Despite their threatens for Egyptian stone monuments, A few studies focused on using biocontrol agents against deteriorative fungi and bacteria instead of using chemical assays that leave residuals leading to human toxicity and environmental pollution. This work aims to isolate and identify fungal and bacterial isolates that showed deteriorative activities from stone monuments in Temple of Hathor, Luxor, Egypt, as well as determine the inhibitory activity of metabolites produced byStreptomyces exfoliatusSAMAH 2021 against the identified deteriorative fungal and bacterial strains. Moreover, studying the spectral analysis, toxicological assessment of metabolites produced byS. exfoliatusSAMAH 2021 against health human cell fibroblast, and colorimetric measurements on the selected stone monuments. Ten samples were collected from Temple of Hathor, Luxor, Egypt. Three fungal isolates and one bacterial isolate were obtained and identified asA. nigerisolate Hathor 2,C. fioriniaestrain Hathor 3,P. chrysogenumstrain HATHOR 1, andL. sphaericusstrain Hathor 4, respectively. Inhibitory potential of the metabolites in all concentrations used (100–25%) against the recommended antibiotics (Tetracycline 10 µg/ml and Doxycycline (30 µg/ml) showed an inhibitory effect toward all tested deteriorative pathogens with a minimum inhibition concentration (MIC) of 25%. Cytotoxicity test confirmed that microbial filtrate as the antimicrobial agent was safe for healthy human skin fibroblast with IC50of < 100% and cell viability of 97%. Gas chromatography analysis recorded the existence of thirteen antimicrobial agents, Cis-vaccenic acid; 1,2-Benzenedicarboxylic acid; ç-Butyl-ç-butyrolactone and other compounds. Colorimetric measurements confirmed no color or surface change for the limestone-treated pieces. The use of the metabolite of microbial species antimicrobial as a biocontrol agent raises contemporary issues concerning the bio-protection of the Egyptian monuments to reduce chemical formulas that are toxic to humans and pollute the environment. Such serious problems need further investigation for all kinds of monuments.