Monday, February 20, 2017

Dry Column Vacuum Chromatography (DCVC) - The Movie!

I have on several occasions been asked to make a DCVC video tutorial and quite liked the idea of doing so. Thus, I have started my acting career as you can see in the video below. I think the video will be a useful guide for first time DCVCers. For more info you should consult this and this blog post on DCVC. Many thanks to the University of Copenhagen's Communication Department, in particular Jacob Lejbach Sørensen, for investing some time in making this possible. D!


21 comments:

Unknown said...

Its great, gonna watch it soon (After I set up reactions :D )
Is there any chance you will post more again on your blog? Lot of interesting stuff here but it seems that it slowly died :(

Daniel Sejer said...

@Mike Krigg: I doubt that I will ever get back to posting on the level I used to. I simply have too many things to do these days. However, I have been considering if I should start sharing my groups research on the blog if nothing else. Let's see if I get inspired. I have on many occassions encouraged students to write blog posts but they aren't really interested. However, it is still a nice resource blog that get's many visits (around 150 hits/day) so I'll keep it up and running and will continue to answer peoples questions. D!

emunfred said...

In my previous job I used DCVC a lot. I had the glassblower modify some glass filter funnels to be taller. I used them in several diameters, up to about 10 cm, and separated up to about 50 grams of crude per run (as I recall). I also used the system with butanol-acetic acid-water to separate some very polar compounds.

Unknown said...

Hello from Oregon USA, and thank you so much for taking the time to pass along this great video. It turns out that I had resorted to this idea on chance as I was purifying some plant extractions. I tried with my hobby grade chromatography tube to get seperation but needed a way to do this much more efficiently and scalable to my process with normally requires 28 grams or so at a time to be purified.

In Oregon it is perfectly legal to grow marijuanna and also to obtain the extractions of marijuanna done by supercritical fluid extraction. The problem I have as a medical user is that at best the dispensary grade extracts test at 70-80% THC nominal. This leaves significant contamination of the target medicine in he extract meant ultimately for inhalation via a vaporizing device. Respiratory irritation is a huge problem for most users needing relief of this medicine. I have learned to remove a good share of the irritants, mostly Volitile Organic Compounds (VOCs) which are known by users in Oregon as terpenes. This GREATLY enhances the potency and medical efficacy of the product. I noticed during chromatography runs in my standard chromatography tubes that there was significant material being held up on the alumina from the extract when isopropyl alcohol was used as the mobile phase.

So as a filter idea only I placed a few inches of alumina into my buchner funnel equipped with a fine frit. The process outshined ANY other filtration method for purifying that I had tried but moreover I saw the potential for seperation and not just filtration in the process. Then my internet search on the idea produced your excellent video! Nice!

I know in great detail the problems that processors have run into trying to produce a purified product here in Oregon. I can tell you without hesitation that the process you so thoroughly document will without question become the standard against which other pufification techniques are judged. In just minutes the filtration process using alumina and isoproply alcohol/water accomplished what can take many hours to do otherwise and does so MUCH better. I realize that my use of alumina deviates from your video but at the moment it works very well and is extremely inexpensive to,purchase from Amazon in an abrasive grade bulk container. The isopropyl alcohol is also inexpensive on Amazon and has a huge side benefit when combined wih water and used in purification but that topic is a different subject altogether.

Thank you so much for advancing the knowledge!

xmac1x said...

Great video and great to see you back Daniel. Your blog was great help to me during my PhD and I tried DCVC a number of times!

Daniel Sejer said...

There is an interesting discussion on DCVC going on at Reddit worth checking out: https://www.reddit.com/r/chemistry/comments/5v5qq1/dry_column_vacuum_chromatography/
D!

Nicolau said...

Hey Daniel, thanks a ton for your very detailed explanations and video. I am interested in trying DCVC, but I can't seem to find the correct kind of silica. In your published article, you use Merck 15111 silica gel 60 (15–40 um), but the product seems to have been discontinued since then. Can you provide a link to an alternative that would work equally well?

Daniel Sejer said...

@Nicolau: I don't have a quick answer to your question. When I Google it I find vendors that have this type of silica in my part of the world. As long as you get the particle size about right it's worth a shot. Some brands prefer to use mesh or microns to describe the silica so you should find a conversion table online and go hunting for the right type of silica. D!

Unknown said...

@Nicolau, I am not a chemist but enjoy the seperations. I routinely use the method shown here on this blog for refining plant extracts that are legal here in Oregon. I found suitable gel on Amazon although not as small as Daniel demonstrates in particule size. So long as I take care in packing the column well the seperations are excellent on an extract with such a great many compounds present. There is often 5 or more color bands that develop as the DCVC gradient progresses. The large molecules like green chlorophyls are among the last to elute. My only variation is because pure EA is not enough to get some of the lighter colors after the chlorophyls to elute. At that point I switch to a 10% methanol to 90% EA and solvent mix to get the really sticky stuff off the column. The last bit is a tiny fracion by weight and generally the target compound elutes in the third or fourth 20 ml fraction. I start with hexane.

Unknown said...
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Unknown said...

Hi all, I've used this technique on small scales 1-5g and I find it works well enough. However, when I've tried larger scales 50-100g I'm finding that there is very wide bands and co elution of spots that are quite well separated by TLC. I've loaded 50g of crude onto a 12cm sinter with a round 6cm of well packed silica. Gradient elution was carried out by increasing from 100% heptane to 80% ethylacetate heptane. However, I'm just not getting that razor sharp separation. are 200 mL fractions too large or too small? Is it possible that the compound just isn't clearing the column on time due to fraction size, or am I missing something else fundamental? Thanks!

Unknown said...
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Nicolau said...

Hello again, Daniel! It is my great pleasure to say that in the past half a year since I posted here, I've started a movement in my lab towards DCVC (Merck still sells 115111 silica after all). After a lot of work convincing labmates to try it out (and supervisors to invest in the idea), it's starting to take hold! You can count on multiple future citations and positive word-of-mouth.

I've had great success on separations with ~1 g crude, and now am starting to explore scales up to ~25 g. Do you have any useful tricks for making larger scale separations more convenient?

Also, occasionally I see compounds "bleeding" past the top of the silica and forming fangs, even during elution with neat pet spirits/cyclohexane, where the compounds have an Rf of zero (see for example these pictures: https://imgur.com/a/vruJM ). Do you know what causes this? Should I use an even higher celite-to-crude ratio? What is the rough celite/crude mass ratio you typically use? For reference, in the photos I had loaded 26 g of crude onto 50 g of celite from a DCM solution (DCM elutes the compounds quickly). The loaded celite was a little clumped when I added it to the column, though I put on the rotovap for 2 h and no solvent seemed to remain. My compounds are viscous resins when pure.

Thank you so much once again! This is an amazing technique which for me has turned columns from a dreadful slog into something I enjoy tinkering with.

Daniel Sejer said...

@Nicolau: That is awesome. It's good to hear that the people around had an open mind and where willing to give it a shot. It really is an excellent purification technique to add to the synthetic organist chemists purification tool box. I'm looking forward to the citations ;-)
Regarding large scale purifications these are just more of a hassle. The biggest "problem" is to pack your column well. You may need a more serious pump and definitely more patience. Other wise it just a question of bigger glassware and figuring out how to set it up in a smart way.
Regarding bleeding of compound on the column this can be due to:
1) Over-loading the column
2) Poor packing of the column edges
3) Residual polar organic solvent
Adding more Celite will not improve the situation. Its function is merely to facilitate loading and you simply use what you find convenient. In my experience the problem is the residual organic solvent. In my experience I have always had residual solvent in there after rotavapping even for prolonged time with heating. To get around this I have always put my compound+celite on a high vacuum pump for about 30 min prior to columning. Even a small amount of dichloromethan can mess things up. I hope that helps and thanks for the kind words. Best of luck with DCVC'ing. D!

Unknown said...

Hi, and thanks for the great video.

Are drawings available to guide a glass blower for making the DCVC adapter for the Buchner funnels suitable for smaller funnels (1-8 cm id) and larger ones (10-20 cm id). By adapter I mean the custom glassware that combines the functions of a sep funnel, vacuum take-off and Buchner adapter that fits the rubber bung.

Finally, there is a reference by Douglass Taber that is close to the DCVC technique that may be worth noting as a precursor to it. He uses finer Sigel (5-15 um) and pressure for small column but vacuum for larger columns. The citation is: Taber JOC 1982 47 1351 TLC Mesh chromatography.

Best regards,
Don Stewart

thrdel said...

Hello Daniel, I'm not sure if you still watch this blog but I figured it doesn't hurt to try so, I love the video, watched it quite a few times just to make sure I didn't miss any subtle details or anything (like a normal chemistry novice would do ) so , thanks Daniel , it's a great help. I do have a question in regards with running the column though. You say you always use heptane but would that work if the compound is only soluble in really polar solvents like water and maybe methanol ? Would it be alright to use methanol fractions ? I've seen reports of others using water saturated butanol . Water saturated heptane isn't an option, is it ?
What options are there other than ethyl acetate / heptane ? Should I try 10% MeOH in Heptane ?
All the best

Zary said...

Hello
first of all, thank you for the impressive video. I did the same for fractionating my 10 gr plant extract. Although I fractioned the extract with much less solvent and silica than Flash, I had some trouble in the process; the bands didn't come down in a straight line! they came down as a curve which made the separation much poorer than it should be. does this relate to how I poured the solvent on the silica bed? or even the vacuum strength of my vacuum pump? I also have two other questions. 1) Is this method also practical for separating two compounds with near RF? or just fractionating the crude extract is desirable in this method 2)what happens if we use an isocratic solvent system? Is it like prep TLC which can be run multiple times in the same solvent system? 3) For small scales, is the resolution improved compared to Flash? 4)can we apply this method on SPE cartridges using a vacuum manifold?

Greg said...

I am trying separations on C18 silica with an ethanol: water:acetic acid solvent in order to be green. Is there a rule of thumb to size the fraction and sample size vs the CV of the reverse phase system with known rf values?

Daniel Sejer said...

Hi Greg, the short answer is no. From a practical aspect the fraction size needs to increase with increasing column diameter. As you can see in the video ca. 25 ml works well for a column id of 3-5 cm. For id 5-10 cm you have to go up to about 50 ml and so on. It will be self evident when you try it out whether the fraction size is too small. How large you can go with the fraction size depends on many factors. For an easy separation you can use larger volumes but there is no reliable rule of thumb. It's trial and error. Be aware that C18 columns with water mixtures usually elute slowly so make sure you have a tight system and a good vacuum. D!

Solidspin said...

Greetings, Daniel -

This is truly an invaluable resource you have created. I am doing larger volumes, having been inspired by you... At one point in the https://www.reddit.com/r/chemistry/comments/5v5qq1/dry_column_vacuum_chromatography/, you mentioned that you have done upwards of 200g. I’m curious- do you distribute the crude over the entire surface of your silica, so as to promote even flow down the column?

Regards,

SAB, PhD

Daniel Sejer said...

Hi SAB, great that you are finding it useful. Yes I always distribute the crude sample evenly on the surface and almost always after absorbing it on Celite. D!