Chromatography Forum

Dear All,
Why does the detection wavelength changes for the same drug product for a particular analysis( say, dissolution for instatnce) when analysed via HPLC euipped with UV detector and secondly analysed on UV-Visible Spectrophotometer.
Thanx in advance.

Dear All,
Why does the detection wavelength changes for the same drug product for a particular analysis( say, dissolution for instatnce) when analysed via HPLC euipped with UV detector and secondly analysed on UV-Visible Spectrophotometer.
Thanx in advance.

Ideally it should not change if your diluent (blank) for the drug is kept same for both HPLC-UV and UV analysis.
lambda max varies if you are changing the diluent.
i.e. if you are taking acidic diluent you will get some lambda max but for the same drug lambda max will vary if you chose the basic diluent

Its not necessarily b/c lambda max is changing. The HPLC method may have been optimized by changing the wavelength. It may be that lambda max caused high baseline noise. Maybe the wavelength selected in the HPLC analysis was not the lambda max for the active but useful for evaluating other related compounds or excipients allowing additional evaluation in the analysis without sacrificing the detection sensitivity of the active.

the dude

Harsha,

The differences you are seeing seem to be related to two instruments (Uv detector, Uv-Vis Spectrophotometer). Therefore thevwavelenght accuracy is likely to be different in each. Obviously, large differences, such as 10-20 nm, are most likely due to solvent effects, temperature, etc.

I hope these comments help you.

josebenjamin

Method Optimisation( ie. to change the wavelength of detection) to get a good look at impurities in case of RS is understood. but in case of dissolution .......? I dont think for the same reason wavelength would be changed when one goes from HPLC-UV to UV-Visisble spectrophotometer.
Actually i hv seen one method, that for a drug product when dissolution was done via HPLC-UV detector the wavelength used to be set at 254 nm and the same analysis when done by UV-Visisble spectrophotometer the absorbance used to be measure at 298 nm with purified water as blank.
Why would this be....? Mind you i am talking abt Dissolution and not RS where one is more interseted in the parent peak irrespective of other peaks.

Actually i hv seen one method, that for a drug product when dissolution was done via HPLC-UV detector the wavelength used to be set at 254 nm and the same analysis when done by UV-Visisble spectrophotometer the absorbance used to be measure at 298 nm with purified water as blank.
Why would this be....? Mind you i am talking abt Dissolution and not RS where one is more interseted in the parent peak irrespective of other peaks.

Dissolution of formulated products has to avoid any other components, so if the analysis was on a UV Spectrophotometer without chromatography, the detection wavelength may be longer, but not at the optimum. Also, some dissolution buffers may absorb at lower wavelengths.

For HPLC, the interfering comounds are separated, so a lower wavelength can be used, perhaps even the compound peak.

254 nm was the standard fixed detector wavelength because a Hg lamp was used ( Zn gave 214 nm ), so many methods used that wavelength , regardless of where the peak was. Note that the selected spectral band width can also affect results.

Bruce Hamilton