Agriculture Reference
In-Depth Information
well. It was reported that the use of methanol may result in poor chromatographic
separation or lack of retention for some analytes [38,39].
Compared with reversed-phase liquid chromatography (RPLC), HILIC provides
unique bene
ts for the separation of polar and/or hydrophilic compounds. Due to the
high organic content in the mobile phases, HILIC results in a lower operating back
pressure, which allows higher
flow rates for high-throughput analysis. The high
organic solvent concentration in the mobile phase also leads to a higher sensitivity for
LC
cantly increased.
Although the mechanism of HILIC is still being debated [40], partitioning theory is
well accepted, that is, HILIC involves the partitioning of an analyte between a
predominantly polar organic mobile phase and a water-enriched layer of the mobile
phase that is partially immobilized on the stationary phase. However, some scientists
believe that the separation mechanism involves both partition and
-
MS analyses because the ionization ef
ciency can be signi
”
processes [39]. As with any other chromatographic separation mode, HILIC depends
on the different interaction of solutes between the mobile phase and the stationary
phase. These interaction forces include hydrogen bonding, which depends on the
acidity or basicity of the solutes, electrostatic interactions, and dipole
“
adsorption
dipole inter-
actions, which rely on the dipole moments and polarity of molecules [40]. Based on
the mechanism of HILIC, the formation and stability of water-enriched layers are very
important. Usually, at least 3% water is needed in the mobile phase for suf
-
cient
hydration of the stationary-phase particles so that a stable water-enriched layer
forms [38,41]. Because the mobile phase is an aqueous
organic solvent, it has
good solubility for polar, hydrophilic analytes. Furthermore, these molecules are more
retained and thus better separated in HILIC than they would be by conventional RPLC
with water-rich mobile phases. Of speci
-
c note, HILIC columns found wide
applicability for detecting melamine in different matrices during China
s melamine
milk crisis because melamine and its analog cyanuric acid are very polar and have
very poor retention on regular reversed-phase columns [42,43]. The use of HILIC in a
broad range of food analyses, including the detection of vitamins, marine toxins, and
amino acids in a variety of matrices, has recently been detailed in two topics [37,44].
'
Monolithic Columns
Monolithic media have been used for various niche applica-
tions in GC and LC for a long time. Only recently did they acquire a major importance
in HPLC. Unlike conventional particle-based construction, monolithic columns
contain a continuous network (monolith) of porous silica or organic polymer.
Although the preparation and use of these types of columns was
first reported in
1967 [45], they suffered from poor
flow characteristics resulting in little interest in this
idea as a feasible approach for producing separation media until the 1990s [29]. At
this point, improvements in manufacturing approaches resulted in monoliths with
better performance characteristics and usefulness for high-throughput assays. They
are prepared using a sol
gel process, either
in situ
or in a manufacturing mold, which
enables the formation of a highly porous material, containing both macropores and
mesopores in its structure. Compared with particle-based columns, the most important
advantage of monolithic columns is low back pressure due to macropores (2
μ
m)
throughout the network that allows high
-
flow rates with relatively
at van Deemter
