CN109917028B - The method for building up and detection method of agricultural chemical compound electronic identity database in edible agricultural product based on LC-Q-Orbitrap - Google Patents
The method for building up and detection method of agricultural chemical compound electronic identity database in edible agricultural product based on LC-Q-Orbitrap Download PDFInfo
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/86—Signal analysis
- G01N30/8696—Details of Software
Abstract
Agricultural chemical compound electronic identity database and detection method in the invention discloses a kind of edible agricultural product based on LC-Q-Orbitrap.Electronic identity database includes set, intelligent Matching value and the collision energy of Multiple Pesticides compound electronic ID card information, and according to being ranked up in electronic ID card by retention time, wherein electronic ID card includes agricultural chemical compound information, retention time, adduct ion information, fragment ion information, collision energy and optimal full scan mass spectrogram；Detection method include sample pre-treatments step, LC-Q-Orbitrap operating condition setting and sample in pesticide residue screening process；Wherein LC-Q-Orbitrap operating condition includes that suitable chromatographic condition and Mass Spectrometry Conditions are arranged, agricultural chemical compound electronic identity database is searched first with retention time during pesticide screening, corresponding electronic ID card information is extracted if matching, then intelligent matching value, if they are the same and recording and displaying is as a result, screening is completed.
Description
Technical field
The present invention designs agricultural chemical compound electronic identity database and inspection in the edible agricultural product based on LC-Q-Orbitrap
Survey method, the invention are able to achieve for the remaining non-target, multi objective of 500 Multiple Pesticides in multiple eating agricultural product, quick sieve
It looks into.
Background technique
Altogether early in the World Health Organization (WHO) in 1976, food and agricultural organization (FAO) and United Nations Environment Programme (UNEP)
It is same to establish global environment detection system/food item (Global Environment Monitoring System, GEMS/
Food), it is intended to grasp member state's food pollution situation, understand food contaminant intake, protect human health, give a impetus to trade hair
Exhibition.Now, countries in the world all promote food safety the strategic position to national security.Pesticide Residue is food safety mark
One of standard and international trade access threshold.Meanwhile to show kind more and more for the requirement to pesticide residue, limit the quantity more next
Stringenter development trend, that is, the Pesticide Residue threshold that international trade is set up are higher and higher.For example, European Union, Japan
It has been formulated respectively with the U.S. 169068 (481 kinds of pesticides), 44340 (765 kinds of pesticides), 13055 (395 kinds of pesticides) agricultures
Medicine residue limits standard, China have issued 4140 MRL standards of 433 kinds of pesticides for 2016.Currently, generally use in the world
Uniform limit limitation is 10 μ g/kg.Therefore, high-throughput quickly Detecting Pesticide skill is all called in food safety and international trade
Art, this undoubtedly also provides opportunities and challenges to vast pesticide residue analysis worker.In current numerous pesticide residue analysis
In technology, hydrolysis and condensation is to realize that high throughput remains the optimized analysis means quickly detected more.
Pesticide residue analysis is mostly with gas-chromatography, liquid chromatography, gas chromatography-mass spectrum and liquid phase chromatogram-mass spectrometry combination at present
Based on technology.These detection techniques all carry out firstly the need of standard sample of pesticide control qualitative.For example, the inspection to 100 kinds of pesticides
Survey just needs to prepare corresponding 100 kinds of standard sample of pesticide control, and the pesticide except this 100 kinds can be all missed.It is residual in pesticide
It stays in the real work in laboratory, most laboratories do not reserve hundreds of standard sample of pesticide, the reason is that pesticide mark
Quasi- product are not only expensive, but also validity period only has 2,3 years, needs overlapping investment.The standing standard sample of pesticide of common laboratory is only
There are tens kinds, the pesticide species of daily monitoring are also just only limited to this tens kinds, thereby result in food safety monitoring loophole.
The present inventor team is based on LC-Q-Orbitrap, has developed the essence of 500 Multiple Pesticides by concentrating on studies for many years
True mass spectrometry database and pesticide residue screening technical method, realize and are not required to standard control, can be simultaneously to agricultural production
500 Multiple Pesticides remain rapid screening in product, meet the urgent need that pesticide residue high throughput quickly detects in Agricultural Products.
Summary of the invention
The present invention is for cannot achieve in current pesticide residue screening technical method to Multiple Pesticides while quickly detecting
Problem develops a kind of based on high throughput high resolution liquid chromatography-quadrupole rod-electrostatic field Orbitrap mass (LC-Q-
Orbitrap) the edible agricultural product agricultural chemical compound electronic identity database and detection method of new technology, realizes the standard of being not required to
Product control it is residual can to meet pesticide in Agricultural Products simultaneously to 500 Multiple Pesticides residual rapid screening detection in agricultural product
Stay the high-throughput urgent need quickly detected.
Technical scheme is as follows:
A kind of electronic identity database of the agricultural chemical compound based on LC-Q-Orbitrap, including multiple agricultural chemical compounds
Electronic ID card, the electronic ID card include agricultural chemical compound information, retention time, adduct ion information, fragment ion letter
Breath, collision energy and optimal full scan mass spectrogram；
The agricultural chemical compound information includes compound name, compound molecule formula；
By LC-Q-Orbitrap instrument in Full MS/ddMS2The agricultural chemical compound is measured under mode specifies chromatography
Retention time under Mass Spectrometry Conditions determines ionized form (+H ,+NH under the source the agricultural chemical compound ESI4,+Na) and change
Adduct molecule formula obtains the accurate mass number of agricultural chemical compound adduct ion；
Under multiple different collision energies, the full scan mass spectrogram of fragment ion is acquired, selects ion informative most
Excellent full scan mass spectrogram, the optimal full scan mass spectrogram refer to that adduct ion abundance ratio is 10%-20%, described optimal complete
It scans selection abundance of ions in mass spectrogram and records the impact energy magnitude than maximum 3-5 fragment ion；
The fragment ion information be fragment ion theory accurate mass number under the optimal full scan mass spectrogram and
Abundance ratio；
Signal intensity ratio of the abundance than referring to the most strong fragment ion of the ion and signal in mass spectrogram.
The database by retention time according to being ranked up in electronic ID card.
Further, the electronic identity database further includes intelligent Matching model, and Matching Model is in the electronic ID card
Middle increase intelligent Matching value Pm, computation model are as follows:
Wb+Wq=1；
Wherein MbFor the theoretical accurate mass number for summing it up ion, MiFor the accurate mass number of i-th of fragment ion, WiIt is
The weight of i fragment ion, IiFor the abundance of ions ratio of i-th of fragment ion, fragment ion puts in order as abundance of ions
Ratio is from big to small；WbFor the weight for summing it up ion, WqFor the comprehensive weight of fragment ion, n is the number of fragment ion.
Further, the Wb, WqIt can be adjusted according to the variation of intelligent Matching model, general value is Wb=Wq=
0.5。
Further, the determination method of the theoretical accurate mass number of the fragment ion are as follows:
1) according to compound molecule formula, fragment ion element composition is specified；
2) according to the mass number M of fragment ion in mass spectrogram, possible fragment ion element group is obtained in column by calculating
Table；
Wherein: XiFor i-th of fragment ion element mass number, n is first prime number of fragment ion, yiFor i-th of fragment ion
The number of corresponding element.
3) it is made up of the splitting mechanism of molecular structure from fragment ion element and selects reasonable fragment ion member in list
Element composition, and calculate its theoretical accurate mass M '.
M '=X1y′1+X2y′2+…+Xny′n
Wherein: X1、X2……XnFor the fragment ion element mass number, y '1、y′2、……y′nFor preferred fragment ion
The number of the corresponding element of element composition.
Further, the chromatographic mass spectrometry condition are as follows:
Chromatographic condition: being separated by liquid chromatographic system, equipped with reverse-phase chromatographic column (Accucore aQ 150 ×
2.1mm, 2.6 μm)；Mobile phase A is -0.1% formic acid of ammonium acetate-water of 5mM；Mobile phase B is 0.1% formic acid-methanol；Gradient is washed
De- program, 0min:1%B, 3min:30%B, 6min:40%B, 9min:40%B, 15min:60%B, 19min:90%B,
23min:90%B, 23.01min:1%B run 4min afterwards；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample volume: 5 μ L；
Mass Spectrometry Conditions: scan pattern: Full MS-ddMS2；Full MS scan range:70-1050m/z；
Resolution:70,000, Full MS；17,500,MS/MS；AGC:Full MS, 1e6；MS/MS,1e5；Max IT:Full
MS,200ms；MS/MS,60ms；Loop count:1；MSX count:1；Isolation width:2.0m/z；NCE
(Stepped NCE): 40 (50%)；Under fill ratio:1%；Apex trigger:2-6s；Dynamic
Exclusion:5s；Mass spectrum testing result is acquired and handled by TraceFinder software.
Agricultural chemical compound detection method in a kind of edible agricultural product based on LC-Q-Orbitrap, comprising:
1) sample to be tested Acidifying acetonitrile is homogenized and is extracted, after being dehydrated and be centrifuged, being concentrated, then through solid-phase extraction column
(SPE) it purifies, acetonitrile+toluene solution elutes remains of pesticide, and sample solution is made after concentrated, filtering；
2) by LC-Q-Orbitrap instrument in Full MS/ddMS2Sample solution is obtained under mode specifies chromatographic mass spectrometry
Under the conditions of chromatogram；
3) by LC-Q-Orbitrap instrument in Full MS/ddMS2Sample solution is obtained under mode specifies chromatographic mass spectrometry
Under the conditions of chromatogram and mass spectrogram, to obtain the accurate mass number information of the retention time and adduct ion in chromatogram,
And the fragment ion and mass spectrogram obtained under corresponding optimum collision energy, and record the electricity of the corresponding unknown material of the retention time
Sub- identity card；
4) successively by the electronic ID card of each agricultural chemical compound in unknown material electronic ID card and electronic identity database
Compare, if Δ T≤0.3 and Δ P≤10%, record the agricultural chemical compound, if Δ T≤0.3 and 10% < Δ P≤
30%, then judge whether to include the agricultural chemical compound by the height of mass spectra peak in mass spectrogram and registration degree of certainty, otherwise into
Enter the comparison of next unknown material electronic ID card；
5) detection is completed, and shows the information of contained agricultural chemical compound in sample solution；
Wherein Δ T is the absolute of unknown material retention time and the difference of agricultural chemical compound retention time any in database
Value,Wherein PcFor unknown material intelligent Matching value, PiAny agricultural chemical compound intelligent Matching value in database.
Further, the sample further includes following pre-treatment:
10.0g sample (being accurate to 0.01g) is weighed in 100mL centrifuge tube, 30~40mL Acidifying acetonitrile extracting solution is added,
10000~11000rpm homogeneous, 1~2min；Anhydrous magnesium sulfate and sodium chloride (mass ratio, 4:1) is added, vibrates 8~10min；
After 4200rpm is centrifuged 5~7min, take 15~20mL supernatant in 150 mL chicken heart bottles, 40 DEG C of heating water bath rotary evaporations to 1
~2mL, it is to be clean；
Using CarbonNH2 column, the anhydrous sodium sulfate of about 1~2cm high is added in CarbonNH2 column, with 5~6mL second
Nitrile-toluene solution prewashing SPE decontaminating column, while bubble in decontaminating column discharge column is touched, efflux is thrown aside below decontaminating column.To liquid
When face is slightly above at the top of sodium sulphate, concentrate is transferred to decontaminating column, under connect 50mL chicken heart bottle.It is molten with 2~3mL acetonitrile-methylbenzene
Liquid rinses the chicken heart bottle for speckling with sample, and cleaning solution is transferred in decontaminating column, repeats 2~3 times；25mL liquid storage device is connect on column, with
The elution of 25~30mL acetonitrile-methylbenzene solution.For rotary evaporation to about 0.5mL, nitrogen is blown to close dry, addition 1mL acetonitrile-after collection
Aqueous solution is filtered through 0.22 μm of nylon membrane after ultrasonic dissolution.
Beneficial effects of the present invention:
1, the electronic ID card information of agricultural chemical compound is obtained using LC-Q-Orbitrap technology and utilize electronics body
Part card information assembly agricultural chemical compound electronic identity database, innovatively establishing 500 Multiple Pesticides accurate masses can reach
The database of 0.00001m/z, with electronic standard replace pesticide material standard, realize high-precision, high efficiency, economize on resources it is non-
Target Detecting Pesticide.Pesticide material standard is replaced to do reference by the screening method of electronic standard of pesticide electronic ID card
Conventional identification method realizes the development that jumps of non-target the Detection Technologies of Pesticide Residues.
2, it innovatively establishes with compounds Information in Mass Spectra such as high-resolution accurate mass number, abundance of ions ratios as identification mark
Quasi-, according to agricultural chemical compound electronic identity database 500 Multiple Pesticides are carried out with the LC-Q-Orbitrap of screening and confirmation
Technical method；This technology revolutionizes original using compound standard object as the qualitative model of reference, is that one kind does not need to mark
Quasi- object control, quick, high throughput, accurately and reliably Detecting Pesticide new technology.It eliminates standard items and does reference, and with electronics
Criteria Qualitative identification, realizes the conventional method that material standard is replaced with electronic ID card, while also achieving and detecting from targeting
To the tech development of non-targeted screening.Resource is saved, pollution is reduced, analysis speed is improved, fullys meet green
Development, environmental-friendly and clean and effective requirement.
3, when the LC-Q-Orbitrap residual screening technical method that the present invention establishes can be according to the reservation of target compound
Between, the information such as accurate mass number, abundance of ions ratio, collision energy, by with chemical combination in agricultural chemical compound electronic identity database
The corresponding informance of object, which is retrieved, to be compared, and the matching degree of target compound is provided.According to the match condition of target compound, realize to agriculture
The qualitatively screening of medicine.It is innovative in the database to increase collision energy, it is realized by the adjustment of collision energy to optimal complete
Scan mass spectrogram acquisition and data extract, improve the accuracy of data, optimum collision energy selection when, select adduction from
Sub- abundance ratio is that the mass spectrogram of 10%-20% is optimal mass spectrogram, and it is related broken both to ensure that adduction ion produces after colliding in this way
Piece ion also guarantees the presence of adduction ion.
5, the present invention establish LC-Q-Orbitrap residual screening technical method, using Full MS/dd MS2 mode into
Sample analysis can be obtained chromatogram and mass spectrum under the specified chromatography Mass Spectrometry Conditions of 500 Multiple Pesticides by single injected sampling analysis
Figure, shortens sample analysis time, improves sample detection efficiency.
6, the LC-Q-Orbitrap residual screening technical method that the present invention establishes, while in 500 Multiple Pesticides of screening,
Pesticide screening sensitivity more than 80% is lower than 10 micro- g kgs of uniform limit, betters meet various countries pesticide residue MRL level
The requirement of screening, this screening technical quality precision significantly reduce false positive detection as a result, preferable within 5ppm
Meet more residuals, high-precision pesticide residue screening requirement.
7, the present invention is the intelligent Matching value that every kind of compound calculates fast automatic comparison, and intelligent Matching value has taken into account essence
The information of true mass number and abundance of ions ratio, and according to the abundance of ions of differentiation between adduction ion and different fragment ions than prominent
It goes on business the influence away from biggish fragment ion, original basis is changed by the introducing of intelligent Matching value and is artificially judged not
Foot can be realized accurate braking matching, really realize the automation of detection.
Detailed description of the invention
Fig. 1 LC-Q-Orbitrap chemistry of pesticide polluter electronic identity model of a syndrome
[M+H]+first mass spectrometric figure of Fig. 2 Benalaxyl
Fig. 3 Benalaxyl [M+H]+normalization method energy NCE typical second order ms figure when being 20
Fig. 4 Benalaxyl [M+H]+normalization method energy NCE typical second order ms figure when being 40
Fig. 5 Benalaxyl [M+H]+normalization method energy NCE typical second order ms figure when being 60
Fig. 6 Benalaxyl [M+H]+ladder normalization method energy Step NCE typical second order ms when being 20,40,60
Figure
Fig. 7 LC-Q-Orbitrap pesticide residue screening process
Specific embodiment
The present invention will be further explained below with reference to the attached drawings and specific examples.
Fig. 1 shows LC-Q-Orbitrap chemistry of pesticide pollutant mass spectrometry database Establishing process, Summary is detailed
It describes in detail bright, below by taking Benalaxyl as an example, describes in detail to the establishment process of agricultural chemical compound electronic ID card:
Chromatographic condition: being separated by liquid chromatographic system, equipped with reverse-phase chromatographic column (Accucore aQ 150 ×
2.1mm, 2.6 μm)；Mobile phase A is -0.1% formic acid of ammonium acetate-water of 5mM；Mobile phase B is 0.1% formic acid-methanol；Gradient is washed
De- program, 0min:1%B, 3min:30%B, 6min:40%B, 9min:40%B, 15min:60%B, 19min:90%B,
23min:90%B, 23.01min:1%B run 4min afterwards；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample volume: 5 μ L；
Mass Spectrometry Conditions: scan pattern: Full MS-ddMS2；Full MS scan range:70-1050m/z；
Resolution:70,000, Full MS；17,500,MS/MS；AGC:Full MS, 1e6；MS/MS,1e5；Max IT:Full
MS,200ms；MS/MS,60ms；Loop count:1；MSX count:1；Isolation width:2.0m/z；NCE
(Stepped NCE): 40 (50%)；Under fill ratio:1%；Apex trigger:2-6s；Dynamic
Exclusion:5s；Mass spectrum testing result is acquired and handled by TraceFinder software.
In Full MS/dd MS2Its solvent standard is measured under mode, molecular formula C20H23NO3 extracts it
Primary information finds that its adduct ion is [M+H]+Peak, accurate mass number are shown in Fig. 2 for 326.17507.In order to understand
The secondary fragment information of Benalaxyl, respectively at NCE=20 (see Fig. 3), NCE=40 (see Fig. 4), NCE=60 (see Fig. 5) and
StepNCE=20,40,60 (see Fig. 6) Shi Yunhang acquisition methods, according to the chemical property of Benalaxyl, in conjunction with it in difference
Second level spectrogram under NCE may infer that the secondary fragment of its 5 practical measurements is respectively 148.11212,91.05415,
121.08865,208.13303 and 294.14871 can be to its 5 secondary fragments to integrated structure formula information and molecular formula
Theoretical value be determined, respectively 148.11208 (C10H14N, abundance is than 100.00%), 91.05423 (C7H7, abundance ratio
85.34%), 121.0886 (C8H11N, abundance is than 47.17%), 208.13364 (C12H18O2N, abundance than 13.40%) and
294.14886(C19H20O2N, abundance is than 5.65%).Calculate the intelligent Matching value P of BenalaxylmAre as follows:
Its electronic ID card is established according to Fig. 1 and is stored in electronic identity database.
In Fig. 7 expression Pesticides Testing electronization method proposed by the present invention, sample is once prepared, it can screening more than 500 simultaneously
Kind pesticide；It eliminates standard items and does reference, and with electronic standard Qualitative Identification, it realizes and material standard is replaced with electronic ID card,
It is also achieved simultaneously from targeting detection to the tech development of non-targeted screening.Resource is saved, pollution is reduced, is improved point
Speed is analysed, Green Development, environmental-friendly and clean and effective requirement are fully meet.
Table 1 gives in LC-Q-Orbitrap electronic identity database and represents agricultural chemical compound electronic ID card example for 5 kinds
(being free of molecular formula), table 2 gives 500 Multiple Pesticides inventories in LC-Q-Orbitrap electronic identity database.
5 kinds of 1 LC-Q-Orbitrap of table represents agricultural chemical compound electronic ID card example (without molecular formula)
2 500 Multiple Pesticides inventory of table
Embodiment
Embodiment 1
(pesticide as indicated above) the LC-Q-Orbitrap screening of 500 Multiple Pesticides and confirmation technology are real in commercially available apple
Apply example.
1, the specific steps of Sample Pretreatment Technique:
1.1 apple samples take edible part to shred, and mix, and label is indicated in sealing；
1.2 weigh 10g apple sample (being accurate to 0.01g), in 100mL centrifuge tube, 40mL1% acetate acetonitrile are added and mentions
Take liquid, 10000rpm homogeneous 1min.Anhydrous magnesium sulfate and sodium chloride (mass ratio, 4:1) is added, vibrates 10min.4200rpm from
After heart 5min, take 20mL supernatant in 150mL chicken heart bottle, 40 DEG C of heating water bath rotary evaporations are to be clean to 1mL；
1.3 are added the anhydrous sodium sulfate of about 2cm high in SPE column, with 5mL acetonitrile-methylbenzene solution prewashing SPE decontaminating column,
Bubble in decontaminating column discharge column is touched simultaneously, and efflux is thrown aside below decontaminating column.It, will be dense when liquid level is slightly above at the top of sodium sulphate
Contracting liquid is transferred to decontaminating column, under connect 50mL chicken heart bottle.The chicken heart bottle for speckling with sample is rinsed with 2mL acetonitrile-methylbenzene solution, will be washed
Liquid is transferred in decontaminating column, is repeated 2 times；25mL liquid storage device is connect on column, with the elution of 25mL acetonitrile-methylbenzene solution.After collection
Rotary evaporation is to about 0.5mL.
1.4 nitrogen are blown to close dry, addition 1mL acetonitrile-aqueous solution, filter after ultrasonic dissolution through 0.22 μm of nylon membrane, for LC-Q-
Orbitrap detection.
2, LC-Q-Orbitrap operating condition
Compound is separated by liquid chromatographic system, equipped with reverse-phase chromatographic column (Accucore aQ 150 ×
2.1mm, 2.6 μm)；Mobile phase A is -0.1% formic acid of ammonium acetate-water of 5mM；Mobile phase B is 0.1% formic acid-methanol；Gradient is washed
De- program, 0min:1%B, 3min:30%B, 6min:40%B, 9min:40%B, 15min:60%B, 19min:90%B,
23min:90%B, 23.01min:1%B run 4min afterwards；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample volume: 5 μ L.
Mass Spectrometry Conditions: scan pattern: Full MS-ddMS2；Full MS scan range:70-1050m/z；
Resolution:70,000, Full MS；17,500,MS/MS；AGC:Full MS, 1e6；MS/MS,1e5；Max IT:Full
MS,200ms；MS/MS,60ms；Loop count:1；MSX count:1；Isolation width:2.0m/z；NCE
(Stepped NCE): 40 (50%)；Under fill ratio:1%；Apex trigger:2-6s；Dynamic
Exclusion:5s.Mass spectrum testing result is acquired and handled by TraceFinder software, apple sample is obtained and specifies chromatography
Chromatogram under Mass Spectrometry Conditions.
3, the accurate mass number of the retention time in extraction chromatography figure and corresponding adduction ion in order, retrieves electronic identity
In database, the electronic ID card information of record and the retention time and corresponding adduction exact mass of ion number then uses data
Corresponding collision energy bombards to obtain mass spectrogram in library, establishes the electronics of the corresponding doubtful pesticide of apple sample of all retention times
Identity card；
4) successively by each agricultural chemical compound in the electronic ID card of the doubtful pesticide of apple sample and electronic identity database
Electronic ID card compare, if Δ T≤0.3 and Δ P≤10%, record the agricultural chemical compound, otherwise enter it is next
The comparison of the electronic ID card of doubtful pesticide；
5) detection is completed, and shows the information of contained agricultural chemical compound in sample solution；
LC-Q-Orbitrap screening results in certain provincial capital's apple sample:
18, the commercially available apple sample of certain provincial capital is acquired, 500 Multiple Pesticides are remained using LC-Q-Orbitrap technology
Screening is carried out, 15 kinds of pesticide residues are detected, amounts to 62 frequencys, is related to 14, sample, concrete outcome is shown in Table 3.
Pesticide residue LC-Q-Orbitrap screening results in 3 somewhere apple sample of table
Embodiment 2
(pesticide as indicated above) the LC-Q-Orbitrap screening of 500 Multiple Pesticides and confirmation technology are real in commercially available lemon
Apply example.
Pesticide residue screening process is referring to real in sample pre-treatments step, LC-Q-Orbitrap operating condition and sample
Apply example 1.
LC-Q-Orbitrap screening results in certain provincial capital's lemon sample:
13, the commercially available lemon sample of certain provincial capital is acquired, 500 Multiple Pesticides are remained using LC-Q-Orbitrap technology
Screening is carried out, 9 kinds of pesticide residues are detected, amounts to 53 frequencys, is related to 10, sample, concrete outcome is shown in Table 4.
Pesticide residue LC-Q-Orbitrap screening results in 4 somewhere lemon sample of table
Embodiment 3
(pesticide as indicated above) the LC-Q-Orbitrap screening of 500 Multiple Pesticides and confirmation skill in commercially available cabbage
Art embodiment.
Pesticide residue screening process is referring to real in sample pre-treatments step, LC-Q-Orbitrap operating condition and sample
Apply example 1.
LC-Q-Orbitrap screening results in certain provincial capital's cabbage sample:
25, the commercially available cabbage sample of certain provincial capital is acquired, using LC-Q-Orbitrap technology to more than 500 kinds of agricultures
Medicine residual carries out screening, detects 18 kinds of pesticide residues, amounts to 121 frequencys, is related to 21, sample, concrete outcome is shown in Table 5.
Pesticide residue LC-Q-Orbitrap screening results in 5 somewhere cabbage sample of table
The series of detailed descriptions listed above only for feasible embodiment of the invention specifically
Protection scope bright, that they are not intended to limit the invention, it is all without departing from equivalent implementations made by technical spirit of the present invention
Or change should all be included in the protection scope of the present invention.
Claims (6)
1. the foundation side of the electronic identity database of agricultural chemical compound in a kind of edible agricultural product based on LC-Q-Orbitrap
Method, which is characterized in that the database includes multiple agricultural chemical compound electronic ID cards, and the electronic ID card includes pesticide
Close object information, retention time, adduct ion information, fragment ion information, collision energy and optimal full scan mass spectrogram；
The agricultural chemical compound information includes compound name, compound molecule formula；
By LC-Q-Orbitrap instrument in Full MS/ddMS2The agricultural chemical compound is measured under mode specifies chromatographic mass spectrometry item
Retention time under part determines+H ,+NH under the source the agricultural chemical compound ESI4,+Na and compound molecule formula, obtain pesticide
The accurate mass number of compound adduct ion；The chromatographic mass spectrometry condition are as follows:
Chromatographic condition: being separated by liquid chromatographic system, is furnished with reverse-phase chromatographic column Accucore 150 × 2.1mm of aQ,
2.6μm；Mobile phase A is -0.1% formic acid of ammonium acetate-water of 5mM；Mobile phase B is 0.1% formic acid-methanol；Gradient elution program,
0min:1%B, 3min:30%B, 6min:40%B, 9min:40%B, 15min:60%B, 19min:90%B, 23min:90%
B, 23.01min:1%B run 4min afterwards；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample volume: 5 μ L；
Mass Spectrometry Conditions: scan pattern: Full MS-ddMS2；Full MS scan range:70-1050m/z；Resolution:
70,000, FullMS；17,500, MS/MS；AGC:FullMS, 1e6；MS/MS, 1e5；Max IT:Full MS, 200ms；MS/
MS, 60ms；Loop count:1；MSX count:1；Isolation width:2.0mz；NCE:40；Under fill
Ratio:1%；Apex trigger:2-6s；Dynamic Exclusion:5s；Mass spectrum is examined by TraceFinder software
Survey results acquisition and processing；
Under multiple different collision energies, the full scan mass spectrogram of fragment ion is acquired, selects ion informative optimal complete
Mass spectrogram is scanned, the optimal full scan mass spectrogram refers to that adduct ion abundance ratio is 10%-20%, in the optimal full scan
It selects abundance of ions than maximum 3-5 fragment ion in mass spectrogram, records the impact energy magnitude；
The fragment ion information is fragment ion theory accurate mass number and abundance under the optimal full scan mass spectrogram
Than；
Signal intensity ratio of the abundance than referring to the most strong fragment ion of the ion and signal in mass spectrogram；
The database by retention time according to being ranked up in electronic ID card；
The electronic identity database further includes intelligent Matching model, and Matching Model increases intelligent in the electronic ID card
With value Pm, computation model are as follows:
Wb+Wq=1；
Wherein MbFor the theoretical accurate mass number of adduct ion, MiFor the accurate mass number of i-th of fragment ion, WiIt is broken for i-th
The weight of piece ion, IiFor the abundance of ions ratio of i-th of fragment ion, fragment ion put in order for abundance of ions ratio from
It arrives greatly small；WbFor the weight of adduct ion, WqFor the comprehensive weight of fragment ion, n is the number of fragment ion.
2. the electronics of agricultural chemical compound in a kind of edible agricultural product based on LC-Q-Orbitrap according to claim 1
The method for building up of identity database, which is characterized in that the Wb, WqIt is adjusted according to the variation of intelligent Matching model, value is
Wb=Wq=0.5.
3. the electronics of agricultural chemical compound in a kind of edible agricultural product based on LC-Q-Orbitrap according to claim 1
The method for building up of identity database, which is characterized in that the determination method of the theoretical accurate mass number of the fragment ion are as follows:
1) according to compound molecule formula, fragment ion element composition is specified；
2) according to the mass number M of fragment ion in mass spectrogram, possible fragment ion element composition list is obtained by calculating；
Wherein: XiFor i-th of fragment ion element mass number, n is first prime number of fragment ion, yiIt is corresponding for i-th of fragment ion
The number of element；
3) it is made up of in list the splitting mechanism of molecular structure from fragment ion element and selects reasonable fragment ion element group
At, and calculate its theoretical accurate mass M '；
M '=X1y′1+X2y′2+…+Xny′n
Wherein: X1、X2......XnFor the fragment ion element mass number, y '1、y′2、......y′nFor preferred fragment ion
The number of the corresponding element of element composition.
4. agricultural chemical compound detection method in a kind of edible agricultural product based on LC-Q-Orbitrap, comprising:
1) sample to be tested Acidifying acetonitrile is homogenized and is extracted, after being dehydrated and be centrifuged, being concentrated, then purified through solid-phase extraction column SPE,
Acetonitrile+toluene solution elutes remains of pesticide, and sample solution is made after concentrated, filtering；
2) by LC-Q-Orbitrap instrument in Full MS/ddMS2Sample solution is obtained under mode to specify under chromatography Mass Spectrometry Conditions
Chromatogram and mass spectrogram, to obtain the accurate mass number information of the retention time and adduct ion in chromatogram and right
The fragment ion and mass spectrogram obtained under optimum collision energy is answered, and records the electronic identity of the corresponding unknown material of the retention time
Card；
3) successively by the electricity of each agricultural chemical compound in the electronic identity database in unknown material electronic ID card and claim 2
Sub- identity card compares, if Δ T≤0.3 and Δ P≤10%, record the agricultural chemical compound, otherwise enters next unknown
The comparison of object electronic ID card；
4) detection is completed, and shows the information of contained agricultural chemical compound in sample solution；
Wherein Δ T is the absolute value of the difference of any agricultural chemical compound retention time in unknown material retention time and database,Wherein PcFor unknown material intelligent Matching value, PiAny agricultural chemical compound intelligent Matching value in database.
5. agricultural chemical compound detection method in a kind of edible agricultural product based on LC-Q-Orbitrap as claimed in claim 4,
It is characterized in that, in step 4) if Δ T≤0.3 and 10% < Δ P≤30%, pass through the height of mass spectra peak in mass spectrogram
Judge whether to include the agricultural chemical compound with registration degree of certainty.
6. agricultural chemical compound detection method in a kind of edible agricultural product based on LC-Q-Orbitrap as claimed in claim 4,
It is characterized in that,
The sample further includes following pre-treatment:
10.0g sample is weighed, 0.01g is accurate to, in 100mL centrifuge tube, 30~40mL Acidifying acetonitrile extracting solution is added,
10000~11000rpm homogeneous, 1~2min；Anhydrous magnesium sulfate and sodium chloride, mass ratio, 4: 1,8~10min of oscillation is added；
After 4200rpm is centrifuged 5~7min, take 15~20mL supernatant in 150mL chicken heart bottle, 40 DEG C of heating water bath rotary evaporations to 1
~2mL, it is to be clean；
Using CarbonNH2 column, the anhydrous sodium sulfate of 1~2cm high is added in CarbonNH2 column, with 5~6mL acetonitrile-methylbenzene
Solution prewashing SPE decontaminating column, while bubble in decontaminating column discharge column is touched, efflux is thrown aside below decontaminating column；It is slightly higher to liquid level
When at the top of the sodium sulphate, concentrate is transferred to decontaminating column, under connect 50mL chicken heart bottle；It is rinsed with 2~3mL acetonitrile-methylbenzene solution
The chicken heart bottle for speckling with sample, cleaning solution is transferred in decontaminating column, is repeated 2~3 times；25mL liquid storage device is connect on column, with 25~
The elution of 30mL acetonitrile-methylbenzene solution；To about 0.5mL, nitrogen is blown to closely to be done rotary evaporation after collection, and it is molten that 1mL acetonitrile-water is added
Liquid is filtered through 0.22 μm of nylon membrane after ultrasonic dissolution.
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| PCT/CN2018/121001 WO2019200947A1 (en) | 2018-04-16 | 2018-12-14 | Electronic identity database of pesticide compounds in edible agricultural products based on lc-q-orbitrap, and detection method |
| US16/314,599 US11169128B2 (en) | 2018-04-16 | 2018-12-14 | Electronic ID database and detection method for pesticide compound in edible agro-products based on LC-Q-Orbitrap |
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| CN201810337240.9A CN108760909A (en) | 2017-04-17 | 2018-04-16 | A kind of edible Practice for Pesticide Residue in Agricultural Products is non-target, multi objective, rapid detection electronic method |
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| WO2019200946A1 (en) * | 2018-04-16 | 2019-10-24 | 中国检验检疫科学研究院 | Gc-q-orbitrap-based electronic identification database of pesticide compounds in edible agricultural products and detection method |
| WO2019200947A1 (en) * | 2018-04-16 | 2019-10-24 | 中国检验检疫科学研究院 | Electronic identity database of pesticide compounds in edible agricultural products based on lc-q-orbitrap, and detection method |
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