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Topic: Help with Solvents for Fat Soluble Vitamins  (Read 4352 times)

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Offline sleepdeprived

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Help with Solvents for Fat Soluble Vitamins
« on: November 16, 2013, 09:16:25 AM »
I'm developing an HPLC assay for quantifying vitamins. I'm having a difficult time getting the fat soluble vitamins into solution. My job purchases the raw materials as spray dried powders, and I have a feeling that's part of the problem. I have tried 75/25 ACN/MEOH, Chloroform, Methylene Chloride, and absolute ethanol, 50/50 MEOH/Chloroform. I sonicate each solution for 10 minutes. The powders are just flocculated throughout the solvent. That, and the lack on peaks on my chromatograms, tells me nothing is going into solution. I have read to try ether, but instinct is telling me that if I'm having trouble with a lll of the solvents I've already tried, ether will be no different.

Any ideas or help would be greatly appreciated. 

Offline Archer

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #1 on: November 16, 2013, 09:23:04 AM »
Do the reference materials dissolve ok in these solvents?
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Offline Arkcon

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #2 on: November 16, 2013, 09:29:32 AM »
I hate to seem to be jumping ahead, but ... what are the eluents you're going to use for the HPLC assay?  You have to be sure your analyte won't precipitate inside the injector or column, so maybe you can start with those reagents to dissolve your sample?  Its best to not switch solvents dramatically from sample to eluent, in any case.
« Last Edit: November 16, 2013, 03:23:23 PM by Arkcon »
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Offline Arkcon

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #3 on: November 16, 2013, 09:43:28 AM »
Here's a reference I found online (there are likely hundreds of others) http://www.sigmaaldrich.com/catalog/product/supelco/g005860?lang=en&region=US  It seems fat soluble vitamins are readily soluble in acetonitrile, and your 25% methanol is probably not disastrous.  What is the nature of this flocculat -- is this a binder or water soluble vitamin fraction?  Like Archer: asked, do the standards dissolve in any choice of solvent?  Or do you have to extract your sample in a strong solvent?  If so, can you tell, either by loss of weight of remaining sample, or by evaporating solvent, that you have extracted something?
Hey, I'm not judging.  I just like to shoot straight.  I'm a man of science.

Offline sleepdeprived

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #4 on: November 16, 2013, 12:06:20 PM »
I hate to seem to be jumping ahead, but ... what are the eluents you're going to use for the HPLC assay?  You have to be sure your analyte won't precipitate inside the injector or column, so maybe you can start with those reagents to dissolve your sample?  Its best to not switch solvents dramatically from sample to eluent any way.

The 75/25 ACN/MEOH is my eluent. It's an isocratic run of 75% ACN, 25% Methanol. The standards do dissolve nicely in it. But the standards are in their natural form (most fat soluble vitamins are liquids.) In the industry, however, they're bought and sold as a spray dried product. I've spray dried eggs and fat in the past, so I know what's involved, but I remember a 50/50 aqueous/organic solvent being enough to dissolve it. For these, I'm in the dark.

Thank you very much for the Sigma-Aldrich reference. I'll look at that sight more to see if they have anything that can help me.

Also, I received a suggestion to try vegetable oil, but I would think I'd get a messy chromatogram.
« Last Edit: November 16, 2013, 12:22:49 PM by sleepdeprived »

Offline Archer

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #5 on: November 16, 2013, 01:30:53 PM »
I too have some experience of spray drying but we used MD-20 to support the water soluble product on so no issues with solubility in an HPLC mobile phase.

Is there any information as to what these materials are spray dried with?
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Offline Arkcon

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Re: Help with Solvents for Fat Soluble Vitamins
« Reply #6 on: November 16, 2013, 03:29:35 PM »
Well, are you sure your unknown samples are the same substance as your reference materials?  For example, tocopheryl acetate is a water soluble ester of tocopheryl (Vitamin E), and the fat soluble vitamins in your sample may likewise be derivatives of some kind, if you're having trouble dissolving them in solvent.  Are you trying to dissolve by sonication for a brief time?  Because that can really help, if the spray-drying process has really tightly bound up your samples.
Hey, I'm not judging.  I just like to shoot straight.  I'm a man of science.

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