CN108613939A - A kind of detection method of small peptide micro-element chelate rate - Google Patents
A kind of detection method of small peptide micro-element chelate rate Download PDFInfo
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- CN108613939A CN108613939A CN201810811488.4A CN201810811488A CN108613939A CN 108613939 A CN108613939 A CN 108613939A CN 201810811488 A CN201810811488 A CN 201810811488A CN 108613939 A CN108613939 A CN 108613939A
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- small peptide
- chelate
- value range
- extraction
- zinc
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- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 130
- 239000013522 chelant Substances 0.000 title claims abstract description 91
- 238000001514 detection method Methods 0.000 title claims abstract description 24
- 239000010949 copper Substances 0.000 claims abstract description 69
- 229910052802 copper Inorganic materials 0.000 claims abstract description 64
- 239000011701 zinc Substances 0.000 claims abstract description 64
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 claims abstract description 61
- 230000009920 chelation Effects 0.000 claims abstract description 59
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 54
- 229910052725 zinc Inorganic materials 0.000 claims abstract description 47
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 claims abstract description 46
- 238000000034 method Methods 0.000 claims abstract description 38
- 238000000605 extraction Methods 0.000 claims abstract description 33
- 239000002904 solvent Substances 0.000 claims abstract description 20
- 238000003808 methanol extraction Methods 0.000 claims abstract description 8
- 239000006228 supernatant Substances 0.000 claims description 25
- 238000010521 absorption reaction Methods 0.000 claims description 21
- 239000011573 trace mineral Substances 0.000 claims description 10
- 235000013619 trace mineral Nutrition 0.000 claims description 10
- 238000000703 high-speed centrifugation Methods 0.000 claims description 4
- 229910021645 metal ion Inorganic materials 0.000 abstract description 45
- 238000005259 measurement Methods 0.000 abstract description 17
- 238000012360 testing method Methods 0.000 abstract description 17
- 238000004519 manufacturing process Methods 0.000 abstract description 5
- 238000007781 pre-processing Methods 0.000 abstract description 2
- 239000007788 liquid Substances 0.000 description 23
- 239000000523 sample Substances 0.000 description 23
- 239000000499 gel Substances 0.000 description 22
- 239000003480 eluent Substances 0.000 description 17
- 238000001641 gel filtration chromatography Methods 0.000 description 13
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 12
- 238000003556 assay Methods 0.000 description 12
- 238000011084 recovery Methods 0.000 description 12
- 238000010828 elution Methods 0.000 description 10
- 102000004196 processed proteins & peptides Human genes 0.000 description 10
- 229910052751 metal Inorganic materials 0.000 description 9
- 238000002474 experimental method Methods 0.000 description 8
- 239000002184 metal Substances 0.000 description 8
- 238000001556 precipitation Methods 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 229920005654 Sephadex Polymers 0.000 description 7
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 7
- 239000003960 organic solvent Substances 0.000 description 7
- 239000003513 alkali Substances 0.000 description 6
- 150000001413 amino acids Chemical class 0.000 description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-M hydroxide Chemical compound [OH-] XLYOFNOQVPJJNP-UHFFFAOYSA-M 0.000 description 6
- 229910052742 iron Inorganic materials 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 239000000243 solution Substances 0.000 description 6
- 239000012507 Sephadex™ Substances 0.000 description 5
- PTFCDOFLOPIGGS-UHFFFAOYSA-N Zinc dication Chemical compound [Zn+2] PTFCDOFLOPIGGS-UHFFFAOYSA-N 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 239000000725 suspension Substances 0.000 description 5
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 4
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 4
- 239000004473 Threonine Substances 0.000 description 4
- 230000002378 acidificating effect Effects 0.000 description 4
- 238000004587 chromatography analysis Methods 0.000 description 4
- TVZPLCNGKSPOJA-UHFFFAOYSA-N copper zinc Chemical compound [Cu].[Zn] TVZPLCNGKSPOJA-UHFFFAOYSA-N 0.000 description 4
- 150000002500 ions Chemical class 0.000 description 4
- 238000011068 loading method Methods 0.000 description 4
- 229930182817 methionine Natural products 0.000 description 4
- 239000012086 standard solution Substances 0.000 description 4
- 229910021511 zinc hydroxide Inorganic materials 0.000 description 4
- 239000000654 additive Substances 0.000 description 3
- 230000000996 additive effect Effects 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 239000008367 deionised water Substances 0.000 description 3
- 229910021641 deionized water Inorganic materials 0.000 description 3
- 238000010494 dissociation reaction Methods 0.000 description 3
- 230000005593 dissociations Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 239000011521 glass Substances 0.000 description 3
- 229920001184 polypeptide Polymers 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- UGZADUVQMDAIAO-UHFFFAOYSA-L zinc hydroxide Chemical compound [OH-].[OH-].[Zn+2] UGZADUVQMDAIAO-UHFFFAOYSA-L 0.000 description 3
- 229940007718 zinc hydroxide Drugs 0.000 description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 2
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 229910052791 calcium Inorganic materials 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 229910000366 copper(II) sulfate Inorganic materials 0.000 description 2
- 239000007863 gel particle Substances 0.000 description 2
- 239000003446 ligand Substances 0.000 description 2
- 229910052749 magnesium Inorganic materials 0.000 description 2
- 239000011777 magnesium Substances 0.000 description 2
- 239000001301 oxygen Substances 0.000 description 2
- 229910052760 oxygen Inorganic materials 0.000 description 2
- 229910052700 potassium Inorganic materials 0.000 description 2
- 239000011591 potassium Substances 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 238000004062 sedimentation Methods 0.000 description 2
- 229910052708 sodium Inorganic materials 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- 230000008961 swelling Effects 0.000 description 2
- 239000003643 water by type Substances 0.000 description 2
- 229910000368 zinc sulfate Inorganic materials 0.000 description 2
- 239000011686 zinc sulphate Substances 0.000 description 2
- KFSJYZYQSZKRRQ-BYPYZUCNSA-N (2s)-2-(hydroxyamino)-4-methylsulfanylbutanoic acid Chemical compound CSCC[C@H](NO)C(O)=O KFSJYZYQSZKRRQ-BYPYZUCNSA-N 0.000 description 1
- 241000040710 Chela Species 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000003674 animal food additive Substances 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- 238000004380 ashing Methods 0.000 description 1
- 125000004429 atom Chemical group 0.000 description 1
- CREXVNNSNOKDHW-UHFFFAOYSA-N azaniumylideneazanide Chemical group N[N] CREXVNNSNOKDHW-UHFFFAOYSA-N 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000010835 comparative analysis Methods 0.000 description 1
- AEJIMXVJZFYIHN-UHFFFAOYSA-N copper;dihydrate Chemical compound O.O.[Cu] AEJIMXVJZFYIHN-UHFFFAOYSA-N 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000007071 enzymatic hydrolysis Effects 0.000 description 1
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 1
- 235000019441 ethanol Nutrition 0.000 description 1
- 210000003608 fece Anatomy 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000005227 gel permeation chromatography Methods 0.000 description 1
- 238000002523 gelfiltration Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 239000008236 heating water Substances 0.000 description 1
- 229910001385 heavy metal Inorganic materials 0.000 description 1
- 150000003840 hydrochlorides Chemical class 0.000 description 1
- ZHUXMBYIONRQQX-UHFFFAOYSA-N hydroxidodioxidocarbon(.) Chemical group [O]C(O)=O ZHUXMBYIONRQQX-UHFFFAOYSA-N 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- 235000000396 iron Nutrition 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910017604 nitric acid Inorganic materials 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 239000003895 organic fertilizer Substances 0.000 description 1
- 230000010355 oscillation Effects 0.000 description 1
- 244000144977 poultry Species 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000013441 quality evaluation Methods 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 239000013049 sediment Substances 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
- G01N21/3103—Atomic absorption analysis
Landscapes
- Physics & Mathematics (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The present invention relates to a kind of detection methods of small peptide micro-element chelate rate.The detection method detaches free metal ion and Chelating state metal ion by methanol extraction method, provide best solvents ratio, extraction temperature and extraction time, by the combination of three's preferred parameter, realizing reduces the meltage of small peptide copper chelate in methyl alcohol to improve accuracy in detection.The present invention it is easy to operate, need not to product pre-processing, quickly, testing result is accurate, be suitable for small peptide copper chelate and small peptide chelates of zinc chelation percent measurement, be satisfied with production in quick and precisely detect needs.
Description
Technical field
The present invention relates to Element detection fields, and in particular to a kind of detection side of small peptide micro-element chelate rate
Method.
Background technology
Small peptide microelement chelate is with small peptide peptide bond amino nitrogen atom and carboxyl oxygen atom by metal ion by matching
Position bond, which is closed, forms the chelate with cyclic structure, is another novel organic micro after Chelates of Amino Acids And Trace Elements
Element additive.With the inorganics such as other hydrochlorides, sulfate trace element or organic acid trace element compared with, on the one hand it
Have the characteristics that absorptivity is high, biology utilization rate is high, the stable structure in gastro-intestinal tract；Low-level in another aspect feed
Additive amount has the effects that growth promotion, improves intestinal environment, improves that body is anti-oxidant and immune function.Therefore it is added in feed low
Horizontal small peptide microelement chelate can not only play growth-promoting effect, but also can reduce the content of copper zinc in feces of livestock and poultry, keep away
Exempt from the heavy metal pollution of soil problem caused by the high zinc of organic fertilizer high-copper, is a kind of copper zinc source additive having a extensive future.
In recent years small peptide microelement chelate as a kind of inorganic high zinc source additive substitute of high-copper in feed
Using more and more common.Often using chelation percent as the quality evaluation index of this substance, small peptide microelement chelate in practice
Chelation percent, i.e., the ratio of metallic element total content is accounted for the Chelating state metal element content that small peptide is combined by coordinate bond.But
The detection method that yet there are no small peptide micro-element chelate rate refers to Chelates of Amino Acids And Trace Elements more in practice
The assay method of chelation percent common are the methods of dextran gel filtration chromatography and organic solvent extractionprocess.
Have the method for the amino acid trace element chelated rate of gel filtration chromatography,《Feed addictive methionine iron
The measurement gel filtration chromatography of (copper, manganese, zinc) chelation percent》(GB/T 13080.2-2005) is surveyed using gel filtration chromatography
Fixed amino acid trace element chelated rate, but occur the problems such as same sample measurement result is unstable in its practical application.First, exist
It needs to elute Chelating state metal ion using different alkaline eluants in chromatographic column elution separation process, to reach point
Purpose from Chelating state and non-Chelating state metal ion, but since the difference such as product composition and its production technology is so as to cause it
It is different to chelate intensity, part Chelating state metal ion is dissociated into free state in the case where alkalinity acts on, and leads to the inaccuracy of measurement result
Property.Hu Xiaobo (2009) is shown using Preliminary Analysis Results when gel filtration chromatography threonine zinc chelation percent：Threonine
The stability of zinc is smaller than the stability of zinc hydroxide, and when gel chromatography sample introduction is the alkaline environment of pH9.0, soluble threonine zinc
It is changed into zinc hydroxide precipitation, soluble threonine zinc cannot be separated with free zinc ion, cannot get the standard of chelation percent
Really value, secondly entire detection process is complicated for operation, and time-consuming for experiment, takes around 4~5 hours, is not suitable in actual production.
Solubility very little of the Chelating state metal ion in the organic solvents such as methanol, ethyl alcohol, and non-Chelating state metal ion
It is soluble in organic solvent, therefore principle can use organic solvent extractionprocess to measure chelation percent accordingly.Yi Kai (2007) and week
The experiment for building group (2007) et al. is shown when the method measures amino acid micro-element chelate rate with preferable essence
Exactness.But Chelating state metal ion still has certain solubility in organic solvent, the accuracy measured to result has one
Fixed influence.Therefore how to reduce influence of the Chelating state metal ion solubility for measurement result is an important problem.
It is more and more for the demand of small peptide microelement chelate currently on the market, but product quality is very different,
The content of Chelating state metal ion is very low in many products.The chelation percent of small peptide microelement chelate is to reflect such product product
The characteristics index of matter, therefore the detection method for establishing small peptide micro-element chelate rate is of great significance.
Invention content
The present invention exists for existing small peptide micro-element chelate rate detection method Small Peptides microelement chelate
Organic solvent still has the inaccuracy being partly dissolved so as to cause measurement result when extracting, and it is trace element chelated to provide a kind of small peptide
The detection method of object chelation percent, using this method measurement result accuracy higher, and detection process is easy to operate, easy to spread to answer
With.
To achieve the above object, the detection method of small peptide micro-element chelate rate provided by the invention includes following
Step：
Step 1：It is measured using atomic absorption spectrophotometer method micro- in small peptide microelement chelate sample
Content is W；
Step 2：The another small peptide microelement chelate to be measured for taking quality to be m, which is dissolved in the methanol that volume is V, to be extracted
It takes, is shaken in the case where extraction temperature is the constant temperature of T, after extraction time t1, by its high speed centrifugation t2 minutes, take supernatant
It is C to measure its free microelement concentration using atomic absorption spectrophotometer method；
Step 3：The chelation percent measured under different extraction conditions is calculated by following formula：
In formula：
X：Small peptide micro-element chelate rate, unit %
W：Mass percent in small peptide microelement chelate sample shared by trace element, unit %
m：The another example weight for taking a small amount of small peptide microelement chelate for methanol extraction, unit mg
C：Microelement concentration in supernatant, unit mg/L
V：The volume of extract liquor, unit L
Wherein, the value range of the quality m of the small peptide microelement chelate to be measured described in step 2 be 25~
1200mg；The variation range of the extraction temperature T is 0~50 DEG C；The value range of the extraction time t1 be 5~
90min；The value range of the ultracentrifugal centrifugal speed is 12000r/min；The high speed centrifugation time t2 is
15min；The volume V of the extract liquor is 3 × 10-3L, solvents than value range 25:3~1200:3mg/mL.
Further, the detection method of small peptide micro-element chelate rate of the present invention, is particularly suitable for small peptide
Copper chelate and/or small peptide chelates of zinc.
Particularly, when measuring small peptide copper chelate chelation percent, copper content is in 13%~16% range in small peptide copper chelate
When, the value range of the m is 600~1000mg, and the value range of the extraction temperature T is 2~15 DEG C, the extraction
The value range of time t1 be 10~40min, solvents than value range 600:3~1000:3mg/mL.
Particularly, when measuring small peptide chelates of zinc chelation percent, Zn content is in 14%~17% range in small peptide chelates of zinc
When, the value range of the m is 550-800mg, and the value range of the extraction temperature T is 15~25 DEG C, the extraction
The value range of time t1 be 15-40min, solvents than value range 550:3~800:3mg/mL.
The present invention is soluble in methanol organic solvent according to free metal ion under normal circumstances, and Chelating state metal ion is not
It is soluble in the characteristic of methanol, free metal ion and Chelating state metal ion, free state after centrifugation are detached with methanol extraction method
Metal ion is present in supernatant, and Chelating state metal ion is present in precipitation.But routinely methanol extraction method, Chelating state gold
Belong to ion still has certain solubility, Chelating state metal ion and free metal ion to be not completely separated out in methyl alcohol.
This Chelating state metal ion being partially dissolved in methanol affects the accuracy of chelation percent detection.By studying and experiment table
Bright, extraction temperature, extraction time and solvents compare the dissolubility of Chelating state metal ion in methanol extraction method extraction process
Have a great impact, The present invention gives best solvents ratio, extraction temperature and extraction times, pass through three's preferred parameter
Combination, realizing reduces the meltage of small peptide copper chelate in methyl alcohol to improve accuracy in detection.
The detection method of small peptide micro-element chelate rate provided by the invention, has the following advantages that：
(1) The present invention gives best solvents ratio, extraction temperature and extraction time, the groups of this three's preferred parameter
Conjunction can effectively reduce the meltage of small peptide copper chelate in methyl alcohol to improve accuracy in detection.
(2) present invention it is easy to operate, need not to product pre-processing, quickly, testing result is accurate, be suitable for small peptide copper
The measurement of chelate and small peptide chelates of zinc chelation percent is satisfied in production and quick and precisely detects needs.
Description of the drawings
Fig. 1 is the influence that solvents compare small peptide copper chelate chelation percent testing result；
Fig. 2 is influence of the extraction temperature to small peptide copper chelate chelation percent testing result；
Fig. 3 is influence of the extraction time to small peptide copper chelate chelation percent testing result；
Fig. 4 is the influence that solvents compare small peptide chelates of zinc chelation percent testing result；
Fig. 5 is influence of the extraction temperature to small peptide chelates of zinc chelation percent testing result；
Fig. 6 is influence of the extraction time to small peptide chelates of zinc chelation percent testing result.
Specific implementation mode
With reference to specific embodiment, the present invention is described in detail.Following embodiment will be helpful to the technology of this field
Personnel further understand the present invention, but the invention is not limited in any way.It should be pointed out that the ordinary skill of this field
For personnel, without departing from the inventive concept of the premise, it can also make certain adjustments and improvements.These belong to the present invention
Protection domain.
Embodiment 1
The chelation percent assay method of small peptide copper chelate：
Copper content in small peptide copper chelate sample is measured with atomic absorption spectrophotometer method；Separately take 800mg small peptides to be measured
Copper chelate is added in the methanol that 3mL temperature is 5 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 10min,
Supernatant is taken, the free copper content in supernatant is measured, copper content testing uses atomic absorption spectrophotometer, solvents ratio
It is 800:3 mg/mL.
It is 94.97% to be calculated according to formula and obtain the chelation percent of small peptide copper chelate.
Embodiment 2
The chelation percent assay method of small peptide chelates of zinc：
Zn content in small peptide chelates of zinc sample is measured with atomic absorption spectrophotometer method；Separately take 600mg small peptides to be measured
Chelates of zinc is added in the methanol that 3mL temperature is 20 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 30min,
Supernatant is taken to measure the free Zn content in supernatant, zinc-content determination uses atomic absorption spectrophotometer, solvents ratio
It is 600:3 mg/mL.
It is 99.65% to be calculated according to formula and obtain the chelation percent of small peptide chelates of zinc.
Embodiment 3
The chelation percent assay method of small peptide copper chelate：
Copper content in small peptide copper chelate sample is measured with atomic absorption spectrophotometer method；Separately take 600mg small peptides to be measured
Copper chelate is added in the methanol that 3mL temperature is 2 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 30min,
Supernatant is taken, the free copper content in supernatant is measured, copper content testing uses atomic absorption spectrophotometer, solvents ratio
It is 600:3 mg/mL.
It is 90% to be calculated according to formula and obtain the chelation percent of small peptide copper chelate.
Embodiment 4
The chelation percent assay method of small peptide chelates of zinc：
Zn content in small peptide chelates of zinc sample is measured with atomic absorption spectrophotometer method；Separately take 550mg small peptides to be measured
Chelates of zinc is added in the methanol that 3mL temperature is 15 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 15min,
Supernatant is taken to measure the free Zn content in supernatant, zinc-content determination uses atomic absorption spectrophotometer, solvents ratio
It is 550:3 mg/mL.
It is 99.32% to be calculated according to formula and obtain the chelation percent of small peptide chelates of zinc
Embodiment 5
The chelation percent assay method of small peptide copper chelate：
Copper content in small peptide copper chelate sample is measured with atomic absorption spectrophotometer method；Separately take 1000mg to be measured small
Peptide copper chelate is added in the methanol that 3mL temperature is 15 DEG C, and 12000r/min is centrifuged after constant temperature oscillator vibrates 40min
15min takes supernatant, measures the free copper content in supernatant, and copper content testing uses atomic absorption spectrophotometer, solute
Solvent ratio is 1000:3mg/mL.
It is 94.96% to be calculated according to formula and obtain the chelation percent of small peptide copper chelate
Embodiment 6
The chelation percent assay method of small peptide chelates of zinc：
Zn content in small peptide chelates of zinc sample is measured with atomic absorption spectrophotometer method；Separately take 800mg small peptides to be measured
Chelates of zinc is added in the methanol that 3mL temperature is 25 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 40min,
Supernatant is taken to measure the free Zn content in supernatant, zinc-content determination uses atomic absorption spectrophotometer, solvents ratio
It is 800:3 mg/mL.
It is 99.65% to be calculated according to formula and obtain the chelation percent of small peptide chelates of zinc
Embodiment 7
The chelation percent assay method of small peptide copper chelate：
Copper content in small peptide copper chelate sample is measured with atomic absorption spectrophotometer method；Separately take 25mg small peptides to be measured
Copper chelate is added in the methanol that 3mL temperature is 0 DEG C, and 12000r/min centrifuges 15min after constant temperature oscillator vibrates 90min,
Supernatant is taken, the free copper content in supernatant is measured, copper content testing uses atomic absorption spectrophotometer, solvents ratio
It is 25:3 mg/mL.
It is 61.20% to be calculated according to formula and obtain the chelation percent of small peptide copper chelate
Embodiment 8
The chelation percent assay method of small peptide chelates of zinc：
Zn content in small peptide chelates of zinc sample is measured with atomic absorption spectrophotometer method；Separately take 1200mg to be measured small
Peptide chelates of zinc is added in the methanol that 3mL temperature is 50 DEG C, and 12000r/min is centrifuged after constant temperature oscillator vibrates 5min
15min, takes supernatant to measure the free Zn content in supernatant, and zinc-content determination uses atomic absorption spectrophotometer, solute
Solvent ratio is 1200:3 mg/mL.
It is 99.60% to be calculated according to formula and obtain the chelation percent of small peptide chelates of zinc.
Embodiment 9
《Measurement --- the gel filtration chromatography of feed addictive methionine iron (copper, manganese, zinc) chelation percent》(GB/T
13885-2003) it is unique country standard about amino-acid metal chelate, the present embodiment is mainly according to above-mentioned national standard
Using gel filtration chromatography prove this method measure small peptide metallo-chelate chelation percent feasibility, with it is using the present invention
Method measurement result is compared.
According to GB/T 13885-2003, assay method includes the following steps：
1. the preparation of gel chromatographic columns
According to《The measurement gel filtration chromatography of feed addictive methionine iron (copper, manganese, zinc) chelation percent》(GB/T
13080.2-2005) method choice " room temperature swelling method " is recommended to activate G-15 gels.Weigh 15g or so sephadexes
G-15 is added in 150mL ultra-pure waters, is configured to appropriate sticky suspension, is stored at room temperature, gently with glass bar per the intervals 5h
Stirring makes suspension be uniformly mixed, and settles 20min, until gel suspension volume is 60ml (gel G-15 swelling volumes：
2.5-3.5ml/g gels dry powder) left and right i.e. illustrate that gel has activated.It is carefully poured out after gel activation and swims in the broken of the water surface
Broken gel particle and impurity prevent from blocking gel column, and water is added to stir into suspension, and ultrasonic vibration excludes in gel after air
Gel suspension is once poured into glass chromatography column along glass bar, chromatographic column effluent switch is opened, liquid is made slowly to flow out,
Make gel slowly sedimentation naturally.Gel section length is controlled in 20cm or so.With about 50~80mL (3~4 times of column journey volumes)
The eluent balanced gel chromatographic column of pH7.0.
2. sample pretreatment
Small peptide copper chelate (small peptide chelates of zinc) is weighed using 50ml centrifuge tubes (preventing from waiting for test sample and pan paper adhesion)
Sample to be tested about 0.5g, accurate recording weigh weight M1, are accurate to 0.0001 g, and 15mL deionized water dissolvings are added, and vortex is mixed
After even device mixing 3min and sonic oscillation 5min in 37 DEG C (temperature should not be too high, prevents destruction of the high temperature to chelate structure,
Influence the accuracy of chelation percent measurement result) heating water bath 30min, during which per 5min, stirring once makes sample fully dissolve, and fills
12000r/min centrifuges 20min after dividing dissolving, is carefully moved supernatant in people's volumetric flask (specification 25mL) with pipette, precipitates
Centrifugation 3 times is washed with 3mL deionized waters (37 DEG C) respectively, supernatant all moves in people's 25mL volumetric flasks, and constant volume obtains solubility
Small peptide copper chelate (small peptide chelates of zinc) and free copper (zinc) solion, referred to as A liquid.
Small peptide copper chelate to be measured (small peptide chelates of zinc) 100mg or so is weighed, it is M2 that accurate recording, which weighs weight, accurately
To 0.0001g, so that sample is carbonized (temperature should not be too high, prevents sample from burning) completely on electric hot plate using crucible, then will
It is transferred to 3~4h of ashing in 550 DEG C of muffle furnaces, is dissolved with 10mL nitric acid after being cooled to room temperature, 120 DEG C of heater plate 1h make Cu
(Zn) all dissolving exists in the form of an ion, transfers it to 50mL volumetric flasks, and deionized water washs crucible 3-4 times, will
Solution is transferred completely into volumetric flask, is cooled to room temperature, deionized water constant volume, referred to as B liquid.
3. loading elution separation
It opens chromatographic column bottom piston gel surface is made in chromatographic column to be bordering on and drain off that (stringent control switch, prevents gel table
Face dried liquid stream), the eluent of closure piston, outflow is collected with 100ml volumetric flasks；200 μm of solution are accurately drawn with liquid-transfering gun
A keeps pipette tips apart from the positions gel surface 1mm, is slowly loaded, solution A is made uniformly to penetrate into gel surface.Then into chromatographic column
A small amount of alkaline eluant is added and carefully washs post jamb and gel surface, opens bottom piston eluent is made to be bordering on and drain off, it is at this moment molten
Liquid A all penetrates into gel surface, closure piston.The elution of 100ml alkaline eluants, eluent flow rate control are continuously added later
In 6~8 drops/min, about 100mL eluents, 100mL volumetric flask constant volumes are collected, this solution is known as C liquid.Wait for that alkaline eluant is received
Addition pH 2.0HCl acidic effluent liquid 100mL in backward chromatographic column are collected, have collected eluent about 100mL, 100mL volumetric flasks are fixed
Hold, this solution is known as D liquid.Chromatographic column is both needed to the alkaline eluant balanced gel column with corresponding pH value before each loading, until
The eluent ph values of outflow are equal with alkaline eluant.
4. metal ion content measures
According to《The measurement atomic absorption spectrography (AAS) of calcium, copper, iron, magnesium, manganese, potassium, sodium and Zn content in animal feed》(GB/T
Pre-treatment 13885-2003) is carried out to sample, is respectively diluted to A liquid, B liquid, C liquid, D liquid properly according to concentration of metal ions
Concentration, upper machine measure.
5. chelation percent measures
Metal ion is mainly by Chelating state and free state (the mainly presence in the form of sulfate) in small peptide metallo-chelate
Two kinds of forms exist.When the pre-treatments such as the heated dissolving of small peptide metallo-chelate, centrifugation, free metal ion-solubility therein
In water；Since different polypeptide fractions different solubilities, a part are not readily dissolved in the chelating of water in small peptide metallo-chelate
State metal ion is present in lower sediment, and the soluble Chelating state metal ion of a part is dissolved in water.In certain alkalinity
Eluent under the conditions of, free state metal ion is converted into hydroxide precipitation and is gathered into macromolecular and rests on gel color in supernatant
Spectrum top end can not elute, and Chelating state metal ion can not be combined with hydroxyl due to the protection of coordinating group and generate hydrogen-oxygen
Change precipitation, can be eluted through alkaline eluant, realizes the separation of Chelating state and free metal ion.In acidic effluent liquid
Elution under, hydroxide precipitation dissolving, free metal ion is eluted collection.
With aas determination alkaline eluant C, acidic effluent liquid D and B liquid concentration can calculate free state and
Chelating state metal ion content, to calculate chelation percent, formula is as follows：
In formula：
X：Small peptide metallo-chelate chelation percent, %
M1：Weigh small-peptide chelated copper sample weight, mg
M2：Weigh small-peptide chelated copper sample weight, mg
VA：The volume of A liquid, L
CB：Concentration of metal ions in B liquid, mg/L VB：The volume of B liquid, L
CC：Concentration of metal ions in C liquid, mg/L VC：The volume of C liquid, L
CD：Concentration of metal ions in D liquid, mg/L VD：The volume of D liquid, L
6. the selection of alkaline eluant pH value
In order to exclude influence of the sample solution metal ion content to sedimentation effect in alkaline eluant elution process, metal from
The concentration of substandard metal ion in solution is not less than the concentration of metal ions in small peptide metallo-chelate loading to be measured, is computed
CuSO4Standard solution and ZnSO4Concentration of standard solution is respectively 600mg/L, 800mg/L.Draw 200 μm of CuSO4Standard solution
Or ZnSO4Standard solution according to 3.2.3 method loading, Cu select respectively pH for 8.0,8.5,9.0,9.5 alkaline eluant
Elution, Zn select pH to be eluted for 7.0,8.0,9.0,10.0 alkaline eluant, then use pH 2.0HCl acidic effluent liquids respectively
Elution, measured the rate of recovery of Cu or Zn ions after column, and determined the Optimal pH of alkaline eluant.
In formula：
X：The rate of recovery of metal ion, %
CC：Concentration of metal ions in alkali eluent, mg/L VC：The volume of alkali eluent, L
CD：Concentration of metal ions in sour eluent, mg/L VD：The volume of sour eluent, L
In formula：
X：The dissociation yield of small peptide metallo-chelate, %
CC：Concentration of metal ions in alkali eluent, mg/L VC：The volume of alkali eluent, L
CD：Concentration of metal ions in sour eluent, mg/L VD：The volume of sour eluent, L
3 interpretation of result of embodiment is as follows：
1. sephadex specification in gel filtration chromatography column
Common sephadex is broadly divided into G-10, G-15, G-25, G-35, G-50, G-75, different size in experiment
Sephadex due to degree of cross linking difference, separating ranges are different, main separating ranges and purposes such as table 1.Enzymatic hydrolysis of soybean egg
Polypeptide molecular weight in white powder<1000D, therefore the sephadex of G-15 specifications is selected in this experiment.
The separating ranges of the different sephadex specifications of table 1
2. the selection of alkaline eluant pH
As shown in Table 2, with the increase of alkaline eluant pH value, Cu2+The rate of recovery gradually increase.As pH=8.0 its
The rate of recovery is less than 99%, illustrates the Cu that dissociates under this alkaline condition2+It cannot be precipitated completely, dissociate Cu for part2+By alkaline eluant
It elutes；Work as pH>Its rate of recovery reaches 99.5% or more when 8, illustrates Cu at this time2+Substantially hydroxide precipitation can be formed, inhaled
Chromatography top end is attached to not to be eluted；Work as pH>Its rate of recovery increases after 9, but difference is not notable, and it is basic to increase pH
The rate of recovery, but the too strong structure that may destroy gel particle of alkalinity will not be significantly improved, therefore, attached gel column is to the resistance to of alkali
By property, the alkaline eluant of pH9 is selected to be eluted.The Optimal pH that this is selected with Yang Lin etc. in experiment in (2003) is consistent.
As shown in Table 2, with the increase of alkaline eluant pH value, Zn2+The rate of recovery gradually increase.Work as pH<When 9, Zn2+
The rate of recovery is less than 99%, illustrates the Zn that dissociates under this alkaline condition2+It cannot be precipitated completely, as pH=9.0, the rate of recovery is more than
99.5%, illustrate Zn at this time2+Substantially hydroxide precipitation can be formed, chromatography top end is adsorbed onto and is not eluted；But work as pH>
The rate of recovery has a declining tendency when 9, and analysis possible cause is Zn (OH)2It is a kind of amphoteric hydroxide, works as OH-When concentration excess
Zn(OH)2It can occur to dissolve and washed away by alkaline eluant, so as to cause Zn2+The rate of recovery declines, therefore selects the alkalinity of pH9
Elution Zn2+。
The selection of 2 alkali eluent Optimal pH of table
3. the influence of two kinds of small peptide metallo-chelates of alkaline condition pair dissociation
The results are shown in Table 3.Under the alkaline eluant elution requirement of pH9 known to result, 92.4% small peptide copper chelating
Object dissociates, and 99.7% small peptide chelates of zinc dissociates.This experiment waits for that test sample is small peptide metallo-chelate after purification,
It waits for being substantially free of free metal ion in test sample.Be soluble chelating state metal in supernatant in pretreatment process from
Son, theoretically the chelation percent of gel filtration chromatography should be close to 100%, but the alkalinity in pH9 is understood by experimental result
Small peptide metallo-chelate is largely dissociated under the conditions of elution.Therefore this experimental result declaratives small peptide copper chelating
Object, small peptide chelates of zinc stability be less than its corresponding Kocide SD, zinc hydroxide stability, pH9 alkalinity to two kinds
The structure of small peptide metallo-chelate has very strong destructiveness, leads to inaccuracy (table 4) of chelation percent measurement result
The degree of dissociation of two kinds of small peptide metallo-chelates under the conditions of 3 alkaline eluant pH9 of table
Metal ion-chelant rate in 4 small peptide copper chelate of table and small peptide chelates of zinc
Comparative analysis：Different chelation percent assay methods measure the chelation percent results contrast of two kinds of small peptide metallo-chelates
It is measured using gel filtration chromatography (embodiment 9) and the improved methanol extraction method (embodiment 1＆2) of the present invention small
The chelation percent of peptide metallo-chelate, as shown in Table 5, gel filtration chromatography measure small peptide copper chelate, small peptide chelates of zinc
Chelation percent is respectively 19.2%, 4.65%, the chelation percent that the improved methanol extraction method of the present invention measures is respectively 94.97%,
99.65%.The result difference that two methods measure is notable.Existing National Standard Method " gel filtration chromatography " is not suitable for small peptide copper (zinc)
The chelation percent of chelate measures, and reason is as follows：1) in metal ion and polypeptide ligand cohesive process, due to different ligands and metal
The chelating intensity of ion is different, and the stability of part small peptide metallo-chelate is poor, causes small peptide metallo-chelate structure by hydrogen-oxygen
Root brokenization forms hydroxide precipitation to which metal ion is replaced out, Chelating state metal and free state is caused not to divide completely
From keeping testing result less than normal.2) entire test experience, which takes around, spends the 5h times, and time-consuming and complex steps grasp experiment
It is required higher, the quick detection needs in unsuitable actual production.
The method of the invention is simple, is applicable in the small peptide metallo-chelate of complicated component.
5 different measuring methods of table measure small peptide copper chelate, small peptide chelates of zinc chelation percent results contrast
Note：With column data shoulder mark difference lowercase letter indication difference significantly (P<0.05), same letter or without letter indicate
Not notable (the P of difference>0.05).
Bibliography
1. Yang Lin, Yang Haipeng, Qian Qin wait measurement [J] feed industries of hydroxy methionine irons, copper, manganese, zinc chelation percent,
2003(6):34-36.
2.GB/T 13885-2003, calcium in animal feed, copper, iron, magnesium, manganese, potassium, sodium and Zn content measurement atom inhale
Receive spectroscopic methodology [S]
3.GB/T 13080.2-2005, the measurement gel filtration of feed addictive methionine iron (copper, manganese, zinc) chelation percent
Chromatography [S]
4. Hu Xiaobo thanks to chelation percent assay method summary [J] food section that bright brave organic trace elements network (chela) closes object
It learns, 2009,30 (23):508-511.
Claims (4)
1. a kind of detection method of small peptide micro-element chelate rate, which is characterized in that this method includes the following steps：
Step 1：Content micro- in small peptide microelement chelate sample is measured using atomic absorption spectrophotometer method
For W；
Step 2：The another small peptide microelement chelate to be measured for taking quality to be m, which is dissolved in the methanol that volume is V, to be extracted, and is being extracted
It takes and is shaken under the constant temperature that temperature is T, after extraction time t1, by its high speed centrifugation t2 minute, supernatant is taken to use atom
It is C that absorption spectrophotometer method, which measures its free microelement concentration,；
Step 3：The chelation percent measured under different extraction conditions is calculated by following formula：
In formula：
X：Small peptide micro-element chelate rate, unit %
W：Mass percent in small peptide microelement chelate sample shared by trace element, unit %
m：The another example weight for taking a small amount of small peptide microelement chelate for methanol extraction, unit mg
C：Microelement concentration in supernatant, unit mg/L
V：The volume of extract liquor, unit L
Wherein, the value range of the quality m of the small peptide microelement chelate to be measured described in step 2 is 25~1200mg；Institute
The variation range of the extraction temperature T stated is 0~50 DEG C；The value range of the extraction time t1 is 5~90min；Described
The value range of ultracentrifugal centrifugal speed is 12000r/min；The high speed centrifugation time t2 is 15min；The extraction
It is 3 × 10 to take the volume V of liquid-3L, solvents than value range 25:3~1200:3mg/mL.
2. the detection method of small peptide micro-element chelate rate according to claim 1, which is characterized in that described
Small peptide microelement chelate is small peptide copper chelate and/or small peptide chelates of zinc.
3. the detection method of small peptide micro-element chelate rate according to claim 1 or 2, which is characterized in that when
Small peptide copper chelate chelation percent is measured, copper content is in 13%~16% range in small peptide copper chelate, the value range of the m
Value range for 600~1000mg, the extraction temperature T is 2~15 DEG C, and the value range of the extraction time t1 is
10~40min, solvents than value range 600:3~1000:3mg/mL.
4. the detection method of small peptide micro-element chelate rate according to claim 1 or 2, which is characterized in that when
Small peptide chelates of zinc chelation percent is measured, Zn content is in 14%~17% range in small peptide chelates of zinc, the value range of the m
Value range for 550-800mg, the extraction temperature T is 15~25 DEG C, and the value range of the extraction time t1 is
15-40min, solvents than value range 550:3~800:3mg/mL.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111693644A (en) * | 2020-07-19 | 2020-09-22 | 广州天科生物科技有限公司 | Method for detecting content of free zinc in amino acid zinc complex |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101368903A (en) * | 2008-09-19 | 2009-02-18 | 广州天科生物科技有限公司 | Detection method for measuring amino acid micro-element chelate rate |
| CN107286219A (en) * | 2016-03-30 | 2017-10-24 | 中国海洋大学 | Photometry and XRD combinations determine micro- compound amino acid/small-peptide chelated thing synthetic ratio and the quick estimating and measuring method of chelation percent |
-
2018
- 2018-07-23 CN CN201810811488.4A patent/CN108613939B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101368903A (en) * | 2008-09-19 | 2009-02-18 | 广州天科生物科技有限公司 | Detection method for measuring amino acid micro-element chelate rate |
| CN107286219A (en) * | 2016-03-30 | 2017-10-24 | 中国海洋大学 | Photometry and XRD combinations determine micro- compound amino acid/small-peptide chelated thing synthetic ratio and the quick estimating and measuring method of chelation percent |
Non-Patent Citations (5)
| Title |
|---|
| LAETITIA L.S. CANABADY-ROCHELLE等: "SPR screening of metal chelating peptides in a hydrolysate for their antioxidant properties", 《FOOD CHEMISTRY》 * |
| 乔月建: "小肽螯合锌的制备及对公猪生产性能、血液生理化指标的影响", 《中国优秀硕士学位论文全文数据库》 * |
| 王子怀等: "肽- 金属离子螯合物的研究进展", 《食品工业科技》 * |
| 薛爱爱: "氨基酸和二肽金属螯合物的研究", 《中国优秀硕士学位论文全文数据库》 * |
| 邢颖: "氨基酸金属离子螯合物合成条件及测定方法的研究", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111693644A (en) * | 2020-07-19 | 2020-09-22 | 广州天科生物科技有限公司 | Method for detecting content of free zinc in amino acid zinc complex |
| CN111693644B (en) * | 2020-07-19 | 2022-09-20 | 广州天科生物科技有限公司 | Method for detecting content of free zinc in amino acid zinc complex |
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