CN112268967A - Method for testing content of inorganic arsenic in food - Google Patents
Method for testing content of inorganic arsenic in food Download PDFInfo
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Abstract
The invention relates to a method for testing inorganic arsenic content in food, which comprises the following steps: (1) sample pretreatment: treating the sample with an oxidizing acid solution, extracting by a hot-extraction method, centrifuging, taking supernatant, fixing the volume, and testing on a machine; (2) and (3) chromatographic column separation: separating the extract by gradient elution with anion exchange chromatographic column as analytical column and phosphate as mobile phase; (3) and detecting and analyzing various forms of arsenic in the sample by ICP-MS. The invention adopts an extraction system of nitric acid-hydrogen peroxide, optimizes the concentration of nitric acid and hydrogen peroxide, namely fully extracts various arsenic forms in a sample, completes the conversion of As (III) to As (V), does not oxidize organic arsenic, avoids the interference of the organic arsenic to inorganic arsenic during sample measurement, particularly the interference of high-concentration arsenic betaine to the inorganic arsenic, and is beneficial supplement for the detection of the inorganic arsenic in the existing food.
Description
Technical Field
The invention belongs to the field of analytical chemistry, and particularly relates to a method for testing inorganic arsenic content, and more particularly relates to a method for testing inorganic arsenic content in animal seafood.
Background
Arsenic is a common toxic element, but its toxicity depends mainly on its chemical form. The inorganic form of arsenic is highly carcinogenic, and recent epidemiological studies have found that inorganic arsenic exposure is associated with heart disease and diabetes. The semi-lethal doses of arsenic compounds were, in order: arsenite (As)3+)>Arsenate (As)5+)>Methyl arsenic acid (MMA)>Dimethyl arsenic acid (V). The world health organization, australia/new zealand, china and the european union have established a limited standard for the content of inorganic arsenic in rice, fish, aquatic animals and algae.
Seafood is an important source of nutrients and an important component of a healthy, balanced diet. Over 10 million people worldwide use seafood as the primary source of animal protein. While seafood has some benefits to human nutrition, it can accumulate a large amount of potentially toxic elements (such as arsenic) in tissues, which can pose a risk to human health. Arsenic is ingested from aquatic products, vegetables, seaweeds and rice in an amount of about 96% of the total arsenic intake.
The arsenic form in fish seafood is mainly arsenic betaine (AsB), the arsenic form in shellfish seafood is mainly arsenic betaine and arsenic sugar, which are considered non-toxic, and the presence of arsenic in seafood is almost harmless to human health; the limit amounts of inorganic arsenic in aquatic animals and products thereof (except fishes and products thereof), fishes and products thereof, infant canned auxiliary food (products taking aquatic products and animal livers as raw materials) are respectively 0.5mg/kg, 0.1mg/kg and 0.3mg/kg according to the regulations of GB2762-2017 limit amounts of pollutants in national food safety standards. When an anion exchange chromatography column is used As the analysis column, the low arsenite (As) content is affected because the tailing of high-concentration arsenobetaine in marine products tends to be generated in the chromatography column relative to low-concentration arsenite (As3+)3+) The accurate determination of the content of arsenic sugar in the shellfish sample, and the arsenic acid salt (As)3 +) Close or identical results in the appearance of false positives. Therefore, in discussing the toxicity of arsenic in such food products, the amount of inorganic arsenic is an important consideration. National food safety Standard applied on 21/09/2015 and 21/03/2016The determination of total arsenic and inorganic arsenic in food (GB 5009.11-2014) has problems in determining inorganic arsenic in marine animal products and canned supplementary food for infants, and measurement deviation is often caused.
The prior art has matured the test development aiming at different forms of arsenic, and the test development comprises high performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP-MS), Atomic Fluorescence Spectrometry (AFS), Atomic Absorption Spectrometry (AAS), a high performance liquid chromatography and hydride generation-atomic absorption combined technology (HPLC-HG-ASS), a capillary electrophoresis and inductively coupled plasma mass spectrometry combined technology (CE-ICP-MS) and the like, wherein the high performance liquid chromatography-inductively coupled plasma mass spectrometry has the advantages of high sensitivity, low detection limit, wide linear range and the like, and is widely applied to morphological research of foods and environments. However, one problem that has been difficult to overcome in measuring arsenic in different forms in aquatic products or canned auxiliary foods and other samples is that the interference of organic arsenic on inorganic arsenic and the conversion of organic arsenic to inorganic arsenic during the test process lead to inaccurate measurement. Therefore, it is needed to develop a method for accurately and rapidly testing and evaluating inorganic arsenic in food, which can avoid the interference of organic arsenic, and provide technical support for food safety and healthy diet.
Disclosure of Invention
In order to overcome the defect of inaccurate inorganic arsenic content test in food, particularly animal aquatic products or infant canned auxiliary food in the prior art, the method can prevent the organic arsenic form from being converted into the inorganic arsenic form in the sample pretreatment process, and simultaneously, the inorganic arsenic and the organic arsenic can be fully separated by chromatographic separation, so that the interference of the organic arsenic is avoided. The method is rapid and accurate in detection, and is a method for detecting inorganic arsenic in food samples which is expected to be upgraded to a national standard method. The method adopts an oxidation hot leaching method to extract inorganic arsenic in the sample, fully oxidizes arsenite into arsenate, and simultaneously keeps the form of organic arsenic unchanged. Solves the problem that high-content arsenic betaine and arsenic sugar interfere with the determination of arsenite, and can accurately determine inorganic arsenic in food, especially in animal marine products and infant canned auxiliary food.
In order to solve the technical problem, the invention provides a method for testing the content of inorganic arsenic in food, which comprises the following steps:
(1) sample pretreatment: treating the sample with an oxidizing acid solution, extracting by a hot-extraction method, centrifuging, taking supernatant, fixing the volume, and testing on a machine;
(2) and (3) chromatographic column separation: taking an anion exchange chromatographic column as an analytical column, and taking a mobile phase as phosphate, and carrying out gradient elution on the sample extracting solution;
(3) and detecting and analyzing various forms of arsenic in the sample by ICP-MS.
Further, the oxidizing acid solution in the step (1) is HNO3-H2O2An aqueous solution of (a). Preferably, the concentration of nitric acid is 0.1-0.2mol/L, H2O2The concentration is 0.3-0.6 wt%; more preferably, the concentration of nitric acid is 0.15 to 0.18mol/L, H2O2The concentration is 0.4-0.5 wt%.
At present, in arsenic morphological analysis, the main pretreatment methods are water, water-acetic acid, nitric acid-water, methanol-water, phosphoric acid-water, acetonitrile-water, acetic acid-water, hydrochloric acid-water and other systems to extract samples. The invention adopts HNO3-H2O2The compound oxidizing acid solution is used for extracting the sample, the extraction efficiency is high, the organic arsenic cannot be changed to further interfere the test result, the easily interfered As (III) is oxidized into As (V), and the content of the inorganic arsenic in the sample can be obtained by testing the amount of the As (V) in the final sample.
Furthermore, the oxidizing acid solution in the step (1) also contains 0.01-0.02mol/L of methanesulfonic acid. A certain amount of methanesulfonic acid is added, so that the oxidation and chemical reaction of organic arsenic can be inhibited in the process of oxidizing As (III) into As (V), the interference of the organic arsenic on inorganic arsenic can be avoided to a greater extent, and the accuracy of detecting the inorganic arsenic is further improved.
Further, the mobile phase A and the mobile phase B in the step (2) are 8-25mmol/L (NH)4)2HPO4Provided that in phase B (NH)4)2HPO4The concentration is higher than that of the A phase; preferably, phase B and phase A (NH)4)2HPO4The concentration ratio of (A) to (B) is 1.5-3:1, preferably 1.8-2.5: 1.
In one embodiment of the invention, phase A (NH)4)2HPO4Concentration of 8-15mmol/L, B phase (NH)4)2HPO4The concentration is 18-25 mmol/L.
Further, the mobile phase A and B contain 1-5% of methanol, preferably 1-2% of methanol.
The inventors have found that methanol is used to enhance the signal intensity of the arsenic form in the HPLC-ICP-MS technique. But the concentration of the methanol is not easy to be too high, on one hand, the concentration of the methanol is continuously increased, the signal intensity enters a plateau stage, and the improvement is not obvious; on the other hand, methanol entering the ICP can be converted into carbon elements, and the carbon elements are deposited and enriched on the sampling cone and the interception cone so as to influence the measurement result.
In a preferred embodiment of the invention, the phase B also contains 1-2mmol/L sodium pyrophosphate. The inventor unexpectedly finds that the separation effect of As (V) and organic arsenic can be enhanced by adding a small amount of sodium pyrophosphate into the B phase, the As (V) peak-out time is shortened, the peak shape is narrowed, and the sensitivity is improved.
In another preferred embodiment of the invention, the pH of phase A and phase B to the mobile phase is in the range of 7 to 9, preferably 7.2 to 8, most preferably 7.5 to 7.6.
In another preferred embodiment of the invention, the flow rate of the mobile phase is 1-1.5mL/min, preferably 1-1.2 mL/min.
Preferably, in the step (2), the gradient elution means that the A phase is 80-100% and the B phase is 0-20% in 0-5 min; in 5-11min, the phase A is 0% -20%, and the phase B is 80-100%; after 11min, the phase A is 80-100%, and the phase B is 0-20%; preferably, within 0-5min, the phase A is 90-100%, and the phase B is 0-10%; in 5-11min, the phase A is 0% -10%, and the phase B is 90-100%; after 11min, the phase A is 90-100% and the phase B is 0-10%.
Further, the anion exchange chromatography column in step (2) includes, but is not limited to, Dionex IonPac AS7, Dionex IonPac AS14, Dionex IonPac AS19, Hamilton PRP-X100, and the like. Preferably Hamilton PRP-X100, the chromatographic column is relatively stable in a wide pH value range and strong in universality, and the chromatographic column is combined with phosphate flow to analyze the arsenic form successfully.
Through the optimized chromatographic column conditions, the method can finish the separation of arsenic in different forms in the sample in a short time (15min), and in aquatic products with high organic arsenic content, As (V) and various organic arsenic are separated effectively without interference.
The food sample of the present invention is not particularly limited, and examples thereof include meat, eggs, drinks, health products, medicinal materials, dairy products, and the like. However, the method disclosed by the invention fully eliminates the interference of organic arsenic, and is particularly suitable for detecting samples which are required to measure inorganic arsenic in food with high content of organic arsenic (particularly arsenic sugar and arsenic betaine), such as animal aquatic products, or some foods taking animal livers and viscera as raw materials, such as food pollutant limit of current national food safety standards (GB2762-2017) such as infant supplementary food and the like.
Compared with the prior art, the invention has the following beneficial effects:
firstly, the invention adopts an extraction system of nitric acid-hydrogen peroxide, and the concentration of nitric acid and hydrogen peroxide is optimized, namely, various arsenic in the sample is fully extracted, the conversion from AS (III) to AS (V) is completed, organic arsenic is not oxidized, and the interference of organic arsenic in the sample on inorganic arsenic is avoided.
In the chromatographic column, proper mobile phase conditions and gradient elution conditions are optimized, so that various forms of arsenic in the sample can be effectively separated quickly, and the interference of organic arsenic, especially high-concentration arsenic betaine, on inorganic arsenic is avoided.
The invention can detect various arsenic form contents, especially inorganic arsenic, in food, especially animal aquatic products rich in organic arsenic and infant canning auxiliary food thereof, can effectively avoid the interference of the organic arsenic to the inorganic arsenic in the test process, accurately judge the content of the inorganic arsenic in the sample, and avoid the false positive condition caused by the organic arsenic. Is a beneficial supplement for detecting arsenic in the existing food.
Drawings
FIG. 1 shows the effect of methanol concentration on the signal intensity of 5 arsenic morphological compounds in a mobile phase.
FIG. 2 is a chromatogram (10. mu.g/L) of a mixed standard solution of 5 arsenic forms.
FIG. 3 is a comparison chromatogram of background and labeled blue willow clam.
FIG. 4 is a cod background and spiked comparison chromatogram.
FIG. 5 is a graph showing the effect of hydrogen peroxide addition on inorganic arsenic spiking recovery (5. mu.g/L spiking).
FIG. 6 shows the effect of hydrogen peroxide addition on inorganic arsenic spiking recovery (100. mu.g/L spiking).
Detailed Description
The following examples are provided to further illustrate the present invention for detecting inorganic arsenic in food such as aquatic products, but it should be understood that the contents of the examples are for better understanding of the present invention, and are not intended to limit the scope of the present invention.
Laboratory apparatus and reagent
Model 1260 high performance liquid chromatograph and model 7700x inductively coupled plasma mass spectrometer (Agilent, usa); hamilton PRP-X100 anion exchange column (250 mm. times.4.1 mm, 10 μm) and its guard column (Hamilton, USA); milliplus2150 ultrapure water treatment system (millirobo, usa); drying ovens (MMM group products, germany); high speed centrifuges (Beckman Corp., USA).
Ultrapure water: resistivity of 18.2 M.OMEGA.cm; ammonia (NH)4·H2O) and hydrogen peroxide (H)2O230%) are premium grade pure and purchased from chemical reagents of national drug group, ltd; diammonium hydrogen phosphate [ (NH)4)2HPO4](analytical purity, chemical reagents of national drug group, Ltd.); methanol (chromatographically pure, Sigma company, usa); nitric acid (purity 65%, Merck, Germany). Arsenite (As)3+) Arsenate (As)5+) Arsenic standards such as monomethylarsenic acid (MMA), dimethylarsenic acid (DMA) and arsenobetaine (AsB) were purchased from the national institute of metrology and science.
Solution preparation
1. Arsenic form standard stock (10.0mg/L, As): accurately absorbing a certain amount of arsenite radical, arsenate radical, monomethyl arsenic, dimethyl arsenic and arsenic betaine solution standard substances respectively, and diluting the standard substances into single standard stock solution of 10.0mg/L by using a proper amount of water. It can be stored at 4 deg.C for 6 months.
2. Arsenic form mixed standard intermediate (1.00mg/L, As): accurately sucking 1.0mL of arsenite standard stock solution, arsenate standard stock solution, monomethylarsenic standard stock solution, dimethylarsine standard stock solution and arsenic betaine standard stock solution respectively into a 10mL volumetric flask, adding water for dilution, and fixing the volume to the scale. It is prepared as before use.
3. Arsenic form mixed standard series solution: accurately sucking 0.00mL, 0.01mL, 0.05mL, 0.10mL, 0.20mL, 0.50mL and 1.00mL of arsenic form mixed standard intermediate solution (1.00mg/L) into a 10mL volumetric flask, and diluting with water to a constant volume. The concentrations of arsenic forms in the standard series of solutions were 0.0. mu.g/L, 1.0. mu.g/L, 5.0. mu.g/L, 10.0. mu.g/L, 20.0. mu.g/L, 50.0. mu.g/L, and 100. mu.g/L, respectively. It is prepared as before use.
Note: and (4) properly adjusting the mass concentration of each arsenic form in the standard series solution according to the concentration of each arsenic form in the actual sample, and re-measuring the arsenic form concentration in the sample after dilution if the arsenic form concentration in the sample exceeds the upper limit of the standard curve.
Example 1
Preparing a solution:
1. nitric acid-hydrogen peroxide extract (0.15mol/L nitric acid + 0.45% hydrogen peroxide): 10mL of nitric acid and 15mL of hydrogen peroxide (30%) were weighed and diluted to 1000mL with water.
2. Mobile phase a (10mmol/L diammonium phosphate + 2% methanol, pH 7.5): accurately weighing 1.320g of diammonium hydrogen phosphate, placing the diammonium hydrogen phosphate in a 1000mL volumetric flask, dissolving the diammonium hydrogen phosphate in water, adding 20mL of methanol, diluting the mixture with water to a constant volume to scale, adjusting the pH value to 8.0 by ammonia water, and uniformly mixing. Filtering with 0.45 μm water system filter membrane, and ultrasonic degassing in ultrasonic water bath for 30 min.
3. Mobile phase B (20mmol/L diammonium phosphate + 2% methanol, pH 7.6): 2.640g of diammonium hydrogen phosphate is accurately weighed, placed in a 1000mL volumetric flask, dissolved in water, added with 20mL of methanol, diluted with water to a constant volume to a scale and mixed evenly. Filtering with 0.45 μm water system filter membrane, and ultrasonic degassing in ultrasonic water bath for 30 min.
First, the sensitizing effect of different concentrations of methanol in the mobile phase was examined, and the mobile phases containing 0%, 1%, 2%, 3%, 4% and 5% of methanol were prepared, and the influence of the methanol concentration on the signal intensity (expressed by the integral area) of 5 arsenic form compounds was examined, and the results are shown in fig. 1. The result shows that after 1% of methanol is added, the signal intensity of the five arsenic forms is increased by 1 time compared with the signal intensity without methanol, and when the addition amount of the methanol is between 2% and 5%, the sensitization effect is not obvious when the arsenic form enters a plateau stage. The methanol introduced into ICP is converted into carbon element, and deposition and enrichment are carried out on the sampling cone and the interception cone so as to influence the determination result, so that methanol with higher concentration cannot be selected as a sensitizer. This example selects 2% methanol as the sensitizer to be added to the mobile phase. Under optimized conditions, chromatograms of 5 arsenic morphologies mixed standard solutions (10. mu.g/L) are shown in FIG. 2, with AsB, As (III), DMA, MMA and As (V) achieving baseline separation within 15 min.
Then, the mobile phase was examined to prepare 10mmol/L (NH)4)2HPO4As the mobile phase, the effect of pH 5 to 9 on the separation effect of As (III), As (V), MMA, DMA and AsB was examined. Experiments show that when the mobile phase is acidic, AsB is overlapped with As (III); AsB was completely separated from As (III) at pH 7.2 to 8, and as (III) and As (III) were gradually shifted backwards with almost no change in the peak times of AsB, DMA and As (V) as the pH increased. As (III) and DMA did not achieve baseline separation when the mobile phase was between 8 and 9. At a mobile phase pH of 7.5, the separation of the 5 arsenic forms is best and the sensitivity is also in a better state.
10mmol/L(NH4)2HPO4When isocratic elution was performed as a mobile phase (pH 7.5), as (v) peaked at about 17.5min, and the analysis time was long, the peak width was broad, and the peak height was short, affecting the sensitivity. In order to shorten the analysis time and improve the sensitivity of As (V), 10mmol/L (NH) is adopted4)2HPO4(pH 7.5) as phase A, 20mmol/L (NH)4)2HPO4(pH 7.6) as B phase. The method can complete the separation of 5 arsenic forms within 15 min.
Example 2
The major arsenic form in willow clam is AsB, and contains small amounts of unknown arsenide 1(U1), DMA, unknown arsenide 2(U2), and As (V). The 5 arsenic form mixed standard solutions of 20 mu g/L are added into the Qingliu clam, and the standard recovery rates of AsB, As (III), DMA, MMA and As (V) are 94.2%, 105.2%, 101.0%, 99.8% and 103.6%, respectively. AsB the recovery rate is relatively low compared with As (III), and the AsB tailing with high concentration leads to a small amount of overlap with As (III) as can be seen from comparison of the chromatogram before and after labeling (figure 3), thereby causing quantitative deviation; if the sample contains a small amount of As (III), the AsB tailing at a high concentration will also interfere with its accurate determination, resulting in the appearance of false negatives. Similarly, arsenic sugar is also present in shellfish samples, and the presence of arsenic sugar also interferes with the determination of As (III), resulting in the appearance of false positives. The predominant arsenic form in cod is AsB, with a small amount of DMA. Similar to the meretrix results, the high concentration of AsB tail affected the accurate determination of as (iii) (fig. 4).
Therefore, in order to solve the problem of interference of high-concentration AsB tailing and arsenic sugar on As (III), the invention adopts hydrogen peroxide to oxidize As (III) into As (V), and the content of As (V) in a sample is measured to represent the content of inorganic arsenic. 5 mu g/L and 100 mu g/L arsenic form mixed standard solutions are respectively added into a sample, and the influence of the addition amount of the hydrogen peroxide on the oxidation effect of the low and high concentration As (III) is studied, and the ratio of the measured value of As (V) to the sum of the addition amounts of As (III) and As (V) is taken as a consideration standard. Respectively preparing 0.15mol/L nitric acid solution containing 0.4-2% hydrogen peroxide (30%), adding the extractive solution into the sample, and hot-extracting at 90 deg.C for 3 hr. The results showed that the recovery of inorganic arsenic was highest (more than 90%) when the amount of hydrogen peroxide in the extract was 0.45%, while the chromatogram showed that all of As (III) was converted to As (V).
In order to ensure that only As (III) is oxidized in the extraction process, single standard solutions of AsB, DMA and MMA, which are 10 mu g/L and 100 mu g/L, are respectively prepared by using 0.15mol/L nitric acid-0.45% hydrogen peroxide extracting solution, the single standard solutions are treated by using the same extraction temperature and time, the influence on the form of the organic arsenic in the extraction process is examined, and the result is shown in Table 1. Results show that the standard recovery rate ranges of the low-concentration AsB, the DMA and the MMA are respectively 99.8-102.3% and 97.6-101.2%; meanwhile, As (III) is not detected, and a small amount of As (V) is shown, which indicates that the organic arsenic exists stably in the pretreatment process. However, the recovery of spiked standards at high concentrations of AsB, DMA and MMA was still slightly insufficient.
TABLE 1 ASB, DMA and MMA Single Standard solution plus Standard Oxidation test (%)
In order to further suppress the conversion of organic arsenic into inorganic arsenic during oxidation, the inventors have unexpectedly found that a small amount of methanesulfonic acid, specifically, 0.15mol/L nitric acid + 0.45% hydrogen peroxide +0.01mol/L methanesulfonic acid, was further added to the extract solution, and the effect on the form of organic arsenic was examined in the same manner as described above, and the results are shown in Table 2. Results show that the standard recovery rate ranges of the low-concentration AsB, the DMA and the MMA are respectively 100.2-102.6% and 100.1-104.0%; meanwhile, As (III) and As (V) are not detected, which indicates that the organic arsenic exists stably in the form and inorganic arsenic does not appear in the pretreatment process. Recovery was satisfactory at high concentrations of AsB, DMA and MMA.
TABLE 2 ASB, DMA and MMA Single Standard solution plus Standard Oxidation test (%)
In conclusion, further experiments were carried out by using a mixed acid solution of 0.15mol/L nitric acid, 0.45% hydrogen peroxide and 0.01mol/L methanesulfonic acid and carrying out hot leaching at 90 ℃ for 3 hours.
Example 3
Sample pretreatment:
accurately weighing about 1g (accurate to 0.001g) of cod sample into a 50mL polypropylene centrifuge tube, adding 20mL of a mixed solution of 1% nitric acid, 0.45% hydrogen peroxide and 0.01mol/L methanesulfonic acid, hot-extracting at 90 ℃ for 3h, and shaking once every 30 min. After extraction, taking out and cooling to room temperature, centrifuging for 10min at 8000r/min, taking supernatant, and filtering with 0.22 μm water system filter membrane while performing blank experiment.
ICP-MS conditions：
RF incident power: 1550W, carrier gas: high purity argon, carrier gas flow rate: 0.65L/min, make-up airflow rate: 0.45L/min, collision gas flow rate (He): 4.5mL/min, radio frequency voltage: 1.70V, sampling depth: 8.0mm, peristaltic pump speed: 0.3 r/s.
Chromatographic conditions：
A chromatographic column: hamilton PRP-X100 anion exchange column (250 mm. times.4.1 mm, 10 μm); mobile phase: phase A is 10mmol/L (NH)4)2HPO4(containing 2% methanol, pH 7.5) and 20mmol/L (NH) of phase B4)2HPO4(2% methanol at pH 7.5) gradient elution procedure as shown in table 3; data acquisition time: 15 min; sample introduction volume: 25 μ L.
TABLE 3 gradient elution procedure
Linear range and detection limit:
5 kinds of arsenic form mixed standard solutions of 0.5, 1.0, 5.0, 10, 20, 50 and 100 mug/L are prepared respectively, and under optimized experimental conditions, the linear range and the detection limit are examined, and the results are shown in Table 4. Within the concentration range of 0.5-100 mug/L, the correlation coefficient r is larger than 0.999, which indicates that 5 arsenic forms meet the linear requirement.
The signal-to-noise ratios (S/N) of 5 arsenic species were measured at different concentrations by the stepwise dilution method, and the solution concentrations were found to be detection limits at S/N of 3, and the detection limits of AsB, As (III), DMA, MMA and As (V) were found to be 0.05, 0.1 and 0.1. mu.g/L, respectively.
TABLE 4 method Linear Range and detection limits
Method precision and spiking recovery test
Selecting a cod sample with high concentration of AsB, performing precision and standard addition recovery tests by adopting an optimized separation test method and a sample pretreatment method, adding 5 arsenic form mixed standard solutions with three concentration levels, preparing 6 samples in parallel to determine arsenic form content, and calculating precision and standard addition recovery. The results are shown in Table 5, where DMA, MMA and inorganic arsenic were measured, and the recovery and RSD were all at satisfactory levels.
TABLE 5 cod sample precision and spiking recovery test results
Example 4
Other conditions and operations were the same as those in example 3 except thatChromatographic conditionsIn (2), the mobile phase: phase A is 10mmol/L (NH)4)2HPO4(containing 2% methanol, pH 7.5) and 18mmol/L (NH) of phase B4)2HPO4+1mmol/LNa4P2O7(containing 2% methanol, pH 7.5). Namely, compared with the example 3, 1mmol/L sodium pyrophosphate is additionally added in the B phase of the mobile phase as an auxiliary reagent. The detection of various forms of arsenic was carried out on the same batch of high-concentration cod samples according to the same method, and the results are shown in table 6 below:
TABLE 6 cod sample precision and spiking recovery test results
From a comparison of example 3 and example 4, it can be seen that a small amount of Na was added to the B phase of the mobile phase4P2O7The method can be more beneficial to the separation of inorganic arsenic and organic arsenic in the chromatogram, the detection accuracy is improved, the recovery rate and the RSD% are improved, and the improvement is more obvious particularly for the detection of the inorganic arsenic.
Example 5
Other conditions and operations were the same as those in example 3 except thatChromatographic conditionsIn (b), the mobile phase is: phase A is 10mmol/L (NH)4)2HPO4(containing 2% methanol, pH 8.0), phase B18 mmol/L (NH)4)2HPO4(containing 2% methanol, pH 8.0). The pH of the mobile phase was changed from 7.5 to 8.0, and the same batch of high-concentration cod samples were tested for arsenic in each form under otherwise unchanged conditions, with the results shown in table 7 below:
TABLE 7 cod sample precision and spiking recovery test results
Example 6
Other conditions and operations were the same as those in example 3 except thatChromatographic conditionsIn, the flow phase: phase A is 10mmol/L (NH)4)2HPO4(containing 2% methanol, pH 7.2) and 18mmol/L (NH) of phase B4)2HPO4(containing 2% methanol, pH 7.2). The pH of the mobile phase was changed from 7.5 to 7.2, and the same batch of high-concentration cod samples were tested for arsenic in each form under otherwise unchanged conditions, with the results shown in table 8 below:
TABLE 8 cod sample precision and spiking recovery test results
The invention establishes the HPLC-ICP-MS method for measuring inorganic arsenic in animal marine products and infant canned supplementary food thereof by optimizing sample pretreatment conditions, liquid chromatography conditions and inductively coupled plasma mass spectrometry conditions. The method has the advantages of simple pretreatment, wide linear range, low detection limit, high sensitivity, accurate and reliable result and the like.
Claims (10)
1. A method for testing the content of inorganic arsenic in food comprises the following steps:
(1) sample pretreatment: treating the sample with an oxidizing acid solution, extracting by a hot-extraction method, centrifuging, taking supernatant, fixing the volume, and testing on a machine;
(2) and (3) chromatographic column separation: taking an anion exchange chromatographic column as an analytical column, and taking a mobile phase as phosphate, and carrying out gradient elution on the sample extracting solution;
(3) and detecting and analyzing various forms of arsenic in the sample by ICP-MS.
2. The test method of claim 1, wherein the oxidizing acid solution in step (1) is HNO3-H2O2An aqueous solution of (a).
3. The test method of claim 2, wherein HNO3The concentration of (A) is 0.1-0.2mol/L, H2O2The concentration is 0.3-0.6 wt%; preferably, HNO3The concentration of (A) is 0.15-0.18mol/L, H2O2The concentration is 0.4-0.5 wt%.
4. The test method according to any one of claims 1 to 3, wherein the oxidizing acid solution in the step (1) further contains 0.01 to 0.02mol/L of methanesulfonic acid.
5. The method of claim 1, wherein the mobile phases of phase A and phase B in step (2) are 8-25mmol/L (NH)4)2HPO4Provided that in phase B (NH)4)2HPO4Higher concentration than phase a.
6. The test method according to claim 5, wherein the phases B and A (NH) are4)2HPO4The concentration ratio of (A) to (B) is 1.5-3:1, preferably 1.8-2.5: 1; more preferablyPhase A (NH)4)2HPO4Concentration of 8-15mmol/L, B phase (NH)4)2HPO4The concentration is 18-25 mmol/L.
7. The test method according to claim 5, wherein the mobile phases A and B contain 1 to 5 wt.% of methanol, preferably 1 to 2 wt.% of methanol.
8. The test method according to any one of claims 5 to 7, wherein the B phase of the mobile phase further contains 1 to 2mmol/L of sodium pyrophosphate.
9. The test method according to any one of claims 5 to 8, wherein the mobile phase has a pH of 7 to 9 for phase A and phase B, preferably 7.2 to 8, most preferably 7.5 to 7.6.
10. The test method according to any one of claims 1 to 9, wherein the food is a food with a high content of organic arsenic (arsenic sugar, arsenic betaine), such as an animal aquatic product, or an infant supplementary food with animal liver and kidney as raw materials.
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