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SUMMARY: A steroid fraction obtained from lemon juice was found to be an essential metabolite for Paramcium aurelia, var. 4, stock 51.7 (s), in axenic culture. This steroid is closely related to, or is identical with, either ,&sitosterol or y-sitosterol. Paramecium aurelia has been established in pure culture in a heat-sterilized medium, composed of equal parts of a 0.5 yo yeast autolysate (Basamin-Busch) and a 24 hr. culture of Aerobacter aerogems in lettuce extract (van Wagtendonk & Hackett, 1949). Lettuce or other plant extracts were shown to be essential since no growth of P. aurelia could be obtained when these extracts were omitted. An axenic medium, consisting of a salt solution, dialysed yeast extract, a solution of water-soluble vitamins, proteose peptone, and guanylic and cytidilic acid has been developed (van Wagtendonk, Miller & Conner, 1952; van Wagtendonk, Conner, Miller & Rao, 1958). P. uurelia is unable to grow in this medium unless plant extracts are added. An essential metabolite for P. aurelia must therefore be present in plants. The isolation and identifi- cation of this metabolite is described. EXPERIMENTAL Method of assay P. uzcrelia, variety 4, stock 51.7, sensitive (Sonneborn, 1949) is not able to grow in either axenic or bacterized media in the absence of plant extracts. Rapid screening assays for activity are desirable in an extraction of biologically active material. The test in axenic medium is slow, and for this reason fractions were screened using a bacterized medium. The basal medium for the bacterial assay consisted of: NaC1, 0.8 g.; KC1, 0-05 g.; NaaPO,, 0-1 g.; N&PO,, 0.1 g.; CaC12.2H20, 0-05 g.; MgS0,.7H20, 0-01 g.; FeSO,(NH,),SO,. 24H20, 0.125 g. ; double distilled water, 1 1. ; HC1 to pH 6.5. The extracts to be tested were prepared in serial dilutions, using the salt solution as a diluent. These dilutions were autoclaved at 120° for 15 min, They were then inoculated with A. aerogmes 24 hr. before use for the assay, as in the procedure routinely used for the culture of P. aurelia (Sonneborn, 1950).

The Isolation from Lemon Juice of a Growth Factor of Steroid Nature required for the Growth of a Strain of Paramecium aurelia