Introduction
Vitamins are essential nutrients to all living organisms and are organic compounds required in small amounts for healthy physiological activity. The class of fat-soluble vitamins is insoluble in water but readily dissolve in oil and thus when consumed in excess may have a detrimental impact on the human body, as they are difficult to excrete.
This application note describes the analysis of fat-soluble vitamins using UHPLC and PDA detection with a high speed data sampling rate of 100 spectra/sec.
Experimental
Equipment
| Pump: | X-LC 3185PU x 2 |
| Degasser: | X-LC 3080DG |
| Mixer: | X-LC 3180MX |
| Column Oven: | X-LC 3067CO |
| Autosampler: | X-LC 3159AS |
| Detector: | X-LC 3110MD |
Conditions
| Column: | ZORBAX SB-C18 (3.0 mmID x 50 mmL, 1.8 µm) |
| Eluent: | Acetonitrile/Methanol (70/30) |
| Flow Rate: | 0.8 mL/min |
| Column Temp.: | 30 ºC |
| Wave Length: | 200-500 nm |
| Injection Volume: | 1 µL |
| Standard Sample: | 10 fat-soluble vitamin standards |
Keywords
UHPLC, Fat-soluble vitamin, Vitamin K3, Vitamin A, Retinol acetate, Vitamin K2, d-tocopherol, Vitamin D2, Vitamin D3, a-tocopherol, Tocopherol-a-acetate, Vitamin K1, C18 Column, PDF detector
Results
A contour plot and chromatogram of the fat-soluble vitamin standard mixture are shown in Figure 1. Excellent separation of the 10 vitamins was obtained in under 4 minutes.
The on-peak spectra of fat-soluble vitamin standard mixture are shown in Fig. 2.
| 1: Vitamin K3 (Menadione) | 5: Vitamin E ( -Tocopherol) | 9: Vitamin E acetate |
| 2:Vitamin A (Retinol) | 6: Vitamin D2 (Ergocalciferol) | 10: Vitamin K1 (Phytonadione) (0.1 mg/mL each) |
| 3: Vitamin A Acetate (Retinol acetate) | 7: Vitamin D3 (Cholecalciferol) | |
| 4: Vitamin K2 (Menaquinone) | 8: Vitamin E ( -Tocopherol) |