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GB 5009.248-2016

Chinese Standard: 'GB 5009.248-2016'
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BASIC DATA
Standard ID GB 5009.248-2016 (GB5009.248-2016)
Description (Translated English) Determination of lutein in milk powder--HPLC-UV method
Sector / Industry National Standard
Classification of Chinese Standard X09
Word Count Estimation 8,876
Date of Issue 2016-08-31
Date of Implementation 2017-03-01
Older Standard (superseded by this standard) GB/T 23209-2008
Regulation (derived from) Announcement of the State Administration of Public Health and Family Planning 2016 No.11

GB 5009.248-2016
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard -
Determination of lutein in foods
食品安全国家标准
食品中叶黄素的测定
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... 6 
5 Analytical procedures ... 6 
6 Analysis results expression ... 9 
7 Precision ... 9 
8 Other ... 9 
Appendix A Method for correction of standard solution concentration ... 11 
Appendix B Liquid chromatogram of standard solution ... 12 
Foreword
This Standard replaces GB/T 23209-2008 “Determination of lutein in milk
powder - HPLC-UV method”.
As compared with GB/T 23209-2008, the main changes of this Standard are as
follows.
- CHANGE the standard’s name to “National Food Safety Standard -
Determination of lutein in foods”;
- EXPAND the scope of application; ADD the liquid chromatography for the
determination of lutein in frozen drinks, rice and flour products, bakery
products, jams, jellies, and beverages;
- MODIFY the “Principles” of the former standard. The former “USE acetone
as a solvent to extract” is changed to “USE diethyl ether-n-hexane-
cyclohexane solvent system to extract”;
- ADD the points worthy of notice for the operation process;
- ADD a saponification step for samples with high fat content including milk
powder; and PROVIDE different pretreatment methods to accommodate
various sample matrix analysis needs;
- ADD the correction requirement for the concentration of lutein standard
solution;
- For the phenomenon that lutein may be isomerized during the test operation,
ADD the qualitative and quantitative requirements for cis-lutein produced
due to isomerization.
National Food Safety Standard -
Determination of lutein in foods
1 Scope
This Standard specifies the liquid chromatography for the determination of
lutein in foods.
This Standard applies to liquid chromatography determination of lutein in infant
formula, dairy products, frozen drinks, rice and flour products, bakery products,
jams, jellies, and beverages.
2 Principles
After foods with high fat content (fat content of not less than 3% on a dry basis)
are saponified with potassium hydroxide solution at room temperature to free
lutein, then USE diethyl ether-n-hexane-cyclohexane (40+40+20, volume ratio)
to extract; USE liquid chromatography to separate; USE UV-detector or diode
array detector to detect; and USE external standard method for quantification.
Lutein in the samples of the other foods is directly extracted by diethyl ether-n-
hexane-cyclohexane (40+40+20, volume ratio). After the extract is purified by a
neutral alumina solid phase extraction cartridge, USE liquid chromatography to
separate; USE UV-detector or diode array detector to detect; and USE external
standard method for quantification.
During the extraction and analysis of sample, the trans-structured lutein may
be isomerized and converted to cis-lutein. The cis-lutein produced by the
transformation can be qualitatively determined by retention time and quantified
by peak area adduction.
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 Cyclohexane (C6H12). chromatographically pure.
cream, bakery nuts, etc.)
Accurately WEIGH 2 g (accurate to 0.01 g) of uniform sample in a 50 mL
polypropylene centrifuge tube; ADD about 0.2 g of BHT (3.1.6) and 10 mL of
ethanol (3.1.4), and MIX well. ADD 10 mL of 10% potassium hydroxide solution
(3.2.1); vortex oscillate for 1 min, and MIX well. Oscillate and saponify at room
temperature for 30 min in the dark; USE 10 mL of extraction solvent (3.2.3) to
vortex oscillate and extract in the dark for 3 min; CENTRIFUGE at 4500 r/min
for 3 min; repeatedly EXTRACT twice; and combine the extracts. USE 10 mL
of water to wash; CENTRIFUGE at 4500 r/min for 3 min for delamination; wash
once again. The organic phases are combined and concentrated under reduced
pressure at room temperature until nearly dry. USE 0.1% BHT ethanol solution
(3.2.4) to vortex oscillate to dissolve the residue and dilute to 5 mL; pass
through a 0.45 μm filter membrane (3.6), for liquid chromatography
determination.
Liquid milk. Accurately WEIGH 10 g (accurate to 0.01 g) of the sample in a 50
mL polypropylene centrifuge tube; ADD about 0.2 g of BHT (3.1.6) and 10 mL
of ethanol (3.1.4), and MIX well. ADD 2 mL of 20% potassium hydroxide solution
(3.2.2); vortex oscillate for 1 min, and MIX well. Oscillate and saponify at room
temperature for 30 min in the dark; USE 10 mL of extraction solvent (3.2.3) to
vortex oscillate and extract in the dark for 3 min; CENTRIFUGE at 4500 r/min
for 3 min; repeatedly EXTRACT twice; and combine the extracts. USE 10 mL
of water to wash; CENTRIFUGE at 4500 r/min for 3 min for delamination; wash
once again. The organic phases are combined and concentrated under reduced
pressure at room temperature until nearly dry. USE 0.1% BHT ethanol solution
(3.2.4) to vortex oscillate to dissolve the residue and dilute to 25 mL; pass
through a 0.45 μm filter membrane (3.6), for liquid chromatography
determination.
5.2.2 Other foods (such as rice, flour products, jams, etc.)
Accurately WEIGH 5 g (accurate to 0.01 g) of uniform sample in a 50 mL
polypropylene centrifuge tube. USE 10 mL of extraction solvent (3.2.3) to vortex
oscillate and extract in the dark for 3 min; CENTRIFUGE at 4500 r/min for 3
min; repeatedly EXTRACT twice. The extracts are combined and concentrated
under reduced pressure at room temperature until nearly dry. USE 3 mL of
extraction solvent (3.2.3) to vortex oscillate and dissolve; REPEAT the
operation once; combine the extraction solvents, and mix well, for purification.
The above solution, at a flow rate of about 1 mL/min, is passed through an
activated neutral alumina solid phase extraction cartridge (3.5). USE 3 mL of
extraction solvent (3.2.3) to elute. The effluent and eluent are combined and
concentrated under reduced pressure at room temperature until nearly dry.
USE 0.1% BHT ethanol solution (3.2.4) to vortex oscillate to dissolve the
mL of ethanol solution of iodine (3.2.5), shake well; and PLACE the mixture under
sunlight or fluorescent lamp for 30 min. Cis-structured lutein can be obtained. The
thus prepared lutein containing cis-structure, when testing, can be used as a
reference substance. The chromatogram of standard solution of trans-lutein
isomerized by light iodine is shown in Appendix B, Figure B.2.
6 Analysis results expression
The content of lutein in the sample shall be calculated according to formula (1).
Where.
X - The content of lutein in the sample, in micrograms per hundred grams
(μg/100 g);
c - The content of the standard in the sample solution obtained from the
standard curve, in micrograms per milliliter (μg/mL);
V - The final constant volume of sample, in milliliters (mL);
m - Sample weighing amount,......
Related standard:   GB 5009.241-2017  GB 5009.242-2017
Related PDF sample:   GB 5009.245-2016  GB 5009.241-2017
   
 
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