Column Dimensions
Particle Size and Column Length
When choosing a column, the first two
parameters that should be considered are
the particle diameter and column length.
These two parameters are the major con-
tributors to separation efficiency (N), also
known as theoretical plates. The number
of theoretical plates is directly proportion-
al to the length of the column over the
diameter of the particle.
Particle Diameter
Particle diameter (dp), is commonly
expressed in micrometers (µm), and has
an inverse relationship to the efficiency of
the separation. As the particle diameter
decreases, the efficiency of the separation
increases proportionately. If all other
parameters remain equal, a 3 µm particle
diameter offers an approximate 60%
increase in efficiency over a 5 µm particle,
and a 1.9 µm particle diameter offers an
additional 60% over a 3 µm particle.
System backpressure also increases pro-
portionally as particle size decreases.
Selecting the proper particle diameter is a
way of controlling separation efficiency,
and even analysis speed, but is limited by
the pressure capabilities of the system.
Often, particle diameters are determined
by instrumentation. Table I is a guideline
for selecting the optimal particle size,
based upon pressure capability for com-
mon mobile phases.
When choosing a particle diameter, it is not recommended to operate significantly below the optimal linear velocity, as losses in effi-
ciency can be observed due to axial dispersion. As a quick estimate of particle diameter usability, check the optimal linear velocity for
the organic solvent used and ensure maximum pressures observed are within the pressure specifications of your instrument. Please
note that these are maximum pressures observed during gradient analyses. Isocratic mobile phases of lesser viscosity will operate with
less back pressure.
Column Length
Column length (L) directly relates to efficiency. Increasing column length increases efficiency. It is important to note that column
length is not an ideal way to increase resolution. Doubling the column length yields only a 1.4x gain in resolution (efficiency is a
square root term in the resolution equation), while doubling both analysis time and system backpressure. Shorter column lengths are
suitable for fast gradients and higher sample throughput, while longer column lengths are more suitable for higher peak capacity and
shallow gradients.
Column Internal Diameter
Column internal diameter (ID) is the inner diameter of the column hardware holding the packing material, and is commonly
expressed in millimeters (mm). Column ID is ultimately related to efficiency and flow rate through the van Deemter equation. This
chromatographic concept relates column efficiency (often called band broadening) to linear velocity. Linear velocity is the distance
mobile phase travels per unit time, while flow rate is the volume of mobile phase per unit time. A specific linear velocity has a flow
rate that is dependent upon the internal diameter of the column. As column ID is lowered, a lower flow rate is needed to
maintain the same linear velocity. Flow rate is the volume of mobile phase needed to create the desired liner velocity. It is important
to note that as particle size decreases, optimal linear velocity increases. Columns with smaller particle sizes, namely 1.9 and 2.2 µm,
are capable of running much higher flow rates and therefore creating higher sample throughput. Table II (next page) can be used to
find the optimal flow rate, as it relates to particle size and internal diameter, and is a good starting point for method development.
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Table I
Emperically determined maximum pressures exhibited for
acetonitrile and methanol gradients for various particle sizes and flow rates
Flow rate
Pressure (psi)
Flow rate
Pressure (psi)
(mL/min.) Acetonitrile @ 25°C
(mL/min.)
Methanol @ 25°C
1.9µm
2.2µm
3µm
1.9µm
2.2µm
3µm
0.2
2436
1755
1045
0.2
3198
2304
1371
0.3
3655
2633
1567
0.3
4797
3455
2057
0.4
4873
3510
2090
0.4
6395
4607
2743
0.5
6091
4388
2612
0.5
7994
5759
3429
0.55
6700
4826
2873
0.55
8794
6335
3771
0.6
7309
5265
3135
0.6
9593
6911
4114
0.7
8527
6143
3657
0.7
11192
8062
4800
0.8
9745
7020
4180
0.8
12791
9214
5486
0.9
10964
7898
4702
0.9
14390
10366
6171
1
12182
8775
5224
1
15989
11518
6857
Data are for 2.1 x 50 mm columns using a gradient of 5% B to 95% B (A: water, B: organic solvent).
See Table II for optimal flow rates for alternate column internal diameters.
Bold blue numbers represent optimal linear velocity for the given particle size and ID. For longer
column lengths, the approximate pressure corresponds to the increase in column length. A 2-fold
increase in column length yields a 2-fold increase in back pressure.
Equation 1
The resolution equation defines variables affecting separations.
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