Voices of Sample Management - Episode 7: The Impact of Hydration on DMSO Sample Integrity

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The Summary:

In Episode 7 of the Voices of Sample Management podcast, Toby Winchester, Product Manager at Titian, and Stuart Naylor, Owner of Roylan Developments, discuss the sample integrity challenges that come with potential effect of hydration of DSMO stocks on scientific assays. 

From DMSO, the universal solvent, to flooding storage with nitrogen, Toby and Stuart cover a wide range of solutions to avoiding hydration problems in samples. 

"This game is about doing what you can, where you can, to reduce that moisture absorption." Stuart Naylor

If you have any topics or ideas for our future episodes, or you're interested in taking part, don't hesitate to get in touch with us at info@titian.co.uk

Available as a video or podcast, you can also view the transcript below:

The Transcript:

Toby Winchester

Hello everybody. Welcome to another episode of Voices of Sample Management.

Today, I have a long term colleague from the world of sample management, Stuart Naylor. Could you introduce yourself?

Stuart Naylor

Sure. So, I guess I've been in the industry for 25 years or so.
It's flown by. I started out working for a company called the Technology Partnership, which metamorphosized into SPT Labtech.

I had an awful lot to do with their compound storage system called comPOUND.

Did a stint with TekCel who ended up becoming Hamilton Storage and since 2008, I have been running my own business called Roylan Developments.

And we make, I guess pertinent to the topic of this conversation, nitrogen storage systems for compound management, which do a pretty good job of keeping moisture out of samples.

Toby Winchester

OK, thank you very much. So yes, the topic of today is, you know, the impact of DMSO on our wonderful small molecule integrity, mainly small molecules.

But I think we should probably just start with DMSO. It's known by some as the universal solvent because it dissolves both polar and you know, non polar samples quite nicely. But can you explain what's so special about DMSO, other than it's solubility abilities?

Stuart Naylor

Well, as you say, it's the universal solvent. Practically every lab uses it to solubilize their compound collections. And provided it's free of moisture, it does a very, very good job of that.

Toby Winchester

So why does it absorb moisture so readily? I suppose is it part of its structure that helps that or?

Stuart Naylor

I think so, yes.

Toby Winchester

Yeah. I suspect there's a lot of hydrogen bonds in the DMSO, isn't there? Which is what water likes to combine with

Stuart Naylor

Yes. And it makes it aggressively hydroscopic.

I remember back when I first came into the industry, I did an experiment where I took a glass speaker and filled it up with the quantity of DMSO, and stood that on a scale and left it for 24 hours and came back and it gained about 20% weight just sucking moisture in from the atmosphere.

And then came back another day later and I think it had gone up to more like 33%

OK, which over a one to two day, is obviously a huge change in what was in that beaker

Toby Winchester

And that's a beaker of DMSO, so, that had sort of a large volume, whereas, you know, small wells in a plate probably have a worse surface area to volume ratio.

So it's even quicker with smaller samples.

Stuart Naylor

Yeah. But then even more incredibly I guess than that, there have been a number of customers that I visited over the years that have been using sealed plates.

There was one in particular I remember early on in my time in the industry that was actually using sealed deep wall blocks and they were investigating into the contents of them.

And bear in mind these are sealed can polypropylene containers. And they were finding the wells around the outside of the plate actually gain significant volume and particularly the corner wells and what they were finding, and this has since been looked at in various studies, that polypropylene is actually quite porous

Toby Winchester

Right.

Stuart Naylor

Combined with that aggressive hydroscopic nature of DMSO, over time it will actually suck moisture in through the side wall of your plastic container.

But, obviously that takes quite some time, but then some of these samples can be stored for quite some time.

And I think the picture that I remember most, was on a few of these deep well blocks, the foil seal had actually busted off in a few of the corners, due to the sheer amount of moisture that had got in.

Toby Winchester

Wow. That’s, so it's like a, you know, a gone off bottle of wine or something. The cork blows off. Yeah, absolutely. Wow.

So yes, so obviously water gets absorbed into DMSO quite readily, at least to the, you know, 30% factor. And then things slightly change because the phases of the mixture changed slightly.

But what effect does that have on the precious potential new drugs that are in dissolved in that DMSO?

Stuart Naylor

Well, it can have… I guess several effects and when you put them all together it can be quite bad.

But running through them, I mean, you know, one of the easy ones is dilution.

Toby Winchester

The volume’s gone up so it's not the concentration you thought it was.

Stuart Naylor

So yeah, if you've had your sample diluted to say a 70% DMSO concentration with the rest being water or moisture that's ingressed, obviously that's a chunk of dilution off the bat.

But I would say worse than that is you don't need much moisture absorption into your DMSO… for it’s solubilizing capabilities to start getting depressed.

And that really accelerates the more the moisture gets in.

Again, using this sort of 70% DMSO purity level, at that point, nominally, you've slightly more than halved the solubilizing capability of DMSO.

Toby Winchester

Right. And that would be different from between molecule and molecule you're trying to solubilize.

But yeah, and there's an overall effect that's quite marked, isn't it?

Stuart Naylor

Yeah, yeah, yeah.

Toby Winchester

Things will start falling out of solution into the bottom of the tube.

Stuart Naylor

Yeah. And then if you're into freezing your compounds, that's where the effect of freeze thaw cycles really become damaging because that moisture absorption has taken effect.

Obviously the freezing points of water and DMSO are slightly different.

You can get these crystallisation effects as your solution starts to freeze, which again can become damaging to your compounds.

But you know, if you put that effect one side for a moment and just look at the first two, let's say that you lose about half the compound in your well to precipitation.

And then you've had about 30% dilution, when you actually look at what you're gonna be pipetting out sample wise to go into an assay, it's, you know, kind of a mile away from really where you really want it to be.

Toby Winchester

Yeah, definitely not ideal. Experimental error coming in for, I suppose that's one argument maybe for using fresh samples each time you know, neat, solubilised, the DMSO and then straight away you're cutting down that time for hydration.

But most research companies don't make an awful lot of sample so that would be quite wasteful in chemistry time to make enough bulk in order to go that way.

You can see the advantage maybe, when you know, maybe in the clinical study area you can get away with that sort of thing.

But in the discovery area where amounts of small molecules isn't very high, you're always going to be, you know, solubilising and then keeping that sample of solute for a while at least.

Stuart Naylor

Yeah. Well, you know, the approach that you mentioned is one way of dealing with it, but you know, there are some other methods and that would be to just try and stop the moisture getting into the DMSO in the first place or even when it's probably a talking point for a future podcast, using a method to take that moisture back out of the DMSO.

Toby Winchester

So, you're recovering the water and hopefully recovering your small molecules back into solution.

Stuart Naylor

Yeah

Toby Winchester

That'd be that'd be good topic. Yes.

I suppose we've talked about the freeze thaw cycles may be affecting the solubility further, but also, you know, the chemical structure of your small molecule, which can be quite complex at times and maybe not the most stable molecule because it's still a research sample.

Is there also decomposition of the small molecule caused by the fact that non ideal DMSO solute and freeze thaw cycles affecting the chemistry?

Stuart Naylor

That absolutely happens. I mean, there are unfortunately some compounds that will break down no matter what you do given enough time.

But yes, the effect that you just mentioned is very real. And another one to worry about.

So, I guess that's, you know, a fourth issue to consider.

Toby Winchester

Yes. So basically DMSO and water, you don't mix until you want to mix it.

So I suppose in your cell assay, you do want to mix the DMSO with water, but that's at the final stage of the process, yeah, rather than at the beginning stage of the process.

So, I suppose yeah, different ways of, well, I'm just thinking one of the other systems that I think has this effect of DMSO is, you know, the world of discovery has been, especially in the assay world, has been taken over by acoustic dispensers by a small company called Beckman and they didn't used to be Beckman, but anyway, there's another story.

But the liquid class, if you like, of a 100% DMSO acoustically sample is different from a 70% DMSO sample. So, you know, your amounts being transferred, you know, can change as well depending on how much hydration of the DMSO you've had.

Stuart Naylor

Yeah. I mean, I remember when the Echo first came along, sold by Labcyte, as it then was, and it was just a revolutionary piece of equipment. You know, the sales of it just really accelerated off the bat. I don't know how many are out there now in use, but it's got to be in the thousands, and you know, it allowed companies to hugely minimise what they were doing, shrink their library size amounts of labware that they were using, reformat down into higher density formats.

But it also had this feature, which was completely new, where you could accurately survey your compound collections in your storage plates and be told what the difference was between the DMSO and the moisture content.

Toby Winchester

Yes, yeah.

Stuart Naylor

Whilst before that, you know, it was certainly in the bigger pharma, widely known that moisture absorption was bad, it made that information available to a much broader audience.

As you say though, in terms of moisture absorption being bad, as that occurs in Echo plates, it can start to throw that dispensing off a bit.

And not with the very latest generation of Echo, but all the legacy Echoes are out there, of which there are many.

Once you drop below that 70% DMSO purity level, the dispenser will actually stop firing.

Which is a another huge reason to try and keep moisture out of your plates.

Toby Winchester

Yes. And I think, you know, labs like Beckman have realised this problem because of course, you know, even if you are keeping your samples in a dry store somewhere. And I'm sure we'll come onto automation systems and manual systems soon, but their cabinets off their latest systems are built to have dry air blown through them, so even when the plate is exposed, it's kept in a dry environment or a better dry environment.

Maybe not perfect, but it's, you know, they're trying to minimise that moisture uptake in their systems as well.

Stuart Naylor

Yeah, well, and this is a game of trying to do whatever, you can where you can.

I mean, if you assume for one moment that you've got your plate stored in the perfect environment and we can maybe come on to what that might look like as we progress in this podcast, at some point, you've got to take those samples out to use them.

Toby Winchester

Yes, yes.

Stuart Naylor

So it follows no matter what you do, over time you are going to have some level of progressive moisture absorption.

Toby Winchester

I suppose, does it make a difference where you are in the world as well? Cause some, you know, some places are more moist than others.

Stuart Naylor

I mean, we've got one customer in particular in Helsinki in Finland. And in the middle of winter when it gets absolutely freezing out there, it just sucks all the moisture out of the air.

Toby Winchester

Right, okay.

Stuart Naylor

And they really don't have much problem at all with moisture absorption, it occurs, but it's a heavily reduced rate.

Whereas when they go to summer and things warm up and you know, humidity levels, ambient can be 50% plus.

Actually they see some quite aggressive moisture absorption.

Another customer that I remember, I guess I can mention their name because they don't exist as that anymore, but in years gone by there was a company called Tripos in Tucson, Arizona, which is a very arid, dry environment, and again was they did see some moisture absorption. It was far, far lower than most other areas in the United States.

Toby Winchester

Right. So, it's almost, you know, if you're if you're a Scandinavian company, then you need to do all your small molecule assays over winter and not during the summer.

And that's why they have such long summer holidays.

Stuart Naylor

And then another thing to think about is really wherever you are in the world, if your building has HVAC managing the environment, you know, so that's full environmental control.

Most HVAC systems are set so that they'll have a 50 to 60% relative humidity level.

And that's for human being comfort, so you know, so your lips don't start cracking or things like that if the air is way, way dry.

But the downside of that in a lab is that's a humidity level where moisture absorption will just happen with gay abandon.

Toby Winchester

So that they're almost pumping in moisture.

Stuart Naylor

Yeah. Yeah. Quite literally.

Toby Winchester

Yes. There's a thought for people designing labs in the future.

I suppose we've talked a little about the Echo measuring hydration.

Are there any other, I mean, I suppose there must have been classical ways other than you put in your DMSO beaker on a weighing balance.

But you know, are there other ways to measure without having to buy a fairly expensive…

Stuart Naylor

If you look way back into the past, I'm not so sure, and probably some of that is to do with when I came into the industry.

But in more recent times, another method was this tube dowser that Ziath launched.

Albeit that's only for tubes.

You know, if you're wanting to look at what's going on in a plate, then, you know, really it's the Echo or nothing else.

Toby Winchester

Yes. Yeah. OK, cool.

So, I suppose there, you know, the obvious thing to do is make sure you haven't got any moisture where you're storing your samples.

And I suppose in things like, you know, you talked about the SPT comPOUND stores and the, you know, Hamilton devices, et cetera, they all use compressed gases, I would say, compressed air is what you would expect it to be being used, but I suppose there's a push not to use that within…

Stuart Naylor

Yeah, I mean you've mentioned a couple of options there.

There's a another one, a CRO that I know that's actually using a Kardex system for storage.

You know, so they're picking manually.

And in that case they're just flooding the room with dehumidified air.

Toby Winchester

Right, okay.

Stuart Naylor

So, you know, not as good as inner environments, which in most cases are nitrogen. You can also go to the dehumidified air route.

Some people that come and talk to us will sometimes ask about argon.

That is another option.

Toby Winchester

Bit more expensive, I would have thought.

Stuart Naylor

Well, but honestly, I wouldn't advise it. I mean, yes, there's the expense of the gas, hugely more expensive than nitrogen, but actually of more concern than that is it's a asphyxiation risk.

You know, it’s a gas that will, you know, if I had a pipe here and released a load down onto the floor next to where I'm sitting, it would just stay there.

You know, it clears incredibly slowly.

And so certainly here with what we do, we view it as somewhat of a dangerous gas, you know, particularly if you're needing to shift a further amount through a storage system to get the conditions that you want.

And we've actually refused to sell systems to customers that want to use argon.

Toby Winchester

Right, okay, very wise.

Stuart Naylor

It’s for that safety reason.

Toby Winchester

Yes. Yeah. Once you open the big door, then you're suddenly flooding your lab with argon, aren’t you?

Yes, Yeah.

Not ideal.

So, yeah, I suppose, you know, flooding an automated store or even a manual store with dry air. And then I suppose we need to come onto your product.

You’re, I suppose, you're not at the auto store area and you're much more in the in the manual zone, but that's where an awful lot of small molecules are still stored even within the big pharma world.

I'll let you do a little bit of sales pitch, but I'm more interested in why does it work?

You know, what’s the tech route?

Stuart Naylor

So, we make products called storage pods. This is a small one here.

This lid can be removed. Now go let me under these latches.

So, there is a draw in here in which you can place plates or tube racks.

Toby Winchester

Okay, yeah.

Stuart Naylor

And then, you can seal this shut, you can see a couple of fittings here on the front, and we'll then flush out this environment with nitrogen.

We've got two different what we call controller options. So, they're like the bench top box that controls the gas flushing.

We've got one controller that will flush a single storage pod.

You disconnect from it and then you can flush the next storage pod and the next one and the next one.

And then we have a much more advanced product called MultiPod, that can do all this small storage pod flushing, but it can also be permanently connected to up to twenty big storage pods that will hold like 250 plates each and continually monitor the environments.

Toby Winchester

And so is important to keep changing the nitrogen within the pod? I mean, once a day?

Stuart Naylor

I’ll come onto that, so obviously what we're doing is creating a, you know, a sealed environment.

And we were talking about risks of moisture ingressing through plastics. This is a metal construction, you know, so once it's sealed shut, nothing's getting in, nothing's getting out.

And so with the smaller storage pods, over time, what we've learned is you just need to flush it and then you can leave it.

We've got customers doing that for extended periods of time, like over a year plus.

Toby Winchester

So that's very different from, you know, going back to the automated store environment, which is just flushing the environment all the time and using nitrogen all the time.

Stuart Naylor

You've got be careful with that because even DMSO can evaporate if the conditions are dry enough.

So, there's a 40% difference in what's called the vapour pressure between DMSO and water.

Toby Winchester

Right.

Stuart Naylor

And that's the point at which things want to evaporate.

Conveniently, it's in favour of water wanting to evaporate first.

So water’s forty more times willing to evaporate than DMSO, but if conditions are dry enough and we've seen this with our MultiPod product where our customers totally overcooked the settings, you can get DMSO to evaporate as well.

Toby Winchester

OK, so you do actually have to be lenient with your flushing.

Stuart Naylor

Well, you need to, I think, with any storage system that uses inert gas.

If the system allows, you need to be sensible with the environmental settings.

Toby Winchester

Right, okay, that's interesting. I did not know that.

Stuart Naylor

Yeah, with the MultiPod product that I mentioned where it's continually monitoring these bigger storage pods and topping it up, there, the situation is slightly different and rather than going down to a really low level of oxygen and moisture content, which is what we do with these smaller pods, we'll run it at a slightly higher level. What we've found empirically, it's dry enough that you're really keeping moisture away from the samples.

And if there is some moisture in the samples, very gently over time, the system will start taking it out, but not so dry that you're getting into the realms of risking evaporating DMSO.

Toby Winchester

Right, okay, cool.

Wow. Well, I've learned a lot today. So is there any wrap ups you want to mention?

Stuart Naylor

Actually, there is one other thing that I thought of because it was sitting on top of the tube rack that I picked up to show our storage pod.

One other thing that's out there that can help with reducing moisture absorption is the Labcyte or Beckman microcline lid.

I've got one here that a customer gave me.

It's, you know, if you can see in the camera, but it's got this layer of foam inside. And it's designed, let me get my plate back…

Designed to sit on top of a microplate and this can provide a somewhat short term barrier to moisture absorption into your plate.

So, the idea here is that you add a certain amount of DMSO into the lid, which gets soaked up into this absorbent foam.

And if the plates exposed, so it's going through robotic system where you've got lids on top of all of your plates, the DMSO in the lid will absorb moisture first.

Toby Winchester

Right, protecting your samples.

Stuart Naylor

It will in effect, delay the time to the samples in the plates starting to get exposed to moisture absorption.

It's really simple idea, but it does have a positive effect, you know, and again, coming back to the comment I made, this game is about doing what you can, where you can, to reduce that moisture absorption.

This is quite a handy step if you're doing plate processing.

Toby Winchester

OK, Thank you.

Yeah, a plug for Beckman as well. Okay.

Stuart Naylor

I’ll invoice them at the end of the month too.

Toby Winchester

Yeah. So yeah, thank you very much for speaking.

I think we are probably going do a follow up podcast when some new data is coming out with one of your partners, I believe, so I look forward to that as well.

But Stuart, thank you very much for joining us.

Stuart Naylor

No worries. A pleasure. Thank you. Look forward to talking next time.

Toby Winchester

Yes, indeed. Cheers. Thank you very much. Thank you everybody for listening in.

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Voices of Sample Management

Episode 7: The Impact of Hydration on DMSO Sample Integrity