TH438A - Antibiotic preparation process A / 16686 Factors A2 and antibiotic A / 16686 co-occurring factors A 1 and A3. - Google Patents

Abstract

The invention has to do with antibiotics that contain chlorine. It was named antibiotic A / 16686 factor A2 in its prime pure form. The science also deals with a process for the production of antibiotic A / 16686 factor A2 by cultivating actinoflens and producing antibiotics A / 16686 factor A1 and A. 3 also coalesce with antibiotics A / 16686 factor A 1, A2 and A3 as well as their corresponding salts. Formed by combining with acids, which are physiologically acceptable, are non-toxic, are anti-microbial, especially anti-gram-positipbacteria, patent drug

Claims (6)

1.Process for the production of purified antibiotic A / 16686 factor A2 and salts formed by combining with acetic acid. Physiologically non-toxic based on the main criteria for separation: a) is a white amorphous powder, molten with decomposition approx. 210-220 ํ B), highly soluble in water, dimethyl. Formamead, aqueous methanol, and 0.1 N HCl; Sparingly soluble in absolute ethanol and normal-butanol, and precipitated from aqueous solution by adding NaHCO3 saturation solution; C) analysis for The elements show approximate values. They contain 54.57% carbon, 6.19% hydrogen, 10.88% nitrogen and 1.37% chlorine. Good), infrared absolution spectrums in Nujol. The highest abstraction displays are 3290, 2930 and 2860 (Nujol), 1765, 1635, 1510,1455 and 1375 (Nujol), 1240, 1175, 1130, 1060, 1015, 975, 840 and 815 cm. Exponentiation-1E), Altraviolet Absorption Sunspectrum shows the highest absolution (1) in neutral 234 nm methanol (## Chemical formula) 268 nano. M (## chemical formula) (2) in baked methan containing 0.1 N HCl 233 nm (## chemical formula) 271 nm (## chemical formula) (3) in methanol containing 0.1 N NaOH 251 nm (## Chemical formula) 300 nm (bend to the shoulder) f) specification Rotation (## Chemical formula G) Here is the Rf value made with perchromaticygraph on the watchman paper No.1, using B. Subtilis ATC 6633. These organisms are used to determine the elution system (vol / volume / volume), Rf 1), saturated norm-butanol. Sorane Chain Buffer, pH 6.0 0.00 2) Normal-Butanol It is saturated with water containing 2% paratluene sulfonic acid 0.00 3) norm-butanol. Saturated with water containing 2% ammonium hydroxide 0.00 4) Shorenchain buffer, pH 6.0, saturated with normal-butanol 0.005). Normal-Butanol: Methanol: Water 4: 1: 2 0.48 6) Ethyl acetate, saturated with water 0.00 7) Normal-Butanol: Acetic press : Water 2: 1: 1 0.44H) The following is the Rf value made in the silica gel thin-layer chromatography system. Shown below, Elution system (volume / volume / volume) Rf 1) 2.5% HCOONH4: CH3CN 65:35 in water 0.36 2) norm-butanol: acetic acid: water 4: 2. : 5 0.80 3) Normal - Butanol: Acetic Acid: Water 4: 2: 5 0.52 4) Normal - Propanol: Normal - Butanol: IN NH4 OH2: 3: 4 (upper cycle) 0.12 5) norm-butanol: acetic acid: water 4: 1: 5 0.15 1,2- on the silane, silica gel 60 F 254 3, 4,5 on silica gel 60 F 254 (standard to which it is inserted: Caphene Rf 0.60, cortichon Rf 0.35, Kake Shamethachon 0.30) i). Sid after hydrolysis with 6N hydrochloric acid at 110 ํ for 6 h, shown at least the following amino acids are present: alanine, lucine, glycine. Aspartic acid, phenylalanine, ornithene, para-hydroxyphenyl glycine and hydroxyl, chloro-substrate, phenylglycine. (Scene J) carbohydrate analysis from an acid hydrolyzate obtained after 2 h in 2N H2 SO4 at 100, showing the presence of d-mannose (K.) as follows: Characteristic reactions: ninhydrin (3% ethanol solution) plus malic plus biurate plus millon minus 1% FeCl3-1. % K3Fe (CN) 6 in green water KMnO4 (acidic) plus H2SO4 concentration minus L) Retention time (tR) 124.43 cm (4 \ '99/100 ") when s Fortified by reverse phase HPLC using - Bonda Pat C18 column (3.9 mm, diameter X300 mm) and HCOONH4 0.025 M / CH3CN 60/40 (volume / volume). It is a moving cycle with a flow rate of 2 ml / min. (Standard included: anthracene tR 1067.26 cm (35 \ '18 / 100 ") alpha-nitrone naphthalene tR 335.64 cm (11 \ '14/100") Poe 245.47 cm (8 \ '64 / 100 ") and M) Luene as hydrochloride. The melting point is approximately 250 ํ (decomposition), where this production method consists of culturing actinoplanes SP. ATCC 33076 under the fermentation of this microorganism to the oxygen in the medium. There are supplements in the water that provide carbon and nitrogen sources. And absorbable inorganic salts are included. Until the meat production reaches a level that shows antibiotic activity Antibiotic A / 16686 Fatter A2 was applied in complex form with two other minor factors named Antibiotic A / 16686 Facttech. The A-1 and A-3 from the fermentation medium separated the A / 16A / 16686 fadter A2 from the complex antibiotic and separated them into each of the major pure antibiotic compounds. 2. Process as set out in claim 1, which uses Antibiotic activity was removed from the final fermentation process by extraction of the fibrous mass of fungal cells with solvent selection. Organic from small molecules alkanol and acetone 3. Process according to claim 2, in which the extraction solvent is methanol. 4. The process defined in claim 1, which excludes antibiotic A / 16686 factor A 2 by the chromatographic column, uses the "Amber Light rR XAD-2" column. And a mixture of water and acetone nitrile Or a mixture of ammonium formate in water and acetonitril As a substance for elute 5. The process set out in claim 1 includes the use of all three separate characteristics, A 1, A2 and A3, further separated by 5A). 16686 Factors A1 in free bases and salts formed by combination with acceptable acids. Physiologically non-toxic substances (A) are white amorphous powder. Melted ready to decompose at approximately 210-220 ° C b) very soluble in water, dimethyl formaldehyde, and methanol in water, dissolved in methanol and ethanol. , But insoluble in ethyl ether, petroleum ether and benzene, C). Elemental analysis shows approximate values: 50.31% carbon, 6.00% hydrogen, 9.92% nitrogen, 0.61% chlorine (total). , Chlorine ions 0.9%, Remaining 6.07% Good) Infrared Absorption Sunspectrum in Nujol shows the highest absofson: 3290,2930 and 2860 (Nujol), 1765, 1630. , 1510,1455 and 1375 (Nujol), 1260,1235,1175, 1150, 1130,1060,1015,975,840 and 810 cm-1e), altraviolet absop, sunspect. Rum expressed the highest absolution (1) in 233 nm of neutral methanol (E 1% 1 cm = 202) 266 nm (E1% 1 cm = 106) (2) in methanol contained. 0.1 N HCI 232 nm (E1% 1 cm = 219) 270 nm (E1% 1 cm = 110) (3) in methanol containing 0.1 N NaOH 251 nm (E1% 1 cm = 264) ~ 290 nm (shoulder bends) f) specific rotations (chemical formula) (C = 0.51, H2O) G). Here is the Rf of perchromatrona. Phip Nok, Paper, Waterman No. 1 uses B.Subtilis ATCC It is an elution system (volume / volume / volume) 1) norm-butanol saturated with sorenzane buffer pH 6.0 0.00 2) norm-butanol saturated with water. Containing 2% para-toluene sulfoniazide 0.00 3) norm-butanol saturated with water containing 2% ammonium hydroxide 0.00 4) sorensen , PH 6.0, saturated with Normal-butanol 0.05 5) normal-butanol; Methanol: water 4: 1: 2 0.48 6) ethyl acetate saturated with water 0.00 7) norm-butanol; Acetic acid: water 2: 1: 1 0.44H), the following is the Rf value made in the silica gel thin-layer chromatographic system, shown below the elution system (volume / Volume / volume) 1) 2.5% HCOONH4: CH3CN65: 35 in water 0.40 2) norm-butanol; Acetic acid: water 4: 2: 5 0.80 3) norm-butanol; Acetic Acid: Water 4: 2: 5 0.52 4) Normal - Propanol: Normal - Butanol 1N, NH4OH 2: 3: 4 (Upper Cycle) 0.12 5) North Al-butanol: Acetic acid: water 4: 1: 5 0.15 1,2 made on silica gel plates, silica gel 60 F 254 3,4,5 on silica plates. Gel 60 F 254 (standard filled with Caphene Rf 0.60, cortisone Rf 0.35; Texamethasone Rf 0.30) i) Amino acid analysis after hydrolysis with 6N hydrochloric acid at 110 ° C for 6 h, indicating that at least it contained amino A. The following zids are with alanine, lucine, glycine, aspartic acid, phenylalanine, orniline, para-hydroxyphenyl glycine. , And hydroxychloro-substrate (phenylglycine) J) carbohydrate analysis from Acid hydrolyzate obtained 2 hours later in 2N H2-SO4 at 100 ° C, indicating the presence of d-mannose K) retained suntime (tR) 93.57 cm (3) \ '84 "100) When analyzed by the Versphase HZ PLC method," u-Bondapac RC18 "columns (3.9mm dia.x 300mm) and HCOONH 4 0.025M / CH3CN mixtures were used. 60/40 (volume / volume) is a moving phase with a flow rate of 2 ml / min (standard included: anthracene tR 1067.26 cm (35 \ '18/100) (chemical symbol - nitronate Phthalene tR 335.64 cm (11 \ '14 \' / 100), toluene tR 245.47 cm (8 \ '64 / 100)) 5 b) Criteria for isolation of antibiotic A / 16686 factor. A3 in free base form and salt formed by combining with accepted acids It is used in physiological nontoxic substances that is A) white amorphous powder. Melted with decomposition at approx. 220 ° C b) Very soluble in water, dimethyl formate, and methanol in water, dissolved in methanol and ethanol, But insoluble in ethyl ether, petroleum ether and benzene, C). Show approximate values: Carbon 48.41%, Hydrogen 5.84%, Nitrogen 9.09%, Chlorine (Total) 1.68%, Chlorine Ions 1.16%, Remaining 9.1% Good), Infrared Absorption Spectroscopy in Nujol. Sopsun is as tall as 3290,2930 and 2860 (Nujol), 1765,1635,1510,1455 and 1375 (Nujol), 1260,1240,1175,1100,1060,1020,975,840, and 850 cm. 1 e) Altraviolet Absorption Sunspectrum shows the highest absolution (1) in neutral methanol 234 nm (E 1% 1 cm = 147) 267 nm. M (E 1% 1 cm = 74) (2) in methanol containing 0.1 N HCl 232 nm (E1% 1 cm = 132) 270 nm (E1% 1 cm = 76) (3) in. Methanol containing 0.1 N NaOH 270 nm (E1% 1 cm = 202) ~ 295 nm (curvature of the shoulder) F) specific Rotation (chemical formula) (C = 0.48, HCl N / 100) G) Here is the Rf value made with perchromatography on paper, Waterman No. 1, using B. Subtilis ATCC 6633 as a living organism. The elution system (volume / volume / volume) 1) norm-butanol saturated with sorenzane buffer pH 6.0 0.00 2) norm-butanol saturated with water. Containing 2% para-toluene sulphonic acid 0.00 3 ) Normal-butanol is saturated with water containing 2% ammonium hydroxide 0.00 4) Sorensen, pH 6.0, saturated with Normal-butanol 0.05 5) normal-butanol; Methanol: water 4: 1: 2 0.48 6) ethyl acetate saturated with water 0.00 7) norm-butanol; Acetic acid: water 2: 1: 1 0.44H), the following is the Rf value made in the silica gel thin-layer chromatographic system, shown below the elution system (volume / Volume / volume) 1) 2.5% HCOONH4: CH3CN65: 35 in water 0.40 2) norm-butanol; Acetic acid: water 4: 2: 5 0.80 3) norm-butanol; Acetic Acid: Water 4: 2: 5 0.52 4) Normal - Propanol: Normal - Butanol 1N, NH4OH 2: 3: 4 (Upper Cycle) 0.12 5) North Al-butanol: Acetic acid: water 4: 1: 5 0.15 1,2 made on silica gel plates, silica gel 60 F 254 3,4,5 on silica plates. Gel 60 F 254 (standard filled with Caphene Rf 0.60, cortisone Rf 0.35; Texamethasone Rf 0.30) i) Amino acid analysis after hydrolysis with 6N hydrochloric acid at 110 ° C for 6 h, indicating that at least it contained amino A. The following zids are with alanine, lucine, glycine, aspartic acid, phenylalanine, ornithine, parahydroxyphenyl glycine, And hydroxyl, chlorosthitide, phenylglycine J). Carbohydrate analysis from Acid hydrolyzate obtained 2 hours later in 2N H2-SO4 at 100 ° C, indicating the presence of d-mannose K) retained suntime (tR) 93.57 cm (3) \ '84 "100) When analyzed by the Versphase HZ PLC method," u-Bondapac RC18 "columns (3.9mm diameter x 300mm) and HCOONH 4 0.025M / CH3CN mixtures were used. 60/40 (volume / volume) is a moving phase with a flow rate of 2 ml / min (standard included: anthracene tR 1067.26 cm (35 \ '18/100) (chemical symbol - nitronate Phthalene tR 335.64 cm (11 \ '14 \' / 100), toluene tR 245.47 cm (8 \ '64 / 100)) 6. The process of claim 1 or 5 was successful. Isolation of antibiotics A / 16686 factor A 1, A2 and A3 by reverse phase HPLC using u-bonkapac R C18 "column and ammonium mixture. Formation in water And azetonitril in Convertible ratio as a mobile cycle (6 claims, 7 pages, 7 photos)