This record contains raw data related to Low-Intensity Pulsed Ultrasound: a new gut liquid biopsy approach to predict disease activity in Ulcerative Colitis Endoscopy coupled to biopsy histological analysis remains the current gold standard for diagnosis and assessment of disease activity in ulcerative colitis (UC). This study explored the potential of Low-Intensity Pulsed Ultrasound (LIPUS) to enhance the release of mucosal miRNAs carried by extracellular vesicles (EVs), as non-invasive biomarkers for UC disease activity. Utilizing a simulation framework to translate in-vivo the optimized energy dosage determined in-vitro, we targeted the colon of healthy and colitic mice, causing no significant lesions macroscopically or microscopically after LIPUS treatment. LIPUS stimulation (38kHz, 150mW/cm², 180s) effectively increased miRNA release, reflecting both acute and chronic inflammatory states in experimental colitis models. Ex-vivo LIPUS stimulation of UC patient gut biopsies not only confirmed its ability to enhance EV release, but also revealed nine miRNAs (let-7g-5p, miR151a-5p, miR-30b-5p, miR-15b-5p, miR-296-5p, miR-339-5p, miR-4291, miR-200a-3p, miR-204a-3p) reflective of the mucosal status, strongly correlating with partial and/or endoscopic Mayo score, that were undetectable in patient’s blood samples. Overall, these findings suggest that LIPUS is a safe, non-invasive method for enriching tissue-derived EV-miRNA biomarkers in blood, offering a promising approach for gut liquid biopsy and UC disease activity monitoring, thus avoiding invasive procedures.