This study investigates the influence of hyporheic exchange on dissolved organic matter (DOM) dynamics in two contrasting 100-m stream reaches within the Isère catchment, French Alps Piedmont. One reach (Correncon, Disconnected) exhibited no significant hyporheic exchange due to semi-permeable silty sand sediment, while the other (La Fauge, Connected) had a relative connectivity of 0.49 ± 0.014, indicating active hyporheic flow through permeable clean sand. At 25 evenly spaced transects, hyporheic wells (1.6 cm PVC, 0.4 cm mesh, 25 cm depth) were installed and sampled over five days. A conservative tracer (NaCl) was added upstream to quantify hyporheic exchange via electrical conductivity (EC) dynamics. Surface and hyporheic water samples were collected at background and plateau conditions, with Dissolved Organic Carbon (DOC) measured via TOC analyzer and DOM composition assessed using excitation-emission matrices (EEMs). EEMs were corrected for instrument bias, inner filter effects, and calibrated to Raman Units. Spectroscopic metrics—fluorescence index (FI), biological index (BIX), and humification index (HIX)—were calculated to assess DOM source, age, and degree of humification. Data include EC time series, DOC concentrations, EEMs, and derived DOM metrics.