GB/T 28717-2012 English PDF (GBT28717-2012)

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GB/T 28717-2012: Determination of malonaldehyde in feed -- High performance liquid chromatography
GB/T 28717-2012
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
ICS 65.120
B 46
Determination of Malonaldehyde in Feed - High
Performance Liquid Chromatography
ISSUED ON: SEPTEMBER 03, 2012
IMPLEMENTED ON: FEBRUARY 01, 2013
Issued by: General Administration of Quality Supervision, Inspection and
Quarantine;
Standardization Administration of the People’s Republic of China.
Table of Contents
Foreword ... 3
1 Scope ... 4
2 Normative References ... 4
3 Principle ... 4
4 Reagents ... 4
5 Apparatus ... 5
6 Preparation of Specimen ... 6
7 Analysis Procedure ... 6
8 Calculation and Presentation of Results ... 7
9 Repeatability ... 7
Appendix A (Informative) Chromatogram of Malondialdehyde and Thiobarbituric Acid
Complex ... 8
Determination of Malonaldehyde in Feed - High
Performance Liquid Chromatography
1 Scope
This Standard specifies the high-performance liquid chromatography detection method for
malondialdehyde in feed.
This Standard is applicable to the determination of malondialdehyde in compound feed,
concentrated feed and feed raw materials.
The detection limit of this Standard method is 0.015mg/kg, and the quantification limit is
0.05mg/kg.
2 Normative References
The following documents are essential to the application of this Document. For the dated
documents, only the versions with the dates indicated are applicable to this Document; for the
undated documents, only the latest version (including all the amendments) is applicable to this
Document.
GB/T 6682 Water for analytical laboratory use - Specification and test methods
GB/T 14699.1 Feeding Stuffs – Sampling
3 Principle
After malondialdehyde in the sample is extracted by trichloroacetic acid, react with
thiobarbituric acid (TBA) to form a pink complex. The supernatant is collected by
centrifugation; and detect by high-performance liquid chromatography and fluorescence
detector. Use external standard method to calculate the content of malondialdehyde.
4 Reagents
Unless otherwise specified, only reagents confirmed to be of analytical grade and Class-I water
specified in GB/T 6682 are used in the analysis.
6 Preparation of Specimen
According to the provisions of GB/T 14699.1, take at least 500g of representative feed samples;
reduce by at least 100g through quartering method; grind them; pass through a 0.9mm aperture
sample sieve; mix them evenly and put them in an airtight container; and store them at low
temperature in the dark for later use.
7 Analysis Procedure
7.1 Extraction
Weigh about 2g of specimen (weigh about 1g of feed oil, accurate to 0.001g); put it into a
250mL conical flask; accurately add 50mL of trichloroacetic acid mixture solution (4.4); shake
at 180r/min for 30min; take about 20mL of extract into a 50mL centrifuge tube and centrifuge
at 5000r/min for 5min.
7.2 Derivatization
Accurately pipette 5mL of the supernatant of the above specimen solution (7.1) and 5mL of
malondialdehyde standard series solution (4.7) into 25.0mL colorimetric tubes; add 5.0mL of
TBA solution (4.3); mix well; and place in a 90°C water bath to keep warm for 20 min. Take it
out; cool it quickly in an ice-water bath; transfer it to a centrifuge tube; centrifuge at 12000r/min
for 5 min. Take the supernatant and measure it on a computer; and the malondialdehyde
standard series and specimens are derivatized simultaneously.
7.3 Determination
7.3.1 Liquid chromatography reference conditions
7.3.1.1 Chromatographic column: C18 column, 250mm long, 4.6mm inner diameter, 5μm
particle size, or equivalent performance.
7.3.1.2 Column temperature: 35°C.
7.3.1.3 Mobile phase: Acetonitrile (4.1) + potassium dihydrogen phosphate solution (4.2) = 18
+ 82 (V + V); pass through a 0.45μm filter membrane before use.
7.3.1.4 Flow rate: 1.0mL/min.
7.3.1.5 Sample-injection volume: 10μL.
7.3.1.6 Excitation wavelength: 525nm.
7.3.1.7 Emission wavelength: 560nm.