18 JOURNAL OF COSMETIC SCIENCE Shimadzu, Kyoto, Japan). For plotting the calibration curve, gallic acid (standard) solutions at different concentrations (0, 50, 100, 150, 250, and 500 mg/l) were prepared in ethanol. The total phenolic content of each EO was expressed as gallic acid equivalent (mg GAE/100 g sample). Each experiment was repeated three times. Cytotoxicity assay. The in vitro cytotoxic activity of the EOs (oregano, cinnamon, lemon, tea tree, laurel, lavender, sandalwood, and eucalyptus) was assessed by MTT assay in NIH 3T3 cell line (derived from mouse embryonic fibroblasts) in 24, 48, 72, and 96-hour time frames. Cells were grown in Dulbecco’s Modified Eagle’s Medium seeded into 96-well plate and incubated at 37°C in a 5% CO 2 incubator for 24 hours before adding EO. Serial dilutions ranging from 2% to 0.0625% (v/v) of EOs were prepared. The absorbance was measured at 570 nm using a microplate reader (HTX Synergy, BioTek, Winooski, VT, USA). The cytotoxic activity was expressed as IC 50 (50% cell growth inhibition). STATISTICAL ANALYSIS For disc diffusion assay, the results were performed by one-way ANOVA and Tukey test (comparison with controls) using SPSS Statistics 19 version. For the MTT assay, the data was performed by ANOVA test using GraphPad Prism version 6.0. The significance value of all data was considered at p 0.05. RESULTS AND DISCUSSION PHYTOCHEMICAL CONSTITUENTS OF ESSENTIAL OILS The chromatogram results of each EO are presented in Figure 1. In our research, the various chemical compounds such as terpenes, terpene alcohols, ethers, ketones, aldehydes, and phenol derivatives were determined by gas chromatography–mass spectrometry (GCMS), and major compounds are given in Table I. When we compared our results to previous studies, the percentages of the main components of lemon (27), tea tree (28), lavender (29– 31), laurel (32), eucalyptus (33,34), sandalwood (35), cinnamon (36), and oregano (37,38) EOs were different. These differences depend on environmental conditions like the season, geographical location, plant genotype, and plant growth (39). ANTIBACTERIAL ACTIVITY OF SELECTED ESSENTIAL OILS AGAINST ACNE-INDUCING BACTERIA Agar disc diffusion assay. The antimicrobial activity results of 16 EOs against strains are displayed in Table II. Oregano, cinnamon, lavender, and tea tree EOs showed excellent antibacterial activity. Ginger, orange, and coconut EOs did not show any inhibitory activity against either bacterium. The inhibitory activity wasn’t seen in chamomile, rosemary, garlic, sage, and clove EOs against S epidermidis and lemon EO against C acnes. The negative control Tween 80 exhibited no inhibition zone. Previous studies have reported that 2 bacteria were susceptible to oregano with IZD: 16 to 32 mm (40). Moderate activity was observed in tea tree EO (IZD: 17 mm). Lavender and chamomile EOs showed no inhibition. Oregano (IZD: 30 mm), lavender (IZD: 18 mm), and lemon (IZD: 14 mm) EOs showed antibacterial activity against S epidermidis (41). Cinnamon (IZD: 33.5 mm), chamomile (IZD: 10 mm), lavender and lemon (IZD: 10–15 mm) (42), eucalyptus

Figure 1. Cell viability (%)of 3T3 cells treated with EOs (2–0.0625% v/v) for (A) 24h, (B) 48h, (C) 72h, and (D) 96h p 0.05*. 19 CHARACTERIZATION AND ACTIVITY OF ESSENTIAL OILS