CN108057096A - A kind of method of the extraction with antitumor activity alkaloid from oat bran - Google Patents
A kind of method of the extraction with antitumor activity alkaloid from oat bran Download PDFInfo
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- CN108057096A CN108057096A CN201810111099.0A CN201810111099A CN108057096A CN 108057096 A CN108057096 A CN 108057096A CN 201810111099 A CN201810111099 A CN 201810111099A CN 108057096 A CN108057096 A CN 108057096A
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- alkaloid
- oat bran
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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)
- A61K36/88—Liliopsida (monocotyledons)
- A61K36/899—Poaceae or Gramineae (Grass family), e.g. bamboo, corn or sugar cane
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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
- A61K2236/35—Extraction with lipophilic solvents, e.g. Hexane or petrol ether
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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
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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
- A61K2236/53—Liquid-solid separation, e.g. centrifugation, sedimentation or crystallization
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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
- A61K2236/55—Liquid-liquid separation; Phase separation
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Abstract
The present invention provides the application of a kind of method of extraction from oat bran with antitumor activity alkaloid and the alkaloid of extraction in exploitation health food and antitumor drug.The extracting method is to crush oat bran, and overnight, filter residue extracts 25 times repeatedly after filtered through gauze for methanol/water/glacial acetic acid room temperature extraction；It collects whole extracting solutions to be concentrated under reduced pressure, concentrate removes lipid with n-hexane extraction, obtains oat bran alkaloid crude extract；Crude extract is chromatographed into further enrichment removal of impurities through 8 macroreticular resins of AB, oat bran alkaloid is obtained after obtained eluent vacuum drying.Using the biology alkali process colorectal cancer cell and normal intestinal epithelial cell, it is found that it has apparent lethal effect to colon cancer cell, to normal cell without apparent cytotoxicity, can be applied in exploitation health food and antitumor drug.
Description
Technical field
The present invention relates to alkaloid extractive technique fields, and in particular to one kind extraction from oat bran has antitumor work
The method of property alkaloid.
Background technology
Oat (Avena sativa L.) is grass,《Compendium of Materia Medica》In be referred to as bromegrass, wild oat fruit.It is one
Kind low sugar, high nutrition, high energy food, Shanxi is one of main place of production of oat.Oat contains multiple biological activities ingredient, as β-
Glucan, saponins, flavonoids, alkaloids etc.；These active ingredients have abundant nutrition and medical value.Study table
Bright, avenabeta glucosan can control blood glucose, be effectively improved diabetes B；Serum cholesterol can be reduced simultaneously, reduce painstaking effort
Pipe disease risk；As a kind of dietary fiber, beta glucan can also promote to digest, and prevent enterogastric diseases.U.S. north
The researcher in card functional food and human health laboratory has found 16 kinds of saponins substances in oat, wherein 2 kinds have it is micro-
The effect of weak inhibitor against colon carcinoma cells.
Oat bran also known as oat bran or oat bran have enteron aisle toxin expelling, relief of constipation and the effect for promoting fat metabolism.
Oat alkaloid A venanthramides (AVNs) is a kind of secondary metabolite of oat, is existed by Collins FW et al.
It is found in the wheat bran of oat, confirms that it is primarily present in oat bran afterwards.AVNs is a kind of N- substitutions cinnamoyl neighbour's ammonia
Yl benzoic acid compound, Collins FW etc. found in 1989 and is determined that more than 20 plant AVNs, and wherein content is enriched the most
It is AVN A, B and C.Someone has confirmed that AVNs has certain anti-inflammatory effect.Alkaloid is extracted from oat bran to be used in combination
It is had not been reported in antitumor activity.
The content of the invention
The method with antitumor activity alkaloid is extracted from oat bran it is an object of the invention to provide a kind of, with
And application of the alkaloid in antitumor drug is prepared.
A kind of alkaloid with antitumor activity provided by the invention is through methanol/water/glacial acetic acid room by oat bran
Warm extracted many times, n-hexane remove lipid, AB-8 macroporous resin purifications, the alkaloid being dried in vacuo.
A kind of method of the extraction with antitumor activity alkaloid from oat bran provided by the invention, including walking as follows
Suddenly：
The first step：Oat bran is crushed, with 1:The solid-liquid ratio of 8-12 adds in methanol/water/glacial acetic acid room temperature extraction overnight,
Wherein the volume ratio of methanol/water/glacial acetic acid is 50-90:10:1；With filtered through gauze extracting solution, filter residue repeats extraction 2-5 times, receives
The whole filtrates of collection, 50 DEG C are concentrated under reduced pressure into 1/10 volume；
Second step：Concentrate removes lipid impurities to get to oat bran alkaloid crude extract through n-hexane extraction 3 times；
3rd step：By crude extract through macroporous resin purification, with 95% ethanol elution, eluent vacuum drying is to get to tool
There is the oat bran alkaloid of active anticancer.
Solid-liquid ratio is 1 in the first step:10.
Extracting solution is that methanol/water/glacial acetic acid volume is 80 in the first step:10:1.
It is 3 times that filter residue, which repeats extraction time, in the first step.
Macroreticular resin is AB-8 macroreticular resins in 3rd step.
Beneficial effects of the present invention compared with prior art：
The oat bran alkaloid antitumor activity is detected using mtt assay, the results show alkaloid is to colorectal cancer
Cell has apparent lethal effect, and IC50 values are 38 μ g/ml, and illustrate oat bran without apparent cytotoxicity to normal cell
Skin alkaloid can be applied in antitumor particularly drugs against colon cancer is prepared.
The method of extraction oat bran alkaloid provided by the invention is at low cost, and easy to operate, purifying process is simple, suitable for work
Industry metaplasia is produced.
Description of the drawings
Fig. 1 oat bran alkaloid optimization for extracting condition single factor experiments
Fig. 2 oat brans alkaloid (AVNs) sample
Influence of Fig. 3 oat brans alkaloid to colon cancer cell and normal cell proliferation
Specific embodiment
Embodiment 1：Oat bran alkaloid extracting technique optimizes
(1) appropriate oat bran is taken, is crushed, sieving weighs 5 parts of 10g powder, is separately added into methanol/water/glacial acetic acid ratio
For (50:10:1；60:10:1；70:10:1；80:10:1；90:10:1) extracting solution room temperature extraction is overnight.Filtered through gauze is collected
Filtrate, 50 DEG C are concentrated under reduced pressure into 1/10 volume；Concentrate is through 3 removing lipid impurities of n-hexane extraction, obtained solution vacuum
It is dry, it weighs, calculates yield (see a in Fig. 1).
(2) appropriate oat bran is taken, is crushed, sieving weighs 5 parts of 10g powder, with 1:4；1:6；1:8,1:10,1:12
Solid-liquid ratio is separately added into methanol/water/glacial acetic acid as 80:10:The extraction of 1 room temperature is overnight.Filtered through gauze collects filtrate, 50 DEG C of decompressions
It is concentrated into 1/10 volume；Concentrate through 3 removing lipid impurities of n-hexane extraction, weigh (see figure by obtained solution vacuum drying
In 1 b).
(3) appropriate oat bran is taken, is crushed, sieving weighs 4 parts of 10g powder, with 1:10 solid-liquid ratio addition methanol/water/
Glacial acetic acid is 80:10:The extraction of 1 room temperature is overnight.Filtered through gauze, filter residue repeat extraction 2-5 times, collect whole filtrates, 50 DEG C of decompressions
It is concentrated into 1/10 volume；Concentrate through 3 removing lipid impurities of n-hexane extraction, weigh (see figure by obtained solution vacuum drying
In 1 c).
Embodiment 2：Oat bran alkaloid extracting method
(1) appropriate oat bran is weighed, is crushed, sieving, with 1:10 solid-liquid ratio adds in methanol：Water：Glacial acetic acid=(80:
10:1) room temperature extraction is stayed overnight.With filtered through gauze extracting solution, filter residue repeats extraction 3 times, collects whole filtrates, 50 DEG C are concentrated under reduced pressure
To 1/10 volume；
(2) concentrate removes lipid impurities to get to oat bran alkaloid crude extract through n-hexane extraction 3 times；
(3) by crude extract through AB-8 macroporous resin purifications, with 95% ethanol elution, eluent vacuum drying is to get to shallow
Yellow powder has the oat bran alkaloid of active anticancer (see Fig. 2).
Embodiment 3：Oat bran alkaloid antitumor activity detects
HCT116, DLD1 and the FHC cell of exponential phase are chosen, adjustment cell density is per hole 8 × 104A/ml, often
Hole adds in 100 μ l into 96 well culture plates, in 37 DEG C, 5%CO2It is incubated overnight in incubator.Culture is taken out after cell attachment
Plate replaces the culture medium containing various concentration (0,8,20,40,80,120 μ g/ml) oat bran alkaloid, if 5 parallel holes.After
After when continuous culture 48 is small, liquid in hole is abandoned in suction, and new culture medium is added in after PBS washings, adds 20 μ l of MTT solution (5mg/ml) per hole,
Continue to terminate culture after being incubated 4h.Careful inhale abandons culture supernatant in hole, and 150 μ l DMSO are added per hole, vibrates 10min, makes crystallization
Object fully melts.490nm wavelength measures each hole absorbance value on enzyme linked immunological monitor (see Fig. 3).
Gained oat bran alkaloid of the invention, anti tumor activity in vitro detection is carried out to Human colorectal cancer cells strain,
Under the various concentration alkaloid is intervened, growth of tumour cell is significantly inhibited, and to normal cell without apparent cytotoxicity.
Claims (9)
- It is through methanol/water/glacial acetic acid room temperature by oat bran 1. a kind of alkaloid with antitumor activity, is characterized in that Extracted many times, n-hexane remove lipid, AB-8 macroporous resin purifications, the alkaloid being dried in vacuo.
- A kind of 2. method of alkaloid of the extraction with antitumor activity from oat bran, which is characterized in that including following step Suddenly：The first step：Oat bran is crushed, with 1:The solid-liquid ratio of 8-12 adds in methanol/water/glacial acetic acid room temperature extraction overnight, wherein The volume ratio of methanol/water/glacial acetic acid is 50-90:10:1；With filtered through gauze extracting solution, filter residue repeats extraction 2-5 times, collects complete Portion's filtrate, 50 DEG C are concentrated under reduced pressure into 1/10 volume；Second step：Concentrate removes lipid impurities to get to oat bran alkaloid crude extract through n-hexane extraction 3 times；3rd step：By crude extract through macroporous resin purification, with 95% ethanol elution, eluent vacuum drying is to get to anti- The oat bran alkaloid of cancer activity.
- 3. extracting the method for the alkaloid with antitumor activity from oat bran as claimed in claim 2, feature exists In solid-liquid ratio described in the first step is 1:10.
- 4. extracting the method for the alkaloid with antitumor activity from oat bran as claimed in claim 2, feature exists In extracting solution described in the first step is that methanol/water/glacial acetic acid volume ratio is 80:10:1.
- 5. extracting the method for the alkaloid with antitumor activity from oat bran as claimed in claim 2, feature exists In filter residue described in second step repeats extraction 3 times.
- 6. extracting the method for the alkaloid with antitumor activity from oat bran as claimed in claim 2, feature exists In macroreticular resin described in the 3rd step is AB-8 macroreticular resins.
- 7. application of the oat bran alkaloid as described in claim 1 in antitumor drug is prepared.
- 8. application of the alkaloid in antitumor drug is prepared made from the either method as described in claim 2-6.
- 9. application of the alkaloid as claimed in claim 8 in antitumor drug is prepared, the tumour is colon cancer.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201810111099.0A CN108057096A (en) | 2018-02-02 | 2018-02-02 | A kind of method of the extraction with antitumor activity alkaloid from oat bran |
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| CN201810111099.0A CN108057096A (en) | 2018-02-02 | 2018-02-02 | A kind of method of the extraction with antitumor activity alkaloid from oat bran |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101798585A (en) * | 2010-04-08 | 2010-08-11 | 天津科技大学 | Method for high-yield production of oat polysaccharide with naked oat bran as raw material |
| CN102369289A (en) * | 2009-03-27 | 2012-03-07 | 加拿大农业及农业食品部 | Method for increasing concentration of avenanthramides in oats |
| CN107412600A (en) * | 2017-08-15 | 2017-12-01 | 广州无添加主义化妆品有限公司 | A kind of extracting method of oat alkaloid, spray and preparation method thereof |
-
2018
- 2018-02-02 CN CN201810111099.0A patent/CN108057096A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102369289A (en) * | 2009-03-27 | 2012-03-07 | 加拿大农业及农业食品部 | Method for increasing concentration of avenanthramides in oats |
| CN101798585A (en) * | 2010-04-08 | 2010-08-11 | 天津科技大学 | Method for high-yield production of oat polysaccharide with naked oat bran as raw material |
| CN107412600A (en) * | 2017-08-15 | 2017-12-01 | 广州无添加主义化妆品有限公司 | A kind of extracting method of oat alkaloid, spray and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| JORDAN HASTINGS ETAL: "venanthramide-C reduces the viability of MDA-MB-231 breast cancer cells through an apoptotic mechanism", 《CANCER CELL INTERNATIONAL》 * |
| 任祎 等: "燕麦生物碱的提取及其抗氧化活性研究", 《农业工程学报》 * |
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