Functional Groups in Pharmaceutical Contaminants Reshape Sludge Fermentation Pathways: Substrate Selectivity and Microbial Stress Responses

Abstract

The post-COVID-19 era has led to a dramatic increase in the consumption of respiratory pharmaceuticals, leading to their inevitable accumulation in waste-activated sludge. Although the impact of these contaminants on anaerobic resource recovery has been recognized, the specific contribution of their functional groups, which are fundamental determinants of chemical reactivity, remains largely overlooked. This study challenges traditional generalized assessments by using two structural analogues, hydroxyl-functionalized ambroxol hydrochloride (AMB) and tertiary amine-functionalized bromhexine hydrochloride (BRO), to reveal the role of functional groups in altering volatile fatty acid (VFA) production. AMB demonstrated a biphasic dose response, whereas BRO showed a relatively stable enhancement pattern, reflecting distinct fermentation profiles driven by functional group variation. The hydroxyl group of AMB exhibited a low promotion and high inhibition dose response, wherein this moiety interacted with proteins via hydrogen bonds, enriching proteolytic bacteria. However, excessive accumulation induced severe oxidative stress, forcing microorganisms to shift their energy from acidogenesis to multidrug efflux pumps for survival. Conversely, the tertiary amine group of BRO drove a continuous and monotonic enhancement of the VFA yields. This group targeted polysaccharides through hydrophobic interactions, enriching the glycolytic bacteria. Consequently, upregulated carbohydrate metabolism provided key precursors for membrane repair, thereby sustaining metabolic activity, even under high stress.