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SUMMARY: Saccharomyces rouxii cells were maintained on yeast extract/neopeptone/glucose medium (YNG). Experimental medium was depleted of endogenous metal ions by repeated passage of YNG over a chelating ion-exchange resin. This treatment did not compromise the glucose or vitamin content but provided a basal medium to which essential salts could be added in defined concentrations. Accordingly it was possible to demonstrate growth requirements by this yeast species for K+, Mg2+, Zn2+ and Fe3+. Na+ would not replace K+. Additions of 1.0 μ;M-Ca2+, NH4+ and Na+, or 0.1 μ;M-Ni2+, Sn2+, Al3+, Co2+ or Cr3+ to a minimal, reconstituted medium were without effect. Cu2+ and Mn2+ additions were not required for growth but, like Mg2+, Zn2+ and Fe3+, they positively affected the biosynthesis of periplasmic acid phosphatase. Cells from iron-depleted medium had only 1 unit (g dry wt)-1 of native acid phosphatase activity although this could be stimulated 10-fold by addition of FeCl3 to washed cell suspensions before enzyme assay. Additions of 10-60 μ;M-FeCl3 to untreated YNG medium progressively increased the concentration of acid phosphatase after 3 d of culture to an upper value of 75 units (g dry wt)-1, and also increased the amount secreted into the growth medium. While unfortified YNG supported cells with only 28% of their acid phosphatase expressed (the bulk being subject to activation by Fe3+ in the assay medium) the cells from YNG fortified with 60 μ;M-FeCl3 displayed fully activated acid phosphatase. Cells from Mg2+-depleted medium exhibited bizarre, elongated cell shapes and those from Zn2+-deficient medium appeared somewhat angular by light microscopy. Fe3+-deficient cells had apparently normal morphology.