Thiamine Mononitrate vs HCl HIGHIER Recovery Issue

Discussions about HPLC, CE, TLC, SFC, and other "liquid phase" separation techniques.

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TLChromGuest wrote:
Hello Everyone,

I have been working on a HPLC method for Water Soluble vitamins and achieved the separation for all of them. All the HPLC parameters looks good, including; linearity, recovery, accuracy and precision for all of them except for Thiamine where I use Thiamine HCl (USP) Reference Standard to analyze the % of Thiamine Mononitrate in sample solution. I using the molecular weight conversion formula as specified in USP monograph for Thiamine, but still the % of thiamine in sample always hewer around 120% of Thiamine HCl.

I know, I am missing something here. Did anyone had similar issue? Can anyone help me out here as what could be wrong.

Thank You!
Hi TLChromGuest,

To preface, I am not familiar with these methods or analytes and am just giving a simple guess to what might be the issue you're facing. First off, some advice: if you want more people to respond to your posts then you need to be as specific as possible when posting. It is very difficult to understand an issue when it is described in (what boils down to) three sentences. Reposting your own question with no additional information, while it's still on the front page, will not get you anywhere.

Where I think there might be an issue is your conversions or your standard. It seems like there are a few different types of Thiamine being used which could change the mass/mole of your standards or injected samples (I'm not sure exactly what you're injecting here). I assume you're injected thiamine mononitrate but using a thiamine hcl reference standard which may change the concentration of the both of them depending on the difference in mass. Either way, on the HPLC you're going to be reading the Thiamine peak exclusively. Is there any chance that type of thiamine you're using is different than the one in the USP? Is there any chance that the standard you're using could be incorrect? Mayhaps try a new one?

I'm sorry I cannot be more of assistance and these solutions are mainly guesses. I hope that someone with more experience can elucidate the issues you're facing better than I can. Hopefully this gets the ball rolling.

Tyler Smith
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