Column Operation and Storage

Good column care requires little additional time and will extend the lifetime of your HPLC columns. Column care for SIELC brands of columns involves operating in a suitable mobile phase and storing column in an appropriate solvent system.

Mobile Phases for Column Operation and Storage

TRY TO AVOID USING ALCOHOLS WITH PRIMESEP A, PRIMESEP 100, Primesep 200, Primesep C and Primesep P columns as you can esterify carboxylic acid attached to the surface of silica. The esterification might significantly effect reproducibility and retention time of some basic analytes.

Mobile Phases for Column Operation 
General Guidelines

•  Choose a mobile phase for your sample that falls within the recommended pH limits for particular PRIMESEP column. PRIEMSEP columns can be operated at the pH range 1.5-7.5. Primesep B, Primesep AP and Primesep D recommended pH limits are 1.5-4.5 Remember to determine the mobile phase pH on the aqueous component alone, before mixing with organic modifiers.

•  Because PRIMESEP columns use AcOH and TFA in the mobile phase there is no buffer to precipitate in the column. Equilibrate the column with the mobile phase before analyzing your sample. Check that your solvents are miscible with the shipping solvents before equilibration.

Mobile Phases for Column Storage  

General Guidelines for storing PRIMESEP columns.          

•  Do not store PRIMESEP HPLC columns in pH outside of recommended storage range (pH=5-7.5).

•  Primesep B, Primesep AP and Primesep D columns require acidic or buffered mobile phase within pH range 1.5-4.5 for all operation and storage.

•  Do not store PRIMESEP HPLC columns in solvents that degrade easily -- tetrahydrofuran (THF), triethylamine (TEA), and trifluoroacetic acid (TFA). Unstabilized THF can form peroxides, which may degrade the column. TEA and TFA are more likely to become contaminated from the lab environment and should be refrigerated during storage. Contamination may change the chromatography or change the column. Be aware of the quality issues with these mobile phase components when you use them for your chromatography

Short Term Storage              

•  Flush the column with your mobile phase minus any additives (acetic or trifluoroacetic acids, triethylamine) for short term storage
i.e.    Mobile Phase: 50 % Acetonitrile: 50% water: 0.05% H 2 SO 4
Flush: 50% Acetonitrile: 50% Water
This will eliminate the risk of stationary phase degradation over prolonged storage time and will minimize re-equilibration time when the column is used next time.

•  For overnight storage, keep the mobile phase flowing at 0.1 - 0.2 mL/min as it reduces re-equilibration time the following day. This slow purge will consume less than 100 mL of the mobile phase.

•  Store Primesep B column in weak acidic solution. Storage of Primesep B column in water can reduce performance of the column.

Long Term Storage             

•  For long-term storage of PRIMESEP columns we recommend 100% acetonitrile. If you use a mobile phase with modifier (AcOH or TFA), first flush the column with the mobile phase minus modifier, then with 100% acetonitrile. In addition, a 100% ACN flush prior to long-term storage can help remove contaminants from the column.

•  Store Primesep B column in weak acidic solution. Storage of Primesep B column in water can reduce performance of the column.

Other Operational Guidelines           

•  It is important to operate PRIMESEP column below the specified maximum temperature limits in order to obtain the longest possible lifetime. At high temperatures columns will degrade faster. This can involve loss of bonded phase or silica dissolution, depending on pH of the mobile phase. It is important to control temperature for the most reproducible results, but not to exceed the maximum recommended operating temperature for any mobile phase condition. The operating temperature for PRIMESEP columns is 0-50 C.

•  It is important to operate PRIMESEP column below the specified maximum pressure limits (5000 psi). You should note the column pressure when you start to use a column with your mobile phase. If you are running a gradient the column pressure will change during the gradient. In many cases, if the starting pressure increases during column operation, the column inlet frit may be plugged.

•  Choose a flow rate that will allow you to keep the column pressure below the specified limits. Typical flow rates for PRIMESEP columns are 0.5 - 2 mL/min for 4.6 mm i.d. columns, 0.2 – 1.0 mL/min for 3.0 mm i.d. columns and 0.1 -0.5 mL/min for 2.1 mm i.d. columns.

•  An arrow on the PRIEMSEP column label is used to specify direction of flow through the column. Typically you should operate the column in the direction marked.

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