CN1908156B - Monascus novel strain and traditional Chinese medicine monascus prepared by fermenting the same - Google Patents
Monascus novel strain and traditional Chinese medicine monascus prepared by fermenting the same Download PDFInfo
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- CN1908156B CN1908156B CN2005100213916A CN200510021391A CN1908156B CN 1908156 B CN1908156 B CN 1908156B CN 2005100213916 A CN2005100213916 A CN 2005100213916A CN 200510021391 A CN200510021391 A CN 200510021391A CN 1908156 B CN1908156 B CN 1908156B
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Abstract
the invention discloses a new strain of Monascus Purpureus Went and preparing method of traditional Chinese medicine antihyperglycemic Monascus color, which is composed of 15-30mg/g open, close ring structured Luovastatin, 3-6mg/g unsaturated fatty acid and not more than 100ppb orange mycin, wherein the content of open-structured Luovastatin is 15-90%.
Description
Technical field
The present invention relates to the Chinese medicine Antilipemic monascus that red colouring agent for food, also used as a Chinese medicine belongs to new bacterial strain and fermentative preparation thereof, relate in particular to a kind of Chinese medicine Antilipemic monascus of the red colouring agent for food, also used as a Chinese medicine strain fermentation through screening domestication, belong to biological technical field.
Background technology
Raising and aging population along with people's living standard, (mainly show as serum cholesterol TC, triglyceride level TG raises hyperlipidaemia, a kind of whole body lipid metabolism that high density lipoprotein cholesterol HDL-C reduces, low-density lipoprotein (LDL) raises is unusual) sickness rate rise year by year, because of the hyperlipidaemia that lipid metabolism disorders causes, become the important pathogenic factors of common diseases such as fatty liver, coronary heart disease and atherosclerosis.
The drug main of reducing blood-fat commonly used will be divided into following a few class clinically at present: 3-hydroxy-3-methylglutaryl-coenzyme A (3-hydroxy-3-methylglutaryl-coenzyme A, HMG-CoA) reductase inhibitor (Statins, statin), bile acid chelating agent (bile acid sequestrants), nicotinic acid (nicotinic acid, niacin) and derivative, fibrate or claim the special class (fibrates) of shellfish, fish oil preparation (polyunsaturated fatty acid) etc.The ideal lipid lowerers is two kinds of medication combined medications that the mechanism of action is different.Statins has become the most frequently used prescription drug of treatment hypercholesterolemia at present.The statins that now gone on the market has: lovastatin (lovastatin), Simvastatin (simvastatin), Pravastatin (pravastatin), atorvastatin (atovastatin) etc.At present, in order to make effect for reducing fat more comprehensively, often need to take the fat-reducing medicament of two kinds of different mechanisms of action.Curative effect of medication as statins triglyceride reducing, high density lipoprotein increasing cholesterol is not ideal enough, needs the special class treatment of associating shellfish, has increased treatment cost and side effect (rhabdomyolysis).
Red colouring agent for food, also used as a Chinese medicine is one of traditional Chinese medicinal materials of China, is to be that raw material is made through fermentation with monascus and polished rice, has invigorating the spleen and regulating the stomach and helps digestion, and the effect of promoting blood circulation, removing blood stasis and relieving pain has long medicinal and edible history in China.1979, contain Mo Na Kelin (Monacolin) series material in the red colouring agent for food, also used as a Chinese medicine of certain special monascuses of discovery such as Endo, the effect that suppresses hydroxyl first glutaryl CoA reductase (HMG-CoA reductase enzyme) in the body's cholesterol biosynthetic process is arranged, this red colouring agent for food, also used as a Chinese medicine is called as Antilipemic monascus, is also referred to as functional Monascus.Wherein Monacolin K (lovastatin, it is the strongest lovastatin) to suppress cholesterol Synthesis effect, kind of Monacolin K analog surplus discovery at present has ten approximately, the statin substance open loop structure is the activity form of Monacolin class lipid-regulation medicine.Monacolin K open loop structure only is present in the Red kojic rice of minority fermentation, it can directly combine with the HMG-CoA reductase enzyme and suppress in vivo synthetic of cholesterol, and do not need in the body alcohol acid lipase to participate in hydrolysis, do not increase burden of liver, effect for reducing fat can take place, and is one of main effective constituent of red colouring agent for food, also used as a Chinese medicine.There is bigger individual difference in the ability that human body produces alcohol acid lipase, somebody even do not have fully, so the effect for reducing fat of closed loop is in different patients difference great disparity on one's body.
The Japan scholar studies show that the lipid acid that the monascus culture mainly contains is Palmiticacid, oleic acid, linolic acid.Song Hongtao adopts gas chromatographic analysis, and the result shows that unsaturated fatty acid content reaches 64%-77%, and polyene unsaturated fatty acid content reaches 16%-27%.Polyunsaturated fatty acid is that a class contains two or more pairs key, and carbonatoms is the straight chain fatty acid of 16-22, and human body must lipid acid be linolic acid, linolenic acid, arachidonic acid for three kinds.In recent years studies show that polyunsaturated fatty acid have synthetic, the prevention rheumatoid arthritis of reducing blood-fat, reducing cholesterol, inhibition fat, prophylaxis of tumours and diabetes, and many aspects such as immunity of organism effect is widely arranged, the content of the necessary lipid acid of three-type-person's body is very low in the daily bread, is not enough to satisfy needed by human body.Linolic acid is converted into linolenic acid under the effect of 6-desaturase in human body.Juzlova etc. find that monascus can produce lipid acid when the meta-bolites of research monascus, 22 kinds of saturated fatty acids are wherein arranged, 14 kinds of monoenoic acids, and two kinds of diolefinic acids and linolic acid, wherein linolic acid accounts for 80% of whole lipid acid.
Kuroda etc. find cholesterol synthetic influence experiment by external more than 40 kinds of lipid acid: a lot of lipid acid can both suppress the synthetic of cholesterol, unsaturated fatty acids with two keys suppresses active strong than saturated fatty acid to the cholesterol synthetic, two keys are many more, and activity is strong more.Unsaturated fatty acidss such as oleic acid, linolic acid and Ara can promote the operation of cholesterol in blood, are deposited on possibility on the vessel wall with minimizing, reach the arteriosclerotic purpose of control.The Chinese medicine that at present existing a lot of effective constituents are polyunsaturated fatty acid is applied to clinical as anticholesteremic agent, as Thistle oil, γ-seed of Radix Oenotherae erythrosepalae oil, sea-buckthorn wet goods.
Nineteen ninety-five, people such as French scholar Blanc have found the Citrinin (Citrinin) in red colouring agent for food, also used as a Chinese medicine liquid state fermentation, the solid state fermentation.Citrinin belongs to mycotoxins, has renal toxicity, toxicity isogonic hypertoxicity placed in the middle, also has teratogenecity simultaneously.According to both domestic and external, monascus parpureus Went and red monascus bacterial classification are considered to produce the bacterial classification of Citrinin.
Patent CN1059463 discloses the purplish red aspergillus of new bacterial strain, Monascus anka Nakazawa et sato and preparation method thereof, contains lovastatin 8mg/g; CN1061537C discloses the red colouring agent for food, also used as a Chinese medicine pharmaceutical preparation of the purplish red aspergillus of new bacterial strain, Monascus anka Nakazawa et sato system, and said preparation is a blood fat reducing, reducing cholesterol again, and side effect is low, contains the lovastatin of open loop; CN1065432C discloses nineteen Monascus anka Nakazawa et sato bacterial classification, with and the Antilipemic monascus of fermentative preparation; CN1373208 discloses high yield lovastatin bacterial strain, in the Hongqu powder (red colouring agent), contains lovastatin 15-20mg/g; CN1448503 discloses a kind of preparation of no citrinin high luminance relay valency functional red koji powder, contains lovastatin 4-12mg/g, open loop lovastatin 70-90%; CN1448503 has filtered out and has not produced Citrinin or the extremely micro-bacterial strain of Citrinin output, lovastatin 3-16mg/g in the Hongqu powder (red colouring agent), lovastatin 9-12.5mg/g in the purplish red aspergillus Hongqu powder (red colouring agent); CN1580272 discloses lovastatin 12mg/g in the Antilipemic monascus powder of high yield lovastatin; CN1513451 is open to be the Chinese medicine red colouring agent for food, also used as a Chinese medicine and the preparation thereof of main component with lovastatin salt.
In sum, also do not possess lovastatin content height, open loop ratio height, content of polyunsaturated fatty acid height simultaneously, contain the monascus fermentative production Chinese medicine Antilipemic monascus report of Citrinin hardly.
Summary of the invention
Technical scheme of the present invention provides a kind of Chinese medicine Antilipemic monascus of red koji strain fermentation, technical scheme to be solved by this invention provides a kind of through screening domestication monascus parpureus Went bacterial strain, and the present invention also provides the preparation method and the purposes of this Chinese medicine Antilipemic monascus.
The new bacterial strain of monascus provided by the invention is monascus parpureus Went (Monascus Purpureus Went), unsaturated fatty acids in its tunning is 3-6mg/g, citrinin content is smaller or equal to 100ppb, lovastatin 15-30mg/g, wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
Red colouring agent for food, also used as a Chinese medicine provided by the invention belongs to the new bacterial strain of monascus parpureus Went, is that the preserving number by China typical culture collection center C CTCC preservation is the bacterial strain of CCTCC M 205073.Monascus parpureus Went belongs to mycota (Fungi), Ascomycota (Ascomycetes), Plectomycetes (Plectomycetes), Eurotiale (Eurotiales), red colouring agent for food, also used as a Chinese medicine section (Monascaceae), red colouring agent for food, also used as a Chinese medicine genus (Monascus).
Monascus strain of the present invention is to separate from soil to obtain, and is reference strain with the purple red colouring agent for food, also used as a Chinese medicine bacterial strain (Monascus Purpureus Went) of Sichuan University's bioengineering dept microorganism classification chamber preservation, and qualification result is as follows:
Morphological specificity:
(1), will identify bacterial strain and reference strain (being reference culture), be connected to respectively on the wort agar flat board and (respectively connect 5 wares), cultivate in 25 ℃-28 ℃ and compared observation in 12 days, its result is as follows:
(1) bacterium colony φ 2.8-3.2cm, the middle part is purplish red to be striven, the green orange in limit, villous shape aerial hyphae, the epithelium shape that has gauffer occurs, the back side is maroon.
(2) mycelia has every, multinuclear, mycelia and connects by having located tangible podocyte with conidiophore or capsule stalk
(3) cleistothecium sphere, tool handle, the handle cleistothecium ball φ 8-15cm that closes different in size, the redness during children is orange after the maturation.
(4) ascus sphere includes 8 thecaspores, and ascus disappears when closing the capsule maturation, is difficult for observing.
(5) thecaspore ellipse, the school conidium is little, and look shallow, the single monolayer wall.
(6) conidium pyriform.The tool double wall, big than thecaspore, single giving birth to or 2-3 bunchiness.
(7) growth temperature range is 15 ℃-42 ℃, does not grow below 14 ℃ He more than 43 ℃, 25 ℃-32 ℃ of thermophilics.
Bacterial strain of the present invention in shape, the size of form, color, quality, mycelial structure, cleistothecium, the mode of giving birth to, ascus and thecasporous shape size, number, decorative pattern and conidial size, form, decorative pattern and to give birth to mode all similar to reference strain.
Physiological and biochemical property (to the utilization of matrix):
Conform to reference culture, identify bacterial strain well-grown on wort, tomato juice, soya bean substratum; Growth is general on Semen Maydis powder, PDA, wheat skin substratum; On the Gao Shi substratum, can grow, limitation is arranged; On czapek'S medium, do not grow.
Think according to interpretation and to be identified that bacterial strain should be monascus parpureus Went (Monascus Purpureus Went), with reference strain relatively, do not have obvious variation state.
When using strain fermentation of the present invention to prepare the Chinese medicine red colouring agent for food, also used as a Chinese medicine, contain unsaturated fatty acids 3-6mg/g in the tunning, also contain lovastatin 20-40mg/g, wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure, and Citrinin is smaller or equal to 100ppb, all inequality with the fermentation strain of the existing Chinese medicine red colouring agent for food, also used as a Chinese medicine of reporting, be the new monascus strain of a strain therefore.
The present invention also provides a kind of Chinese medicine red colouring agent for food, also used as a Chinese medicine, it is characterized in that containing unsaturated fatty acids 3-6mg/g, lovastatin 15-30mg/g, and wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
Described Chinese medicine red colouring agent for food, also used as a Chinese medicine contains following main effective constituent: lovastatin 15-30mg/g, and unsaturated fatty acids 3-6mg/g, wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
Further, polyunsaturated fatty acid 2-5mg/g in the unsaturated fatty acids.Described unsaturated fatty acids is mainly oleic acid and linolic acid.The Chinese medicine red colouring agent for food, also used as a Chinese medicine contains Citrinin smaller or equal to 100ppb.
Described Chinese medicine red colouring agent for food, also used as a Chinese medicine is by the monascus fermentative preparation.The preparation method comprises the steps:
A, seed liquor preparation
Monascus parpureus Went (Monascus Purpureus Went), deposit number by China typical culture collection center (CCTCC) preservation is the bacterial strain of CCTCC M 205073, use seed as producing, in 30～40 ℃, 180～220rpm cultivates 8～24h as first order seed or fermentor tank seed liquor in the PDA substratum;
B, fermentation: adopt liquid state fermentation or solid state fermentation, drying gets Hongqu powder (red colouring agent) of the present invention.
Wherein, the culture medium prescription of the described liquid state fermentation of step b is (g/L):
Carbon source (sucrose) 10～40, nitrogenous source (peptone) 10～45, extractum carnis 0.5～3, CaCl
20.05～1, pH6.5 ± 1.0,121 ℃ sterilization 20min, wherein, carbon source can be used replacements such as glucose, glycerine, Zulkovsky starch, molasses, and nitrogenous source can be used corn steep liquor, yeast extract paste, extractum carnis, plant or animal proteinum peptone;
Fermentation process is: the seed liquor for preparing is inserted in the fermention medium according to 5%～20% inoculum size.Fermentation culture is based on 30～40 ℃, and 180～220rpm cultivates 5d～7d, finishing 24h from cultivating, adds open loop stabilization liquid, the centrifugal supernatant of removing, and auxiliary material is dosed in the mycelium fragmentation, and drying obtains Hongqu powder (red colouring agent) of the present invention.
The component of the nutritive medium of described solid state fermentation is (g/L): glycerine 50～300, peptone 50～300, extractum carnis 50～100, Na
2HPO
412H
2O 5～20, NaH
2PO
42H
2O 5～20, CaCl
20.5～10, MgSO
47H
2O 0.5～10, Na
2CO
30.5～10.
Process for solid state fermentation is: will obstruct rice sterilization back and spray nutritive medium, weight proportion is that every 1000g rice sprays nutritive medium 20～120ml, obtain substratum in 100 ℃ of boilings, after the cooling seed liquor for preparing inserted according to 5%～20% inoculum size and be mixed with in the substratum of nutritive medium; Cultivation is based on 30～45 ℃, humidity 〉=80%, and 8d and 12d add the nutritive medium of solid state fermentation by 5% (liquid-solid ratio) after cultivation respectively, finish in back 14 days in fermentation, and drying obtains red colouring agent for food, also used as a Chinese medicine of the present invention.
The present invention also provides a kind of pharmaceutical composition, and it is that described red colouring agent for food, also used as a Chinese medicine is an activeconstituents, and Hongqu powder (red colouring agent) is broken to the 80-200 order, adds the medicament that acceptable accessories or complementary composition are prepared from.
Described medicament is: tablet, capsule, dispersion agent, micronization preparation, micropill.
The present invention also provides purposes and the purposes in the medicine of preparation treatment fatty liver of aforementioned pharmaceutical compositions in the medicine of preparation treatment and preventing hyperlipidemia and relative disease thereof.
In sum, the Chinese medicine Antilipemic monascus powder that the invention provides the monascus ruber fermentation has lovastatin content height, open loop ratio height, content of polyunsaturated fatty acid height simultaneously, does not contain Citrinin.This Chinese medicine Antilipemic monascus is compared with simple lovastatin, and the various active composition has synergy, and having overcome existing medicine needs drug combination, medicine valency height, shortcoming that side effect is big.The present invention is also by to the screening of monascus parpureus Went bacterial strain domestication, and screening has been tamed out to have stable high lovastatin productive rate, high yield polyunsaturated fatty acid, do not produce and the bacterial strain of utmost point low-yield citrinin.And the cycle is short, efficient height, the production fermentation process that cost is low.The present invention do not use solvent extraction and adopt convection drying post-processing technology, guarantee in the aftertreatment that the lovastatin loss is little and make open loop in the product, closed loop proportional stable.
In a word, the Chinese medicine Antilipemic monascus of monascus parpureus Went of the present invention fermentation with compare with the lovastatin of dosage have good effect, security is higher, advantage such as cheap.
Embodiment
The preparation of embodiment 1 Chinese medicine Antilipemic monascus of the present invention: (numbering A)
The bacterial classification that the present invention uses is to separate from soil to obtain, by Sichuan University's monascus parpureus Went that bio-engineering research is accredited as (Monascus Purpureus Went) bacterium, China typical culture collection center (CCTCC) gives preservation, and deposit number is CCTCC M 205073.
Behind the work seed lot bacterial classification breakdown, the PDA nutrient agar is dull and stereotyped to be cultivated, be stored in the PDA test tube slant, after form, purity, throughput passed examination, [promptly on cultured PDA test tube slant, collude the culture smear that takes a morsel, microscopically is observed behind the gramstaining, form unanimity under all mirrors, and the seed of no living contaminants is a certified seed.The throughput inspection is the throughput (being the open loop content of every milliliter of fermented liquid) that the fermentation of chamber level by experiment detects bacterial strain, and all throughput is certified seed greater than 1ug/ml].Use seed as producing, seed is in 30～40 ℃, and 180～220rpm cultivates 24h as first order seed in the PDA substratum, also can be used as the fermentor tank seed liquor.
Solid state fermentation nutrient solution prescription (g/L): glycerine 50, peptone 50, extractum carnis 50, Na
2HPO
412H
2O 5, NaH
2PO
42H
2O 5, CaCl
20.5, MgSO
47H
2O 0.5, Na
2CO
30.5.
A nutritive medium (weight proportion is that every 1000g rice sprays nutritive medium 120ml) is sprayed in stalk rice sterilization back, obtains substratum in 100 ℃ of boilings, after the cooling seed liquor for preparing is inserted according to 20% inoculum size to be mixed with in the substratum of nutritive medium.Cultivation is based on 30～45 ℃, humidity 〉=80%; Ferment after 5 days and to stir 2 times every day, to keep fermentation evenly; 8d and 12d finished in fermentation by the nutritive medium of 5% (liquid-solid ratio) adding solid state fermentation in back 14 days after cultivation respectively, put 70 ℃ of temperature dryings 5 hours, had both obtained the high natural Antilipemic monascus powder of open loop structure content of the present invention.
The preparation of embodiment 2 Chinese medicine Antilipemic monascus of the present invention:
The bacterial classification that the present invention uses is to separate from soil to obtain, by Sichuan University's monascus parpureus Went that bio-engineering research is accredited as (Monascus Purpureus Went) bacterium, China typical culture collection center (CCTCC) gives preservation, and deposit number is CCTCC M 205073.
Behind the work seed lot bacterial classification breakdown, the PDA nutrient agar is dull and stereotyped to be cultivated, be stored in the PDA test tube slant, after form, purity, throughput passed examination, [promptly on cultured PDA test tube slant, collude the culture smear that takes a morsel, microscopically is observed behind the gramstaining, form unanimity under all mirrors, and the seed of no living contaminants is a certified seed.The throughput inspection is the throughput (being the open loop content of every milliliter of fermented liquid) that the fermentation of chamber level by experiment detects bacterial strain, and all throughput is certified seed greater than 1ug/ml].Use seed as producing, seed is in 30～40 ℃, and 180～220rpm cultivates 24h as first order seed in the PDA substratum, also can be used as the fermentor tank seed liquor.
Solid state fermentation nutrient solution prescription (g/L): glycerine 300, peptone 300, extractum carnis 100, Na
2HPO
412H
2O20, NaH
2PO
42H
2O 20, CaCl
210, MgSO
47H
2O 10, Na
2CO
310.
A nutritive medium (weight proportion is that every 1000g rice sprays nutritive medium 20ml) is sprayed in stalk rice sterilization back, obtains substratum in 100 ℃ of boilings, after the cooling seed liquor for preparing is inserted according to 5% inoculum size to be mixed with in the substratum of nutritive medium.Cultivation is based on 30～45 ℃, humidity 〉=80%; Ferment after 5 days and to stir 2 times every day, to keep fermentation evenly; 8d and 12d finished in fermentation by the nutritive medium of 5% (liquid-solid ratio) adding solid state fermentation in back 14 days after cultivation respectively, put 70 ℃ of temperature dryings 5 hours, had both obtained the high natural Antilipemic monascus powder of open loop structure content of the present invention.
The preparation of embodiment 3 Chinese medicine Antilipemic monascus of the present invention:
Behind the work seed lot bacterial classification breakdown, the PDA nutrient agar is dull and stereotyped to be cultivated, be stored in the PDA test tube slant, after form, purity, throughput passed examination, [promptly on cultured PDA test tube slant, collude the culture smear that takes a morsel, microscopically is observed behind the gramstaining, form unanimity under all mirrors, and the seed of no living contaminants is a certified seed.The throughput inspection is the throughput (being the open loop content of every milliliter of fermented liquid) that the fermentation of chamber level by experiment detects bacterial strain, and all throughput is certified seed greater than 1ug/ml].Use seed as producing, seed is in 30～40 ℃, and 180～220rpm cultivates 8～24h as first order seed in the PDA substratum, also can be used as the fermentor tank seed liquor.
Solid state fermentation nutrient solution prescription (g/L): glycerine 150, peptone 150, extractum carnis 80, Na
2HPO
412H
2O12, NaH
2PO
42H
2O 15, CaCl
27, MgSO
47H
2O 6, Na
2CO
38.
A nutritive medium (weight proportion is that every 1000g rice sprays nutritive medium 100ml) is sprayed in stalk rice sterilization back, obtains substratum in 100 ℃ of boilings, after the cooling seed liquor for preparing is inserted according to 10% inoculum size to be mixed with in the substratum of nutritive medium.Cultivation is based on 30～45 ℃, humidity 〉=80%; Ferment after 5 days and to stir 2 times every day, to keep fermentation evenly; 8d and 12d finished in fermentation by the nutritive medium of 5% (liquid-solid ratio) adding solid state fermentation in back 14 days after cultivation respectively, put 70 ℃ of temperature dryings 5 hours, had both obtained the high natural Antilipemic monascus powder of open loop structure content of the present invention.
The preparation of embodiment 4 Chinese medicine Antilipemic monascus of the present invention: (numbering B)
(1) production bacterial classification amplification cultivation
1. the preparation of seed culture medium:
Potato 200g, glucose 20g, agar 20g adds water 100ml, boils 30 minutes, and 1.0Kg/cm is put in packing
2Sterilized 20 minutes, promptly.
2. expanding species:
Under aseptic condition, to produce with bacterial classification Monascus parpureus Went China typical culture collection center (CCTCC) and give preservation, deposit number is CCTCC M 205073, be seeded on the cultivation basis of potato glucose sugar agar, relative humidity is controlled at 40-60%, in 32-34 ℃ of amplification cultivation 5-7 days, obtain seeding.
(2) fermentation, drying
1. the preparation of nutritive medium:
Peptone 50g, yeast extract paste 20g, potassium primary phosphate 2g adds water to 1000ml, promptly.
2. fermentation:
Select for use rice to should be fresh, do not have the Sichuan rice that goes mouldy, carry out washing, elutriation before the fermentation earlier, add the 1000ml nutritive medium by every 1Kg rice, mixing places 121 ℃ of (1.0Kg/cm
2/) vapor sterilization 25 minutes, standby.The fermentation inoculation should be dissolved in seeding in the sterilized water earlier, inoculates in sterilized rice mixing.Leavening temperature is controlled at 32-34 ℃, and indoor relative humidity is controlled at 40-60%, fermentation time 15 days, and stir 2 the every day of fermenting after 5 days,
To keep fermentation evenly.
3. dry:
The red colouring agent for food, also used as a Chinese medicine that will ferment is put tray (thickness 5cm is following), puts 80 ℃ of temperature dryings 4 hours, has both obtained the natural Antilipemic monascus powder of open loop structure content 10-30% of the present invention.
The assay of lovastatin composition in the embodiment 5 Chinese medicine Antilipemic monascus of the present invention
Take by weighing above-mentioned Chinese medicine Antilipemic monascus 100g, 500ml methanol extraction three times, behind the concentrating under reduced pressure, C18 reversed-phase silica gel column chromatography on the extract, methyl alcohol: the water gradient elution gets 2 components, concentrates, leaves standstill, crystallization.The I that weighs is 0.42g, and II is 0.38g.
HPLC: ultraviolet detection 238nm, UV spectrum: solvent is the methyl alcohol infrared spectra: the KBr compressing tablet,
1H-NMR:TMS is interior mark, CDCl
3Be solvent, mass spectrum: ionizer 70eV, acceleration voltage 6000V.
The spectral data of the open and close ring structure compound of the Monacolin K that result that Spectrum Analysis obtains and document are reported contrasts, and determines that I is the open loop structure of Monacolin K, and II is the closed-loop structure of Monacolin K.
Above-mentioned Chinese medicine Antilipemic monascus sample, mortar grinds, and crosses 80 mesh sieves.Precision takes by weighing 2g (being accurate to 0.001g), with methanol constant volume to the 10ml volumetric flask, supersound extract 20min, the centrifugal 10min of 6000rpm draws the supernatant sample introduction.
The employing reversed-phase HPLC chromatography carries out, and chromatographic condition is: chromatographic column is that (4.6 * 150mm * 5um), moving phase is methyl alcohol to Shim-pack VP-ODS C: 18mM phosphoric acid solution 77: 23 (v/v), flow velocity 0.6ml/min, detect wavelength 238nm, 30 ℃ of column temperatures, sampling volume are 20 μ l.
The preparation of reference substance solution: it is an amount of that precision takes by weighing the reference substance of lovastatin open and close ring structure, adds 75% ethanol and make the solution that contains 0.1mg among every 1ml, promptly.
Accurate respectively above-mentioned reference substance and each 20 μ l of need testing solution of drawing inject hplc determination, according to lovastatin open and close ring structure content in the color atlas calculated by peak area Chinese medicine Antilipemic monascus.
Lovastatin open loop structure formula:
Lovastatin opening structure chemistry formal name used at school is 1,2,6,7,8,8 α-6 hydrogen δ-and, β-two hydroxyls-2, two methyl-8-[2-methyl]-benzene-butyryl acyloxy]-1-naphthalene-3,5-second hydroxyl-enanthic acid.
Detect lovastatin content in embodiment 1, embodiment 4 each 5 batches of fermented tcm Antilipemic monascus, open and close ring structure content.The results are shown in Table 1.
Lovastatin content in the table 1 Chinese medicine Antilipemic monascus
The research of fatty acid component in the embodiment 6 Chinese medicine Antilipemic monascus of the present invention:
1, the extraction of lipid acid in the Chinese medicine Antilipemic monascus
The receiving flask of apparatus,Soxhlet's is cleaned, dried by the fire 2 hours down in 105 ℃, taking-up is put and is cooled to room temperature in the moisture eliminator, weighs, and precision takes by weighing powder 2-5g in the Chinese medicine Antilipemic monascus in cellulose thimble, and wrap up with absorbent cotton at filter paper nozzle, a tube end; Cellulose thimble is put into extraction tube, in the receiving flask of having weighed, pour the ether of 1/3-1/2 into, connect the apparatus,Soxhlet's each several part, in water-bath, heat, bath temperature is 70-80 ℃, and the control ether splashes into cellulose thimble from condenser 150 droplets/minute of speed are advisable, and make the continuous refluxing extraction of ether; Extracted 5 hours, and till no oil droplet after the effusive solvent evaporation of extractor, stopped heating after extracting finishes, receiving flask is tilted in cooling, and ether flow in the receiving flask, at last receiving flask is linked to each other with common prolong, boils off ether; Receiving flask is dry in baking oven, place 80 ℃ of moisture eliminators to be cooled to room temperature again, claim to constant weight.
2, adopting sulfuric acid-methyl alcohol method is corresponding fatty acid methyl ester with fatty acid derived:
Each lipid acid standard specimen is taken by weighing in right amount, mix, place port grinding bottle, add sulfuric acid one methanol solution [getting the 1ml vitriol oil (proportion is 1.84) is added drop-wise in the 98ml anhydrous methanol] of 10 times of amounts, jolting is even, load onto the condenser that has calcium chloride tube, 60-80 ℃ of reflux 2h boils off methyl alcohol, and adding distil water is an amount of, pour in the separating funnel after the jolting, add the sherwood oil jolting, tell ether liquid, with distilled water repetitive scrubbing ether liquid, be neutral to washing lotion, spend the night with anhydrous sodium sulfate drying, filter steaming petroleum ether, use a small amount of sherwood oil constant volume again, get the fatty acid methyl ester mixed standard solution.
3, detect lipid acid with Gas Chromatography-mass Spectrometer (GCMS)
Chromatographic condition:
Instrument: HP6890/5973 GC/MS chromatograph-mass spectrometer coupling system.
Chromatographic column: HP-5 30m * 0.3mm * 0.25um
Column temperature:
Carrier gas: He, 0.9ml/min; Injector temperature: 260 ℃; Splitting ratio: 20: 1; Sample size: 1 μ l; Mass spectrum condition ion source temperature: 280 ℃; Ionization mode: electron-bombardment (EI); Electron energy: 70ev; Quadrupole temperature: 280 ℃; Mass range: 30-400u
Measure: the GC-MS for determining sample gets the mass spectrum of each component through gas-chromatography, and the machine examination rope gets each component result as calculated.
4. gas chromatographic detection:
Chromatographic condition: gas chromatographic column is that (30m * 0.25mmi.d), thickness of liquid film is 0.25 μ m to the crosslinked quartz column of PE-225 kapillary, uses N
2Do carrier gas, carrier/5.0 psi, oven:208 ℃, injector temperature: 250 ℃, the FID temperature: 300 ℃, splitting ratio 20: 1, H
2: 45ml/min, air: 450ml/min.
Measure: draw reference substance solution and need testing solution 1 μ l respectively, inject gas chromatograph is measured by above-mentioned gas phase condition, with the external standard standard measure.
Detect embodiment 1, embodiment 4 each 5 batches of fermented tcm Antilipemic monascus, the content of its lipid acid is 4.36 ± 1.01mg/g, and its unsaturated fatty acids accounts for 87.79%, and saturated fatty acid accounts for 12.21%, saturated fatty acid/unsaturated fatty acids=0.139.See the following form 2 concrete the composition.
Lipid acid in the table 2 Chinese medicine Antilipemic monascus is formed table
The fatty acid analysis experimental result shows, have 21 fatty acid methyl ester chromatographic peaks in the Chinese medicine Antilipemic monascus sample, its hexadecanoic acid (palmitinic acid), octadecanoic acid (stearic acid), 9-octadecenoic acid (oleic acid), 9, the peak area of 12-octadecadienoic acid (linolic acid), punicic acid (linolenic acid), tetracosa carbon alkanoic acid (tar acid holds with both hands with both hands) is bigger, and these lipid acid account for more than 97% of total fatty acid content.Content of polyunsaturated fatty acid accounts for 40%.
The mensuration of Citrinin in the embodiment 7 Chinese medicine Antilipemic monascus of the present invention:
1, the Citrinin extracting mode is in the red koji fermentation sample:
Accurately take by weighing red colouring agent for food, also used as a Chinese medicine 0.2g, the aqueous hydrochloric acid 0.5ml acidifying 10min with pH is 2 adds toluene, the mixed extraction agent 2ml of ethyl acetate again, adopt sonic oscillation to extract, time 15min, centrifugal (6000rpm, 10min), draw in another centrifuge tube of solvent stratification, blowing air volatilizes in 45 ℃ of water-baths, and residue dissolves with methyl alcohol 100 μ l, centrifugal (6000rpm, 10min), it is standby to get supernatant.
2, Citrinin HPLC testing conditions is:
Chromatographic column: Irrogulour-H C18 post (250 * 4.6mm, 10 μ m); Pre-column: Phenomenex C18; Moving phase: acetonitrile-water (15: 85) (phosphoric acid is transferred pH to 2.8); Detect wavelength: UV-detector, λ=254nm; Fluorimetric detector: λ ex=331nm, λ em=500nm; Flow velocity: 1ml/min; Column temperature: 25 ℃
The preparation of standard stock solution: precision takes by weighing Citrinin reference substance 14.64mg, dissolve with methanol and constant volume 10.0ml, and concentration is 1.464mg/ml, in contrast the product stock solution.
The preparation of reference substance solution: accurate absorption stock solution is a certain amount of, dilutes constant volume with methyl alcohol, is mixed with the standardized solution of different concns by the test demand.
High performance liquid chromatography
With reference to 2000 editions two ones of Chinese Pharmacopoeias (appendix VI D) high effective liquid chromatography for measuring.
Typical curve: accurate absorption Citrinin stock solution is a certain amount of, and being configured to concentration with moving phase is 7.32 μ g/ml, 21.96 μ g/ml, 65.88 μ g/ml, 144.94 μ g/ml, the standardized solution of 219.6 μ g/ml; All with 10 μ l sample introductions, with peak area standard substance concentration is carried out linear regression, regression equation is Y=1078+4098X, r=0.99989.Be that sample size is good in 7.00-220ng scope internal linear relation.
The mensuration of Citrinin in the red colouring agent for food, also used as a Chinese medicine sample
Adopt the Citrinin detection method of having set up that embodiment 1, embodiment 3 each 5 batch sample are detected result's following (table 3):
The content of unsaturated fatty acids, Citrinin in the table 3 Chinese medicine Antilipemic monascus
"-" do not detect
Isolating Citrinin is lemon-yellow needle-like prismatic crystallization from the Cai Shi nutrient solution of Penicillium citrinum (Penicillium citrinium), acidity.Fusing point: 172.12 ℃, decomposition point is 172.49 ℃; Specific optical rotation: [α]
23 5461-41.7 ,-43.7; Citrinin is insoluble in water, is dissolved in chloroform, acetone, ethyl acetate, ethanol, is slightly soluble in ether, and the sodium salt of Citrinin is soluble in water; Ultimate analysis: C 62.31%; H 5.56%; O 32.17%; Molecular formula: C
13H
14O
5Relative molecular mass is 251; Its molecular structural formula is as follows:
(3R, 4S)-4,6-dihydro-8-hydroxyl-3,4, the structure iron of 5-trimethylammonium-6-oxygen-3H-2-benzene pyrrole-7-carboxylic acid Citrinin
The production bacterial strain of Antilipemic monascus preparation of the present invention has the ability of not producing with utmost point low-yield citrinin through domestication, detects citrinin content in 10 batches of Chinese medicine Antilipemic monascus far below the regulation of Japan; We have also detected isolating 20 strain monascuses from environment simultaneously, and its content is higher, reaches 100-500 μ g/g, same culture condition, and different strains produces the Citrinin ability so difference, mainly is that the character of bacterial strain own determines.Demand to oxygen in the fermenting process is very big, therefore needs fully to support (improve air flow, accelerate stirring velocity).Wherein oxygen-supply quantity, temperature and nutrient media components are crucial, cross low and too high Ta Ting and the lipid acid output of all can having a strong impact on.
The preparation method of embodiment 8 Antilipemic monascus tablets of the present invention:
The material red colouring agent for food, also used as a Chinese medicine 273.0g that gets it filled, drying is pulverized, cross the 60-100 mesh sieve, add the fine little plain 37.45g micropowder silica gel 10.50g of crystallite, stearic acid 17.5g, lactose 10.50g, mix and pushed 14 mesh sieves, granulate, put 60 ℃ of dryings, cool cold, particle is crossed the whole grain of 16 mesh sieves, qualified particle adds Magnesium Stearate 1.05g, and mixing is pressed into 1000, film-making, promptly.
The capsular preparation method of embodiment 9 Antilipemic monascus of the present invention:
Red colouring agent for food, also used as a Chinese medicine 273g
Micropowder silica gel 7g
Make 1000
The material red colouring agent for food, also used as a Chinese medicine of getting it filled, drying is pulverized, and crosses the 60-200 mesh sieve, adds micro mist, and medicine and powder mixes is even, and the Capsules of packing into is made 1000, promptly.
Embodiment 10 Antilipemic monascus preparation of soft capsule methods of the present invention:
Red colouring agent for food, also used as a Chinese medicine 273g
Soybean oil 320g
Beeswax 7g
Make 1000
Get red colouring agent for food, also used as a Chinese medicine, drying was pulverized 80 mesh sieves, added soybean oil and beeswax, and mixing is made soft capsule, makes 1000 altogether, promptly.
The preparation method of embodiment 11 Antilipemic monascus micropills of the present invention:
Get Chinese medicine Antilipemic monascus 546g, drying is pulverized, and crosses the 60-100 mesh sieve, add auxiliary material 254g starch powder mixing, the ethanol of adding 70% is wetting agent, makes softwood, softwood micropill mechanism ball, wet feed pushed the 0.8mm sieve aperture, the wet grain of strip cuts off round as a ball, and drying and moulding is crossed 16～20 mesh sieves and selected ball.Dose is a 0.8g, 2 times on the one.
Below prove beneficial effect of the present invention by pharmacodynamics test, toxicology test.
Test example 1 Chinese medicine Antilipemic monascus preparation of the present invention is to the influence of rat food hyperlipidaemia
The contriver confirms that by the pharmacological testing comparative study to this Antilipemic monascus preparation and single lovastatin the treatment that this Antilipemic monascus preparation is used for hyperlipidaemia has significant more pharmacologically active.
Get 50 male SD rats and be divided into normal control group (10) and modeling group (totally 50) at random by empty stomach serum cholesterol and body weight.The modeling group is by following prescription: 5% lard, 2% cholesterol, 1% cholate, 92% basal feed are fed and were raised for 5 weeks, the 28th late fasting, cut tail and get blood morning next day, measure serum TC on an empty stomach, raise normal diet rat TC relatively with feeding, the difference significance is promptly determined modeling success (TC＞6.00mmol/l), select 40 altogether.Be divided into 4 groups (10 every group) by serum cholesterol and body weight principle at random then, promptly control group, hyperlipidemia model group, lovastatin group (2mg/kg), Chinese medicine Antilipemic monascus A03 organize (0.1g/kg); Administering mode is for irritating stomach, 1ml/100g, 1 time/day; During administration, except that the normal control group, its excess-three group is still given high lipid food.(medicine suspends with 0.5%CMC, and to be made into 8mg/ml concentration frozen standby in dissolving, faces with before adding 0.5%CMC and dilute, and is made into 0.2mg/ml concentration).
Table 4 couple hyperlipidemia rats serum blood fat TC, LDL-C, HDL-C, TG, influence
Compare with control group: * p＜0.05; * p＜0.01
Antilipemic monascus of the present invention reduces active, the active lovastatin that obviously is better than same dose of HDL-C rising to hyperlipidemia rats serum blood fat TC, LDL-C, TG.
The comparison of test example 2 Chinese medicine Antilipemic monascus preparations of the present invention and lovastatin treatment hyperlipidaemia:
All patients who meets the hyperlipidaemia Case definition, keep usual diet, twice of two all class blood sampling, as detect twice serum cholesterol 〉=6.5mmol/L or serum triglyceride 〉=1.8mmol/L or high density lipoprotein cholesterol (HDL-C) the value male sex≤1.04mmol/L, women≤1.17mmol/L, Chinese medical discrimination belongs to that temper is double holds that phlegm is turbid, the syndrome of blood stasis person under the arm, is selected object.Get rid of following situation: once suffered from Acute Myocardial Infarction, cerebrovascular accident in (1) half a year, severe trauma or capital operation, pregnant woman and lactating women.(2) nephrotic syndrome, thyroprivia, chronic liver and gall diseases patient of acute favour and diabetic subject.(3) by medicine (as phenothiazines, beta-Blocking agent, adrenocortical steroid and some contraceptive bian etc.) hyperlipidaemia that causes and the subtype hypercholesterolemiapatients patients of isozygotying.(4) using heparin, Tiroidina curative and other to influence the patient of blood lipid metabolism medicine and the patient that nearly two all classes once adopted other lipopenicillinase measures.150 examples that meet inclusion criteria are divided into two groups, control group 1 (lovastatin group 20mg/ days, every day a slice, continuously around), treatment group 1 (Antilipemic monascus of the present invention, total lovastatin content be less than 12mg/ days, clothes at twice, continuous 4 weeks).
Treatment group 2 (Antilipemic monascus of the present invention, total lovastatin content is 20mg/ days, obeys continuous 4 weeks at twice)
Curative effect determinate standard: (1) produce effects: reach following any 1 person, TC descends 〉=20%; TG descends 〉=40%; HDL-ch rising 〉=0.26mmol/L; AI descends 〉=20%.(2) effective: TC decline 〉=10-19%; TG decline 〉=20-39%; HDL-ch rising＞0.104mmol/L-0.25mmol/L; AI decline 〉=10-19%.(3) invalid: as not reach effective standard.The result is as follows:
The total effective rate of table 5 Chinese medicine Antilipemic monascus of the present invention preparation and lovastatin treatment hyperlipidaemia
Above clinical study shows that treatment group 1 (every day dose in lovastatin amount less than 12mg) curative effect is equivalent to take every day lovastatin tablet 20mg approximately, and treatment is organized 2 TG total effective rates and obviously is better than control group.Content of polyunsaturated fatty acid is far smaller than effective dose (thousandth that is about effective dose) in the red colouring agent for food, also used as a Chinese medicine, the lipopenicillinase activity of Antilipemic monascus of the present invention obviously is better than the lovastatin of same dose, illustrates that the various active composition has synergistic function in the Antilipemic monascus of the present invention.Especially the effect that has overcome the statins triglyceride reducing is not ideal enough, needs the shortcoming of drug combination.
The clinical trial explanation, pharmaceutical composition of the present invention has the effect of treatment fatty liver.
Test of the influence of example 3 Antilipemic monascus tablets of the present invention to fatty liver:
Collect 35 routine Patients with Fatty Liver (Case definition: ultrasound diagnosis diagnosis fatty liver, reference " practical digestion is sick to be learned "; Laboratory examination is with reference to " new Chinese medicine clinical guidance principle ", triglyceride level (TG)＞1.7mmol/L, cholesterol (TC)＞5.7mmol/L, serum glutamic pyruvic transminase (ALT)＞60IU/L), take the capsule of embodiment 1, usage is each 3, twice on the one, 4 weeks were a course of treatment, treated 6 courses of treatment continuously.
Curative effect determinate standard: formulate standard with reference to " new Chinese medicine clinical guidance principle ", " practical digestion is sick to be learned " and pertinent literature: recovery from illness: subjective symptoms disappearance, TC≤5.7mmol/L, TG≤1.7mmol/L/L, ALT≤60IU/L, B ultrasonic show liver sign liver luminous point, its back court echo recovery normally.Produce effects: symptom is clearly better, TC descends＞and 20%, TG descends＞40%, and ALT descends 〉=50% or normal, and B ultrasonic liver densities of points of light is clearly better, and echo attenuation thereafter obviously alleviates.Effectively: symptom take a favorable turn, TC decline 10%-20%, and TG decline 20%-40%, ALT descend＜50% or normal, the more preceding improvement of B ultrasonic liver sign.Invalid or worsen: subjective symptoms still exists or increases the weight of, TC and (or) TG and (or) ALT and (or) B ultrasonic acoustic inspection no change or increase the weight of.
Treatment result: treat 35 examples, 5 examples of fully recovering, produce effects 12 examples, effective 11 examples, invalid 7 examples, total effective rate 80%.Blood fat obtains desirable control as a result, and it is normal that ALT, AST and cholinesterase index are recovered, and it is normal that the patient of B ultrasonic image 60% recovers substantially.
The clinical trial explanation, the effect with treatment fatty liver of the present invention.
The acute toxicity test of test example 4 Antilipemic monascus tablets of the present invention
Get the ICR19-21g mouse and (provide animal rank: one-level by the military region, Chengdu medicine group first research department's Pharmacology Lab.Conformity certification number: real moving Guan Zhidi-No. 1 of river) 30, male and female half and half, successive administration is twice in 8 hours, each gastric infusion 0.8ml (Antilipemic monascus tablet of outward appearance reddish-brown of the present invention, it is standby to be made into the 0.3g/ml soup with 0.7%CMC-Na) be equivalent to 400 times of clinical dosages, observe a week continuously after the administration, see that animal has or not death.
By above test as seen, animal one day administration 0.48g-20g (being equivalent to 400 times of clinical dosages) observes none death of week continuously, there are not significantly other toxic reactions, animal in order, and we can not increase administration volume and concentration again, so think that Chinese medicine red colouring agent for food, also used as a Chinese medicine of the present invention is that a kind of toxicity is little, safe drugs.
By above-mentioned pharmacodynamics test and toxicological test, prove absolutely mutual synergy between each component of Chinese medicine red colouring agent for food, also used as a Chinese medicine of the present invention, obviously be better than lovastatin, to the those skilled in the art, the drug effect of Chinese medicine red colouring agent for food, also used as a Chinese medicine of the present invention is not apparent.Craft science of the present invention is reasonable, adopts the drug use dosage of preparation of drug combination method gained of the present invention little, and raw materials cost is low, and clinical efficacy is remarkable, and toxic side effect is little, and is easy to use, and steady quality is controlled, and its potency ratio meets national conditions.
Red colouring agent for food, also used as a Chinese medicine provided by the invention belongs to the new bacterial strain of monascus parpureus Went, be by " China. Wuhan. Wuhan University ", the typical culture collection center C CTCC of China preservation, preservation date on July 16th, 2005, preserving number is CCTCC M 205073, the bacterial strain of classification called after Monascus Purpureus Went Diao 5.029.
Claims (17)
1. red koji strain is characterized in that: described red koji strain is that the preserving number by China typical culture collection center C CTCC preservation is the bacterial strain of CCTCC M 205073.
2. red koji strain according to claim 1, described red koji strain is monascus parpureus Went (MonascusPurpureus Went), the lovastatin 15-30mg/g that contains the open and close ring structure in its tunning, unsaturated fatty acids is 3-6mg/g, and wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
3. red koji strain according to claim 2 is characterized in that: the citrinin content in the described strain fermentation product is smaller or equal to 100ppb.
4. Chinese medicine red colouring agent for food, also used as a Chinese medicine, it is that the preparation method comprises the steps: by the arbitrary described red koji strain fermentative preparation of claim 1-3
A, seed liquor preparation
Monascus parpureus Went (Monascus Purpureus Went), deposit number by China typical culture collection center (CCTCC) preservation is the bacterial strain of CCTCC M 205073, use seed as producing, in 30～40 ℃, 180～220rpm cultivates 8～24h as first order seed or fermentor tank seed liquor in the PDA substratum;
B, fermentation: adopt liquid state fermentation or solid state fermentation, drying gets red colouring agent for food, also used as a Chinese medicine.
5. Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 4 is characterized in that: contain unsaturated fatty acids 3-6mg/g, and lovastatin 15-30mg/g, wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
6. Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 5 is characterized in that: contain following main effective constituent: unsaturated fatty acids 3-6mg/g, and lovastatin 20-30mg/g, wherein to account for the per-cent of total lovastatin be 10-90% to the lovastatin of open loop structure.
7. Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 6 is characterized in that: contain following main effective constituent: unsaturated fatty acids 3-6mg/g, and lovastatin 20-25mg/g, wherein to account for the per-cent of total lovastatin be 50-90% to the lovastatin of open loop structure.
8. according to each described Chinese medicine red colouring agent for food, also used as a Chinese medicine of claim 4～7, it is characterized in that: the amount of polyunsaturated fatty acid is 2-5mg/g in the described unsaturated fatty acids.
9. Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 8 is characterized in that: described unsaturated fatty acids is mainly oleic acid and linolic acid.
10. according to each described Chinese medicine red colouring agent for food, also used as a Chinese medicine of claim 4～7, it is characterized in that: the Chinese medicine red colouring agent for food, also used as a Chinese medicine contains Citrinin smaller or equal to 100ppb.
11. a method for preparing the described Chinese medicine red colouring agent for food, also used as a Chinese medicine of claim 4, it comprises the steps:
A, seed liquor preparation
Monascus parpureus Went (Monascus Purpureus Went), deposit number by China typical culture collection center (CCTCC) preservation is the bacterial strain of CCTCC M 205073, use seed as producing, in 30～40 ℃, 180～220rpm cultivates 8～24h as first order seed or fermentor tank seed liquor in the PDA substratum;
B, fermentation: adopt liquid state fermentation or solid state fermentation, drying gets red colouring agent for food, also used as a Chinese medicine.
12. the preparation method of Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 11 is characterized in that: the culture medium prescription of the described liquid state fermentation of step b is:
Carbon source 10～40g/L, nitrogenous source 10～45g/L, extractum carnis 0.5～3g/L, CaCl
20.05～1g/L, pH 6.5 ± 1.0,121 ℃ of sterilization 20min, and wherein, carbon source is sucrose, glucose, glycerine, Zulkovsky starch or molasses, the N source is corn steep liquor, yeast extract paste, extractum carnis, plant or animal proteinum peptone;
Fermentation process is: the seed liquor for preparing is inserted in the fermention medium according to 5%～20% inoculum size, fermentation culture is based on 30～40 ℃, 180～220rpm cultivates 5d～7d, finishing 24h from cultivating, add open loop stabilization liquid, the centrifugal supernatant of removing, mycelium fragmentation, dose auxiliary material, drying obtains Hongqu powder (red colouring agent).
13. the preparation method of Chinese medicine red colouring agent for food, also used as a Chinese medicine according to claim 11 is characterized in that: the component of the nutritive medium of the described solid state fermentation of step b is: glycerine 50～300g/L, peptone 50～300g/L, extractum carnis 50～100g/L, Na
2HPO
412H
2O 5～20g/L, NaH
2PO
42H
2O 5～20g/L, CaCl
20.5～10g/L, MgSO
47H
2O0.5～10g/L, Na
2CO
30.5～10g/L;
Its fermentation process is: nutritive medium is sprayed in polished rice sterilization back, weight proportion is that every 1000g rice sprays nutritive medium 20～120ml, obtain substratum in 100 ℃ of boilings, after the cooling seed liquor for preparing inserted according to 5%～20% inoculum size and be mixed with in the substratum of nutritive medium; Cultivation is based on 30～45 ℃, humidity 〉=80%, and 8d and 12d add the nutritive medium of solid state fermentation by liquid-solid ratio 5% after cultivation respectively, finished in back 14 days in fermentation, high-temperature sterilization, drying obtains red colouring agent for food, also used as a Chinese medicine.
14. a pharmaceutical composition, it is to be activeconstituents by each described red colouring agent for food, also used as a Chinese medicine of claim 4-10, and Hongqu powder (red colouring agent) is broken to the 80-200 order, adds the medicament that acceptable accessories or complementary composition are prepared from.
15. pharmaceutical composition according to claim 14 is characterized in that: described medicament is: tablet, capsule, dispersion agent, micronization preparation or micropill.
16. the purposes of the described pharmaceutical composition of claim 14 in the medicine of preparation treatment or preventing hyperlipidemia.
17. the purposes of the described pharmaceutical composition of claim 14 in the medicine of preparation treatment fatty liver.
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| CN102925509A (en) * | 2012-11-14 | 2013-02-13 | 广东药学院 | Method for increasing output of microbial lovastatin based on quorum sensing mechanism |
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| CN102010831B (en) * | 2010-05-26 | 2012-04-18 | 云南农业大学 | Lovastatin-generating fungus and application thereof in production of Pu'er tea |
| CN102199544B (en) * | 2011-03-18 | 2012-07-04 | 天津科技大学 | Monascus sp. M3 strain with efficient cholesterol degrading capability |
| CN102885303B (en) * | 2012-10-09 | 2014-04-02 | 西藏月王生物技术有限公司 | High-r-aminobutyric-acid-content highland-barley red yeast and preparation method thereof |
| CN103933078A (en) * | 2013-01-20 | 2014-07-23 | 杭州桐君堂生物科技有限公司 | Traditional Chinese medicine red yeast rice producing technology |
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| CN103829090A (en) * | 2014-02-21 | 2014-06-04 | 四川农业大学 | Application of red yeast rice to reduction of methane discharged by ruminant such as sheep |
| CN104099254B (en) * | 2014-07-11 | 2017-02-15 | 无锡超科食品有限公司 | Bacterial strain for producing polyunsaturated fatty acids and screening method thereof |
| CN104922160A (en) * | 2015-05-22 | 2015-09-23 | 舟山三合生物科技有限公司 | Composition prepared by compounding statin with Omega-3 fatty acid and coenzyme Q10 and application of composition |
| CN108342327B (en) * | 2018-03-09 | 2021-06-01 | 福建省微生物研究所 | Monascus ruber and application thereof in preparation of lipid-lowering drugs |
| CN109517740B (en) * | 2018-12-05 | 2020-11-27 | 广东天益生物科技有限公司 | Monascus purpureus mutant strain, water-soluble functional red yeast and preparation method and application thereof |
| CN109793217A (en) * | 2019-01-29 | 2019-05-24 | 北京工商大学 | A kind of preparation method of red yeast rice sea-buckthorn seed cake powder |
| CN112852646B (en) * | 2021-03-17 | 2023-06-13 | 江南大学 | Monascus purpureus H5-3 with high lovastatin yield and application thereof |
| CN118546790A (en) * | 2023-02-24 | 2024-08-27 | 北京北大维信生物科技有限公司 | Monascus mutant strain and method for preparing monascus or monacolin K |
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