Metabolism of daunomycin (NSC-82151) in vitro and the chemotherapeutic activity of isolated metabolites in vivo.

Daunomycin (D1) was quantitatively converted to a metabolite, D2 (> 98% conversion), by an aerobic, cell-free system containing nicotinamide adenine dinucleotide phosphate, reduced form (NADPH) and the 100,000 x g supernatant fluid of rat kidneys. Under lowered oxygen concentrations, cell homogenates plus NADPH converted D, quantitatively to an unknown metabolite, D(x), with D2 as a possible intermediate. The chromatographic properties of D(x) differed from those of D2 and the aglycones of D1 and D2 (D4 and D3 respectively). Sufficient quantities of D2 and D(x) were prepared using NADPH-supplemented cell-free preparations from the kidneys of rats to assay their chemotherapeutic activity in vivo. The metabolite D2 was as effective as daunomycin (D1) against P388 leukemic cells growing in mice, but only one sixth as active against L1210 leukemic cells growing in suspension culture. The metabolite D(x) was much less active than either D2 or D1 when assayed against L1210 cells growing in culture and was inactive against P388 cells growing in mice.

Effect of divalent cations on the uptake and oxidation of substrates by Pseudomonas aeruginosa.

Evidence is presented that Zn(2+) and certain other multivalent cations inhibit the uptake and oxidation of substrates by Pseudomonas aeruginosa and by other bacterial cells. Tris(hydroxymethyl)aminomethane augmented this effect. Inhibition by these multivalent cations could be prevented by incubation of cellular suspensions of P. aeruginosa in phosphate buffer. In the case of other microorganisms, incubation in the presence of ethylenediaminetetraacetate also prevented the inhibition. Inhibition by Zn(2+) of the uptake and of the oxidation of substrates could be reversed by the addition of Mg(2+). Zn(2+) was shown to inhibit noncompetitively the uptake of the three substrates studied, l-alanine, d-glucose, and citrate.

Role of Multivalent Cations in the Organization, Structure, and Assembly of the Cell Wall of Pseudomonas aeruginosa.

Asbell, Mary A. (University of Georgia, Athens), and R. G. Eagon. Role of multivalent cations in the organization, structure, and assembly of the cell wall of Pseudomonas aeruginosa. J. Bacteriol. 92:380-387. 1966. -Incubation of Pseudomonas aeruginosa with ethylenediaminetetraacetate induced the formation of osmotically fragile rods termed osmoplasts. These could be restored to osmotically stable forms by multivalent cations. Only those cells restored by divalent cations normally found in the cell wall were capable of multiplication. The respiration of restored cells, however, was unimpaired, irrespective of whether they were capable of multiplication. Moreover, the permeability characteristics of osmoplasts and restored cells were unimpaired. When multivalent cations were chelated from the cell wall and replaced by sodium, a weakened cell wall and an osmotically fragile cell resulted. This was apparently caused by the absence of cross-linkages in the cell wall via multivalent cations. Tris(hydroxymethyl)aminomethane buffer compounded the lethal effects of ethylenediaminetetraacetate. The lipopolysaccharide component was inferred to be the site of attack by ethylenediaminetetraacetate. A mechanism for the synthesis of the lipopolysaccharide sacculus was proposed whereby negatively charged subunits are "trapped" by forming ionic and coordinate bonds intermediated by multivalent cations.