GB 5009.36-2023_English: PDF (GB5009.36-2023)
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National Food Safety Standard - Determination of Cyanide in Foods
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GB 5009.36-2023
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Standards related to: GB 5009.36-2023
| Standard ID | GB 5009.36-2023 (GB5009.36-2023) | | Description (Translated English) | (National food safety standards Determination of sucralose (sucralose) in food) | | Sector / Industry | National Standard | | Classification of Chinese Standard | X09 | | Word Count Estimation | 15,174 | | Date of Issue | 2023-09-06 | | Date of Implementation | 2024-03-06 | | Issuing agency(ies) | National Health Commission of the People's Republic of China, State Administration for Market Regulation | | Summary | This standard specifies the high performance liquid chromatography and high performance liquid chromatography-tandem mass spectrometry methods for the determination of sucralose (sucralose) in foods. This standard applies to the determination of sucralose (sucralose) in food. | GB 5009.36-2023
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard - Determination of Cyanide
in Foods
ISSUED ON: SEPTEMBER 6, 2023
IMPLEMENTED ON: MARCH 6, 2024
Issued by: National Health Commission of the People’s Republic of China;
State Administration for Market Regulation.
Table of Contents
Foreword ... 4
1 Scope ... 5
Method I - Spectrophotometry ... 5
2 Principle ... 5
3 Reagents and Materials ... 5
4 Instruments and Equipment ... 7
5 Analytical Procedures ... 7
6 Expression of Analysis Results ... 10
7 Precision ... 12
8 Others ... 12
Method II - Gas Chromatography ... 12
9 Principle ... 12
10 Reagents and Materials ... 13
11 Instruments and Equipment ... 14
12 Analytical Procedures ... 14
13 Expression of Analysis Results ... 16
14 Precision ... 17
15 Others ... 17
Method III - Gas Chromatography - Mass Spectrometry ... 17
16 Principle ... 17
17 Reagents and Materials ... 18
18 Instruments and Equipment ... 18
19 Analytical Procedures ... 18
20 Expression of Analysis Results ... 20
21 Precision ... 20
22 Others ... 20
Method IV - Ion Chromatography ... 20
23 Principle ... 20
24 Reagents and Materials ... 20
25 Instruments and Equipment ... 22
26 Analytical Procedures ... 22
27 Expression of Analysis Results ... 24
28 Precision ... 24
29 Others ... 24
Method V - Flow Injection / Continuous Flow - Spectrophotometry ... 25
30 Principle ... 25
31 Reagents and Materials ... 25
32 Instruments and Equipment ... 27
33 Analytical Procedures ... 27
34 Expression of Analysis Results ... 28
35 Precision ... 28
36 Others ... 29
Appendix A Reference Conditions of Headspace Analysis ... 30
Appendix B Standard Gas Chromatogram of Cyanogen Chloride ... 31
Appendix C Setting of Ion Chromatography Conditions and Examples of Standard
Chromatograms ... 33
Appendix D Cyanide Ion Determination Chart of Distilled Liquor and Its Integrated
Alcoholic Beverages under Flow Injection / Continuous Flow ... 35
National Food Safety Standard - Determination of Cyanide
in Foods
1 Scope
This Standard specifies the method for the determination of cyanide in foods.
The first method “spectrophotometry” is applicable to the determination of cyanide in distilled
liquor and its integrated alcoholic beverages, edible alcohol, tapioca flour, packaged drinking
water and mineral water.
The second method “gas chromatography” and the third method “gas chromatography - mass
spectrometry” are applicable to the determination of cyanide in liquor, edible alcohol, grain,
packaged drinking water and mineral water.
The fourth method “ion chromatography” is applicable to the determination of cyanide in
distilled liquor and its integrated alcoholic beverages, edible alcohol, packaged drinking water,
mineral water and beverages (based on almonds).
The fifth method “flow injection / continuous flow - spectrophotometry” is applicable to the
determination of cyanide in distilled liquor and its integrated alcoholic beverages, edible
alcohol, packaged drinking water and mineral water.
Method I - Spectrophotometry
2 Principle
Heat the specimen under alkaline conditions to remove high-boiling-point organic matters, or
heat and distill it under acidic conditions. After using sodium hydroxide solution to dissolve or
absorb it, under the condition of pH = 7.0, use chloramine T to convert cyanide to cyanogen
chloride, then, react it with isonicotinic acid-pyrazolone to generate a blue dye, which can be
quantitatively compared with the standard series.
3 Reagents and Materials
Unless it is otherwise specified, the reagents used in this Method are all analytically pure, and
the water is Grade-3 water specified in GB/T 6682.
3.1 Reagents
3.1.1 Absolute ethanol (C2H6O).
distillation device (for water cooling, a condenser is required to adjust the temperature to 10 C
~ 15 C for distillation), insert the lower end of the condenser tube under the liquid level of a
100 mL volumetric flask containing 10 mL of sodium hydroxide solution (20 g/L) (if the room
temperature is higher than 25 C, use an ice pack to cool it down, so that the temperature of the
distillate is between 20 C ~ 25 C), and perform distillation. When the distillate collected is
nearly 100 mL, use water to reach a constant volume to the scale. Accurately draw 1.00 mL of
the solution into a 10 mL colorimetric tube, add 2 g/L of sodium hydroxide solution to 5 mL,
shake it well, and use it as a specimen solution.
5.1.1.2 Add 2 drops of phenolphthalein indicator solution to the specimen solution, add acetic
acid solution to adjust it, until the red color fades, then, use 20 g/L sodium hydroxide solution
to adjust it to nearly red color. Add 2 mL of phosphate buffer solution, then add 0.2 mL of
chloramine T solution, shake it well and let it stand for 3 min. Add 2 mL of isonicotinic acid-
pyrazolone solution, add water to reach a constant volume to the scale. Add a stopper and shake
to evenly mix it. Place it in a constant-temperature water bath at 37 C for 40 min and reserve
it for testing.
5.1.2 Distilled liquor and its integrated alcoholic beverages, edible alcohol
5.1.2.1 Accurately draw 1.00 mL of specimen (for sesame-flavor liquor, Maotai-flavor liquor
and special-flavor liquor specimens, draw 0.10 mL) into a 25 mL beaker, add 5 mL of sodium
hydroxide solution (2 g/L), and leave it for 10 min. Then, place it on an electric heating plate at
160 C ~ 180 C and heat it at low temperature, until about 1 mL of the solution remains.
Remove it and place it at room temperature. Use 2 g/L sodium hydroxide solution to transfer it
to a 10 mL colorimetric tube with a stopper. Finally, add 2 g/L sodium hydroxide solution to 5
mL.
5.1.2.2 If the liquor specimen is turbid or colored, take 25.0 mL of specimen into a 250 mL
distillation flask, add 100 mL of water, add a few glass beads, and dropwise add a few drops of
methyl orange indicator. Insert the lower end of the condenser tube under the liquid level of the
colorimetric tube containing 10 mL of sodium hydroxide solution (20 g/L), add 1 g ~ 2 g of
tartaric acid, the solution changes from orange to orange-red. Quickly connect the distillation
device to perform distillation, and collect nearly 50 mL of the distillate, then, use water to reach
a constant water of 50 mL, and evenly mix it. Take 2.00 mL of the distillate and operate in
accordance with 5.1.2.1.
5.1.2.3 Same as 5.1.1.2.
5.1.3 Packaged drinking water and mineral water
5.1.3.1 Measure-take 250 mL of water specimen (when the cyanide content exceeds 20 g, take
an appropriate amount of water specimen and add water to 250 mL) and place it into a 500 mL
distillation device. Add 1 ~ 2 drops of methyl orange indicator, then, add 5 mL of zinc acetate
solution, and add 1 g ~ 2 g of tartaric acid, the solution changes from orange to orange-red.
Quickly connect the distillation device, insert the lower end of the condenser tube under the
liquid level of a 50 mL colorimetric tube with a stopper containing 5.0 mL of sodium hydroxide
solution (20 g/L). By adjusting the temperature, control the distillation speed at 2 mL/min ~ 3
mL/min, collect nearly 50 mL of the distillate, then, use water to reach a constant water of 50
mL, and evenly mix it. Take 10.0 mL of the distillate and place it in a 25 mL colorimetric tube
with a stopper.
5.1.3.2 Add 5.0 mL of phosphate buffer solution to the colorimetric tube containing the
specimen, place it in a constant-temperature water bath at 37 C, then, add 0.25 mL of
chloramine T solution, add a stopper to mix it, leave it for 5 min. Then, add 5.0 mL of
isonicotinic acid-pyrazolone solution, add water to 25 mL and mix it well. Place it in a constant-
temperature water bath at 37 C for 40 min and reserve it for testing.
5.2 Drawing of Standard Working Curve
5.2.1 Standard curve of tapioca flour
Respectively and accurately draw 0 mL, 0.40 mL, 0.80 mL, 1.20 mL, 1.60 mL and 2.00 mL of
cyanide ion standard solution (1.0 g/mL) into a 10 mL colorimetric tube [equivalent to the
mass of cyanide (CN): 0 g, 0.40 g, 0.80 g, 1.20 g, 1.60 g and 2.00 g], add 2 g/L sodium
hydroxide solution to 5 mL. In accordance with 5.1.1.2, conduct synchronous operation with
the sample below. Take the prepared standard curve solution to be tested, use a 1 cm cuvette,
and take 2 g/L sodium hydroxide solution as the blank solution to adjust the zero point. At a
wavelength of 638 nm, determine the absorbance; take the mass of cyanide ions as the x-
coordinate and the absorbance of cyanide ions as the y-coordinate, and draw a standard curve.
5.2.2 Standard curve of distilled liquor and its integrated alcoholic beverages, edible
alcohol
Respectively and accurately draw 0 mL, 0.40 mL, 0.80 mL, 1.20 mL, 1.60 mL and 2.00 mL of
cyanide ion standard solution (1.0 g/mL) into a 50 mL beaker [equivalent to the mass of
cyanide (CN): 0 g, 0.40 g, 0.80 g, 1.20 g, 1.60 g and 2.00 g], add 5 mL of sodium
hydroxide solution (2 g/L) and leave it for 10 min. In accordance with 5.1.2.1 and 5.1.2.3,
conduct synchronous operation with the sample below. Take the prepared standard curve
solution to be tested, use a 1 cm cuvette, and take 2 g/L sodium hydroxide solution as the blank
solution to adjust the zero point. At a wavelength of 638 nm, determine the absorbance; take
the mass of cyanide ions as the x-coordinate and the absorbance of cyanide ions as the y-
coordinate, and draw a standard curve.
5.2.3 Standard curve of packaged drinking water and mineral water
Respectively and accurately draw 0 mL, 0.10 mL, 0.20 mL, 0.40 mL, 0.80 mL and 1.00 mL of
cyanide ion standard solution (1.0 g/mL) into a 25 mL colorimetric tube [equivalent to the
mass of cyanide (CN): 0 g, 0.10 g, 0.20 g, 0.40 g, 0.80 g and 1.00 g], add 2 g/L sodium
hydroxide solution to 10.0 mL. In accordance with 5.1.3.2, conduct synchronous operation with
the sample below. Take the prepared standard curve solution to be tested, use a 3 cm cuvette,
and take 2 g/L sodium hydroxide solution as the blank solution to adjust the zero point. At a
wavelength of 638 nm, determine the absorbance; take the mass of cyanide ions as the x-
coordinate and the absorbance of cyanide ions as the y-coordinate, and draw a standard curve.
cyanide in the specimen, under acidic conditions, use chloramine T to derive it into cyanogen
chloride. Cyanogen chloride reaches equilibrium in the gas phase and liquid phase, and the gas
phase part is introduced into the gas chromatograph for separation. Use an electron capture
detector for detection and the external standard method for quantitative determination.
10 Reagents and Materials
Unless it is otherwise specified, the reagents used in this Method are all analytically pure, and
the water is Grade-2 water specified in GB/T 6682.
10.1 Reagents
10.1.1 Chloramine T (C7H7ClNNaO2S 3 H2O).
10.1.2 Phosphoric acid (H3PO4): 85%.
10.1.3 Sodium hydroxide (NaOH).
10.2 Reagent Preparation
10.2.1 Chloramine T solution (10 g/L): weigh-take 0.1 g of chloramine T, use water to dissolve
it and reach a constant volume of 10 mL. Prepare it right before use.
10.2.2 Phosphoric acid solution (1 + 5): measure-take 10 mL of concentrated phosphoric acid,
add it to 50 mL of water, and evenly mix it.
10.2.3 0.1% sodium hydroxide solution: weigh-take 1.0 g of sodium hydroxide, use water to
dissolve it and reach a constant volume of 1 L.
10.2.4 0.01 mol/L sodium hydroxide solution: weigh-take 0.4 g of sodium hydroxide, use water
to dissolve it and reach a constant volume of 1 L.
NOTE: when the prepared chloramine T solution is turbid, it needs to be replaced with newly
prepared chloramine T.
10.3 Reference Material
Cyanide standard solution (50.0 g/mL, calculated as CN): a standard substance certified by
the state and awarded a reference material certificate. Please refer to the reference material
certificate for shelf life and storage conditions.
10.4 Preparation of Standard Solutions
10.4.1 Cyanide ion (calculated as CN) standard intermediate solution: accurately transfer-take
2.00 mL of cyanide standard solution (50.0 g/mL) into a 10.0 mL volumetric flask; use 0.1%
sodium hydroxide solution to reach a constant volume. The mass concentration of this solution
is 10.0 mg/L. Prepare it right before use.
10.4.2 Cyanide ion (calculated as CN) standard working solutions: transfer-take an appropriate
amount of cyanide ion (calculated as CN) standard intermediate solution (10.0 mg/L), and use
0.01 mol/L sodium hydroxide solution to dilute it and prepare working solutions respectively
with a mass concentration of 0.001 mg/L, 0.005 mg/L, 0.050 mg/L, 0.100 mg/L and 0.200 mg/L.
Prepare it right before use.
NOTE: since cyanide is a highly toxic substance, please operate in a fume hood.
11 Instruments and Equipment
11.1 Gas chromatograph: equipped with an electron capture detector (ECD).
11.2 Headspace sampler.
11.3 Headspace bottle: 20 mL.
11.4 Vortex mixer.
11.5 Analytical balance: with a division value of 0.0001 g.
11.6 Ultrasonic cleaner.
11.7 Centrifuge: with a rotational speed 4,000 r/min.
12 Analytical Procedures
12.1 Pre-treatment of Sample
12.1.1 Specimen preparation
Take about 500 g of solid specimen, use a sample crushing device to make it into powder, put
it into a clean container, seal it, and store it at 0 C ~ 4 C. Take about 500 mL of liquid specimen,
thoroughly mix it, put it into a clean container, seal it, and store it at 0 C ~ 4 C.
NOTE: during the operation of specimen preparation, samples must be prevented from being
contaminated.
12.1.2 Determination of specimen solution
12.1.2.1 Liquor and edible alcohol
Accurately transfer-take 0.20 mL of specimen into a headspace bottle, add 9.80 mL of 0.01
mol/L sodium hydroxide solution, let it stand for 5 min, add 0.2 mL of phosphoric acid solution.
Then, add 0.2 mL of chloramine T solution, immediately cap seal it, and reserve it for testing.
12.1.2.2 Grain
24.1.4 Methanol (CH3OH).
24.1.5 Acetonitrile (C2H3N).
24.2 Reagent Preparation
24.2.1 Eluent: 100 mmol/L sodium hydroxide + 500 mmol/L sodium acetate + 0.5%
ethylenediamine (volume ratio): weigh-take 41.02 g of sodium acetate and dissolve it in 1 L of
water; pass it through a 0.22 m nylon microporous filter membrane, add 5.00 mL of
ethylenediamine and 5.24 mL of 50% sodium hydroxide.
24.2.2 500 mmol/L sodium hydroxide solution: draw 2.62 mL of 50% sodium hydroxide, add
it to 100 mL of water, and evenly mix it.
24.3 Reference Material
Cyanide standard solution (50.0 g/mL, calculated as CN): a standard substance certified by
the state and awarded a reference material certificate. Please refer to the reference material
certificate for shelf life and storage conditions.
24.4 Preparation of Standard Solution
24.4.1 Preparation of distilled liquor and its integrated alcoholic beverages, edible alcohol
standard series solutions
24.4.1.1 Cyanide (calculated as CN) standard intermediate solution (1.00 mg/L): draw 0.20
mL of cyanide standard solution (50.0 g/mL) into a 10 mL volumetric flask, add water to reach
a constant volume to the scale, and evenly mix it. Prepare it right before use.
24.4.1.2 Cyanide (calculated as CN) standard series working solutions: respectively draw
0.050 mL, 0.10 mL, 0.20 mL, 0.50 mL and 1.00 mL of cyanide (calculated as CN) standard
intermediate solution (1.00 mg/L) into a 10 mL volumetric flask, add water to reach a constant
volume to the scale, and evenly mix it. The mass concentration of the cyanide (calculated as
CN) standard series working solutions is respectively 0.0050 mg/L, 0.010 mg/L, 0.020 mg/L,
0.050 mg/L and 0.100 mg/L. Prepare it right before use.
24.4.2 Preparation of packaged drinking water and mineral water standard series
solutions
24.4.2.1 Cyanide (calculated as CN) standard intermediate solution (1.00 mg/L): draw 0.20
mL of cyanide standard solution (50.0 g/mL) into a 10 mL volumetric flask, add water to reach
a constant volume to the scale, and evenly mix it. Prepare it right before use.
24.4.2.2 Cyanide (calculated as CN) standard series working solutions: respectively draw
0.050 mL, 0.10 mL, 0.20 mL, 0.50 mL and 1.00 mL of cyanide standard solution (1.00 mg/L)
into a 10 mL volumetric flask, add water to reach a constant volume to the scale. Then, add
0.10 mL of sodium hydroxide solution (500 mmol/L), and evenly mix it. The mass
concentration of the cyanide (calculated as CN) standard series working solutions is
Take about 500 mL of sample, thoroughly mix it, put it into a clean container, seal it, and store
it at 0 C ~ 4 C.
26.1.2 Specimen dilution, purification and pH adjustment
26.1.2.1 Packaged drinking water and mineral water
Accurately transfer-take 10.0 mL of specimen into a colorimetric tube, add 0.1 mL of sodium
hydroxide solution (500 mmol/L), evenly mix it, and reserve it for testing.
26.1.2.2 Distilled liquor and its integrated alcoholic beverages, edible alcohol
Accurately transfer-take 1.00 mL of specimen into a 10.0 mL colorimetric tube, use water to
dilute to reach a constant volume to the scale, and evenly mix it. Then, filter it through 0.22 m
nylon microporous filter membrane and reserve it for testing. When the sample color is
relatively dark or the sugar content is relatively high, firstly, pass the test solution through the
IC-RP solid-phase extraction column, control the entire flow rate at 4 mL/min, discard the initial
3 mL or 6 mL (selected in accordance with the column specifications), then, filter it through the
0.22 m nylon microporous filter membrane, and reserve it for testing.
26.1.2.3 Beverages (based on almonds)
Accurately transfer-take 4.00 mL of specimen into a 10.0 mL colorimetric tube, use acetonitrile
to reach a constant volume to the scale, perform vortex mix for 2 min. Transfer it to a centrifuge
tube, let it stand, then, centrifuge for 10 min. Accurately transfer-take 2.5 mL of the supernatant
and use water to reach a constant volume to 10.0 mL. Take an appropriate amount of the above-
mentioned solution, firstly, pass it through the IC-RP solid phase extraction column, control the
entire flow rate at 4 mL/min, discard the initial 3 mL or 6 mL (selected in accordance with the
column specifications), then, filter it through the 0.22 m nylon microporous filter membrane,
and reserve it for testing.
26.2 Reference Conditions of Instruments
Chromatographic column: the filler is an anion protection column and separation column of
polystyrene divinylbenzene copolymer with alkyl quaternary ammonium groups, or those with
equivalent performance.
Eluent: 100 mmol/L sodium hydroxide + 500 mmol/L sodium acetate + 0.5% ethylenediamine
(volume ratio), perform isocratic elution.
Flow rate: 1 mL/min.
Column temperature: 30 C.
Detector: ampere detector.
Injection volume: 250 L for beverages (based on almonds); 50 L for others.
31.2 Reagent Preparation
31.2.1 Flow injection method
31.2.1.1 Sodium hydroxide solution (1.0 g/L): weigh-take 1.0 g of sodium hydroxide, dissolve
in water, and reach a constant volume of 1,000 mL. Prepare it right before use. Before use, it
needs to be degassed. This solution is also used for the standard curve and the alkaline
hydrolysis dilution of liquor samples.
31.2.1.2 Distillation reagent (pH = 3.8): weigh-take 13.2 g of tartaric acid into a 1,000 mL
beaker, add 800 mL of water, then, weigh 3.5 g of sodium hydroxide and add it to the above-
mentioned solution to dissolve, then, use water to reach a constant volume in a 1,000 mL
volumetric flask.
31.2.1.3 Buffer solution (pH = 4.0): weigh-take 97.0 g of MKP (KH2PO4), use water to reach a
constant volume of 1,000 mL. It shall remain valid for 1 month.
31.2.1.4 Chloramine T solution: weigh-take 2.0 g of chloramine T and reach a constant volume
in 500 mL of water. Place it in a brown glass bottle and prepare it right before use. Before use,
it needs to be degassed.
31.2.1.5 Isonicotinic acid / barbituric acid color developer: weigh-take 12.0 g of sodium
hydroxide, add 500 mL of water to dissolve it, add until it is completely dissolved. Then, weigh-
take 16.8 g of 1,3-dimethylbarbituric acid and add to it. After it is completely dissolved, add
13.6 g of isonicotinic acid, use water to reach a constant volume to 1,000 mL, and store it in a
brown glass bottle. It shall remain valid for 1 month (if there are insoluble substances in the
solution, before use, it needs to be filtered with a filter paper). Before use, it needs to be
degassed.
31.2.2 Continuous flow method
31.2.2.1 0.5 mol/L sodium hydroxide solution: weigh-take 20 g of sodium hydroxide and
dissolve it in 500 mL of water, then, use water to reach a constant volume of 1,000 mL.
31.2.2.2 Distillation reagent: add 250 mL of phosphoric acid to 500 mL of water, add 50 mL of
hypophosphorous acid, evenly mix it, and use water to reach a constant volume of 1,000 mL.
31.2.2.3 0.1 mol/L sodium hydroxide solution: weigh-take 4 g of sodium hydroxide and
dissolve it in 500 mL of water, use water to reach a constant volume of 1,000 mL.
31.2.2.4 pH 5.2 buffer solution: dissolve 13.6 g of MKP and 0.34 g of DKP in 800 mL of water,
use 0.5 mol/L NaOH to adjust pH = 5.2; use water to reach a constant volume of 1,000 mL, add
3 mL of Brij35 and evenly mix it. Place it in a brown bottle and store in the refrigerator at 4 C.
31.2.2.5 Chloramine T solution: weigh-take 1 g of chloramine T and dissolve in 500 mL of
water.
31.2.2.6 Color developing reagent: weigh-take 4.4 g of sodium hydroxide and dissolve in 400
mL of water, add 6.8 g of barbituric acid and 6.8 g of isonicotinic acid. At 30 C, stir it for 1 h;
use 0.5 mol/L sodium hydroxide solution to adjust pH = 5.5, and reach a constant volume of
500 mL.
31.3 Reference Material
Same as 3.3.
31.4 Preparation of Standard Solution
Same as 3.4.
32 Instruments and Equipment
32.1 Analytical balance: with a division value of 0.001 g.
32.2 pH meter.
32.3 Fully automatic flow injector or flow analyzer: automatic sampler, chemical analysis unit,
600 nm detector and data processing unit.
33 Analytical Procedures
33.1 Pre-treatment of Sample
33.1.1 Packaged drinking water and mineral water
The samples can be directly tested on the machine.
33.1.2 Distilled liquor and its integrated alcoholic beverages, edible alcohol
Accurately draw 1.00 mL of specimen (for sesame-flavor liquor, Maotai-flavor liquor and
special-flavor liquor specimens, draw 0.25 mL) into a 25 mL volumetric flask, use sodium
hydroxide solution (1.0 g/L) to reach a constant volume to the scale, evenly mix it and let it
stand for 10 min. After alkaline hydrolysis, inject the specimen for determination.
33.2 Instrument Operation
Refer to the instrument manual. Turn on the instrument, adjust the flow path system, and select
corresponding analysis conditions to preheat the instrument.
33.3 Determination
After the flow path system becomes stable, successively determine the standard series and
samples.
33.3.1 Drawing of standard curve
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