PREPRINTS OF THE 1997 ANNUAL SCIENTIFIC SEMINAR 65 Recognizing these community and phenotypic mechanisms requires one to abandon the concept that bacteria exist in evenly distributed suspensions of single cells. Instead, organisms exist as biofilm communities or as aggregates and associations. Microorganisms particularly aggregate and form biofilms when exposed to biocides 2. 6. As a result, our reliance on lab methods using planktonic organisms at the worst overestimates the activity of the biocide or preservative in nature at best, they simply do not reflect or predict reality. In order to design a test that would more closely mimic reality, we should be using techniques employing cork•tant-depth film fermentors/Robbins devices to establish biofilms and confocal laser microscopy coupled with fluorescent physiological stains to visualize the biofilms •B, •9. 20. Such methods are routinely used with the study of plaque for the development of mouthrinses and dentifrices 2•. We should adapt such techniques to preservative efficacy tests (PETs). Preservative Efficacy Tests: Indicators of Bioavailability not Consumer Potential for Contamination We should also reject the sophistry of claiming current PETs are predictive of consumer contamination. My own research included •2, the claim of predictive ability by any PET based on planktonic unicells is just a serendipitous correspondence, not a correlation. To claim a "predictive" PET one only needs to conduct the in-use test so that well-preserved products do not become contaminated before the poorly preserved ones do. So what are we left with when we have developed an excellent, well-controlled, statistically-correct PET that uses planktonically-grown, lab-tamed, ATCC cultures? We do not have a test that can be used to predict real- world exposure. However, since planktonic organisms are much more sensitive to biocides than biofilm organisms, it could be argued that a product failing such a test certainly represents a risk to the public. This is an incorrect argument since no PET considers the effect that the container has in protecting the product from consumer contamination. Thus, no PET can be developed with a truly predictive ability in order to enforce a recall. Hoxvever, such a test will be able to rank all products on an equal plane with respect to bioavailability of preservative in the formula. When a product fails such a test, it does not substantiate a claim that it is a risk to the population it only indicates that it may not provide as much available biocide as another product that passes the test. If it can be shown that such a product derives no protection from the container, then one has a reason to do some field studies retrieving consumer-used product to see if the product has become contaminated. Once the field saudies show that the product is a risk, then one has grounds for recall. However, even products that pass the PET are not necessarily safe since the organisms do not represent the kinds of organisms it will be exposed to in the real world. Field testing is the only method that can truly represent how a product will behave. Thus, PETs are simply indicators of bioavailability of preservative but they are not predictors of consumer-contamination potential. Nor are they predictive of the potential for manufacturing contamination. In order for an in vitro test to be a valid predictor of reality, it must first discard the old ways of doing microbiology and come into the nexv age of biofilms and the methods used to study them. Acknowledgements: Thanks to the foilroving for critical reviews: Drs. Doug Caldwell, Murry Cooper, Bill Costerton, Phil Gels, Rich Mulhall, Don Singer, Scott Sutton. All responsibility, however, rests entirely on the authors. References Cited: 1. Cosierton, J. W., Z. Lewandowski, D. E. Caldwell, D. R. Korber, H. M. Lappin-Scott, Ann. Rev. Mtcrobtol., 49, 711 (1995). 2. Yu, F. P. andG. A_ McFeters, AppI. Envtron. Mtcrobtol., 60, 2462 (1994). 3. Favero, M. S., W. W. Bond, N.J. Peterson, and E. H. Cook, Jr., Proceedings of the International S),mpostum on Portdone Iodme (13. A_ Digenis and J. Arisell, eds.), University of Kentucky Press, pp 158-166 (1983 ). 4. Brown, M. L., H. C. Aldrich, and J. J. Gauthier, Appl. Environ. Mtcrobtol. 61, 187 (1995). 5. deBeer, D., R. Srinivasan, and P.S. Stewa• Appl. Environ. Microbtol. 60, 4339 (1994). 6. Brannan, D. K., J. Soc. Cosmet. Chem., 46, 199 (1995). 7. Foley, I., A_ Jocob, and P. Gilbert, ASM Abstracts, New Orleans ASM National Meeting, (1996). 8. Dibdin, G.H., S.J./Lssinder, W.W. Nichols, and P.A_ Lambert, J. Anttmicrob. Chetnother., 38, 757 (1996). 9. Gefiic, S. G., H. Heymann, and F. W. Adair, Appl. Environ. Mtcrobiol., 37, 505 (1979). 10. Gilbert, P.. P. J. Collier, and M. R. W. Brown, ,4nttrnicrob. Ag. Chemother. 34, 1865 (1990). I 1. Allison, D. 13. and P. Gilbert, J. lndust. Mtcrobiol. 15, 311 (1995). 12. Chapman, J. S., M. A_ DieM, and R. C. Lyreart, J. lndust. Mtcrobtol., 12, 403 (1993). 13. Deretic, V., M. J. Schutt, J. C. Boucher, and D. W. Martin, J. Bactertol. 176, 2773 (1994). 14. Brozel, V. S. and T. E. Cloete, J. Appl. Bacteriol. 76, 576 (1994). 15. Nikaido, H., Science, 264, 382 (1994). 16. Sabate, J., A_ Vilanueva, and M. J. Prieto, FEMSMicrobiol. Lett. 17. Russell, A_ D., J. AppL BactertoL, 71, 191 (1991). 18. Caldwell, D.E., D.R. Korber, and J.R. Lawrence, Advances inMlcrobialEcology, Plenum Pr•..s, New York, 12, 1-67 (1992). 19. Caldxvell, D.E., G.M. Wolfaardh D.R. Korber, and J.R. Lawrence, Manual ofEnvtronmentalMicrobiology, ASM Press, Washington. I)(2, pp 79-90 (1996). 20. Tsien, R.Y., and A_ Waggoner, Handbook of ConfocalMicroscop), (ed. J.B. PawIcy), Plenum, New York, pp 169-78 (1990). 21. Kinniment, S.D, J.W. Wirepenny, D. 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66 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS NATURAL PRESERVATIVES Anthony C. Dweck Research Director, Peter Black Medicare Ltd., White Horse Business Park, Aintree Avenue, Trowbridge, Wiltshire, UK. BA14 OXB INTRODUCTION The subject of natural preservatives is one that probably has more academic interest than practical or economic virtue. However, it does have a marketing angle which may justify the higher raw material costs. LEGAL POSITION No preservative may be used which does not appear in:- Annex VI Part 1 or 2 of the EEC Cosmetic Directive 76/768/EEC - including 19th. amending Commission Directive 94/32/EC. However, there is no legislation for those natural materials, which, when used for their beneficial effect on the skin, may coincidentally have a positive effect on the total preservative requirement of the formulation. Of course, no material appearing in Annex II may be considered. SUGAR High levels of sugar can preserve against spoilage organisms, this may be seen in jams, preserves, certain sweet pickles and marmalades. This is also an important factor in the preservation of boiled sweets and chocolates etc. Increasingly, it will be noticed that many products now have to be kept in the refrigerator of freezer once opened, because sugar has been replaced by artificial sweetener which is cheaper and healthier(?) to eat, but which compromises the self-preservation of the product. HONEY Honey in its undiluted form is also a natural preservative and, indeed, there are many learn6d papers citing honey as a viscous barrier to bacteria and infection. ALCOHOL Not all organisms are bad! The production of alcohol from sugar by yeast is an industry in its own right. A wine carefully produced using sterilised equipment and fermented to 13% by volume will just about resist further infection from external organisms, once the ferment has completed. It is during the time of the fermentation process that the fermenting must is vulnerable to infection. The naturally produced fermentation grade alcohol can be concentrated by distillation and used as a natural preservative in toners, aftershaves and colognes. HEAT Heating, cooking and pasteurisation is another natural form of preservation that will sterilise products, especially where that product is designed as a one-shot use product - for example, a phial or a sachet. Alternatively, once opened, the product can be stored in the fridge of freezer to prevent microbiological degradation. DESICCATION Removing water from a product or making it totally dehydrated will greatly reduce the possibility of spoilage however, it must be recognised that the presence of spore-bearing organisms could become active once that water is reintroduced. ANHYDROUS In a similar vein, one could make products with materials that do not contain any traces of water, i.e. to deliberately design and formulate a totally anhydrous product. However, creams that can be finished by the consumer, by introducing water to the blend of oils, fats and waxes are prone to the same restrictions as the dessicated products.