2.3. Determination of Acesulfame Potassium, Sodium Saccharin Dihydrate and Aspartame

The determination of acesulfame potassium, sodium saccharin dihydrate and aspartame has been carried out according to HRN EN 12856:2000 [33].

The acetonitrile and methanol, HPLC grade, as well as potassium dihydrogen phosphate, were purchased from Merck Co. (Darmstadt, Germany). The standards of acesulfame potassium and aspartame were bought from Sigma Aldrich (St. Louis, MO, USA), while sodium saccharin dihydrate was purchased from Merck Co. (Darmstadt, Germany).

Deionized water (0.05 mS cm⁻¹) was prepared from the ultrapure water system Omnia Pure (Stakpure GmbH, Niederahr, Germany). Standard stock solutions of acesulfame potassium, sodium saccharin dihydrate and aspartame (1000 mg L⁻¹) were prepared by dissolving in deionized water; working solutions were prepared by serial dilution of the stock solution.

Carbonated samples were degassed before preparation (15 min at room temperature) in an ultrasonic bath Sonis 4GT (Iskra, Ljubljana, Slovenia). In total, 20 mL of the homogenized sample was diluted to a volume of 100 mL with deionized water and filtered through a 0.45 μm syringe filter (Labex Ltd., Budapest, Hungary). Prepared samples were immediately analyzed on HPLC.

The analysis was carried out on the Agilent 1200 HPLC system (Agilent Technologies Singapore (International) Pte. Ltd., Singapore) equipped with a diode array detector (190–400 nm) and OpenLab CDS Chemstation Software (Agilent, Singapore, rev. B.04.03) and the Agilent 1260 HPLC system (Agilent Technologies Singapore (International) Pte. Ltd., Singapore) equipped with a diode array detector (190–900 nm) and OpenLab CDS Chemstation Software (Agilent, Singapore, rev.C.01.05). For the separation, an Agilent Zorbax reversed-phase HPLC column (250 × 4.6 mm; 5 µm) was used with a flow of 1.0 mL min⁻¹ and an injection volume of 20 μL. The mobile phase was 0.05 M solution of potassium dihydrogen phosphate (A) and acetonitrile (B); (90:10, v/v, gradient elution—0 min: 90% A:10% B/10 min: 60% A:40% B). Potassium dihydrogen phosphate (0.05 M) was prepared by dissolving in deionized water; the pH was adjusted to 3 with 5% phosphoric acid. The maximum wavelength was 210 nm for aspartame and sodium saccharin dihydrate and 226 nm for acesulfame potassium.

The limit of quantitation (LOQ) was 2.5 mg L⁻¹ for all three analytes. The mean recovery was ≥ 95.0% with a linearity (R²) of 0.999 for all analytes. All samples were analyzed in duplicate, with an RSD < 5%. The results are reported as the mean value.