CN106556585A - A kind of ochratoxin A detection method of content - Google Patents
A kind of ochratoxin A detection method of content Download PDFInfo
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- CN106556585A CN106556585A CN201611103356.3A CN201611103356A CN106556585A CN 106556585 A CN106556585 A CN 106556585A CN 201611103356 A CN201611103356 A CN 201611103356A CN 106556585 A CN106556585 A CN 106556585A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/38—Diluting, dispersing or mixing samples
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- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
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Abstract
The invention discloses a kind of ochratoxin A detection method of content, comprises the following steps：Sample pre-treatments：Take vegetables and fruits sample to pulverize, add extractant, mix, using ultrasound wave 30～60min of Rapid Extraction, extraction temperature is 30～40 DEG C, and centrifugation takes supernatant, supernatant is crossed into ochratoxin A affine in immunity column purification, mycotoxin cleaning buffer solution, water wash immune affinity column are used successively, then with methanol-eluted fractions, collect eluent, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant is the mixed liquor of acetonitrile and tetrahydrofuran.The present invention can be used for the ochratoxin A of trace group in complicated substrate and quantitatively quickly determine, and as a result more accurately, is more beneficial for the development of existing market and promotes.
Description
Technical field
The present invention relates to a kind of detection method, specifically a kind of ochratoxin A detection method of content.
Background technology
The ochratoxin toxic metabolic products that to be aspergillus similar with one group of structure of the toxigenic bacterium strain parameter such as Penicillium,
It is widely present in various food, feedstuff and agricultural product.Ochratoxin includes 7 kinds of compounds for having similar chemical constitution, its
Middle ochratoxin A is distributed in nature most extensively, and toxicity is most strong, affects maximum to the mankind and animals and plants.Toxicologic study
Show, ochratoxin A has various poison such as Toxicity of Kidney, hepatotoxicity, immunotoxicity and teratogenesis and mutagenic action
Property, there is very big potential hazard to animal and health.Therefore the detection to ochratoxin A and control are paid attention in countries in the world
System, has formulated related limit standard, in order to guaranteeing food safety and eliminating the technology barriers in international trade.Conventional at present
Determination Methods for Ochratoxin A has thin layer chromatography, gas chromatography, capillary electrophoresis technique（CE）, enzyme linked immunosorbent assay
(ELISA) etc..These detections of conventional detection method to ochratoxin A play very important effect, but these sides
More or less there are some defects such as in method：Sample pre-treatments are complicated, strong, low sensitivity, poor reproducibility poor for applicability the problems such as,
And the development with science and technology and the raising of detection level, the requirement to the ochratoxin A limit standard in food
Also improve constantly, traditional detection method can not meet present detection demand.Hence set up a kind of quick, high pass
The good detection method of amount, high sensitivity, low cost, specificity, so as to effectively be supervised to mycotoxin present in agricultural product
Control is very necessary.
The content of the invention
It is an object of the invention to provide a kind of ochratoxin A detection method of content, to solve in above-mentioned background technology
The problem of proposition.
For achieving the above object, the present invention provides following technical scheme：
A kind of ochratoxin A detection method of content, comprises the following steps：Sample pre-treatments：Take vegetables and fruits sample to pulverize, add extraction
Agent is taken, is mixed, using ultrasound wave 30～60min of Rapid Extraction, extraction temperature is 30～40 DEG C, and centrifugation takes supernatant, by supernatant mistake
Ochratoxin A affine in immunity column purification, successively with mycotoxin cleaning buffer solution, water wash immune affinity column, then uses first
Alcohol eluting, collects eluent, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant is acetonitrile and tetrahydrochysene
The mixed liquor of furan；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up the straightening die of quantitative analyses
Type；Institute's positive model for school building is judged with testing sample solution, and verify the reliability of calibration model；With calibration model to vegetable
In fruit actual sample, the content analysis of ochratoxin A are determined.
As further scheme of the invention：Sample pre-treatments are additionally included in the range of linearity prepares ochratoxin A
Correction sample, checking sample, take extract, and its three-dimensional excitation-emission are gathered under the scanning wavelength and sweep spacing for having optimized
Fluorescence data；Mathematics is carried out using parallel factor method (PARAFAC) to resulting three-dimensional data battle array and separates parsing, and set up
Calibration model.
As further scheme of the invention：The consumption of the extractant is 0.4～0.6g/ to make vegetables and fruits sample concentration
ml。
As further scheme of the invention：The extractant be acetonitrile and tetrahydrofuran by volume（0.8～1.2）：
（0.8～1.2）The mixed liquor of mixing.
As further scheme of the invention：The extractant is acetonitrile and tetrahydrofuran by volume 1：1 mixing it is mixed
Close liquid.
As further scheme of the invention：The centrifugation is centrifuged 8～12min for 1800～2200r/min.
As further scheme of the invention：Contain 25g/L Sodium Chloride, 5g/L carbon in the mycotoxin cleaning buffer solution
Sour hydrogen sodium, 0.01%v/v polysorbas20s, balance of water.
As further scheme of the invention：The flow velocity during drip washing is 1 drop/s.
As further scheme of the invention：The concrete operations of the filtration are：By 0.22 μm of organic facies of eluent Jing
Membrane filtration.
Compared with prior art, the invention has the beneficial effects as follows：The present invention can be used for the reddish brown song of trace group in complicated substrate
Syphilis element A is quantitatively quickly determined, and as a result more accurately, is more beneficial for the development of existing market and is promoted.
Specific embodiment
Below the technical scheme in the embodiment of the present invention is clearly and completely described.
In the embodiment of the present invention, a kind of ochratoxin A detection method of content is comprised the following steps：Sample pre-treatments：Take
Vegetables and fruits sample pulverizes, and adds extractant, mixes, and using ultrasound wave 30～60min of Rapid Extraction, extraction temperature is 30～40 DEG C,
Centrifugation, takes supernatant, supernatant is crossed ochratoxin A affine in immunity column purification, successively with mycotoxin cleaning buffer solution, water wash
Immune affinity column, then with methanol-eluted fractions, collects eluent, eluent is filtered, filtrate is taken and is obtained testing sample solution；It is described
Extractant is the mixed liquor of acetonitrile and tetrahydrofuran；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up
The calibration model of quantitative analyses；Institute's positive model for school building is judged with testing sample solution, and verify the reliability of calibration model
Property；The content analysis of ochratoxin A in vegetables and fruits actual sample are determined with calibration model.Sample pre-treatments are additionally included in linearly
The correction sample of latitude of formulation ochratoxin A, checking sample, take extract, and in the scanning wavelength and scanning room for having optimized
Every lower its three-dimensional fluorescence excitation-emission matrix spectrum data of collection；Parallel factor method (PARAFAC) is adopted to resulting three-dimensional data battle array
Carry out mathematics and separate parsing, and set up calibration model.The consumption of the extractant is 0.4～0.6g/ to make vegetables and fruits sample concentration
ml.The extractant be acetonitrile and tetrahydrofuran by volume（0.8～1.2）：（0.8～1.2）The mixed liquor of mixing.The extraction
Agent is taken for acetonitrile and tetrahydrofuran by volume 1：The mixed liquor of 1 mixing.It is described centrifugation for 1800～2200r/min centrifugation 8～
12min.Contain 25g/L Sodium Chloride, 5g/L sodium bicarbonate, 0.01%v/v polysorbas20s in the mycotoxin cleaning buffer solution, it is remaining
Measure as water.The flow velocity during drip washing is 1 drop/s.The concrete operations of the filtration are：By 0.22 μm of organic faciess filter of eluent Jing
Membrane filtration.
Embodiment 1：
Ochratoxin A detection method of content of the present invention, comprises the following steps：Sample pre-treatments：Take vegetables and fruits sample to pulverize, add
Extractant, mixes, and using ultrasound wave Rapid Extraction 30min, extraction temperature is 30 DEG C, and centrifugation takes supernatant, supernatant is crossed Aspergillus ochraceus
Toxin A affine in immunity column purifications, successively with mycotoxin cleaning buffer solution, water wash immune affinity column, then with methanol-eluted fractions,
Eluent is collected, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant is the mixed of acetonitrile and tetrahydrofuran
Close liquid；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up the calibration model of quantitative analyses；With to be measured
Sample solution is judged to institute's positive model for school building, and verifies the reliability of calibration model；With calibration model to the actual sample of vegetables and fruits
In product, the content analysis of ochratoxin A are determined.Sample pre-treatments are additionally included in the range of linearity school for preparing ochratoxin A
Positive sample, checking sample, take extract, and it is glimmering that its three-dimensional excitation-emission is gathered under the scanning wavelength and sweep spacing for having optimized
Light spectroscopic data；Mathematics is carried out using parallel factor method (PARAFAC) to resulting three-dimensional data battle array and separates parsing, and set up school
Positive model.The consumption of the extractant is 0.4g/ml to make vegetables and fruits sample concentration.The extractant is that acetonitrile and tetrahydrofuran are pressed
Volume ratio 0.8：The mixed liquor of 0.8 mixing.The extractant is acetonitrile and tetrahydrofuran by volume 1：The mixed liquor of 1 mixing.
The centrifugation is centrifuged 8min for 1800r/min.Contain 25g/L Sodium Chloride, 5g/L carbonic acid in the mycotoxin cleaning buffer solution
Hydrogen sodium, 0.01%v/v polysorbas20s, balance of water.The flow velocity during drip washing is 1 drop/s.The concrete operations of the filtration are：Will
0.22 μm of organic phase filter membrane of eluent Jing is filtered.
Embodiment 2：
Ochratoxin A detection method of content of the present invention, comprises the following steps：Sample pre-treatments：Take vegetables and fruits sample to pulverize, add
Extractant, mixes, and using ultrasound wave Rapid Extraction 60min, extraction temperature is 40 DEG C, and centrifugation takes supernatant, supernatant is crossed Aspergillus ochraceus
Toxin A affine in immunity column purifications, successively with mycotoxin cleaning buffer solution, water wash immune affinity column, then with methanol-eluted fractions,
Eluent is collected, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant is the mixed of acetonitrile and tetrahydrofuran
Close liquid；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up the calibration model of quantitative analyses；With to be measured
Sample solution is judged to institute's positive model for school building, and verifies the reliability of calibration model；With calibration model to the actual sample of vegetables and fruits
In product, the content analysis of ochratoxin A are determined.Sample pre-treatments are additionally included in the range of linearity school for preparing ochratoxin A
Positive sample, checking sample, take extract, and it is glimmering that its three-dimensional excitation-emission is gathered under the scanning wavelength and sweep spacing for having optimized
Light spectroscopic data；Mathematics is carried out using parallel factor method (PARAFAC) to resulting three-dimensional data battle array and separates parsing, and set up school
Positive model.The consumption of the extractant is 0.4～0.6g/ml to make vegetables and fruits sample concentration.The extractant is acetonitrile and tetrahydrochysene
Furan by volume 1.2：The mixed liquor of 1.2 mixing.The extractant is acetonitrile and tetrahydrofuran by volume 1：1 mixing it is mixed
Close liquid.The centrifugation is centrifuged 12min for 2200r/min.Contain 25g/L Sodium Chloride, 5g/ in the mycotoxin cleaning buffer solution
L sodium bicarbonate, 0.01%v/v polysorbas20s, balance of water.The flow velocity during drip washing is 1 drop/s.The concrete operations of the filtration
For：0.22 μm of organic phase filter membrane of eluent Jing is filtered.
Embodiment 3：
Ochratoxin A detection method of content of the present invention, comprises the following steps：Sample pre-treatments：Take vegetables and fruits sample to pulverize, add
Extractant, mixes, and using ultrasound wave Rapid Extraction 40min, extraction temperature is 35 DEG C, and centrifugation takes supernatant, supernatant is crossed Aspergillus ochraceus
Toxin A affine in immunity column purifications, successively with mycotoxin cleaning buffer solution, water wash immune affinity column, then with methanol-eluted fractions,
Eluent is collected, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant is the mixed of acetonitrile and tetrahydrofuran
Close liquid；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up the calibration model of quantitative analyses；With to be measured
Sample solution is judged to institute's positive model for school building, and verifies the reliability of calibration model；With calibration model to the actual sample of vegetables and fruits
In product, the content analysis of ochratoxin A are determined.Sample pre-treatments are additionally included in the range of linearity school for preparing ochratoxin A
Positive sample, checking sample, take extract, and it is glimmering that its three-dimensional excitation-emission is gathered under the scanning wavelength and sweep spacing for having optimized
Light spectroscopic data；Mathematics is carried out using parallel factor method (PARAFAC) to resulting three-dimensional data battle array and separates parsing, and set up school
Positive model.The consumption of the extractant is 0.5g/ml to make vegetables and fruits sample concentration.The extractant is that acetonitrile and tetrahydrofuran are pressed
Volume ratio 1：The mixed liquor of 1 mixing.The centrifugation is centrifuged 10min for 2000r/min.Contain in the mycotoxin cleaning buffer solution
There are 25g/L Sodium Chloride, 5g/L sodium bicarbonate, 0.01%v/v polysorbas20s, balance of water.The flow velocity during drip washing is 1 drop/s.Institute
The concrete operations for stating filtration are：0.22 μm of organic phase filter membrane of eluent Jing is filtered.
It is obvious to a person skilled in the art that the invention is not restricted to the details of above-mentioned one exemplary embodiment, Er Qie
In the case of spirit or essential attributes without departing substantially from the present invention, the present invention can be realized in other specific forms.Therefore, no matter
From the point of view of which point, embodiment all should be regarded as exemplary, and be nonrestrictive, the scope of the present invention is by appended power
Profit is required rather than described above is limited, it is intended that all in the implication and scope of the equivalency of claim by falling
Change is included in the present invention.Moreover, it will be appreciated that although this specification is been described by according to embodiment, not each
Embodiment only includes an independent technical scheme, and this narrating mode of description is only this area for clarity
Technical staff should using description as an entirety, the technical scheme in each embodiment can also Jing it is appropriately combined, form this
Art personnel may be appreciated other embodiment.
Claims (9)
1. a kind of ochratoxin A detection method of content, it is characterised in that comprise the following steps：Sample pre-treatments：Take vegetables and fruits sample
Product pulverize, and add extractant, mix, and using ultrasound wave 30～60min of Rapid Extraction, extraction temperature is 30～40 DEG C, and centrifugation takes
Supernatant is crossed ochratoxin A affine in immunity column purification by supernatant, successively with mycotoxin cleaning buffer solution, water wash immunity parent
And post, then with methanol-eluted fractions, eluent is collected, eluent is filtered, filtrate is taken and is obtained testing sample solution；The extractant
For acetonitrile and the mixed liquor of tetrahydrofuran；Based on parallel factor method (PARAFAC), prepare testing sample solution to set up quantitative point
The calibration model of analysis；Institute's positive model for school building is judged with testing sample solution, and verify the reliability of calibration model；With school
Positive model is determined to the content analysis of ochratoxin A in vegetables and fruits actual sample.
2. ochratoxin A detection method of content according to claim 1, it is characterised in that sample pre-treatments also include
Correction sample in linear latitude of formulation ochratoxin A, checking sample, take extract, and the scanning wavelength for optimize with
Its three-dimensional fluorescence excitation-emission matrix spectrum data is gathered under sweep spacing；Parallel factor method is adopted to resulting three-dimensional data battle array
(PARAFAC) carry out mathematics and separate parsing, and set up calibration model.
3. ochratoxin A detection method of content according to claim 1, it is characterised in that the consumption of the extractant
It is 0.4～0.6g/ml to make vegetables and fruits sample concentration.
4. ochratoxin A detection method of content according to claim 1, it is characterised in that the extractant is acetonitrile
With tetrahydrofuran by volume（0.8～1.2）：（0.8～1.2）The mixed liquor of mixing.
5. ochratoxin A detection method of content according to claim 1, it is characterised in that the extractant is acetonitrile
With tetrahydrofuran by volume 1：The mixed liquor of 1 mixing.
6. ochratoxin A detection method of content according to claim 1, it is characterised in that the centrifugation is 1800～
2200r/min is centrifuged 8～12min.
7. ochratoxin A detection method of content according to claim 1, it is characterised in that the mycotoxin cleaning
Contain 25g/L Sodium Chloride, 5g/L sodium bicarbonate, 0.01%v/v polysorbas20s, balance of water in buffer.
8. ochratoxin A detection method of content according to claim 1, it is characterised in that the flow velocity during drip washing
For 1 drop/s.
9. ochratoxin A detection method of content according to claim 1, it is characterised in that the concrete behaviour of the filtration
As：0.22 μm of organic phase filter membrane of eluent Jing is filtered.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112198250A (en) * | 2020-09-27 | 2021-01-08 | 常州金坛江南制粉有限公司 | Detection method of ochratoxin in glutinous rice |
| CN114235765A (en) * | 2021-12-10 | 2022-03-25 | 江西业力医疗器械有限公司 | Method for detecting dermatophyte |
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|---|---|---|---|---|
| CN103525927A (en) * | 2013-10-11 | 2014-01-22 | 南京师范大学 | Method for detecting ochratoxin A |
| CN103808716A (en) * | 2014-01-07 | 2014-05-21 | 江西省农业科学院农产品质量安全与标准研究所 | Method for portably and rapidly detecting ochratoxin A |
| CN104390946A (en) * | 2014-11-13 | 2015-03-04 | 中国农业大学 | Method for determining content of ochratoxin A in juice |
| CN104820032A (en) * | 2015-04-29 | 2015-08-05 | 深圳市宇驰检测技术有限公司 | Method for detecting ochratoxin A in vegetables and fruits |
| CN105543376A (en) * | 2016-01-24 | 2016-05-04 | 湖南科技大学 | Rapid detection method for ochratoxin A |
| CN105842441A (en) * | 2016-03-18 | 2016-08-10 | 南昌大学 | Detection method for ochratoxin A |
-
2016
- 2016-12-05 CN CN201611103356.3A patent/CN106556585A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103525927A (en) * | 2013-10-11 | 2014-01-22 | 南京师范大学 | Method for detecting ochratoxin A |
| CN103808716A (en) * | 2014-01-07 | 2014-05-21 | 江西省农业科学院农产品质量安全与标准研究所 | Method for portably and rapidly detecting ochratoxin A |
| CN104390946A (en) * | 2014-11-13 | 2015-03-04 | 中国农业大学 | Method for determining content of ochratoxin A in juice |
| CN104820032A (en) * | 2015-04-29 | 2015-08-05 | 深圳市宇驰检测技术有限公司 | Method for detecting ochratoxin A in vegetables and fruits |
| CN105543376A (en) * | 2016-01-24 | 2016-05-04 | 湖南科技大学 | Rapid detection method for ochratoxin A |
| CN105842441A (en) * | 2016-03-18 | 2016-08-10 | 南昌大学 | Detection method for ochratoxin A |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112198250A (en) * | 2020-09-27 | 2021-01-08 | 常州金坛江南制粉有限公司 | Detection method of ochratoxin in glutinous rice |
| CN114235765A (en) * | 2021-12-10 | 2022-03-25 | 江西业力医疗器械有限公司 | Method for detecting dermatophyte |
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Application publication date: 20170405 |