Methicillin-resistant Staphylococcus aureus (MRSA) continues to represent a significant clinical challenge, characterized by consistently elevated rates of morbidity and mortality. Care regimen success is still difficult and necessitates assessing new antibiotics as well as supplemental services, including source control and searching for alternative approaches to combating it. Hence, we propose to synthesize silver nanoparticles (Ag-NPs) by employing a cell-free filter (CFF) of Streptomyces sp. to augment antibiotic activity and combat biofilm-forming MRSA. Seven bacterial isolates from clinical samples were identified, antibiotics were profiled with Vitek-2, and the phenotypic detecting of biofilm with Congo red medium and microplate assay was carried out. The PCR technique was used for detecting genes (icaA and icaD) coded in biofilm forming. The characterization of Ag-NPs was performed using several analytical methods, such as UV spectroscopy, dynamic light scattering (DLS), zeta potential measurement, transmission electron microscopy (TEM), X-ray diffraction (XRD), and Fourier transform infrared spectroscopy (FTIR). The antibacterial properties of Ag-NPs and oxacillin–Ag-NPs were assessed against standard strains and clinical isolates by employing the agar well diffusion technique and the microdilution assay. The biogenic synthesis Ag-NPs resulted in uniformly spherical particles, with an average size of 20 nm. These Ag-NPs demonstrated significant activity against biofilm-forming MRSA, with minimum inhibitory concentrations (MICs) ranging from 12 to 15 μg/mL. Additionally, Ag-NPs completely impede biofilm formation by MRSA at sublethal doses of 0.75 MICs. The expression levels of the icaA and icaD genes were reduced by 1.9- to 2.2- and 2.4- to 2.8-fold, respectively. A significant synergistic effect was noted when Ag-NPs were used in combination with oxacillin, leading to reduced MICs of 1.87 μg/mL for oxacillin and 4.0 μg/mL for Ag-NPs against MRSA. The FICi of 0.375 further validated the synergistic relationship between oxacillin and Ag-NPs at the concentrations of 1.87 and 4 μg/mL. Findings from the time-kill test demonstrated the highest reduction in log₁₀ (CFU)/mL of the initial MRSA inoculum after 12-hour exposure. The cytotoxicity analysis of Ag-NPs revealed no significant cytotoxic effects on the human skin cell line HFB-4 at low concentrations, with IC₅₀ values of 61.40 µg/mL for HFB-4 and 34.2 µg/mL for HepG-2. Comparable with oxacillin–Ag-NPs, Ag-NPs showed no cytotoxic effects on HFB-4 at different concentrations and exhibited an IC₅₀ value of 31.2 against HepG-2-cells. In conclusion, the biosynthesis of Ag-NPs has demonstrated effective antibacterial activity against MRSA and has completely hindered biofilm formation, suggesting a valuable alternative for clinical applications.

Background: Nigella sativa (black cumin) seeds are renowned for their ethnomedicinal significance and are rich in bioactive phytochemicals, which contribute to food preservation and the prevention of various diseases through their antimicrobial and antioxidant properties. Accordingly, this study aimed to characterize the phytochemical composition of N. sativa seed extracts, isolate thymoquinone, and assess their antibacterial, antibiofilm, antioxidant, anti-inflammatory and antidiabetic activities. Methods: Nigella sativa seed extracts were prepared using solvents of increasing polarity and analyzed for phytochemical content. Metabolite profiling was performed using UHPLC/QTOF-MS. Thymoquinone, the major constituent, was isolated via thin-layer chromatography (TLC), further purified using semi-preparative reverse-phase high-performance liquid chromatography (RP-HPLC), and evaluated in vitro for antibacterial, antibiofilm, antioxidant, anti-inflammatory, and antidiabetic activities. Results: Extraction yields ranged from 5.5% to 8.4% (w/w), with methanol yielding the highest phenol (6.34 ± 0.31 mg GAE/mL) and flavonoid (5.12 ± 0.26 mg QE/mL) contents. UHPLC/QTOF-MS revealed a chemically diverse profile dominated by thymoquinone (58% relative abundance), alongside p-cymene, carvacrol, longifolene, and nigellidine. Thymoquinone (Rf = 0.56) was initially isolated from the methanolic extract with a yield of 270 mg/g and further purified from preparative TLC fractions using semi-preparative RP-HPLC, affording 82 mg of >95% pure compound with a 68.3% recovery, suitable for subsequent biological assays. It inhibited Gram-positive and Gram-negative bacteria, with MICs of 62.5 µg/mL against Staphylococcus aureus, Bacillus subtilis, and Listeria monocytogenes; 125–250 µg/mL against Escherichia coli and Salmonella typhimurium; and 500 µg/mL against Pseudomonas aeruginosa. Thymoquinone reduced biofilm formation (>80% at 25–50 µg/mL; MBIC₅₀ ≈ 5.4–11.6 µg/mL), exhibited antioxidant activity (DPPH IC₅₀ = 52.3 ± 2. 1 µg/mL; ABTS IC₅₀ = 41.6 ± 1.9 µg/mL), stabilized erythrocyte membranes (IC₅₀ ≈ 14.8 µg/mL), and inhibited carbohydrate-hydrolyzing enzymes, with stronger inhibition of α-glucosidase (~92%) than α-amylase (~84%) at 128 µg/mL. Conclusions: Thymoquinone is a major bioactive constituent of N. sativa seeds, exhibiting consistent multi-target in vitro activity. These findings highlight its functional relevance and in vivo investigations to establish therapeutic potential.

Background: Rosmarinus officinalis L. is a medicinally important aromatic plant rich in bioactive secondary metabolites with diverse therapeutic properties. This study aimed to characterize the phytochemical profile of R. officinalis leaf extracts, isolate carnosic acid as a major bioactive diterpene, and evaluate its biological activities. Methods: Leaf extracts were prepared using solvents of increasing polarity and analyzed by phytochemical screening and UHPLC/QTOF-MS. Carnosic acid was isolated by thin-layer chromatography and assessed for antibacterial, antibiofilm, antioxidant, anti-inflammatory, antidiabetic, and antiproliferative activities using in vitro assays. Expression of apoptosis-related genes was also investigated. Results: Methanolic and ethanolic extracts exhibited the highest abundance of phenolic compounds and secondary metabolites, whereas the hexane extract showed lower phytochemical content. UHPLC/QTOF-MS identified seven major metabolites, including phenolic acids, flavonoids, and abietane-type diterpenes. Purified carnosic acid demonstrated potent antibacterial activity (MIC: 10–23 μg/mL) and inhibited biofilm formation by up to 90%. Strong antioxidant activity was observed, with DPPH and ABTS radical-scavenging IC₅₀ values of 125 and 130 μg/mL, respectively. The compound also exhibited notable anti-inflammatory activity and markedly inhibited α-amylase and α-glucosidase activities. Furthermore, carnosic acid exhibited dose-dependent antiproliferative activity against MCF-7, HepG2, and MCF-10A cells, reducing cell viability to 10.8%, 16.9%, and 70.4 ± 1.8%, respectively, at 250 μg/mL, with corresponding IC₅₀ values of 28.3, 37.8, and >250 μg/mL, respectively. Gene expression analysis revealed upregulation of BAX and downregulation of BCL2, indicating activation of mitochondrial-mediated apoptosis. Conclusions:R. officinalis leaves represent a valuable source of multifunctional phytochemicals, particularly carnosic acid. Its broad-spectrum biological activities and apoptosis-inducing potential support its promising application in pharmaceutical, nutraceutical, and biomedical fields.