PHA Microplastic Aging Decreases N₂O Sink Capacity: Released γ-Butyrolactone Decouples Denitrifying Electron Transfer and Oxidative Phosphorylation

Bacterial denitrification is a main pathway for soil N₂O sinks, which is crucial for assessing and controlling N₂O emissions. Biobased polyhydroxyalkanoate (PHA) microplastic particles (MPs) degrade slowly in conventional environments, remaining inert for extended periods. However, the impacts of PHA microplastic aging on the bacterial N₂O sink capacity before degradation remain poorly understood. Here, the soil model strain <i>Paracoccus denitrificans</i> was exposed to 0.05-0.5% (w/w) virgin and aged PHA MPs. Although no significant changes in the molecular weights were observed, aged PHA MPs hindered cell growth and N₂O reduction rates, leading to a surge in N₂O emissions. ¹H NMR spectroscopy and UPLC-QTOF-MS analysis identified γ-butyrolactone as the key component released from aged PHA MPs. Metabolic verifications at the cellular level confirmed its inhibition on the N₂O sink and ATP synthesis. The γ-butyrolactone that protonated and hydrolyzed spontaneously in the periplasm would compete for protons with ATPase and destroy the coupling between denitrifying electron transfer and oxidative phosphorylation. Consequently, energy-deficient cells reduced the electron supply for N₂O reduction, which did not contribute to energy conservation. This work unveils a novel mechanism by which PHA microplastic aging impairs the bacterial N₂O sink and highlights the need to consider environmental risks posed by biobased microplastic aging.