Page 40 - BR20828_E Accucore 0713S

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Figure 1: Comparison of the resolution of a critical pair (peaks 2 and 3) on 5 µm fully porous C18 and 4 µm Accucore XL C18 HPLC
columns. Experimental conditions: columns – 150 x 4.6 mm; mobile phase – acetonitrile / methanol (80:20 v/v); flow rate – 1 mL/min;
column temperature – 30 °C; UV detection - 280 nm; injection volume – 5 μL
Resolution Comparison in Isocratic Mobile
Phase Conditions
Figure 1 illustrates the separation of six fat soluble
vitamins under isocratic mobile phase conditions.
On the Accucore XL C18 4 µm HPLC column, the
resolution is greater than or equal to 2.5 for all
compounds. Resolution of the critical pair (vitamin
D2 and D3) increased by 30% to 2.5 when using the
Accucore XL C18 4 µm HPLC column compared to
1.92 for the 5 µm fully porous C18 column. This is a
result of the improved efficiency provided by the
Accucore XL HPLC column, which is visually evident
from the peak widths in the chromatograms. Table 1
shows that the Accucore XL HPLC column almost
doubles the efficiency in all cases compared to the fully
porous material. This was achieved under the same
chromatographic conditions, with no changes in system
configuration and with only a small backpressure
increase (from 47 bar with the 5 µm fully porous column
to 62 bar for the 4 µm Accucore XL HPLC column).
mAU
Time (min)
-20
0
300
50
100
150
200
250
0
2
1
2 3
4 5
6
4 6 1 3 5 7 9 11 8 10 12 13
Rs = 2.50 N = 26179 plates
1 Vitamin K2
2 Vitamin D2
3 Vitamin D3
4 Vitamin E
5 Vitamin E acetate
6 Vitamin K1
Table 1: Efficiency values for the six fat soluble vitamins
mAU
Time (min)
-20
0
300
50
100
150
200
250
0
2
1
2 3
4 5
6
4 6 1 3 5 7 9 11 8 10 12 13
Rs = 1.96 N = 12776 plates
Fully porous C18 5 μm
Accucore XL C18 4μm
Efficiency (USP)
Accucore XL 4 μm Fully porous 5 μm
Vitamin K2 23826 13599
Vitamin D2 25566 13963
Vitamin D3 25710 13985
Vitamin E 22788 13288
Vitamin E acetate 24568 13880
Vitamin K1 26179 12776