CN103472124A - Fingerprint identification method of marine organisms as well as products thereof - Google Patents
Fingerprint identification method of marine organisms as well as products thereof Download PDFInfo
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- CN103472124A CN103472124A CN201310388947XA CN201310388947A CN103472124A CN 103472124 A CN103472124 A CN 103472124A CN 201310388947X A CN201310388947X A CN 201310388947XA CN 201310388947 A CN201310388947 A CN 201310388947A CN 103472124 A CN103472124 A CN 103472124A
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Abstract
The invention discloses a fingerprint identification method of marine organisms as well as products thereof. The fingerprint identification method comprises the following steps of freeze-drying collected marine organisms as well as products thereof and samples of baits in an appliance without the interference of stable isotopes, and grinding into powder by the appliance without the interference of stable isotopes; detecting the isotopes; calculating the result; and establishing a fingerprint library of the stable isotopes of marine organisms as well as products thereof of different types at different sea areas through the obtained result. According to the invention, the fingerprint library of stable isotopes is established through the result, and can be used for directly comparing the components of the stable isotopes of the marine organisms as well as the products thereof at different sea areas and further judging the organism types. The differences between marine organisms as well as products thereof in peculiar sea areas and marine organisms as well as products thereof in other sea areas are analyzed by virtue of a stable isotope technique, and the fingerprint library of the stable isotopes of the marine organisms as well as the products thereof at different sea areas is established through comparing the values of Delta 13C and Delta 15N of the same type of animals at different areas. The fingerprint identification method provided by the invention is beneficial to detection on differences of precious seafood as well as products thereof at different sea areas.
Description
Technical field
The present invention relates to the authentication method of a kind of sea life and goods thereof, particularly a kind of fingerprint identification method of sea life and goods thereof.
Background technology
In recent years, due to blind import and to reasons such as the background of introduced variety are unclear, cause that Dalian precious marine product endemic species incubation rate and survival rate reduce, the germplasm degradation phenomenas such as the speed of growth slows down, resistance variation constantly occur; Have many businessmans to utilize the similar characteristics of outward appearance such as precious marine product shape, color, with other, kind inferior pretends to be the Dalian precious marine product to be speculated again, and this has greatly damaged the image of Dalian characteristic biology and goods.At present halobiontic classification is mainly carried out for the polymorphism of protein (enzyme) and DNA, utilize DNA sequence dna and hereditary information to differentiate the sea life kind, especially in recent years the development that molecular biotechnology is advanced by leaps and bounds, a series of molecular genetic markers emerge in large numbers in succession, make people obtain abundanter hereditary information at DNA level, but no matter be protein labeling or molecular labeling, this procedure more complicated, and the cycle is longer, spend also higher.
Different biosomes have different isotopic ratios due to the difference of food source and the impact of isotopic fractionation, can be used for judging the isotopics of a certain histoorgan in the zones of different biosome, and then analyze the difference that between different biosomes, stable isotope forms.Stable isotope technology stablizing, the cycle is short and spend the advantages such as less, aspect the evaluation of sea life and goods, can play a significant role, so it is imperative to set up the stable isotopic fingerprint storehouse of sea life and goods.
Summary of the invention
The problems referred to above that exist for solving prior art, the present invention will design a kind of sea life and the goods fingerprint identification method thereof that testing process is simple, sense cycle is short, testing cost is low.
To achieve these goals, technical scheme of the present invention is as follows: a kind of sea life and goods fingerprint identification method thereof comprise the following steps:
A, pre-treatment: the sample of the sea life of collecting and goods and bait is contained in the utensil disturbed without stable isotope after freeze drying, uses the utensil disturbed without stable isotope to be ground into powder;
The described utensil disturbed without stable isotope is the utensil by glass material system; Described freeze drying temperature is-60～-50 ℃, sublimation drying with the sample bone dry till; The order number of described powder was 80 purpose aperture sieve;
B, isotope detection: taking out sample, deliver in elemental analyser through automatic sampler after wrapping with tin can, is pure CO by carbon in sample and nitrogen cycling of elements
2gas and N
2after gas, then, through dilution instrument dilution and purification, finally enter Delta V Advantage mass spectrometer and detected, obtain carbon stable isotope δ
13c and nitrogen stable isotope δ
15the value of N, and set up sea life and goods stable isotopic fingerprint storehouse and application system;
Described stable isotope is carbon and nitrogen stable isotope; Carbon and nitrogen stable isotope form the foundation for sea life and goods stable isotopic fingerprint storehouse and application system; The carbon stable isotope that described fingerprint base is different waters sea life and goods and nitrogen stable isotopic fingerprint;
C, result are calculated:
The natural abundance of internationally recognized Stable Carbon and nitrogen isotope is expressed as:
δ X=([R
sample/ R
standard]-1) * 10
3
Wherein X representative
13c or
15n; The R representative
13c/
12c or
15n/
14n; δ
13the C value is the natural abundance with respect to the PDB standard, δ
15the N value is the abundance of nitrogen in relative air; Analysis precision δ
13c<0.10 ‰, δ
15n<0.20 ‰; The stable isotope abundance is the stable isotope value;
D, measurement result:
Computing formula according to step C, by computer, mass spectrometric spectrogram result is calculated according to blas, gained is the measurement result that nitrogen and carbon isotope ratios forms, and by acquired results, sets up different waters variety classes sea life and goods stable isotopic fingerprint storehouse.
In step B of the present invention, the running parameter of each instrument is as follows:
Elemental analyser: the oxidation furnace temperature is 950 ℃, and the reduction furnace temperature is 650 ℃, and injector helium purge flow is 300ml/min, and oxygen flow is 180ml/min, and the reference gas flow is 250ml/min;
The dilution instrument of ConFloIV type: helium pressure is 0.6bar;
Delta V Advantage mass spectrometer: the analytical column temperature is 45 ℃; With USGS24, be δ
13c
pDB=-16.00 ‰ demarcate CO
2steel cylinder is δ with IAEAN1
15n
air=0.4 ‰ demarcates N
2steel cylinder; With the steel cylinder of demarcating as standard.
Compared with prior art, the present invention has following beneficial effect:
1, the present invention is dried, is pulverized by Marine Biological Samples, directly carries out the stable isotope composition analysis, and the sample drying time is 24h, and each sample stable isotope detects as 10min, and each sample detection consumes about RMB 5～10.Set up the stable isotopic fingerprint storehouse by result, can be used for directly comparing different waters sea life and goods stable isotope forms, and then differentiation biological species, different biosomes have different isotopic ratios due to the difference of food source and the impact of isotopic fractionation, can be used for judging the isotopics of a certain histoorgan in the zones of different biosome, and then analyze the difference that between different biosomes, stable isotope forms.Utilize stable isotope technology to analyze the otherness between specified sea areas sea life and goods and other marine site, the δ by more same species in zones of different
13c and δ
15the value of N, set up different waters sea life and goods stable isotopic fingerprint storehouse.
2, the present invention selects glass mortar or the cutter disturbed without stable isotope, Marine Biological Samples is cut and grinds, and replace the method for 60 ℃ of oven dry with freeze drying, with the isotopic exchange of same stable in stable isotope and sample in the minimizing air.That the present invention has is stable, the cycle is short and spend the advantages such as less, has guaranteed the accuracy of monitoring information and has enriched degree, contributes to detect the otherness of different waters precious marine product and goods.
The accompanying drawing explanation
The present invention is 1 of drawings attached only, wherein:
Fig. 1 is process flow diagram of the present invention.
Embodiment
Below in conjunction with specific embodiment, the present invention will be further described, but the present invention is not limited to following examples.
The clam sample (being respectively reference sample 1 and reference sample 2) that collect in July, 2011 in marine site, Yingkou, the Bohai Sea and Huanghai Sea black stone reef marine site is taken back to laboratory, ultrapure water cleans up, get described sample musculature with the glass blade, be contained in glassware and be placed on freeze drier drying under-50 ℃, after drying completes, described reference sample 1 and reference sample 2 are put into to glass mortar to be ground, make powder cross 80 order aperture sieve, complete described reference sample 1 and the pre-treatment operation while detecting with reference to 2 stable isotope.
By the above-mentioned reference sample be disposed 1 and reference sample 2, measure by the following method wherein carbon, nitrogen stable isotope and form.
1, mensuration program:
Get the 2mg sample, tin can is delivered in elemental analyser (Flash EA1112 type) through automatic sampler after wrapping, in this sample, carbon and nitrogen cycling of elements are after pure CO2 gas and N2 gas, the dilution instrument that is the ConFloIV type through model again, finally enter Delta V Advantage mass spectrometer and detected.
The specific works parameter is as follows:
Elemental analyser: the oxidation furnace temperature is 950 ℃, and the reduction furnace temperature is 650 ℃, and injector helium purge flow is 300ml/min, and oxygen flow is 180ml/min, and the reference gas flow is 250ml/min.
The dilution instrument of ConFloIV type: helium pressure is 0.6bar.
Delta V Advantage mass spectrometer: the analytical column temperature is 45 ℃; Use USGS24(δ
13c
pDB=-16.00 ‰) demarcate CO
2steel cylinder, use IAEAN1(δ
15n
air=0.4 ‰) demarcate N
2steel cylinder.With the steel cylinder of demarcating as standard.
2, result is calculated:
The natural abundance of internationally recognized stable carbon and nitrogen isotopes is expressed as:
δ X=([R
sample/ R
standard]-1) * 10
3
Wherein X representative
13c or
15n.The R representative
13c/
12c or
15n/
14n.δ
13the C value is the natural abundance with respect to the PDB standard, δ
15the N value is the abundance of nitrogen in relative air.Analysis precision δ
13c<0.10 ‰, δ
15n<0.20 ‰.
3, measurement result:
According to the computing formula of step 2, by computer, mass spectrometric spectrogram result to be calculated according to blas, the measurement result that the gained nitrogen and carbon isotope ratios forms is as shown in table 1.
Table 1, with reference to sample carbon, nitrogen stable isotope composition measuring result
Annotate: P means utmost point conspicuousness detection level, and different letter representations have significant difference.
As shown in Table 1, use detection method provided by the invention, after the clam of two different waters is processed, it is extremely remarkable that resulting carbon, nitrogen stable isotope form difference, shows analytical approach accurate and effective provided by the invention.
Claims (2)
1. sea life and goods fingerprint identification method thereof is characterized in that: comprise the following steps:
A, pre-treatment: the sample of the sea life of collecting and goods and bait is contained in the utensil disturbed without stable isotope after freeze drying, uses the utensil disturbed without stable isotope to be ground into powder;
The described utensil disturbed without stable isotope is the utensil by glass material system; Described freeze drying temperature is-60～-50 ℃, sublimation drying with the sample bone dry till; The order number of described powder was 80 purpose aperture sieve;
B, isotope detection: taking out sample, deliver in elemental analyser through automatic sampler after wrapping with tin can, is pure CO by carbon in sample and nitrogen cycling of elements
2gas and N
2after gas, then, through dilution instrument dilution and purification, finally enter Delta V Advantage mass spectrometer and detected, obtain carbon stable isotope δ
13c and nitrogen stable isotope δ
15the value of N, and set up sea life and goods stable isotopic fingerprint storehouse and application system;
Described stable isotope is carbon and nitrogen stable isotope; Carbon and nitrogen stable isotope form the foundation for sea life and goods stable isotopic fingerprint storehouse and application system; The carbon stable isotope that described fingerprint base is different waters sea life and goods and nitrogen stable isotopic fingerprint;
C, result are calculated:
The natural abundance of internationally recognized Stable Carbon and nitrogen isotope is expressed as:
δ X=([R
sample/ R
standard]-1) * 10
3
Wherein X representative
13c or
15n; The R representative
13c/
12c or
15n/
14n; δ
13the C value is the natural abundance with respect to the PDB standard, δ
15the N value is the abundance of nitrogen in relative air; Analysis precision δ
13c<0.10 ‰, δ
15n<0.20 ‰; The stable isotope abundance is the stable isotope value;
D, measurement result:
Computing formula according to step C, by computer, mass spectrometric spectrogram result is calculated according to blas, gained is the measurement result that nitrogen and carbon isotope ratios forms, and by acquired results, sets up different waters variety classes sea life and goods stable isotopic fingerprint storehouse.
2. a kind of sea life according to claim 1 and goods fingerprint identification method thereof, it is characterized in that: in step B, the running parameter of each instrument is as follows:
Elemental analyser: the oxidation furnace temperature is 950 ℃, and the reduction furnace temperature is 650 ℃, and injector helium purge flow is 300ml/min, and oxygen flow is 180ml/min, and the reference gas flow is 250ml/min;
The dilution instrument of ConFloIV type: helium pressure is 0.6bar;
Delta V Advantage mass spectrometer: the analytical column temperature is 45 ℃; With USGS24, be δ
13c
pDB=-16.00 ‰ demarcate CO
2steel cylinder is δ with IAEAN1
15n
air=0.4 ‰ demarcates N
2steel cylinder; With the steel cylinder of demarcating as standard.
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| CN104678019A (en) * | 2015-03-01 | 2015-06-03 | 云南农业大学 | Panax notoginseng root tuber production place identification method based on stable isotope fingerprints |
| CN105628781A (en) * | 2015-12-24 | 2016-06-01 | 张敏 | Isotopic tracing method for fate characteristics of organic substances in aquaculture pond |
| CN106248774A (en) * | 2016-08-31 | 2016-12-21 | 成都科源生物技术有限公司 | A kind of method carrying out L leucine product analysis |
| CN106970408A (en) * | 2017-04-11 | 2017-07-21 | 上海海洋大学 | Stable isotope determination pre-treating method in siphonopods spectacle lens |
| CN111879846A (en) * | 2020-07-30 | 2020-11-03 | 华南农业大学 | Method for identifying authenticity of bird's nest by using elemental analysis-stable isotope mass spectrometry and application |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104678019A (en) * | 2015-03-01 | 2015-06-03 | 云南农业大学 | Panax notoginseng root tuber production place identification method based on stable isotope fingerprints |
| CN105628781A (en) * | 2015-12-24 | 2016-06-01 | 张敏 | Isotopic tracing method for fate characteristics of organic substances in aquaculture pond |
| CN106248774A (en) * | 2016-08-31 | 2016-12-21 | 成都科源生物技术有限公司 | A kind of method carrying out L leucine product analysis |
| CN106970408A (en) * | 2017-04-11 | 2017-07-21 | 上海海洋大学 | Stable isotope determination pre-treating method in siphonopods spectacle lens |
| CN111879846A (en) * | 2020-07-30 | 2020-11-03 | 华南农业大学 | Method for identifying authenticity of bird's nest by using elemental analysis-stable isotope mass spectrometry and application |
| CN111879846B (en) * | 2020-07-30 | 2021-08-20 | 华南农业大学 | Method for identifying authenticity of bird's nest by using elemental analysis-stable isotope mass spectrometry and application |
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Application publication date: 20131225 |