Chromatography Forum

recently I am working an assay method for dextran sulfate, target LOQ ug/mL. So far, tried SEC, ion exchange, not very rewarding.

Are there any methods available without derivitization?

Thanks

Nick

Use a Refrective Index (RI) detector.

I have a method to analyse DEAE Dextran by SEC and RI detection. Works great for me since all the other components in the sample have very low RI 'adsorbance'. Just remember that all Dextrans have a wide size range and therefore you will get a wide peak in SEC.

I tried RI, signal was even worse than low UV. I used 20 mM phosphate in SEC. can you share with your RI parameters and mobile phase?

Thanks

nick

Use a Refrective Index (RI) detector.

I have a method to analyse DEAE Dextran by SEC and RI detection. Works great for me since all the other components in the sample have very low RI 'adsorbance'. Just remember that all Dextrans have a wide size range and therefore you will get a wide peak in SEC.

I use 50 mM Phosphate and 0.5 M NaCl pH 6.0 for my analysis. The column is a BioSep SEC-S3000 column from Phenomenex. The RI detecter is a Waters 2414 and the parameters are: Positive, Sensitivity 128, sampling rate 1, filter time 1 and internal temp 30 C.
It probably works for well for me since I have quite high amounts of DEAE-Dextran in my samples. I never load less than 100 ug but that gives quite a high peak (~500 mV). The LOQ was never established for this method but I would imagine it to be less than 10 ug.
How low levels of Dextran Sulphate are you looking at?

I am targeting ppm level ug/mL.

Thanks for your help.

Nick

I use 50 mM Phosphate and 0.5 M NaCl pH 6.0 for my analysis. The column is a BioSep SEC-S3000 column from Phenomenex. The RI detecter is a Waters 2414 and the parameters are: Positive, Sensitivity 128, sampling rate 1, filter time 1 and internal temp 30 C.
It probably works for well for me since I have quite high amounts of DEAE-Dextran in my samples. I never load less than 100 ug but that gives quite a high peak (~500 mV). The LOQ was never established for this method but I would imagine it to be less than 10 ug.
How low levels of Dextran Sulphate are you looking at?

I am targeting ppm level ug/mL.

The relevant question for detectability is not so much concentration as mass-on-column. 1 ppm is 1 nanogram/microliter. Assuming an injection volume of 100 microliters, that means you will have to detect 100 ng. That should be (barely) possible with a good RI detector (and very good temperature and flow control!) or with one of the evaporative detectors.

agree. I gave up RI and ELSD for now. CAD is promising now without complex matrix.

I am targeting ppm level ug/mL.

The relevant question for detectability is not so much concentration as mass-on-column. 1 ppm is 1 nanogram/microliter. Assuming an injection volume of 100 microliters, that means you will have to detect 100 ng. That should be (barely) possible with a good RI detector (and very good temperature and flow control!) or with one of the evaporative detectors.