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This standard replaces GB/T 22253-2008 Determination of Alitame in Foods and GB/T 22254-2008 Determination of Aspartame in Foods.
The following main changes have been made with respect to GB/T 22253-2008 and GB/T 22254-2008:
——the standard name is changed to "食品安全国家标准 食品中阿斯巴甜和阿力甜的测定 (National Food Safety Standard Determination of Aspartame and Alitame in Foods)";
——the application scope of the standard is added.
National Food Safety Standard
Determination of Aspartame and Alitame in Foods
1 Scope
This standard specifies the determination method of aspartame and alitame in foods.
This standard is applicable to the determination of aspartame and alitame in foods.
2 Theory
As aspartame and alitame are soluble in water, and polar solvents like methanol and ethanol are insoluble in fat-soluble solvents, extract the specimens of vegetable and its product, fruit and its product, edible mushroom, alga, cereal and its product, bakery product, baked food and jelly with methanol aqueous solution under ultrasonic oscillation; extract the specimens of fruitade, carbonated beverage, solid beverage, condiment and other candy (except for gum-based candy) with water; extract the specimens of milk product, milk beverage and frozen drink with ethanol aqueous solution after precipitating the protein with ethanol; extract the specimens of gum-based candy with water after dissolving the gum base with n-hexane; extract the specimens of fat emulsified product, cocoa product, chocolate and its product, nut, seed, aquatic product and its product and egg product with water and then degrease them with n-hexane. Separate the above-mentioned extract on liquid-phase chromatographic C18 reversed-phase column and detect at place with wavelength of 200nm, conduct qualitative analysis according to the retention time of chromatographic peak, and then quantitate them by external standard method.
3 Reagents and Materials
Unless otherwise specified, analytically pure reagents and laboratory Grade 1 water (specified in GB/T 6682) shall be adopted.
3.1 Reagents
3.1.1 Methanol (CH3OH): chromatographically pure.
3.1.2 Ethanol (CH3CH2OH): guaranteed reagent.
3.2 Standards
3.2.1 Alitame standard (C14H25N3O4S, CAS No.: 80863-62-3): purity ≥99%.
3.2.2 Aspartame standard (C14H18N2O5, CAS No.: 22839-47-0): purity ≥99%.
3.3 Preparation of standard solutions
3.3.1 The standard stock solutions (0.5mg/mL) of aspartame and alitame: respectively weigh 0.025g (to the nearest 0.0001g) of aspartame and alitame, dissolve them with water and transfer the solutions into two 50mL volumetric flasks, then dilute the solutions to the scale, and store the solutions in the refrigerator at about 4℃, with the validity period of 90d.
3.3.2 Preparation of mixed standard working solution series of aspartame and alitame: dilute the standard stock solutions of aspartame and alitame to the mixed standard series with water step by step to make the standard working solution series with the concentration of aspartame and alitame reach 100μg/mL, 50μg/mL, 25μg/mL, 10.0μg/mL and 5.0μg/mL respectively, and store them in the refrigerator at about 4℃, with the validity period 30d.
4 Apparatuses
4.1 Liquid chromatograph: equipped with diode array detector or ultraviolet detector.
4.2 Ultrasonic oscillator.
4.3 Balance: with a sensibility of 1mg and 0.1mg.
4.4 Centrifuge: rotating speed ≥4,000r/min.
5 Analytical Procedures
5.1 Specimen preparation and pretreatment
5.1.1 Carbonated beverage, fruitade, solid beverage, table condiment and other candy (except for gum-base candy)
Weigh about 5g (to the nearest 0.001g) of carbonated beverage specimen into a 50mL beaker, remove carbon dioxide in water bath at 50℃, completely transfer the specimen into a 25mL volumetric flask for subsequent use; weigh about 2g (to the nearest 0.001g) of fruitade specimen into a 25mL volumetric flask for subsequent use; weigh about 1g (to the nearest 0.001g) of solid beverage or condiment or ground candy specimen into a 50mL beaker, add 10mL of water, extract the solution under ultrasonic vibration for 20min, then transfer the extract into a 25mL volumetric flask, add another 10mL of water into the beaker, extract the solution under ultrasonic vibration for 10min, then transfer the extract into the same 25mL volumetric flask for subsequent use. Dilute the solutions in the above-mentioned volumetric flasks with water, mix well, centrifuge for 5min at 4,000r/min, and filter the supernatant through 0.45μm hydrophilic membrane for chromatographic analysis.
5.1.2 Milk product, milk beverage and frozen drink
Liquid milk product containing solid pulp shall be subjected to homogenate in a food processor, while solid milk product like cheese shall be subjected to homogenate according to the mass ratio (1:4) of specimen to water in a food processor.
Respectively weigh about 5g (to the nearest 0.001g) of homogenate specimens of liquid milk product, milk beverage, frozen drink and solid milk product into a 50mL centrifuge tube, add 10mL of ethanol and put on the stopper; as for specimens of milk beverage and frozen drink, gently turn centrifuge tube upside down (no shaking) for 5 times at first; as for milk product, well mix the solution in centrifuge tube by vortexing it for 10s at first, then keep it still for 1min, centrifuge it for 5min at 4,000r/min, filter the supernatant into a 25mL volumetric flask, wash the precipitate with 8mL of ethanol-water (2+1), after centrifugation, transfer the supernatant into the same 25mL volumetric flask, dilute the solution with ethanol-water (2+1), and filter the supernatant through 0.45μm organic membrane for chromatographic analysis.
5.1.3 Jelly
Well mix the sucking jelly and transparent jelly with glass rod; conduct homogenate to fruit pulp jelly in a food processor.
Weigh about 5g (to the nearest 0.001g) of jelly specimen prepared uniformly into a 50mL colorimetric tube, add 25mL of 80% methanol aqueous solution, heat it in water bath at 70℃ for 10min, take out the colorimetric tube, transfer the extract into a 50mL volumetric flask while it is hot, wash the colorimetric tube in twice with 15mL of 80% methanol aqueous solution, shake the tube for about 10s at each time, transfer the solution into the same 50mL volumetric flask, cool to room temperature, then dilute it with 80% methanol aqueous solution to scale, mix well, centrifuge it for 5min at 4,000r/min, and filter the supernatant through 0.45μm organic membrane for chromatographic analysis.
5.1.4 Vegetable and its product, fruit and its product, edible mushroom and alga
As for the specimen of fruit and its product, the pit (if any) shall be removed first.
As for relatively dry and hard specimens, carry out homogenate in a food processor according to the ratio (1:4) of specimen to water mass, weigh about 5g (to the nearest 0.001g) of homogenate specimen into a 25mL centrifuge tube, add 10mL of 70% methanol aqueous solution, shake well, carry out ultrasound treatment for 10min, centrifuge it for 5min at 4,000r/min, transfer the supernatant into a 25mL volumetric flask, add 8mL of 50% methanol aqueous solution and repeat the operation once, then transfer the supernatant into the same 25mL volumetric flask, dilute it with 50% methanol aqueous solution, and filter the supernatant through 0.45μm organic membrane for chromatographic analysis.
As for relatively sticky and soft specimens with high sugar content, carry out homogenate in a food processor according to the ratio (1:2) of specimen to water mass, weigh about 3g (to the nearest 0.001g) of homogenate specimen into a 25mL centrifuge tube; as for other specimens, carry out homogenate in a food processor according to the ratio (1:1) of specimen to water mass, weigh about 2g (to the nearest 0.001g) of homogenate specimen into a 25mL centrifuge tube, then add 10mL of 60% methanol aqueous solution, shake well, carry out ultrasound treatment for 10min, centrifuge it for 5min at 4,000r/min, transfer the supernatant into a 25mL volumetric flask, add 10mL of 50% methanol aqueous solution and repeat the operation once, then transfer the supernatant into the same 25mL volumetric flask, dilute it with 50% methanol aqueous solution, and filter the supernatant through 0.45μm organic membrane for chromatographic analysis.
Foreword I
1 Scope
2 Theory
3 Reagents and Materials
4 Apparatuses
5 Analytical Procedures
6 Expression of Analysis Results
7 Precision
8 Others
Annex A Chromatograms
