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Introduction
HyperSep Retain PEP
Using a 30mg 1mL SPE Cartridge
(part number 60107-201)
1. Condition with 1mL methanol
followed by 1mL water
2. Load 1mL of 500ng/mL sample
in water
3. Wash with increasing strengths of
methanol in water, starting with
0% methanol/100% water to
90% methanol/10% water, increasing
methanol content by 10% each time
4. Elute 4 times with 0.5mL of
100% methanol
HyperSep Retain PEP Wash/Elution Profile
The results show an optimum wash profile of 30% methanol/70% water before compounds of interest
start to be eluted. An elution volume of 1.5mL in three aliquots of 500µL eluted off the compounds of
interest to give a high recovery level.
The results show an optimum wash profile of 5% ammonia in 50% methanol/50% water before
compounds of interest start to be eluted. An elution volume of 1.5mL in three aliquots of 500µL eluted
off the compounds of interest to give a high recovery level.
The results show an optimum wash profile of 20% methanol/80% water before compounds of interest
start to be eluted. An elution volume of 1.5mL in four aliquots of 500µL eluted off the compounds of
interest to give a high recovery level.
HyperSep Retain-CX
Using a 30mg 1mL cartridge
(part number 60107-301)
1. Condition with 1mL of 0.1% formic
acid in methanol followed by
re-equilibrium using 1mL of
0.1% formic acid in water
2. Load 1mL of 500ng/mL samples in
0.1% formic acid in water
3. Wash with 1mL of 0.1% formic acid
in water
4. Wash with 1mL of 0.1% formic acid
in methanol
5. Wash with increasing strengths of
5% ammonia in methanol/water, starting
with 20% methanol/80% water to
80% methanol /20% water, increasing
methanol content by 10% each time
6. Elute 4 times with 0.5mL of
5% ammonia in methanol
HyperSep Retain-CX Wash/Elution Profile
HyperSep Phenyl
Using 100mg 1mL cartridge
(part number 60108-386)
1. Condition with 1mL methanol
followed by 1mL of water
2. Load 1mL of 500ng/mL sample
in water
3. Wash with increasing strengths of
methanol from 10% to 90%
4. Elute 4 times with 0.5mL
100% methanol
HyperSep Phenyl Wash/Elution Profile
Conclusion
•
For second (organic) wash, choose the strongest solution where no compound breakthrough occurs
•
For elution step, use a solution stronger than where all the compound of interest is eluted
•
NB: when choosing these solutions allow some margin for error