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This work describes the configuration and operation of a bioreactor system especially designed for Tetrahymena cultivation and its use for milk improvement, particularly cholesterol elimination by the action of this cell. An advantage of the proposed method is the re-use of the growth medium; thus, the medium is used twice to provide two batches of Tetrahymena biomass without the need of further inoculation. This makes the procedure of producing the cell biomass faster and more economical. Cells are concentrated in the culture vessels by sedimentation at room temperature and then transferred to milk suspensions, where they can further grow for at least one generation with the benefit of reducing steeply cholesterol level. Milk treated according to this process is separated from the biomass by centrifugation. Under these conditions, less than 5% of the cells remain in the milk, and cholesterol elimination amounts to 75 +/- 10% of that initially present. No changes in sensorial properties of the milk, such as clotting or butyric odor, were observed as a result of this treatment. In addition, the bioreactor allows the aseptic recovery of the spent growth medium, which contains diverse enzymes of interest, and the cell pellets, to exploit particular lipids like phosphonolipids, abundant poly-unsaturated fatty acids and co-enzyme Q(8).

The nonpathogenic ciliate Tetrahymena thermophila converts cholesterol from foodstuffs into provitamin D compounds in high yields. However, prolonged incubation with wild-type strain CU-399 at high densities results in a final deterioration of milk properties, possibly as a result of secreted hydrolases. Here we attempted to solve this problem using MS-1 Tetrahymena strain, a stable mutant with a low rate of hydrolase secretion. Densities of to 2 x 10( 6 ) cells/ml can be incubated for up to 5 h in milk, without any clotting or change in appearance. Moreover, centrifugation of this suspension eliminates most of the cells, and results in an about 75% +/- 10 (n = 10) decrease of the initial cholesterol. Sterols are recovered in the cell pellets, which show that Tetrahymena is able to avidly capture them from the medium. Therefore, this mutant strain is optimal for milk cholesterol depletion, avoiding unfavorable sensory alterations.

The exceptional nutritional values of milk have long been recognized. However, important drawbacks of milk and dairy products are their relatively high cholesterol contents and their low proportion of polyunsaturated fatty acids. Hypersensitivity to milk proteins can also draw the public away from fluid milk consumption. The nonpathogenic ciliate Tetrahymena thermophila has a unique set of enzymes that converts cholesterol into provitamin D3 and related compounds. It also contains delta 12 fatty acid desaturases and releases various hydrolases, including proteases, lipases, and phospholipases. We investigated the potential use of these peculiar traits to improve milk by short-term incubation with live ciliate suspensions. We found that cholesterol can be reduced to 50% of its initial content in whole milk by a 12-h period of incubation with T. thermophila cells, mostly by conversion to delta 7-desaturated cholesterol derivatives, including provitamin D3. In addition, a considerable increase in gamma-linolenate (18:3, n-6) takes place. The protein profile is markedly changed by proteolysis, with a striking reduction of casein and enhancement in total amino acid content. Despite these changes, no marked alterations in flavor or odor were observed at the end of 12 h of incubation. Thus, Tetrahymena appeared ideally suited to correct for milk draw-backs, making this important source of nutrients more acceptable to the health-conscious public.