HPLC Column Care, Storage, and Restoration
An HPLC column has a finite lifetime. It is recommended that you routinely monitor a column's retention characteristic and performance using standards. If you plan to store the column for a prolonged period of time, use a mobile phase that inhibits microbial growth (a mobile phase containing sodium azide or high concentrations of methanol, acetonitrile, etc.). For optimum performance and column life, follow these recommendations.
- Routinely monitor the column's performance.
- Switch only between miscible mobile phases.
- Avoid precipitating salts in the column.
- Use filtered and degassed mobile phases.
- Do not allow the column to dry out. Keep it tightly capped when not in use.
- For prolonged storage, use a mobile phase that inhibits bacterial and mold growth.
- Use the column in the direction indicated on the label. An unusually high operating pressure may indicate a plugged inlet frit and may be cleared by reversing flow through the column for 10-20 mL.
- Use Hamilton guard columns to remove particulate matter or impurities that may permanently bind to the analytical column.
Restoring Column Performance
The procedures listed below may restore the performance of a column that has become fouled, but the procedures will not improve the performance of a column that is damaged or beyond its useful life. Nor can all stationary phases be restored. The unique supports in PRP-X500 and PRP-X600 columns make it impossible to restore performance with a washing procedure. For technical assistance regarding these columns, please Contact HPLC Support. NOTE: Before performing a restoration procedure, always make sure that the mobile phases are miscible and that precipitation will not occur. If necessary, wash the column with a suitable intermediate solvent before changing over to the new mobile phase. A minimum of five column volumes of intermediate solvent should be used.
Column Restoration Procedures
Follow the appropriate restoration procedure listed below using a flow rate of 1.0 or 2.0 mL/min. unless otherwise specified. Evaluate column performance using the test mix and conditions listed on the Performance Test Report shipped with each column.
| Packing Material | Restoration Procedure |
|---|---|
| PRP-C18 | Flush with 40:40:20 (ACN:IPA:H20) for 5-10 column volumes. Equilibrate with starting mobile phase for 5-10 column volumes. |
| PRP-1 | Flush with 40:40:20 (ACN:IPA:H20) for 5-10 column volumes. Equilibrate with starting mobile phase for 5-10 column volumes. |
| PRP-3 | Flush with 40:40:20 (ACN:IPA:H20) for 5-10 column volumes. Equilibrate with starting mobile phase for 5-10 column volumes. |
| PRP-h5 | Flush with 40:40:20 (ACN:IPA:H20) for 5-10 column volumes. Equilibrate with starting mobile phase for 5-10 column volumes. |
| HxSil C8 | Flush with 40:40:20 (ACN:IPA:H20) for 5-10 column volumes. Equilibrate with starting mobile phase for 5-10 column volumes. |
| PRP-X100 | Pump approximately 50 mL of methanol with 1% 6 N nitric acid. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| PRP-X200 | Inject several times with 100 µL of 1 N nitric acid. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| PRP-X300 | Inject several times with 100 µL of 1 N nitric acid. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| PRP-X400 | Inject Several times with 100 µL of 0.1 M potassium EDTA. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| RCX-10 | Flush with 50 mL 0.1 N sodium hydroxide. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| RCX-30 | Flush with 150 mL 0.1 N sodium hydroxide. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| HC-40 | Flush with 1% calcium chloride at 0.1 mL/min overnight. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| HC-75 H+ | Flush with 0.1 N sulfuric acid at 0.1 mL/min overnight. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| HC-75 Ca+2 | Flush with 1% calcium chloride at 0.1 mL/min overnight. Equilibrate with starting mobile phase for ~ 200 column volumes. |
| HC-75 Pb+2 | Flush with 1% lead nitrate at 0.1 mL/min overnight. Equilibrate with starting mobile phase for ~ 200 column volumes. |
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