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Hydraulic fracturing, or fracking, requires large amounts of water to extract fossil fuel from rock formations. As a result of hydraulic fracturing, the briny wastewater, often termed back-produced fracturing or fracking water (FW), is pumped into holding ponds. One of the biggest challenges with produced water management is controlling microbial activity that could reduce the pond water’s reusable layer and pose a significant environmental hazard. This study focuses on the characterization of back-produced water that has been hydraulically fractured using chemical and biological analysis and the development of a high-throughput screening method to evaluate and predict the antimicrobial effect of four naturally and commercially available acidic inhibitors (edetic acid, boric acid, tannic acid, and lactic acid) on the growth of the FW microbiome. Liquid cultures and biofilms of two laboratory model strains, the vegetative Escherichia coli MG1655, and the spore-forming Bacillus atrophaeus (also known as Bacillus globigii, BG) bacteria, were used as reference microorganisms. Planktonic bacteria in FW were more sensitive to antimicrobials than sessile bacteria in biofilms. Spore-forming BG bacteria exhibited more sensitivity to acidic inhibitors than the vegetative E. coli cells. Organic acids were the most effective bacterial growth inhibitors in liquid culture and biofilm.