CN105911157A - Novel method for rapid detection of aflatoxin in food - Google Patents

Novel method for rapid detection of aflatoxin in food Download PDF

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CN105911157A
CN105911157A CN201610216595.3A CN201610216595A CN105911157A CN 105911157 A CN105911157 A CN 105911157A CN 201610216595 A CN201610216595 A CN 201610216595A CN 105911157 A CN105911157 A CN 105911157A
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aflatoxin
solution
organic acid
food
new method
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CN105911157B (en
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焦扬
杨亚玲
赵娇
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YUNNAN LUNYANG TECHNOLOGY Co.,Ltd.
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YUNNAN JIANNIU BIOLOGICAL TECHNOLOGY Co Ltd
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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
    • 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
    • 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
    • G01N30/14—Preparation by elimination of some components
    • 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
    • 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
    • G01N2030/062—Preparation extracting sample from raw material
    • 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
    • G01N30/14—Preparation by elimination of some components
    • G01N2030/143—Preparation by elimination of some components selective absorption

Abstract

The invention discloses a novel method for rapid detection of aflatoxin in food. The novel method studies extraction, purification and enrichment of aspertoxin in food and detects aflatoxin in food by using a combination of high performance liquid chromatography, on-line post-column photochemical derivatization and a fluorescence detector. The method employs a combination of dispersive magnetic solid-phase micro-extraction and dispersive liquid-liquid micro-extraction techniques for extraction, purification and enrichment of aspertoxin in a sample. Compared with a conventional determination method for aflatoxin, the method provided by the invention has the characteristics of a small amount of an organic solvent used in detection, short analysis time, simple operation, low detection cost, accuracy, etc.

Description

The new method of aflatoxin in a kind of quick detection food
Technical field
The present invention relates to the new method of aflatoxin in a kind of quick detection food.Belong to food Detection field.
Background technology
Aflatoxin (AFT) is the compound that a class chemical constitution is similar, is dihydro furan Mutter the derivant of coumarin.Aflatoxin (Aflatoxin, it is called for short AFT) it is 20th century 60 The fungus metabolite of a kind of severe toxicity found at the beginning of the age, mainly by Aspergillus flavus (Aspergillus Flaws) and aspergillus parasiticus bacterium (Aspergillus parasiticus) etc. infect agricultural product after produce, Being to be currently known one of the strongest carcinogen, toxicity is extremely strong.Within 1993, aflatoxin is by generation Boundary's health organization Agency for Research on Cancer delimited as one-level carcinogen.
Along with in the world with the frequent generation of China aflatoxin contamination event, various countries are at food The monitoring in field is more and more stricter.At present, commonly used detection method includes that enzyme linked immunological is inhaled Attached method (ELASA), immune affinity column-spectrofluorophotometer method (SFB), immune affinity column -high performance liquid chromatography (IAC-HPLC) and HPLC-MS/MS method etc..The highest Effect liquid phase chromatogram (HPLC) method measures accurately, and resolution is high, can measure multiple yellow bent simultaneously Mould toxic components, completes qualitative, quantitative determination and is widely used.Owing to food samples containing There is a large amount of compound, need that measuring samples is carried out purified treatment and contratoxin is enriched with, Just can carry out detection to analyze.
The sample that in the food that standard GB/T/T 18979-2003 uses, aflatoxin measures Purification method is the immune affinity column method of purification, uses the method GB to do experiment a collection of inspection product experiment Cost about needs 160 yuan, and the time-consuming about 1.5h of experiment pre-treatment completes a collection of inspection product.Patent of invention CN Although 104914196A has been also carried out improving, make scavenging material with HC-C18+PSA, but not Specify material therefor, and there is separation difficulty and the problem of clean-up effect difference.
Summary of the invention
It is an object of the invention to provide a kind of testing cost low, sample extraction good purification, The assay method of aflatoxin in the food that the detection time is short.
The new method of aflatoxin in a kind of quick detection food, comprises the following steps:
(1) preparation of aflatoxin extracting solution to be measured in food samples
1. for rice, Semen Maydis, Semen Tritici aestivi, Semen arachidis hypogaeae, Semen Juglandis, Semen Armeniacae Amarum and goods extracting method thereof For: accurately weigh the sample 2～5g after pulverizing homogenizing, be accurate to 0.0001g, add volume Mark is the methanol aqueous solution 20～50mL of 70～80%, and acutely vibrate 1min, 50 ± 2 DEG C of water Bath supersound extraction 20～30min, is centrifuged 5～10min with 3500～5000r/min, in taking-up Layer solution, repeats to extract 2～3 times, merges supernatant, stand-by；
2. for soy sauce, vinegar, Vegetable oil lipoprotein extracting method it is: accurately weigh 5.00～10.00g Sample, addition 2～5g NaCl and the methanol aqueous solution 20～50mL that volume fraction is 70～80%, Vortex mixed 1～3min, quantitative filter paper filters, and accurately draws 5.00mL filtrate, adds 10mL ultra-pure water dilutes, mixing, stand-by.
(2) extracting solution purifies
Addition 50～100 μ L ionic liquids in step (1) extracting solution, vortex mixed 1～2min, Form homogeneous phase solution, add the ultra-pure water of 2～5 times of volumes, add organic acid prepared by the present invention Coated magnetic nano material 2.0～10.0mg, vortex mixed 1～3min, place 10～20min, Separate under externally-applied magnetic field effect, collect magnetic polymer；Add 5～10mL ultra-pure waters, whirlpool Rotation mixing 1min, separates under externally-applied magnetic field effect, collects magnetic polymer；Add eluant, Vortex mixed 1～3min, separates under externally-applied magnetic field effect, repeats 2～3 times, merges eluting Liquid, eluent, through 0.45 μm membrane filtration, obtains testing sample solution；
(3) measure
Respectively take standard working solution and testing sample solution 50 μ L, at the liquid phase chromatogram condition set Lower sample introduction, uses high performance liquid chromatography-online Post-column photochemical derivatization-fluorescence detector detection, Obtain aflatoxin content in testing sample.
The preparation of the organic acid coated magnetic nano material described in step (1) comprises the following steps: Weigh appropriate divalent iron salt and trivalent iron salt, use 100mL deionized water dissolving, be configured to Fe2+ And Fe3+Mixed solution, under inert gas shielding, heating in water bath mechanical agitation 10～15min, Add acetone or the methanol solution of organic acid, after continuing stirring 5min, add precipitant dense Ammonia, simultaneously after stirring 30min, naturally cools to room temperature；Under externally-applied magnetic field effect, solid-liquid divides Respectively wash 2～3 times from, product deionized water/methanol remove unnecessary organic acid acetone or Methanol solution, i.e. obtains organic acid coated magnetic nanometer 60～80 DEG C of vacuum drying 24～48h Material.
In described food samples, aflatoxin to be measured includes: AFB1, B2, G1、G2。
In described organic acid coated magnetic nano material organic acid be castor oil acid, palmitoleic acid, One of oleic acid, pelargonic acid, n-capric acid are planted.
Described extraction and cleaning intermediate ion liquid include 1-butyl-3-methyl-imidazoles hexafluorophosphate, 1-amyl group-3-methyl-imidazoles hexafluorophosphate, 1-hexyl-3-methyl-imidazoles hexafluorophosphate, 1- Heptyl-3-methyl-imidazoles hexafluorophosphate, the one of 1-octyl group-3-methyl-imidazoles hexafluorophosphate.
In described extraction and cleaning, eluant is acetonitrile, and consumption is 0.5～1mL.
The described liquid phase chromatogram condition set as: C18 chromatographic column (4.6mm × 250mm, 5.0 μm), mobile phase methanol-water (45: 55), flow velocity 1.0mL/min, sample size 50 μ L, Column oven 30 DEG C.Detector is fluorescence detector, excitation wavelength 360nm, launches wavelength 440 nm；High performance liquid chromatograph (outfit fluorescence detector)；Photochemical derivatization device.
Fe described in the preparation of described organic acid coated magnetic nano material2+And Fe3+Mixing In solution, Fe2+With Fe3+Mol ratio be 1:1～5.
Described in the preparation of described organic acid coated magnetic nano material, noble gas is nitrogen, Mechanical agitation speed is 600～1000rpm, and water bath heating temperature is 70～85 DEG C.
The acetone of organic acid is contained described in the preparation of described organic acid coated magnetic nano material Or methanol is molten specifically, contain 100～400 μ g organic acid, precipitant in 5mL acetone or methanol The consumption of strong aqua ammonia is 10～20mL.
The present invention uses dispersion magnetic solid-phase microextraction to be combined with dispersive liquid-liquid microextraction technology, dispersion The magnetic Nano material of liquid-liquid micro-extraction intermediate ion liquid and preparation has extraction to aflatoxin And the dual function of absorption so that it is the clean-up effect that existing good effect of extracting has had again；Due to Preparing material is nanoscale, and material usage is few, only needs several milligrams, with low cost, only needs several units Money；The superparamagnetism that material has, can be with sharp separation at externally-applied magnetic field, the sample purification time Short, only need a few minutes.Technology used by these features all present invention is in aflatoxin measures There is big advantage.
It is an advantage of the current invention that:
1. the present invention uses dispersion magnetic solid-phase microextraction to be combined with dispersive liquid-liquid microextraction technology, Aflatoxin is had by the magnetic Nano material of dispersive liquid-liquid microextraction intermediate ion liquid and preparation Extraction and the dual function of absorption so that it is the clean-up effect that existing good effect of extracting has had again.
2. the organic carboxyl acid modified magnetic nano material that prepared by the present invention has spy to aflatoxin Opposite sex absorption, simultaneously because specific surface is big, consumption is low in use, simultaneously during eluting, first uses Wash polar composition, then with acetonitrile eluting aflatoxin, clean-up effect is obvious.
3. the superparamagnetism of material is prepared in utilization, collects the extraction of extraction object by external magnetic field Agent, lock out operation is simple, quick, and consumption of organic solvent is minimum.
4. combine high performance liquid chromatography-online Post-column photochemical derivatization-fluorescence detector detection food Aflatoxin in product, detection limit is low, highly sensitive, and recovery of standard addition is high.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is further described, but protection scope of the present invention It is not limited to this.
Embodiment 1 utilizes the present invention to detect the content of Aflatoxin in Peanut byHigh, and step is:
1, the preparation of organic acid coated magnetic nano material: weigh 4.10g Ferrous ammonium sulfate ((NH4)2Fe(SO4)2·6H2O), 13.52g iron chloride (FeCl3·6H2O), 100mL is used After deionized water dissolving, forwarding in 250mL three-necked bottle, under nitrogen protection, water-bath adds Hot to 80 DEG C, mechanical agitation 10min (rotating speed is 1000rpm), it is subsequently adding 5mL containing 200 μ L The methanol solution of pelargonic acid, continues stirring 5min, adds 10mL strong aqua ammonia, and mixed liquor continues After continuous reaction 30min, after naturally cooling to room temperature, collect product with externally-applied magnetic field and discard reaction Liquid, magnetic material 100mL deionized water wash 2 times, 100mL methanol washs 2 times, 100mL deionized water wash 3 times, i.e. obtains pelargonic acid cladding magnetic at 60 DEG C of vacuum drying 48h Property nano material.
2, take appropriate aflatoxin reference substance storing solution, obtain concentration with 70% methanol dilution and divide It is not the reference substance solution of 0.02,0.05,0.1,0.3,0.5,1.0 μ g/mL, mistake The organic filter membrane of 0.45m, C18 chromatographic column (4.6mm × 250mm, 5.0 μm), flow phase Methanol-water (45: 55；V/v), flow velocity 1.0mL/min, sample size 50 μ L, column oven 30 DEG C. Detector is fluorescence detector, excitation wavelength 360nm, launches wavelength 440nm；Agilent 1200 high performance liquid chromatographs (quaternary pump, automatic sampler are equipped with fluorescence detector)；Light Chemically derived device.With concentration as abscissa, with peak area as vertical coordinate, draw standard curve, Calculating regression equation, correlation coefficient, relative standard deviation, the range of linearity etc. are shown in Table 1, sample Middle recovery of standard addition is shown in Table 2.
The linear equation of table 1 aflatoxin, correlation coefficient, relative standard deviation, the range of linearity
The recovery of standard addition of aflatoxin in table 2 blank sample
3, sample treatment: accurately weigh the peanut sample 2g after pulverizing homogenizing, be accurate to 0.0001g, adds the methanol aqueous solution 20mL that volume fraction is 70%, and acutely vibrate 1min, 50 ± 2 DEG C of water bath sonicator extract 20min, are centrifuged 10min with 3500r/min, take out upper strata molten Liquid, repeats to extract 2 times, merges supernatant；It is added thereto to the 1-octyl group-3-methyl of 50 μ L -limidazolium hexafluorophosphate, vortex mixed 1min, form homogeneous phase solution, add 70mL ultrapure Water, pelargonic acid coated magnetic nano material 2.0mg of addition step 1 preparation, vortex mixed 1min, Place 10min, separate under externally-applied magnetic field effect, collect magnetic polymer；Add 5mL to surpass Pure water, vortex mixed 1min, separate under externally-applied magnetic field effect, collect magnetic polymer；Add Enter acetonitrile 0.3mL, vortex mixed 1min, separate under externally-applied magnetic field effect, be repeated 2 times, Merge acetonitrile, through 0.45 μm membrane filtration, obtain testing sample solution.
4, measure: by the testing sample solution handled well at chromatographic condition and the identical bar of step 2 Sample detection under part, draws the content of aflatoxin, wherein aflatoxin B 2, G1, G2 does not detects, and AFB1 is 5.49 μ g/kg.
Embodiment 2 utilizes the present invention to detect the content of aflatoxin in Semen Maydis, and step is:
1, the preparation of organic acid coated magnetic nano material: weigh 2.00g ferrous chloride (FeCl2·4H2O), 5.40g iron chloride (FeCl3·6H2O), water-soluble with 100mL deionization Xie Hou, forwards in 250mL three-necked bottle, and under nitrogen protection, mechanical agitation 15min (turns Speed is 600rpm) and heating in water bath to 70 DEG C, be subsequently adding 5mL containing 300 μ L n-capric acids Methanol solution, continues stirring 5min, adds 15mL strong aqua ammonia, and mixed liquor continues reaction 30min After, after naturally cooling to room temperature, collect product with externally-applied magnetic field and discard reactant liquor, magnetic material With 100mL deionized water wash 2 times, 100mL methanol washs 2 times, 100mL go from Sub-water washs 2 times, i.e. obtains n-capric acid coated magnetic nano material at 80 DEG C of vacuum drying 24h.
2, appropriate aflatoxin reference substance storing solution is taken same as in Example 1.
3, sample treatment: accurately weigh the corn sample 3g after pulverizing homogenizing, be accurate to 0.0001g, adds the methanol aqueous solution 30mL that volume fraction is 75%, and acutely vibrate 1min, 50 ± 2 DEG C of water bath sonicator extract 25min, are centrifuged 8min with 4000r/min, take out upper strata molten Liquid, repeats to extract 3 times, merges supernatant；It is added thereto to the 1-heptyl-3-first of 100 μ L Base-limidazolium hexafluorophosphate, vortex mixed 2min, form homogeneous phase solution, add 120mL and surpass Pure water, adds n-capric acid coated magnetic nano material 5.0mg of step 1 preparation, vortex mixed 2min, places 15min, separates under externally-applied magnetic field effect, collects magnetic polymer；Add 8mL Ultra-pure water, vortex mixed 2min, separate under externally-applied magnetic field effect, collect magnetic polymer； Add acetonitrile 0.4mL, vortex mixed 1min, separate under externally-applied magnetic field effect, be repeated 2 times, Merge acetonitrile, through 0.45 μm membrane filtration, obtain testing sample solution.
4, measure: by the testing sample solution handled well at chromatographic condition and the identical bar of step 2 Sample detection under part, draws the content of aflatoxin, and wherein aflatoxin B 2, G2 are not Detection, AFB1 is 30.1 μ g/kg, aflatoxin G 1 is 21.0 μ g/kg.
Embodiment 3 utilizes the present invention to detect the content of aflatoxin in Semen Juglandis, and step is:
1, the preparation of organic acid coated magnetic nano material: weigh 2.00g ferrous chloride (FeCl2·4H2O), 5.62g iron sulfate (Fe2(SO4)39H2O), 100mL deionization is used After water dissolution, forward in 250mL three-necked bottle, under nitrogen protection, mechanical agitation 12min (rotating speed is 800rpm) heating in water bath, to 85 DEG C, are subsequently adding 5mL containing 400 μ L oleic acid Acetone soln, continue stirring 5min, add 20mL strong aqua ammonia, mixed liquor continue reaction After 30min, after naturally cooling to room temperature, collect product with externally-applied magnetic field and discard reactant liquor, magnetic Property material 100mL deionized water wash 2 times, 100mL methanol washs 3 times, 100mL Deionized water wash 2 times, i.e. obtains Coated with Oleic Acid magnetic Nano material at 70 DEG C of vacuum drying 30h Material.
2, appropriate aflatoxin reference substance storing solution is taken and with embodiment 1.
3, sample treatment: accurately weigh the Semen Juglandis sample 5g after pulverizing homogenizing, be accurate to 0.0001g, adds the methanol aqueous solution 50mL that volume fraction is 80%, and acutely vibrate 1min, 50 ± 2 DEG C of water bath sonicator extract 25min, are centrifuged 5min with 5000r/min, take out upper strata molten Liquid, repeats to extract 2 times, merges supernatant；It is added thereto to the 1-hexyl-3-methyl of 80 μ L -limidazolium hexafluorophosphate, vortex mixed 2min, form homogeneous phase solution, add 250mL ultrapure Water, the Coated with Oleic Acid magnetic Nano material 10.0mg of addition step 1 preparation, vortex mixed 3min, Place 20min, separate under externally-applied magnetic field effect, collect magnetic polymer；Add 10mL to surpass Pure water, vortex mixed 3min, separate under externally-applied magnetic field effect, collect magnetic polymer；Add Enter acetonitrile 0.5mL, vortex mixed 1min, separate under externally-applied magnetic field effect, be repeated 2 times, Merge acetonitrile, through 0.45 μm membrane filtration, obtain testing sample solution.
4, measure: by the testing sample solution handled well at chromatographic condition and the identical bar of step 2 Sample detection under part, draws the content of aflatoxin, AFB1 be 1.24 μ g/kg, Aflatoxin B 2 is 0.61 μ g/kg, aflatoxin G 1 is 1.15 μ g/kg, Aspergillus flavus poison Element G2 is 0.51 μ g/kg.
Embodiment 4 utilizes the present invention to detect the content of aflatoxin in soy sauce, and step is:
1, the preparation of organic acid coated magnetic nano material: weigh 4.10g Ferrous ammonium sulfate ((NH4) 2Fe (SO4) 2 6H2O), 11.24g iron sulfate (Fe2(SO4)39H2O), use After 100mL deionized water dissolving, forward in 250mL three-necked bottle, under nitrogen protection, Mechanical agitation 10min (rotating speed is 1000rpm) heating in water bath, to 80 DEG C, are subsequently adding The 5mL acetone soln containing 300 μ L castor oil acids, continues stirring 5min, adds 15mL dense Ammonia, after mixed liquor continues reaction 30min, after naturally cooling to room temperature, receives with externally-applied magnetic field Collection product discards reactant liquor, magnetic material 100mL deionized water wash 2 times, 100mL Methanol washs 3 times, 100mL deionized water wash 2 times, at 80 DEG C of vacuum drying 24h i.e. Obtain castor oil acid coated magnetic nano material.
2, appropriate aflatoxin reference substance storing solution is taken and with embodiment 1.
3, sample treatment: accurately weigh soy sample 5g, is accurate to 0.0001g, adds 2g NaCl and the methanol aqueous solution 50mL that volume fraction is 70%, vortex mixed 1min, quantitatively Filter paper filtering, accurately draws 5.00mL filtrate, adds the dilution of 10mL ultra-pure water, mixing； It is added thereto to the 1-amyl group-3-methyl-imidazoles hexafluorophosphate of 50 μ L, vortex mixed 1min, Form homogeneous phase solution, add 150mL ultra-pure water, add the castor oil acid bag of step 1 preparation Cover magnetic Nano material 4.0mg, vortex mixed 1min, place 10min, externally-applied magnetic field effect Lower separation, collects magnetic polymer；Addition 10mL ultra-pure water, vortex mixed 3min, outward Add and separate under the action of a magnetic field, collect magnetic polymer；Addition acetonitrile 0.5mL, vortex mixed 1min, Separate under externally-applied magnetic field effect, be repeated 2 times, merge acetonitrile, through 0.45 μm membrane filtration, Obtain testing sample solution.
4, measure: by the testing sample solution handled well at chromatographic condition and the identical bar of step 2 Sample detection under part, draws the content of aflatoxin, wherein aflatoxin G 1, G2 Not detecting, AFB1 is 5.30 μ g/kg, aflatoxin B 2 is 0.86 μ g/kg.
Embodiment 5 utilizes the present invention to detect the content of aflatoxin in Oleum Arachidis hypogaeae semen, and step is:
1, the preparation of organic acid coated magnetic nano material: weigh 4.10g Ferrous ammonium sulfate ((NH4) 2Fe (SO4) 2 6H2O), 8.10g iron chloride (FeCl3·6H2O), 100mL is used After deionized water dissolving, forwarding in 250mL three-necked bottle, under nitrogen protection, machinery stirs Mix 12min (rotating speed is 800rpm) heating in water bath to 75 DEG C, be subsequently adding 5mL containing 500 μ L The acetone soln of palmitoleic acid, continues stirring 5min, adds 20mL strong aqua ammonia, mixed liquor After continuing reaction 30min, after naturally cooling to room temperature, collect product with externally-applied magnetic field and discard instead Answer liquid, magnetic material 100mL deionized water wash 2 times, 100mL methanol washs 3 times, 100mL deionized water wash 2 times, i.e. obtains palmitoleic acid cladding at 80 DEG C of vacuum drying 24h Magnetic Nano material.
2, appropriate aflatoxin reference substance storing solution is taken and with embodiment 1.
3, sample treatment: accurately weigh Oleum Arachidis hypogaeae semen sample 10g, be accurate to 0.0001g, adds 5g NaCl and the methanol aqueous solution 20mL that volume fraction is 80%, vortex mixed 1min, fixed Amount filter paper filtering, accurately draws 5.00mL filtrate, adds the dilution of 10mL ultra-pure water, mixed Even；It is added thereto to the 1-butyl-3-methyl-imidazoles hexafluorophosphate of 70 μ L, vortex mixed 1min, forms homogeneous phase solution, adds 100mL ultra-pure water, adds the Petiolus Trachycarpi of step 1 preparation Coated with Oleic Acid magnetic Nano material 7.0mg, vortex mixed 1min, place 10min, additional magnetic Separate under field action, collect magnetic polymer；Addition 10mL ultra-pure water, vortex mixed 3min, Separate under externally-applied magnetic field effect, collect magnetic polymer；Add acetonitrile 0.5mL, vortex mixed 1min, separates under externally-applied magnetic field effect, is repeated 2 times, and merges acetonitrile, filters through 0.45 μm Membrane filtration, obtains testing sample solution.
4, measure: by the testing sample solution handled well at chromatographic condition and the identical bar of step 2 Sample detection under part, draws the content of aflatoxin, AFB1 be 3.18 μ g/kg, Aflatoxin B 2 is 0.82 μ g/kg, aflatoxin G 1 is 1.60 μ g/kg, Aspergillus flavus poison Element G2 is 0.61 μ g/kg.
Above example measures with aflatoxin in GB/T 18979-2003 food and compares, knot Fruit is shown in Table 3.
Table 3 measurement result compares (μ g/kg)
"-" represents and does not detects
As seen from the results in Table 3: with micro-extraction purification method of the present invention combine high performance liquid chromatography- In online Post-column photochemical derivatization-fluorescence detector detection food aflatoxin with GB/T18979-2003 assay method is compared, and result is consistent, but few because processing step, used Time is short, and processing cost is low, consumption of organic solvent the most more advantage.

Claims (10)

1. the new method of aflatoxin in a quick detection food, it is characterised in that the method Comprise the following steps:
(1) preparation of aflatoxin extracting solution to be measured in food samples
1. to be measured for rice, Semen Maydis, Semen Tritici aestivi, Semen arachidis hypogaeae, Semen Juglandis, Semen Armeniacae Amarum and goods thereof Sample carries out extracting method: accurately weigh the to be measured all product 2～5g after pulverizing homogenizing, accurately To 0.0001g, add the methanol aqueous solution 20～50mL that volume fraction is 70～80%, acutely Vibration 1min, 50 ± 2 DEG C of water bath sonicator extract 20～30min, with 3500～5000r/min from The heart 5～10min, takes out upper solution, repeats to extract 2～3 times, merges supernatant, stand-by； Or
2. soy sauce, vinegar, Vegetable oil lipoprotein are treated that test sample extracting method is: accurately weigh 5.00～10.00g all product to be measured, add 2～5g NaCl and volume fraction is 70～80% Methanol aqueous solution 20～50mL, vortex mixed 1～3min, quantitative filter paper filters, accurately inhales Take 5.00mL filtrate, add the dilution of 10mL ultra-pure water, mix, stand-by.
(2) extracting solution purifies
In step (1) extracting solution add 50～100 μ L ionic liquids, vortex mixed 1～ 2min, forms homogeneous phase solution, adds the ultra-pure water of 2～5 times of volumes, adds prepared by the present invention Organic acid coated magnetic nano material 2.0～10.0mg, vortex mixed 1～3min, place 10～ 20min, separates under externally-applied magnetic field effect, collects magnetic polymer；Add 5～10mL ultrapure Water, vortex mixed 1min, separate under externally-applied magnetic field effect, collect magnetic polymer；Add Eluant, vortex mixed 1～3min, separate under externally-applied magnetic field effect, repeat 2～3 times, Merging eluent, eluent, through 0.45 μm membrane filtration, obtains testing sample solution；
(3) measure
Respectively take standard working solution and testing sample solution 20 μ L, in the liquid chromatograph set Under the conditions of sample introduction, use high performance liquid chromatography-online Post-column photochemical derivatization-fluorescence detector inspection Survey, obtain aflatoxin content in testing sample.
In a kind of quick detection food the most according to claim 1, aflatoxin is new Method, it is characterised in that the preparation bag of step (1) described organic acid coated magnetic nano material Include following steps: weigh appropriate divalent iron salt and trivalent iron salt, water-soluble with 100mL deionization Solve, be configured to Fe2+And Fe3+Mixed solution, under inert gas shielding, heating in water bath machine Tool stirring 10～15min, adds precipitant strong aqua ammonia, is simultaneously introduced acetone or the first of organic acid Alcoholic solution, after stirring 30min, naturally cools to room temperature；Under additional the action of a magnetic field, solid-liquid divides From, product deionized water/methanol washs the acetone or first removing unnecessary organic acid 3～5 times Alcoholic solution, i.e. obtains organic acid coated magnetic nanometer material 60～80 DEG C of vacuum drying 24～48h Material.
In a kind of quick detection food the most according to claim 1, aflatoxin is new Method, it is characterised in that in described food samples, aflatoxin to be measured includes: aflatoxin B1、B2、G1、G2。
In a kind of quick detection food the most according to claim 1, aflatoxin is new Method, it is characterised in that in described organic acid coated magnetic nano material organic acid be castor oil acid, One of palmitoleic acid, oleic acid, pelargonic acid, n-capric acid are planted.
In a kind of quick detection food the most according to claim 1, aflatoxin is new Method, it is characterised in that described extraction and cleaning intermediate ion liquid includes 1-butyl-3-methyl-imidazoles Hexafluorophosphate, 1-amyl group-3-methyl-imidazoles hexafluorophosphate, 1-hexyl-3-methyl-imidazoles six Fluorophosphate, 1-heptyl-3-methyl-imidazoles hexafluorophosphate, 1-octyl group-3-methyl-imidazoles hexafluoro The one of phosphate.
In a kind of quick detection food the most according to claim 1, aflatoxin is new Method, it is characterised in that in described extraction and cleaning, eluant is acetonitrile, consumption is 0.5～1mL.
In a kind of quick detection food the most according to claim 2, aflatoxin is new Method, it is characterised in that the described liquid phase chromatogram condition set is as C18 chromatographic column (4.6 Mm × 250mm, 5.0 μm), mobile phase methanol-water (45: 55), flow velocity 1.0mL/min, Sample size 50 μ L, column oven 30 DEG C.Detector is fluorescence detector, excitation wavelength 360nm, Launch wavelength 440nm；High performance liquid chromatograph (outfit fluorescence detector)；Photochemical derivatization Device.
In a kind of quick detection food the most according to claim 2, aflatoxin is new Method, it is characterised in that Fe described in the preparation of described organic acid coated magnetic nano material2+With Fe3+Mixed solution in, Fe2+With Fe3+Mol ratio be 1:1～5.
In a kind of quick detection food the most according to claim 2, aflatoxin is new Method, it is characterised in that inertia described in the preparation of described organic acid coated magnetic nano material Gas is nitrogen, and mechanical agitation speed is 600～1000rpm, water bath heating temperature be 70～ 85℃。
In a kind of quick detection food the most according to claim 2, aflatoxin is new Method, it is characterised in that contain described in the preparation of described organic acid coated magnetic nano material The acetone of machine acid or methanol are molten specifically, organic containing 100～400 μ g in 5mL acetone or methanol Acid, the consumption of precipitant strong aqua ammonia is 5～10mL.
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