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By WK on Thursday, July 29, 2004 - 05:59 am:
Does anyone use a 250 or 300mm x 3um x 4mmid C18 column routinely?
Can you get 1.5 times the plates out of it compared to a 5um particle? (With same flow/dimensions)
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By DR on Thursday, July 29, 2004 - 06:28 am:
You could, in theory, get those kinds of plate numbers with a 3µ packing in a standard jacket, but you would also, in theory, have to find a pump capable of pushing eluent through >8000psi to get anything approaching a useful flow rate.
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By Uwe Neue on Tuesday, August 3, 2004 - 04:32 pm:
The best plate count on these columns is reached at a backpressure of around 2000 psi. For maximum performance, the backpressure itself is not really the problem. The real problem is that the plate-count optimum comes at a slow velocity. In water, the retention time of an unretained peak would be around 7 minutes, making an analysis awfully long.
PS: the expected plate count is above 40 000 plates for a 30 cm column.
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By JA on Monday, August 9, 2004 - 05:41 am:
where can we find the optimum (HETP) flow rates for a 150 x 4.6mm, 5um and 3um column?
are there any difference for different lengths of column?
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By Alex on Tuesday, August 10, 2004 - 12:59 am:
Uwe, I disagree. Years ago I have tested a YMC 3µm 25 cm column at 0.7 ml/min and lower flow rates (down to 0.07 ml/min). At 0.7 I had a back pressure of ca. 380 bar. Plate count went down for lower flow rates.
Alex
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By tom jupille on Tuesday, August 10, 2004 - 10:18 am:
To JA: If you know the linear velocity at which you have the minimum HETP, you simply multiply that by the cross-sectional area of the column to get the flow rate. As to finding the optimum linear velocity, that depends on analyte molecular weight, mobile phase viscosity, temperature, stationary phase pore size distribution, stationary phase chemistry. Usually much easier to simply measure efficiency at different flow rates. As Uwe pointed out, in many cases the optimum HETP occurs at such a low linear velocity (flow rate) that run times are unacceptably long.
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By JA on Tuesday, August 10, 2004 - 03:07 pm:
ty Tom. looks like a couple of variables I didn't take account of before posting, lol.
My original question was also angled towards whether or not there's a different "typical" flow rate for 3um vs. 5um particle columns, in a similar way to how we scale flow rates down for smaller i.d's.
In our lab we generally kick off development with 1.0 ml/min for 4.6mm and 0.2 ml/min for 2.1mm. Should we therefore initially try 0.7 ml/min for 3.9mm columns, or are these not really ideal?
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By tom jupille on Tuesday, August 10, 2004 - 09:56 pm:
Okay, I see where you're coming from. To grossly oversimplify, if you're dealing with small molecules, the HETP vs linear velocity curve is flatter for smaller particles. For 3-micron packings, you just don't see much of a plate loss as you increase the flow rate from the optimum.
Bottom line is to run at the highest flow rate you can get away with based on your pressure limitations.
As to column diameter, what you're doing is exactly right: scaling the flow rate to the square of the diameter maintains constant linear velocity .
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By Uwe Neue on Sunday, August 15, 2004 - 06:46 pm:
Alex,
The estimate that I made above assumed that the performance of the column is reasonable, and that the molecular weight of the sample is low (I assumed a MW of 100). For a MW of 300, the optimum is at about 1000 psi. Only for a MW of over 1000 do I get into the pressure range that you described. The assumptions underlying this estimate is that the interaction of the analyte with the stationary phase is normal, and does not show hindered mass transfer due to secondary effects. If there is hindered mass transfer, then it is quite easily possible that the pressure and the velocity for maximum plate count is lower.
Bottom line: your findings are quite possible, but I would not consider them to be normal for a normal small molecule.
Uwe
Does anyone use a 250 or 300mm x 3um x 4mmid C18 column routinely?
Can you get 1.5 times the plates out of it compared to a 5um particle? (With same flow/dimensions)
-------------------------------------------------------------------------------------------------------
By DR on Thursday, July 29, 2004 - 06:28 am:
You could, in theory, get those kinds of plate numbers with a 3µ packing in a standard jacket, but you would also, in theory, have to find a pump capable of pushing eluent through >8000psi to get anything approaching a useful flow rate.
-------------------------------------------------------------------------------------------------------
By Uwe Neue on Tuesday, August 3, 2004 - 04:32 pm:
The best plate count on these columns is reached at a backpressure of around 2000 psi. For maximum performance, the backpressure itself is not really the problem. The real problem is that the plate-count optimum comes at a slow velocity. In water, the retention time of an unretained peak would be around 7 minutes, making an analysis awfully long.
PS: the expected plate count is above 40 000 plates for a 30 cm column.
-------------------------------------------------------------------------------------------------------
By JA on Monday, August 9, 2004 - 05:41 am:
where can we find the optimum (HETP) flow rates for a 150 x 4.6mm, 5um and 3um column?
are there any difference for different lengths of column?
-------------------------------------------------------------------------------------------------------
By Alex on Tuesday, August 10, 2004 - 12:59 am:
Uwe, I disagree. Years ago I have tested a YMC 3µm 25 cm column at 0.7 ml/min and lower flow rates (down to 0.07 ml/min). At 0.7 I had a back pressure of ca. 380 bar. Plate count went down for lower flow rates.
Alex
-------------------------------------------------------------------------------------------------------
By tom jupille on Tuesday, August 10, 2004 - 10:18 am:
To JA: If you know the linear velocity at which you have the minimum HETP, you simply multiply that by the cross-sectional area of the column to get the flow rate. As to finding the optimum linear velocity, that depends on analyte molecular weight, mobile phase viscosity, temperature, stationary phase pore size distribution, stationary phase chemistry. Usually much easier to simply measure efficiency at different flow rates. As Uwe pointed out, in many cases the optimum HETP occurs at such a low linear velocity (flow rate) that run times are unacceptably long.
-------------------------------------------------------------------------------------------------------
By JA on Tuesday, August 10, 2004 - 03:07 pm:
ty Tom. looks like a couple of variables I didn't take account of before posting, lol.
My original question was also angled towards whether or not there's a different "typical" flow rate for 3um vs. 5um particle columns, in a similar way to how we scale flow rates down for smaller i.d's.
In our lab we generally kick off development with 1.0 ml/min for 4.6mm and 0.2 ml/min for 2.1mm. Should we therefore initially try 0.7 ml/min for 3.9mm columns, or are these not really ideal?
-------------------------------------------------------------------------------------------------------
By tom jupille on Tuesday, August 10, 2004 - 09:56 pm:
Okay, I see where you're coming from. To grossly oversimplify, if you're dealing with small molecules, the HETP vs linear velocity curve is flatter for smaller particles. For 3-micron packings, you just don't see much of a plate loss as you increase the flow rate from the optimum.
Bottom line is to run at the highest flow rate you can get away with based on your pressure limitations.
As to column diameter, what you're doing is exactly right: scaling the flow rate to the square of the diameter maintains constant linear velocity .
-------------------------------------------------------------------------------------------------------
By Uwe Neue on Sunday, August 15, 2004 - 06:46 pm:
Alex,
The estimate that I made above assumed that the performance of the column is reasonable, and that the molecular weight of the sample is low (I assumed a MW of 100). For a MW of 300, the optimum is at about 1000 psi. Only for a MW of over 1000 do I get into the pressure range that you described. The assumptions underlying this estimate is that the interaction of the analyte with the stationary phase is normal, and does not show hindered mass transfer due to secondary effects. If there is hindered mass transfer, then it is quite easily possible that the pressure and the velocity for maximum plate count is lower.
Bottom line: your findings are quite possible, but I would not consider them to be normal for a normal small molecule.
Uwe
