The polyene antibiotic candicidin produces a rapid efflux of K + ions from a suspension of Candida albicans . Onset of K + leakage depends on the culture age, stationary-phase yeasts leaking K + more slowly than exponential-phase yeasts. The time taken for potassium leakage to begin represents the time taken by the antibiotic to cross the cell wall and produce membrane damage. It was shown that there were factors in the cell wall of C. albicans that increased their total binding capacity and their affinity for candicidin during growth. An attempt was made to relate changes in the lipid content of the yeast cell with the increased time taken to produce membrane damage.

S U M M A R YWith a liquid membrane potassium-sensitive electrode it was shown that lethal concentrations of the polyene antibiotics amphotericin B, candicidin and nystatin caused leakage of 90% of non-bound potassium ions from Candida albicans. Accurate and rapid determinations of rates of potassium leakage were made directly in suspensions of Candida. Leakage patterns for the three polyenes were very similar, suggesting a common site of action. Differences were noted in the rates of potassium efflux with organisms from cultures of differing ages. I N T R O D U C T I O NAmphotericin B, candicidin and nystatin are polyene antibiotics important in the treatment of human mycotic infections, particularly vaginitis caused by Candida albicans. The binding of polyenes by membrane sterols has been known for some time and is well documented (Lampen, I 966).This paper considers the effect of the binding of polyene antibiotics to the membrane on membrane permeability and specifically the selective permeability towards potassium. M E T H O D SOrganism and method of culture. Candida albicans N C T C~I~ was grown on Sabouraud dextrose agar slopes at 37 "C for 24 h and maintained at 4 "C. From these slopes approximately 106 organisms were inoculated into 10 ml Sabouraud dextrose broth (S.D.B.) and incubated with shaking for 18 h at 37 "C. After centrifuging and washing three times in double distilled water standardized suspensions were made (using a Cecil 303 spectrophotometer Eig;: = 1.0, equivalent to 23-7 x I O~ organisms/ml or 0.92 mg dry wtlml).Polyene antibiotics. Candicidin, mol. wt I173 (No. 681 NOF2) of potency 218 % (manufacturer's bioassay, where original candicidin made by S. B. Penick Ltd was given an activity of IOO %) was supplied by Pharmax Ltd, Bexley, Kent. Amphotericin B,'mol. wt 959 (No. 2B 903) and nystatin, mol. wt 927 (4740 unitslmg) were supplied by E. R. Squibb Ltd, Twickenham, Surrey. They were dissolved in dimethyl sulphoxide (DMSO) and then diluted with water or buffer to give a final DMSO concentration of I %. Fresh solutions were prepared daily. The potency of these antibiotics as measured by microbiological assay (Simpson, 1963) did not change during the experimental period. Aqueous solutions of the antibiotics were protected from light, to which they are sensitive, wherever practicable.Detection of potassium loss. Leakage of potassium ions was detected with a Phillips 560K

The polyene antibiotic candicidin is a potent membrane active agent, the action of which can be inhibited by the presence of certain ions. The destruction of the selective permeability of yeast membranes by candicidin allows small molecules to leak into the environment. Loss of intracellular potassium ions inhibits yeast glycolysis. This inhibition may be reversed by extracellular concentrations of potassium or ammonium ions. Monovalent ions did not prevent antibiotic absorption or protect yeast growth from the action of the antibiotic. Divalent ions did not protect yeast glycolysis from the action of candicidin, but were able to reduce antibiotic‐induced membrane damage and allowed yeast growth in the presence of antibiotic. It is suggested that divalent ions may interact with membrane sterols creating steric hindrance to subsequent candicidin absorption.

SUMMARYThe polyene antibiotic candicidin is a potent antifungal agent acting upon the cell membrane of Candida albicans. Destruction of selective permeability by polyenes allows cations to leak into the environment and permits the entry of protons to neutralize the charge so created, causing internal acidification. Quantitive studies on proton entry reveal that K+ leakage alone is not sufficient to account for the degree of acidification recorded. Extracts of untreated C. albicans, when acidified to the same extent, precipitated. Electron micrographs of candicidin treated C. albicans cells confirm this precipitation.

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