Peptide Reconstitution

Peptides are short stretches of amino acids often referred to as the building blocks of proteins. Peptides often mimic the function of essential proteins or hormones and help to establish the same chemical action, albeit in a more controlled way, of their parent proteins.

 

Lyophilized Peptides

Manufacturers sell peptides in freeze-dried powder, or lyophilized form. Lyophilization is the technique whereby water is removed from a solution upon freezing. The use of vacuum allows ice to be directly converted into vapor from solid without passing through the intermediate liquid phase. The powder either has a granular appearance or looks fluffy, depending on the technique used for lyophilization.

 

Peptides Reconstitution

Lyophilized peptides need to be dissolved in a solvent before use. There is no universal solvent that can be used for all the peptides. The choice of solvent is guided by peptide stability and compatibility of the specific solvent with bioassays. In this respect, sterile water or bacteriostatic water is often the first solvent choice. Despite being the first choice, sterile or bacteriostatic water cannot dissolve every peptide. Hence experimentation with different solvents is often required for optimizing the solvent most relevant for employment in research studies. Sodium chloride solution in water is not advised to be used for dissolving peptides to avoid precipitation with acetate salts.

The polarity or native charge of a peptide is the key determinant for its solvent. Basic peptides require acidic solvents, whereas acidic molecules require basic solvents for optimal dissolution. The hydrophobic and neutral molecules dissolve best in organic solvents like DMSO, propanol, and acetic acid. The powder should be dissolved in a small volume of the organic solvent and further diluted in sterile water. It is important to note that peptides with methionine or free cysteine should not be done in DMSO to avoid side-chain oxidation or being rendered inactive.

 

Peptides Reconstitution Guidelines

The selection of appropriate solvent demands sterile water, 0.1% acetic acid, or any solvent that can be easily removed by lyophilization. Dissolving a small amount of the peptide in the chosen solvent is recommended before dissolving the complete vial. The molecule should always be dissolved to form a higher stock concentration than that required for the actual assays. It can be diluted further in the appropriate assay buffer to derive the appropriate concentration in the bioassays.

 

Sonication

Upon resuspension, visible particles sometimes persist in the peptide solution. The vial can then be sonicated to enhance the dissolution of the particles to yield a clear solution. Sonication does not modify the solubility of the peptide but expedites the breaking up of the particles to enhance solubility. Post sonication, the researcher has to inspect for the presence of any surface layer, cloudy appearance, or gel-like consistency. If so, then the peptide has merely been suspended but not dissolved. Such a case would indicate the need for a more potent solvent.

 

Practical Implementation in the Laboratory

In this segment, we have outlined simple guidelines for the effective dissolution of peptides in the laboratory. Most peptides have recommended storage temperatures as lyophilized powder and solution and should be adhered to. If the powder form of the substance is stored in refrigerated condition, it should be equilibrated to room temperature initially before resuspension. The peptide solution can be filtered through a 0.2 µm filter to avoid any possible bacterial contamination since it is a rich source of amino acids.

 

Disclaimer: The products mentioned are not intended for human or animal consumption. Research chemicals are intended solely for laboratory experimentation and/or in-vitro testing.  Bodily introduction of any sort is strictly prohibited by law.  All purchases are limited to licensed researchers and/or qualified professionals. All information shared in this article is for educational purposes only.