Workup Tricks

Reaction Roadmap: Workup

 Problem (Workup Nightmare #1): Upon combination of organic and aqueous solutions,

a gooey or insoluble precipitate appears, which floats between the two layers and obscures
the border.
 Solution: Keep washing with water until most of the goo is removed. Then use copious
drying agent, and with luck, the goo will be absorbed and you will be able to filter it away.

 Problem (Workup Nightmare #2): When an aqueous solvent is added to the diluted

reaction mixture, an emulsion forms. All efforts to resolve the layers fail, your solution has
swollen to gargantuan proportions, and you can't find or can't lift a separatory funnel large
enough to hold it
 Solutions:
o If an emulsion forms during workup of a reaction, next time try evaporating the
solvent first, then taking the residue up in the solvent for the extraction.
o Wait half an hour to see if separation will occur on its own. Then think twice before
introducing more aqueous solution.
o Add solid NaCl to the emulsion- very briny solutions sometimes separate better.
o Last resort- especially when the solution is already dilute- Try diluting the organic
layer significantly- 5X or 10X.
o Filter the whole thing through Celite

Problem (Workup Nightmare #3): During Acid/Base Workup, the expected precipitate

does not form.

 Solution: See Acid/Base Workup.

 Problem (Workup Nightmare #4): Upon addition of aqueous bicarbonate, the organic

layer becomes a graceful fountain, coating the inside of your fume hood.
 Solution: When you repeat the procedure, add aqueous bicarbonate carefully, shake
gently, and vent the separatory funnel often.

 Problem (Workup Nightmare #5): Addition of aqueous solution to your black organic

reaction mixture leads to a uniform black mixture. The solution in the addition funnel is
opaque, and you can't see the border between the organic and aqueous layers.
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 Solution: Try adding ice, which will float on the water, between the layers.

 Problem (Workup Nightmare #6): The aqueous solution you use to wash the organic
layer turns yellow, orange, brown or pink- the first ten times you try it.
 Solution: If the color is caused by excess halogen reagent from your reaction mixture, try
washing with sodium thiosulfate. The solution should become instantly clear- if it does not
and you still suspect halide, try stirring vigorously in an Erlenmeyer for 10-15 minutes to
complete the has-text-dangeruction of halogen molecules.

 Problem: (Workup Nightmare #7): After workup, you can't find your product.
 Explanations and Solutions:
o Your product may be soluble in the aqueous layer- check it (you still have it, right?)

o Your product may be volatile: check the solvent in the rotovap trap.
o Your product may have gotten stuck in filtration media- if you had a filtration step,
suspend the solid in an appropriate solvent and TLC the suspension.

Troubleshooting Thin-Layer Chromatography

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 Problem: My reaction is in a high boiling solvent (DMF, Pyridine, DMSO, Amine
solvents) and TLC just looks like a huge smear.
 Solutions: Spot the plate as usual and then put it in a flask under high vacuum for a few
minutes, and then run it, OR use TLC tip #1.

 Problem: My reactant and my product have very similar Rfs. I can't really tell what is
happening. How do I know when the reaction is complete?
 Solutions:
1. The cospot could help. If it looks like a snowman, your reaction is complete.
2. Try changing solvent systems.
3. Try staining with anisaldehyde. Compounds are different (bright) colors. Sometimes
color differences are also visible with molybdenum stains.

Problem: My compound might not be stable to silica gel (acid sensitive compounds can
be a problem.) How can I find out if it is stable to silica or not?

 Solution: Run a 2D TLC (TLC tip #4):

1. Using a square silica plate, spot the sample in one corner.
2. Run the plate in one direction (all components of the sample will appear in a vertical
line of spots).
3. Turn the plate 90 degrees (your "line of spots" should be at the bottom) and run the
plate again.
4. If the sample is stable to silica, all spots will appear on the diagonal. If a compound is
decomposing, it will appear below the diagonal.

Problem: My compound is unstable on silica. How can I monitor the reaction accurately?

 Solution: Try TLC tip 2, 3 or 4.

Problem: I don't want to expose my reaction to air- how do I get a TLC sample without
opening the reaction vessel?

 Solution: Set up your reaction flask such that it has a septum and is not under strong
positive pressure of an inert gas. Thread a capillary spotter into a 20 gauge disposable
needle, and put the needle into the septum. Use the spotter to get your sample, reinstall
your inert gas, and remove the needle. This procedure involves minimal exposure to air.

Problem: When I dip my capillary into my reaction to take a TLC, the reaction mixture
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 clogs the pipette and I can't spot it onto a plate. How do I TLC a heterogeneous or viscous
reaction mixture?

 Solution: TLC tip #1.

 Problem: My reaction looks streaky on TLC.

 Explanations and Solutions: This could mean a number of things.
o Massive decomposition. However, don't assume this just because your TLC is streaky.
o One of the reagents streaks: try TLC tip #1.
o Your product is intact, but unstable to silica. Try a 2D TLC (TLC tip #4).
o Your high boiling solvent is interfering with the TLC. See TLC tip #3.

Problem: My compound is very polar and stays on the baseline. I can't see what is happening
during the reaction.
Solutions: Try solvent systems effective for polar compounds, or use reverse phase silica gel
plates.

Problem: The TLC of my product mixture changed after work-up!

Explanation/Solution #1: Your product may not tolerate exposure to acid, base, air or water
and a reaction is occurring during workup. You can test small samples of your reaction
mixture before you quench it to figure out what causes the problem, which may allow you
to prevent the reaction.
Explanation/Solution #2: An unknown contaminant may have entered your material at some
point. Purify and see if things improve.

Troubleshooting Flash Chromatography

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 Problem: My compound is not stable to silica gel (if you want to find out if your
compound is stable to silica, see TLC tip #4. How can I purify it by chromatography if it
decomposes on silica?
 Solution: If the separation is an easy one, you could try purifying on florisil (200 mesh) or
alumina. For more challenging separations, it is possible to deactivate silica gel (reduce its
acidity) so that it is less damaging to your compound.

 Problem: Your compound should be off the column by now, but you're still collecting
fractions with no sign of it.
 Solution: A number of things could have happened here.
o The compound decomposed on the column and will never come off. Test your
compound for silica stability.
o You are not using the solvent system you thought you were using. Double-check the
bottles you used to prepare the solvent system and consider whether you could have
reversed the polar and the nonpolar element during preparation.
o Your compound came off in the solvent front. Check the first fraction.
o Your compound did come off, but the fractions are so dilute you haven't detected it.
Try concentrating fractions in the range you expected see your compound, then you
will be able to detect it.

 Problem: I cannot separate the components of my reaction mixture using column

chromatography, even though there is a large Rf difference. All the fractions are mixed. I
don't understand why this is happening.
 Explanations and Solutions:
One reason for this behavior is that you are being misled by thin layer chromatography.
You see two compounds, but one may be the degradation product of the other, and the
degradation occurs on silica gel. During a column, this process goes on during elution, and
all fractions have both product and degradation product. Check that your compound is
stable to silica (2D TLC).
Sometimes the wrong choice of solvent will also encourage this behavior- if the compound
with low Rf dissolved well in the eluant, and the compound with high Rf does not, you
may observe this behavior. Try to find a solvent system that dissolves both compounds
well.

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 Problem: My compound is very nonpolar and does not have an Rf as low as 0.3-0.4 in any
solvent system. What do I do?
 Solution: If it's an easy separation, a high Rf might not be a problem. If it is, you will want
to consider other purification methods. If the compound is a solid, you may be able
to purify by crystallization, or you are working on large scale, you could use distillation. If
none of these methods work, but your compound is mostly pure, think about the future of
the compound. Are you about to convert it to a more polar species that will be easier to
purify? Depending on the chemistry, sometimes it's possible to go on to the next reaction
with your mostly-pure compound and purify successfully at a later stage.

 Problem: My compound does not have any major impurities in it and I hate
chromatography. Do I have to run a column on it?
 Solution: It depends on the next step of the reaction sequence. The best answer is to try
the next step without purification, on a small scale, to see how it goes. Another option is to
run the compound through a short plug of silica, which will remove baseline impurities
and call allow you to skip the fraction-collecting stage of chromatography.

 Problem: My compound is very polar: it does not run up the plate even eluting with 100%
EtOAc. How do I purify it by chromatography?
 Solution: You could use reverse-phase silica for your column, or try a more aggressive
solvent systems to move it off the baseline. Solvent systems containing ammonia can be
useful in this context: prepare a stock solution of 10% ammonium hydroxide in methanol,
and try using 1-10% of this in dichloromethane for very polar compounds.

 Problem: I have less than 50 mg of compound, how do I purify it?

 Solution: You can run a miniature flash column by packing a short (6'') pipette with
cotton, sand and silica, just like you'd run a normal column. Choose a solvent system that
places your compound at an Rf of ~0.2. You will have to add solvent to the column every
1-2 fractions, but it's simple to apply pressure with a pipette bulb, and you'll be finished in
no time. This method works better than you might expect.

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 Problem: Your compound takes FOREVER to come off the column, it starts coming off at
a reasonable point in the elution, but it doesn't stop coming off until many many fractions
later.
 Solution: When the compound starts coming off the column, increase the polarity of your
eluting solvent. If there are no impurities at lower Rf, you can increase it significantly and
avoid the annoying trailing effect. It is important to keep the same two solvents, and just
increase the percentage of the polar component.

 Problem: I have two compounds very close in Rf, how do I separate them?
 Solution: Although it's not very much fun, the best way is to use a solvent gradient during
elution. You begin with a solvent system in which your compound runs at an Rf of about
0.2 (or less) and increase the percentage of the polar component of the solvent system
slightly each time you replenish the solvent in the column. You will need to experiment
with degrees of gradient for each difficult case before you find conditions that give optimal
separation. Some people find it's easier to run two columns instead of one. For difficult
separations, you will often have some mixed fractions- rather than throw them out or
contaminate pure fractions, keeping mixed fractions "for a rainy day" is recommended.
You can always purify your mixed sample later, if you need it.

 Problem: Your crude reaction mixture is not soluble in the solvent system you plan to use
to elute your column. You don't know how to load it on the column gracefully. (note: this
happens most often with the ethyl acetate/ hexane solvent system, and usually is only a
problem on large reaction scale.)
 Solutions:
o You can switch another solvent system like dichloromethane/ hexane or acetone/
hexane.
o A (rather risky) method is to dissolve your sample in very small volume of another
solvent (like dichloromethane). There is a fine line here between success and having to
repeat the column due to poor elution behavior.
o If you suspect that a reaction byproduct is the cause of the solubility problems, you
can try a silica filtration to remove the offending solid. Use a short plug and a solvent
that efficiently dissolves your product but not the impurity. You may want to collect
big fractions to achieve the best rough separation possible, before running the real
column.
o You can infuse your crude reaction mixture into a small quantity of silica, and load
your sample as a solid. Pack your column as usual with solvent-infused silica, but do
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 not top with a layer of sand. Using a large round-bottomed flask, dissolve your
compound in an appropriate solvent and add enough silica gel to form a thin (not
thick) layer on the top of your column. Remove the solvent by rotoevaporation. You
should have a flask of free-flowing silica gel that you can pour on the top of your
column. Add a layer of sand and elute as usual.

 Problem: Your compound has clogged the silica somehow- elution is very slow
 Explanations and "Solutions": This is not very common, but when it happens, it's a
major headache. The problem arises when either the compound or an impurity crystallizes
in the column, forming a solid barrier that blocks solvent flow. The prognosis is not good-
console yourself by noting the problem so you can avoid loading the loathsome mixture
onto a normal column in the future. You will either need to employ some pre-
chromatography purification technique or use a very wide column and a lot of silica.
o Take a thin wire and push up on the cotton plug from the spout of the column. If
you're lucky, the blockage is at the narrow point where the cotton is and can be
cleared.
o Take a long glass pipette, rod or similar implement and try "stirring" the silica slurry
in the column, just to get the solvent moving again. You'll have to start over to purify
it, obviously.
o If these methods fail, you'll have to pour the whole mess out through the top of the
column and perform a crude filtration to extract your sample from the silica. It is
possible this step will remove the insoluble material, and you can carry on with
chromatography.

Weighing Reactants and Reagents

 Problem: I need to weigh 0.5 mg of catalyst. The balance is not accurate for such small
weights.
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 Solution: Make a solution of (say) 5 mg of the solid in (say) 1 mL of solvent, and then
syringe out 100 microliters.

 Problem: I do not know the density of a liquid- how do I know how much to add via
syringe?
 Solutions:
o Weigh an empty syringe, fill the syringe (start by assuming the density is 1g/mL), and
weigh again. Adjust until the weight is accurate.
o Measure the density of the liquid by weighing a known volume and doing the
appropriate division
o For small amounts of viscous liquids, weigh an empty pipette, dip the tip in the
substance, and weigh again.

 Problem: When I try to use a syringe to measure a certain liquid, nothing happens because
it is too viscous to be drawn into the syringe.
 Solutions:
o Dilute the liquid with your reaction solvent, and then try using a syringe.

o For small amounts of viscous liquids, weigh an empty pipette, dip the tip in the
substance, and weigh again.

Problem: I have a small amount of an oil (say, 40 mg) in a flask. I only want to use some
of it (say 10 mg), but I do not know how to weigh it accurately.
 Solution: For the example given, dissolve the material in 4 mL of solvent, and remove 1
mL. You can remove the solvent to check the weight. This method is more accurate than
you might think.

 Problem: I want to weigh a hygroscopic solid, but it "melts" as I weigh it, and then I can
not easily tranfer it to my reaction.
 Solutions:
o Weigh the reagent in a glove box, if one is available.

o Weigh the reagent directly into your reaction flask, if practical.

o Weigh the reagent in a glass vessel, and then dissolve it in an appropriate solvent for
transfer.

 Problem: I want to weigh a reagent with a melting point near room temperature. It keeps
changing states on me- it clogs a syringe, but melts on weighing paper! (t-BuOH is one
example).
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 Solution: Warm it up slightly (if it is heat- stable) and syringe quickly, before it cools
enough to solidify.

 Problem: I want to syringe a liquid that evaporates at room temperature, and I never get
an accurate volume.
 Solution: Cool it to a low temperature before syringing it.

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