CN105367494A - Sinomenine derivative and preparing method thereof - Google Patents
Sinomenine derivative and preparing method thereof Download PDFInfo
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- CN105367494A CN105367494A CN201410442733.0A CN201410442733A CN105367494A CN 105367494 A CN105367494 A CN 105367494A CN 201410442733 A CN201410442733 A CN 201410442733A CN 105367494 A CN105367494 A CN 105367494A
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- Prior art keywords
- tuduranine
- sinomenine
- fermentation
- fermented liquid
- sinomenine derivate
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- BJJRDCBDIOEIKD-UHFFFAOYSA-N COc1ccc(CC2NCCC3(CC4=O)C2C=C4OC)c3c1O Chemical compound COc1ccc(CC2NCCC3(CC4=O)C2C=C4OC)c3c1O BJJRDCBDIOEIKD-UHFFFAOYSA-N 0.000 description 1
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Abstract
The invention discloses a sinomenine derivative and a preparing method thereof. The preparing method for the sinomenine derivative comprises the steps that absidia ramosa is inoculated into a culture medium to carry out liquid state fermentation; after absidia ramosa is fermented for 4-5 days, sinomenine is added into fermentation liquor to carry out bioconversion; after bioconversion is completed, solid-liquid separation is carried out on the fermentation liquor, and separation and purification are carried out. The microbial conversion method has the advantages of being high in yield, safe and reliable in process, low in environmental pollution, easy and convenient to operate, and suitable for large-scale industrialized production.
Description
Technical field
The present invention relates to biological technical field, relate to a kind of Sinomenine derivate and preparation method thereof particularly.
Background technology
Tuduranine (Sinomenine) extracts from the dry root of sinomenium acutum (SinomeniumAcutumRehderettWilson) isoquinoline alkaloid obtained.Modern pharmacology experiment proves, its have dispel rheumatism, the effect of promoting blood circulation and removing obstruction in channels, anti-inflammatory analgetic, inducing diuresis to remove edema, multiplexly at present make antiheumatic special efficacy bulk drug.The long-term clinical test of pesticide effectiveness proves that it has the multiple physiologically actives such as anti-inflammatory, immunity, analgesia, step-down, anti-arrhythmia.Be used for the treatment of in addition in recent years chronic nephritis, anti-oxidant, antitumor, drug rehabilitation report.Tuduranine pharmaceutical preparation is used for the treatment of rheumatoid arthritis and irregular pulse clinically, and obtains good efficacy.The result of sinomenine gels preparation in animal Transdermal absorption and test of pesticide effectiveness research shows that tuduranine has substantial connection to the synthesis of Topical Prostaglandin (GP) and release.Tuduranine also has the effect of Selective depression cyclooxygenase 2 (COX-2), and this may be that it has one of main mechanism that anti-inflammatory and analgesic effect is strong and side effect is relatively little.Research shows that tuduranine plays anti-inflammatory, anti rheumatism action mainly through regulating cellular inflammation medium and controlling cytokine synthesis, but, tuduranine clinically its dosage is bigger than normal, there is obvious histamine release effect, thus cause supersensitivity side effect, therefore, carry out modifying to the structure of tuduranine and optimize, significant with the Sinomenine derivate of new generation seeking high-efficiency low-toxicity.
Absidia rasmosa (Absidiaramosa), belong to absidia (Absidia), its mycelium is similar to head mold, have creeper and rhizoid, but sporangiophore is in the middle of creeper, not with rhizoid to life.Most 2 ~ 5 clusters of sporangiophore, normal in colyliform or irregular branch branch.Sporocyst base portion has obvious apophysis, columella taper or semisphere.Zygospore raw on creeper, and this belongs to bacterium and is distributed widely in soil, distiller's yeast and various ight soil, is the contaminated bacteria that alcoholic is produced, and some can be pathogenic bacterias of people, animal in the kind of 37 degrees Celsius of growths.How several research at present for the conversion of absidia rasmosa strain microorganism is in steroide field, are the critical strain transforming steroide.Also there is no the report of absidia rasmosa in microbial transformation at present.
Summary of the invention
The object of this invention is to provide a kind of Sinomenine derivate and preparation method thereof.
A first aspect of the present invention provides a kind of Sinomenine derivate, and its structure is as shown in the formula shown in I:
Another aspect provides a kind of preparation method of Sinomenine derivate, comprise step:
(1) thalline fermentation
Absidia rasmosa (Absidiaramosa) is inoculated in substratum and carries out liquid state fermentation;
(2) bio-transformation
Fermentation absidia rasmosa, after 4-5 days, adds tuduranine, carries out bio-transformation in fermented liquid;
(3) separation and purification
After bio-transformation completes, fermented liquid solid-liquid separation, carries out separation and purification.
Preferably, the absidia rasmosa in step (1) is CGMCC3.3390 bacterial strain.
Preferably, in the fermentation culture process in step (1), in substratum, Carbon and nitrogen sources is respectively sucrose and soyflour.
Preferably, in step (1), in fermention medium, sucrose content is 2-3% (w/v), and soyflour content is 1-2% (w/v).
Preferably, in step (2), the addition of tuduranine is 0.5-3.0g/L fermented liquid.
More preferably, in step (2), the addition of tuduranine is 1-2g/L fermented liquid.
Preferably, in step (2), the pH in biotransformation is 6.5-8.5, and more preferably pH is 7.5.
Preferably, in step (2), invert point is 25-37 DEG C, is more preferably 27-30 DEG C.
Preferably, in step (3), macroporous adsorbent resin is adopted to carry out separation and purification.
Another aspect provides the purposes of Sinomenine derivate, for the preparation of the medicine of anti-inflammatory, antiviral, antitumor, bacterial-infection resisting, wherein, described Sinomenine derivate, its structure is as shown in the formula shown in I:
We are to tuduranine and the derivative research continued without having carried out going deep into thereof, sinomenium acutum extract is added in absidia rasmosa fermented liquid and cultivates, be surprised to find that in fermented liquid and occurred a kind of new material, detect through LC-MS, molecular weight lacks a methyl compared with tuduranine, warp
1hNMR identifies that its structure is such as formula shown in I.Although the structure of this compound has been reported, be and be prepared by the method for chemosynthesis, it is high that microbial conversion process of the present invention has yield, process safety is reliable, low in the pollution of the environment, the advantage such as simple to operation, is applicable to carrying out large-scale industrial production.
Should be understood that within the scope of the present invention, above-mentioned each technical characteristic of the present invention and can combining mutually between specifically described each technical characteristic in below (eg embodiment), thus form new or preferred technical scheme.As space is limited, tiredly no longer one by one to state at this.
Accompanying drawing explanation
Fig. 1 shows the collection of illustrative plates that Sinomenine derivate of the present invention and standard substance HPLC detect jointly.
Embodiment
Below in conjunction with specific embodiment, set forth the present invention further.Should be understood that these embodiments are only not used in for illustration of the present invention to limit the scope of the invention.The experimental technique of unreceipted actual conditions in the following example, the usually conveniently conditioned disjunction condition of advising according to manufacturer.
Unless otherwise defined, all specialties used in literary composition and scientific words and one skilled in the art the same meaning be familiar with.In addition, any method similar or impartial to described content and material all can be applicable in the inventive method.The use that better implementation method described in literary composition and material only present a demonstration.
Embodiment 1
Bacterial strain: absidia rasmosa CGMCC3.3390 bacterial strain is purchased from Beijing North Na Kaichuan Bioisystech Co., Ltd.
Slant medium: solid potato culture medium (PDA), potato 20% (w/v), glucose 2.5% (w/v), agar 2% (w/v), pH7.0, cultivate 4-5 days for 25 DEG C.
Seed culture medium: sucrose 2.5% (w/v), yeast powder 0.3% (w/v), peptone 1.0% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.05% (w/v), pH=6.8, cultivate 1-2 days for 25 DEG C.
Fermention medium: sucrose 3% (w/v), soyflour 1.5% (w/v), KH
2pO
40.15% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.2% (w/v), pH=6.8,28 DEG C of cultivations.
Fermentation culture is after 4 days, add tuduranine 2g/L (purchased from Pa Nier bio tech ltd, Shaanxi) in the fermentation medium, and regulate fermented liquid pH to be 7.5, sample after bio-transformation 6h, with methyl alcohol 1: 1 volume mixture, after soaking 1h, centrifugal, supernatant HPLC detection by quantitative, calculating molar yield is 84%.
The condition of HPLC detection method is as follows:
Moving phase:
Phosphate buffered saline buffer (dipotassium hydrogen phosphate 0.01mol/L, adds 1% triethylamine, and phosphoric acid adjusts pH3.0): acetonitrile=80: 20;
Column type number: C18,150 × 4.60mm, 3 μm;
Column temperature: 40 DEG C;
Determined wavelength: 265nm.
Molar yield %=Sinomenine derivate mole number/substrate tuduranine mole number × 100%.
After conversion completes, isopyknic 95% ethanol is added in conversion fluid, soak centrifuging in 2 hours,, use HP20 macroporous adsorbent resin to carry out purifying, products obtained therefrom and standard substance (purchased from Yan Yu bio tech ltd, Shanghai) to mix by weight at 1: 1, HPLC is sample detection altogether, as shown in Figure 1, the HPLC peak shape of standard substance and products obtained therefrom of the present invention overlaps the HPLC collection of illustrative plates detected completely, proves that products obtained therefrom of the present invention is consistent with standard substance.LC-MS,
1hNMR verifies that structure of title compound formula is:
Embodiment 2
Absidia rasmosa strain culturing
Slant medium: solid potato culture medium (PDA), potato 20% (w/v), glucose 2.5% (w/v), agar 2% (w/v), pH7.0, cultivate 4-5 days for 25 DEG C.
Seed culture medium: sucrose 2.5% (w/v), yeast powder 0.3% (w/v), peptone 1.0% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.05% (w/v), pH=6.8, cultivate 1-2 days for 25 DEG C.
Fermention medium: sucrose 2.0% (w/v), soyflour 2.0% (w/v), KH
2pO
40.15% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.2% (w/v), pH=6.8,28 DEG C of cultivations.
Fermentation culture is after 5 days, add tuduranine 3%g/L (purchased from Pa Nier bio tech ltd, Shaanxi) in the fermentation medium, and regulate fermented liquid pH to be 8.0, sample after bio-transformation 6h, with methyl alcohol 1: 1 volume mixture, after soaking 1h, centrifugal, supernatant HPLC detection by quantitative, molar yield is 72%.
Embodiment 3
Absidia rasmosa strain culturing
Slant medium: solid potato culture medium (PDA), potato 20% (w/v), glucose 2.5% (w/v), agar 2% (w/v), pH7.0, cultivate 4-5 days for 25 DEG C.
Seed culture medium: sucrose 2.5% (w/v), yeast powder 0.3% (w/v), peptone 1.0% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.05% (w/v), pH=6.8, cultivate 1-2 days for 25 DEG C.
Fermention medium: sucrose 3.0% (w/v), soyflour 1.0% (w/v), KH
2pO
40.15% (w/v), K
2hPO
40.1% (w/v), MgSO
47H
2o0.2% (w/v), pH=6.8,28 DEG C of cultivations.
Fermentation culture is after 5 days, add tuduranine 1%g/L (purchased from Pa Nier bio tech ltd, Shaanxi) in the fermentation medium, and regulate fermented liquid pH to be 6.5, sample after bio-transformation 6h, with methyl alcohol 1: 1 volume mixture, after soaking 1h, centrifugal, supernatant HPLC detection by quantitative, molar yield is 88%.
The all documents mentioned in the present invention are quoted as a reference all in this application, are just quoted separately as a reference as each section of document.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (10)
1. a Sinomenine derivate, is characterized in that, described Sinomenine derivate structure is as shown in the formula shown in I:
2. a preparation method for Sinomenine derivate, is characterized in that, comprises step:
(1) thalline fermentation
Absidia rasmosa (Absidiaramosa) is inoculated in substratum and carries out liquid state fermentation;
(2) bio-transformation
Fermentation absidia rasmosa, after 4-5 days, adds tuduranine, carries out bio-transformation in fermented liquid;
(3) separation and purification
After bio-transformation completes, fermented liquid solid-liquid separation, carries out separation and purification.
3. method as claimed in claim 2, it is characterized in that, the absidia rasmosa in step (1) is CGMCC3.3390 bacterial strain.
4. method as claimed in claim 2, it is characterized in that, in the fermentation culture process in step (1), in substratum, Carbon and nitrogen sources is respectively sucrose and soyflour.
5. method as claimed in claim 4, is characterized in that, in step (1), in fermention medium, sucrose content is 2-3% (w/v), and soyflour content is 1-2% (w/v).
6. method as claimed in claim 2, is characterized in that, in step (2), the addition of tuduranine is 0.5-3.0g/L fermented liquid.
7. method as claimed in claim 2, is characterized in that, in step (2), the addition of tuduranine is 1-2g/L fermented liquid.
8. method as claimed in claim 2, it is characterized in that, in step (2), the pH in biotransformation is 6.5-8.5, and more preferably pH is 7.5.
9. method as claimed in claim 2, it is characterized in that, in step (2), invert point is 25-37 DEG C, is more preferably 27-30 DEG C.
10. a purposes for Sinomenine derivate, is characterized in that, for the preparation of the medicine of antigout, antitumor, bacterial-infection resisting, wherein, described Sinomenine derivate structure is as shown in the formula shown in I:
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410442733.0A CN105367494A (en) | 2014-08-27 | 2014-08-27 | Sinomenine derivative and preparing method thereof |
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| CN201410442733.0A CN105367494A (en) | 2014-08-27 | 2014-08-27 | Sinomenine derivative and preparing method thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107325097A (en) * | 2017-08-04 | 2017-11-07 | 北京师范大学 | Sinomenine derivate and preparation method and application |
| CN107337672A (en) * | 2017-08-04 | 2017-11-10 | 北京师范大学 | A kind of Sinomenine derivate and preparation method and application |
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2014
- 2014-08-27 CN CN201410442733.0A patent/CN105367494A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107325097A (en) * | 2017-08-04 | 2017-11-07 | 北京师范大学 | Sinomenine derivate and preparation method and application |
| CN107337672A (en) * | 2017-08-04 | 2017-11-10 | 北京师范大学 | A kind of Sinomenine derivate and preparation method and application |
| CN107325097B (en) * | 2017-08-04 | 2019-05-28 | 北京师范大学 | Sinomenine derivate and the preparation method and application thereof |
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Application publication date: 20160302 |
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