CN114076806A - Fingerprint spectrum detection method of linseed oil and application thereof - Google Patents

Fingerprint spectrum detection method of linseed oil and application thereof Download PDF

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CN114076806A
CN114076806A CN202111265401.6A CN202111265401A CN114076806A CN 114076806 A CN114076806 A CN 114076806A CN 202111265401 A CN202111265401 A CN 202111265401A CN 114076806 A CN114076806 A CN 114076806A
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王进英
韩玉泽
王兴瑞
董国鑫
甘生睿
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Abstract

The invention discloses a fingerprint detection method of linseed oil and application thereof, wherein the detection method adopts a fingerprint to carry out detection and comprises the following steps: constructing the phytosterol control fingerprint spectrums of different varieties of linseed oil by adopting a gas chromatography-mass spectrometry combined method; the detection conditions of the gas chromatograph comprise: a chromatographic column: 5% phenyl-95% methylpolysiloxane column; the split ratio is as follows: 38-43: 1; temperature rising procedure: the initial temperature is 160-200 ℃, the temperature is kept for 0.5-2 min, the temperature is raised to 280-320 ℃ at the speed of 1-8 ℃/min, and the temperature is kept for 20-30 min. The method can be used for distinguishing and identifying different varieties of linseed oil, can realize the quick and effective identification of the linseed oil, and has certain significance for the quality control and adulteration identification of the linseed oil.

Description

Fingerprint spectrum detection method of linseed oil and application thereof
Technical Field
The invention belongs to the technical field of detection and analysis, and particularly relates to a fingerprint spectrum detection method of linseed oil and application thereof.
Background
Flax is one of the important special oil crops in China and is mainly distributed in northeast, northwest and northwest regions. Due to the special geographical and climatic characteristics of the plateau, the Qinghai flax becomes an oil crop with cultivation value. Due to reasonable fatty acid composition, rich nutrients and medical and health care effects, linseed oil is the first choice for many families, and the higher price also promotes the adulteration of linseed oil, so that a linseed oil adulteration identification method must be developed to control the product quality.
Different vegetable oils are distinguished primarily by their fatty acid composition. However, the fatty acid composition may be satisfactory when other, low-cost vegetable oils are blended with the vegetable oil. Therefore, a certain leak exists only by depending on the fatty acid composition to judge whether the vegetable oil is pure, and a riding opportunity is provided for illegal merchants to adulterate. In addition to fatty acids, trace nutrients such as vitamin E, phytosterols, squalene, carotenoids, etc. are present in vegetable oils. Among them, phytosterol is an important metabolite present in almost all animal and vegetable fats, and the content and composition of phytosterol mainly depend on plant species and also vary with species, geography, climatic conditions and processing techniques. Compared with other components such as fatty acid, the sterol has stronger characteristics, can represent the authenticity of oil and is a powerful tool for identifying the authenticity of the linseed oil.
At present, the sterol determination method in vegetable oil reported in literature mainly comprises gas chromatography, gas chromatography-mass spectrometry, high performance liquid chromatography and liquid chromatography-mass spectrometry, and scholars at home and abroad use chromatography technology and chemometrics to perform research on adulteration identification of vegetable oil. Xu et al (Characterization and analysis of four odoriferous oil using free phytosteriol profiles estableshied by GC-GC-TOF/MS [ J ]. AnalMethod,2014,6(17):12-19.) extract phytosterol from peanut oil, rapeseed oil, soybean oil and sunflower oil by solid phase extraction, and distinguish 4 edible vegetable oils by combining main component analysis, fractional cluster analysis and random forest statistical method after multi-dimensional gas chromatography tandem flight time mass spectrometry, and can identify 5% of soybean oil adulterated with pseudo-peanut oil by using free phytosterol as index. Al-Ismail et Al (Detection of olive oil addition with a needle plant oil by GLC analysis of sterol using polar column [ J ]. Food Chemistry,2010,121(4):1255-1259.) can identify that 5% of soybean oil, corn oil and sunflower seed oil are blended into olive oil according to the change of the sum of the contents of campesterol and stigmasterol before and after blending. However, the method has a single identification mode for adulteration samples and a limited application range, and the fingerprint spectrum technology can effectively represent the relationship between the complex components of natural substances and the quality of the complex components, can effectively research the overall situation of complex substance systems such as food and the like, and realizes the comprehensive evaluation of the quality and authenticity of products, so that the method is widely applied to food adulteration identification research.
Widely known incense and the like (research on corn germ oil adulteration detection technology based on fingerprint similarity [ J ]. Hunan agricultural science, 2019 (08)) adopt gas chromatography to establish a fatty acid fingerprint of corn oil, and detect the plant oil doped in the corn oil through similarity analysis; in the forest and the like (fingerprint spectra [ J ] of 4 edible vegetable oils are analyzed by gas chromatography in combination with chemometrics. analysis and test report, 2016,35 (04)), fatty acid compositions and contents of 4 types of vegetable oils (olive oil, peanut oil, rapeseed oil and soybean oil) are determined by gas chromatography, chromatographic fingerprint spectra of the vegetable oils are constructed, and the 4 types of vegetable oils are classified and identified by a pattern recognition method. The method judges whether the vegetable oil is pure or not by depending on the fatty acid composition, and has low accuracy.
Disclosure of Invention
The invention aims to provide a fingerprint detection method of linseed oil and application thereof, the method can be used for distinguishing and identifying different varieties of linseed oil, is particularly suitable for identifying and identifying the types of adulterated oil when the linseed oil is doped with rapeseed oil, peanut oil and sunflower oil with the concentration of more than 20% and is doped with sesame oil with the concentration of more than 30%, and can realize the quick and effective identification of the linseed oil.
In order to achieve the purpose, the invention adopts the following technical scheme:
a fingerprint detection method of linseed oil is characterized in that fingerprint detection is carried out, and the detection method comprises the following steps: (1) constructing the phytosterol control fingerprint spectrums of different varieties of linseed oil by adopting a gas chromatography-mass spectrometry combined method; the detection conditions of the gas chromatograph comprise:
a chromatographic column: 5% phenyl-95% methylpolysiloxane column;
the split ratio is as follows: 38-43: 1;
temperature rising procedure: the initial temperature is 160-200 ℃, the temperature is kept for 0.5-2 min, the temperature is raised to 280-320 ℃ at the speed of 1-8 ℃/min, and the temperature is kept for 20-30 min.
In a specific embodiment of the present invention, the detection conditions of the gas chromatography include:
a chromatographic column: 5% phenyl-95% methylpolysiloxane column;
the split ratio is as follows: 40: 1;
temperature rising procedure: the initial temperature is 180 deg.C, and the temperature is maintained for 1min, and the temperature is raised to 300 deg.C at 4 deg.C/min, and maintained for 25 min.
Further, the detection conditions of the gas chromatography further include one or more of the following i to v:
i specification of chromatographic column: 20 to 60m × 0.10 to 0.32mm × 0.18 to 0.50 μm;
ii carrier gas: an inert gas;
iii flow rate: 0.5-1.5 ml/min;
iv, sample inlet temperature: 300-335 ℃;
v sample size: 0.5-2 μ L.
In a specific embodiment of the present invention, the detection conditions of the gas chromatography further include one or more of the following i to v:
i specification of chromatographic column: 30 m.times.0.32 mm.times.0.50 μm;
ii carrier gas: helium gas;
iii flow rate: 1 ml/min;
iv, sample inlet temperature: 320 ℃;
v sample size: 1 μ L.
Further, the mass spectrometry conditions include one or more of the following i to v:
i ionization source: an EI ion source;
ii electron energy: 65-75 eV, preferably 70 eV;
iii ion source temperature: 240-260 ℃, preferably 250 ℃;
iv transmission line temperature: 290-320 ℃, preferably 300 ℃;
v ion scan range: 30 to 700 amu.
In a specific embodiment of the invention, the mass spectrometry conditions comprise one or more of the following i to v:
i ionization source: an EI ion source;
ii electron energy: 70 eV;
iii ion source temperature: 250 ℃;
iv transmission line temperature: 300 ℃;
v ion scan range: 50 to 650 amu.
Further, the fingerprint spectrum detection method of the linseed oil further comprises the following steps:
(2) determining similarity threshold values of linseed oil and a contrast fingerprint spectrum thereof through similarity analysis;
(3) and (3) carrying out similarity analysis on the fingerprint of the linseed oil sample to be detected and the comparison fingerprint of the phytosterol, and carrying out linseed oil identification by comparing the similarity threshold values of the linseed oil sample and the comparison fingerprint.
Further, the phytosterol comprises one or more of campesterol, stigmasterol, beta-sitosterol and cycloartenol;
further, the adulterant is one or more of rapeseed oil, peanut oil, sunflower oil and sesame oil.
The invention also provides an identification method of linseed oil adulteration, which is used for detecting and identifying the adulteration by combining a stoichiometric method; the chemometric method comprises one of cluster analysis and discriminant analysis.
Further, the discriminant analysis can identify one or more of rapeseed oil, peanut oil and sunflower oil with adulteration concentration of more than 20% and sesame oil with concentration of more than 30%.
The invention has the following beneficial effects:
(1) the identification method has good precision, repeatability and stability; through analyzing the composition of the linseed oil phytosterol, the fact that 40 linseed oils of the linseed oil contain 4 monomers of campesterol, stigmasterol, beta-sitosterol and cycloartenol is found, the beta-sitosterol content is the highest in the linseed oils of different production places, the cycloartenol is the second highest in the linseed oils, the stigmasterol content is the lowest, and the adulteration identification of the linseed oil is realized according to the characteristics of the different sterol contents in the linseed oils.
(2) The method is simple and easy to implement, has wide applicability and reliable result, and can realize the quick and effective identification of the linseed oil, thereby standardizing the market order of the edible oil, protecting the benefits and physical health of consumers and having obvious economic and social benefits.
Drawings
FIG. 1 is a chromatogram obtained by mixing standard solutions under different detection conditions;
FIG. 2 is a chromatogram of phytosterols from different vegetable oils;
FIG. 3 is a schematic representation of the phytosterol overlay of the linseed oil from Qinghai Linum;
FIG. 4 is a Qinghai linseed oil phytosterol standard fingerprint;
FIG. 5 is a dendrogram of clustering analysis of linseed oil samples and adulterated samples;
fig. 6 is a typical discriminant analysis diagram.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the accompanying drawings, and it should be understood that the described embodiments are some, but not all embodiments of the present invention. All other embodiments, which can be obtained by a person skilled in the art without any inventive step based on the embodiments of the present invention, are within the scope of the present invention.
Example 1
1 materials and methods
1.1 materials and reagents
40 kinds of flaxseed, rapeseed, peanut, sunflower seed, sesame (see table 1 for the production area information); cleaning the vegetable oil, removing impurities, baking at 160-180 deg.C for 20min, preparing oleum Lini, oleum Rapae, oleum Arachidis Hypogaeae, oleum Helianthi, and oleum Sesami by screw squeezing, centrifuging at 4500r/min for 15min, and storing at-4 deg.C.
And (3) standard substance: 5 alpha-cholestane-3 beta-alcohol, campesterol, stigmasterol, beta-sitosterol and cycloartenol reference substances (purity is more than or equal to 98%)
Ethyl acetate and n-hexane are used as chromatographic purity, potassium hydroxide, absolute ethyl alcohol and alkaline alumina are used as analytical purity.
TABLE 1 sample origin information
Figure BDA0003326763410000051
1.2 instruments and devices
QP 2020NX GC-MS instrument: shimadzu, Japan; magnetic force heating agitating unit: supelco, USA; JA1003 electronic balance: shanghai Liangping instruments and meters, Inc.; LKTC-B1-T digital display constant temperature water bath: guo hua electric appliances limited;
1.3 Experimental methods
1.3.1 solution preparation
Accurately weighing 10mg of stigmasterol, beta-sitosterol and cycloartenol respectively, and 20mg of campesterol and dihydrocholesterol respectively in a10 mL brown volumetric flask, adding ethyl acetate to a constant volume to scale, and preparing into 1mg/mL single-standard stock solution of stigmasterol, beta-sitosterol, cycloartenol, 2mg/mL campesterol and internal standard dihydrocholesterol.
And (4) transferring a proper amount of single-standard stock solution, gradually diluting the single-standard stock solution into a sterol mixed standard solution with a proper mass concentration by using ethyl acetate, and storing the sterol mixed standard solution in a refrigerator at 4 ℃.
1.3.2 sample pretreatment
Referring to the national standard method and slightly modifying, 0.5 +/-0.001 g of oil sample is precisely weighed into a ground triangular flask, 8mL of 0.5mol/mL KOH-ethanol solution and 0.5mL of 0.5mg/mL internal standard solution are added, the mixture is magnetically stirred at 80 ℃ and condensed and refluxed for 15min, and 8mL of absolute ethyl alcohol is added to dilute the sample solution when the reaction is finished. Adding 10g of alumina into 20mL of ethanol, pouring the suspension into a glass column, naturally settling the alumina, opening a piston to discharge the solvent, adding the prepared sample mixed solution when the liquid level reaches the top layer of the alumina, and opening the piston to discharge the solvent until the liquid level reaches the top layer of the alumina. Eluting unsaponifiable matter with 10mL of ethanol, eluting with 30mL of petroleum ether at a flow rate of about 2mL/min, concentrating the collected eluate by rotary evaporation to remove solvent, diluting the concentrated solution to 2mL, filtering with 0.45 μm filter membrane, and performing gas chromatography-mass spectrometry.
1.3.3 optimization of phytosterol assay conditions
Chromatographic conditions: a chromatographic column: inter Cap 5M/Sil capillary column (30 m.times.0.32 mm, 0.50 μ M); carrier gas: high purity helium (99.999% pure); flow rate: 1.0mL/min, split ratio: 40: 1; sample inlet temperature: the sample size is 1 mu L at 320 ℃; temperature rising procedure: the initial temperature is 180 deg.C, and the temperature is maintained for 1min, and the temperature is raised to 300 deg.C at 4 deg.C/min, and maintained for 25 min.
Mass spectrum conditions: an EI ionization source; electron energy 70 eV; ion source temperature: 250 ℃ and the transmission line temperature is 300 ℃. The ion scanning range is 50-650 amu.
Chromatographic conditions: a chromatographic column: inter Cap 5M/Sil capillary column (30 m.times.0.32 mm, 0.50 μ M); carrier gas: high purity helium (99.999% pure); flow rate: 1.0mL/min, split ratio: 20: 1; sample inlet temperature: the sample volume is 1 mu L at 280 ℃; temperature rising procedure: the initial temperature is 200 deg.C, and the temperature is maintained for 1min, and the temperature is increased to 300 deg.C at 10 deg.C/min, and maintained for 9 min.
Mass spectrum conditions: an EI ionization source; electron energy 70 eV; ion source temperature: 250 ℃ and the transmission line temperature is 250 ℃. The ion scanning range is 50-550 amu.
③ chromatographic conditions: a chromatographic column: inter Cap 5M/Sil capillary column (30 m.times.0.32 mm, 0.50 μ M); carrier gas: high purity helium (99.999% pure); flow rate: 1.0mL/min, split ratio: 10: 1; sample inlet temperature: the sample size is 1 mu L at 270 ℃; temperature rising procedure: the initial temperature is 100 ℃, the temperature is raised to 280 ℃ at the speed of 12 ℃/min, and the temperature is kept for 10 min.
Mass spectrum conditions: an EI ionization source; electron energy 70 eV; ion source temperature: 230 ℃ and a transmission line temperature of 280 ℃. The ion scanning range is 45-550 amu.
Chromatographic conditions: a chromatographic column: inter Cap 5M/Sil capillary column (30 m.times.0.32 mm, 0.50 μ M); carrier gas: high purity helium (99.999% pure); flow rate: 0.8mL/min, split ratio: 20: 1; sample inlet temperature: the sample size is 1 mu L at 320 ℃; temperature rising procedure: the initial temperature is 240 ℃, the temperature is kept for 5min, the temperature is increased to 255 ℃ at the speed of 4 ℃/min, and the temperature is kept for 30 min.
Mass spectrum conditions: an EI ionization source; electron energy 70 eV; ion source temperature: 250 ℃ and the transmission line temperature is 300 ℃. The ion scan range is 50-650 amu.
The chromatographic condition: a chromatographic column: inter Cap 5M/Sil capillary column (30 m.times.0.32 mm, 0.50 μ M); carrier gas: high purity helium (99.999% pure); flow rate: 1.0mL/min, split ratio: 40: 1; sample inlet temperature: the sample size is 1 mu L at 320 ℃; temperature rising procedure: the initial temperature is 180 deg.C, and the temperature is maintained for 1min, and the temperature is raised to 300 deg.C at 20 deg.C/min, and the temperature is maintained for 25 min.
Mass spectrum conditions: an EI ionization source; electron energy 70 eV; ion source temperature: 250 ℃ and the transmission line temperature is 300 ℃. The ion scan range is 50-650 amu.
Analyzing the samples respectively by adopting the determination conditions of firstly, secondly, thirdly, fourthly and fifthly, obtaining the chromatogram result graphs as shown in the figures 1a, b, c, d and e respectively, according to the results, the figure 1a can well separate a plurality of phytosterol types in the linseed oil, and the finally selected chromatogram condition is firstly.
1.3.4 construction of Qinghai linseed oil fingerprint
Processing oil sample according to 1.3.2 method, obtaining gas chromatogram of 40 batches of linseed oil phytosterol by GC-MS analysis, introducing chromatographic signals into a traditional Chinese medicine chromatography fingerprint similarity evaluation system (2004A) to obtain an original spectrum overlay chart, and obtaining 40 batches of linseed oil phytosterol comparison fingerprints by multipoint correction and automatic matching.
1.3.5 preparation of adulterated oil samples
Four kinds of common vegetable oil (rapeseed oil, peanut oil, sunflower oil and sesame oil) are respectively doped into the linseed oil according to different proportions (0%, 10%, 20%, 30%, 40% and 50%) to prepare adulterated linseed oil samples which are respectively named as CZ1-CZ5, HS1-HS5, KH1-KH5 and ZM1-ZM5, and the adulterated linseed oil samples are uniformly mixed and stored at low temperature.
1.3.6 data processing
All data were averaged over 3 sets of parallel experimental data, and the differences were analyzed using SPSS 23.0 software and plotted using Origin 2018.
2 results and analysis
2.1 qualitative and quantitative analysis of Qinghai Linseed oil sterols
According to the sample injection analysis under the chromatographic condition of 1.3.3, the better analysis effect can be achieved by mixing the phytosterol compounds in the standard product (figure 1), and the composition of the linseed oil phytosterol from campesterol, stigmasterol, beta-sitosterol and cycloartenol can be determined by carrying out qualitative analysis on the phytosterol in the linseed oil according to the appearance sequence and retention time of each component.
The phytosterol in the sample is quantitatively analyzed by adopting an internal standard method, a standard curve is drawn by taking the mass concentration ratio of the reference substance to the internal standard substance as an abscissa (X) and the peak area ratio of the reference substance to the internal standard substance as an ordinate (Y), and the result is shown in table 2.
TABLE 2 Linear regression equation and correlation coefficient for each component of mixed standard solution working solution
Figure BDA0003326763410000081
QH-32 (Haidongmin and 6) was selected for the four phytosterols for phytosterol methodological evaluation. And continuously injecting the liquid to be detected after sample pretreatment for 6 times according to the condition of 1.2.4, and detecting the peak area of each sterol component and the peak area of the internal standard in the oil sample for inspecting the precision. 6 groups of sample solutions to be tested are prepared in parallel according to the method of 1.3.2, and are subjected to sample injection analysis for inspecting repeatability. And (3) respectively placing the prepared oil sample at room temperature for 0, 2, 4, 8, 12 and 24 hours, then carrying out sample injection analysis, and calculating the RSD of the peak area of each sterol and the peak area of the internal standard at different time points for investigating the stability of the method. The results are shown in Table 3.
As can be seen from Table 3, the precision RSD of the sample is 0.73-1.76%, the repeatability RSD is 0.63-3.32%, and the stability RSD is 1.46-4.01%, which indicates that the gas chromatography mass spectrometer used in the experiment has good precision, and the method has good repeatability and stability and meets the requirements.
TABLE 3 precision test results for linseed oil phytosterol
Figure BDA0003326763410000082
2.2 five vegetable oil phytosterol composition analysis
The phytosterol is a main part of unsaponifiable matters in edible oil, is also a very important natural active ingredient, is widely present in various vegetable oils, nuts and plant seeds, and plays an important role in reducing blood cholesterol, inhibiting tumors, regulating immunity and maintaining body health. The composition and content of phytosterol in 40 batches of linseed oil were determined and the results are shown in table 4. Analysis shows that the linseed oil is rich in phytosterol, and 40 linseed oils contain 4 monomers of campesterol, stigmasterol, beta-sitosterol and cycloartenol. The linseed oil in different production areas has the highest beta-sitosterol content, the beta-sitosterol content is as high as 101.62-152.38 mg/100g and accounts for nearly 50% of total sterols, the cycloartenol content is 78.98-137.81 mg/100g, the campesterol content is 55.09-81.55 mg/100g, and the stigmasterol content is the lowest and is only 12.75-24.36 mg/100 g.
The total phytosterol content of the 40 linseed oils is 267.02-370.83 mg/100g, and the average content is 316.42mg/100 g. Moreover, QH-40 (Haidonghua 2) has the highest phytosterol content (370.83mg/100g) and QH-8 (6 in Xining hybrid) content (267.02mg/100g), indicating that both flaxseed varieties and production areas have a large impact on unsaponifiable content. From the analysis of the contents of campesterol and beta-sitosterol which are easy to be absorbed by human bodies, the contents of the campesterol and the beta-sitosterol in QH-40 (Haidonghua 2) are obviously higher than those of other linseed oils, thereby being beneficial to the effective absorption and utilization of the human bodies. In addition to the differences in phytosterol content caused by the genetic background, the harvest maturity stage also has some effect on the phytosterol enrichment in flaxseed.
TABLE 4 Phytosterol content in linseed oil
Figure BDA0003326763410000091
Figure BDA0003326763410000101
After 5 vegetable oils are processed according to the method of 1.3.2, 5 vegetable oil phytosterol chromatograms (figure 2) are obtained by GC-MS sample injection analysis, the sterol in the vegetable oil is quantified by an internal standard method and differential analysis is carried out, and the results are shown in Table 5. Campesterol, beta-sitosterol and stigmasterol are main common components of five edible vegetable oils, in terms of content, the beta-sitosterol accounts for the highest proportion in the vegetable oil, the percentage content of the beta-sitosterol is between 40.0 and 80.4 percent, and the campesterol and the stigmasterol are used secondarily.
Due to different plant species of different oil crops, the content of each sterol monomer is greatly different, except three total phytosterols (campesterol, stigmasterol and beta-sitosterol), cycloartenol which is not common in other vegetable oils also exists in linseed oil, and the difference with other vegetable oils is obvious (P is less than 0.05). Cycloartenol belongs to 4, 4' -dimethyl sterol, and research shows that cycloartenol can be used as a marked sterol of linseed oil to identify the quality of oil so as to prevent the oil from being adulterated. In addition, the content of beta-sitosterol and campesterol in rapeseed oil and sesame oil is the highest, wherein the content of beta-sitosterol accounts for more than 50% of the total sterol content in the two vegetable oils, and the determination results are consistent with the reports of the literature (i.e. the content of squalene and four phytosterols [ J ] in vegetable oil is determined by using a reversed phase polymer solid phase extraction-gas chromatography-mass spectrometry method, 2021,47(04):231 and 236.) in the food and fermentation industry. In the total amount, the sesame oil content is the highest and is 516.35mg/100g, while the peanut oil content is the lowest and is only 157.95mg/100g, which indicates that the types of raw materials are key factors influencing the content and proportion of phytosterol in vegetable oil, so that the effective distinction of linseed oil and vegetable oil such as rapeseed oil and soybean oil can be realized according to the content and proportion of each phytosterol.
TABLE 55 vegetable oil phytosterol composition and content
Figure BDA0003326763410000111
Note: capital letters indicate multiple comparison results of different phytosterols in the same vegetable oil, and lowercase letters indicate multiple comparison results of different phytosterols in different vegetable oils.
2.3 construction and application of Qinghai linseed oil sterol standard fingerprint
And (4) sequentially carrying out sample injection and determination on 40 batches of linseed oil samples, and recording a chromatogram. Introducing the obtained chromatogram signals into a traditional Chinese medicine chromatogram fingerprint similarity evaluation system to obtain a linseed oil phytosterol original spectrum superposition map (figure 3), selecting an S1 chromatogram as a reference map, setting the width of a time window to be 0.1, fitting the fingerprint by adopting an average vector method, obtaining a common mode (namely a comparison fingerprint R) of 40 batches of linseed oil fingerprints through the steps of multipoint correction, chromatogram peak matching, comparison generation and the like, and identifying each phytosterol peak in the comparison fingerprint through the retention time of a comparison product, wherein the result is shown in figure 4. Similarity is calculated by adopting an included angle cosine method, the similarity of 40 samples is 0.982-1, the linseed oil fingerprint comparison spectrum established by the method and the calibrated 4 peaks can reflect the fingerprint characteristics of the linseed oil phytosterol, and the method has identification and evaluation significance.
In the evaluation of the fingerprint applied to the authenticity identification, the average value or the minimum value of the similarity of different samples and the comparison fingerprint is generally used as a threshold value. In this study, the average of the similarity between 40 batches of linseed oil samples and the control fingerprint is 0.995, so that 0.995 can be determined as the similarity threshold of the Qinghai linseed oil, and the linseed oil with the similarity lower than 0.995 can be considered as the adulterated linseed oil.
The similarity analysis is carried out on the adulterated oil sample by applying a traditional Chinese medicine chromatogram fingerprint similarity evaluation system, and the result is shown in a table 6. The greater the adulteration concentration of the low-price vegetable oil is, the lower the similarity of the adulteration sample and the Qinghai pure linseed oil compared with the fingerprint is, and the similarity is lower than the similarity threshold value (0.995) of the Qinghai linseed oil, so that the adulteration oil sample and the pure linseed oil can be distinguished, and the identification of the adulteration linseed oil sample is realized.
TABLE 6 similarity of the control fingerprints between adulterated samples and linseed oil
Figure BDA0003326763410000121
2.4 identification of linseed oil adulteration by phytosterol composition in combination with chemometrics
2.4.1 Cluster analysis for distinguishing pure linseed oil from non-linseed oil
And (3) performing cluster analysis on 40 linseed oil samples and 20 adulterated samples by using group connection by using SPSS 23.0 software, and obtaining a cluster pedigree diagram consisting of the oil-like phytosterol by taking the squared Euclidean distance as a measurement standard. As can be seen from fig. 5, when the euclidean distance is 6, all oil samples are grouped into 6 types, 40 linseed oil samples are grouped into one type separately, and 20 adulterated linseed oil samples are grouped into five different types respectively, at which time 40 pure linseed oil samples can be distinguished from 20 adulterated samples.
2.4.2 discrimination analysis and identification of the type of the adulterated vegetable oil
Discriminant analysis is a common classification method and is widely applied to food authenticity identification. 40 batches of pure linseed oil are named as type 1(YMZ1-YMZ40), adulterated linseed oil mixed with different proportions of rapeseed oil, soybean oil, peanut oil, sunflower seed oil, corn oil and sesame oil are named as type 2(CZ1-CZ5), type 3(HS1-HS5), type 4(KH1-KH5) and type 5(ZM1-ZM5), and the phytosterol indexes of 5 types of edible oil samples are subjected to discriminant analysis by using SPSS 23.0, and the results are shown in FIG. 6 and Table 7.
TABLE 7 discriminant analysis Classification results
Figure BDA0003326763410000122
From the classification scatter plot obtained from the canonical discriminant function (see fig. 6), 40 pure linseed oils partially overlapped with the adulterated linseed oil sample. In 40 pure linseed oils, 6 samples are misjudged, the identification accuracy rate is 85%, and the identification accuracy rates among adulterated linseed oil samples doped with rapeseed oil, peanut oil, sunflower oil and sesame oil in different proportions are 75%, 77%, 69% and 59% respectively. Further analysis finds that the adulteration concentrations of the misjudged oil samples are mainly concentrated on 10% and 20%, the identification accuracy rates of the rapeseed oil, the peanut oil and the sunflower oil are respectively 89.5%, 85.5% and 96%, 97.5% and 96.5% under the higher adulteration concentration, the identification accuracy rate is still low when the sesame oil adulteration concentration is 30% and is 78.5%, the misjudged oil samples are mainly identified as that the sunflower oil is doped into the linseed oil, and the identification accuracy rate is 100% when the adulteration concentration is 40%, therefore, the discriminant analysis can identify that the rapeseed oil, the peanut oil and the sunflower oil are doped into the linseed oil at the concentration of more than 20%, and the vegetable oil type can be accurately identified when the sesame oil doping concentration reaches more than 30%.
In conclusion, the composition and content of the phytosterol in the linseed oil of Qinghai flax are analyzed by adopting solid-phase extraction and GC-MS (gas chromatography-mass spectrometry), and the four phytosterols, namely campesterol, stigmasterol, beta-sitosterol and cycloartenol, are detected in the linseed oil, wherein the cycloartenol is the linseed oil marker sterol. In the identification of the simulated adulterated linseed oil, software of a traditional Chinese medicine chromatographic fingerprint similarity evaluation system is utilized to establish a linseed oil phytosterol fingerprint, and 3 methods of similarity evaluation, cluster analysis and discriminant analysis are combined to perform mode identification on the linseed oil sample and the adulterated sample phytosterol fingerprint. The similarity evaluation result shows that the fingerprints of 40 batches of linseed oils of different varieties and different production places have good consistency, the similarity of the adulterated sample and the comparison fingerprint is lower than the similarity threshold (0.995) of the Qinghai linseed oil, and the similarity evaluation and the cluster analysis can realize the distinction of the pure linseed oil sample and the adulterated sample. When the proportion of the rapeseed oil, the peanut oil and the sunflower oil doped in the linseed oil is more than 20 percent and the proportion of the sesame oil doped in the linseed oil is more than 30 percent, the discriminant analysis model can accurately identify the type of the adulterated vegetable oil. Therefore, the phytosterol fingerprint spectrum is combined with chemometrics analysis, and can be used for the adulteration identification and quality control of the Qinghai linseed oil.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.

Claims (10)

1. A fingerprint detection method of linseed oil is characterized in that fingerprint detection is carried out, and the detection method comprises the following steps: (1) constructing the phytosterol control fingerprint spectrums of different varieties of linseed oil by adopting a gas chromatography-mass spectrometry combined method; the detection conditions of the gas chromatograph comprise:
a chromatographic column: 5% phenyl-95% methylpolysiloxane column;
the split ratio is as follows: 38-43: 1;
temperature rising procedure: the initial temperature is 160-200 ℃, the temperature is kept for 0.5-2 min, the temperature is raised to 280-320 ℃ at the speed of 1-8 ℃/min, and the temperature is kept for 20-30 min.
2. The fingerprint detection method according to claim 1, wherein the detection conditions of the gas chromatograph include:
a chromatographic column: 5% phenyl-95% methylpolysiloxane column;
the split ratio is as follows: 40: 1;
temperature rising procedure: the initial temperature is 180 deg.C, and the temperature is maintained for 1min, and the temperature is raised to 300 deg.C at 4 deg.C/min, and maintained for 25 min.
3. The fingerprint detection method according to claim 1, wherein the detection conditions of the gas chromatography further comprise one or more of the following i to v:
i specification of chromatographic column: 20 to 60m × 0.10 to 0.32mm × 0.18 to 0.50 μm;
ii carrier gas: an inert gas;
iii flow rate: 0.5-1.5 ml/min;
iv, sample inlet temperature: 300-335 ℃;
v sample size: 0.5-2 μ L.
4. The fingerprint detection method according to claim 3, wherein the detection conditions of the gas chromatography further include one or more of the following i to v:
i specification of chromatographic column: 30 m.times.0.32 mm.times.0.50 μm;
ii carrier gas: helium gas;
iii flow rate: 1 ml/min;
iv, sample inlet temperature: 320 ℃;
v sample size: 1 μ L.
5. The fingerprint detection method of claim 1, wherein the mass spectrometry conditions comprise one or more of the following i-v:
i ionization source: an EI ion source;
ii electron energy: 65-75 eV, preferably 70 eV;
iii ion source temperature: 240-260 ℃, preferably 250 ℃;
iv transmission line temperature: 290-320 ℃, preferably 300 ℃;
v ion scan range: 30 to 700 amu.
6. The fingerprint detection method of claim 5, wherein the mass spectrometry conditions comprise one or more of the following i-v:
i ionization source: an EI ion source;
ii electron energy: 70 eV;
iii ion source temperature: 250 ℃;
iv transmission line temperature: 300 ℃;
v ion scan range: 50 to 650 amu.
7. The fingerprint detection method according to any one of claims 1 to 6, further comprising the steps of:
(2) determining similarity threshold values of linseed oil and a contrast fingerprint spectrum thereof through similarity analysis;
(3) and (3) carrying out similarity analysis on the fingerprint of the linseed oil sample to be detected and the comparison fingerprint of the phytosterol, and carrying out linseed oil identification by comparing the similarity threshold values of the linseed oil sample and the comparison fingerprint.
8. The fingerprint detection method according to claim 1, wherein the phytosterol comprises one or more of campesterol, stigmasterol, beta-sitosterol, cycloartenol;
further, the adulterant is one or more of rapeseed oil, peanut oil, sunflower oil and sesame oil.
9. A method for identifying linseed oil adulteration, which adopts the fingerprint detection method of any one of claims 1-8 to detect and identifies the adulteration by combining a chemometry method; the chemometric method comprises one of cluster analysis and discriminant analysis.
10. The method according to claim 9, wherein the discriminant analysis can identify one or more of rapeseed oil, peanut oil and sunflower oil with a adulteration concentration of 20% or more and sesame oil with a concentration of 30% or more.
CN202111265401.6A 2021-10-28 2021-10-28 Fingerprint spectrum detection method of linseed oil and application thereof Pending CN114076806A (en)

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