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What happens to semaglutide after twelve weeks at 30 °C in solution?

Asked 5 May 2024Modified 2.1 years agoViewed 24k times
36

The case in front of me: semaglutide · twelve weeks · 30 °C.

This is one of those things that everyone repeats and nobody derives.

This matters practically, not just academically, because it changes what I would do next.

Why does this happen, and what would falsify the usual explanation?

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askedines_delacruz16k165 May 2024
Is there a printed date on the vial, and do you know what it was derived from? – tess_amankwah 2 months ago
8Voting to keep this open — it is more specific than it first looks. – ben_akintola 14 days ago
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5 Answers

Accepted answer first, then by votes
89

Accepted answer

twelve weeks is 84 days, which at 30 °C is on the order of 475 refrigerated days. 30 °C is 25 kelvin above the 5 °C middle of a 2–8 °C refrigerator. The ten-degree rule of thumb — degradation rate roughly doubling per 10 K — makes that about 5.7 times the refrigerated rate, which is an order-of-magnitude statement and not a shelf life. In solution the routes that matter are hydrolysis of the backbone, deamidation at Asn, and physical association — the first two cost content, the third costs neither until it precipitates. Over 84 days at 30 °C you should expect all three to have moved, and a purity figure to have noticed only some of them. Reconstituted material has no certificate; the one in the box describes the powder.

In practice, asparagine and glutamine are the deamidation risk, and methionine is the oxidation risk.

Oxidation targets methionine, cysteine and tryptophan, adding sixteen daltons per oxygen. It is catalysed by trace metals and promoted by dissolved oxygen and by light.

Aggregation is physical: peptides unfold at air-liquid interfaces and associate. Shaking maximises that interface, which is why swirling and shaking produce visibly different outcomes on the same vial.

Metal-catalysed oxidation of methionine is documented across peptide and protein formulations and is why chelators appear in some formulations.

Swirl, never shake. Aggregation is a handling problem more than a time problem.

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LM
answered · acceptedleonid_marchuk19k276 Jul 2024
Does the same reasoning apply to material already in solution, or is that a different curve? – marta_szymanska 10 months ago
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80

The underlying point is that this is answerable from the chemistry rather than from anecdote, which is unusual and welcome.

Adsorption onto glass and plastic is significant at low concentrations — micrograms per millilitre — and negligible at milligrams per millilitre. It is the usual explanation for an apparent loss in a dilute preparation.

A mass spectrum resolves most of this: minus eighteen is dehydration or succinimide, plus one is deamidation, plus sixteen is oxidation, and an unchanged mass with a shifted retention time is an isomer.

Nothing here is medical advice, and research-use compounds are not approved for human use.

A mass spectrum names the pathway. Plus one, plus sixteen, minus eighteen.

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MF
answeredmeniscus_film32k2725 Jun 2024
4Thank you — this is the answer I was looking for. – plate_count_9k 9 months ago
5Small correction: it is the number of cycles rather than the freezer temperature that does the damage. – lukas_sedlacek 20 days ago
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39

Start with the sequence, because which pathways are available depends on which residues are present.

Light exposure matters for tryptophan-containing sequences and for anything with a chromophore. Amber vials and a closed box are free mitigations.

Hydrolysis cleaves the backbone, most readily at aspartate-proline and aspartate-glycine sequences, and is acid-catalysed. In a dry solid it barely proceeds at all.

Cold, dry, dark, still. Those four words cover most of the mitigation.

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VR
answeredv_ramaswamy68k573 Jun 2024
31

Aggregation is a physical process and is the one most often caused by handling rather than by time.

Freeze-thaw cycling drives aggregation through concentration at the ice interface and pH shifts as buffer components crystallise out at different rates. Each cycle costs something.

Deamidation via the succinimide intermediate is well characterised, with sequence-dependent rates highest for asparagine-glycine motifs.

At dilute concentrations, suspect adsorption before you suspect chemistry.

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HN
answeredhalvard_ness69k4714 Jun 2024
30

Answering this needs the physical state, since a dry powder is protected from most of these and a solution is protected from none.

Deamidation converts asparagine or glutamine to the corresponding acid via a succinimide intermediate, adding one dalton. It is base-catalysed, accelerates above neutral pH and is the dominant aqueous pathway for many peptides.

Sequence determines which pathways apply, so general statements are general.

Sequence decides which pathways are even available. Check the residues.

edited 9 Jun 2024 by halvard_ness — added the method parameters

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HN
answeredhalvard_ness69k4712 May 2024
Any published figure for how much a collapsed cake actually retains? – Dr_Colm_Fitzhenry 4 days ago
Is there a reason to prefer minus eighty here, or is minus twenty genuinely enough? – t_oyelaran 8 months ago
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Your answer

Ask PeptideStack is a static archive. Posting is closed, but the norms are worth stating: answer the question that was asked, show your working, cite the trial or the certificate, and say plainly where the evidence runs out.

Not medical advice. Research-use-only compounds are not approved for human use.