Get yourself a little assay…..

So what is one of those?

One of the things I do (for my sins) is run a stability testing service out of a laboratory in the city.  The testing is quite routine really – just prepare the samples in final packaging and in glassware, place them into their appropriate conditions – oven, room temperature, fridge etc then watch, periodically getting them out, weighing and measuring then putting them back.  Changes do sometimes happen and when they are dramatic such as a huge shift in pH or a collapse of an emulsion everyone agrees that there is a problem, but when something subtly changes it can take a bit more sciencing to work out if the changes are likely to wreck your efficacy or not.  That’s where assays come in.

An assay is the name for an analytical test that is carried out to establish if a chemical is present in a formula or not.  Sometimes the assay can only qualitatively account for your active – tell you if it is present or not – while in other cases it can quantitatively measure its presence too, informing you of how much is still present. This can be very helpful when you are making claims that revolve around a specific active – the product is unlikely to work if said active has disappeared somewhere or changed its chemistry.  I usually request assays be carried out on products that rely on, or feature a particular active in the product name or (of course) in products that have to be assayed such as TGA products or some APVMA formulations.   For cosmetics we are mostly assaying for vitamin A, C and/or E as well as some antioxidants such as Ubiquinone (Coenzyme Q10) or Epigallocatechin Gallate (EGCG) from Green Tea.

Assaying for vitamin activity is usually relatively simple and regularly done by analytical test labs, mainly because these things frequently turn up in food and pharmaceuticals and they are both industries that require formula validation via analytical measures.  However, in cosmetics we don’t always use the vitamins in their simple form.  For example, vitamin C is not always added as Ascorbic Acid, sometimes it is Sodium Ascorbyl Phosphate and sometimes it might be Ascorbyl Glucoside.  I have tried to get products assayed for different types of vitamin C and failed. Maybe it is my fault for not asking hard enough but in my experience, most labs testing for vitamin C will assay against Ascorbic Acid so if your vitamin C is in another form it won’t show up. This can be an issue.  This is true for other vitamins too,  there is more than one form of Vitamin A used in cosmetic science.  Understanding what can and can’t be tested for is key to a successful assay.  I’ve had a case once where we wanted to analyse for the presence of retinaldehyde, a very active form of Vitamin A.  I thought this would be simple but it turned out that this molecule is so very unstable (and expensive to buy to boot) that a ‘good’ (as in accurate) result was not easy at all to achieve.

Vitamins and Antioxidants are both generally good candidates for assaying as they are usually put into a cosmetic product as part of the products ‘actives’ and therefore should be present in the formula throughout its shelf life but also these things are quite likely to break down over time.  Antioxidants are natures little soldiers, fighting free-radical pollution  on your skin and in your product.  Just like regular human soldiers these things get worn out and need a break, sometimes they even expire – you can’t go on soldiering for ever, no matter how much you want to.  As such, assaying these things at the end of a shelf life test using the aged 40C sample VS the room temperature sample or the spec (if you already have that established) is a good idea.  So  that’s what I do.

When assays go wrong.

I’ve already mentioned that assays are chemical specific in as much as you set up the test to ‘find’ the exact thing you are looking for.  So if you want to assay for vitamin C you set the machine up to look for Ascorbic Acid and nothing else.  What I haven’t mentioned yet though is the issue that is often faced when a product formula is complicated.  IT is all well and good when there is one distinct chemical to assay in a noiseless base but when you have lots of actives, some of which might look similar on the graphical trace (if you use a GCMS test) you can end up with difficulties in reading your results.  Quite literally the trace can look like peak hour on the main motorway out of the city – chaotic!

Another issue that can be faced with assays is their sensitivity.  Getting a good trace depends on the sensitivity of the machine and how it has been set, so that you get a good, clear separation of the peaks.  If you have a lot of actives at very low levels you might not get a good enough peak to quantify, or the peaks might differ slightly every time you run the trace – analytical labs might repeat the assay 2-3 times to get an average result. This is quite a normal way to approach science.

When they work well.

When things go well you can end up with a fully validated stability study that proves, through the weekly oven/ fridge/ RT samples that your formula is physically stable and proves through the analytical chemistry testing that the product is chemically stable also. You can use your assay results as evidence to back up product efficacy claims, at least to a point, and the results can also help you quantify improvements in stability between successive versions of a formula.  This is one thing I’ve been doing with my high strength ascorbic acid formula – assaying every month or so to gain a better understanding of how it stands up over time.

Assays can also work well when you want to validate the safety of your formula with regards to the potential for it to cause irritation. This is especially so when you have a lot of vegetable oils that could turn rancid and increase their acid values (free oleic acid) or where you have a high level of essential oils that can degrade to release by-products that are more likely to cause reactions.  Assaying to get an idea of the chemical going’s on in your formula can save money on human trials and can also help you hone in on what is and isn’t working in your formula or what is the root cause of instability (as oxidative changes usually have a cascading effect through a formula, once one thing changes, it affects other things too).

Assays can also work well when you’ve extracted or created something – say a vegetable or essential oil or a herbal extract – and want to know if you have captured the ‘essence’ of it.  The assay might be set to look for one particular ‘active’ or might give you a general finger print of the thing.  Your test lab can usually help you choose the right test.  I assay the essential oils that I distill, just to see if they meet the standard for that particular plant.  I also used the assay results to help me identify the type of Eucalyptus I had gathered.  It’s awesome!

But can’t you simply see/ smell or feel when a product has gone bad?

Sure, you can see when something has gone critically bad but it is much harder to just see (or sense) when something is slowly degrading, especially (as with many cosmetics) the product formula is quite complex – it could be any one of a number of things that are causing the visual change for example, some of which are not that critical to the key performance of a product such as chlorophyl bleaching in a cream due to UV.

So what can we assay for?

Quite a few things but my most common tests are:

• Acid Value of a vegetable oil containing formula.
• Presence and quantity of named actives.
• Stability of essential oils (looking for presence of oxidative by-products and/or quantitative analysis of whole oil)
• Peroxide Value of vegetable oil formulations

And who does these tests?

As much as I’d love to have my own analytical lab I don’t so I use established and NATA accredited facilities.  Just google ‘cosmetic analytical laboratories’ and I’m sure around 5-6 will pop up – there are a few.  On top of that I use a government laboratory for oil quality analysis so when I want to assess the quality or shelf-life changes of a vegetable oil blend or neat oil and I also refer lots of people to the Southern Cross University here in Australia for essential oil assays including oil characterisation (if you want to identify an oil) and herbal extract analysis.

What does it cost?

It really depends on how the assay is carried out (what equipment), how expensive the reagent is, how long the assay takes and how many samples you tend to submit but the tests I commission usually cost between $45 – $250 each so not always cheap but not prohibitively expensive.

And when should I assay my formulations?

Well, that depends. Some brands will go for ever without running a single assay while others will send products off all the time.  I am tending to rely on them more and more as I find the crisp, non-disputable data gleaned form an analytical test very helpful and re-assuring, especially for brands that are making ingredient claims for natural formulations and want these formulations to have a shelf life of 30 months.

If in doubt just ask.

and if you are a complete nerd like me then assay away – it’s like having a little chemical spy peep into your formula and measure stuff for you and that, my friends, is awesome.

So that’s the intel on assays.

Have fun in the lab 🙂

Amanda

PS: I did my honours degree project in analytical chemistry way back when I had no idea about cosmetic science.  I have always enjoyed the investigative part of science but I’m not really neat enough to do this type of analysis for my full-time job.  I spill too much!