In this present study, commercial Pharmacopeia (PhEur) grade chamomile essential oil (Mairicariae aetheroleurn) was combined with different antimicrobial agents including ampicillin sodium, cefuroxime acetyl, tetracycline hydrochloride, fluconazole and nystatin. All combinations were evaluated in vitro against pathogenic standard and clinical resistant Gram-negative (Escherichia coli) and Gram-positive (Staphylococcus aureus) bacterial isolates as well as against Candida albicans for their broad antimicrobial effectiveness. Furthermore, the essential oil was fractioned by column chromatography using n-hexane, diethyl ether, dichloromethane and methanol, respectively. Additionally, all fractions of essential oil were tested in combinations for their minimum inhibitory concentrations (MIC) as well as for their fractional inhibitory concentrations (FIC) against the resistant microbial pathogens. Antimicrobial activities were evaluated by microdilution method and antimicrobial interactions were assayed using the checkerboard method. Cytotoxicity of compounds were evaluated using Cytotox-XTT-1 1:rameter Kit in WS1 cells and Aliivibrio fischeri bioluminescence toxicity assay. The analyses proved that alpha-bisabolol oxide A (47.7%), (E)-beta-famesene (21.5%), alpha-bisabolol oxide B (6.2%), alpha-bisabolone oxide A (5.8%), chamazulene (4.1%) and alpha-bisabolol (2.2%), respectively were the major compounds and in compliance with PhEur. The essential oil combination of fluconazole and nystatin showed "synergic and additive inhibitory effects" against the clinical Candida strain. According to the IC50 values obtained, the inhibitory concentrations of combinations against the clinical Candida strain can be considered to be selective when compared with its effect on WS1 cells. Additionally, the essential oil combination of fluconazole and nystatin showed low toxicity against A. fischeri.