<p><b>BACKGROUND: </b>Physical activity fragmentation represents the frequency of transitioning from an active to sedentary state. The prognostic information of physical activity fragmentation is unclear in Hispanics/Latinos. This study examined the association of PA fragmentation with all-cause mortality in Hispanic/Latino adults.</p><p><b>METHODS: </b>We investigated 11,992 participants from the Hispanic Community Health Study/Study of Latinos (HCHS/SOL) (18-74 yr; 52.2% women), from four United States urban communities (Bronx, New York; Chicago, Illinois; Miami, Florida; San Diego, California), that wore an accelerometer for one week. Physical activity fragmentation was calculated using the active-to-sedentary transition probability (ASTP) as the reciprocal of the average active bout duration. Daily total log-transformed activity count (TLAC) was used as a measure of total physical activity. The residual of ASTP regressed on TLAC (TLAC-adjusted ASTP) was explored to investigate the association of ASTP independent of total physical activity. Deaths were identified from annual follow-up interviews, obituary searches, or matches to the National Death Index through December 31, 2021. Cox regression models were fitted according to physical activity fragmentation.</p><p><b>FINDINGS: </b>There were 745 deaths (6.2%) over a mean follow-up of 11.2 (SD 2.2) years. The highest compared to the lowest tertile of ASTP showed a HR of 1.45 (95% CI 1.10-1.92) of all-cause mortality after accounting for confounders. The mortality risk also increased for each 0.10-unit increase of ASTP, as a continuous variable, by 22% (HR 1.22; 95% CI 1.07-1.39). The results were similar considering TLAC-adjusted ASTP.</p><p><b>INTERPRETATION: </b>Among Hispanic/Latino adults, more fragmented physical activity was associated with elevated all-cause mortality, independent of total physical activity volume.</p><p><b>FUNDING: </b>HCHS/SOL was supported by the National Institutes of Health.</p>