Chromatography Forum

I am facing a problem in related substances method. Carry over of main peak is observing (about 5000 area) in blank after the sample solutions injection. In second "blank injection" from a fresh vial it's reducing to 2000 area. However, there is no problem with bracketing standard(standard area is 40000) in this case.
But, when iam injecting a number of samples (eg 10 samples), in that case 4 to 5 "blank injections" are needed and i can't give guarantee to pass the bracketing standard. This API is soluble in THF and MDC. My diluent is 0.1%OPA and Acetonitrile(50:50). It is not soluble in methanol, ACN and water (mixtures of these solvents) as such. Please suggest me, how to solve this problem?

check your needle-wash procedure. If you're not currently using one, try turning it on. If you are using a needle wash, make sure that the solution you are using is appropriate for the sample. It sounds as though the analyte has some definite solubility issues, and if it's not freely soluble in the needle wash, then the needle wash will not be effective.

Thanks for responding.
yes, I used THF as a needle wash, in which it is freely soluble. But, still i got the carry over of main API peak. Please help me to find out the solution.

It might be helpful for us if you would provide us more detailed information about your instrument, software and application.

When I observe a carry-over problem, I usually solve the problem with a “blank”-injection (double injection volume) of acetone from a completely filled vial. For short methods it is possible that you will see an acetone-peak in the following chromatogram. In this case add a real blank-injection or program the autosampler to perform both injections.

Hello

1. If you're using needle wash options (Agilent?) make sure that vial for it has no septa (just cap)
2. Are you using "delay volume reduction" - are you changing sampler valve position after injection (mainpas to bypass)?

Regards

Tomasz Kubowicz

In both the HPLC systems (Waters and Agilent) i tried.
In agilent without septa also i tried.
In "Waters" HPLC all the modes (normal, extended and double) of needle wash also tried.
Different needle washes, like diluent, THF also tried.
My buffer is phosphate(5mM)+0.1% TEA, pH adjusted 6.8 with OPA. In this buffer sample is not soluble.
Run time is long 50 min.

Hi.

You can use Universal solvent for needle wash:
25 ACN-25 MeOH-25 2-propanol-25 Water- 0.1%Formic Acid
In most of our carry over and contamination issues it helps. I don't know if this help you after 2.5 years.

Hì,
If no way to be found, you should try to take off the needle and bring it to sonic bath for cleaning after every injection. Of course if the needle can be easily taken off