CN103725033B - Method for extracting Enteromorpha pigment - Google Patents
Method for extracting Enteromorpha pigment Download PDFInfo
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- CN103725033B CN103725033B CN201310724512.8A CN201310724512A CN103725033B CN 103725033 B CN103725033 B CN 103725033B CN 201310724512 A CN201310724512 A CN 201310724512A CN 103725033 B CN103725033 B CN 103725033B
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- ethyl acetate
- sherwood oil
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Abstract
The invention discloses a method for extracting an Enteromorpha pigment, which comprises the following steps: washing Enteromorpha, drying, pulverizing, and extracting by immersing in alcohol; filtering the extracting solution to obtain a filtrate, recovering the alcohol, and extracting with ethyl acetate to obtain a pigment solution; and passing the pigment solution through a silica gel resin chromatographic column, eluting by using petroleum ether or a petroleum ether-ethyl acetate mixed solution or ethyl acetate as an eluting agent, collecting the pigment component, and concentrating to remove the solvent. According to the method, the reagents used in the pigment crude extracting solution working process are water and alcohol, and the alcohol is recycled, so no environmental pollution is generated; the organic solvents used in the pigment purification process can also be recycled; and the extracted pigment has the advantages of stable properties and low production cost, is simple to operate, and has wide prospects when being developed as a food additive or staining agent.
Description
Technical field
The invention belongs to field of processing of aquatic products, especially algae processing, be specifically related to Enteromorpha pigment preparation.
Background technology
Enteromorpha belongs to Chlorophyta, Chlorophyceae, Ulvales, Ulvaceae, is a kind of large ocean green alga.Enteromorpha has extremely strong degeneration-resistant energy for growth, and break out the green tide of 7 years continuously in Yellow Sea of China marine site, its sociales are Enteromorpha.Enteromorpha itself is nontoxic, but the green tide formed may cause breaking out of insect, and algae deposition seabed can cause water hypoxia to cause hydrobiont dead, and changes sedimental physico-chemical property, produces have a strong impact on ocean Benthic ecology system.But biomass resource contained by Enteromorpha can be exploited.Enteromorpha can be processed into tongue bar and eat, or extracts the nutrition such as polysaccharide.When preparing polysaccharide from Enteromorpha, needs are prior decolours Enteromorpha to improve sugared purity, the invention provides a kind of method extracting Enteromorpha pigment, is intended to utilize the above-mentioned Enteromorpha pigment removed.
Summary of the invention
The object of this invention is to provide a kind of method extracting Enteromorpha pigment.
Technical scheme is, a kind of method extracting Enteromorpha pigment, is by after dry for Enteromorpha washing and pulverizing, uses alcohol-pickled extraction; Leaching filtrate crossed by vat liquor, is extracted with ethyl acetate and obtains pigment solution after recovered alcohol; By silica column chromatographic column loading, the mixed solution of sherwood oil or sherwood oil and ethyl acetate, as washing composition wash-out, collects colour component, concentrated removal solvent.
Concrete step comprises:
(1) gather in Enteromorpha, reject impurity, fresh water washing is no more than 0.02ms to rinsing liquid specific conductivity;
(2) moisture also 40 ~ 75 DEG C of oven dry are removed, preferably at 60 DEG C of heating, dryings;
(3) Enteromorpha is pulverized the fragment for below 3mm; Be preferably, adopt stamp mill dry method to smash to pieces to fragment at below 3mm;
(4) 75% ~ 95%(volumetric concentrations) alcohol-pickledly to spend the night, vat liquor 200-800 order silk cover filtering, get filtrate, filtrate is rotary evaporation recovered alcohol at 40-60 DEG C; Filter residue can be further used for extracting polysaccharide or being processed as Enteromorpha mud;
(5) remaining liquid is extracted with ethyl acetate pigment, gets upper solution, obtains pigment solution;
(6) to suspend silica column with sherwood oil, packing; The pigment solution silica gel of extraction into ethyl acetate blots, loading;
(7) by the mixed solution of sherwood oil or sherwood oil and ethyl acetate or ethyl acetate as washing composition wash-out, collect colour component, concentratedly remove solvent;
Concrete, with sherwood oil as washing composition wash-out, collect colour component, obtain zeaxanthin class pigment;
Or, with sherwood oil be eluted to colourless after, then (volume ratio is 4:1,7:3 to use sherwood oil and ethyl acetate mixtures, 3:2,1:1) as eluent successively wash-out, collect colour component, obtain phoeophytin acid esters a successively, chlorophyllide a, chlorophyll a and xenthophylls, finally use ethyl acetate as eluent, collects and obtain chlorophyll oxide compound.
And then with acetone, methylene dichloride or methyl alcohol as elutriant, the chlorophyll oxide compound that wash-out obtains brown color can be continued.
Present method agents useful for same in the pigment crude extract course of processing is water and alcohol, and alcohol recycle, non-environmental-pollution; Organic solvent used in pigment pufification process is also recyclable.The prepared pigment proterties extracted is stablized, and low production cost is easy and simple to handle, has broad prospects as foodstuff additive or staining agent exploitation.Byproduct after pigment extracts is the Enteromorpha of decolouring, can be processed into sea grass polysaccharide, Enteromorpha mud etc. further.The present invention can make full use of the economic worth of Enteromorpha, reduces waste.
Accompanying drawing explanation
Fig. 1 is in embodiment 1, the absorption spectrum of gained zeaxanthin class pigment.
Fig. 2 is in embodiment 2, the absorption spectrum of gained phoeophytin acid esters a pigment.
Fig. 3 is in embodiment 6, the absorption spectrum of gained chlorophyll oxide pigment.
Embodiment
The technique means realized to make the present invention, creation characteristic, reaching object and effect is easy to understand, setting forth the present invention further below.
Embodiment 1
(1) gather in Enteromorpha, reject impurity, be no more than 0.02ms with fresh water washing to the specific conductivity of rinsing liquid.(2) moisture is removed, 60 DEG C of heating, dryings.(3) adopt stamp mill dry method, the Enteromorpha of oven dry is smashed to pieces to fragment and be of a size of 3mm.(4) spend the night with volumetric concentration 75% is alcohol-pickled, vat liquor 600 order silk cover filterings, get filtrate, by filtrate rotary evaporation recovered alcohol at 50 DEG C.(5) remaining liquid is extracted with ethyl acetate pigment, gets upper solution, obtains pigment solution.
(6) to suspend silica column with sherwood oil, packing.The pigment solution silica gel of extraction into ethyl acetate blots, and is placed in silica gel chromatographic column loading.
(7) collect colour component with sherwood oil as washing composition wash-out, be spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with sherwood oil, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
| Pigment | Color | Actual measurement maximum absorption band (nm) | Rf value |
| Zeaxanthin | Yellow | 450 | 0.53 |
Absorption spectrum as shown in Figure 1.Qualification result is zeaxanthin class pigment.With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become yellow.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
Embodiment 2
(1) gather in Enteromorpha, reject impurity, be no more than 0.02ms with fresh water washing to the specific conductivity of rinsing liquid.(2) moisture is removed, 60 DEG C of heating, dryings.(3) adopt stamp mill dry method, the Enteromorpha of oven dry is smashed to pieces to fragment and be of a size of 3mm.(4) spend the night with volumetric concentration 75% is alcohol-pickled, vat liquor 600 order silk cover filterings, get filtrate, by filtrate rotary evaporation recovered alcohol at 45 DEG C.(5) remaining liquid is extracted with ethyl acetate pigment, gets upper solution, obtains pigment solution.
(6) to suspend silica column with sherwood oil, packing.The pigment solution silica gel of extraction into ethyl acetate blots, and is placed in silica gel chromatographic column loading.
(7) be first eluted to as washing composition colourless with sherwood oil, then collect colour component with the petrol ether/ethyl acetate mixed solvent wash-out that volume ratio is 4:1, be spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with the mixed solvent of petrol ether/ethyl acetate=3/1, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
Absorption spectrum as shown in Figure 2.Qualification result is that (phoeophytin acid esters a) for chlorophyll class pigment.With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become grey green-yellow.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
Embodiment 3
In embodiment 2, by sherwood oil and volume ratio be the petrol ether/ethyl acetate mixed solvent of 4:1 as after washing composition successively wash-out, then with volume ratio be 7:3 petrol ether/ethyl acetate mixed solvent wash-out collect colour component, be spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with the mixed solvent of petrol ether/ethyl acetate=3/1, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
| Pigment | Color | Actual measurement maximum absorption band (nm) | Rf value |
| Chlorophyll class (chlorophyll | Bluish-green | 475 | 0.45 |
| Acid esters a) |
Qualification result is that (chlorophyllide a) for chlorophyll class pigment.With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become blue-greenish colour.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
Embodiment 4
After embodiment 3 loading wash-out, then continuation volume ratio is the petrol ether/ethyl acetate mixed solvent wash-out of 3:2, collects colour component, is spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with the mixed solvent of petrol ether/ethyl acetate=3/1, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
| Pigment | Color | Actual measurement maximum absorption band (nm) | Rf value |
| Chlorophyll class (chlorophyll a) | Bluish-green | 435 | 0.40 |
Qualification result is chlorophyll class pigment (chlorophyll a).With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become blue-greenish colour.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
Embodiment 5
In embodiment 4, volume ratio is after the petrol ether/ethyl acetate mixed solvent wash-out of 3:2, then continuation volume ratio is the petrol ether/ethyl acetate mixed solvent wash-out collection colour component of 1:1, is spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with the mixed solvent of petrol ether/ethyl acetate=3/1, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
| Pigment | Color | Actual measurement maximum absorption band (nm) | Rf value |
| Xenthophylls | Deep yellow | 450 | 0.34 |
Qualification result is xenthophylls.With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become deep yellow.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
Embodiment 6
In embodiment 5, after making wash-out with the petrol ether/ethyl acetate mixed solvent that volume ratio is 1:1, then collect colour component with eluent ethyl acetate, be spin-dried for concentrated.Thin-layer chromatography is done in sampling, dissolves with extraction agent (acetone/sherwood oil=3/2 volume ratio), and developping agent (sherwood oil/acetone=3/1 volume ratio) launches, and measures Rf value.Sampling scanning absorption spectrum, do solvent and blank with the mixed solvent of petrol ether/ethyl acetate=3/1, sweep limit is 300-700nm, interval 1nm.Result preliminary evaluation is as follows:
| Pigment | Color | Actual measurement maximum absorption band (nm) | Rf value |
| Chlorophyll oxide compound | Pale brown | 410 | 0 |
Absorption spectrum as shown in Figure 3.Qualification result is chlorophyll oxide compound.With this pigment dye white cloth, bolting silk and waterproof face-cloth, after solvent evaporates, cloth, bolting silk and waterproof face-cloth become brown color.Pigment water cannot wash away, and wash more than 50 times, or be soaked in water more than 5 days, color is constant, tentatively shows that this pigment has Color.
After eluent ethyl acetate, continuation acetone, methylene dichloride or methyl alcohol, as elutriant, can continue the chlorophyll oxide compound that wash-out obtains brown color.
More than show and describe ultimate principle of the present invention, principal character and advantage of the present invention.The technician of the industry should understand; the present invention is not restricted to the described embodiments; what describe in above-described embodiment and specification sheets just illustrates principle of the present invention; the present invention also has various changes and modifications without departing from the spirit and scope of the present invention, and these changes and improvements all fall in the claimed scope of the invention.Application claims protection domain is defined by appending claims and equivalent thereof.
Claims (1)
1. extract a method for Enteromorpha pigment, it is characterized in that, comprise the following steps:
(1) washing of Enteromorpha fresh water is no more than 0.02ms/cm to rinsing liquid specific conductivity;
(2) remove moisture, dry at 40 ~ 75 DEG C;
(3) dried Enteromorpha is pulverized the fragment for below 3mm;
(4) 75% ~ 95% alcohol-pickledly to spend the night, vat liquor 200-800 order silk cover filtering, gets filtrate, rotary evaporation recovered alcohol at 40 ~ 60 DEG C;
(5) remaining liquid is extracted with ethyl acetate, and gets upper solution, obtains pigment solution;
(6) to suspend silica column with sherwood oil, packing; In step (5), the pigment solution silica gel of extraction into ethyl acetate blots, loading;
(7) use sherwood oil as washing composition wash-out, collect colour component, concentrated removal solvent, obtains zeaxanthin class pigment;
Sherwood oil be eluted to colourless after, then with volume ratio be the sherwood oil of 4:1 and ethyl acetate mixtures as eluent, collect colour component, concentratedly remove solvent, obtain phoeophytin acid esters a;
Again with volume ratio be 7:3 sherwood oil and ethyl acetate mixtures as eluent, collect colour component, concentratedly remove solvent, obtain chlorophyllide a;
Again with volume ratio be 3:2 sherwood oil and ethyl acetate mixtures as eluent, collect colour component, concentratedly remove solvent, obtain chlorophyll a;
Again with volume ratio be 1:1 sherwood oil and ethyl acetate mixtures as eluent, collect colour component, concentratedly remove solvent, obtain xenthophylls;
Use ethyl acetate as eluent again, collect colour component, concentrated removal solvent, obtains chlorophyll oxide compound;
Continuation acetone, methylene dichloride or methyl alcohol, as elutriant, collect colour component, and concentrated removal solvent, obtains chlorophyll oxide compound.
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| CN201310724512.8A CN103725033B (en) | 2013-12-24 | 2013-12-24 | Method for extracting Enteromorpha pigment |
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| CN201310724512.8A CN103725033B (en) | 2013-12-24 | 2013-12-24 | Method for extracting Enteromorpha pigment |
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| CN105542509A (en) * | 2016-01-25 | 2016-05-04 | 苏州印丝特纺织数码科技有限公司 | Natural dye for spinning and preparation method thereof |
| CN111948319B (en) * | 2020-08-18 | 2022-04-01 | 宁波大学 | Efficient separation and analysis method for active ingredients in enteromorpha |
| CN111848360B (en) * | 2020-08-18 | 2023-01-06 | 宁波大学 | Preparation method of active ingredients in enteromorpha |
| CN115679689A (en) * | 2022-12-12 | 2023-02-03 | 杨开健 | Flame-retardant fabric finishing agent and fabric finishing method |
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| CN100383054C (en) * | 2006-05-26 | 2008-04-23 | 中国海洋大学 | Method for preparing enteromorpha extract for inhibiting red tide algae and its use |
| CN101843261B (en) * | 2010-05-05 | 2013-05-08 | 秦旭坤 | Method and application for extracting alexandrium tamarense inhibitor from enteromorpha |
| CN103012414A (en) * | 2012-12-04 | 2013-04-03 | 青岛艾华隆生物科技有限公司 | Process for enzymatic preparation of enteromorpha chlorophyll |
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Effective date of registration: 20220610 Address after: 201318 4 building, 1 Lane 99, Shen Mei Road, Pudong New Area, Shanghai, 4 Patentee after: SHANGHAI SONGHAO BIOTECHNOLOGY Co.,Ltd. Address before: 201306, No. 999, Huan Cheng Road, Pudong New Area, Shanghai Patentee before: SHANGHAI OCEAN University |