| Summary: | The use of phytochemicals is gaining increasing attention for treating cancer morbidity with minimal burden side effects. This study evaluated the cytotoxicity and antimicrobial activities of <i>Pinus roxburghii</i> branch (<i>P. roxburghii</i>) and <i>Nauplius graveolens</i> (<i>N. graveolens</i>) extracts <i>in vitro</i>. Cell viability was estimated using MTT assay. DNA fragmentation was determined to detect apoptotic pathway initiation. Mechanistically, the apoptotic pathway was tracked by estimating the relative mRNA expression levels of the Bcl-2, Bax, Cas3, NF-κB, and PI3k genes by qRT-PCR. <i>P. roxburghii</i> exhibited moderate antioxidant activity, while <i>N. graveolens</i> possessed highly significant (<i>p</i> < 0.05) scavenging activity against DPPH and ABTS assays. HPLC analysis demonstrated that catechin and chlorogenic acid were the predominant polyphenolic compounds in <i>P. roxburghii</i> and <i>N. graveolens</i>, respectively. The <i>P. roxburghii</i> and <i>N. graveolens</i> extracts inhibited the viability of HCT-116 cells with IC<sub>50</sub> values of 30.6 µg mL<sup>−1</sup> and 26.5 µg mL<sup>−1</sup>, respectively. DNA fragmentation analysis showed that the proposed extracts induced apoptosis in HCT-116 cells. Moreover, the IC<sub>50</sub> doses of the selected extracts significantly (<i>p</i> < 0.05) upregulated Bax and cleaved Cas-3, and downregulated Bcl-2, NF-κB, and PI3k genes versus the GAPDH gene as a housekeeping gene in comparison to the control group. The Bax/Bcl-2 ratio was raised upon treatment. The mentioned extracts exhibited antimicrobial action against all tested bacteria and fungi. The highest antibacterial effect was recorded against <i>E. coli</i>, with inhibition zones of 12.0 and 11.2 mm for <i>P. roxburghii</i> and <i>N. graveolens</i>, respectively. On the other hand, the highest antifungal action was registered for <i>Penicillium verrucosum</i> and <i>A. niger</i>, with inhibition zones of 9.8 and 9.2 mm for the tested extracts, respectively. In conclusion, the outcomes of this study indicate that <i>P. roxburghii</i> and <i>N. graveolens</i> extracts could potentially be used as anticancer, antibacterial, and antifungal agents.
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