CN104846030A - Preparation method of moxidectin - Google Patents
Preparation method of moxidectin Download PDFInfo
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- CN104846030A CN104846030A CN201510229838.2A CN201510229838A CN104846030A CN 104846030 A CN104846030 A CN 104846030A CN 201510229838 A CN201510229838 A CN 201510229838A CN 104846030 A CN104846030 A CN 104846030A
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Abstract
The invention discloses a preparation method of moxidectin. The preparation method comprises the following steps: 1) inoculating a strain in a strain culture medium for culture to obtain spore suspension M1; 2) inoculating the spore suspension M1 in a seed culture medium for culture to obtain seed solution M2; 3) inoculating the seed solution M2 in a fermentation culture medium for culture to obtain primary extract solution M3; 4) performing protection reaction, oxidation reaction, deprotection reaction and oximation reaction to the primary extract solution M3 to obtain the moxidectin, wherein the strain is Streptomyces cyaneogriseus and/or Streptomyces hygroscopicus. By adopting the method, the total yield of the prepared moxidectin is enabled to be obviously higher and thus the effects of simpler and more convenient production manner and higher yield are realized.
Description
Technical field
The present invention relates to the manufacture field of moxidectin, particularly, relate to a kind of preparation method of moxidectin.
Background technology
Moxidectin is a kind of novel antiparasitic is by the macrolide antibiotics of the single component of streptomycete fermentation.It is widely used in the basic agricultural such as livestock industry, aquaculture as a kind of antiparasitic, and along with the development in the fields such as agricultural, the demand of international antiparasitic keeps sustainable growth always.This produce market annual requirement is about 10 tons at present, and market scale is more than 200,000,000 dollars.
The 1970's, China once synthesized medicines such as producing hexachloroparaxylene (HPX), amoscanate, bithionol, niclosamide, and the animal parasite disease that China is generally occurred obtains effectively preventing to the eighties.Since generation nineteen ninety, the sound development of development to China's livestock industry, aquaculture that China has carried out avermectin and Ivermectin HCL parasiticide new drug creates active effect.Anti-parasite medicine is Ivermectin HCL the most widely in the world at present, is secondly avermectin, doractin etc.But, through the application of more than ten years, the product such as avermectin, Ivermectin HCL creates resistance to some extent, and drug effect declines comparatively obvious, and selling price is also on a declining curve.
New Zealand one is about the global report of survey display of cattle and sheep parasite resistance, and older generation's antiparasitic drug resistance problems such as avermectin expand just rapidly, product curative effect continuous decrease.And the production technique of the moxidectin of routine is often comparatively complicated, be thus difficult to meet large-scale demand.
Therefore, provide a kind of mode of production easier, and the preparation method of the higher moxidectin of output is the problem that the present invention needs solution badly.
Summary of the invention
For above-mentioned prior art, the object of the invention is to overcome parasite in prior art, for conventional antiparasitic, there is higher resistance, the property of medicine is obviously declined, and new antiparasitic is often produced comparatively complicated, output is lower, and cost costly, cannot meet the problem of extensive demand, thus provide a kind of mode of production easier, and the preparation method of the higher moxidectin of output.
To achieve these goals, the invention provides a kind of preparation method of moxidectin, wherein, described preparation method comprises:
1) strain inoculation is cultivated in bacterium culture medium, obtain spore suspension M1;
2) described spore suspension M1 is inoculated in seed culture medium cultivates, obtained seed liquor M2;
3) described seed liquor M2 is inoculated in fermention medium cultivates, obtain just extract M3;
4) by first extract M3 obtained moxidectin after upper protective reaction, oxidizing reaction, deprotection reaction and oximation reaction; Wherein,
Described bacterial classification is cyaneogriseus streptomyces (Streptomyces cyaneogriseus) and/or streptomyces hygroscopicus (Streptomyces hygroscopicus).
Pass through technique scheme, the present invention is by being inoculated in obtained spore suspension M1 on bacterium culture medium by cyaneogriseus streptomyces (Streptomycescyaneogriseus) and/or streptomyces hygroscopicus (Streptomyces hygroscopicus), then spore suspension M1 is inoculated in seed culture medium and cultivates, obtained seed liquor M2, again seed liquor M2 is inoculated in fermention medium and carries out fermentation culture, obtain just extract M3, finally by first extract M3 through upper protective reaction, oxidizing reaction, moxidectin is obtained after deprotection reaction and oximation reaction, thus make fermenting process comparatively simple by the way, and the moxidectin output obtained is higher, and then it is easier to achieve the mode of production, and the effect that output is higher.
Other features and advantages of the present invention are described in detail in embodiment part subsequently.
Embodiment
Below the specific embodiment of the present invention is described in detail.Should be understood that, embodiment described herein, only for instruction and explanation of the present invention, is not limited to the present invention.
The invention provides a kind of preparation method of moxidectin, wherein, described preparation method comprises:
1) strain inoculation is cultivated in bacterium culture medium, obtain spore suspension M1;
2) described spore suspension M1 is inoculated in seed culture medium cultivates, obtained seed liquor M2;
3) described seed liquor M2 is inoculated in fermention medium cultivates, obtain just extract M3;
4) by first extract M3 obtained moxidectin after upper protective reaction, oxidizing reaction, deprotection reaction and oximation reaction; Wherein,
Described bacterial classification is cyaneogriseus streptomyces (Streptomyces cyaneogriseus) and/or streptomyces hygroscopicus (Streptomyces hygroscopicus).
Above-mentioned design is by being inoculated in obtained spore suspension M1 on bacterium culture medium by cyaneogriseus streptomyces (Streptomyces cyaneogriseus) and/or streptomyces hygroscopicus (Streptomyces hygroscopicus), then spore suspension M1 is inoculated in seed culture medium and cultivates, obtained seed liquor M2, again seed liquor M2 is inoculated in fermention medium and carries out fermentation culture, obtain just extract M3, finally by first extract M3 through upper protective reaction, oxidizing reaction, moxidectin is obtained after deprotection reaction and oximation reaction, thus make fermenting process comparatively simple by the way, and the moxidectin output obtained is higher, and then it is easier to achieve the mode of production, and the effect that output is higher.Described upper protective reaction, oxidizing reaction, deprotection reaction and oximation reaction can operate according to this area ordinary operation mode, seldom repeat at this.
Bacterium culture medium described above, seed culture medium and fermention medium can be prepared in a conventional manner, such as in the present invention, described bacterium culture medium can comprise yeast powder, glucose, Semen Maydis powder, sodium-chlor, Sodium orthomolybdate, sodium-acetate, potassium primary phosphate and dipotassium hydrogen phosphate, described seed culture medium can comprise glucose, maltodextrin, starch, yeast extract paste and soybean cake powder, and described fermention medium can comprise glucose, lactose, cottonseed muffin, soya-bean oil, fermentation defoaming agent, calcium carbonate and magnesium sulfate.Certainly, the substratum that other this area routines use also can use at this, and the composition between each raw material can be configured according to practical situation, seldom repeats at this.
Step 1) in cultivation can cultivate according to this area cellar culture mode, incubation time and culture temperature can regulate according to actual needs, such as, one of the present invention preferred embodiment in, in order to make spawn culture better effects if, the spore suspension miospore content obtained is higher, step 1) in incubation time can be chosen as 216-264h further, culture temperature is chosen as 25-30 DEG C.
Similarly, of the present invention another preferred embodiment in, step 2) in incubation time be 20-30h, culture temperature is 25-30 DEG C.
In order to make seed liquor M2 ferment better in the fermentation medium, one of the present invention preferred embodiment in, step 3) in can also comprise that seed liquor M2 is placed in temperature is be inoculated in fermention medium after cultivating 45-65h under the condition of 25-30 DEG C.
Similarly, one of the present invention more preferred embodiment in, step 3) in incubation time be 200-300h, culture temperature is 25-30 DEG C.
In order to make the output of the moxidectin obtained higher, one of the present invention preferred embodiment in, step 4) can also comprise by first extract M3 filtration, wash-out, extraction and concentrated after carry out upper protective reaction again.
The filter operation mode that described filter operation can use according to this area routine operates, such as, one of the present invention preferred embodiment in, in order to make filter effect better, described filtration procedure can comprise first extract M3 through Perlite filter.
Similarly, of the present invention another preferred embodiment in, described elution process can comprise by filter after first extract M3 carry out wash-out through resin.
Described concentration process can operate according to the condensing mode of this area routine, such as, can be the mode such as air distillation, certainly, in order to make concentrated effect better, one of the present invention preferred embodiment in, described concentration process can be concentrating under reduced pressure.Such as, can be underpressure distillation.Other modes that can realize also can use at this, seldom repeat.
The mode that described oximation reaction can adopt according to this area routine operates, certainly, one of the present invention preferred embodiment in, in order to output after making oximation reaction is higher, described oximation reaction reacts 15-30h under can being defined as the environment being placed in 20-25 DEG C.
Similarly, in order to make the output of the moxidectin obtained higher, of the present invention another preferred embodiment in, can also concentrating under reduced pressure be comprised after described oximation reaction.
In order to avoid introducing new bacterium in operation, make product occur deviation, one of the present invention preferred embodiment in, described bacterium culture medium, described seed culture medium and described fermention medium can also comprise before use and carry out sterilization operation.The mode that described sterilization operation adopts according to this area routine operates, and does not illustrate one by one at this.
Below will be described the present invention by embodiment.In following examples, described cyaneogriseus streptomyces is conventional commercial product, described yeast powder, glucose, Semen Maydis powder, sodium-chlor, Sodium orthomolybdate, sodium-acetate, potassium primary phosphate and, dipotassium hydrogen phosphate, maltodextrin, starch, yeast extract paste, soybean cake powder, lactose, cottonseed muffin, soya-bean oil, fermentation defoaming agent, calcium carbonate, magnesium sulfate, perlite and resin be conventional commercial product.Described bacterium culture medium is prepared according to the preparation method of preparation example 1; Described seed culture medium is prepared according to the preparation method of preparation example 2, and described fermention medium is prepared according to the preparation method of preparation example 3.
Preparation example 1
1000ml is settled to after 3g yeast powder, 10g glucose, 3g Semen Maydis powder, 0.5g sodium-chlor, 0.05g Sodium orthomolybdate, 0.05g sodium-acetate, 0.05g potassium primary phosphate and 0.05g dipotassium hydrogen phosphate being dissolved in water, and be placed on the pressure kettle mesohigh sterilizing 20min that temperature is 121 DEG C, obtained bacterium culture medium.
Preparation example 2
Be settled to 1000ml after 5g glucose, 20g maltodextrin, 1g starch, 10g yeast extract paste and 2g soybean cake powder being dissolved in water, and be placed on the pressure kettle mesohigh sterilizing 20min that temperature is 121 DEG C, obtained seed culture medium.
Preparation example 3
Be settled to 1000ml after 10g glucose, 25g lactose, 25g cottonseed muffin, 1g soya-bean oil, 0.5g fermentation defoaming agent, 3g calcium carbonate and 1g magnesium sulfate being dissolved in water, and be placed on the pressure kettle mesohigh sterilizing 20min that temperature is 121 DEG C, obtained fermention medium.
Embodiment 1
1) 1g strain inoculation being placed in 15ml bacterium culture medium temperature is cultivate 216h under the condition of 25 DEG C, obtains spore suspension M1;
2) described spore suspension M1 is inoculated in 30ml seed culture medium to be placed in temperature be cultivate 20h under the condition of 25 DEG C to cultivate, obtained seed liquor M2;
3) described seed liquor M2 being placed in temperature is that to be inoculated in 20ml fermention medium after cultivating 45h under the condition of 25 DEG C and to be placed in temperature be cultivate 200h under the environment of 25 DEG C, obtains just extract M3;
4) by first extract M3 after Perlite filter through resin elution; then carry out again carrying out concentrating under reduced pressure again after oximation reaction 15h under the condition extracting and carry out upper protective reaction, oxidizing reaction, deprotection reaction after concentrating under reduced pressure and be placed in 20 DEG C, obtain moxidectin A1.(total recovery of moxidectin is 40.6%)
Embodiment 2
1) 1g strain inoculation being placed in 30ml bacterium culture medium temperature is cultivate 264h under the condition of 30 DEG C, obtains spore suspension M1;
2) described spore suspension M1 is inoculated in 80ml seed culture medium to be placed in temperature be cultivate 30h under the condition of 30 DEG C to cultivate, obtained seed liquor M2;
3) described seed liquor M2 being placed in temperature is that to be inoculated in 50ml fermention medium after cultivating 65h under the condition of 30 DEG C and to be placed in temperature be cultivate 300h under the environment of 30 DEG C, obtains just extract M3;
4) by first extract M3 after Perlite filter through resin elution; then carry out again carrying out concentrating under reduced pressure again after oximation reaction 30h under the condition extracting and carry out upper protective reaction, oxidizing reaction, deprotection reaction after concentrating under reduced pressure and be placed in 25 DEG C, obtain moxidectin A2.(total recovery of moxidectin is 42.5%)
Embodiment 3
1) 1g strain inoculation being placed in 25ml bacterium culture medium temperature is cultivate 240h under the condition of 28 DEG C, obtains spore suspension M1;
2) described spore suspension M1 is inoculated in 50ml seed culture medium to be placed in temperature be cultivate 25h under the condition of 28 DEG C to cultivate, obtained seed liquor M2;
3) described seed liquor M2 being placed in temperature is that to be inoculated in 35ml fermention medium after cultivating 55h under the condition of 28 DEG C and to be placed in temperature be cultivate 240h under the environment of 28 DEG C, obtains just extract M3;
4) by first extract M3 after Perlite filter through resin elution; then carry out again carrying out concentrating under reduced pressure again after oximation reaction 23h under the condition extracting and carry out upper protective reaction, oxidizing reaction, deprotection reaction after concentrating under reduced pressure and be placed in 22 DEG C, obtain moxidectin A3.(total recovery of moxidectin is 41.7%)
Embodiment 4
Be prepared according to the preparation method of embodiment 1, unlike, described step 4) in do not comprise filtration, wash-out, extraction and concentrated, obtained moxidectin A4.(total recovery of moxidectin is 35.2%)
Embodiment 5
Be prepared according to the preparation method of embodiment 2, unlike, do not carry out concentrating under reduced pressure after oximation reaction, obtained moxidectin A5.(total recovery of moxidectin is 36.1%)
Comparative example 1
Be prepared according to the preparation method of embodiment 3, unlike, without step 2), obtained moxidectin D1.(total recovery of moxidectin is 12.5%)
Comparative example 2
Be prepared according to the preparation method of embodiment 3, unlike, without step 3), obtained moxidectin D1.(total recovery of moxidectin is 11.0%)
Can be found out by above-mentioned, the total recovery of the moxidectin that preparation method within the scope of the present invention obtains is obviously higher, but the total recovery of the moxidectin obtained outward in the scope of the invention then reduces greatly, and the total recovery of moxidectin that the preparation method in preferable range of the present invention obtains is then higher, thus can find out, achieve fermenting process by the way comparatively simple, and the moxidectin output obtained is higher, and then it is easier to achieve the mode of production, and the effect that output is higher.
More than describe the preferred embodiment of the present invention in detail; but the present invention is not limited to the detail in above-mentioned embodiment, within the scope of technical conceive of the present invention; can carry out multiple simple variant to technical scheme of the present invention, these simple variant all belong to protection scope of the present invention.
It should be noted that in addition, each concrete technical characteristic described in above-mentioned embodiment, in reconcilable situation, can be combined by any suitable mode, in order to avoid unnecessary repetition, the present invention illustrates no longer separately to various possible array mode.
In addition, also can carry out arbitrary combination between various different embodiment of the present invention, as long as it is without prejudice to thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (10)
1. a preparation method for moxidectin, is characterized in that, described preparation method comprises:
1) strain inoculation is cultivated in bacterium culture medium, obtain spore suspension M1;
2) described spore suspension M1 is inoculated in seed culture medium cultivates, obtained seed liquor M2;
3) described seed liquor M2 is inoculated in fermention medium cultivates, obtain just extract M3;
4) by first extract M3 obtained moxidectin after upper protective reaction, oxidizing reaction, deprotection reaction and oximation reaction; Wherein,
Described bacterial classification is cyaneogriseus streptomyces (Streptomyces cyaneogriseus) and/or streptomyces hygroscopicus (Streptomyces hygroscopicus).
2. preparation method according to claim 1, wherein, step 1) in incubation time be 216-264h, culture temperature is 25-30 DEG C.
3. preparation method according to claim 1, wherein, step 2) in incubation time be 20-30h, culture temperature is 25-30 DEG C.
4. preparation method according to claim 1, wherein, step 3) in also comprise that seed liquor M2 is placed in temperature is be inoculated in fermention medium after cultivating 45-65h under the condition of 25-30 DEG C.
5. preparation method according to claim 1, wherein, step 3) in incubation time be 200-300h, culture temperature is 25-30 DEG C.
6. preparation method according to claim 1, wherein, step 4) also comprise first extract M3 filtration, wash-out, extraction and carry out upper protective reaction again after concentrating.
7. preparation method according to claim 6, wherein, described filtration procedure comprises first extract M3 through Perlite filter;
Described elution process comprises the first extract M3 after by filtration and carries out wash-out through resin.
8. preparation method according to claim 6, wherein, described concentration process is concentrating under reduced pressure.
9. preparation method according to claim 1, wherein, described oximation reaction is react 15-30h under the environment being placed in 20-25 DEG C;
Preferably, also concentrating under reduced pressure is comprised after described oximation reaction.
10. preparation method according to claim 1, wherein, described bacterium culture medium, described seed culture medium and described fermention medium also comprise before use and carry out sterilization operation;
Preferably, described bacterium culture medium at least comprises yeast powder, glucose, Semen Maydis powder, sodium-chlor, Sodium orthomolybdate, sodium-acetate, potassium primary phosphate and dipotassium hydrogen phosphate;
Described seed culture medium at least comprises glucose, maltodextrin, starch, yeast extract paste and soybean cake powder;
Described fermention medium at least comprises glucose, lactose, cottonseed muffin, soya-bean oil, fermentation defoaming agent, calcium carbonate and magnesium sulfate.
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Cited By (1)
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| CN106075458A (en) * | 2016-06-08 | 2016-11-09 | 芜湖福民生物药业有限公司 | Moxidectin preparation and preparation method thereof |
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| AU2006100662A4 (en) * | 2006-05-08 | 2006-09-07 | Wyeth | Intermediates and processes using them |
| CN101372492A (en) * | 2007-06-29 | 2009-02-25 | 浙江海正药业股份有限公司 | Method for preparing high-purity moxidectin |
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Application publication date: 20150819 |