CN110590967A - Method for recovering organic matters rich in sea cucumber polysaccharide from sea cucumber blanching liquid - Google Patents
Method for recovering organic matters rich in sea cucumber polysaccharide from sea cucumber blanching liquid Download PDFInfo
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- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
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- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
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- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
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- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
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- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
- C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
- C08B37/0069—Chondroitin-4-sulfate, i.e. chondroitin sulfate A; Dermatan sulfate, i.e. chondroitin sulfate B or beta-heparin; Chondroitin-6-sulfate, i.e. chondroitin sulfate C; Derivatives thereof
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Abstract
The invention discloses a method for recovering organic matters rich in sea cucumber polysaccharide from a sea cucumber blanching liquid, which comprises the steps of properly adjusting the pH value of the sea cucumber blanching liquid, adding a certain amount of protamine for flocculation, directly centrifuging the flocculated liquid, collecting the centrifuged dregs, and freeze-drying to obtain a large amount of organic matters containing sea cucumber polysaccharide. The organic matter rich in the sea cucumber polysaccharide is recovered from the sea cucumber blanching liquid, the recovery rate of the organic matter is more than 70%, wherein the content of the sea cucumber polysaccharide exceeds 60%, and the content of the protein exceeds 30%. The method has simple steps, is easy for industrial production, has high content of the sea cucumber polysaccharide in the recovered organic matters, and is suitable for being processed into corresponding functional food.
Description
Technical Field
The invention relates to the technical field of treatment of sea cucumber processing liquid, in particular to a method for recovering organic matters rich in sea cucumber polysaccharide from sea cucumber blanching liquid.
Background
After the fresh sea cucumbers are caught, because the fresh sea cucumbers have proteolytic enzymes such as autolytic enzymes, the skin of the fresh sea cucumbers is easy to melt and autolyze. For the convenience of preservation, fresh sea cucumbers are generally preserved as raw materials after being subjected to hot water blanching treatment. According to incomplete statistics, 0.6-1.2 tons of sea cucumber blanching liquid can be generated when 1 ton of fresh sea cucumbers are processed. Along with the scale and intensification of sea cucumber processing, the produced sea cucumber blanching liquid is more and more, the sea cucumber blanching liquid which is not added with the treatment can not reach the sewage discharge standard, and extra economic burden is added for enterprises. How to utilize the sea cucumber blanching liquid resource with high value, change waste into valuable, develop the sea cucumber blanching liquid resource into functional food with extremely high utilization value, extend the sea cucumber industrial chain, and gradually become a subject which is commonly concerned by large-scale sea cucumber processing enterprises and scientific researchers engaged in sea cucumber research.
At present, the extraction of sea cucumber saponin, glycoprotein, phospholipid and sea cucumber oil from sea cucumber blanching liquid has been reported in documents. For example, chinese patent application CN 105266075a published in 2016, 1, 27 discloses a method for extracting holothurian saponins by using a holothurian blanching liquid, which comprises the steps of firstly concentrating the holothurian blanching liquid to a concentrated liquid with a solid content of 20-45%, and then directly spray-drying to obtain a first solid powder. And then removing grease by adopting a supercritical extraction technology to obtain second solid powder, adding ethanol into the second solid powder for extraction, repeatedly extracting a filter cake by using the ethanol, and then concentrating the extracting solution. Precipitating the concentrated solution with water, removing precipitate with horizontal screw, disc and tubular centrifuge to obtain supernatant, subjecting the supernatant to macroporous adsorbent resin column adsorption chromatography, eluting the macroporous adsorbent resin column with ethanol solution to obtain eluate, and vacuum freeze drying the eluate to obtain sea cucumber saponin extract. Chinese patent CN102406048B also discloses a method for preparing sea cucumber glycoprotein by using sea cucumber blanching liquid, and particularly discloses a method for preparing sea cucumber glycoprotein by using sea cucumber blanching liquid as a raw material, performing reduced pressure concentration, alcohol precipitation, precipitation drying, preparing into powder, performing alcohol washing and desalination on the powder, re-dissolving the powder in purified water, performing enzymolysis, centrifuging, performing tube filtration on supernatant, performing ultrafiltration, and drying to obtain a dried sea cucumber glycoprotein. Although the two methods are optimized compared with the prior art, the two methods have higher requirements on equipment, more complex processing mode and large amount of ethanol, and other documents have similar problems in extracting functional active ingredients from the sea cucumber blanching liquid.
At present, the method for recovering active substances from the sea cucumber blanching liquid has no selectivity, and consumes a large amount of ethanol and the like. Therefore, it is necessary to provide a recovery method with no pollution, simple process, low cost and high extraction rate.
Disclosure of Invention
The invention aims to provide a method for recovering organic matters rich in sea cucumber polysaccharide from a sea cucumber blanching liquid without adding ethanol, which is environment-friendly, free of chemical investment and pollution.
In order to achieve the purpose, the invention provides the following scheme:
the invention provides a method for recovering organic matters rich in sea cucumber polysaccharide from sea cucumber blanching liquid, which comprises the following steps:
(1) adjusting the pH value of the sea cucumber blanching liquid;
(2) adding protamine into the sea cucumber blanching liquid after pH adjustment, and stirring;
(3) centrifuging, and collecting the residues after centrifuging;
(4) directly freeze-drying the dregs to obtain the organic matter rich in sea cucumber polysaccharide.
Further, in the step (1), the pH value of the sea cucumber blanching liquid is adjusted to 4-6.
Further, the final concentration of the protamine added in the step (2) is 0.5 g/L.
Further, the method comprises the following steps: if the pH value of the sea cucumber blanching liquid is not adjusted in the step (1), the final adding concentration of the protamine in the step (2) is 1 g/L.
Further, in the step (2), the stirring is continuously performed for 8-12min under the condition of 200 rpm.
Further, the centrifugation in the step (3) is performed for 1min at the rotating speed of 4800 rpm. The centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid.
Further, in the step (4), the residue is freeze-dried until the water content is below 10%.
The extraction process of protamine is as follows:
1. extraction of DNP (deoxyribonuclein) from testis tissue
Taking 100g of mature salmon and carp testis tissues (milt) in a tissue triturator, adding 200mL of NaCl solution with the concentration of 0.14mol/L, fully homogenizing for 1min, continuously stirring for 20min under the action of an electric stirrer, standing for 10min after stirring, centrifuging for 10min under the condition of 4000rpm, pouring out supernatant, collecting precipitate, repeating the operation once, collecting precipitate, and combining the two precipitates;
2. extraction of sulfuric acid
Adding H with the concentration of 7.5% into the precipitate obtained in the step 12S04Fully crushing 300mL of solution in a tissue crusher, stirring and extracting for 3h under the action of an electric crusher, centrifuging for 10min under the condition of 4000rpm, discarding the precipitate, collecting the supernatant, and filtering the supernatant to remove insoluble substances;
3. ethanol precipitation
Adding 900mL of cold ethanol with volume fraction of 95% into the supernatant obtained in the step 2, precipitating and standing for 30min under the ice bath condition, centrifuging for 10min under the condition of 4000rpm, and pouring out the supernatant to leave a precipitate;
4. protein washing
And (4) adding an acetone solution into the precipitate obtained in the step (3), repeatedly washing for two to three times, drying to obtain a protamine crude product, and weighing.
The protamine for recycling the organic matters of the sea cucumber blanching liquid has the basic amino acid content of not less than 45 percent.
The invention provides a method for recovering organic matters rich in sea cucumber polysaccharide from a sea cucumber blanching liquid, which comprises the steps of properly adjusting the pH value of the sea cucumber blanching liquid, adding a certain amount of protamine for flocculation, directly centrifuging the flocculated liquid, collecting the centrifuged dregs, and freeze-drying to obtain a large amount of organic matters containing sea cucumber polysaccharide. The method has simple steps, is easy for industrial production, has high content of the sea cucumber polysaccharide in the recovered organic matters, and is suitable for being processed into corresponding functional food.
Protamine is a protein rich in basic amino acids and with positive charges, and sea cucumber polysaccharide takes sea cucumber fucosan sulfate, sea cucumber chondroitin sulfate and the like as main components, is rich in sulfate groups and has strong negative charges, so that the protamine basic amino acids with positive charges can be effectively combined and flocculate sulfated polysaccharide, thereby realizing recovery. The organic matter rich in the sea cucumber polysaccharide is recovered from the sea cucumber blanching liquid, the recovery rate of the organic matter is more than 70%, wherein the content of the sea cucumber polysaccharide exceeds 60%, and the content of the protein exceeds 30%.
Moreover, the recovery method of the invention aims at the negative charge sea cucumber polysaccharide, and has strong selectivity; secondly, after flocculation, the collection cost is greatly reduced, and ethanol is not needed; and thirdly, the protamine is a natural protein product, is environment-friendly, has no chemical investment and has no pollution.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The extraction process of protamine in the embodiment of the invention is as follows:
1. extraction of DNP (deoxyribonuclein) from testis tissue
Taking 100g of mature salmon and carp testis tissues (milt) in a tissue triturator, adding 200mL of NaCl solution with the concentration of 0.14mol/L, fully homogenizing for 1min, continuously stirring for 20min under the action of an electric stirrer, standing for 10min after stirring, centrifuging for 10min under the condition of 4000rpm, pouring out supernatant, collecting precipitate, repeating the operation once, collecting precipitate, and combining the two precipitates;
2. extraction of sulfuric acid
Adding H with the concentration of 7.5% into the precipitate obtained in the step 12S04Fully crushing 300mL of solution in a tissue crusher, stirring and extracting for 3h under the action of an electric crusher, centrifuging for 10min under the condition of 4000rpm, discarding the precipitate, collecting the supernatant, and filtering the supernatant to remove insoluble substances;
3. ethanol precipitation
Adding 900mL of cold ethanol with volume fraction of 95% into the supernatant obtained in the step 2, precipitating and standing for 30min under the ice bath condition, centrifuging for 10min under the condition of 4000rpm, and pouring out the supernatant to leave a precipitate;
4. protein washing
And (4) adding an acetone solution into the precipitate obtained in the step (3), repeatedly washing for two to three times, drying to obtain a protamine crude product, and weighing.
Protamine with the content of basic amino acid not less than 45 percent is used for recovering organic matters rich in polysaccharide from the sea cucumber blanching liquid.
Example 1
Accurately measuring 1L of the sea cucumber blanching liquid, adjusting the pH value of the sea cucumber blanching liquid to 4, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 0.5g/L, and continuously stirring for 10min at 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In the example, the recovery rate of the organic matter is 80.39%, the polysaccharide accounts for 60.05% and the protein accounts for 32.62%. The calculation method is as follows:
the recovery rate of organic matters (percent) is that the mass of the organic matters in the flocculation dregs/the mass of the organic matters in the sea cucumber blanching liquid multiplied by 100 percent is 7.91g/9.84g multiplied by 100 percent is 80.39 percent.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.75 g/7.91g × 100% — 60.05%.
The protein content (%) is 2.58 g/7.91g × 100% is 32.62% of the mass of protein in the flocculated sludge/the mass of organic matter in the flocculated sludge.
Example 2
Accurately measuring 1L of the sea cucumber blanching liquid, adjusting the pH value of the sea cucumber blanching liquid to 5, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 0.5g/L, and continuously stirring for 8min at 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In the embodiment, the recovery rate of the organic matter is 70.33%, the polysaccharide accounts for 62.43%, and the protein accounts for 30.20%. The calculation method is as follows:
the recovery rate of organic matters (mass percent) of the organic matters in the flocculation dregs/mass of the organic matters in the sea cucumber blanching liquid multiplied by 100 percent is 6.92g/9.84g multiplied by 100 percent is 70.33 percent.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.32g/6.92g × 100% — 62.43%.
The protein content (%) is 2.09 g/6.92g × 100% is 30.20% of the mass of protein in the flocculated sludge/the mass of organic matter in the flocculated sludge.
Example 3
Accurately measuring 1L of the sea cucumber blanching liquid, adjusting the pH value of the sea cucumber blanching liquid to 6, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 0.5g/L, and continuously stirring for 12min under the condition of 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In the example, the recovery rate of the organic matter is 76.42%, the polysaccharide content in the organic matter is 63.70%, and the protein content in the organic matter is 31.78%. The calculation method is as follows:
the recovery rate (%) of the organic matters is 7.52g/9.84g × 100% of the organic matters in the sea cucumber blanching liquid and 76.42% of the organic matters in the flocculation dregs.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.79 g/7.52g × 100% — 63.70%.
The protein content (%) is 2.39 g/7.52g × 100% 31.78% based on the mass of protein/mass of organic matter in the flocculated sludge.
Example 4
Accurately measuring 1L of the sea cucumber blanching liquid, adjusting the pH value of the sea cucumber blanching liquid to 4.5, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 0.5g/L, quickly stirring at 300rpm, and then continuously stirring for 12min at 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In this example, the recovery rate of organic matter was 74.09%, the ratio of polysaccharide in organic matter was 68.45%, and the ratio of protein in organic matter was 29.77%. The calculation method is as follows:
the recovery rate of organic matters (mass percent) of the organic matters in the flocculation dregs/mass of the organic matters in the sea cucumber blanching liquid multiplied by 100 percent is 7.29g/9.84g multiplied by 100 percent is 74.09 percent.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.99 g/7.29g × 100% — 68.45%.
The protein content (%) is 2.17g/7.29g × 100% is 29.77% of the mass of protein in the flocculated sludge/the mass of organic matter in the flocculated sludge.
Example 5
Accurately measuring 1L of sea cucumber blanching liquid, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 1.0g/L, quickly stirring at 300rpm, and then continuously stirring for 12min at 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In the embodiment, the recovery rate of the organic matter is 71.24%, the polysaccharide accounts for 62.34%, and the protein accounts for 33.81%. The calculation method is as follows:
the recovery rate of organic matters (mass percent) of the organic matters in the flocculation dregs/mass of the organic matters in the sea cucumber blanching liquid multiplied by 100 percent is 7.01g/9.84g multiplied by 100 percent is 71.24 percent.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.37 g/7.01g × 100% — 62.34%.
The protein content (%) is 2.37 g/7.01g × 100% 33.81% based on the mass of protein/mass of organic matter in the flocculated sludge.
Example 6
Accurately measuring 1L of sea cucumber blanching liquid, adding a certain amount of protamine into the sea cucumber blanching liquid to enable the final concentration to reach 1.0g/L, quickly stirring at 300rpm, and then continuously stirring for 10min at 200 rpm. Centrifuging the stirred sea cucumber blanching liquid at 4800rpm for 1min, recovering dregs at the lower layer, wherein the centrifuged dregs are cakes with compact structures, and the supernatant is clear and transparent liquid. Directly freeze-drying the dregs until the water content is below 10 percent to obtain the organic matter rich in sea cucumber polysaccharide. Determination of organic matter mass: continuously drying the sea cucumber blanching liquid and the flocculation dregs at 60 ℃ for 2 days to ensure that the water is completely dried and weighed (the mass is M1), then putting the dried sample in a muffle furnace at 550 ℃ for calcining for 2 hours and weighing to obtain the mass of ash (M2), and obtaining the mass of organic matters which is M1-M2. In this example, the recovery rate of the organic matter was 76.32%, the polysaccharide content in the organic matter was 62.32%, and the protein content was 32.89%. The calculation method is as follows:
the recovery rate (%) of the organic matters is 7.51g/9.84g × 100% of the organic matters in the sea cucumber blanching liquid and 76.32% of the organic matters in the flocculated dregs.
The ratio of polysaccharide (%) — sea cucumber polysaccharide mass in flocculation dreg/organic matter mass in flocculation dreg 4.68 g/7.51g × 100% — 62.32%.
The protein content (%) is 2.47g/7.51g × 100% is 32.89% of the mass of protein in the flocculated sludge/the mass of organic matter in the flocculated sludge.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.
Claims (7)
1. A method for recovering organic matters rich in sea cucumber polysaccharide from sea cucumber blanching liquid is characterized by comprising the following steps: the method comprises the following steps:
(1) adjusting the pH value of the sea cucumber blanching liquid;
(2) adding protamine into the sea cucumber blanching liquid after pH adjustment, and stirring;
(3) centrifuging, and collecting the residues after centrifuging;
(4) directly freeze-drying the dregs to obtain the organic matter rich in sea cucumber polysaccharide.
2. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: in the step (1), the pH value of the sea cucumber blanching liquid is adjusted to 4-6.
3. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: the final concentration of the protamine added in the step (2) is 0.5 g/L.
4. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: the method further comprises the steps of: if the pH value of the sea cucumber blanching liquid is not adjusted in the step (1), the final adding concentration of the protamine in the step (2) is 1 g/L.
5. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: in the step (2), the stirring is continuously carried out for 8-12min under the condition of 200 rpm.
6. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: and (4) centrifuging at the rotating speed of 4800rpm for 1min in the step (3).
7. The method for recovering the organic matter rich in the sea cucumber polysaccharide from the sea cucumber blanching liquid according to claim 1, which is characterized in that: in the step (4), the residue is lyophilized until the water content is below 10%.
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Application publication date: 20191220 |