CN101392272A - Method for enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system - Google Patents
Method for enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system Download PDFInfo
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- CN101392272A CN101392272A CNA2008102022531A CN200810202253A CN101392272A CN 101392272 A CN101392272 A CN 101392272A CN A2008102022531 A CNA2008102022531 A CN A2008102022531A CN 200810202253 A CN200810202253 A CN 200810202253A CN 101392272 A CN101392272 A CN 101392272A
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Abstract
The invention relates to a method for producing caffeic acid phenethyl ester by carrying out the enzymatic synthesis of a solvent-free system, which comprises the steps: (1) a 4-type molecular sieve and phenethyl alcohol are placed in a triangular flask according to a mass ratio of 1 to 5 and protected form light; and after shaking on a shaker for 48h, the molecular sieve is removed, thus obtaining anhydrous phenethyl alcohol; (2) caffeic acid reacts with the phenethyl alcohol for 32h to 60h under the catalytic reaction of Novozym 435(lipase from C.antarctica on macroporous acrylic resin) lipase; the mole ratio of the caffeic acid and the phenethyl alcohol is 1: 86 to 1: 96, and reaction temperature is controlled to be 60 DEG C to 70 DEG C; and (3) after reaction, a biological catalyst is removed by filtering; the residual phenethyl alcohol is removed by rotary evaporation, thus obtaining caffeic acid phenethyl ester after the isolation and purification of column chromatography. The synthesizing method of the invention promotes the concentration of substrates and products and reaction selectivity; the yield rate of the caffeic acid phenethyl ester is up to 90 percent approximately; the reaction condition of the method is safe and mild and purification process is easy and has few steps, thus being convenient to industrialized production.
Description
Technical field
The invention belongs to the preparation field of CAPE, particularly relate to a kind of method of enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system.
Background technology
CAPE is the major ingredient of propolis, also is present in the Chinese medicines such as Radix Angelicae Sinensis, Ligusticum wallichii simultaneously.Coffic acid content is also higher in food raw material such as coffee, chinese cymbidium beans, husk, bagasse, beet pulp, wheat bran and rice bran.
Recently, CAPE in a plurality of fields such as food by extensive studies and application.This mainly is because CAPE not only has good oxidation-resistance, and physiologically active is widely arranged, as anti-inflammatory, pain relieving, adjusting body immunity, simultaneously can be promoting blood circulation and removing obstruction in channels, dispersing the stagnated live-QI to relieve the stagnation of QI, strengthening spleen and nourishing stomach, and multiple effect such as stimulate circulation.And CAPE also is international cognitive in recent years anti-cancer material, causes that common people gaze at.
Except that being widely used in pharmaceutical industries, coffic acid also enlarges day by day in the application of food service industry.Because CAPE has anti-oxidant activity and bacteriostatic action, it also can be used as the antisepsis antistaling agent of food, and safety non-toxic; CAPE also little by little is applied to prepare food glue and edible packing membrane as linking agent; In addition, it promotes that as antioxidant and function material can be applicable in the motion food.
Yet up to the present obtaining of CAPE still adopted the method for extracting from natural product.Have many difficulties in this process, long as extraction time, leaching process with an organic solvent causes serious harm to environment in a large number; In addition, cause costing an arm and a leg because productivity ratio is low, and can not satisfy the needs in market to a great extent, therefore a lot of research groups are devoted to prepare CAPE by the synthetic method.
Adopt the synthetic CAPE of chemical method also to run into huge obstacle: the reaction conditions that the chemical synthesis by product is many, product yield is low, violent usually can destroy the structure of natural compounds.In addition, in the process of chemosynthesis, need to use expendable heavy metal catalyst, not only toxic to workman's health, and also also very unfriendly to environment, do not meet the green friendly chemical theme of advocating now.
Summary of the invention
The method that the purpose of this invention is to provide a kind of enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system, synthetic method of the present invention has improved the selectivity of substrate and production concentration and reaction, the productive rate of CAPE is up to about 90%, this method reaction conditions safety, gentleness, purge process is easy, step is few, is convenient to suitability for industrialized production.
The method of a kind of enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system of the present invention comprises:
(1) will
Type molecular sieve and phenylethyl alcohol place airtight triangular flask by mass ratio 1:5, and lucifuge is removed molecular sieve behind the shaking table vibration 48h, obtains dry-out benzene ethanol;
(2) with coffic acid and phenylethyl alcohol 1:86-1:96 in molar ratio, reaction 32-60h under Novozym435 lipase (lipase from C.antarctica on macroporous acrylic resin) catalysis, temperature of reaction is controlled at 60-70 ℃;
(3) after the reaction, remove by filter biological catalyst, rotary evaporation is removed remaining phenylethyl alcohol, through column chromatographic isolation and purification, obtains CAPE.
Under purifying in the step (3) be purification by silica gel column chromatography, solvent is a normal hexane, wash-out stage by stage, the elutriant ratio is from benzene: ether is that 10:3 increases to benzene gradually: the ether ratio is 1:10.
Concrete reaction formula is as follows:
Solvent-free system enzymic catalytic reaction of the present invention is under the condition that does not contain organic solvent simple mixing of reactant to be carried out enzymic catalytic reaction (substrate exists with solid or fusing form), it has whole advantages of organic phase enzymic catalytic reaction, as: the thermodynamics of reactions balance is moved to compound direction by hydrolysis, enzyme has the stability of height etc., it is big simultaneously to have overcome organic solvent toxicity again, inflammable volatile, environment is polluted, reclaim and recycle shortcomings such as cost height.Simultaneously, solvent-free synthesize the reaction the new branch subenvironment different with conventional solvent is provided, in solvent-free system, enzyme directly acts on substrate.
Enzyme catalysis efficient height of the present invention exceeds 10 than general chemical catalysis
6～10
13Doubly.In enzymic catalytic reaction, avoided radical protection and de-protected operation in chemosynthesis, shortened the synthetic step, improved the yield of target product, simultaneously, reduced synthetic cost.Productive rate with the synthetic CAPE of enzyme process catalysis has been brought up to about 90% by about 30% of chemical synthesis, and the while experimental procedure is compared chemical synthesis and is simplified.
Beneficial effect
(1) synthetic method of the present invention has improved the selectivity of substrate and production concentration and reaction, and the productive rate of CAPE is up to about 90%;
(2) this method reaction conditions safety, gentleness, control easily, side reaction is few, and suitability for industrialized production is convenient in the environmental pollution of having avoided the organic solvent in the chemical synthesis to bring simultaneously.
Description of drawings
Fig. 1 is the structural representation of CAPE of the present invention;
Fig. 2 is the CAPE tlc analysis figure before the purifying;
Fig. 3 is the CAPE tlc analysis figure behind the purifying.
Embodiment
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in and limit the scope of the invention.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after the content of having read the present invention's instruction, these equivalent form of values fall within the application's appended claims institute restricted portion equally.
(1) gets 50 gram phenylethyl alcohols and 10 grams
Molecular sieve places in the 100mL tool plug triangular flask, is placed in 50 ℃ of air shaking tables, and 180rpm fully vibrates behind the 48h, removes by filter molecular sieve, obtains dry-out benzene ethanol;
(2) enzymic catalytic reaction adds the 2.0mmoL coffic acid in the tool plug triangular flask of 50mL, the 184mmoL phenylethyl alcohol, and 120mg Novozym435 lipase, vibration 50r/min reaction 48h is bathed in oil bath under 70 ℃ of degree;
(3) rotary evaporation was removed unreacted phenylethyl alcohol after reaction stopped, and purifying.Purification condition: dress post: get 25～30g silica gel G, take the normal hexane as solvent wet method dress post, the control flow velocity is 1/s.Chromatography condition: adopt the method for wash-out stage by stage, elutriant ratio control flow velocity 5min connects effluent liquid, and from benzene: ether=gradually increase to benzene at 10: 3: the ether ratio is 1:10.
Every pipe 5mL; Every pipe sample is got 5 μ L by test tube number order point sample successively on same block of thin layer plate, is scanning and processing data under the 254nm with dual wavelength thin layer chromatography scanner at wavelength.Merge test tube solution in the CAPE peak scope, obtain white crystalline CAPE behind the purifying behind the rotary evaporation, the tlc analysis result sees Fig. 2 before and after the purifying.
Claims (2)
1. the method for an enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system comprises:
(1) will
Type molecular sieve and phenylethyl alcohol place airtight triangular flask by mass ratio 1:5, and lucifuge is removed molecular sieve behind the shaking table vibration 48h, obtains dry-out benzene ethanol;
(2) with coffic acid and phenylethyl alcohol 1:86-1:96 in molar ratio, at the lipase-catalyzed reaction down of Novozym 435 (lipase from C.antarcticaon macroporous acrylic resin) 32-60h, temperature of reaction is controlled at 60-70 ℃;
(3) after the reaction, remove by filter biological catalyst, rotary evaporation is removed remaining phenylethyl alcohol, through column chromatographic isolation and purification, obtains CAPE.
2. the method for a kind of enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system according to claim 1, it is characterized in that: the purifying in the affiliated step (3) is a purification by silica gel column chromatography, solvent is a normal hexane, wash-out stage by stage, elutriant ratio are from benzene: ether is that 10:3 increases to benzene gradually: the ether ratio is 1:10.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2008102022531A CN101392272A (en) | 2008-11-05 | 2008-11-05 | Method for enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system |
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| CNA2008102022531A CN101392272A (en) | 2008-11-05 | 2008-11-05 | Method for enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system |
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| CN101392272A true CN101392272A (en) | 2009-03-25 |
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| CNA2008102022531A Pending CN101392272A (en) | 2008-11-05 | 2008-11-05 | Method for enzymatic synthesis of caffeic acid phenethyl ester by solvent-free system |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102321692A (en) * | 2011-09-27 | 2012-01-18 | 江苏科技大学 | Preparation method for catalytic synthesis of caffeic acid phenethyl ester by using lipase in ionic liquid |
| CN103088078A (en) * | 2013-02-04 | 2013-05-08 | 江苏科技大学 | Lipase synthesis-promoting method of propyl caffeate |
-
2008
- 2008-11-05 CN CNA2008102022531A patent/CN101392272A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102321692A (en) * | 2011-09-27 | 2012-01-18 | 江苏科技大学 | Preparation method for catalytic synthesis of caffeic acid phenethyl ester by using lipase in ionic liquid |
| CN103088078A (en) * | 2013-02-04 | 2013-05-08 | 江苏科技大学 | Lipase synthesis-promoting method of propyl caffeate |
| CN103088078B (en) * | 2013-02-04 | 2015-07-01 | 江苏科技大学 | Lipase synthesis-promoting method of propyl caffeate |
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Application publication date: 20090325 |