CN110229225B - Preparation method of sulfur-rich and/or cesium-rich protein in antarctic krill - Google Patents
Preparation method of sulfur-rich and/or cesium-rich protein in antarctic krill Download PDFInfo
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Abstract
The invention provides a preparation method of sulfur-rich and/or cesium-rich protein in antarctic krill, which comprises the steps of extracting total protein from the antarctic krill through aqueous solution extraction, and experiments show that the total protein sulfur and cesium extracted by the aqueous solution extraction method are higher in content in an aqueous solution extraction method, a dilute acid solution extraction method, a dilute alkali solution extraction method, a dilute salt solution extraction method and an alcohol solution extraction method. Meanwhile, the antarctic krill sulfur-rich protein or the antarctic krill cesium-rich protein can be further obtained by a sephadex chromatographic separation method.
Description
Technical Field
The disclosure belongs to the technical field of food and chemical engineering, relates to the technical field of extraction and separation of sulfur-rich or cesium-rich protein, and particularly relates to a preparation method of sulfur-rich and/or cesium-rich protein in euphausia superba.
Background
The statements herein merely provide background information related to the present disclosure and may not necessarily constitute prior art.
Antarctic krill (Euphausia superba), also known as krill, is a large number of crustaceans living in the Antarctic sea area. Antarctic krill is rich in nutritive value, and contains abundant fatty acids (essential fatty acids), proteins, vitamins, trace elements and bioactive substances. The protein content of the fresh antarctic krill is about 11.9% -15.4%, and the euphausia superba is a high-quality protein resource bank and has high nutritional value and research value. Proteins are one of the bases of living organisms and are also important components necessary for the growth and metabolism of organisms. With the progress of research on genomics, structural biology, molecular biology and the like, scientists have increasingly recognized the biological activity, structure and function of proteins, and many researchers have turned the focus of biological research from genome research to protein structure and function research. To study the structure and function of protein, the high-purity protein with biological activity to be studied is first separated out, and the efficient extraction, separation and purification method and technology are one of the basis and key for studying the activity performance of protein.
Sulfur elements are ubiquitous in various cells, and sulfur elements are not only elements required by the human body but also one of the constituent elements constituting amino acids. The sulfur-rich protein has important functions in the aspects of nutritive value, physiological function, human metabolism and the like. Sulfur-rich proteins (metallothioneins) which bind to metals are available by binding to toxic heavy metalsSo as to effectively reduce the poison of heavy metal to the organism, thereby realizing the detoxification function. The sulfur-rich protein food is rich in sulfur-containing amino acids, so that the food is different in color, fragrance and taste and has important effects in aspects of nutritive value, browning inhibition, aging prevention, oxidation resistance and the like. The nutritional function and physiological regulation function of sulfur-rich protein in food are summarized mainly in the following aspects. First, methionine is an essential amino acid in the human body, i.e., it is not synthesized by the human body and must be taken in through the diet. Secondly, there are many methods for preventing food browning, and the widely used sulfite is a reagent commonly used in chemical treatment, and although sulfite can effectively inhibit food browning, sulfite can cause certain adverse effects, so that a safe and reliable sulfite substitute is required to be found. The research finds that the sulfur-containing amino acid has strong inhibiting effect on enzymatic browning and non-enzymatic browning, and the adding effect is similar to the effect of sulfite. Sulfur-containing amino acids, particularly cysteine, inhibit maillard reactions of proteins or other amino acids with sugars, and thus can prevent a reduction in the nutritional value of food proteins due to browning. Thirdly, cystine and cysteine have strong antioxidation effect, and the sulfhydryl group of cysteine is a very strong nucleophilic group which can remove active oxygen in the form of reducing agent, thereby achieving the effect of preventing food oxidative putrefaction and body oxidative aging, and preventing V in foodC、VEAnd polyunsaturated fatty acids and phenolic bioactive substances are oxidized, so that the nutritional value of the food is indirectly improved.
The metal cesium is golden yellow, soft and light and low-melting-point active metal, is very easy to oxidize in air, and can react with water violently to generate hydrogen and explode. Cesium iodide reacts with bismuth triiodide to form insoluble bright red double salts, which are used for qualitative and quantitative determination of cesium; the flame of cesium turns purple-red and can be used to detect cesium. The cesium has no elementary substance form in nature, and is rarely distributed in the land and the sea in the form of salt. The radionuclide Cs-137 is one of radioactive contamination leaked out of the first nuclear power plant of fukushima, japan. Cesium is also an important material for manufacturing vacuum devices, phototubes, etc., and can be used as a catalyst chemically. Cesium can generate an outstanding photoelectric effect, is very easy to ionize to release electrons, and is a main material of a photoelectric tube; in recent years, the method has new application in ion rockets, magnetohydrodynamic generators, thermoelectric transducers and the like.
Disclosure of Invention
The purpose of the present disclosure is to provide a method for preparing sulfur-rich and/or cesium-rich proteins in antarctic krill, which can extract sulfur-rich and/or cesium-rich proteins from antarctic krill.
In order to achieve the purpose, the technical scheme of the disclosure is as follows:
on one hand, the method for extracting the total protein of the antarctic krill by the aqueous solution comprises the steps of placing shrimp paste of the antarctic krill in water, uniformly dispersing the shrimp paste to obtain a mixed solution, carrying out ultrasonic extraction on the mixed solution, and removing precipitates in the mixed solution after the ultrasonic extraction to obtain the total protein of the antarctic krill.
According to the application, the content of amino acids in the hydrolysate of the euphausia superba protein is rich through research, wherein the content of sulfur-containing amino acids is high, the liver cirrhosis, senile dementia and the like can be weakened due to the fact that the sulfur-containing amino acids participate in processes of transcription, translation and the like, and the sulfur-rich protein is also helpful for maintaining the health and luster of skin and nails, maintaining oxygen balance and helping normal operation of brain functions. In order to extract sulfur-rich protein from antarctic krill, the total protein in the antarctic krill is extracted from the antarctic krill by respectively adopting an aqueous solution extraction method, a dilute acid solution extraction method, a dilute alkali solution extraction method, a dilute salt solution extraction method and an alcohol solution extraction method in the research process, and qualitative and quantitative detection of sulfur elements is carried out by inductively coupled plasma emission spectrometry (ICP-5000), so that the highest content of the sulfur elements in the total protein of the antarctic krill extracted from the aqueous solution is found.
In another aspect, total protein of antarctic krill is obtained by the above method.
Adding water into dry powder of total protein of antarctic krill, uniformly mixing the dry powder on a vortex mixer, removing filter residues through centrifugal separation, collecting supernatant, separating and purifying the supernatant by using water as an eluent and adopting a Sephadex G-75 gel chromatography method to obtain a plurality of protein samples, and performing qualitative and quantitative detection on sulfur or cesium of the protein samples by using inductive coupling plasma emission spectrometry (ICP-5000) to obtain the protein sample with the highest sulfur or cesium content, namely the antarctic krill sulfur-rich protein or the antarctic krill cesium-rich protein.
The disclosure finds that, through further research, Antarctic krill sulfur-rich protein or Antarctic krill cesium-rich protein can be further extracted from total protein of Antarctic krill.
In a fourth aspect, the sulfur-rich protein of the antarctic krill or the cesium-rich protein of the antarctic krill is obtained by the preparation method.
The beneficial effect of this disclosure does:
1. the disclosure provides a method for extracting total protein of antarctic krill by using an aqueous solution, wherein the content of sulfur element in the total protein of the antarctic krill extracted by the method is highest.
2. The method comprises the steps of separating total protein of the antarctic krill by using a glucan gel chromatography, and detecting sulfur or cesium by using an inductively coupled plasma emission spectrometry (ICP-5000) to obtain the sulfur-rich protein of the antarctic krill with high sulfur content or the cesium-rich protein of the antarctic krill with high cesium content.
Detailed Description
It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.
It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of example embodiments according to the present disclosure. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise.
The present disclosure provides a preparation method of sulfur-rich and/or cesium-rich protein in antarctic krill, which can extract sulfur-rich and/or cesium-rich protein from antarctic krill.
The typical embodiment of the disclosure provides a method for extracting total protein of antarctic krill by using an aqueous solution, which comprises the steps of placing shrimp paste of the antarctic krill into water, uniformly dispersing the shrimp paste to obtain a mixed solution, carrying out ultrasonic extraction on the mixed solution, and removing precipitates in the mixed solution after the ultrasonic extraction to obtain the total protein of the antarctic krill.
According to the application, the content of amino acids in the hydrolysate of the euphausia superba protein is rich through research, wherein the content of sulfur-containing amino acids is high, the liver cirrhosis, senile dementia and the like can be weakened due to the fact that the sulfur-containing amino acids participate in processes of transcription, translation and the like, and the sulfur-rich protein is also helpful for maintaining the health and luster of skin and nails, maintaining oxygen balance and helping normal operation of brain functions. In order to extract sulfur-rich protein from antarctic krill, the total protein in the antarctic krill is extracted from the antarctic krill by respectively adopting an aqueous solution extraction method, a dilute acid solution extraction method, a dilute alkali solution extraction method, a dilute salt solution extraction method and an alcohol solution extraction method in the research process, and qualitative and quantitative detection of sulfur elements is carried out by inductively coupled plasma emission spectrometry (ICP-5000), so that the highest content of the sulfur elements in the total protein of the antarctic krill extracted from the aqueous solution is found.
In one or more embodiments of this embodiment, the steps are as follows:
(1) unfreezing and dehydrating frozen fresh antarctic krill, and crushing to obtain fresh antarctic krill shrimp paste;
(2) adding triple-distilled water into fresh euphausia superba mud, and magnetically stirring;
(3) carrying out ultrasonic extraction;
(4) centrifuging and collecting supernatant;
(5) and (4) performing rotary evaporation until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
In this series of examples, a high-speed pulverizer was used for the crushing in step (1). The high-speed pulverizer helps to increase the contact area with the solvent, and can completely extract the total protein in the antarctic krill.
In the series of embodiments, in the step (2), the volume ratio (material-liquid ratio) of the mass of the shrimp paste to the volume of the triple distilled water is 1: 5-20. When the volume ratio of the shrimp paste to the triple distilled water is 1:15, the extraction rate of the total protein is higher, and the extraction rate is not obviously increased when the feed-liquid ratio is increased. When the ratio of the feed liquid to the feed liquid is too large, the reaction of the materials is incomplete, and the extraction rate is low due to incomplete protein extraction; when the ratio of the material to the liquid is too small, the material is completely extracted, but the subsequent extraction work is not facilitated.
In this series of examples, in the step (2), the protein extraction rate was high when the stirring time was 80 min. When the extraction time is short, the protein is not completely dissolved, and the extraction is not complete; when the extraction time is too long, the protein extraction rate is not obviously increased, and the workload of subsequent rotary evaporation treatment is increased.
In this series of examples, in step (3), the conditions of ultrasonic extraction were: the temperature is 20-40 ℃, the power is 200-300W, the frequency is 30-50 kHz, and the ultrasonic extraction time is 10-120 min. By ultrasonic extraction, the aqueous solution is fully immersed into the crushed tissue of the antarctic krill, and the extraction rate of the aqueous solution is maximized. When the total protein extraction rate is measured to be 17.76% +/-0.02% under the conditions of 25 ℃, 250W and 40kHz for 40min by ultrasonic extraction. The protein extraction rate is higher when the extraction time is 40 min. When the extraction time is short, the protein is not completely dissolved, and the extraction is not complete; when the extraction time is too long, the structure and properties of the protein are damaged to different degrees, which not only reduces the extraction rate of the protein, but also affects the performance of the protein.
In this series of examples, in step (4), the centrifugation conditions were: the rotating speed is 1000-5000 r/min, and the time is 5-15 min. The conditions for better centrifugation effect are as follows: 3000r/min, 10 min.
In the series of embodiments, in the step (5), the temperature of the rotary evaporation is 25-45 ℃. Total protein extraction was better when the temperature of the rotary evaporation was 40 ℃.
In another embodiment of the present disclosure, a total protein of antarctic krill obtained by the above method is provided.
The third embodiment of the disclosure provides a preparation method of sulfur-rich or cesium-rich protein in antarctic krill, which includes the steps of adding water into dry powder of total protein in antarctic krill, placing the dry powder on a vortex mixer, uniformly mixing, removing filter residues through centrifugal separation, collecting supernatant, then separating and purifying the supernatant by taking water as an eluent through Sephadex G-75 gel chromatography to obtain a plurality of protein samples, and performing qualitative and quantitative detection on sulfur or cesium of the plurality of protein samples through inductive coupling plasma emission spectrometry (ICP-5000), wherein the protein sample with the highest content of sulfur or cesium is the antarctic krill sulfur-rich protein or the krill cesium-rich protein.
In one or more embodiments of this embodiment, the steps are as follows:
dissolving dry powder of total protein of Antarctic krill in triple distilled water, uniformly mixing on a vortex mixer, centrifuging the mixed solution by using a high-speed centrifuge, removing filter residue, and collecting supernatant;
eluting the supernatant by using water as an eluent and adopting a Sephadex G-75 gel chromatography method, taking a certain amount of labels every other amount, and respectively carrying out rotary evaporation and concentration to obtain corresponding protein powder after taking a certain amount of protein liquid;
and <3> carrying out qualitative and quantitative detection on sulfur or cesium of a plurality of parts of protein powder by using inductively coupled plasma emission spectrometry to obtain protein liquid corresponding to the protein powder with the highest content of sulfur or cesium, namely the sulfur-rich protein of the antarctic krill or the cesium-rich protein of the antarctic krill.
In the series of embodiments, in the step <1>, the mixture is mixed on a vortex mixer for 1-5 min, the mixed solution is centrifuged for 1-10 min by a high-speed centrifuge at 1000-5000 r/min, and then filter residues are removed to collect supernatant. Mixing for 2min in a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge of 3000r/min, and removing filter residue to collect supernatant, wherein the extraction effect is better.
In the series of examples, in the step <2>, the method for preparing the chromatographic column by the Sephadex G-75 gel chromatography comprises the following steps:
adding Sephadex G-75 dry powder into eluent, and absorbing water to swell to obtain a gel solution;
<2-2> the gel solution was added to the column tube, and naturally settled, and after the gel was settled, the solvent was discharged while the column bed was stabilized by adding the solvent.
In this series of examples, in step <2-1>, a boiling water bath was used for swelling. The swelling time can be reduced to several hours and bacteria and moulds can be killed and any air bubbles which are trapped in the gel can be eliminated.
In the series of examples, in the step <2-1>, the swelling ratio is 1:15 to 20. When the water absorption swelling is carried out at room temperature, the water absorption swelling is fully carried out. When the swelling is carried out in boiling water bath, the swelling time is 60-180 min. When the swelling ratio is 1:15, the swelling time of the boiling water bath is 120 min.
In this series of examples, in the step <2-2>, 2 to 3 bed volumes of the solvent are added to stabilize the bed.
In this series of examples, in step <2>, the loading process into the Sephadex G-75 gel chromatography column was: adding the supernatant to a Sephadex G-75 gel chromatographic column, opening a pipe clamp at the bottom of the gel chromatographic column to make the supernatant flow into the column, and controlling the adding amount and flow rate of the supernatant.
In the series of embodiments, the adding amount of the supernatant is 1-3 mL, and the flow rate is 2-5 mL/10 min. When the adding amount of the supernatant is 2mL and the flow rate is 3mL/10min, the separation effect is better.
In the series of examples, in the step <3>, one part of the protein solution is taken at intervals of 30-50 mL, and 8-20 parts of the protein solution are respectively concentrated by rotary evaporation. When one part is taken every 50mL as one part of protein liquid, and 10 parts are taken, the antarctic krill sulfur-rich protein or the antarctic krill cesium-rich protein is easier to distinguish and obtain.
According to a fourth embodiment of the present disclosure, a sulfur-rich protein in antarctic krill or a cesium-rich protein in antarctic krill obtained by the above preparation method is provided.
In order to make the technical solutions of the present disclosure more clearly understood by those skilled in the art, the technical solutions of the present disclosure will be described in detail below with reference to specific examples and comparative examples.
Example 1: preparation of sulfur-rich protein of Antarctic krill
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (3) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dry protein powder of the antarctic krill, wherein the extraction rate of the protein is 17.76%.
The sulfur content in the dry protein powder was 54.1435ppm by qualitative and quantitative determination of sulfur element by inductively coupled plasma emission spectrometry (ICP-5000).
2. The method for separating the sulfur-rich protein from the Antarctic krill by sephadex chromatography comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step (1) in triple distilled water, mixing for 2min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge at 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 120min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 3 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) carrying out qualitative and quantitative detection on sulfur elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the sulfur element content in the 5 th protein solution is the highest (14.92ppm, and the other 9 parts are below 10 ppm), namely the 5 th protein solution is used as sulfur-rich protein of antarctic krill.
Example 2: preparation of sulfur-rich protein of Antarctic krill
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 20 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (3) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dry protein powder of the antarctic krill, wherein the extraction rate of the protein is 16.37%.
2. The method for separating the sulfur-rich protein from the Antarctic krill by sephadex chromatography comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step (1) in triple distilled water, mixing for 1min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge at 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 120min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 2 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) carrying out qualitative and quantitative detection on sulfur elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the sulfur element content in the 5 th protein solution is the highest (14.89ppm, and the other 9 parts are below 10 ppm), namely the 5 th protein solution is used as sulfur-rich protein of antarctic krill.
Example 3: preparation of sulfur-rich protein of Antarctic krill
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (3) performing rotary evaporation at 10 ℃ until the dried powder is completely dried to obtain the water-soluble dry protein powder of the antarctic krill, wherein the extraction rate of the protein is 17.07%.
2. The method for separating the sulfur-rich protein from the Antarctic krill by sephadex chromatography comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step (1) in triple distilled water, mixing for 4min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge at 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 60min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 3 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) carrying out qualitative and quantitative detection on sulfur elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the sulfur element content in the 5 th protein solution is the highest (14.87ppm, and the other 9 parts are below 10 ppm), namely the 5 th protein solution is used as sulfur-rich protein of antarctic krill.
Example 4: preparation of sulfur-rich protein of Antarctic krill
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Unfreezing and dehydrating frozen fresh antarctic krill, and crushing by using a high-speed tissue crusher;
(2) taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1: 10, and stirring for 80min by a magnetic stirrer;
(3) ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min;
(4) centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min;
(5) and (3) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dry protein powder of the antarctic krill, wherein the extraction rate of the protein is 16.58%.
2. The method for separating the sulfur-rich protein from the Antarctic krill by sephadex chromatography comprises the following steps:
(1) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step 1 in triple distilled water, mixing for 2min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge at 3000r/min, removing filter residues, and collecting supernatant;
(2) selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 60min to kill bacteria and mold, and eliminate the air bubbles in the gel;
(3) the column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 2 bed volumes of solvent;
(4) loading the sample, wherein the sample adding amount is 2mL, and the flow rate is 3mL/10 min;
(5) elution was carried out using a pure aqueous solution as the eluent for the experiment. Taking one part of the mark every 50mL, taking 10 parts of the protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use;
(6) and (3) carrying out qualitative and quantitative detection on sulfur elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the sulfur element content in the 5 th protein solution is the highest (14.88ppm, and the other 9 parts are below 10 ppm), namely the 5 th protein solution is used as sulfur-rich protein of antarctic krill.
Example 5: preparation of euphausia superba caesium-rich protein
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
2. A sephadex chromatographic separation method of euphausia superba caesium-rich protein comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step 1 in triple distilled water, mixing for 3min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 120min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 3 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) carrying out qualitative and quantitative detection on cesium elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the cesium element content in the 2 nd protein solution is the highest (20.06ppm, and the other 9 parts are below 19 ppm), namely the 2 nd protein solution is used as the cesium-rich protein of the antarctic krill.
Example 6: preparation of euphausia superba caesium-rich protein
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 20 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
2. A sephadex chromatographic separation method of euphausia superba caesium-rich protein comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step 1 in triple distilled water, mixing for 3min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 120min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 2 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) carrying out qualitative and quantitative detection on cesium elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the cesium element content in the protein solution of the 2 nd part is the highest (20.05ppm, and the other 9 parts are below 19 ppm), namely, the protein solution of the 2 nd part is used as the cesium-rich protein of the antarctic krill.
Example 7: preparation of euphausia superba caesium-rich protein
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 10 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
2. A sephadex chromatographic separation method of euphausia superba caesium-rich protein comprises the following steps:
(1) and (3) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step 1 in triple distilled water, mixing for 3min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge 3000r/min, removing filter residues, and collecting supernatant.
(2) Selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 60min to kill bacteria and mold, and eliminate air bubbles in the gel.
(3) The column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 3 bed volumes of solvent.
(4) The sample is loaded, the sample adding amount is 2mL, and the flow rate is 3mL/10 min.
(5) Elution was carried out using a pure aqueous solution as the eluent for the experiment. And marking one part every 50mL, taking 10 parts of protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use.
(6) And (3) qualitatively and quantitatively detecting cesium elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the content of the cesium elements in the protein solution of the 2 nd part is the highest (20.03ppm, and the other 9 parts are below 19 ppm), namely, the protein solution of the 2 nd part is used as cesium-rich protein of antarctic krill.
Example 8: preparation of euphausia superba caesium-rich protein
1. Aqueous solution extraction method of total protein of Antarctic krill
(1) Unfreezing and dehydrating frozen fresh antarctic krill, and crushing by using a high-speed tissue crusher;
(2) taking 20g of fresh antarctic krill shrimp paste into a beaker, adding triple distilled water, and mixing the antarctic krill (W) and the triple distilled water (V) in a proportion of 1: 10, and stirring for 80min by a magnetic stirrer;
(3) ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min;
(4) centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min;
(5) and (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
2. A sephadex chromatographic separation method of euphausia superba caesium-rich protein comprises the following steps:
(1) re-dissolving the water-soluble dry protein powder of the euphausia superba extracted in the step (5) in the step 1 in triple distilled water, mixing for 2min on a vortex mixer, centrifuging the mixed solution for 5min by using a high-speed centrifuge at 3000r/min, removing filter residues, and collecting supernatant;
(2) selecting Sephadex G-75 as Sephadex gel, collecting dry powder, pouring the weighed dry powder into eluent, selecting pure water as eluent, and standing at room temperature to make it fully absorb water and swell with swelling ratio of 1: 15. Care should be taken not to over stir to prevent particle breakage. Heating in boiling water bath for 60min to kill bacteria and mold, and eliminate the air bubbles in the gel;
(3) the column is packed by using common column chromatography tube, the gel of thick slurry is poured into the column once to make it naturally settle, and it is necessary to pay attention that there is no air bubble between particles, and it is not necessary to use fractional packing method or dilute gel suspension to pack the column. After the gel had settled, the solvent was drained and the bed was stabilized by 2 bed volumes of solvent;
(4) loading the sample, wherein the sample adding amount is 2mL, and the flow rate is 3mL/10 min;
(5) elution was carried out using a pure aqueous solution as the eluent for the experiment. Taking one part of the mark every 50mL, taking 10 parts of the protein solution, and respectively carrying out rotary evaporation and concentration to 2mL for later use;
(6) and (3) qualitatively and quantitatively detecting cesium elements in the eluted protein solutions by using inductively coupled plasma emission spectrometry (ICP-5000), wherein the content of the cesium elements in the protein solution of the 2 nd part is the highest (20.04ppm, and the other 9 parts are below 19 ppm), namely, the protein solution of the 2 nd part is used as cesium-rich protein of antarctic krill.
Comparative example 1: preparation of sulfur-rich protein of Antarctic krill
1. Diluted acid extraction method for total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, and adding 0.1mol/L CH3COOH solution, Antarctic krill (W) and 0.1mol/L CH3COOH solution (V) at 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
The sulfur content in the dry protein powder was 40.3002ppm by qualitative and quantitative determination of sulfur element by inductively coupled plasma emission spectrometry (ICP-5000).
Comparative example 2: preparation of sulfur-rich protein of Antarctic krill
1. Dilute alkali extraction method for total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) Taking 20g of fresh antarctic krill shrimp paste into a beaker, adding 0.1mol/L sodium hydroxide solution, mixing the antarctic krill (W) and the 0.1mol/L sodium hydroxide solution (V) in a proportion of 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
The sulfur content in the dry protein powder was 34.3026ppm by qualitative and quantitative determination of sulfur element by inductively coupled plasma emission spectrometry (ICP-5000).
Comparative example 3: preparation of sulfur-rich protein of Antarctic krill
1. Method for extracting diluted salt from total protein of Antarctic krill
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) 20g of fresh antarctic krill shrimp paste is put into a beaker, 1% sodium chloride solution in parts by weight is added, and the weight ratio of the antarctic krill (W) to the 1% sodium chloride solution (V) is 1: adding 10-20 parts of the mixture, and stirring for 80min by using a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
The sulfur content in the dry protein powder was 15.5492ppm by qualitative and quantitative determination of sulfur element by inductively coupled plasma emission spectrometry (ICP-5000).
Comparative example 4: preparation of sulfur-rich protein of Antarctic krill
1. Method for extracting total protein of Antarctic krill by using alcoholic solution
(1) Thawing frozen fresh antarctic krill, removing water, and crushing with high-speed tissue pulverizer.
(2) 20g of fresh antarctic krill shrimp paste is put into a beaker, 95% ethanol solution in parts by volume is added, and the volume ratio of the antarctic krill (W) to the 95% ethanol solution (V) is 1:15, and stirring for 80min by a magnetic stirrer.
(3) Ultrasonic extraction in a numerical control ultrasonic cleaning machine is carried out under the following conditions: ultrasonic extracting at 25 deg.C and 250W and 40kHz for 40 min.
(4) Centrifuging by using a high-speed centrifuge, and collecting supernatant under the centrifugation conditions: 3000r/min, 10 min.
(5) And (4) performing rotary evaporation at 40 ℃ until the dried powder is completely dried to obtain the water-soluble dried protein powder of the antarctic krill.
The sulfur content in the dry protein powder was 7.0660ppm by qualitative and quantitative determination of sulfur element by inductively coupled plasma emission spectrometry (ICP-5000).
The above description is only a preferred embodiment of the present disclosure and is not intended to limit the present disclosure, and various modifications and changes may be made to the present disclosure by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present disclosure should be included in the protection scope of the present disclosure.
Claims (16)
1. A preparation method of sulfur-rich protein in antarctic krill is characterized by comprising a method for extracting the total protein of the antarctic krill by using an aqueous solution, adding water into dry powder of the total protein of the antarctic krill, placing the dry powder on a vortex mixer, uniformly mixing, removing filter residues by centrifugal separation, collecting supernatant, then separating and purifying the supernatant by using the water as an eluent and adopting a Sephadex G-75 gel chromatography method to obtain a plurality of protein samples, and carrying out qualitative and quantitative detection on sulfur elements in the protein samples by using an inductive coupling plasma emission spectrometry to obtain the protein sample with the highest sulfur element content, namely the sulfur-rich protein in the antarctic krill;
the method for extracting the total protein of the Antarctic krill by using the aqueous solution comprises the following steps:
(1) unfreezing and dehydrating frozen fresh antarctic krill, and crushing to obtain fresh antarctic krill shrimp paste;
crushing by adopting a high-speed crusher in the step (1);
(2) adding triple-distilled water into fresh euphausia superba mud, and magnetically stirring;
in the step (2), the volume ratio of the mass of the shrimp paste to the volume of the triple distilled water is 1: 5-20;
(3) carrying out ultrasonic extraction;
in the step (3), the ultrasonic extraction conditions are as follows: the temperature is 20-40 ℃, the power is 200-300W, the frequency is 30-40 kHz, and the ultrasonic extraction time is 10-120 min;
(4) centrifuging and collecting supernatant;
in the step (4), the centrifugation conditions are as follows: the rotating speed is 1000-5000 r/min, and the time is 5-15 min;
(5) rotary evaporating to completely dry to obtain water-soluble dry protein powder of Euphausia superba;
in the step (5), the temperature of rotary evaporation is 25-45 ℃.
2. The method according to claim 1, wherein the volume ratio of the mass of the mashed shrimp to the triple distilled water in the step (2) is 1: 15.
3. The method according to claim 1, wherein the ultrasonic extraction is performed at 25 ℃ and 250W at 40kHz for 40min in the step (3).
4. The method according to claim 1, wherein the centrifugation conditions in the step (4) are as follows: 3000r/min, 10 min.
5. The method according to claim 1, wherein the temperature of the rotary evaporation in the step (5) is 40 ℃.
6. The method of claim 5, comprising the steps of:
dissolving dry powder of total protein of Antarctic krill in triple distilled water, uniformly mixing on a vortex mixer, centrifuging the mixed solution by using a high-speed centrifuge, removing filter residue, and collecting supernatant;
eluting the supernatant by using water as an eluent and adopting a Sephadex G-75 gel chromatography method, taking a certain amount of labels every other amount, and respectively carrying out rotary evaporation and concentration to obtain corresponding protein powder after taking a certain amount of protein liquid;
and <3> carrying out qualitative and quantitative detection on sulfur elements of a plurality of protein powder parts by using an inductively coupled plasma emission spectrometry to obtain a protein liquid corresponding to the protein powder with the highest sulfur element content, namely the sulfur-rich protein of the Antarctic krill.
7. The preparation method according to claim 6, wherein in the step <1>, the mixture is mixed on a vortex mixer for 1-5 min, the mixture is centrifuged for 1-10 min by using a high-speed centrifuge at 1000-5000 r/min, and then the supernatant is collected after removing filter residue.
8. The method according to claim 7, wherein in the step <1>, the mixture is mixed for 2min on a vortex mixer, and the mixture is centrifuged for 5min by using a high speed centrifuge of 3000r/min, and then the supernatant collected from the residue is removed.
9. The method according to claim 6, wherein the Sephadex G-75 gel chromatography is performed using a column chromatography method according to step <2> comprising:
adding Sephadex G-75 dry powder into eluent, and absorbing water to swell to obtain a gel solution;
<2-2> the gel solution was added to the column tube, and naturally settled, and after the gel was settled, the solvent was discharged while the column bed was stabilized by adding the solvent.
10. The method according to claim 9, wherein the swelling is carried out in a boiling water bath in the step <2-1 >.
11. The method according to claim 9, wherein the swelling ratio in the step <2-1> is 1:15 to 20.
12. The method according to claim 9, wherein in the step <2-2>, the solvent is added in an amount of 2 to 3 bed volumes to stabilize the bed.
13. The method according to claim 6, wherein the step <2> comprises loading the Sephadex G-75 gel column with the following: adding the supernatant to a Sephadex G-75 gel chromatographic column, opening a pipe clamp at the bottom of the gel chromatographic column to enable the supernatant to flow into the column, and controlling the adding amount and the flow rate of the supernatant;
or the adding amount of the supernatant is 1-3 mL, and the flow rate is 2-5 mL/10 min;
or, in the step <3>, one part of the protein solution is taken every 30-50 mL, 8-20 parts of the protein solution are taken, and the protein solution is respectively subjected to rotary evaporation concentration.
14. The method according to claim 13, wherein in the step <2>, the supernatant is added in an amount of 2mL at a flow rate of 3mL/10 min.
15. The method according to claim 13, wherein in the step <2>, one portion is taken every 50mL and 10 portions are taken as one portion of the protein solution.
16. Antarctic krill sulfur-rich protein obtained by the preparation method of any one of claims 1 to 5.
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| CN101585879A (en) * | 2009-05-19 | 2009-11-25 | 中国科学院亚热带农业生态研究所 | Method of quickly inducing pork liver metallothionein |
| EP1694692B1 (en) * | 2003-12-16 | 2010-04-28 | QIAGEN North American Holdings, Inc. | Formulations and methods for denaturing proteins |
| CN102524510A (en) * | 2010-12-14 | 2012-07-04 | 大连工业大学 | Preparation method of low-fluorine Euphasia superb protein base stock |
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| EP1694692B1 (en) * | 2003-12-16 | 2010-04-28 | QIAGEN North American Holdings, Inc. | Formulations and methods for denaturing proteins |
| CN101585879A (en) * | 2009-05-19 | 2009-11-25 | 中国科学院亚热带农业生态研究所 | Method of quickly inducing pork liver metallothionein |
| CN102524510A (en) * | 2010-12-14 | 2012-07-04 | 大连工业大学 | Preparation method of low-fluorine Euphasia superb protein base stock |
Non-Patent Citations (3)
| Title |
|---|
| 冻藏过程中南极磷虾蛋白理化特性和品质的变化;郭慧等;《食品工业科技》;20181231;第39卷(第23期);第1.2.1-1.2.2节 * |
| 蛋白理化特性和品质的变化;刘志东等;《食品与发酵工业》;20170731;第43卷(第7期);表4、7 * |
| 郭慧等.冻藏过程中南极磷虾蛋白理化特性和品质的变化.《食品工业科技》.2018,第39卷(第23期), * |
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