Emerging investigator series: Phthalate esters are present at elevated concentrations in floor dust, and resuspension of dust represents a major source for human exposure to chemicals. Biodegradation of phthalates occurs in aquatic systems and soils but has not been demonstrated in house dust. The goal of this study was to quantify indoor phthalate ester degradation through both biotic and abiotic mechanisms. Worn carpet squares were embedded with dust and incubated for one to six weeks at equilibrium relative humidity (ERH) levels of 50, 80, 85, 90, 95, and 100%, and nine phthalates were measured. Removal was observed for DEHP, BBzP, DINP, DiDP, and DMP (p < 0.05) when incubated under elevated relative humidity conditions. Abiotic and biotic losses were examined separately using dust spiked with deuterated di(2-ethylhexyl)phthalate (d-DEHP) that was embedded in carpet and incubated at 100% ERH. Abiotic processes resulted in a 10.1% (±1.1%, standard error) to 69.6% (±4.8%) decrease in total d-DEHP after one week (p = 0.03) and a 27.2% (±1.4%) to 52.0% (±2.1%) decrease after three weeks (p = 0.008). Biodegradation resulted in a decrease in total d-DEHP after one week, ranging from 5.9% (±8.9%) to 8.5% (±1.7%) (p = 0.07) and a 1.7% (±3.9%) to 10.3% (±4.5%) decrease after three weeks (p = 0.044). Metatranscriptomic-based analysis indicates that fungi found in carpet dust express genes capable of degrading phthalate esters via various biochemical processes (including β-oxidation and hydrolysis). Overall, these results support the hypothesis that phthalate losses in floor dust are due to a combination of abiotic and microbial degradation at ≥80% ERH.