Cosmetic Microbiologist Here. I Can Help Answer Your Germ Questions
The_Microbiologist
Member
Hi Everybody,
My name is Ben Tanner and I am the microbiologist in charge of Cosmetic Test Labs in Texas. My lab specializes in cosmetic microbiology, such as preservative challenge tests and aerobic plate counts.
I was on the old forum for some time as "CosmeticTestLabs." Anyway, I think there is a great group of people on this forum and so I like to share my microbiological expertise when I can. Feel free to ask me any germ/preservation/contamination/infection/test questions you might have. I usually check in every few days.
Happy formulating and testing!
- Ben
My name is Ben Tanner and I am the microbiologist in charge of Cosmetic Test Labs in Texas. My lab specializes in cosmetic microbiology, such as preservative challenge tests and aerobic plate counts.
I was on the old forum for some time as "CosmeticTestLabs." Anyway, I think there is a great group of people on this forum and so I like to share my microbiological expertise when I can. Feel free to ask me any germ/preservation/contamination/infection/test questions you might have. I usually check in every few days.
Happy formulating and testing!
- Ben
For testing help visit us at Cosmetic Test Labs
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http://www.americanpharmaceuticalreview.com/Featured-Articles/38885-Antimicrobial-Preservatives-Part-Two-Choosing-a-Preservative/ specifically.....
Benzyl alcohol and phenoxyethanol should not be formulated with non-ionic surfactants eg polysorbate 80. They will interact and such interactions may not involve conventional chemical transformation, but concern more subtle phenomena e.g. hydrogen bonding and complex formation. Thus the overall level of preservative in the product may not change, but unless the preservative is available in the “free” form its efficacy may be compromised.
The article also states:-
EDTA can interact with benzoic acid ....and..
Adsorption onto excipients, especially those with large surface areas or on to container / closure systems can also remove preservative(s) from solution for example
- benzoic acid by kaolin
- phenoxyethanol by pvc plastic, cellulose based excipients
- sorbic acid/sorbates by plastics (polyproylene, PVC and polyethylene)
The antimicrobial efficacy of phenoxyethanol is reduced in the presence of the cellulosic suspending agents such as HPMC.
I'm wondering what your experiences are with the above and whether you agree with points raised in the article?
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MakingSkinCare - That article is a gem. I looked over it briefly and will read it in full later. In the meantime, I can confirm that polysorbate 80 does neutralize certain classes of preservatives. In fact, we use it in our all-purpose neutralizing broth for that very purpose in the lab. Here is the product insert and formula if you are interested. In that document (and in lots of scientific literature), it says the purpose of polysorbate 80 is to neutralize substituted phenolics. I don't think phenoxyethanol is technically a substituted phenolic (it would seem to me to be more like a monophenyl ether), but it is structurally similar to many substituted phenolics so the two chemicals probably should not be present in the same cosmetic product if the goal is to make it microbiologically stable.
I am not as familiar with adsorption of preservatives onto plastics, though I know some long-time formulators who consider the plastic type of their packaging to be effectively one more formulation variable.
As for the cellulosics, I would anticipate some interference with preservative effectiveness, but in a minor way. We see this with almost all thickening agents. Maybe the best example comes from a contact lens solution maker several years ago. I don't have all the information, but I think that basically what happened in that instance is they added a thickening agent to their lens solution to aid in eye feel that bound up some portion of their preservative, thereby knocking out some efficacy and allowing some pathogenic fungi to gain a foothold.
Most antimicrobial agents carry a positive charge - and this is just a broad theory of mine - which physically leads them to the negatively charged cell wall, where the rest of the molecule does the destruction. Since cellulose is a huge molecule with lots of charge variation over its surface, it would not surprise me to see it bind up some preservatives. However I would not expect them to be too tightly bound so the effect may not be a formula-killer.
milliachemist - some non-ionic surfactants do interfere with antimicrobial efficacy but usually in a minor way and the effect can be expected to vary depending on other aspects of the formulation such as pH. I don't have a list in my head of which ones should be avoided, but I have definitely seen instances where removal, substitution, or scale-down of a non-ionic surfactant improves antimicrobial activity.
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Perhaps I can kill two birds with one stone with this statement about "natural" preservatives (including Leucidal):
Can a cosmetic formulation be microbiologically stabilized using one or more "natural" preservatives like Leucidal, sorbic acid, and lactic acid? Yes. Is it easy? No! Is it cheap? No! Will it always work with "tough" formulations? No!
And about grapefruit seed extract and vitamin E, I have never been wowed by them. That's not to say they don't work, just that I don't recall ever seeing it happen, and I vaguely recall some scandal where grapefruit seed extract appeared to have been spiked with quaternary ammonium compounds.
We get a lot of samples through the lab for preservative challenge testing, many from customers we haven't worked with before who are submitting formulations to us that are as new to them as they are to us. When they use traditional preservatives (even retailer-preferable traditional preservatives like phenoxyethanol), their success rate on challenge tests is very high. I estimate they fail only one in every 5 or 10 tests. That's great, because challenge tests take a long time to conduct and are relatively spendy.
On the other hand, when they use natural preservatives it often takes a few rounds of testing and reformulation for them to pass. That's good for the lab but bad for them, and my heart goes out to them when I see them give up altogether rather than throw in a tiny amount of synthetic preservative and go to market
So have I seen Leucidal work, yes. Do I think it is as efficacious as most of the traditional preservatives, no. But on the other hand, many traditional preservatives are being unfairly vilified. So unfair or not, there's a great big and trendy marketing trade-off to be considered.
Frequently we see customers use antifungal preservatives such as sorbic acid in combination with Leucidal. That seems to be a pretty good place to start and I'm sure the folks at Active Micro Technologies (makers of Luecidal) could give some other suggestions, too.
MakingSkinCare: As for polysorbate 60, I doubt it interferes but am not certain. As for the proposed formulation 1% GC/D GMCY and p-Anisic/Dermostoft, I just don't know. I am pretty sure that your formulation expertise - even as related to preservation - exceeds mine! I am a "test it and let's see" sort of guy, which I suppose is a good fit for a testing lab owner
Tasiaashton: Thanks for giving us a shot!
Thanks all for the great discussions.
Robert Zonis was posting regarding Leucidal before the site crashed. Through him and several other sources I heard rumblings regarding issues with Leucidal in the last quarter of 2013. Primarily the issues were with Yeast/mold coverage so Potassium sorbate is the way to address that.
Grapeseed Extract is an over hyped "preservative." I will not open that can of worms, but I would not recommend it to anyone.
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http://www.personalcaremagazine.com/Print.aspx?Story=4108
Chelating agents are able to enhance
the efficacy of most preservatives. This occurs as the chelator removes
metal ions from cell walls of the microbes. The weakened walls then
allow the biocide to penetrate and destroy the microorganisms. Although
the boosting effect of chelating agents on preservatives is well known,
the environmental fate of these materials has been debated. To avoid the
environmental discussion about chelating agents, readily biodegradable
alternatives have been introduced to the market.
I use a chelator with PE9010 each time to boost its efficacy. This need was passed onto me several years ago by Wolfgang Siegert at Schulke. It can be challenging as EDTA is a big no no in most "Natural" standards (NPA, NSG, WF's). In that case I will generally use Dermofeel PA-3 from Dr. Straetmans (Kinetik in the US).
The only micro tests typically done on finished product are aerobic plate counts for bacteria and fungi. Those tests tell a person how many bacteria and how many fungi are present in a given gram or milliliter of product, which in turn indicates whether or not the finished product meets a company's specifications for acceptable quality for release/sale. Less than 100 bacteria or fungi per gram or milliliter is a common specification, though less than 10 bacteria or fungi per milliliter or gram is more typical for eye-area cosmetics.
I haven't heard of that practice before...probably means I spend too much time in the lab
Anyhow, I am sure it's fine for the day or two. Then after that I'm pretty sure it would turn into germ soup unless as Bob says the pH is very high.
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Time Point S.aureurs P.aeruginosa C.albicans A.brasiliensis
CFU/g CFU/g CFU/g CFU/g
Inoculum 7.0 x 105 6.7 x 105 6.9 x 105 1.7 x 105
0 hour 6.8 x 105 6.1 x 105 6.1 x 105 1.9 x 105
7 days < 10 < 10 [NA] [NA]
14 days < 10 < 10 1.0 x 105 150
28 days < 10 < 10 < 10 < 10
What is your opinion regarding this result? How I could reduce the
survival of yest (C.albicans)? Your help will be really appreciated.
thanks
I notice the "n/a" for Candida and Aspergillus at day 7. I presume those organisms were just not analyzed at that timepoint. Is that correct? Was this by chance an ISO method?
Nevertheless this appears to be a well-preserved formula in general. I see the same "odd" data that you do with regard to Candida. It's strange for that organism to have what we would call a "shoulder" die-off curve, meaning slight reduction followed by precipitous reduction. Usually Candida reduction is much more linear, meaning you'll loose a log (90%) or so each week on the way down to a final result of non-detection. It's possible that the 14 day data was artificially increased by chance, by clumps of organism within the test material or something else.
If USP <51> criteria were applied it would "pass" so you are in good shape. If you want to be extra diligent, you could re-test just the Candida and see if next time you see a more linear (sort of more predictable and thus more dependable) die-off. If you wanted to be even more diligent, you could do that at a different lab. If two labs get the same result, it's almost certainly "real."
Here is my last note, please verify with your lab that they did separate controls to verify neutralization and recovery from the test sample. Ask them for the data for that, too. They should have it and it should be quantitative. If they don't, then they're not doing the test right and it calls into question much of the data. Feel free to email me privately if that comes up and I'll help you work through it and re-interpret the data.
I have a question. I have been making lotion for my family to use. I am currently preserving it with 2% leucicidal SF liquid and it seems to last 4 months. Do you have any experience with adding aspen bark for additional preservative coverage? I was reading about combining each at 2%.
Thanks!
I don't have much experience with aspen bark, sorry. I don't know how you are determining the 4 month shelf life but if it's based on aesthetics the lotion is probably microbially contaminated long before you can perceive it visually.
As a point of reference, one can put 100,000 bacteria into a milliliter of water and the water will appear to the naked eye to be crystal clear and usually won't smell bad, either. That all changes once you get to about 10,000,000 per milliliter. Most of the cosmetics we test in our lab that have counts ranging into the tens of thousands or millions of cells per milliliter have subtle or no aesthetic differences from sterile samples.
You may be determining shelf life with lab tests - I am just sharing for the benefit of the forum in general.
Hopefully someone with experience with aspen bark can chime in too.
Are the colonies that always appear on your plates the same type type, or do they vary? Do they "run with the dilutions" meaning that there are more contaminants in plates that got more neutralized product and fewer in plates that got less? If you haven't done a dilution series it's easy, just put 1/10 of what you would normally put onto a plate onto an agar plate and incubate, then compare with the ordinary plate.
If the contaminants "run with dilution" that's a good indication they're actually in the product (though it may also indicate they're in your neutralization medium) and you're just detecting them where other labs are missing them. There are all sorts of ways labs miss contaminants....no/poor neutralization of preservatives, too high of incubation temps, not long enough duration of incubation, etc.
If the colonies appear on every plate regardless of dilution and especially if they're morphologically varied, then you probably have some sort of lab contamination going on and the product is in fact clean. The contamination could be from air, technique in need of improvement, or even non-sterile plasticware marketed as sterile. It happens. Part of the art of microbiology is reducing the frequency with which such contaminants pop up and learning to tell environmental contaminants from product contaminants.
The other explanation is that the plates are too wet, so you have a situation where microorganisms grow in the fluid on the surface (as if growing in broth) and then that fluid gets spread around the plate during movement and seeds other parts of the plate.
There is no need to send anything out to get it ID'd at this point. For practical purposes if you can't tell them apart on the basis of colony morphology, you can consider them the same organism.
And another one ... If I dry a slurry full of germs (or leave a small amount of water) and leave it like that for several months... Adding water do they "wake up" again?
Hi ben
whats different between parabens and isoiazlinon preservatives? are they longevity different?
Yes, microbes will definitely cause loss of optical transmittance in a water based solution. In fact, we use spectrophotometers to quickly estimate microbial concentrations in broth cultures. Each little cell is sort of like a glass sphere when it comes to light, they cause the light to refract and go in all different directions, which means it doesn't wind up at the sensor (that is, it's not "transmitted")
To answer your second question, I would have to know which microbes you're using and specifically what you're doing (what temperature, humidity, vessel type, etc).
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Thank you so much for your answer. That solved a big problem I had. Relative to the type of microbe or details of production sadly I do not have access to that information
Microbes are notorious for propagating from seemingly out of nowhere...when you have liquid around nutrients, you can count on them showing up at the party.
In fact, they pop up so quickly and reliably that people literally used to think that they spontaneously generated (appeared from nothing).
I'm glad my comment was helpful.
Ben
I have been using the leucidal liquid and Phytocide Elderberry Extract at 2% and 0.5% respectively for my oil in water emulsions. This combo has been very effective though I am in a quandary for 2 reasons: 1. Needing to use this high amount means that the preservative costs is quite high, especially in relation to the rest of the formula. 2. Is it better to use a 'natural' preservative at this higher % or a synthetic preservative at a much smaller %?
I want to keep my customer base that want paraben free and I am looking at my options to reduce the costs and preservative. I would be happy for any information and feedback. I am wondering if I can use these at a lower rate with other combinations - potassium sorbate? phenoxyethanol?
I have just purchased Geogard Ultra to experiment with and am finding that it too needs at least 1.5% and am seeking information if I should combine it with something else.
I have tried Opiphen plus, but I wasn't happy with the end result.
Thank you,
Regards, Kathy
First and foremost I cringe whenever someone throws out "natural" vaguely. It isn't a defined term at heart and it needs to be qualified. If you are trying to meet your clients demand for "safer" products. I would investigate the Natural Standards (NSF, NPA, USDA), learn one, endorse it and then use it as a qualifier for your ingredients. This is simpler, it gives you third party validation and you can move onto what is more important, the performance of your product. If you leave it undefined you will either be too non-compliant (green washing) or too scared to use anything. Using the Internet/Google as your ingredient validation is dangerous. Nobody will agree and you can argue all day long. You can make most Cosmetic Chemists grumble by even mentioning the "paraben scare" and you can infuriate the super strict "natural good, synthetic bad" crowd by telling them about naturally occurring parabens. Endorse a standard and get out of the whole muck pile.
There are numerous preservative systems with long records of success and lower costs available and endorsed under the Natural standards. Their inclusion is generally not controversial and both camps (strict, mainstream industry) can generally agree.
That said, can I recommend any ONE preservative that you can buy, use in every product and buy exclusively? ABSOLUTELY NOT!!! When selecting a preservative the entire Formulation must be taken into account. If it is a surfactant system, will a preservative for a basic cream work? Not necessarily. I encourage you to take a look at the FormulaProtect site that Lonza has and run a Formula through their decision engine. In the end it spits out a Lonza preservative of course, but you will be able to see how many factors will come together to suggest a best preservative on a case by case basis.
Other factors contribute to the preservation strategy as well. Some packaging is more secure than others. For example a lotion pump would provide more protection than a jar. Final pH of the product has an effect. For the informed Formulator adding a glycol can augment the preservative system.
Sorry I didn't answer your question outright, but I think my point is that preservation takes some due diligence and research. Jane at Making Skincare (she can chime in) has some documents that will give you an overview of preservation. Again, it is not impossible to learn preservation but it does require more information than could be included efficiently in a single or even multiple discussions. I read 4 Journals a month plus multiple RSS Feeds and Professional blogs. I would estimate that at least 30% of that reading is in the area of preservation and new preservatives.
Lots of information. I hope I didn't discourage you or come off as pedantic. It is simply a topic that needs more than time and effort than a blog can deliver. As for wanting just ONE preservative for everything the best analogy I can give you is in food. I am letting you buy ONE condiment for every meal you cook. To paraphrase some arguments I have heard in preservation "I only want to pay for 1 condiment, paprika is too expensive, etc." You can have salt, but not pepper also, You might want Paprika but you can't get it retail. Any good chef would balk at that scenario. You can't limit yourself to one preservative as well as a universal preservation panacea. It doesn't work.
Hello everyone! I am new in microbiology and have a lot of question to ask, especially in PET test.
1. How to do a micro challenge test for the W/O cosmetics? I have read the summary of CTFA-6(through this link http://cosmetictestlabs.com/ctfa_m-6_summary.html), but it just mentioned one method as that:"reduced the inoculum volume or oil based carrier system", I want to know how to reduced the inoculum(centrifuge?)? And oil based carrier means I need to put the microorgnisim into oil? What kind of oil?
2. CTFA-6 also mentioned molds surface inoculation for W/O soild products, I want to konw how to inoculate and how do the judge? I think spray or inoculate with a disc of media may be practical method, but I do not know if it is right?
3. Do you have good research artical or report to reconmend?
Thank you so much!
Thank y'all.