Try this method. However with a C18 column (rather than a C1 column) you will see more peaks.

https://appslab.thermofisher.com/App/1882/single-peak-quantitation-tween-20-by-hplc-corona-charged-aerosol-detection

Thank you and what diluent can be recommended ?

They use water, but the answer depends on your sample matrix!

Thank you and would be the other columns if that column is not available?

Dear Srikanth Voruganti,

we have developed a method for the quantification of Tween20 and Tween80 in a vaccine solution in our laboratory some years ago. However, this method was based on LC-MS with MS^3 transition.

We tested the following columns:

Agilent Polaris Amid 150x3mm, 3µm

Agilent Carboxen 150x3mm, 5µm (no longer available)

Merck Chromolith C18e 100x3mm

However, you can also test other phases like YMC Triart C18

Our eluent was isocratic 90% methanol to suppress clustering. Under these conditions, Tween20 elutes on the Polaris Amide column after about 2.7 min and shows only few peaks as the structural variants (different EO chain length and fatty acid chain length) are pushed together.

The suppression of the clusters was essential for us, because we detected molecule mass-dependently and could only generate linear calibrations in this way.

We validated this method in the GMP environment.

Please feel free to contact me for further information.

Best regards

Joachim Horst

Thank you and can you please provide method parameters if possible for the method developed . Can I use C18 column with your method parameters?.