Chlorobutanol has some strong antimicrobial properties, and it can kill both bacteria as well as fungus.
It's most commonly found as a preservative in eyedrops, or as the active ingredient in some mouthwashes.
However over the years, its use has become more and more limited, because it's not super stable at room temperature and
degrades over long-term storage.
If chlorobutanol is ingested,
it has a strong sedative effect,
but it's also quite toxic to the liver and can cause some other nasty side effects.
In the past it was used as a drug, but it's been completely replaced by safer alternatives
There is a related chemical called chloral hydrate,
which is also a sedative, but it's not nearly as hard on the liver,
and it doesn't cause as many side effects. However, it still is somewhat toxic and there are other drugs that do a better job
So it's not really prescribed very often. As I just mentioned earlier,
it's effective against both bacteria as well as fungus; it interferes with and
destabilizes cell membranes which eventually leads to the rupturing of the cell and the death of the microbe.
Unfortunately though,
it's also damaging to animal cells which includes human cells. However, when it's used externally and in low concentrations,
people seem to tolerate it pretty well.
Anyway, even though chlorobutanol isn't used that much anymore,
I still thought it could be interesting to make and to test out some of its properties.
What I think is interesting is that it's pretty easy to synthesize and all I need is some acetone,
chloroform, and potassium or sodium
hydroxide. The chloroform is the hardest ingredient to get and I actually made it myself in a previous video.
It's one of my very early videos and if you're interested in checking it out, there's a link in the description.
I start things off by adding 80 mL of acetone to a suitable flask.
The acetone needs to be cooled to between 0-5 °C and to do this I set up an ice bath.
Once it cooled, I went ahead and added 10 mL of chloroform.
When the chloroform is added, it warms up to about 5 °C. So again I wait for it to cool down.
A few minutes later, it's back to around 0 °C, and now it's time to add two grams of crushed potassium hydroxide.
It's important to add it in small amounts, because with each addition the temperature is going to rise.
In some of the additions, the temperature spiked to around 8 °C,
but I never let it go higher than that.
As more potassium hydroxide is added, the solution slowly gets more and more opaque until it's eventually completely white.
So what's going on here is a base-catalyzed reaction between
acetone and chloroform. The acetone is used in a large excess,
and it dissolves the chlorobutanol as it forms. The solid white stuff that we see floating around is just finely powdered
potassium hydroxide.
I couldn't find a proper resource for the mechanism,
but this is probably what's going on. In the first step, the hydroxide ion from the potassium hydroxide
reacts with the slightly acidic hydrogen of chloroform. This leads to the production of a negatively charged intermediate and
water as a side product. This intermediate then attacks a molecule of acetone, and the carbonyl opens up.
The negative charge on the oxygen
picks up a hydrogen from the water that formed earlier, which regenerates the base catalyst.
When I'm done adding the potassium hydroxide, I leave it in the ice bath, and I continue to stir it for about an hour.
When it's done, I remove the thermometer and it's time to filter things off.
Remember that our product is dissolved into the solution, and all of this white stuff is actually just potassium hydroxide.
It can also be filtered off using a paper filter, but in either case the powder is really fine,
and some is probably going to make it through. The majority of it is trapped by the filter,
but I don't like how cloudy it looks.
To clean it up, I quickly filtered it through some celite.
Celite is basically just a silicon dioxide-based powder that works really well as a filter.
Anyway, it was able to get rid of all of the small particles, and I was left with a nice and crystal clear solution.
After everything passes through, I washed the flask with a little bit of acetone, and I also clean up the walls of the funnel.
Now we need to isolate the chlorobutanol, and to do this
I'm going to boil off all of the acetone and any unreacted chloroform.
To make sure that all of the solvent is gone,
I'm going to heat it right up to the melting point of chlorobutanol which is around 95 °C.
This is well above the boiling points of acetone and chloroform which both boil around 60 °C.
When it got to a volume of about 80 mL, I transferred it to a smaller beaker and then continued to boil things off.
I reached the 90 °C mark when the volume was around 20 mL, and I took it off the hot plate.
While still hot, everything was dumped into 75 mL of ice-cold water.
Very quickly we see something crystallize out and this is the chlorobutanol.
I mix things as thoroughly as possible, and you can see that we have quite a bit of white solid now.
I then use a vacuum filtration to separate the chlorobutanol from the water.
The beaker was washed with water three or four times to make sure that I transferred everything.
To try to dry things up, I left the vacuum on for several minutes, but in the end
it was still a little bit damp. When the vacuum was turned off, the strong minty and camphor like odor of chlorobutanol
became pretty evident.
At this point, I think the chlorobutanol is relatively pure, but I decided to recrystallize it anyway.
It's often recommended to use a mixture of ethanol and water to do this, but the proportions are never given,
so I'm going to have to figure it out myself.
This really isn't too hard though,
and it's just a matter of playing back and forth with the ethanol and the water.
So the first thing that I need to do is dissolve all of the chlorobutanol in a minimal amount of boiling or near boiling
ethanol. When everything is dissolved, I start adding water which will reduce the solubility.
The basic idea is to find a balance where the chlorobutanol is just barely
soluble in the hot mixture. This way, when the solution cools down, the
solubility of the chlorobutanol will decrease, and it should crystallize out.
I ended up adding a little bit too much water, so I had to add a bit more ethanol to redissolve everything.
Eventually I got to what I felt was a decent balance between the ethanol and the water, so I took it off the hot plate,
the stir bar was removed, and I let it cool to room temperature.
Unfortunately, it didn't really crystallize. A major problem with chlorobutanol
is that it often hates to be crystallized, and it instead oils out.
I tried placing it in the freezer,
but even then it still stayed as an oil layer at the bottom.
In my mind, this was kind of a disaster
and I figured the only way to salvage things would be to try to recrystallize it again
and hope that this time it would crystallize instead.
I didn't get footage of this because I was honestly surprised,
but when I dropped in a stir bar and started stirring it, everything immediately
crystallized in the beaker. Here the chlorobutanol was on the bottom,
but the layer above it was an ice-cold mixture of ethanol and water.
So once the stirring was turned on, in my mind
this was exactly like we did before in the synthesis when the chlorobutanol was dumped directly into ice-cold water.
It was the fast change in temperature that caused crystals to form instead of the oil.
With this idea in mind, I decided I would try recrystallizing it again, but I would do things a little bit differently.
I turn on the hot plate, and I heat it until everything dissolves, and it's just on the verge of boiling,
then I quickly take it off and place it in an ice bath.
It's kind of hard to see here, but the chlorobutanol is actually crystallizing instead of oiling out.
After a couple minutes, I replenish the ice bath with fresh ice and I let it chill for another 15 or 20 minutes.
When it was done, I had some pretty nice crystals,
definitely a lot better than the oil I had last time.
Anyway, just like before I separate them off using vacuum filtration.
The crystals are washed with a very small amount of cold water, and
then I leave the pump on for about 30 minutes to try to dry it up.
Even after 30 minutes on the vacuum filter, the crystals were still a little bit damp. To fully dry it up,
I put it in a vacuum desiccator for a couple days.
It should be pointed out though that the vapor pressure of chlorobutanol is quite high.
By keeping it under vacuum for a couple days like I did, there's going to be a pretty big loss.
When I took it out, it was a lot drier,
but it still kind of looked damp. This isn't because of solvents or anything though,
and it's just because chlorobutanol tends to be oily even in its crystal form.
In the end, my final yield of chlorine all was 8 grams, which corresponds to a percent yield of about 36%.
My yield here is really low, and I think it's because I lost a lot in the vacuum desiccator.
I was concerned about the vapor pressure of the chlorobutanol,
so I weighed it before putting it in, and it had a mass of 14.2 g
So between putting it in and taking it out,
I lost 6.2 g,
and I highly doubt this was all solvent.
The amount of solvent left over after the vacuum filtration was probably no more than a gram or two.
Anyway, now I'd like to move on and test the antibacterial and antifungal properties of the chlorobutanol.
To do this, I just have to set up some simple agar petri dishes.
Into a beaker, I dumped in 2.3 g
of the agar, followed by about 100 mL of water. At room temperature
it's not really going to dissolve, and it needs to be heated either using a hot plate or a microwave. In either case,
it's heated to its boiling point, and then it's boiled continuously for another minute. All of the agar dissolves,
and now we need to wait for it to cool to around 50 or 60 °C.
We don't need a thermometer or anything for this, and we just need to wait until it stops hurting to touch it.
When I reach this point, it's ready to be added to the petri dishes.
I fill each dish about halfway,
and then I quickly cover the top. I do this because I don't want any potential
contaminants falling in and I also don't want the water evaporating off. As
the plates cool, the agar will slowly solidify.
When it reaches room temperature, I'm ready to set up my test.
Little microbes are everywhere so to infect these plates, I really just have to touch them.
I infected both plates with each hand to try to get an even distribution.
This definitely isn't the best setup, but it should get the job done.
Just to be completely sure that both plates get infected, I also added two drops of milk to each one.
There's a bunch of bacteria in milk, and I'm just adding it here to really make sure that I get things growing.
With the bacterial samples loaded onto the plates, I cover the left one and I mark it as the control.
To the plate on the right,
I sprinkle in a whole bunch of chlorobutanol. The amount that I add is completely overkill,
but the chlorobutanol tends to sublimate away, so I decided to use an excess.
It was covered with a lid labeled "test" and both of the dishes were moved somewhere warm and dark.
It's important to keep it out of sunlight.
It took about a week to get to this point, and two weeks to get here,
which is what I have now. The dish on the left,
which is our control, clearly has a whole bunch of stuff growing on it.
But our test dish which had the chlorobutanol is pretty clean.
This wasn't exactly the best designed and cleanest test but from these results,
I think it's reasonable to conclude that the chlorobutanol does have antimicrobial
properties. One important thing to point out and you may have noticed is that there's no chlorobutanol left on the test plate.
I mentioned before that it has a high vapor pressure, and over the course of about a week it kind of all disappeared.
I think before it all disappeared though
it was able to completely sterilize the plate which is why nothing was able to grow even over the course of two weeks.
Anyway, that's all I have to say about chlorobutanol. I hope you guys enjoyed this video, and I'll see you on the next one.
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