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Small Molecules Solubility
The solubility of small molecule products, especially those that are hydrophobic can sometimes be a limiting factor in experimental design. Ideally one would like to deliver all treatments in water or a simple buffered aqueous solution, but this is often not possible.
How should I dissolve the compound I purchased from you?
The product datasheet supplied with a product will include information on the compound’s solubility. Many compounds that are not directly soluble in water can be dissolved in alcohols, or solvents such as DMSO or DMF to create a concentrated stock solution. These stock solutions are often reasonably stable for storage, and can be diluted to the desired final treatment concentration with water or aqueous media at the time of use.
We strongly recommend using anhydrous solvents whenever working with a compound that is not water soluble. Solvents like DMSO are hygroscopic, and if the solvent has picked up water from the atmosphere it can be problematic. Even very tiny amounts of water can significantly impact the limit of solubility of hydrophobic compounds.
In the majority of cases, solubility in either ethanol or DMSO will be high enough to allow for a 1/500 to 1/1000 dilution with water or aqueous buffer to achieve the final treatment concentration desired.
Small amounts of residual organic solvent (for example 0.1% DMSO) do not normally affect biological systems, however we strongly recommend that solvent controls be run alongside all experiments as controls. For those compounds where these common techniques are not sufficient, the datasheets will provide additional instructions specific to the particular compound in question.
How do I dissolve my peptide?
Reconstitute peptides in sterile, distilled water. A few drops of ammonium hydroxide for acidic peptides or 10% acetic acid for basic peptides will facilitate dissolution. If necessary the solution can be sonicated briefly. With extremely hydrophobic or neutral peptides, solubilize using a minimal amount of dimethylsulfoxide (DMSO) or dimethylformamide (DMF) then add water or buffer. In order to avoid serious solubility problems, fully dissolve peptides before adding buffer or saline. Use oxygen-free solutions and reducing agents with peptides containing Trp, Met or free Cys to avoid oxidation.
The datasheet says the compound will dissolve at this concentration, but I still see solid material. What should I do?
In most cases the speed at which a crystalline solid will dissolve is dependent upon three main factors, the affinity of the compound for the solvent in question, the size of the crystals within the sample, and the temperature in the room. Warming the solution is often the easiest way to speed up solubilization. Most products can tolerate being warmed in a 37°C water bath for brief periods without damage, and this alone, or combined with vortexing or sonication is often enough to dissolve the compound. If the solid does not dissolve after brief warming with agitation or sonication we suggest that you contact our technical support for further assistance tailored to the specific compound you are working with.
My compound dissolved well in DMSO, but when I tried to dilute the DMSO stock solution with aqueous media a precipitate formed.
It is common for compounds to precipitate when a stock solution in a solvent such as DMSO is diluted with aqueous media. Most of these precipitates will redissolve with a few minutes of vortexing or sonication, or brief warming in a 37°C water bath with sonication. While this type of precipitation will not harm the compound, it is important to take the time to be sure the precipitate has completely redissolved before using the solution.
My stock solution was initially clear, but when I removed it from the freezer to use it a second time, a precipitate formed.
The process of a solution freezing and thawing can sometimes cause chemicals to crystallize or precipitate out of solution. This is most common in solutions at concentration near the upper limit of solubility. These precipitates are temporary, and will normally redissolve on their own as the solution warms up to room temperature. If the precipitate is slow to redissolve, a few minutes of warming in a 37°C water bath with sonication will normally solve the problem.
My datasheet says to add BSA or serum to the aqueous solution before attempting to dilute my stock solution. Why is this suggested?
BSA and other serum albumins are natural carrier proteins for hydrophobic small molecules such as hormones and fatty acids. They are highly water soluble yet contain distinct hydrophobic binding domains that bind non-specifically to many hydrophobic molecules. The association of a hydrophobic small molecule with BSA can often help to keep the hydrophobic small molecule in solution.
In general, BSA will form a 1:1 association with a hydrophobic small molecule, so the molar concentration of the BSA should be the same as, or slightly higher than the molar concentration you are attempting to achieve with the small molecule. As with the organic chemical solvents, it is important to carry out appropriate solvent controls with your experiments.
How can I dissolve lipids for use in cell culture or other biological experiments?
Most lipids are strongly hydrophobic compounds, and working with them in an aqueous media such as for cell or tissue culture can be difficult. Most lipids can be complexed with carrier proteins such as BSA as described above to allow for some limited use in predominantly aqueous media, at least at lower concentrations. Other options would be to deliver the lipid in suspension rather than as a true solution (vortex or sonicate the suspension immediately before dispensing to make sure the suspension is as uniform as possible), or to deliver the lipid in micelle form.
