Naringenin is a 5,7,4′-trihydroxyflavanone naturally occurring mainly in citrus fruits, characterized by a wide spectrum of biological activity. Chemical modifications based on alkylation and oximation in most cases increase its bioactivity. The aim of our research was to evaluate the antiproliferative activity and influence on selected representatives of the human gut microbiota of new synthesizedO-alkyl derivatives (A1–A10) and their oximes (B1–B10), which contain hexyl, heptyl, octyl, nonyl and undecyl chains attached to the C-7 or to both the C-7 and C-4′ positions in naringenin. To the best of our knowledge, compoundsA3,A4,A6,A8–A10andB3–B10have not been described in the scientific literature previously. The anticancer activity was tested on human colon cancer cell line HT-29 and mouse embryo fibroblasts 3T3-L1 using the sulforhodamine B (SRB) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays. We also determined the impacts of all compounds on the growth of Gram-positive and Gram-negative bacterial strains, such asStaphylococcus aureus,Enterococcus faecalisandEscherichia coli. The antimicrobial activity was expressed in terms of minimal inhibitory concentrations (MIC) and minimal bactericidal concentrations (MBC) values. For 7,4′-di-O-hexylnaringenin (A2), 7-O-undecylnaringenin (A9) and their oximes (B2,B9), which were safe for microbiota (MIC > 512 µg/mL) and almost all characterized by high cytotoxicity against the HT-29 cell line (A2: IC50> 100 µg/mL;A9: IC50= 17.85 ± 0.65 µg/mL;B2: IC50= 49.76 ± 1.63 µg/mL;B9: IC50= 11.42 ± 1.17 µg/mL), apoptosis assays were performed to elucidate their mechanisms of action. Based on our results, new compoundB9induced an apoptotic process via caspase 3/7 activation, which proved its potential as an anticancer agent.