Chromatography Forum

Hello,

I am PhD student and working with CAD Corona from ESA company and I have a problem regarding Oxytetracycline detection,I tried many methods but still don't get peaks.

Also, I am using Agilent series 1100 ,coloum Zorbax eclipse XDB-C8 4.5 x 150 mm , the flow rate is 1 ml/min and the injection volum is 10 μl as recommended in the manual.

I would be grateful if you help me to solve my problem or guide me to someone who can help.

Thanks a lot in advance

Emi

At this point you have no idea if you have a sample, injector, column, software, or detector problem. A little basic trouble shooting is in order. Myself, to start, I would make up a unretained sample that I know I can see with the CAD, maybe sucrose or NaCl , inject at least 100 ng on column and see what you get. Where you go next depends on the results of that experiment. Try it and let us know what happens.

Why are you using a CAD detector for oxytet? It absorbs very strongly at 355 nm and some lower wavelengths.

I use the following conditions:
Column: 4.6 X 75 mm Prontosil C18 EPS (3 μm particles) operated at 35 °C.
Mobile phase: 20% methanol in 80% 0.1% phosphoric acid
Injection volume: 5 μL
Flow rate: 1.5 mL/min.
Detector: 355 nm
Retention time: 3 minutes

This compound is unstable unless stored in acidic solution.

Thanks a lot for your help I will try your method for sure and I will update you with results.

The reason that I use ACD that I have it in my lab and my supervisor asked me to make it work as he said it is very sensitive detector.


Emi

Thank you very much ,I tried what you said before and I didn't find problem regarding those things.

I got new column and I will try it with Dr Merlin's method

Thanks again

Emi

Hello Emi;

The Corona CAD can easily see the Oxytetracycline the mw is 460.
To confirm this remove the column and inject the standard.
You should check the background of your mobile phase firt which should be below 2pA. the detection limit should be low ng on column.
Hope this helps

Dave Clark

Hello,

I am PhD student and working with CAD Corona from ESA company and I have a problem regarding Oxytetracycline detection,I tried many methods but still don't get peaks.

Also, I am using Agilent series 1100 ,coloum Zorbax eclipse XDB-C8 4.5 x 150 mm , the flow rate is 1 ml/min and the injection volum is 10 μl as recommended in the manual.

I would be grateful if you help me to solve my problem or guide me to someone who can help.

Thanks a lot in advance

Emi

Since tetracyclines absorb at 355nm (as Merlin pointed out) - UV is
is a really nice method. Problems associated with matrix interference
(specificity) should be greatly reduced at that wavelength.

Once one decides to move on to ELSD, CAD, or MS detection, things
become tricky. Volatile additives must now be used - and only low-bleed
phases should be used with these detectors. In addition - tetracyclines
are known to have chelating characteristics - so a highly intert stationary
phase (low heavy metal content) is required.

Below is an LC-MS application for tetracylines on Cadenza CW-C18 (30nm pore size):
http://www.imtakt.com/TecInfo/TI411E.pdf