Biomedical subjects
Publications and source records attributed to E D Etingov.
Solubility of sodium novobiocin in binary mixtures of ethyl alcohol with ehtyl acetate, methylene chloride, diethyl ether, carbon tetrachloride or n-heptane was studied. Deviation in the sodium novobiocin solubility from the additivity was observed depending on the nature of the second component of the solvent mixture.
An actinomyceteous strain LIA-0185 producing a heptaenic non-aromatic antibiotic of the candidin type was isolated from a soil sample taken in the Georgian SSR under the programme of screening antifungal antibiotics. The taxonomic study of the strain showed that it belonged to the series of viridoflavum and had the following main taxonomic features: the sporophores in the whorls, straight, remote: the aerial mycelium from yellow to dark-olive-grey; the substrate mycelium olive; the soluble pigment absent; the melanine pigment was produced on the peptone medium; the culture formed H2S; assimilated glucose, mannose, inozide and to a lesser extent fructose; did not assimilate arabinose, xylose, sucrose, lactose, ramnose and raffinose. The strain inhibited the growth of yeast and fungi, grampositive bacteria and actinomycetes and produced a complex of non-aromatic heptaenic antibiotics. The actinomycete differed from the other whorl cultures. It was classified as a new species Sv. griseoviridum sp. nov. The antibiotic complex was a mixture of 2 components, i. e. I and II present approximately in equal amounts. Component II was analogous to candidin. Component I was a new original substance.
A marked decrease in the coefficient of oleandomycin distribution in the system of the fermentation broth filtrate-butyl acetate was observed during the transfer from the 1st to the following extraction stages. It was supposed that the oleandomycin fermentation broth filtrate contained admixtures affecting significantly the antibiotic distribution between the aqueous phase and organic solvent.
A thin-layer chromatographic method for separation of novobiocin, isonovobiocin and descarbamylnovobiocin using Silufol plates was developed. The method is simple and rapid. It provided clear separation of the components and using of diethyl ether, a simple individual separating solvent. The method allowed determination of novobiocin and the products of its isomerization in the fermentation broth, extracts and dry substances, as well as performance of operative control and regulation of the technological processes of fermentation and chemical purification of novobiocin.
A possibility of increasing the distribution coefficient of oleandomycin on its extraction from the aqueous solution into the organic phase containing an electron carrier-acceptor was shown. It was found that the level of the oleandomycin equilibrium shift into the organic phase depended on the carrier nature, i. e. the donor-acceptor interaction of the carrier and the antibiotic. It was shown that the oleandomycin fermentation broth contained non-identified admixtures, the interaction of which with the antibiotic was higher than its interaction with the acceptor-carriers tested.
It was shown on model experiments that the microbiological method was not applicable for determination of levorin content in industrial intermediate products containing in addition levoristatin, since the presence of the latter made higher the results of the microbiological assay. Because of this till to the present date the quantitative content of levorin in the industrial intermediate products was determined photometrically using alcohol (pure solvent) as the reference solution. Still, this method also made higher the results of the assay, especially when the content of levorin was determined in the fermentation broth. In the solid phase levorin is contained in the mycelium which occupies only 1 to 2 per cent of the fermentation broth, while the liquid phase or the fermentation broth filtrate occupies 98 to 99 per cent. It was found that the fermentation broth filtrate contained protein admixtures which coagulated on addition of alcohol to the fermentation broth and formed fine colloid solutions. As a result the absorption values became higher. In the present study not the pure solvent but an extract of the fermentation broth filtrate containing neither levorin, nor levoristatin was used as the reference solution. Such a differential method provided elimination of all errors due to the presence of various metabolites in the fermentation broth filtrate which varied both qualitatively and quantitatively.
Dependence of the oleandomycin distribution coefficient on pH of the acqueous phase and temperature in the system of butylacetate extract-water acidified with orthophosphoric acid was studied. With a purpose of intensification of the process of oleandomycin reextraction, decreasing the antibiotic inactivation and evaporation of the organic solvent it was proposed to perfom oleandomycin extraction at pH 4.0--5.0 accompanied by simultaneous decreasing of the temperature.
Dependence of the oleandomycin distribution coefficient on the antibiotic concentration, duration of the filtrate storage, extraction temperature and salting-out agents was studied. The distribution coefficient of oleandomycin in the system of fermentation broth filtrate--butylacetate was much lower (about 3 times) than that in the system of oleandomycin phosphate aqueous solution--butylacetate. Addition of sodium sulfate to the aqueous phase provided a significant increase in the oleandomycin distribution coefficient.
It was shown that "nepheline coagulant"--Al2 (SO4)3-18H2O was effective in coagulating the admixtures in oleandomycin fermentation broth. The use of the "nepheline coagulant" in an amount of 1 to 3 per cent (by the weight) of the volume of the oleandomycin fermentation broth provided butyl acetate extraction of the antibiotic without formation of stable emulsion. Addition of the "nepheline coagulant" in an amount of 1 to 2 per cent (by the weight) to the fermentation broth markedly increased the rate of the fermentation broth filtration.
Complexes of polyenic antibiotics with polyvinylpyrrolidone (PVP) can be used for preparing effective pharmaceutical forms soluble in water and consisting of fine dispersions. Studies were carried out; they are of great importance for revealing the mechanism of polyen interaction with neutral polymers, as well as for development of the technological processes for production of the pharmaceutical forms. The sorption isoterms of PVP with the molecular weight of 10 000 and 35 000 on nystatin and amphotericin B were obtained in the process of precipitation in the system of dimethylformamide-ethylacetate. The constants of the strength of the antibiotic binding with the polymer in a complex were calculated. It was shown that the complex strength increased with a rise in the relative amount of the precipitant in the system. The temperature dependence of the binding strength constant was studied. The process of the complex forming was shown to be exothermic, the activation energy of the complex being 26-30 kcal per a mole of the antibiotic. No significant differences in the binding strength of nystatin and amphotericin B were observed. On the basis of the experimental data, a scheme of the complex structure explaining the binding process by formation of a number of hydrogen bonds between the antibiotic hydroxyl groups and the PVP tertiaryamide groups is proposed.
Deep effect of gamma-rays on polyenic antibiotics was studied. It was shown that gamma-radiation induced radiation-chemical oxidation of the substances. The chromatographic analysis showed that the levorin degradation products were identical to the polyenic products of the antibiotic oxidative destruction. As for mycoheptin and amphotericin B, destruction of their molecules to non-polyenic products was observed. It was found that toxicity of the levorin aromatic heptaen did not practically change after gamma-irradiation in high doses. The toxicity of mycoheptin and amphotericin B, non-aromatic heptaens increased after exposure to high doses of gamma-rays.
Experimental differences in the curves of the optic rotation dispersion (ORD) of cystrans-heptaenic antibiotics were found. The ORD curves of amphotericin B, mycoheptin, levorin components and isolevorin A2, components of criptomycin and candidin were registered. The curves of the ORD which were smooth had been prepared in dimethylsulphoxide in the spectral range at 450 to 600 nm. In the spectral range at 300 to 420 nm the ORD curves appeared to be anomal with a complex Kotton effect, they were prepared in methyl alcohol. The Kotton effect was probably due to asymmetry of the electron membranes of polyenic chromophore induced by the other part of the polyen molecule. This was evident from the fact that the curve of the Kotton effect was situated in the same spectral range as the absorption bond of the polyenic chromophore. The oscillating structure in the absorption spectrum and the curve of the complex Kotton effect were analogous.
Acid-base properties of amphotericin B, polyenic antibiotic in aqueous solutions was studied. A special procedure provided the use of potentiometric titration for investigation of ionization of the groups of the water-insoluble substance. The ionization constants of the carboxylic and amine groups of the antibiotic were determined at several temperatures. It was found that ionization of the acid group did not depend on the temperature. At the same time the heat effect of the amine group ionization was significant and amounted to about 10 kcal/mole. Thermodynamic analysis of the ionization process of amphotericin B in aqueous solutions was performed. Integral components defining the process energetics were calculated.
An actinomycetous culture designated as LIA-0721 was isolated from a soil sample. It was close to Act. fulvoviolaceus by its morphologo-cultural features and differed from it in production of melanoid pigments and the spectrum of carbohydrate assimilation. This justified its classification as Act. fulvoviolaceus var. achromogenes var. nov. The culture produced new aromatic heptaens, i. e. fulvomycins A, B and C. Their physico- chemical and biological characteristics are presented.
Association of nystatin and amphotericin B in non-aqueous systems was studied with the method of equilibria dialysis. A specially treated celophane membrane arresting colloid associats and macromolecules with a molecular weight of more than 30000 in the systems of dimethylformamide-ethylacetate was used for the dialysis. Relation between the dialysis rate and the difference of the concentrations at every side of the membrane was used for estimation of the antibiotic colloid association level. It was found that nystatin formed stable associates of the colloid type in the system of dimethylformamide-ethylacetate, close by the composition to the critical one or that providing precipitation of the antibiotic. Unlike nystatin, amphotericin B formed not colloid but larger conglomerates which precipitated. Neither of the antibiotics formed colloid associates in dimethylformamide. The level of the nystatin colloid association increased with a rise in the solution concentration and reached 80%. On the basis of the results obtained the following supposition concerning the mechanism of formation of the antibiotic complexes with polyvinylpyrrolidone (PVP) in non-aqueous systems is possible: sorption of PVP on the colloids formed or larger associates of the polyenic antibiotics must take place during coprecipitation which is accompanied by formation of a precipitate of the sorption complex.
The acid-base characteristics of polyenic antibiotics, such as nystatin, mycoheptin and levorin in aqueous solutions were studied. A special procedure provided the use of potentiometric titration for investigation of ionization of the groups of vater-insoluble substances. The ionization constants of the carboxylic and amine groups of the antibiotics at several temperatures were determined. It was found that ionization of the acid group did not practically depend on the temperature. At the same time the heat effect of the amine group ionization was significant and amounted to about 10 kcal/mole. Thermodynamic analysis of the ionization process of the polyenic antibiotics in aqueous solutions was performed. Integral components defining the process energetics were calculated.