ALTERNATIVES TO COSMETICS PRESERVATION 109 to those of parabens (23). Although Lonicera caprifoleum and Lonicera japonica extracts are well known for their antimicrobial properties (24–26), there are no studies in the litera- ture regarding their incorporation as preservatives in cosmetic formulations. In order to evaluate the preservative effi cacy of these multifunctional ingredients, we performed challenge tests (preservative effi cacy tests, PETs) according to the standards proposed by the European Pharmacopoeia. Furthermore, we examined the microbial purity of the formulations in two different states of use (the intact product and following use) because few published papers refer to the effi cacy of preservative systems contained in cosmetic products during their use by consumers (27,28). MATERIALS AND METHODS COSMETIC FORMULATIONS A series of aqueous formulations, i.e., tonic lotion, shampoo, and shower gel, and O/W cosmetic formulations such as conditioning cream, anticellulite cream, cleansing milk, and peeling cream was prepared. Tonic lotion. Water, Syringa vulgaris (lilac) extract, lactic acid, cinnamyl alcohol, hydroxy- citronellal, and preservative systems I, II, III, or IV (Table III) were used as the ingredi- ents in the tonic lotion formulation. Shampoo. Water, sodium cocoyl isethionate, lauryl glycoside, cocamidopropyl betaine, cocobetaine, glyceryl oleate, coco glycoside, hydrolyzed milk protein, sodium phytate, Urtica dioica leaf water, Rosmarinus offi cinalis (rosemary) leaf water, Salix alba (white willow) Table I MIC Values of the Mixture Consisting of Lonicera caprifoleum and Lonicera Japonica Extracts (Plantservative WSr) (25) Microorganism MIC* (% w/v) Staphylococcus aureus 0.125 Escherichia coli 0.125 Pseudomonas aeruginosa 0.125 Aspergilus niger 0.20 Candida albicans 0.10 * Minimum inhibitory concentration. Table II Activity of the Alternative Preservatives Against Bacteria and Fungi (18,19) Alternative preservative Recommended dosage (% w/w) Gram+ Gram- Yeasts Molds Glyceryl caprylate 0.5–1.0 +++ +++ +++ + p-Anisic acid 0.05–0.3 ++ ++ ++ +++ Levulinic acid 0.2–0.3* +++ +++ ++ +++ +++ Very good activity. ++ Good activity. + Activity depends on compatibility or dosage. * As 2–3% w/w Dermosoft 1388®

JOURNAL OF COSMETIC SCIENCE 110 leaf water, Ginkgo biloba leaf water, citric acid, fragrance, linalool, and preservative system V (Table III) were used as the shampoo ingredients. Shower gel. The ingredients used in the shower gel formulation were: water, sodium cocoyl isethionate, lauryl glycoside, cocamidopropyl betaine, sodium lauryl glutamate, glyceryl oleate, cocoglucoside, cocobetaine, sodium phytate, Aloe barbadensis (Aloe vera) extract, Avena sativa (oat) leaf extract, Calendula offi cinalis leaf water, Arnica montana leaf water, Lavandula angustifolia (lavender) leaf water, hydrolyzed milk protein, parfum (fragrance), citric acid, D-limonene, and preservative system V (Table III). Conditioning cream. The conditioning cream ingredients were: water, hydroxypropyl starch phosphate, cetyl alcohol, dioleyloylethyl hydroxyethylammonium methosulfate, sucrose laurate, cetearyl alcohol, Macadamia ternifolia seed oil, glycerin, polyglyceryl-10 laurate, meadowfoam (Limnantes alba) seed oil, fragrance, stearyl stearate, glycine soja, phospholipids, soy sterol, sodium phytate, ethanol, lauryl glycoside, tocopheryl acetate, Urtica dioica (nettle) leaf water, Rosmarinus offi cinalis (rosemary) leaf water, Gingko biloba leaf water, Salix alba (white willow) leaf water, hydrolyzed milk protein, tocopherol, and preservative system V (Table III). Anticellulite cream. The ingredients used in the anticellulite cream formulation were: water, sodium stearoyl lactylate, caffeine, glycerin, tricaprylin, dicaprylyl carbonate, iso- propyl myristate, polyglyceryl-3 stearate, dicaprylyl ether, cetyl alcohol, glyceryl stear- ate, panthenol, behenyl alcohol, glyceryl stearate, lecithin, glycine soja (soybean) sterols, Lactobacillus/Trifolium pratense (clover) fl ower ferment extract, Lactobacillus/Theobroma cacao (cocoa) ferment extract, Lactobacillus/Camellia sinensis leaf ferment extract, Vitis vinifera (grape) seed oil, Prunus armeniaca (apricot) kernel oil, sodium phytate, xanthan gum, es- cin, tocopherol, citric acid, Olea europaea (olive) fruit extract, parfum (fragrance), linalool, benzyl benzoate, benzyl salicylate, farnesol, geraniol, eugenol, and preservative system V, VI, or VII (Table III). Cleansing milk. The cleansing milk ingredients were: water, isopropyl myristate, glyceryl stearate, polyglyceryl 3-stearate, myristyl myristate, glycerin, sodium stearoyl lactylate, tricaprylin, caprylic/capric triglyceride, Calendula offi cinalis oil, cetearyl alcohol, bisab- olol, Prunus armeniaca (apricot) kernel oil, tocopherol, xanthan gum, sodium phytate, Aloe barbadensis extract, sodium hydroxide, citric acid, Chamomilla recutita (Matricaria) extract, and preservative system V, VI, VII, or VIII (Table III). Table III Preservative Systems Preservative systems Concentration of preservative system in tested formulation (w/w) I Lonicera extracts 0.2 % II Lonicera extracts 0.2 % + p-anisic acid 0.1 % III Lonicera extracts 0.2 % + levulinic acid 0.3 %* IV Lonicera extracts 0.2 % + ethanol 5 % V Lonicera extracts 0.2 % + glyceryl caprylate 1 % VI Lonicera extracts 0.2 % + glyceryl caprylate 1 % + p-anisic Acid 0.1 % VII Lonicera extracts 0.2 % + glyceryl caprylate 1 % + levulinic acid 0.3 % VIII Lonicera extracts 0.2 % + glyceryl caprylate 1 % + levulinic acid 0.1 %** * As 3 % w/w Dermosoft 1388. ** As 1 % w/w Dermosoft 1388.