CN107619803A - A kind of preparation of the sour and dark yellow strepto- ketone of dark yellow strepto- and its medical usage - Google Patents
A kind of preparation of the sour and dark yellow strepto- ketone of dark yellow strepto- and its medical usage Download PDFInfo
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Abstract
A kind of very dark yellow streptomycete provided by the invention, the separation acquisition from marine animal taeniae flesh sea anemone (Haliplanella lineata).Classification And Nomenclature is Streptomycesfulvissimus ZZ406, by the center preservation of China typical culture collection center Wuhan, deposit number:CCTCC M 2017435, preservation day：On August 3rd, 2017.The dark yellow strepto- of compound sour (1) and dark yellow strepto- ketone (2) obtained using the very dark yellow streptomycete has anticol matter tumor activity, the propagation of different brain glioblastoma cells and the obvious protein expression level for reducing glioma cell glycolysis metabolism key enzyme can significantly be suppressed, Anticancer Effect and Mechanism with uniqueness, the application in treating glioma medicine can prepared.The structural formula of the dark yellow strepto- of compound sour (1) and dark yellow strepto- ketone (2) is as follows.
Description
Technical field
The invention belongs to field of medicaments, is related to from the very dark yellow streptomycete of marine actinomycete
(StreptomycesfulvissimusZZ406) the sour and dark yellow strepto- ketone of the dark yellow strepto- of active compound for anti tumor is prepared in
Method and they prepare treat glioma medicine in terms of application.
Background technology
Glioma is most common in the brainpan and high brain tumor of the death rate.Because glioma is located at the important work(of many brains more
Energy area so that operating difficulty is big and is not easy tumour complete resection, so the treatment of radiotherapy and medicine to glioma is more
It is important.But anticol matter tumor medicine famine at present, mainly including traditional card chlorine mustard (carmustine), Lomustine
(lomustine), the minority such as procarbazine (procarbazine) and the Temozolomide (temozolomide, TMZ) of the second generation
Anticol matter tumor medicine, wherein Temozolomide are unique selectable first-line drugs for being individually used for Treatment for Glioma.Moreover, including
Existing most treatment colloid tumor medicines including Temozolomide are all cytotoxic alkylating agents, how effective in cure limited, toxic side effect
Big and serious drug resistance deficiency.Therefore, clinically there is an urgent need to better efficacy, that security is higher and mechanism of action is unique is new
Type anticol matter tumor medicine.Secondary metabolite from marine microorganism is antineoplastic or antineoplastic lead compound hair
Existing valuable source.Tumour cell metabolism reprogramming provides new target drone for the research and development of new anticol matter tumor medicine.
The excessive glycolysis (Glycolysis) of tumour cell is that tumour is thin to meet the quick unconfined propagation of its cell
Born of the same parents are different from one of protrusion metabolic characteristics of normal cell.Tumour cell absorbs a large amount of glucose in glycolysis ring from microenvironment
Section is specifically relied on by glioma cell and multiple key enzymes of high expression include Hexokinase 2 (HK2), phosphofructokinase/fruit
Sugared 2,6- diphosphatases (PFKFB3), pyruvate kinase M2 (PKM2) and lactic dehydrogenase 5 (LDH5) etc. participate in producing greatly jointly
It is the important initiation material of tumour cell synthesising biological macromolecular to measure intermediate product and end-product lactic acid, these intermediate products, and
Lactic Acid Secretion suppresses Scavenging activity of the immune cell to tumour cell to extracellular, tumour cell can be promoted to spread, high
The glycolysis key enzyme HK2 of expression can also improve tumour cell to radiotherapy and the resistance of Temozolomide chemotherapy.It can be appreciated that it is exactly
Tumour cell height is promoted using the ability of its metabolic intermediate synthesising biological macromolecular DNA, RNA, protein and biomembrane
It is quickly unrestrictedly bred, and the special key enzyme of modulate tumor cell glycolysis can effectively suppress the propagation of tumour cell.Cause
This, the medicine that target tumor is metabolized the multiple target effect of special key enzyme may have more preferable antitumor curative effect.Dark yellow strepto-
Sour and dark yellow strepto- ketone is separated from the very dark yellow streptomycete of marine bacteria (StreptomycesfulvissimusZZ406)
Two noval chemical compounds arrived, their propagation to a variety of different glioma cells have significant inhibitory action, can significantly reduced
Important special key enzyme HK2, PFKFB3, PKM2 and LDH5 of glioma cell glycolysis protein expression level, have uniqueness
Anticancer Effect and Mechanism, therefore, the sour and dark yellow strepto- ketone of dark yellow strepto- in terms of preparing anticol matter tumor medicine have application before
Scape.
The content of the invention
It is an object of the invention to provide a kind of very dark yellow streptomycete, Classification And Nomenclature Streptomycesfulvissimus
ZZ406, by China typical culture collection center-Wuhan center preservation, deposit number:CCTCC M 2017435, preservation
Day：On August 3rd, 2017.The very dark yellow streptomycete is from marine animal taeniae flesh sea anemone (Haliplanella lineata)
Separation, is separately cultured by following steps to obtain：
(1) very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) is separately cultured
A certain amount of fresh taeniae flesh sea anemone (Haliplanella lineata) is taken to clean three with the natural sea-water of sterilizing
It is homogenized after secondary, the suspension of various concentrations is made.The uniform suspension of a certain amount of various concentrations is taken to be distributed to containing solid culture
In the culture dish of base, after cultivating certain time at ambient temperature, different bacterium colonies is transferred to respectively another containing solid training
In the culture dish for supporting base, continue to cultivate certain time at ambient temperature.Finally well-grown single bacterium colony (ZZ406) is connect
Kind saves backup to the rearmounted 4 DEG C of refrigerators of slant medium culture.
Described taeniae flesh sea anemone (Haliplanella lineata) is obtained from the rock seam at Zhejiang Mount Putuo seabeach
, natural sea-water is obtained from the Area of The East China Sea near Zhoushan Zhejiang；The various concentrations of the suspension are 1 × 10-4~1
×10-1g/mL；The sampling amount of described sample suspension is 100~300 μ L；Solid medium contained by the culture dish is height
Family name's agar (Gause ' s agar) culture medium or other solid mediums；Described slant medium be Gao Shi agar mediums or
Other solid slope culture mediums；Described incubated at room temperature temperature is 22~28 DEG C；Described incubation time is 5~15 days.
(2) strain idenfication of very dark yellow streptomycete (Streptomycesfulvissimus ZZ406)
Above-mentioned steps (1) are separately cultured the 16S rDNA sequences that obtained bacterial strain ZZ406 is generally used with current laboratory
Row analysis method identifies its species, is defined as very dark yellow streptomycete, Classification And Nomenclature Streptomycesfulvissimus
ZZ406, by China typical culture collection center-Wuhan center preservation, deposit number:CCTCC M 2017435.
Second object of the present invention is to provide two dark yellow strepto-s of compounds sour (1) with anticol matter tumor activity and secretly
Yellow strepto- ketone (2), the sour and dark yellow strepto- ketone of described dark yellow strepto- is noval chemical compound, and its chemical structural formula is：
Third object of the present invention is to provide the preparation method of dark yellow strepto- sour (1) and dark yellow strepto- ketone (2), by with
Lower step is realized：
(1) preparation of very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) zymocyte liquid
The colony inoculation of very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) is taken to containing a certain amount of
In the big conical flask of liquid spawn culture medium, by the nutrient solution containing ZZ406 strains, shaken cultivation is certain at ambient temperature
Strain liquid is obtained after time.Finally strain liquid is transferred in the big conical flask containing a certain amount of liquid fermentation medium,
Under room temperature condition after shaken cultivation certain time, the ZZ406 nutrient solution with antitumor activity is obtained.
Described bacterium culture medium is liquid Gao Shi culture mediums, and described dosage is 150-250mL；Described liquid fermentation
Culture medium is SC fluid nutrient mediums (10 grams of soluble starch, 0.3 gram of casein, 2 grams of potassium nitrate, 0.5 gram of magnesium sulfate, phosphoric acid hydrogen
2 grams of dipotassium, 0.02 gram of calcium carbonate, 0.01 gram of ferrous sulfate, 1 liter of natural sea-water, pH value 7.2)；Described big triangle blake bottle is
500mL；Described incubated at room temperature temperature is 22~28 DEG C；The rotating speed of the vibration is 160~180rpm；Described incubation time
For 5~15 days.
(2) extraction separation and purification of the sour and dark yellow strepto- ketone of dark yellow strepto-
Bacterial strain ZZ406 fermentation mycelium is extracted with methanol, and methanolic extract is extracted with ethyl acetate to obtain acetic acid second again
Ester extract.Acetic acid ethyl ester extract is first separated with normal-phase silica gel column chromatography, with ethyl acetate and hexamethylene mixed solvent gradient
Elution, and with thin layer chromatography analysis, obtain nine components (Fr.1~Fr.9).Component Fr4 and component Fr 8 is again respectively with preparation
Type high performance liquid chromatography (HPLC) instrument separates, and obtains the dark yellow strepto- of pure compound sour (1) and dark yellow strepto- ketone (2).
The purification on normal-phase silica gel dosage of the column chromatography and the sample size ratio of upper amount are 30~50g:1.0g；Described efficient liquid
Phase separation is:Permanent CXTH-3000 high performance liquid chromatographs, Fuji C are led in innovation18CT-30 chromatographic columns (280 × 30mm, 10 μ
M), first alcohol and water is mobile phase, Detection wavelength 220nm, flow velocity 10.0mL/min.
(3) Structural Identification of the sour and dark yellow strepto- ketone of dark yellow strepto-
The structure of the sour and dark yellow strepto- ketone of dark yellow strepto- is a peacekeeping two-dimentional NMR spectrum, the high resolution mass spectrum according to them
The methods of data and ECD are calculated and determine.
Fourth object of the present invention is to provide the sour and dark yellow strepto- ketone of dark yellow strepto- and is preparing treatment glioma medicine
In application.The sour and dark yellow strepto- ketone of described dark yellow strepto- can significantly inhibit the propagation of a variety of glioma cells, reduce glue
The protein expression of multiple key enzymes in matter oncocyte glycolytic pathway, there is unique Anticancer Effect and Mechanism.
The medicine is that dark yellow strepto- is sour or dark yellow strepto- ketone is independent, or the combination of both, or alone or in combination with
Other active materials together, with pharmaceutically acceptable carrier composition medicine.
Very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) provided by the invention, indulges from marine animal
Separation obtains in bar flesh sea anemone (Haliplanella lineata).Research shows, utilizes the change of very dark yellow streptomycete acquisition
The dark yellow strepto- of compound sour (1) and dark yellow strepto- ketone (2) have anticol matter tumor activity, can significantly suppress different brain glioblastoma cells
Propagation and the obvious protein expression level for reducing glioma cell glycolysis metabolism key enzyme, there is unique antitumor action
Mechanism, the application in treating glioma medicine can prepared.
Brief description of the drawings
Fig. 1 is very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) bacterium colony figure.
Fig. 2 is the hydrogen spectrum of dark yellow strepto- acid.
Fig. 3 is the carbon spectrum of dark yellow strepto- acid.
Fig. 4 is the HMBC spectrums of dark yellow strepto- acid.
Fig. 5 is the main HMBC accompanying drawings of dark yellow strepto- acid.
Fig. 6 is the high resolution mass spectrum of dark yellow strepto- acid.
Fig. 7 is the hydrogen spectrum of dark yellow strepto- ketone.
Fig. 8 is the carbon spectrum of dark yellow strepto- ketone.
Fig. 9-10 is dark yellow strepto- ketone1H-1HCOSY is composed.
Figure 11 is the hsqc spectrum of dark yellow strepto- ketone.
Figure 12 is the HMBC spectrums of dark yellow strepto- ketone.
Figure 13 is that dark yellow strepto- ketone is main1H-1H COSY and HMBC accompanying drawings.
Figure 14 is the high resolution mass spectrum of dark yellow strepto- ketone.
Figure 15 be the sour and dark yellow strepto- ketone of dark yellow strepto- to U87-MG cell different metabolic key enzymes HK2, PFKFB3,
The influence of PKM2, LDH5, ACO2 and CytoC protein expression level.
Embodiment
The present invention is described in further detail below in conjunction with drawings and examples.But the invention is not restricted to these realities
Apply example.
Embodiment 1
Very dark 1. yellow streptomycete (Streptomycesfulvissimus ZZ406's) is separately cultured
Fresh 5 grams of taeniae flesh sea anemone (Haliplanella lineata) is taken, after being cleaned three times with the natural sea-water of sterilizing
Homogenate, concentration is made as 1 × 10-1G/mL suspension.By 1 × 10-1G/mL suspension is diluted to concentration as 1 × 10 successively-2、1×10-3With 1 × 10-4G/mL suspension.The μ L of suspension 200 of each concentration are taken to evenly spread to containing Gao Shi agar
In the culture dish of (Gause ' s agar) solid medium, after being cultivated 5 days under the conditions of 28 DEG C, different bacterium colonies is shifted respectively
Into another culture dish containing Gao Shi agar (Gause ' sagar) solid medium, continue 5 days under the conditions of 28 DEG C.Finally
It is standby that well-grown single bacterium colony (ZZ406) is inoculated into the rearmounted 4 DEG C of refrigerators preservation of Gao Shi agar solid slope culture medium cultures
With.
2. the strain idenfication of very dark yellow streptomycete (Streptomycesfulvissimus ZZ406)
Bacterial strain ZZ406 species is obtained using the identification of 16S rDNA sequence analysis methods.
2.1 experiment reagents and instrument
PCR reagent：EX Taq enzymes (TaKaRa), dNTP (TaKaRa), primer (Invitrogen synthesis), primer sequence
It is:
SEQ ID No：1：AGAGTTTGATCCTGGCTCAG
SEQ ID No：2：GGTTACCTTGTTACGACTT
Marker：DL5000
Laboratory apparatus：Centrifuge, electrophoresis apparatus, PCR instrument, ABI 3730XL sequenators.
2.2 experimental procedure
Bacterial genomes DNA is extracted
Electrophoresis detection
PCR is expanded
A.PCR reaction systems
B.PCR reaction conditions
C. electrophoresis detection
D. it is sequenced:Cut glue purification sequencing
E. analysis result:Splice sequence.
2.3 experimental result
Spliced sequence is SEQ ID No：3：
TGCAGTCGACGATGAAGCCGCTTCGGTGGTGGATTAGTGGCGAACGGGTGAGTAACACGTGGGCAATCT
GCCCTTCACTCTGGGACAAGCCCTGGAAACGGGGTCTAATACCGGATAATACTCCTGCCTGCATGGGTGGGGGTTGA
AAGCTCCGGCGGTGAAGGATGAGCCCGCGGCCTATCAGCTTGTTGGTGGGGTAATGGCCTACCAAGGCGACGACGGG
TAGCCGGCCTGAGAGGGCGACCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGA
ATATTGCACAATGGGCGAAAGCCTGATGCAGCGACGCCGCGTGAGGGATGACGGCCTTCGGGTTGTAAACCTCTTTC
AGCAGGGAAGAAGCGCAAGTGACGGTACCTGCAGAAGAAGCGCCGGCTAACTACGTGCCAGCAGCCGCGGTAATACG
TAGGGCGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGAGCTCGTAGGCGGCTTGTCACGTCGGATGTGAAAGCCCG
GGGCTTAACCCCGGGTCTGCATTCGATACGGGCTAGCTAGAGTGTGGTAGGGGAGATCGGAAATTCCTGGTGTAGCG
GTGAAATGCGCAGATATCAGGAGGAACACCGGTGGCGAAGGCGGATCTCTGGGCCATTACTGACGCTGAGGAGCGAA
AGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGTTGGGAACTAGGTGTTGGCGACATTC
CACGTCGTCGGTGCCGCAGCTAACGCATTAAGTTCCCCGCCTGGGGAGTACGGCCGCAAGGCTAAAACTCAAAGGAA
TTGACGGGGGCCCGCACAAGCAGCGGAGCATGTGGCTTAATTCGACGCAACGCGAAGAACCTTACCAAGGCTTGACA
TATACCGGAAAGCATCAGAGATGGTGCCCCCCTTGTGGTCGGTATACAGGTGGTGCATGGCTGTCGTCAGCTCGTGT
CGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTTCTGTGTTGCCAGCATGCCTTTCGGGGTGATGGG
GACTCACAGGAGACTGCCGGGGTCAACTCGGAGGAAGGTGGGGACGACGTCAAGTCATCATGCCCCTTATGTCTTGG
GCTGCACACGTGCTACAATGGCCGGTACAATGAGCTGCGATGCCGTGAGGCGGAGCGAATCTCAAAAAGCCGGTCTC
AGTTCGGATTGGGGTCTGCAACTCGACCCCATGAAGTCGGAGTTGCTAGTAATCGCAGATCAGCATTGCTGCGGTGA
ATACGTTCCCGGGCCTTGTACACACCGCCCGTCACGTCACGAAAGTCGGTAACACCCGAAGCCGGTGGCCCAACCCC
TTGTGGGAGGGA
Compared with U.S. NIH NCBI GenBank databases, its result shows 16S rDNA sequences achieved above:Bacterium
Strain ZZ406 16S rDNA sequences and the 16S of the Streptomyces fulvissimus DSM 40593 in GenBank storehouses
RDNA sequences have 99% similitude (accession number:NC 021177.1).Therefore, the marine bacteria strain ZZ406 that the present invention is obtained determines
For very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) (accompanying drawing 1).The very dark yellow streptomycete obtained
The bacterial strain of (Streptomycesfulvissimus ZZ406) by China typical culture collection center-Wuhan center preservation,
Deposit number:CCTCC M 2017435, preservation day：On August 3rd, 2017.
The preparation of 2 very dark yellow streptomycete of embodiment (Streptomycesfulvissimus ZZ406) zymocyte liquid
Take the very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) of Gao Shi agar solid slope culture mediums
Bacterium be inoculated into the 500mL conical flasks containing 250mL liquid Gao Shi culture mediums, the nutrient solution containing ZZ406 strains is existed
Strain liquid is obtained after (180rpm) shaking culture being rotated under the conditions of 28 DEG C 5 days.5mL strain liquids are transferred to the SC containing 250mL
Fluid nutrient medium (10 grams of soluble starch, 0.3 gram of casein, 2 grams of potassium nitrate, 0.5 gram of magnesium sulfate, 2 grams of dipotassium hydrogen phosphate, carbon
Sour 0.02 gram of calcium, 0.01 gram of ferrous sulfate, 1 liter of natural sea-water, pH value 7.2) 500mL conical flasks in, under the conditions of 28 DEG C
(180rpm) shaking culture 13 days is rotated, obtains the ZZ406 zymocyte liquid with antitumor activity.
Embodiment 3
1. the extraction separation and purification of the sour and dark yellow strepto- ketone of dark yellow strepto-
The zymocyte liquid (70.0 liters) that embodiment 2 obtains is filtrated to get mycelium.Mycelium is obtained three times with methanol extraction
Methanolic extract.Methanolic extract is extracted with ethyl acetate to obtain acetic acid ethyl ester extract (8.0 grams), acetic acid ethyl ester extract again
With silica gel (250 grams) column chromatography for separation, with hexamethylene and ethyl acetate (4:1 to 1:1) gradient elution, it is each with thin layer chromatography analysis
Eluent, merges the component containing identical component, and 9 components (Fr.1~Fr.9) are always obtained.(30 milligrams) use of component Fr.4
Semipreparative high performance liquid chromatography instrument separates (instrument：Permanent CXTH-3000 is led in innovation；Chromatographic column：Fuji C18CT-30,280×
30mm,10μm；Mobile phase：Methanol/water, 75/25；Detection wavelength:220nm, flow velocity:10.0mL/min), it is dark yellow to obtain compound
Strepto- acid (1,9.3mg, tR31min).Equally, with identical high performance liquid chromatography separation condition (but different mobile phase:First
Alcohol/water, 21/79) separation component Fr.8 (20mg) obtains dark yellow strepto- ketone (2,8.0mg, the t of pure compoundR 35min)。
2. the Structural Identification of the sour and dark yellow strepto- ketone of dark yellow strepto-
Dark yellow strepto- is sour (1)：Yellow powder；Molecular formula C16H10O6；UV absorption UV (MeOH) λmax(logε)217
(4.32),279(4.22),411(3.66)nm；High resolution mass spectrum (HRESIMS) is m/z [M-H]-297.0407 (calculated values
C16H9O6297.0399).According to dark yellow strepto- acid1H spectrums (accompanying drawing 2),13C spectrum (accompanying drawing 3), HMBC spectrum (accompanying drawing 4, accompanying drawing 5) and
High resolution mass spectrum (accompanying drawing 6), it is determined that the structure of dark yellow strepto- acid, is a noval chemical compound, its13C and1H NMR signals belong to
It is shown in Table one.
Table one, dark yellow strepto- acid13C and1H NMR data (solvents：Deuterated dimethyl sulfoxide DMSO-d6)
Dark yellow strepto- ketone (2)：Yellow powder；Molecular formula C16H16O6；[α]D 25-5.6(c 0.20,MeOH)；UV absorption UV
(MeOH)λmax(logε)224(4.09),302(3.63)nm；High resolution mass spectrum (HRESIMS) is m/z [M+H]+305.0998
(calculated value C16H17O6305.1025).According to dark yellow strepto- ketone1H spectrums (accompanying drawing 7),13C spectrums (accompanying drawing 8),1H-1H COSY are composed
(accompanying drawing 9, accompanying drawing 10, accompanying drawing 13), hsqc spectrum (accompanying drawing 11), HMBC spectrums (accompanying drawing 12, accompanying drawing 13), high resolution mass spectrum (accompanying drawing 14)
Compose and calculate with ECD, it is determined that the structure of dark yellow strepto- ketone, it is a noval chemical compound, dark yellow strepto- ketone13C and1H NMR signals
Ownership is shown in Table two.
Table two, dark yellow strepto- ketone (2)13C and1H NMR data (solvents：Deuterated dimethyl sulfoxide DMSO-d6)
The activity of the sour and dark yellow strepto- ketone of 4 dark yellow strepto- of embodiment
1. the sour and dark yellow strepto- ketone of dark yellow strepto- suppresses the effect of glioma
Human glioma U251 cells DMEM and 10%FBS culture mediums, and glioma U87MG cells, SHG44 cells and
Normal human glial cell HA uses MEM culture mediums, RPMI-1640 culture mediums and AM culture mediums respectively.All cells at 37 DEG C and
In the incubator of 5% carbon dioxide the experimental study for the present invention is cultivated by three generations.
Suppression of the dark yellow sour and dark yellow strepto- ketone of strepto- to tumor cell proliferation is determined with Sulforhodamine B (SRB) method to live
Property, adriamycin compares for positive drug.Cell is inoculated in 96 orifice plates, and the test compound of various concentrations is added after adherent 24h,
Dyed after incubator culture 72h with SRB, with the absorbance value at ELIASA measure 515nm, detect the survival of tumour cell
Rate, calculate the IC that each test compound suppresses glioma50Value.Test result indicates that：The sour and dark yellow chain of dark yellow strepto-
Mould ketone significantly inhibits the propagation of three kinds of different glioma cells, its IC50It is worth for 4.7-25.8 μM, wherein the activity of dark yellow strepto- acid
It is stronger, its IC50It is worth for 4.7-8.1 μM (table three).It is thin to normal colloid also to determine the sour and dark yellow strepto- ketone of dark yellow strepto- simultaneously
The cytotoxicity of born of the same parents (HA), their CC50Value is all higher than 100 μM, the selectivity index (CC of dark yellow strepto- acid50/IC50) it is big
In 12.3 to 21.3, and the selectivity index (CC of dark yellow strepto- ketone50/IC50) it is more than 3.8 to 4.6, with positive control drug Ah
Mycin is similar.As a result show：Dark yellow strepto- acid is higher than positive control drug adriamycin to the selectively acting of glioma cell, changes
Yan Zhi, dark yellow strepto- acid will be less than comparison medicine adriamycin (table three) to the toxicity of normal spongiocyte.
The sour and dark yellow strepto- ketone of table three, dark yellow strepto- suppresses the effect of glioma cell and normal glia proliferation
2. influence of the sour and dark yellow strepto- ketone of dark yellow strepto- to glioma cell metabolic enzyme
The preparation of protein sample：People's glioma U87MG cells are with MEM and 10%FBS culture mediums in 37 DEG C and 5% titanium dioxide
Cultivated in the incubator of carbon, the cell by three generations's culture is used for the experimental study of the present invention.Cell (1.5 × 107) be inoculated in
In 10 centimetres of culture dish, dark yellow strepto- sour (1,30 μM), dark yellow strepto- ketone (2,60 μM) or 2- deoxidations are added after adherent 24h
After glucose (2-DG, 6mM) co-cultures 48 hours, first washed twice with ice-cold PBS, afterwards with ice-cold lysis buffer
(200 μ L) is cracked 15 minutes.Lysate centrifuges through 4 DEG C of low-temperature and high-speeds (11200rpm), and supernatant is protein sample.
The measure of protein content：Use the protein content of each protein sample of BCA kit measurements.By seminal plasma fructose detection kit
A and B are with 50：1 ratio is made into working reagent liquid, and standard items BCA dilutions are made into various concentrations with bovine serum albumin(BSA) (BSA)
BCA standard liquids.10 μ L standard liquids or protein sample liquid and 200 μ L working reagents liquid are taken at 37 DEG C to hatch 30 points after mixing
Clock artemia hatching solutions ELIASA determines trap in 562nm wavelength.Using BCA amounts as abscissa, trap is that ordinate draws standard
Curve, calculate regression equation.From the protein content of each protein sample liquid of regression equation calculation.
Western Blotting：10% dodecyl sodium sulfate polyacrylamide of each sample containing equal protein (15 μ g)
Amine gel electrophoresis (SDS-PAGE) is separated, and gel electrophoresis protein graphical spectrum is gone on polyvinylidene fluoride film (PVDF), and pvdf membrane is used
The 0.1%TBST of also 5% degreasing milk is closed 2 hours at room temperature.Pvdf membrane after closing is first and the primary antibody of tested enzyme is 4
DEG C be incubated overnight, wash film with TBST；The secondary antibody with HRP marks was in incubation at room temperature 2 hours afterwards.After TBST washes film three times, with enhancing
Chemical illuminating reagent detects immunoreactivity, then develops, and washes, and is fixed, and washing, dries, observation experiment result.Moved with β-flesh
Albumen (β-actin) compares as internal reference.
Experimental result (accompanying drawing 15) shows：Compare with negative control group (without the U87MG cells of drug-treated, CON), secretly
Yellow strepto- sour (1) and dark yellow strepto- ketone (2) can significantly reduce tumour cell glycolysis key enzyme HK2, PFKFB3, PKM2 and
LDH5 protein expression.But the dark yellow sour and dark yellow strepto- ketone of strepto- is to tricarboxylic acid cycle enzyme aconitase 2 (ACO2)
Unobvious are influenceed with the protein expression level of oxidative phosphorylation enzyme cromoci (CytoC).CON：U87-MG cell controls groups；
CON：Negative control group；1：Dark yellow strepto- acid treatment group；2：Dark yellow strepto- ketone treatment group；2-DG:2- deoxyglucoses；HK2:Oneself
Sugared kinases 2：PFKFB3:Phosphofructokinase/fructose 2,6- diphosphatases；PKM2:Pyruvate kinase M2；LDH5:Lactic dehydrogenase
Enzyme 5；ACO2:Aconitase 2；CytoC:Cromoci；β-actin:Beta-actin, internal reference).Known tricarboxylic acids
Circulation and oxidative phosphorylation are the leading metabolic pathways of normal cell, and ACO2 and CytoC be respectively krebs cycle pathway and
One of key enzyme of oxidative phosphorylation approach.Result above and analysis shows：Sour and dark yellow two activity of strepto- ketone of dark yellow strepto-
Compound, which may be by selectivity, to be influenceed multiple key enzymes of glioma glycolysis and produces its anticol matter tumor activity, is had only
Special Anticancer Effect and Mechanism.
In a word, the present invention is separated to a kind of ocean from marine animal taeniae flesh sea anemone (Haliplanella lineata)
The very dark yellow streptomycete of actinomyces (Streptomycesfulvissimus ZZ406), the bacterium can produce anticol matter tumor activity
The sour and dark yellow strepto- ketone of the dark yellow strepto- of compound.The invention provides the isolated culture method of very dark yellow streptomycete, reactive compound
The extraction separation and purification method of the sour and dark yellow strepto- ketone of dark yellow strepto- and the anti-glioma of the sour and dark yellow strepto- ketone of dark yellow strepto-
Activity.Because the sour and dark yellow strepto- ketone of dark yellow strepto- has good anticol matter tumor activity and unique modulate tumor cell metabolism
The Anticancer Effect and Mechanism of enzyme, so, the sour and dark yellow strepto- ketone of dark yellow strepto- has in terms for the treatment of glioma medicine is prepared
Application prospect.
Sequence table
<110>Zhejiang University
<120>A kind of preparation of the sour and dark yellow strepto- ketone of dark yellow strepto- and its medical usage
<160> 3
<170> SIPOSequenceListing 1.0
<210> 1
<211> 20
<212> DNA
<213>Engineer (Unknown)
<400> 1
agagtttgat cctggctcag 20
<210> 2
<211> 19
<212> DNA
<213>Engineer (Unknown)
<400> 2
ggttaccttg ttacgactt 19
<210> 3
<211> 1390
<212> DNA
<213> Streptomyces fulvissimus ZZ406
<400> 3
tgcagtcgac gatgaagccg cttcggtggt ggattagtgg cgaacgggtg agtaacacgt 60
gggcaatctg cccttcactc tgggacaagc cctggaaacg gggtctaata ccggataata 120
ctcctgcctg catgggtggg ggttgaaagc tccggcggtg aaggatgagc ccgcggccta 180
tcagcttgtt ggtggggtaa tggcctacca aggcgacgac gggtagccgg cctgagaggg 240
cgaccggcca cactgggact gagacacggc ccagactcct acgggaggca gcagtgggga 300
atattgcaca atgggcgaaa gcctgatgca gcgacgccgc gtgagggatg acggccttcg 360
ggttgtaaac ctctttcagc agggaagaag cgcaagtgac ggtacctgca gaagaagcgc 420
cggctaacta cgtgccagca gccgcggtaa tacgtagggc gcaagcgttg tccggaatta 480
ttgggcgtaa agagctcgta ggcggcttgt cacgtcggat gtgaaagccc ggggcttaac 540
cccgggtctg cattcgatac gggctagcta gagtgtggta ggggagatcg gaaattcctg 600
gtgtagcggt gaaatgcgca gatatcagga ggaacaccgg tggcgaaggc ggatctctgg 660
gccattactg acgctgagga gcgaaagcgt ggggagcgaa caggattaga taccctggta 720
gtccacgccg taaacgttgg gaactaggtg ttggcgacat tccacgtcgt cggtgccgca 780
gctaacgcat taagttcccc gcctggggag tacggccgca aggctaaaac tcaaaggaat 840
tgacgggggc ccgcacaagc agcggagcat gtggcttaat tcgacgcaac gcgaagaacc 900
ttaccaaggc ttgacatata ccggaaagca tcagagatgg tgcccccctt gtggtcggta 960
tacaggtggt gcatggctgt cgtcagctcg tgtcgtgaga tgttgggtta agtcccgcaa 1020
cgagcgcaac ccttgttctg tgttgccagc atgcctttcg gggtgatggg gactcacagg 1080
agactgccgg ggtcaactcg gaggaaggtg gggacgacgt caagtcatca tgccccttat 1140
gtcttgggct gcacacgtgc tacaatggcc ggtacaatga gctgcgatgc cgtgaggcgg 1200
agcgaatctc aaaaagccgg tctcagttcg gattggggtc tgcaactcga ccccatgaag 1260
tcggagttgc tagtaatcgc agatcagcat tgctgcggtg aatacgttcc cgggccttgt 1320
acacaccgcc cgtcacgtca cgaaagtcgg taacacccga agccggtggc ccaacccctt 1380
gtgggaggga 1390
Claims (7)
1. a kind of very dark yellow streptomycete, Classification And Nomenclature is Streptomycesfulvissimus ZZ406, is trained by Chinese Typical Representative
Support thing collection-Wuhan center preservation, deposit number:CCTCC M 2017435, preservation day：On August 3rd, 2017.
2. the preparation method of the very dark yellow streptomycete described in claim 1, it is characterised in that obtained by following steps：
(1) very dark yellow streptomycete is separately cultured
Take fresh taeniae flesh sea anemone (Haliplanella lineata) to be cleaned with the natural sea-water of sterilizing to be homogenized afterwards three times, be made
The suspension of various concentrations, take uniform suspension to be distributed in the culture dish containing solid medium, cultivate at ambient temperature
Afterwards, different bacterium colonies is transferred in another culture dish containing solid medium respectively, continues to cultivate at ambient temperature, most
Well-grown single colony inoculation is saved backup to the rearmounted 4 DEG C of refrigerators of slant medium culture afterwards；
Described taeniae flesh sea anemone (Haliplanella lineata) is obtained from the rock seam at Zhejiang Mount Putuo seabeach, day
Right seawater is obtained from the Area of The East China Sea near Zhoushan Zhejiang；The concentration of the suspension is 1 × 10-4~1 × 10-1g/
mL；The sampling amount of described sample suspension is 100~300 μ L；Solid medium contained by the culture dish is Gao Shi agar
Culture medium or other solid mediums；Described slant medium is Gao Shi agar mediums or other solid slope culture mediums；
Described incubated at room temperature temperature is 22~28 DEG C；Described incubation time is 5~15 days；
(2) strain idenfication of very dark yellow streptomycete
Above-mentioned steps (1) are separately cultured obtained bacterial strain and identify its species using 16S rDNA sequence analysis methods, are defined as
Very dark yellow streptomycete, Classification And Nomenclature is Streptomycesfulvissimus ZZ406, by China typical culture collection
The heart-Wuhan center preservation, deposit number:CCTCC M 2017435.
3. the sour and dark yellow strepto- ketone of the dark yellow strepto- of compound that very dark yellow streptomycete obtains according to claim 1, its feature
It is, described sour (1) and dark yellow strepto- ketone (2) chemical structural formula of dark yellow strepto- is respectively：
4. the preparation method of the dark yellow strepto- acid of compound and the dark yellow strepto- ketone of compound described in claim 3, it is characterised in that
Realized by following steps：
(1) preparation of very dark yellow streptomycete zymocyte liquid
The colony inoculation of the very dark yellow streptomycete (Streptomycesfulvissimus ZZ406) described in claim 1 is taken to arrive
In big conical flask containing liquid spawn culture medium, it will be obtained after the nutrient solution containing strain at ambient temperature shaken cultivation
Strain liquid, finally strain liquid is transferred in the big conical flask containing liquid fermentation medium, at ambient temperature shaken cultivation,
Obtain that there is very dark yellow streptomycete zymocyte liquid；
Described bacterium culture medium is liquid Gao Shi culture mediums, and described dosage is 150-250mL, described liquid fermentation and culture
Base is SC fluid nutrient mediums：10 grams of soluble starch, 0.3 gram of casein, 2 grams of potassium nitrate, 0.5 gram of magnesium sulfate, dipotassium hydrogen phosphate 2
Gram, 0.02 gram of calcium carbonate, 0.01 gram of ferrous sulfate, 1 liter of natural sea-water, pH value 7.2；Described big triangle blake bottle is 500mL；
Described incubated at room temperature temperature is 22~28 DEG C；The rotating speed of the vibration is 160~180rpm；Described incubation time be 5~
15 days；
(2) extraction separation and purification of the sour and dark yellow strepto- ketone of dark yellow strepto-
Bacterial strain ZZ406 fermentation mycelium is extracted with methanol, and methanolic extract is extracted with ethyl acetate to obtain ethyl acetate extraction again
Take thing.Acetic acid ethyl ester extract is first separated with normal-phase silica gel column chromatography, with ethyl acetate and hexamethylene mixed solvent gradient elution,
And with thin layer chromatography analysis, nine component Fr.1~Fr.9, component Fr4 and component Fr 8 are obtained again respectively with the efficient liquid of preparative
Chromatography separates, and obtains the sour and dark yellow strepto- ketone of the dark yellow strepto- of pure compound；
The purification on normal-phase silica gel dosage of the column chromatography and the sample size ratio of upper amount are 30~50g:1.0g；Described efficient liquid phase point
It is from condition:Permanent CXTH-3000 high performance liquid chromatographs, Fuji C are led in innovation18CT-30 chromatographic columns, 280 × 30mm, 10 μm, first
Alcohol and water is mobile phase, Detection wavelength 220nm, flow velocity 10.0mL/min；
(3) Structural Identification of the sour and dark yellow strepto- ketone of dark yellow strepto-
The structure of the sour and dark yellow strepto- ketone of dark yellow strepto- is a peacekeeping two-dimentional NMR spectrum according to them, high resolution mass spectrum data
The methods of being calculated with ECD and determine.
5. the sour and dark yellow strepto- ketone of the dark yellow strepto- of compound according to claim 3 is in treatment glioma medicine is prepared
Application.
6. application according to claim 5, it is characterised in that the medicine is independent by the sour and dark yellow strepto- ketone of dark yellow strepto-
Both combinations are made with pharmaceutically acceptable carrier.
7. application according to claim 5, it is characterised in that the medicine is independent by the sour and dark yellow strepto- ketone of dark yellow strepto-
Both combinations are made with other active materials and pharmaceutically acceptable carrier.
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| CN110172485A (en) * | 2019-05-13 | 2019-08-27 | 浙江大学 | The preparation and use of anticol matter tumor activity substance pyrroles's spiral shell ketone alkali G |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002000261A (en) * | 2000-06-28 | 2002-01-08 | Ozeki Corp | Antibacterial agent |
| CN102978144A (en) * | 2012-12-24 | 2013-03-20 | 湖南农业大学 | Streptomyces fulvissimus |
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| JPS5824538A (en) * | 1981-08-06 | 1983-02-14 | Sanraku Inc | Novel anthracyclinone compound |
| FR2759079B1 (en) * | 1997-02-03 | 2002-06-14 | Girex | PROCESS FOR THE PREPARATION OF SUBSTITUTED ANTHRAQUINONES AND APPLICATION TO THE PREPARATION OF RHEINES |
| JP2000229908A (en) * | 1999-02-15 | 2000-08-22 | Kyowa Hakko Kogyo Co Ltd | Cht22 compound |
| AUPQ008299A0 (en) * | 1999-04-30 | 1999-05-27 | G.J. Consultants Pty Ltd | Isoflavone metabolites |
| GB0016020D0 (en) * | 2000-06-29 | 2000-08-23 | Inst Biomar Sa | New polyciclic xanthones and their use |
| CN1748675A (en) * | 2005-07-11 | 2006-03-22 | 丛晓东 | Composition of rheinic acid compounds and preparing method and use for treating diabetes |
| CN105112322B (en) * | 2015-08-07 | 2018-07-06 | 浙江大学 | Grey mold quinone A and B and preparation method thereof and medical usage |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002000261A (en) * | 2000-06-28 | 2002-01-08 | Ozeki Corp | Antibacterial agent |
| CN102978144A (en) * | 2012-12-24 | 2013-03-20 | 湖南农业大学 | Streptomyces fulvissimus |
Non-Patent Citations (1)
| Title |
|---|
| MYRONOVSKYI M.等: "Complete genome sequence of streptomyces fulvissimus", 《J. BIOTECHNOL.》 * |
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| CN110172485A (en) * | 2019-05-13 | 2019-08-27 | 浙江大学 | The preparation and use of anticol matter tumor activity substance pyrroles's spiral shell ketone alkali G |
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