CN105675702A - GC-Q-TOF/MS detection technology of 708 pesticide residues in edible mushrooms - Google Patents
GC-Q-TOF/MS detection technology of 708 pesticide residues in edible mushrooms Download PDFInfo
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- G—PHYSICS
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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
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Abstract
The invention relates to the field of pesticide residue detection and provides a detection method for screening 708 pesticide residues in edible mushrooms. The method includes the steps of: a) under a first-stage mode of GC-Q-TOF/MS, collecting a fragment ion full-scanning mass spectrogram of pesticide reference substances under certain conditions to obtain a chemical formula, retention time, accurate mass numbers of fragment ions, and ion abundance ratio of the pesticide; b) introducing the data to PCDL software, correlating the data with corresponding pesticide information to obtain a database; c) under the same preset conditions, detecting the full-spectrum data of the to-be-detected samples; and d) performing accurate mass database retrieval to the sample detection results, and accurately identifying the detected pesticide satisfying identification basis to obtain the types of the pesticide residues in the edible mushrooms. By means of the GC-Q-TOF/MS high-resolution mass spectrum, the detection method can detect the pesticide residues in the edible mushrooms, preferably, with combination of special pre-treatments, so that the method is quick, high-throughput, high-precision and high-reliability. The method is free of a reference substance and can be used for qualitatively detecting and identifying the 708 pesticides in edible mushrooms.
Description
Technical field
The present invention relates to the residual detection field of agriculture, apply, particularly to a kind of, the method for detecting that in GC-Q-TOF/MS examination edible fungi, 708 kinds of agricultures are residual.
Background technology
Fruit and vegetable is closely bound up with the life of people, and especially the nutritive value of edible fungi (mushroom, Pleurotus eryngii, Lentinus Edodes, Flammulina velutiper (Fr.) Sing and Pleurotus ostreatus etc.) enjoys the favor of consumer. But, at present in the plantation of edible fungi, growth and preserving process is likely to use pesticide and chemicals more than 1100 kinds. But, with the widely using of pesticide, it is excessively used, abuse and the abnormal activity such as misuse also produces therewith so that pesticide residues are in agricultural product, thus causing potential threat to human health, also the foreign trade of agricultural product is created adverse effect simultaneously.
Global environment detection system/food item (GlobalEnvironmentMonitoringSystem is jointly established as far back as World Health Organization (WHO) (WHO) in 1976, food and agricultural organization (FAO) and United Nations Environment Programme (UNEP); GEMS/Food); it is intended to grasp member state's food pollution situation; understand food contaminant intake; protection health, give a impetus to trade development. Now, countries in the world all rise to food safety the strategic position of national security. Pesticide Residue is one of food security standard, is also international trade access threshold. Meanwhile, the requirement of pesticide residues presenting kind and gets more and more, increasingly stricter development trend of limiting the quantity, the Pesticide Residue threshold that namely international trade is set up is more and more higher. Present European Union has formulated 162248 MRL standards of 839 kinds of pesticide, and the U.S. has formulated 39147 MRL standards of 351 kinds of pesticide, and Japan has formulated 51600 multinomial MRL standards of 579 kinds of pesticide, and China has issued 3650 MRL standards of 381 kinds of pesticide for 2014. At present, commonly used in the world 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 provides opportunities and challenges also to undoubtedly vast pesticide residue analysis worker. 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.
Current pesticide residue analysis is many based on gas chromatogram, liquid chromatograph, gas chromatography-mass spectrum and LC-MS-MS.These detection techniques all carry out qualitative firstly the need of standard sample of pesticide comparison. Such as, it is accomplished by preparing corresponding 100 kinds of standard sample of pesticide comparison to the detection of 100 kinds of pesticide, and the pesticide outside these 100 kinds all can be missed. In the real work of agricultural and veterinary chemicals results from residue tests room, most laboratorys are all without laying in hundreds of standard sample of pesticide, and its reason is that standard sample of pesticide is not only expensive, and effect duration only has 2-3, it is necessary to overlapping investment. The standing standard sample of pesticide of common laboratory only has tens kinds, and the pesticide species of its daily monitoring is also just only limited to these tens kinds, thereby results in food safety monitoring leak.
Summary of the invention
In order to overcome drawbacks described above, the present inventor team concentrates on studies through for many years, have developed the GC-Q-TOF/MS detection techniques of 708 kinds of pesticide residues in the accurate mass data base of 708 kinds of pesticide and a kind of fruit and vegerable, achieve and do not need standard control, in edible fungi 708 kinds of pesticide residues can be carried out rapid detection detection, meet the urgent need that Agricultural Products Pesticide Residues high flux quickly detects.
On the one hand, the invention provides a kind of GC-Q-TOF/MS and detect the method for detecting of 708 kinds of pesticide residues in edible fungi, it is characterised in that: set up 708 kinds of pesticide TOF/MS data bases, in edible fungi 708 kinds of pesticide are carried out qualitative detecting and confirmation simultaneously; The method comprises the following steps:
(1) standard sample of pesticide is configured to respectively the standard solution of 1-5mg/L, impose a condition the lower full modal data measuring every kind of standard solution through the full modal data of GC-QTOF/MS first class mode, respectively obtains the chemical molecular formula of every kind of pesticide, retention time, fragment ion accurate mass number, abundance of ions ratio;
(2) the full modal data of first class mode that will obtain in step (1), fragment ion full scan mass spectrum after verified imports in PCDL software, and be associated with corresponding pesticide information, build up TOF/MS accurate mass data base, for data retrieval analysis;
(3) the full modal data measuring testing sample Pesticides of edible fungi under the imposing a condition identical with step (1), obtains the chemical molecular formula of its pesticide contained, retention time, fragment ion accurate mass number, abundance of ions ratio;
(4) to sample determination result through accurate mass database retrieval, the detection pesticide meeting appraisal basis is carried out precise Identification, thus the residual kind of agriculture obtained in edible fungi.
Preferably, by edible fungi carrying out pre-treatment to obtain the mensuration testing sample of edible fungi, described pre-treating method includes following procedure:
Sample preparation: edible fungi sample extracts through 0.5-5 volume % acetate acetonitrile homogenate, after dehydration and centrifugal, concentration, then through Carbon/NH2 column purification, acetonitrile+toluene eluting residual pesticide, through reconcentration, filter after make and treat sample measuring liquid a.
The present invention is had the advantages that
1, the present invention is with compound information such as high-resolution fragment ion accurate mass number, retention time, abundance of ions ratios for criterion of identification, the 708 kinds of pesticide detectings set up and confirmation technical method, revolutionize original qualitative model being reference with compound standard thing, it is that a kind of reference material that do not need compares, quick, high flux, accurately and reliably Detecting Pesticide new technique.
2, the present invention is under the premise not needing reference material to compare, it is achieved that the qualitative detecting of 708 kinds of pesticide residues and confirmation, it is possible to be greatly saved the cost buying reference material, also more environmentally-friendly, safety.
3, the present invention can complete the detecting of 708 kinds of pesticide in 1 hour, qualitative with confirmation, compared with traditional detection method, it is possible to improve work efficiency hundreds times.
4, the present invention can accomplish that single injected sampling detects, compared with traditional detection method to 708 kinds of disposable extraction and cleanings of pesticide residues in edible fungi sample, it is possible to saves testing cost, improve work efficiency hundreds times.
5, the edible fungi 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 pesticide, cinerins pesticide and other kinds of pesticide and metabolite.
6, the GC-QTOF/MS detection techniques index that the present invention sets up: sweep limits 50-600m/z; The accurate mass measured is up to 0.0001m/z; Within Mass accuracy can be controlled in 20ppm; Scanning speed ranges for 5 times per second at 50-600m/z.
7, in a kind of application gas chromatogram level Four bar flight time mass spectrum (GC-Q-TOF/MS) rapid screening edible fungi provided by the invention, the high resolution mass spectrum detection techniques of 708 kinds of pesticide residues obtains promotion and application at 7 pesticide residue analysis laboratorys, through the detecting of the 30 multiple commercially available edible fungi samples of Duo Ge provinces and cities is verified, achieve important residual detecting data.
8, in the 708 kinds of pesticide simultaneously detected, detecting sensitivity, without exception lower than the micro-g kg of standard 10, meets the requirement of the horizontal examination of various countries pesticide residues MRL.
Other features and advantages of the present invention will be described in detail in detailed description of the invention part subsequently.
Accompanying drawing explanation
Accompanying drawing is used to provide a further understanding of the present invention, and constitutes the part of description, is used for explaining the present invention, but is not intended that limitation of the present invention together with detailed description below. In the accompanying drawings:
Fig. 1 is Beijing area commercially available mushroom sample GC-Q-TOF/MS technology frequency the highest pesticide (phthalimide) result example.
Fig. 2 is the result that the Information in Mass Spectra of the highest pesticide of the frequency (phthalimide) of Beijing area commercially available mushroom sample GC-Q-TOF/MS technology detection is compared with data base.
Fig. 3 is Chongqing region commercially available mushroom sample GC-Q-TOF/MS technology frequency the highest pesticide (bioresmethrin) result example.
Fig. 4 is the result that the Information in Mass Spectra of the highest pesticide of the frequency (bioresmethrin) of Chongqing region commercially available mushroom sample GC-Q-TOF/MS technology detection is compared with data base.
Fig. 5 is In Chengdu commercially available mushroom sample GC-Q-TOF/MS technology frequency the highest pesticide (propisochlor) result example
Fig. 6 is the result that the Information in Mass Spectra of the highest pesticide of the frequency (propisochlor) of In Chengdu commercially available mushroom sample GC-Q-TOF/MS technology detection is compared with data base.
Detailed description of the invention
Hereinafter the specific embodiment of the present invention is described in detail. It should be appreciated that detailed description of the invention described herein is merely to illustrate and explains the present invention, it is not limited to the present invention.
The invention provides a kind of GC-Q-TOF/MS and detect the method for detecting of 708 kinds of pesticide residues in edible fungi, it is characterised in that: set up 708 kinds of pesticide TOF/MS data bases, in edible fungi 708 kinds of pesticide are carried out qualitative detecting and confirmation simultaneously; The method comprises the following steps:
(1) standard sample of pesticide is configured to respectively the standard solution of 1-5mg/L, impose a condition the lower full modal data measuring every kind of standard solution through the full modal data of GC-QTOF/MS first class mode, respectively obtains the chemical molecular formula of every kind of pesticide, retention time, fragment ion accurate mass number, abundance of ions ratio;
(2) the full modal data of first class mode that will obtain in step (1), fragment ion full scan mass spectrum after verified imports in PCDL software, and be associated with corresponding pesticide information, build up TOF/MS accurate mass data base, for data retrieval analysis;
(3) the full modal data measuring testing sample Pesticides of edible fungi under the imposing a condition identical with step (1), obtains the chemical molecular formula of its pesticide contained, retention time, fragment ion accurate mass number, abundance of ions ratio;
(4) to sample determination result through accurate mass database retrieval, the detection pesticide meeting appraisal basis is carried out precise Identification, thus the residual kind of agriculture obtained in edible fungi.
Concrete, illustrate that for phenthoate dimephenthoate cidial (Phenthoate) the GC-QTOF/MS data base of the present invention is set up by following method:
1) at data acquisition interface, phenthoate dimephenthoate cidial (Phenthoate) reference material (1-5mg/L) is carried out fragment ion full scan mass spectrum collection through GC-QTOF/MS first class mode under specifying chromatograph Mass Spectrometry Conditions.
Chromatographic condition is: chromatographic column: VF-1701MS (30m × 0.25mm × 0.25um) (J&WScientific, Folsom, CA); Chromatographic column heating schedule: 30-50 DEG C keeps 0.5-5min, then to be warming up to 200-280 DEG C with 3-10 DEG C/min again, then it is warming up to 110-150 DEG C with 20-50 DEG C/min, it is warming up to 200-280 DEG C again with 3-10 DEG C/min, it is warming up to 250-320 DEG C subsequently with 5-15 DEG C/min, keep 3-10min, then be cooled to 200-290 DEG C with 5-20 DEG C/min, keep 1-5min; Carrier gas: helium; Purity >=99.999%; 0.5-2mL/min; Injector temperature: 230-320 DEG C; Sample size 1 μ L; Input mode: Splitless injecting-Sample, solvent delay 2-10min. Preferably, chromatographic condition: chromatographic column: VF-1701MS (30m × 0.25mm × 0.25um) (J&WScientific, Folsom, CA); Chromatographic column heating schedule: 40 DEG C keep 1min, are then warming up to 130 DEG C with 30 DEG C/min, then are warming up to 250 DEG C with 5 DEG C/min, it is warming up to 300 DEG C with 10 DEG C/min subsequently, keeps 5min, then be cooled to 280 DEG C with 10 DEG C/min, keep 2min carrier gas: helium, purity >=99.999%; Flow velocity: 1.2mL/min; Injector temperature: 290 DEG C; Sample size 1 μ L; Input mode: Splitless injecting-Sample, solvent delay 6min.
Mass Spectrometry Conditions is: ion source: EI (electron impact ionization source); Ion source temperature: 230-330 DEG C, electron bombardment ionization source energy: 50-100eV; Gas phase-mass spectrometer interface temperature: 200-320 DEG C; Scan mode: carry out TOF pattern full scan, mass number scope 50-600amu in the way of 2-10 per second opens spectrogram. Preferably, Mass Spectrometry Conditions: ion source: EI (electron impact ionization source); Ion source temperature: 280 DEG C, electron bombardment ionization source energy: 70eV; Gas phase-mass spectrometer interface temperature: 290 DEG C; Scan mode: carry out TOF pattern full scan, mass number scope 50～600amu. in the way of 5 spectrograms per second.
The described mass spectrum information that fragment ion full scan mass spectrum collection is phenthoate dimephenthoate cidial (Phenthoate) reference material, including compound chemistry molecular formula, retention time, fragment ion accurate mass number, abundance of ions ratio.
2) in the qualitative software of AgilentMassHunter (instrument carries), invocation target pesticide (Phenthoate) gathers file, and determines target pesticide chromatographic peak.
3), after opening target pesticide mass spectrum background correction, use low spectrum storehouse (instrument carries) of differentiating of NIST11 to scan for obtaining the information of compound, and verify library searching to compound whether be consistent with actual.
4) mass spectrum is verified, target compound mass spectrum can show composes, through low resolution of NIST11, fragment ion molecular formula and the accurate mass number that storehouse matches, by " EditPeakAnnotion " window, ion information is carried out the amendment of necessity, the ion that such as accurate mass number deviation is excessive, might mean that the Formula structure that software calculates automatically is wrong, now manually can input correct information in " Formula " string, or " IronType " is changed to Unknown.
5) the fragment ion full scan mass spectrum after verified imports in PCDL software (instrument carries), and is associated with phenthoate dimephenthoate cidial information, builds up TOF/MS data base. The mass spectrum information of 708 kinds of pesticide in data base, for details see attached table 1.
Preferably, described pesticide is 708 shown in table a kind pesticide. The method forming these pesticide can carry out according to the mode of the TOF/MS data base of phenthoate dimephenthoate cidial as formed above (Phenthoate).
According to the present invention, the mass spectrum information of the reference material obtained can being carried out according to PCDL software document with the corresponding pesticide method of being mutually related by PCDL software, the present invention does not repeat them here.
According to the invention it is preferred to, described edible fungi selects in the group that free Rhizoma Solani tuber osi, Radix Raphani, Radix Dauci Sativae, Fructus Colocasiae Esculentae, Rhizoma Steudnerae Henryanae, Radix Platycodonis, Rhizoma Dioscoreae esculentae, yacon, Rhizoma Zingiberis Recens and Rhizoma Dioscoreae form.
Table 1
According to the present invention, and set up the full modal data measuring testing sample Pesticides of edible fungi under identical imposing a condition, obtain the chemical molecular formula of its pesticide contained, retention time, fragment ion accurate mass number, abundance of ions ratio; Call the TOF/MS data base set up, according to TOF/MS data base to target compound retrieval by header, search argument retention time deviation is defined to ± 0.15min, accurate mass deviation is defined to ± 20ppm, and software can provide the retrieval matching score value of each detection ion according to the measured value of the key elements such as every kind of Compound Retention time, accurate mass with the deviation of theoretical value in TOF/MS data base automatically. Detection ion score averages is the comprehensive score of this compound, specify each compound score value > 60 and the detection of at least three ion, it is positive detection, according to the method accuracy of the qualitative detecting Residual Pesticides in Farm Produce of TOF/MS data base, reaches more than 90%.
According to the present invention, by edible fungi carrying out pre-treatment to obtain the mensuration testing sample of edible fungi, described pre-treating method preferably includes following procedure:
Edible fungi sample is extracted through acetonitrile solution (the be called for short 0.5-5 volume % acetate acetonitrile) homogenate containing 0.5-5 volume % acetic acid, after dehydration and centrifugal, concentration, then through Carbon/NH2Column purification, acetonitrile+toluene eluting residual pesticide, make after reconcentration, filtration and treat sample measuring liquid;
Wherein, it is preferred that described edible fungi sample size is 5-20g, the consumption that consumption is 30-50mL of the acetate acetonitrile of described 0.5-5 volume % is preferably 35-45ml; And the fruit and vegerable sample adding acetate acetonitrile is placed in centrifuge tube and carries out homogenized. Described homogenate extraction conditions is preferably: rotating speed 12000-15000rpm, time 0.5-2min.
Wherein, described through dehydration preferably by the sodium chloride adding 0.5-5g in acetate acetonitrile extracting solution, the anhydrous magnesium sulfate of 2-8g, and vibrate 4-6min realize. Then the dehydrated sample centrifugal 1-10min under 4000-4500rpm that will obtain, obtains supernatant, and the supernatant 10-30mL rotary evaporation in the water-bath of 20-60 DEG C obtained is concentrated into 0.5-2mL.
Wherein, described Carbon/NH2Column purification step is preferably as follows: at Carbon/NH2Post adds the preferred 2cm height anhydrous sodium sulfate of about 1-3cm. First with 2-6mL preferred 4mL acetonitrile+toluene (preferred 2.5-3.5: 1, more preferably 3: 1, v/v) drip washing SPE post, and discard effluent, when liquid level arrive sodium sulfate top time, rapidly sample concentration liquid is transferred on decontaminating column, under connect new Cor Gigeriae Galli bottle graft receive. Each 2-6mL preferred 2mL acetonitrile+toluene (preferred 2.5-3.5: 1, more preferably 3: 1, v/v) washs sample liquid bottle three times again, and is moved into by cleaning mixture in SPE post. Again with 20-30mL preferred 25mL acetonitrile+toluene (preferred 2.5-3.5: 1, more preferably 3: 1, v/v) eluting pesticide, merge eluent in Cor Gigeriae Galli bottle.
Wherein, described treat sample measuring liquid it is preferred that be prepared in the following way, by eluent obtained as described above spin concentration extremely about 0.5-2mL in 30-50 DEG C of water-bath, and concentrated solution is placed under nitrogen and dries up, add acetonitrile+0-5% formic acid water (2+ (7-9), it is preferable that 2+8, the v/v) mixing of 1-5mL, constant volume after 0.18-0.22 μm preferably 0.2 μm of membrane filtration, obtains treating sample measuring liquid.
The specific embodiment of the present invention described in detail below.
Embodiment 1
The present embodiment is for illustrating the method for detecting that in GC-Q-TOF/MS examination edible fungi provided by the invention, 708 kinds of agricultures are residual
(1) 10g Beijing area mushroom sample (being accurate to 0.01g) is weighed, in 80ml centrifuge tube, add the acetonitrile solution of 40mL1% acetic acid, with high-speed homogenization machine 13500r/min, 1min is extracted in homogenate, adding 1g sodium chloride, 4g anhydrous magnesium sulfate, vibrate 5min, centrifugal 5min under 4200r/min, taking supernatant 20mL, in 40 DEG C of water-baths, rotary evaporation is concentrated into about 1mL, to be clean.
(2) 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 sample concentration liquid be transferred in SPE post, under connect new Cor Gigeriae Galli bottle graft and receive. Wash sample liquid bottle three times with 2mL acetonitrile+toluene (3+1, v/v), and cleaning mixture is moved in SPE post every time again. Post connects 50mL liquid reservoir, with 25mL acetonitrile+toluene (3+1, v/v) eluting pesticide and related chemicals, is incorporated in Cor Gigeriae Galli bottle, and in 40 DEG C of water-baths spin concentration to about 0.5mL.
(3) concentrated solution is placed under nitrogen and dries up, and adds acetonitrile+water (2+8, v/v) mixing of 2mL, constant volume after 0.2 μm of membrane filtration, obtains testing sample solution.
(4) GC-Q-TOF/MS operating condition, separates and obtains the credit minor of each fragment ion of each pesticide in mushroom sample, retention time, fragment ion accurate mass number, abundance of ions ratio.
Chromatographic condition: chromatographic column: VF-1701MS (30m × 0.25mm × 0.25um) (J&WScientific, Folsom, CA); Chromatographic column heating schedule: 40 DEG C keep 1min, are then warming up to 130 DEG C with 30 DEG C/min, then are warming up to 250 DEG C with 5 DEG C/min, be warming up to 300 DEG C with 10 DEG C/min subsequently, keep 5min, then are cooled to 280 DEG C with 10 DEG C/min, keep 2min; Carrier gas: helium, purity >=99.999%; Flow velocity: 1.2mL/min; Injector temperature: 290 DEG C; Sample size 1 μ L; Input mode: Splitless injecting-Sample, solvent delay 6min.
Mass Spectrometry Conditions: Mass Spectrometry Conditions: ion source: EI (electron impact ionization source);Ion source temperature: 280 DEG C, electron bombardment ionization source energy: 70eV; Gas phase-mass spectrometer interface temperature: 290 DEG C; Scan mode: carry out TOF pattern full scan, mass number scope 50～600amu. in the way of 5 spectrograms per second.
(5) according to the data base of table 1 to acquisition information retrieval by header in step (4), search argument retention time deviation is defined to ± 0.5min, accurate mass deviation is defined to ± 100ppm, detection at least three fragment ion, and according to the measured value of every kind of each key element of compound and the deviation of theoretical value in table 1 data base, automatically provide retrieval matching score value, the compound of retrieval matching score value > 60, namely confirm to detect this target compound. Testing result is in Table 2.
For the phthalimide that frequency is higher, as shown in Figure 1, mushroom detects 5 fragment ion information, with database retrieval, with phthalimide comparison, 5 fragment ion molecular weight deviation scores are all more than 99 points, retention time deviation is within 0min, the fragment ion masses deviation that abundance is the highest is 1.56ppm (as shown in Figure 2), it is possible to confirms, has phthalimide to detect in mushroom.
Embodiment 2
The present embodiment is for illustrating the method for detecting that in GC-Q-TOF/MS examination edible fungi provided by the invention, 708 kinds of agricultures are residual
Detect with the agriculture in batch mushroom sample is residual according to the method in embodiment 1, the difference is that, pre-treatment step is as follows:
(1) accurately weighing 10.00g sample in centrifuge tube, add 50mL water, vibration is until sample is fully infiltrated by water. Pipette 50mL acetonitrile to centrifuge tube after freezing 10min, then centrifuge tube is placed on vortex mixing shaker, with 2000rpm rate oscillation 1min. Centrifuge tube is placed under-10 DEG C of conditions and keeps 10min, then in centrifuge tube, add 20g anhydrous magnesium sulfate and 5g sodium chloride, 5g sodium citrate and 2.5g monobasic sodium citrate, immediately on whirlpool mixing shaker, with 2000rpm rate oscillation 2min, then with the centrifugal 10min of 4000rpm speed, supernatant is obtained.
(2) pipette supernatant 1.0mL in 1.5mL centrifuge tube, add the PSA adsorbent of 150mg anhydrous magnesium sulfate and 25mg, with 2000rpm rate oscillation 2min on whirlpool mixing shaker, with the centrifugal 2min of 6000rpm speed. Aspirate supernatant, through 0.22 μm of organic facies membrane filtration, pipettes 200 μ L, adds 100 μ L acetonitriles, is diluted to 1.0mL with ultra-pure water, obtains testing sample solution.
Testing result is in Table 2.
Table 2
By data above it can be seen that the method adopting the present invention, especially preferred method more effective can detect the kind that agricultural product middle peasant is residual, and also has less detection limit.
Embodiment 3
The present embodiment is for illustrating the method for detecting that in GC-Q-TOF/MS examination edible fungi provided by the invention, 708 kinds of agricultures are residual
Detect the agriculture in edible fungi is residual according to the method in embodiment 1, the difference is that, sample is Chongqing region mushroom sample.
Gathering 10, the commercially available mushroom sample in Chongqing region, detect 3 kinds of pesticide residues, amount to 3 frequencys, relate to 1, sample, concrete outcome is in Table 3.
Table 3
For the bioresmethrin that frequency is higher: as shown in Figure 3, mushroom detects 6 fragment ion information, with database retrieval, with bioresmethrin comparison, 6 fragment ion coupling marks are all more than 96 points, and retention time deviation is within 0.008min, and the fragment ion masses deviation that abundance is the highest is-1.79ppm (as shown in Figure 4), it has been confirmed that mushroom there is bioresmethrin detect.
Embodiment 4
The present embodiment is for illustrating the method for detecting that in GC-Q-TOF/MS examination edible fungi provided by the invention, 708 kinds of agricultures are residual
Detect the agriculture in edible fungi is residual according to the method in embodiment 1, the difference is that, sample is In Chengdu mushroom sample.
Gathering 13, the commercially available mushroom sample in In Chengdu, detect 3 kinds of pesticide residues, amount to 3 frequencys, relate to 3, sample, concrete outcome is in Table 4.
Table 4
For the propisochlor that frequency is higher: as shown in Figure 5, mushroom detects 7 fragment ion information, with database retrieval, with propisochlor comparison, 7 fragment ion molecular weight deviation scores are all more than 63 points, and retention time deviation is 0.008min, and the fragment ion masses deviation that abundance is the highest is-5.38ppm (as shown in Figure 6), it has been confirmed that mushroom there is propisochlor detect.
The preferred embodiment of the present invention described in detail above; but, the present invention is not limited to the detail in above-mentioned embodiment, in the technology concept of the present invention; technical scheme can being carried out multiple simple variant, these simple variant belong to protection scope of the present invention.
It is further to note that each the concrete technical characteristic described in above-mentioned detailed description of the invention, in reconcilable situation, it is possible to be combined by any suitable mode. In order to avoid unnecessary repetition, various possible compound modes are no longer illustrated by the present invention separately.
Additionally, can also carry out combination in any between the various different embodiment of the present invention, as long as it is without prejudice to the thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (10)
1. a GC-Q-TOF/MS detects the method for detecting of 708 kinds of pesticide residues in edible fungi, it is characterised in that: set up 708 kinds of pesticide TOF/MS data bases, in edible fungi 708 kinds of pesticide are carried out qualitative detecting and confirmation simultaneously; The method comprises the following steps:
(1) standard sample of pesticide is configured to respectively the standard solution of 1-5mg/L, impose a condition the lower full modal data measuring every kind of standard solution through the full modal data of GC-QTOF/MS first class mode, respectively obtains the chemical molecular formula of every kind of pesticide, retention time, fragment ion accurate mass number, abundance of ions ratio;
(2) the full modal data of first class mode that will obtain in step (1), fragment ion full scan mass spectrum after verified imports in PCDL software, and be associated with corresponding pesticide information, build up TOF/MS accurate mass data base, for data retrieval analysis;
(3) the full modal data measuring testing sample Pesticides of edible fungi under the imposing a condition identical with step (1), obtains the chemical molecular formula of its pesticide contained, retention time, fragment ion accurate mass number, abundance of ions ratio;
(4) to sample determination result through accurate mass database retrieval, the detection pesticide meeting appraisal basis is carried out precise Identification, thus the residual kind of agriculture obtained in edible fungi.
2. method according to claim 1, wherein, described pesticide is 708 shown in table a kind pesticide.
3. method according to claim 1, wherein, described edible fungi selects in the group that free mushroom, Pleurotus eryngii, Lentinus Edodes, Flammulina velutiper (Fr.) Sing and Pleurotus ostreatus form.
4. method according to claim 1, wherein, the condition of the full modal data measuring every kind of standard solution Pesticides includes:
(1) chromatographic condition: chromatographic column: VF-1701MS; Chromatographic column heating schedule: 30-50 DEG C keeps 0.5-5min, then it is warming up to 110-150 DEG C with 20-50 DEG C/min, it is warming up to 200-280 DEG C again with 3-10 DEG C/min, it is warming up to 250-320 DEG C subsequently with 5-15 DEG C/min, keep 3-10min, it is cooled to 200-290 DEG C with 5-20 DEG C/min again, keeps 1-5min;Carrier gas: helium; Flow velocity: 0.5-2mL/min; Injector temperature: 230-320 DEG C: Splitless injecting-Sample, solvent delay 2-10min;
(2) Mass Spectrometry Conditions: ion source: EI; Ion source temperature: 230-330 DEG C, electron bombardment ionization source energy: 50-100eV; Gas phase-mass spectrometer interface temperature: 200-320 DEG C; Scan mode: carry out TOF pattern full scan, mass number scope 50-600amu in the way of 2-10 per second opens spectrogram.
5. method according to claim 1, wherein, TOF/MS accurate mass data base can will measure the measured value of the retention time of the fragment ion obtained, accurate mass number, abundance of ions ratio from testing sample, with the corresponding key element theoretical value retrieval comparison of fragment ion in TOF/MS accurate mass data base, provide the score of each fragment ion, and then provide the matching degree score value of average mark and target compound; Wherein, the retention time deviation setting of fragment ion is at ± 0.5min, and accurate mass number deviation setting is ± 100ppm.
6. method according to claim 5, wherein: by the following method the detection pesticide meeting appraisal basis is carried out precise Identification:
By characteristic ion being detected situation and goodness of fit, the score value provided, pesticide is carried out qualitative recognition, its method program is: for the detecting of target compound in sample, the retention time of the compound that flight time mass spectrum is recorded, the result of 7 fragment ion accurate mass numbers that abundance is the highest, fragment ion abundance ratio, with TOF/MS database retrieval comparison, result is detection at least three fragment ion, and the comprehensive score pesticide more than 60 is confirmed as detection pesticide.
7. method according to claim 1, by edible fungi carrying out pre-treatment to obtain the mensuration testing sample of edible fungi, described pre-treating method includes following procedure:
Sample preparation: edible fungi sample extracts through 0.5-5 volume % acetate acetonitrile homogenate, after dehydration and centrifugal, concentration, then through Carbon/NH2Column purification, acetonitrile+toluene eluting residual pesticide, make after reconcentration, filtration and treat sample measuring liquid a.
8. method according to claim 7, wherein: described edible fungi sample size is 5-20g;
Preferably, the consumption of the acetate acetonitrile of described 0.5-5 volume % is 30-50mL;
Preferably, described homogenate extraction conditions is: rotating speed 12000-15000r/min, and the time is 0.5-5min;
Preferably, described referring to through dehydration adds 0.5-5g sodium chloride in 0.5-5 volume % acetate acetonitrile extracting solution, 2-8g anhydrous magnesium sulfate, and vibrate 1-10min;
Preferably, described centrifugal referring to is centrifuged 1-10min under 3000-5000r/min;
Preferably, described concentration refers to and takes centrifuged supernatant 10-30mL, and in 20-60 DEG C of water-bath, rotary evaporation is concentrated into about 0.5-2mL.
9. method according to claim 8, wherein: described Carbon/NH2Column purification step is as follows: at Carbon/NH2Post adds about 1-3cm height anhydrous sodium sulfate, first with a certain proportion of acetonitrile of 2-6mL and toluene mixed solution drip washing SPE post, and discards effluent, when liquid level arrives the top of sodium sulfate, rapidly sample concentration liquid is transferred on decontaminating column, under connect container reception; Wash sample liquid bottle three times with 2-6mL acetonitrile and toluene mixed solution more every time, and cleaning mixture is moved in SPE post. Again with the certain volume acetonitrile of preferred 20-30ml and toluene mixed solution eluting pesticide, merge eluent in container;
Preferably, described a certain proportion of acetonitrile and toluene mixed solution, acetonitrile: volume of toluene ratio is for 2.5-3.5: 1.
10. method according to claim 8, wherein: described making after reconcentration, filtration treats that sample measuring liquid a refers to the eluent that claim 9 is obtained, spin concentration extremely about 0.5-2mL in 30-50 DEG C of water-bath, and concentrated solution is placed under nitrogen and dries up, add a certain proportion of acetonitrile and the mixing of 0-5 volume % formic acid water mixed solution of 1-5mL, constant volume after 0.18-0.22 μm of membrane filtration, obtains testing sample solution a.
Preferably, described a certain proportion of acetonitrile and 0-5 volume % formic acid water mixed solution, acetonitrile: 0.1-5 volume % formic acid water 2: 7-9.
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