SU892278A1 - Method of determination of aminoglycoside antibiotics in biological liquids - Google Patents

Description

(5) METHOD FOR DETERMINING AMINO-GLYCOSIDE ANTIBIOTICS IN BIOLOGICAL The invention relates to analytical chemistry and can find practical application, for example, in clinical analysis in determining the level of amino-glycoside antibiotics in various biological fluids of humans and animals. A known method for the determination of new-glycosidic antibiotics in biological fluids, including the treatment of the prbba with a reagent followed by colorimetration, with the thiobarbituric acid periodate used as the reagent. However, the known method is not accurate enough since its sensitivity is 2.5 µg / ml. The aim of the invention is to increase the sensitivity and accuracy of detection. This goal is achieved by using a mixture of aqueous solutions of the salt of praseodymium and phthalexone-S at their 2: 8 ratio as a reagent, and the treatment is carried out in a water-ethanol medium at pH 5.3-5.8. As a result of the reaction, a compound colored blue (the absorption band maximum at 619 nm) with a high molar absorption coefficient is formed. For example, for the compound monomycin with praseodymium and phthalexone-S, this value is equal to 1600000 for gentamicin, 155000 for zygomycin, when carrying out the reaction in aqueous ethanol (1: 1), the interacting components enter into an antibiotic: praseodymium: phthallexone 1: 2: 8. Sulfates, nitrates, chlorides, acetates of alkali and alkaline earth ions, most of the metal ions at PH 5, carbohydrates, and amino acids like glycine, analy, tyrosine tryptophan, novocaine, benzylpenicillin, tetracycline antibiotics (if their content in the analyzed sample not more than 1000 times the amount of amino-glycosidic antibiotics). The interfering effect of iron (lit), aluminum, copper, calcium, zinc is eliminated by introducing into the test solution up to 20 kg of quantities with respect to the praseodymium of sodium fluoride. Example 1. Determination of amino-glycosidic antibiotics in the blood. To 0.2 ml of blood is poured 2 ml of water; 2 ml of 30% aqueous solution of trichloroacetic acid, stirred and centrifuged at 5000 rpm for 10 minutes. The centrifugate is transferred to a 20 ml flask, 0.2 ml of O, an aqueous solution of ascorbic acid is added; 0.1 ml of 0.01 M sodium fluoride; 0.2 ml of 10% aqueous ammonia; 5 ml of acetate buffer with pH 5i5; 1 ml of 0.002 M phthalicone Sona-S; 0.5 ml 0.001 M of the right-, zeodim chloride and 96% ethanol to 20 ml. After 2 min, the optical density of the solution in a 2 cm cell at 619 nm relative to water is measured. The content of amino-glycoside antibiotics is found on a calibration chart constructed by processing blood samples, to which different amounts of one or another antibiotic are added. . Example 2. Determination of amine glycosidic antibiotics in urine. To 0.5 ml of urine pour 0.5 ml of water; 0.2 ml of 0, aqueous solution of ascorbic acid; 0.1 ml OjOl M sodium fluoride; 10 ml of acetate buffer solution with a pH of 1 ml of 0.002 M aqueous solution of phthalexone-S; 0.5 ml of 0.001 M praseodymium chloride is mixed and 96% ethanol is added to 20 ml. After 2 m, the optical density of the 84 ra in a two-meter cuvette at b19 nm relative to water is measured. The content of amino-glycosidic antibiotics is found according to a calibration graph constructed by processing urine samples, in which increasing amounts of one or another amino-glycoside antibiotics are added. Example 3. Determination of amino-glycosidic antibiotics in milk. To 0.2 ml of milk pour 2 ml of water; 0.2 ml of trichloroacetic acid solution, stirred and centrifuged at 5000 rpm for 10 minutes, the centrifugate was filtered into a 20 ml flask, and the filter cake was washed with 5 ml of acetate buffer with a pH in the flask was added 0.2 ml 0.01 % aqueous solution of ascorbic acid; 0.1 ml of 0.01 M sodium fluoride; 0.3 ml of aqueous ammonia, mix the contents of the flask, pour 1 ml of 0.002 M aqueous solution of phthalexone-S; 0.5 ml of 0.001 M praseodymium chloride and ethanol to 20 ml. After 2 min, the optical density of the solution in a 2 cm dish at 61 9 nm relative to water was measured. The content of antibiotics is found according to the calibration schedule, building by processing samples of milk in which increasing quantities of one or another antibiotic are added. The results of the determination of amino-glycosidic antibiotics in various materials are presented in the table. The metrological characteristics of the method for the determination of amino-glycoside antibiotics in biological materials are as follows: the number of experiments, the student's criterion, with a degree of reliability, 95. X is the arithmetic mean of the n definitions, S is the quadratic error. 0.1 μg of antibiotic was administered in 1 ml of the test material.

Monomitsin 0.09810.002 0.097 ± 0.003 Kanamycin 0.098 ± 0.002 0.09710.003

0, 002 0.099 ± 0.002

Zygomycin 0.098 + 0.002 0, 003 Gentamicin 0.099tO, 002 0,, 003

The invention makes it possible to determine 0.01 to 50 µg of monomycin, kinacin, neomycin, zygomycin, naromycin, gentamicin, streptomycin and hydroximicin in blood, milk and urine of humans and animals. The results are identical, regardless of which object (human, dog, rabbit, or mouse) the material was taken for the study.

The sensitivity of the method is 0.01 μg / ml.

Claims (1)

1. Fukumoto Hisako J.Hyg.Chem., 1976, Po22, 380-38 o