CN101914043B - Method for synchronously extracting and preparing taurine, biological zinc and polypeptide from oysters - Google Patents
Method for synchronously extracting and preparing taurine, biological zinc and polypeptide from oysters Download PDFInfo
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- CN101914043B CN101914043B CN 201010234452 CN201010234452A CN101914043B CN 101914043 B CN101914043 B CN 101914043B CN 201010234452 CN201010234452 CN 201010234452 CN 201010234452 A CN201010234452 A CN 201010234452A CN 101914043 B CN101914043 B CN 101914043B
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Abstract
The invention discloses a method for extracting and preparing taurine, biological zinc and polypeptide from oysters, comprising the following steps of: separating biological zinc and free amino acid in oyster meat through ultrasonic crushing; poaching the uniformly pulped oyster meat to release the taurine from cells, and then carrying out resin purification and concentrated crystallization to obtain high-purity taurine; and hydrolyzing the residual oyster meat in a bienzyme hydrolysis mode to obtain an oyster extractive rich in polypeptide. The extraction method not only greatly reserves natural active substances in the oysters, but also solves the problem of inapplicability for partial crowds because the oysters do not contain easy-sensitivity albumen and purine, and lays the foundation for the application and the development of the oyster extractive in beverages or foods.
Description
Technical field
The extraction that the present invention relates to three kinds of active substances such as biological zinc, taurine, polypeptide separates preparation, relate in particular to a kind of from oyster the method for these three kinds of active substances of simultaneous extraction.
Background technology
Oyster is the water warm bivalve, and its delicious meat is nutritious, and it is rich in the compositions such as glycogen, protein, amino acid, VITAMIN, biological zinc, trace element, has very high nutritive value.
Taurine is one of amino acid that content is the highest in the oyster.Taurine is called again 2-aminoethyl sulfonic acid, is a kind of special acid of nonprotein structure, is one of important amino acid of needed by human, is prevalent in various tissues in the animal body with the form of total free aminoacids.Taurine is the protective material of oxidative damage, can remove oxyradical, suppresses lipid peroxidation, keeps the Premeabilisation of cells stabilizing cell membrane, to regulate cell calcium stable, simultaneously or the central neurotransmitter, can regulate central nervous system.The effects such as myocardial preservation, anti-arrhythmia are arranged in cardiovascular systems.Taurine is being protected the cerebral ischemia-reperfusion cell, prevention, and the lowering blood glucose level, reducing blood-fat, hypotensive, atherosclerosis and prevention geriatric disease aspect also play an important role.At present both at home and abroad mainly rely on chemosynthesis to produce taurine, but have the serious problems such as the large and environmental pollution of material toxicity.And oyster output is high, and content of taurine is abundant, becomes undoubtedly one of safe natural taurine resource.
Contain the zinc that many effective constituents are not lost in the oyster, and zinc is as a kind of trace element of needed by human, has in vivo different physiological roles, most important to growing of child, can activate kind more than 300 in the organism enzyme, promote acid base equilibrium in body development and bone growth, the adjusting cell, promote sense of taste sense of smell normal formation, keep skin health, strengthen immunity function, prevent from wearing out etc.When lacking zinc in the body, will occur underdevelopment or hypoevolutism, dysgeusia, bone growth slowly, the symptoms such as immunity degradation, congenital abnormality.Zinc participates in the metabolism of body protein and nucleic acid, and is most important to neurocyte, also is the important nutrition of keeping male reproductive system health.
Biologically active peptides has special physiologically active, is mainly reflected in immunocompetence, hypertension, anti-lipid, growth-promoting activity, tumor suppression activity etc., often is present in the amino acid long-chain that consists of protein with inactive form.The nutrient composition content such as protein is very high in the oyster, is the good oceanic resources of preparation peptide class functional foodstuff, by biotechnology, can extract from oyster and prepares water miscible oyster polypeptide.
Summary of the invention
The technical problem that the present invention solves provides a kind of method that adopts the modern techniquies such as ultrasonic disruption, ion-exchange, double-enzyme hydrolysis, lyophilize to extract synchronously three kinds of active substances such as biological zinc, taurine, polypeptide from oyster.
The technical solution used in the present invention may further comprise the steps:
A. biological zinc extraction: Oyster is put into container, add the pure water of 1～3 times of volume, then carry out fragmentation with the ultrasonic disruption instrument, power is 60w～80w, and the time is 15～25min.Because biological zinc mainly is free being present in the oyster fresh meat, adopt the oyster fresh meat of the direct broken monoblock of ultrasonic wave to extract this moment, and the part total free aminoacids also is extracted simultaneously.
B. the separation of biological zinc is concentrated: supernatant liquor is got in described oyster liquid centrifugation behind ultrasonic disruption, the supernatant liquor rotary evaporation is concentrated, then can obtain the concentrated powder of biological zinc by cryodesiccated mode, freeze temperature is-30 ℃.
C. prepare the oyster slurries: in described Oyster, add the pure water of 1～3 times of volume, smash homogenate to pieces.
D. poach extracts taurine: because taurine is soluble small molecular, mainly be free being distributed in the cell, therefore poach after the oyster homogenate can be extracted.The oyster slurries are controlled at 60 ℃ of boiling 2h.With described oyster slurries centrifuging and taking supernatant liquor, and then add 1～3 times of volume pure water, get supernatant after 60 ℃ of boilings, repeat 3 times, supernatant liquor is merged.
E. taurine is concentrated: described supernatant liquor is adopted the mode concentrating under reduced pressure of rotary evaporation, get concentrated solution.
F. the preparation of taurine crude extract: owing to also containing the impurity components such as protein, polysaccharide, inorganic salts except taurine in the extracting solution; therefore need to carry out purifying to sample: add the trichoroacetic acid(TCA) solution that is equivalent to concentrated solution 1/10th volumes in described concentrated solution, concentration is 10%.Leave standstill postprecipitation and get supernatant, then supernatant liquor is transferred pH to 4～5.
G. resin absorption purifying: the Zeo-karb of described supernatant liquor after with activation purified, use the distilled water wash-out, collect taurine chromatographic solution partly.
H. the crystallization purifying of taurine: after described chromatographic solution carried out concentrating under reduced pressure with Rotary Evaporators; add dehydrated alcohol; make the final concentration of ethanol reach 60%～70%; then leave standstill crystallization in 4 ℃; namely get the taurine crystal; repeatedly can obtain highly purified white, needle-shaped crystals behind the recrystallization, drying is processed and is namely got highly purified natural taurine.
I. the preparation of polypeptide 1.: in the centrifugal oyster meat mincing that obtain, add the pure water of 3 times of volumes in steps d.Regulate pH to 6.0～8.5, add Sumizyme MP or neutral protease after being heated to 40～55 ℃, the add-on of proteolytic enzyme is 4%～8% (mass ratio) of Oyster Protein concentration.Enzymolysis time is controlled at 2h, and temperature and pH remain unchanged in the whole process.After enzymolysis is finished enzymolysis solution is heated to 90 ℃～95 ℃, keeps 10min, enzyme goes out.
J. the preparation of polypeptide 2.: described enzymolysis solution pH is adjusted to 3.0～5.0, adds aspartic protease when maintaining the temperature at 40 ℃～55 ℃, the add-on of proteolytic enzyme is 4%～8% (mass ratio) of Oyster Protein concentration.Enzymolysis time is controlled at 2h, and temperature and pH remain unchanged in the whole process.After enzymolysis is finished enzymolysis solution is heated to 90 ℃～95 ℃, keeps 10min, enzyme goes out.
K. the separation of polypeptide: with described two steps enzymolysis solution centrifugation, get supernatant liquor, regulate pH to 6.0～7.0.Then concentrated by the Rotary Evaporators reduction vaporization, namely get the polypeptide concentrated solution.The polypeptide concentrated solution is carried out frozen dried namely get pulverous oyster polypeptide.
The used proteolytic enzyme of described i step can also can for papoid also can be neutral protease, preferably be taken as trypsinase for trypsinase.
In the process of described prepared slarry, the volume ratio of described Oyster and pure water is preferably 1: 3.
In described ultrasonic disruption process, described hyperacoustic preferred power is 80w, and the described treatment time is preferably 20mm.
The invention has the advantages that and from a collection of oyster, to separate simultaneously preparation three seed oyster active substances; extract the amino-acid zinc (biological zinc) of high-content by hyperacoustic crushing effect; prepare highly purified natural taurine by ion-exchange; in the preparation of polypeptide, then adopt the mode of double-enzyme hydrolysis; not only degree of hydrolysis is high; and can obtain high-content amino acid and polypeptide; take full advantage of the effective constituent in the oyster, for oyster is from now on laid a good foundation in the application of the aspects such as food, beverage interpolation.
Embodiment
Embodiment 1
A kind of method for preparing biological zinc, natural taurine, polypeptide of extracting from oyster may further comprise the steps:
A. biological zinc extraction: take by weighing the Oyster of 1000g, carry out fragmentation with the ultrasonic disruption instrument after putting into the pure water that adds 3000g behind the container, power is 80w, and the time is 20min.
B. the separation of biological zinc is concentrated: supernatant liquor is got in the oyster liquid centrifugation after the fragmentation, the supernatant liquor rotary evaporation is concentrated, then carry out lyophilize at-30 ℃ and process, obtain biological zinc powder 11.19g, wherein containing zinc is 63.02%, total free aminoacids 12.83%.
C. prepare the oyster slurries: add the pure water of 3000g in the Oyster after centrifugation, smash homogenate to pieces.
D. poach extracts taurine: the oyster homogenate is controlled at 60 ℃ of boiling 2h.With the oyster slurries centrifuging and taking supernatant liquor after the boiling, and then the pure water of adding 3000g, get supernatant after 60 ℃ of boilings, repeat 3 times, supernatant liquor is merged.
E. taurine is concentrated: the supernatant liquor after will merging adopts the mode concentrating under reduced pressure of rotary evaporation, gets concentrated solution.
F. the preparation of taurine crude extract: add the trichoroacetic acid(TCA) solution that is equivalent to concentrated solution 1/10th volumes in concentrated solution, concentration is 10%.Leave standstill postprecipitation and get supernatant, then supernatant liquor is transferred pH to 4～5.
G. resin absorption purifying: the Zeo-karb of the supernatant liquor after will processing after with activation purified, and use the distilled water wash-out, collects taurine chromatographic solution partly.
H. the crystallization purifying of taurine: after chromatographic solution carried out concentrating under reduced pressure with Rotary Evaporators, add dehydrated alcohol, make the final concentration of ethanol reach 60%, then leave standstill crystallization in 4 ℃, obtain taurine 3.02g after recrystallization and drying treatment, purity is 98.0%.
I. the preparation of polypeptide 1.: in the centrifugal oyster meat mincing that obtain, add the pure water of 3000g in steps d.Regulate pH to 8.0, add trypsinase after being heated to 55 ℃, the add-on of proteolytic enzyme is 6% (mass ratio) of Oyster Protein concentration.Enzymolysis time is controlled at 2h, and whole process temperature remains on 55 ℃, constantly adds NaOH solution and keeps the pH of solution to be stabilized in 7.5～8.5.After enzymolysis is finished enzymolysis solution is heated to 95 ℃, keeps 10min, enzyme goes out.
J. the preparation of polypeptide 2.: the pH of above-mentioned enzymolysis solution is adjusted to 3.0, adds aspartic protease when maintaining the temperature at 45 ℃, the add-on of proteolytic enzyme is 6% (mass ratio) of Oyster Protein concentration.Enzymolysis time is controlled at 2h.After enzymolysis is finished enzymolysis solution is heated to 95 ℃, keeps 10min, enzyme goes out.
K. the separation of polypeptide: with the enzymolysis solution centrifugation of two steps, get supernatant liquor, regulate pH to 6.0～7.0.Then concentrated by the Rotary Evaporators reduction vaporization, namely get the polypeptide concentrated solution.The polypeptide concentrated solution is carried out frozen dried namely get pulverous oyster polypeptide products 61.87g, wherein contain polypeptide 31.04%, total free aminoacids 13.41%.
Embodiment 2
Substantially the same manner as Example 1, difference is in the ultrasonication process, and hyperacoustic power is 70w, and the treatment time is 25min; The enzyme that adds in the first step enzymolysis process in polypeptide preparation is papoid, and temperature is controlled at 55 ℃, and pH is 6.0.Obtain at last biological zinc 10.25g, wherein containing zinc is 58.29%, total free aminoacids 13.50%; Taurine 2.95g, purity is 97.5%; Polypeptide products 65.70g wherein contains polypeptide 38.57%, total free aminoacids 10.54%.
Embodiment 3:
Substantially the same manner as Example 1, when difference is to extract biological zinc, prepared slarry, preparation polypeptide, all add the pure water of 2000g.Obtain at last biological zinc 8.55g, wherein contain zinc 60.40%, total free aminoacids 11.20%; Taurine 1.53g, purity is 98.5%; Polypeptide products 58.75g wherein contains polypeptide 36.65%, total free aminoacids 12.70%.
Embodiment 4:
Substantially the same manner as Example 1, difference is that hyperacoustic treatment time is 15min, adopts the PROTAMEX compound protease to carry out an one-step hydrolysis during preparation polypeptide, and temperature is controlled at 60 ℃, and pH is controlled at 7.0, and the enzyme digestion reaction time is 4h.Obtain at last biological zinc 9.36g, wherein contain zinc 55.60%, total free aminoacids 10.29%; Taurine 2.73g, purity is 98.0%; Polypeptide products 43.76g wherein contains polypeptide 23.40%, total free aminoacids 9.35%.
Embodiment 5:
Substantially the same manner as Example 1; difference is that hyperacoustic broken power is 60w, and the taurine crystallization is that the alcohol concn that adopts is 70%, and the enzyme that adds in the first step enzymolysis process in polypeptide preparation is neutral protease; temperature is controlled at 45 ℃, and pH is 7.0.Obtain at last biological zinc 7.38g, wherein contain zinc 55.70%, total free aminoacids 9.28%; Taurine 1.84g, purity is 97.8%; Polypeptide products 60.75g wherein contains polypeptide 34.05%, total free aminoacids 12.47%.
Claims (3)
1. the method for simultaneous extraction biological zinc, taurine and polypeptide from an oyster, comprise that ultrasonic disruption extracts the step of biological zinc, the step of water-boiling method coupled ion exchange separation and purification taurine, double-enzyme hydrolysis prepares the step of polypeptide, it is characterized in that: the temperature that adopts when adopting water-boiling method to extract taurine is 60 ℃, and the best electrical conductivity scope that accesses the taurine chromatographic solution is 900 μ s/cm～78 μ s/cm; When double-enzyme hydrolysis prepares polypeptide, at first use neutrality or Sumizyme MP, then adopt aspartic protease to carry out enzymolysis, enzyme concentration is 4%～8% of protein content, mass ratio in the enzymolysis process; The concentrated powder of the biological zinc that extraction obtains and polypeptide concentrated solution adopt-30 ℃ of cryodesiccated methods to carry out preservation.
2. the method for claim 1, the preferred trypsinase of described Sumizyme MP.
3. the method for claim 1, described enzyme concentration is preferably 6% of protein content, mass ratio.
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Families Citing this family (8)
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| CN102643368B (en) * | 2012-05-07 | 2014-04-09 | 连云港职业技术学院 | Method for synchronously extracting taurine, chitin and polypeptide from shrimp heads and shrimp leftovers |
| CN103408473B (en) * | 2013-07-22 | 2016-01-06 | 中国科学院海洋研究所 | A kind of method extracting natural taurine from scallop splanchna |
| CN103387521A (en) * | 2013-08-03 | 2013-11-13 | 北海市源龙珍珠有限公司 | Method for extracting taurine from shellfish meat |
| CN103445165B (en) * | 2013-09-04 | 2016-11-23 | 百丽晨（上海）生物科技有限公司 | A kind of for product improving sub-health state and combinations thereof thing |
| CN104783234B (en) * | 2015-05-06 | 2018-06-12 | 北海市银海区蓝海牧场贝类养殖农民专业合作社 | A kind of preparation method of ostreae testa pulverata |
| CN105476014B (en) * | 2015-12-29 | 2018-06-26 | 广西潘达生物科技开发股份有限公司 | A kind of method that biological zinc is prepared using oyster |
| CN110283106A (en) * | 2019-05-28 | 2019-09-27 | 广西大学 | A method of it is synchronous from Pinctada martensii to extract biological zinc and taurine |
| CN110467669A (en) * | 2019-07-22 | 2019-11-19 | 福建福瑞康信息技术有限公司 | The extracting method of Zinc metal sulfur protein zinc in one seed oyster |
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| CN101347460A (en) * | 2007-07-16 | 2009-01-21 | 青岛啤酒股份有限公司 | Oyster extract and method of preparing the same |
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Owner name: YONGAN PHARMACEUTICAL CO., LTD., QIANJIANG Free format text: FORMER OWNER: GUANGXI ACADEMY OF SCIENCE Effective date: 20140522 |
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Effective date of registration: 20140522 Address after: 433132 Hubei Province, Qianjiang City Economic Development Zone, bamboo Ze Road No. 16 Patentee after: Yongan Pharmaceutical Co., Ltd., Qianjiang Address before: 530007 Guangxi Academy of Sciences, 98 Da Ling Road, Nanning, the Guangxi Zhuang Autonomous Region Patentee before: Guangxi Academy of Science |