Chromatography Forum

It seems you can have 50 different HPLC methods to analyze a particular analyte.

But, some just seem to work better than others.

Question, does anybody out there have a really nice stability indicating method for Benzalkonium Chloride?

Right now I am using 0.5% Acetic acid with MeCN on a C18 column. It really stinks. Peaks are tailing etc.

What's your matrix? Possibly you could titrate as a cationic with standard anionic to dimidium bromide-sisulphine blue endpoint. If the BAC degrades, it won't titrate as a cationic.

That won't work out for my application. This is something that will have to be validated and quantitated in a QC laboratory with little analyst involvement. HPLC is what I am looking for.

Thanks for your response.

It would be helpful to tell us what you have done to make your method suck less. has the method been used for a while, or is it new. Which expected degradation products does your current method monitor - assuming it's been validated.

Because Benzalkonium chloride is a mixture of homologues, there are several possible ways of approaching the analysis.

Some methods use cyano columns with a mobile phase of 55-60% acetonitrile and 40-45% 0.075-01 M acetate buffer pH 5 at 1.0 ml min.

There's several examples in the literature on Pubmed and elsewhere, you don't have to buy the articles, kind authors often put the gory details in the free abstracts.

Bruce Hamilton

Some methods use cyano columns with a mobile phase of 55-60% acetonitrile and 40-45% 0.075-01 M acetate buffer pH 5 at 1.0 ml min.

There's several examples in the literature on Pubmed and elsewhere, you don't have to buy the articles, kind authors often put the gory details in the free abstracts.

Bruce Hamilton

Yeah, I have noticed that.

Well, it seems that a pH of 5 gives me separation from the oxidation degs.

Anything below pH 4 and they co-elute.

But, the lower the pH the better the peak shape...makes sense becuase this is a quaternary ammonium compound.

This compound seems to have problems eluting for me. What I mean is its almost 100% organic to get them out??? I am a bit baffled about this but thats how it is.

Honestly, I started this new job and they threw this method in my lap. So I tried it out and I was surprised this thing actually passed validation????? I am thinking a smaller particle size may help with the peak shape?? Not sure.

Also, thinking a polar embedded column?? may speed up retention?

Any thoughts?

Go to Waters webside, look for the Library / browse eApplication Notebook / chemistry applications under B for benzalkonium...

The tailing is due to silanol activity on the silica column. Try a different type of column. The Acclaim Surfactant column has very good peak shapes for quaternary compounds at pH 5 with ammonium acetate buffer, but its selectivity is somewhat different than C18. I would be willing to run a sample for you.

http://www1.dionex.com/en-us/webdocs/25 ... et_V24.pdf

The tailing is due to silanol activity on the silica column. Try a different type of column. The Acclaim Surfactant column has very good peak shapes for quaternary compounds at pH 5 with ammonium acetate buffer, but its selectivity is somewhat different than C18. I would be willing to run a sample for you.

http://www1.dionex.com/en-us/webdocs/25 ... et_V24.pdf

What is the stationary phase on this column? It says Silica?

Thats why I was thinking polar embedded column for the peak shape?

The Surfactant column is based on a silica substrate. The surface bonding has a balanced mix of hydrophobic, polar and ionic functions that neutralize the interaction between quaternary amines and the silica substrate. The details are proprietary, but there is a technical article in Journal of Chromatography A, 1118 (2006) 29–34.

Dear Skipjack:
by mechanism, there is not much reason causing your peak tailing unless there were some overloading; the tailing actually was coeluting; the column happened to be too old; or you really need more proton to protonate the silanol.
If none of these, I suggest to try different brand C-18/8.
Good Luck from AL

Dear all:

I just found a reference for a method for BAC in PDA Journal of Pharmaceutical Science and Technology, titled "Simple HPLC Determination of Benzalkonium Chloride in Ophthalmic Formulations Containing Antazoline and Tetrahydrozoline." (Vol 59, No. 5, Sep. 2005). Author: Pornchai Rojsitthisak et al.

Comments on this method:
Pro: can have high recovery (almost 100%) and precision.
Con: not observed peak of C16 homolog (only C12 and C14). Also suffered from bad tailing (visual inspection of chromatogram in the publication)

We have created an in-house method that could give peaks of C12, C14, and C16- homologs. However, for one matrix our method did not give acceptable recovery (e.g. mean recovery is below 90% or above 110%). Considered proprietary, I can’t provide details here.

Any one had better luck with this assay? (I am not asking column mfg’s such as Dionex. We have got the Acclaim surfactant column already!).

Alfred.