CN102321596B - Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave - Google Patents
Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave Download PDFInfo
- Publication number
- CN102321596B CN102321596B CN2011102603283A CN201110260328A CN102321596B CN 102321596 B CN102321596 B CN 102321596B CN 2011102603283 A CN2011102603283 A CN 2011102603283A CN 201110260328 A CN201110260328 A CN 201110260328A CN 102321596 B CN102321596 B CN 102321596B
- Authority
- CN
- China
- Prior art keywords
- laver
- lypase
- yeast displaying
- gene
- cell
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/52—Improvements relating to the production of bulk chemicals using catalysts, e.g. selective catalysts
Landscapes
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Enzymes And Modification Thereof (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a method for crushing laver cells by using yeast display lipase cooperated with supersonic wave, which comprises the following steps: adding laver dry powder, yeast display lipase into water and well mixing, performing an enzymatic hydrolysis reaction under the temperature of 35 DEG C-37 DEG C for 30-45 minutes, and then performing ultrasonic crushing by using the 730W-750W of power under the temperature of 30-32 DEG C for 65-70 minutes. The invention also discloses a method for extracting laver protein and laver amylase, which is characterized in that the cell crushed liquid of crushed laver cells is separated. The invention also discloses a method for preparing laver antihypertensive peptide, which is characterized in that the separated laver protein is performed the enzymatic hydrolysis reaction by using protease under the temperature of 36 DEG C-37 DEG C for 130-133 minutes. According to the invention, the yeast display lipase is used to degrade cell membranes of the laver cells and cooperated with the ultrasonic treatment, so that the method of the invention is capable of high efficiently crushing the laver cells, raising the laver cell crushing rate, increase the yields of laver amylase, laver protein and laver antihypertensive peptide.
Description
Technical field
The present invention relates to technical field of bioengineering, be specifically related to the method for the broken laver cell of a primary yeast displaying type lypase synergistic supersonic wave.
Background technology
Laver contains abundant polysaccharide and protein, can be used for preparing active polysaccharide and active polypeptide with physiologically active effect.Laver amylose is the polygalacturonic acid sulfuric ester, has constituted the staple of laver cell wall.Laver cell contains abundant protein, and the laver cell content is flowed out, and can obtain laver albumen.Therefore, laver amylose and laver albumen be obtained fully, laver cell wall and membrane structure should be destroyed as far as possible.
The ultrasonication of the broken mainly employing of laver cell at present.The cavitation effect that ultrasonication produces can destroy laver cell wall and membrane structure, obtains laver amylose and purple laver protein.But use ultrasonication merely, laver cell is kept perfectly basically, only produces a small amount of space, thereby influences the laver cell crushing efficiency, is unfavorable for the proteic extraction of laver amylose and laver.
Summary of the invention
The invention provides the method for the broken laver cell of a primary yeast displaying type lypase synergistic supersonic wave, improve the laver cell percentage of damage, increase the yield of laver amylose, laver albumen and peptide of decrease blood pressure in laver.
The method of the broken laver cell of one primary yeast displaying type lypase synergistic supersonic wave comprises:
Laver dry powder, yeast displaying type lypase are added mixing in the entry, and in 35～37 ℃ of following enzyme digestion reaction 30～45min, under 30～32 ℃, utilizing power again is ultrasonic disruption 65～70min of 730～750W;
Described yeast displaying type lypase prepares through following method:
The recombinant plasmid of linearization process is changed among pichia spp (Pichia pastoris) GS115; The gained transformant is inoculated in the BMMY substratum; Centrifugal collection thalline behind inducing culture 72～144h, thalline makes yeast displaying type lypase through flushing and lyophilize;
Described recombinant plasmid is by initial carrier pPIC9K and insert the lipase gene in MF α 1 signal peptide gene downstream among the initial carrier pPIC9K successively and the cell walls α agglutinin gene of pichia spp GS115 is formed.
Described yeast displaying type lipase gene can be selected for use and be the sequence of AF229435 Genbank number, and pichia spp (Pichia pastoris) GS115 is a commercially produced product, can buy from Invitrogen company.Genbank number of its cell walls α agglutinin gene sequence is M28164.
Carrier pPIC9K is commercially produced product (like an Invitrogen company), and there is MF α 1 signal peptide sequence in it, and (Genbank number: M17301), there is the AOX1 promotor in the signal peptide sequence upper reaches (Genbank number: Z46233) in this carrier simultaneously.The purpose of recombinant plasmid linearization process be for born of the same parents in genome generation homologous recombination, improve expression stability.
In weight part, 1000 parts in water, 20～25 parts in laver dry powder, 1～2 part in yeast displaying type lypase.
Enzyme digestion reaction stirs reaction solution simultaneously, and stir speed (S.S.) is 100～200 commentaries on classics/min.
The present invention also provides a kind of extraction laver amylose or the proteic method of laver, comprising:
Laver dry powder, yeast displaying type lypase are added mixing in the entry; 35～37 ℃ of following enzyme digestion reaction 30～45min; Under 30～32 ℃, utilizing power again is ultrasonic disruption 65～70min of 730～750W, gets cytoclasis liquid, and separation and purification gets laver amylose or laver albumen.
The proteic separation purification method of said laver amylose or laver: said cytoclasis liquid is got supernatant after centrifugal, saltout, 4 ℃ leave standstill behind the 12h centrifugally, and supernatant is a laver amylose solution, is precipitated as laver albumen.
The present invention provides a kind of method for preparing peptide of decrease blood pressure in laver again, comprising:
Laver dry powder, yeast displaying type lypase are added mixing in the entry, and in 35～37 ℃ of following enzyme digestion reaction 30～45min, under 30～32 ℃, utilizing power again is ultrasonic disruption 65～70min of 730～750W, and separation and purification obtains laver albumen; Laver albumen, yeast displaying type proteolytic enzyme are added mixing in the entry,, get peptide of decrease blood pressure in laver in 36～37 ℃ of following enzyme digestion reaction 130～133min;
Described yeast displaying type proteolytic enzyme prepares through following method:
To change over to through the recombinant plasmid of linearization process among pichia spp (Pichia pastoris) GS115; The gained transformant is inoculated in the BMMY substratum; Centrifugal collection thalline behind inducing culture 72～144h, thalline makes yeast displaying type proteolytic enzyme through flushing and lyophilize;
Said recombinant plasmid is by initial carrier pPIC9K and insert the proteinase gene in MF α 1 signal peptide gene downstream among the initial carrier pPIC9K successively and the cell walls α agglutinin gene of pichia spp GS115 is formed.
Described yeast displaying type proteinase gene can be selected for use and be the sequence of NM_012708.2 Genbank number.
In weight part, 1～1.1 part in 1000 parts in water, laver protein 20～22 part, yeast displaying type proteolytic enzyme.
The lypase degradable destroys the phospholipid bilayer of forming the laver cell film; Handle with lypase enzymolysis synergistic supersonic wave, lypase destroys the cavitation effect that can further improve follow-up ultrasonication generation to the degraded of the phospholipid bilayer of composition laver cell film, promotes whole laver cell structure deteriorate; Cellular constituent is discharged as far as possible; Laver amylose and laver albumen and laver cell chip separation then through the processing of saltouing, can obtain higher laver amylose and the laver albumen of purity in high yield ground; Again laver albumen is handled with yeast displaying type protease hydrolyzed, obtained peptide of decrease blood pressure in laver.
The present invention is through importing lipase gene and cell walls α agglutinin gene in the pichia spp GS115 cell; Pichia spp GS115 cell induction is expressed justacrine outside born of the same parents with lypase after cultivating; Utilize cell walls α lectin that this lypase is fixed on cell surface simultaneously, improve the hydrolysis vigor of lypase.Utilize this yeast display lipase that the cytolemma of laver cell is degraded; And and then synergistic supersonic wave handle; The efficiency crushing laver cell is to discharge cellular constituent as much as possible, and centrifugal removal post precipitation is measured laver amylose and purple laver protein content in the supernatant, and the laver amylose yield reaches 40.8～41.9%; Laver albumen yield reaches 47.5～48.5%, and then laver amylose and purple laver protein yield only are respectively 28.7～30.28% and 30.6～32.3% with ultrasonication separately.After lypase enzymolysis synergistic supersonic wave handled contain laver amylose and the proteic solution of laver saltout handle after, centrifugal, supernatant is laver amylose solution, yield is 35.8-36.2%, precipitates to be laver albumen, yield is 43.5～46.1%.Laver albumen after by yeast displaying type protease hydrolyzed peptide of decrease blood pressure in laver, yield reaches 21.5%, is 83.5% to Zinc metallopeptidase Zace1 (ACE) inhibition vigor, has remarkable hypotensive vigor.
Embodiment
Embodiment 1 preparation yeast displaying type lypase and yeast displaying type proteolytic enzyme
Method through synthetic; Synthetic fat enzyme gene (Genbank number: AF229435), proteinase gene (Genbank number: NM_012708.2) with the cell walls α agglutinin gene of pichia spp GS115 (Genbank number: M28164); Add connection peptides sequence GSSGGSGGSGGSGGSGS (linker) at lipase gene (or proteinase gene) C end simultaneously; Obtain nucleotide sequence LIP-linker-α-agglutinin (or PRO-linker-α-agglutinin) after the connection; Add EcoR I and Not I restriction enzyme site simultaneously at the sequence two ends; Wherein LIP is lipase gene (PRO is a proteinase gene), and α-agglutinin is a cell walls α agglutinin gene.
Be template with above-mentioned artificial synthesized sequence respectively, utilize following primer right, carry out pcr amplification,
Primer to lypase is right:
Upstream primer: 5 '-AAGGAAAAAAGAATTCGTTCCAGTTTCTGG～3 ';
Downstream primer: 5 '-TTTTCCTTTTGCGGCCGCTAATGAAACG～3 '
Primer to proteolytic enzyme is right:
Upstream primer: 5 '-CCTTCGCCGCACGCCGCTGCTGATTGGAAA～3 ';
Downstream primer: 5 '-TTTTCCTTTTGCGGCCGCTAATGAAACG～3 '
The PCR reaction system is: template DNA is 1 μ l, high-fidelity DNA polymerase 0.5 μ l, and dNTP (50mM) 0.4 μ l, each 0.5 μ l of upstream and downstream primer, 10 * PCR damping fluid, 5 μ l add water to 50 μ l.
The PCR operational conditions is: 94 ℃ of 3min, 35 circulations (94 ℃ of 30s, 60 ℃ of 1min, 72 ℃ of 30s), 72 ℃ of 10min.
With EocR I and Not I simultaneously enzyme cut PCR product and pPIC9K plasmid, and under the effect of T4 ligase enzyme, spend the night and be connected to form pPIC9K-LIP plasmid (or pPIC9K-PRO plasmid), through electrophoresis check and recovery plasmid.For making goal gene and pichia spp GS115 that His 4 unit points displacement reorganization take place, with Sal I pPIC9K-LIP plasmid (or pPIC9K-PRO plasmid) is carried out linearization for enzyme restriction and handle.The about 15 μ l of goal gene that linearization for enzyme restriction is handled well join in the previously prepd pichia spp GS115 competent cell; Change in the electric revolving cup; Ice bath 15min, the 10ms that under 1500V, 400 Ω, 25uF condition, shocks by electricity then, and the sorbyl alcohol of the about 1ml precooling of adding.The electricity of the above-mentioned mixing about 400 μ l of thing that change the line of production are applied on the MD flat board, and the screening positive transformant is applied to positive transformant on the G418 flat board of different concns then, the resistance screening of G418 is gone out the positive recombinant bacterial strain of multiple copied of Mut phenotype according to positive transformant.
The positive recombinant bacterial strain of multiple copied is seeded in fermentation culture 30h in the BMGY substratum, centrifugal collecting cell; Again cell is placed the BMMY substratum inducing culture 144h that contains 0.5% (volume percent) methyl alcohol; Centrifugal collecting cell after the water flushing, is seeded to 30 ℃ of cultivation 120h in the YGC substratum; The centrifugal 5min of 3000g collects thalline then; Again with the washing of 50mM pH7.0 phosphoric acid buffer, again through the dry 24h of German Christ vacuum freeze drier, obtain yeast displaying type lypase and yeast displaying type proteolytic enzyme after-80 ℃ of following pre-freezes behind the distilled water wash.
The broken laver cell of embodiment 2 yeast displaying type lypase synergistic supersonic waves
Instance 1 water 1000 grams, laver dry powder 20 grams, yeast displaying type lypase 1 gram are mixed with mixing suspension, and the system stir speed (S.S.) is 100 commentaries on classics/min; Temperature is 35 ℃; Treatment time is 30min, supersound process then, and ultrasonic power 730W, supersound process time are that 65min, treatment temp are 30 ℃; Obtain cytoclasis liquid, centrifugal (4000g 30min) goes post precipitation to survey the yield of laver albumen and laver amylose in the supernatant.
Instance 2 water 1000 grams, laver dry powder 25 grams, yeast displaying type lypase 2 grams are mixed with mixing suspension, and the system stir speed (S.S.) is 200 commentaries on classics/min; Temperature is 37 ℃; Treatment time is 45min, supersound process then, and ultrasonic power 750W, supersound process time are that 65min, treatment temp are 30 ℃; Obtain cytoclasis liquid, centrifugal (4000g 30min) goes post precipitation to survey the yield of laver albumen and laver amylose in the supernatant.
Embodiment 3 uses the ultrasonic disruption laver cell separately
Instance 1 water 1000 grams, laver dry powder 20 grams; Be mixed with mixing suspension; Supersound process then; Ultrasonic power 730W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) goes post precipitation to survey the yield of laver albumen and laver amylose in the supernatant.
Instance 2 water 1000 grams, laver dry powder 25 grams; Be mixed with mixing suspension; Supersound process then; Ultrasonic power 750W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) goes post precipitation to survey the yield of laver albumen and laver amylose in the supernatant.
The broken laver cell of embodiment 4 yeast displaying type lypase synergistic supersonic waves extracts laver amylose
Instance 1 water 1000 grams, laver dry powder 20 grams, yeast displaying type lypase 1 gram are mixed with mixing suspension, and the system stir speed (S.S.) is 100 commentaries on classics/min; Temperature is 35 ℃, and the treatment time is 30min, supersound process then; Ultrasonic power 730W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) gets supernatant with cytoclasis liquid; In supernatant, add ammonium sulfate concentrations to 20% (W/V) again, 4 ℃ leave standstill 12h after, centrifugal (4000g 30min) removes deposition; Supernatant is laver amylose solution, surveys the yield of laver amylose in the laver amylose solution.
Instance 2 water 1000 grams, laver dry powder 25 grams, yeast displaying type lypase 2 grams are mixed with mixing suspension, and the system stir speed (S.S.) is 200 commentaries on classics/min; Temperature is 37 ℃, and the treatment time is 45min, supersound process then; Ultrasonic power 750W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) gets supernatant with cytoclasis liquid; In supernatant, add ammonium sulfate concentrations to 25% (W/V) again, 4 ℃ leave standstill 12h after, centrifugal (4000g 30min) removes deposition; Supernatant is laver amylose solution, surveys the yield of laver amylose in the laver amylose solution.
The broken laver cell of embodiment 5 yeast displaying type lypase synergistic supersonic waves extracts laver albumen
Instance 1 water 1000 grams, laver dry powder 20 grams, yeast displaying type lypase 1 gram are mixed with mixing suspension, and the system stir speed (S.S.) is 100 commentaries on classics/min; Temperature is 35 ℃, and the treatment time is 30min, supersound process then; Ultrasonic power 730W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) gets supernatant with cytoclasis liquid; In supernatant, add ammonium sulfate concentrations to 20% (W/V) again, 4 ℃ leave standstill 12h after, centrifugal (4000g 30min) removes supernatant; Deposition is laver albumen, surveys the proteic yield of laver, and present embodiment gained laver albumen can be used for preparing peptide of decrease blood pressure in laver.
Instance 2 water 1000 grams, laver dry powder 25 grams, yeast displaying type lypase 2 grams are mixed with mixing suspension, and the system stir speed (S.S.) is 200 commentaries on classics/min; Temperature is 37 ℃, and the treatment time is 45min, supersound process then; Ultrasonic power 750W, supersound process time are that 65min, treatment temp are 30 ℃, obtain cytoclasis liquid, and centrifugal (4000g 30min) gets supernatant with cytoclasis liquid; In supernatant, add ammonium sulfate concentrations to 25% (W/V) again, 4 ℃ leave standstill 12h after, after centrifugal (4000g 30min) removes supernatant; Deposition is laver albumen, surveys the proteic yield of laver, and present embodiment gained laver albumen can be used for preparing peptide of decrease blood pressure in laver.
Embodiment 6 yeast displaying type protease hydrolyzeds are handled laver protein Preparation peptide of decrease blood pressure in laver
Water 1000 grams, laver protein 20 gram, yeast displaying type proteolytic enzyme 1 gram are configured to mixing suspension.The system stir speed (S.S.) is 160 commentaries on classics/min during processing, and temperature is 36.5 ℃, and the treatment time is 130min; Centrifugal (4000g 30min) gets supernatant; In supernatant, add ammonium sulfate concentrations to 25% (W/V) again, 4 ℃ leave standstill 12h after, after centrifugal (4000g 30min) removes supernatant; Deposition is peptide of decrease blood pressure in laver, surveys the yield and the hypotensive vigor of peptide of decrease blood pressure in laver.
Laver albumen and peptide of decrease blood pressure in laver Determination on content adopt ultraviolet spectrophotometry among the above embodiment, are standard substance with the casein.The laver amylose Determination on content adopts the phenolsulfuric acid colourimetry, is standard substance with D～semi-lactosi, and test result is as shown in the table:
Annotate: the proteic extraction yield of laver amylose or laver is defined as the ratio of laver amylose in the extracting solution or proteic amount of laver and laver raw material dry weight; The rate of being prepared into of peptide of decrease blood pressure in laver is defined as the ratio of the amount and the laver raw material dry weight of the peptide of decrease blood pressure in laver for preparing.
The hypotensive vigour-testing method of peptide of decrease blood pressure in laver is following among the above embodiment 6:
The Zinc metallopeptidase Zace1 (ACE) of measuring prepared peptide of decrease blood pressure in laver suppresses active, estimates its hypotensive vigor.Contain 5mM Hip-His-Leu in the 50 μ l reaction mixtures as reaction substrate, 0.3M NaCl, 5mU ACE and 50mM borate buffer solution (pH 8.3).50 μ l samples join in the above-mentioned reaction mixture, after 37 ℃ are reacted 3h down, add 250 μ l 1.0N HCl termination reactions, in reaction system, add 1.5ml ETHYLE ACETATE again.In the vortex oscillator concussion 2min after, the centrifugal 15min of 2500g, leave standstill 10min after; Draw 1.0ml ethyl acetate layer liquid in other test tube with pipettor; Dry 60min in loft drier adds 3.0ml zero(ppm) water again it is dissolved, and measures this solution light absorption value at the 228nm place.
ACE inhibiting rate (%)=(A-B)/(A-C) * 100%
In formula; A is for there not being peptide of decrease blood pressure in laver but the absorbancy when having Zinc metallopeptidase Zace1 (ACE); The absorbancy of B when having peptide of decrease blood pressure in laver and Zinc metallopeptidase Zace1 (ACE), C is peptide of decrease blood pressure in laver and Zinc metallopeptidase Zace1 (ACE) absorbancy when not existing.
According to aforesaid method the peptide of decrease blood pressure in laver for preparing is detected, it is 83.5% to Zinc metallopeptidase Zace1 (ACE) inhibition vigor, has remarkable hypotensive vigor.
Claims (5)
1. the method for the broken laver cell of a primary yeast displaying type lypase synergistic supersonic wave comprises:
Laver dry powder, yeast displaying type lypase are added mixing in the entry, and in 35～37 ℃ of following enzyme digestion reaction 30～45min, under 30～32 ℃, utilizing power again is ultrasonic disruption 65～70min of 730～750W;
Said yeast displaying type lypase prepares through following method:
To change over to through the recombinant plasmid of linearization process among pichia spp (Pichia pastoris) GS115; The gained transformant is inoculated in the BMMY substratum; Centrifugal collection thalline behind inducing culture 72～144h, thalline makes yeast displaying type lypase through flushing and lyophilize;
Said recombinant plasmid is by initial carrier pPIC9K and insert the lipase gene in MF α 1 signal peptide gene downstream among the initial carrier pPIC9K successively and the cell walls α agglutinin gene of pichia spp GS115 is formed;
The Genbank of said lipase gene number is AF229435; Genbank number of said cell walls α agglutinin gene is M28164.
2. method according to claim 1 is characterized in that, in weight part, and 1000 parts in water, 20～25 parts in laver dry powder, 1～2 part in yeast displaying type lypase.
3. one kind is extracted laver amylose or the proteic method of laver, comprising:
Laver dry powder, the described yeast displaying of claim 1 type lypase are added mixing in the entry; In 35～37 ℃ of following enzyme digestion reaction 30～45min; Under 30～32 ℃, utilizing power again is ultrasonic disruption 65～70min of 730～750W; Get cytoclasis liquid, separation and purification gets laver amylose or laver albumen.
4. method according to claim 3 is characterized in that, the proteic separation purification method of said laver amylose or laver is: said cytoclasis liquid is got supernatant after centrifugal; Saltout; 4 ℃ leave standstill behind the 12h centrifugally, and supernatant is a laver amylose solution, is precipitated as laver albumen.
5. method for preparing peptide of decrease blood pressure in laver comprises:
Laver dry powder, the described yeast displaying of claim 1 type lypase are added mixing in the entry, and in 35～37 ℃ of following enzyme digestion reaction 30～45min, under 30～32 ℃, utilizing power is ultrasonic disruption 65～70min of 730～750W, and separation and purification gets laver albumen; Laver albumen, yeast displaying type proteolytic enzyme are added mixing in the entry,, get peptide of decrease blood pressure in laver in 36～37 ℃ of following enzyme digestion reaction 130～133min;
Described yeast displaying type proteolytic enzyme prepares through following method:
To change over to through the recombinant plasmid of linearization process among pichia spp (Pichia pastoris) GS115; The gained transformant is inoculated in the BMMY substratum; Centrifugal collection thalline behind inducing culture 72～144h, thalline makes yeast displaying type proteolytic enzyme through flushing and lyophilize;
Said recombinant plasmid is by initial carrier pPIC9K and insert the proteinase gene in MF α 1 signal peptide gene downstream among the initial carrier pPIC9K successively and the cell walls α agglutinin gene of pichia spp GS115 is formed;
The Genbank of said proteinase gene number is NM_012708.2; Genbank number of said cell walls α agglutinin gene is M28164.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011102603283A CN102321596B (en) | 2011-09-05 | 2011-09-05 | Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011102603283A CN102321596B (en) | 2011-09-05 | 2011-09-05 | Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102321596A CN102321596A (en) | 2012-01-18 |
| CN102321596B true CN102321596B (en) | 2012-11-14 |
Family
ID=45449432
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011102603283A Expired - Fee Related CN102321596B (en) | 2011-09-05 | 2011-09-05 | Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102321596B (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101270148A (en) * | 2008-01-29 | 2008-09-24 | 南京农业大学 | Preparation for high-purity laver phycoerythrin with one-step chromatography |
| CN101624613A (en) * | 2009-08-06 | 2010-01-13 | 南京农业大学 | Bio-modification preparation method for improving activity of laver phycoerythrin |
| CN102154423A (en) * | 2011-01-12 | 2011-08-17 | 淮海工学院 | Method for preparing active peptide of laver |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS58149666A (en) * | 1982-02-26 | 1983-09-06 | Nagoya Seiraku Kk | Beverage containing sea weeds |
-
2011
- 2011-09-05 CN CN2011102603283A patent/CN102321596B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101270148A (en) * | 2008-01-29 | 2008-09-24 | 南京农业大学 | Preparation for high-purity laver phycoerythrin with one-step chromatography |
| CN101624613A (en) * | 2009-08-06 | 2010-01-13 | 南京农业大学 | Bio-modification preparation method for improving activity of laver phycoerythrin |
| CN102154423A (en) * | 2011-01-12 | 2011-08-17 | 淮海工学院 | Method for preparing active peptide of laver |
Non-Patent Citations (3)
| Title |
|---|
| JP昭58-149666A 1983.09.06 |
| 何荣海等.超声波在酶法生产紫菜降血压肽过程中的应用.《江苏大学学报（自然科学版）》.2007,第28卷(第1期),第4-7页. * |
| 段杉等.利用纤维素酶辅助提取紫菜藻红蛋白的研究.《水产科学》.2009,第28卷(第5期),第280-283页. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102321596A (en) | 2012-01-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102154423B (en) | Method for preparing active peptide of laver | |
| CN105399855A (en) | Method for preparing oat beta-glucan through ball milling | |
| CN102212601B (en) | Method for performing catalytic synthesis of vitamin A palmitate by using yeast display lipase | |
| CN102321596B (en) | Method for crushing laver cells by using yeast display lipase cooperated with supersonic wave | |
| CN102321598B (en) | Method for disrupting seaweed cells by using yeast display pectinase cooperated with ultrasonic waves | |
| CN102296101B (en) | Method for preparing micro capsulate embedded antihypertensive peptide | |
| CN102994476B (en) | Saccharifying enzyme | |
| CN106434711A (en) | Method for improving enzyme activity of feruloyl esterase | |
| CN105838685A (en) | Method for extracting SOD (superoxide dismutase) from mulberry wine lees | |
| CN102212586B (en) | Method for catalytically synthesizing starch sodium alkenylsuccinate (SSAS) with yeast display lipase | |
| CN103013956A (en) | Saccharifying enzyme and recombinant expression strain thereof | |
| CN103031289A (en) | Lactase and recombinant expression engineering bacterium thereof | |
| CN102212584B (en) | Method for catalyzing and synthesizing starch acetate through yeast show lipase | |
| CN102286110B (en) | Method for extracting porphyra polysaccharide through ultrasonic treatment with cooperation of yeast display type polygalacturonase | |
| CN102212583B (en) | Method for catalyzing and synthesizing octenyl succinic anhydride modified starch ester through yeast show lipase | |
| CN102212593B (en) | Method for performing catalytic synthesis of maltose stearate by using yeast display lipase | |
| CN102212580B (en) | Method for catalytic synthesis of glucose laurate by utilizing yeast display lipase | |
| CN105087519A (en) | Genetic engineering inulase and method for preparing crystalline fructose with jerusalem artichoke as raw material | |
| CN102212578B (en) | Method for synthesizing fructose palmitate by catalysis of yeast display lipase | |
| CN202081041U (en) | Ultrasonic auxiliary extracting system for cake protein | |
| CN102212579B (en) | Method for catalyzing and synthesizing glucose myristate through yeast show lipase | |
| CN102212585B (en) | Method for catalytic synthesis of starch phosphate by utilizing yeast display lipase | |
| CN104263651A (en) | Semi-continuous culture method for spores recovered from cordyceps sinensis fermentation liquor | |
| CN102226209B (en) | Method for catalyzing and synthesizing vitamin E oleate by using lipase displayed on yeast | |
| CN102212588B (en) | Method for performing catalytic synthesis of starch phosphate monoester by using yeast display lipase |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20121114 Termination date: 20150905 |
|
| EXPY | Termination of patent right or utility model |