Despite ethanol's penetration into urban transportation, observational evidence quantifying the consequence for the atmospheric particulate burden during actual, not hypothetical, fuel-fleet shifts, has been lacking. Here we analyze aerosol, meteorological, traffic, and consumer behavior data and find, empirically, that ambient number concentrations of 7–100-nm diameter particles rise by one-third during the morning commute when higher ethanol prices induce 2 million drivers in the real-world megacity of São Paulo to substitute to gasoline use (95% confidence intervals: +4,154 to +13,272 cm −3). Similarly, concentrations fall when consumers return to ethanol. Changes in larger particle concentrations, including US-regulated PM2.5, are statistically indistinguishable from zero. The prospect of increased biofuel use and mounting evidence on ultrafines' health effects make our result acutely policy relevant, to be weighed against possible ozone increases. The finding motivates further studies in real-world environments. We innovate in using econometrics to quantify a key source of urban ultrafine particles.