CN110013477A - A kind of new application of the secondary metabolites of Enteromorpha source fungi - Google Patents
A kind of new application of the secondary metabolites of Enteromorpha source fungi Download PDFInfo
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Abstract
The present invention provides a kind of new application of the secondary metabolites of Enteromorpha source fungi, belongs to microbial medicine technical field, is preparing the new application in alpha-glucosidase restrainer and/or the drug for treating diabetes；Above-mentioned secondary metabolites the preparation method comprises the following steps: being inoculated in Enteromorpha source fungi containing fermenting in TSA and enoxolone fermentation liquid, separate, purify to get targeted secondary metabolin.Secondary metabolites of the present invention are good to the inhibitory effect of alpha-glucosidase, slow down the generation and absorption of glucose, prevent postprandial hyperglycemia, to the preferable effect of diabetes tool is prevented and treated, are easy to be absorbed by organisms；The preparation method of secondary metabolites of the present invention can improve the yield of secondary metabolites, promote the proliferation activity of Enteromorpha source fungi, improve the yield of its secondary metabolite.
Description
Technical field
The invention belongs to microbial medicine technical fields, and in particular to a kind of secondary metabolites of Enteromorpha source fungi it is new
Purposes.
Background technique
To the research history of terrestrial microorganism last 100 years, it was found that a large amount of chemical structure diversity and bioactivity are significant
Natural products, greatly pushed the development of biotin drug, such as: penicillin, vancomycin, streptomysin.To microorganism
More and more concerns lead to the repetition research of microorganism and the machine of known compound overlapping development, noval chemical compound discovery naturally
Rate reduces.Therefore people begin to focus on that microorganism reserves are bigger, the more marine-derived microorganisms of specific diversity.Ocean is micro-
Biology is completely different with terrestrial microorganism since its unique living environment (high pressure, with high salt, low temperature, oligotrophic) causes it to have
Metabolic pathway, thus be more also easy to produce the secondary metabolite different from terrestrial microorganism, and then it is living to show good biology
Property such as: inhibit quorum sensing activity, bacteriostatic activity, antiviral activity, protein kinase inhibiting activity, cytotoxic activity, tumour thin
Born of the same parents' cycle arresting activity etc., thus increasingly become the important natural product chemistry resource of marine drug research worker favor.Sea
The abundance of microorganism reaches 10 in water6/ mL, bottom sediment bacterial abundance are even more to have reached 109/mL.However Oceanic Samples are adopted
Collection difficulty is high, only isolated lower than 5% marine microorganism under typical laboratory conditions, even if the microorganism obtained exists
The metabolite of its all biosynthesis pathway under marine environment can not be expressed under the conditions of laboratory cultures completely.Therefore
How to develop these hard-won Marine Microorganisms to the greatest extent, can to greatest extent by human use, at
For the vital task and project of scientific worker.Aspergillus fungi is all the important bacterium of natural products circle research all the time
Kind, secondary metabolite structure novel skeleton is changeable, in addition to the common structures types such as conventional steroidal, sequiterpene, anthraquinone with
Outside, contain toward contact: a variety of skeleton such as alkaloids, peptides, polyketone class, sesterterpene.The compound of these structure novels
Containing cell toxicant, antibacterial toward contact, the various actives such as antiviral become one of the important sources of marine drug lead compound,
Cause the extensive concern of scholar in the industry.
Summary of the invention
It is good that the purpose of the present invention is to provide the inhibitory effects of a kind of pair of alpha-glucosidase, slow down glucose generation and
It absorbs, prevents postprandial hyperglycemia, to the preferable effect of diabetes tool is prevented and treated, the Enteromorpha for being easy to be absorbed by organisms comes
The new application of the secondary metabolites of source fungi.
The technical solution that the present invention is taken to achieve the above object are as follows:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is preparing alpha-glucosidase restrainer and/or is controlling
Treat the new application in the drug of diabetes.
Preferably, secondary metabolites are compound, its stereoisomer or its officinal salt and pharmacy shown in Formulas I
Upper acceptable carrier or excipient,
Preferably, half-inhibitory concentration (IC of the secondary metabolites to alpha-glucosidase50) value be 1.2 μM.Deoxidation is wild
The IC of buttocks mycin and acarbose50Value is respectively 191.7 μM and 555.1 μM, illustrates targeted secondary metabolin to alpha-glucosaccharase
The inhibitory effect of enzyme is better than 1-Deoxynojirimycin and acarbose.
Preferably, the K of secondary metabolites pairiValue is 1.42 μM.This shows reactive compound to alpha-glucosidase
Inhibitory effect is better than acarbose, inhibits body alpha-glucosidase activity, slows down the generation and absorption of glucose, prevent
Postprandial hyperglycemia is preferably acted on diabetes tool is prevented and treated, especially diabetes B.
Preferably, alpha-glucosidase restrainer and/or the drug for treating diabetes further include pharmaceutically acceptable
Auxiliary material.
Preferably, alpha-glucosidase restrainer and/or treat diabetes drug further include compound shown in Formula II,
Its stereoisomer or its officinal salt and pharmaceutically acceptable carrier or excipient,
IC of the formula compound II to K562 cell50Value is 13.0 μM, to the IC of MCF-7 cell50Value is 10.1 μM, and right
Alpha-glucosidase does not have inhibiting effect, but compound II can play gain effect with compound I, to treatment diabetes
Effect be better than individual compound I.
Preferably, diabetes are type 1 diabetes or diabetes B.
Another object of the present invention is to provide a kind of versatility related gene expressions for promoting Enteromorpha source fungi, improve
The yield of secondary metabolites, while promoting the proliferation activity of Enteromorpha source fungi, the yield of its secondary metabolite is improved, is micro-
Biomedicine field provides the preparation method of the secondary metabolites of the Enteromorpha source fungi of new approach.
The technical solution that the present invention is taken to achieve the above object are as follows:
A kind of preparation method of the secondary metabolites of above-mentioned Enteromorpha source fungi, Enteromorpha source fungi is inoculated in containing TSA
It ferments, separates in enoxolone fermentation liquid, purify to get targeted secondary metabolin.TSA and enoxolone can be in fermentation liquid
The facilitation that collaboration is generated to the fermentation of Enteromorpha source fungi, carried out under different cell-signaling pathways effects cytoplasm and
Nucleus shuttles, and becomes slack high degree of coagulation Chromatin domains, removes the acetyl group on histone ε-N-acetyllysine residue
Group, so that DNA is more closely wrapped on histone, and it is compound to form co-suppression with other transcription factor regulatory proteins
Object, enhancing and the amino acid residue at enzyme active sites edge interact, and inhibitory effect and the selectivity to enzyme are improved, to press down
Gene expression processed enhances the degree of acetylation of histone, promotes transcriptional activation, causes transcriptional level to increase, do not changing gene
The selective expression of controlling gene, promotes the versatility related gene expression of Enteromorpha source fungi in the case where group DNA sequence dna,
The yield of secondary metabolites is improved, while the expression of Enteromorpha source fungi nitrogen metabolism related gene can be regulated and controled, is adjusted intracellular
The level of glutamine synthelase promotes absorption and utilization of the fungi to nitrogen source, and then promotes the proliferation of Enteromorpha source fungi living
Property, the yield of its secondary metabolite is improved, new approach is provided for microorganism field of medicaments.
Preferably, Enteromorpha source fungi is Aspergillus terreus OUCMDZ-2739.
Preferably, containing 9-11 μM of TSA and 0.8-1.3 μM of enoxolone in fermentation liquid.
Compared with prior art, the invention has the benefit that
Half-inhibitory concentration (the IC of the secondary metabolites alpha-glucosidase of Enteromorpha source of the present invention fungi50) value be 1.2 μ
The IC of M, 1-Deoxynojirimycin and acarbose50Value is respectively 191.7 μM and 555.1 μM, illustrates targeted secondary metabolin to α-
The inhibitory effect of glucuroide is better than 1-Deoxynojirimycin and acarbose, and the K of secondary metabolites pairiValue is 1.42 μM.
This shows that reactive compound is better than acarbose to the inhibitory effect of alpha-glucosidase, inhibits body alpha-glucosaccharase enzyme activity
Property, slow down the generation and absorption of glucose, prevent postprandial hyperglycemia, preferably acts on diabetes tool is prevented and treated,
Especially diabetes B；Simultaneously Enteromorpha source of the present invention fungi secondary metabolites from microorganism secondary metabolite,
Belong to natural alpha-glucosidase restrainer, is easy to be absorbed by organisms.The system of the secondary metabolites of Enteromorpha source of the present invention fungi
Preparation Method promotes the versatility related gene expression of Enteromorpha source fungi, improves the yield of secondary metabolites, while promoting Enteromorpha
The proliferation activity of source fungi improves the yield of its secondary metabolite, and new approach is provided for microorganism field of medicaments.
Present invention employs above-mentioned technical proposals to provide a kind of new application of the secondary metabolites of Enteromorpha source fungi, makes up
The deficiencies in the prior art, reasonable design, easy operation.
Specific embodiment
In the following, being described further in conjunction with specific embodiments to embodiment of the present invention.
Embodiment 1:
A kind of preparation method of the secondary metabolites of Enteromorpha source fungi, including,
1) Enteromorpha source fungi isolates and purifies: isolating fungi from Qingdao Stone old man bathing beach Enteromorpha sample
Enteromorpha sample is successively used antiseptic sea water, 75% ethyl alcohol and sterile water washing by bacterial strain Aspergillus terreus OUCMDZ-2739, then, with grinding
Alms bowl smashs sample to pieces, then deposits in PDA culture medium (every liter of 200g containing potato extract, Portugal containing 100mg/mL chloramphenicol
Grape sugar 20g, agar 15g and seawater 1L) in, it is cultivated at 28 DEG C to there is single culture, which is transferred to another PDA
Culture medium is simultaneously stored in 4 DEG C, passes through polyphase sort research, colonial morphology and the analysis of 18S rRNA sequence, strain development tree
It establishes, is identified as Aspergillus terreus OUCMDZ-2739；
2) Enteromorpha source fungi is inoculated in fermentation liquid and is fermented 30 days at 25 DEG C, fermentation liquid by glucose (10g/L),
Maltose (20g/L), mannitol (20g/L), monosodium glutamate (10g/L), KH2PO4(0.5g/L)、MgSO4·7H2O(0.3g/
L), yeast extract (3g/L), SEA salt (33g/L), TSA (10 μM) and tap water (1L, pH 7) composition；
3) the extraction separation and purifying of the secondary metabolites of Enteromorpha source fungi: fermentation liquid is filtered by garrha to divide
From filtrate and mycelia, the ethyl acetate extraction of filtrate equivalent volume obtains ethyl acetate solution, 80% acetone of mycelia three times
Three times, then acetone soln obtains ethyl acetate solution through being concentrated under reduced pressure with ethyl acetate extraction three times for extraction, will be above-mentioned two
Ethyl acetate solution is combined, and is concentrated under reduced pressure, and 31.2g ethyl acetate solution extract is obtained；Then by acetic acid
Ethyl ester extract is gradually eluted using decompression silica gel column chromatography, and eluent is petroleum ether-CH2Cl2(1:1 and 0:1) and
CH2Cl26 main fractions of surrender gradient (fraction 1~6) of-MeOH (100%~0%), the fraction 3 of 6.7g pass through Sephadex
LH-20(1:1CH2Cl2- MeOH) it is further purified, 1.4g fraction 3.3 and 1.1g fraction 3.4 are obtained, wherein fraction 3.3 continues to use
HPLC purifying, eluent 70%MeOH-H2O obtains 40.3mg targeted secondary metabolin (tR=11.5min) and 13.6mgization
Close object II (tR=16.4min).
Embodiment 2:
A kind of preparation method of the secondary metabolites of Enteromorpha source fungi, including,
1) Enteromorpha source fungi isolates and purifies: isolating fungi from Qingdao Stone old man bathing beach Enteromorpha sample
Enteromorpha sample is successively used antiseptic sea water, 75% ethyl alcohol and sterile water washing by bacterial strain Aspergillus terreus OUCMDZ-2739, then, with grinding
Alms bowl smashs sample to pieces, then deposits in PDA culture medium (every liter of 200g containing potato extract, Portugal containing 100mg/mL chloramphenicol
Grape sugar 20g, agar 15g and seawater 1L) in, it is cultivated at 28 DEG C to there is single culture, which is transferred to another PDA
Culture medium is simultaneously stored in 4 DEG C, passes through polyphase sort research, colonial morphology and the analysis of 18S rRNA sequence, strain development tree
It establishes, is identified as Aspergillus terreus OUCMDZ-2739；
2) Enteromorpha source fungi is inoculated in containing fermenting 30 days at 25 DEG C in TSA and enoxolone fermentation liquid, fermentation liquid
By glucose (10g/L), maltose (20g/L), mannitol (20g/L), monosodium glutamate (10g/L), KH2PO4(0.5g/L)、
MgSO4·7H2O (0.3g/L), yeast extract (3g/L), SEA salt (33g/L), TSA (10 μM), enoxolone (1.0 μM) and
Tap water (1L, pH 7) composition；
3) the extraction separation and purifying of the secondary metabolites of Enteromorpha source fungi: fermentation liquid is filtered by garrha to divide
From filtrate and mycelia, the ethyl acetate extraction of filtrate equivalent volume obtains ethyl acetate solution, 80% acetone of mycelia three times
Three times, then acetone soln obtains ethyl acetate solution through being concentrated under reduced pressure with ethyl acetate extraction three times for extraction, will be above-mentioned two
Ethyl acetate solution is combined, and is concentrated under reduced pressure, and 37.5g ethyl acetate solution extract is obtained；Then by acetic acid
Ethyl ester extract is gradually eluted using decompression silica gel column chromatography, and eluent is petroleum ether-CH2Cl2(1:1 and 0:1) and
CH2Cl26 main fractions of surrender gradient (fraction 1~6) of-MeOH (100%~0%), the fraction 3 of 8.1g pass through Sephadex
LH-20(1:1CH2Cl2- MeOH) it is further purified, 1.78g fraction 3.3 and 1.49g fraction 3.4 are obtained, wherein fraction 3.3 continues
Purified with HPLC, eluent 70%MeOH-H2O obtains 67.8mg targeted secondary metabolin (tR=11.5min) and 18.2mg
Compound II (tR=16.4min).TSA and enoxolone can be to the hairs of Enteromorpha source fungi in the preparation method fermentation liquid
Ferment generates the facilitation of collaboration, carries out cytoplasm under different cell-signaling pathways effects and nucleus shuttles, make height
Condensed chromatin region becomes slack, and the acetyl group on histone ε-N-acetyllysine residue is removed, so that DNA is more closely
It is wrapped on histone, and co-suppression compound, enhancing and enzyme active sites can be formed with other transcription factor regulatory proteins
The amino acid residue at edge interacts, and inhibitory effect and the selectivity to enzyme is improved, thus inhibition of gene expression, enhancing group egg
White degree of acetylation promotes transcriptional activation, causes transcriptional level to increase, downward the case where not changing genomic dna sequence
The selective expression for controlling gene, promotes the versatility related gene expression of Enteromorpha source fungi, improves obtaining for secondary metabolites
Rate, while the expression of Enteromorpha source fungi nitrogen metabolism related gene can be regulated and controled, adjust the water of intracellular glutamine synthelase
It is flat, promote absorption and utilization of the fungi to nitrogen source, and then promote the proliferation activity of Enteromorpha source fungi, improves the production of its cometabolism
The yield of object provides new approach for microorganism field of medicaments, and furthermore gained targeted secondary metabolin is mentioned in ethyl acetate solution
It takes accounting in object to greatly increase, further explains TSA and enoxolone in fermentation liquid and the fermentation of Enteromorpha source fungi is generated
The facilitation of collaboration.
Secondary metabolites are compound shown in Formulas I:
Compound II is compound shown in Formula II:
Embodiment 3:
The active testing of targeted secondary metabolin
1. targeted secondary metabolin tests the inhibition of alpha-glucosidase:
The preparation of alpha-glucosaccharase enzyme solutions: enzyme freeze-dried powder 0.1%BSA solution is dissolved, the enzyme of 100U/mL is made into
Liquid is placed in -20 DEG C of refrigerator and freezes.Before experiment, 100U/mL enzyme solution liquid-transfering gun is drawn on a small quantity, it is first molten with 0.1%BSA
Liquid is diluted to 20U/mL, then with 0.1%BSA solution to be diluted to 0.2U/mL spare.
PNPG solution is prepared: weighing a certain amount of PNPG solid, the phosphate buffer for being 6.8 with 0.1mol/L pH
(PBS) it dissolves, being made into concentration is 10mmol/L, then spare with the centrifuge tube packing of 1.5mL.
Using the activity of colorimetric method for determining alpha-glucosidase.Active ingredient to be measured is dissolved with dimethyl sulfoxide (DMSO)
Object prepares the stock solution that mass concentration is 10mg/mL.Stock solution is diluted with PBS again, is configured to the to be measured molten of required concentration
Liquid.Active compounds solution (experimental group), acarbose solution (positive controls) or PBS (yin are added in the every hole of 96 orifice plates
Property control group) 10 μ L, 50 μ L of PBS, 20 μ L of enzyme solutions, shake up, be placed in 37 DEG C of constant temperature water bath 10min, then be added PNPG solution
20 μ L, shake up, and 37 DEG C of reaction 10min add Na2CO330 μ L of terminate liquid terminates reaction, measures absorbance at 405nm immediately
Value.The inhibiting rate of alpha-glucosidase is calculated as follows in sample: inhibiting rate (%)=[(negative control group absorbance-experiment
Group absorbance)/negative control group absorbance] × 100%.When inhibiting rate of the sample to alpha-glucosidase is 50%, by sample
Quality concentration is set to half-inhibitory concentration (IC50) value.Targeted secondary metabolin is measured to inhibit the half of alpha-glucosidase
Concentration (IC50) value be 1.2 μM.The IC of 1-Deoxynojirimycin and acarbose50Value is respectively 191.7 μM and 555.1 μM, explanation
Targeted secondary metabolin is better than 1-Deoxynojirimycin and acarbose to the inhibitory effect of alpha-glucosidase.And above-mentioned secondary generation
The Ki value for thanking to object pair is 1.42 μM.This shows that reactive compound is better than acarbose to the inhibitory effect of alpha-glucosidase, suppression
Body alpha-glucosidase activity processed slows down the generation and absorption of glucose, postprandial hyperglycemia is prevented, to prevention and treatment
The preferable effect of diabetes tool, especially diabetes B.Compound II does not have inhibiting effect to alpha-glucosidase.
2. the cytotoxic activity of targeted secondary metabolin is tested:
Respectively using human chronic myeloblastic leukemia K562 cell and MCF-7 cell as model, adriamycin is positive control drug, is adopted
It is tested with cytotoxic activity of the MTT colorimetric method to targeted secondary metabolin.
The preparation method of MTT solution: weighing MTT0.5g, is dissolved in the phosphate buffer (PBS) of 100mL or without phenol red training
It supports in base, with 0.22 μm of membrane filtration to remove the bacterium in solution, puts 4 DEG C of refrigerators and be kept in dark place.It is preparing and is saving
During, container is encased with aluminium-foil paper, and the fluorescent lamp on super-clean bench is closed when experiment to be protected from light.
Method: cell density is adjusted to every milliliter 2 × 10 by the tumour cell of logarithmic growth phase5A/mL, by every hole 200
μ L is added in 96 porocyte culture plates, is passed through 5%CO in 37 DEG C2Incubator in cultivate 4h.5 concentration ladders are set separately in sample
Degree, each concentration set 3 Duplicate Samples, while setting positive, negative control, and every hole adds sample liquid or each 2 μ L of blank solution, cultivate 72h,
Then every hole adds 10 μ L of MTT liquid, continues to cultivate 4h, 37 DEG C, 2000r/min centrifugation 8min suck supernatant.Dimethyl is added in every hole
Each 100 μ L of sulfoxide, vibrates 15min on micro oscillator, until microplate reader measures at every hole 570nm after crystallization is completely dissolved
Light absorption value (OD value).Take the average absorbance value in three holes, and by IR%=(OD blank control-OD sample)/OD blank control ×
100% formula calculates the inhibiting rate of sample cell proliferation, and calculates half inhibiting rate IC using bliss method50.Measure target
The IC of secondary metabolites50Value is 9.5 μM, IC of the compound II to K562 cell50Value is 13.0 μM, to the IC of MCF-7 cell50
Value is 10.1 μM.
Embodiment 4:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is used for the medicine of alpha-glucosidase restrainer in preparation
Purposes in object.
The preparation of tablet: take 40g targeted secondary metabolin, 0.6g compound II, 37g edible cellulose, 17g lactose,
0.1g sodium citrate, 1.2g magnesium stearate and appropriate additive of tablet are uniformly mixed, by well known tablet manufacturing technology and equipment
Tablet is made, product specification is 0.5g/ piece.
Embodiment 5:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is preparing the use in the drug for treating diabetes
On the way.
The preparation of granule: targeted secondary metabolin 36g, 0.7g compound II, edible cellulose 67g and appropriate are taken
Granula auxiliary material is uniformly mixed, and is pelletized using drying process with atomizing.
Embodiment 6:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is preparing the use in the drug for treating diabetes
On the way.
The preparation of capsule: 25g targeted secondary metabolin, 0.9g compound II, 45g microcrystalline cellulose and appropriate capsule are taken
Agent auxiliary material is uniformly mixed, and fills capsule, and filling specification is 0.45g/.
Embodiment 7:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is preparing the use in the drug for treating diabetes
On the way.
The preparation of oral liquid formulations: targeted secondary metabolin 10g, 1.0g compound II, honey 8g and appropriate mouth are weighed
It takes liquor auxiliary material, the solution of 100ml is configured to purified water, filter, is filling, high-temperature short-time sterilization to obtain the final product.
Comparative example 1:
A kind of new application of the secondary metabolites of Enteromorpha source fungi is preparing the use in the drug for treating diabetes
On the way.
The preparation of capsule: taking 25g targeted secondary metabolin, 45g microcrystalline cellulose and appropriate capsule auxiliary material, and mixing is equal
It is even, capsule is filled, filling specification is 0.45g/.
Embodiment 9:
The therapeutic effect of 1 gained drug of embodiment 6 and comparative example
The building of diabetes B rat model: 100 SD rats (male, 150-180g) is chosen, is randomly divided into 2 groups: its
In 20 be Normal group, fed using normal diet；80 are high sugared group high in fat, (contain 10% using high-sugar-fat-diet
Lard, 20% sucrose, 2.5% cholesterol) raising.Record food ration daily records weekly a changes of weight.It is high after 1 month
High sugar group intraperitoneal injection streptozotocin (STZ, the sigma) 40mg/Kg of rouge, takes caudal vein blood monitoring blood glucose and remembers before injection
Record.Injection continuously monitored blood glucose 2 days after 3 days, and it is 2 type glycosurias that it is modeling success that blood glucose value, which is above 16.7mmol/L, twice
Disease model group.
Daily food ration and Avoirdupois monitoring be as the result is shown: the average food ration for 24 hours of high sugar group high in fat is 22.14 ± 2.77g/
Only, it is higher than normal group (19.24 ± 2.34g/ is only)；The weight of high sugar group high in fat is also higher than normally organizing, but fasting blood sugar still belongs to
In in range of normal value.After injection STZ tri- days, the weight of diabetic model group is begun to decline, and fasting blood-glucose reaches 20.8 ±
1.1mmol/L is higher than diabetes standard value 16.7mmol/L, and difference has significant (P < 0.05) compared with normal group.
And have typical diabetic disorders in behaviouristics: fur is uninteresting, burnout, drink, more food, diuresis, cloudy urine, syntexis more.
Show that diabetes B rat model is successful.
Administration: diabetes B model group by not give 1 gained drug of embodiment 6 and comparative example, be as administered 1 group and be administered 2
Group, dosage are respectively reactive compound 50mg/kg.
Physiochemical indice measurement: after animal is administered 48 days, fasting 12 hours, docking took whole blood in EDTA-K2 anticoagulant blood-collecting pipe
In (be purchased from BD), take 10 μ L anticoagulated whole bloods measurement glycosylated hemoglobin concentration and hemoglobin concentration, the two ratio is as saccharification
Content of hemoglobin (kit is purchased from RNADOX).
After administration 50 days, mouse plucks eyeball and takes blood, after being stored at room temperature 2 hours, in 15 DEG C, 3000rpm, is centrifuged 10min, point
From serum, with full automatic biochemical apparatus (Hitachi 7100) and biochemistry detection kit (purchased from middle raw north control) measurement glutamic-pyruvic transaminase
(ALT), glutamic-oxalacetic transaminease (AST), triglycerides (TG), total cholesterol (CHO), low density lipoprotein cholesterol (LDL-C), urine
Plain (UREA).Creatinine (CRE) carries out manual measurement using all-wave length microplate reader, measures total bile acid using all-wave length microplate reader
(TBA), total bilirubin (T-Bil), bilirubin direct (D-Bil) measure used kit by hand and are purchased from nine Johnson & Johnson's object of Beijing.Blood
Clear insulin content utilizes ELISA kit (being purchased from Merck Millipore) measurement.Various blood purification methods on serum advanced glycation
AGEs content utilizes ELISA kit (being purchased from cell biolabs).
After administration 50 days, control animals serum insulin content is 0.05ng/ml, model group animal blood serum insulin contains
Amount is 13.56ng/ml, 1 group of animal blood serum insulin content of administration is 2.33ng/ml, 2 groups of animal blood serum insulin contents of administration
For 3.72ng/ml.This explanation is in diabetes B mouse model, after administration 50 days, model group animal and normal group animal
It is significantly increased compared to insulin content, each administration group has different degrees of reduction compared with model group, this illustrates targeted secondary
Metabolin can improve hyperinsulinism state in animal body, can be used in treating diabetes B.And the degree of 2 groups of reductions is administered
Better than being administered 1 group, this illustrates that the drug of embodiment 6 is better than the drug of comparative example 1, specific mechanism to the effect for the treatment of diabetes
It is even unclear.
The prior art of routine techniques dawn known to those skilled in the art in above-described embodiment, therefore herein no longer in detail
It repeats.
The above embodiments are only used to illustrate the present invention, and not limitation of the present invention, the ordinary skill people of this field
Member can also make a variety of changes and modification without departing from the spirit and scope of the present invention.Therefore, all equivalent
Technical solution also belong to scope of the invention, scope of patent protection of the invention should be defined by the claims.
Claims (10)
1. a kind of new application of the secondary metabolites of Enteromorpha source fungi, it is characterised in that: inhibit preparing alpha-glucosidase
New application in the drug of agent and/or treatment diabetes.
2. a kind of preparation method of the secondary metabolites of Enteromorpha source fungi according to claim 1, it is characterised in that: institute
Stating secondary metabolites is compound, its stereoisomer or its officinal salt and pharmaceutically acceptable carrier shown in Formulas I
Or excipient,
3. a kind of preparation method of the secondary metabolites of Enteromorpha source fungi according to claim 1 or 2, feature exist
In: half-inhibitory concentration (IC of the secondary metabolites to alpha-glucosidase50) value be 1.2 μM.
4. a kind of preparation method of the secondary metabolites of Enteromorpha source fungi according to claim 1 or 2, feature exist
In: the K of the secondary metabolites pairiValue is 1.42 μM.
5. a kind of new application of the secondary metabolites of Enteromorpha source fungi according to claim 1, it is characterised in that: described
Alpha-glucosidase restrainer and/or the drug for treating diabetes further include pharmaceutically acceptable auxiliary material.
6. a kind of new application of the secondary metabolites of Enteromorpha source fungi according to claim 1, it is characterised in that: described
Alpha-glucosidase restrainer and/or treat diabetes drug further include compound shown in Formula II, its stereoisomer or its
Officinal salt and pharmaceutically acceptable carrier or excipient,
7. the new use of the secondary metabolites of Enteromorpha source fungi made from any one of -6 preparation methods according to claim 1
On the way, it is characterised in that: the diabetes are type 1 diabetes or diabetes B.
8. a kind of preparation method of the secondary metabolites of the Enteromorpha source any one of claim 1-7 fungi, it is characterised in that:
Enteromorpha source fungi is inoculated in containing fermenting in TSA and enoxolone fermentation liquid, is separated, is purified to get targeted secondary metabolin.
9. a kind of preparation method of the secondary metabolites of Enteromorpha source fungi according to claim 8, it is characterised in that: institute
Stating Enteromorpha source fungi is Aspergillus terreus OUCMDZ-2739.
10. a kind of preparation method of the secondary metabolites of Enteromorpha source fungi according to claim 8, it is characterised in that:
Contain 9-11 μM of TSA and 0.8-1.3 μM of enoxolone in the fermentation liquid.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111393421A (en) * | 2020-04-07 | 2020-07-10 | 自然资源部第三海洋研究所 | Butenolide derivative and preparation method and application thereof |
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| CN109045020A (en) * | 2018-09-04 | 2018-12-21 | 浙江海洋大学 | The purposes of Aspergillus terreus secondary metabolite extract |
| CN109280041A (en) * | 2018-09-04 | 2019-01-29 | 浙江海洋大学 | A kind of preparation method of the separation and Extraction noval chemical compound from A.terreus secondary metabolite |
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2019
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109045020A (en) * | 2018-09-04 | 2018-12-21 | 浙江海洋大学 | The purposes of Aspergillus terreus secondary metabolite extract |
| CN109280041A (en) * | 2018-09-04 | 2019-01-29 | 浙江海洋大学 | A kind of preparation method of the separation and Extraction noval chemical compound from A.terreus secondary metabolite |
Non-Patent Citations (3)
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| SUN, KUNLAI ET AL.: "Chemical-epigenetic method to enhance the chemodiversity of the marine algicolous fungus, Aspergillus terreus OUCMDZ-2739", 《TETRAHEDRON》 * |
| TAOTAO MA ET AL.: "A potential adjuvant chemotherapeutics, 18β-glycyrrhetinicacid,inhibits renal tubular epithelial cells apoptosis via enhancing BMP-7 epigenetically through targeting HDAC2", 《SCIENTIFIC REPORTS》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111393421A (en) * | 2020-04-07 | 2020-07-10 | 自然资源部第三海洋研究所 | Butenolide derivative and preparation method and application thereof |
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