Thermo Hypercarb column high back pressure

[HaruYap]

Its quite logical from your explanation, Mueller. Thanks. I will try to check again the entire system itself for other external factors contributing to such discrepancies in the run.

FYI, prior to this sudden increase in pressure, the system runs just fine. The pressure increased happened one day and ever since, it has been in the region of 1400psi; up from the initial 1100psi that I've mentioned. I am just wondering how come the pressure just changed like that and has remained ever since? It seems that; like what I've mentioned. something is blocking the column; thus the pump works harder! I didnt changed my solvent systems nor changed my sample prep method.

Bruce, my sample prep would be by using solvent solvent extraction. Solvent of concern is diethyl ether; aqueous layer will be acidified water to remove intereferences from the sample. The organic layer will be dried under a stream of purified N2 before reconstitution using water. The sample extract will then be filtered through a 0.20um syringe filter before injection into the HPLC.

[Bruce Hamilton]

If the method has worked fine in the past and suddenly increased one day, it's most likely either a dirty sample or mobile phase. It's unlikely to be your analyte or other component binding, as that would be a gradual increase.

It seems like you had a custard day, and something went wrong. I'm not sure what you initial mobile phase composition is, but I would make up samples in that before filtering, then it's less likely you'll carry some junk through.

I'd also use a guard column. If it leaked, return it for credit, or if you assembled it and the seals are OK, go to the gym and purchase larger spanners .

Bruce Hamilton

[rhaefe]

Well, we don't really know if there is a flow restriction. According to the first post, back pressure noted on the QC chromatogram was 1100psi and now it is 1400psi which in my experience is not very high.
The problem is disappearing peaks if I read the posts correctly.

cheers,

[HaruYap]

You were right, Rhaefe. I was wondering if the high pressure is due to something blocking the column hence the 'disappearing peaks' from the chromatogram. Anyway, can the column be 'recondition' again to lower backpressure? I have posted in the earlier posts about cleaning the column with ipa: water as well as THF but the pressure still remains. So; just hoping if anyone can tell me a better way than to change the frit since Thermo did not recommend that. I have not tried flowing it in the reverse order; a little doubt that I might blocked the frit end as well. As for guard column; Bruce, maybe I'll have to go for the GYM and get a BIGGER spanar... if the guard turned out to be fine.

Disappearing peaks? ermm...can anyone give me some insights??

[HW Mueller]

Robert, did a problem of definition creep up? I differentiated between flow restriction and flow rate, a flow restriction (increase of resistance) being compensated by the pump so that the flow rate stays constant.
I think Uwe already told Haru Yap to look elsewhere for the source of the problem.
Repeating myself: It could be due to a drastic change in cations on the stat. phase, or sticky dirt, or a stability problem. Now one may add that the extraction may be faulty? Or do fresh standards at different concentrations also disappear on the column? You can check on the integrity of the standards via a UV spec. (for instance in the detector by injection without the column, careful that you don´t get air in the cell!).

[rhaefe]

Hans,

no problem here. The pump (if working properly) will of course deliver the correct flow rate and if a flow restriction exists the back pressure will rise. I agree with you that it might be a contaminant that interacts (binds) with the analyte.
If the contaminant would be inert towards the analyte HaruYap would still see peaks (although they would be probably distorted).

cheers

[Harry Ritchie]

I have observed that sometimes when columns are disconnected and re-connected, the tightening of the fittings can sometimes crimp the tubings in the flowpath. This can have the effect of decreasing the internal diameter of the flow channel and thus increase observed pressure.

Just something else to check.