Repeat injection issues

[Pear]

Hi,

I'm noticing a strange behaviour from my HPLC when I inject the same sample repeatedly.

I have my system calibrated with know standards at:
0.1 ppm
0.2 ppm
0.3 ppm
0.4 ppm
0.5 ppm

The calibration is very accurate and gives good results.

However, when I run any sample, for example I did my 0.3ppm today, repeatedly, my results vary.

Real results from today:

0.3ppm calibration result = 0.3098ppm

But when I run this sample 3 times, I get the following:

1: 0.3020 ppm
2: 0.2910 ppm
3: 0.2830 ppm

If I continue, the sample gets lower and lower. Even if I do sample > blank > sample > blank, the results are the same.

Can anybody offer any input? Thankful for any help at all.

[Rndirk]

Welcome to the forum.

Can you tell us what compound you are analyzing?

If you reinject a calibration standard after repeated sample injections, does it give a lower area as well?

[Pear]

Welcome to the forum.

Can you tell us what compound you are analyzing?

If you reinject a calibration standard after repeated sample injections, does it give a lower area as well?

Hi Rndirk, thanks for the reply.

I'm analysing formaldehyde. The calibration standard is the re-injection 'sample' that gives a lower result each time.

[uzman]

If you are using NASH reagent for post-column derivatisation , you have to protect your reagent from light , it is light sensitive.

[Pear]

If you are using NASH reagent for post-column derivatisation , you have to protect your reagent from light , it is light sensitive.

Hi uzman,

We do pre-injection derivitisation. The reagent is kept in the dark at all times.

[uzman]

Hi Pear ,

Is your injection solvent weaker than your mobile phase ; if you are using a RP column ?
If not , you may try to dissolve your sample in mobile phase and inject.

[Pear]

Hi Pear ,

Is your injection solvent weaker than your mobile phase ; if you are using a RP column ?
If not , you may try to dissolve your sample in mobile phase and inject.

Hi uzman,

my mobile phase is 0.1% phosphoric acid + ACN: 0.1% phosphoric acid + water (45:55), the sample is added to 9:1 ACN:water then injected.

[uzman]

Hi Pear ,

Your injection solvent is stronger than your mobile phase , you may try mobile phase as injection solvent.

[Pear]

Hi Pear ,

Your injection solvent is stronger than your mobile phase , you may try mobile phase as injection solvent.

Hi uzman,

Thanks again for your help. We tried injections using mobile phase as the sample solvent, yet we are still seeing a decrease in result every injection.

Scratching my head over this one.

[uzman]

Some more suggestions :

First , try to backflush your column with pure acetonitrile , accumulated junk on the column might be the reason.

You may also switch to methanol as your mobile phase , instead of acetonitrile.

Formaldehyde ( as methylene glycol , in aqeous solutions ) is more stable in methanol.

[lmh]

just out of curiosity, if you take two vials, put the same standard in both, put them both in the autosampler, but inject from only one (repeatedly), if after a load of injections you then inject from the other, is it low or normal? I.e., is this a stability problem, or is it connected with making injections from the vial?