If you are working with an established method, we would recommend for you to:

• Check all solutions used for the SPE method to ensure that nothing is expired or has been mislabelled.
• Check the process. Was flow potentially too fast during loading or elution? Could the cartridges have been allowed to dry between equilibration and sample loading?

It is generally quickest to simply repeat the extraction, taking care of these points.

If, however, you are developing a new method, it might be necessary to “track down” your analyte. The first thing we would recommend is to make up a solution of the analyte without sample matrix (to use as an ideal sample). Then work through the sample method capturing the eluent from each step potentially containing the analyte:

1. Condition the Cartridge.

2. Equilibrate the Cartridge.

3. Load the sample and collect the flow through in a vial.

4. Analyze this for analyte content. If analyte is found in this fraction it would suggest:

• Sample solvent is too strong, or the pH is incorrect.
• Loading flow rate is too high.
• You have breakthrough because either sample load is too high, or sorbent mass is too low.

5. Wash the cartridge, collecting each wash fraction in a separate vial.

6. Again, analyze for analyte content. If analyte is found in any wash fraction, this suggests:

• Wash solvent is too strong.
• Wash solvent is not at the correct pH.

7. If the analyte has not been found in the load or wash fractions, then the analyte must be retained on the cartridge.

• Try using a stronger elution solvent.
• Ensure pH of the elution solvent is correct.
• If the first 2 suggestions don’t help, try using a less retentive sorbent (in reversed phase, try switching a C18 cartridge for a C8 if the analytes are very hydrophobic).