How to Calculate Void Volume in HPLC
The void volume is the volume that the mobile phase occupies in a column. For instance, if 40% of the entire column's capacity is filled up with stationary phase material, the void volume would be 60% of the total column volume.
The void volume can be calculated using the formula below:
V0 = π × (d/2)2 × L × pore volume
or
V0 = π × r2 × L × pore volume
where:
d = column internal diameter in mm
r = column internal radius in mm
L = column length in mm
pore volume = 0.70 (for fully porous packing) or 0.50 (for superficially porous packing)
To determine the void volume of a column experimentally, you must inject an unretained compound that has no or little retention. An example of this is Uracil for reversed phase HPLC columns. Following this injection, use the following formula to calculate its void volume (Vo):
V0 = Retention time of unretained compound × Flow rate
Example: If the unretained compound elutes at 2.00 min, and the flow rate is 0.40 mL/min, then:
V0 = 2.00 min × 0.40 mL/min
V0 = 0.80 mL
If you are unable to perform an experiment, quickly calculate your column’s void volume using our void volume calculator below!
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