This standard supersedes Method for the Determination of Neotame in Foods - High Performance Liquid Chromatography (GB/T 23378-2009).
Compared with GB/T 23378-2009, this standard has the following main changes:
- the standard name was revised as "National Food Safety Standard - Determination of Neotame in Foods".
- the application scope was changed to the determination of neotame in beverage, preserved fruit, pastry, roasted seeds and nuts, pickled vegetable, candy, jam, jelly and compound flavouring.
National Food Safety Standard
Determination of Neotame in Foods
食品安全國家標準
食品中紐甜的測定
1 Scope
This standard specifies the high performance liquid chromatography for the determination of neotame content in foods.
This standard is applicable to the determination of neotame in foods such as beverage, preserved fruit, pastry, roasted seeds and nuts, pickled vegetable, candy, jam, jelly and compound flavouring.
2 Principle
Specimen is determined with high performance liquid chromatograph after it is extracted with mixed extracting solution and purified with solid-phase extraction column. Qualitative analysis is conducted by retention time and quantitative analysis is conducted by external standard method of peak area.
3 Reagents and Materials
Unless otherwise specified, the reagents adopted in this method are all analytically pure, and the water is Grade 1 water specified in GB/T 6682.
3.1 Reagents
3.1.1 Acetonitrile (CH3CN): chromatographically pure.
3.1.2 Sodium 1-octanesulfonate (C8H17NaO3S): chromatographically pure.
3.1.3 Phosphoric acid (H3PO4).
3.1.4 Formic acid (CH3COOH): chromatographically pure.
3.1.5 Methanol (CH3OH): chromatographically pure.
3.1.6 Triethylamine (C6H15N): chromatographically pure.
3.2 Reagent preparation
3.2.1 Mixed extracting solution: pipet 0.8mL of formic acid and 2.5mL of triethylamine respectively and add water to bring the volume to 1 000mL to make the pH of about 4.5.
3.2.2 Ion pair reagent buffer solution: weigh 2.00g of sodium 1-octanesulfonate and dissolve it with 500mL of water, then add 1.0mL of phosphoric acid and add water to bring the volume to 1 000mL.
3.3 Standard substance
Neotame (C20H30N2O5, CAS No.: 165450-17-9), with purity greater than or equal to 99.0%.
3.4 Preparation of standard solution
3.4.1 Standard stock solution: accurately weigh 0.100 0g of neotame standard substance, dissolve in mixed extracting solution and bring the volume to 100ml; the neotame content in this solution is 1.00mg/mL.
3.4.2 Standard working solution: respectively pipet appropriate amount of neotame standard stock solution to prepare series of standard working solutions in concentrations of 0.2μg/mL, 1.0μg/mL, 5.0μg/mL, 10.0μg/mL ,50.0μg/mL and 100.0μg/mL with mixed extracting solution.
3.5 C18 solid-phase extraction column: 6mL, 500mg or equivalent; activate with 5mL of methanol and 10mL of water successively prior to use.
3.6 0.45μm filter membrane, organic system.
4 Instruments and Apparatus
4.1 Liquid chromatograph: equipped with ultraviolet detector or diode array detector.
4.2 Ultrasonic cleaner.
4.3 Analytical balance: with sensibility of 0.000 1g and 0.01g.
4.4 Tissue blender.
4.5 Swirl oscillator.
4.6 Pressured gas blowing concentrator.
4.7 Solid-phase extraction device.
4.8 Centrifuge: with rotation speed greater than or equal to 4 000r/m.
5 Analytical Procedures
5.1 Specimen preparation
5.1.1 Solid sample
Weigh 10g (to the nearest of 0.01g) evenly pulverized specimen and put it into a 50mL plastic centrifuge tube with stopper, add 30mL of mixed extracting solution, oscillate for 10min in swirling way, treat is for 30min with ultrasound, and then bring the volume to the scale with mixed extracting solution; if the solution is turbid, centrifuge for 10min at the speed of no less than 4 000r/min and filter for use.
5.1.2 Liquid sample
Accurately measure 10.0mL of specimen and put it into a 50mL plastic centrifuge tube with stopper, add 30mL of mixed extracting solution, oscillate and mix it uniformly. After 15min ultrasonic treatment, bring the volume to the scale with mixed extracting solution; if the solution is turbid, centrifuge for 10min at the speed of no less than 4 000r/min and filter for use.
Note: For the sample containing gas, such as carbonated beverage and steam-water, subject it to tepidity and stirring to remove the carbon dioxide or carry out ultrasonic degasification firstly, then accurately weigh specimen.
5.2 Specimen purification
Pipet 10.0mL of filtered solution and make it pass through solid-phase extraction column at the flow speed of 1mL/min~2mL/min, after the filtrate flows out entirely, rinse the extraction column with 5mL of mixed extracting solution at the flow speed of 1mL/min~2mL/min, discard all effluent, elute with 5mL of methanol at the flow speed of 1mL/min, concentrate the eluent with pressured gas blowing concentrator at 40℃ water bath, bring the volume to 2.0mL with mixed extracting solution, filter via 0.45μm filter membrane to serve as to-be-determined solution for the analysis with liquid chromatograph.
Foreword i
1 Scope
2 Principle
3 Reagents and Materials
4 Instruments and Apparatus
5 Analytical Procedures
6 Expression of Analysis Results
7 Precision
8 Others
Appendix A Chromatogram
