Agriculture Reference
In-Depth Information
hardware with higher pressure limits for using even smaller particles in combination
with longer columns. Based on the theories of van Deemter et al., then Giddings, and
finallyKnox, the use of small particles is one of the best solutions in the quest to improve
chromatographic performance [28]. Over the last three decades, the size of standard
high-ef
ciency particles has decreased from the 5
-
10
μ
m range in the 1980s to the
3
-
5
μ
m range in the 1990s and to the 1
-
3
μ
m range more recently for use in ultrahigh-
ef
ciency separations [29].
However, small particles induce a high pressure drop because, based on Darcy
s
law, the pressure drop is inversely proportional to the square of particle size at the
optimum linear velocity. The traditional 400 bar pump systems have been
the standard instrumentation since the 1970s [29]. In 2004, Waters introduced
the ACQUITY UPLC
TM
System. A new ultrahigh pressure/performance era began
with this launch. Most LC vendors have identi
'
ed HPLC systems as
ultrahigh-pressure liquid chromatography (UHPLC) systems. UHPLC systems can
reliably deliver solvents up to 1300 bar with routine operation in the 500
ed their modi
-
1000
bar range.
Compared with a traditional HPLC system, a UHPLC system has much lower
extracolumn variance ranging from 4 to 9
l
2
. The HPLC system can contribute
μ
40
-
∼
l
2
. In order to maximize sub-2
200
μ
μ
m column ef
ciency, the UHPLC system should
be well con
gured and operated in the optimized conditions such as using a smaller
volume needle seat capillary, narrower and shorter connector capillary tubes, and
a smaller volume detector cell. Otherwise, the column ef
ciency can lose as much
as 60% [30].
The sub-2
m particle columns and UHPLC systems offer much shorter analysis
times with higher resolution and greatly reduced solvent consumption. Some
separations can be performed within minutes or even seconds using UHPLC
systems. UHPLC systems have been widely used for routine bioanalysis, food
safety, and in other
μ
fields. Many applications and reviews have been published
recently [31
34]. UHPLC has become one of the most advanced techniques for LC
in the past decade.
-
Hydrophilic Interaction Liquid Chromatography
Hydrophilic interaction liquid
chromatography (HILIC) is a variation of normal-phase chromatography with the
advantage of using organic solvents that are miscible with water. It uses polar materials
such as amino, cyano, diol, and silanol as the stationary phase. Thus, HILIC is
sometimes called
“
reverse reversed-phase
”
or
“
aqueous normal phase
”
chromatogra-
phy. The HILIC concept was
first introduced by Dr. Andrew Alpert in his 1990
paper [35]. A large number of papers on this subject have been published since
then [36,37].
A hydrophilic stationary phase and an aqueous
organic solvent mobile phase with
high organic solvent content are used in HILIC. Like normal phase LC, retention
increases when the polarity/hydrophilicity of the analytes and/or the stationary phase
increases. However, retention also increases as the polarity of the mobile phase
decreases. The most common organic solvent for HILIC is acetonitrile due to its low
viscosity [38]. Methanol, tetrahydrofuran, and other organic solvents can be used as
-
