By Anonymous on Monday, March 8, 2004 - 10:04 am:
I'm new in the area of LC/MS. I was told that if a LC method uses a 4.6 mm ID at a flow rate of 1 ml/min, one needs to switch to a narrow bore column at a reduced flow rate or use a flow splitter for LC/MS applications. However, in the manual of the instrument I want to use, typical values of vaporiser temperature are given for different flow rates, including a flow rate of 1 ml/min in APCI mode. Does this mean that in some cases, a 4.6 mm ID column can be used at a flow rate of 1 ml/min for LC/MS in APCI mode?
By Basil on Tuesday, March 9, 2004 - 06:10 am:
I'm using a 4.6 mm ID with ESI and APCI only for qualifying new impurities (development process or Stress Testing impurities)
If you want to quantify it would be better to use a narrower column.
By mgoodwin on Tuesday, March 9, 2004 - 07:01 am:
Electrospray works better with slow flow rates. (I believe due to the formation of smaller droplets that can readily undergo coulombic dissociation)
Conversely, APCI works well with normal flow rates like 1 ml/min. (more solvent available to ionized)
In fact, at 0.2mL/min flow, I experienced a noise problem. When calling the tech support, they told me a minimum flow of 0.5 mL/min is needed to adequately operate the APCI.
By Tony Larson on Tuesday, March 9, 2004 - 07:22 am:
Hello,
I have recently tried scaling different HPLC analyses between 250 x 4.6mm and 150 x 2.0mm columns, using a scaling factor for the flow rate based on the formula flow2=flow1 x length2/length1 x (radius2/radius1)^2. By doing this, I get the same retention times when scaling the flow rates appropriately for the 2 columns, but peak widths are no narrower and resolution is compromised for the narrower bore columns (I am observing loading limits for the narrower column and keeping injection volumes at 5 uL for both). I thought this might be due to extra column dead volume, but even at the same flow rates (the lower rates), the resolution is better on the larger columns (with markedly longer run times). My question is this: is there an effect of increased cross sectional area of the stationary phase in a given column diameter that accounts for improved peak resolution in wider-bore columns? If not, what else might be the reason for the lower resolution in the narrower-bore column?
thanks
Tony
By Tom Mizukami on Tuesday, March 9, 2004 - 11:12 am:
Hi Tony,
Did you also go to a smaller particle diameter when you moved to a shorter column? If not, you would expect a lower plate count and poorer resolution with the shorter column. Normally you you just scale flow rate with the cross sectional area to keep the linear velocity constant. You would have less retention and shorter runtimes with the shorter column.
There is no direct effect between the cross sectional area and resolution. However as the cross sectional area decreases the volume you can inject without affecting resolution also decreases. 5uL may be too much for your narrower column for optimum performance. Also as you noted as column diameter drops so does the flow rate and extra column volume has a greater effect. Calculate your peak volume for peaks on your narrow id column and compare to extra column volume and detector cell volume. You may need to a smaller volume cell and narrower bore tubing to see the best performance at the low flow rates. Good luck.
By RH on Thursday, March 11, 2004 - 02:33 am:
APCI in fact works well with high flowrates. Electrospray is ideal for lower flow rates as the detection is concentration dependent and so a down-scale will improve sensitivity due to lower peak widths and higher signal to noise ratio. But as Tom stated downscaling chromatography is sometimes troublesome. With shorter columns the particle size must be reduced and with narrow-bore columns the lower flow rates can bring along adverse extra-column effects.
For LCMS there was a special issue of LCGC Europe in Dec. 2001:
http://lcgceurope.adv100.com/lcgceurope/issue/issueDetail.jsp?id=538