CN110746284A - Extraction and separation method of β -acid homolog in hops - Google Patents
Extraction and separation method of β -acid homolog in hops Download PDFInfo
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- CN110746284A CN110746284A CN201911126750.2A CN201911126750A CN110746284A CN 110746284 A CN110746284 A CN 110746284A CN 201911126750 A CN201911126750 A CN 201911126750A CN 110746284 A CN110746284 A CN 110746284A
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
- C07C45/78—Separation; Purification; Stabilisation; Use of additives
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C2601/00—Systems containing only non-condensed rings
- C07C2601/12—Systems containing only non-condensed rings with a six-membered ring
- C07C2601/16—Systems containing only non-condensed rings with a six-membered ring the ring being unsaturated
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Abstract
The invention relates to a method for extracting, separating and purifying β -acid homologues in hops, belongs to the technical field of natural medicine extraction and separation, and relates to the technical field of extraction and separation of active ingredients of natural products, in particular to a method for extracting, separating and purifying β -acid from carbon dioxide extracts of hops to further obtain β -acid homologues with different ratios.
Description
Technical Field
The invention relates to the technical field of extraction and separation of active ingredients of natural products, in particular to a separation method for extracting, separating and purifying β -acid from a hop carbon dioxide extract so as to obtain β -acid homologues with different proportions.
Background
Hops (academic name:Humulus lupulusl.) is one of the basic raw materials for brewing beer, can endow the beer with the effects of bacteriostasis, corrosion prevention, foam retention enhancement and the like, and provides unique bitterness and aroma characteristics for the beer. In addition, the hop has stronger physiological activity, the medicinal value research history of the hop at home and abroad is long, and the hop is clinically proved to have the characteristics of good sedation, sleep promotion and the like. As early as in Ben Cao gang mu, China records that the hop has good food therapy effect and is considered to have the effects of preventing and treating tuberculosis, reducing phlegm and relieving cough and the like.
Hops are mainly composed of soft/hard resins, essential oils, proteins, polyphenols, waxes, celluloses, amino acids, etc., wherein the soft resin is the most main bitter component in hops, and mainly consists of α -acid and β -acid.
α -acid is the most bitter substance in hops, and β -acid is considered to have small bitter contribution to beer and is often separated from hops as a byproduct, but a large number of research results show that β -acid has biological activities of bacteriostasis, antisepsis, anti-depression and the like, wherein the bacteriostasis and antisepsis performance is mainly due to the fact that β -trione groups existing in β -acid have bacteriostasis and can influence ion permeability of fungus cell walls.
β -acid radical has three kinds of homologues, namely co-lupulone (co-lupulone), n-lupulone (n-lupulone) and adlupulone (ad-lupulone) according to different acyl side chains in a molecular structure, because the content ratio of the ad-lupulone in the β -acid in the hop is relatively stable, and the separation difficulty of the n-lupulone and the ad-lupulone is higher, the n-lupulone and the ad-lupulone are generally researched as a whole, although the three kinds of homologues of β -acid have similar structures, the related research shows that the three kinds of homologues have certain difference in activity, at present, the separation method for the three kinds of homologues of β -acid mainly takes preparation type liquid chromatography separation as a main part, but has the following defects that an instrument is expensive, the obtained product quantity is small, and the value for practical application is not high.
Disclosure of Invention
The invention aims to overcome the defects of the prior art for separating homologs, and provide a method for extracting and separating β -acid with low cost and high yield and obtaining β -acid homolog products with different proportions, aiming at the characteristics that α -acid in a hop extract has strong bitter taste and can not be directly applied to most food systems, β -acid has strong bacteriostatic activity and no bitter taste and can be applied to a plurality of food systems except beer after separation and purification, and aiming at the difference of bacteriostatic activity among the hop homologs, the obtained homologs with different proportions can be applied to corresponding systems according to the characteristics, so that the application potential is improved.
In order to achieve the above object, the present invention is achieved by the following technical means.
A method for extracting and separating hop β -acid homologue is characterized by comprising the following steps:
(1) in the hop CO2Adding ethanol with the mass 0.5-1 times of the extract into the extract, placing the mixture into a water bath at 50-60 ℃, uniformly mixing, then dropwise adding 10-20% of alkaline aqueous solution under stirring to adjust the pH to 12-14, stopping dropwise adding, and then adding CO2Fully stirring distilled water 8-15 times the mass of the extract, measuring the pH value of the solution, adjusting the pH value to 12-14 by using alkaline water solution with the mass concentration of 5-25%, and standing for 20-40 min; filtering, separating and removing insoluble oily substances in the solution to obtain clear filtrate; slowly introducing CO into the filtrate under stirring2Continuously stirring for 20-40min, standing for 30min, filtering, collecting separated solid, washing with distilled water until filtrate is neutral, drying, and collecting separated solid which is β -acid crude extract;
(2) placing the β -acid crude extract in step (1) in a clean beaker, slowly dropwise adding 90% methanol aqueous solution on a water bath at 50 ℃ until β -acid crude extract is dissolved, quickly filtering, standing the filtrate at 4 ℃ for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with 90% methanol aqueous solution at 50 ℃, and repeating the crystallization step for 2 times to obtain β -acid homolog I;
(3) taking β -acid homolog I obtained in the step (2), slowly dropwise adding 90% methanol aqueous solution at 50 ℃ again until β -acid homolog I is completely dissolved, standing at room temperature for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with 90% methanol aqueous solution at 50 ℃ again, and repeating the crystallization step for 2 times to obtain β -acid homolog II;
(4) taking β -acid homologue I in the step (2), slowly dripping 90% methanol aqueous solution at 50 ℃ again until homologue I is completely dissolved, and then slowly introducing CO2Regulating pH to 7-7.5, stopping introducing gas after a small amount of crystals are separated out, continuing stirring for 10-30min, standing for 30min, filtering to obtain crystals, washing the crystals with distilled water until the filtrate is neutral, and drying to obtain β -acid homolog III;
(5) taking a proper amount of β -acid crude extract obtained in the step (1), slowly dripping n-hexane solution at 50 ℃ until β -acid crude extract is dissolved, quickly filtering, standing filtrate at 4 ℃ for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with n-hexane at 50 ℃ and recrystallizing for 2 times to obtain β -acid homolog IV.
The extraction and separation method of the hop β -acid homologue is characterized in that CO is slowly introduced in the steps (1) and (4)2The flow rate of the gas is 0.1-1.0 mL/min.
The extraction and separation method of the hop β -acid homologues is characterized in that the mass fraction of β -acid in the β -acid crude extract obtained by separation in the step (1) is not less than 80%.
The extraction and separation method of the hop β -acid homologue is characterized in that the mass fraction of β -acid in the homologue I in the step (2) is not less than 98%, wherein the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 1: 1.
The extraction and separation method of the hop β -acid homologue is characterized in that the mass fraction of β -acid in the homologue II in the step (3) is not less than 98%, wherein the mass ratio of the homologue co-lupulone to the homologue n + ad-lupulone is 10: 1.
The extraction and separation method of the hop β -acid homologue is characterized in that the mass fraction of β -acid in the homologue III in the step (4) is not less than 98%, wherein the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 1: 4.
The extraction and separation method of the hop β -acid homologue is characterized in that the mass fraction of β -acid in the homologue IV in the step (5) is not less than 98%, wherein the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 20: 1.
The extraction and separation method of the hop β -acid homologue is characterized in that the alkaline aqueous solution in the step (1) is an aqueous solution of NaOH or KOH, preferably an aqueous solution of KOH, and the preferred concentration is 10-20% by mass.
The separated product β -acid crude extract and the homolog I, homolog II, homolog III and homolog IV thereof obtained by the method for extracting and separating the hop β -acid homolog have the activity of inhibiting the food spoilage bacteria, have different bacteriostatic activities of the homologs, and can be added into different foods or material systems to be used as bacteriostatic agents according to needs.
The beneficial effects obtained by the invention are as follows:
the method adopted by the invention is simple, cheap and easy to operate, and is very suitable for extracting and separating a large amount of β -acid homologue products with different proportions, and β -acid obtained by the invention is separated and discarded as a crop byproduct in the deep processing process of the hop, the waste of raw materials can be greatly reduced by utilizing the method, and the production cost of enterprises is reduced.
Detailed Description
The present invention will be specifically explained below by way of specific examples, but the present invention is not limited to the examples.
Example 1
The liquid CO of the hop containing α -acid 21.46% and β -acid 66.24% by analysis2Adding 60mL of anhydrous ethanol into 100g of the extract, and placing in a water bath at 50 ℃ for pre-treatmentHeating to a flowing state, dropwise adding a KOH aqueous solution with the mass concentration of 10% while stirring to adjust the pH value to 13, stopping dropwise adding, adding 800mL of distilled water, fully stirring, adjusting the pH value to 13 again by using the KOH aqueous solution with the mass concentration of 10%, standing for 30min, filtering, and removing insoluble substances in the solution to obtain a filtrate; introducing CO at a flow rate of 0.6mL/min into the filtrate2And (3) stopping aeration when the pH value is adjusted to 8.4 by gas, separating out β -acid in the filtrate, and filtering to obtain 50g of β -acid crude extract, wherein the β -acid content is 87% by analysis.
Example 2
Taking 15g of β -acid crude extract, dripping 90% methanol aqueous solution at 50 ℃ until β -acid crude extract is completely dissolved, quickly filtering, standing the filtrate at 4 ℃ for 1.5h, filtering, collecting precipitated crystals, dripping 90% methanol aqueous solution at 50 ℃ again to the obtained crystals for redissolution, and repeating the crystallization process for 2 times to obtain homolog I.
Example 3
5g of homologue I is taken, 90 percent methanol aqueous solution at 50 ℃ is dripped until the homologue I is completely dissolved, then the homologue I is recrystallized for 1.5h at room temperature, crystal is obtained by filtration, and the process is repeated for 2 times to obtain the homologue II.
Example 4
2g of homologue I is taken, 90 percent methanol aqueous solution at 50 ℃ is dripped until the homologue I is completely dissolved, and then CO with the flow rate of 0.6mL/min is introduced2And (4) stopping aeration when the pH is adjusted to 7.2, and filtering to obtain the homologue III.
Example 5
5g of β -acid crude extract is taken, n-hexane solution at 50 ℃ is dripped into β -acid crude extract for dissolving, the mixture is quickly filtered, filtrate is kept still for 1.5h at 4 ℃, the filtered solution is filtered, precipitated crystals are collected, the obtained crystals are dripped into 90% methanol water solution at 50 ℃ again for redissolution, and the crystals are repeated for 2 times to obtain homolog IV.
Example 6
The compositions of the homologues I-homologues IV obtained in examples 2 to 5, as analyzed by liquid chromatography, are shown in table 1 for the respective homologue samples:
TABLE 1 composition of the components of homologue I-homologue IV
Example 7
The bacteriostatic action of methanol solutions of homologues I-IV with different concentrations in examples 2-5 on staphylococcus aureus and escherichia coli is examined by using a zone of inhibition method, which shows that β -acid samples with different proportions and the same family have certain difference in bacteriostatic activity, the size of the bacteriostatic activity is expressed by the diameter of the zone of inhibition, the sample is considered to have the inhibitory action on strains when the diameter is more than 6.5mm, and the data are shown in table 2:
TABLE 2 results of bacteriostatic action of the samples
While particular embodiments of the present invention have been illustrated and described, the present invention is not limited to the particular embodiments described above, and modifications or changes may be made by those skilled in the art in light of the present teachings without departing from the spirit of the present invention. The scope of the invention should be determined from the following claims, which are to be construed in accordance with the accompanying claims.
Claims (9)
1. A method for extracting and separating β -acid homologues from hops is characterized by comprising the following steps:
in the hop CO2Adding ethanol with the mass 0.5-1 times of the extract into the extract, placing the mixture into a water bath at 50-60 ℃, uniformly mixing, then dropwise adding 5-25% of alkaline aqueous solution under stirring to adjust the pH to 12-14, stopping dropwise adding, and then adding CO2Fully stirring distilled water 5-20 times the mass of the extract, measuring the pH value of the solution, adjusting the pH value to 12-14 by using alkaline aqueous solution with the mass concentration of 5-25%, and standing for 20-40 min; filtering, separating and removing insoluble oily substances in the solution to obtain clear filtrate; slowly introducing CO into the filtrate under stirring2Gas, adjusting pH to 8.0-8.5, stopping ventilation when the filtrate is turbid and a small amount of precipitate is separated out, continuing stirring for 20-40min, standing for 30min, filtering, collecting separated solid, washing with distilled water until the filtrate is neutral, drying, and collecting the separated solid, namely β -acid crude extract;
placing the β -acid crude extract in step (1) in a clean beaker, slowly dropwise adding 90% methanol water solution on a water bath at 50 ℃ until β -acid crude extract is dissolved, quickly filtering, standing the filtrate at 4 ℃ for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with 90% methanol water solution at 50 ℃, and repeating the crystallization step for 2 times to obtain β -acid homolog I;
taking β -acid homolog I obtained in the step (2), slowly dropwise adding 90% methanol aqueous solution at 50 ℃ again until β -acid homolog I is completely dissolved, standing at room temperature for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with 90% methanol aqueous solution at 50 ℃ again, and repeating the crystallization step for 2 times to obtain β -acid homolog II;
taking β -acid homologue I in the step (2), slowly dripping 90% methanol aqueous solution at 50 ℃ again until homologue I is completely dissolved, and then slowly introducing CO2Regulating pH to 7-7.5, stopping introducing gas after a small amount of crystals are separated out, continuing stirring for 10-30min, standing for 30min, filtering to obtain crystals, washing the crystals with distilled water until the filtrate is neutral, and drying to obtain β -acid homolog III;
taking a proper amount of β -acid crude extract obtained in the step (1), slowly dripping n-hexane solution at 50 ℃ until β -acid crude extract is dissolved, quickly filtering, standing filtrate at 4 ℃ for 1-2h, separating out crystals, filtering to obtain crystals, redissolving the crystals with n-hexane at 50 ℃ and recrystallizing for 2 times to obtain β -acid homolog IV.
2. The method of claim 1, wherein the slow CO introduction in step (1) and step (4)2The flow rate of the gas is 0.1-1.0 mL/min.
3. The method as claimed in claim 1, wherein the mass fraction of β -acids in the β -acid crude extract separated in step (1) is not less than 80%.
4. The method of claim 1, wherein the mass fraction of β -acid in the homologue I in the step (2) is not less than 98%, and the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 1: 1.
5. The method of claim 1, wherein the mass fraction of β -acid in the homolog II in the step (3) is not less than 98%, and the mass ratio of the homolog co-lupulone to the n + ad-lupulone is 10: 1.
6. The method of claim 1, wherein the mass fraction of β -acid in the homologue III in the step (4) is not less than 98%, and the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 1: 4.
7. The method of claim 1, wherein the mass fraction of β -acid in the homologue IV in the step (5) is not less than 98%, and wherein the mass ratio of the homologue co-lupulone to the n + ad-lupulone is 20: 1.
8. The method according to claim 1, wherein the alkaline aqueous solution in step (1) is an aqueous solution of NaOH or KOH, preferably an aqueous solution of KOH, preferably with a concentration of 10-20% by mass.
9. The method of claims 1-8, wherein the obtained β -acid crude extract and its homologues I, II, III and IV have activity in inhibiting food spoilage bacteria, and the homologues have different antibacterial activity, and can be added into different food or material systems as required to be used as antibacterial agent.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112409151A (en) * | 2020-11-03 | 2021-02-26 | 晨光生物科技集团股份有限公司 | Industrial method for extracting resin acid and xanthohumol from hops |
Citations (4)
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| JPH04202138A (en) * | 1990-11-30 | 1992-07-22 | Asahi Breweries Ltd | Hop extract having active oxygen scavenging action and its utilization |
| CN101768068A (en) * | 2010-01-06 | 2010-07-07 | 刘玉英 | Method utilizing hop residues to extract xanthohumol |
| CN104404058A (en) * | 2014-11-05 | 2015-03-11 | 中国科学院遗传与发育生物学研究所 | Protein for preparing beta-picric acid, and application thereof |
| CN104447264A (en) * | 2014-11-08 | 2015-03-25 | 商洛学院 | Method for extracting and separating lupulone from lupulus |
-
2019
- 2019-11-18 CN CN201911126750.2A patent/CN110746284A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH04202138A (en) * | 1990-11-30 | 1992-07-22 | Asahi Breweries Ltd | Hop extract having active oxygen scavenging action and its utilization |
| CN101768068A (en) * | 2010-01-06 | 2010-07-07 | 刘玉英 | Method utilizing hop residues to extract xanthohumol |
| CN104404058A (en) * | 2014-11-05 | 2015-03-11 | 中国科学院遗传与发育生物学研究所 | Protein for preparing beta-picric acid, and application thereof |
| CN104447264A (en) * | 2014-11-08 | 2015-03-25 | 商洛学院 | Method for extracting and separating lupulone from lupulus |
Non-Patent Citations (3)
| Title |
|---|
| CAROLIN SCHULZ 等: "Quantification of co-, n-, and ad-lupulone in hop-based dietarysupplements and phytopharmaceuticals and modulation of theircontents by the extraction method", 《JOURNAL OF PHARMACEUTICAL AND BIOMEDICAL ANALYSIS》 * |
| 刘玉梅: "啤酒花的分析评价及六氢蛇麻酮类的合成与应用", 《江南大学》 * |
| 李玉晶等: "啤酒花活性成分与抗氧化活性的相关性", 《食品科学》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112409151A (en) * | 2020-11-03 | 2021-02-26 | 晨光生物科技集团股份有限公司 | Industrial method for extracting resin acid and xanthohumol from hops |
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Application publication date: 20200204 |