Page 52 - Application Handbook - Liquid Chromatography

LAAN-A-LC-E199

Application High Performance Liquid Chromatography

News High Speed, High Resolution Analysis (Part 38)

Analysis of Artificial Colorings with the Nexera
No.L421 UHPLC System

High-speed analysis of artificial colorings was previously Chromatography) System together with the
introduced in Application News No. L349. In this issue, Phenomenex Kinetex C18 high-speed, high-resolution
we present an example of ultra-high-speed analysis analytical column, developed with the goal of achieving
using the Nexera UHPLC (Ultra High Performance Liquid even greater analysis speeds.

n Analysis of 12 Artificial Colorings Table 1 Analytical Conditions
We conducted simultaneous analysis of 12 kinds of tar Column : Phenomenex Kinetex 2.6 μm C18 100 Å
synthetic dyes. The Phenomenex Kinetex-C18 (particle (50 mm L. × 3.0 mm I.D., 2.6 μm)
size 2.6 μm) was used for the separation. This is a Core- Mobile Phase : A; 10 mmol/L Ammonium acetate
Shell column consisting of a 1.9 μm solid core coated B; 10 mmol/L Ammonium acetate/Acetonitrile
= 1/1 (v/v)
with a bonded multilayer (0.35 μm thick) of porous Flowrate : 2.5 mL/min
silica gel micro particles. Monitoring was conducted Time Program : B Conc. 5 % (0.0 min) → 100 % (1.1-1.6 min)
→ 5 % (1.61-2.4 min)
using the maximum absorbance (MAX plot) from 400 • Mixer: 20 μL
to 700 nm with the SPD-M20A photodiode array Column Temp. : 40 °C
detector. Injection Volume : 1 μL
Fig. 1 shows the chromatogram of a standard mixture Detection : SPD-M20A Max Plot 400-700 nm
Flow Cell
: Semi-micro cell
of the 12 artificial colorings (each at 50 mg/L in
aqueous solution), in addition to a contour plot. Table 1 Table 2 Repeatability of 12 Artificial Colorings (n=6)
shows the analytical conditions.
The 12 dyes were separated within 1 minute using Peak Retention Time Peak Area
these analytical conditions. In addition, excellent No. %RSD %RSD
repeatability of retention time and peak area were 1 0.103 0.117
obtained for all of the components, using 6 consecutive 2 0.069 0.161
injections (1-μL injections), as shown in Table 2. 3 0.047 0.289
4 0.068 0.219
5 0.091 0.116
mAU 6 0.057 0.203
7 0.066 0.170
10
300 7 8 11 8 0.056 0.206
9 0.021 0.073
200 9 12 10 0.034 0.117
11 0.032 0.140
5 6
100 123 4 12 0.037 0.148
mAU
0.0
750
0.00 0.25 0.50 0.75 1.00 min
nm
300 500
400
250
500
600
0.0
700
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 min 0.00 0.25 0.50 0.75 1.00 min
■Peaks
1. Y4 (Tartrazine), 2. R2 (Amaranth), 3. B2 (Indigo Carmine),
4. R102 (New Coccine), 5. Y5 (Sunset Yellow FCF),
6. R40 (Allura Red AC), 7. G3 (Fast Green FCF),
8. B1 (Brilliant Blue FCF), 9. R3 (Erythrosine), 10. R106 (Acid Red),
11. R104 (Phloxine), 12. R105 (Rose Bengale)

Fig. 1 Chromatogram and Contour Plot of a Standard Mixture
of 12 Artificial Colorings (50 mg/L each)