The kinetics of cholesterol oxidase synthesis by Nocardia rhodocrous

Abstract

The production of cholesterol oxidase by 3 liter batch cultures of Nocardia rhodocrous growing on a glycerol/yeast extract medium was investigated. Cholesterol was shown to be a good inducer of the enzyme. The optimum time for cholesterol addition and the quantity to be added were determined, resulting in a 15-fold yield increase. Cholesterol oxidase synthesis was influenced by the dissolved oxygen tension. Maximum cholesterol oxidase production was obtained at 30-40% air saturation. The effect of growth conditions on the extraction of cholesterol oxidase by Triton X-100 was investigated. The scale-up of the fermentation to 800 liters in a pilot-plant fermenter is described.

Rhodococcus erythropolis ATCC 25544 as a suitable source of cholesterol oxidase: cell-linked and extracellular enzyme synthesis, purification and concentration

“…Under this conditions the cell yield was doubled (9.5 mg/ml vs. 4.8 mg/ml dry cell weight) and the cultivation time was reduced to one third (60 vs. 180 hours) as compared with shaken flasks. These results are in good agreement with the literature [12]. We found that addition of 2 g/l cholesterol to the culture broth [12], prepared as an aqueous emulsion with the aid of Tween 80 at a weight ratio 2:1 results in a high yield of COX production [9], but the preparation procedure of that emulsion had a marked influence in the final enzyme yield, although not on the cell weight, as seen in Table 1.…”

“…The kinetics of enzyme synthesis at both bench and large scale by Nocardia rhodochrous (renamed as Rhodococcus rhodochrous ), a strain that produces only a cell-bound COX, has been studied and the growing conditions for bacterial enzyme synthesis in fermentor were defined [12]. …”

The influence of dissolved oxygen tension on the synthesis of the antibiotic difficidin by bacillus subtilis

“…Critical DOT values below which synthesis is reduced have been reported for other antibiotics, such as cephalosporins,6 cephamycin C,'5,2' and candicidin. l 2 In large-scale fermentations, due to inefficient bulk mixing and changes in hydrostatic pressure, microorganisms are exposed to a continually changing environment as they pass through different regions in a fermenter. Dissolved oxygen concentration gradients have been observed for an unspecified 19 m3 fermentati~n'~ and for chlorotetracycline production by strains of Streptomyces aureofaciens in a 1 12-m3 fermenter."…”

Some enzymatic properties of cholesterol oxidase produced by Brevibacterium sp

“…Cholesterol may be completely oxidized by microbial cells to carbon dioxide and water by the action of a complex enzyme system in which cholesterol oxidase is the first enzyme involved. Cell-free enzymes and microbial cells have been investigated for reduction of cholesterol level in foods (2,3,14,10,26,28), for precursors production in manufacturing pharmaceutical steroids from cheap sterols (15) and for clinical assay of serum cholesterol (7,25). Microbial cholesterol oxidases have received much attention in recent years, mainly due to its large use in medical practice for determination of free and bound cholesterol.…”

“…Cholesterol oxidases can be intrinsic membranebound enzymes located on the outside of the cell, as produced by Nocardia rhodochrous (8,7,27), Nocardia erythropolis (19), and Mycobacterium species (22), or can be isolated from broth filtrate as in cultivation of Streptomyces violascens (23), Brevibacterium sterolicum (24), Streptoverticilium cholesterolicum (15), Rhodococcus equi n o 23 (26), Mycobacterium ATCC 19652 (9) and R. erythropolis (22).…”