Chemistry Reference
In-Depth Information
4.1 Mobile Phase Solvent Effects and Small Molecule
Retention Mechanisms
For decades, RPLC has been one of the most widely used techniques for the
separation and analysis of chemical mixtures, but there remains significant debate
about the molecular level details of numerous aspects of the RPLC process. For
example, it has been observed that a dramatic loss of retention can occur when the
concentration of water in the mobile phase exceeds a certain threshold [
12
,
41
].
One explanation for this phenomenon is that the alkyl chains of the stationary phase
collapse in the presence of highly aqueous solvents [
41
-
43
]. A more recent,
competing explanation for this retention loss is that a highly aqueous mobile
phase does not enter a substantial fraction of the smaller pores in the silica particles
due to its higher surface tension and hence is not able to bring the analytes in
contact with the stationary phase chains [
12
]. With regard to solvent effects, another
well known phenomenon in RPLC is the tendency of the organic component of the
mobile phase to solvate preferentially the stationary phase. However, it is not fully
resolved if this preferential solvation occurs solely through the formation of an
organic layer atop the stationary phase [
44
,
45
] or if penetration of the organic
modifier into the stationary phase is also important [
46
,
47
].
Perhaps the biggest and most longstanding debate in RPLC is on the retention
mechanism. Here there are conflicting views as to whether analyte molecules
adsorb at the stationary phase/mobile phase interface or fully partition into the
stationary phase and to what extent various chromatographic parameters, such as
mobile phase composition, affect this mechanism [
48
-
51
]. Even if partitioning is
taken to be the dominant mechanism of retention, it is not clear if the process can be
modeled accurately by bulk liquid-liquid (e.g., oil-water) partitioning [
48
,
52
]orif
partitioning into the constrained hydrocarbon chains of the RPLC stationary phase
would involve a different molecular mechanism [
53
,
54
]. Furthermore, it is dis-
puted whether the thermodynamic driving forces for analyte retention (transfer
from mobile to stationary phase) are primarily from solvophobic interactions with
the mobile phase [
50
] or lipophilic interactions with the stationary phase [
48
,
52
].
To illustrate the effects of mobile phase composition, results for four different
water/acetonitrile mobile phase compositions are discussed here: (1) pure water, (2)
33% molfraction acetonitrile, (3) 67% molfraction acetonitrile, and (4) pure aceto-
nitrile, (hereafter referred to as systems WAT, 33A, 67A, and ACN, respectively;
the results for system WAT are taken from [
32
] and those for the other three
compositions from [
34
]). Each system contained 1,200 solvent molecules and 16
analytes (2 each of C1 to C4 normal alkanes and alcohols) and utilized a stationary
phase with a surface coverage of 2.9
mol/m
2
(9 ODS chains on each surface)
which resulted in a residual silanol density of 4.8
m
mol/m
2
(15 silanols on each
surface). The temperature and pressure used in this study were 323 K and 1 atm,
respectively. For each solvent system, four independent simulations were carried
out. Each simulation was equilibrated for 2
m
10
5
Monte Carlo (MC) cycles (one
MC cycle corresponds to
N
MC moves, where
N
is the total number of molecules in
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