Home Cosmetic Science Talk General Cosmetic Microbiologist Here. I Can Help Answer Your Germ Questions

  • mikebavington

    Member
    February 15, 2014 at 5:47 pm

    What is naticide composed of? The INCI just says: Fragrance or Parfum.

  • labtechnician

    Member
    February 16, 2014 at 7:22 pm

    @The_Microbiologist Thanks a lot for your kind help. yes this preservative efficacy test was performed according to British Pharmacopoeia (5.1.3)  method. I have asked lab for data regarding separate controls and will email you asap. thanks once again.

    @mikebavington.. Naticide is a blend of natural extracts formulated by SINGERA research. company says it is a fragrance that inhibits bacterial growth in cosmetic formulations. it is a good natural preservative

  • Anonymous

    Guest
    February 18, 2014 at 11:56 am

    Hi Ben,

    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!

  • the_microbiologist

    Member
    February 18, 2014 at 9:45 pm

    Hi Compounder,

    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.

  • Anonymous

    Guest
    June 4, 2014 at 9:42 am

    Hi Ben, @The_Microbiologist

    I’m a fairly new microbiologist in cosmetics and have many questions that range from basic to philosophical in microbiology since I have started. I could talk about microbiology all day and the exciting bacterial growths I have come across but before I go on a tangent I have some quick questions of concern.
    First, I have had some growth of bacteria and it appears 100% of the time. I mean that i test the product 5 out of 5 times or sometimes 10 out of 10 and I get growth for this particular product “A”. Also I have lets say product “B” which I have also had the same occurrence. When I send it out to 3rd party labs they say its clean. <10cfu/g. There was one instance they found more than 1,000 cfu in product A but the 2nd time around they found nothing.
    Hypothesis:
    - Test are in USP methods and I use BD Letheen broth that has Licithin and Polysorbate 80 to neutralize preservatives so it says. From what i heard 3rd party labs are asked to do USP methods as well and probably use phosphate buffer as their broth. 
    Would this be a problem?  I have to probably reread USP <61> and see if neutralizing preservatives is just recommended and not mandatory. 
    Another idea i have is probably sample size used in testing. I use 10g to 90mL. 
    I have many more questions but i have run out of time for now. I hope you can help me and share your experience so I can be enlightened. I’ll greatly appreciate it. 
    Thank you,
    Josh.
  • the_microbiologist

    Member
    June 4, 2014 at 11:32 am

    Welcome to the forum!

    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.

  • Anonymous

    Guest
    June 5, 2014 at 12:30 am
    Thanks for the greetings and reply. 
    I would assume they are the same but i might have to send it out to get ID. Its a gram positive rod and this product A is a powdery/grainy like dry product. I have diluted the sample 1/10 and 1/100 dilution and then spread a 1mL on to a TSA plate. 
    The 1/10 and 1/100 TSA plates looked the similar with confluent growth. I’ll take pictures when I can. 
    My results:
    TSA: TNTC and Gram stain it http://imgur.com/a/i5pzf (link has pictures of all plates and gram stain)
    MSA plate: 2 plates (from TSA) and there was yellow colonies one plate. 2nd plate yellow colonies but my streak looked like mucous and the colonies were gelatinous goo. 

    It is a great mystery that 3 other labs were unable to find growth. I do about 60 different samples a day and when product A shows up I come across the confluent growth again. I’m sort of out of ideas how to explain this. I did an side experiment with just using 90mL of sterile water and performed a 1/10 dilution. I still got growth on my TSA plate. HA. 

    Contaminants:
    I do all my work under Biosafety cabinet class 2 A2 and have positive and negative controls during my testing. It can be from environment since opening the drum causes white powdery mist that can be felt when breathing. Maybe some spores? 
    Thanks again.


     
     
  • the_microbiologist

    Member
    June 5, 2014 at 5:31 pm

    Wow, the pictures really help!  What you are considering to be “confluent growth” I think is probably more along the lines of a “spreader,” meaning a single or a few colonies that rapidly take over the plate either by rapid multiplication (which gram+ rods are great at) or other means.  The way you can check this is to look at the plate after just 12 or 18 hours incubation, then again at the end of the incubation period.  My guess is that you will have one or a few colonies only to begin with, then a couple of days later I imagine those colonies will have spread over the entire plate.  Do note the zones that on the plates in question that do not show growth.  If the product was very contaminated and the spread plating covered that part of the plate, I would expect growth there.

    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.

  • Anonymous

    Guest
    June 6, 2014 at 12:24 pm

    Yeah about the spreading… that might fall on my part since I am rushed to do some of the testing. X_X. I’ll see about retesting it and doing a better job at spreading it. Then i’ll check on it within time intervals. I was also going to try the pour plate method and see if that stops it from spreading.

     “Confluent growth” is the term that was coined for growth that spreads over the plate. I am new to this spreader type of bacteria definition. I know of Proteus likes to swarm over the plate so would that also be considered a spreader? 
    It’s good to know that gram + rods are great at being spreaders. thank you. 
    If it is just a few colonies causing this spreading action would it be safe to assume my 1/100 dilution results be greater than 100cfu/g ? 
    I was curious about my MSA results that it managed to have yellow colonies and ferment the mannitol. I am beginning to see a trend that selective plate results have interesting results. I had EMB agar once with a contamination of Burkholderia cepacia and it had the metallic green sheen appearance similar appearance of E. coli  but I went back and forth plating with centrimide agar and EMB agar to confirm it wasn’t. http://imgur.com/a/HTsnL  .I couldn’t find anything about B. cepacia fermenting lactose on EMB. Sometimes i get jungle of multiple organisms on plate. 
    If possible one more quick questions. In testing cosmetics would the type of neutralizing broth (D/E or LB) matter for testing yeast and mold? or would extra things need to be added to the broth? I assume that as long as its plated on a SDA plate it would be fine to grow any possible yeast or mold. 
    Thank you Ben, I greatly appreciate you helping me! :)
  • Eli

    Member
    June 16, 2014 at 2:09 pm

    Hello Ben, I have maybe two very stupid questions but … Do saturation of germs can cause lost of transmittance in a water based solution?

    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?

  • nasrins

    Member
    June 17, 2014 at 3:01 am

    Hi ben

    whats different between parabens and isoiazlinon preservatives? are they longevity different?

  • the_microbiologist

    Member
    June 18, 2014 at 10:23 am

    Hi Eli,

    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).

  • Zoe

    Member
    June 19, 2014 at 10:23 am

    Hi Ben 

    I am looking for some advice about effective preservatives to use with face masks containing Clay and in facial exfoliators when using exfoliating grains (such as bamboo) in a cream (water& oil) based formula. In the past I have used Phenoxyethanol and Ethylhexyglycerine, but I have since found out that these aren’t effective enough against the Gram+ bacteria when going though microbial testing. Is there something I could add to assist this preservative? Ideally I would like to go down the natural route and have looked at using Dermosoft 1388 eco along with GMCY, do you think this would be effective? But equally I am not opposed to using other preservatives. I have a small scale uk based skincare company but not from a scientific background, so learning all the time! I would love to hear your thoughts…..many thanks Zoe
  • MakingSkincare

    Member
    June 19, 2014 at 10:56 am
    Zoe, no-one can tell you whether a preservative will be effective or not.  You will need a lab test for that.

    What I can say is that the preservatives you mention (Phenoxyethanol and Ethylhexyglycerin and Dermosoft 1388 eco/GMCY), being the more “natural” type preservatives, will prove a real challenge to preserve the clay/bamboo.  Even if the clay has been irridiated, clays are problematic because they adsorb spores onto them and the large surface area makes it difficult for your preservative to cover everything.  Clays also tend to deactivate formaldehyde donor preservatives.

    Do bear in mind that there’s a lot more to preservation than just adding a preservative eg reducing water activity, using packaging which minimises contamination in use, microbiological testing of raw ingredients and process water, etc and adding 0.1% disodium EDTA (heated water phase) and glycols will help. Another major factor is the amount of “bug food” in the formula - this often gives overlooked.  I see many people putting tea, goat’s milk, honey, hydrosols, floral waters, aloe vera, extracts, protein, powders, starches etc in their formulas which will really challenge the preservative system - reduce these to a tiny % (eg 0.1%).

    More info including reviews of 27 preservatives - http://makingskincare.com/preservatives/
  • Eli

    Member
    June 30, 2014 at 1:24 pm

    Hello Ben,

    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 :( but it is nice to know that there might be some type of microbe that can “come back to life” somehow… Thanks again. !

  • the_microbiologist

    Member
    July 1, 2014 at 6:56 pm

    Hi Eli,

    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

  • botanicalsecrets

    Member
    July 11, 2014 at 3:11 am

    Hello,

    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

  • microformulation

    Member
    July 11, 2014 at 11:51 am

    Dear 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.

  • Eli

    Member
    November 27, 2014 at 4:09 am

    Hello everybody! I was wondering some things about microbiology because i went to a cosmetic event yesterday where one of the speakers said: Maybe in the future cosmetics will be treated as food, and they will be store in the fridge ! and that phrase keeps moving in my mind .. Is it really possible? maybe it is actually a good idea but too expensive to create the chain of cold.. from supermarkets and distributors etc.. 

    My question is, does every cosmetic form need a preservative system ? like powders (dry shampoo), or oils (i guess oils can also develop microbes). Would it be useful to use the fridge once we open a cosmetic product and start to use it ?
    Thanks!
  • bobzchemist

    Member
    November 28, 2014 at 2:14 am

    Anything that is used on skin needs a preservative system, just to be safe. The stupidity of consumers should never be underestimated (spit in mascara? Really?).

    I would say that the only way to make a completely safe cosmetic without preservation would be to make it sterile, and then package it aseptically in refrigerated single-use containers - but guaranteed, someone would try to stretch it out, and put a half-empty open container back into the refrigerator.
    The difference between food and cosmetics is that the human digestive system is designed to cope with a wide variety of micro-organisms, so food can be a bit contaminated, and still safe to eat - but the human eye is not designed that way, and a little contamination can be disastrous.
    Then, we must also consider legal liability and consumer lawsuits - if a consumer claims injury from a product, and it tests positive for bacteria? The company that made it, and everyone who had a hand in formulation and QC, may be in big trouble.
  • Eli

    Member
    November 28, 2014 at 3:13 am

    Thank you very much for the answer Bobzchemist, pointing out the exact difference btw food and cosmetics made me realize it is a serious thing when we talk about safety in cosmetics.. you are right ! The eye doesn’t have defense systems like the digestive track.. great example!

    About lawsuits it is still a mystery how a consumer can prove the bacteria comes from the use of that specific cosmetic? If consumer is always right, then big companies must be scare about them. .. 
  • Microinjala

    Member
    December 2, 2014 at 2:02 am

    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!

  • braveheart

    Member
    December 31, 2014 at 3:53 pm

    Really really fantastic thread!

    Rich information from all the experts.
    Thank y’all.
  • oldperry

    Member
    April 15, 2015 at 2:37 pm

    This really was an excellent thread.  I’ll have to seek other experts to do a Q&A

  • mart

    Member
    September 8, 2015 at 2:21 am

    I have a question for the microbiologist. I am working with a product that has benzyl alcohol at 0.08% . Is this enough to preserve it? The PH is 4 and it is about 50%.water I am debating on adding something extra but dont want to if not needed. Maybe PG ? What would you recommend ? Parabens are unacceptable. I can drop the PH , slightly. Thank You.

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