CN110068622A - Acetamide-group herbicides efficiently separates method and rapid detection method in miscellaneous beans - Google Patents
Acetamide-group herbicides efficiently separates method and rapid detection method in miscellaneous beans Download PDFInfo
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- CN110068622A CN110068622A CN201910238919.7A CN201910238919A CN110068622A CN 110068622 A CN110068622 A CN 110068622A CN 201910238919 A CN201910238919 A CN 201910238919A CN 110068622 A CN110068622 A CN 110068622A
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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
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
- G01N2030/065—Preparation using different phases to separate parts of sample
Abstract
The invention discloses acetamide-group herbicides in miscellaneous beans to efficiently separate method and rapid detection method, wherein, separation method was the following steps are included: miscellaneous beans were clayed into power 80 meshes, miscellaneous bean powder end and water are mixed and adjust pH=7, oscillation, continue to add extractant (ethyl acetate mixes in equal volume with n-hexane) and NaCl, ultrasonic extraction 2min, then it is centrifuged at 0 DEG C with 15000rpm, supernatant is transferred in the centrifuge tube containing adsorbent MIL-101 (Zn), it is stirred, then it is centrifuged with 10000rpm, remove supernatant, methanol is added into centrifuge tube, ultrasonic elution is carried out to analyte.The invention has the beneficial effects that: (1) it is nonpolar using ethyl acetate and n-hexane isometric mix reagent as extractant, acetamide-group herbicides can be extracted from miscellaneous beans；(2) absorption that acetamide-group herbicides are carried out with MIL-101 (Zn), realizes the separation of acetamide-group herbicides, fat, protein etc.；(3) ultrasound elution realizes the purification of acetamide-group herbicides.
Description
Technical field
The present invention relates to the separation method of compound and detection methods, and in particular to acetamide-group herbicides is effective in miscellaneous beans
Separation method and rapid detection method, belong to detection technique field.
Background technique
Acetamide-group herbicides are one of the herbicides largely used in the world.Currently, about acetamide-group herbicides vegetables,
Detection method and residue dynamics on the crops such as peanut, green soy bean have some researchs, but its residue detection in coarse cereals is ground
It is less to study carefully report.
Currently, the biggish kind of acetamide-group herbicides sales volume has: propanil, metazachlor, pretilachlor, butachlor.These are removed
The herbicidal effect of careless agent is good, but it is careless slightly easily there is phytotoxicity, such as: pretilachlor has the arthropods such as honeybee, shrimp, crab
Stronger toxicity, butachlor can cause the red blood cell of catfish make a variation, induce people chromatid deformity.
Acetamide-group herbicides have very strong water-soluble and mobility and lower adsorption by soil constant, are easy to leaching,
It is easy to be transferred in phreatic water by infiltration, or enters in surface water with rainfall runoff, cause water pollution.
Many researchs have confirmed that many herbicides are all harmful to human body and biotic environment.Such as: Hill etc. is reported
The sister chromosomes that the metabolins such as two hydrocarbon benzoquinone imines of acetamide-group herbicides can induce the Human Lymphocytes manually cultivated produce
Raw exchange.
Currently, China only defines the maximum residue limit of a small number of herbicides, for the acyl in most of especially coarse cereals
There is no provide for amine herbicide maximum residue limit.
Summary of the invention
The first purpose of this invention is that the acetamide-group herbicides in miscellaneous beans can be efficiently separated out by providing one kind
The method come.
Second object of the present invention is to provide a kind of amide that can rapidly and effectively detect in miscellaneous beans sample
The method of class herbicide.
In order to realize above-mentioned first aim, the present invention is adopted the following technical scheme that:
The method of acetamide-group herbicides is efficiently separated from miscellaneous beans, which comprises the following steps:
Step1: miscellaneous beans are clayed into power, and cross 80 meshes；
Step2: miscellaneous bean powder end and water are added in centrifuge tube, adjust pH value of solution=7, oscillation；
Step3: continue that extractant and NaCl, ultrasonic extraction 2min are added into centrifuge tube, then at 0 DEG C with 15000rpm
Centrifugation, aforementioned extractant are that ethyl acetate mixes in equal volume with n-hexane, and the ratio of miscellaneous bean powder end and extractant is 1g:2mL；
Step4: supernatant being transferred in the centrifuge tube of another Zhi Hanyou adsorbent, be stirred, then with
10000rpm centrifugation, preceding adsorption agent are MIL-101 (Zn), and the ratio of miscellaneous bean powder end and adsorbent is 100g:14g；
Step5: removing supernatant, methanol be added into centrifuge tube, carries out ultrasonic elution to analyte.
In order to realize above-mentioned second target, the present invention is adopted the following technical scheme that:
Quickly detect the method for acetamide-group herbicides in miscellaneous beans sample, which comprises the following steps:
Step1: miscellaneous beans sample is clayed into power, and crosses 80 meshes；
Step2: the miscellaneous bean powder end 0.4g and 0.4mL water are added in centrifuge tube, adjust pH value of solution=7, with forced oscillation, after
Continue addition 0.8mL extractant and 40mg NaCl, ultrasonic extraction 2min into centrifuge tube to be then centrifuged at 0 DEG C with 15000rpm
Supernatant is transferred in the centrifuge tube of another Zhi Hanyou adsorbent, is stirred by 5min, is centrifuged 3min with 10000rpm, removes
Supernatant is removed, 2ml methanol is added into centrifuge tube, ultrasonic elution is carried out to analyte, wherein aforementioned extractant is ethyl acetate
It is mixed in equal volume with n-hexane, preceding adsorption agent is MIL-101 (Zn)；
Step3: eluent is transferred in chicken heart bottle, rotates the obtained solution filter with 100 μ L methanol back dissolvings after being evaporated
Film filtering, finally analyzes sample with high performance liquid chromatograph.
Method above-mentioned, which is characterized in that in Step2, adjust pH value of solution using 0.5mol/LNaOH.
Method above-mentioned, which is characterized in that in Step3, rotating the temperature being evaporated is 35 DEG C.
Method above-mentioned, which is characterized in that in Step3, what aforementioned filter membrane was selected is 0.22 μm of teflon membrane filter.
Method above-mentioned, which is characterized in that in Step3, chromatographic condition are as follows:
Chromatographic column: Eclipse XDB-C18 column, 150mm × 4.6mm, 3.5 μm；
Column temperature: 30 DEG C；
Mobile phase A: water；
Mobile phase B: acetonitrile；
Linear gradient elution program: 0-5min, 40-60%B；5-9min, 60-80%B；9-13.5min 80-79.5%
B；13.5-14.7min 79.5%B；14.7-16.0min 79.5-79.4%B；16.0-21.0min 79.4-79.0%B；
21-25min, 79-60%B；
Flow rate of mobile phase: 0.50mL/min；
Sample volume: 20 μ L.
The invention has the beneficial effects that:
(1) method of the acetamide-group herbicides in miscellaneous beans is separated
Using ethyl acetate and the isometric mix reagent of n-hexane as extractant, which is nonpolar, so can
Acetamide-group herbicides to be extracted from miscellaneous beans；
Since fat content and protein content are all very high in miscellaneous beans, these fat and protein also can be by nonpolar extractions
It takes agent to extract together, so joined adsorbent MIL-101 (Zn) in our supernatants after the extraction, uses MIL-101
(Zn) absorption for carrying out acetamide-group herbicides, realizes the separation of acetamide-group herbicides, fat, protein etc.；
We have carried out ultrasonic elution to analyte again after separation, just only remaining acetamide-group herbicides in the solution after elution
, the purification of acetamide-group herbicides is thereby realized, the stroma contaminants such as fat and protein is avoided and enters liquid chromatogram
The detection of acetamide-group herbicides is interfered in instrument, this lays a good foundation for effective detection of subsequent acetamide-group herbicides.
(2) method of the acetamide-group herbicides in miscellaneous beans sample is detected
On the basis of effectively extracting, separating, purify the acetamide-group herbicides in miscellaneous beans, using high performance liquid chromatograph
Sample is analyzed, so the acetamide-group herbicides in impurity removal beans sample can be detected rapidly and effectively.
Detailed description of the invention
Fig. 1 is the composition principle figure of MIL-101 (Zn)；
Fig. 2 (a) is the XRD spectrum of the MIL-101 (Zn) of synthesis；
Fig. 2 (b) is the electron-microscope scanning figure of the MIL-101 (Zn) of synthesis；
Fig. 3 is influence diagram of the extractant type to effect of extracting；
Fig. 4 is influence diagram of the ultrasonic extraction time to effect of extracting；
Fig. 5 is influence diagram of the adsorbent amount to effect of extracting；
Fig. 6 is influence diagram of the eluent dosage to four kinds of acetamide-group herbicides rate of recovery；
Fig. 7 (a) is the chromatogram of the hybrid standard working solution of 10 μ g/mL；
Fig. 7 (b) is the chromatogram of blank sample；
Fig. 7 (c) is the chromatogram of mark-on sample.
Specific embodiment
Specific introduce is made to the present invention below in conjunction with the drawings and specific embodiments.
One, capital equipment
Agligent1260 type high performance liquid chromatograph: Agilent company, the U.S..
Eclipse XDB-C18 column (150mm × 4.6mm, 3.5 μm): Agilent company, the U.S..
Allegra 64R type high speed freezing centrifuge: U.S. Bake Mann.
RE-52AA vacuum rotary evaporator: Shanghai Yarong Biochemical Instrument Plant.
Millipore-q water purification system: Millipore company, the U.S..
Rigaku D/max-2550X ray powder diffractometer: Japanese Rigaku company, using Cu target (K α,)。
Tecnai G2F20S-Twin transmission electron microscope: Dutch Philips company.
Other: it is Philip HR-2870 grinder, diode array detector, automatic sampler, quaternary gradient pump, stainless
Steel reaction kettle (band polytetrafluoroethylliner liner).
The ultrapure water of test is all derived from Millipore-q water purification system.
Two, Primary Chemical and material
Propanil, metazachlor, pretilachlor, butachlor: country, BeiJing, China medicine bioengineering product controls research institute.
Acetonitrile (chromatographically pure): Fisher company, the U.S..
Hydrofluoric acid (HF, 40%): Nanjing chemical reagent factory.
Terephthalic acid (TPA) (H2BDC): fine chemistry industry research institute is recovered in Tianjin.
Zn(NO3)2·9H2O: Xilong Chemical Co., Ltd.
N,N-dimethylformamide (DMF), dehydrated alcohol, n-hexane: Tianjin Tian Tai fine chemicals Co., Ltd.
Other reagents are that analysis is pure, are purchased from Beijing Chemical Plant.
Three, standard reserving solution and hybrid standard working solution are prepared
Standard reserving solution is made with methanol in propanil, metazachlor, pretilachlor, butachlor respectively.
The hybrid standard working solution that concentration is 100 μ g/mL is made with standard reserving solution.
The lower hybrid standard working solution of concentration (0.05 μ g/ is made with the hybrid standard working solution that concentration is 100 μ g/mL
mL、0.1μg/mL、0.5μg/mL、1.0μg/mL、5.0μg/mL、10.0μg/mL、25.0μg/mL)。
All standard reserving solutions and hybrid standard working solution are all kept in dark place at 4 DEG C spare.
Four, prepare sample
In August, 2018, we have purchased semen sojae atricolor, red bean, kidney bean in the market of farm produce of Heilongjiang Province of China Daqing locality
Three kinds of miscellaneous beans.
These three miscellaneous beans have been worn into powder with Philip HR-2870 grinder by us respectively, and powder has been crossed 80 mesh
Sieve, obtains three powdered samples, is respectively as follows: semen sojae atricolor powder sample, red bean powder sample, kidney bean powder sample.
Fresh mark-on sample: mixing mark is added into semen sojae atricolor powder sample, red bean powder sample, kidney bean powder sample respectively
Quasi- working solution (pitch-based sphere is shown in Table 2), then vibrates 3min, in order to ensure acetamide-group herbicides can be relatively uniform be dispersed in
In sample powder, we also joined a certain amount of third into semen sojae atricolor powder sample, red bean powder sample, kidney bean powder sample
Then ketone carefully stirs evenly, be dried overnight at room temperature for 24 hours before extraction.
Five, MIL-101 (Zn) is synthesized
We are with Zn (NO3)2·6H2O and H2BDC is raw material, heated, and MIL-101 (Zn), MIL-101 has been made in reaction
(Zn) composition principle is as shown in Figure 1.
The synthesis process of MIL-101 (Zn) is specific as follows:
By 800mg Zn (NO3)2·6H2O、322mg H2BDC, 0.1mL HF and 9.5mL H2O is placed in stainless steel cauldron
Polytetrafluoroethylliner liner in, stir evenly, stainless steel cauldron be placed into 160 DEG C of baking oven and reacted after lid lid sealing
Product is washed repeatedly with DMF and dehydrated alcohol after reaction and (removes the complete H of unreacted at least three times by 8h2BDC), so
5min is centrifuged with the speed of 10000rpm afterwards, final product is finally put into baking oven 150 DEG C of dryings for 24 hours.
We have then carried out XRD analysis and morphology characterization to the MIL-101 of synthesis (Zn) and have analyzed, as a result such as Fig. 2 (a) and
Shown in Fig. 2 (b).
By Fig. 2 (a) it is found that the simulation map of the XRD spectrum for the product that we synthesize and MIL-101 reported in the literature (Zn)
Unanimously, illustrate that we are successfully prepared MIL-101 (Zn).
By Fig. 2 (b) it is found that the product MIL-101 (Zn) that we synthesize is cubic system crystal.
Since MIL-101 (Zn) has biggish aperture and hole window, so that the acetamide-group herbicides of small molecule are preferentially absorbed into
Enter in hole, forms hole diffusion, the unsatuated metal site phase interaction in hetero atom and MIL-101 (Zn) in acetamide-group herbicides
With the π-π reciprocation between phenyl ring and the ligand of MIL-101, the delocalizedπelectron and metal in acetamide-group herbicides in, amide
The Zn- π in unsaturated site interacts, these interact so that the acetamide-group herbicides in apolar extraction agent are by MIL-
101 (Zn) absorption, to be separated with matrix such as protein, fat in nonpolar solvent, so we select to use MIL-101
(Zn) the improvement adsorbent of QuEChERs (Quick, Easy, Cheap, Effective, Rugged, Safe) method is made,
QuEChERs method is a kind of quick sample pretreatment technology for detection of agricultural products that latest development is got up in the world in recent years,
We improve QuEChERs method, so that can more effectively isolate acetamide-group herbicides from miscellaneous beans.
Six, the acetamide-group herbicides in miscellaneous beans are separated
We using in liquid-phase extraction-miscellaneous beans of solid absorbent dispersive solid-phase extraction acetamide-group herbicides (such as: enemy
Barnyard grass, metazachlor, pretilachlor, butachlor), specific as follows:
Step1: semen sojae atricolor is clayed into power, and crosses 80 meshes；
Step2: 0.4g semen sojae atricolor powder and 0.4mL water are added in centrifuge tube, the NaOH tune for being 0.5mol/L with concentration
PH value of solution=7 are saved, with forced oscillation 20s；
Step3: continue that 0.8mL extractant and 0.04gNaCl, ultrasonic extraction 2min are added into centrifuge tube, then at 0 DEG C
With 15000rpm be centrifuged 5min, wherein extractant be ethyl acetate mixed in equal volume with n-hexane (ethyl acetate 0.4mL, just oneself
Alkane 0.4mL)；
Step4: supernatant is transferred in the centrifuge tube of another Zhi Hanyou 56mg adsorbent, is stirred 2min, then
3min is centrifuged with 10000rpm, wherein adsorbent is MIL-101 (Zn)；
Step5: removing supernatant, and 2mL methanol is added into centrifuge tube, carries out ultrasonic elution to analyte, collects elution
Liquid.
The method of the separating amide class herbicide provided by the invention from miscellaneous beans, is mixed in equal volume with ethyl acetate and n-hexane
It closes reagent and makees extractant, wherein n-hexane is nonpolar solvent, can effectively be penetrated into sample, and ethyl acetate is medium
Polar solvent, it is preferable to the solubility of the acetamide-group herbicides in sample, after ethyl acetate and n-hexane mixing, can more have
The extraction of the realization acetamide-group herbicides of effect.
The method of the separating amide class herbicide provided by the invention from miscellaneous beans, the mixing of salt is replaced with a small amount of NaCl
Object effectively avoids the salt agglomeration in solid sample.
The conventional method of separating amide class herbicide from miscellaneous beans has: solid phase extraction, liquid-liquid extraction method, QuEChERs
Method, but these methods exist using organic reagent amount is more, consuming time is long, complex steps, serious interference, clean-up effect are bad etc.
Disadvantage.
The method of the separating amide class herbicide provided by the invention from miscellaneous beans is on the basis of traditional QuEChERs method
The improvement of upper progress, improved method have following some advantages:
(1) it selects the mix reagent of nonpolar ethyl acetate and n-hexane as extractant, replaces acetonitrile, nonpolarity is molten
Agent effectively can carry out good extraction to stroma contaminants such as fat, protein；
(2) it selects MIL-101 (Zn) as adsorbent, replaces PSA (primary secondary amine), acetamide-group herbicides
In hetero atom and the interaction of (Zn) the unsatuated metal site MIL-101, in amide between phenyl ring and the ligand of MIL-101
The Zn- π of π-π reciprocation, the delocalizedπelectron in amide herbicide and the unsaturated site of metal interacts, these phase interactions
It is adsorbed with the promotion semipolar acetamide-group herbicides of apolar extraction agent are stronger by MIL-101 (Zn), and fat, protein etc.
Stroma contaminants will not be then adsorbed, and be stayed in apolar extraction agent, to realize the separation of acetamide-group herbicides and stroma contaminants；
(3) a small amount of sodium chloride replaces the mixture of salt, can condense in avoid salt in miscellaneous beans sample.
It can be seen that the method provided by the invention from separating amide class herbicide in miscellaneous beans can effectively will be in miscellaneous beans
Acetamide-group herbicides extraction, separate, by purification after can carry out subsequent detection, for it is subsequent it is effective detection establish
Basis.
Acetamide-group herbicides are to detect amide-type herbiciding in miscellaneous beans sample from the purpose separated in miscellaneous beans by we
The content of agent (propanil, metazachlor, pretilachlor, butachlor), so subsequent detection for convenience, we and the elution to collection
Liquid has carried out concentration, filtration treatment, specific as follows:
Eluent is transferred in chicken heart bottle, 35 DEG C of rotations be evaporated after with 100 μ L methanol back dissolvings, obtained solution is with 0.22
The filtering of μm teflon membrane filter, may wait for that sample introduction is analyzed later.
Seven, sample is analyzed
Sample is analyzed using high performance liquid chromatograph, chromatographic condition is as follows:
Chromatographic column: Eclipse XDB-C18 column (150mm × 4.6mm, 3.5 μm).
Column temperature: 30 DEG C.
Mobile phase A: water.
Mobile phase B: acetonitrile.
Linear gradient elution program: 0-5min, 40-60%B；5-9min, 60-80%B；9-13.5min 80-79.5%
B；13.5-14.7min 79.5%B；14.7-16.0min 79.5-79.4%B；16.0-21.0min 79.4-79.0%B；
21-25min, 79-60%B.
Flow rate of mobile phase: 0.50mL/min.
Sample volume: 20 μ L.
DAD Detection wavelength: 228nm.
Reference wavelength: 360nm.
Bandwidth: 4nm.
Eight, extraction conditions is studied
Effect of extracting directly affects the analysis result of target analytes.In order to obtain optimal extraction efficiency, in determination
It states before the method for separating amide class herbicide in miscellaneous beans, we have done detailed research to extraction conditions, comprising: extractant
Influence of the influence, ultrasonic extraction time of type and dosage to effect of extracting to effect of extracting, adsorbent amount are to effect of extracting
Influence.
1, the influence of extractant type and dosage to effect of extracting
We test investigated methanol (MA), n-hexane (HA), ethyl acetate (EA), chloroform (CF), ethyl acetate/just oneself
Alkane (VEA:VHA=1:1) five kinds of organic single extractants and mixed extractant effect of extracting.It is as shown in Figure 3 to investigate result.
We analyze result is investigated, specific as follows:
(1) methanol and chloroform due to polarity it is stronger, it is unfavorable so cannot effectively penetrate into high protein and fat sample
In the extraction of analyte.
(2) n-hexane is nonpolar, although can effectively penetrate into sample, centering polar amide class herbicide
Solubility is limited, causes the analyte rate of recovery lower.
(3) with ethyl acetate/n-hexane (VEA:VHA=1:1) be extractant when, can be improved propanil, metazachlor, pretilachlor,
The rate of recovery of these four acetamide-group herbicides of butachlor in miscellaneous beans, chromatogram is also cleaner clear, the baseline noise of chromatography
It is lower.
Therefore, we finally select ethyl acetate/n-hexane (VEA:VHA=1:1) as propanil, metazachlor, pretilachlor,
The extractant of these four acetamide-group herbicides of butachlor.
We, which also test, has investigated extractant ethyl acetate/n-hexane (VEA:VHA=1:1) in 0.4mL to 2.0mL range
The interior influence to propanil, metazachlor, pretilachlor, butachlor these four acetamide-group herbicides recovery of extraction.Investigate result are as follows: with
Extractant ethyl acetate/n-hexane (VEA:VHA=1:1) volume increase, the recovery of extraction of four kinds of acetamide-group herbicides exists
It is basically unchanged after initial increase, as extractant ethyl acetate/n-hexane (VEA:VHA=1:1) volume be 0.8mL when, extract back
Yield highest, in order to extract sufficiently, we finally select extractant ethyl acetate/n-hexane (VEA:VHA=1:1) dosage be
0.8mL。
2, influence of the ultrasonic extraction time to effect of extracting
Ultrasonic radiation can accelerate mass transfer and the diffusion of substance.The present invention selects ultrasound to carry out assisted extraction object.
We assess the ultrasonic extraction time to amide-type herbiciding by carrying out experiment of single factor in 0.5min to 4min
The influence of the agent rate of recovery.As a result as shown in Figure 4.
As shown in Figure 4, when the ultrasonic extraction time extending to 2min from 0.5min, the analyte rate of recovery is significantly improved, but
As the ultrasonic extraction time continues to extend, the analyte rate of recovery is basically unchanged.Therefore, we finally select the ultrasonic extraction time be
2min。
3, influence of the adsorbent amount to effect of extracting
We have studied the dosages of adsorbent MIL-101 (Zn) (40-72mg) to propanil, metazachlor, pretilachlor, butachlor
The influence of recovery of extraction of these four acetamide-group herbicides in miscellaneous beans.Result of study is as shown in Figure 5.
As shown in Figure 5, increase to 56mg, four kinds of acetamide-group herbicides from 40mg with adsorbent MIL-101 (Zn) dosage
The rate of recovery gradually increase, due to the limited sorption capacity of adsorbent MIL-101 (Zn), when the use of adsorbent MIL-101 (Zn)
When amount is less than 56mg, not exclusively, when the dosage of adsorbent MIL-101 (Zn) is 56mg, four kinds of acetamide-group herbicides exist for extraction
Rate of recovery highest in miscellaneous beans.To guarantee adequately to extract, we finally determine that the dosage of adsorbent MIL-101 (Zn) is
56mg。
Nine, elution requirement is studied
Before determining above-mentioned chromatographic condition, we study eluent type and dosage.
Target analytes propanil, metazachlor, pretilachlor, butachlor belong to middle polar compound in organic compound and strong
Good to the solubility of the target analytes and compatibility with chromatogram flow phase of polar compound, methanol and acetonitrile is good, therefore, we
It is that eluant, eluent is studied, and has studied 0.5-2.0mL eluent different amounts pair simultaneously with methanol (MA) and acetonitrile (ACN)
The influence of four kinds of acetamide-group herbicides rate of recovery.Result of study is as shown in Figure 6.
It will be appreciated from fig. 6 that the rate of recovery of propanil, metazachlor, pretilachlor, butachlor these four acetamide-group herbicides is with eluent
Increase of the volume between 0.5-1.5mL and increase, when eluent is greater than 1.5mL, the recycling of these four acetamide-group herbicides
Rate variation is little.
It is also known by Fig. 6, propanil, metazachlor, pretilachlor, the Methanol Recovery rate of butachlor these four acetamide-group herbicides are equal
Slightly above acetonitrile.This is because containing hydroxyl in methanol, hydroxyl can be with the unsaturation metal of MIL-101 (Zn) as proton acceptor
Site chelating, weakens the interaction of analyte and unsaturation metallic site, promotes the elution of analyte.Although containing in acetonitrile
Cyanogen, but cyanogen is proton donor, cannot interact with the unsaturation metallic site of MIL-101 (Zn), rely only on analysis
Solubility of the object to eluate.
In addition, methanol is cheaper, toxicity is smaller.Comprehensively considered and analyzed, we finally select the conduct of 1.5mL methanol and wash
De- liquid.
Ten, analysis measurement method
1, working curve, LODs and LOQs
According to the relationship of the target analyte concentration extracted in the chromatographic peak peak area of measurement and semen sojae atricolor powder, correction is established
Curve.
We are having studied the linear relationship between each parameter, equation of linear regression and phase relation under optimal pretreatment condition
Number is as shown in table 1.
Table 1 analyzes performance
As can be seen from Table 1, in the case where related coefficient > 0.9974, good linear relationship has been obtained.
After measured, in semen sojae atricolor four kinds of acetamide-group herbicides detection limit (LODs) 0.58-1.78 μ g/kg range, this
A little low LODs values demonstrate the overall feasibility of accurate, the reasonable measurement acetamide-group herbicides of detection method provided by the invention.
After measured, in semen sojae atricolor four kinds of acetamide-group herbicides quantitative limit (LOQs) 2.02-5.94 μ g/kg range,
Lower than maximum detectability (MRLs), this shows that detection method provided by the invention has preferable practicability.
2, precision and the rate of recovery
By measuring withinday precision, day to day precision, the precision of detection method provided by the invention is evaluated.
By carrying out 5 analyses to 1 sample within 1 working day, the withinday precision on the same day is obtained.
Continuous analysis daily is carried out within 5 working days to same sample, obtains day to day precision.
By calculating the average recovery rate (n=5) of each herbicide under three kinds of spiked levels, to detection provided by the invention
The repeatability of method is evaluated.
Withinday precision, day to day precision, average recovery rate, standard deviation calculated result be shown in Table 2.
The analysis result of 2 sample of table
As shown in Table 2, it by carrying out actual analysis to three semen sojae atricolor, red bean, kidney bean samples, is respectively removed under three kinds of spiked levels
The average recovery rate of careless agent is between 86.9-119.0%, and standard deviation is between 1.14-2.80%, wherein withinday precision
Acceptable standard deviation value range is 1.1-2.3%, and the acceptable standard deviation value range of day to day precision is 1.8-
2.8%.
The result shows that detection method provided by the invention has good repeatability and applicability.
3, sample analysis
By taking semen sojae atricolor powder sample as an example.
The chromatogram of hybrid standard working solution (10 μ g/mL), blank sample and mark-on sample is respectively such as Fig. 7 (a), Fig. 7 (b)
With shown in Fig. 7 (c).
By Fig. 7 (a), Fig. 7 (b) and Fig. 7 (c) it is found that in detection limit, the semen sojae atricolor come is bought back from the market of farm produce at us
In acetamide-group herbicides including propanil, metazachlor, pretilachlor and butachlor are not detected.
Compare between 11, method
Detection method provided by the invention is compared with reported LLSPE, SPE, QuEChERs, investigates the present invention
The advantage performance of the detection method of offer.Comparison result is shown in Table 3.
Comparison result between 3 method of table
【1】China Food and Drug Administration.National food safety standards
Determination ofacetanilide herbicide residues in cereals and oil seeds Gas
chromatography-mass spectrometry:GB 23200.1-2016[S/OL].Beijing:Standards
Press of China,2017:1-12(2017-02-09)[2017-04-02].http://www.doc88.com/p- 8436315147394.html.
【2】A.R.Mughari,P.P.Vázquez,M.M.Galera,Analysis of phenylurea and
propanil herbicides by solid-phase microextraction and liquid chromatography
combined with post-column photochemically induced fluorimetry derivatization
and fluorescence detection,Anal.Chim.Acta 593(2007)157-163.
【3】Y.M.Zhou,B.Chen,H.Zh.Tian,Determination ofresidues of fifteen
herbicides in potato by HPLC,Chin.J.of Agrochem.48(2009)749-754.
As shown in Table 3, compared with other are with reference to method, detection method provided by the invention is in selectivity, sensitivity, absorption
Agent and extractant dosage, sample preparation time and operating procedure etc. all have a clear superiority.
On the one hand, MIL-101 (Zn) has biggish aperture and hole window, so that the acetamide-group herbicides of small molecule are absorbed
In access aperture, hole diffusion is formed.Amide herbicide is adsorbed by MIL-101 (Zn), mainly due to the hetero atom in amide and not
In π-π reciprocation, amide herbicide in the interaction of saturation metallic site, amide between phenyl ring and the ligand of MIL-101
Delocalizedπelectron and the unsaturated site of metal Zn- π interaction.
On the other hand, these four herbicides of propanil, metazachlor, pretilachlor and butachlor are all polar compounds.There is document
The adsorption capacity of polar compound can be enhanced in unsatuated metal site in report metal-organic framework, and Huang etc. is reported
MIL-101 has stronger absorption property to containing heteroatomic volatile organic compounds, especially to aminated compounds, Fu etc.
People's report, in nonpolar solvent system, the phase in unsatuated metal site and the polar compound rich in electron group in MIL-101
Interaction is stronger.Therefore, unsatuated metal site (electron acceptor) and hetero atom (electron donor) are matched in acetamide-group herbicides
Position interaction may be the main drive of absorption.
In conclusion the method for the separating amide class herbicide provided by the invention from miscellaneous beans, can effectively extract, divide
From the acetamide-group herbicides in, miscellaneous beans of purification, extracting and purifying efficiency and enrichment times are improved, experimental cost is reduced, are reduced
The consumption of organic solvent, simplifies operating procedure, shortens pretreatment time.
The method of acetamide-group herbicides in the miscellaneous beans sample of detection provided by the invention, in effective extraction, separation, purification
On the basis of acetamide-group herbicides in miscellaneous beans, sample is analyzed using high performance liquid chromatograph, thus can quickly and
The content of acetamide-group herbicides, has very positive application value in effective detection impurity removal beans sample.
It should be noted that the above embodiments do not limit the invention in any form, it is all to use equivalent replacement or equivalent change
The mode changed technical solution obtained, falls within the scope of protection of the present invention.
Claims (6)
1. efficiently separating the method for acetamide-group herbicides from miscellaneous beans, which comprises the following steps:
Step1: miscellaneous beans are clayed into power, and cross 80 meshes；
Step2: miscellaneous bean powder end and water are added in centrifuge tube, adjust pH value of solution=7, oscillation；
Step3: continuing that extractant and NaCl, ultrasonic extraction 2min are added into centrifuge tube, then 0 DEG C with 15000rpm from
The heart, the extractant are that ethyl acetate mixes in equal volume with n-hexane, and the ratio of miscellaneous bean powder end and extractant is 1g:2mL；
Step4: supernatant being transferred in the centrifuge tube of another Zhi Hanyou adsorbent, be stirred, then with 10000rpm from
The heart, the adsorbent are MIL-101 (Zn), and the ratio of miscellaneous bean powder end and adsorbent is 100g:14g；
Step5: removing supernatant, methanol be added into centrifuge tube, carries out ultrasonic elution to analyte.
2. quickly detecting the method for acetamide-group herbicides in miscellaneous beans sample, which comprises the following steps:
Step1: miscellaneous beans sample is clayed into power, and crosses 80 meshes；
Step2: the miscellaneous bean powder end 0.4g and 0.4mL water are added in centrifuge tube, adjust pH value of solution=7, with forced oscillation, continue to
0.8mL extractant and 40mg NaCl, ultrasonic extraction 2min are added in centrifuge tube, 5min is then centrifuged with 15000rpm at 0 DEG C,
Supernatant is transferred in the centrifuge tube of another Zhi Hanyou adsorbent, is stirred, 3min is centrifuged with 10000rpm, removes supernatant
2ml methanol is added into centrifuge tube for liquid, carries out ultrasonic elution to analyte, wherein the extractant be ethyl acetate with just oneself
Alkane mixes in equal volume, and the adsorbent is MIL-101 (Zn)；
Step3: eluent is transferred in chicken heart bottle, is rotated after being evaporated with 100 μ L methanol back dissolvings, obtained solution filter membrane mistake
Filter, finally analyzes sample with high performance liquid chromatograph.
3. according to the method described in claim 2, it is characterized in that, adjusting solution using 0.5mol/L NaOH in Step2
pH。
4. according to the method described in claim 2, it is characterized in that, rotating the temperature being evaporated is 35 DEG C in Step3.
5. according to the method described in claim 2, it is characterized in that, what the filter membrane was selected is 0.22 μm poly- four in Step3
Vinyl fluoride filter membrane.
6. according to the method described in claim 2, it is characterized in that, in Step3, chromatographic condition are as follows:
Chromatographic column: Eclipse XDB-C18 column, 150mm × 4.6mm, 3.5 μm；
Column temperature: 30 DEG C；
Mobile phase A: water；
Mobile phase B: acetonitrile；
Linear gradient elution program: 0-5min, 40-60%B；5-9min, 60-80%B；9-13.5min 80-79.5%B；
13.5-14.7min 79.5%B；14.7-16.0min 79.5-79.4%B；16.0-21.0min 79.4-79.0%B；21-
25min, 79-60%B；
Flow rate of mobile phase: 0.50mL/min；
Sample volume: 20 μ L.
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| CN103308641A (en) * | 2013-06-08 | 2013-09-18 | 国家烟草质量监督检验中心 | High performance liquid chromatography-tandem mass spectrometry measuring method of three amide herbicides in tobacco and tobacco products |
| CN107907606A (en) * | 2017-10-27 | 2018-04-13 | 吉林省农业科学院 | The method of a variety of herbicides in a kind of while quick detection soil |
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| CN103308641A (en) * | 2013-06-08 | 2013-09-18 | 国家烟草质量监督检验中心 | High performance liquid chromatography-tandem mass spectrometry measuring method of three amide herbicides in tobacco and tobacco products |
| CN107907606A (en) * | 2017-10-27 | 2018-04-13 | 吉林省农业科学院 | The method of a variety of herbicides in a kind of while quick detection soil |
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