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Gradient separation-Calculating final hold time
Posted: Tue Apr 05, 2011 1:32 pm
by LMNAIR
Can anyone help me how to calculate the final hold time (100%B) in a gradient separation? Just give me a general idea how it is calculated.
Thanks,
LMN
Re: Gradient separation-Calculating final hold time
Posted: Tue Apr 05, 2011 4:18 pm
by skunked_once
A good rule of thumb is to flush with 10 column volumes at 100% B. You will need to know the column volume and the flow rate to determine how much time to flush. For example, a 1.5-mL column volume at at flow rate of 1 ml/minute would require a 15 minute final hold time. You could reduce the time by doubling the flow at the end of the run if the double flow does not exceed the pressure limit of your procedure.
Re: Gradient separation-Calculating final hold time
Posted: Tue Apr 05, 2011 5:43 pm
by LMNAIR
Thank you very much.
LMN
Re: Gradient separation-Calculating final hold time
Posted: Tue Apr 05, 2011 8:33 pm
by JGK
Can anyone help me how to calculate the final hold time (100%B) in a gradient separation? Just give me a general idea how it is calculated.
Thanks,
LMN
I'd set up an injection of a typical sample and treble the run time. run with the desired gradient and leave running at 100% B until finished. You should be able to assess when the last of the material from the injection is removed from the column from this.