Translator for HPLC HINTS and TIPS for Chromatographers

Saturday, April 21, 2018

The HPLC Restriction Capillary; Troubleshooting, Qualification and Running Without A Column:

Most types of HPLC pumps will not operate properly without 30 or more bars of back-pressure on their outlets to prevent cavitation and excessive pulsation. Columns play a vital role in stabilizing the baseline during an analysis.

When we want to closely replicate the operation of an HPLC system under "normal" conditions and do not want to use an HPLC column in-line (because a column adds variability), we install a "restrictor" such as a restriction capillary in its place. A restriction capillary is often a very narrow ID section of long tubing (capillary) which will restrict the flow of mobile phase through it. For most HPLC systems, a restrictor which is sized to provide about 1,000 to 2,000 psi (~ 70 to 140 Bars) of back-pressure will closely replicate normal operating conditions. The restrictor can be chosen based on length, ID, volume and your flow rate to create this level of back-pressure. You could place a high pressure rated, zero-dead-volume union its place, but in doing so, the system back-pressure may be extremely low ( a few bars) and show poor pump performance. We need to replicate actual analysis conditions during testing or the results obtained may be invalid and unscientific. An HPLC column, with its densely packed small particles inside acts as a pressure pulse buffer and adds a great deal of back-pressure to the HPLC system. That back-pressure greatly improves the stability of the pump operation and overall baseline. HPLC Columns prevents pulsations by acting as a dampener and/or system buffer.

There will be times when you need to operate the HPLC system without an HPLC column installed.

For Example: 
  • Troubleshooting sources of contamination, carryover or artifact peaks on a column;
  • Measuring the HPLC system delay volume (gradient delay);
  • Testing the performance of the injector;
  • Testing the performance of the pump (measure % ripple); 
  • Testing the performance of a detector module (measure S/N);
  • Running HPLC Operational Qualification Tests (OQ);
  • Running HPLC Installation Qualification Tests (IQ);
  • Running Performance Verification Tests on a Module (PV);
  • Running many of the ASTM Tests (e.g. "Baseline Noise & Drift Test").
Example of a commercially available Restriction Capillary (Agilent P/N G1312-67500). You will want to include any needed details of the restriction capillary chosen for your work in the SOP's that you write which utilize it as part of any test (P/N, source, dimensions, volume...).