Current trends in scientific studies focus on the development of smartphone-based biosensors via green nanoparticle for clinical diagnosis, food, and environmental monitoring. In this study, we developed a novel portable smartphone-based biosensor via green dendrimer-coated matcha extract/silver nanoparticles (ME-Ag NPs) enriched with polyphenol for detecting hydrogen peroxide (H₂O₂). Also, we investigated the biological evaluation of the nanostructure as a safe preservative for use in biomedical applications. Ag NPs were prepared using a green sonochemical method and were characterized to determine surface and chemical properties by different techniques such as scanning electron microscopy—energy-dispersive X-ray, transmission electron microscope, Fourier transform infrared spectroscopy, atomic force microscopy, X-ray diffraction, and Brunauer–Emmett–Teller. Furthermore, antimicrobial and antifungal properties of ME-Ag NPs were investigated against pathogenic microorganisms such as Staphylococcus aureus, Pseudomonas aureginosa, Escherichia coli, Candida albicans, and Aspergillus brasiliensis. The experimental sensor methodology was based on the detection of H₂O₂ by analysis of images of novel silver nanostructure-coated papers and processing of color histograms with a RGB (red–green–blue) analyzer software. Consequently, the smartphone-based biosensor exhibited high sensitivity with detection limits of 0.82 μM response time of 5 s. The smartphone-based biosensor via ME-Ag NPs provided a rapid and selective detection of H₂O₂.