36 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS EXPERIMENTAL We are exploring this approach in both a simple model system and in cosmetic products as they are formulated for the market. The objective in the model system work is to test Evans' treatment of simple emulsions. The system is a simple olive oil/water emulsion aseptically prepared with a minimum level of emulsifier (to permit representative sampling after mixing). Typical compositions studied to date are given in Table III. Table III Composition of Oil Water Emulsions (%) 50/50 Oil/Water 20/80 Oil/Water 1.2% 1.2% 0.6% 0.6% Control Methyl Butyl Control Methyl Butyl Olive Oil a 49.90 49.35 49.35 19.98 19.86 19.86 Water 49.90 49.35 49.35 79.92 79.44 79.44 Myrj 52 b 0.05 0.05 0.05 0.05 0.05 0.05 Brij 72 b 0.05 0.05 0.05 0.05 0.05 0.05 Methyl Paraben c 0.00 1.20 0.00 0.00 0.60 0.00 Butyl Paraben c 0.00 0.00 1.20 0.00 0.00 0.60 100.00 100.00 100.00 100.00 100.00 100.00 •Progresso. •ICI, Americas Inc. CWenneco. Methyl and butyl paraben were compared at 0.6 and 1.2%, just below the measured solubilities of the former, 0.65 and 1.25% in 20 and 50% oil mixtures respectively. In these same mixtures, the butyl paraben solubilities are much greater, 3 and 7%, and consequently this preservative is present in both comparisons at less than half its saturation concentration. Fifty (50) gram aliquots in 125 m/ Erlenmeyer flasks were inoculated with a 24-hr culture of Pseudomonas cepacia adjusted to give a count of either 1.8 X 103 or 1.8 X 105/ml. The capped flasks were mixed continuously on a gyratory shaker. On day 14 respective aliquots were inoculated again at 2.2 X 102 or 2.2 X 104/ml and on day 25 at 4 X 102 or 4 X 104/m/. Standard plate counts were performed on each work day from days 0 through 31. In both unpreserved 20% oil mixtures (low and high inoculation levels) the recoveries were 107 to 108/ml throughout the test period. In the unpreserved 50% oil mixtures the recovery increased to 108/ml over the first few days and remained at that level thereafter. The emulsions with methyl paraben near its saturation concentration gave no recoverable counts at any time during the test period. The undersaturated butyl paraben emulsions gave somewhat reduced counts on day one, but by day two the counts had risen to 106/ml and increased to greater than 107 throughout the test. Overall, the butyl paraben systems were not markedly different from the unpreserved controls. In a similar test of the 20% olive oil emulsion with methyl and butyl parabens at levels corresponding to the same saturation fractions (as estimated from our solubility measurements), we found methyl paraben very effective (no recoveries) at 0.5 and 0.4%

PARABENS 37 and marginally effective at 0.3% the last is below half saturation. Butyl paraben, on the other hand, gave no significant reduction in count relative to the unpreserved control even at 2.75%, or about 90% of saturation. This result is in conflict with expectation based on Evans' analysis but it supports, albeit inexplicably, our qualitative rule that the less soluble preservative is preferable. In the standard challenge test of products being developed, we inoculate' with resuspended 24-hr cultures of bacteria grown in nutrient broth and fungi growt• on low pH Mycophil agar. Bacterial samples are incubated for 18-48 hr at 32øC, fungi for five days at 20-23øC. The challenge organisms include Pseudomonas aeruginosa, Enterobacter hafnia, Aspergillus niger and Enterobacter agg[omerans. In some cases, we use both ATCC strains and wild strains isolated from cosmetics. Following are summaries of observations in several cases Where products in develop- ment were compared at equal weight concentrations of single and mixed parabens: 1. A heavy night cream with 0.8% methyl parben gave no recoveries at any tes• date in a three inoculation study ove• 11 weeks. The same product with the conventional 0.5% methyl paraben, 0.3% propyl paraben gave high counts in all tests after the fourth week and with 0.8% propyl paraben high counts were recovered throughout the test. 2. An eyeliner with 0.8% methyl paraben gave no recoveries at any point in a two-inoculation study while its counterpart witl• 0.5% methyl paraben, 0.3% propyl paraben, gave high counts in all samples. 3. A glycerine/rose water hand cream, which gave high mold counts from the outset and high bacterial counts after the fourth week when preserved with 0.2% methyl paraben and 0.15% propyl paraben, gave only low mold counts at week fot•t and no counts on any other test date when preserved with 0.35% methyl paraben alone. 4. A moisturizing lotion with 0.5% methyl paraben and 0.3% propyt paraben and its counterpart with 0.8% propyl paraben both gave high counts throughout a two- inoculation test. The same product with 0.8% methyl paraben gave no counts during the first three weeks of the test but gave high counts at the fourth week and thereafter. Although we have not always succeeded in satisfactorily preserving problem products with a single paraben, we have not yet found an instance of better preservation with a mixed paraben system than with an equal weight of a single pataben. DISCUSSION The "problem" products are those with high levels of vegetable oils (fragrance oils probably contribute to the problem as indicated in reference 13) emulsified with nonionic surfactants and thickened with weak acid polyelectrolytes at pH's above 7.0. The generally greater efficacy of methyl paraben in such systems is probably attributable, in part, to its lower solubility in vegetable oils as shown in Figure 1. We have also found methyl paraben much less strongly adsorbed on talc and less soluble in shampoos and liquid bubble baths, products with high concentrations of solubiliz- ing surfactants. Perhaps all of these observations on methyl paraben are reflections of the fact that it is the most water soluble member of the paraben series and, hence, the least susceptible to deactivation in many emulsion systems. This would suggest that a somewhat more