The application of biological materials in synthesizing nanoparticles has become significant issue in nanotechnology. This research was designed to assess biogenic silver nanoparticles (AgNPs) fabricated using two aqueous extracts of Acacia arabica (Arabic Gum) (A-AgNPs) and Opophytum forsskalii (Samh) seed (O-AgNPs), which were used as reducing and capping agents in the NPs development, respectively. The current study is considered as the first report for AgNP preparation using Opophytum forsskalii extract. The dynamic light scattering, transmission electron microscopy, and scanning electron microscopy were employed to analyze the size and morphology of the biogenic AgNPs. Fourier transform infrared (FTIR) spectroscopy and chromatography/mass spectrometry (GC-MS) techniques were used to identify the possible phyto-components of plant extracts. The phyto-fabricated NPs were assessed for their antibacterial activity and also when combined with some antibiotics against Staphylococcus aureus (Gram-positive) and Pseudomonas aeruginosa and Escherichia coli (Gram-negative) and their anticandidal ability against Candida albicans using an agar well diffusion test. Furthermore, cytotoxicity against LoVo cancer cell lines was studied. The results demonstrated the capability of the investigated plant extracts to change Ag+ ions into spherical AgNPs with average size diameters of 91 nm for the prepared O-AgNPs and 75 nm for A-AgNPs. The phyto-fabricated AgNPs presented substantial antimicrobial capabilities with a zone diameter in the range of 10–29.3 mm. Synergistic effects against all tested strains were observed when the antibiotic and phyto-fabricated AgNPs were combined and assessed. The IC50 of the fabricated O-AgNPs against LoVo cancer cell lines was 28.32 μg/mL. Ten and four chemical components were identified in Acacia arabica (Arabic Gum) and Opophytum forsskalii seed extracts, respectively, by GC-MS that are expected as NPs reducing and capping agents. Current results could lead to options for further research, such as investigating the internal mechanism of AgNPs in bacteria, Candida spp., and LoVo cancer cell lines as well as identifying specific molecules with a substantial impact as metal-reducing agents and biological activities.