Rapid analysis of water-soluble vitamins using Smartline HPLC

Instrumentation
KNAUER Smartline HPLC system equipped with Autosampler 3900, Pump 1000 with 10 ml pump head, SmartMix
static mixer, Manager 5000 with low pressure gradient and degasser modules, Column Oven 4000 and UV
Detector 2600 with analytical flow cell.
Results
Since each vitamin has a slightly different optimal absorption wavelength (Fig. 1), a wavelength switching program
was used to increase the sensitivity of quantification for each vitamin. Figure 2 depicts the chromatogram obtained
for the micronutrient tablet sample. Analysis results are listed in Table 1.
- ascorbic acid
- nicotinamide
- pyridoxine
- riboflavin
- thiamine
200
|
220
|
240| 260| | | | |
210 230 250 270 280 290 300 310 320 330 340
nm
200 220 240 260 280 300 320 340 nm
Fig. 1 Wavelength spectrum of 5 water-soluble vitamins.
mAU
2000
1800
1600
1400
1200
1000
800
600
400
200
oxalic acid, λ = 246 nm
thiamine (vitamin B1), λ = 246 nm
ascorbic acid (vitamin C), λ =2 40 nm
nicotinamide (vitamin B3), λ = 260 nm
pyridoxine (vitamin B6), λ = 290 nm
riboflavin (vitamin B2), λ = 267 nm
0
-200
Fig. 2
0,0 0,5 1,0 1,5 2,0 2,5 3,0 3,5 4,0 4,5 5,0 5,5 6,0 6,5 7,0 7,5
Minutes
Separation of five vitamins in a micronutrient tablet.
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