Introduction <p>Poor cleaning practices for healthcare workers’ mobile phones can promote microbial contamination, posing a latent risk of infections for patients. Despite widespread mobile phone use in healthcare environments, evidence on the effectiveness of disinfection methods in low and middle-income countries remains limited.</p> Objective <p>To evaluate the disinfectant capacity of 70% isopropanol on the surface of healthcare workers’ smartphones in a Peruvian hospital in 2023.</p> Methods <p>We designed a pre-post study to collect demographic, occupational, and phone usage information, along with microbiological culture analysis results before and after cleaning with 70% isopropanol.</p> Results <p>We evaluated 178 mobile phones from healthcare workers, 90.5% (95%CI: 85.1–94.0%) of which showed microbiological contamination. The most frequently isolated microorganism was <i>Staphylococcus aureus</i> (41.0%), followed by <i>Enterococcus sp</i>. (14.9%) and <i>Staphylococcus epidermidis</i> (9.9%). Antibiogram analysis revealed high resistance rates to ampicillin. No factors were associated with microbial contamination on phones. A significant reduction in microbiological contamination was observed (90.4% to 14.0%, <i>p</i> &lt; 0.001) after cleaning with 70% isopropanol. Colony counts significantly decreased from 73.4 ± 31.8 CFU/cm² to 3.9 ± 11.0 CFU/cm² (<i>p</i> &lt; 0.001).</p> Conclusion <p>The disinfectant capacity of 70% isopropanol on healthcare workers’ smartphone surfaces is highly effective, resulting in an approximately fivefold reduction in contamination rates.</p>