GB 5009.181-2016 English PDF

GB 5009.181-2016: National food safety standard - Determination of malondialdehyde in food
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GB 5009.181-2016
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard -
Determination of malondialdehyde in food
ISSUED ON. AUGUST 31, 2016
IMPLEMENTED ON. MARCH 01, 2017
Issued by. National Health and Family Planning Commission of the PRC
Table of Contents
Foreword ... 3 
1 Scope ... 4 
2 Principles ... 4 
3 Reagents and materials ... 4 
4 Instruments and equipment... 5 
5 Analytical procedures ... 6 
6 Analysis results expression ... 7 
7 Precision ... 7 
8 Principles ... 7 
9 Reagents and materials ... 8 
10 Instruments and equipment... 9 
11 Analytical procedures ... 9 
12 Analysis results expression ... 9 
13 Precision ... 10 
14 Other ... 10 
Appendix A Chromatogram of malondialdehyde thiobarbituric acid derivative
standard solution ... 11 
National Food Safety Standard -
Determination of malondialdehyde in food
1 Scope
This Standard specifies the method for determination of malondialdehyde in
food.
Method 1 of this Standard is applicable to the determination of malondialdehyde
in food. Method 2 of this Standard is applicable to the determination of
malondialdehyde in animal and vegetable fats and oils.
Method 1 High performance liquid chromatography
2 Principles
The sample is first extracted with acid solution. Then the extract is reacted with
thiobarbituric acid (TBA) to form a colored compound. USE high performance
liquid chromatography-diode array detector to determine; and external standard
method to quantify.
3 Reagents and materials
Unless otherwise stated, the reagents used in this method are of analytically
pure; the water is Grade I water specified in GB/T 6682.
3.1 Reagents
3.1.1 Methanol (CH3OH). chromatographically pure.
3.1.2 Trichloroacetic acid (C2HCl3O2).
3.1.3 Ethylene diamine tetraacetic acid (C10H14N2Na2O8 · 2H2O).
3.1.4 Thiobarbituric acid (TBA) (C4H4N2O2S).
3.2 Preparation of reagents
3.2.1 Ammonium acetate solution (0.01 mol/L). WEIGH 0.77 g of ammonium
acetate; ADD water to dissolve and dilute to 1000 mL; and USE a 0.45 μm filter
to filter.
3.2.2 Trichloroacetic acid mixture. accurately WEIGH 37.50 g (accurate to 0.01
g) of trichloroacetic acid (3.1.2) and 0.50 g (accurate to 0.01 g) of ethylene
diamine tetraacetic acid (3.1.3); USE water to dissolve, dilute to 500 mL.
3.2.3 Aqueous solution of thiobarbituric acid (TBA). accurately WEIGH 0.288 g
(accurate to 0.001 g) of thiobarbituric acid (3.1.4) dissolved in water and diluted
to 100 mL (If not easy to dissolve, it can be heated by ultrasonic until completely
dissolved. After cooling, DILUTE to volume of 100 mL), equivalent to 0.02 mol/L.
3.3 Standard
1,1,3,3-tetraethoxypropane (also known as malondialdehyde acetal, C11H24O4,
CAS number. 122-31-6). Purity≥97%.
3.4 Preparation of standard solutions
3.4.1 Standard stock solution of malondialdehyde (100 μg/mL). Accurately
TRANSFER 0.315 g (accurate to 0.001 g) of 1,1,3,3-tetraethoxypropane to a
1000 mL volumetric flask; after dissolving in water, DILUTE to 1000 mL; SET in
a refrigerator at 4 °C for storage. It is valid for 3 months.
3.4.2 Standard use solution of malondialdehyde (1.00 μg/mL). accurately
PIPETTE 1.0 mL of standard stock solution of malondialdehyde (3.4.1); USE
trichloroacetic acid mixture (3.2.2) to dilute to 100 mL; SET in a refrigerator at
4 °C for storage. It is valid for 2 weeks.
3.4.3 Standard series solution of malondialdehyde. accurately PIPETTE 0.10
mL, 0.50 mL, 1.0 mL, 1.5 mL, 2.5 mL of standard use solution of
malondialdehyde in 10 mL volumetric flasks; ADD trichloroacetic acid mixture
(3.2.2) to dilute to volume. The concentration of the series of standard solutions
is 0.01 μg/mL, 0.05 μg/mL, 0.10 μg/mL, 0.15 μg/mL, 0.25 μg/mL. Prepared
when used.
4 Instruments and equipment
4.1 High performance liquid chromatography. equipped with a diode array
detector.
4.2 Balance. The sensitivity is 0.0001 g, 0.01 g.
4.3 Thermostatic oscillator.
4.4 Thermostatic water bath.
to the standard curve, the concentration of malondialdehyde in the solution to
be tested is obtained. The liquid chromatogram of malondialdehyde standard is
shown in Appendix A.
6 Analysis results expression
The content of malondialdehyde in the sample shall be calculated according to
formula (1).
Where.
X - The content of malondialdehyde in the sample, in milligrams per kilogram
(mg/kg);
c - The concentration of malondialdehyde in the sample solution obtained from
the standard series curve, in micrograms per milliliter (μg/mL);
V - The constant volume of sample solution, in milliliters (mL);
m - The mass of sample represented by the final sample solution, in grams (g);
1000 - Conversion factor.
The calculation result is expressed as the arithmetic mean of the two
independent determination results obtained under repeated conditions. The
result retains two significant figures.
7 Precision
The absolute difference between the two independent determination results,
obtained under repeated conditions, shall not exceed 10% of the arithmetic
mean.
Method 2 Spectrophotometry
8 Principles
After the malondialdehyde is extracted by trichloroacetic acid solution, it reacts
with thiobarbituric acid (TBA) to form a pink compound. Its absorbance at 532
nm of wavelength is measured and compared with the standard series to
to dilute to volume. The concentration of the series of standard solutions is 0.01
μg/mL, 0.05 μg/mL, 0.10 μg/mL, 0.15 μg/mL, 0.25 μg/mL. Prepared when used.
10 Instruments and equipment
10.1 Spectrophotometer.
10.2 Balance. The sensitivity is 0.0001 g, 0.01 g.
10.3 Thermostatic oscillator.
10.4 Thermostatic water bath.
11 Analytical procedures
11.1 Sample preparation
WEIGH 5 g (accurate to 0.01 g) of the sample; PLACE it in a 100 mL conical
flask with stopper; accurately ADD 50 mL of trichloroacetic acid mixture; SHAKE
it well; and USE the stopper to seal. PLACE on a thermostatic oscillator to
shake at 50 °C for 30 min; REMOVE, and COOL to room temperature. USE
double-layer quantitative slow filter paper to filter; DISCARD the primary filtrate;
and the subsequent filtrate is standby.
Accurately PIPETTE 5 mL of the above filtrate and of standard series solution
respectively; PLACE them in 25 mL colorimetric tube with stopper. TAKE
another 5 mL of trichloroacetic acid mixture as a sample blank; ADD 5 mL of
aqueous solution of thiobarbituric acid (TBA) respectively; COVER the stopper;
and MIX well. PLACE in a 90 °C water bath to react for 30 min; TAKE out; and
COOL to room temperature.
11.2 Determination
USE the sample blank to adjust the zero point. At 1 cm optical path at a
wavelength of 532 nm, the absorbance of the sample solution and the standard
series solution (9.4.3) is determined. USE the mass concentration of the
standard series solution as the abscissa. USE the absorbance value as the
ordinate. The standard curve is drawn.
12 Analysis results expression
The content of malondialdehyde in the sample shall be calculated according to
formula (2).
GB 5009.181-2016
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard -
Determination of malondialdehyde in food
ISSUED ON. AUGUST 31, 2016
IMPLEMENTED ON. MARCH 01, 2017
Issued by. National Health and Family Planning Commission of the PRC
Table of Contents
Foreword ... 3 
1 Scope ... 4 
2 Principles ... 4 
3 Reagents and materials ... 4 
4 Instruments and equipment... 5 
5 Analytical procedures ... 6 
6 Analysis results expression ... 7 
7 Precision ... 7 
8 Principles ... 7 
9 Reagents and materials ... 8 
10 Instruments and equipment... 9 
11 Analytical procedures ... 9 
12 Analysis results expression ... 9 
13 Precision ... 10 
14 Other ... 10 
Appendix A Chromatogram of malondialdehyde thiobarbituric acid derivative
standard solution ... 11 
National Food Safety Standard -
Determination of malondialdehyde in food
1 Scope
This Standard specifies the method for determination of malondialdehyde in
food.
Method 1 of this Standard is applicable to the determination of malondialdehyde
in food. Method 2 of this Standard is applicable to the determination of
malondialdehyde in animal and vegetable fats and oils.
Method 1 High performance liquid chromatography
2 Principles
The sample is first extracted with acid solution. Then the extract is reacted with
thiobarbituric acid (TBA) to form a colored compound. USE high performance
liquid chromatography-diode array detector to determine; and external standard
method to quantify.
3 Reagents and materials
Unless otherwise stated, the reagents used in this method are of analytically
pure; the water is Grade I water specified in GB/T 6682.
3.1 Reagents
3.1.1 Methanol (CH3OH). chromatographically pure.
3.1.2 Trichloroacetic acid (C2HCl3O2).
3.1.3 Ethylene diamine tetraacetic acid (C10H14N2Na2O8 · 2H2O).
3.1.4 Thiobarbituric acid (TBA) (C4H4N2O2S).
3.2 Preparation of reagents
3.2.1 Ammonium acetate solution (0.01 mol/L). WEIGH 0.77 g of ammonium
acetate; ADD water to dissolve and dilute to 1000 mL; and USE a 0.45 μm filter
to filter.
3.2.2 Trichloroacetic acid mixture. accurately WEIGH 37.50 g (accurate to 0.01
g) of trichloroacetic acid (3.1.2) and 0.50 g (accurate to 0.01 g) of ethylene
diamine tetraacetic acid (3.1.3); USE water to dissolve, dilute to 500 mL.
3.2.3 Aqueous solution of thiobarbituric acid (TBA). accurately WEIGH 0.288 g
(accurate to 0.001 g) of thiobarbituric acid (3.1.4) dissolved in water and diluted
to 100 mL (If not easy to dissolve, it can be heated by ultrasonic until completely
dissolved. After cooling, DILUTE to volume of 100 mL), equivalent to 0.02 mol/L.
3.3 Standard
1,1,3,3-tetraethoxypropane (also known as malondialdehyde acetal, C11H24O4,
CAS number. 122-31-6). Purity≥97%.
3.4 Preparation of standard solutions
3.4.1 Standard stock solution of malondialdehyde (100 μg/mL). Accurately
TRANSFER 0.315 g (accurate to 0.001 g) of 1,1,3,3-tetraethoxypropane to a
1000 mL volumetric flask; after dissolving in water, DILUTE to 1000 mL; SET in
a refrigerator at 4 °C for storage. It is valid for 3 months.
3.4.2 Standard use solution of malondialdehyde (1.00 μg/mL). accurately
PIPETTE 1.0 mL of standard stock solution of malondialdehyde (3.4.1); USE
trichloroacetic acid mixture (3.2.2) to dilute to 100 mL; SET in a refrigerator at
4 °C for storage. It is valid for 2 weeks.
3.4.3 Standard series solution of malondialdehyde. accurately PIPETTE 0.10
mL, 0.50 mL, 1.0 mL, 1.5 mL, 2.5 mL of standard use solution of
malondialdehyde in 10 mL volumetric flasks; ADD trichloroacetic acid mixture
(3.2.2) to dilute to volume. The concentration of the series of standard solutions
is 0.01 μg/mL, 0.05 μg/mL, 0.10 μg/mL, 0.15 μg/mL, 0.25 μg/mL. Prepared
when used.
4 Instruments and equipment
4.1 High performance liquid chromatography. equipped with a diode array
detector.
4.2 Balance. The sensitivity is 0.0001 g, 0.01 g.
4.3 Thermostatic oscillator.
4.4 Thermostatic water bath.
to the standard curve, the concentration of malondialdehyde in the solution to
be tested is obtained. The liquid chromatogram of malondialdehyde standard is
shown in Appendix A.
6 Analysis results expression
The content of malondialdehyde in the sample shall be calculated according to
formula (1).
Where.
X - The content of malondialdehyde in the sample, in milligrams per kilogram
(mg/kg);
c - The concentration of malondialdehyde in the sample solution obtained from
the standard series curve, in micrograms per milliliter (μg/mL);
V - The constant volume of sample solution, in milliliters (mL);
m - The mass of sample represented by the final sample solution, in grams (g);
1000 - Conversion factor.
The calculation result is expressed as the arithmetic mean of the two
independent determination results obtained under repeated conditions. The
result retains two significant figures.
7 Precision
The absolute difference between the two independent determination results,
obtained under repeated conditions, shall not exceed 10% of the arithmetic
mean.
Method 2 Spectrophotometry
8 Principles
After the malondialdehyde is extracted by trichloroacetic acid solution, it reacts
with thiobarbituric acid (TBA) to form a pink compound. Its absorbance at 532
nm of wavelength is measured and compared with the standard series to
to dilute to volume. The concentration of the series of standard solutions is 0.01
μg/mL, 0.05 μg/mL, 0.10 μg/mL, 0.15 μg/mL, 0.25 μg/mL. Prepared when used.
10 Instruments and equipment
10.1 Spectrophotometer.
10.2 Balance. The sensitivity is 0.0001 g, 0.01 g.
10.3 Thermostatic oscillator.
10.4 Thermostatic water bath.
11 Analytical procedures
11.1 Sample preparation
WEIGH 5 g (accurate to 0.01 g) of the sample; PLACE it in a 100 mL conical
flask with stopper; accurately ADD 50 mL of trichloroacetic acid mixture; SHAKE
it well; and USE the stopper to seal. PLACE on a thermostatic oscillator to
shake at 50 °C for 30 min; REMOVE, and COOL to room temperature. USE
double-layer quantitative slow filter paper to filter; DISCARD the primary filtrate;
and the subsequent filtrate is standby.
Accurately PIPETTE 5 mL of the above filtrate and of standard series solution
respectively; PLACE them in 25 mL colorimetric tube with stopper. TAKE
another 5 mL of trichloroacetic acid mixture as a sample blank; ADD 5 mL of
aqueous solution of thiobarbituric acid (TBA) respectively; COVER the stopper;
and MIX well. PLACE in a 90 °C water bath to react for 30 min; TAKE out; and
COOL to room temperature.
11.2 Determination
USE the sample blank to adjust the zero point. At 1 cm optical path at a
wavelength of 532 nm, the absorbance of the sample solution and the standard
series solution (9.4.3) is determined. USE the mass concentration of the
standa...