Problem with retention and separation

[mk_3249]

Hello
I've a drug molecule named Dimenhydrinate. We are following as per BP / EP. There is one test called Related substance (By HPLC) in the monograph of this product.
I am analysing as per the BP monogram using prescribed conditions but still unable to get the said retention time. Moreover, there are two product peaks at different retention time but both are seperating very nearby eachother.

The conditions are as under :
Column size : L = 0.25 m , d = 4.6 mm.
Stationary phase : end-capped octadecylsilyl silica gel for chromatography
(5 µm). I am using Merck C18 ODS column with the same parameters.
Temperature : 30 C.
Mobile phase :
Mobile phase A : Dissolve 10.0 g. Triethylamine in 950 ml of water, adjust the pH to 2.05 with Phosphoric acid and dilute to 1000 ml with water.
Mobile phase B : Acetonitrile
Solvent mixture: Mixture of Acetonitrile & Water (18:82 v/v)
Test solution : Dissolve 0.100 g. of the sample in the solvent mixture and dilute to 100.0 ml with the solvent mixture.
Gradient Program is :


Detection : Spectrophotometer at 225 nm.
Injection : 10 µl.

Retention time of two product peak are:
8-chlorotheophylline is about 8.0 min. and for Diphenhydramine, it is about 13.0 min.
But I am getting 8-CT on 2.5 min. and Diphenhydramine on 4.0 min.
I am unable to get the problem. Column is new and I am also getting good results with the other products using this column.

Thank you

[HPLCaddict]

Not all C18 columns are equal. There can be HUGE selectivity differences between different columns although, formally, they're both just C18 columns. Therefore the fact that other products look good using this column doesn't mean that this particular analysis of dimenhydrinate must look good too. We all would be very happy if we could use just one C18 column for every analysis .

You don't have enough retention. I'd suggest you double-check that the mobile-phases are prepared correct and that the method parameters (flow-rate, temperature, etc.) are correct, too. Then prepare a fresh batch of mobile phase and try again. If you still have low retention, the column is simply not suited.
I had a look at EP which seems to contain the same procedure for dimenhydrinate related substances. Suggested column for this analysis is a Luna C18 from Phenomenex (see https://extranet.edqm.eu/4DLink1/4DCGI/ ... w/mono/601). Get this column and try again.

[mk_3249]

Not all C18 columns are equal. There can be HUGE selectivity differences between different columns although, formally, they're both just C18 columns. Therefore the fact that other products look good using this column doesn't mean that this particular analysis of dimenhydrinate must look good too. We all would be very happy if we could use just one C18 column for every analysis .

You don't have enough retention. I'd suggest you double-check that the mobile-phases are prepared correct and that the method parameters (flow-rate, temperature, etc.) are correct, too. Then prepare a fresh batch of mobile phase and try again. If you still have low retention, the column is simply not suited.
I had a look at EP which seems to contain the same procedure for dimenhydrinate related substances. Suggested column for this analysis is a Luna C18 from Phenomenex (see https://extranet.edqm.eu/4DLink1/4DCGI/ ... w/mono/601). Get this column and try again.

Thanks for your suggestion.
I always prepares fresh mobile phase and have checked 2 to 3 times by preparing fresh mobile phase. Even also lookafter about temperature, pH, flow rate. but still not getting good results.

[prepcolumn]

As HPLCaddict stated try Phenomenex Luna which is offered by EDQM for dimenhydrinate..