Chromatography Forum

Dear all
I am assay the impurity of a drug product,and use the
column:Agilent ZORBAX SB-C18,4.6*150mm,5um,
buffer:0.2%dibasic potassium phosphate adjust to pH6.0 with diluent phosphoric acid;
mobilephase A:buffer-ACN(95:5)
mobilephase B:ACN
and mobilephase follow as below:

min A B
0-3 100 0
19 79 21
20 100 0
25 100 0

this method come from PF.
when I have injected about 200 ,the back pressure grows up about 30bars,and the perfoemence lose,I clean the column with ACN,methonal and back-flush it,the back pressure still high,
there is hypromellose in my test solution,is it cause the high back pressure?I am so puzzled.who can help me?
if you have suggestions,you can give me Email,thank you very much!

it is the hypromellosee that's causing trouble. Get rid of it prior to loading the sample on the column.
A simple filtration might do the trick. The hypromellose is not in actual solution - it's a suspention.

Best Regards

it is the hypromellosee that's causing trouble. Get rid of it prior to loading the sample on the column.
A simple filtration might do the trick. The hypromellose is not in actual solution - it's a suspention.

Best Regards

thank you！
but i have filtrated the sample before injection.

OK. Filtered using wich kind of filters?
Btw. what is the sample solvent?

Best Regards

OK. Filtered using wich kind of filters?
Btw. what is the sample solvent?

Best Regards

the solvent is buffer-ACN(95:5)
and the filter is oganic filter,0.45um

Try 0.22 um pore size filter. Also make sure the filter material is somewhat hyrophobic.
If that doesn't work you might like to consider SPE.

Best Regrds