ver.211228E
SHOWA DENKO K.K. (https://www.shodex.com/)
4
7. Column Usage Procedure
7.1 HPLC System Preparation
Wash entire LC system prior to the column installation, including all flow-lines and sample loop by switching the
valve, and then replace the washing solution with the eluent to be used. If desired new eluent has low
miscibility/solubility to the eluent of previous analysis, first use the eluent that is miscible/soluble to both eluents,
and then replace it with the desired eluent.
e.g. When replacing chloroform to water, first run methanol and then replace it with water.
e.g. When replacing highly concentrated buffer solution or salt solution to water/acetonitrile, first run water and
then replace it with water/acetonitrile.
Attention!
· If the eluent left in the system is not compatible with the column to be used, it may damage
the column.
· A drastic change in the eluent compositions may remove substances adsorbed on the
system and they may enter and deteriorate the column.
7.2 Column Installation
(1) Connect the column to LC system by following the “flow direction arrow” (
➡
) indicated on the column name
tag. If guard column is used, position the guard column in front (before the inlet) of the analytical column.
(2) Make sure to insert the tubing all the way to the end fitting and secure it with the male nut. It is important that
there is no extra space between the tubing and the column side of the end fitting. Presence of an extra space
will let the sample to spread out and may result in wide peaks.
(3) Set the initial flow rate at less than 0.2 mL/min and start the system. If using the column at an elevated
temperature, keep a low flow rate until the temperature of the column reaches to the set temperature, and
then gradually increase the flow rate to the desired.
(4) Multiple number of columns can be connected in series. When connecting multiple columns with each having
different exclusion limits, set the column with higher exclusion limit at the upper stream position. When
connecting GF-7M HQ, use multiple number of the same columns in series. This is because
GF-7M HQ is
prepared by using several different gels with different pore sizes, designed to provide linear calibration
curves over a set molecular weight range. If a GF-7M HQ was used together with columns with different
pore sizes, this changes the mixing ratio of the gels and consequently it may deform the linear calibration
curve.
Caution!
· Verify that there is no solvent leak. It may cause electronic leakage, rust, and/or chemical
injury.
Attention!
· Make sure not to let air bubbles enter the column while installing the column. The air
bubbles may damage the column.
· When restarting the system after column installation or after holding the eluent flow, start
the system at less than 0.2 mL/min. A rapid increase in pressure can damage the column.
· If the column was used at an elevated temperature, lower the flow rate to less than
0.2 mL/min at the end of analysis. Then, turn off the column oven, and let the column
temperature return to room temperature before stopping the pump. If the pump was
stopped while the eluent inside the column was still hot, as the eluent temperature
decreases, its volume also decreases. This may result in creating an empty space in the
column and deteriorates the column.
Note
· It is recommended to set the pump limiter to avoid exceeding the maximum pressure.