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Discussions about HPLC, CE, TLC, SFC, and other "liquid phase" separation techniques.
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Can someone refresh my memory about who makes a couple of products for polishing water prior to use with HPLC? One is an in-line organic filter that can be used with either low- or -high-pressure-mixing systems. I thought this one was from Whatman, but can't locate it in their info. The second is a device that can be placed on top of the reservoir, something like a giant C18 SPE cartridge. Water is poured through this to give a final extraction of organics before it goes in the reservoir. Someone from Waters mentioned this one to me, but I can't find info on it, either. Terrible when the brain cells quit working. Thanks in advance -- John Dolan
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Both 3M and Alltech make extraction disks which can be used to clean up aqueous mobiel phases. These are used in place of a mobile phase filter.
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Andy, do you happen to have part numbers or a product name? Where are these mounted in the flow stream? Thanks - John Dolan
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John, are you sure that you don´t add more junk than you remove with those things?
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HW: I've also been somewhat skeptical about these things, but I've heard enough people claim success that I would like to mention them as options in a book chapter I'm writing. We've used guard columns to cleanup water prior to LC use, so there is no reason to think that the in-line RP cartridge shouldn't do the same thing.
Anyone out there who has had success with either type of device? -- John Dolan
Anyone out there who has had success with either type of device? -- John Dolan
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I am glad to see you here, Mr. Dolan. What is the book you wrote this time? New edition of Troubleshooting LC System or Practical HPLC Method Development?
Regards, SYX
Regards, SYX
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The use of in line C18 cartridges for our proteomic applications succesfully decreased/eliminated problems from occasional mobile phase contamination (mainly from the aqueous mobile phase). The cartridges we use are self packed and withstand very high pressures (we use home build UPLC systems).
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Right after finishing browsing through the forum I will compare MilliQ water, recently obtained, to an older batch, using my UV detector. (Actually it was not easy to find a method to do this, but now the water, etc., is placed in a ~100 mL empty column, evacuated for 10+ minutes, then forced through the detector for at least 20 min by applying N2 gas. Incidentally, this method showed that "degassing", really trans-gassing, with He causes strong scattering which looks like an absorption. This was discussed some time ago.)
So: Just wondering how people using pre-something (especially filtering type devices) know whether, first of all, their device is clean, and secondly, when they have breakthrough of organics.
So: Just wondering how people using pre-something (especially filtering type devices) know whether, first of all, their device is clean, and secondly, when they have breakthrough of organics.
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According to my personal experience, the best way to minimize baseline spikes (due to water issues) is to pretreat the water destined to go into MP "A" with a C-18 membrane (vacuum filter it through). Breakthrough is prevented by filtering the water 1L at a time and flushing a bit of MeOH w/ a water chaser through the membrane between liters of water.
A guard column between the "A" pump and mixing chamber (high pressure mixing gradient systems only) works, but not as well, especially if the A phase has more than a trace of organic solvent in it.
I have seen large improvements in baselines by employing this water pretreatment regimen, especially when running at or below 220nm. It takes a while to pull the water through these membranes, but it works very well.
A guard column between the "A" pump and mixing chamber (high pressure mixing gradient systems only) works, but not as well, especially if the A phase has more than a trace of organic solvent in it.
I have seen large improvements in baselines by employing this water pretreatment regimen, especially when running at or below 220nm. It takes a while to pull the water through these membranes, but it works very well.
Thanks,
DR

DR

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DR,
what do you mean by "large improvement"? Less spikes or a lower baseline level? How do you get spikes anyway with organics in H2O? Are you talking about gradient stuff or some switching of some kind?
what do you mean by "large improvement"? Less spikes or a lower baseline level? How do you get spikes anyway with organics in H2O? Are you talking about gradient stuff or some switching of some kind?
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I'm speaking of those annoying little (sometimes not-so-little) peaks that show up over the course of gradient runs. The ones that get larger if you equilibrate at initial conditions for a longer period of time between injections - they are from junk in the water (probably water polishing system detritus that made it through the final filter - an occasional styrene monomer or such thing).
I have seen such peaks get markedly smaller as a result of Empore disk pretreatment the water going into a pot of A phase. The gradient profile for blank injections looks the same, except for the shrinkage of the water related peaks that usually show up near the end of the gradient.
PS - I tend to use Empore Extraction Disks (3M product, available through Varian Chromatography).
I have seen such peaks get markedly smaller as a result of Empore disk pretreatment the water going into a pot of A phase. The gradient profile for blank injections looks the same, except for the shrinkage of the water related peaks that usually show up near the end of the gradient.
PS - I tend to use Empore Extraction Disks (3M product, available through Varian Chromatography).
Thanks,
DR

DR

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DR,
maybe you got rid of air? Or did you do MS and know these peaks are organic? (I imagine you would have to be very careful and take an MS just before the start or after the end of peaks as well?)
Or did you get a UV spec "proving" organics?
maybe you got rid of air? Or did you do MS and know these peaks are organic? (I imagine you would have to be very careful and take an MS just before the start or after the end of peaks as well?)
Or did you get a UV spec "proving" organics?
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Do peaks that are the result of air get larger for blank injections if the system is left running at initial conditions longer between injections? Would vacuum filtering water through an Empore disk remove that much more air than vacuum filtering mobile phases through 0.5µ nylon membranes? Would this difference continue to be obvious even after running both through inline vacuum degassers on LCs from 3 different vendors?
If you have not seen the benefits of pretreating water with a C-18 filter, you should count yourself lucky that you have excellent water for your mobile phases before you doubt that others have reaped the benefit of these prep. tactics in the face of inferior water sources.
If you have not seen the benefits of pretreating water with a C-18 filter, you should count yourself lucky that you have excellent water for your mobile phases before you doubt that others have reaped the benefit of these prep. tactics in the face of inferior water sources.
Thanks,
DR

DR

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Hi DR,
May I get to know what water source you've been using?
Just in case that some water source sounds good but actually needs to get pretreated as you mentioned.
Thanks!
kiknos
May I get to know what water source you've been using?
Just in case that some water source sounds good but actually needs to get pretreated as you mentioned.
Thanks!
kiknos
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- Joined: Mon Aug 30, 2004 7:17 am
DR,
I don´t know the answers to your questions, though if you did everything that your questions imply it appears unlikely that air is the primary factor for your peaks. However, having spend all kinds of time tracking the source of peaks which turned out to be due to gas, or disturbances, and knowing that if one changes a procedure (in your case the inclusion of the disk..) one often inadvertendly changes other things as well, I would feel more convinced if MS or at least UV spectra of the peaks were obtained which are also seen in the water.
I don´t know the answers to your questions, though if you did everything that your questions imply it appears unlikely that air is the primary factor for your peaks. However, having spend all kinds of time tracking the source of peaks which turned out to be due to gas, or disturbances, and knowing that if one changes a procedure (in your case the inclusion of the disk..) one often inadvertendly changes other things as well, I would feel more convinced if MS or at least UV spectra of the peaks were obtained which are also seen in the water.
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