GB/T 5009.17-2003 Determination of total mercury and organic mercury in food
Some standard content:
ICS 67.040
National Standard of the People's Republic of China
GB/T 5009.17—2003
Replaces GB/T5009.17--1996, partially replaces GB/T5009.45--1996 Determination of total mercury and organic-mercury in foods
Determination of total mercury and organic-mercury in foodsPublished on August 11, 2003
Ministry of Health of the People's Republic of China
Standardization Administration of the People's Republic of China
Implemented on January 1, 2004
GB/T 5009.17-2003
This standard replaces 4.6 methylmercury in GB/T5009.17-1996 "Determination of total mercury in food" and GB/T5009.45-1996 "Analytical method for hygienic standards of aquatic products". Compared with GB/T5009.17-1996 and GB/T5009.45-1996 "Analysis Methods for Hygienic Standards of Aquatic Products", this standard has the following major revisions:
The Chinese name of the standard has been revised, and the Chinese name of the standard has been changed to "Determination of Total Mercury and Organic Mercury in Food"; - The structure of the original standard has been revised in accordance with GB/T20001.42001 "Standard Preparation Rules Part 4: Chemical Analysis Methods";
- Added hydride atomic fluorescence spectrometry as the first method for the determination of total mercury; 4.6 methyl mercury in GB/T5009.45-1996 "Analysis Methods for Hygienic Standards of Aquatic Products" is used as the determination of methyl mercury. This standard is proposed and managed by the Ministry of Health of the People's Republic of China. The first method for the determination of total mercury in this standard was drafted by the Food Hygiene Inspection Institute of the Ministry of Health, Beijing Food Hygiene Inspection Institute, Sichuan Food Hygiene Inspection Institute, and Beijing Imported Food Hygiene Inspection Institute. The second method (I) for the determination of total mercury in this standard was drafted by Shanghai Food Hygiene Inspection Institute, Institute of Nutrition and Food Hygiene, Chinese Academy of Preventive Medicine, and Food Hygiene Inspection Institute of the Ministry of Health. The second method (II) for the determination of total mercury in this standard was drafted by Food Hygiene Inspection Institute of the Ministry of Health. The third method for the determination of total mercury in this standard was drafted by Jiangsu Provincial Health and Epidemic Prevention Station. The determination of methyl mercury in this standard was drafted by Shanghai Food Hygiene Inspection Institute, Jiangsu Provincial Health and Epidemic Prevention Station, Hangzhou Health and Epidemic Prevention Station, Food Hygiene Inspection Institute of the Ministry of Health, Qinghai Provincial Health and Epidemic Prevention Station, and Fujian Provincial Health School. The main drafters of the first method for the determination of total mercury in this standard are: Yang Huifen, Huang Liusheng, Mao Hong, Qiang Weiguo, and Yan Jun. This standard was first issued in 1985, revised for the first time in 1996, and this is the second revision. 130
1 Scope
Determination of total mercury and organic mercury in food
Determination of total mercury
This standard specifies the determination method of total mercury in various types of food. This standard applies to the determination of total mercury in various foods GB/T 5009.17--2003
Atomic fluorescence spectrometry: detection limit 0.15ug/kg, standard curve optimal linear range 0ug/L～60μg/L; cold atomic absorption method detection limit: pressure digestion method is 0.4μg/kg, other digestion methods are 10μg/kg; colorimetry is 25μg/kg. Method 1 Atomic fluorescence spectrometry
2 Principle
After the sample is digested by acid heating, in the acidic medium, the mercury in the sample is reduced to atomic mercury by potassium borohydride (KBH) or sodium borohydride (NaBH.), and is carried into the atomizer by the carrier gas (argon). Under the irradiation of a special mercury hollow cathode lamp, the ground state mercury atoms are excited to a high energy state. When deactivated and returned to the ground state, they emit fluorescence of a characteristic wavelength. Its fluorescence intensity is proportional to the mercury content, and is quantitatively compared with the standard series. 3 Reagents
3.1 Nitric acid (superior grade).
3.2 30% hydrogen peroxide.
3.3 Sulfuric acid (superior grade).
3.4 Sulfuric acid + nitric acid + water (1+1+8): Measure 10mL nitric acid and 10mL sulfuric acid, slowly pour into 80mL water, and mix carefully after cooling.
3.5 Nitric acid solution (1+9): Measure 50mL nitric acid, slowly pour into 450mL water, and mix. 3.6 Potassium hydroxide solution (5g/L): Weigh 5.0g potassium hydroxide, dissolve in water, dilute to 1000mL, and mix. 3.7 Potassium borohydride solution (5g/L): Weigh 5.0g potassium borohydride, dissolve in 5.0g/L potassium hydroxide solution, and dilute to 1000mL, mix, and prepare before use.
3.8 Mercury standard stock solution: Accurately weigh 0.1354g of dried mercuric dichloride, add sulfuric acid + nitric acid decahydrate mixed acid (1+1+8) to dissolve, then transfer to a 100mL volumetric flask, dilute to scale, mix well, this solution is equivalent to 1mg of mercury per milliliter. 3.9 Mercury standard working solution: Use a pipette to draw 1mL of mercury standard stock solution (1mg/mL) into a 100mL volumetric flask, dilute to scale with nitric acid solution (1+9), mix well, the concentration of this solution is 10μg/mL. Draw 1mL and 5mL of 10μg/mL mercury standard solution into two 100mL volumetric flasks respectively, dilute to scale with nitric acid solution (1+9), mix well, the solution concentrations are 100ng/mL and 500ng/mL, respectively, used to measure low-concentration samples and high-concentration samples, and prepare standard curves. 4 Instruments
4.1 Dual-channel atomic fluorescence photometer.
4.2 High-pressure digestion tank (100mL capacity). 4.3 Microwave digestion furnace.
GB/T5009.17—2003
5 Analysis steps
5.1 Sample digestion
5.1.1 High-pressure digestion method
This method is applicable to the determination of total mercury in grains, beans, vegetables, fruits, lean meat, fish, eggs, milk and dairy products. 5.1.1.1 Dry samples of grains and beans: Weigh 0.2g~1.00g of dry samples that have been crushed and mixed and passed through a 40-mesh sieve, place them in a polytetrafluoroethylene plastic inner tank, add 5mL nitric acid, mix and leave overnight, then add 7mL hydrogen peroxide, cover the inner cover and place it in a stainless steel jacket, tighten and seal. Then place the digester in a drying oven and heat it to 120℃ and keep the temperature constant for 2h~3h until the digestion is complete, and then cool it to room temperature naturally. Quantitatively transfer the digestion solution with nitric acid solution (1+9) and make it up to 25mL, shake it. At the same time, do a reagent blank test. 5.1.1.2 Vegetables, lean meat, fish and eggs with high moisture content are beaten into a slurry with a masher. Weigh 1.00g~5.00g of the slurry and place it in a polytetrafluoroethylene plastic inner tank. Cover it with a lid and leave a gap. Place it in a 65℃ forced air drying oven or a general oven to dry until it is nearly 100% water. Take it out and proceed according to 5.1.1.1 from "add 5mL nitric acid". 5.1.2 Microwave digestion method
Weigh 0.10g~0.50g of sample and add 1mL~~5mL nitric acid and 1mL~2mL hydrogen peroxide to the digestion tank. After covering the safety valve, place the digestion tank into the microwave digestion system. Set the optimal analytical conditions of the microwave digestion system according to different types of samples (see Tables 1 and 2) until the digestion is complete. After cooling, use nitric acid solution (1+9) to quantitatively transfer and dilute to 25mL (low-content samples can be diluted to 10 mL), mix and test.
Table 1 Microwave analysis conditions for grain, vegetable, fish and meat samples Steps
Power/(%)
Pressure/kPa
Rise time/min
Pressure holding time/min
Exhaust volume/(%)
Power/(%)
Pressure/kPa
Rise time/min
Pressure holding time/min
Exhaust volume/%）
5.2 Preparation of standard series
Table 2 Microwave analysis conditions for oil and sugar samples 2
5.2.1 Low concentration standard series: Pipette 0.25, 0.50, 1.00, 100 ng/mL mercury standard solution respectively. 2.00, 2.50mL in a 25mL volumetric flask, dilute to the mark with nitric acid solution (1+9), and mix. Each is equivalent to a mercury concentration of 1.00, 2.00, 4.00, 8.00, 10.00ng/mL. This standard series is suitable for general sample determination. 5.2.2 High concentration standard series: Pipette 0.25, 0.50, 1.00, 1.50, 2.00mL of 500ng/mL mercury standard solution in a 25mL volumetric flask, dilute to the mark with nitric acid solution (1+9), and mix. Each is equivalent to a mercury concentration of 5.00, 10.00, 20.00, 30.00, 132
40.00ng/mL. This standard series is suitable for the determination of fish and samples with high mercury content. 5.3 Determination
GB/T 5009.17—2003
5.3.1 Instrument reference conditions: Photomultiplier tube negative high voltage: 240 V; Mercury hollow cathode lamp current: 30 mA; Atomizer: temperature: 300°C, height 8.0 mm; Argon gas flow rate: carrier gas 500 mL/min, shielding gas 1000 mL/min; Measuring method: standard curve method, Reading method: peak area, Reading delay time: 1.0 s; Reading time: 10.0 s; Potassium borohydride solution addition time: 8.0 s; Standard solution or sample solution addition volume: 2 mL.
Note: AFS series atomic fluorescence analyzers such as 230, 230a, 2202, 2202a, 2201 and other instruments are fully automatic or sequential flow instruments, and are all equipped with the operating software of this instrument. The instrument analysis conditions should be set to the analysis conditions prompted by this instrument. After the instrument is stable, measure the standard series until the correlation coefficient of the standard curve is r>0.999, then test the sample. Sample pretreatment can be applied to any type of atomic fluorescence analyzer. 5.3.2 Determination method: Select any of the following methods according to the situation. 5.3.2.1 Concentration determination method measurement: Set the optimal conditions of the instrument, gradually increase the furnace temperature to the required temperature, and start measuring after stabilizing for 10min~20min. Continuously inject nitric acid solution (1+9), wait for the reading to stabilize, and then switch to standard series measurement and draw a standard curve. Switch to sample measurement, first inject nitric acid solution (1+9) to make the reading basically return to zero, and then measure the sample blank and sample digestion solution respectively. The injector should be cleaned before measuring each different sample. The sample determination result is calculated according to formula (1). 5.3.2.2 Measurement by the instrument automatically calculating the result: Set the optimal conditions of the instrument, enter the following parameters in the sample parameter screen: sample mass (g or mL), dilution volume (mL), and select the concentration unit of the result, gradually raise the furnace temperature to the required temperature, and measure after stabilization. Continuously inject nitric acid solution (1+9), wait for the reading to stabilize, switch to standard series measurement, and draw a standard curve. Before switching to sample determination, enter the blank value measurement state, inject the sample blank digestion solution, and let the instrument take its average as the blank value for deduction. Then the sample can be measured according to the law. After the determination is completed, select "Print Report" to automatically print the determination results. 6 Result Calculation
The mercury content in the sample is calculated according to formula (1). X (c=co)XV×1 000
m×1000 ×1000
Wherein:
X is the mercury content in the sample, in milligrams per kilogram or milligrams per liter (mg/kg or mg/L); c is the mercury content in the sample digestion solution, in nanograms per milliliter (ng/mL); co is the mercury content in the reagent blank solution, in nanograms per milliliter (ng/mL); V is the total volume of the sample digestion solution, in milliliters (mL); m is the mass or volume of the sample, in grams or milliliters (g or mL). The calculation result retains three significant figures.
7 Precision
The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 10% of the arithmetic mean. The second method is cold atomic absorption spectrometry
8 Principle
Mercury vapor has a strong absorption effect on the resonance line with a wavelength of 253.7nm. The sample is digested with acid or catalytic acid to convert mercury into ion state, and then reduced to elemental mercury with stannous chloride in a strong acid medium. With nitrogen or dry air as a carrier, elemental mercury is blown into the mercury analyzer for cold atomic absorption determination. Within a certain concentration range, its absorption value is proportional to the mercury content, and is quantitatively compared with the standard series. 133
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9 Reagents
9.1 Nitric acid.
9.2 Hydrochloric acid. wwW.bzxz.Net
9.3 Hydrogen peroxide (30%).
(-) Pressure digestion method
9.4 Nitric acid (0.5 + 99.5): Take 0.5mL nitric acid and slowly add it to 50mL water, then dilute it to 100mL with water. 9.5 Potassium permanganate solution (50g/L): Weigh 5.0g potassium permanganate and place it in a 100mL brown bottle, dissolve it in water and dilute it to 100mL. 9.6 Nitric acid-potassium dichromate solution: Weigh 0.05g potassium dichromate and dissolve it in water, add 5mL nitric acid, and dilute it to 100mL with water. 9.7 Stannous chloride solution (100g/L): Weigh 10g stannous fluoride and dissolve it in 20mL hydrochloric acid, dilute it to 100mL with water, and prepare it before use.
9.8 Anhydrous calcium chloride.
9.9 Mercury standard stock solution: Accurately weigh 0.1354g mercuric dioxide dried in a desiccator and dissolve it in nitric acid-potassium dichromate solution, transfer it to a 100mL volumetric flask, and dilute it to the mark with nitric acid-potassium dichromate solution. Mix well. This solution contains 1.0mg mercury per milliliter. 9.10 Mercury standard working solution: 1.0 mg/mL mercury standard stock solution is diluted with nitric acid-potassium dichromate solution to 2.0 ng/mL, 4.0 ng/mL, 6.0 ng/mL, 8.0 ng/mL, and 10.0 ng/mL mercury standard working solution. Prepare it before use. 10 Instruments
All glass instruments used must be soaked in nitric acid (1 + 5) overnight, rinsed repeatedly with water, and finally rinsed with deionized water. 10.1 Double-beam mercury analyzer (with gas circulation pump, gas drying device, mercury vapor generator and mercury vapor absorption bottle). 10.2 Constant temperature drying oven.
10.3 Pressure digester, pressure digestion tank or pressure dissolution bomb. 11 Analysis steps
11.1 Sample pretreatment
11.1.1 During the sampling and preparation process, care should be taken not to contaminate the sample. 11.1.2 After removing impurities from grains and beans, grind them, pass through a 20-mesh sieve, store them in a plastic bottle, and store them for later use. 11.1.3 Fresh samples with high moisture content such as vegetables, fruits, fish, meat and eggs are beaten into a slurry with a food processor or homogenizer, stored in a plastic bottle, and stored for later use.
11.2 Sample digestion (any of the following methods can be used for digestion according to laboratory conditions) Pressure digestion tank digestion method: weigh 1.00g~3.00g of sample (dry sample, sample with high fat content <1.00g, fresh sample <3.00g or weigh the sample according to the instruction manual of the pressure digestion tank) in a polytetrafluoroethylene inner tank, add 2mL~4ml of nitric acid. Soak overnight. Then add 2mL~3mL of hydrogen peroxide (30%) (the total amount cannot exceed one-third of the tank volume). Cover the inner cover, tighten the stainless steel jacket, put it in a constant temperature drying oven, keep it at 120℃140℃ for 3h~4h, cool it naturally to room temperature in the oven, use a dropper to wash or filter the digestion solution into a 10.0mL volumetric flask (depending on the salt content of the sample after digestion), wash the tank with a small amount of water several times, combine the washing solution in the volumetric flask and make up to the scale, mix and set aside; at the same time, make a reagent blank.
11.3 Determination
11.3.1 Instrument conditions: Turn on the mercury analyzer, preheat it for 1h~2h, and adjust the instrument performance to the best state. 11.3.2 Drawing of standard curve: Take 5.0mL of the above prepared mercury standard solution 2.0, 4.0, 6.0, 8.0, 10.0ng/mL (equivalent to 10.0ng, 20.0ng, 30.0ng, 40.0ng, 50.0ng) and place it in the reduction bottle of the mercury vapor generator of the recorder, add 1.0mL of reducing agent stannous chloride (100/1), quickly cover the bottle stopper, and then bubbles will be generated. Measure its absorption value from the highest point displayed by the instrument reading, then open the three-way valve on the absorption bottle to absorb the generated mercury vapor in potassium permanganate solution (50g/L), and perform the next measurement when the reading on the mercury meter reaches zero. And obtain the linear regression equation of the relationship between the absorbance value and the mass of mercury. 134
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11.3.3 Sample determination: Take 5.0 mL of sample solution and reagent blank solution respectively and place them in the reduction bottle of the mercury vapor generator of the mercury analyzer. The following is carried out according to 11.3.2 starting from "add 1.0 mL of reducing agent stannous chloride...". Substitute the measured absorption value into the linear regression equation of the standard series to obtain the mercury content in the sample solution. 12 Calculation of results
The mercury content in the sample is calculated according to formula (2). X - (A1- A)×(V//V2) ×1 000m×1000
Wherein:
The mercury content in the sample, in micrograms per kilogram or microgram per liter (pg/kg or μg/L); A,——The mass of mercury in the sample digestion solution, in nanograms (ng); A2--The mass of mercury in the reagent blank solution, in nanograms (ng); Vi--—The total volume of the sample digestion solution, in milliliters (mL); V,—The volume of the sample digestion solution used for determination,The unit is milliliter (mL); m
The mass or volume of the sample is in grams or milliliters (g or mL). The calculation result shall retain two significant figures.
13 Precision
The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 20% of the arithmetic mean. (II) Other digestion methods
14 Reagents
14.1 Nitric acid.
14.2 Sulfuric acid.
14.3 Stannous chloride solution (300g/L): Weigh 30g of stannous chloride (SnCl2·2H.O), add a small amount of water, and add 2mL of sulfuric acid to dissolve, then dilute with water to 100mL and store in a refrigerator. 14.4 Anhydrous calcium chloride: for drying.
14.5 Mixed acid (1+1+8): Measure 10mL sulfuric acid, add 10mL nitric acid, slowly pour into 50mL water, cool and dilute to 100mL with water
14.6 Vanadium pentoxide.
14.7 Potassium permanganate solution (50g/L): Boil for 10min after preparation, let stand overnight, filter, and store in a brown bottle. 14.8 Hydroxylamine hydrochloride solution (200g/1). 14.9 Mercury standard stock solution: Accurately weigh 0.1354g of mercuric dichloride dried in a desiccator, add mixed acid (1+1+8) to dissolve, transfer to a 100mL volumetric flask, dilute to scale, mix well, and this solution is equivalent to 1.0mg of mercury per milliliter. 14.10 Mercury standard solution: Take 1.0 mL of mercury standard stock solution and place it in a 100 mL volumetric flask. Add mixed acid (1+1+8) to dilute to the mark. Each mL of this solution is equivalent to 10.0 μg of mercury. Take another 1.0 mL of this solution and place it in a 100 mL volumetric flask. Add mixed acid (1+1+8) to dilute to the mark. Each mL of this solution is equivalent to 0.10 μg of mercury. Prepare it before use. 15 Instruments
15.1 Digestion device.
15.2 Mercury analyzer, with gas drying and exhaust devices. 15.3 Mercury vapor generator, see Figure 1.
GB/T5009.17—2003
16 Analysis steps
16.1 Sample digestion
16.1.1 Reflux digestion method
Figure 16 0mL mercury vapor generator
16.1.1.1 Grain or food with little moisture: Weigh 10.00g sample, place it in the conical flask of the digestion device, add a few glass beads, 45mL nitric acid and 10mL sulfuric acid, and rotate the conical flask to prevent local carbonization. After installing the condenser, heat on low heat, stop heating when bubbling begins, and heat to reflux for 2h after bubbling stops. If the solution turns brown during heating, add 5 mL of nitric acid and continue to reflux for 2 h. After cooling, carefully add 20 mL of water from the upper end of the condenser and continue heating and reflux for 10 min. After cooling, rinse the condenser with an appropriate amount of water, and add the washing liquid to the digestion solution. Filter the digestion solution through glass wool into a 100 mL volumetric flask, wash the conical flask and filter with a small amount of water, add the washing liquid to the volumetric flask, add water to the scale, and mix well. Perform a reagent blank test in the same way.
16.1.1.2 Vegetable oil and animal fat: Weigh 5.00 g of the sample, place it in the conical flask of the digestion device, add a few glass beads, add 7 mL of sulfuric acid, carefully mix until the solution turns brown, then add 40 mL of nitric acid, install the condenser, and follow the steps in 16.1.1.1 from "heating on low heat" onwards.
16.1.1.3 Potatoes and bean products: Weigh 20.00g of the mashed and mixed sample (the potatoes must be washed and dried in advance), place it in the conical flask of the digestion device, add a few glass beads and 30mL nitric acid and 5mL sulfuric acid, and rotate the conical flask to prevent local carbonization. After installing the condenser, follow the steps in 16.1.1.1 from "heating with low heat" to operate according to the law. 16.1.1.4 Meat and eggs: Weigh 10.00g of the mashed and mixed sample, place it in the conical flask of the digestion device, add a few glass beads and 30mL nitric acid and 5mL sulfuric acid, and rotate the conical flask to prevent local carbonization. After installing the condenser, follow the steps in 16.1.1.1 from "heating with low heat." to operate according to the law.
16.1.1.5 Milk and dairy products: Weigh 20.00g of milk or yogurt, or dairy products equivalent to 20.00g of milk (2.4g of whole milk powder, 8g of sweetened condensed milk, 5g of light condensed milk), place in a conical flask of a digestion device, add a few glass beads and 30mL of nitric acid, add 10mL of sulfuric acid to milk or yogurt, and add 5mL of sulfuric acid to dairy products, and rotate the conical flask to prevent local carbonization. After installing the condenser, follow the steps in 16.1.1.1 from “Heat on low heat…”.
16.1.2 Vanadium pentoxide digestion method
This method is applicable to aquatic products, vegetables, and fruits. 16.1.2.1 Take the edible part, wash, dry, chop, and mix. Take 2.50g of aquatic products or 10.00g of vegetables and fruits, place them in a 50mL~~100ml conical flask, add 50mg of vanadium pentoxide powder, add 8mL of nitric acid, shake, leave for 4h, add 5mL of sulfuric acid, mix well, then move to a 140℃ sand bath to heat, the effect is violent at the beginning, and then gradually slows down. When there is basically no brown gas escaping from the bottle mouth, rinse the bottle mouth with a small amount of water, heat for another 5min, cool, add 5mL of potassium permanganate solution (50g/L), leave for 4h (or overnight), add hydroxylamine hydrochloride solution (200g/L) dropwise to fade the purple color, shake, leave for several minutes, move to a volumetric flask, and dilute to the scale. Vegetables and fruits are 25mL, and aquatic products are 100mL.
16.1.2.2 Carry out a reagent blank test in the same way. 16.2 Determination
16.2.1 Sample digestion solution prepared by reflux digestion method 16.2.1.1 Take 10.0mL of sample digestion solution and place it in a mercury vapor generator. Connect the exhaust device and quickly add 3mL of stannous chloride solution (300g/L) along the wall. Immediately pass nitrogen gas or air treated with activated carbon at a flow rate of 1.0I./min to allow the mercury vapor to pass through the calcium chloride drying tube and enter the mercury meter. Read the maximum reading on the mercury meter and perform a reagent blank test at the same time. 16.2.1.2 Take 0, 0.10, 0.20, 0.30, 0.40, 0.50 mL of standard mercury solution (equivalent to 0, 0.01, 0.02, 0.03, 0.04, 0.05 μg of mercury) and place in a test tube. Add 10 mL of mixed acid (1+1+8) to each tube. Then proceed as per 16.2.1.1 starting from “Place in mercury vapor generator” to draw a standard curve. 16.2.2 Sample digestion solution prepared by vanadium pentoxide digestion method 16.2.2.1 Take 10.0 mL of sample digestion solution and proceed as per 16.2.1.1. 16.2.2.2 Take 0, 1.0, 2.0, 3.0, 4.0, 5.0 mL of the standard mercury solution (equivalent to 0, 0.1, 0.2, 0.3, 0.4, 0.5 μg of mercury) and place them in 6 50 mL volumetric flasks. Add 1 mL of sulfuric acid (1+1) and 1 mL of potassium permanganate solution (50 g/L) to each flask. Add 20 mL of water and mix well. Add hydroxylamine hydrochloride solution (200 g/L) dropwise to make the purple color fade. Add water to the scale and mix well. Take 10.0 mL (equivalent to 0, 0.02, 0.04, 0.06, 0.08, 0.10 μg of mercury) respectively. The following operations are carried out according to 16.2.1.1 starting from "Place in the mercury vapor generator..." to draw the standard curve. 17 Calculation of results
The mercury content in the sample is calculated according to formula (3). (A —A2) ×1 000
mX(V2/V) X 1 000
Wherein:
The content of mercury in the sample, in milligrams per kilogram (mg/kg); The mass of mercury in the sample digestion solution for determination, in micrograms (μg); The mass of mercury in the reagent blank solution, in micrograms (μg); The mass of the sample, in grams (g);
The total volume of the sample digestion solution, in milliliters (mL); The volume of the sample digestion solution for determination, in milliliters (mL). The calculation result shall be rounded to two significant figures.
18 Precision
The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 15% of the arithmetic mean. Method 3: Disulfide colorimetric method
19 Principle
· (3)
After the sample is digested, mercury ions can react with disulfide in an acidic solution to form an orange-red complex, which is dissolved in chloroform and compared with the standard series for quantification.
20 Reagents
20.1 Nitric acid.
GB/T 5009.17--2003
20.2 Sulfuric acid.
20.3 Ammonia water.
20.4 Chloroform: should not contain oxides. 20.5 Sulfuric acid (1+35): Measure 5 mL of sulfuric acid and slowly pour it into 150 mL of water. After cooling, add water to 180 mL. 20.6 Sulfuric acid (1+19): Take 5 mL of sulfuric acid and pour slowly into water. After cooling, add water to 100 mL. Hydroxylamine hydrochloride solution (200 g/L): Blow clean air to remove trace mercury in the solution. 20.7
20.8 Bromovanillin blue-ethanol indicator solution (1 g/L). 20.9 Disulfide-trifluoromethane solution (0.5 g/L), store in a refrigerator and purify using the following method if necessary. Weigh 0.5g of finely ground dithiocarbamide and dissolve it in 50mL chloroform. If it is not completely dissolved, filter it with filter paper in a 250mL separatory funnel and extract it three times with ammonia water (1+99), 100mL each time. Filter the extract with cotton into a 500mL separatory funnel and adjust it to acidity with hydrochloric acid (1+1). Extract the precipitated dithiocarbamide with chloroform 2 to 3 times, 20mL each time. Combine the chloroform layers, wash twice with an equal amount of water, discard the washing liquid, and evaporate the chloroform in a 50℃ water bath. Put the refined dithiocarbamide in a sulfuric acid desiccator and dry it for later use, or extract the precipitated dithiocarbamide with 200, 200, and 100mL chloroform three times, and combine the chloroform layers to form the dithiocarbamide solution. 20.10 Dithiocarbamide working solution: Take 1.0mL of dithiocarbamide solution, add chloroform to 10mL, and mix. Use a 1 cm colorimetric cup and adjust the zero point with chloroform. Measure the absorbance (A) at a wavelength of 510 nm. Use formula (4) to calculate the number of milliliters (V) of disulfide solution required to prepare 100 mL of disulfide working solution (70% transmittance). V 10(2 = g70) 1. 55
+....-( 4 )
20.11 Mercury standard solution: Accurately weigh 0.1354 g of mercuric dichloride dried in a desiccator, add sulfuric acid (1+35) to dissolve it, transfer it to a 100 mL volumetric flask, and dilute to the mark. This solution is equivalent to 1.0 mg of mercury per milliliter. 20.12 Mercury standard working solution: Take 1.0 mL of mercury standard solution, place it in a 100 mL volumetric flask, add sulfuric acid (1+35) to dilute to the mark. This solution is equivalent to 10.0 μg of mercury per milliliter. Then take 5.0mL of this solution into a 50mL volumetric flask, add sulfuric acid (1+35) to dilute to the mark. Each mL of this solution is equivalent to 1.0g of mercury. 21 Instruments
21.1 Digestion apparatus.
21.2 Visible spectrophotometer.
22 Analysis steps
22.1 Sample digestion
22.1.1 Grain or food with little moisture: Weigh 20.00g of sample, place it in the conical flask of the digestion apparatus, add a few glass beads and 80mL nitric acid and 15mL sulfuric acid, and rotate the conical flask to prevent local carbonization. After installing the condenser, heat on low heat, stop heating when bubbling begins, and heat to reflux for 2h after bubbling stops. If the solution turns brown during heating, add 5mL nitric acid, continue to reflux for 2h, cool, wash the condenser with appropriate amount of water, add the washing liquid into the digestion solution, remove the conical flask, and add water to the total volume of 150mL. Perform a reagent blank test in the same way. 22.1.2 Vegetable oils and animal fats: Weigh 10.00 g of the sample, place it in a conical flask of a digestion device, add a few glass beads and 15 mL of sulfuric acid, carefully mix until the solution turns brown, then add 45 mL of nitric acid, install a condenser, and proceed according to 22.1.1 starting from "heating over low heat...".
22.1.3 Vegetables, fruits, potatoes, and bean products: Weigh 50.00 g of the crushed and mixed sample (soy products are sampled directly, and the edible part of other samples is washed and dried), place it in a conical flask of a digestion device, add a few glass beads and 45 mL of nitric acid and 15 mL of sulfuric acid, and rotate the conical flask to prevent local carbonization. After installing a condenser, proceed according to 22.1.1 starting from "heating over low heat". 22.1.4 Meat, eggs, and aquatic products: Weigh 20.00 g of mashed and mixed sample, place in a conical flask of a digestion device, add several glass beads, 45 mL of nitric acid, and 15 mL of sulfuric acid, install a condenser, and proceed as per 21.1.1 from “heating over low heat.” 22.1.5 Milk and dairy products: Weigh 50.00 g of milk, yogurt, or dairy products equivalent to 50.00 g of milk (6 g of whole milk powder, 138
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20 g of sweetened condensed milk, 12.5 g of evaporated milk), place in a conical flask of a digestion device, add several glass beads and 45 mL of nitric acid, add 15 mL of sulfuric acid to milk and yogurt, and add 10 mL of sulfuric acid to dairy products, install a condenser, and proceed as per 22.1.1 from “heating over low heat…” 22.2 Determination
22.2.1 Take the digestion solution (full volume) of 21.1.1 to 21.1.5, add 20 mL of water, boil on an electric stove for 10 min to remove nitrogen dioxide, etc., and cool.
22.2.2 Add potassium permanganate solution (50 g/L) to the sample digestion solution and reagent blank solution until the solution turns purple, then add hydroxylamine hydrochloride solution (200 g/L) to fade the purple, add 2 drops of thymol blue indicator solution, and adjust the pH with ammonia water to change the orange-red color to orange-yellow (pH 1 to 2). Quantitatively transfer to a 125 mL separatory funnel. 22.2.3 Take 0, 0.5, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0uL of standard mercury solution (equivalent to 0, 0.5, 1.0, 2.0, 3.0, 4.0, 5.0, 6.0μg of mercury) and place them in a 125mL separatory funnel, add 10mL of sulfuric acid (1+19), then add water to 40mL and mix. Then add 1mL of hydroxylamine hydrochloride solution (200g/L) to each, let stand for 20min, and shake from time to time. 22.2.4 Add 5.0 mL of dithiothreitol working solution to the separatory funnel after shaking and cooling the sample digestion solution, reagent blank solution and standard solution, shake vigorously for 2 minutes, let stand and separate, filter the chloroform layer into a 1 cm colorimetric cup through absorbent cotton, adjust the zero point with chloroform, measure the absorbance at a wavelength of 490 nm, subtract the absorbance of the zero tube from the absorbance of the standard tube, and draw a standard curve. 23 Result calculation
The mercury content in the sample is calculated according to formula (5). X-(At-A,)X1 000
mX 1 000
Where:
X—mercury content in the sample, in milligrams per kilogram (mg/kg); A,——the mass of mercury in the sample digestion solution, in micrograms (μg); A—the mass of mercury in the reagent blank solution, in micrograms (μg); m—the mass of the sample, in grams (g). The calculation result shall retain two significant figures.
24 Precision
The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 10% of the arithmetic mean. Determination of methylmercury
25 Scope
This standard specifies the determination method of methylmercury in aquatic products. This standard is applicable to the determination of methylmercury in aquatic products. Gas chromatography (acid extraction thiol cotton method) 26 Principle
· (5)
The methylmercury in the sample is ground with sodium chloride and then added with hydrochloric acid (1+11) containing Cu2+. After complete extraction (Cu2+ exchanges with CH4Hg bound to the tissue), the supernatant is adjusted to a certain acidity by centrifugation or filtration, adsorbed with thiol cotton, and eluted with hydrochloric acid (1+5). Finally, the methylmercury is extracted with benzene and analyzed by a gas chromatograph with an electron capture detector. 27 Reagents
27.1 Sodium chloride.
GB/T5009.17—2003
27.2 Benzene: No impurity peaks on the chromatogram, otherwise it should be redistilled and purified. 27.3 Anhydrous sodium sulfate: Extract with benzene, the concentrate has no impurity peaks on the chromatogram. 27.4 Hydrochloric acid (1+5): Take high-grade pure hydrochloric acid, add an equal volume of water, and distill at constant boiling temperature. The hydrochloric acid distilled is (1+1), and diluted and prepared. 27.5 Copper chloride solution (42.5g/L).
27.6 Sodium hydroxide solution (40g/L): Weigh 40g of sodium hydroxide and dilute to 1000mL with water. 27.7 Hydrochloric acid (1+11): Take 83.3mL of hydrochloric acid (high-grade pure) and dilute to 1000mL with water. 27.8 Eluent (pH 3.0-3.5): Use hydrochloric acid (1+11) to adjust the pH of water to 3.0-3.5. 27.9 Mercapto cotton: Add 35 mL of acetic anhydride, 16 mL of glacial acetic acid, 50 mL of thioglycolic acid, 0.15 mL of sulfuric acid, and 5 mL of water to a 250 mL stoppered conical flask, mix well, cool, add 14 g of absorbent cotton, keep turning and pressing to make the cotton completely soaked, cover the stopper, and place in a constant temperature incubator at (37 ± 0. Keep warm at 5℃ for 4 days (be careful not to exceed 40℃), take it out and wash it with water until it is nearly neutral, remove the moisture and spread it flat on a porcelain plate, then dry it in a constant temperature box at (37±0.5)℃, put the finished product in a brown bottle and store it in the refrigerator for later use (before use, the adsorption efficiency of the cotton wool on methyl mercury should be determined to be above 95% before use). Note: All reagents are extracted with benzene, and the extract should not show methyl mercury peaks on the gas chromatograph. 27.10 Methyl mercury standard solution: Accurately weigh 0.1252g of methylmercury chloride is dissolved in benzene in a 100mL volumetric flask, and diluted to the mark with benzene. Each milliliter of this solution is equivalent to 1.0mg of methylmercury. Store in a refrigerator. 27.11 Methylmercury standard working solution: Take 1.0mL of methylmercury standard solution, place it in a 100mL volumetric flask, and dilute to the mark with benzene. Each milliliter of this solution is equivalent to 10μg of methylmercury. Take 1.0mL of this solution, place it in a 100mL volumetric flask, and dilute to the mark with hydrochloric acid (1+5). Each milliliter of this solution is equivalent to 0.10pg of methylmercury. Prepare it fresh before use. 27.12 Methyl orange indicator solution (1g/L).
28 Instruments
28.1 Gas chromatograph: with 3Ni electron capture detector or xenon source electron capture detector. 28.2 Acidity meter.
28.3 Centrifuge: with 50mL~～80mL centrifuge tubes. 28.4 Universal cotton tube: Use a glass dropper with an inner diameter of 6mm, a length of 20cm, and a thin end (inner diameter 2mm) to fill 0.1g~0.15g of thiol cotton, fill it evenly, and install it before use.
28.5 Glass instruments: Soak them in nitric acid (1+20) for a day and night, and rinse them with water. 29 Analysis steps
29.1 Gas chromatography reference conditions
29.1.163Ni electron capture identifier: column temperature 185℃, identifier temperature 260℃, vaporization chamber temperature 215℃. 29.1.2 Xenon source electron capture identifier: column temperature 185℃, identifier temperature 190℃, vaporization chamber temperature 185℃. 29.1.3 Carrier gas: high purity nitrogen, flow rate 60mL/min (select the best conditions for the instrument). 29.1.4 Chromatographic column: A glass column with an inner diameter of 3 mm and a length of 1.5 m, filled with 60-80 mesh chromosorb WAWDMCS coated with 7% by mass PEGylated polyester (PEGS) or 1.5% by mass 0V-17 and 1.95% QF-1 or 5% by mass diethylene glycol succinate (DEGS) stationary phase. 29.2 Determination
29.2.1 Weigh 1.00g-2.00g of peeled, boneless and minced fish meat (weigh 5g of shrimp and grind it), add an equal amount of sodium chloride, grind it into a paste in a mortar, add 0.5mL of cupric chloride solution (42.5g/1.), grind it gently, transfer it into a 100mL stoppered conical flask with 30mL of hydrochloric acid (1+11) in batches, shake it vigorously for 5min, let it stand for 30min (it can also be shaken on an oscillator for 30min), transfer all the sample liquid into a 50mL centrifuge tube, rinse the conical flask with 5mL of hydrochloric acid (1+11), combine the washing liquid and the sample liquid, centrifuge it for 10min (speed is 2000r/min), transfer all the supernatant into a 100mL separatory funnel, add 10mL of hydrochloric acid (1+11) to the residue, stir it evenly with a glass rod and centrifuge it again, and combine the two centrifuged solutions.
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