CN103304801A - Method for preparing theabrownin - Google Patents
Method for preparing theabrownin Download PDFInfo
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- CN103304801A CN103304801A CN2012100668771A CN201210066877A CN103304801A CN 103304801 A CN103304801 A CN 103304801A CN 2012100668771 A CN2012100668771 A CN 2012100668771A CN 201210066877 A CN201210066877 A CN 201210066877A CN 103304801 A CN103304801 A CN 103304801A
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Abstract
The invention provides a method for preparing a similar substance of theabrownin by taking gallic acid as a raw material. The method particularly comprises the following steps of: 1, adding the gallic acid into ammonia water for reaction to obtain crude theabrownin; 2, extracting the crude theabrownin by ethyl acetate to obtain a water layer; 3, extracting the water layer by normal butanol to obtain a water layer; and 4, drying to obtain the theabrownin. By adopting the method, the problem of high consumption of puer tea for extracting the theabrownin in a conventional process is solved.
Description
Technical field:
The present invention relates to a kind of preparation method of theabrownin, particularly is the method for feedstock production theabrownin with the gallic acid.
Background technology:
Polyphenols in the tealeaves is the important active substances in the tealeaves, it is the general name of multiple phenolic compound in the tealeaves, it is the main moiety of fresh leaves of tea plant, be its main body composition with catechin wherein, account for 60%～80% of polyphenols total amount, with soup look, flavour and the fragrance of tea close relationship is arranged.
The water-soluble products of polyphenols mainly is theoflavin, thearubigins and theabrownin.Theabrownin is the general name of the very complicated product of a class formation of catechin oxypolymerization formation, 80% theoflavin and thearubigins oxidation, polymerization in the course of processing of Leaf of Assam Tea, form theabrownin, and its content is multiplied, thereby the convergency of millet paste and bitter taste are obviously reduced, add higher sugar and solubility water extraction content, this flavour that has just formed Leaf of Assam Tea is mellow, the basic substance that the soup look reddish brown.
From the soup look of tea, theoflavin is the important component of toning " bright ", but theoflavin a large amount of minimizing owing to oxypolymerization in the tea course of processing; Thearubigins is the important component of red liquor color " red ", the essential substance of flavour intensity, and relevant with the concentration of millet paste; Theabrownin is soup look dark major cause, can present reddish brown bright qualitative characteristics when theabrownin content reaches 6%～8%, Tang Se.When theabrownin content is lower than 5%, mean insufficient fermentation, the soup look presents the bright feature of blood orange.
Chinese patent 200910094074 report, theabrownin are to be raw material with black tea or Leaf of Assam Tea, extract the natural product that obtains by method of extraction from tealeaves.Pharmacological tests shows that theabrownin has the function of arrestin matter tyrosine phosphatase significantly.Can be used for treatment or prevention type ii diabetes, treatment or prevention of obesity diabetes, treatment or prevention of obesity disease.Owing to be the pure natural substance of from tealeaves, extracting, have and have no side effect, take safe and reliable advantage.
Relevant document has also been reported the preparation method of theabrownin, and as chemistry of forest product and the article of industrial magazine the 5th phase of October in 2009, the extraction process of theabrownin of Pu'er tea and the preliminary study of physico-chemical property have been described the method that obtains theabrownin from Leaf of Assam Tea.
Gallic acid chemical name: 3,4, the 5-trihydroxybenzoic acid, have another name called gallic acid, it has following effect: 1. anti-bacteria and anti-virus: external streptococcus aureus, sarcina, α-type streptococcus, Neisseria gonorrhoeae, Pseudomonas aeruginosa, bacterium flexneri, Corynebacterium diphtheriae Hd, paratyphosum Bacterium A etc. are had restraining effect, its Mlc is 5mg/ml.External, under 3% concentration, 17 kinds of fungies there is bacteriostatic action, influenza virus also there is certain restraining effect.2. antitumor: the mouse lung adenoma that morpholine is added due to the sodium nitrite has strong restraining effect.
The present invention is through research, and finding a kind of is the method for preparing the theabrownin similar substance of raw material with gallic acid, has avoided obtaining from Leaf of Assam Tea the problem of theabrownin mass consumption Leaf of Assam Tea.
Summary of the invention:
The invention provides a kind of preparation method of theabrownin, this method may further comprise the steps:
Step 2, theabrownin crude product ethyl acetate extraction obtains water layer;
Step 3, water layer are used n-butanol extraction again, obtain water layer;
Preparation method of the present invention, wherein
Step 2, ethyl acetate extraction 1-3 time of theabrownin crude product, preferred 2 times,
Step 3, water layer are used n-butanol extraction 1-3 time again, and preferred 2 times,
Wherein gallic acid can be l-Epicatechol, catechin or pyrogallol.
The particularly preferred preparation method of the present invention sees embodiment.
It is similar that the theabrownin that theabrownin similar substance and the prior art that the present invention obtains obtains is compared physico-chemical property, can be used as the substitute of natural theabrownin.Below test illustrates the consistence of theabrownin of the present invention and natural theabrownin.
One, character simultaneous test
1, pH value determination test
1.1 the preparation of need testing solution
Take by weighing traditional theabrownin and self-control theabrownin (theabrownin of the embodiment of the invention 1 method preparation), add water and make the mixing solutions that every 1ml contains 1mg.
1.2pH pH-value determination pH
1.2.1 instrument calibration
Schedule of operation is undertaken by the instrument specification sheets, and sample preparation liquid and standard buffer solution are transferred to same temperature.
1.2.2 sample determination
In glass or plastic containers, add sample preparation liquid, make the enough submergence electrodes of its capacity, measure need testing solution with the pH determinator, and record pH value, result such as following table;
Table 1pH pH-value determination pH result
| The sample title | The pH value | Temperature |
| The tradition theabrownin | 6.22 | 21.6℃ |
| The self-control theabrownin | 6.32 | 21.6℃ |
2, theabrownin Infrared spectroscopy
Tradition theabrownin and self-control theabrownin in the Fourier transformation infrared spectrometer scanning analysis, scan wave-number range 450～4000cm respectively through the KBr compressing tablet
-1, resolving power 4cm
-1, scanning times 16, sweep velocity 1.0cm/s, the gained infrared spectrogram is seen Fig. 1 and Fig. 2.
3, ultraviolet full wavelength scanner
3.1 the preparation of need testing solution
Traditional theabrownin and self-control theabrownin respectively with the dissolved in distilled water that boils, are prepared into certain density solution, take out in right amount, with the n-butanol extraction of equivalent, water intaking layer 2ml is settled to scale with 95% ethanol in the 25ml volumetric flask, be need testing solution after shaking up.
3.2 ultraviolet full wavelength scanner
The need testing solution that makes in 3.1 is packed in the cuvette in right amount, do blank solution with 95% ethanol, put into the ultraviolet-visible spectrophotometer, (200nm～700nm) the results are shown in Figure 3 and Fig. 4 to carry out full wavelength scanner.
4, GPC stratographic analysis
4.1 method:
Analyze and adopt gel permeation chromatography in ten anistree laser light scattering instruments and the coupling of differential detector.Adding 0.2% sodiumazide with the 0.1mol/L sodium nitrate solution is moving phase, 40 ℃ of column temperatures; Differential refraction detector; 40 ℃ of temperature; Laser detection wavelength 658nm; Flow velocity is 0.5mL/min; Sampling volume: 200 μ L.
4.2 result:
As stated above traditional theabrownin component (removing albumen and polysaccharide) and self-control theabrownin component are carried out gpc analysis, the gained color atlas is seen Fig. 5 and Fig. 6, and molecular weight distribution the results are shown in Table 2
The traditional theabrownin component of table 2 (removing albumen and polysaccharide) and self-control theabrownin component Mr distribute
Two, interpretation of result
1, pH pH-value determination pH
As shown in Table 1, the pH value of traditional theabrownin and self-control theabrownin similar substance is respectively 6.22 and 6.32, and is very approaching, and all becomes slightly acidic.
2, Infrared spectroscopy
As depicted in figs. 1 and 2, the theabrownin mixture is at 3374cm
-1About the strong wide absorption peak located be the O-H stretching vibration peak, have intermolecular and intramolecular hydrogen bond, at 3600～3000cm
-1The broad peak that occurs also may comprise the N-H stretching vibration; 2937cm
-1The place is the C-H stretching vibration peak of methylene radical; All there is the eigen vibration of above-mentioned functional group in two samples.1614cm
-1The place is the stretching vibration peak of C=0.In conjunction with 1531cm
- 1The absorption peak at place can judge that this place may also comprise the aromatic ring frame vibration and cause absorption; 1403cm
-1The place is the symmetrical stretching vibration peak of COO-; 1306cm
-1The place should be aromatic C-N stretching vibration peak, 1242cm
- 1The place is the C-O stretching vibration peak of phenol; 1069,1046cm
-1Two places are the stretching vibration peak of C-O-C, should be the characteristic absorbance of glucosides; Two samples differ greatly at this wave band, and the self-control theabrownin does not have the glycosidic link vibration performance, because the participation of self-control theabrownin material such as sugar-free and albumen in preparation process; 784,701cm
-1Two places are the C-H out-of-plane deformation vibration of phenyl ring, all have the eigen vibration of above-mentioned functional group in two samples.
3, ultraviolet full wavelength scanner
As shown in Figure 3 and Figure 4, two kinds of materials all have maximum absorption at the 342nm place, deducibility thus, and the two has identical chromophoric group, functional group or conjugated system.
4, GPC stratographic analysis
As shown in Figure 5 and Figure 6, two kinds of materials all contain weight-average molecular weight and are the material of (1.070e+4), and the weight-average molecular weight that contains in traditional theabrownin is M
w=3.374e+6 and Mw=3.302e+4 material should be the high molecular weight materials that combination sugar and albumen produce, and make by oneself in the theabrownin analogue owing to sugar-free and albumen participation, so there is not this peak.
Two, conclusion
By inferring with simultaneous test, self-control theabrownin similar substance and traditional theabrownin have similar physico-chemical property.
The invention has the advantages that,
1, theabrownin means that a class can be water-soluble and be insoluble to the brown pigment of ethanol, ethyl acetate, propyl carbinol and chloroform, also is the very complicated non-dialysis high polymeric compound of a class.Theabrownin mixture in the ripe tea of Leaf of Assam Tea has accounted for about 15% of tea dry measure greatly, and the height of theabrownin content is the important indicator of estimating the Leaf of Assam Tea quality.The blood fat reducing function that theabrownin wherein is good becomes the focus of scholar's concern in recent years.The preparation method of theabrownin similar substance of the present invention is if can realize industrialization, and following advantage is arranged: the inventive method is simple, and used starting material are cheap and easy to get, and preparation time is short, greatly reduces Pu'er raw tea is fermented and then obtains time of Folium camelliae assamicae (processed); Saved time cost greatly;
2, product structure of the present invention is clearer, can be considered as the exploitation that reference material carries out theabrownin content assaying method in the Leaf of Assam Tea, fills up the method blank of at present theabrownin being carried out more accurately assay.
3, the theabrownin similar substance that obtains of the present invention belongs to oligomeric or middle polyacetylene compound, is convenient to absorption and utilization in the body more.
Description of drawings:
Fig. 1 tradition theabrownin infrared spectrogram
Fig. 2 makes theabrownin analogue infrared spectrogram by oneself
Fig. 3 tradition theabrownin
Fig. 4 makes theabrownin analogue ultraviolet full wavelength scanner by oneself
Fig. 5 tradition theabrownin component (removing albumen and polysaccharide) GPC color atlas
Fig. 6 makes theabrownin similar substance GPC color atlas by oneself
Embodiment:
Further specify the present invention by the following examples, but not as limitation of the present invention.
Precision takes by weighing the 422mg gallic acid, places the 250ml round-bottomed flask, adds 100ml water, and 0.5ml ammoniacal liquor at room temperature stirs 48h, gets the theabrownin crude product solution.Solution is transferred in the 250ml separating funnel, uses ethyl acetate extraction 2 times, and water is used the equal-volume n-butanol extraction 2 times again, and the water drying under reduced pressure derives from theabrownin 402mg processed.
Embodiment 2
Precision takes by weighing the 200mg gallic acid, places the 250ml round-bottomed flask, adds 40ml water, and 0.3ml ammoniacal liquor at room temperature stirs 24h, gets the theabrownin crude product solution.Solution is transferred in the 250ml separating funnel, uses ethyl acetate extraction 1 time, and water is used the equal-volume n-butanol extraction 1 time again, and the water drying under reduced pressure derives from theabrownin 200mg processed.
Embodiment 3
Precision takes by weighing the 600mg gallic acid, places the 250ml round-bottomed flask, adds 600ml water, and 0.6ml ammoniacal liquor at room temperature stirs 48h, gets the theabrownin crude product solution.Solution is transferred in the 250ml separating funnel, uses ethyl acetate extraction 3 times, and water is used the equal-volume n-butanol extraction 3 times again, and the water drying under reduced pressure derives from theabrownin 600mg processed.
Precision takes by weighing the 300mg gallic acid, places the 250ml round-bottomed flask, adds 30ml water, and 0.3ml ammoniacal liquor at room temperature stirs 96h, gets the theabrownin crude product solution.Solution is transferred in the 250ml separating funnel, uses ethyl acetate extraction 3 times, and water is used the equal-volume n-butanol extraction 2 times again, and the water drying under reduced pressure derives from theabrownin 285mg processed.
Precision takes by weighing the 200mg gallic acid, places the 250ml round-bottomed flask, adds 150ml water, and 0.25ml ammoniacal liquor at room temperature stirs 12h, gets the theabrownin crude product solution.Solution is transferred in the 250ml separating funnel, uses ethyl acetate extraction 2 times, and water is used the equal-volume n-butanol extraction 3 times again, and the water drying under reduced pressure derives from theabrownin 195mg processed.
Claims (8)
1. the preparation method of a theabrownin is characterized in that, this method may further comprise the steps:
Step 1, gallic acid add ammoniacal liquor reaction 12-96 hour, get the theabrownin crude product;
Step 2, theabrownin crude product ethyl acetate extraction obtains water layer;
Step 3, water layer are used n-butanol extraction again, obtain water layer;
Step 4 obtains theabrownin after the drying.
2. according to the preparation method of claim 1, it is characterized in that,
Wherein, step 1 reaction times is 24-48 hour.
3. according to the preparation method of claim 1, it is characterized in that,
Wherein, step 1, the amount of gallic acid are the 2-6 weight part, and the amount of ammoniacal liquor is the 3-7 parts by volume, are reflected in the water to carry out, and the add-on of water is 100-1000 times of ammoniacal liquor add-on.
4. according to the preparation method of claim 1, it is characterized in that,
Step 2, ethyl acetate extraction 1-3 time of theabrownin crude product.
5. according to the preparation method of claim 1, it is characterized in that,
Step 2, ethyl acetate extraction 2 times of theabrownin crude product.
6. according to the preparation method of claim 1, it is characterized in that,
Step 3, water layer are used n-butanol extraction 1-3 time again.
7. according to the preparation method of claim 1, it is characterized in that,
Step 3, water layer are used n-butanol extraction 2 times again.
8. a theabrownin analogue is characterized in that, is prepared from either party's method of claim 1-5.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103652090A (en) * | 2013-12-18 | 2014-03-26 | 张松波 | Method for directly extracting theabrownin with pu'er tea serving as raw material |
| CN104365912A (en) * | 2014-11-27 | 2015-02-25 | 桂林瑞丰环保微生物应用研究所 | Method for preparing and extracting theabrownin |
| CN104474138A (en) * | 2014-11-25 | 2015-04-01 | 吉林大学 | Application of Puer tea or Puer tea extract in preparation of medicines or heath products inhibiting epilepsy |
| CN104614335A (en) * | 2015-02-12 | 2015-05-13 | 张进 | Method for rapidly identifying theabrownin |
| CN107372933A (en) * | 2017-07-06 | 2017-11-24 | 杭州丹娜生物科技有限公司 | A kind of preparation method of the compound theabrownin of feature |
| CN110887713A (en) * | 2019-12-04 | 2020-03-17 | 湖南农业大学 | Dark tea brown reference substance and application thereof |
| CN117357593A (en) * | 2023-10-27 | 2024-01-09 | 云南省农业科学院农产品加工研究所 | Application of theabrownin of puer tea in regulating circadian rhythm disorder |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103652090A (en) * | 2013-12-18 | 2014-03-26 | 张松波 | Method for directly extracting theabrownin with pu'er tea serving as raw material |
| CN103652090B (en) * | 2013-12-18 | 2015-04-29 | 张松波 | Method for directly extracting theabrownin with pu'er tea serving as raw material |
| CN104474138A (en) * | 2014-11-25 | 2015-04-01 | 吉林大学 | Application of Puer tea or Puer tea extract in preparation of medicines or heath products inhibiting epilepsy |
| CN104365912A (en) * | 2014-11-27 | 2015-02-25 | 桂林瑞丰环保微生物应用研究所 | Method for preparing and extracting theabrownin |
| CN104614335A (en) * | 2015-02-12 | 2015-05-13 | 张进 | Method for rapidly identifying theabrownin |
| CN107372933A (en) * | 2017-07-06 | 2017-11-24 | 杭州丹娜生物科技有限公司 | A kind of preparation method of the compound theabrownin of feature |
| CN110887713A (en) * | 2019-12-04 | 2020-03-17 | 湖南农业大学 | Dark tea brown reference substance and application thereof |
| CN117357593A (en) * | 2023-10-27 | 2024-01-09 | 云南省农业科学院农产品加工研究所 | Application of theabrownin of puer tea in regulating circadian rhythm disorder |
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Application publication date: 20130918 |