Determination of Acesulfame in Carbonated Beverages by Wayeal High Performance Liquid Chromatography

Acesulfame is a food additive with the chemical name acesulfame potassium, also known as A-K sugar. It appears as a white crystalline powder and is an organic synthetic salt. Its taste is similar to that of cane sugar. It is readily soluble in water and slightly soluble in alcohol. It is the fourth generation of synthetic sweeteners currently used in the world.

Acesulfame is one of the six approved artificial sweeteners. It is 200 times sweeter than sugar but has zero calories. Acesulfame is defined as a sweetener in the national standard of the People's Republic of China, the National Food Safety Standard — Standard for the Use of Food Additives (GB 2760-2024). The standard stipulates strict regulations on its maximum usage levels and the names of the food products to which it may be added.

Acesulfame is a chemically synthesized sugar substitute sweetener. Compared to sodium cyclamate, it is less harmful to the human body. However, excessive intake of acesulfame may still have adverse effects on the gastrointestinal tract, metabolism, and sweetness dependence.

In this experiment, the content of acesulfame in a certain carbonated beverage was determined using Wayeal's high performance liquid chromatograph LC3400 series equipped with a UV detector, in accordance with GB 5009.140-2023 "National Food Safety Standard — Determination of Acesulfame Potassium in Foods".

Keywords:Acesulfame, Sweetener, High-performance Liquid Chromatography.

1. Instrument and Reagents

1.1 Configuration List of High-performance Liquid Chromatograph

Table 1 Table of Instrument Configuration

1.2 Reagents and Standards

Table 2 List of Reagents and Standards

1.3 Experiment Material and Auxiliary Equipment

Analytical balance;

Ultrasonic cleaner;

Vortex mixer;

Centrifuge.

2. Experiment Method

2.1 Reagents Preparation

2.1.1 Standard Working Solutions: Precisely transfer 1.0mL of acesulfame standard solution (in water) into a 10mL volumetric flask. Add an appropriate amount of water to dissolve, dilute to the mark, and mix well to obtain the standard stock solution (100mg/L). Precisely transfer appropriate volumes of the standard stock solution into separate volumetric flasks of different capacities, dilute to the mark with water, and mix well to obtain standard working solutions at concentrations of 0.2mg/L, 1.0mg/L, 5.0mg/L, 10.0mg/L, 20.0mg/L, 50.0mg/L, and 100.0mg/L.

2.1.2 Test Solution: Accurately weigh approximately 5g of the test sample (degas the sample by ultrasonication for 5 minutes before use) into a 50mL stoppered centrifuge tube. Add approximately 20mL of water, vortex to mix, ultrasonicate for 20 minutes, and centrifuge at 10,000r/min for 5 minutes. Transfer the supernatant to a 50mL volumetric flask. Add another 20mL of water to the centrifuge tube, mix well, ultrasonicate for 5 minutes, and centrifuge at 10,000r/min for 5 minutes. Combine the supernatant, dilute to the mark with water, and mix well. Transfer 5.00mL of the extract into an activated neutral alumina solid-phase extraction (SPE) column, and collect the eluate. Wash the SPE column twice with 2 mL of eluent each time, collect the eluate, combine the three eluates, and dilute to 10mL with the eluent, then mix well. Filter through a 0.45μm microporous membrane.

3. Experiment Result

3.1 Linearity and Range

Table 3 Standard Curve Concentration Table

Fig1 Acesulfame Standard Curve

The experimental results show that within the concentration range of 0.2 to 100.0mg/L for acesulfame, the linear correlation coefficient is greater than 0.9999, indicating good linearity.

3.2 Precision

Fig 2 Test Overlap Chromatogram of Acesulfame Standards (10.0mg/L)

Table 4 Test Data of Acesulfame Standards (10.0mg/L)

The experimental results show that for six consecutive injections of the 10.0mg/L standard solution, the repeatability RSD of retention time is 0.020%, and the repeatability RSD of peak area is 0.316%, indicating good instrument precision.

3.3 Sample Test

Fig 3 Overlaid Chromatograms of a Certain Carbonated Beverage Test Solution

Calculated by the external standard method, the content of acesulfame in a certain carbonated beverage is 0.0174%.

4. Conclusion

In this experiment, the content of acesulfame in a certain carbonated beverage was determined using Wayeal's high performance liquid chromatograph LC3400 series equipped with a UV detector, in accordance with GB 5009.140-2023 "National Food Safety Standard — Determination of Acesulfame Potassium in Foods". The experimental results showed that for six consecutive injections of the 10.0mg/L standard solution, the repeatability RSD of retention time was 0.020%, and the repeatability RSD of peak area was 0.316%, indicating good instrument precision. Within the concentration range of 0.2 to 100.0mg/L, the linear correlation coefficient was greater than 0.9999, demonstrating good linearity. Acesulfame was detected in a certain carbonated beverage with a content of 0.0174%. All the above data meet the instrument requirements specified in the standard method.