CN107646764A - It is a kind of grow cultured pearls before change Pteria penguin mother's shellfish color and luster method and Pteria penguin mother's shellfish color and luster detection method - Google Patents
It is a kind of grow cultured pearls before change Pteria penguin mother's shellfish color and luster method and Pteria penguin mother's shellfish color and luster detection method Download PDFInfo
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- CN107646764A CN107646764A CN201710937715.3A CN201710937715A CN107646764A CN 107646764 A CN107646764 A CN 107646764A CN 201710937715 A CN201710937715 A CN 201710937715A CN 107646764 A CN107646764 A CN 107646764A
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- 241000183300 Pteria penguin Species 0.000 title claims abstract description 104
- 235000015170 shellfish Nutrition 0.000 title claims abstract description 97
- 239000002932 luster Substances 0.000 title claims abstract description 48
- 238000000034 method Methods 0.000 title claims abstract description 39
- 238000001514 detection method Methods 0.000 title claims abstract description 22
- 239000010984 cultured pearl Substances 0.000 title claims abstract description 19
- 230000008859 change Effects 0.000 title claims abstract description 15
- BEJNERDRQOWKJM-UHFFFAOYSA-N kojic acid Chemical compound OCC1=CC(=O)C(O)=CO1 BEJNERDRQOWKJM-UHFFFAOYSA-N 0.000 claims abstract description 35
- 229960004705 kojic acid Drugs 0.000 claims abstract description 35
- WZNJWVWKTVETCG-UHFFFAOYSA-N kojic acid Natural products OC(=O)C(N)CN1C=CC(=O)C(O)=C1 WZNJWVWKTVETCG-UHFFFAOYSA-N 0.000 claims abstract description 35
- 239000000049 pigment Substances 0.000 claims abstract description 35
- 239000011049 pearl Substances 0.000 claims abstract description 21
- 238000009313 farming Methods 0.000 claims abstract description 15
- 235000013305 food Nutrition 0.000 claims abstract description 12
- 239000013535 sea water Substances 0.000 claims abstract description 5
- 238000009360 aquaculture Methods 0.000 claims abstract description 4
- 244000144974 aquaculture Species 0.000 claims abstract description 4
- 230000029087 digestion Effects 0.000 claims abstract description 4
- 230000000284 resting effect Effects 0.000 claims abstract description 3
- XUMBMVFBXHLACL-UHFFFAOYSA-N Melanin Chemical compound O=C1C(=O)C(C2=CNC3=C(C(C(=O)C4=C32)=O)C)=C2C4=CNC2=C1C XUMBMVFBXHLACL-UHFFFAOYSA-N 0.000 claims description 28
- 150000002500 ions Chemical class 0.000 claims description 15
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 12
- 239000012043 crude product Substances 0.000 claims description 11
- 238000011534 incubation Methods 0.000 claims description 10
- 239000006228 supernatant Substances 0.000 claims description 10
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 8
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 claims description 7
- 238000007254 oxidation reaction Methods 0.000 claims description 7
- 238000001914 filtration Methods 0.000 claims description 6
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 claims description 6
- 230000003647 oxidation Effects 0.000 claims description 6
- 230000008569 process Effects 0.000 claims description 6
- 238000012360 testing method Methods 0.000 claims description 6
- 238000006243 chemical reaction Methods 0.000 claims description 5
- 230000014759 maintenance of location Effects 0.000 claims description 5
- 239000000047 product Substances 0.000 claims description 5
- KKJUPNGICOCCDW-UHFFFAOYSA-N 7-N,N-Dimethylamino-1,2,3,4,5-pentathiocyclooctane Chemical compound CN(C)C1CSSSSSC1 KKJUPNGICOCCDW-UHFFFAOYSA-N 0.000 claims description 4
- 108090000526 Papain Proteins 0.000 claims description 4
- 239000004365 Protease Substances 0.000 claims description 4
- 238000004458 analytical method Methods 0.000 claims description 4
- 239000007864 aqueous solution Substances 0.000 claims description 4
- 239000007789 gas Substances 0.000 claims description 4
- 238000002347 injection Methods 0.000 claims description 4
- 239000007924 injection Substances 0.000 claims description 4
- 229940055729 papain Drugs 0.000 claims description 4
- 235000019834 papain Nutrition 0.000 claims description 4
- 239000008363 phosphate buffer Substances 0.000 claims description 4
- 239000000243 solution Substances 0.000 claims description 4
- 238000004704 ultra performance liquid chromatography Methods 0.000 claims description 4
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 claims description 3
- 241000195649 Chlorella <Chlorellales> Species 0.000 claims description 3
- 241000272194 Ciconiiformes Species 0.000 claims description 3
- 241000196324 Embryophyta Species 0.000 claims description 3
- 230000033228 biological regulation Effects 0.000 claims description 3
- 238000004807 desolvation Methods 0.000 claims description 3
- 235000019253 formic acid Nutrition 0.000 claims description 3
- SVWLIIFHXFGESG-UHFFFAOYSA-N formic acid;methanol Chemical compound OC.OC=O SVWLIIFHXFGESG-UHFFFAOYSA-N 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 3
- 239000012044 organic layer Substances 0.000 claims description 3
- 238000005070 sampling Methods 0.000 claims description 3
- 239000007921 spray Substances 0.000 claims description 3
- 239000003643 water by type Substances 0.000 claims description 3
- 239000012670 alkaline solution Substances 0.000 claims description 2
- 238000001816 cooling Methods 0.000 claims description 2
- 238000012258 culturing Methods 0.000 claims description 2
- 239000000284 extract Substances 0.000 claims description 2
- 238000002552 multiple reaction monitoring Methods 0.000 claims description 2
- 241000605372 Fritillaria Species 0.000 claims 1
- 239000002002 slurry Substances 0.000 claims 1
- 238000000638 solvent extraction Methods 0.000 claims 1
- 230000036564 melanin content Effects 0.000 abstract description 10
- 230000004083 survival effect Effects 0.000 abstract description 10
- 238000012544 monitoring process Methods 0.000 abstract description 4
- 230000002829 reductive effect Effects 0.000 abstract description 3
- 239000012071 phase Substances 0.000 description 13
- 230000000052 comparative effect Effects 0.000 description 9
- 238000012545 processing Methods 0.000 description 7
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 6
- 238000005119 centrifugation Methods 0.000 description 5
- 229910000027 potassium carbonate Inorganic materials 0.000 description 4
- 238000009395 breeding Methods 0.000 description 3
- 230000001488 breeding effect Effects 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000001556 precipitation Methods 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- 241000195958 Dicksoniaceae Species 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- 229940124200 Melanin inhibitor Drugs 0.000 description 2
- 239000012491 analyte Substances 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000004047 hole gas Substances 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 241000272814 Anser sp. Species 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000692870 Inachis io Species 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 235000019219 chocolate Nutrition 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 150000002170 ethers Chemical class 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 238000003760 magnetic stirring Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
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- 238000004445 quantitative analysis Methods 0.000 description 1
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- 239000011734 sodium Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
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- 231100000419 toxicity Toxicity 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K61/00—Culture of aquatic animals
- A01K61/50—Culture of aquatic animals of shellfish
- A01K61/54—Culture of aquatic animals of shellfish of bivalves, e.g. oysters or mussels
- A01K61/56—Culture of aquatic animals of shellfish of bivalves, e.g. oysters or mussels for pearl production
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
-
- 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
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
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- General Health & Medical Sciences (AREA)
- Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- General Physics & Mathematics (AREA)
- Environmental Sciences (AREA)
- Zoology (AREA)
- Animal Husbandry (AREA)
- Biodiversity & Conservation Biology (AREA)
- Marine Sciences & Fisheries (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The present invention relates to before a kind of grow cultured pearls change Pteria penguin mother's shellfish color and luster method and Pteria penguin mother's shellfish color and luster detection method, it is described grow cultured pearls before change Pteria penguin mother's shellfish color and luster method comprise the following steps：S1：Pteria penguin mother shellfish is taken out from aquaculture sea area, is put in artificial farming pond and temporarily supports, feed food；S2：Pause, which is fed, makes it that food digestion is complete；S3：Kojic acid culture is launched to artificial farming pond；S4：Pteria penguin mother shellfish is placed in seawater after resting can be used to plant core grow cultured pearls.It is provided by the invention grow cultured pearls before change the method for Pteria penguin mother's shellfish color and luster and can obviously reduce the melanin content of Pteria penguin mother shellfish, survival rate is high, melanin content highest is reduced to 0.08ug/mg in Pteria penguin mother's shellfish outer embrane, reduce 89.5%, it can be applied to cultivate the unusual high quality pearl of color and luster on a large scale, be with a wide range of applications.Simultaneously Pteria penguin mother shellfish color and luster provided by the invention detection method can Accurate Determining Pteria penguin mother shellfish pigment content, so as to realize detection and monitoring to Pteria penguin mother's shellfish color and luster.
Description
Technical field
The present invention relates to Pteria penguin to cultivate field, and it is female to change Pteria penguin before being grown cultured pearls more particularly, to one kind
The method and Pteria penguin mother's shellfish color and luster detection method of shellfish color and luster.
Background technology
The color and luster of pearl is the important indicator for judging its value, and the color and luster of pearl is mainly by the nacre of mother pearl shellfish secretion
Determine, and nacre is mainly secreted by the outer embrane of female shellfish, its color and luster also depends primarily on the species of female Bei Suohan pigments and contained
Amount.Pteria penguin（Pteria penguin）The preferable female shellfish for cultivating large-scale high-quality sea water pearls, produce pearl pearl thickness,
Diameter is big, is considered China " revitalizing southern pearl " most potential kind by industrial circle.At present, Pteria penguin seeling industry, into
Shellfish cultivates to the artificial breeding technique of attached shell pearl and has been mature on the whole, highly specialized, industrial chain is complete.Pteria penguin is produced
Pearl is based on black and bronze-colored dark color series, gold, chocolate, peacock green and other rare color and lusters of high value
Pearl ratio it is smaller.The thing expected using the Pteria penguin rare color and luster pearl of cultivation high-quality as jewelry circle.
Melanin is one of pigment that distribution is most wide in biochrome, not only not soluble in water but also organic insoluble in major part
Solvent.Melanin is divided into two kinds：Eumelanin（eumelanin）With de- melanin（phemelanin）, eumelanin makes life
Brown-black is presented in object, and de- melanin makes organism that yellow-rufous be presented.Pteria penguin colony shell is homogeneous
Black, its primary pigments composition is eumelanin.By the way of xenobiontics are added, suppress or strengthen early stage planting core
The expression of Pteria penguin mother's shellfish melanin, it is that acquisition shell color is special so as to regulate and control the color and luster of female shellfish and its produced pearl
The effective way of high-quality pearl, the very long cycle of conventional hybridization breeding can be shortened in production, be fine-variety breeding and high-quality
Pearl cultivating provides reserve supply.At present, it is rarely reported on regulating and controlling the method for Pteria penguin color and luster.
Therefore, developing a kind of method for changing Pteria penguin mother's shellfish color and luster before grow cultured pearls has important Research Significance and answers
With value.
The content of the invention
It is an object of the invention to overcome the defects of rare color and luster of Pteria penguin is less in the prior art, there is provided Yi Zhongyu
Change the method for Pteria penguin mother's shellfish color and luster before pearl.Method provided by the invention can obviously reduce the black of Pteria penguin mother shellfish
Pigment content, it can be applied to cultivate the unusual high quality pearl of color and luster on a large scale, be with a wide range of applications.
Another object of the present invention is to provide the detection method of above-mentioned Pteria penguin mother shellfish color and luster.
For achieving the above object, the present invention adopts the following technical scheme that：
It is a kind of grow cultured pearls before change Pteria penguin mother's shellfish color and luster method, it is characterised in that methods described comprises the following steps：
S1：Pteria penguin mother shellfish is taken out from aquaculture sea area, is put in artificial farming pond and temporarily supports, feed food；
S2：Pause, which is fed, makes it that food digestion is complete；
S3：Kojic acid, culture are launched to artificial farming pond；Kojic acid concentration is 1 ~ 10g/L in the artificial farming pond, and incubation time is
Kojic acid concentration is 10g/L in 18h, or the artificial farming pond, and incubation time is kojic acid in 6 ~ 18h, or the artificial farming pond
Concentration is 10 ~ 100g/L, cultivates 6h；
S4：Pteria penguin mother shellfish is placed in seawater after resting can be used to plant core grow cultured pearls.
The inventors found that kojic acid is a kind of effective Pteria penguin mother's shellfish melanin inhibitor, launched
The melanin content of Pteria penguin mother shellfish is effectively reduced in the artificial farming Chi Zhongke of Pteria penguin mother shellfish.The present invention is logical
Crossing control kojic acid concentration and incubation time can not only ensure to effectively reduce the pigment content of Pteria penguin mother shellfish, can also keep away
Exempt from the influence that kojic acid is survived to Pteria penguin mother shellfish.
Preferably, the food fed in S1 is chlorella, and temporarily the foster time is more than 1 week.
Pteria penguin mother shellfish is temporarily supported to the ring that may be such that Pteria penguin mother shellfish adapts to artificial farming completely for more than one week
Border.
Preferably, 2 days of the time fed are suspended in S2.
Pteria penguin mother Bei Nei food is containing pigment mostly, and pause launches food and may be such that Pteria penguin mother
Food in shellfish is digested completely, and internal pigment is not disturbed by food.
Preferably, kojic acid concentration is 10g/L, incubation time 18h in the artificial farming pond.
When kojic acid concentration is 10g/L, incubation time is 18h, it both can ensure that Pteria penguin melanin content was remarkably decreased,
Its survival rate is not influenceed again.
A kind of detection method of Pteria penguin mother shellfish color and luster, methods described comprise the following steps：
S1：Take Pteria penguin mother's shellfish melanin sample to be dissolved with methanol aqueous solution, test sample is obtained after filtering；
S2：Test sample injection LC-MS is subjected to analysis detection, wherein, mobile phase A is that the formic acid that volume fraction is 0.1% is water-soluble
Liquid, Mobile phase B are the formic acid methanol solution that volume fraction is 0.1%；Elution program is 0 ~ 6min, mobile phase A and B ratio
For 90%:10%；6 ~ 8min, the ratio of Mobile phase B is 100%；8 ~ 12min, mobile phase A and B ratio are 90%:10%.
The present invention, can under above-mentioned flowing phase composition and elution program by the exploration discovery to mobile phase and elution program
The content of the pigment of Accurate Determining Pteria penguin mother shellfish, so as to realize detection and monitoring to Pteria penguin mother's shellfish color and luster.
Preferably, the volume fraction of methanol aqueous solution is 50% in S1.
Preferably, setting MRM tuner parameters are in S2：PDCA：Parent ion is 155.98, product ion 138.01, is protected
It is 2.9min to stay the time；PTCA ：Parent ion is 199.99, product ion 182.09, retention time 4.0min.
Preferred by being carried out to MRM tuner parameters, particularly retention time is preferred, may be such that testing result is more smart
Really.
Preferably, flow rate of mobile phase is 0.2ml/min in S2, and sampling volume is 10 μ l.
Preferably, chromatographic column is Waters ACQUITY UPLC HSS T3 in S2,40 DEG C of column temperature.
Preferably, MS conditions are：Xevo TQD systems；Ionization pattern：Electric spray ion source positive ion mode；How anti-MRM is
Answer monitoring pattern；Capillary voltage：3.0kV；Taper hole voltage：30.0 V；Source temperature： 150 ℃；Desolvation temperature：550
℃；Desolventizing gas：1100 L·h-1；Taper hole gas velocity： 50 L·h-1。
Preferably, Pteria penguin melanin sample is prepared by the following procedure method and obtained in S1：
S1：The refinement of Pteria penguin pigment crude product：Pteria penguin outer embrane mantle district's groups are knitted after being processed into homogenate, added
Phosphate buffer containing papain, regulation pH are 7.0 ~ 7.5, are stirred, supernatant is abandoned in enzymolysis, centrifugal filtration, and solvent carries
Take, filter, being dried in vacuo and to obtain pigment crude product；
S2：The oxidation processes of Pteria penguin mother's shellfish pigment：Pigment crude product is taken, adds alkaline solution agent and hydrogen peroxide in 100 ± 5
Oxidation processes are carried out at DEG C, terminating reaction after cooling, adjust pH to 0.8 ~ 1.2, centrifuging and taking supernatant；Organic layer is extracted to obtain, very
Empty dry Pteria penguin mother's shellfish melanin sample.
Pigment during effectively Pteria penguin mother's shellfish outer embrane mantle district's groups can be knitted by enzyme digestion reaction extracts.
Preferably, the amount ratio that phosphate buffer is knitted with mantle district's groups in S1 is organized for 15mL/1g, papain
Mass fraction is more than 2%, and hydrolysis temperature is 50 ~ 60 DEG C, and enzymolysis time is 18 ~ 24h.
Preferably, S2 neutral and alkalis solution is K2CO3, the K2CO3Concentration be 1mol/L；The quality of the hydrogen peroxide point
Number is 3%；Add the Na that mass fraction is 10%2SO3Terminating reaction, the K2CO3, hydrogen peroxide and Na2SO3Knitted with mantle district's groups
Amount ratio be respectively 8.6mL/10mg, 0.8mL/10mg and 0.4mL/10mg.
Compared with prior art, the present invention has the advantages that：
It is provided by the invention grow cultured pearls before change Pteria penguin mother's shellfish color and luster method can obviously reduce Pteria penguin mother shellfish
Melanin content, survival rate is high, and melanin content highest is reduced to 0.08ug/mg in Pteria penguin mother's shellfish outer embrane, reduces
89.5%, it can be applied to cultivate the unusual high quality pearl of color and luster on a large scale, be with a wide range of applications.The present invention provides simultaneously
Pteria penguin mother's shellfish color and luster detection method can Accurate Determining Pteria penguin mother shellfish pigment content, so as to realize pair
The detection and monitoring of Pteria penguin mother's shellfish color and luster.
Brief description of the drawings
Fig. 1 is influence of the various concentrations treated with kojic acid different time to Pteria penguin mother's shellfish survival rate；
Fig. 2 is Pteria penguin mother's shellfish overcoat membranochromic pigments crude product；
Fig. 3 is Pteria penguin mother shellfish pigment P DCA provided by the invention MS collection of illustrative plates；
Fig. 4 is Pteria penguin mother shellfish pigment P TCA provided by the invention MS collection of illustrative plates；
Fig. 5 is melanin content standard curve；
Fig. 6 is Pteria penguin mother's shellfish pigment LC-MS collection of illustrative plates, and wherein A is standard items；B is control group；C is treated with kojic acid group（It is real
Apply example 4）.
Embodiment
The present invention is expanded on further with reference to embodiment.These embodiments are merely to illustrate the present invention rather than limitation
The scope of the present invention.The experimental method of unreceipted actual conditions in lower example embodiment, generally according to this area normal condition or is pressed
The condition suggested according to manufacturer；Used raw material, reagent etc., unless otherwise specified, being can be from the business such as conventional market
The raw material and reagent that approach obtains.The change for any unsubstantiality that those skilled in the art is done on the basis of the present invention
And replace and belong to scope of the present invention.
Embodiment 1 ~ 5 and comparative example 1 ~ 5 change Pteria penguin mother's shellfish color and luster method before growing cultured pearls
The method for changing Pteria penguin mother's shellfish color and luster before what the present embodiment 1 ~ 5 and comparative example 1 ~ 5 provided grow cultured pearls is as follows：By penguin
Dicksoniaceae shellfish is taken out from aquaculture sea area, and it is temporarily foster one week in propagate in pond to be divided into 4 groups, during which feeds chlorella.Processing
A few days ago stop feeding, make its digestion complete.Kojic acid is launched into 4 culturing pools respectively, after each group immersion culture certain time,
It is placed in seawater and continues culture 3 days.Kojic acid concentration and kojic acid incubation time are as follows in embodiment 1 ~ 5 and comparative example 1 ~ 5：
Kojic acid concentration and kojic acid incubation time in the embodiment 1 ~ 5 of table 1 and comparative example 1 ~ 5
The color and luster of Pteria penguin mother shellfish determines with survival rate
After method processing of the Pteria penguin mother shellfish through being provided in embodiment 1 ~ 5 and comparative example 1 ~ 5, pigment content measure is carried out,
Its result is as shown in table 2：1g/L treated with kojic acid 18h（Such as embodiment 1）Pteria penguin mother's shellfish pigment synthesis can be generated
Obvious inhibitory action, melanin content is from the near 0.67 μ g/mg of 0.78 μ g/mg in outer embrane；When kojic acid concentration be 10g/L and
During 100g/L, melanin content can be all significantly inhibited when acting on 6h, 12h and 18h, when its inhibition is with kojic acid concentration and effect
Between increase and increase.Wherein, 10g/L treated with kojic acid 18h and 100g/L treated with kojic acid 18h inhibition is best, is respectively
0.08 μ g/mg and 0.02 μ g/mg.This explanation kojic acid is a kind of effective Pteria penguin mother's shellfish melanin inhibitor.
In order to determine feasibility of the treated with kojic acid during Pteria penguin mother Bei Yuzhu, we analyze kojic acid to enterprise
The influence of goose Dicksoniaceae shellfish survival rate.As a result such as table 2 and Fig. 1 are shown, after 1 ~ 10g/L treated with kojic acid 6h, 12h and 18h,
The survival rate of Pteria penguin mother shellfish is not all significantly affected；When kojic acid concentration is 100g/L, processing 6h is not detected by
Survival rate is had a significant effect, but the survival rate of Pteria penguin is remarkably decreased after processing 12h and 18h.Which imply that 1 ~ 10g/
L kojic acid is a kind of safely and effectively pigment inhibitor to Pteria penguin mother shellfish, precious to penguin when kojic acid concentration is 100g/L
Certain toxicity be present in pearl shellfish.It is thus determined that optium concentration and time that kojic acid uses are that 10g/L handles 18h.
After the Pteria penguin mother shellfish laboratory handled more than is rested 1 day, put back to marine site and rest 3 days or so, can carry out
Plant core and grow cultured pearls.This method for changing Pteria penguin mother's shellfish color and luster before core is planted, can be applied to cultivate color and luster on a large scale strange
Different high quality pearl, is with a wide range of applications.
The Pteria penguin mother shellfish pigment content and survival rate measurement result of the embodiment 1 ~ 5 of table 2 and comparative example 1 ~ 4
The detection method of the Pteria penguin mother's shellfish color and luster of embodiment 6
The color and luster for the Pteria penguin mother shellfish that the method provided embodiment 1 ~ 5 and comparative example 1 ~ 5 obtains detects, specific behaviour
Make as follows：
First, the refinement of Pteria penguin mother shellfish pigment crude product：
（1）5 individuals are randomly selected respectively, the mantle area of clip outer embrane, it is female to weigh appropriate Pteria penguin before and after the processing
Shellfish outer embrane mantle district's groups are knitted, and are homogenized using multi-functional homogenizer.
（2）The phosphate buffer that 15mL/1g organizes pH7.0- pH 7.5 is added, wherein containing 2% papain（Quality
Fraction）, 50 ~ 60 DEG C of 18 ~ 24h of stirring enzymolysis on magnetic stirring apparatus.
（3）After enzymolysis, 10000rpm centrifugation 10min, supernatant is abandoned.
（4）Add petroleum ether（2ml/1g is organized）Suspend precipitation, 10000rpm centrifugation 10min, abandons supernatant, is repeated 3 times.
（5）Add absolute ethyl alcohol（2ml/1g is organized）Suspend precipitation, 10000rpm centrifugation 10min, abandons supernatant, repeats 3
It is secondary.
（6）Add distilled water（2ml/1g is organized）Suspend precipitation, 10000rpm centrifugation 10min, abandons supernatant, is repeated 3 times,
（7）Deposit vacuum drying after will be washed can obtain pigment crude product（See Fig. 2）.
2nd, the oxidation processes of Pteria penguin mother shellfish pigment
（1）Precision weighs appropriate melanin crude product, and every 10 mg melanin crude product is according to addition volumes below various reagents.Add
8.6 mL 1mol/L K2CO3With the H of 0.8 mL 3%2O2Oxidation processes are carried out, 100 DEG C of water-baths are incubated 20 ~ 30min.
（2）Flowing water is cooled to normal temperature, adds the Na of 0.4 mL 10%2SO3Terminating reaction.
（3）Add the mol/L HCl of 5 mL 6 regulations pH to 0.8-1.2.
（4）9500 centrifugation 10min, take supernatant.
（5）The extraction of 70 mL absolute ethers is added into supernatant, upper organic layer is collected, is repeated 2 times.
（6）Merge all organic phases, vacuumize after drying with 50% methanol aqueous solution（V/V）Redissolve, 0.45 μm of organic film
Filtering, prepare injection LC-MS analyses.
3rd, test sample injection LC-MS is subjected to analysis detection
（1）UPLC conditions
Sample after oxidation is detected with LC-MS, and liquid phase chromatogram condition is：Chromatographic column： Waters ACQUITY UPLC HSS T3
(2.1 x 50 mm, 1.7 μm), 40 DEG C of column temperature.Mobile phase A is 0.1% aqueous formic acid（V/V）, Mobile phase B is 0.1% formic acid
Methanol solution（V/V）.Elution program is 0 ~ 5.9min, and mobile phase A and B ratio are 90%:10%；6.0 ~ 8min, Mobile phase B
Ratio is 100%；8.1 ~ 12min, mobile phase A and B ratio revert to 90%:10% （It is shown in Table 3）.Flow velocity is 0.2ml/min,
Sampling volume is 10 μ l.
The chromatographic column gradient elution program of table 3
（2）MS conditions
Xevo TQD systems；Ionization pattern： ESI+（Electric spray ion source positive ion mode）；MRM multiple-reaction monitoring patterns；
Capillary voltage：3.0kV；Taper hole voltage：30.0 V；Source temperature： 150℃；Desolvation temperature：550℃；Desolventizing gas：
1100 L/h；Taper hole gas velocity： 50 L/h.
（3） LC-MS/MS
Select and optimize the MRM ion pairs of analyte, such as table 4.
The MRM tuner parameters and retention time of the analyte of table 4
4th, Pteria penguin mother shellfish pigment content determines
（1）The qualitative analysis of Pteria penguin mother's shellfish pigment
The Mass Spectrometer Method result of Pteria penguin mother shellfish after the method processing that example 1 provides by contrast is as shown in Figure 3：m/z：[M-
H]-Quasi-molecular ion peak is 155.97, with known PDCA (C6H5O4N1) relative molecular mass 155 is consistent, it can determine whether the peak group
It is divided into PDCA；As shown in figure 4, m/z：[M-H]-Quasi-molecular ion peak is 199.92, with known PTCA (C7H5O6N1) relative point
Protonatomic mass 197 is consistent, and can determine whether the peak component is PTCA.
（2）The quantitative analysis of Pteria penguin mother's shellfish pigment
Melanin standard product concentration gradient is set, prepares the standard curve of melanin content and LC-MS peak areas（See Fig. 5 and figure
6）, calculate the pigment content that each embodiment and comparative example determine（Melanin quality μ g/overcoat film quality mg）.
Pigment content is obtained by this detection method after the method processing in each embodiment and comparative example in table 2.
Claims (10)
1. it is a kind of grow cultured pearls before change Pteria penguin mother's shellfish color and luster method, it is characterised in that methods described comprises the following steps：
S1：Pteria penguin mother shellfish is taken out from aquaculture sea area, is put in artificial farming pond and temporarily supports, feed food；
S2：Pause, which is fed, makes it that food digestion is complete；
S3：Kojic acid, culture are launched to artificial farming pond；Kojic acid concentration is 1 ~ 10g/L in the artificial farming pond, and incubation time is
Kojic acid concentration is 10g/L in 18h, or the artificial farming pond, and incubation time is kojic acid in 6 ~ 18h, or the artificial farming pond
Concentration is 10 ~ 100g/L, cultivates 6h；
S4：Pteria penguin mother shellfish is placed in seawater after resting can be used to plant core grow cultured pearls.
2. change the method for Pteria penguin mother's shellfish color and luster before growing cultured pearls according to claim 1, it is characterised in that fed in S1
Food be chlorella, temporarily the foster time is more than 1 week.
3. change the method for Pteria penguin mother's shellfish color and luster before growing cultured pearls according to claim 1, it is characterised in that suspend in S2
2 days of the time fed.
4. change the method for Pteria penguin mother's shellfish color and luster before growing cultured pearls according to claim 1, it is characterised in that described artificial
Kojic acid concentration is 10g/L, incubation time 18h in culturing pool.
A kind of 5. detection method of any Pteria penguin mother shellfish color and luster of claim 1 ~ 4, it is characterised in that methods described
Comprise the following steps：
S1：Take Pteria penguin mother's shellfish melanin sample to be dissolved with methanol aqueous solution, test sample is obtained after filtering；
S2：Test sample injection LC-MS is subjected to analysis detection；Wherein, mobile phase A is that the formic acid that volume fraction is 0.1% is water-soluble
Liquid, Mobile phase B are the formic acid methanol solution that volume fraction is 0.1%；Elution program is 0 ~ 6min, mobile phase A and B ratio
For 90%:10%；6 ~ 8min, the ratio of Mobile phase B is 100%；8 ~ 12min, mobile phase A and B ratio are 90%:10%.
6. the detection method of Pteria penguin mother shellfish color and luster according to claim 5, it is characterised in that set MRM to adjust in S2
Humorous parameter is：PDCA：Parent ion is 155.98, product ion 138.01, retention time 2.9min；PTCA ：Parent ion is
199.99, product ion 182.09, retention time 4.0min.
7. the detection method of Pteria penguin mother shellfish color and luster according to claim 5, it is characterised in that flow rate of mobile phase in S2
For 0.2ml/min, sampling volume is 10 μ l.
8. the detection method of Pteria penguin mother shellfish color and luster according to claim 5, it is characterised in that chromatographic column is in S2
Waters ACQUITY UPLC HSS T3,40 DEG C of column temperature.
9. the detection method of Pteria penguin mother shellfish color and luster according to claim 5, it is characterised in that MS conditions are in S2：
Xevo TQD systems；Ionization pattern：Electric spray ion source positive ion mode；MRM multiple-reaction monitoring patterns；Capillary voltage：
3.0kV；Taper hole voltage：30.0V；Source temperature： 150℃；Desolvation temperature：550℃；Desolventizing gas：1100 L·h-1；Cone
Gas flow hole speed： 50 L·h-1。
10. the detection method of Pteria penguin mother shellfish color and luster according to claim 5, it is characterised in that penguin pearl in S1
Fritillaria shellfish melanin sample is prepared by the following procedure method and obtained：
S1：The refinement of Pteria penguin mother's shellfish pigment crude product：Pteria penguin mother's shellfish outer embrane mantle district's groups are knitted be processed into it is even
After slurry, the phosphate buffer containing papain is added, regulation pH is 7.0 ~ 7.5, is stirred, supernatant is abandoned in enzymolysis, centrifugal filtration
Liquid, solvent extraction, filtering, it is dried in vacuo to obtain pigment crude product；
S2：The oxidation processes of Pteria penguin mother's shellfish pigment：Pigment crude product is taken, adds alkaline solution and hydrogen peroxide in 100 ± 5 DEG C
Lower carry out oxidation processes, terminating reaction after cooling, adjust pH to 0.8 ~ 1.2, centrifuging and taking supernatant；Extract to obtain organic layer, vacuum
Dry Pteria penguin mother's shellfish melanin sample.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101057927A (en) * | 2006-04-17 | 2007-10-24 | 吴英萍 | Medicinal composition and its preparation method and quality control method |
| CN101776659A (en) * | 2010-02-03 | 2010-07-14 | 北京市理化分析测试中心 | Method for detecting anthocyanin in red radish through high-performance liquid chromatography |
| CN102757664A (en) * | 2012-07-03 | 2012-10-31 | 福建农林大学 | Extracting method of passion flower fruit pigment |
-
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101057927A (en) * | 2006-04-17 | 2007-10-24 | 吴英萍 | Medicinal composition and its preparation method and quality control method |
| CN101776659A (en) * | 2010-02-03 | 2010-07-14 | 北京市理化分析测试中心 | Method for detecting anthocyanin in red radish through high-performance liquid chromatography |
| CN102757664A (en) * | 2012-07-03 | 2012-10-31 | 福建农林大学 | Extracting method of passion flower fruit pigment |
Non-Patent Citations (2)
| Title |
|---|
| 于非非等: "熊果苷和丙戊酸影响企鹅珍珠贝贝壳色泽", 《基因组学与应用生物学》 * |
| 孙亚真等: "乌骨鸡黑色素的高校液相色谱法测定", 《分析测试学报》 * |
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