CN105738460A - LC-Q-TOF/MS detection technology for 544 pesticide residues in leaf vegetables - Google Patents
LC-Q-TOF/MS detection technology for 544 pesticide residues in leaf vegetables Download PDFInfo
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
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- G—PHYSICS
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- 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
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
Abstract
The invention provides an LC-Q-TOF/MS detection technology for 544 pesticide residues in leaf vegetables. The technology includes the steps of: 1) under a TOF/MS mode in the LC-Q-TOF/MS, respectively measuring retention time of each pesticide reference substance under the certain chromatography-mass spectrometry conditions; 2) determining the ionization formation and chemical formula of the compound with an ESI source to obtain accurate mass number of a parent ion of each compound to form a TOF/MS database; 3) under a Q-TOF/MS mode, respectively collecting fragment ion mass spectrograms under 3-5 different collision energy of each pesticide reference substance, and introducing the collected information to PCDL software to obtain a Q-TOF/MS database; 4) comparing the retention time, primary mass spectrum and secondary mass spectrum of the sample of the leaf vegetables to determine whether the pesticide residues exist in the sample or not, and meanwhile, performing secondary determination to the compound having higher primary score, and if secondary score is high, determining existence of the pesticide residues. The technology is quick, high-throughput, high-precision and high-reliability, and can be used for accurately screening the pesticides in the leaf vegetables.
Description
Technical field
The present invention relates to food hygiene, agricultural product security detection field, particularly to one for leaf vegetables
In the method for detecting of 544 kinds of pesticide residues.
Background technology
As far back as World Health Organization (WHO) (WHO) in 1976, food and agricultural organization (FAO) and United Nations Environment Programme
(UNEP) global environment detecting system/food item (Global Environment is jointly established
Monitoring System, GEMS/Food), it is intended to grasp member state's food pollution situation, understand food
Pollutant intake, protects health, and give a impetus to trade development.Now, countries in the world all propose food safety
It is raised to the strategic position of national security.Pesticide Residue is one of food security standard, is also that international trade is accurate
Enter threshold.Meanwhile, the requirement of pesticide residues is presented kind and gets more and more, the strictest development of limiting the quantity
Trend, the Pesticide Residue threshold that namely international trade is set up is more and more higher.European Union formulates now
162248 MRL standards of 839 kinds of pesticide, the U.S. has formulated 39147 MRL marks of 351 kinds of pesticide
Standard, Japan has formulated 51600 multinomial MRL standards of 579 kinds of pesticide, and China has issued 381 in 2014
Plant 3650 MRL standards of pesticide.At present, the most commonly used uniform limit limitation is 10 μ g/kg.
Therefore, the quick the Detection Technologies of Pesticide Residues of high flux is all called in food safety and international trade, and this most also gives wide
Big pesticide residue analysis worker provides opportunities and challenges.In current numerous Analytical Techniques of Pesticide Residues,
Hydrolysis and condensation is to realize the optimized analysis means that the many residuals of high flux quickly detect.
Pesticide residue analysis is many with gas chromatogram, liquid chromatograph, gas chromatography-mass spectrum and liquid chromatograph-matter at present
Spectrum multiple techniques is main.These detection techniques all carry out qualitative firstly the need of standard sample of pesticide comparison.Such as, right
The detection of 100 kinds of pesticide is accomplished by preparing corresponding 100 kinds of standard sample of pesticide comparison, and outside these 100 kinds
Pesticide all can be missed.In the real work of agricultural and veterinary chemicals results from residue tests room, most laboratorys are all without storage
Standby hundreds of standard sample of pesticide, its reason is that standard sample of pesticide is the most expensive, and effect duration only has 2,
3 years, need overlapping investment.The standing standard sample of pesticide of common laboratory only has tens kinds, the agriculture of its daily monitoring
Medicine kind is the most just only limited to these tens kinds, thereby results in food safety monitoring leak.
Summary of the invention
The present invention on the basis of have rated the world and commonly use the mass spectrum behavior characteristics of 1000 Multiple Pesticides, novelty
Establish 544 kinds of pesticide TOF/MS and Q-TOF/MS accurate mass data base；Establish innovatively
With compound Information in Mass Spectras such as high-resolution accurate mass number, Isotopic Distribution and isotope abundances as criterion of identification
, according to TOF/MS and Q-TOF/MS accurate mass data base, 544 kinds of pesticide are detected and confirmed
LC-Q-TOF/MS technical method；Develop matched leaf vegetables Pesticide Residues innovatively
High flux sample preparation technology.Achieve have only to sample prepare, single injected sampling detection, it is possible to leaf
The rapid detection of 500 Multiple Pesticides in dish class vegetable, and the pesticide detected can be carried out Q-TOF/MS
Data base confirms, and revolutionizes original qualitative model with compound standard thing as reference, is that one need not
Reference material compares, quick, high flux, accurately and reliably Detecting Pesticide new technique.
One, novelty of the present invention is:
1, establish 544 kinds of pesticide accurate masses innovatively and can reach the accurate mass data of 0.0001m/z
Storehouse, including TOF/MS data base and Q-TOF/MS data base.
2, TOF/MS data base comprise the English name of 544 kinds of pesticide, chemical molecular formula, retention time,
The full scan mass spectrum letter of accurate mass number, Isotopic Distribution, isotope abundance and parent ion and fragment ion
Breath, contains much information, the most comprehensively.
3, the full scan mass spectrum information of every kind of pesticide fragment ion in Q-TOF/MS data base, is by right
Under eight different collision energies, full scan mass spectrum is preferred, informative four the different impact energies of ion obtained
The lower fragment ion full scan figure of amount.
4, establish innovatively with chemical combination such as high-resolution accurate mass number, Isotopic Distribution and isotope abundances
Material spectrum information is criterion of identification, according to TOF/MS and Q-TOF/MS accurate mass data base to 544
Plant the LC-Q-TOF/MS technical method that pesticide carries out detecting and confirming, revolutionize original with compound mark
Quasi-thing is the qualitative model of reference, is that one need not reference material comparison, quickly, high flux, accurately and reliably
Detecting Pesticide new technique.
5, the residual detection techniques method that the present invention sets up can according to the retention time of target compound, accurately
The information such as mass number, Isotopic Distribution and isotope abundance, by with compound in TOF/MS data base
Corresponding informance retrieval comparison, provides the matching degree score value of target compound.According to the score value of target compound,
Realize the qualitative detecting to pesticide.
6, the detection techniques method that the present invention sets up can be according to the full scan mass spectrum of target compound fragment ion
Figure information, by with the full scan mass spectrum information retrieval of compound fragment ion in Q-TOF/MS data base
Comparison, provides the matching degree score value of target compound, it is achieved the qualitative confirmation to target compound.Compare biography
The detection qualitative confirmation method of system, owing to have employed full scan mass spectrum information comparison, has higher qualitative standard
Exactness.
7, develop the Residual Pesticides in Farm Produce high flux sample supporting with remaining detection techniques method and prepare skill
Art.Achieve have only to sample prepare, twice sample detection, it is possible to more than 500 kind of agriculture in agricultural product
The rapid detection of medicine and confirmation.
8, LC-Q-TOF/MS main detection techniques index: sweep limits 50-1600m/z；Measure is accurate
Quality is up to 0.0001m/z；Within Mass accuracy can be controlled in the 10ppm of molecular mass；Scanning speed exists
50-1600m/z scope is 4 times per second.LC-Q-TOF/MS main detection techniques feature: sample is once prepared,
544 kinds of pesticide detectings, qualitative and confirmation can be completed in 1 hour；More than 80% pesticide confirmation point reaches 10
More than individual；In the 544 kinds of pesticide detected at the same time, 327 kinds of pesticide detecting sensitivity are less than uniform limit 10
Micro-g kg, accounting 64.1%, substantially meet the requirement of the horizontal examination of various countries' pesticide residues MRL.
Two, the technical solution used in the present invention:
1, accurate mass data base is set up
The foundation of 1.1TOF/MS data base
Under TOF/MS pattern, measure the reservation under specifying chromatograph Mass Spectrometry Conditions of the every kind of pesticide standard thing respectively
Time, determine the ionized form (+H ,+NH under this compound ESI source4,+Na) and chemical formula, obtain every
Plant the accurate mass number of compound parent ion, isotopic peak distribution and abundance ratio.By the title of 544 kinds of pesticide,
The information such as the distribution of retention time, molecular formula, accurate mass number, isotopic peak and abundance ratio import to data base
In CSV software document, form TOF/MS data base.
The foundation of 1.2Q-TOF/MS data base
544 kinds of pesticide in TOF/MS data base are sequentially input at Agilent MassHunter data acquisition interface
Parent ion, under Q-TOF/MS pattern, to 544 kinds of pesticide respectively under 8 collision energies, carry out
Fragment ion full scan mass spectrum gathers.Broken under informative 3-5 the different collision energies of ion preferably wherein
Sheet ion full scan mass spectrum imports in database software file, and is associated with corresponding pesticide information, builds up
Q-TOF/MS data base.
2, set up with compound information such as high-resolution accurate mass number, Isotopic Distribution and isotope abundances for knowing
Other standard, 544 kinds of pesticide LC-Q-TOF/MS detectings and confirmation technical method, technical step includes:
Prepared by 2.1 samples: leaf vegetables sample extracts through 1% acetate acetonitrile, Carbon/NH2Column purification,
Acetonitrile+toluene (3+1, v/v) eluting residual pesticide, concentrated, dissolve and filter after make and treat sample measuring liquid；
Described leaf vegetables includes Herba Apii graveolentis, Herba Spinaciae, Plantula Brassicae chinensis, Caulis et Folium Chrysanthemi segeti, Caulis et Folium Lactucae Sativae, Chinese cabbage, green vegetable, Herba Amaranthi tricoloris
Dish, leaf mustard, Radix Pachyrhizi Erosi leaf, Folium Lycii, Caulis et Folium Ipomoeae aquaticae, baby cabbage, Lettuce, Brassica campestris L, Herba basellae rubrae, cabbage,
Herba Lactucae versicoloris, Caulis et Folium Foeniculi, Caulis et Folium Lactucae sativae, Herba Cleomis, youngster's dish, spring vegetable, crow dish, Caulis et Folium Brassicae junceae and taro；
2.2 chromatographic mass spectrometry detections: use and be furnished with reversed phase chromatographic column (ZORBAX SB-C18 column of 2.1
Mm × 100mm, 3.5 μm) liquid chromatograph separate the pesticide treating in sample measuring liquid, be furnished with electric spray ion source
LC-Q-TOF-MS detects；
The 2.3 qualitative method for detecting of TOF/MS data base: call in the qualitative software of Agilent MassHunter
The TOF/MS data base set up, according to TOF/MS data base to target compound retrieval by header, retrieval ginseng
Number retention time deviation is defined to ± 0.5min, and accurate mass deviation is defined to ± 10ppm, and ionized form selects
+ H ,+NH4,+Na pattern, software can be according to the reality of the key elements such as every kind of Compound Retention time, accurate mass
The deviation of theoretical value in measured value and TOF/MS data base, and Isotopic Distribution and isotope ratio four ginseng
Number, by scientifically setting each key element weight (wherein accurate mass number deviation in matching degree score calculates
Weight the highest), provide retrieval matching score value, the retrieval higher compound of matching score value, for doubtful inspection
Go out compound, according to the method accuracy of TOF/MS data base qualitative detecting Residual Pesticides in Farm Produce, reach
More than 90%；
2.4 Q-TOF/MS data base qualitative confirmation method: under Targeted MS/MS drainage pattern, input
The parent ion of doubtful detection compound, retention time and optimal collision energy, treat sample measuring liquid and again detect,
By detection mass spectrum and the full scan mass spectrum of compound fragment ion in Q-TOF/MS data base, at mirror image
Mate under the conditions of contrast, the most consistent with spectrum storehouse according to characteristic ion distribution, secondary fragment accurate mass number deviation
Providing score with the ratio of fragment ion, its matching score value is higher, i.e. confirms to detect this target compound.Should
Confirmation method use full scan mass spectrum coupling confirmation, make more than 80% pesticide confirmation point can reach 10 with
On, ratio 4 confirmation point requirements of standard, there is higher qualitative accuracy.
Three, the technology contents of the present invention:
The present invention provides the accurate mass data base of two kinds of 544 kinds of pesticide and one for detecting leaf vegetables vegetables
544 kinds of pesticide residues detection techniques in dish.
1, two kinds of accurate mass data bases that the present invention provides are TOF/MS data base and Q-TOF/MS data
Storehouse.
1.1 TOF/MS database technology contents include: the chemical formula of 544 kinds of pesticide, accurate mass number, retain
The information such as time and pesticide English name.In TOF/MS data base, 3 kinds of compounding techniques content example are as follows:
1.2 Q-TOF/MS database technology contents include: 544 kinds of pesticide fragment under 4 collision energies
Ion information mass spectrum.
2, the invention provides 544 kinds of pesticide LC-Q-TOF/MS detectings and confirmation in a kind of leaf vegetables
Technical method.Detecting and confirmation technical method content include the following:
Prepared by 2.1 samples
2.1.1 leaf vegetables sample takes edible part chopping, mixing, seals, indicates labelling.
2.1.2 weigh 10g leaf vegetables sample (being accurate to 0.01g), in 80mL centrifuge tube, add
40mL 1% acetate acetonitrile, with high-speed homogenization machine 13500r/min, homogenate is extracted 1min, is added 1g chlorination
Sodium, 4g anhydrous magnesium sulfate, vibrate 5min, and under 4200r/min, centrifugal 5min, takes supernatant 20mL,
In 40 DEG C of water-baths, rotary evaporation is concentrated into about 1mL, to be clean.
2.1.3 in Carbon/NH2 post, add about 2cm height anhydrous sodium sulfate.First with 4mL acetonitrile+toluene
(3+1, v/v) drip washing SPE post, and discard effluent, when liquid level arrives the top of sodium sulfate, rapidly will
Sample concentration liquid is transferred on decontaminating column, under connect new Cor Gigeriae Galli bottle graft receive.The most every time with 2mL acetonitrile+toluene (3+1,
V/v) washing sample liquid bottle three times, and cleaning mixture is moved in SPE post.Post connects 50mL liquid reservoir,
Again with 25mL acetonitrile+toluene (3+1, v/v) eluting pesticide, it is incorporated in Cor Gigeriae Galli bottle, and 40 DEG C of water-baths
Middle spin concentration is to about 0.5mL.
2.1.4 concentrated solution is placed under nitrogen and dries up, add the acetonitrile+0.1% formic acid water (2+8, v/v) of 2mL
Mixing, constant volume after 0.2 μm membrane filtration, obtain testing sample solution a.
2.2 LC-Q-TOF/MS detections
2.2.1 liquid chromatograph separates: testing sample solution a is by being furnished with reversed phase chromatographic column (ZORBAX SB-C18
Column of 2.1mm × 100mm, 3.5 μm) liquid chromatographic system separate；Mobile phase A is the second of 5mM
Acid ammonium-0.1% formic acid-water；Mobile phase B is acetonitrile；Liquid chromatograph gradient elution program is: 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
℃；Sample size: 10 μ L.
2.2.2 Mass Spectrometer Method: Agilent 6530 LC-Q-TOF/MS is at electron spray ionisation positive ion mode
(ESI+)；Capillary voltage: 4000V；Dry temperature: 325 DEG C；Dry gas stream amount 10L/min, sheath
Gas flow velocity 11L/min, sheath gas temperature is 325 DEG C；Atomization gas pressure 40psi, taper hole voltage 60V, broken
Split voltage 140V.Full scan karyoplasmic ratio scope is 50-1600m/z, and uses internal standard reference solution to instrument matter
Accuracy of measurement carries out real time correction.By Agilent MassHunter Workstation Software (version
B.05.00) to mass spectrometric data collection and process.
The 2.3 qualitative detectings of TOF/MS data base
In the qualitative software of Agilent MassHunter, call the TOF/MS data base set up, retrieval is set
Parameter retention time deviation is ± 0.5min, and accurate mass deviation is ± 10ppm, ionized form selection+H,
+NH4,+Na pattern；It is fixed to carry out the sample solution mass spectrometric data of above-mentioned collection according to TOF/MS data base
Property retrieval, go out every kind of compound retrieval matching score value by computed in software, retrieval matching score value ＞ 70
Compound, for doubtful detection compound.
The 2.4 qualitative confirmations of Q-TOF/MS data base
Under Targeted MS/MS drainage pattern, input the parent ion of doubtful detection compound, retention time
With optimal collision energy, testing sample solution a is detected again, will detection mass spectrum and Q-TOF/MS
The full scan mass spectrum of compound fragment ion in data base, coupling confirmation under the conditions of mirror image contrasts.Its coupling
Score value ＞ 70, i.e. confirms to detect this target compound.
The present invention is had the beneficial effect that compared to existing technology
1, the present invention with compound information such as high-resolution accurate mass number, Isotopic Distribution and isotope abundances is
Criterion of identification, 500 Multiple Pesticides detectings of foundation and confirmation technical method, revolutionize original with compound
Reference material is the qualitative model of reference, is that one need not reference material comparison, quickly, high flux, accurately and reliably
Detecting Pesticide new technique；
2, the present invention is on the premise of being not required to reference material comparison, it is achieved that qualitative the detecing of 500 Multiple Pesticides residuals
Survey and confirmation, the cost buying reference material, the most more environmentally-friendly, safety can be greatly saved；
3, the present invention can complete 500 Multiple Pesticides detectings, qualitative and confirmation in 1 hour, with traditional detection
Method compares, and can improve work efficiency hundreds times；
4, the present invention can accomplish 500 Multiple Pesticides in leaf vegetables sample are remained disposable extraction and cleaning,
Single injected sampling detect, compared with traditional detection method, can save testing cost, improve work efficiency hundreds of
Times；
5, in 544 kinds of pesticide that the present invention detects simultaneously, there are 327 kinds of pesticide detecting sensitivity less than 10 micrograms
/ kilogram, accounting 64.1%, substantially meet the requirement of the horizontal examination of various countries' pesticide residues MRL；
6, the present invention is on the premise of being not required to the comparison of pesticide standard thing, according to accurate mass data base to targeted
Qualitative and the confirmation of compound, makes more than 80% pesticide confirmation point reach more than 10, and whole pesticide meet European Union
Requirement to compound 4 confirmation point, qualitative results is more accurately and reliably；
7, the leaf vegetables sample Pesticides that the present invention can test includes the class in following material or a few class:
Organophosphorus insecticide, carbamate chemicals for agriculture, Ion pairing, sulfonylurea pesticide, nicotinoids agriculture
Medicine, cinerins pesticide and other kinds of pesticide and metabolite；
8, the detection techniques index that the present invention sets up: sweep limits 50-1600m/z；The accurate mass measured can
Reach 0.0001m/z；Within Mass accuracy can be controlled in the 10ppm of molecular mass；Scanning speed is at 50-600m/z
Scope is 4 times per second；
9, accurate mass data base and the one of two kinds of 544 kinds of pesticide that the present invention provides is used for detecting leaf vegetables
In class vegetable, 544 kinds of pesticide residues detection techniques are promoted at 7 pesticide residue analysis laboratorys and answer
With, through the detecting checking to more than 500 commercially available leaf vegetables samples of 30 provinces and cities, achieve important residual
Stay detecting data.
Accompanying drawing explanation
Fig. 1 Herba Apii graveolentis sample full scan pattern TOF shot chart；
Two grades of acquisition method exemplary plot of doubtful pesticide in Fig. 2 Herba Apii graveolentis sample；
Fig. 3 Herba Apii graveolentis sample detects two grades of TOF/MS of pesticide and confirms shot chart；
Fig. 4 leaf mustard sample full scan pattern TOF shot chart；
Two grades of acquisition method exemplary plot of doubtful pesticide in Fig. 5 leaf mustard sample；
Fig. 6 leaf mustard sample detects two grades of TOF/MS of pesticide and confirms shot chart；
Fig. 7 Lettuce sample full scan pattern TOF shot chart；
Two grades of acquisition method exemplary plot of doubtful pesticide in Fig. 8 Lettuce sample；
Fig. 9 Lettuce sample detects two grades of TOF/MS of pesticide and confirms shot chart.
Detailed description of the invention
Embodiment 1
In leaf vegetables (as a example by Herba Apii graveolentis) 544 kinds of pesticide (being shown in Table 1) LC-Q-TOF/MS detecting and
Confirmation technology implementation example, comprises the steps:
1, the concrete steps of Sample Pretreatment Technique:
1.1 Herba Apii graveolentis samples take edible part chopping, mixing, seal, indicate labelling；
1.2 weigh 10g Herba Apii graveolentis sample (being accurate to 0.01g), in 80mL centrifuge tube, add 40mL 1%
Acetate acetonitrile, with high-speed homogenization machine 13500r/min, 1min is extracted in homogenate, adds 1g sodium chloride, 4g without
Water magnesium sulfate, vibrate 5min, and under 4200r/min, centrifugal 5min, takes supernatant 20mL, at 40 DEG C
In water-bath, rotary evaporation is concentrated into about 1mL, to be clean.
1.3 at Carbon/NH2Post adds about 2cm height anhydrous sodium sulfate.First with 4mL acetonitrile+toluene
(3+1, v/v) drip washing SPE post, and discard effluent, when liquid level arrives the top of sodium sulfate, rapidly will
Sample concentration liquid is transferred in SPE post, under connect new Cor Gigeriae Galli bottle graft receive.The most every time with 2mL acetonitrile+toluene (3+1,
V/v) washing sample liquid bottle three times, and cleaning mixture is moved in SPE post.Post connects 50mL liquid reservoir,
With 25mL acetonitrile+toluene (3+1, v/v) eluting pesticide and related chemicals, it is incorporated in Cor Gigeriae Galli bottle, and
In 40 DEG C of water-baths, spin concentration is to about 0.5mL.
Concentrated solution is placed under nitrogen and to dry up by 1.4, adds acetonitrile+water (2+8, v/v) mixing of 2mL, warp
Constant volume after 0.2 μm membrane filtration, obtains testing sample solution.
2, LC-Q-TOF/MS operating condition
Chromatographic condition: liquid chromatogram mobile phase A is ammonium acetate-0.1% formic acid-water of 5mM；Mobile phase B is
Acetonitrile；Gradient elution program is: 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, after
Run 4min；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample size: 10 μ L.
The mass spectrometric capillary voltage of Mass Spectrometry Conditions: Agilent 6530 LC-Q-TOF/MS: 4000V；It is dried
Temperature: 325 DEG C；Dry gas stream amount 10L/min, sheath gas flow velocity 11L/min, sheath gas temperature is 325
℃；Atomization gas pressure 40psi, taper hole voltage 60V, fragmentation voltage 140V.Full scan karyoplasmic ratio scope is
50-1600m/z, and use internal standard reference solution that instrument quality precision is carried out real time correction.Pass through Agilent
Mass spectrum testing result is gathered and processes by MassHunter Workstation Software (version is B.05.00).
3 leaf vegetableses (as a example by Herba Apii graveolentis) Pesticide Residues is detected
3.1 measure sample solution under full scan pattern, detecting result are compared with TOF data storehouse,
Go out 1 grade of TOF score, see Fig. 1.
3.2 meet the compound of requirement for score, set up two grades of acquisition methods in software, refer to Fig. 2.
3.3, under MS/MS pattern, rerun sample solution, it is thus achieved that sample fragment ion full scan data,
It is compared with fragment ion information in second order ms picture library, obtains two grades of QTOF scores, see Fig. 3.
In 4 certain provincial capital's Herba Apii graveolentis sample, LC-Q-TOF/MS detects result
Gathering 21, certain provincial capital commercially available Herba Apii graveolentis sample, application LC-Q-TOF/MS technology carries out 544 kinds
Pesticide residues are detected, LC-Q-TOF/MS technology detection 18 kinds of pesticide residues, altogether 65 frequencys, relate to sample
Product 19, concrete outcome is shown in Table 2.
1 544 kinds of pesticide LC-Q-TOF/MS detection parameters tables of table
In the Herba Apii graveolentis sample of table 2 somewhere, LC-Q-TOF/MS detects result
Embodiment 2
544 kinds of pesticide (being shown in Table 1) LC-Q-TOF/MS detectings in leaf vegetables (as a example by leaf mustard)
With confirmation technical em-bodiments.
Sample pre-treatments step, LC-Q-TOF/MS operating condition are all with reference to the treatment conditions in embodiment 1.
Under full scan pattern, measure sample solution, detecting result is compared with TOF data storehouse, draws 1
Level TOF score, is shown in Fig. 4；Score is met to the compound of requirement, sets up two grades of acquisition methods in software,
Refer to Fig. 5；Under MS/MS pattern, rerun sample solution, it is thus achieved that sample fragment ion full scan number
According to, it is compared with fragment ion information in second order ms picture library, obtains two grades of QTOF scores, see figure
6。
In certain provincial capital's leaf mustard sample, LC-Q-TOF/MS detects result: gather the commercially available leaf of certain provincial capital
12, Caulis et Folium Brassicae junceae sample, application LC-Q-TOF/MS technology carries out 544 kinds of pesticide residues detectings,
LC-Q-TOF/MS technology detection 27 kinds of pesticide residues, altogether 83 frequencys, relate to 12, sample, specifically tie
Fruit is shown in Table 3.
In the leaf mustard sample of table 3 somewhere, LC-Q-TOF/MS detects result
Embodiment 3
544 kinds of pesticide (being shown in Table 1) LC-Q-TOF/MS detectings in leaf vegetables (as a example by Lettuce)
With confirmation technology implementation example.
Sample pre-treatments step, LC-Q-TOF/MS operating condition are all with reference to the treatment conditions in embodiment 1.
Under full scan pattern, measure sample solution, detecting result is compared with TOF data storehouse, draws 1
Level TOF score, is shown in Fig. 7；Score is met to the compound of requirement, sets up two grades of acquisition methods in software,
Refer to Fig. 8；Under MS/MS pattern, rerun sample solution, it is thus achieved that sample fragment ion full scan number
According to, it is compared with fragment ion information in second order ms picture library, obtains two grades of QTOF scores, see figure
9。
In certain provincial capital's Lettuce sample, LC-Q-TOF/MS detects result: gather the commercially available oil of certain provincial capital
9, wheat dish sample, application LC-Q-TOF/MS technology carries out 544 kinds of pesticide residues detectings, LC-Q-TOF/MS
Technology detection 11 kinds of pesticide residues, altogether 23 frequencys, relate to 5, sample, and concrete outcome is shown in Table 4.
In the Lettuce sample of table 4 somewhere, LC-Q-TOF/MS detects result
Claims (10)
1. one kind is used for detecting the method for detecting of 544 kinds of pesticide residues in leaf vegetables, it is characterised in that:
By setting up TOF/MS data base and the Q-TOF/MS data base of 544 kinds of pesticide, by target compound
Chromatographic mass spectrometry information is mated with the information retrieval of TOF/MS data base and Q-TOF/MS data base, it is achieved that
Compare without reference material, more than in agricultural product 500 kinds pesticide are carried out qualitative detecting and confirmation simultaneously；Described
544 kinds of pesticide be the pesticide of numbered 1-544 in table 1.
2. a TOF/MS data base as claimed in claim 1, it is characterised in that: by the following method
Build data base: under TOF/MS pattern, measure every kind of pesticide standard thing respectively and specifying chromatographic mass spectrometry bar
Retention time under part, determines the ionized form under this compound ESI source, selection+H ,+NH4,+Na, and
Chemical formula, obtains the accurate mass number of every kind of compound parent ion, and isotopic peak distribution and abundance ratio, by 544
Plant the title of pesticide, retention time, molecular formula, accurate mass number, isotopic peak distribution and abundance ratio information to lead
Enter in data base's CSV software document, form TOF/MS data base.
3. a Q-TOF/MS data base as claimed in claim 1, it is characterised in that: by with lower section
Method builds data base: sequentially input TOF/MS data base as stated in claim 2 at data acquisition interface
In the parent ion of 544 kinds of pesticide, under Q-TOF/MS pattern, to 544 kinds of pesticide respectively 8 collisions
Under energy, carry out fragment ion full scan mass spectrum collection, preferably wherein informative 3-5 the difference of ion
Under collision energy, fragment ion full scan mass spectrum imports in database software file, and with corresponding pesticide information
It is associated, builds up Q-TOF/MS data base.
Method for detecting the most according to claim 1, it is characterised in that: for pesticide residues detecting not
Pesticide standard thing is needed to compare.
Method for detecting the most according to claim 1, it is characterised in that described method includes following procedure:
A prepared by () sample: leaf vegetables sample extracts through acetate acetonitrile homogenate, through being dehydrated and centrifugal, dense
After contracting, then through Carbon/NH2Column purification, acetonitrile+toluene eluting residual pesticide, concentrated, filter after make
Treat sample measuring liquid a；
B () chromatographic mass spectrometry detects: use the liquid chromatograph being furnished with reversed phase chromatographic column, under special color spectral condition,
Separate the pesticide treating in sample measuring liquid a；Use the LC-Q-TOF/MS being furnished with electric spray ion source, in specific matter
Under spectral condition, detect pesticide residues；
(c) TOF/MS data base's examination: call set up as claimed in claim 2 in qualitative software
TOF/MS data base, according to TOF/MS data base to the qualitative inspection of target compound in mass spectrum testing result
Rope, search argument retention time deviation is defined to ± 0.5min, and accurate mass deviation is defined to ± 10ppm, ion
Change formal character+H ,+NH4,+Na pattern, according to measured value and the TOF/MS of every kind of each key element of compound
The deviation of theoretical value in data base, provides retrieval matching score value, retrieves the compound of matching score value ＞ 70,
For doubtful detection compound；
D () Q-TOF/MS data base confirms: under Targeted MS/MS drainage pattern, input doubtful inspection
Go out the parent ion of compound, retention time and optimal collision energy, treat sample measuring liquid a and again detect, will inspection
Survey spectrogram in spectrogram and Q-TOF/MS data base as claimed in claim 3, under the conditions of mirror image contrasts
Join confirmation, its matching score value ＞ 70, i.e. confirm to detect this target compound.
Method for detecting the most according to claim 5, it is characterised in that: in step (a), described leaf vegetables
Class vegetable include Herba Apii graveolentis, Herba Spinaciae, Plantula Brassicae chinensis, Caulis et Folium Chrysanthemi segeti, Caulis et Folium Lactucae Sativae, Chinese cabbage, green vegetable, Amaranthus mangostanus L., leaf mustard,
Radix Pachyrhizi Erosi leaf, Folium Lycii, Caulis et Folium Ipomoeae aquaticae, baby cabbage, Lettuce, Brassica campestris L, Herba basellae rubrae, cabbage, Herba Lactucae versicoloris, Caulis et Folium Foeniculi,
Caulis et Folium Lactucae sativae, Herba Cleomis, youngster's dish, spring vegetable, crow dish, Caulis et Folium Brassicae junceae and taro.
Method for detecting the most according to claim 5, it is characterised in that: in step (a), described
Carbon/NH2Column purification step is as follows: at Carbon/NH2In post add anhydrous sodium sulfate, first with acetonitrile+
Toluene mixture liquid drip washing SPE post, and discard effluent, when liquid level arrives the top of sodium sulfate, rapidly will
Sample concentration liquid is transferred on decontaminating column, under connect new Cor Gigeriae Galli bottle graft receive；Acetonitrile+toluene mixture liquid washing sample liquid again
Bottle three times, and cleaning mixture is moved in SPE post；Again with acetonitrile+toluene mixture liquid eluting pesticide, merge eluting
Liquid is in Cor Gigeriae Galli bottle.
Method for detecting the most according to claim 5, it is characterised in that: in step (a), described through dense
Contracting, filter after make treat sample measuring liquid a refer to claim 7 obtained by eluent, in 40 DEG C of water-baths revolve
Turn and concentrate, and concentrated solution is placed under nitrogen and dries up, add acetonitrile+formic acid water mixing, fixed after membrane filtration
Hold, obtain testing sample solution a.
Method for detecting the most according to claim 5, it is characterised in that: in step (b), described specific
Chromatographic condition refers to: liquid chromatogram mobile phase A is ammonium acetate-0.1% formic acid-water of 5mM；Mobile phase B is
Acetonitrile；Gradient elution program is: 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, after
Run 4min；Flow velocity is 0.4mL/min；Column temperature: 40 DEG C；Sample size: 10 μ L.
Method for detecting the most according to claim 5, it is characterised in that: in step (b), described spy
Determine Mass Spectrometry Conditions to refer to: the mass spectrometric capillary voltage of Agilent 6530LC-Q-TOF/MS: 4000V；Dry
Dryness temperature: 325 DEG C；Dry gas stream amount 10L/min, sheath gas flow velocity 11L/min, sheath gas temperature is 325
℃；Atomization gas pressure 40psi, taper hole voltage 60V, fragmentation voltage 140V, full scan karyoplasmic ratio scope is
50-1600m/z, and use internal standard reference solution that instrument quality precision is carried out real time correction, pass through Agilent
Mass spectrum testing result is gathered and processes by MassHunter Workstation Software.
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| CN106248773A (en) * | 2016-07-19 | 2016-12-21 | 中国检验检疫科学研究院 | Vitamin B in a kind of quickly mensuration multidimensional sheet1with ascorbic method |
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| CN107843660A (en) * | 2017-10-17 | 2018-03-27 | 贵州省农产品质量安全监督检验测试中心 | It is a kind of at the same determine it is dark brownish green in 124 kinds of residues of pesticides method analysis method |
| CN112782317A (en) * | 2021-02-04 | 2021-05-11 | 天津市食品安全检测技术研究院 | Method for simultaneously determining residual quantity of 13 carbamates pesticides in milk |
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