by
Hollow » Mon Apr 25, 2022 9:16 pm
Good monday
Maybe the column dimensions would be a helpful information too.
But...
...washed it with 0.025 M H2SO4 at a flow of 0.2 mL/min for about 24 hours at 85-degrees C.
Swapped it over to nano pure filtered water until it equilibrated. Let it run for another 4 hours. The baseline looked pretty solid.
I loaded a few known samples (Glucose, Xylose, Erythrose) turned the flow up to 0.5 mL/min and dropped the temp to 65-degrees...
...is a bit confusing, especially the last sentence and no clear reference to the pictures.
1) if you changed the temperature and flow just shortly before your injection, then your system is in non-equlibrated state. The flow may be a small issue but the temperature will take a while to reach 65°C.
> the pressure trace may be ok but has increased by about 50% between the two pictures, if looked closely.
>> let the system run at the same temperature as your separation method. Low flow is ok, then ramp up to working flow and wait until the pressure is stable again. Then inject your samples.
I would even keep the temperature for (overnight) standby after the sequence, as thermal equilibration is slow.
2) do you have a backpressure regulator at the RID outlet? This may help for steady baseline and signal.
If not, add a dedicated one (there are posts with links on other topics in this forum) or just connect a piece of smallbore tubing - but make sure it stays within the limits of the RID cell under all working and flushing conditions (flow, temperature, solvent)! You may also want to put a big, red warning label with the maximum flow on the PC screen.
Maybe calculate the length of the tubing with the viscosity of MeOH 50% and only about half of the limit of the cell, to stay on the safe side).
The pressure drop can be calculated by the Hagen-Poisseuille equation or use some online calculators e.g. the one on the Waters website:
https://www.waters.com/webassets/other/ ... itters.htm