CN115980167B - Method for discriminating duck feces fragrance single-cluster tea harvesting season by stable isotope analysis technology and application - Google Patents
Method for discriminating duck feces fragrance single-cluster tea harvesting season by stable isotope analysis technology and application Download PDFInfo
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Abstract
The application belongs to the technical field of duck stool fragrance single tea discrimination, and particularly relates to a method for discriminating a harvesting season of duck stool fragrance single tea by a stable isotope analysis technology and application thereof. The stable isotope analysis technology is applied to distinguishing the Duck feces fragrance Dancong tea in different harvesting seasons. The application takes the stable isotope ratio of hydrogen and oxygen in tea as a characteristic index, and has the advantages of less required index, simple method, less sample consumption and the like. According to the application, the hydrogen and oxygen stable isotope ratio is combined with Fisher discriminant analysis to construct a discriminant model, and the harvesting season of the sample to be detected can be simply and rapidly predicted according to the season corresponding to the maximum value of the classification function by substituting the hydrogen and oxygen stable isotope ratio in tea into the discriminant model. Meanwhile, the Support Vector Machine (SVM) model established by the application also realizes rapid prediction of the harvesting season of the sample by substituting the stable isotope ratio of hydrogen and oxygen into the model.
Description
Technical Field
The application belongs to the technical field of duck stool fragrance single tea discrimination, and particularly relates to a method for discriminating a harvesting season of duck stool fragrance single tea by a stable isotope analysis technology and application thereof.
Background
The duck feces-flavored Dancong tea is a high-value variety with unique flavor in Chaozhou phoenix Dancong tea, and is wide in planting area and popular with consumers. The Duck feces fragrance single tea is produced all the year round, wherein spring tea which is formed by buds She Caizhi of tea trees sprouted for the first time after winter is rich in free amino acids, proteins and other components, and has good quality. Compared with single-cluster tea harvested in other seasons, the spring tea has higher economic value and drinking value. Under the drive of economic benefits, the condition that tea counterfeits in other seasons take advantage of spring tea is common in recent years, and consumer benefits and single tea brand reputation are seriously infringed.
Traditional single-tea-cluster harvesting season identification depends on sensory evaluation, and needs expertise to accurately distinguish, and accuracy is easily influenced by commodity forms and environments. Metabonomics analysis techniques based on tea chemistry have also been applied to harvest season identification. Li Xiaoling (Li Xiaoling; li; zhang Yuanyuan; chen Yufen; lin Xiaorong; chen Zhongzheng. Guangdong phoenix single-cluster three-flavor oolong tea physicochemical and aroma characteristics. Food industry science and technology. 2014.35.302-307.) and the like, the components such as water extract, caffeine, alcohols and olefins of the single-cluster tea are analyzed by adopting a high performance liquid chromatography and a headspace solid-phase microextraction-gas chromatography, and the spring tea and the autumn tea of the Honylang single-cluster tea, the almond single-cluster tea, the Wu She Shancong tea can be roughly distinguished by combining the main component analysis. And, there is a metabonomics analysis technique of patent CN202110285998.4The application of the space-time classification of the art to distinguish the Duck feces aroma single tea discloses the analysis of the characteristic components of the Duck feces aroma single tea by adopting a non-targeting metabonomics technology and the combination of Log 2 17 differential metabolites such as D-proline, eucalyptol, astragalin and the like are screened out by Fold Change, adj.p, VIP values and the like, and the harvesting season of the Duck feces fragrance single tea can be judged by bringing the peak area of the characteristic markers into a Duck feces fragrance single tea space-time classification judgment model. However, metabonomic analysis methods require the simultaneous determination of ten characteristic chemical components, resulting in high requirements for analytical determination conditions and methods. And the chemical component content of the tea can change in the processes of processing, storage and the like, thereby affecting the accuracy of season identification.
The stable isotope analysis technology is an effective analysis means for food traceability research, has the characteristics of higher detection precision and sensitivity, good stability and the like, and the stable isotope composition of the tea becomes an important characteristic index in researches such as tracing the tea production area, distinguishing seasons and the like. Zhou et al (Zhou, P.; li, Z.; ouyang, L.; gong, X.; meng, P.; dai, M.; wang, Z.; wang, Y.Amulti-element stable isotope approach coupled with chemometrics for the determination of Tieguanyin tea geographical origin and harvest seal. Animal. Methods 2019,11 (3), 346-352.) can distinguish between commercially available Tieguanyin spring tea and autumn tea by analysis of the carbon, nitrogen, hydrogen, oxygen stable isotope ratios in the tea leaves, by principal component analysis and hierarchical clustering analysis. Xia et al (Xia, W.; li, Z.; yu, C.; liu, Z.; nie, J.; li, C.; shao, A.; zhang, Y.; rogers, K.; yuan, Y.; understanding processing, maturity and harvest period effects to authenticate early-spring Longjing tea using stable isotopes and chemometric analysis.food control.2021.124.107907.; etc.) analyzed stable isotope composition in Longjing tea at different picking stages (early spring and late spring) based on field experiments, and could differentiate between early spring and late spring Longjing tea based on orthorhombic least squares discriminant analysis, wherein delta 15 The N value is the primary characteristic variable. However, the harvesting season distinguishing research based on stable isotope analysis is limited to one or two seasons, is not performed for the seasonal variation of a wide time scale within one year, and does not give out season distinguishing indexes and models suitable for single tea。
Disclosure of Invention
Aiming at the problems, the application aims to provide a method for distinguishing the harvesting season of the Duck feces-fragrance single-cluster tea by using a stable isotope analysis technology and application thereof, and the single-cluster tea is distinguished for the single-cluster tea in four seasons of spring, summer, autumn and winter based on characteristic indexes of stable isotopes and a distinguishing model, so that the Duck feces-fragrance single-cluster tea in four seasons of spring, summer, autumn and winter is accurately distinguished.
The technical content of the application is as follows:
the application provides an application of a stable isotope analysis technology for distinguishing a harvesting season of Duck feces aroma Dancong tea, wherein the stable isotope analysis technology adopts a stable isotope ratio of hydrogen and oxygen.
The application also provides a method for distinguishing the harvesting season of the Duck feces aroma Dancong tea by using the stable isotope analysis technology, which comprises the following steps:
1) Sample collection and preparation: collecting duck feces fragrance single-cluster tea collected in four seasons of spring, summer, autumn and winter in a single-cluster tea producing area, and recording information of a sample;
crushing and sieving the collected single-cluster tea samples;
2) Stable isotope ratio determination
Measuring the hydrogen and oxygen stable isotope ratio of a tea sample in a known real season by using an elemental analysis-stable isotope ratio mass spectrometer, weighing the tea sample, wrapping the tea sample by a silver cup, balancing the tea sample in a dryer for 2-3 days, and then placing the tea sample in the elemental analysis-stable isotope ratio mass spectrometer for measuring the hydrogen and oxygen stable isotope ratio by a high-temperature cracking reaction;
in the hydrogen-oxygen stable isotope ratio determination, the reaction parameters of the elemental analysis-stable isotope ratio mass spectrometer include: the high-temperature cracking tube temperature is 1380 ℃, the column temperature is 50 ℃, and the helium carrier gas flow rate is 100 L.min -1 The flow rate of helium purge gas is 150 L.min < -1 >, and the purity of helium is 99.999%;
3) Harvesting season discrimination model construction
The method comprises the following steps: according to the sample data adopted in the step 2), establishing a dependent variable recovery season and independent variable stability by using a Fisher discriminant analysis methodDetermining isotope ratio discrimination model and analyzing delta in tea 2 H value and delta 18 A discriminant function and a variance contribution rate established by the O value combination index;
build based on delta in tea 2 H value and delta 18 The model for distinguishing the harvesting seasons of the Duck feces fragrance single-cluster tea with the O value combination index is as follows:
f (season) =a+b×δ 2 H+C×δ 18 O；
Wherein A, B, C is a constant;
the stable isotope ratio data of the tea samples collected in the step 1) are brought into each harvesting season discrimination model to obtain a plurality of F values, and the harvesting season of the duck faeces aroma single-cluster tea samples to be tested is determined according to the maximum F value;
finally, carrying out in-group verification on duck feces fragrance single-cluster tea samples in each harvesting season to evaluate the reliability of the model;
the standard substance was USGS54 (. Delta.) 2 H VSMOW ＝-150.4‰，δ 18 O VSMOW ＝17.79‰)，USGS55(δ 2 H VSMOW ＝-28.2‰，δ 18 O VSMOW ＝19.2‰)，USGS56(δ 2 H VSMOW ＝-44‰，δ 18 O VSMOW =27.23%o) to perform multi-point correction;
the second method is as follows: building a harvesting season discrimination model by using a supervised learning classifier Support Vector Machine (SVM), building an SVM model by using a Gaussian kernel function, and setting the cost to be 2 -10 To 2 10 G is e -2 To e 2 Performing super-parameter optimization in the range of (1), and performing 7-fold cross validation;
and a third method: on the basis of the first method, the carbon and nitrogen stable isotope ratio is increased to carry out season discrimination, and the discrimination model is as follows:
f (season) =a+b×δ 2 H+C×δ 18 O+D×δ 13 C or F (season) =a+b×δ 2 H+C×δ 18 O+D×δ 15 N or F (season) =a+b×δ 2 H+C×δ 18 O+D×δ 13 C+E×δ 15 N；
Wherein A, B, C, D, E is a constant;
the carbon-nitrogen stable isotope ratio is obtained by measuring oxidation-reduction reaction in an elemental analysis-stable isotope ratio mass spectrometer.
The calculation formula of the stable isotope ratio is as follows:
δE＝[(R sample of /R Standard of )-1]；
Wherein E is the target isotope ratio, R Sample of For the abundance ratio of heavy isotope and light isotope in the tea sample to be measured, namely 2 H/ 1 H、 18 O/ 16 O、 13 C/ 12 C、 15 N/ 14 N；R Standard of The abundance ratio of heavy isotope to light isotope in the international universal standard sample; the delta value is multiplied by 1000 to be the unit per mill;
delta using the international standard VSMOW 2 H and delta 18 Confirming the value of O; using international general standard VPDB for delta 13 C, confirming the fixed value of the value C; delta using international standard AIR pair 15 And (5) confirming the value of N.
Further, in step 3), the method one:
f (spring tea) = -168.831-3.602 ×δ 2 H+9.063×δ 18 O；
F (Xia Cha) = -190.629-3.603 x delta 2 H+8.314×δ 18 O；
F (autumn tea) = -228.508-3.885 x delta 2 H+8.542×δ 18 O；
F (winter tea) = -208.019-3.415 ×δ 2 H+10.030×δ 18 O。
Further, in the second method of step 3), the cost value of the SVM model is 1024 and the number of support vectors is 42.
The beneficial effects of the application are as follows:
the method for distinguishing the duck stool fragrance single-cluster tea harvesting season by the stable isotope analysis technology can distinguish the single-cluster tea in four seasons of spring, summer, autumn and winter based on the characteristic indexes of isotopes and the distinguishing model.
The stable isotope analysis technology is applied to distinguishing the duck stool fragrance single tea harvesting seasons. The application takes the stable isotope ratio of hydrogen and oxygen in tea as a characteristic index, and has the advantages of less required index, simple method, less sample consumption and the like. According to the application, the hydrogen and oxygen stable isotope ratio is combined with Fisher discriminant analysis to construct a discriminant model, and the harvesting season of the sample to be detected can be simply and rapidly predicted according to the season corresponding to the maximum value of the classification function by substituting the hydrogen and oxygen stable isotope ratio in tea into the discriminant model. Meanwhile, the Support Vector Machine (SVM) model established by the application also realizes rapid prediction of the harvesting season of the sample by substituting the stable isotope ratio of hydrogen and oxygen into the model.
Drawings
FIG. 1 is a box plot of hydrogen and oxygen stable isotope ratios for duck feces single tea samples;
fig. 2 is a Fisher discrimination score of the actual harvest season of the known Duck feces-fragrance single-cluster tea.
Detailed Description
The application is described in further detail below with reference to specific embodiments and the accompanying drawings, it being understood that these embodiments are only for the purpose of illustrating the application and not for the purpose of limiting the same, and that various modifications of the application, which are equivalent to those skilled in the art, will fall within the scope of the appended claims after reading the present application.
Example 1
Method for distinguishing harvesting season of Duck feces-flavored single-cluster tea by stable isotope analysis technology
1) Sample collection and preparation: collecting duck faeces aroma single-cluster tea collected in four seasons of spring, summer, autumn and winter in a single-cluster tea producing area, wherein 71 samples of spring tea, 18 samples of summer tea, 32 samples of autumn tea and 25 samples of winter tea, and recording season information of the samples;
taking 100g of each collected single tea sample, putting the single tea samples into a grinder for grinding, sieving the single tea samples with a 100-mesh sieve, collecting undersize materials, and storing the undersize materials at 4 ℃ to be measured;
2) Stable isotope ratio determination
Measuring the hydrogen and oxygen stable isotope ratio of a tea sample in a known real season by using an elemental analysis-stable isotope ratio mass spectrometer, weighing 0.2-0.3 mg of the tea sample in a part per million day, wrapping the tea sample by a silver cup, placing the tea sample in a dryer for 2-3 days, and then placing the tea sample in the elemental analysis-stable isotope ratio mass spectrometer to perform hydrogen and oxygen stable isotope ratio measurement by high-temperature cracking reaction;
the reaction parameters of the elemental analysis-stable isotope mass spectrometer include: the high-temperature cracking tube temperature is 1380 ℃, the column temperature is 50 ℃, and the helium carrier gas flow rate is 100 L.min -1 Helium purge gas flow rate is 150 L.min -1 Helium purity was 99.999%;
the standard substance was USGS54 (. Delta.) 2 H VSMOW ＝-150.4‰，δ 18 O VSMOW ＝17.79‰)，USGS55(δ 2 H VSMOW ＝-28.2‰，δ 18 O VSMOW ＝19.2‰)，USGS56(δ 2 H VSMOW ＝-44‰，δ 18 O VSMOW =27.23%o) to perform multi-point correction;
the stable isotope ratio is calculated as:
δE＝[(R sample of /R Standard of )-1]；
Wherein E is the target isotope ratio, R Sample of For the abundance ratio of heavy isotope and light isotope in the tea sample to be measured, namely 2 H/ 1 H、 18 O/ 16 O；R Standard of The abundance ratio of heavy isotope to light isotope in the international universal standard sample; the delta value is multiplied by 1000 to be the unit per mill, and the international general standard VSMOW is adopted for delta 2 H value and delta 18 Confirming the value of O;
as shown in FIG. 1, delta of a known real-season Duck faeces aroma Dancong tea sample is determined by using an elemental analysis-stable isotope ratio mass spectrometer 2 H value and delta 18 The result of the O value reflects delta of the duck feces fragrance single tea samples in different seasons 2 H value and delta 18 The center position of the O value and the data dispersion.
3) Harvesting season discrimination model construction
According to the sample data adopted in the step 2), a Fisher discriminant analysis method is utilized to establish a discriminant model of the stable isotope ratio of the dependent variable and the independent variable in the harvesting season, and the stable isotope ratio delta in the tea is analyzed 2 H and delta 18 Discriminant function and variance contribution rate established by O combination index, wherein discriminant function 1 is y1=11.855+0.188×δ 2 H-0.013×δ 18 O, variance contribution ratio 80.2%, mainly including delta 2 Information of H value; the discriminant function 2 is y1= -15.353-0.074×δ 2 H+0.592×δ 18 O, mainly comprising delta 18 Information of the O value;
build based on delta in tea 2 H value and delta 18 The model for distinguishing the harvesting seasons of the Duck feces fragrance single-cluster tea with the O value combination index is as follows:
f (spring tea) = -168.831-3.602 ×δ 2 H+9.063×δ 18 O；
F (Xia Cha) = -190.629-3.603 x delta 2 H+8.314×δ 18 O；
F (autumn tea) = -228.508-3.885 x delta 2 H+8.542×δ 18 O；
F (winter tea) = -208.019-3.415 ×δ 2 H+10.030×δ 18 O；
And (3) taking stable isotope ratio data of the sample to be detected into each harvesting season discrimination model to obtain a plurality of F values. And determining the harvesting season of the duck feces aroma single-cluster tea sample to be tested according to the maximum F value. And finally, carrying out in-group verification on duck feces fragrance single-cluster tea samples in each harvesting season to evaluate the reliability of the model.
As shown in fig. 2, a score chart of Fisher discrimination of the real harvesting season of the known duck faeces aroma single-cluster tea shows that summer tea and winter tea can be respectively distinguished from tea leaves in other seasons; although the spring tea and the winter tea are relatively close, the group mass centers can be distinguished from each other; autumn tea, although a few discrete samples were close to winter tea, was still distinguishable from winter tea as a whole.
As shown in table 1, stable isotope ratio data of 71 samples of spring tea, 18 samples of summer tea, 32 samples of autumn tea and 25 samples of winter tea were taken into each harvesting season discrimination model to obtain F values. Wherein sample numbers 1-71 are spring tea, 72-89 are summer tea, 90-121 are autumn tea, and 122-146 are winter tea.
Table 1 stable isotope ratio and F-number for single tea samples
The collected duck faeces aroma single tea samples were subjected to in-group verification, and the results are shown in table 2. As can be seen from Table 2, in 146 duck stool fragrance single-cluster tea samples from different harvesting seasons, the discrimination accuracy of summer tea and winter tea is 94.4% and 96.0%, the discrimination accuracy of spring tea is 80.3%, and the discrimination accuracy of autumn tea is slightly lower, but the overall accuracy reaches 82.2%, which shows that the Fisher discrimination model constructed based on the stable hydrogen and oxygen parity ratio can be used for discriminating the harvesting seasons of the duck stool fragrance single-cluster tea.
TABLE 2 Fisher discrimination results based on the stable isotope ratio of hydrogen and oxygen
Example 2
A method for discriminating duck faeces aroma single-cluster tea harvesting seasons by stable isotope analysis technology is the same as that of the embodiment 1, and the difference is that the step 3) harvesting season discriminating model is constructed, and the harvesting season discriminating model is built by adopting a supervised learning classifier Support Vector Machine (SVM).
146 tea samples were randomly divided into training and test sets at a ratio of 7:3. Based on training set data, a Gaussian kernel function is adopted to construct an SVM model, and the cost is 2 -10 To 2 10 G is e -2 To e 2 And (3) performing super-parameter optimization in the range of (1) and performing 7-fold cross validation. The cost value of the finally established SVM model with the best harvesting season performance is 1024, the number of support vectors is 42, and the discrimination result is shown in table 3.
TABLE 3 SVM model discrimination accuracy based on Hydrogen and oxygen stable isotope ratios
As can be seen from table 3, in 146 duck stool flavored single-cluster tea samples from different harvesting seasons, the discrimination accuracy of Xia Cha and spring tea is 100.0% and 88.7%, the discrimination accuracy of autumn tea is 84.4%, and the discrimination accuracy of winter tea is slightly lower, but the overall accuracy reaches 87.0%, which indicates that the SVM model constructed based on the stable hydrogen and oxygen parity ratio can be used for discriminating the harvesting seasons of duck stool flavored single-cluster tea.
Example 3
The method for distinguishing duck faeces aroma single tea harvesting season by stable isotope analysis technology comprises the following operation steps, which are the same as those in the embodiment 1, wherein the difference is that the season distinguishing is carried out by adding carbon and nitrogen stable isotope ratio based on the hydrogen and oxygen stable isotope ratio;
the specific operation is as follows:
1. determination of carbon isotope ratio: about 0.2mg of the tea leaf sample was weighed and measured by oxidation-reduction reaction in a tin cup wrapped and placed in an elemental analysis-stable isotope ratio mass spectrometer. The instrument parameters include: the temperature of the oxidation tube is 960 ℃, the column temperature is 60 ℃, and the flow rate of helium carrier gas is 180 L.min -1 HeliumThe flow rate of the air purge gas is 70 L.min -1 The oxygen flow rate is 250 L.min -1 The oxygen injection time is 3s;
the standard substance was USGS40 (. Delta.) 13 C VPDB ＝-26.4193‰)，UREA((δ 13 C VPDB ＝-43.432‰)，IAEA601((δ 13 C VPDB = -28.506%o) for multi-point correction;
2. determination of nitrogen isotope ratio: about 1.2mg of the tea leaf sample was weighed and measured by oxidation-reduction reaction in a tin cup-wrapped and placed in an elemental analysis-stable isotope ratio mass spectrometer. The instrument parameters include: the temperature of the oxidation tube is 960 ℃, the column temperature is 60 ℃, and the flow rate of helium carrier gas is 180 L.min -1 Helium purge gas flow rate is 70 L.min -1 The oxygen flow rate is 250 L.min -1 The oxygen injection time is 3s;
the standard substance was USGS40 (. Delta.) 15 N AIR ＝-4.19967‰)，UREA(δ 15 N AIR ＝-0.595‰)，IAEA600((δ 15 N AIR = 1.017667%o) to perform multi-point correction;
3. constructing a Fisher discrimination model: according to the collected sample data, a Fisher discriminant analysis method is utilized to establish a discriminant model of the dependent variable harvesting season and the independent variable stable isotope ratio, a discriminant function and an accuracy established based on the multiple stable isotope ratios in the tea are analyzed, and the calculated harvesting season discriminant accuracy and the function are shown as follows.
TABLE 4 Fisher discrimination results based on various stable isotope ratios
The harvesting season discrimination models constructed based on the various stable isotope ratios represented by a, b and c in the table are as follows:
fa (spring tea) = -676.613-2.679 ×δ 2 H+8.386×δ 18 O-39.958×δ 13 C；
Fa (Xia Cha) di-756.558-3.198 Xdelta 2 H+7.600×δ 18 O-42.184×δ 13 C；
Fa (autumn tea) = -777.631-3.456 x delta 2 H+8.839×δ 18 O-41.553×δ 13 C；
Fa (winter tea) = -727.806-3.027 ×δ 2 H+9.346×δ 18 O-40.828×δ 13 C；
Fb (spring tea) = -170.674-3.081 ×δ 2 H+8.974×δ 18 O+2.145×δ 15 N；
Fb (Xia Cha) = -192.140-3.620 ×δ 2 H+8.233×δ 18 O+1.943×δ 15 N；
Fb (autumn tea) = -230.335-3.873×δ 2 H+9.453×δ 18 O+2.136×δ 15 N；
Fb (winter tea) = -209.313-3.430 ×δ 2 H+9.955×δ 18 O+1.798×δ 15 N；
Fc (spring tea) = -677.660-2.693 ×δ 2 H+8.320×δ 18 O-39.931×δ 13 C+1.617×
δ 15 N；
Fc (Xia Cha) = -757.326-3.211 ×δ 2 H+7.543×δ 18 O-42.161×δ 13 C+1.385×
δ 15 N；
Fc (autumn tea) = -778.639-3,470×δ 2 H+8.773×δ 18 O-41.526×δ 13 C+1.587×
δ 15 N；
Fc (winter tea) = -728.445-3.038 ×δ 2 H+9..293×δ 18 O-40.407×δ 13 C+1.263
×δ 15 N；
As can be seen from Table 4, the discrimination results of the model constructed with three stable isotope ratios, δ 2 H、δ 18 O and delta 13 The accuracy of the model constructed in the step C is more than 90% in the discrimination of summer tea and winter tea, and the discrimination accuracy of spring tea and autumn tea is general, but the overall accuracy reaches 82.9%; delta 2 H、δ 18 O and delta 15 The accuracy of the model constructed by N is more than 90% in the discrimination of summer tea and winter tea, and the model constructed by N is discriminated in spring tea and autumn teaThe accuracy is slightly lower, but the overall accuracy reaches 81.5%. At four stable isotope ratios delta 2 H、δ 18 O、δ 13 C and delta 15 The accuracy of the model constructed by N is more than 95% in the discrimination of summer tea and winter tea, and the discrimination accuracy of spring tea and autumn tea is common, but the overall accuracy reaches 83.6%.
Thus, as can be seen by comparing example 1 and example 3, two (δ 2 H、δ 18 O), and the Fisher discrimination model of the Duck feces aroma single-cluster tea season constructed by three or four stable isotope ratios is also stable and reliable.
Example 4
A method for discriminating duck faeces aroma single-cluster tea harvesting season by stable isotope analysis technology is characterized in that the operation steps are the same as those of the embodiment 1, wherein the difference is that a harvesting season discriminating model is constructed in the step 3), and specifically, an SVM model is constructed by adding carbon and nitrogen stable isotope ratios on the basis of hydrogen and oxygen stable isotope ratios.
146 samples were randomly divided into training and test sets at a ratio of 7:3. Based on training set data, a Gaussian kernel function is adopted to construct an SVM model, and the cost is 2 -10 To 2 10 G is e 2 To e 2 Performing super-parameter optimization within the range of (1), performing 7-fold cross validation, and establishing an SVM model with optimal harvesting season performance. For the optimal SVM model constructed by the combination indexes of the isotope ratios, the corresponding parameters are respectively as follows:
(1)δ 2 H、δ 18 o and delta 13 Index combination of C: the cost value is 256, and the number of support vectors is 54;
(2)δ 2 H、δ 18 o and delta 15 Index combination of N: a cost value of 64 and a support vector number of 74;
(3)δ 2 H、δ 18 O、δ 13 c and delta 15 Index combination of N: the cost value is 64 and the number of support vectors is 67.
The accuracy of the harvesting season discrimination model based on the SVM is shown in table 5.
TABLE 5 SVM model discrimination accuracy based on multiple stable isotope ratios
As can be seen from Table 5, the discrimination results, δ, of the model constructed with three stable isotope ratios 2 H、δ 18 O and delta 13 The accuracy of the model constructed in the step C is more than 90% in the discrimination of spring tea, summer tea and winter tea, and the discrimination accuracy of autumn tea is general, but the overall accuracy reaches 90.4%; delta 2 H、δ 18 O and delta 13 The accuracy of the model constructed in the step C is more than 90% in the discrimination of spring tea, summer tea and winter tea, and the discrimination accuracy of autumn tea is slightly low, but the overall accuracy reaches 91.1%. At four stable isotope ratios delta 2 H、δ 18 O、δ 13 C and delta 15 The accuracy of the model constructed by N is more than 90% in the discrimination of spring tea and summer tea, but the discrimination accuracy of spring tea and autumn tea is common, but the overall accuracy reaches 89.7%.
Claims (5)
1. A method for discriminating duck feces fragrance single tea harvesting seasons by stable isotope analysis technology is characterized by comprising the following steps:
1) Sample collection and preparation: collecting duck feces fragrance single-cluster tea collected in four seasons of spring, summer, autumn and winter in a single-cluster tea producing area, and recording information of a sample;
crushing and sieving the collected single-cluster tea samples;
2) Stable isotope ratio determination
Measuring the hydrogen and oxygen stable isotope ratio of a tea sample in a known real season by using an elemental analysis-stable isotope ratio mass spectrometer, weighing the tea sample, wrapping the tea sample by a silver cup, balancing the tea sample in a dryer for 2-3 days, and then placing the tea sample in the elemental analysis-stable isotope ratio mass spectrometer for measuring the hydrogen and oxygen stable isotope ratio by a high-temperature cracking reaction;
3) Harvesting season discrimination model construction
The method comprises the following steps: according to the sample data adopted in the step 2), a Fisher discriminant analysis method is utilized to establish a discriminant model of the ratio of the dependent variable harvesting season to the independent variable stable isotope, and delta in the tea is analyzed 2 H value and delta 18 A discriminant function and a variance contribution rate established by the O value combination index;
build based on delta in tea 2 H value and delta 18 The model for distinguishing the harvesting seasons of the Duck feces fragrance single-cluster tea with the O value combination index is as follows:
f (spring tea) = -168.831-3.602 ×δ 2 H+9.063×δ 18 O；
F (Xia Cha) = -190.629-3.603 x delta 2 H+8.314×δ 18 O；
F (autumn tea) = -228.508-3.885 x delta 2 H+8.542×δ 18 O；
F (winter tea) = -208.019-3.415 ×δ 2 H+10.030 ×δ 18 O；
The stable isotope ratio data of the tea samples collected in the step 1) are brought into each harvesting season discrimination model to obtain a plurality of F values, and the harvesting season of the duck faeces aroma single-cluster tea samples to be tested is determined according to the maximum F value;
finally, carrying out in-group verification on duck feces fragrance single-cluster tea samples in each harvesting season to evaluate the reliability of the model;
the second method is as follows: on the basis of the first method, the carbon and nitrogen stable isotope ratio is increased to carry out season discrimination, and the discrimination model is as follows:
f (season) =a+b×δ 2 H+C×δ 18 O+D×δ 13 C or F (season) =a+b×δ 2 H+C×δ 18 O+D×δ 15 N or F (season) =a+b×δ 2 H+C×δ 18 O+D×δ 13 C+E×δ 15 N；
Wherein A, B, C, D, E is a constant;
the carbon/carbon isotope ratio is measured by oxidation-reduction reaction in an elemental analysis-stable isotope ratio mass spectrometer.
2. The method for discriminating a harvesting season of Duck faeces aroma Dancong tea according to claim 1 wherein in said determination of hydrogen-oxygen stable isotope ratio in step 2), the reaction parameters of the elemental analysis-stable isotope ratio mass spectrometer include: the high-temperature cracking tube temperature is 1380 ℃, the column temperature is 50 ℃, the helium carrier gas flow rate is 100L/min, and the helium purge gas flow rate is 150 L.min -1 Helium purity was 99.999%.
3. The method for discriminating duck feces aroma single tea harvesting season by stable isotope analysis technique according to claim 1 wherein in said method one of step 3), standard substance adopts USGS54 (delta) 2 H VSMOW =-150.4‰，δ 18 O VSMOW =17.79‰)，USGS55(δ 2 H VSMOW = -28.2‰，δ 18 O VSMOW = 19.2‰)，USGS56(δ 2 H VSMOW =-44‰，δ 18 O VSMOW =27.23%o) for multipoint correction.
4. The method for discriminating a duck faeces aroma single tea harvesting season by stable isotope analysis technology according to claim 1, wherein the calculation formula of the stable isotope ratio is:
δE= [(R sample of /R Standard of )-1]；
Wherein E is the target isotope ratio, R Sample of For the abundance ratio of heavy isotope and light isotope in the tea sample to be measured, namely 2 H/ 1 H、 18 O/ 16 O；R Standard of The abundance ratio of heavy isotope to light isotope in the international universal standard sample; the delta value is multiplied by 1000 to be the unit per mill;
delta using the international standard VSMOW 2 H and delta 18 And (5) confirming the value of O.
5. An application of the method according to any one of claims 1-4 in discriminating duck feces single tea harvest seasons by stable isotope analysis technology, wherein the stable isotope analysis technology adopts a stable isotope ratio of hydrogen and oxygen.
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