To Freeze or Not To Freeze: Lyophilized Peptides

[pizzathehutt]

Should you freeze peptides that you’ve yet to reconstitute? I’ve seen partial answers all over.

The old QSC discord was also full of people talking about deep freezing their extras.

The recommendations from US labs for freezing seem to assume that your powder would be free of moisture content, stored at -20C, and that you avoid thawing and refreezing.

While I love our underground friends and have yet to test, I suspect we have moisture in our vials. Enough to be thrashing our peptides if we decide to freeze.

I work with yeast and some other items that are very finicky with freezing without being dry or leveraging glycol. You can quickly degrade your stock with just a short freeze.

Has anyone tested the impact of freezing on underground peptides like Tirz?

 

[Ghoul]

Should you freeze peptides that you’ve yet to reconstitute? I’ve seen partial answers all over.

The old QSC discord was also full of people talking about deep freezing their extras.

The recommendations from US labs for freezing seem to assume that your powder would be free of moisture content, stored at -20C, and that you avoid thawing and refreezing.

While I love our underground friends and have yet to test, I suspect we have moisture in our vials. Enough to be thrashing our peptides if we decide to freeze.

I work with yeast and some other items that are very finicky with freezing without being dry or leveraging glycol. You can quickly degrade your stock with just a short freeze.

Has anyone tested the impact of freezing on underground peptides like Tirz?

I wouldn't. Household freezers are nothing like lab freezers, not getting nearly as cold, but worse, subject to wide temp fluctuations. If moisture content of the peptide is higher than lab standards, you could end up with crystals expanding and contracting within the peptide breaking the proteins chains apart.

Personally, I think dark refrigeration is the best environment. Many reconstituted peptides are good for years in that environment, including Sema and Tirz.

 

[narta]

Just stick it in the fridge and don't buy a truckload. Glps are going down in price all the time until they stabilize lower, look at Sema.

 

[pizzathehutt]

Just stick it in the fridge and don't buy a truckload. Glps are going down in price all the time until they stabilize lower, look at Sema.

For sure.

I already have a stockpile and storing in the fridge has done well for me. I do have a bit more tirzepatide than I'll need for a long time which I may share just so it doesn't go to waste.

I'm mostly curious about if anyone has actually tested, say some QSC peptides and specifically GLP-1 agonists, after freezing. Or, if they haven't, what arguments they have for accepting the risk of freezing.

The last QSC Discord is what sparked this as there were half a dozen peptiders talking about deep freezing. But, I could never catch them to ask about this.

I have no plans to freeze because of what's stated above. I don't have a freezer that doesn't have a thaw cycle -- nevertheless one that will go to -20C. I also suspect that there's a good deal of moisture in our vials.

Someone mentioned they checked for frost but that's a bad "test" as you won't be able to visually inspect for crystallization in their peptide cakes. They also go through lengths with desiccant packs and vacuum containers. But that won't take the moisture out of a vial.

 

[Ghoul]

For sure.

I already have a stockpile and storing in the fridge has done well for me. I do have a bit more tirzepatide than I'll need for a long time which I may share just so it doesn't go to waste.

I'm mostly curious about if anyone has actually tested, say some QSC peptides and specifically GLP-1 agonists, after freezing. Or, if they haven't, what arguments they have for accepting the risk of freezing.

The last QSC Discord is what sparked this as there were half a dozen peptiders talking about deep freezing. But, I could never catch them to ask about this.

I have no plans to freeze because of what's stated above. I don't have a freezer that doesn't have a thaw cycle -- nevertheless one that will go to -20C. I also suspect that there's a good deal of moisture in our vials.

Someone mentioned they checked for frost but that's a bad "test" as you won't be able to visually inspect for crystallization in their peptide cakes. They also go through lengths with desiccant packs and vacuum containers. But that won't take the moisture out of a vial.

The better question is what's to gain by freezing? Laymen think it preserves chems better but that's not always the case. Even dry tablets can be damaged by freezing.

The fact they use the archaic term "deep freezing" tells me they don't know wtf they're talking about. Combo fridge freezers and chest freezers have had the same freezing capabilities for about 40 years now, lol.

 

[deadbeef]

I wouldn't. Household freezers are nothing like lab freezers, not getting nearly as cold, but worse, subject to wide temp fluctuations.

Define "wide temp fluctuations". And why would it matter if the temps never go above the freezing point?

If moisture content of the peptide is higher than lab standards, you could end up with crystals expanding and contracting within the peptide breaking the proteins chains apart.

Why would they expand and contract if the temp never gets above the freezing point?

 

[deadbeef]

I don't have a freezer that doesn't have a thaw cycle -- nevertheless one that will go to -20C.

Any relatively modern freezer should be able to do -20C. That's -4F. My small chest freezer will do -26C (-15F).

 

[pizzathehutt]

Define "wide temp fluctuations". And why would it matter if the temps never go above the freezing point?



Why would they expand and contract if the temp never gets above the freezing point?

It's more of an issue with crystallization in general which may occur during the initial freeze. The potential decrystallization and recrystallization due to fluctuations just makes it worse.

Peptide structures are very delicate.

If underground peptides leave just a lick of moisture in the vial, and are not properly vacuum sealed, crystals may form and physically break the peptide bonds or alter their conformation for their main chains. That's not including the secondary and tertiary structures the crystals can distort -- which will cause misfolding and/or denaturation. Or, aggregation which nullifies their functional props and/or become difficult to reconstitute.

Initial freezing, and especially thawing and refreezing, could introduce moisture as well. If the vials themselves are not sealed well + whatever container you may have them in.

I suppose we'll only know if we spot check the moisture levels in some vials from places like QSC. I may work out a way to do this with some of my extras soon.

Any relatively modern freezer should be able to do -20C. That's -4F. My small chest freezer will do -26C (-15F).

Have you tested to ensure it actually gets that low and maintains that temp? I know some folks that have serious Grafana monitors set up w/alerts and regularly catch variations that could put things at risk.

 

[deadbeef]

It's more of an issue with crystallization in general which may occur during the initial freeze. The potential decrystallization and recrystallization due to fluctuations just makes it worse.

Why would you have decrystallization and recrystallization if it never goes above the freezing point?

If underground peptides leave just a lick of moisture in the vial, and are not properly vacuum sealed, crystals may form and physically break the peptide bonds or alter their conformation for their main chains. That's not including the secondary and tertiary structures the crystals can distort -- which will cause misfolding and/or denaturation. Or, aggregation which nullifies their functional props and/or become difficult to reconstitute.

Moisture is bad for peptides. Which is worse, a single freeze/thaw, or continual exposure to moisture in the fridge? If there is a bad seal, wouldn't bacteria growth be more likely in a fridge vs a freezer?

Initial freezing, and especially thawing and refreezing, could introduce moisture as well. If the vials themselves are not sealed well + whatever container you may have them in.

Again, why would there be more than one freeze/thaw? For the bad seal, you'd have the same issue in the fridge?

I suppose we'll only know if we spot check the moisture levels in some vials from places like QSC. I may work out a way to do this with some of my extras soon.

The testing server has done moisture testing in various tests. I'm in a test that will be doing another moisture test soon. As far as I'm aware, there has not been a widespread moisture issue.

Have you tested to ensure it actually gets that low and maintains that temp? I know some folks that have serious Grafana monitors set up w/alerts and regularly catch variations that could put things at risk.

Yes:



You can see I keep my manual defrost chest freezer below -20C. I don't run it colder because I want to save energy and extend compressor life.

Even my combo fridge/auto defrost freezer doesn't come close to the freezing point:



The spike is the defrost cycle. The max I've logged it at during a defrost cycle is 8F.

 

[pizzathehutt]

Good points, exactly what I was looking for in terms of a discussion with someone experienced with this.

The testing server has done moisture testing in various tests. I'm in a test that will be doing another moisture test soon. As far as I'm aware, there has not been a widespread moisture issue.

I'd be interested in joining in on this. I produce various things such as dry brewing yeast vials where I have to measure and maintain minimal moisture. I have the resources and a set up to contribute.

I also make liquid brewing yeast freezable for long term storage without having to go through a drying process. Things are easier in that world, though, as folks are propagating up from the vials and you can still proceed even if you lost a chunk of viability. My stuff comes out of the freezer quite viable, though.

Yes

Nice set up.

That's something we have for other purposes but not at home. I can't store my stuff there, either.

Unfortunately, I guess I need new freezers at home as mine are not that consistent.

I'm not against freezing and this info is exactly what I'm looking for as I build up stock.

 

[Ghoul]

Freezing in this discussion is the common meaning referencing the freezing point of pure water.

Any contaminant water is in contact with will lower the freezing point. The residual water in lypholized peptides is not only not "pure", the impurities in the vial contents, and no peptide is ever 100%, are largely salts. It's entirely possible the freezing point of the water in a peptide is much lower than pure water, and your freezer fluctuation, while always below 0c "freezing", could cause crystals to form, melt, and then reform many times a day as it repeatedly crosses the lower, unknown freezing point and back above.

It's for this reason the absolute minimum freezer storage temp recommended by peptide manufacturers is no higher than -20c, including fluctuations never rising above that temp. Impossible in a home freezer. The ideal is -80c, only achievable in an advanced lab freezer, which ensures even the most salt saturated water will remain stably frozen.

Keeping the peptide at the lowest possible refrigeration temperature will slow chemical degradation processes to a minimum while avoiding the hazards of freeze/thaw cycles inflicting damage on the peptide chains.

 

[deadbeef]

Freezing references the freezing point of pure water.

I never said the freezing point of water.

Any contaminant it's in contact with will alter that point. The residual water in lypholized peptides is not only not "pure", the portion of the peptide between its measured purity and 100% are largely salts. It's entirely possible the freezing point of the water in a peptide is much lower than pure water, and your freezer fluctuation, while always below 32f "freezing", could cause crystals to form, melt, and then reform many times a day as it repeatedly crosses the lower, unknown freezing point and back above.

As noted above, my freezer stays below -4F (-20C). Some quick googling shows legit peptide manufacturers specify to store their lyophilized peptides at -20 to -80C for long term storage. I doubt they would specify a long term storage temperature where it could be freezing/thawing regularly, especially since they specify freezing/thawing is undesirable.

 

[Ghoul]

I never said the freezing point of water.



As noted above, my freezer stays below -4F (-20C). Some quick googling shows legit peptide manufacturers specify to store their lyophilized peptides at -20 to -80C for long term storage. I doubt they would specify a long term storage temperature where it could be freezing/thawing regularly, especially since they specify freezing/thawing is undesirable.

You have no idea what the freezing point of the water content of the peptides we're supplied is.

You don't even know what the water content is, as purity tests specifically exclude water from that measurement.

As a matter of fact, it's in the UGL manufacturer's interest to package peptides with the highest amount of water content they could get away with, and even if not intentional, many peptides are hydrophilic, rapidly absorbing moisture from exposure to air as it's being packaged. The idea UGL manufacturers are maintaining, or even have the extremely expensive capacity, or motivation to maintain the tightly controlled packaging environments of legitimate manufacturers is laughable.

You can do what you like, but to suggest there's no way repeated freeze thaw cycles could be taking place is nonsense. Even your own freezer, if we're to take your measurements at face value, could stray far from -20c for any number of reasons, including opening the lid, adding contents, or the steady buildup of frost on the coils.

 

[deadbeef]

if we're to take your measurements at face value

I come with data, you come with conjecture. We're done here.

 

[Ghoul]

What manufacturers actually advise, is not to engage in long term storage at all, but to only acquire a sufficient amount of the peptide to conduct whatever experiment is at hand, or, to quickly incorporate the peptide as an API into your pharmaceutical and get it into the supply chain with the appropriate expiration date.

Are lyophilized pharmaceuticals like HGH kept frozen at the pharmacy? Or anywhere in the distribution system, NO.

Somatropin, like all pharma peptides, is kept at 36f while in "long term" storage.

Sema and Tirz are immediately reconstituted in BAC by Novo and Lily and given a 2 year expiration date then stored and shipped at approx 36f.

But you know "better" than the pharma companies do to keep their products in useable condition, maintaining proper clinical potency, for the longest time possible?

The same companies, that, if they supply damaged peptides that aren't properly effective, would be given a manufacturing time out by the FDA and gorged on by trial lawyers? Use some common sense.

 

[rypeptideguy]

Do you think vacuum sealing them before placing in a -20c chest freezer would make a difference? I guess not since the water would already be in the the vial.

 

[qadsan]

Just another point to consider is that moisture contamination can occur when vials of peptides are opened immediately after bringing them out to room temperature from frozen conditions. The peptides tend to absorb moisture from the cool air inside the container or the inside of the container. Hence, the best practice would be to allow the peptide to equilibrate to room temperature before opening.

I would think for long term storage that also keeping peptides in a dessicator would be more ideal in terms of minimizing the chances of moisture. I dessicate my raws for long term storage, but haven’t done so with my peptides.

 

[sushak]

Isn't it all moot if you store your peptides in a good vacuum flask inside the freezer? Keeps the temps of the peptides stable, even during a auto defrost cycle.

I don't have the means, but could you collect data with your sensor inside a thermos flask inside the freezer for comparison?

 

[qadsan]

I don't have the means, but could you collect data with your sensor inside a thermos flask inside the freezer for comparison?

That would be a cool experiment to see the data from this. My dessicator is too large to put in any of my freezers, otherwise I’d give it a try. I don’t have any smaller vacuum containers or spare sensors to try this with right now, but I’ll keep it in mind for later this year if nobody else does this.

Getting a sealed container and adding desiccant and o2 absorbers and data logging freezer temps and humidity both inside and outside the container would be interesting.

 

[TyroneBiggums]

Should you freeze peptides that you’ve yet to reconstitute? I’ve seen partial answers all over.

The old QSC discord was also full of people talking about deep freezing their extras.

The recommendations from US labs for freezing seem to assume that your powder would be free of moisture content, stored at -20C, and that you avoid thawing and refreezing.

While I love our underground friends and have yet to test, I suspect we have moisture in our vials. Enough to be thrashing our peptides if we decide to freeze.

I work with yeast and some other items that are very finicky with freezing without being dry or leveraging glycol. You can quickly degrade your stock with just a short freeze.

Has anyone tested the impact of freezing on underground peptides like Tirz?

I throw everything in the freezer that i know im not using. The peps are lyophilized so they shouldnt contain any moisture. Ive never seen moisture in any of them.