Flexar Analytical / FX-10 UHPLC Pumps
IQ/OQ Rev. D June 2009
2.4.6 Binary / Quaternary Analytical Pump Composition
Accuracy Test
Test Prerequisites
There is a minimum pressure requirement of 350 psi for the check valves to operate properly.
To ensure the valves are operating and seating during the test PerkinElmer runs the test with
the pressure approximately >500 psi. This is accomplished by using a length of 0.007” ID
tubing or a backpressure device that will create the necessary back pressure on the check
valves.
Test Conditions
The Composition Accuracy Test proves that the proportioning valves are operating correctly.
This is accomplished by varying the amounts of solvent entering the pump through the
proportioning valves. Using an acetone solution, which has an absorbance at 265 nm as one
solvent, and water, which does not absorb at 265 nm we look at the absorbance of the
resulting composition. By controlling or proportioning the amount of acetone in the
water/acetone composition we can change the absorbance of the mobile phase composition.
We do this by setting up a pump program to run through several different positions of the
proportioning valves and measure the absorbance of the resultant mobile phase.
Test Steps
•
Connect pump output directly to the backpressure regulator or other device (length of
tubing for example) to create the necessary back pressure.
•
Fill reservoirs A (and C if testing a quaternary model) with HPLC grade water.
•
Pipette 1.0 mL of acetone into approximately 250 mL of HPLC-grade water. Dilute to 500
mL with HPLC grade water. Transfer the mobile phase to the labeled solvent reservoirs.
Place the 0.2% acetone solution into reservoirs B (and D if testing a quaternary model).
•
Ensure that acetone solution is mixed well and degassed.
•
Bypass the column and autosampler and connect pump output directly to the
backpressure regulator or other device (length of tubing for example) to create the
necessary back pressure.
•
Connect the output of the back pressure device to the input of the detector flowcell.
•
Equilibrate and insure the binary or quaternary valves are operating correctly by running
the pumps as follows:
•
Binary pump -
run 50/50 at 5mL/min for 10 minutes
Quaternary pump -
run 25/25/25/25 at 5 mL/min for 10 minutes.
•
Observe the P-counts on the pump (CNFG/CAL/TEST/PCNT). The P-count value should
fall typically in the range of 1300 + 50. Stability is important and the Range value should
be typically less than 12.
•
Set the detector wavelength to 265 nm.
•
Set the flow rate to 5 mL/minute from reservoir A and allow the pressure to stabilize.
Ensure the pump and baseline is stable at 5 mL/min.
•
Create and run the method in Tables 1 and 2 as appropriate with TotalChrom® or
Chromera®, zeroing the detector at the end of step zero. Step 2 is the 100% reference.
