Aminex HPX 87P regenaration

[nguyenthanh]

Dear all
I am newbie in HPLC and we have a pressure problem with our 87P column. Hope someone can help us to find out the solution. We are trying to follow the suggestion from Biorad like reverse flush...However our question is how to prepare the lead nitrate 0.1M with pH 4. Cause when we make this solution, the pH was already around 3 (in accordance to this information http://en.wikipedia.org/wiki/Lead(II)_nitrate). As recommendation from Biorad, user can use nitric acid. It sound strange.
We would really appreciate any help.

Thank you

Merry Christmas and Happy New Year all

[tom jupille]

Just go ahead and use the lead nitrate. The pH is close enough.

[unmgvar]

is it a new column, or an old one that has done already many injections?
if it is a new one, then maybe the column was put to high temperature too fast.
if it is an old column, than maybe nothing will help and you need a new one

[nguyenthanh]

Just go ahead and use the lead nitrate. The pH is close enough.

Thanks Tom Jupille for your suggestion. Actually, the pH range of operation (for this column) is announced by company of 5 -9, therefore I am a little scare with pH 3.

Thanh

[nguyenthanh]

is it a new column, or an old one that has done already many injections?
if it is a new one, then maybe the column was put to high temperature too fast.
if it is an old column, than maybe nothing will help and you need a new one

is it a

Hi unmgvar

The column is quite new, just around 20 injections. We used 60oC for separation of oligosaccharide mixture (with Agilent 1200). I don't think it is too fast heated up.

Thanh

[unmgvar]

how did you storage it?

[nguyenthanh]

I store in fridge, so could not because of microorganism

[tom jupille]

Thanks Tom Jupille for your suggestion. Actually, the pH range of operation (for this column) is announced by company of 5 -9, therefore I am a little scare with pH 3.

The concern with low pH is leaching Pb off the column. That's not an issue when you're using a Pb(NO3)2 solution. Just gently backflush the column.

[nguyenthanh]

thanks a lot Tom. I will try and inform you what is going on

Best

Thanh

[push]

Are you sure that your problem is really high pressure?
Aminex -87H column is a special one. You have to increase the flow rate step by step: 0.1 - to wait for the prssure stabilization - 0.2 ....0.8 ml/min.
Good luck

[nguyenthanh]

Hi Push, I am sure the increase of back pressure.

Guys, what can I do if a little H2SO4 5mM was flushed through the 87P column (the precipitation of PbSO4 will block column). Do you think the column still "alive" ?

Thanks

Thanh

[tom jupille]

That will kill the column.

[nguyenthanh]

Hi Tom and every one

I would like to back this column, maybe you have some advice.

The column, we regenerated with PbNO3 0.1M (as mentioned in manual) and tried to separate the mixture of C5, C6 sugar. The separation look good ) However, the pressure is really high (for example, originally we could use 0.6 ml/min, 60oC and P = 52 bar; but now at the same temperature, even low flow rate 0.25 ml/min, the P was up to 92 bar). It is strange that when I reduced the flow rate to 0.1 ml/min, first the pressure was lower, but then increased gradually.

I supposed in the column it has precipitation of PbSO4 (see my previous post), and therefore during the flow of mobile phase, the precipitation blocked the outlet frit of column.
1. Do you think if it is correct?
2. If yes, could I clean the outlet frit by sonication? or replace the frit?

Thanks for any help

[tom jupille]

The precipitation will occur where the SO4= and the Pb++ ions first come in contact: at the top of the resin bed. If the pressure is so high that the column is not usable, then by all means try to change the frit. It probably won't help and it may make things worse, but you have nothing to lose. If the column is still usable, but you are worried about the pressure, I would continue to use it. Unless you work for free, you can very easily spend more in time to fix the problem than you would in money to get a new column.

"Columns are cheap; your time is valuable."

[nguyenthanh]

Hi Tom

Thanks for your suggestions. Maybe we will use the column, until it is completely " died"

Thanh