Seaweed field wild stichopus japonicus recognition model based on stable isotope and construction method thereof

Abstract

The invention relates to a seaweed field wild stichopus japonicus recognition model based on stable isotopes and a construction method thereof. According to the model provided by the invention, a large batch of stichopus japonicus samples can be identified in a stable isotope detection mode in combination with a parameter calculation method provided by the patent. The model and the identification method can be applied to large-scale rapid and accurate direction identification of wild stichopus japonicus in a seaweed field in an aquatic product market, are easier to popularize, and are beneficial to standard productization development of the wild stichopus japonicus market.

Description

Technical Field

[0001] The invention relates to the technical field of seafood traceability, and in particular to a seaweed field wild sea cucumber identification model based on stable isotopes and a construction method thereof. Background Art

[0002] With the development of society and the improvement of living standards, consumers pay more attention to the quality and reliability of food, and have higher requirements for the transparency and traceability of information on high-quality food, especially various aquatic products and agricultural and sideline ecological products. Wild sea cucumbers are ecological seafood unique to offshore ecosystems. Although they have natural advantages, due to the lack of a standardized certification and traceability system, there are still obstacles for them to realize their own ecological product brand value.

[0003] Mislabeling of seafood and agricultural products is a common phenomenon not only in China but also in the world. According to statistics, 80% of seafood imported by the United States has sales label errors; a large number of seafood products such as fish, shrimp, and shellfish in Australia are often mistakenly labeled as other species. Through investigation and research, the World Wildlife Fund is seeking to use stable isotopes as a way to determine the ecological label of seafood and certify the traceability of seafood ingredients. The demand for traceability of agricultural and sideline products has also prompted the European Union to formulate traceability regulations (178 / 2002 / EC). The European Commission Implementing Regulation (1337 / 2013) requires that at each stage of the production and distribution of agricultural and sideline products, operators must mark the traceability mark, which requires the geographical indication guarantee (Protected Geographic Indication), the name of origin guarantee (Protected Designation of Origin) and the traditional craft guarantee (Traditional Specialties Guarantee). In addition, international organizations such as the Global Aquaculture Stewardship Council and the Global Aquaculture Alliance are also exploring ways to certify seafood using similar methods.

[0004] At present, the traceability and identification of ecological seafood such as wild sea cucumbers is generally in the initial exploration stage. There are still various difficulties to overcome, and there is no feasible and targeted specific traceability and identification method. To solve this problem, it is urgent to seek reliable technical means to identify the "identity" of ecological products such as wild sea cucumbers. As a "natural fingerprint" shaped by various specific habitats during the growth of various food sources for humans, stable isotope abundance has attracted the attention of relevant researchers in identifying the authenticity and traceability of agricultural and sideline products and aquatic products. In recent years, the use of stable isotope analysis to track aquatic products and agricultural and sideline ecological products has become more and more research and gradually mature. Stable isotope analysis methods have gradually become a promising and reliable tool for identifying the authenticity and traceability of agricultural and sideline ecological products. According to theoretical research, stable isotope fingerprints may be able to realize the identification of the geographical source, origin, and production method of biological products, such as identifying wild sources or farmed sources, "organic" free-range or intensive large-scale production. Based on the above situation, developing a practical fingerprint identification method based on stable isotopes can not only reduce information asymmetry and improve consumer trust, but also become a reliable guarantee and promotion basis for building an ecological seafood brand.

[0005] In seaweed fields, wild sea cucumbers mainly use macroalgae as nutrients. Macroalgae include various large seaweeds from the Rhodophyta, Phaeophyta and Chlorophyta phyla, such as kelp, laver, Ulva, Sargassum, etc. Sea cucumbers in seaweed fields usually give priority to large seaweed and its debris, where large seaweed is abundant as a food source and is usually of higher quality than other foods (Wang Xiaoyan et al., 2019). Our food source contribution analysis based on programs such as IsoSource, SIAR and SIBER also shows that the average contribution of large seaweed in seaweed fields to local wild sea cucumbers reached 76.6%, which is higher than other food sources.

[0006] Reference: Wang Xiaoyan, Qiao Hongjin, Huang Bingshan, Wang Chengqiang, Li Peiyu, Li Baoshan, Wang Jiying. Application of five seaweeds in feed for juvenile sea cucumbers. Advances in Fisheries Science, 2019, 40(3): 160–167 Summary of the invention

[0007] In order to solve the above problems, the present invention provides a model for identifying wild sea cucumbers in seaweed fields based on stable isotopes, which is as follows:

[0008]

[0009] in:

[0010] δ 15 N x The nitrogen stable isotope limit value of wild sea cucumbers in the target seaweed field;

[0011] δ 15 N Y is the nitrogen stable isotope value of the sea cucumber to be identified;

[0012] δ 15 N j is the nitrogen stable isotope value of the macroalgae in the target seaweed field;

[0013] Y j The macroalgae trophic level for the target seaweed field.

[0014] Specifically, (1) when the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (2) when the model output result has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (3) when the model output result has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

[0015] The above method for further analysis and confirmation based on the value of carbon stable isotopes is:

[0016] Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field;

[0017] If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0018] If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

[0019] Based on the above model, the present application also provides a method for constructing a model. When constructing the model,

[0020] By collecting multiple wild sea cucumber samples from the target seaweed field for testing, the δ 15 N eigenvalue upper limit δ 15 N x ;

[0021] A meta-analysis method can also be used to collect historical data on stable isotopes of wild sea cucumbers in the target seaweed field, and statistical analysis can be used to obtain the δ 15 N is the upper limit of the eigenvalue.

[0022] By statistically analyzing multiple samples of large seaweed in the target seaweed field, the average value of nitrogen stable isotopes of large algae in the target seaweed field, δ 15 N j .

[0023] The above model can be used to identify wild sea cucumbers in target seaweed fields.

[0024] A method for identifying wild sea cucumbers in seaweed fields based on stable isotopes, wherein the nitrogen stable isotope value of the sea cucumber to be identified is tested and calculated using a model formula;

[0025] (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0026] (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0027] (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

[0028] The above method for further analysis and confirmation based on the value of carbon stable isotopes is:

[0029] Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field;

[0030] If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0031] If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

[0032] Compared with the traditional wild sea cucumber identification method, the present invention, based on the indication of stable isotopes, greatly improves the accuracy and operability of wild sea cucumber identification in seaweed fields. This patent concretizes and indexes various identification indicators.

[0033] The present invention uses various indicators based on stable isotopes as the basis for identification. Compared with traditional sensory identification methods such as appearance, color, smell, and taste, the accuracy has been greatly improved, and the operability of large-scale commodity identification has been achieved. That is, through the method of stable isotope detection, combined with the parameter calculation method provided by this patent, sea cucumber samples can be identified in large quantities. Correspondingly, the stable isotope analysis and testing technology required by the method of the present invention has been gradually promoted internationally, the test accuracy meets the requirements, and the batch testing of sea cucumbers is accurate and fast. Combining the above-mentioned detection basis, analysis basis and identification indicators, this patent can be applied to the rapid and accurate identification of wild sea cucumbers in seaweed fields in the aquatic product market. It is easier to promote and is conducive to the standardized product development of the wild sea cucumber market. DETAILED DESCRIPTION

[0034] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0035] Example 1

[0036] A stable isotope-based identification model for wild sea cucumbers in seaweed fields is as follows:

[0037]

[0038] in:

[0039] δ 15 N x The nitrogen stable isotope limit value of wild sea cucumbers in the target seaweed field;

[0040] δ 15 N Y is the nitrogen stable isotope value of the sea cucumber to be identified;

[0041] δ 15 N j is the nitrogen stable isotope value of the macroalgae in the target seaweed field;

[0042] Y j The macroalgae trophic level for the target seaweed field.

[0043] During identification, (1) when the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (2) when the model output result has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (3) when the model output result has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

[0044] As a specific embodiment, the above method for further analyzing and confirming the value based on carbon stable isotopes is:

[0045] Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field;

[0046] If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0047] If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

[0048] The present application provides a method for constructing the above model. Specifically, when constructing the model,

[0049] By collecting multiple wild sea cucumber samples from the target seaweed field for testing, the δ 15 N eigenvalue upper limit δ 15 N x ;

[0050] A meta-analysis method can also be used to collect historical data on stable isotopes of wild sea cucumbers in the target seaweed field, and statistical analysis can be used to obtain the δ 15 N is the upper limit of the eigenvalue.

[0051] By statistically analyzing multiple samples of large seaweed in the target seaweed field, the average value of nitrogen stable isotopes of large algae in the target seaweed field, δ 15 N j .

[0052] When the identification model for a target seaweed field is built, the relevant identification can be carried out. During the identification, the nitrogen stable isotope value of the sea cucumber to be identified is tested and calculated using the model formula;

[0053] (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0054] (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0055] (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

[0056] The above method for further analysis and confirmation based on the value of carbon stable isotopes is:

[0057] Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field;

[0058] If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0059] If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

[0060] Example 2

[0061] Based on the content of Example 1, the present application provides a specific method for constructing a wild sea cucumber identification model in seaweed fields based on stable isotopes, and the scheme is as follows:

[0062] 1. Preparation of Sea Cucumber Samples

[0063] Sampling sites of wild sea cucumbers in seaweed fields were selected, and 30 sea cucumber samples were collected at each sampling site to ensure that all samples were adult individuals that had reached sexual maturity, with an individual wet weight of more than 200 g. After collection, they were stored in sterile food-grade plastic bags and stored at -20°C before testing.

[0064] Pretreatment: dissect the sea cucumber sample, remove the internal organs, select the body wall, and wash it with pure water. The processed sample is stored at -20℃ for 24 hours to be completely frozen, and then placed in a -50℃ freeze dryer for 48 hours. After drying, it is ground into powder with a grinder, passed through an 80μm sieve, and dried and stored for stable isotope ratio mass spectrometry.

[0065] 2. Stable isotope detection of sea cucumber samples

[0066] The stable isotope test of sea cucumber samples uses the following standard substances: IAEA-600: δ 13 C=-27.71‰;δ 15 N = 1‰; Acetanilide #1: δ 13 C=-26.85‰;δ 15 N = -4.21‰; USGS-40: δ 13 C=-36.39‰;δ 15 N=-4.52‰.

[0067] Instrument conditions include: oxidation tube (fillers from top to bottom are quartz crucible, corundum beads, copper oxide, silver wire, quartz debris) temperature: 950°C, reduction tube (fillers from top to bottom are silver wire, high-purity reduced copper) temperature: 600°C; helium pressure: 0.12MPa; helium flow rate: 200mL / min; oxygen flow time: 20s; oxygen flow rate: 40mL / min;

[0068] Stable nitrogen isotopes (δ 15 N) Detection process:

[0069] About 0.5 mg of freeze-dried sea cucumber powder was accurately weighed and packed in 4×6 tin cups. The stable isotope ratio mass spectrometer (Delta Q Advantages, Thermo Fisher Scientific) was connected to the TM ) to determine the total organic nitrogen (TON) and the total organic nitrogen stable isotope ratio (δ 15 N TON ). Helium was used as carrier gas with a flow rate of 90 mL / min, a reaction tube temperature of 950°C, and a column temperature of 50°C. The nitrogen content in the sample was calculated based on the ratio of the peak area of ​​the sample nitrogen to the peak area of ​​the standard sample nitrogen, and the test accuracy was STD<0.6‰. The nitrogen index was the percentage of organic nitrogen and the nitrogen isotope ratio with atmospheric nitrogen as the reference standard. The stable isotope ratio results were expressed as δ 15N means that the accuracy STD is less than 0.3‰, and the calculation formula is as follows:

[0070]

[0071] 15 N / 14 N sample is the actual nitrogen isotope ratio of the sample; 15 N / 14 N air is the standard atmospheric nitrogen isotope ratio.

[0072] Table 1δ 15 N TON Measurement item index

[0073]

[0074] Carbon stable isotopes (δ 13 C) Detection process:

[0075] About 10 mg of the powdered sample was accurately weighed and packed in a 4×6 tin cup. The total organic carbon content (TOC) and the total organic carbon stable isotope ratio (δ 13 C TOC ). Helium was used as the carrier gas, with a flow rate of 90 mL / min, a reaction tube temperature of 950°C, and a column temperature of 50°C. The carbon index was the organic carbon percentage and the carbon isotope ratio using VPDB (Vienna Peedee Belemnite) as the standard; the stable isotope ratio was expressed as δ 13 C indicates that its test accuracy is STD<0.2‰, and the formula is as follows:

[0076]

[0077] 13 C / 12 C sample is the actual carbon isotope ratio of the sample, 13 C / 12 C VPDB It is the carbon isotope ratio of the international standard VPDB.

[0078] Table 2δ 13 C TOC Measurement item index

[0079]

[0080]

[0081] 3. Construction and analysis of wild sea cucumber identification model

[0082] The wild sea cucumber identification process is the core part of this patent, which is mainly divided into two steps: first, the nitrogen stable isotope values ​​of the wild sea cucumbers in the target seaweed field and the large algae in the seaweed field are used to build a wild sea cucumber identification model suitable for this seaweed field, and then the model is used for identification. During identification, there are mainly two steps. One is to substitute the nitrogen stable isotope value of the sample to be identified into the model for calculation, and determine whether the sample belongs to the wild sea cucumber in this seaweed field based on the calculation result; if so, then compare the carbon stable isotope of the sample to be identified with the statistical threshold range of the carbon stable isotope of the wild sea cucumber in the target seaweed field for further judgment.

[0083] The details are as follows:

[0084] 3.1 Identification of wild sea cucumbers

[0085] 3.1.1 Identification model of wild sea cucumbers relative to macroalgae in seaweed fields

[0086] The model of the present invention is mainly based on the trophic level of wild sea cucumbers in seaweed fields. For the trophic level identification process of wild sea cucumbers in seaweed fields, the trophic level of sea cucumber samples relative to the large algae in the seaweed field is first calculated. The trophic level calculation of sea cucumber samples refers to the segmented trophic level formula, and focuses on the actual situation of wild sea cucumbers in seaweed fields. The calculation range is limited to the nitrogen stable isotope limit (δ 15 N x ) range, and the nitrogen stable isotope values ​​of macroalgae in seaweed fields (δ 15 N j ) is used as the calculation basis for special calculations for the identification of wild sea cucumbers in seaweed fields.

[0087] The above segmented trophic level formula can be found in: Hussey et al., 2014; Reum et al., 2015.

[0088] Specifically, through hierarchical meta-analysis of experimental data, it was found that the fractionation coefficient Δδ between adjacent trophic levels 15 N has a linear relationship with food source, the formula is as follows:

[0089] Δδ 15 N = 5.92 - 0.27 × δ 15 N s ①

[0090] Among them, δ 15 N sis the nitrogen stable isotope value of the food source. The 95% highest posterior median (HPM) uncertainty intervals of the intercept (β0) and slope (β1) of the formula coefficient are [4.55, 7.33] and [-0.41, -0.14], respectively. This formula can predict the δ 15 N value, the trophic level formula can also be further modeled, with the trophic level of the feeder as δ 15 function of N.

[0091] To simulate the predicted δ 15 For the N value, we use formula ① and combine it with von Bertalanffy's δ 15 The improved model of N enrichment model obtains formula ②:

[0092]

[0093] Among them, δ 15 N x is the saturation delta that increases with trophic level 15 N value, δ 15 N j is the delta of the baseline organism 15 N value, k is the delta in each trophic level interval 15 N is close to δ 15 N x The rate, Y ss is the trophic level of the ingestor relative to a benchmark organism.

[0094] To solve Y ss The absolute value of Y i Based on formula ②, we increase Y j Parameters, obtained based on formula ② the trophic level Y of the feeder i Formula ③:

[0095]

[0096] Among them, δ 15 N lim The and k values ​​are calculated based on the aforementioned β0 and β1 values ​​using the following formulas ④ and ⑤:

[0097]

[0098]

[0099] In order to further calculate the identification of wild sea cucumbers in seaweed fields, we made targeted improvements based on formula ③, taking large seaweed in seaweed fields as baseline organisms, that is, the δ 15N value as the reference value δ 15 N j , and obtain the identification model formula ⑥ for wild sea cucumbers in seaweed fields:

[0100]

[0101] in:

[0102] δ 15 N x The nitrogen stable isotope limit value of wild sea cucumbers in the target seaweed field;

[0103] δ 15 N x The δ 15 N eigenvalue upper limit δ 15 N x In addition to this method, a meta-analysis method can also be used, that is, to collect historical data on stable isotopes of wild sea cucumbers in the target seaweed field, and to obtain the δ 15 N eigenvalue upper limit;

[0104] δ 15 N TL is the nitrogen stable isotope value of the sea cucumber to be identified;

[0105] δ 15 N base is the nitrogen stable isotope value of the macroalgae in the target seaweed field;

[0106] This isotope value is the average value of nitrogen stable isotopes of macroalgae in the seaweed field, calculated by statistical analysis of at least 30 samples of macroalgae in the target seaweed field.

[0107] Y j is the trophic level of macroalgae in the target seaweed field. In the seaweed field, macroalgae are the main nutrients of wild sea cucumbers. Macroalgae are primary producers and belong to the first trophic level. Therefore, Y j The value of is 1.

[0108] 3.1.2 Specific identification process

[0109] Based on the above model formula, according to the nitrogen stable isotope value δ 15 N TL : Calculate the specific TL value.

[0110] (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0111] (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0112] (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required;

[0113] (3-1) Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field.

[0114] If the carbon stable isotope value of the sample to be identified is outside the statistical threshold range of the carbon stable isotope of wild sea cucumbers in the target seaweed farm, the sample to be identified does not belong to the wild sea cucumbers in the target seaweed farm;

[0115] If the carbon stable isotope value of the sample to be identified is within the statistical threshold range of carbon stable isotopes of wild sea cucumbers in the target seaweed farm, the sample to be identified belongs to the wild sea cucumbers in the target seaweed farm.

[0116] During the identification process, there will be a certain error rate due to testing, human interference, accidental factors, etc., but since the samples in this identification process are in large quantities, individual slip-throughs can be ignored. If the identification rate of wild sea cucumbers in the seaweed farm reaches more than 95% for the whole batch of samples, the whole batch of samples can be considered wild from the seaweed farm according to probability. If individual adulteration issues are taken into consideration, the identification and certification levels can be further divided for batches with an identification rate of more than 95%, such as 95% for the third level, 95%-98% for the second level, 98%-100% for the first level, and so on.

[0117] Based on the above-mentioned model construction method, an identification model suitable for wild sea cucumbers in a certain area can be constructed for this area, such as the identification model of wild sea cucumbers suitable for the seaweed fields near the Changshan Islands constructed below in this application.

[0118] Specific experimental examples:

[0119] Based on the models and methods of Examples 1 and 2, the present application constructs an identification model for wild sea cucumbers suitable for the seaweed fields near the Changshan Islands, and performs identification based on this model.

[0120] 1. Preparation of Sea Cucumber Samples

[0121] In this case, 50 wild sea cucumber samples collected from the seaweed fields near Changshan Islands were randomly selected to establish the basic model of the target sea area. The samples were collected from four sampling sites in Changshan Islands: 10 samples from Daqin Island, 10 samples from Cheyou Island, 20 samples from Tuoji Island and 10 samples from Gaoshan Island.

[0122] All samples are adult individuals that have reached sexual maturity, with an individual wet weight of more than 200g. After collection, they are stored in sterile food-grade plastic bags and stored at -20℃ before testing. Pretreatment: Dissect the sea cucumber samples, remove the internal organs, select the body wall, and wash them with pure water. After the processed samples are stored at -20℃ for 24 hours to completely freeze, they are placed in a -50℃ freeze dryer for 48 hours. After drying, they are ground into powder with a grinder, passed through an 80μm sieve, and dried and stored for stable isotope ratio mass spectrometry testing.

[0123] 2. The stable isotope test was performed on the sea cucumber samples according to the method of Example 1 or 2. The test results are shown in Table 1.3.

[0124] After testing, the carbon and nitrogen stable isotope data of 50 wild sea cucumber samples from the seaweed fields near the Changshan Islands were obtained (see Table 3). This data served as the basis for modeling and identification.

[0125] Table 3 Stable isotope test data of wild sea cucumber samples from Changshan Islands seaweed field

[0126]

[0127]

[0128] 3.1 Construction of a stable isotope-based identification model for wild sea cucumbers in the Changshan Islands seaweed field

[0129] Based on the statistical data of 50 wild sea cucumber samples collected near the Changshan Islands, the relevant parameters required for constructing the model formula were obtained.

[0130]

[0131] in:

[0132] (1)δ 15 N x It is the limit value of nitrogen stable isotope of wild sea cucumbers in the target seaweed field. Based on the statistical data of 50 wild sea cucumber samples, the upper limit of the statistical value of nitrogen stable isotope of wild sea cucumbers in the target seaweed field is 8.96‰.

[0133] (2)δ 15 N y is the nitrogen stable isotope value of the sea cucumber to be identified;

[0134] (3)δ 15 N j is the nitrogen stable isotope value of the macroalgae in the target seaweed field; that is, the average nitrogen stable isotope value of the macroalgae in the Changdao seaweed field, which is 8.13‰ calculated by statistically analyzing 117 samples of macroalgae in the target seaweed field;

[0135] (4)Yj is the trophic level of the macroalgae in the target seaweed field; that is, the trophic level of the macroalgae in the Changdao seaweed field, where the algae are primary producers and belong to the first trophic level, then Y j Set to 1.

[0136] By supplementing the parameters based on the statistical data of wild sea cucumbers, the following model formula was obtained:

[0137]

[0138] Based on the above model formula, the nitrogen stable isotope value δ 15 N y Get the Y value and then use the following method to identify it.

[0139] (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0140] (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0141] (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required;

[0142] (3-1) Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field.

[0143] If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field;

[0144] If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

[0145] 4. Verification of recognition results

[0146] In this case, 50 bottom-seeded sea cucumber samples, 39 farmed sea cucumber samples, and 30 wild sea cucumber samples collected from the waters near the Changshan Islands were randomly selected to reversely verify the identification accuracy of this patent.

[0147] When verifying recognition, the δ of the samples is tested separately 13 C(‰) and δ 15 N(‰), and the Y value of each sea cucumber sample is obtained by the above-constructed model, as shown in the following table:

[0148] Table 4 Stable isotope detection and Y value of samples to be identified

[0149]

[0150]

[0151]

[0152]

[0153] As can be seen from the table above, the identification model constructed based on this patent corresponds to the nitrogen stable isotope value (δ 15 N), the identification results of 50 bottom-sown sea cucumber samples from Changshan Islands showed that 43 samples did not calculate reasonable values ​​(i.e., these 43 samples did not belong to wild sea cucumbers from Changdao seaweed farms), and 2 of the 7 samples with calculated Y values ​​had Y≤2 (i.e., the other 5 samples did not belong to wild sea cucumbers from Changdao seaweed farms). A total of 2 of the 50 bottom-sown sea cucumber samples temporarily met the identification standards for wild sea cucumbers from seaweed farms of this patent, numbered CDHG 02 and CDHG 06, and entered the next step of the identification procedure.

[0154] Reasonable values ​​were not calculated for 33 of the 39 farmed samples (i.e., these 33 samples did not belong to the wild sea cucumbers from Changdao Seaweed Farm), and Y≤2 was found for 3 of the 6 samples that were calculated (i.e., the other 3 did not belong to the wild sea cucumbers from Changdao Seaweed Farm either). A total of 3 of the 39 farmed sea cucumber samples temporarily met the identification standards for wild sea cucumbers from seaweed farms of this patent, with numbers CDMDYZ 01, CDMDYZ 16, and CDMDYZ 29, and entered the next step of the identification procedure.

[0155] Reasonable values ​​were calculated for all 30 wild samples, and 29 samples with calculated results had Y≤2 (i.e., only 1 was identified as a wild sea cucumber that did not belong to the Changdao seaweed farm); 29 of the 30 wild sea cucumber samples temporarily met the seaweed farm wild sea cucumber identification standards of this patent, with the numbers shown in Table 5, and entered the next step of the identification procedure.

[0156] Table 5 The sea cucumber samples that need further confirmation are as follows

[0157]

[0158]

[0159] 4.2 Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field;

[0160] Carbon stable isotopes (δ 13 C) The statistical threshold range is -22.04‰ to -18.05‰ (from Table 1.3), and the carbon stable isotopes (δ 13C) values, samples numbered CDHG02, CDMDYZ01, CDMDYZ16, CDMDYZ29, CDYS02, and CDYS03 were judged not to be wild sea cucumbers from the Changdao Seaweed Farm. Finally, the samples judged to be wild sea cucumbers from the Changdao Seaweed Farm are shown in the following table.

[0161] Table 6: Samples of wild sea cucumbers from Changdao Seaweed Farm

[0162]

[0163]

[0164] 4.3 Recognition result statistics

[0165] This case study verified the identification of sea cucumbers from the seaweed fields near the Changshan Islands. The results showed that the identification accuracy of 50 bottom-sown sea cucumber samples was 98%; the identification accuracy of 39 farmed samples was 100%; and the identification accuracy of 30 wild sea cucumber samples from the Changshan Islands was 90.0%. A total of 119 samples from the three sources of sea cucumbers were combined, and 115 samples were accurately identified, with an overall identification accuracy of 96.6%.

[0166] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Although the present invention has been disclosed as a preferred embodiment as above, it is not used to limit the present invention. Any technical personnel in this field can make some changes or modify the technical contents disclosed above into equivalent embodiments without departing from the scope of the technical solution of the present invention. However, any brief modifications, equivalent changes and modifications made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention are still within the scope of the technical solution of the present invention.

Claims

1. A stable isotope-based identification model for wild sea cucumbers in seaweed fields, characterized in that: The details are as follows: in: δ 15 N x The nitrogen stable isotope limit value of wild sea cucumbers in the target seaweed field; δ 15 N Y is the nitrogen stable isotope value of the sea cucumber to be identified; δ 15 N j is the nitrogen stable isotope value of the macroalgae in the target seaweed field; Y j The macroalgae trophic level of the target seaweed field.

2. The stable isotope-based wild sea cucumber identification model for seaweed fields according to claim 1, characterized in that: (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

3. The identification model of wild sea cucumbers in seaweed fields based on stable isotopes according to claim 2, characterized in that: The above-mentioned method for further analysis and confirmation based on the carbon stable isotope value is: Compare the carbon stable isotope value of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field; If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

4. The method for constructing a model according to any one of claims 1 to 3, characterized in that: When building the model, By collecting multiple wild sea cucumber samples from the target seaweed field for testing, the δ 15 N eigenvalue upper limit δ 15 N x ; A meta-analysis method can also be used to collect historical data on stable isotopes of wild sea cucumbers in the target seaweed field, and statistical analysis can be used to obtain the δ 15 N is the upper limit of the eigenvalue.

5. The construction method according to claim 4, characterized in that: When building the model, By statistically analyzing multiple samples of large seaweed in the target seaweed field, the average value of nitrogen stable isotopes of large algae in the target seaweed field, δ 15 N j .

6. An identification model for wild sea cucumbers in a target seaweed field constructed by the construction method according to any one of claims 4 to 6.

7. Application of the model described in any one of claims 1 to 3 and claim 6 in identifying wild sea cucumbers in seaweed fields.

8. A method for identifying wild sea cucumbers in seaweed fields based on stable isotopes, characterized in that: Using the model described in claim 1, 2, 3 or 6, during identification, testing the nitrogen stable isotope value of the sea cucumber to be identified, and calculating it using the model formula; (1) When the model output result has no solution, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (2) When the model output has a solution and Y>2, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; (3) When the model output has a solution and Y≤2, further analysis and confirmation based on the value of carbon stable isotopes is required.

9. The identification method according to claim 9, characterized in that: The above method for further analysis and confirmation based on the value of carbon stable isotopes is: Compare the carbon stable isotope of the sample to be identified with the statistical threshold range of carbon stable isotope of wild sea cucumbers in the target seaweed field; If the carbon stable isotope of the sample to be identified is outside these two thresholds, the sample to be identified does not belong to the wild sea cucumber in the target seaweed field; If the carbon stable isotope of the sample to be identified is within these two thresholds, it belongs to the wild sea cucumber from the target seaweed field.

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