Page 42 - BR20828_E Accucore 0713S

4
Productivity Comparison
The high efficiencies of the Accucore XL 4 µm HPLC
column over a wide flow rate range can be used to
reduce analysis time by optimizing flow rate and
gradient conditions. The van Deemter curve for the
Accucore XL 4 μm HPLC column is very flat at high
flow rates [1]. Therefore, a wide range of or flow rates
can be used without losing chromatographic
performance. In Figure 5, a reduction of run time
by half is demonstrated (Figures 5a and 5d),
simultaneously improving the resolution of the critical
pair when using the Accucore XL HPLC column
compared to the fully porous 5 µm column. When
running the fully porous 5 µm and the Accucore XL
4 µm HPLC column under the same conditions, the
retention time of the last eluting peak reduces from 8.62
to 6.56 min, respectively (Figures 5a and 5b), which can
be attributed to the lower column volume of the solid
core material and lower carbon load of the C18 phase.
From the van Deemter curve for a 4.6 mm ID
Accucore XL 4 μm HPLC column, the flow rate that
provides the highest efficiency is 1.3 mL/min [1]. When
increasing the flow rate, the gradient time needs to
be adjusted to keep the same gradient through the
column. The original 10 min gradient was reduced to
7.5 min (scaled gradient), which enabled a reduction
in the retention time of the last peak from 8.62 to 5.07
min (Figures 5a and 5c). The resolution of the critical
pair is still >3 under the scaled gradient conditions.
Optimizing the gradient by making it faster (4 min)
enables a reduction in analysis time to just over 4 min
(Figure 5d).
Figure 4. Comparison of the resolution of a critical pair (peaks 4 and 5) on 5 µm fully porous C8 and 4 µm Accucore XL C8 HPLC columns.
Experimental conditions: columns – 150 x 4.6 mm; mobile phase A – water + 0.1% formic acid; mobile phase B – methanol + 0.1%
formic acid; gradient: 20% to 50% in 15 min; flow rate – 1 mL/min; column temperature – 25 °C; UV detection – 280 nm; injection
volume – 5 μL
mAU
Minutes
1 0 2 3 4 5 6 7 8 9 10 11 12
1
2
3
4
5
6
7
4
5
Rs = 1.17
Rs = 2.68
1
2
3
6
7
Table 2: Peak widths for the seven catechins
Peak width (5% height)
Accucore XL 4 μm Fully porous 5 μm
Epigallocatechin 0.094 0.157
Catechin 0.101 0.173
Epigallocatechin gallate 0.132 0.201
Epicatechin 0.124
Partial co-elution
Gallocatechin gallate 0.149
Epicatechin gallate 0.151 0.228
Catechin gallate 0.158 0.227
Accucore XL C8, 4 µm
Fully porous C8, 5 µm 1.0 mL/min
∆P = 182 bar
PW0.05 = 0.197 min
1.0 mL/min
∆P = 241 bar
PW0.05 = 0.130 min 1 Epigallocatechin
2 Catechin
3 Epigallocatechin gallate
4 Epicatechin
5 Gallocatechin gallate
6 Epicatechin gallate
7 Catechin gallate