To study the mechanism of the proposed assimilation of cholesterol, we cultured various strains of LactobaciUlus acidophilus and a Bifidobacterium sp. in the presence of cholesterol and oxgall. During culturing, both cholesterol and bile salts were precipitated. Because of bacterial bile salt deconjugation, no conjugated bile salts were observed in either the culture fluids or the pellets. During incubation, the cell count and optical density decreased. The degree of precipitation of bile salts and of cholesterol was dependent on the culture conditions. If L. acidophilus RP32 was cultured under acidifying conditions, the degree of precipitation of deconjugated bile salts was higher than if the pH was maintained at 6.0. Under acidifying conditions, cholesterol was coprecipitated with the bile salts, whereas in pH-controlled cultures, no coprecipitation of cholesterol was observed. From control experiments with different mixtures of bile salts, it appeared that coprecipitation of cholesterol during culturing was a result of formation of deconjugated bile salts, which have a decreased solubility at pH values lower than 6.0. It is concluded that the removal of cholesterol from the culture medium by L. acidophilus RP32 and other species is not due to bacterial uptake of cholesterol, but results from bacterial bile salt-deconjugating activity.

The determination of the best conditions of preparation of a (tentatively) probiotic starter culture that might be suitable for cheese making composed solely of Bifidobacterium lactis Bo and Lactobacillus acidophilus Ki is critical if a consistently reliable acid production is to be achieved, especially because bifidobacteria have stringent requirements for growth. Therefore, we determined whether B. lactis Bo and L. acidophilus Ki required or benefitted from the addition of milk hydrolyzates (brought about by proteinase or neutrase as the nitrogen source). The growth and acid production of B. lactis in milk were affected by the addition of proteinase-mediated hydrolyzate and, to a lesser extent, by neutrase-mediated hydrolyzate; a higher degree of hydrolysis of either hydrolyzate resulted in greater biomass increase and greater acid production. This result suggests that the poor growth of bifidobacteria in milk is due partially to the lack of small peptides and free amino acids. The rates of growth and acidification by B. lactis were enhanced when cocultured with L. acidophilus (1:1 inoculum ratio). Conversely, the growth rates and acid production of L. acidophilus were not positively affected by the addition of either milk hydrolyzate. Although L. acidophilus grew slowly, its proteolytic system was apparently able to generate its own nitrogen source. Nevertheless, coculture with B. lactis (1:1 inoculum ratio) led to enhanced rates of growth and acidification when compared with that of the single strain, suggesting some degree of symbiosis between the strains.

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