PRESERVATIVES 31


Table 6: Log reduction required vs number of bacteria killed. Inoculation level


Log Reduction Lower limit (105.0 ) 3 log reduction at 7 days 100,000 to 100 2 log reduction 2 days Upper limit (106.0 ) 3 log reduction 7 days


to questions whether the procedure is still the most applicable. The use of single organisms rather than mixtures is at odds with in-use challenges where mixtures of organisms are more likely to be encountered and mixed organisms may assist each other in surviving or colonising a product. It is also unlikely that a product will only be challenged on one occasion soon after manufacture. The sources of microorganisms include the raw materials, production and filling equipment, packaging and the long term use by consumers. Loss of the preservative added to a product may occur quickly, due to incorrect pH or temperature exposure during production, or more slowly due to migration into the oil phase or micelles. Challenging the product several times may show whether there is adequate protection if these problems occur. Standard development procedures should include repeating the preservative efficacy test at the completion of the shelf life period for products stored in the final retail container, but also modifying the test to include additional inoculations or the use of mixed pools of test organisms may be considered to provide a more realistic challenge to the preservative system. The final protocol should take into account the potential susceptibility of the product to become contaminated as outlined in Annex A of ISO 29621.13 The requirements for log10


reduction


indicate that the difficulty in passing the preservative efficacy test can be ranked as BP, ISO 11930 and finally the USP. The BP and ISO methods require the same reduction at 7 days while the BP also requires significant reduction after 2 days contact. The additional contact period required by the BP (2 days) can make it difficult for products preserved with multifunction actives or preservative boosters, commonly referred to as non- preservative preservatives, as they are often slower acting than traditional preservative actives.


The inoculum concentration is stipulated in test methods but includes a range in the numbers or organisms allowed. All tests listed in Table 5 allow an initial concentration load of 105.0


– 106.0


cfu/mL or g of the test sample. Data in April 2020


1,000,000 to 1,000 999,000 (106.0 )


Table 6 shows that the initial inoculum concentration determines the number of organisms that need to be killed to meet the required log reduction, with significantly more organisms needed to be killed to meet the test requirements if the inoculum concentration is towards the upper allowed limit. The data from Table 6 highlights the issue with log reductions in that it shows the variation between the number of organisms added and the number surviving while giving no information on the numbers involved. As there is a tenfold variation in the number of bacteria allowed in the inoculum concentration, there is a corresponding tenfold variation in the number of bacteria required to be killed to pass the test to the same reduction criteria.


What do they really mean? The Preservative Effectiveness Test methods are generally consistent provided a standard method is used and if conducted correctly they will indicate if a sample under test has sufficient preservative available in the water phase to protect the finished product from microbial contamination. However, poor hygiene practices allowing growth of organisms in the process equipment and especially the development of biofilms, changes in the formulation or quality of raw materials in particular, the fragrance or packaging may render the test results invalid. Incorrect maintenance of the test


organisms, introducing variations to the inoculum preparation, including media type, growth phase and inoculum numbers may have a huge effect on the result achieved, failing a good preservative system or passing a poor one.


Conclusion There are many antimicrobial products available on the market including all products containing a preservative and those containing biocides that claim antimicrobial properties. There are many tests available to determine the efficacy of these. However, the particular test method utilised and variations introduced into these method by the testing laboratory may have a huge impact on the result obtained and therefore, the conclusions


99,900 (105.0 ) 1,000,000 to 10,000 990,000 (105.99 ) Reduction 2 log reduction at 2 days 100,000 to 1,000 99,000 (104.99


Bacteria killed )


that can be drawn from those results. An assessment of the method used, the initial inoculum concentration and contact times are the minimum variants that need to be included in determining if log reduction or % killed numbers quoted actually indicate a true antimicrobial effect for the product.


PC


References 1 Regulation (EC) No 1223/2009 of the European


Parliament and of the Council as amended. Annex V, List of preservatives allowed in cosmetic products


2 Biocidal Products Regulation (EU) No 528/2012 of the European Parliament and of the Council of 22 May 2012 concerning the making available on the market and use of biocidal products.


3 Beveridge TJ. Structures of Gram-Negative Cell Walls and Their Derived Membrane Vesicles, J Bacteriology. 1999; 181(16): 4725–4733.


4 Ghai, Ghai S. Exploring bacterial outer membrane barrier to combat bad bugs. Infection and Drug Resistance 2017; 10: 261–273.


5 Luppens SBI, Rombouts FM, Abee T. The Effect of Growth Phase of Staphylococcus aureus on Resistance to Disinfectants in a Suspension Test. Journal of Food Protection 2002; 65(1):124 – 129.


6 Desai M. Increasing resistance of planktonic and biofilm cultures of Burkholderia cepacian to ciprofloxacin and ceftazidime during exponential growth, Journal of Antimicrobial Chemotherapy 1998; 42: 153–160


7 Antimicrobial Finishes on Textile Materials: Assessment of. AATCC Test Method 100 – 2012. American Association of Textile Chemists and Colorists


8 Antimicrobial Products – Test for Antimicrobial activity and efficiency. JIS Z2801 – 2000. Japanese Industrial Standard, Japanese Standards Association.


9 Measurement of antibacterial activity on plastics and other non-porous surfaces ISO 22196:2011(E). International Organization for Standardization[ISO]


10 Cosmetics – Microbiology – Evaluation of the Antimicrobial Protection of a Cosmetic Product. ISO 11930:2019, International Organization for Standardization [ISO]


11 Lee JS, Pfeifer DK. Influence of Recovery Media and Incubation Temperatures on the Types of Microorganisms isolated from Seafoods. Journal Milk Food Technology 1974; 37(11)


12 General requirements for the competence of testing and calibration laboratories ISO 17025 – 17, International Organization for Standardization [ISO]


13 ISO Cosmetics – Microbiology – Guidance for the risk assessment and identification of Microbiologically low-risk products. ISO 29621:2017, International Organization for


PERSONAL CARE EUROPE


Page 1  |  Page 2  |  Page 3  |  Page 4  |  Page 5  |  Page 6  |  Page 7  |  Page 8  |  Page 9  |  Page 10  |  Page 11  |  Page 12  |  Page 13  |  Page 14  |  Page 15  |  Page 16  |  Page 17  |  Page 18  |  Page 19  |  Page 20  |  Page 21  |  Page 22  |  Page 23  |  Page 24  |  Page 25  |  Page 26  |  Page 27  |  Page 28  |  Page 29  |  Page 30  |  Page 31  |  Page 32  |  Page 33  |  Page 34  |  Page 35  |  Page 36  |  Page 37  |  Page 38  |  Page 39  |  Page 40  |  Page 41  |  Page 42  |  Page 43  |  Page 44  |  Page 45  |  Page 46  |  Page 47  |  Page 48  |  Page 49  |  Page 50  |  Page 51  |  Page 52  |  Page 53  |  Page 54  |  Page 55  |  Page 56  |  Page 57  |  Page 58  |  Page 59  |  Page 60  |  Page 61  |  Page 62  |  Page 63  |  Page 64  |  Page 65  |  Page 66  |  Page 67  |  Page 68  |  Page 69  |  Page 70  |  Page 71  |  Page 72  |  Page 73  |  Page 74  |  Page 75  |  Page 76  |  Page 77  |  Page 78  |  Page 79  |  Page 80  |  Page 81  |  Page 82  |  Page 83  |  Page 84  |  Page 85  |  Page 86  |  Page 87  |  Page 88  |  Page 89  |  Page 90  |  Page 91  |  Page 92  |  Page 93  |  Page 94  |  Page 95  |  Page 96  |  Page 97  |  Page 98  |  Page 99  |  Page 100  |  Page 101  |  Page 102  |  Page 103  |  Page 104  |  Page 105  |  Page 106  |  Page 107  |  Page 108  |  Page 109  |  Page 110  |  Page 111  |  Page 112  |  Page 113  |  Page 114  |  Page 115  |  Page 116  |  Page 117  |  Page 118  |  Page 119  |  Page 120  |  Page 121  |  Page 122  |  Page 123  |  Page 124  |  Page 125  |  Page 126  |  Page 127  |  Page 128  |  Page 129  |  Page 130  |  Page 131  |  Page 132  |  Page 133  |  Page 134  |  Page 135  |  Page 136  |  Page 137  |  Page 138  |  Page 139  |  Page 140  |  Page 141  |  Page 142  |  Page 143  |  Page 144  |  Page 145  |  Page 146  |  Page 147  |  Page 148  |  Page 149  |  Page 150  |  Page 151  |  Page 152  |  Page 153  |  Page 154  |  Page 155  |  Page 156  |  Page 157  |  Page 158  |  Page 159  |  Page 160  |  Page 161  |  Page 162  |  Page 163  |  Page 164  |  Page 165  |  Page 166  |  Page 167  |  Page 168  |  Page 169  |  Page 170  |  Page 171  |  Page 172  |  Page 173  |  Page 174  |  Page 175  |  Page 176  |  Page 177  |  Page 178  |  Page 179  |  Page 180  |  Page 181  |  Page 182  |  Page 183  |  Page 184  |  Page 185  |  Page 186  |  Page 187  |  Page 188  |  Page 189  |  Page 190  |  Page 191  |  Page 192  |  Page 193  |  Page 194  |  Page 195  |  Page 196