Analysis of Additives in Soft Drink Using UHPLC (4-wavelengths UV absorption monitoring)

May 24, 2021


Food additives are substances added to food as preservatives, antioxidants, emulsifiers, etc. Recent increase in consumption of processed foods has introduced more additives. Securing food safety demands determination of these additives. High performance liquid chromatography (HPLC) has been used for the measurement of these food additives.

We examined the usefulness of an X-PressPak V-C (2.0 mm I.D. x 50 mm L.) packed with 2 µm diameter packing material for the ultra-high speed separation of food additives. We used a PDA detector for the simultaneous monitoring of these additives at four different wavelengths. The results were examined to determine whether the performance of the column and chromatography system exceeds those of conventional HPLC.


LC-4000 UHPLC system


Chromatographic conditions
Column: X-PressPak V-C18 (2.0 mmID x 50 mmL, 2 µm)
Column temperature: 40°C
Mobile phase A: 0.3% phosphoric acid
Mobile phase B: acetonitrile
Gradient (A/B): 0min(95/5) -> 1.3min(85/15) -> 1.7min(85/15) -> 5min(30/70) -> 5.1 min(10/90) -> 6 min(10/90) -> 6.1 min(95/5)
Injection interval: 8 min
Flow rate: 0.5 mL/min
Detection wavelengths: 210 nm, 240 nm, 270 nm, 300 nm
Injection volume: 1 µL

The method was applied to the analysis of soft drinks. The soft drink was purchased at a grocery store, diluted 5 times with a mixture of 0.3% phosphoric acid /acetonitrile (95/5), and then filtered with 0.2 µm membrane filter.


Figure 1 shows an UHPLC chromatogram of a standard mixture of 11 food additives. The UHPLC system provides 5 times shorter analysis time than conventional HPLC without sacrificing the resolution.

Figure 1. UHPLC chromatogram of a standard mixture of food additives (1=acesulfame K (0.2 mg/mL), 2=p-hydroxybenzoic acid (0.05 mg/mL), 3=caffeine (0.05 mg/mL), 4=aspartame (0.1 mg/mL), 5=vitamin B2 (0.1 mg/mL), 6=benzoic acid (0.05 mg/mL), 7=methylparaben (0.05 mg/mL), 8=propyl gallate (0.05 mg/mL), 9=ethylparaben (0.05 mg/mL), 10=propylparaben (0.05 mg/mL), 11=butylparaben (0.05 mg))

Figure 2 shows an UHPLC chromatogram of the soft drink sample.  Eleven food additives were clearly separated from unidentified peaks.

Figure 2. UHPLC chromatogram of the soft drink (The sample is diluted 5 times with 0.3% phosphoric acid/acetonitrile(95/5), and filtered with 0.2 µm membrane filter. The other conditions are the same as in the Figure 1 caption.)

About the Author

Chromatography Group