CN106038624B - Yantai herba limonii gmelinii anti-cancer effective component extract and its preparation method and application - Google Patents
Yantai herba limonii gmelinii anti-cancer effective component extract and its preparation method and application Download PDFInfo
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- CN106038624B CN106038624B CN201610376979.1A CN201610376979A CN106038624B CN 106038624 B CN106038624 B CN 106038624B CN 201610376979 A CN201610376979 A CN 201610376979A CN 106038624 B CN106038624 B CN 106038624B
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying
Abstract
The invention discloses Yantai herba limonii gmelinii anti-cancer effective component extracts and its preparation method and application.Yantai herba limonii gmelinii anti-tumor effective component extract is by obtaining hexamethylene, ethyl acetate, four n-butanol, water active components after the herba limonii gmelinii alcohol extracting of Yantai.Yantai herba limonii gmelinii active component can inhibit a variety of cancer cell line to be proliferated, it is especially significant to the inhibiting effect of the tumor cell lines such as leukemia HL-60 and K562 cell, MCF-7 Breast Cancer Cell, lung cancer A549 cell, cervical cancer Hela cells, HepG-2 cell line, it is most obvious with ethyl acetate extract anti-tumor activity, anticancer spectrum is wider, and Yantai herba limonii gmelinii active component has the function of inducing leukemia cell HL-60 differentiation, therefore, it can be used for manufacturing anti-tumor drug, have great importance especially for leukemia treating research.
Description
Technical field
The present invention relates to natural drug extractions and application field, in particular to Yantai herba limonii gmelinii with anti-tumor activity to mention
Take object and its preparation method and application.
Background technique
Cancer Mortality constantly increases, and has become the main reason for mankind are because of disease death at present.Wherein, lung
Cancer, liver cancer and gastric cancer are the main entity tumor causes of death, and three accounts for about the 60% of total mortality of malignant tumors, and leukaemia
It is the main malignant disease for endangering hematological system.It is at present radiotherapy, chemotherapy for the main treatment means of malignant tumour, but putting
It treats clinically side effect and is also easy to produce drug resistance greatly, so that clinical efficacy lowers.Therefore, new less toxic, efficient, safe medicine is developed
Object become treatment tumour there is an urgent need to.The traditional medicine of China is with a long history, and many Chinese medicines itself have anti-tumor activity, and
And can play and adjust immunological function repression tumour, have the effect of wholistic therapy.High-efficiency low-toxicity is found from natural drug
The lead compound of anti-tumor activity is always the hot spot of domestic and international anticancer drug research, and effective activated monomer is in
The anticancer compound preparation of herbal medicine active component all has important clinical value.
Yantai herba limonii gmeliniiLimoniumfranchetii(Debx.) Kuntze belong to Plumbaginaceae (Plumbaginaceae, again
Claim lumbaginaceae) Statice (LimoniumIt Miller), is Chinese unique wheat;Because its corolla lavender (sometimes top without
Color), limb fades to white by lilac red, also known as pale reddish brown herba limonii gmelinii.This kind is rare species, is only distributed in Liaoning, Shandong
The peninsula is listed in national second batch rare extinction plants to northeast of Jiangsu Province.There are about 300 for whole world Limonnium resource
Kind, China has 17 kinds, is distributed north and south each province.The platymiscium containing flavones, alkaloid, polysaccharide, organic acid, tannin, volatile oil etc. at
Point, have and enriches blood, stops blooding, dissipating the stasis of blood, antibacterial anti-inflammatory and antitumor and other effects, and it is newest research shows that Limonnium also has
There are the pharmacological activity such as liver protection, antiviral, inhibition alpha-glucosidase.The research of Limonnium chemical component is in recent years
Hot spot, mainly have limonium sinenseKuntze chemical component research, Herba Limonii Gmelinii with yellow flower volatile chemical component research, twocolor sealavander herb
Compounds content measurement and the research of compound extraction process etc..Currently, there is no related Yantai herba limonii gmelinii chemical component and its
The document report of pharmacological activity also has no report related with the Yantai herba limonii gmelinii method for preparing extractive in the present invention, purposes.
Summary of the invention
The technical problems to be solved by the invention aim to overcome that above-mentioned shortcoming, researching and designing Yantai herba limonii gmelinii mention
Take object and its extraction separation method and application in preparation of anti-tumor drugs.
It is an object of the invention to provide a kind of to extract effective portion with antitumor action from the herba limonii gmelinii of Yantai
Position.
It extracts to obtain in the herba limonii gmelinii of Yantai the second purpose of the invention is to provide a kind of simple preparation method and there is anti-swell
The extract of tumor effect.
The third object of the present invention is to provide a kind of using described position as the pharmaceutical composition of effective component.
The fourth object of the present invention is to provide described active component and is preparing the application in antitumor and related drugs.
Technical scheme is as follows:
Yantai herba limonii gmelinii anti-cancer effective component extract, it is Yantai herba limonii gmelinii herb crushed after being dried, and 95% second is added
Alcohol extracts three times, and combined extract is concentrated under reduced pressure, after filtrate boils off ethyl alcohol, distilled water is added to be stirred into suspension, water layer is pressed
According to the sequence that polarity is ascending, hexamethylene, ethyl acetate, extracting n-butyl alcohol are successively used.By hexamethylene, ethyl acetate, positive fourth
After recycling design is concentrated under reduced pressure respectively in alcohol and water extract, the extraction of four positions Yantai herba limonii gmelinii anti-cancer effective component is obtained
Object.
A method of preparing above-mentioned Yantai herba limonii gmelinii anti-cancer effective component extract, it the following steps are included:
(1) Yantai herba limonii gmelinii is raw material, and 95% ethyl alcohol uniform stirring is added in medicinal material Yantai herba limonii gmelinii crushed after being dried, is stood
For 24 hours, in triplicate, it filters, merging obtains extracting solution.
(2) after the extracting solution concentration removal ethyl alcohol of step (1), total medicinal extract is obtained, 2-5 times of volume of water is added to stir into suspension
Liquid extracts Aqueous extracts and cyclohexane volume ratio 1:1, and extraction three times, merges hexamethylene alkane extract.Hexamethylene is recycled,
40 DEG C of vacuum driers are dry, obtain hexamethylene position.
(3) hexamethylene water layer extracted is extracted with ethyl acetate volume ratio 1:1, extraction three times, merges extraction
Liquid.Ethyl acetate is recycled, 30 DEG C of vacuum driers are dry, obtain ethyl acetate extract.
(4) hexamethylene will be had been subjected to and the stripped water layer solution of ethyl acetate extracts with n-butanol volume ratio 1:1
It takes, extracts three times, combining extraction liquid.N-butanol is recycled, 60 DEG C of vacuum driers are dry, obtain n-butanol portion.
(5) water extract is concentrated and dried using the method for freeze-drying, obtains water position.Above four polar fractions are
For Yantai herba limonii gmelinii active principle extract.
In the preparation of antitumor drugs invention further provides Yantai herba limonii gmelinii anti-tumor effective component extract
Using.With Human leukemia cell line HL-60 and K562, HepG2 cell lines, human A459 lung cancer cell line, people
Breast cancer cell MCF-7, human cervical carcinoma cell strain Hela are detection tumor strain, to inhibit tumor cell proliferation ability to refer to as activity
Mark carries out anti-tumor activity experiment.The experimental results showed that four positions can inhibit tumour cell in vitro in cellulotoxic experiment
Proliferation, have cytotoxicity, the anti-tumor activity that ethyl acetate extract performs better than.Also, hexamethylene position and water portion
Position shows as the activity for inducing Human leukemia cell line HL-60 differentiation well, can be used for preparing antineoplaston drug.
The present invention also provides the pharmaceutical compositions of the active component prepared in this way.Described pharmaceutical composition be with
Active component of the invention is effective component, is formed with pharmaceutically acceptable carrier, and to be suitble to medicinal dosage form
It can be tablet, sugar coated tablet, film coated tablet, enteric coated tablet, sustained-release tablet, capsule, hard in the presence of, these dosage forms
Capsule, soft capsule, Duracaps, oral solution, mixture, mouth containing agent, granule, electuary, pill, powder, paste, pellet
Agent, suspension, solution, injection, powder-injection, freeze drying powder injection, suppository, ointment, emplastrum, creme, spray, drop
Agent, pill and patch etc..The drug of above-mentioned various dosage forms can be prepared according to the conventional method of pharmaceutical field.
Beneficial effect
(1) advantages of the present invention and effect are using conventional methods such as solvent extraction, extractions, find that Yantai is enriched blood for the first time
Careless hexamethylene position, ethyl acetate extract, n-butanol portion and water position have anti-tumor activity, which demonstrate,proves through experiment
The bright activity with extracorporeal suppression tumor cell growth.
(2) present invention firstly discovers that the hexamethylene position and water position of Yantai herba limonii gmelinii have induction human leukemia cell
The activity of HL-60 cell differentiation is suitable for developing into Chinese medicine for treating leukemia new drug.
(3) present invention has expanded the raw material channel of anti-tumor drug, expands the purposes of Yantai herba limonii gmelinii, Yantai is made to enrich blood
Grass develops into the new raw material of anti-tumor drug, is remarkably improved the medical value of Yantai herba limonii gmelinii.
Detailed description of the invention
Fig. 1 is the schematic diagram of Yantai herba limonii gmelinii extract anti-tumor activity experiment IC50 value (μ g/mL).
Fig. 2 is the schematic diagram that Wright's staining observation Yantai herba limonii gmelinii extract influences HL-60 cell differentiation.
Specific embodiment
Implementation set forth below facilitates those skilled in the art and more fully understands the present invention, but does not limit in any way
The present invention.
Embodiment 1: the preparation of Yantai herba limonii gmelinii anti-tumor effective component extract
By Yantai herba limonii gmelinii herb drying and crushing, mixture 1500g is taken, 95% ethyl alcohol that 8-10 times is measured is added and impregnates 2-3
It, extracts Alcohol soluble composition, in triplicate, merges No. 3 extracting solutions, and 30 DEG C ~ 40 DEG C are recovered under reduced pressure ethyl alcohol to no alcohol taste, obtain total
Medicinal extract adds distilled water to be stirred into suspension, is fitted into separatory funnel, adds isometric hexamethylene to extract three times, merges ring
Hexane extract is recovered under reduced pressure, and 40 DEG C of vacuum driers are dry to constant weight, obtains hexamethylene position.Remaining aqueous decompression removes
It after removing remaining hexamethylene, is extracted 3 times with isometric ethyl acetate, combined ethyl acetate extract liquor is recovered under reduced pressure, and 30 DEG C true
Empty drying machine is dry to constant weight, obtains ethyl acetate extract.After remaining ethyl acetate is removed under reduced pressure in remaining water layer solution, use
Isometric extracting n-butyl alcohol 3 times merges butanol extraction liquid, is recovered under reduced pressure, and 60 DEG C of vacuum driers are dry to constant weight, obtains
N-butanol portion.Water extract is concentrated and dried using the method for freeze-drying finally, obtains water position.
Embodiment 2: the research of Yantai herba limonii gmelinii extract anti-tumor activity
The Yantai herba limonii gmelinii anti-tumor effective component extract obtained using embodiment 1 takes raw in logarithm as test medicine
(there are several types of Human leukemia cell line HL-60 and K562, people liver for the tumor cell line of use for long-term kinds of tumor cells
Cancer cell line HepG2, human A459 lung cancer cell line, human breast cancer cell line Bcap-37, human cervical carcinoma cell strain Hela), with containing
The RPMI-1640 complete culture solution of 10% calf serum adjusts cell concentration, and 8000, every hole cell, 180 μ L is inoculated in
In 96 porocyte culture plates, 37 DEG C, 5%CO are set2 In incubator overnight, next day dosing.By the sample solution of the different extracted parts
It is configured to the mother liquor that initial concentration is 1mg/mL, two-fold dilution, it is yin with PBS that the 20 μ L of drug diluted, which is added, in every hole
Property control.After sample-adding culture 72h, the 20 μ L of MTT of 5mg/mL is added in every hole, continues to cultivate 4 h, then quick turnover panel method
Discard liquid in hole, every hole adds the DMSO of 100 μ L, 10min is vibrated, in detecting each hole in microplate reader at 570nm wavelength
OD value, and calculate using GraphPad Prism software the IC50 value of each extract part.It the results are shown in Table 1.It can by result
Know, herba limonii gmelinii extract four positions in Yantai have good tumor inhibitory effect, most with ethyl acetate extract antitumous effect
Obviously.
Embodiment 3: influence of the Yantai herba limonii gmelinii extract to the induction differentiation of acute myeloid leukemia HL-60 cell
The Yantai herba limonii gmelinii anti-tumor effective component extract obtained using embodiment 1 is test medicine, with 25 μ g/ of final concentration
The different parts extract-treated of mL is in the HL-60 cell of logarithmic growth phase, with 5 × 104A/mL cell concentration is laid on
24 orifice plates set 37 DEG C, 5%CO25d is cultivated in incubator, collects group of cells, smear, Wright's staining, microscopically observation
The features such as cellular morphology is simultaneously taken a picture, observation cell size, karyoplasmic ratio, nuclear shapes, endochylema dyeing.The result shows that Yantai is mended
After the extract-treated leukemia HL-60 cell 5d at four positions of blood grass, obvious become occurs for light microscopic observation cytomorphology
Change (see Fig. 2).Show as nuclear chromatin condensation, caryoplasm ratio reduces, and core is partial to side, there is a recess and leaflet, and cell is to maturation
It is stage differentiated, it is especially the most obvious with hexamethylene position and ethyl acetate extract, illustrate Yantai herba limonii gmelinii extract to HL-60
Cell has the ability of certain induction differentiation.Therefore, herba limonii gmelinii extract in Yantai can be used for preparing anti-tumor drug, especially
It can be used as treatment of the differentiation-inducing agents for leukaemia.
Claims (3)
1. Yantai herba limonii gmelinii anti-tumor effective component extract application in preparation of anti-tumor drugs, which is characterized in that described
Extract is the ethyl acetate extract of Yantai herba limonii gmelinii；
Its active principle method for preparing extractive includes the following steps:
Step 1: Yantai herba limonii gmelinii herb being crushed, three times with 95% alcohol steep, ethyl alcohol is removed in combined extract, reduced pressure, obtains
Total medicinal extract；
Step 2: total medicinal extract that step 1 is obtained adds distilled water to be stirred into suspension, is fitted into separatory funnel, water layer is pressed
It according to the sequence that polarity is ascending, is successively extracted with hexamethylene, ethyl acetate, butanol solution, by hexamethylene, acetic acid second
Ester, n-butanol and water extract rotary evaporation be concentrated under reduced pressure respectively after to get Yantai herba limonii gmelinii anti-tumor effective component extract；
The Yantai herba limonii gmelinii being extracted is the herb for impregnating and drying by ethyl alcohol, including root, stem, Ye Hehua；
It is described antitumor to inhibit leukemia HL-60 and K562 cell in vitro for Yantai herba limonii gmelinii anti-tumor effective component extract
Proliferation, inducing leukemia cell HL-60 directed differentiation.
2. Yantai herba limonii gmelinii anti-tumor effective component extract according to claim 1 answering in the preparation of antitumor drugs
With, it is characterized in that: the anti-tumor drug be Yantai herba limonii gmelinii anti-tumor effective component extract with it is pharmaceutically acceptable
Carrier or auxiliary material composition.
3. Yantai herba limonii gmelinii anti-tumor effective component extract according to claim 1 answering in the preparation of antitumor drugs
With, it is characterised in that: described pharmaceutical composition is using active component of the invention as effective component, with pharmaceutically acceptable load
Body is formed, and to be suitble to medicinal dosage form to exist, these dosage forms can be tablet, capsule, oral solution, mouth containing
It is agent, granule, electuary, pill, powder, paste, sublimed preparation, suspension, solution, injection, powder-injection, suppository, ointment, hard
Paste, creme, spray, drops, pill and patch.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102920753A (en) * | 2012-11-05 | 2013-02-13 | 江苏省中国科学院植物研究所 | Dichromatism gmelin sealavender herb extractive and application thereof |
| CN105362314A (en) * | 2015-12-09 | 2016-03-02 | 江苏省中国科学院植物研究所 | Novel application of Limonium bicolor ethyl acetate extract |
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- 2016-06-01 CN CN201610376979.1A patent/CN106038624B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102920753A (en) * | 2012-11-05 | 2013-02-13 | 江苏省中国科学院植物研究所 | Dichromatism gmelin sealavender herb extractive and application thereof |
| CN105362314A (en) * | 2015-12-09 | 2016-03-02 | 江苏省中国科学院植物研究所 | Novel application of Limonium bicolor ethyl acetate extract |
Non-Patent Citations (1)
| Title |
|---|
| 补血草属植物野生资源多样性及药理学研究进展;丁鸽等;《时珍国医国药》;20121220;第23卷(第12期);第3113页右边栏第2-3段 * |
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