Chromatography Forum

I've extensively googled this question and found no adequate response. Is it necessary to minimize the volume from the column to the MS when using low flow rates ~0.2ml/min? If it does not, please explain why. thx

Necessary is a strong word but if you're using excessively long tubing it will effect your chromatography and eliminating the extra length might save you headaches later. In practice the tubing tends to be such narrow diameter the volume is ~almost~ negligible but all the problems associated with excessive dead volume are multiplied as flow rate decreases because for easy maths if the sample was in the tube for 1 second at 1ml/min it's in the tube for 5 seconds at 0.2 ml/min.

I've seen some comedy lengths when bench space and source location hasn't been well considered but nothing ridiculously spectacular.

Anyone else seen anything truly ridiculous?

not ridiculous in the sense of so impractical as to send a neutral observer into hysterical laughter, but Thermo scored a daftness-hit with their first couple of ion traps. The DecaXP up to the LTQ had the spray chamber on the front of the instrument, towards the left side, but with the spray probe accessed from the right. If you installed the HPLC to the right of the instrument, the location of the spray chamber demanded an extra 40cm of tubing, if you installed the HPLC to the left of the instrument (which was normal) then the fact the spray probe inlet was on the wrong side of the chamber required an extra 40cm of tubing.

The daft thing was that the spray chamber was otherwise more-or-less symmetrical, and it would have been quite easy to make it accessed from the left. If you're going to design a superbly complex and sensitive bit of kit that sells for the modern equivalent of half a million quid (similarly lots of dollars) you might just as well take the trouble to get simple, easy things right. Thermo were never any good at HPLC; they were always a mass spec company who happened to stick an HPLC on the front.

"I've extensively googled this question and found no adequate response. Is it necessary to minimize the volume from the column to the MS when using low flow rates ~0.2ml/min? If it does not, please explain why. thx "

Google is not an experienced chromatographer so expecting to receive an answer is hopeful at best. A trained chromatographer will tell you that the FLOW rate is not the issue at all. The connection between the Column outlet AND the detector inlet should be: As short as possible AND as low in volume as possible. Column volume will determine how important this is in actual use (A 2.9mL in volume 4.6 x 250mm column is less impacted vs a 2.1 x 100mm column)..
There is a scientific reason for this concern and why we optimize the tubing flow path. Diffusion. You just cleanly separated and resolved those peaks into nice, narrow Gaussian peak(s) of low volume on the column. Now you want to keep them that way when they enter the detector (flow cell or ion chamber). Diffusion is taking place the entire time the sample "slug' is traveling down the tubing towards the detector and the slug is spreading out (This has been studied for decades). Normally, we use a narrow capillary of low internal volume to accomplish this. Please review this article for more info: "HPLC Capillary Tubing Connection Volumes"; https://hplctips.blogspot.com/2012/10/c ... lumes.html
Please keep in mind that if you use a column with very small internal volume, then the use of narrow, short connections everywhere in the flow path (pump to injector, injector to column, column to detector(s)) is critical to get right. Columns with tiny ID's or short lengths means the total column volume is low and the sample slugs that are eluted off them are also tiny so diffusion will work quickly to spread them out and reduce the signal. COLUMN dimensions must always be reviewed to insure they are compatible and OPTIMIZED for the application.This means the HPLC plumbing (all of the tubing IDs and Lengths) need to be optimized for diffusion to be minimized and peak integrity maximized. This is something that we teach to intermediate level users so they can learn how to optimize their HPLC system (or LC-MS system, same thing) to run different applications. Failure to optimize the flow path of an HPLC system for an application often results in poor quality results or loss of separation.

Column Volume: "Determination of HPLC Column Void Volume / Dead Volume, Dead Time (T zero)"; https://hplctips.blogspot.com/2011/05/d ... -time.html

PS: My boss tells me that he has seen many LC and LC-MS systems that connected their column outlet to their MS inlet using two-meter long 0.17mm or 0.25mm ID lines resulting in very poor quality results. *Keep in mind that changing from 0.17mm ID to 0.25mm ID doubles the volume inside the tubing! Moving some modules around and switching the tubing over to one with a narrow bore immediately showed a signal improvement of 4x with better peak shapes, better separation and better results.