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- Posts: 252
- Joined: Sat Nov 07, 2009 6:27 pm
I'm interested in trying different mobile phase as a way to improve selectivity in a multi-step purification scheme.
Peptide standards look fine when run on PLRP with 0.1% TFA and a linear gradient 5-60% MeCN. However, peak shape dramatically deteriorated when 15 mM NH4OH was used in place of TFA. What would cause this?
Perhaps the equilibration time must be longer?
Loading sample on a column equilibrated with 5% MeCN 15 mM NH4OH
My sample volume is 0.4 mL in 10 mM NH4OAc + 250 mM NaClO4, 5% MeCN.
After loading the 0.4 mL sample on to this 1 mm ID PLRP column at 50 ul/min, how long will it take to re-equilibrate with ammonia before beginning the gradient?
Peptide standards look fine when run on PLRP with 0.1% TFA and a linear gradient 5-60% MeCN. However, peak shape dramatically deteriorated when 15 mM NH4OH was used in place of TFA. What would cause this?
Perhaps the equilibration time must be longer?
Loading sample on a column equilibrated with 5% MeCN 15 mM NH4OH
My sample volume is 0.4 mL in 10 mM NH4OAc + 250 mM NaClO4, 5% MeCN.
After loading the 0.4 mL sample on to this 1 mm ID PLRP column at 50 ul/min, how long will it take to re-equilibrate with ammonia before beginning the gradient?
