GB/T 23882-2009 PDF English
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Determination of L-Ascorbyl-2-monophosphate in feeds -- High performance liquid chromatography
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GB/T 23882-2009: PDF in English (GBT 23882-2009) GB/T 23882-2009
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
ICS 65.120
B 46
Determination of L-ascorbyl-2-monophosphate in feeds -
High performance liquid chromatography
ISSUED ON: MAY 26, 2009
IMPLEMENTED ON: OCTOBER 01, 2009
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 or materials ... 4
5 Instruments and equipment ... 5
6 Sampling ... 5
7 Sample preparation ... 6
8 Analysis steps ... 6
9 Calculation of results ... 7
10 Repeatability ... 7
Appendix A (Informative) Chromatograms of L-ascorbyl-2-monophosphate ... 8
Determination of L-ascorbyl-2-monophosphate in feeds -
High performance liquid chromatography
1 Scope
This Standard specifies the high performance liquid chromatography detection method
for L-ascorbyl-2-monophosphate in feeds.
This Standard applies to the determination of L-ascorbyl-2-monophosphate content in
compound feeds, concentrated feeds and additive premixed feed for aquatic products.
The detection-limit of the method is 2 mg/kg (calculated as L- ascorbic acid), and the
quantitation-limit is 10 mg/kg (calculated as L- ascorbic acid).
2 Normative references
The terms in the following documents become the terms of this Standard by reference
to this Standard. For dated references, all subsequent amendments (not including errata
content) or revisions do not apply to this standard. However, parties to agreements that
are based on this Standard are encouraged to study whether the latest versions of these
documents can be used. For undated references, the latest edition applies to this
Standard.
GB/T 6682, Water for analytical laboratory use - Specification and test methods
GB/T 14699.1, Feeding stuffs - Sampling
GB/T 20195, Animal feeding stuffs - Preparation of test samples
3 Principle
The L-ascorbyl-2-monophosphate in the sample shall be extracted with phosphate
buffer solution, centrifuged and filtered; the filtrate shall be measured with high
performance liquid chromatography, and quantified by external standard method.
4 Reagents or materials
Unless otherwise stated, in the analysis, only use reagents confirmed to be analytical
reagents AND water complying with the grade-II water specified in GB/T 6682.
4.1 Methanol: chromatographic pure.
7 Sample preparation
Prepare the sample according to GB/T 20195; grind it; pass it through a 0.45 mm mesh
sieve; mix it evenly; put it in an airtight container; store it at low temperature in the
dark for future use.
8 Analysis steps
8.1 Extraction
Weigh a certain amount of sample [containing about 1 mg of L-ascorbyl-2-
monophosphate (calculated as L- ascorbic acid)], accurate to 0.1 mg; put it in a 100 mL
volumetric flask; add about 70 mL of phosphate buffered saline (4.5); sonicate in an
ultrasonic water bath for 15 minutes; use phosphate buffered saline (4.5) to dilute to the
mark (V). Take an appropriate amount of solution and centrifuge at 3 500 r/min for 5
min. After centrifugation, filter the supernatant through a 0.45 μm filter membrane; use
the filtrate for high-performance liquid chromatography analysis.
8.2 Determination
8.2.1 Liquid chromatography reference conditions
Chromatographic column: C18 column, length 250 mm, inner diameter 4.6 mm, particle
size 5 μm, or equivalent performance.
Column temperature: room temperature.
Flow rate: 1.0 mL/min.
Detection wavelength: 250 nm.
Injection volume: 20 μL.
8.2.2 Drawing of the standard curve
Continuously inject L-ascorbyl-2-monophosphate standard working solution (4.8) into
the HPLC analyzer with a stable baseline, from low concentration to high; plot the peak
area-concentration to obtain the standard curve regression equation.
8.2.3 Quantitative determination
Inject the sample solution to be tested (8.1), in which the peak area response value of
L-ascorbyl-2-monophosphate shall be within the linear range of the standard curve; if
it exceeds the linear range, it shall be diluted and then injected for analysis. According
to the peak area, obtain the concentration (C) of L-ascorbyl-2-monophosphate in the
......
Source: Above contents are excerpted from the PDF -- translated/reviewed by: www.chinesestandard.net / Wayne Zheng et al.
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