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LC_PH0474
phase composed of two phenyl groups bonded
end-to-end, easily achieves retention capacities
similar to, and even greater than, those of a C18
column when used with a highly organic mobile
phase. For this reason, we evaluated the enhanced
retention of the Allure® Biphenyl column for
assaying NSAIDs through aromatic selectivity.
First, we compared the retention characteristics of
a conventional C18 column and an Allure®
Biphenyl column, using acetonitrile as the organic
modifier. As expected, the Allure® Biphenyl col-
umn exhibited similar retention under equivalent
analytical conditions (Figure 3). But, when we
assayed the same analytes, using methanol as the
organic modifier, we found retention on the
Allure® Biphenyl column was
greatly increased
. To
maintain the same retention capacities (k')
between the columns, we had to increase the
organic content by 20% (Figure 3). In addition,
selectivity between the two columns became dra-
matically different. Based on these results, we con-
clude that methanol in the mobile phase enhances
π
-
π
interactions between aromatic compounds
and the biphenyl stationary phase, leading to
greater retention and superior selectivity.
An Allure® Biphenyl column, in combination with
a methanol-containing mobile phase, significantly
improves separations of NSAIDs, or other aromat-
ic drug compounds. Increased retention capacity
creates a need for a higher percentage of organic
solvent in the mobile phase, to elute the analytes in
a timely manner. Increasing the organic content, in
turn, increases sensitivity in LC/MS methods,
because it optimizes the desolvation efficiency in
electrospray interfaces. And this, in turn, makes an
Allure® Biphenyl column the best choice for sepa-
rating aromatics.
Figure 3
The versatility of the Allure® Biphenyl phase makes
it a great alternative to conventional phenyl phase columns,
especially in method development.
Conditions:
Mobile phase:
0.5% formic acid in water (pH 2.25):0.1% formic acid in acetonitrile, 50:50 (v/v)
Flow:
1.0mL/min.
Temp.:
ambient
Det.:
UV @ 254nm
Column:
C18
Dimensions:
150mm x 4.6mm
Particle size:
5µm
Pore size:
100Å
1. sulindac
2. piroxicam
3. ketoprofen
4. diclofenac
B) C18, 50% acetonitrile
LC_PH00411
LC_PH00414
LC_PH00415
Sample:
Inj.:
5µL
Conc.:
~300µg/mL each component
Sample diluent: mobile phase
Conditions:
Mobile phase:
0.5% formic acid in water (pH 2.25):0.1% formic acid in methanol, 30:70 or 10:90 (v/v)
Flow:
1.0mL/min.
Temp.:
ambient
Det.:
UV @ 254nm
Column:
C18
Dimensions:
150mm x 4.6mm
Particle size:
5µm
Pore size:
100Å
Column:
Allure
®
Biphenyl (cat.# 9166565)
Dimensions:
150mm x 4.6mm
Particle size:
5µm
Pore size:
60Å
A) Allure® Biphenyl,
90% methanol
B) C18, 70% methanol
In methanol, retention capacity & selectivity of NSAIDs are much greater on an
Allure® Biphenyl column, compared to a C18 column, and
elution order changes.
In acetonitrile, retention of NSAIDs on an Allure® Biphenyl column is
comparable to retention on a C18 column and
elution order is the same.
0
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Column:
Allure
®
Biphenyl (cat.# 9166565)
Dimensions:
150mm x 4.6mm
Particle size:
5µm
Pore size:
60Å
A) Allure® Biphenyl, 50% acetonitrile
LC_PH00410
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1 2
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Allure® Biphenyl Columns (USP L11)
Physical Characteristics:
5µm Column, 4.6mm
cat. #
price
150mm
9166565
particle size: 5µm, spherical
pore size: 60Å
carbon load: 23%
endcap: yes
pH range: 2.5 to 7.5
temperature limit: 80°C
Allure® Guard Cartridges
Allure Biphenyl
qty.
cat. #
price
10 x 2.1mm
3-pk. 916650212
10 x 4.0mm
3-pk. 916650210
20 x 2.1mm
2-pk. 916650222
For other dimensions of these columns, visit our website at
www.restek.com