CN103645265B - The irradiation-induced degradation facture of a kind of FT and ochratoxin - Google Patents
The irradiation-induced degradation facture of a kind of FT and ochratoxin Download PDFInfo
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Abstract
The present invention relates to the irradiation-induced degradation facture of a kind of FT and ochratoxin; Comprise the following steps: (1) standard items preparing standard solution to FT FB1 and ochratoxin OTA, be the radiation treatment of 0-200kGy through irradiation range to standard solution, measured by the content of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (2) after the irradiation that irradiation range is 0-9kGy, pre-service is carried out to the malicious sample of band, measured by the degradation effect of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (3) choose the highest FB1 and the OTA standard solution of concentration as sample, after the radiation treatment that irradiation range is 0-200kGy, carry out detection by liquid chromatography-tandem mass spectrometry analyze catabolite; Solve research and the environmental protection application to lacking irradiation-induced degradation FB1 and OTA catabolite; Good degrading effect and realize to agricultural product refuse environmental protection detoxification treatment of going mouldy.
Description
Technical field
The invention belongs to analytical chemistry field, be specifically related to the irradiation-induced degradation facture of a kind of FT and ochratoxin.
Background technology
FT and ochratoxin belong to mycotoxin, mycotoxin is the toxic metabolic products produced under adapt circumstance condition by Toxigenic fungi, FT and ochratoxin endanger especially greatly the mankind and pollution frequency is high, have formulated strict limit standard both at home and abroad to the FT in Cereals, food and feed and ochratoxin.
In Chinese patent 201110000805.2 " immune affinity column purification high performance liquid chromatography detects the method for ochratoxin A in Chinese medicine tincture ", disclose and by tablets by HPLC-MS, positive findings is confirmed; In Chinese patent 201010209190.X " a kind of method detecting nivalenol and deoxynivalenol toxin in Chinese medicine ", disclose and under full scan pattern, confirm toxin by tablets by HPLC-MS; In Chinese patent 201110370420.5 detection method of several mycotoxin levels " in the wheat ", disclose and detected by tablets by HPLC-MS, by the regression equation calculation DON of toxin concentration for peak area, the content of T-2 toxin; In Chinese patent 201210054250.4 method of mycotoxin levels " in the LC-MS/MS Araliaceae ", disclosing can be accurate, measures the multicomponent residual quantity of mycotoxin in Araliaceae delicately; In Chinese patent 201210361145.5 " method with Algae toxins in electron beam irradiation-induced degradation water ", disclose when radiation dose is 5kGy, the generation of Algae toxins can be suppressed; In Chinese patent 2009100787702.0 " method of degrading fumonisins ", disclose irradiation can make fumonisins B
1degradation rate reaches 63.47%; In Chinese patent 200910078703.5 " a kind of method of degrading fumonisins ", disclose irradiation can make fumonisins B
1degradation rate reaches 59.03%; Irradiation detoxification is disclosed without industrial pollution in Chinese patent 200910078708.8 " method of aflatoxin degradation "; In Chinese patent 200910078707.3 " a kind of method of aflatoxin degradation ", disclose irradiation can make AFB
1degradation rate reaches 71.51%.
Existing correlation technique has: Zhang Xichun etc. report that the degraded of irradiation dose to mould and mycotoxin of 3kGy ~ 5kGy has obvious effect; Poplar waits quietly studying and shows that mass concentration is that aflatoxin B1 solution degradation rate when 4kGy of 0.1mg/L fumonisins B1 solution degradation rate when 10kGy that degradation rate reaches 96%, 0.1mg/L when can reach more than 80%, 6kGy reaches more than 90%; The result of study of Yin Qinggang shows when irradiation dose is 18kGy, in powdered form corn, zearalenone degradation rate can reach 74.9%, when irradiation dose is 10kGy, in whole kernel corn, zearalenone degradation rate can reach 94.1%, when irradiation dose is 5kGy, the degradation rate of 30.0mg/kg zearalenone aqueous solution reaches 90%; The people such as Chi Lei have delivered the paper of gamma-rays to the irradiation-induced degradation of ochratoxin A, mention the possible mechanism of the method for catabolite structure elucidation, degraded, but do not report the product of degraded in literary composition.
Summary of the invention
The object of this invention is to provide a kind of good degrading effect, realize the irradiation-induced degradation facture of FT to agricultural product refuse environmental protection detoxification treatment and ochratoxin.
The technical scheme realizing the object of the invention is the irradiation-induced degradation facture of a kind of FT and ochratoxin; Comprise the following steps: (1) standard items preparing standard solution to FT FB1 and ochratoxin OTA, be the radiation treatment of 0-200kGy through irradiation range to standard solution, measured by the content of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (2) after the irradiation that irradiation range is 0-9kGy, pre-service is carried out to the malicious sample of band, measured by the degradation effect of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (3) choose the highest FB1 and the OTA standard solution of concentration as sample, after the radiation treatment that irradiation range is 0-200kGy, carry out detection by liquid chromatography-tandem mass spectrometry analyze catabolite.
Preferred described irradiation bomb is gamma-rays, the chromatographic condition of described liquid chromatography-tandem mass spectrometry: adopt ThermoFinnigan liquid chromatographic system, it is SepaxBR-C18 post 100 × 2.1mm that its liquid-phase condition comprises chromatographic column, 5 μm; Mobile phase is A: methyl alcohol, B: Cu Suan An – aqueous formic acid, under flow velocity 250 μ L/min and sample size 25 μ L condition, carries out gradient elution; Concrete gradient is:
| Time | A% | B% |
| 0.00 | 15 | 85 |
| 2.00 | 15 | 85 |
| 2.10 | 90 | 10 |
| 5.00 | 90 | 10 |
| 5.10 | 15 | 85 |
| 7.00 | 15 | 85 |
The Mass Spectrometry Conditions of described liquid chromatography-tandem mass spectrometry: adopt ThermoTSQUltra triple level Four bar mass spectrometer system, it is ESI that its Mass Spectrometry Conditions comprises Ionization mode, and spray voltage is 3000V, and polar mode is positive ion, atomization temperature is 400 DEG C, sheath gas is 48arb, and assisted gas is 10arb, and ion transfer tube temperature is 400 DEG C, sweep length is 0.01m/z, sweep time is 0.1s, and resolution is Q1=0.7FWHM, Q3=0.7FWHM; Ion pair information, wherein * is quota ion pair:
Standard model in preferred described step (1) is
Wherein the compound method of FB1 standard solution selects the purity of Enzo company to be 98.0%, 5mg*2 bottle, and variable concentrations has concentration to be 0.8mg/mL respectively, draws in 315 μ L to 25mL volumetric flasks, dissolve constant volume with acetonitrile+water (volume ratio 50:50); Concentration is 10.0 μ g/mL, draws in 1.0mL to 10mL volumetric flask, dissolves constant volume with acetonitrile+water (volume ratio 50:50); Concentration is 1.0 μ g/mL, draws in 1.0mL to 10mL volumetric flask, dissolves constant volume with acetonitrile+water (volume ratio 50:50); Concentration is 100.0ng/mL, draws in 500 μ L to 10mL volumetric flasks, dissolves constant volume with acetonitrile+water (volume ratio 50:50); Concentration is also had to be 50.0ng/mL; Wherein the compound method of OTA standard solution selects the purity of Enzo company to be 98.0%, 5mg*1 bottle, and variable concentrations has concentration to be 0.7mg/mL respectively, draws in 360 μ L to 25mL volumetric flasks, dissolve constant volume with methyl alcohol; Concentration is 10.0 μ g/mL, draws in 500 μ L to 5mL volumetric flasks, dissolves constant volume with methyl alcohol; Concentration is 1.0 μ g/mL, draws in 500 μ L to 5mL volumetric flasks, dissolves constant volume with methyl alcohol; Concentration is 100.0ng/mL, draws in 500 μ L to 5mL volumetric flasks, also has concentration to be 10.0ng/mL; 0.8mg/mL to FB1 concentration, OTA concentration is that the standard solution of 0.7mg/mL adopts irradiation dose 0,3,5,7,9,20,50,100 and 200kGy process respectively, adopts irradiation dose 0,3,5,7 and 9kGy process respectively to the standard solution of other concentration.
In preferred described step (2), the malicious sample of band has 6, is go mouldy 2, corn seed sample (being numbered A, B), rice sample 2 (being numbered C, D), wheat (being numbered E) and soya bean sample each 1 (being numbered F) respectively; Preprocess method accurately takes sample 5.00g (± 0.05g) in the centrifuge tube of 50mL, adds 10mL86% acetonitrile solution, vortex 30s, ultrasonic 30min, the centrifugal 5min of 4000r/min, transfer supernatant, in test tube, extracts with 10mL86% acetonitrile solution again, merge extracted twice liquid, dry up at 45 DEG C with nitrogen, then use acetonitrile+water (volume ratio 50:50) to be settled to 1mL, vortex 30s, with 0.22 μm of filtering with microporous membrane, in order to sample introduction; Irradiation dose is 0,3,5,7 and 9kGy.
To the standard solution of the highest 0.7mg/mL of 0.8mg/mL and the OTA concentration that FB1 concentration is the highest in preferred described step (3), after adopting irradiation dose 0,3,5,7,9,20,50,100 and 200kGy to carry out radiation treatment respectively, detect by full scan pattern, observe the formation with or without characteristic peak.
Preferred when irradiation dose is 3-9kGy, detoxification treatment can be carried out to having commercial value agricultural product, when irradiation dose is 20-200kGy, can to without commercial value go mouldy agricultural product and or other material that goes mouldy carry out concentrated detoxification treatment.
Preferred when irradiation dose is 7-9kGy, detoxification treatment can be carried out to having commercial value agricultural product, when irradiation dose is 100-200kGy, can to without commercial value go mouldy agricultural product and or other material that goes mouldy carry out concentrated detoxification treatment.
The present invention has positive effect: the radiation treatment of (1) 3-9kGy has degradation effect to FB1, and during 9kGy, concentration is that the degradation rate of the FB1 of 0.8mg/mL, 10.0ug/mL, 1.0ug/mL and 50ng/mL is respectively 22.5%, 51.0%, 59.0% and 64.8%; The radiation treatment of 3-9kGy can be degraded OTA, but DeGrain.When irradiation dose is 100kGy, when the degradation rate of FB1, OTA all meets or exceeds 90%, 200kGy, FB1 and OTA almost all degrades.
(2) screening 6 may with poison sample in have the testing result of 2 samples to be positive, be respectively A and B sample.Detected FB1 in A, B sample, the change that in 2 samples, FB1 content produces with the increase of irradiation dose is substantially identical, and dosage lower than there being the trend raised with the increase of dosage during 9kGy, but significantly reduces again during 9kGy.
(3) feature catabolite is had no after FB1 radiation treatment, compare entirely sweeping collection of illustrative plates with the collection of illustrative plates of control sample, the collection of illustrative plates of irradiation sample creates change, doubtful have catabolite to be formed, but owing to containing acetonitrile in the solvent that FB1 standard items adopt, so compare with the collection of illustrative plates of solvent blank, find that the peak produced is solvent peak; New characteristic peak is had to be formed after OTA standard solution irradiation, the material of m/z=398.19 is created by analysis after radiation treatment, this material starts when irradiation dose is 50kGy to occur, during 100kGy, response is the highest, close when response during 200kGy and 100kGy.
(4) detoxification process can be made pollution-free by irradiation-induced degradation, have good environment protecting.Dosage is that the radiation treatment of 3-200kGy has degradation effect to FB1 and OTA, better to FB1 degradation effect under the irradiation dose of 3-9kGy of low concentration.The irradiation dose of 3-9kGy can be used for the agricultural product detoxification having commercial value, as: grain, food, feed etc., preferred 7-9kGy; The irradiation dose of 20-200kGy can do the utilization of environmental protection aspect, for no longer having the concentrated detoxification treatment of go mouldy agricultural product or other material that goes mouldy of commercial value, preferred 50-200kGy, most preferably 100-200kGy.
Accompanying drawing explanation
In order to make content of the present invention more easily be clearly understood, below according to specific embodiment also by reference to the accompanying drawings, the present invention is further detailed explanation, wherein
Fig. 1 is the typical curve of FB1;
Fig. 2 is the typical curve of OTA;
Fig. 3 be FB1 standard solution control sample entirely sweep collection of illustrative plates;
Fig. 4 be FB1 standard solution irradiation sample entirely sweep collection of illustrative plates (200kGy);
Fig. 5 sweeps collection of illustrative plates (200kGy) after FB1 solvent irradiation entirely;
Fig. 6 be OTA standard solution control sample entirely sweep collection of illustrative plates;
Fig. 7 is that collection of illustrative plates (200kGy) swept entirely by OTA standard solution irradiation sample;
Fig. 8 sweeps collection of illustrative plates (200kGy) after OTA solvent irradiation entirely;
Fig. 9 is OTA standard solution control sample collection of illustrative plates (m/z=398.19);
Standard solution irradiation master drawing spectrum (m/z=398.19) of Figure 10 to be OTA dosage be 20kGy;
Standard solution irradiation master drawing spectrum (m/z=398.19) of Figure 11 to be OTA dosage be 50kGy;
Standard solution irradiation master drawing spectrum (m/z=398.19) of Figure 12 to be OTA dosage be 100kGy;
Standard solution irradiation master drawing spectrum (m/z=398.19) of Figure 13 to be OTA dosage be 200kGy;
Embodiment
To the standard model preparing standard solution of FT FB1, ochratoxin OTA, detect the typical curve obtaining toxin, see Fig. 1 and Fig. 2:
FB1：Y＝-4407.4+35124.6*X、R
2＝0.9970，OTA：Y＝116009+673499*X、R
2＝0.9979；
The standard solution of variable concentrations is all respectively after the gamma-ray irradiation process that dosage is 3-9kGy, and the content of contratoxin detects, the sample in contrast of the standard solution without irradiation that each toxin is corresponding, and the response of toxin is in table 1.The result display of table 3, the radiation treatment of 3-9kGy has degradation effect to FB1, and during 9kGy, concentration is that the degradation rate of the FB1 of 0.8mg/mL, 10.0ug/mL, 1.0ug/mL and 50ng/mL is respectively 22.5%, 51.0%, 59.0% and 64.8%; The radiation treatment of 3-9kGy can be degraded OTA, but DeGrain.Due in the dosage range of 3-9kGy, the degradation effect of the standard solution of each toxin maximum concentration is remarkable, so the irradiation dose adding them is to 20-200kGy, response is in table 2.The result display of table 3, when irradiation dose is 100kGy, when the degradation rate of FB1, OTA all meets or exceeds 90%, 200kGy, FB1 and OTA almost all degrades.
The response of table 1 toxin standard solution after various dose irradiation (3-9kGy)
The response of table 2 high concentration toxin standard solution after various dose irradiation (20-200kGy)
The degradation rate (%) of the toxin of table 3 standard solution after various dose irradiation
External standard method is adopted to the detection of the recovery of the detection method of toxin in the malicious sample of band; It is FB1:10.0 μ g/kg, OTA:5.0 μ g/kg that sample A-F adds scalar; The method recovery is respectively FB1:53.7-87.2%, OTA:43.0-57.8%.
Screening 6 may with poison sample in have the FB1 testing result of 2 samples to be positive, be respectively A and B sample.The change that in 2 samples, FB1 content produces with the increase of irradiation dose is substantially identical, and dosage lower than there being the trend raised with the increase of dosage during 9kGy, but significantly reduces again during 9kGy.
The irradiation-induced degradation effect of toxin in table 4 sample
Using the highest standard solution of FB1 and OTA concentration as sample, FB1:0.8mg/mL, OTA:0.7mg/mL, respectively after the radiation treatment of irradiation dose 0,3,5,7,9,20,50,100 and 200kGy, detect by the pattern of full scan, catabolite is analyzed.
Feature catabolite is had no after FB1 radiation treatment.As can be seen from entirely sweeping collection of illustrative plates, compare with the collection of illustrative plates (Fig. 3) of control sample, the collection of illustrative plates of irradiation sample creates change (Fig. 4), doubtful have catabolite to be formed, but owing to containing acetonitrile in the solvent that FB1 standard items adopt, so compare with the collection of illustrative plates (Fig. 5) of solvent blank, find that the peak produced is solvent peak.
Formed as can be seen from there being new characteristic peak after Fig. 6-8, OTA standard solution irradiation.By analysis (Fig. 9-13), create the material of m/z=398.19 after radiation treatment, this material starts when irradiation dose is 50kGy to occur, during 100kGy, response is the highest, close when response during 200kGy and 100kGy.
Above-described specific embodiment; object of the present invention, technical scheme and beneficial effect are further described; be understood that; the foregoing is only specific embodiments of the invention; be not limited to the present invention; within the spirit and principles in the present invention all, any amendment made, equivalent replacement, improvement etc., all should be included within protection scope of the present invention.
Claims (5)
1. the irradiation-induced degradation facture of a FT and ochratoxin; It is characterized in that: comprise the following steps: (1) standard items preparing standard solution to FT FB1 and ochratoxin OTA, be the radiation treatment of 0-200kGy through irradiation range to standard solution, measured by the content of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (2) after the irradiation that irradiation range is 0-9kGy, pre-service is carried out to the malicious sample of band, measured by the degradation effect of liquid chromatography-tandem mass spectrometry to FB1 and OTA; (3) choose the highest FB1 and the OTA standard solution of concentration as sample, after the radiation treatment that irradiation range is 0-200kGy, carry out detection by liquid chromatography-tandem mass spectrometry analyze catabolite; When irradiation dose is 3-9kGy, detoxification treatment can be carried out to having commercial value agricultural product, when irradiation dose is 20-200kGy, can to without commercial value go mouldy agricultural product and or other material that goes mouldy carry out concentrated detoxification treatment; The irradiation bomb of described irradiation is gamma-rays, the chromatographic condition of described liquid chromatography-tandem mass spectrometry: adopt ThermoFinnigan liquid chromatographic system, it is SepaxBR-C18 post 100 × 2.1mm that its liquid-phase condition comprises chromatographic column, 5 μm; Mobile phase is A: methyl alcohol, B: Cu Suan An – aqueous formic acid, under flow velocity 250 μ L/min and sample size 25 μ L condition, carries out gradient elution; Concrete gradient is:
The Mass Spectrometry Conditions of described liquid chromatography-tandem mass spectrometry: adopt ThermoTSQUltra triple level Four bar mass spectrometer system, it is ESI that its Mass Spectrometry Conditions comprises Ionization mode, and spray voltage is 3000V, and polar mode is positive ion, atomization temperature is 400 DEG C, sheath gas is 48arb, and assisted gas is 10arb, and ion transfer tube temperature is 400 DEG C, sweep length is 0.01m/z, sweep time is 0.1s, and resolution is Q1=0.7FWHM, Q3=0.7FWHM; Ion pair information, wherein * is quota ion pair:
2. the irradiation-induced degradation facture of FT according to claim 1 and ochratoxin, is characterized in that: the standard solution in described step (1) is
wherein the compound method of FB1 standard solution selects the standard model of Enzo company, standard model purity is 98.0%, 5mg*2 bottle, variable concentrations has concentration to be 0.8mg/mL respectively, draw in 315 μ L to 25mL volumetric flasks, with acetonitrile and the water-soluble solution constant volume of volume ratio 50:50, obtain the solution that concentration is 10.0 μ g/mL; Concentration is 10.0 μ g/mL, draws in 1.0mL to 10mL volumetric flask, with acetonitrile and the water-soluble solution constant volume of volume ratio 50:50, obtains the solution that concentration is 1.0 μ g/mL; Concentration is 1.0 μ g/mL, draws in 1.0mL to 10mL volumetric flask, with acetonitrile and the water-soluble solution constant volume of volume ratio 50:50, obtains the solution that concentration is 100.0ng/mL; Concentration is 100.0ng/mL, draws in 5.0mL to 10mL volumetric flask, with acetonitrile and the water-soluble solution constant volume of volume ratio 50:50, obtains the solution that concentration is 50.0ng/mL; Wherein the compound method of OTA standard solution selects the standard model of Enzo company, and standard model purity is 98.0%, 5mg*1 bottle, variable concentrations has concentration to be 0.7mg/mL respectively, draw in 360 μ L to 25mL volumetric flasks, dissolve constant volume with methyl alcohol, obtain the solution that concentration is 10.0 μ g/mL; Concentration is 10.0 μ g/mL, draws in 500 μ L to 5mL volumetric flasks, dissolves constant volume with methyl alcohol, obtains the solution that concentration is 1.0 μ g/mL; Concentration is 1.0 μ g/mL, draws in 500 μ L to 5mL volumetric flasks, dissolves constant volume with methyl alcohol, obtain the solution that concentration is 100.0ng/mL; Concentration is 100.0ng/mL, draws in 500 μ L to 5mL volumetric flasks, dissolves constant volume with methyl alcohol, obtain the solution that concentration is 10.0ng/mL; 0.8mg/mL to FB1 concentration, OTA concentration is that the standard solution of 0.7mg/mL adopts irradiation dose 0,3,5,7,9,20,50,100 and 200kGy process respectively, adopts irradiation dose 0,3,5,7 and 9kGy process respectively to the standard solution of other concentration.
3. the irradiation-induced degradation facture of FT according to claim 1 and ochratoxin, it is characterized in that: the malicious sample of band has 6 in described step (2), is go mouldy that 2, corn seed sample is numbered A, B, rice sample 2 is numbered C, D, wheat is numbered E and soya bean sample number into spectrum is each 1 of F respectively; Preprocess method accurately takes sample 5.00g ± 0.05g in the centrifuge tube of 50mL, adds 10mL86% acetonitrile solution, vortex 30s, ultrasonic 30min, the centrifugal 5min of 4000r/min, transfer supernatant, in test tube, extracts with 10mL86% acetonitrile solution again, merge extracted twice liquid, dry up at 45 DEG C with nitrogen, then be settled to 1mL, vortex 30s with the acetonitrile of volume ratio 50:50 and water, with 0.22 μm of filtering with microporous membrane, in order to sample introduction; Irradiation dose is 0,3,5,7 and 9kGy.
4. the irradiation-induced degradation facture of FT according to claim 1 and ochratoxin, it is characterized in that: to the standard solution of the highest 0.7mg/mL of 0.8mg/mL and the OTA concentration that FB1 concentration is the highest in described step (3), after adopting irradiation dose 0,3,5,7,9,20,50,100 and 200kGy to carry out radiation treatment respectively, detect by full scan pattern, observe the formation with or without characteristic peak.
5. the irradiation-induced degradation facture of FT according to claim 1 and ochratoxin, it is characterized in that: when irradiation dose is 7-9kGy, detoxification treatment can be carried out to having commercial value agricultural product, when irradiation dose is 100-200kGy, can to without commercial value go mouldy agricultural product and or other material that goes mouldy carry out concentrated detoxification treatment.
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| CN103931970A (en) * | 2014-05-14 | 2014-07-23 | 河南工业大学 | Method for reducing deoxynivalenol in wheat with gibberellic disease |
| CN104017679A (en) * | 2014-05-18 | 2014-09-03 | 中国食品发酵工业研究院 | Method for reducing OTA (ochratixin A) in beer malting process |
| CN104900295B (en) * | 2015-05-28 | 2018-04-20 | 官爱平 | Irradiation apparatus and its method for fluid processing |
| CN106721926A (en) * | 2016-11-09 | 2017-05-31 | 中国农业科学院农产品加工研究所 | The method of irradiation-induced degradation ochratoxin |
| CN108225884B (en) * | 2017-12-15 | 2020-10-27 | 江南大学 | Combined degradation method of zearalenone and ochratoxin A in solution |
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