Tip # 120: Procedure to Adjust Mobile Phase Flow Rate to
Maintain HPLC Column Linear Velocity When Column Diameter Is Changed
Lots of 2.1mm ID chromatography columns are appearing on the market right now. Since
most of us are using 4.6 mm ID columns to develop HPLC and UHPLC methods, use of these
smaller ID columns requires a few adjustments be made to the method. If gradient elution
is used, then the gradient profile often must be changed to compensate for changes in void
volume of the column and dwell volume of the system. Injection volume must also be
adjusted in a linear fashion too. Additionally, to maintain the same initial mobile phase
linear velocity through the column as we had before (to obtain the same approximate
retention times), the flow rate must also be adjusted. *We will discuss how to calculate
the change in flow rate for identical particle sized supports in this installment.
In order to reproduce your original method, we must first adjust the flow rate for the
new, narrower bore column. The formula to do this is very simple. We decrease the flow
rate by using the square of the ratios of the column diameters times the original flow
rate.
Formula:
( C1 / C2 )2
x original flow rate (ml/min) = new flow rate (ml/min)
Where:
C1 = Diameter (mm) of new (smaller) column;
C2 = Diameter (mm) of the original column.
Example #1: Find the new linear flow rate if we use a 2.1 mm ID column in place
of a 4.6mm column with an initial flow rate of 1.000 ml/min.
(
2.1 / 4.6 ) 2 x 1.000 = 0.208 (208
ul/min)
Example #2: Find the new linear flow rate if we use a 2.1 mm ID column in place
of a 4.6mm column with an initial flow rate of 2.000 ml/min.
(
2.1 / 4.6 ) 2 x 2.000 = 0.416 (416
ul/min)
Example #3: Find the new linear flow rate if we use a 1.0 mm ID column in place
of a 4.6mm column with an initial flow rate of 1.500 ml/min.
(
1.0 / 4.6 ) 2 x 1.500 = 0.071 (71
ul/min)
Note: If we assume that the original flow rate is 1.000 ml/min then we can also use
this table to get an idea of how the flow rate changes with decreasing column diameter.
Column I.D. (mm)
Flow Rate (ul/min)
4.6
1,000
2.1
208
1.0
47
0.3
4
0.15
1
>
Bill Letter, 04/01/2013
