Waters Certified Sep-Pak - Manual (Page 4)

[ Care and Use ManUal ]
Certified Sep-Pak Cartridges 4
5. Elute
Once the interferences are washed off the cartridge, a strong solvent is introduced to elute the analytes of interest. The volume and flow rate
of the eluting solvents should be precisely controlled as in the load step to ensure reproducible results.
Reversed Phase Normal Phase Ion Exchange
Analyte
Low to moderate polarity/
hydrophobic
Moderate to highly polar/uncharged Charged or ionized
Matrix Aqueous Non-polar organic solvent Aqueous/low ionic strength
Condition/Equilibrate
1. Solvate polar organic
2. Water
Non-polar organic Low ionic strength buffer
Wash Aqueous/buffer Non-polar Low ionic strength buffer
Elute
Increase polar organic content
in steps
Increase moderate to high polarity
organic content in steps
Stronger buffers ionic strength or
pH to neutralize the charge
Table 2. Guidelines on the Various Types of Separation Mechanisms
IV. FACtoRs AFFeCtInG sePARAtIon eFFICIenCY
Separation efficiency varies with flow rate and sample load. Here
are some guidelines to consider when optimizing your separation.
a. Flow Rate
If the sample flow rate is too high, components may not interact
sufficiently with he sorbent. The result is loss of resolution, analyte
breakthrough, and poor recovery. To condition the cartridge, use a
flow rate of 5-10 mL/min. To load and elute Certified Sep-Pak Vac 3
cc 200 mg, and 500 mg cartridges, use a flow rate or 0.2-1 mL/min.
To load and elute all other cartridges, use a flow rate or 2-10 mL/min.
b. Sample Load
Cartridge capacity depends upon the mass of all components in the
sample, volume of solvent, solvent strength, sorbent type, sorbent
quality, and retention factor (k) of components. Overloading results in
sample breakthrough which leads to variability in sample recovery.
V. stoRAGe AnD DIsPosAL oF UseD CARt RIDGes
Cartridges stored in their original sealed pouch remain stable
for long periods. Once opened, special care must be taken to
ensure that the SPE cartridges are not exposed to the laboratory
environment for long periods. The highly active sorbents will readily
accumulate background contaminants from the environment and
will affect the quality of the device. Waters recommends that you
open and use only those cartridges that will be needed. If this is not
possible, store the exposed cartridges in clean contaminant free
container such as a desiccator.
Dispose of used cartridges safely in accordance with applicable
government or local regulations.