414 JOURNAL OF COSMETIC SCIENCE S. aureus P. aeruginosa C. albicans A. niger Figure 2. Minimum inhibitory concentration (pg/ml) of methyl paraben during six months. BF: Before freeze-drying. F: After freeze-drying. d: days. culture. Moreover, the culture of microorganisms can also suffer some modification of characteristics (10,18). The minimum inhibitory concentration (MIC) values for methyl paraben (Figure 2) and for Glydant plus © (Figures 3 and 4) were the same before and after the freeze-drying process. This performance indicated that the microorganisms did not lose resistance after the process. Propyl paraben also presented this behavior, except for the A. niger. Fur- thermore, all the results (Figures 2, 3, 4, and 5) showed variability from one month S. aureus P. aeru•nos• Figure 3. Minimum inhibitory concentration (pg/ml) of Glydant pins © for bacteria during six months. BF: Before freeze-drying. F: After freeze-drying. d: days.

PRESERVATIVE EFFICACY TESTING METHOD 415 C. albicans A. niger Fiõure 4. Minimum inhibitory concentradon (pg/ml) of Glydant plus © for fungi during six months. Before freeze-drying. F: After freeze-drying. d: days. •]BF 5o0 .': i::-•:?"i:: ?: :?-i' ?:.if'•'7':-•: _'.:•. '. :::::::::[::::)':5 ii -•'::'?!::•?•: : !!:!':!::i'• : -':::::.::!::: [] F E •60d • • • -'-= .... :'"•::! i•i:•' ::177:.::::__:__-'-i:120d•::'--:-::':•'*:• S. aureus p. aeruginosa C. albicans A. niger FJõure •. Minimum inhibkory contentradon (pg/ml) of propy! paraben during six months. B•: Before freeze-drying. F: After freeze-drying. d: days. to another, which could be attributed to the method, since the oscillations were unim- portant. The adequate support agent must be used to increase the survival level, and to protect the microorganisms against damage that could occur during the freeze-drying process (19). In this research, skim milk added to inositol was used since it is known to be a good agent to preserve the viability of microorganisms (20). It would be possible to maintain a better viability for the freeze-dried microorganisms. Some authors demonstrated that the use of ampoules instead of vials could show better results (17,21). The option of using vials was adopted in order to make the process easier, but the study of freeze-dried microorganisms in ampoules is being performed. The microorganisms were tested by the linear regression method (13) and by the official ones (6-1o).