CN108164415A - A kind of method that EPA and DHA are kept completely separate from fish oil - Google Patents
A kind of method that EPA and DHA are kept completely separate from fish oil Download PDFInfo
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- CN108164415A CN108164415A CN201711459972.7A CN201711459972A CN108164415A CN 108164415 A CN108164415 A CN 108164415A CN 201711459972 A CN201711459972 A CN 201711459972A CN 108164415 A CN108164415 A CN 108164415A
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- dha
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C51/00—Preparation of carboxylic acids or their salts, halides or anhydrides
- C07C51/42—Separation; Purification; Stabilisation; Use of additives
- C07C51/48—Separation; Purification; Stabilisation; Use of additives by liquid-liquid treatment
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Abstract
The present invention relates to a kind of method that EPA and DHA are kept completely separate from fish oil, including：By one shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；Fish oil extract is dissolved in upper phase, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, removes mobile phase, obtains the mixture of EPA and DHA；By two shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；Mixture is dissolved in upper phase, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, removes mobile phase, respectively obtains EPA and DHA products.The all recyclable recycling of reagent used by two dicyandiamide solutions of the present invention, theoretically EPA and DHA can 100% recycling.
Description
Technical field
The invention belongs to unrighted acid processing technique field, more particularly to it is a kind of be kept completely separate from fish oil EPA and
The method of DHA.
Background technology
DHA (docosahexaenoic acid, Docosahexaenoic Acid) is a kind of ω -3 essential fatty acids.Research shows that
DHA is the brain development of people, the important component of growth；Have emphatically improving senile dementia, anti-aging, antitumor etc.
The physiological regulation function wanted.
EPA (eicosapentaenoic acid, Eicosapentaenoic Acid) be the common several omega-fatty acids of human body it
One.EPA can help to reduce the content of cholesterol and triglycerides, promote internal saturated fat acid metabolic.Fat is prevented in blood vessel
The deposition of wall, the angiocardiopathies such as prevention cerebral thrombus, cerebral hemorrhage, hypertension.
Contain abundant DHA and EPA in fish oil.Successively invention employs low temperature staging, solvent extraction method, urea to people
The methods of inclusion method, molecularly distilled and supercritical gas extraction method, extracts DHA and EPA from organism.
The study found that EPA can destroy the balance of DHA and EPA in human body during infant's early development.Therefore, have
EPA and DHA are kept completely separate and come by necessity.
High speed adverse current chromatogram (HSCCC) technology is a kind of new separating and purifying technology based on liquid liquid distribution principle, it is not
Any solid state support or carrier are needed, stationary phase and mobile phase are all liquid, no Irreversible Adsorption.
Invention content
The technical problems to be solved by the invention are to provide a kind of method that EPA and DHA are kept completely separate from fish oil, use
High-speed countercurrent chromatography, in two steps dicyandiamide solution EPA and DHA are kept completely separate from fish oil, wherein dicyandiamide solution one can be complete
The mixture of fully separating EPA and DHA and other impurity；Dicyandiamide solution two can be kept completely separate EPA and DHA；Obtain the EPA of high-purity
And DHA.
A kind of method that EPA and DHA are kept completely separate from fish oil of the present invention, including：
(1) by one shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；Fish oil extract is dissolved in
In upper phase, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, obtain containing only EPA and
The mixture of DHA and the mixed liquor of mobile phase remove mobile phase, obtain the mixture of EPA and DHA；Wherein dicyandiamide solution one by
N-hexane or normal heptane, acetonitrile, methanol are 3~6 by volume：2~4：1~2 is obtained by mixing；
(2) by two shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；Step (1) is obtained mixed
Object is closed be dissolved in phase, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, respectively
To the mixed liquor and DHA of EPA and mobile phase and the mixed liquor of mobile phase, mobile phase is removed, respectively obtains EPA and DHA products；Its
Middle dicyandiamide solution two is 15~30 by volume by n-hexane or normal heptane, methanol, deionized water：10~20：2~4 mixing and
.
The upper and lower two-phase that separate collection obtains in the step (1) and (2) carries out ultrasonic degassing process.
The condition of step (1) the high speed adverse current chromatogram is：It turns to rotate forward, rotating speed 800rpm；Column temperature is 25 DEG C；
The flow velocity of mobile phase is 5mL/min；The Detection wavelength of detector is 214nm.
The process conditions of removal mobile phase in the step (1) are：Rotation steaming is carried out under the conditions of 45 DEG C, -0.08MPa
Hair vacuum drying.
The condition of step (2) the high speed adverse current chromatogram is：It turns to rotate forward, rotating speed 800rpm；Column temperature is 25 DEG C；
The flow velocity of mobile phase is 10mL/min；The Detection wavelength of detector is 214nm.
The process conditions of removal mobile phase in the step (2) are：Rotation steaming is carried out under the conditions of 50 DEG C, -0.08MPa
Hair vacuum drying.
Advantageous effect
(1) present invention for the first time using high-speed countercurrent chromatography from fish oil simultaneously isolate and purify to obtain the EPA of high-purity
And DHA.
(2) high-speed countercurrent chromatography of the invention has no sample loss, pollution-free, efficient, big preparation amount, and solvent can
The advantages that recycling.
(3) all recyclable recycling of the reagent of two dicyandiamide solutions used in the present invention, theoretically EPA and DHA can be real
Existing 100% recycling.
Specific embodiment
With reference to specific embodiment, the present invention is further explained.It should be understood that these embodiments are merely to illustrate the present invention
Rather than it limits the scope of the invention.In addition, it should also be understood that, after reading the content taught by the present invention, people in the art
Member can make various changes or modifications the present invention, and such equivalent forms equally fall within the application the appended claims and limited
Range.
The reagent used in the embodiment of the present invention and instrument are as follows：
Reagent：N-hexane, normal heptane, acetonitrile, methanol are that the analysis of Sinopharm Chemical Reagent Co., Ltd.'s production is pure
Reagent, water are deionized water, and fish oil extract is commercial product；
Instrument：High-speed counter-current chromatograph is high for the TBE-300C models of Shanghai Tauto Biotechnology Co., Ltd.'s production
Fast counter-current chromatograph.
Embodiment 1
(1) by n-hexane, acetonitrile, methanol is 6 by volume：4：1 is mixed to prepare dicyandiamide solution one, adds in separatory funnel
In, shake well stands split-phase, obtains two-phase mixtures liquid, and separate collection upper and lower phase solvent is respectively placed in supersonic oscillations
Ultrasonic degassing process is carried out in device, fish oil extract is dissolved in upper phase solvent, then above is mutually stationary phase, and lower phase is stream
Dynamic phase, is detached, wherein chromatographic condition is set as using high speed adverse current chromatogram：It rotates forward, rotating speed 800rpm；25 DEG C of column temperature；
Flow rate of mobile phase 5mL/min；The Detection wavelength of detector is 214nm, obtains containing only the mixture and mobile phase of EPA and DHA
Mixed liquor, be placed in rotary evaporator, under the conditions of 45 DEG C of bath temperature, vacuum pressure -0.08MPa carry out rotary evaporation it is true
Sky is dry, removes mobile phase, obtains the mixture of EPA and DHA.
(2) by n-hexane, methanol, deionized water is 20 by volume：20：3 are mixed to prepare dicyandiamide solution two, add in liquid separation
In funnel, shake well stands split-phase, obtains two-phase mixtures liquid, and separate collection upper and lower phase solvent is respectively placed in ultrasonic wave
Ultrasonic degassing process is carried out in oscillator, the EPA and the mixture of DHA that step (1) is obtained are dissolved in upper phase solvent, then
It is mutually above stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, wherein chromatographic condition is set as：It rotates forward,
Rotating speed is 800rpm；25 DEG C of column temperature；Flow rate of mobile phase 10mL/min；The Detection wavelength of detector is 214nm, respectively obtains EPA
It with the mixed liquor and DHA of mobile phase and the mixed liquor of mobile phase, is placed in rotary evaporator, in 50 DEG C of bath temperature, vacuum pressure
Rotary evaporation vacuum drying is carried out under the conditions of -0.08MPa by force, removes mobile phase, removes mobile phase, respectively obtains EPA and DHA productions
Product.
The purity of EPA and DHA products that the present embodiment obtains is analyzed by HPLC, is as a result shown：EPA purity is
99.62%；DHA purity is 99.60%.
Embodiment 2
(1) by normal heptane, acetonitrile, methanol is 6 by volume：4：1 is mixed to prepare dicyandiamide solution one, adds in separatory funnel
In, shake well stands split-phase, obtains two-phase mixtures liquid, and separate collection upper and lower phase solvent is respectively placed in supersonic oscillations
Ultrasonic degassing process is carried out in device, fish oil extract is dissolved in upper phase solvent, then above is mutually stationary phase, and lower phase is stream
Dynamic phase, is detached, wherein chromatographic condition is set as using high speed adverse current chromatogram：It rotates forward, rotating speed 800rpm；25 DEG C of column temperature；
Flow rate of mobile phase 5mL/min；The Detection wavelength of detector is 214nm, obtains containing only the mixture and mobile phase of EPA and DHA
Mixed liquor, be placed in rotary evaporator, under the conditions of 45 DEG C of bath temperature, vacuum pressure -0.08MPa carry out rotary evaporation it is true
Sky is dry, removes mobile phase, removes mobile phase, obtains the mixture of EPA and DHA.
(2) by normal heptane, methanol, deionized water is 20 by volume：20：3 are mixed to prepare dicyandiamide solution two, add in liquid separation
In funnel, shake well stands split-phase, obtains two-phase mixtures liquid, and separate collection upper and lower phase solvent is respectively placed in ultrasonic wave
Ultrasonic degassing process is carried out in oscillator, the EPA and the mixture of DHA that step (1) is obtained are dissolved in upper phase solvent, then
It is mutually above stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, wherein chromatographic condition is set as：It rotates forward,
Rotating speed is 800rpm；25 DEG C of column temperature；Flow rate of mobile phase 10mL/min；The Detection wavelength of detector is 214nm, respectively obtains EPA
It with the mixed liquor and DHA of mobile phase and the mixed liquor of mobile phase, is placed in rotary evaporator, in 50 DEG C of bath temperature, vacuum pressure
Rotary evaporation vacuum drying is carried out under the conditions of -0.08MPa by force, mobile phase is removed, respectively obtains EPA and DHA products.
The purity of EPA and DHA products that the present embodiment obtains is analyzed by HPLC, is as a result shown：EPA purity is
96.57%；DHA purity is 97.18%.
Claims (6)
1. a kind of method that EPA and DHA are kept completely separate from fish oil, including：
(1) by one shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；Fish oil extract is dissolved in upper phase
In, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, obtains containing only EPA's and DHA
The mixed liquor of mixture and mobile phase removes mobile phase, obtains the mixture of EPA and DHA；Wherein dicyandiamide solution one is by n-hexane
Or normal heptane, acetonitrile, methanol are 3~6 by volume：2~4：1~2 is obtained by mixing；
(2) by two shake well of dicyandiamide solution, split-phase, the upper and lower two-phase of separate collection are stood；The mixture that step (1) is obtained
Be dissolved in phase, more than mutually for stationary phase, lower phase is mobile phase, is detached using high speed adverse current chromatogram, respectively obtains EPA
With the mixed liquor and DHA of mobile phase and the mixed liquor of mobile phase, mobile phase is removed, respectively obtains EPA and DHA products；It is wherein molten
Agent system two is 15~30 by volume by n-hexane or normal heptane, methanol, deionized water：10~20：2~4 are obtained by mixing.
2. a kind of method that EPA and DHA are kept completely separate from fish oil according to claim 1, it is characterised in that：The step
Suddenly the upper and lower two-phase that separate collection obtains in (1) and (2) carries out ultrasonic degassing process.
3. a kind of method that EPA and DHA are kept completely separate from fish oil according to claim 1, it is characterised in that：The step
Suddenly the condition of (1) high speed adverse current chromatogram is：It turns to rotate forward, rotating speed 800rpm；Column temperature is 25 DEG C；The flow velocity of mobile phase is
5mL/min；The Detection wavelength of detector is 214nm.
4. a kind of method that EPA and DHA are kept completely separate from fish oil according to claim 1, it is characterised in that：The step
Suddenly the process conditions of removal mobile phase are in (1)：Rotary evaporation vacuum drying is carried out under the conditions of 45 DEG C, -0.08MPa.
5. a kind of method that EPA and DHA are kept completely separate from fish oil according to claim 1, it is characterised in that：The step
Suddenly the condition of (2) high speed adverse current chromatogram is：It turns to rotate forward, rotating speed 800rpm；Column temperature is 25 DEG C；The flow velocity of mobile phase is
10mL/min；The Detection wavelength of detector is 214nm.
6. a kind of method that EPA and DHA are kept completely separate from fish oil according to claim 1, it is characterised in that：The step
Suddenly the process conditions of removal mobile phase are in (2)：Rotary evaporation vacuum drying is carried out under the conditions of 50 DEG C, -0.08MPa.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
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| CN201711459972.7A CN108164415B (en) | 2017-12-28 | 2017-12-28 | Method for completely separating EPA and DHA from fish oil |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201711459972.7A CN108164415B (en) | 2017-12-28 | 2017-12-28 | Method for completely separating EPA and DHA from fish oil |
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| CN108164415B CN108164415B (en) | 2021-07-09 |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110590545A (en) * | 2019-09-11 | 2019-12-20 | 上海同田生物技术股份有限公司 | Method for completely separating oleic acid and linoleic acid |
| CN111362790A (en) * | 2020-04-17 | 2020-07-03 | 辽宁科技大学 | Chromatographic method for separating EPA and DHA |
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| CN102285880A (en) * | 2011-06-14 | 2011-12-21 | 国家海洋局第三海洋研究所 | Method for preparing ethyl eicosapentaenate (EPA) and ethyl docosahexaenoate (DHA) |
| CN103086873A (en) * | 2013-01-11 | 2013-05-08 | 国家海洋局第三海洋研究所 | Preparation method of high-purity DHA (Docosahexaenoic Acid) by means of high-speed counter-current chromatography separation |
| CN103787862A (en) * | 2012-10-31 | 2014-05-14 | 江苏汉邦科技有限公司 | Preparation method for separating and purifying EPA and DHA |
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2017
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Patent Citations (3)
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| CN103787862A (en) * | 2012-10-31 | 2014-05-14 | 江苏汉邦科技有限公司 | Preparation method for separating and purifying EPA and DHA |
| CN103086873A (en) * | 2013-01-11 | 2013-05-08 | 国家海洋局第三海洋研究所 | Preparation method of high-purity DHA (Docosahexaenoic Acid) by means of high-speed counter-current chromatography separation |
Non-Patent Citations (3)
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110590545A (en) * | 2019-09-11 | 2019-12-20 | 上海同田生物技术股份有限公司 | Method for completely separating oleic acid and linoleic acid |
| CN110590545B (en) * | 2019-09-11 | 2023-08-29 | 上海同田生物技术股份有限公司 | Method for completely separating oleic acid and linoleic acid |
| CN111362790A (en) * | 2020-04-17 | 2020-07-03 | 辽宁科技大学 | Chromatographic method for separating EPA and DHA |
| CN111362790B (en) * | 2020-04-17 | 2022-12-13 | 辽宁科技大学 | Chromatographic method for separating EPA and DHA |
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| CN108164415B (en) | 2021-07-09 |
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Address after: Room 101-2102-1301-1, 326 Edison Road, Zhangjiang hi tech park, China (Shanghai) pilot Free Trade Zone, Pudong New Area, Shanghai, 201203 Patentee after: SHANGHAI TAUTO BIOTECH Co.,Ltd. Address before: Room 101-2102-1301-1, 326 Edison Road, Zhangjiang hi tech park, China (Shanghai) pilot Free Trade Zone, Pudong New Area, Shanghai, 201203 Patentee before: SHANGHAI TAUTO BIOTECH Co.,Ltd. |