Storing peptides & shelf life: the laboratory principles

Research context (RUO): This guide describes how peptides are stored in a laboratory environment and what determines their shelf life. All substances mentioned are supplied exclusively for in-vitro laboratory research (Research Use Only) and are not intended for human or animal use.

Storing peptides & shelf life: the laboratory principles

Storing peptides is one of the most underestimated steps in working with research peptides: how a peptide is stored largely determines whether it is still intact and analysable weeks or months later. Peptides are short chains of amino acids, and it is precisely that amino acid chain that is sensitive to heat, moisture, oxygen and light. This article describes how researchers store lyophilisate (freeze-dried powder) and reconstituted (dissolved) peptides, why degradation occurs and which factors determine the shelf life after dissolving. The focus is purely laboratory handling, not use advice.

Lyophilisate versus reconstituted: two very different shelf lives

Peptides are almost always supplied as lyophilisate: a freeze-dried, dry powder in a sealed vial. In that dry form a peptide is chemically relatively stable, because the main degradation reactions require water. According to the storage guidelines of manufacturers such as GenScript, lyophilised peptides usually remain stable for several years under the right conditions. As soon as a peptide is reconstituted — dissolved in a liquid such as bacteriostatic or sterile water — that picture changes drastically. In solution, water-driven degradation processes get going and the shelf life drops from years to, depending on the peptide and the storage temperature, usually days to a few weeks. Which liquid is used when is compared in bacteriostatic water, sterile water and acetic acid.

The practical principle that researchers derive from this: a peptide is kept as dry lyophilisate for as long as possible and is only dissolved just before use in an experiment. More on that step is in the guide on reconstituting peptides.

Storing peptides as lyophilisate: temperature, moisture and light

For dry lyophilisate, three key points apply. First, temperature: storage guidelines advise storing lyophilised peptides at approximately −20 °C; for long-term storage, −20 °C to −80 °C is often maintained. For brief handling in the lab, refrigeration (2–8 °C) is usually sufficient, but the lower the temperature, the slower every chemical reaction proceeds.

Second, moisture. Many peptides are hygroscopic: they attract moisture from the air. When a cold vial is opened immediately, air humidity condenses on the cold glass and powder. That is why researchers first let a vial come to room temperature before the cap comes off, and lyophilisate is stored in a well-sealed vial, often with a drying agent (desiccant). Peptides with amino acids such as aspartic acid, glutamic acid, lysine, arginine or histidine are extra moisture-sensitive and, according to the GenScript guidelines, are stored in a closed vial in a desiccator.

Third, light. UV and bright light can damage sensitive amino acids, which is why vials are stored in the dark or in a closed box. As long as these three conditions — cold, dry, dark — are met, dry lyophilisate is the most stable storage form.

Degradation: why peptides lose their integrity

Shelf life ultimately comes down to chemical and physical degradation. A technical article by MilliporeSigma on peptide stability describes several degradation routes that are relevant during storage:

• Oxidation: amino acids such as cysteine (Cys), methionine (Met) and tryptophan (Trp) react with oxygen. Peptides with these residues are therefore stored in tightly closed vials and as oxygen-free as possible.
• Deamidation: asparagine and glutamine residues can convert, which changes the structure of the peptide.
• Hydrolysis: in solution, peptide bonds can be broken by water; this is the main reason that dissolved peptides have a shorter shelf life.
• Aggregation: peptide molecules can clump together, especially with repeated temperature fluctuations.

These processes proceed faster the higher the temperature and the more water, oxygen or light is present. Correct storage essentially comes down to slowing down all these reactions as much as possible.

Shelf life after dissolving: storing reconstituted peptides

Once a peptide is dissolved, cooling is not a luxury but a necessity. Storage guidelines advise against keeping peptides in solution for long periods; if it is nevertheless necessary, the solution is kept at −20 °C, or preferably −80 °C. Two practical techniques then determine the shelf life:

• Aliquoting: the dissolved stock is divided over several small vials. According to the guidelines, aliquoting reduces both the number of freeze-thaw cycles and the amount of air exposure per portion.
• Avoid freeze-thaw cycles: peptides that are repeatedly frozen and thawed are susceptible to degradation. Each cycle is an opportunity for hydrolysis and aggregation, so researchers thaw only what an experiment requires.

The choice of solvent also plays a role. Bacteriostatic water contains 0.9% benzyl alcohol, which inhibits bacterial growth and thus supports a longer shelf life of a solution, whereas sterile water contains no preservative. Which liquid suits which research protocol is set out in the guide dissolve peptide: which liquid. The required materials — BAC water and sterile water — can be found in the category peptide accessories.

Quality & purity as a starting point

Correct storage starts with a pure starting product. A peptide that already contains impurities or degradation products on receipt ages unpredictably. At Peplife, every relevant batch is independently HPLC-tested by an external laboratory; the accompanying certificate of analysis (CoA) is publicly verifiable per batch. This way it is known what purity and composition are assumed before the storage conditions start to determine the shelf life.

Frequently asked questions about storing peptides

How long do lyophilised peptides keep?
Under the recommended conditions — cold (approximately −20 °C), dry and dark in a sealed vial — many lyophilised research peptides usually remain stable for several years according to manufacturer guidelines. The exact shelf life depends on the amino acid composition.

How long does a reconstituted peptide keep?
Considerably shorter than lyophilisate. In solution, storage at −20 °C to −80 °C is advised, and the shelf life is usually spoken of as days to a few weeks, depending on the peptide, the liquid and the temperature. Aliquoting and avoiding freeze-thaw cycles are decisive here.

Should peptides go in the freezer or is the refrigerator enough?
For long-term storage of both lyophilisate and dissolved peptides, guidelines point to the freezer (−20 °C, or −80 °C for the long term). The refrigerator (2–8 °C) is mentioned in the literature for short-term handling; lower temperatures slow down all degradation reactions.

Why must a vial come to room temperature before it is opened?
Because many peptides are hygroscopic. If a cold vial is opened immediately, air humidity condenses on the cold powder, which accelerates moisture-driven degradation. Letting it warm up to room temperature before opening limits this.

Are all peptides light-sensitive?
Not to the same extent, but bright and UV light can damage sensitive amino acids. As a precaution, vials are stored in the dark. Peptides with cysteine, methionine or tryptophan are also oxidation-sensitive and are stored as oxygen-free and closed as possible.

Why is aliquoting recommended?
Aliquoting divides a dissolved stock over small portions, so that only one vial needs to be thawed per experiment. This reduces both the number of freeze-thaw cycles and the air exposure of the rest of the stock, and thus supports the shelf life.

Read more & research at Peplife

• Research BAC water at Peplife
• Sterile water at Peplife
• View all peptide accessories
• Reconstituting peptides: step by step
• Dissolving peptides: which liquid

Sources: GenScript — Peptide Storage and Handling Guidelines · MilliporeSigma — Peptide Stability · Ó’Fágáin & Colliton — Storage and Lyophilization of Pure Proteins (Methods Mol Biol, 2023)

Research Use Only. All products are supplied exclusively for in vitro laboratory research. Not intended for diagnostic or therapeutic use in humans or animals, and not approved by the EMA or FDA.