• 专利附录
  • 专利说明
  • 权利要求
  • 类似技术
  • 同族专利
  • 引用文献
  • 法律状态
  • 优先权
    • 专利摘要:
    Antibiotic A-21978 mixtures, in particular the A-21978C mixture, comprising microbiologically active, related factors Co, C1, C2, C3, C4, and Cs. A-21978 mixture and A-21978C mixture are produced by submerged aerobic fermentation of Streptomyces roseosporus NRRL 11379. The individual A-29178C factors are separated and isolated by chromatography. The A-21978 and A-2f978C mixtures; the A-21978C factors; and pharmaceutically acceptable salts thereof are antibacterial agents and improve growth promotion in poultry.
    公开号:SU1071226A3
    申请号:SU792822004
    申请日:1979-10-11
    公开日:1984-01-30
    发明作者:Л.Хэмилл Роберт；Мартин Хоен Марвин
    申请人:Эли Лилли Энд Компани (Фирма)；
    IPC主号:
    专利说明:

    15
    4000 3SOO then 2500 20001800 JSOO then CCW woo SOOWO 1 (00250
    (CM-f
    & 7V 9 10 21f1S1S202S30 0
    (pue.f
    The invention relates to the production of antibiotic A-21978 mixtures, mainly to the preparation of a mixture A-21978 e containing microbiologically active related factors GO, C, C3, C4 and Su. Mixtures A-21978 and A-21978 C by submerged aerobic fermentation of strain 11379. Individual A-21978 C factors are separated and isolated by chromatography. A-21978 and A-21978 C mixture, A-21978 C factors and their pharmaceutically applicable salts are antibacterial agents and promote the growth of poultry.
    A-21978 C blends are related acid peptide antibiotics. This class of antibiotics includes crystallomycin, amphomycin, zaomycin, aspartocin, and gluamycin t1J.
    Among these antibiotics, Cerexin A, Cerexin B and Brevestin are believed to be antibiotics most related to the newer A-21978 antibiotics.
    A known method for producing an antibiotic mixture, including cultivating strain 5758 or 8092 in a culture medium containing sources of carbon, nitrogen and inorganic salts, under aerobic conditions, until a significant antibiotic activity of NW is achieved.
    However, antibiotics obtained by this method do not possess growth-promoting activity for poultry.
    The purpose of the invention is to expand the range of antibiotics.
    The goal is achieved by the fact that according to the method of obtaining the antibiotic mixture, the strain Streptomyces roseoscope NPPLH379 is cultivated under aerobic conditions in a medium containing sources of carbon, nitrogen and mineral salts, followed by the separation of the individual components of the mixture.
    An example, Factor C A-21978 is designated as a mixture A-21978 C. Salts of mixtures A-21978 and A-21978 C, as well as individual A-21978. Factors C, C, C, Cj, C, and Cf form part of the invention.
    In A-21978 C, the mixture of factors C, Cg and Cj are major, and the factors Cjj, 04, and Su are factors present in small quantities.
    Antibiotic substances obtained by the proposed method, arbitrarily designated as A-21978 antibiotics. The term A-21978 antibiotics is used to abbreviate A-2197 and A-21978 C mixture, A-21978 factors Co, C, C4 and Cg, as well as their pharmaceutically applicable salts.
    The proposed method provides A-21978 antibiotic mixture as a result of submerged aerobic cultivation of the Streptomyces roseosporus NPPL 11379 schtam or its A-2197B producing mutant, as well as the antibiotic mixture A-21978 C and the factors Cd, C, Cj and C, which are its components.
    The invention also provides a method for producing an A-21978 mixture, which consists in cultivating the strain of Streptomyces roseosplus NPPL 11379 or A-21978 producing a mutant in a culture medium containing assimilable sources of carbohydrate, nitrogen and inorganic salts, under submerged aerobic fermentation to obtain significant amount of antibiotic activity.
    After a sufficient level of antibiotic activity A-2197 is obtained, the mixture is separated by filtration of the fermentation broth g and the pH is lowered. the filtrate to a value below 3, the mixture is allowed to precipitate and the resulting mixture is separated by filtration. The precipitated mixture can be subjected to additional purification using an extraction technique. Chromatographic separation is required to separate the individual A-21978 ° C and factors. A-21978 antibiotics dnguyut the growth of pathogenic organisms, especially gram-positive bacteria.
    FIG. 1-7 shows the IR spectra (tablets КВ2) of the following antibiotics A-21978 antibiotics (in the form of sodium salts): FIG. 1 - A-21978 mixture; FIG. 2 A-21978 C; the factor of FIG. 3 А-21978 С factor С-; figure 4 - А-21978 factor Сз; FIG. 5 - А-21978 С ФАЯЯ тор Ср; FIG. b - A-21978 factor of FIG. 7 - A-21978 C factor Su,.
    A-21978 C factors are the related related pibad antibiotics. As a result of fermentation, up to six antibiotic factors are regenerated and an A-21.978 mixture is formed. The individual factors CQ, C, C, Cj, C, and Cg are released as 0 of the individual compounds.
    A-21978 factors are related acid-related, cyclic polypeptide antibiotics that carry a fatty acid acial group at the terminal amino group. During hydrolysis, each of the factors forms the following amino acids, mol: Aspartic acids 4
    Glycine2
    Alanin1
    Serine1
    Theonii1
    Tryptophan1 Ornithine Kinurenin 3-Methylglutamic acid One of them is aspargin. May be formed from 3-methylglutamine. Each of the A-21978 factors contains a fatty acid. In tab. 1 summarizes the carbon content and identifies the fatty acids contained in each of the factors A-21978 C. Table 1 of 1 8-Methyllecanic Acid 10-Methylundecano KISMl. ,
     -N-Trp-Asx-Asx-TliP-bBu-OM-Asx-Aeoi-Asx-ue f-Ser-MeGfx-Kyn
    ..ON II I C W s; -N -T p-Asx-Ttir-G ev Orn-A3X-A a-Asx-Asx-E Eu-Sef-Me C UE Enzyme hydrolysis of A-21978 Su factor when using carboxypeptidase Y confirmed that kinurerin represents With a C-terminal amino acid and a C-terminal Soon group can esterify the hydroxyl group of the threonine moiety. Based on the above data, the following structure of A-21978 C antibiotics is suggested. L-ASPN iL-llaGJu L-ASpD-Ser L-OrnSlObGl L; rW (i.-Thr-0 cL-ASp d - ASp f d. - Tlvr in which 3MG is L Tpeo-3-methylglutamic acid Continuing Table 1 Edman's degradation reactions indicate that tryptophan is a ot-terminal amino acid, and a fragment of aspartic acid is the next neighboring amino acid .. Gas chromatographic and mass spectral study of A-21978 C of factor C2 shows that one of the following formulas can Describing the & structure of this factor (Asx denotes sparic acid or aspargin, and MeGlx - 3-methylglutamic acid Or 3-methyl glutamine) and R, is a special fragment of fatty acid, and special R-rpynyns. A-21978 C factors have the following value A-21978 e mixtures and factors (in the form of sodium salts) are soluble in acidic alkaline solutions except for solutions with mi rI 3.5, in lower alcohols, such as methanol, ethanol, propanol and butanol, as well as in dimethylformamide, dimethyl sulfoxide, dioxane and tetrahydrofuran; but only slightly soluble or completely insoluble in acetone, chloroform, diethyl ether, benzene, ethyl acetate, and carbon solvents. The salt forms of A-21978 C cm1gsi and factors are soluble in water, methanol, dimethylformamide and dimethyl sulfoxide, but insoluble in solvents such as ethanol, butanol and dioxane.
    In tab. 2 summarizes the approximate elementary composition of the sodium salt of Ks1 of A-21978 C factors,
    table 2
    The IR spectra of the A-2l978 C mixture and factors (in the form of Na salts) in tablets 45 are shown in FIG. 1-7. . 3 summarizes the most significant absorption maxima for each of these substances. Table 3
    Approximate molecular weights and molecular formulas of three basic-. nzkh A-21978 With factors summarized
    at . four,. „“ -,
    Ta b l and C a 4
    Table 5 summarizes the absorption maxima of the UV absorption spectra of the three main A-21978 factors (in the form of Na salts) in neutral ethanol. .
    Table3
    Continued table. 3
    Continuation of table.5
    thirty
     Below are summarized electrometric titration data, determined in a 66% aqueous solution of dimethyl; form of amide, for the three main
    А-21978 С factors and А-21978 С mixture
    5 in the form of rja salt), pK values:
    C, 5.7; 5.9; 7.2; 7.6 C | 5.8; 5.93; 7.6; 7.63 „
    5.73; 5.75; 7.54; 7.58
    50
    A mixture of 5.62; 7.16
    All have smaller groups at 11.5-12.
    Two definitions 55 Below are the rotation $ .- 21978 ° C factors (Na salts), totj when determining in water:
    , 9 (c.0,7;)
    +16.9 (s. O)
    0
    + 18.6 ° (s. O)
    + 20.9 ° (c 0.4; NDO)
    + 14.8 ° (c.0,)
    4 C5
    +17.9 (s.O, 7; HjO)
    65
    A-21978 C factors can be separated by liquid chromatography at elevated pressure (HPLC) using the following conditions:
    column: glass, 1 x 21 cm;
    gasket: silica gel (Quanti in LP-1);
    solvent: water: mgetanol: acetonitrile (95:30:75), containing 0.2% acetic acid and 0.2% pyridine;
    UV detector at 285 nm;
    pressure 100 psi.
    The retention times for factors A-21978 C (Na salt) are summarized in Table 2. 6
    T a b l and c a b
    A-21978 C, the mixture can be separated from factors A-21978 A, V. and E when using silica gel thin-layer chromium, tography. The preferred system of solvents is ay, a mixture of (aetonitrile: water (3: 1)), and the preferred method of detection is bioavtographic.
    Approximate values of Rl (chromatographic data from the indicated A-21978 G factors (as Na salts) are given below:
    A0,66
    With a mixture of 0.31
    D0,51
    E0.48
    Factors A-21978 C mixtures can be separated from each other using silica gel T C with a reversible phase (QuantiOT), The preferred solvent system is a water: meta-;: nol: acetonitrile (45:15:40) system, which contains 0.2 pyridine and O, 2% acetic acid. Long-wavelength UV light (365 nm) can be used for detection.
    The approximate values of the RP A-21978 C factors (in the form of Na s. Olir
    in such a system can be written as follows:
    0.71
    -0 0.64 C, 0.56
    C2 C, 0.47 0.63
    -four
    Cs0,53
    A-21978 C factors and A-21978 C mixture are stable in solutions having a pH of 2-9, at 5 ° and for at least 7 days. They are unstable at a pH of 11 after 4 hours (complete deactivation) at both 5 and 25 ° C.
    A-21978 C and A-21978 mixtures, as well as individual A-21978 C factors
    0 С
    G-.
    C4 and Cj
    capable of
    Z chz
    ABOUT
    salt formation. These salts are used, for example, to separate and purify mixtures and individual factors. In addition, especially. Cain% 1I are pharmaceutically applicable salts in which the toxicity of the compound n6 relative to warm-blooded animals does not exceed the toxicity of the non-salt form A
    A-21978 antibiotics contain up to five free carboxyl groups that can form salts, therefore the invention encompasses partial, mixed, and full salts of such compounds. When obtaining these salts, pH values above 10 should be avoided due to the instability of the antibiotic. otikov with such; pH values.
    A-21978 antibiotics also contain two free amino groups and therefore can form mono- and dis-acid addition salts. Particularly valuable are pharmaceutically applicable alkali metal, alkaline earth metal salts, ammonia, and also acid addition salts. Examples of such alkali metal and alkaline earth metal salts of antibiotics Y-21978 are sodium, potassium, lithium, cesium, rubidium, barium, calcium and magnesium salts. Suitable amine salts of antibiotics A-21978 include ammonium salts, primary, secondary and tertiary salts of C .4 alkylammonium, as well as hydroxy - C d-alkylammonium salts. As an illustration of the amine salts can be cited; obtained by the reaction of antibiotic A-21978 with ammonium hydroxide, methylamine, sec, butylMin, isopropylamine, diethylamine, diisopropylamine ethanolamine, t-ethylamine, 3-amkno-1-propanol, etc.
    Salts of A-21978 antibiotics, in which alkaline and alkaline-earth metals serve as cations, are prepared according to conventional methods used to obtain cationic salts. Thus, for example, the free acid form A-21978 GJ is dissolved in a suitable solvent, warm methanol or ethanol. To this solution is added a solution containing a stoichiometric amount of the desired inorganic base in a water solution of methanol. The salt thus obtained can be isolated by conventional methods, such as filtering or evaporating the solvent. Salts obtained using organic amines can be obtained in a similar way. Thus, for example, gaseous or liquid amine can be added to solution A-21978 of C fact C in a suitable solvent, such as acetone, and the solvent and excess of amine can be removed by fusing. Examples of suitable salts for the addition of A-21978 acids to antibiotics can be salts prepared by standard reaction with an organic or inorganic acid, for example, hydrochloric, sulfuric, phosphoric, acetic, succinic, citric, lactic, miloic, fumaric, palmitic, cholic , pamow, mucus, D-glutamic, D-camphor, glu: tar, glycolic, phthalic, tartaric, lauric, syaric, salits: fishing, methanesulfonic, benzenesulfonic, sorbic, picric benzo, cinnamic, etc. Vvet-erinar pharmacy good from estno that commonly used form of direct e antibiotic is not critical in the treatment of an animal after the antibiotic in most cases the conditions existing in the animal body is converted to drug pho mu different from that in which it was used. Therefore, the salt form in which the medicine is used is not essential. However, the salt form can be chosen / guided by considerations of energy, convenience and toxicity. ; Identification of the A-21978-producing strain. . The morphology of culture A-21978.b, which forms A-21978 antibiotics, consists of sporophores, which belong to the Rectus Flexibilis classification. Spore chains have 10 spores per chain. The surface of the dispute is smooth .. Culture A-21978.b characterizes with the production of mainly spore mass of red color with a reddish-brown reversible shade. There is also a light brown water-soluble pigment. These characteristics are obtained on three of 14 agar media in bowls (T5P # 2, ISP # 7, SRW). The three media are precisely those that confirm abundant air and vegetative growth. Two. Agar media in thickets, 15RJ4 and lglucose-paraffin agar produce a whitish-gray airborne spore mass with a yellow reversible shade. Water-soluble pigment is not observed. These two environments showcase-. They are good, but not abundant vzdushny and vegetative growth. Nine other agar media are also used, which ... however, give poor growth or do not give rise to growth and sporulation at all. The air color, when present, is xrt and is bad, but is related to a whitish-gray color series. Melanoid pigments are absent. The main constituents of the cell wall are LL-DAP, glycine, glucose and ribose. All of this points to a cell wall related to type 1 and to a sugar sample of type C / R (E. Buchanon, NE Gibbons. Running, geys Manual of Determinatire Bacteriology, Williams, Wilkins Gom. Ed. 8th, 1974, p. 658. The following five cultures are compared in laboratory tests with A-21978.6 Streptomyces albovinaceons I P5136; ATCG.15833 S. candidus I.P5141; ATCC 19891 S. moderatus I P5529; ATCC 23443 S. roseosporus 1 P5122; ATCC 239.58 S. setonii IP5395; ATCC 25497: These cultures belong to white and red-color series, have a sporophore morphology of the RF type / spore smooth surface and are melanin-negative according to ISP descriptions and are not clearly you -. reversible color or water-soluble pigments. These characteristics together with the sample for carbon utilization and other secondary distinguishing features of the present and culture A-21978.b. During laboratory testing of these cultures when compared with A-21978.b four of them give negative results . S. candidus and S. setonii demonstrate the formation of yellowish air spore mass in many media, as a result of which they differ from culture A-21978.b. S. albo-vinaceons and S. moderatus exhibit a distinct black reversible color, form water-soluble pigments and produce melanoid pigments. All of this is different from the A-21978 culture. B. According to the prescription of the ISP, the S. moderatus variety belongs, with a reddish-brown or dark brown reversible UBeTyj, however, these characteristics are not related to the S. albovlnaceons variety. None of the above categories is a melanin-positive. Therefore, the A-21978.6 culture has been found. classified by F lcaode Morias and Dalio Maio in 1961. Kai gatdm Streptomyces: ros eowroz j. The Tada classification is based on a comparison with published and direct laboratory comparisons. The following cultural characteristics summarize the direct comparative studies. Cultural characteristics. Morphology. A-21978.6 S, rbseos porus. Straight to flexible sporoforms (RF) without hooks, loop or spirals. Chains of more than 10 spores. The surface of the spores is smooth according to the data obtained using a scanning electron microscope. Disputes b.prodolgovaty From oblong oval; : up to the average size 0.85x1.78 M1.01x2.47 Interval 0.65-0.97x0.97-2.6 M О, 97-1.3x1 3.25 M Growth Color Growth Color Color Other; Gray-Morocre good Kovna No Yellow-ic brown mass Vegeta-Korichne-Choroshoytivny: vyy, raschy solution plentiful soluble pigment pigment from the absence Potato mass Airy Mr. Grayish-No Ze: good pink brown Vegetative Brown-Medium-Orangish, dark chewing. no-brown pigment soluble coping from ISP 1 non-pigment (tryptone yeast strategic agar) Air: White Poor White (W) and moderate (W) a VegetativeZheltov- Yellow Yellow-green- X10B2) green-native , wound, shy (10A1) flammable pigment cop otpigmen sunt - no vyusvuet Growth Color Growth Color (yeast-malt extract on agar J Aero-yellow Yellow-Obil-Orange is a rosy yellow-yellow (R) 5 cb, gy vy (R) Ssa Krasov a-Obil Vegetative: abundant brown nevy, (127) It Olive (5DLO) light brown VITAL, light brown pigment soluble pigment (oats on flour on agar) ISP 3 Air: Vely Poor White (W) is a basic (W) and Yellowhetale. roman Otsu stipe pigment ISP 4 (inorganic salts starch on agar) air: White Good Orange R) 3c2 vato (W). yellow etetivat- Orange-Obil-Sulfa11 good yellow, (10В1) green- (1115) solution , light-colored rodent kornich- otsut. Neutral Vuv soluble ISP 5 pigment (glycerin-aszarginovy agar) Purpur- Znach-Bely but-and-iachi spruce (K): Za egetiva- YellowGood Grayish-. (10С2) yellow, s: pryavatoy (ZV7) rososvetlovy,. brown pink soluble solution pipchente A-21978.6 ISP # 7 (S, roseosporus), (tyrosive agar) ost Color Growth Color air: Yellow-rich Yellow-to-rose toy-ro) 5fr5 «v (N) The 5th luminous autonomic-Krasnova-Obyl-Zheltovy: (7Y2) the one-that-brown-brown face is brown; (11E5) the nevalous, oval telyur-korotlorichny brown-brownish-brown; solution-pigment; the pigment; The modified Bennett 5 agar; Abundant Yellowish-to-rose (R) 5C8 vegetative Vegetative- Yellowish-Obil- Grayish: poorly dried yellow, brown I, (110) light astvori- korichnemy the pigment vy rastment not vorimy obrazuet- pigment Calcium malate agar Air: No Bad White no. (W) and the Vegetiva-Krasnova Poor Yellowish: a well-brown-green-20, a distinct nevy, a ny, a GELS7Y2) light brown-green green plant sour mold. 25.. . pigment. Chapek solution on agar air:. Poor (W) and White None Vegative, ny: bad White None 30 Soluble pigment does not form Emerson's Agar Air: bad Abundant Yellow. (R) 5 cb 35; vegetative- Ample Yellow; niy: plentiful (1115) light (13b6) .bright .new A-21978.6 Growth Color GrowthColor Glucose-aspartic agar Air: White - Mean-White 45 good body (W) b (W) b Vegetiva- Yellow, GoodHeltovany: co-solution-. (12В2) that is the greenest Roman, 50 (12В2) pigment, the pigment does not form the formed pigment. Glycerol-glycine, agar Vozltiny: Obil-Belyi bad ny (I) b Vegeta.tiv-Green-Obil-Yellow, Ny: cotton light {8Ы2) brown (10b3) brown-oyed, brown brown-colored soluble pigment pigment-65 Age No Vehygian Ros Ros Aimi (R) Vehirony solution- Romey, pigment Mets IS same (IS ISP i, however, Nutritious agar of the ear: Significant-White. Tel-tative-Yellow-Bad Yellow-yellow: full-gray, then gray-washed pigment does not form a pigment. Color Growth Color of tomato paste and wheat flour on agar ear: Yellow-.Ob- Yellow yellow-pink top-hooded (H) 5cY YOU take the Zelenova-Obil-Zheltova: Abundant-to-white-to-measles h (8Y2) brown, (12L7) nevy, brown: brown plant soluble soluble pigment Carbon utilization Substrate A-21978.6 L-Arabinose -f D-Fructose + D-Galactose, + D-Glukoza + 1-Inositol - Mannitol + D-raffinose, L-Ramnose, Salicin Sucrose, D-Xylose Note, +, poloelnoy intake; - - rs-on-calorie intake. Characterization A-21978.6 anoid pigments: # 1 (three-yeast extract) - - uton yeast on ezistom agar) -. . 7 (tyrosine agar) - # 7 modif, (13P # 7 minus tyrosine) - - Tyrazine test - Gelatin liquefaction + + Effect of sn - Minor milk hydrolysis Hydrolysis of starch, + + Interval pH 5-11 5-11 per day , ° C25-40. 25-45 Nitrate Reduction - +
    Tolerance to the action of NaCl growth, up to,% 10
     - stable (there are no zones of inhibition). Some characteristics of the A-21978 strain of the S. roseosporus NRRL 11379 strain are different from the S. roseosporus characteristics. Culture A-21978., B differs from the indicated strain in spore size, growth in carrots and potato masses, tolerance to NaCl and nitrate reduction. Streptomyces roseosporus is a cult pa used to obtain A-21978 antibiotics, deposited is part of the raw material complex. Culture Northern Regional Research Center, US Department of Agriculture. Agricultural Research Serviel Peoria, Illinoisi61604, from where it is used for use as number 11379.. As in the case of shch) microorganisms, the characteristics of the A-21978-producing Streptomyces ros osporus NRRL 11379 culture can be subjected to changes. For example, artificial variants and mutants of the NRRL 11379 strain can be obtained as a result of processing various mutagens, such as UV radiation, X-rays, high-frequency waves, radioactive exposures and chemical treatments. In the present invention, all natural and artificial variants and mutants of Streptomyces roseosporus NRRL 11379 which are produced by A-21978 antibios can be used. The culture medium is used to grow Streptomyces roseosporu
    Antibiotic sensitivity of antibiotics is presented in Table. 7,
    Table 7 NHRL 11379 and may be selected from a large number of media. However, in order to save production, obtain optimal yield and ease of product isolation, some culture media are preferred. For example, tapioca dextrin is a preferred carbon source for large-scale fermentation, although glucose, fructose, galactose, maltose, mannose, cottonseed oil, methyl oleate, glycerin, refined soybean oil, and the like can also be used. The preferred nitrogen source is enzyme-hydrolyzed casein, although soluble meat peptone, soybean flour, soy hydrolyzate, soybean meal, yeast, amino acids such as L-aspargin and DL-leucine, etc. can also be used. . The nutritional inorganic salts that can be added to the culture medium are soluble salts capable of forming potassium ammonium ions, chloride, sulfate, nitrate, and the like. Among these salts, kgSOj is particularly useful for the preparation of antibiotics. Molasses as molded dialysate and synthetic mineral mixtures can also be used. To obtain A-21978 antibiotics, it is preferred to use distilled or deionized water in a fermentation medium. Some of the minerals contained in tap water, such as calcium and carbonate, for example, do not contribute to antibiotics. The culture medium should also include trace elements necessary for the growth and development of the organism. Such trace elements are usually found as impurities in other ingredients of the medium in quantities sufficient to meet the requirements of microorganism growth. niema Sometimes during large-scale fermentation, small amounts must be added (for example, 0.2 ml / antifoaming agent such as polypropylene glycol if foaming becomes a problem. For the production of significant quantities of A-21978 antibiotics, the preferred method is immersed for aerobic fermentation in the reservoir. Small amounts of antibiotics A-21978 can be obtained by culture in a coil flask, due to the time lag in the preparation of an antibiotic usually associated with inoculation of antibiotics. vegetative graft material, which is obtained by inoculating He6oJibmoro of the culture medium with the spore shape of the microorganism’s micellar fragments to form a fresh, actively growing microorganism culture, then the vegetative graft material is transferred to a large reservoir. A-21978 is a producing microorganism. can be grown at temperatures lying in the range of 20-37 ° C. Optimal production of A-21978 C takes place in the temperature range of 30-32 C. Kai typically, sterile air is dispersed through it in the processes of aerobic submerged fermentation of the culture medium. For efficient production. A-21978 of anti-ribiotics, the percentage of air saturation at. during the cutting process, the sound should have a value higher, preferably higher than 30% (at 30 ° C and a pressure of 1 atm). When fermenting in a tank, it is believed to maintain the pH level in the fermentation medium in the range of 6.5-7.0. Such regulation can be achieved by adding appropriate amounts, for example, sodium HYDROXIDE (in the early stages of the process) and hydrochloric acid ( later stages). 3 caps A-21978 antnbiotics during fermentation can be monitored by testing samples of broth or solid micelle extracts for antibiotic activity against microorganisms that are sensitive to VIA deist antibiotics. One of the microorganisms for testing such antibiotics is Micrococcus luteus. The biological analysis is preferably carried out using a disk-disc technique on agar plates. . After receiving A-21978 submerged aerobic fermentation, antibiotics can be isolated from the fermentation medium by methods known in the art. The antibiotic activity obtained during the A-21978-producing fermentation is usually contained in broth. Therefore, the maximum regeneration of A-21978 antibiotics is achieved as a result of the initial filtration in order to remove the mycelial mass. The filtered broth can be purified in various ways to form an A-21978 mixture. The preferred method is extraction and precipitation to form an A-21978 mixture. Further purification and separation of the A-21978 C mixture and individual A-21978 C factors consists in additional adsorption and extraction. Suitable adsorption material for cleaning A-21978 C mixture and factors include anion-exchange resins: strongly basic (polystyrene, Bio-RAD AG 1 and 2, Bio-Rex, Dowex 1 and 2, Amberlite 1RA 400, 401, 410) moderately basic epoxy polyamine (Bio Rex 5 and Duolit AZOV) and weakly basic polystyrene or fonnel polyamines (Bio-Rad AG3, Duolit AB, A-7, Amberlite 1RA.68, 1R 45, 1R 4B), silica gel, phlorizil, polymeric adsorbents (XAD 2 and 4) , high porous polymer (Diaion NR-20). Sephadex G-10, G-25 and G-50, Bio-Gel P-2 and R-10, reversible phase resins (silicon. C-JQ and Cd), carbon, DEAE cellulose and DEAE Sephadex, polyamide, alumina and microcellulose. Sources: Bio-Rad and Bio-Gel resin - Bio-Rad lab. Richmond, California; Amberlite and XAD Resins Rohra and Haas So Philadelphia, Ra .; duolite resins - Diamond Shamrock Chemical, Redwood City, Cal .; cephadex resins - PharmaciaFine Chem., AB, Uppsala Sweden; duox resins- - Dow Chemical Co., Midland, Mich .; Diaion - Mitsubishi Chemical Industies Ltd., .Tokyo, Japan. XAD resin, silica gel and Cd - E Merck. Darrastadt, Germany. On the other hand, culture solids, including components among and mycelium, can be used without extraction and separation, but preferably after removal of water, as a source of A-21978 antibiotics. For example, the culture medium can be dried by lyophilization.
    and directly mix in promix.
    A-21578 mixture and individual A-21978 ° C factors are always in the form of a sodium salt. A-21978 and A-21978 C mixture and individual A-21978 C antibiotic factors Co, C-f, Cg / € 4 and C inhibit
    . NT
    not tested. The minimum inhibitory concentrations at which A-21978 C is a mixture of „n pipppnmv A-P1CH7Ya S L tgtooi yiriyi And the main A-21978 C factors are inhibiOne of the most important aspects and tabulated. 10 summarized values
    the shadow is the fact that A-21978MIK (diluted on agar) A-21978 ° C
    antibiotics inhibit the growth of microorganisms Cd, C. |, C2, Cj, C, and Cj of proganisms, which are resistant to 60tin of some microorganisms when compared to other antibiotic use of dilution methods on
    tikam.agare (10).
    the growth of some pathogens, odo6eHHo gram-positive bacteria. The minimum inhibitory concentrations (MICs), at which A-21978 factors and A-21978 C mixture inhibit selected bacteria according to the established standard dilution test on agar, are summarized in Table. eight.
    Ta 9litza8
    Table 9 Some bacteria are tested according to the dilution test on agar, summarized in Table 9. Pop A pt OA-21978 C antibiotics also inhibit the growth of some aerobic bacteria. In tab. And the activities of A-21978 C mixture and A-21978 C
    Symbio-
    128
    necrop 4
    A-21978 C factors demonstrate in vivo antibacterial activity against experimental bacterial infections. In that case, when two doses of the test compound are applied per g of plush, illustrating the infection, the observed activity is measured as the EDj-0 value (effective
    0,220,13 0,0b4 93
    21978 С
    0.5 21978 Cj
    0.160.13 0.032 59
    0.13 21978 C,
    0.06
    0.080.06 0.032. 66
    V4
    16 32
    128
    128
    .
    one . dose, mg / kg, protecting 50% of the tested animals, see article W, Wick et al. I. Bacterial., 81,233-235, 1961). ,
    The observed values are EDg-o for. A-21978 C mixture and A-21978 C factors C, C, Cj, C, and C are presented in table. 12. ..
    T and b l and% and 12
    0,130,3
    0,130,14
    0.030.09 C, C / C, and C factors against various anaerobic bacteria using standard agar dilution test. , Table .11 H1US, mg / ml, dilution on aga MIC, subcutaneous administration, MIC, oral administration, A-21978 C factors and A-21978 C mixture are effective agents for treating pyelonephritis. Thus, for example, with experimental inheritance transmitted by pyelonephritis in A-21978 rats, the factors provide protection against this disease, which is superior to vanomitsin. In this case, the bacterial culture of Streptococcus faecal is used (G.UIE), which is grown on Trypticase soy .agare (BBL) ,. suspended in cerebrospinal broth; 1 liter of oil (BBL) V taken 0.2 gl portions and frozen in liquid nitrogen. Bacterial suspensions for inoculation of rats are prepared daily by sowing 60 ml Trypticase flasks with soy broth (BBL) from frozen ampoules and growing the culture overnight with shaking. The culture is diluted to a concentration of 5-10 colonies forming units per ml. Test compounds are administered subcutaneously once a day for 7 days. All compounds are suspended in an O, 125% solution of carboxymethylcellulose. Experimental infection of rats was carried out according to the following procedure. Females of randomly selected white rats (Coxr-wistar) weighing 190-210 g were anesthetized by intra-syrup injection of 12 t of methohexital sodium. An experimental model of pyelonephritis is based on the study of Guz and Buson, according to which the left
    Continued table. 12
    The metering device is blocked for 20 minutes, after which 0.5 ml of the test organism is injected into the femoral vein. Anti-Microbial therapy is started 4-5 hours after infection. Four hours after the last treatment, the rats are killed and the left-sided kidney is removed and homogenized in a Dual1 dispersing device containing 9 ml of physiological saline. This treatment is a 10-fold dilution of kidney tissue. An additional ten-fold dilution is based on the expected number of bacterial cells present in the tissue of the disgenerate. Finally, some of the dilutions are placed on duplicate agar plates that are incubated overnight at 37 ° C. The therapeutic results are expressed in two ways: (i) the percentage of rats whose kidneys are less than 10 per 1 g of renal tissue, denoted as cured. , and. (ii) the percentage of rats with at least 4-1od by lowering the bacterial titer compared with the infected control kidneys. The control of the rats was treated with 0.125% carboxymethyl cellulose solution. The number of viable kidney tissues in the control rats 3 infected with S. faecalis, Ted is in the range of 1, 6x10 per g of homogenized tissue. The results of these studies are summarized in Table. 13.
    ; vitro susceptibility of S. faecal is Guze strain. Subcutaneous use. Data on the toxicity of the main A-21978 C factors and A-21978 C cm are summarized in the table, 14 / Tabl and c a 14
    Cl 250
    479 ± 32d
    365
    C 150-250 175 204 ± 17
    Sz 50
    70-75,160
    175-190 169 + 10 150
    In the case when A-21978 C mixture or D-21978 C factor is used as an antibacterial agent, the latter can be used
    Control A-21978 C
    25
    T-a blitz 13
    notion of MIC and ED
    and
    50
    toxicity data considered in conjunction with factors such as the patient or the host, as well as the infectious microorganism, A-21978 antibiotics can also be used as agents that promote animal growth. For example, in A-21978 chickens, the mixture increases the growth weight and efficiency of food consumption. Table 15 summarizes the results of two tests demonstrating such activity,
    Table 15
    1, 773
    - 734
    1.80 1.741 4.10 750 orally or parenterally. The A-21978 C mixture or factor can usually be used in conjunction with a pharmaceutically acceptable carrier or diluent. The dosage of the A-21978 C mixture or factor depends on a large number of circumstances, such as, for example, the nature and severity of the particular infection to be treated. Appropriate dosage intervals and / or dosage units for the application of morut should be determined on the basis of a note. Value in processing value control
    In this test, A-21978 C, the mixture was administered to animals with a concentration of 25 g / ton of food. An antibiotic is given in four repetitions: eight to eight birds, with each wire; a doubled study of time in series (a total of eight repetitions on eight birds or 64 birds). The test period is 21 days from 7 to 28 days of bird life. Data on increasing growth efficiency (weight gain, food intake, and nutritional efficiency) are compared with data obtained from 40 repetitions of a simultaneous control treatment, brkhki,.
    Typically, A-2197S antibiotics are effective in promoting the growth of poultry in the event that they are used in conjunction with food of animals in an amount of 1-100 g of A-21978 antibiotic per 1 ton of food of the animal.
    Example 1; Fermentation A-21978 C in a flask with shaking. Lyophilized granule of Streptormyces rbseosporus NRRL 11379 solution: T in 1-2 ml of sterilized water. The resulting solution is used to inoculate an oblique agar having the following composition.%: Glucose .0,5
    Yeast extract 0,2 CaCOj0,3
    Vegetable Juice 20.0 Agar2.0
    Unknown pH value is 6.1; pH value after autoclave process is 5.9.
    U / 8, juice, Campbell Soup Go .. Inoculated bevel is incubated for 70 days. Mature culture on the oblique agar is poured with sterile deionized water (10 ml). and scraped off with a sterile pipette in order to loosen the spores. A portion (1 ml) of the resulting spore suspension is used to inoculate a vegetative medium having the following composition,% Trypticase soy broth 3.0. , Dextrin 2.5 Water (deionized)
    Baltimore Biological Lab ,, Cocheuville, Md.
    32
    1071226
    Continued table. 15
    The inoculated vegetative medium was incubated in a 250 ml Erlen Meyer flask at 30 ° C for 48 hours in a shaking device rotating through an arc of 5 cm in diameter at a rate of 25.0 rpm.
    This incubated vegetative medium (0.5 ml) is used for 0, inoculating with 50 ml of production medium having the following composition, g / l: Glucose 7.5
    Tapioca dextrin 30.0 Enzymatic hydrolyzate 5 casein.5,0
    Enzyme-hydrolyzed .casein 5.0
    K2SO4.17,4
    Anhydrous Asparagine 1,320 Deionized water - Up to 1 l Stadex 11, A.E. S.taley, Co,
    Decatur, iii. NL Amine A, Sheffild Chemical
    Co, N.I.
    5 Amber EHC, Amber Lab., Luneau,. Wise.
    The inoculated production medium is incubated in a 250 ml Erlenmeyer flask at 30 ° C for
    0 DN, in a shaker, rotating through an arc of 5 cm in diameter at a speed of 250 rpm.
    PRI me R 2 Fermentation in the tank A-21 978 C, so that
    to obtain a larger volume of inoculum; 10 ml of the incubated vegetative medium is used to inoculate 400 ml of medium in the 2nd stage of vegetative growth, which has the same composition as the vegetative
    0 Wednesday. This medium of the 2nd stage tea is incubated in a 2 l flask for 48 hours while in a shaker rotating through an arc with a diameter of 5 cm at a speed of 250 rpm,
    5 The incubated vegetative medium in the 2nd growth stage (800 ml) was used to inject 100 liters of sterile production medium having the same composition as in Example 1. The inoculated production medium is fermented in a 165 liter fermentation tank for 6 -8 days at. The fermentation medium aerates. Sterile. by air
    5 under pressure of 1 atm in order to improve. maintain 30% boiling saturation, with the spirit while mixing with conventional stirrers at a speed of 3,200 rpm. Separation of A-21978 C antibiotic mixture. The entire fermentation broth (1600 gallons), obtained according to: measure 1, is filtered on a filter press using 3% filter media (Celite 545, lohns Manville). The filter cake is washed with water to form a filtrate in the amount of 4100 l, having a concentration of 230 units / ml, the pH of the filter is maintained at 3.5 with the aid of HC1 and the acidified filtrate is heated at room temperature for 16 hours in order to precipitate the active factors. A filtering agent (0.75% celite 545) is added to the suspension, and the resulting precipitate is separated by filtration. The filter cake is extracted twice with 410 L of methanol, each time stirring for 1 before filtering. Water was added to the combined meta-extracts (720 L) (0.1 volume or 72 L). The pH of this solution is maintained at 6.5-7.0 with NaOH. The resulting solution is concentrated in vacuo to 1/20 volume (30 L) in order to remove the methanol, and distilled water is added to the concentration as needed. With stirring, n-butanol (3/4 volume or 22 L) is added. The pH of the resulting solution is maintained at 3.0 with HC1, the phases are separated, and the α-butanol phase containing the activity is concentrated in vacuo to a residue. This residue is diluted with a minimum amount of methanol-methanol solution added to acetone (30 volumes for the purpose of precipitating the main part A- 21978 ° C. The precipitate is separated. By filtration and dried to form 247 g with a mixture of A-21978 ° C mixture (780 units, / mg). The methanol acetone filtrate containing the remaining portion of A-21978 mixture (Factor A and B) is concentrated to a residue that is dissolved in tert-butanol: (5: 1) and dried in the cold to form 169GA21078 secMix. -21978cX: mixtures. Crude A-21978 mixture (734 g) obtained in Example 2 was suspended in water (25 L), pH of the suspension and equilibrium 6.5 is established with 5 N, NaOH in order to completely dissolve the material. This solution is passed through a column containing 27 liters of ion exchange medium (acetate acetate resin cycle (IRA-68, Rohm Haas Co)). The column is washed with water (4 volumes or 108 L) and then 0.1 N acetic acid (5 volumes or 135 L). The active material is eluted with 0.5 N acetic acid, approximately 120 L fractions are collected and each is tested for biological activity. The highly active fractions are combined and dried in the cold to form a 278 g A-21978 mixture, which has a brown color (1100 units / mg). Fractions with low activity are combined to form 238 g of brown A-21978 C mixture (880 units / mg1 Further purification of A-21978 C mixture. Part of the more active A-21978 C mixture (150 g) eluted from the column with IRA -68, suspended in water (600 ml), the pH maintained at 6.5 to completely dissolve the suspended preparation, and a sufficient amount of dry silica gel (Grace, grade 62) is added in order to absorb the aqueous solution. the preparation is placed in a 30 l. silica gel column (Grace 62.10x375 cm) treated with acetrinyl (with the silica gel is pre-washed with water to remove small particles, then the column is packed with silica gel suspended in water and the silica gel column is washed with 30 l of acetonitrile). After loading, the column is washed with acetonitrile (15 l), then develop with acetonitrile: water (4 : 1) and collect 4 liters of the fraction. The elution was monitored by biological analysis, and the silica gel of those (CH) bioautogram was removed. The fractions containing only A-21078 C mixture (fractions 43-60) are combined, concentrated in vacuo and dried in the cold to form 86.2 g of yellowish purified A-21978 C complex (1160 units / mg). The fractions containing factors D and C are combined and dried in the cold with the formation of 13 g of a yellowish powder with low biological activity. The purified A-21978 C mixture (30 g) thus obtained is subjected to additional provision by suspending 30 g of the mixture in a minimal amount of water and mixing with a small amount of gel-gel (type P-1, 10-20 µm Quantmn Industres, 341, Kaplan Drive, Fairfield, NI 07006) to absorb the solution. The wet silica gel mixture is suspended in acetonitrile: methanol (4: 1) and filled with a glass-lead column {external diameter cm) attached to a glass column (external diameter 4x30 cm) containing 2.8 l of silica gel. The silica gel suspended in acetonitrile: methanol (4: 1) is pre-washed with water and then the mixture
    acetonitrile: methanol (4: 1). The column is filled with silica gel in acetonitrile: methanol (4: 1) at a pressure of 50-60 psi. The lead and base columns are washed with 3 L of acetonitrile: methanol (4: 1) at a pressure of 5 psig. The active material was eluted with acetonitrile: methanol: water (55:20:25) and 300 ml fractions were collected. Over the course of elution, barely, d t according to the data of biological analysis of u (micrpcoccus luteus). Fractions 14-25, possessing the highest activity, are combined, concentrated and dried on cold to form 24 g of light yellow, pure A-21978 C mixture with sodium salt (1200 units / mg). Less active fractions 26-32 are combined, concentrated and dried in the cold to form 1.6 g of the less pure A-21978 C mixture (780 units / mg).
    Separation of A-21978 C factors. The purified A-21978 C mixture (2 g) obtained in Example 3 was dissolved in water (40 ml) and pumped through a pump (FMI, LAB Pump, Fluid Metering, Jnc. 48, Summit, NI 11771) under pressure at 60 pounds / inch 4.1 x 60 cm column with reversible silica gel phase Quantmn LP -1 silica gel (C d), water: methanol: acetonitrile (100: 15: 85) containing 305 containing 0.15% acetic acid and 0.15% pyridine. This column is developed under a pressure of 65 psi with a solvent, while collecting 25 ml fractions. The zlation of factors is monitored according to the UV-spectrum of an Oscopy at 280 nm and using biological analysis. , Indi-1 fractions are analyzed on an analytical column for factor purity.
    The data of a typical separation is as follows: fractions contain factor C, fractions 45-53 - factor C,: 45 fractions 75-92 - factor €, fractions 112-134 - factor Cj, fractions 54-74 -. factors C, C2 and Cj, and fractions 93111 - factors C2, Cj and Su. The fractions containing mixed factors, repeat -, /, but are passed through the column in order to obtain additional quantities of factors. Fractions containing a single factor are combined,. concentrated in vacuo and dried n-a .. cold with the formation of light yellow powders of each of the factors (in the ni-. de Na salts). The following yields are obtained from 60 g of the mixture: factor C 5.55 g; factor C2 10 g g Factor C 6.61 g. Fractions containing mixed factors are recycled through a column with an ion exchange resin with a reversible phase and the formation of additional yields: factor Co 550 Mg; factor C-, 1.29 g; factor C 1.99 g; factor 65
    C, 443 mg; factor C4, 512 mg; factor Cj 384 mg.
    Large-scale purification and separation of A-21978 C factors. On a larger scale, the factors are separated by reversible phase chromatography. The pure A-21978 C mixture (6 g) obtained in Example 3 is dissolved in water (80 ml), the pH of the solution is set to 4.4 with acetic acid and tetrahydrofuran (20 ml) is added. The resulting solution is pumped under reduced pressure (Lapp Pump) through a steel column (4.8 X 100 cm) containing 1.77 liters of silica gel (C gtQuantxom LP-1, 10-20 μm, silylated with octadecyltrichlorosilane) in a system: water: tetrahydrofuran (THF) (4: 1). Such a column is washed under a pressure of about 100 psig for 150 ml of OGTGF (4: 1) .. The column is eluted with water: methanol: acetonitrile (47.5: 15: 37.5) containing 0.2% pyridine and 0 , 2% acetic acid at a pressure of about 100 psi, at a flow rate of 35 ml / mi, with a 175 ml fraction being taken. Elution is continuously recorded on a recording device using a UV detector at a wavelength; 280 nm. The fractions containing individual factors according to the peaks in the graph are additionally analyzed on an analytical column with a reversible phase. Fractions containing one factor are combined and dried in the cold. Typical experiments have the following results: fractions 12-16 contain Co factor, fractions 20-26 - factor C fraction 38-50 - factor C2, and fraction 63-78 - factor Cj. Fractions 2737, containing factors C and C, and fractions 51-62, containing factors C and Cj, are recycled and through the column to obtain pure factors C4 and C. The load is 612 g. The following yields are obtained from A-21978 C , g: C 1.9; C 3.27; C2 4.97; Cd 1.94. Increased yields of individual factors are obtained by recycling fractions containing mixed factors using appropriate HPL C solvent systems. The choice of system varies to depends on the individual groups, as well as the nature of the reverse phase resin and columns.
    The following are the systems used to separate the A-21978 C factors.
    Analytical systems.
    Water: methanol: aceTonitrile (50: 15: 35), containing 0.2% acetic acid (HOAc), maintained at a pH value of 5.5 with pyridine. Water: methanol: acetonitrile (50: 15: 35), containing 0.2% HOAc and 0.20% pyridine. Ox: methanol: acetonitrile (50: 15: 35) containing 0.55% ammonium formate. Water: methanol: acetonitrile (95: 30 J 75), containing 0.2% NOA and 0.2% pyridine; . Water: methanol: acetonitrile (105: 15: 80), containing 0.2% NOA and 0.2% pyridine and Water: Methanol: THF (99: 15: 25) containing 0.5% HOAc and 0.5% pyridium Water: methanol: THF (60: 15g 25) containing 0.5% ammonium formate, Preparative systems. Water: methanol: acetonitrile (95: 20:85), containing 0.15% HOAc and 0.15% pyridine. Water; methanol: acetonitrile (100: 15: 85), containing 0.15% HOAc and 0.15% pyridine. Water: methanol: acetonitrile (50: 10: 40), containing 0.1% HOAc and Of 1% pyridine. Water: methanol: acetonitrile (50: 15: 35), containing 0.75% ammonium formate. Water: methanol: acetonitrile (55; 10; 35), containing. 0.2% HOAc and 0.8% pyridine. Water: methanol: THF (52.5: 15: 32.5) containing 0.6% ammonium formate. Water: methanol: THF (50: 15: 35), containing 0.6% ammonium formate. The advantage of the acetic acid system (pyridine compared to ammonium formate) is that the first can be removed in an ice-cream, while ammonium formate should be removed by x-ray chromatography (Sephadex C-25). . Variable removal of A-21978-C mixture. The whole fermentation broth (97 l) obtained in example 1 is filtered using filtering agent (Hyflo Super-gel). The resulting filtrate (80 l) is mixed with 2 l of non-ionic macroporous styrene copolymer cross-linked with divinylbenzene HP-20 (ion exchange resin. Mitsubishi Chem. Ind, Lim, (Tokyo, Japan) for 2 h The upper layer is decanted, the resulting resin is washed with water (8 L), and the water is also desiccated. The resin is then mixed with 8 L of acetonitrile: water (15: 8 for 15 minutes and the solvent is removed by filtration. Then A-21978 the mixture was eluted from the resin by stirring with 8 l of acetonitrile: water (2: 3) for 1 h and filtered. The test is repeated until the A-21978 C is completely removed. The two filtrates are combined and concentrated in vacuo to form an oil. The resulting oil is dissolved in a minimum volume of water, methanol (2 volumes) is added at maturation and then acetone ( 30 volumes) to precipitate A-21978 C mixture. The precipitate is separated by filtration and dried in Vacuum to form 13.6 g of raw material A-21978 C mixture (570 units / mg). Crude A-21978 C mixture is purified by silica gel column chromatography. The resulting mixture (1 g) is dissolved in a minimal amount of water, silica gel (Grace-62) is added for absorption, and the absorbent is slurry in acetonitrile. The resulting sludge is passed through a column of 1.5x40 cm with silica gel (Grace-62) in acetonitrile. The column is then washed with acetonitrile. The activity was eluted with acetonitrile: water (4: 1), and 25 ml of the fraction was collected. The fractions are analyzed as in Example 3. The fractions 21-46f containing most of the A-21978 C mixture are combined, concentrated to a small volume in vacuum and dried in the cold to form 605 mg of purified A-21978 C mixture (Na salt) (900 units / mg). Preparation of A-21978 C mixture (acidic. Form). A-21978 ° C. The mixture in the form of a sodium salt (7 g) was dissolved in water (150 ml) and n-butanol (150 ml) was added. The pH of the solution is maintained at 3.4 with 2 CH2 while stirring for 1 hour. The n-but-nol phase is separated and concentrated to a residue in BaKys Me. The residue obtained is dissolved in water and dried in the cold to form b g A -21978 C mixture (acid form), Individual A-21978 C factor in the form of a salt is converted into the corresponding acid form using the same procedure. ; Preparation of A-21978 C mixture in the form of a sodium salt from A-21978 C mixture in an acid form. A-21978 with a mixture in acid form (50 mg). dissolved in warm absolute ethanol (5 ml), added dropwise is 1N. NaOH solution until the solution reaches a pH of 9.4. The resulting solution was incubated at room temperature overnight. The sediment is then filtered and dried under vacuum to form 32 mg A-21978 C mixture (sodium form). The resulting salt contains 8% sodium according to an atomic absorption assay. Using the technique indicated above, A-21978 C, a mixture in the form of an alci salt is obtained by adding CaClj ethanol to an ethanol solution of -21978 C mixture in acid form. Microbiological Analysis of A-21978 Embedded and Allocated Images. To quantify the A-21978 activity in the fermentation broth and in the isolated samples, a paper disk system is used to determine the diffusion on the agar using Micrococcus tu teu5.
    The seeded agar-diffusion bowls are prepared by inoculating the nutrient agar medium with an appropriate concentration of the test culture, pouring 8 ml of agar into each of the plastic Petri dishes (20 x 100 mm).
    The reference standard for this analysis is A-21978 .5
    100 W SB W 20
    then 3500 then 25oo 2oooo im sho then IIQQ yooo 900 soofoozso
     5 6 7 81910 izmeismsmo
    2.5 3
    100 80
    SO 40 20
    3500 3000 2500 20001800 THIS WOW 800 600 t00250
    si, which is used per unit of activity. High-purity A-2197 B mixture contains about 1250 units / m. The result is a response curve of a standard dose containing 150-75-40-20-10 units / ml. As a diluent for standard and samples, 0.1 M Phosphate buffer having a pH of 6.0 is applied.
    Solutions of the sample and standard are transferred onto 12.7 NIM paper discs using automatic pipette. Incubation is carried out for 16-18 hours.
    S 7 8 3 1012 n ismmmo
    (
    (pUf: 2
    fie.W (CM-t)
    100 80
    SB
    w
    20
    o 3500 3000 2SOO 2000 mo 1SO014001100 1000 800 SOO 400250. IPU {. four
     - t
    (cM-f}
    3f 5 6 7 8 3 10 1g 1lHSm0253010
      1 I r I I 1 I;
    SHO 3500 woo 2500 20001800160014001200.1000 fpus. five
    2.5
    100 80 SO 2Q
    mo 3500 mo 2500 20001500 tm mo im woo soo soo 400250
    Fig.e.
    800 f 00
    (CM-}
    ff 7 8 ff S12 n 1Sl8202Sm5
    (CMf)
    h 5
    5 3
    yo
    80
    so w
    20
    s 7 8 9 JO 12 n cpgo25zo
    -1111-t111 II I J
    0
    woo 3500 3000 2500 20001800 mO W012001000 SOO f 00100250
    FIG. 7
    权利要求:
    Claims (2)
    [1]
    METHOD FOR PRODUCING AN ANTIBIOTIC MIXTURE; namely, 1 that carry out the cultivation of the strain: Streptomyces roseosporus NPPL 1137 9; under aerobic conditions in an environment containing sources of carbon, nitrogen in ° mineral salts, followed by the allocation of individual components of the mixture
    7 8 3 10 ^ 2 14161820253040
    100
    [2]
    2.5
    • '' G ........ ..... ι 1----- " ' .1 1 — I - g - 1 1 II II II. •x 4 -'ch- •.• .1 ------ t ------------ 1 ί. '; H Λ · ... · j ..... A ', ·, 1 1 ...... 1. ,, .1
    (cm-η Fig. 1 0 4000 3500 3000 250020001800 16001400 1200K
    类似技术:
    公开号 | 公开日 | 专利标题
    SU1071226A3|1984-01-30|Method of prerating antibiotic mixture
    EP0055070B1|1986-10-29|Antibiotic a-4696 factors b1,b2,b3,c1a,c3 and e1
    EP0055069B1|1985-01-23|Derivatives of actaplanin
    SU716524A3|1980-02-15|Method of preparing substances possessing antiparasitic activity
    EP0182315B1|1989-04-05|Novel antibiotic nk84-0218 pharmaceutical compositions containing it and process for the production of the same
    PT726902E|2000-06-30|POLYCYCLICAL AGENTS ANTIPARASITARIAN ITS PROCESSES AND PREPARATION AND USES OF PREPARATION AND USE
    US4331594A|1982-05-25|A-21978 Antibiotics and process for their production
    USRE32333E|1987-01-20|A-21978 Antibiotics and process for their production
    USRE32455E|1987-07-07|A-21978 antibiotics and process for their production
    US4415557A|1983-11-15|Antibiotic compound, its production and its medicinal use
    US4174390A|1979-11-13|Antibiotic A-7413 and process for preparation thereof
    US3780174A|1973-12-18|Antibiotic a477 and process for preparation thereof
    USRE31396E|1983-09-27|A-21978 Antibiotics and process for their production
    FI101402B|1998-06-15|A method for preparing a new antibiotic, balhimycin
    RU2134694C1|1999-08-20|Aminooligosaccharide "sk-4416", method of its synthesis, inhibitor of saccharide hydrolase and antibacterial agent
    US20090023665A1|2009-01-22|Antibiotics ge 81112 factors a, b, b1, pharmaceutically acceptable salts and compositions, and use thereof
    KR830001069B1|1983-06-01|A-21978 Preparation of antibiotics
    KR970002492B1|1997-03-05|Novel tripedtide derivatives
    US5565561A|1996-10-15|Natural substance cyclamenol and chemical derivatives
    SU632307A3|1978-11-05|Method of producing polypeptide antibiotic complex exibiting antifungus activity
    KR830002817B1|1983-12-26|Method for preparing biologically active FR-900156 material
    US4482488A|1984-11-13|Antibiotic A53868 and process for production thereof
    EP0385594B1|1996-05-01|Acidic polycyclic ether antibiotic
    SU470964A3|1975-05-15|The method of obtaining antibiotic
    IE45014B1|1982-06-02|Antibiotic a-7413 and process for preparation thereof
    同族专利:
    公开号 | 公开日
    ZA795435B|1981-05-27|
    DK148918B|1985-11-18|
    JPS6455184A|1989-03-02|
    GT196000041A|1981-04-04|
    EP0010421B1|1990-10-10|
    BE879401A|1980-04-15|
    IL58448A|1983-03-31|
    PH16248A|1983-08-16|
    MY8500568A|1985-12-31|
    PT70298A|1979-11-01|
    DK434479A|1980-05-14|
    FI793190A|1980-04-17|
    HK42987A|1987-06-05|
    PL218977A1|1980-06-16|
    AR221738A1|1981-03-13|
    DD146618A5|1981-02-18|
    ATA669979A|1981-05-15|
    IE48564B1|1985-03-06|
    NZ191822A|1982-09-14|
    EP0010421A1|1980-04-30|
    DOP1979002809A|1987-12-03|
    GB2031437A|1980-04-23|
    YU247079A|1982-10-31|
    IE791948L|1980-04-16|
    IL58448D0|1980-01-31|
    LU81794A1|1980-05-07|
    US4208403A|1980-06-17|
    FI65631C|1984-06-11|
    PL122756B1|1982-08-31|
    FR2444079A1|1980-07-11|
    EG14631A|1985-03-31|
    GR73146B|1984-02-10|
    AU5175779A|1980-04-24|
    GB2031437B|1982-10-27|
    ES485074A1|1980-06-16|
    FR2444079B1|1983-08-05|
    HU180746B|1983-04-29|
    FI65631B|1984-02-29|
    RO77097A|1981-06-22|
    JPH0123479B2|1989-05-02|
    JPS5592353A|1980-07-12|
    BG41138A3|1987-04-15|
    AT365235B|1981-12-28|
    AU531616B2|1983-09-01|
    DK148918C|1986-05-12|
    CS219259B2|1983-03-25|
    CA1137900A|1982-12-21|
    DE2967701D1|1990-11-15|
    CH655127A5|1986-03-27|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    US4482487A|1982-05-21|1984-11-13|Eli Lilly And Company|A-21978C cyclic peptides|
    USRE32310E|1982-05-21|1986-12-16|Eli Lilly And Company|Derivatives of A-21978C cyclic peptides|
    USRE32311E|1982-05-21|1986-12-16|Eli Lilly And Company|Derivatives of A-21978C cyclic peptides|
    US4399067A|1982-05-21|1983-08-16|Eli Lilly And Company|Derivatives of A-21978C cyclic peptides|
    US4537717A|1982-05-21|1985-08-27|Eli Lilly And Company|Derivatives of A-21978C cyclic peptides|
    US4524135A|1982-05-21|1985-06-18|Eli Lilly And Company|A-21978C cyclic peptides|
    IL68700D0|1982-05-21|1983-09-30|Lilly Co Eli|Improvements relating to a-21978c cyclic peptide derivatives and their production|
    US4396543A|1982-05-21|1983-08-02|Eli Lilly And Company|Derivatives of A-21978C cyclic peptides|
    IL76608A|1984-10-09|1991-03-10|Lilly Co Eli|Process for the production of a-21978c derivatives|
    US4800157A|1985-09-09|1989-01-24|Eli Lilly And Company|Process for producing the A-21978C antibiotics|
    US4874843A|1987-12-03|1989-10-17|Eli Lilly And Company|Chromatographic purification process|
    US4994270A|1988-04-11|1991-02-19|Eli Lilly And Company|A54145 antibiotics and process for their production|
    US5028590A|1988-04-11|1991-07-02|Eli Lilly And Company|Derivatives of A54145 cyclic peptides|
    US4977083A|1988-04-11|1990-12-11|Eli Lilly And Company|Processes for preparing A54145 compounds|
    JP2863934B2|1989-07-24|1999-03-03|塩野義製薬株式会社|Antibiotic plus basin|
    US7235651B2|2001-12-26|2007-06-26|Cubist Pharmaceuticals, Inc.|Genes and proteins involved in the biosynthesis of lipopeptides|
    CN101165168B|2006-10-20|2010-05-12|上海爱普香料有限公司|Streptomycete and method for producing vanillin by using the same to biologically transform ferulic acid|
    IT201600127655A1|2016-12-16|2018-06-16|Gnosis Spa|PROCESS FOR THE PURIFICATION OF LIPOPOLIPEPTIDIC ANTIBIOTICS|
    法律状态:
    优先权:
    申请号 | 申请日 | 专利标题
    US05/951,695|US4208403A|1978-10-16|1978-10-16|A-21978 Antibiotics and process for their production|
    [返回顶部]