Method for determining an operating flow rate for a chromatographic column in an HPLC system
Disclosed is a method for determining an operating flow rate for a chromatographic column ( 4 ) in an HPLC system ( 1 ). The method comprises: measuring/calculating a pressure of the system ( 1 ) without the chromatographic column ( 4 ) for one or more flow rates; fitting a function to the flow rate(s) and corresponding pressure(s), calculating from the function and a predetermined recommended flow rate for the chromatographic column ( 4 ) a system pressure drop at the predetermined recommended flow rate. An operating flow rate is determined by summing the system pressure drop and a maximal column pressure limit, and determining a contribution of the system pressure drop to the summed pressure. If this contribution exceeds 1% an operating flow rate for the column is determined to a flow rate that corresponds to a pressure at a pressure monitor arranged before the column that is lower than the predetermined maximum column pressure limit.
1. A method for determining an operating flow rate for a chromatographic column in a high-performance liquid chromatographic system, the system comprising:
a liquid reservoir for a liquid medium,
a chromatographic column in fluid communication with the liquid reservoir, said chromatographic column having a predetermined recommended flow rate and a predetermined maximal column pressure limit,
a system pump arrangeable to force liquid from the liquid reservoir through the chromatographic column at a certain flow rate,
a fluid flow path connecting the liquid reservoir, the system pump and the chromatographic column, and
a pressure monitor arranged before said chromatographic column,
the method comprising:
a) measuring or calculating a pressure of the system without the chromatographic column for one or more flow rates,
b) fitting a function to said flow rate(s) and corresponding measured or calculated pressure(s),
c) calculating from said function and said predetermined recommended flow rate for the chromatographic column a system pressure drop at said predetermined recommended flow rate, and
d) determining an operating flow rate for the chromatographic column by summing the system pressure drop and the predetermined maximal column pressure limit, and determining a contribution of the system pressure drop to the summed pressure, wherein if this contribution exceeds 1 % an operating flow rate for the chromatographic column is determined to a flow rate that corresponds to a pressure at the pressure monitor that is lower than the predetermined maximum column pressure limit.
2. The method of claim 1 , wherein the pressure of the system is measured by means of the pressure monitor for one or more flow rates.
3. The method of claim 2 , wherein the chromatographic column is replaced or bypassed with tubings.
4. The method of claim 1 , wherein the pressure of the system is calculated for one or more flow rates by subtracting pressure(s) of the system measured by means of the pressure monitor, wherein the chromatographic column is disconnected from the system, from pressure(s) of the system measured by means of the pressure monitor.
5. The method of claim 1 , wherein the pressure of the system is calculated for one or more flow rates without the chromatographic column using the Bernoulli equation:
Δ P =(128 *L*Q*η )/(π* d 4)
wherein, d is an inner diameter of the fluid flow path (mm), Q is the flow rate (ml/min), L is a length (mm) of the flow path and η is the viscosity (cP) of the liquid medium.
6. The method of claim 1 , wherein the pressure of the system is measured or calculated without the chromatographic column for at least two different flow rates.
7. The method of claim 1 , wherein if the determined contribution of the system pressure drop to the summed pressure exceeds 5%, 10%, 15% or 20%, an operating flow rate for the chromatographic column is determined to a flow rate that corresponds to a pressure at the system pump which is lower than the predetermined maximum column pressure limit.
8. The method of any claim 1 , further comprising:
calculating/measuring a delta-pressure signal for the chromatographic column in the system by measuring a pressure by means of the pressure monitor at the predetermined recommended flow rate and subtracting therefrom a measured or calculated pressure of the system without the chromatographic column at the predetermined recommended flow rate.
9. A high performance liquid chromatographic system comprising: a) a liquid reservoir for a liquid medium, a chromatographic column in fluid communication with the liquid reservoir, said chromatographic column having a predetermined recommended flow rate and a predetermined maximal column pressure limit, at least one system pump arrangeable to force liquid from the liquid reservoir through the chromatographic column at a certain flow rate, a fluid flow path connecting the liquid reservoir, the system pump and the chromatographic column, and a pressure monitor arranged before said chromatographic column, and b) a system controller for determining an operating flow rate for the chromatographic column of the system, wherein the system controller is arranged to perform steps of claim 1 .
10. A computer program comprising program code means stored on a non-transitory computer readable medium for performing the method of claim 1 , when the program is run on a computer.
11. A computer program product comprising program code means stored on a non-transitory computer readable medium for performing the method of claim 1 , when the program is run on a computer.
12. The method of claim 1 , wherein the pressure of the system is measured or calculated without the chromatographic column for at least three different flow rates.