FI89368B - Polycyclic ether and the use thereof as a growth promoting agent in cattle and pig feed composition - Google Patents

Abstract

The invention relates to a novel polycyclic ether of the formula <IMAGE> and the physiologically acceptable cationic salts thereof. The compounds can be used in promoting growth in pigs and ruminants and/or to improve efficiency of feed utilization.

Description

1 89368

Polycyclic ether and its use in feed composition for cattle and pigs as a growth promoter

Divided by application 873326 5

The present invention relates to a novel polycyclic ether of the formula OCH ^ 10 eOH OH yV'0'xl> ^ CH3 CH3 i_CH Cn3

f / V— / \ —L .Joint CH

^ ^ OH H * H ^ £ 0 A »A

C00H CH3 ch3 h ch3 H H H oh and its physiologically acceptable cationic salts and the use of these compounds in the composition of cattle or pig feed-20 as growth-promoting and / or food-enhancing agent.

The above-mentioned polycyclic ether according to the invention belongs to the group of acidic polycyclic ethers, the compounds of which are biologically characterized in that they affect the cation transfer between cell membranes. This group of antibiotics includes well-known antibiotics such as monensin, nigericin, grisorxin, dianemycin, meduramycin, narasin, salinomycin, 1α-saloside, mutalomycin, ionomycin and leuceramycin. 30 This topic is discussed in Westley, "Polyether Antibiotics", Adv. Appl. Microbiol., 22, 177, 1977.

The ether antibiotics listed above act on gram-positive bacteria, fungi, and protozoa. In particular, these antibiotics show an effective coccidial effect. Therefore, they have been used with 2 39368 varying degrees of success in the treatment of various animal infections.

Increasing production capacity (increased growth rate and / or increased food utilization efficiency) in ruminants, such as cattle, and monogastric animals, such as pigs, is the desired goal in veterinary medicine. Of particular interest is the improved production capacity achieved by improving the efficiency of food utilization. The mechanism for utilizing the major nutrient portion 10 of ruminant feed is well known. Microorganisms in the rumen of an animal break down carbohydrates to make monosaccharides, and then convert these monosaccharides to pyruvate compounds. In microbiological processes, pyruvate is converted via metabolism to acetates, butyrates or propionates, known as volatile fatty acids. For a more detailed review, see publications: Leng, "Physiology of Digestion and Metabolism in the Ruminant", Phillipson et al., eds., Oriel Press, Newcastle-upon-Tyne, England, 20 1970, pp. 408-410.

The corresponding efficiency of volatile fatty acid utilization is discussed in McCullough, "Feed-stuffs," June 19, 1971, p. 19; Eskeland et al., J. An. Sci., 33 (1971) p. 282; and Church et al., "Digestive 25 Physiology and Nutrition of Ruminants," Vol. 2, 1971, pp. 622 and 625. Although acetates and butyrates are utilized, the efficiency of propionate utilization is higher. Thus, the beneficial compound causes the animals to produce a higher proportion of propionates than the carbohydrates, thereby increasing the efficiency of carbohydrate utilization.

EP-A-0169011 describes an acidic polycyclic ether antibiotic designated UK-58852 having the formula li 3 09368

αΛ * OCH

Y ^ ywm OCH

C »3 k Λ 5 O or 3 yXO ^^ CH3

CH 2 ^ 2 .OCH. 1 O CH

__. V ^ 3yY

\ ~ L · Λί'-Ο ^ '..... CH3 I OH H A H 0 | fNo ^ I ΙΟ * 4 ^ 1

10 cooh ch3 ch3 h ch3 H H H OR

wherein both R and R1 are hydrogen. It is prepared by aerobic immersion propagation in an aqueous nutrient solution containing the microorganism Actinomadura roseorufa Huang sp. Nov., ATCC 39697, isolated from a sample taken from Japanese soil. It has now been found that hydrolysis of UK-58852 under closely controlled conditions results in the cleavage of a single glucone ring in the molecule to give the aforementioned novel compound of formula I which is superior to the UK-58852 compound due to its lower toxicity. This novel compound of formula I of the invention is designated UK-61689.

UK-61689 and its cationic salts can be prepared by a process comprising controlled hydrolysis of UK-58852 or a cationic salt thereof.

The hydrolysis of UK-58852 to prepare a compound of formula I (UK-61689) is preferably performed using para-toluenesulfonic acid in an acetonitrile / aqueous solvent. The preferred acetonitrile / water composition contains 5 to 0.5% water. Para-toluenesulfonic acid is preferably used in 1.1 molar equivalents per molar equivalent of the sodium salt of UK-58852.

4 89368

Other useful solvent systems include acetonitrile alone or t-butanol / water or acetone / water. Alternative acids include mineral acids such as hydrochloric acid, acetic acid or a strongly acidic resin. In general, the reaction is carried out at room temperature and monitored by thin layer chromatography until the yield of the desired product is optimal. The reaction time is generally about one to three hours for a para-toluenesulfonic acid hydrolysis system in acetonitrile / water, but may vary depending on the acid and solvent system used. When the reaction is substantially complete, the reaction mixture is neutralized with excess sodium bicarbonate, concentrated and extracted with diethyl ether or dichloromethane and then purified using standard silica gel chromatography methods. The final product can then be recrystallized as the free acid or the appropriate cationic salt.

Alternatively, compound UK-61689 can be prepared from untreated fermentation extracts containing UK-58852. In this case, the methyl isobutyl ketone (MIBK) fermentation extract is concentrated and preferably dissolved in acetonitrile / water and treated with para-toluenesulfonic acid. The preferred aqueous acetonitrile composition contains 5% water, and the ratio of crude fermentation oil to para-toluenesulfonic acid is about 9: 1; these ratios are averages and may vary depending on the fermentation extract. Another solvent system that can be used is methanol / water, but the hydrolysis can also be performed by treating the crude MIBK extract with para-toluene-nisulfonic acid in the absence of added solvent. Alternative acids include mineral acids such as hydrochloric acid.

The value of animal feed is usually determined directly by feeding the animal. GB Patent Publication 1,197,826 details an in vitro rumen method which

II

With the help of 5 y 9 368, the changes in the feed caused by microorganisms are measured most easily and very accurately when evaluating animal feed. This method uses an apparatus in which the digestive processes of animals are performed and studied in vitro. Animal feed, rumen inoculum, and various growth promoters are fed into and removed from the laboratory unit under closely controlled conditions, and changes are closely and continuously monitored during the use of the micro-organisms.

The increase in propionic acid content of the rumen indicates that the growth promoters in the feed composition have produced the desired response in the overall performance of the ruminant. The change in propionic acid content is expressed as a percentage of the propionic acid content of the reference rumen found as a percentage of 15 to 15%. Long-term in vivo feeding studies are used to demonstrate a reliable correlation between increased propionic acid content in rumen fluid and improved feed efficiency.

20 Rumen fluid is collected from a cow that has been rumen and fed commercial fattening feed and hay. The rumen is immediately filtered through the cheese cloth and 10 ml of liquid is added to a 50 ml conical flask containing 400 mg of standard substrate 25 (68% maize starch + 17% cellulose + 15% extracted soy flour), 10 ml of pH 6 buffer. 8, as well as the test compound. Oxygen-free nitrogen gas is bubbled into the flasks for about two minutes and incubated in a shaking water bath at 39 ° C for about 16 hours. All experiments 30 are performed in triplicate.

After incubation, 5 ml of the sample is mixed with 1 ml of 25% metaphosphoric acid. After 10 minutes, 0.25 ml of formic acid is added and the mixture is centrifuged at 1500 rpm. For 10 minutes 35. The samples are then analyzed by gas-liquid chromatography according to the method of DH Kellog, J. Dairy Science, 52 (1969) p. 1690. Peak concentrations of acetic, propionic and butyric acid are determined from untreated and treated incubation flasks. samples obtained.

When tested by this in vitro method, UK-61689 at a ratio of 20 micrograms per milliliter increased propionic acid production by about 80% compared to propionic acid production in a control solution to which no compound UK-61689 had been added. Compounds that stimulate the production of PPH (rumen propionic acid) are known to improve food utilization in ruminants such as cattle and sheep and may additionally have a similar effect on monogastric animals such as pigs. Yh-15 diste UK-61689 can be administered to an animal by mixing it into the feed composition as either the free acid or a salt, e.g. the sodium or potassium salt, or a mixture thereof. Alternatively, the crude product containing UK-61689 or the dried fermentation broth can be mixed into the feed composition in the desired concentrations. Compound UK-61689 may also be administered in the desired dosages using a suitable sustained release device designed to release the active ingredient in standard amounts.

The present invention is illustrated by the following preambles.

Example 1

To a solution of 15.0 g (0.0147 mol) of pure UK-58,852 antibiotic (prepared as described in EP-A-169011) in acetonitrile / water 30 (95: 5) was added 3.07 g (0.0161 mol) of ) p-toluenesulfonic acid. The reaction was monitored by thin layer chromatography until the yield of the desired product was optimal (about three hours). The reaction mixture was treated with solid bicarbonate or an aqueous solution thereof in excess and concentrated to dryness in vacuo. The resulting solid was dissolved in diethyl ether and washed with saturated aqueous sodium bicarbonate, the bicarbonate washings were extracted with diethyl ether, and all ether layers were combined and washed successively with water and saturated sodium chloride. The ether solution was dried over anhydrous sodium sulfate and evaporated to dryness in vacuo. The resulting crude reaction product contained UK-61689 as the predominant product but with varying amounts of starting material, UK-58852, and other by-products. The crude product was purified by silica gel column chromatography, followed by recrystallization from isopropyl ether to give 4.2 g (32%) of UK-61689 as the sodium salt, m.p. 175-176 ° C; [α] D 25 = +19.3 (c = 0: 5, MeOH); C-13 NMR (CDCl 3): 179.15 ppm, 107.45, 103.18, 97.74, 96.96, 86.92, 84.60, 84.20, 82.27, 82.01, 80.88, 80.20, 79.86, 74.80, 74.55, 73.07, 70.06, 67.71, 66.89, 59.11, 56.81, 45.40, 39, 82, 38.93, 36.46, 33.81, 33.71, 33.54, 33.42, 33.13, 32.44, 20 32.26, 30.56, 27.58, 26.90 , 26.84, 26.11, 23.23, 18.40, 17.53, 16.9.9, 12.13, 11.04, 10.42.

Example 2 The crude fermentation extract (MIBK) (1 L) containing the antibiotic UK-58852 (approximately 25 g) was concentrated in vacuo to 680 g. A thick, dark oil was added to an acetonitrile / water mixture (95: 5) (5.6 L), and the resulting milky emulsion was treated with p-toluenesulfonic acid (78.2 g) in one portion. As the reaction progressed, the emulsion gradually separated into two separate layers, which were stirred vigorously for a total of 1.25 hours. The reaction mixture was poured into a separatory funnel, and the oily base layer was removed and suspended by the addition of acetonitrile / water (95: 5). The acetonitrile and water layers were combined and treated with saturated aqueous sodium bicarbonate (400 mL). The mixture was then concentrated to a thick syrup, dissolved in diethyl ether / ethyl acetate (3: 1) (2 L) and washed with saturated aqueous sodium bicarbonate. The organic layer was dried over sodium sulfate and concentrated to a thick brown oil (432 g). Purification was most easily performed using countercurrent extraction methods. Hexane / ethyl acetate (1: 1) was used as the non-polar phase and methanol / water (3: 2) was used as the polar aqueous phase. Evaporation of the organic extracts gave 10 semi-solids which were triturated to dryness in isopropyl ether to give UK-61689 as the sodium salt, 10 g, m.p. 169-170 ° C. This material corresponded to UK-61689 prepared by the method of Example 1 when comparing TLC and NMR results.

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Claims (2)

1. A polycyclic ether of the formula: OCH 2 O H OH ° ^> L \ / \ -L ,,,,,, CH3 OH H in H ° g ρο / Τί * cooh ch3 ch3 h ch3 HKH OH (I) or a physiologically acceptable cation salt thereof. Use of a polycyclic ether according to claim 1 or a physiologically acceptable cation salt thereof in a feed composition for cattle or pigs as a growth promoting and / or feed utilizing increasing substance. li