GBZ/T 316.1-2018Determination of lead in blood -- Part 1: Graphite furnace atomic absorption spectrometry method (English PDF)

3.1 Principle

Blood samples (hereinafter referred to as blood samples) are deproteinized with a nitric acid solution and measured by graphite furnace atomic absorption spectrometry at a wavelength of 283.3 nm.

Determine the lead content.

3.2 Instrument

3.2.1 Volumetric flask, 10 mL.

3.2.2 stoppered polyethylene centrifuge tube, 1.5mL.

3.2.3 Vortex mixer.

3.2.4 Centrifuge, the speed is greater than 10000r/min.

3.2.5 Micropipettes, the range is 20µL~200µL, 100µL~1000µL.

3.2.6 Atomic absorption spectrometer with graphite furnace, Zeeman or xenon lamp background correction device and lead hollow cathode lamp.

3.3 Reagents

3.3.1 Deionized water.

3.3.2 Nitric acid. rho20=1.42g/mL, excellent grade pure.

3.3.3 Nitric acid solution. 1% (volume fraction).

3.3.4 Nitric acid solution. 5% (volume fraction).

3.3.5 Bovine blood. Heparin anticoagulation, stored at -20 °C, shake it at room temperature. Low background human blood, sheep blood, etc. can also be used. Background lead content should be less than 50µg/L.

3.3.6 Standard solution, using lead single element certified standard substance.

3.4 Sample collection, transportation and storage

According to GBZ/T 295.

The collected samples and sample blanks are placed in a clean container for refrigerated transport.

The sample can be stored for half a year at -20 °C.

3.5 Analysis steps

3.5.1 Instrument Operation Reference Conditions

Dry 80 ° C ~ 150 ° C 55s Ashing 300 ° C ~ 350 ° C 20s Atomization 1600 ° C 5s stop gas Clear 2400 ° C 3s

3.5.2 Preparation of blood lead standard working curve series

The lead single element standard solution is diluted with a 1% nitric acid solution into a 50.0 µg/mL lead standard application solution, and then a 1% nitric acid solution is used to prepare a concentration of 0 µg/mL, 1.25 µg/mL, 2.50 µg/mL, 5.00 µg/mL, 10.00 µg/mL, and 12.50 µg/mL lead standard solution series. Take another 6 10mL Volumetric flasks, numbered from 1 to 6. Add 0.40mL of lead standard solution series with concentration of 0µg/mL~12.50µg/mL, respectively, and then use the bovine blood to make up the volume.

To the scale, the blood lead working curve is prepared at concentrations of 0µg/L, 50µg/L, 100µg/L,.200µg/L, 400µg/L, and 500µg/L.

Quasi-solution series. The preparation method is shown in Table 1.

Table 1 Blood lead working curve standard series configuration

Volumetric flask number 1 2 3 4 5 6 Lead standard solution series/(µg/mL) 0 1.25 2.50 5.00 10.00 12.50 Lead standard solution series/mL 0.40 0.40 0.40 0.40 0.40 0.40 Take bovine blood/mL 9.60 9.60 9.60 9.60 9.60 9.60 Lead standard solution in blood/(µg/L) 0 50 100.200 400 500

3.5.3 Blood lead standard working curve solution series, sample and sample blank pretreatment method

3.5.3.1 Blood lead standard working curve solution series pretreatment method. take 0.15mL blood lead standard solution working curve series respectively in an ethylene centrifuge tube, add 0.60 mL of 5% nitric acid solution to each tube, cover immediately, shake vigorously, and then shake on a vortex mixer.

5 min, centrifuged at 10,000 r/min for 5 min, and the supernatant was used for measurement.

3.5.3.2 Sample Pretreatment Method. Remove the frozen blood sample and return to the laboratory temperature. Shake well and mix well, remove 0.15mL, place in a 1.5 mL stoppered polyethylene centrifuge tube, the remaining treatment steps are the same as above.

3.5.3.3 Sample blank pretreatment method. 2.0 mL water was taken with a blood collection needle and placed in a blood collection tube, and shaken. The remaining treatment steps were the same as above.

3.5.4 Determination of blood lead standard working curve solution series, sample and sample blank

3.5.4.1 Determination of blood lead standard working curve solution series. adjust the atomic absorption spectrometer to the best according to the instrument operation reference conditions.

For the measurement state, 15 µL of the supernatant was injected, and each standard series was measured, and each concentration was repeatedly measured three times. 2 to 6 absorbance value minus 1 After the absorbance value of the number, draw a working curve or calculate a regression equation for the corresponding lead concentration (µg/L).

3.5.4.2 Determination of sample and sample blanks. Determine the sample and sample blank solution using the operating conditions of the standard series of assays.

The result should be less than the detection limit. When the test result is greater than the detection limit, it indicates that the sample is contaminated during the collection, transportation and storage process.

Should be void. The measured absorbance value is calculated from the working curve or the regression equation for the concentration of lead (µg/L).

3.6 Calculation

Calculate the concentration of lead in the blood sample according to formula (1).

C = C0 (1)

In the formula.

C--the concentration of lead in blood, in micrograms per liter (µg/L);

C0--The concentration of lead in a blood sample obtained from the working curve or regression equation in micrograms per liter (µg/L).

3.7 Description

3.7.1 The detection limit of this method is 7µg/L, the lower limit of quantification is 20µg/L; the measurement range is 20µg/L~500µg/L. Zeeman background corrected the relative standard deviation of the atomic absorption spectrometer ranges from 1.6% to 2.8% (n=6), and the background calibration of the atomic absorption spectrometer with xenon lamp is relatively standard deviation.

The difference range is 1.5% to 5.3% (n=6).

3.7.2 The injection volume of this method should be determined according to the specific conditions of the instrument, generally choose 10µL ~ 20µL.

3.7.3 In this method, the matrix has an influence on the measurement. The sample should be treated in the same way as the working curve series solution. If the lead concentration in the sample Exceeding the measurement range, the blood lead working curve range can be increased to 800µg/L or 1000µg/L, and the standard series and samples are diluted by 10 times.

After the method was measured, 0.1 mL of blood was taken, and 5% nitric acid solution was added to 1.0 mL.

3.7.4 The EDTA anticoagulation tube cannot be used for blood collection tubes.

3.7.5 Quality control of the testing process shall be carried out in accordance with the requirements of GBZ/T 295.

4.1 Principle

Blood samples (hereinafter referred to as blood samples) are diluted with a mixture of Triton and Nitric Acid and then absorbed by graphite furnace atom at a wavelength of 283.3 nm.

Spectrometric determination of the concentration of lead.

4.2 Instrument

4.2.1 Volumetric flask. 5mL.

4.2.2 Plugged polyethylene centrifuge tube. 1.5mL.

4.2.3 Micropipette. The range is 20µL~200µL, 100µL~1000µL.

4.2.4 Atomic Absorption Spectrometer. with graphite furnace, Zeeman background correction device and lead hollow cathode lamp.

4.3 Reagents

4.3.1 The experimental water is deionized water.

4.3.2 Nitric acid. rho20=1.42g/mL, excellent grade pure.

4.3.3 Nitric acid solution. 1% (volume fraction).

4.3.4 Triton X-100. Analytically pure.

4.3.5 Triton X-100 solution. 1% (volume fraction).

4.3.6 Diluent. Mix 20 mL of nitric acid solution (1%), 10 mL of Triton X-100 solution (1%) with 70 mL of water.

4.3.7 Bovine blood. Heparin anticoagulation, stored at -20 ° C, shake it at room temperature. Low background human blood, sheep blood, etc. can also be used. Background lead the amount should be less than 50 µg/L.

4.3.8 Standard solution, using lead single element standard solution.

4.4 Sample collection, transportation and storage

According to GBZ/T 295.

The collected samples and sample blanks are placed in a clean container for refrigerated transport.

The sample can be stored for half a year at -20 °C.

4.5 Analysis steps

4.5.1 Instrument Operation Reference Conditions

Dry 80~150°C, 10s

Ashing 500~600°C, 30s, keep 10s Atomization 1800 ° C, 5 s, gas Purification 2400 ° C, 3 s

4.5.2 Preparation of blood lead standard working curve series

The lead single element standard solution is diluted with a 1% nitric acid solution to a standard application solution of 50.0 µg/mL, and then a concentration of 1% nitric acid solution is used.

0 µg/mL, 1.25 µg/mL, 2.50 µg/mL, 5.00 µg/mL, 10.00 µg/mL, 20.00 µg/mL, 25.00 µg/mL standard solution series.

Take 7 5mL volumetric flasks, numbered 1~7. Add 0.20mL of lead standard solution series with concentration of 0µg/mL~25.00µg/mL, respectively.

The volume is set to 0µg/L, 50µg/L, 100µg/L,.200µg/L, 400µg/L, 800µg/L, 1000µg/L.

The blood lead standard working curve solution series. See Table 2 for details.

Table 2 blood lead standard working curve series preparation

Volume bottle number 1 2 3 4 5 6 7 Lead standard solution series/(µg/mL) 0 1.25 2.50 5.00 10.00 20.00 25.00 Lead standard solution series/mL 0.20 0.20 0.20 0.20 0.20 0.20 0.20 Take bovine blood/mL 4.80 4.80 4.80 4.80 4.80 4.80 4.80 Blood lead standard solution series/(µg/L) 0 50 100.200 400 800 1000

4.5.3 Treatment of blood lead standard working curve solution series, sample and sample blank

4.5.3.1 Blood lead standard working curve solution series treatment. blood lead standard working curve series respectively take 0.10mL in the stoppered polyethylene Add 0.90 mL of diluent to each of the hearts, and shake well to mix.

4.5.3.2 Treatment of the sample. Remove the frozen blood sample and return to the laboratory temperature. Shake well and mix well, take 0.10mL and place in 1.5mL in a stoppered polyethylene centrifuge tube, add 0.90 mL of diluent and mix well by shaking.

4.5.3.3 Sample blank treatment. 2.0 mL water was taken from the blood collection tube and placed in the blood collection tube. The remaining treatment steps were the same as above.

4.5.4 Determination of blood lead standard working curve solution series, sample and sample blank

4.5.4.1 Determination of blood lead standard working curve solution series. adjust the atomic absorption spectrometer to the best according to the instrument operation reference conditions.

For the measurement state, 15 µL of the supernatant was injected, and each standard series was measured, and each concentration was repeatedly measured three times. 2 to 7 absorbance value minus 1 After the absorbance value of the number, draw a working curve or calculate a regression equation for the corresponding lead concentration (µg/L).

4.5.4.2 Determination of sample and sample blanks. Determination of samples and blank sample solutions using the operating conditions of the standard series of assays, blank determination of knots the result should be less than the detection limit. When the test result is greater than the detection limit, it indicates that the sample is contaminated during the collection, transportation and storage process.

Should be void.

4.6 Calculation

Calculate the concentration of lead in the blood sample according to formula (2).

C = C0 (2)

In the formula.

C--the concentration of lead in blood, in micrograms per liter (µg/L);

C0--The lead concentration of the blood sample obtained from the standard curve or the regression equation in micrograms per liter (µg/L).

4.7 Description

4.7.1 Atomic absorption spectrometers with background correction of xenon lamps are not suitable for the determination of blood lead in this method.

4.7.2 The detection limit of this method is 7µg/L, the lower limit of quantification is 20µg/L; the measurement range is from 20µg/L to 1000µg/L. The relative standard deviation the range was 1.8%~5.1% (n=6); the recovery rate of blood samples was 99.9%~108.9% (spiking concentration was 100µg/L~800µg/L).

4.7.3 The matrix in this method has an influence on the determination. The sample and the working curve series should be treated in the same way.

4.7.4 The injection volume of this method should be determined according to the specific conditions of the instrument, generally choose 10µL~20µL.

4.7.5 The EDTA anticoagulation tube cannot be used in the blood collection tube.

4.7.6 Quality control of the testing process shall be carried out in accordance with the requirements of GBZ/T 295.