CN107677758B - Method for detecting additive in livestock and poultry meat product - Google Patents
Method for detecting additive in livestock and poultry meat product Download PDFInfo
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- CN107677758B CN107677758B CN201710911432.1A CN201710911432A CN107677758B CN 107677758 B CN107677758 B CN 107677758B CN 201710911432 A CN201710911432 A CN 201710911432A CN 107677758 B CN107677758 B CN 107677758B
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Abstract
The invention discloses a method for detecting additives in livestock and poultry meat products, which aims at the current situation of additive detection, adopts a pretreatment method of ethanol aqueous solution extraction with volume concentration of 50% and neutral alumina column purification, and applies ultra-high performance liquid chromatography-tandem quadrupole linear ion trap mass spectrometer to establish a detection method for simultaneously screening and confirming 15 additives in livestock and poultry meat products.
Description
Technical Field
The invention relates to the technical field of additive detection, in particular to a method for detecting an additive in a livestock and poultry meat product.
Background
In the processing engineering of cooked meat products sold in the market, food additives such as preservatives, sweeteners and the like are added for pursuing color and luster, prolonging the shelf life and the like, although China already puts requirements on the addition limit of the food additives, the phenomenon of abusing the food additives is still very serious, and the long-term consumption of food containing a large amount of food additives brings great potential harm to the health of human bodies.
Degradation of the quality of the meat product may result due to decomposition of proteins and fats in the meat product. In order to prolong the shelf life and prevent the deterioration of meat products, preservatives are often added during the processing of meat products. Mouthfeel is the most important indicator in evaluating cooked meat products, which are often used with sweeteners in order to pursue mouthfeel during processing. At present, according to the currently allowable food additive regulations of national standard GB2760-2007, sorbic acid and potassium salt thereof are commonly used as preservatives in meat products. Other preservatives such as benzoic acid are subject to strict restrictions and are not detected by national standards. High-content sweeteners are still widely used in food in China, but according to the regulations of the national standard GB2760-2007 on relevant standards of meat and food additives in meat products, only natural sweeteners are allowed to be added into the meat products, and if artificially synthesized sweeteners are detected in meat product samples, the artificially synthesized sweeteners are illegally added.
Development of additive analysis research and development of related detection methods is a very important step in enhancing additive monitoring. With the increasing emphasis on food safety, food inspection organizations face a lot of detection works, but the detection method at present mainly detects one or more additives, so that the samples need to be repeatedly processed and respectively detected, which is time-consuming and labor-consuming. And the method aims at the detection of target compounds, the screening of various additives cannot be realized, and false positive samples can still be detected.
Therefore, how to develop a reagent that can detect multiple additives simultaneously, especially screening and confirming multiple additives in a sample is a problem to be solved by many additive analysis workers.
Disclosure of Invention
In view of this, the invention discloses a method for detecting additives in livestock and poultry meat products, which at least solves the problems that the prior detection method mainly detects one or more additives, so that the detected samples need to be repeatedly processed and respectively detected, the time and the labor are consumed, the detection efficiency is low, and the like.
The technical scheme provided by the invention is specifically a method for detecting additives in livestock and poultry meat products, which is characterized by comprising the following steps:
1) sample pretreatment:
weighing a sample, placing the sample in a centrifuge tube, adding 50% ethanol water solution by volume concentration, performing oscillation extraction and centrifugation, collecting supernatant in another centrifuge tube, adding 50% ethanol water solution by volume concentration, and repeatedly extracting once to obtain an extracting solution;
purifying the extract by a neutral alumina column, concentrating, and fixing volume for later use;
2) and (3) qualitative analysis:
screening the livestock and poultry meat product sample to be detected after sample pretreatment by using an UPLC-MS/MS-Qtrap method, establishing an MRM-IDA-EPI screening method by adopting an enhanced ion scanning function of a liquid chromatography-tandem quadrupole linear ion trap mass spectrometer, and obtaining MS of each target additive parent ion in the livestock and poultry meat product sample to be detected2Spectra, using standard MS for each additive in a known database2Spectrogram and MS for obtaining target additive parent ions in livestock and poultry meat product sample to be detected2Respectively comparing the spectrograms to obtain similarity values, matching characteristic fragment ions of the spectrograms and the characteristic fragment ions of the spectrograms, and further judging whether the livestock and poultry meat product sample to be detected contains the target additive or not;
3) quantitative analysis:
weighing each additive standard substance respectively, and preparing 10 mug/mL standard stock solution with water respectively for later use;
processing a blank sample according to the sample pretreatment method in the step 1), and diluting the standard stock solution by using a blank sample matrix solution to prepare a mixed standard stock solution;
after the mixed standard stock solution is subjected to constant volume, obtaining a blank sample matrix solution, and preparing a series of matching standard working solutions with the concentration of 0.05 mu g/mL-5.0 mu g/mL;
respectively measuring the series of matching standard working solutions and the livestock and poultry meat product sample to be detected after sample pretreatment by adopting a liquid chromatogram-series quadrupole linear ion trap mass spectrometer to obtain a drawn standard curve with the concentration as a horizontal coordinate and the peak area of the quantitative ions as a vertical coordinate, and calculating the content of the target additive in the livestock and poultry meat product sample to be detected by using the standard curve.
Preferably, the additive is one or more of saccharin sodium, acesulfame potassium, sodium cyclamate, aspartame, sorbic acid, benzoic acid, dehydroacetic acid, ethyl naphthol, salicylic acid, alitame, neotame, methyl paraben, ethyl paraben, propyl paraben and butyl paraben.
More preferably, the sample pretreatment in the step 1) is specifically as follows:
weighing 5.00g of sample, placing the sample in a 100mL centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, shaking for extraction for 20min, centrifuging for 5min at 8000r/min, collecting supernatant in another centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, and repeatedly extracting once to obtain an extracting solution;
passing the extractive solution through neutral alumina-filled glass column, collecting 20mL solution, rotary evaporating at 40 deg.C to near dryness, adding 1.0mL ethanol water solution with solubility of 20% to dissolve residue, and filtering with 0.22 μm filter membrane.
Further preferably, the mass spectrum conditions of the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the step 2) are as follows:
an ion source: an electrospray ion source;
scanning mode: scanning negative ions;
an acquisition mode: monitoring multiple reactions;
information association scanning: intensity threshold 100 cps;
enhancer ion scanning: the mass number acquisition range is m/z 50-1000 Da;
electrospray voltage: -4500V;
atomizing gas pressure: 50 psi;
air curtain pressure: 30 psi;
auxiliary gas pressure: 50 psi;
ion source temperature: 500 ℃;
inlet voltage: -10V;
collision cell exit voltage: -10V;
energy of collision gas: 20. 35, 50V;
expansion of collision energy: 15V.
Further preferably, the chromatographic conditions of the liquid chromatography-tandem quadrupole linear ion trap mass spectrometer in the step 3) are as follows:
a chromatographic column: waters BEH C18A reverse phase chromatography column;
column temperature: 40 ℃;
sample introduction volume: 10 mu L of the solution;
flow rate: 0.2 mL/min;
mobile phase: mobile phase A: 0.02mol/L ammonium acetate aqueous solution, B: methanol, gradient elution procedure: 0min, 90% A; 1min, 90% A; 5min, 10% A; 7min, 10% A; 7.1min, 90% A; 12min, 90% A;
the mass spectrum conditions of the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the step 3) are as follows:
an ion source: an electrospray ion source;
scanning mode: scanning negative ions;
an acquisition mode: monitoring multiple reactions;
electrospray voltage: -4500V;
atomizing gas pressure: 50 psi;
air curtain pressure: 30 psi;
auxiliary gas pressure: 50 psi;
ion source temperature: 500 ℃;
inlet voltage: -10V;
collision cell exit voltage: -10V.
Further preferably, the livestock and poultry meat product sample to be detected is sauced beef or sausage.
The method for detecting the additives in the livestock and poultry meat product provided by the invention adopts a pretreatment method of extracting by using an ethanol aqueous solution with the volume concentration of 50% and purifying by using a neutral alumina column and applies an ultra-high performance liquid chromatography-tandem quadrupole linear ion trap mass spectrometer to establish a detection method for simultaneously screening and confirming the 15 additives in the livestock and poultry meat product, has the characteristics of simple operation, high flux efficiency, accurate qualification, high sensitivity, environmental friendliness and the like, is suitable for screening and confirming various additives in the meat product, can greatly improve the detection efficiency, and avoids the detection of false positive samples.
Drawings
The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the invention and together with the description, serve to explain the principles of the invention.
In order to more clearly illustrate the embodiments or technical solutions in the prior art of the present invention, the drawings used in the description of the embodiments or prior art will be briefly described below, and it is obvious for those skilled in the art that other drawings can be obtained based on these drawings without creative efforts.
FIG. 1 shows MS of suspicious peaks in a livestock and poultry meat product to be detected according to an embodiment of the disclosure2A spectrogram;
FIG. 2 shows a standard MS for rhodamine B as an additive provided in an embodiment of the disclosure2Spectra.
Detailed Description
Reference will now be made in detail to the exemplary embodiments, examples of which are illustrated in the accompanying drawings. When the following description refers to the accompanying drawings, like numbers in different drawings represent the same or similar elements unless otherwise indicated. The embodiments described in the following exemplary embodiments do not represent all embodiments consistent with the present invention. Rather, they are merely examples of methods consistent with certain aspects of the invention, as detailed in the appended claims.
The method for simultaneously detecting the content of 15 additives including the preservative and the sweetener in the livestock and poultry meat product comprises the following steps:
sample pretreatment:
1. sample extraction:
accurately weighing 5.00g (accurate to 0.01g) of sample, placing the sample in a 100mL centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, extracting for 20min by shaking, centrifuging for 5min at 8000r/min, collecting supernatant in another centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, and repeatedly extracting once to obtain extract;
2. sample purification:
passing the extractive solution through neutral alumina-filled glass column, collecting 20mL solution, rotary evaporating at 40 deg.C to near dryness, adding 1.0mL ethanol water solution with solubility of 20% to dissolve residue, and filtering with 0.22 μm filter membrane.
And (3) qualitative analysis:
qualitative determination by UPLC-MS/MS-Qtrap method: the MRM-IDA-EPI screening and confirming method can realize one-time completion of qualitative and quantitative detection and can be applied to analysis of complex matrix samples.
Firstly, a liquid chromatogram-series quadrupole linear ion trap mass spectrometer is adopted to carry out detection scanning on a livestock and poultry meat product sample to be detected after sample pretreatment, and a system automatically judges whether the signal intensity acquired by the detection scanning exceeds each set value in IDA condition setting, namely whether a chromatographic peak appears. When the preset value is satisfied, the system is fast: (<1MS) is automatically switched into a linear ion trap mode, enhanced ion scanning (EPI) is carried out, and high-quality MS of parent ions obtained by detection scanning is obtained2Spectra, using standard MS for each additive in a known database2Spectrogram and MS for obtaining target additive parent ions in livestock and poultry meat product sample to be detected2Respectively comparing the spectrograms to obtain similarity values, matching characteristic fragment ions of the spectrograms and the characteristic fragment ions of the spectrograms, and further judging whether the ions to be detected are to be detectedWhether the target additive is contained in the livestock and poultry meat product sample.
Quantitative analysis:
and (3) determining the matrix matching standard working solution and the sample to be detected, drawing a standard curve by taking the concentration as a horizontal coordinate and the peak area of the quantitative ions as a vertical coordinate, and calculating the content of the target additive in the sample to be detected by using the standard curve.
Wherein, the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the quantitative analysis adopts the chromatogram conditions as follows: acquisty Waters BEH C18 column (2.1X 100mm, 1.7 μm), mobile phase A: 0.02% aqueous ammonium acetate; b: methanol was subjected to gradient elution under the conditions shown in Table 1, at a flow rate of 0.2mL/min, a column temperature of 40 ℃ and a sample injection volume of 10. mu.L.
TABLE 1 liquid chromatography gradient elution procedure
The mass spectrum conditions adopted by the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer are as follows: ionization mode: electrospray negative ion ESI (-); the detection mode is as follows: multiple reaction monitoring scans (MRM); electrospray voltage (IS): -4500V; atomization gas pressure (GS 1): 50 psi; air curtain pressure (CUR): 30 psi; assist gas pressure (GS 2): 50 psi; ion source Temperature (TEM): 500 ℃; inlet voltage (EP): -10V; collision cell exit voltage (CXP): -10V.
The retention times and mass spectrometric acquisition parameters for the 15 additives are shown in table 2.
TABLE 2 Mass Spectrometry parameters of additives tested
Quantification of ions.
Compared with the prior art, the method for detecting the additive in the livestock and poultry meat product provided by the embodiment has the following excellent effects:
(1) the embodiment establishes a UPLC-MS/MS-Qtrap screening and confirming method for analyzing 15 additives in the livestock and poultry meat products, and realizes the simultaneous detection of various additives.
(2) The embodiment adopts a sample pretreatment method of extracting by using an ethanol water solution with the volume concentration of 50% and purifying by using a neutral alumina column, realizes the simultaneous extraction and purification of 15 additives, and has the advantages of simplicity, convenience, high efficiency, rapidness, environmental friendliness and the like.
(3) The extraction solution adopted by the embodiment is an ethanol water solution with the volume concentration of 50%, only one common solution of ethanol needs to be added, and the ethanol has low toxicity and is beneficial to environmental protection. The method adopts a filled neutral alumina glass column purification mode, has low cost and good purification effect, and the pretreatment method has simple and convenient operation, low cost and environment-friendly solvent.
(4) The UPLC-MSMS-Qtrap method which is good in selectivity and strong in anti-interference capability is adopted for detection, stability is good, sensitivity is high, accuracy and precision meet the requirements of a multi-residue analysis method, and meanwhile, the UPLC-MSMS-Qtrap method has screening and confirming functions and can avoid detection of false positive samples. Selecting MRM-IDA-EPI function, setting related judgment standard in IDA, connecting liquid phase, inputting liquid phase collection condition, collecting data, triggering EPI to collect MS by enhanced ion scanning once MRM signal intensity exceeds judgment standard value2MS to open its suspicious peaks (judged by retention time, peak shape, etc.) for the collected maps2And the image is subjected to operations such as searching of a known spectrum library and the like, and is compared with mass spectrum data of the spectrum library to obtain corresponding matching data for detecting judgment residue, so that the aims of rapid screening and confirmation are fulfilled.
(5) The detection limit and the quantification limit of the embodiment are low, the method is high in sensitivity, stable in recovery rate and good in reproducibility, and the requirements of multi-residue analysis are met.
DETAILED DESCRIPTION OF EMBODIMENT (S) OF INVENTION
The present invention is further illustrated by the following specific examples, which should not be construed as limiting the scope of the invention.
The method is adopted to measure the contents of the 15 additives in the livestock and poultry meat products of sausage and sauced beef and screen suspicious peaks.
1. Instruments and reagents
The main apparatus is as follows: ACQUITYTMUltra high performance liquid chromatography (Waters corporation, usa); AB Sciex QTrap 5500 quadrupole-ion trap mass spectrometer (AB, USA).
The main reagents are as follows: saccharin sodium, acesulfame potassium, sodium cyclamate, aspartame, sorbic acid, benzoic acid, dehydroacetic acid, ethyl naphthol, and salicylic acid, the purity of which is more than or equal to 98.0 percent, and the saccharin sodium, the acesulfame potassium, the sodium dehydroacetate, the ethyl naphthol, and the salicylic acid are purchased from Dr.Ehrenstorfer, Germany; alitame and neotame, with purity of more than or equal to 98.0%, purchased from Shanghai Anpu company; methyl p-hydroxybenzoate, ethyl p-hydroxybenzoate, propyl p-hydroxybenzoate and butyl p-hydroxybenzoate, the purity of which is not less than 98.0%, and the compounds are purchased from Dima corporation.
2. Preparing a standard solution:
preparation of standard stock solutions: weighing appropriate amount of each standard substance, preparing standard stock solution with water to about 10 mug/mL, and storing in a refrigerator at 0-4 deg.C for use.
Preparing a substrate matching standard working solution: to eliminate the influence of matrix effects on the quantitation, the mixed standard stock solution was diluted with the blank sample solution. Processing a blank sample according to the same pretreatment mode, diluting the mixed standard stock solution by using the blank sample matrix solution, fixing the volume to obtain the blank sample matrix solution, and preparing series matching standard working solutions with the concentrations of 0.05 mu g/mL, 0.2 mu g/mL, 0.5 mu g/mL, 1.0 mu g/mL, 2.0 mu g/mL and 5.0 mu g/mL.
3. Chromatographic mass spectrometry conditions:
chromatographic analysis conditions:
Waters BEH C18a reverse phase chromatography column; mobile phase A: 0.02mol/L ammonium acetate aqueous solution, B: methanol; gradient elution was used, flow rate 0.2mL/min, gradient elution procedure: 0min, 90% A; 1min, 90% A; 5min, 10% A; 7min10% of A; 7.1min, 90% A; 12min, 90% A; the column temperature is 40 ℃; the injection volume is 10. mu.L.
Mass spectrometry conditions:
an ion source: an electrospray ion source; scanning mode: scanning negative ions; an acquisition mode: multiple Reaction Monitoring (MRM); electrospray voltage (IS): -4500V; atomization gas pressure (GS 1): 50 psi; air curtain pressure (CUR): 30 psi; assist gas pressure (GS 2): 50 psi; ion source Temperature (TEM): 500 ℃; inlet voltage (EP): -10V; collision cell exit voltage (CXP): -10V. The retention times and mass spectrometric acquisition parameters for the 15 additives are shown in Table 2.
4. The sample pretreatment method comprises the following steps:
sample extraction: accurately weighing 5.00g (accurate to 0.01g) of sample, placing in a 100mL centrifuge tube, adding 20mL of ethanol water solution with the volume concentration of 50% of the extract, shaking for 20min, centrifuging at 8000r/min for 5min, collecting the supernatant in another centrifuge tube, adding 20mL of ethanol water solution with the volume concentration of 50% of the extract, and repeatedly extracting once to obtain the extract.
Sample purification: passing the extractive solution through neutral alumina-filled glass column, collecting 20mL solution, rotary evaporating at 40 deg.C to near dryness, adding 1.0mL ethanol-water (volume ratio of 2:8) solution to dissolve the residue, and filtering the sample solution with 0.22 μm filter membrane.
5. Quantitative analysis and method effectiveness evaluation:
linear, detection limit and quantitation limit: and (3) measuring the substrate matching standard solution with the series of concentrations, and drawing a standard curve by taking the concentration as an abscissa and taking the peak area of the quantitative ions as an ordinate. Then, the sample to be tested is measured to obtain the peak areas of the quantitative ions of various additives, and the peak areas are substituted into the standard curve to calculate the content of 15 additives. And adding standard solutions with different masses into the blank sample, and respectively calculating the detection limit and the quantification limit of the method by using a signal-to-noise ratio of 3 times and a signal-to-noise ratio of 10 times. The detection limit of the 15 additives is between 0.002 and 0.06mg/kg, the quantitative limit is between 0.006 and 0.18mg/kg, and the linear regression equation, the detection limit and the quantitative limit of each compound are shown in a table 3.
TABLE 3 Linear regression equation, correlation coefficient r, detection limit, quantitation limit for additives
Accuracy and precision: and performing standard addition recovery experiments at three concentration levels of 0.2, 0.4 and 2.0mg/kg, performing 6 parallel samples at each concentration level, respectively performing sample pretreatment and instrument analysis, and calculating the recovery rate according to the standard addition amount and the measured value. The average recovery and Relative Standard Deviation (RSD) of the 15 additives in the livestock and poultry meat products are shown in Table 4. As can be seen from Table 4, the average recovery rate of the 15 additives in the livestock and poultry meat products of sausage and sauced beef is 75.2-94.8%, and the relative standard deviation is 4.8-17.3%, so that the requirements of multi-residue analysis are met.
Table 4 recovery and relative standard deviation (n ═ 6) at different addition levels in the samples of meat products from livestock and poultry
6. Example of qualitative screening analysis of suspicious peaks
And (3) carrying out high-flux unknown rapid screening by adopting Analyst software. And detecting an actual sample, finding a suspected target object in an MRM scanning mode, and performing compound confirmation by comparing the detection result of the actual sample with information such as fragments, abundance ratio, retention time and the like under three energies in a database.
The mass spectrum conditions are as follows: an ion source: an electrospray ion source; scanning mode: scanning negative ions; an acquisition mode: multiple Reaction Monitoring (MRM); information association scan (IDA): intensity threshold 100 cps; enhancer ion scanning (EPI), mass number acquisition range m/z 50-1000 Da; electrospray voltage (IS): -4500V; atomization gas pressure (GS 1): 50 psi; air curtain pressure (CUR): 30 psi; assist gas pressure (GS 2): 50 psi; ion source Temperature (TEM): 500 ℃; inlet voltage (EP): -10V; collision cell exit voltage (CXP): -10V; collision gas energy (CE): 20. 35, 50V; extended Collision Energy (CES): 15V.
Selecting MS2The spectrogram and maximize the graph, right click on the mouse, click on Search Library, and the software automatically gives the result of the screening. The Fit value (matching value) is a similarity value obtained by comparing a standard substance spectrogram with a suspicious sample spectrogram, the full score is 100, and the higher the Fit value is, the higher the reliability is, the higher the possibility of indicating the compound is; the Revfit value (inverse matching value) is a similarity value obtained by comparing a suspicious sample spectrogram with a standard substance spectrogram; the Purity value (Purity value) is a value obtained by combining the first two results. FIG. 1 is a graph showing MS of a suspicious peak in a sample2Spectrogram, FIG. 2 is MS of rhodamine B standard2The spectrogram, as apparent from table 5, shows that rhodamine B (rhodamine B) in the library has high score and is arranged most forward, and the characteristic fragment ions are all matched with the sample, so that the sample can be judged to contain rhodamine B, the confidence of result confirmation is further improved, and the generation of false positive results is avoided.
Table 5.
Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. This application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims.
It will be understood that the invention is not limited to the precise arrangements described above and shown in the drawings and that various modifications and changes may be made without departing from the scope thereof. The scope of the invention is limited only by the appended claims.
Claims (2)
1. A method for detecting additives in livestock and poultry meat products is characterized by comprising the following steps:
1) sample pretreatment:
weighing a sample, placing the sample in a centrifuge tube, adding 50% ethanol water solution by volume concentration, performing oscillation extraction and centrifugation, collecting supernatant in another centrifuge tube, adding 50% ethanol water solution by volume concentration, and repeatedly extracting once to obtain an extracting solution;
purifying the extract by a neutral alumina column, concentrating, and fixing volume for later use;
2) and (3) qualitative analysis:
screening the livestock and poultry meat product sample to be detected after sample pretreatment by using an UPLC-MS/MS-Qtrap method, establishing an MRM-IDA-EPI screening method by adopting an enhanced ion scanning function of a liquid chromatography-tandem quadrupole linear ion trap mass spectrometer, and obtaining MS of each target additive parent ion in the livestock and poultry meat product sample to be detected2Spectra, using standard MS for each additive in a known database2Spectrogram and MS for obtaining target additive parent ions in livestock and poultry meat product sample to be detected2Respectively comparing the spectrograms to obtain similarity values, matching characteristic fragment ions of the spectrograms and the characteristic fragment ions of the spectrograms, and further judging whether the livestock and poultry meat product sample to be detected contains the target additive or not;
3) quantitative analysis:
weighing each additive standard substance respectively, and preparing 10 mug/mL standard stock solution with water respectively for later use;
processing a blank sample according to the sample pretreatment method in the step 1), and diluting the standard stock solution by using a blank sample matrix solution to prepare a mixed standard stock solution;
after the mixed standard stock solution is subjected to constant volume, obtaining a blank sample matrix solution, and preparing a series of matching standard working solutions with the concentration of 0.05 mu g/mL-5.0 mu g/mL;
respectively measuring a series of matching standard working solutions and a livestock and poultry meat product sample to be detected after sample pretreatment by adopting a liquid chromatogram-series quadrupole linear ion trap mass spectrometer to obtain a drawn standard curve with the concentration as a horizontal coordinate and the peak area of quantitative ions as a vertical coordinate, and calculating the content of the target additive in the livestock and poultry meat product sample to be detected by using the standard curve;
the additive comprises: sodium saccharin, acesulfame potassium, sodium cyclamate, aspartame, sorbic acid, benzoic acid, dehydroacetic acid, ethyl naphthol, salicylic acid, alitame, neotame, methyl paraben, ethyl paraben, propyl paraben, and butyl paraben;
the pretreatment of the sample in the step 1) is specifically as follows:
weighing 5.00g of sample, placing the sample in a 100mL centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, shaking for extraction for 20min, centrifuging for 5min at 8000r/min, collecting supernatant in another centrifuge tube, adding 20mL of 50% ethanol water solution in volume concentration, and repeatedly extracting once to obtain an extracting solution;
passing the extractive solution through neutral alumina-filled glass column, collecting 20mL solution, rotary evaporating at 40 deg.C to near dryness, adding 1.0mL ethanol water solution with volume solubility of 20% to dissolve residue, and passing through 0.22 μm filter membrane;
the mass spectrum conditions of the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the step 2) are as follows:
an ion source: an electrospray ion source;
scanning mode: scanning negative ions;
an acquisition mode: monitoring multiple reactions;
information association scanning: intensity threshold 100 cps;
enhancer ion scanning: the mass number acquisition range is m/z 50-1000 Da;
electrospray voltage: -4500V;
atomizing gas pressure: 50 psi;
air curtain pressure: 30 psi;
auxiliary gas pressure: 50 psi;
ion source temperature: 500 ℃;
inlet voltage: -10V;
collision cell exit voltage: -10V;
energy of collision gas: 20. 35, 50V;
expansion of collision energy: 15V;
the chromatographic conditions of the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the step 3) are as follows:
a chromatographic column: waters BEH C18A reverse phase chromatography column;
column temperature: 40 ℃;
sample introduction volume: 10 mu L of the solution;
flow rate: 0.2 mL/min;
mobile phase: mobile phase A: 0.02mol/L ammonium acetate aqueous solution, B: methanol, gradient elution procedure: 0min, 90% A; 1min, 90% A; 5min, 10% A; 7min, 10% A; 7.1min, 90% A; 12min, 90% A;
the mass spectrum conditions of the liquid chromatogram-tandem quadrupole linear ion trap mass spectrometer in the step 3) are as follows:
an ion source: an electrospray ion source;
scanning mode: scanning negative ions;
an acquisition mode: monitoring multiple reactions;
electrospray voltage: -4500V;
atomizing gas pressure: 50 psi;
air curtain pressure: 30 psi;
auxiliary gas pressure: 50 psi;
ion source temperature: 500 ℃;
inlet voltage: -10V;
collision cell exit voltage: -10V.
2. The method for detecting additives in livestock and poultry meat product according to claim 1, wherein the livestock and poultry meat product sample to be detected is beef with paste or sausage.
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| CN109655567A (en) * | 2018-12-25 | 2019-04-19 | 沈阳出入境检验检疫局检验检疫综合技术中心 | A kind of method of acid stain in detection animal meat products |
| CN109765325A (en) * | 2018-12-28 | 2019-05-17 | 沈阳出入境检验检疫局检验检疫综合技术中心 | A method of detection animal meat products neutral and alkali colorant |
| CN110632194A (en) * | 2019-08-27 | 2019-12-31 | 郑州科技学院 | Method for determining aspartame and alitame in food by reversed phase liquid chromatography mixed standard sample adding method |
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| CN107037150A (en) * | 2017-04-11 | 2017-08-11 | 安徽省公众检验研究院有限公司 | It is a kind of at the same determine food in Determination of Preservatives, sweetener and colouring agent method |
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| CN106645603A (en) * | 2017-01-13 | 2017-05-10 | 广西民族大学 | Method for synchronously detecting content of food additives and heavy metals in food in inverse HPLC-ICP-MS method |
| CN107037150A (en) * | 2017-04-11 | 2017-08-11 | 安徽省公众检验研究院有限公司 | It is a kind of at the same determine food in Determination of Preservatives, sweetener and colouring agent method |
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