Chromatography Forum

Hi All,

In a spot of bother. Not had much experience extracting from oils so any help is appreciated.

I have a sample of Vit D3/Cholecalciferol in oil and would like to extract to aqueous as i have a good RP method waiting. I am looking for the easiest process for extracting the Cholecalciferol from the oil and into an aqueous phase. Anyone who has any experience with oils who would like to help, its appreciated.

HPLC of Vit. D3/ Cholecalciferol extracted from various matrices is a regular topic on this forum.

Enter "Cholecalciferol" into "SEARCH" and you will see the threads.

JMB

The reason it's in oil – in the first place - is its insolubility in water.
So how on earth would anyone expect it to be extracted in water?

Best Regards

I'm biased, but use your RP method to separate the vit from your matrix using a prep chromatography system.

Being insoluble in water, you will need to use an intermediate organic solvent to achieve desired solubility. Perhaps IPA.

I am working on Vitamin D analysis now, attempting to extract it from Dried Plum Powder. Vitamin D is soluble in Acetonitrile, Ethyl Acetate and Methanol, so if any of those would work to extract the Vitamin D from the oil, it could then be diluted into Methanol for injection into the RP analysis.

I was having a terrible time with my extractions. My first simple test extract work great, so to bring up the concentration I decided to try 4g/20ml in each solvent instead of 2g/20ml and I seemed to get worse results. Thinking maybe it was the plastic centrifuge tubes, I tried glass. With glass I got almost no recovery, even on blank spikes. After that I thought maybe it was UV light causing my problems so I tried using 40ml Amber vials, first run gave lower than expected results, second gave almost no recovery at all. Difference was second run was extracted for a longer time. It would appear from this that Vitamin D can adhere to glass.

In frustration, and after reading another paper, I decided to try the Quad Solvent we use for broccoli seed extractions which is equal parts DI water, Acetonitrile, DMF an DMSO. A quickie extraction in plastic centrifuge tubes gave similar results to my very first quickie extraction, but matrix spike was about 50% recovery. Today I am running the test again with a blank, blank spike, sample, sample duplicate and spiked sample to see it the result was a fluke or something genuine. If so this may be some interesting research in that I am finding Vitamin D2 and D3 in fruits and finding that glass can interfere with recovery of Vitamin D.

I wonder if on the oil, you could extract with a mixture of water and acetonitrile, which should not dissolve the oil completely. Then dilute with more water and pass it through a C18 SPE, then elute with acetonitrile or methanol to make an extract for injection?

Vitamin D is extremely non polar. Using a polar solvent to extract will lead to issues.

Over the weekend I found that Plum Powder doesn't contain Vitamin D, but I did get good recovery from the spiked sample. What I found in the past appears to be some kind of interference, though I have yet to figure out what could interfere that would be a match for both the retention time and MRM masses. Either something has gotten contaminated with Vitamin D2 and D3 or there is something in the lab that can mimic it

Over the weekend I found that Plum Powder doesn't contain Vitamin D, but I did get good recovery from the spiked sample. What I found in the past appears to be some kind of interference, though I have yet to figure out what could interfere that would be a match for both the retention time and MRM masses. Either something has gotten contaminated with Vitamin D2 and D3 or there is something in the lab that can mimic it

My money would be on contamination. They put that stuff in hand creams and body lotions, etc. I've run into this issue several times over the years.