may any one send me the procedures beacause the sound is very fast and i cant understand wat he say...please edit the video with writing he did with each procedure....thanx alot
When you pack the column, use small portions of silica. This is to avoid seggregation during packing where the bigger particles will settle at the bottom first and the finer particles at the top. A non uniform column results in a more efficient zone at the top and less efficient at the bottom which is not good chromatography.
If I want to purify a compound and run a column chromatography I choose an eluent that drives out the impurities fast but lets my main compound pretty much just sit on top of the column. If other eluents/mixes won't move my compound will putting it onto a nitrogen canister do any harm to it? The basic idea is to let the pressure just push it out mechanically from the column. Is this method completely stupid, or does it worth a try?
What you're talking about sounds a bit like flash chromatography, where pressure is applied at the top of the column. I suggest that you do a bit of research on flash chromatography before trying this. Always beware when applying pressure or vacuum to glassware as there are risks - make sure you stay safe!
What is the difference between dry packing and wet packing? Are they simply two different techniques used to pack the column or will it affect the separation of the compounds in the mixture.
This is more of a like undergrad o-chem lab video. I advise NOT making a slurry before adding the silica gel. I add silica gel dry, then add hexane to wet it. Also I REFUSE to run a column without a stream of flowing air to push the eluent through. Takes wayyy to long. I can fill up 50 test tubes with an air stream attached to the top in about 20 minutes or less. Yea you do have to work about the column bumping up but if you packed it correctly it is so easy.
Is it possible to take one of the test tubes containing a fraction and obtain measurements of absorbance from a spectrophotomer? Would the eluent not be a suitable optical medium for this procedure?
It can vary depending on the compounds you are working with. We most typically use an acetone:methanol mix in a 2:1 or 4:1 ratio. Different ratios will give you different polarities and most often you perform a column to separate impurities from the compound that you want. Sometimes you have to run a second column if you didn't get enough separation and change the eleunt and make it more or less polar.
It separates impurities from the compound that you want. You check the fractions with TLC to see if you have multiple spots or the correct spot for your compound. The vials that don't have impurities you collect and can evaporate to collect your solid pure product.
Increasing the volume won't increase separation. Most often you run a column and they take between 2-3 hours sometimes longer and you get 100's of test tubes filled. You add more eluent (solvent) as necessary to finish the column. It's the polarity of the eluent that determines if the compounds will separate.
nice video, really helped me to undestand what the books say, im reading the Pavia´s. if u keep submiting vids like this1 im suscribing oh and THX a lot :D
the sand is suppose to be added before the sample is loaded from what i've heard, that way craters aren't formed in the silica. he loaded prior to the sand which may not hurt considering a pippette was used but i'd think that not only elution, but sample loading would disturb the upper silica
It is up to you to decide how much volume you collect for each fraction. If you expect the compounds you are separating to be close together when they elute from the column, you might choose to collect lots a small fractions to prevent contamination. This, of course, means that you have to run lots of TLC plates to identify which fractions contain which compound. I hope that helps a bit!
simple & understandable, good work.
harunoblossom99 3 months ago
may any one send me the procedures beacause the sound is very fast and i cant understand wat he say...please edit the video with writing he did with each procedure....thanx alot
saher018 7 months ago
thank you
ajaykumaralikonda 8 months ago
Thanks! You helped me understand my lab a lot better!
chrissir 1 year ago
What is the eluent used?
paopaomanalansan 1 year ago
When you pack the column, use small portions of silica. This is to avoid seggregation during packing where the bigger particles will settle at the bottom first and the finer particles at the top. A non uniform column results in a more efficient zone at the top and less efficient at the bottom which is not good chromatography.
oomblikkies 1 year ago
thnx alot for sharing it
elsawalhy 1 year ago
If I want to purify a compound and run a column chromatography I choose an eluent that drives out the impurities fast but lets my main compound pretty much just sit on top of the column. If other eluents/mixes won't move my compound will putting it onto a nitrogen canister do any harm to it? The basic idea is to let the pressure just push it out mechanically from the column. Is this method completely stupid, or does it worth a try?
kicsiqki 1 year ago
What you're talking about sounds a bit like flash chromatography, where pressure is applied at the top of the column. I suggest that you do a bit of research on flash chromatography before trying this. Always beware when applying pressure or vacuum to glassware as there are risks - make sure you stay safe!
lowlevelpanic999 1 year ago
@lowlevelpanic999
Will do, thanks!
kicsiqki 1 year ago
the pressure doesnt affect the speed of chromatography process and its quite dangerous
wachuca 1 year ago
What is the difference between dry packing and wet packing? Are they simply two different techniques used to pack the column or will it affect the separation of the compounds in the mixture.
dpmlybsk 2 years ago
i actually find it better to make the slurry to add the absorbant. or else all the fine powder gets stuck to the walls of the column!
acconliffe 2 years ago
This is more of a like undergrad o-chem lab video. I advise NOT making a slurry before adding the silica gel. I add silica gel dry, then add hexane to wet it. Also I REFUSE to run a column without a stream of flowing air to push the eluent through. Takes wayyy to long. I can fill up 50 test tubes with an air stream attached to the top in about 20 minutes or less. Yea you do have to work about the column bumping up but if you packed it correctly it is so easy.
PlatinumPisces 2 years ago
what was the mobile phase and stationary phase being used??
tripq2aztig 2 years ago
Is it possible to take one of the test tubes containing a fraction and obtain measurements of absorbance from a spectrophotomer? Would the eluent not be a suitable optical medium for this procedure?
boomchook 2 years ago
What was the eluent? Does anyone know?
illuminati789 2 years ago
It can vary depending on the compounds you are working with. We most typically use an acetone:methanol mix in a 2:1 or 4:1 ratio. Different ratios will give you different polarities and most often you perform a column to separate impurities from the compound that you want. Sometimes you have to run a second column if you didn't get enough separation and change the eleunt and make it more or less polar.
bigguns175 2 years ago
What exactly does this do...? It looks like all you did was dissolve the crude compound in alot of eluent and took your time doing it.
What exactly did it purify?
HelloBrainMind 2 years ago
It separates impurities from the compound that you want. You check the fractions with TLC to see if you have multiple spots or the correct spot for your compound. The vials that don't have impurities you collect and can evaporate to collect your solid pure product.
bigguns175 2 years ago
it would be useful to put the text of the narrator... I could understand most of the explanation, but i missed some words :)
rodolforibas 2 years ago
If you increase the volume of the mixture loaded will that help with separation of the compounds? What if concentration is increased?
oekakinin 2 years ago
Increasing the volume won't increase separation. Most often you run a column and they take between 2-3 hours sometimes longer and you get 100's of test tubes filled. You add more eluent (solvent) as necessary to finish the column. It's the polarity of the eluent that determines if the compounds will separate.
bigguns175 2 years ago
@bigguns175 " It's the polarity of the eluent that determines if the compounds will separate" - also called the selectivity
oomblikkies 1 year ago
nice video, really helped me to undestand what the books say, im reading the Pavia´s. if u keep submiting vids like this1 im suscribing oh and THX a lot :D
agrippaviar 3 years ago
Does it really matter if the sand is applied on top of the crude product, or can it be applied before the crude product is placed?
caphamer 3 years ago
Yes - the sand is to protect the silica at the top of the column as it may easily be disturbed when you add more solvent.
lowlevelpanic999 3 years ago
@lowlevelpanic999
the sand is suppose to be added before the sample is loaded from what i've heard, that way craters aren't formed in the silica. he loaded prior to the sand which may not hurt considering a pippette was used but i'd think that not only elution, but sample loading would disturb the upper silica
34doodoo 1 year ago
@caphamer Wet silica, i.e. deactivated silica, will also do the job and you can afterwards recover the silica without sand contamination.
oomblikkies 1 year ago
How do you determine when one fraction ends and the next begins? Is it visual like using a separating funnel?
drbroccoli1 3 years ago
It is up to you to decide how much volume you collect for each fraction. If you expect the compounds you are separating to be close together when they elute from the column, you might choose to collect lots a small fractions to prevent contamination. This, of course, means that you have to run lots of TLC plates to identify which fractions contain which compound. I hope that helps a bit!
lowlevelpanic999 3 years ago
It does. Thanks!
drbroccoli1 3 years ago