JPS6127369B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to compositions containing antibiotics. The present invention provides compositions containing the new antibiotic 31559RP or its salts with metals or with nitrogenous bases. 31559RP has valuable anticoccidial, antibacterial, and antimalarial activities. It is also valuable as a growth factor for ruminants and as a drug to prevent dysentery in pigs. The new antibiotic 31559RP, its properties and production will be described later, but they are also described in Japanese Patent Application No. 50-26443 (see Japanese Patent Application Laid-open No. 129796-1982), which corresponds to the original application of the present application. The present invention contains 0.0001 to 99.9% by weight of 31559RP or its salts with metals or with nitrogen-containing bases and a substance consumable by animals (excluding humans; hereinafter the term animal refers to animals other than humans). ) provides a composition for administration. (Of course, any salt used must contain cations that are non-toxic at the concentrations and dosages administered.) Such compositions may be used as mixed animal feeds or concentrates for feeding animals. blend,
31559RP or its salts with metals or with nitrogenous bases, and optionally other anticoccidial agents. These compositions are particularly useful for combating coccidiosis in birds, and particularly in poultry. The dose required to produce a suitable effect can, of course, vary within a fairly wide range, depending on the proportion of feed. Generally speaking, feed given to animals contains 0.005 to 0.04% by weight of 31559RP or its salts.
It is sufficient to include it. 31559RP or its salts can be distributed throughout the complete feed as a uniform dispersion at the above concentrations. It can also be distributed throughout the supplementary feed in doses of 0.025 to 1%, often accompanied by other additives such as vitamins and inorganic salts. These supplementary feeds may be mixed with the main food or may be eaten as is. These usually represent 5 to 20% of the staple food. "Premixes" used to make complete meals or supplementary feeds are diluted in feed extenders.
It usually contains 0.05 to 20% of 31559RP or its salts. These are convenient intermediates that facilitate uniform distribution of the active product throughout the feed. The premix itself consists of 31559RP or its salts 99.9 to 20% with the addition of edible modifiers, such as edible dyes, flavoring substances, dispersants, or anti-agglomeration agents, and feed fillers.
It is generally produced from a concentrate containing %. Concentrates and premixes are generally powders. Supplementary feeds and complete feeds can be either flour or granules produced by conventional techniques. The following examples (percentages expressed by weight) illustrate compositions according to the invention. Example A A basic feed having the following composition is prepared: Grain flour 13.41% Barley flour 13.41% Corn flour 13.41% Wheat flour 31.32% Fish meal 8.92% Soy flour 8.92% Dehydrated forage flour 4.56% Yeast extract 2.33% Dry milk Powder 2.68% Sodium chloride 0.09% Calcium chloride 0.89% Mineral components 0.06% Vitamin complex: Vitamin A 4000 IU/Kg Vitamin D ₃ 1000 IU/Kg Choline chloride 11.5 mg/Kg Riboflavin 2.24 mg/Kg Sodium salt of 31559RP 0.02 % to this feed and distribute it evenly throughout. The present invention provides a composition based on 31559RP or its salts that can be used as a growth factor for ruminant animals (cattle, sheep, and goats) and as a drug for combating dysentery in pigs. These compositions consist of mixed feeds for ruminants or pigs containing 31559RP or its salts and concentrated mixtures for feeding animals. The dose required to produce a suitable effect may, of course, vary within wide limits depending on the species of animal and on the nutritional value of the feed itself. It is particularly advantageous to feed the animals with a daily dose of 50 to 250 mg of 31559RP per animal. 31559RP is preferably distributed throughout the supplementary feed containing 0.01 to 0.1% of the antibiotic, often along with other additives such as vitamins and inorganic salts. These supplementary feeds may be mixed with the main diet or eaten alone, and usually represent approximately 1 to 10% of the daily food intake. In this way, 31559RP is
Corresponds to 0.0001-0.01% by weight. "Premixes" used to make complete rations or supplementary feeds generally contain 0.1 to 5% 31559RP diluted in feed filler. These are convenient intermediates that facilitate uniform distribution of 31559RP throughout the feed. ``Premix'' itself is 31559RP with the addition of edible modifiers, such as edible dyes, flavoring substances, dispersants, or anti-agglomeration agents, and feed fillers.
It is generally produced from concentrates containing 99.9% to 5%. Concentrates and premixes are generally powders. Supplementary feeds can be either flour or granules manufactured according to conventional techniques. The 31559RP or salt thereof in these compositions can be in the form of fine particles, either uncoated or covered with a coating. The following example illustrates the use of 31559RP as a growth factor. Example B Twelve young steers of the Holstein breed are placed in individual pens (their average surface area is 48 m ² ) and then weighed. The feed to be investigated is supplied from day 0, that is, from the start of the experiment. The steers are then weighed after 28 days and then 56 days to determine the weight of feed consumed during this period. 0-28 days and
For each feed investigated within a period of 28-56 days, the average weight gain and average degree of feed conversion are determined. Feed the animals twice a day. Daily food is
Decide so that the steer absorbs the maximum amount of food without leaving anything behind. In this experiment, young steers are divided into three groups. Each animal receives a basal diet consisting of corn, hay, and protein supplement. Each animal in the group receives a basal diet. Each animal in the group received 100 mg of antibiotics with protein supplement
Receive basic feed containing 31559RP. Each animal in the group received 200 mg of antibiotics with protein supplement.
Receive basic feed containing 31559RP. The average daily ration per animal has the following composition:

【table】

【table】 The results obtained are shown in the following table.

【table】

[Table] In this way, in the first 28 days, 1
Animals receiving 100 mg of 31559RP per day gained 3% more weight than control animals and had a lower degree of conversion.
1.3% less, these percentages are 31559RP 100
15.3 for animals receiving mg/head/day, respectively.
% and 13.4%. 100 mg/day in the second period (28-56 days)
Animals receiving 31559RP catch up with animals receiving higher doses. Then, the weight gain is 13.4% higher and the degree of conversion is 9.8% lower than that of the control at a dose of 100 mg/day. and these percentages are 9.6% and 7.6%, respectively, for a dose of 200 mg/day. Over the entire period of the experiment (56 days), at each dose of 31559RP 100 and 200 mg per day, the average daily acquisition was
7.8 and 12.2% improvement, conversion degree is 6.4% and
Improved by 10.2%. 31559RP, which was later fully identified, belongs to the genus Streptomyces and is known as Streptomyces hygroscopicus DS 24367 (NRRL
It is obtained by culturing a new microorganism called 5787). This microorganism has been deposited with the Institute of Microbial Technology as Microtechnology Research Institute No. 4329. 31559RP contains carbon, hydrogen and oxygen. The elemental composition of the sodium salt is approximately as follows: C = 63.2%, H = 8.8%, O = 24.3%, Na = 2.9
% Its neutral equivalent (determined by titration of the sodium salt with perchloric acid in acetic acid) is 836. Its elemental analysis and its proton and carbon
From ^{the 13} C NMR spectrum, the sodium salt of 31559RP probably has the following empirical formula: C ₄₄ H _73-75 O _13-14 Na. Furthermore, the proton and carbon ¹³ C NMR spectra show the presence of four methoxy groups in the molecule. The sodium salt of 31559RP has the following properties. This is a white microcrystalline powder:
[Pharmacopee Francaise 2nd edition, Part 2, 13
(1972)] It is virtually insoluble in water, sparingly soluble in hydrocarbon solvents such as hexane, soluble in alcohols such as methanol, ketones such as acetone, dimethylformamide, and ethyl acetate. Easily soluble in chlorinated solvents such as methylene chloride and chloroform: Its melting point (measured on a Koffler bench) is 220 °C
(with decomposition): Its optical rotation: (C=1.050 in methanol) [α] ²⁰ _D = +51.2° ± 1° [α] ²⁰ ₄₃₆ = +101° ± 1.5° [α] ²⁰ ₃₆₅ = +167゜±2゜: Its ultraviolet spectrum (0.950mg/cc in ethanol
(measured in a solution with a concentration of 210 nm). Its infrared spectrum (measured on a tablet of the mixture with potassium bromide) is shown in Figure 1 of the accompanying drawings, in which the wavelength in microns (upper scale) and wavenumber in cm ^-1 (lower scale)
is plotted on the horizontal axis, and the optical density is plotted on the vertical axis. The main infrared absorption bands (expressed in wave numbers (cm ^-1 )) of this compound are shown in the following table.

【table】

Table: The sodium salt of 31559RP was also characterized by silica gel thin layer ascending chromatography using two mixed solvent systems: (1) ethyl acetate, cyclohexane, water and butanol (50-50-25-5 by volume); ): In this system, the sodium salt of 31559RP has an Rf of 0.5. (2) Methylene chloride and methanol (94− by volume)
6): In this system, the sodium salt of 31559RP is
It has an Rf of 0.7. This sodium salt of 31559RP was found to have the following formula: In Vitro Bacteriostatic Activity 31559RP exhibits bacteriostatic activity, particularly against certain Gram-positive bacteria. against some microorganisms
The bacteriostatic activity of 31559RP sodium salt was determined by one of the dilution methods commonly used for this purpose. For each microorganism, the minimum concentration of active substance that inhibits visible growth in a suitable nutrient culture medium under the specified conditions is determined. The results of various measurements are shown in the following table, where the minimum bacteriostatic concentration is 1 c.c. of the test medium.
It is expressed in micrograms of substance per unit.

[Table] Toxicity The 50% lethal dose (LD ₅₀ ) of 31559RP sodium salt in chicks is 150 mg/Kg as a single dose.
(oral). Anticoccidial activity The anticoccidial activity of 31559RP sodium salt is determined in chicks infected with Eimeria tenella or Eimeria acervulina. Substantial anticoccidial activity is shown when the sodium salt of 31559RP is incorporated into the feed at non-toxic concentrations of 0.005 and 0.04% by weight. The organism producing 31559RP is a Streptomyces strain taken from a soil sample in India and has been given the number DS24367. The samples were taken to the Northern Regional Research Laboratory of the US Department of Agriculture.
Research Laboratory of the USDepartment
of Agriculture), Peoria, Illinois (U.
SA) and recorded here under reference NRRL 5787. This strain was developed using a general method consisting of suspending a small amount of soil in sterile distilled water, diluting this suspension to various concentrations, and spreading a small amount of each dilution onto the surface of a Petri dish containing a nutrient agar medium. isolated by. After a few days of incubation at 25°C to allow microbial growth, colonies (which should preferably be isolated for further investigation) are removed and transferred onto slanted nutrient agar to provide a more abundant culture. Create. Streptomyces strain DS24367 belongs to the species Streptomyces hygroscopicus and its essential characteristics are defined by HD TRESNER and EJ BACKUS (Applied Microbiology 4 ,
243-250, 1956) and by SA WAKSMAN (Actinobacteria, The Williams & Wilkins Company, Baltimore,
1961, pp. 230-231) and eventually Streptomyces hygroscopicus strain DS24367.
displayed by the name. S. hygroscopicus DS24367 has three characteristics that correspond to the three features that defined the species S. hygroscopicus by HD Tresner and EJ Butkus and SA Watzmann: (a) its spore stalks are generally Terminating in a tight helix with two or three coiled coils, these spiral spore stalks are usually clustered along the filament to form a more or less elongated tuft. (b) When the spore-forming aerial microorganism reaches a favorable stage of development, it exhibits a dark gray coloration consistent with that exhibited by S. hygroscopicus species. (c) On certain media that allow good growth of aerial microorganisms, shiny black areas with a moist appearance appear on the sporulated surface over time. In the case of S. hygroscopicus DS24367, the formation of aerial microorganisms often occurs only slightly and may not even appear on certain media. When a gray spore-forming aerial fungal mass appears, the formation of dark areas, if it occurs, generally occurs rather diffusely, rather than over all of the surface of the fungal mass. Airborne bacteria are limited to locally distributed small points or small areas on the veneer. However, it is especially evident and observable on Pridham yeast agar, Pridham oatmeal and tomato paste agar, and glucose-asparagine agar. The strain of S. hygroscopicus mentioned for reference in Actinobacteria by S.A. Watzmann shows some minor differences compared to strain DS24367 on a few of the media whose morphological appearance has been described. However, the most important thing is that the former forms aerial microorganisms on nitrate agar and nutrient agar containing sucrose, whereas the DS24367 strain forms aerial microorganisms on these two media. consists of the fact that it does not form. Furthermore, S. hygroscopicus does not reduce nitrate on media containing sucrose as a carbon source, whereas DS24367 significantly reduces nitrate to nitrite under the same conditions. However, these small differences are not significant enough to distinguish between strain DS24367 and the species S. hygroscopicus, which otherwise has key characteristics. S. hygroscopicus DS24367 forms spore chains that generally terminate in tight helices with a limited number of turns, rarely 3 or 4 turns, but in some cases many turns. It is also possible to observe helices, or a few spore chains simply bent at their terminal parts without making a complete turn, or more or less loose and unwound helices. The spore-producing mechanism has a tuft-like structure, with a spiral spore stalk with a small number of branches inserted along the main filament (its length is quite considerable). The spores are oval to cylindrical and have dimensions of 0.8 to 1.0μ/0.6 to 0.8μ. Microscopic examination showed exactly the same composition of the sporulating organs on Bennett's agar and on Prydham oatmeal-tomato agar. Due to its method of sporulation, S. hygroscopicus DS24367 is a member of the Pridham classification Spira.
section. S. hygroscopicus DS24367 grows well at 25°C, less well at 37°C, and does not grow at 50°C. When cultured at 25°C, it has the following biochemical properties:

[Table] Streptomyces hygroscopicus
The culture characteristics of DS24367 are shown in the following table. Unless otherwise specified, these have reached a good developmental stage, i.e.
Characteristics of culture after approximately 2 to 3 weeks at 25°C. These characteristics were observed on nutrient agar and broth, which are commonly used to determine the morphological characteristics of Streptomyces, and the cultivation on agar medium was performed on agar slants. Some of the media used were ``actinobacteria'' [SA Waxman, 193-
197 pages, Chronica Botanica Company, Waltham,
Massachusetts, USA, 1950]. In this case, the media are designated by the letter W, followed by the number given to them in "Streptomyces".
References for other media or constituents therein are as follows: Reference A - "Bennet's Agar" - SA Waxman - Actinomycetes, Volume 2, Page 331 - No. 30 - The Williams and Wilkins Inc., Baltimore, 1961. Reference B - "Hitsky and Tresner Agar" -
TG Pridham and co Antibiotics
Annual, (Antibiotic Annual Report), 1956-1957, 950
page. Reference C-Formulation W-23, to which 2% agar was added. Reference D - "Yeast Extract Agar" - TG Pridham and co-workers, Annals of Antibiotics, 1956-1957, 950
page. Reference E - "Tomato paste oatmeal agar" -
TG Pridham and co-workers, Annals of Antibiotics, 1956-1957, p. 950. Reference F - "Melaninogenic medium" - SA Waxman - Actinobacteria, vol. 2, p. 333 - No. 42 - The Williams and Wilkins Company, Baltimore, 1961. Reference G-WE GRUNDY and co-workers, Antibiotics and Chem., 2 , 401,
1952. Reference H - "Inorganic Salt Starch Agar" - TG Pridham and co-workers, Annual Review of Antibiotics, 1956-1957,
951 pages. Reference I - Corresponds to formulation W-1, except that 15 g of glucose was used instead of 30 g of sucrose. Reference J - Corresponds to formulation W-1, except that 15 g of glycerin was used instead of 30 g of sucrose. Reference K - Corresponds to formulation W-18, except that 15 g of glucose was used instead of 30 g of sucrose. Reference L - Corresponds to formulation W-18, except that the sucrose was omitted and a piece of paper partially immersed in the liquid was used. Reference M - “Manual of Methods for pure Culture Research”
Study of Bacteria” - Society of American Bacteriologists
American Bacteriologists), Geneva, N.
Y., ₅₀ −18. Reference N - "Basic gelatin" - "Guide to pure bacterial culture research methods" - Society of American
Bacteriologist, Geneva, NY, ₅₀
Prepared according to instructions in -18. Reference P - Commercial skimmed milk powder, reconstituted according to manufacturer's instructions. Reference Q - Medium shown in the following literature for research into H ₂ S production, HD Tresner and F. DANGA - Journal of Bacteriology, 76 , 239 -
244, 1958.

【table】

【table】

【table】

[Table] The growth levels in the table above are classified in decreasing order as follows: very good, good,
Fairly good, moderate, very mild, poor, very poor or none. Streptomyces hygroscopicus
The ability of DS24367 to utilize various carbon and nitrogen sources for its development was demonstrated using the Pridham and Gottlieb method.
method) [J. of Bact., 56 , 107-114, 1948]. The degree of growth was determined on the basal medium as directed by the authors, but with glucose replaced by one of the various tested carbon sources, or (NH ₄ ) ₂ SO ₄ replaced with one of the various tested nitrogen sources. , observed after a suitable incubation period at 25°C. The results are shown in the table below.

【table】

[Table] 31559RP is obtained by culturing Streptomyces hygroscopicus DS24367 or its productive mutant (i.e., a mutant capable of producing 31559RP) aerobically in a medium, and 31559RP produced during the culture. It is produced by isolating it as it is or as a salt. The method for producing 31559RP and its salts was published in a patent application corresponding to the original application of the present application.
No. 50-26443. Streptomyces hygroscopicus
Cultivation of DS24367 can be carried out by either aerobic surface culture or deep culture, the latter being preferred for reasons of convenience. For this purpose, various types of equipment commonly used in the fermentation industry can be used. In particular, the following sequence can be adopted for carrying out the culture: Streptomyces hygroscopicus DS 24367 - stock ↓ Culture on agar ↓ Culture in shake flasks ↓ Inoculum culture in fermenter ↓ Production in fermenter Cultivation The fermentation medium must essentially contain an assimilable carbon source and an assimilable nitrogen source, mineral components, especially chlorides, and optionally growth factors, and these materials must contain well-defined products. or as a complex mixture, as is experienced with biological products of various origins. Suitable assimilable carbon sources include carbohydrates such as glucose, dextrins, starches or other carbohydrate substances such as sugar alcohols (including glycerin), and certain organic acids such as lactic acid or citric acid. . Some animal or vegetable oils, such as lard oil or soybean oil, can be used in place of, or added to, these various carbohydrate sources. Suitable assimilable nitrogen sources are extremely diverse.
These can be very simple chemicals, such as inorganic or organic ammonium salts, urea and certain amino acids. Nitrogen is also present in complex substances containing nitrogen, mainly in the form of proteins, such as casein, lactalbumin, gluten and their hydrolysates, soybean flour, peanut flour, fishmeal, meat extract, yeast extract, distillers solubles and corn. It can also be introduced as a steep. Of the mineral components added, some may have a buffering or neutralizing effect, such as alkali metal or alkaline earth metal phosphates, or calcium or magnesium carbonate. Others, such as alkal metal and alkaline earth metal chlorides and sulfates, provide the ionic balance necessary for the development of Streptomyces hygroscopicus DS24367 and for the production of 31559RP. Finally, some are more specific than Streptomyces hygroscopicus.
Acts as an activator of the metabolic reactions of DS24367 and these include zinc, cobalt, iron, copper and manganese salts. Suitable growth factors are natural products of the vitamin type, such as riboflavin, folic acid and pantothenic acid. The pH of the fermentation medium at the start of culture is 5.8 to 5.8.
7.8, and preferably 6.2 to 7.4. The optimum temperature for the fermentation process is between 25 and 30°C, although satisfactory production is achieved at temperatures between 23 and 33°C. Fermentation aeration can vary between fairly wide values. However, 0.3 to 3 air per 1 culture solution
It has been found that an airflow rate of /min is very suitable. Maximum yields of 31559RP are obtained after 2 to 8 days of culture, and this time essentially depends on the medium used. From these facts, Streptomyces hygroscopicus DS24367 for the production of 31559RP.
The general conditions for culturing can vary widely and can be adapted to specific requirements. 31559RP can be isolated from the product of culture by the following method. The medium is maintained at an acidic pH, generally between 3 and 6, preferably close to 5. The active substance retained in the mass is extracted from this mass with a suitable solvent, such as a lower alcohol, such as methanol, or a chlorinated solvent, such as methylene chloride. After treatment with a suitable alkali, 31559RP is extracted from the above solution after concentration under reduced pressure and optionally subsequent dilution with a non-solvent for the salt or a solvent in which the salt is sparingly soluble and storage in a cool room. For example, it is isolated in the form of the sodium salt by crystallization. 31559RP or its sodium salt can be purified by conventional methods, such as crystallization, chromatography on various adsorbents, or countercurrent partitioning. The following example is a reference example explaining the manufacturing method of 31559RP. Reference Example (A) Fermentation The following ingredients are introduced into the 170 fermenter. Peptone 1200 g Yeast extract 600 g Glucose monohydrate 1200 g Agar 240 g Tap water Sufficient amount to 110 Adjust the pH to 7.30 by adding 10N sodium hydroxide solution (70 c.c.). The medium is sterilized by bubbling 122°C water vapor into it for 40 minutes. After cooling, the volume of the culture solution becomes 120 due to condensation of water vapor during the sterilization process, and the pH
is 6.85. Next, this culture solution was mixed with Streptomyces hygroscopicus, which was grown in a stirred Erlenmeyer flask.
Inoculate with a culture of DS24367 (200c.c.). Culture
Grow for 25 hours at 27°C. Stir and vent with sterile air during this time. This makes it suitable for inoculating production cultures. The production culture is carried out in 800 fermenters introduced with the following substances: Distillers Soluble 2 Kg Bean Granules 10 Kg Glycerin 6 Kg Sodium Chloride 2 Kg Solution of cobalt chloride hexahydrate at a concentration of 20 g/kg
0.4 Amount of tap water to 365 Adjust the pH of the culture medium to 10N sodium hydroxide solution (200
Adjust to 7.50 by adding cc). The culture solution is then sterilized by passing 122°C steam through it for 40 minutes. After cooling, the volume of the culture medium decreases to 390% due to the condensation of water vapor during the sterilization process.
becomes. By adding a sterile aqueous solution (10) containing 4 kg of glucose monohydrate,
Make it. The pH of the medium is 6.65. Add this to 170 above.
Inoculate with inoculum culture (40) from the fermenter. Cultures were grown for 113 hours at 27°C, during which time 205
It is stirred by a turbine rotating at rev/min and aerated with sterile air (with a volume of 20 m ³ /h). At the end of this operation, the pH of the culture is 7.65 and the volume is 380. (B) Extraction The medium (400) obtained as above was diluted with 6N
Bring to pH 5 by adding hydrochloric acid solution (2.8) and then stir for 30 minutes. After adding the supernatant (20Kg), the medium is filtered through a filter press, and the mass is washed on the filter with water (100) (its pH is adjusted to 5 with 6N hydrochloric acid).The solution and washing solution are removed. do. Grind the mass in methanol (300),
The resulting mixture is adjusted to pH 7 by addition of 6N sodium hydroxide solution (200 c.c.) and stirred for 30 minutes. Pass the mixture through a filter press and wash the mass with methanol (80%) on the filter.
Wash with Combine the liquid and washing liquid and evaporate under reduced pressure (5 to 10
mmHg). The concentrate is adjusted to pH 3 by the addition of 6N hydrochloric acid (290 c.c.) and then stirred for 10 minutes in the presence of methylene chloride (30).
Separate the two phases. The aqueous phase is extracted in this manner twice more with methylene chloride (30 times each time). Combine the three methylene chloride extracts and add water (9
) and during this time acidify the aqueous phase to PH3 (20 c.c. of 6N hydrochloric acid). The organic phase is isolated, washed with water (18) and sodium hydroxide solution is added to the aqueous phase until a pH of 9.5 is obtained (20 c.c. of 6N sodium hydroxide solution). The methylene chloride phase was removed under reduced pressure (5
to 10 mmHg). (C) Purification Slowly pour the methylene chloride concentrate obtained above into hexane (10) at room temperature. The solution is filtered and inert insoluble matter (63 g) is removed in this way. Liquid under reduced pressure (5 to 10 mm
Hg) to a volume of approximately 0.5. This solution was stirred for 2 hours at a temperature of 0 °C, then +
Leave at 4°C for 24 hours. The crystals obtained are isolated by centrifugation of the solution, then washed with hexane (250 c.c.) at +4° C. and dried under reduced pressure. The sodium salt of 31559RP (103 g) is thus isolated. (D) Recrystallization Dissolve the sodium salt of 31559RP (57 g) obtained above in acetone (1.2) and stir for 15 minutes. Add decolorizing charcoal (2.5g) washed with hydrochloric acid,
Stir the mixture again for 30 minutes and finally separate the decolorizing charcoal. Add distilled water (750c.c.) to the slowly stirred solution. The sodium salt of 31559RP slowly crystallizes at a temperature of 0°C. Separate the crystals, wash with a mixture of acetone and water (50−50 by volume) (150 c.c.) and store at 35 °C under reduced pressure.
Dry with. Sodium salt of 31559RP (35g)
is obtained. Concentrating the mother liquor to half its original volume and then cooling it to 4° C. yields an additional 13.2 g of the sodium salt of 31559RP.

[Brief explanation of the drawing]

The drawings depict antibiotics used in the compositions of the invention.
This is the infrared absorption spectrum of 31559RP.

Claims (1)

[Claims] 1 31559RP (as an acid or as a salt with a metal or with a nitrogen-containing base) contains from 0.0001 to 99.9% by weight and a substance that can be consumed by non-human animals; (a) Its elemental composition is approximately C=63.2%, H=
8.8%, O = 24.3%, Na = 2.9%; (b) its neutral equivalent (determined by titration of the sodium salt in acetic acid with perchloric acid) is 836; (c) protons and carbon NMR spectrum of ¹³ C shows the presence of 4 methoxy groups in the molecule; (d) White microcrystalline powder, virtually insoluble in water and slightly soluble in hydrocarbon solvents such as hexane. (e) It is soluble in alcohols such as methanol, ketones such as acetone, dimethylformamide and ethyl acetate, and readily soluble in chlorinated solvents such as methylene chloride and chloroform; (e) It has a melting point of 220°C ( decomposition), and its optical rotation (C=1.050, in methanol) is [α] ²⁰ _D = +51.2
゜±1゜, [α] ²⁰ ₄₃₆ = +101゜±1.5゜, [α]
²⁰ ₃₆₅
= +167°±2°; (f) Its ultraviolet spectrum (0.950mg/
(g) Its infrared spectrum (measured in tablets in a mixture with potassium bromide) is as follows: [Table] [Table] and (h) In silica gel thin layer elevation chromatography using two of the following mixed solvent systems: (i) 31559RP in a mixed system of ethyl acetate, cyclohexane, water and butanol (volume ratio, 50-50-25-5); The sodium salt of 31559RP has Rf 0.5 and (ii) in a mixed system of methylene chloride and methanol (volume ratio, 94-5) the sodium salt of 31559RP has Rf 0.7; An anticoccidial composition for feeding to non-human animals, characterized by: