7 Matters in the Operating Procedures of Flash Columns
The Flash liquid chromatography column is packed with ultra-pure silica gel, which has lower impurity content, narrow particle size distribution, strict water content control, and neutral pH, which can meet the higher separation needs of users. The Flash chromatographic column plays a separation role in the chromatography analysis system.
Correct use and maintenance of chromatographic columns are particularly important. Improper use will reduce column efficiency, shorten the service life, or even damage. The following is a brief answer from seven aspects to introduce the issues that should be paid attention to during the use of Flash columns.
1. Avoid sudden changes in pressure and temperature
Sudden changes in temperature or dropping the chromatographic column from a high place will affect the packing conditions in the column; a sudden increase or decrease in column pressure will also impulse the packing in the column. Therefore, operators should adjust the flow rate slowly to keep the stability of the filler in the column.
2. Avoid directly changing the composition of the solvent
When the composition of the test sample solution needs to be changed, the composition of the solvent should be gradually changed, especially in reversed-phase chromatography. Direct change from organic solvents to all water is not allowed, and vice versa. And when washing the chromatographic column with the eluent with strong elution capacity, the displacement of the mobile phase in the flow path system should be gradually transitioned with miscible solvents. Each mobile phase’s volume ought to be about 20 times the column volume. That is, 50~75ml is required for routine analysis.
3. Avoid column backflush
Generally speaking, chromatographic columns cannot be backflushed. Only when the column’s operating instructions indicate that the column can be backflushed, can backflush to remove impurities left on the head of the column.
Otherwise, the effect of backflushing the column is to rapidly reduce column efficiency.
4. Avoid improper use of mobile phase and damage to stationary phase
Choose to use a suitable mobile phase (especially pH) to avoid the destruction of the stationary phase.
Sometimes a pre-column can be connected in front of the sampler. Before entering the analytical column, the mobile phase can be “saturated” with silica gel, if the pre-column is silica gel and the analytical column is bonded silica gel,
5. It is forbidden to directly enter the chromatographic column without treatment.
Injecting samples with complex matrices, especially biological samples, into the column directly should be avoided.
It is necessary to pretreat the samples or connect a guard column between the injector and the chromatographic column. The guard column is generally a short column packed with a similar stationary phase.
6. It is forbidden to store the buffer solution in the column for a long time.
To prevent the solvent from evaporating and drying, fill the column with acetonitrile or methanol, and tighten the column connector when storing the chromatographic column. Do not leave the buffer solution in the column overnight or longer.
7. It is forbidden to seal the column without treatment
After the chromatographic column is used, it must be cleaned with a suitable solvent. For example, the ODS column should be flushed to baseline equilibrium with methanol; when a salt buffer solution is used as the mobile phase, it should be flushed with a salt-free mobile phase after use; containing halogen elements (fluorine, Chlorine, bromine) compounds may corrode stainless steel pipes and should not be in contact with them for a long time; if the chromatographic column installed on the liquid chromatography is not used frequently, it should be turned on for 15 minutes every 4-5 days.