated by the starter culture during curd preparation or the early stages of cheese ripening. In cases where glycolysis has not been completed by the starter, nonstarter lactic bacteria may contribute. The metabolism of lactose was discussed in Section IIIA5. 

The steps for boza production can be summarised as (1) preparation of the raw materials, (2) boiling, (3) cooling and straining, (4) addition of sugar and (5) fermentation. Boza (2-3%) from a previous batch is usually used as a starter culture. The mixture is left to ferment in wooden barrels. The ratio of the starter culture depends on the season and temperature at which it is produced. The inoculated... 

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