Flowcell Care and Cleaning Chromatography Technical Note
庐 庐 TN01
for the CombiFlash Companion
Overview Flowcell Cleaning Solvents
For optimum performance, the CombiFlash Com- The flowcell can be cleaned by pumping a solvent
panion detector flowcell must be clear of any through the system. Generally, the recommended sol-
obstructions or build-up of residual sample material. vent is the highest polarity solvent you have recently
Such problems primarily result from low solvent used. Therefore, ethyl acetate (often solvent B) would
polarity. When solvent polarity is too low, compounds be used. See Table 1 for alternative solvents. Recent syn-
may precipitate or sample material may not be flushed thesis solvents should also be considered.
from the system before the next purification.
If the flowcell is obstructed, or performs poorly as a
Table 1: Suggested Flow Cell Cleaning
result of residual compounds, the Companion displays
Solventsa
an error message. The error message identifies the fault
and prompts the user for a response before continuing
Solvent Description
with the run.
This technical note describes the error messages Methanol Miscible with most LC solvents. It will force other solvents
and corrective actions, and advises of measures that through the system so that they will not interfere with the
can prevent the fault conditions. next sample. Breaks down the tertiary structure of silica
and allows it to flush through.
Flowcell Error Messages Acetone Common organic solvent with high dissolving properties.
Dichloromethane Effective at solubilizing organic molecules.
(DCM)
Flowcell cleaning is recommended. Continued opera-
Dimethylsulfoxide Strong solvent that will clear organic sample.
tion without cleaning may result in a noisy trace.
(DMSO)
Dimethylformamide Strong solvent if other solvents do not produce the desired
If the flowcell is partially obstructed the above mes-
(DMF) results.
sage is displayed. Continuing to operate the system
Tetrahydrofuran Strong solvent.
without servicing the flow cell may cause excessive
(THF)
noise in the trace (see Figure 1) or reduce amplitude of
Mildly acidic 0.5M acetic acid in distilled water can be used to clear
absorbing species.
aqueous solution samples that are not removed by organic solvents.
Mildly basic A dish-type detergent solution can wash through particu-
aqueous solution lates not picked up by the acidic solution. The emulsifica-
tion effect of detergent is an effective tool as well to help
wash through oily samples. Follow the detergent with sev-
eral minutes of clean, distilled water.
Figure 1: Noisy trace
a. Always handle solvents in accordance with the manufacturer鈥檚 or
supplier鈥檚 MSDS.
The detector lamp is not on. You can continue the
Quick Flowcell Cleaning
separation without detection, or cancel. 280@81
Quick cleaning is advantageous because very little
solvent is required and repriming is not necessary. To
When the above message occurs at the beginning of
quick clean the system:
a run, either the lamp is not working at all, or the flow
1. Remove the column from the system.
cell is blocked to a degree that the detector is not
receiving adequate light. The number preceding the 鈥淍鈥? 2. With a syringe, inject the selected solvent into
symbol at the end of the message indicates which of the the lower column mount.
two faults is likely. 3. Insert a bypass tube in place of the column.
鈮? 200: Contact Isco Technical Service. The lamp
鈼?
4. From the PeakTrak menu select the Tools>Manual
may not be working at all.
Control鈥? command and pump 100 mL of Solvent
> 200: The lamp is working but the flow cell
鈼?
A at 100 mL/min.
requires cleaning.
5. Replace the bypass tube with a column and start
In the example the number is 280, indicating that the a run. The Companion will attempt to calibrate
flow cell should be cleaned. the flow cell.
Chromatography Technical Note TN01
Preventive care
If the calibration is successful, the system is
鉂?
ready for use. Often the only maintenance required is to operate
If the alert message persists, refer to the thor-
鉂?
the instrument with a bypass tube in place at a high flow
ough flowcell cleaning solvents and tech- rate with a high polarity solvent.
niques. With typical gradient configurations under normal
phase conditions, the B solvent has the highest polarity.
Thorough Flowcell Cleaning Adding a high percentage of B solvent (much higher
than is necessary to elute the sample) at the end of the
To thoroughly clean the flowcell:
separation ensures that all of the sample material is
1. Remove the column from the system and insert a
flushed through the flowcell. This can be added to the
bypass tube.
Companion鈥檚 default methods so that flowcell cleaning
2. If using an alternative solvent, place the solvent
occurs every run (see Figure 2).
inlet line into a container of the selected solvent.
3. Pump 100 ml of solvent through the system at
100 mL/min. 100
4. If an alternative solvent was used, return the sol- Flowcell and
vent inlet line to the appropriate solvent con- Column
Gradient % Solvent B
75 Flush
tainer.
5. Select the Tools>Auto Prime menu command to
prime the system. 50
End of End of
6. Replace the bypass tube with a column and start Separation
Separation Run
Gradient
a run. The Companion will attempt to calibrate
25
the flow cell.
Equilibrate/
If the calibration is successful, the system is
鉂?
Next Initial %B
ready for use.
Initial %B
0
If the alert message persists, refer to the alter-
鉂?
native flow cell cleaning solvents and tech-
Time or Column Volumes
niques.
Figure 2: Flowcell flushing at end of run
The solvent strength and high flow rate will typically
clear the obstruction and return the flow cell to
optimum performance. Summary
A clean flowcell ensures optimum peak detection
Alternative Cleaning Techniques performance.
If the thorough flow cell cleaning instructions do not Pumping a high polarity solvent through the system
clear the obstruction, the following alternative tech- at the end of each run may prevent flow cell problems.
niques should be considered. If the Companion does detect a flow cell problem,
Pump water/DMF/methanol (50/25/25) for 3鈥?4
鈼?
determine the cause according to the displayed error
minutes followed by pumping straight methanol message, then follow through with the recommended
(or ethanol) for 2鈥?3 minutes. corrective action as soon as possible.
Pump 5鈥?10 mL DMSO followed by methanol,
鈼?
ethyl acetate, then hexane.
After the lines have been filled with solvent,
鈼?
reduce the flow rate or pause up to 18 hours.
Doing so will allow the solvent to remain longer
to dissolve the obstruction.
Last modified July 15, 2005
Teledyne Isco, Inc.
P.O. Box 82531, Lincoln, Nebraska, 68501 USA
Toll-free: (800) 775-2965 鈥? Phone: (402) 464-0231 鈥? Fax: (402) 465-3001
E-mail: service@isco.com
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