AimTo evaluate an antibiotic inactivation strategy to protect the gut microbiome from antibiotic-mediated damage. Methods and ResultsSYN-004 (ribaxamase) is an orally delivered beta-lactamase intended to degrade penicillins and cephalosporins within the gastrointestinal tract to protect the microbiome. Pigs (20kg, n=10) were treated with ceftriaxone (CRO) (IV, 50mgkg(-1), SID) for 7days and a cohort (n=5) received ribaxamase (PO, 75mg, QID) for 9days beginning the day before antibiotic administration. Ceftriaxone serum levels were not statistically different in the antibiotic-alone and antibiotic+ribaxamase groups, indicating ribaxamase did not alter systemic antibiotic levels. Whole-genome metagenomic analyses of pig faecal DNA revealed that CRO caused significant changes to the gut microbiome and an increased frequency of antibiotic resistance genes. With ribaxamase, the gut microbiomes were not significantly different from pretreatment and antibiotic resistance gene frequency was not increased. ConclusionRibaxamase mitigated CRO-mediated gut microbiome dysbiosis and attenuated propagation of the antibiotic resistance genes in pigs. Significance and Impact of the StudyDamage of the microbiome can lead to overgrowth of pathogenic organisms and antibiotic exposure can promote selection for antibiotic-resistant micro-organisms. Ribaxamase has the potential to become the first therapy designed to protect the gut microbiome from antibiotic-mediated dysbiosis and reduce emergence of antibiotic resistance.