Enzyme immunoassay for the quantification of mithramycin using beta-D-galactosidase as a label.

A sensitive enzyme immunoassay for mithramycin (MTM) has been developed by using antibody induced in rabbits, beta-D-galactosidase-labeled MTM, and a double-antibody separation technique, which allowed us to measure accurately as little as 100 pg of MTM per assay tube. MTM-antibody was produced against MTM-bovine serum albumin conjugate prepared by the use of diazotized p-aminobenzoic acid as a cross-linker. The beta-D-galactosidase-labeled MTM conjugate was similarly prepared by a geometric m-isomer of diazotized aminobenzoic acid. This enzyme immunoassay was specific to MTM and showed a very slight cross-reactivity with MTM analogues, chromomycin A3 (5.6%) and olivomycin (2.4%), but no cross-reactivity with drugs commonly used with MTM in combination chemotherapy for cancer treatment. The values of MTM concentrations detected by this assay were comparable to those detected by the high-pressure liquid chromatography method. However, the enzyme immunoassay method was 100 times more sensitive in detecting MTM in lower concentrations. Using this assay, drug levels were easily determined in the blood and urine of rats during 6 h after i.v. administration of MTM in a single dose of 2.0 mg/kg. Since MTM has long been used against a variety of human cancers, the enzyme immunoassay of the drug will be a valuable new tool in clinical pharmacological studies.

Binding of selected drugs to a "treated" inline filter.

The binding of several drugs to an inline i.v. filter that had been treated to inhibit drug-binding was studied. Solutions of mithramycin, vincristine sulfate, digitoxin, insulin, dactinomycin, and nitroglycerin in both 5% dextrose injection and 0.9% sodium chloride injection were allowed to flow through an i.v. administration set containing a 0.22-micron cellulose ester filter that had been treated with a proprietary agent. Actual administration conditions were simulated by using drug concentrations and flow rates commonly employed in clinical practice. The amount of each drug retained by the filter was determined by assaying aliquots of the solutions sampled before and after the solution passed through the filter membrane. In a second experiment, drug binding to the filter membrane was measured by incubating small pieces of the treated membrane in drug solutions and determining concentrations periodically until equilibrium was reached. Untreated filter membrane pieces were used as a control. In the experiment simulating actual i.v. administration, cumulative binding of mithramycin, vincristine sulfate, dactinomycin, and nitroglycerin to the treated filter was less than 6% of the initial dose infused; approximately 8-12% of the initial amounts of digitoxin and insulin were bound to the filter. In the equilibrium binding studies, the untreated filters bound twice as much digitoxin, 5-7 times as much mithramycin, vincristine sulfate, and dactinomycin, and 20 times as much insulin as the treated filters. The amount of nitroglycerin bound to the treated and untreated filters was not substantially different. Insulin had a greater binding tendency in 5% dextrose injection than in 0.9% sodium chloride injection in both experiments regardless of the filter treatment. Treatment of a cellulose ester i.v. filter with the proprietary agent used in this study facilitates drug delivery through this filter.

[Isolation of the new antibiotic variamycin B from the biosynthetic products of a Streptomyces olivovariabilis sp. nov. culture]. / Vydelenie novogo antibiotika variamitsina B iz produktov biosinteza kul'tury Streptomyces olivovariabilis sp. nov.

Variamycin B, a new antitumor antibiotic was isolated from metabolic products of Streptomyces olivovariabilis sp. nov. The empirical formula of variamycin B is C52H76O24, the melting point is 163-165 degrees C (dec.), (alpha)20D--30 +/- 2 degrees (C 0.5, alcohol), the UV-spectrum: lambda max 230, 279, 317, 330, 415 (lg E 4.27, 4.6, 3.83, 3.75, 3.95), IR-spectrum: 1080, 1170, 1380, 1570, 1640, 1720, 3400 cm-1. Variamycin B is classified as belonging to the group of antibiotic analogs of aureolic acid. Chromomycinone, tetrazide and residues of two 2,6-didesoxysugars, i.e. olivose and variose were obtained as a result of complete and partial acid degradation of variamycin B. It was shown with the data of quantitative analysis of the hydrolysates obtained on complete hydrolysis of variamycin B that the ratio between the residues of variose and olivose in the antibiotic molecule was 1 : 4. Oxidation of variamycin B with Fremi salt resulted in formation of chinone having chromomycinone-chinone, 2 residues of olivose and 1 residue of variose in its composition. Investigation of the products of partial hydrolysis provided isolation of a substance having aglicone-chromomycenone and 4 residues of olivose in its composition. A partial structure of variamycin B is proposed.

[C14-variamycin content in normal and tumorous brain tissues of white rats]. / Soderzhanie C14-variamitsina v normal'noi i opukholevoi tkaniakh golovnogo mozga belykh krys

Penetration of variamycin, a new antitumor antibiotic into the normal and tumor tissues of the brain of rats with multiform glioblastoma was investigated. The content of the C14-labeled antibiotic was determined radiometrically. The radioactive label penetrated into the normal and tumor tissues of the brain during the first hours after the drug administration. The level of the radioactivity in the tumor tissue was higher than that in the normal brain tissue during the whole period of the study. The greatest deviation in the contents of the radioactive label in the tumor and normal tissues was observed 2 and 3 hours after administration of the labeled antibiotic, i. e. 4.3 and 3.6 times respectively.

[Quantitative method for determining variamycin]. / Kolichestvennyi metod opredeleniia variamitsina

A quantitative spectrophotmetric method for the assay of variamycin in commercial samples was developed. It was based on the measurement of the optical density of variamycin solutions in 0.01 N hydrochloric acid at a UV spectrum wave length of 412 nm. A method of thin-layer chromatography for a semi-quantitative estimation of tetraside, the main admixture of variamycin was devised. On storage for 18 months the content of variamycin in the preparations did not change as compared to that at the moment of the drug manufacturing and the amount of tetraside in 18 months did not exceed 2 per cent.