Background: The intensive data typically collected by mobile health (mHealth) apps allows factors associated with persistent use to be investigated, which is an important objective given users' well-known struggles with sustaining healthy behavior., Objective: Data from a commercial meditation app (n=14,879; 899,071 total app uses) were analyzed to assess the validity of commonly given habit formation advice to meditate at the same time every day, preferably in the morning., Methods: First, the change in probability of meditating in 4 nonoverlapping time windows (morning, midday, evening, and late night) on a given day over the first 180 days after creating a meditation app account was calculated via generalized additive mixed models. Second, users' time of day preferences were calculated as the percentage of all meditation sessions that occurred within each of the 4 time windows. Additionally, the temporal consistency of daily meditation behavior was calculated as the entropy of the timing of app usage sessions. Linear regression was used to examine the effect of time of day preference and temporal consistency on two outcomes: (1) short-term engagement, defined as the number of meditation sessions completed within the sixth and seventh month of a user's account, and (2) long-term use, defined as the days until a user's last observed meditation session., Results: Large reductions in the probability of meditation at any time of day were seen over the first 180 days after creating an account, but this effect was smallest for morning meditation sessions (63.4% reduction vs reductions ranging from 67.8% to 74.5% for other times). A greater proportion of meditation in the morning was also significantly associated with better short-term engagement (regression coefficient B=2.76, P<.001) and long-term use (B=50.6, P<.001). The opposite was true for late-night meditation sessions (short-term: B=-2.06, P<.001; long-term: B=-51.7, P=.001). Significant relationships were not found for midday sessions (any outcome) or for evening sessions when examining long-term use. Additionally, temporal consistency in the performance of morning meditation sessions was associated with better short-term engagement (B=-1.64, P<.001) but worse long-term use (B=55.8, P<.001). Similar-sized temporal consistency effects were found for all other time windows., Conclusions: Meditating in the morning was associated with higher rates of maintaining a meditation practice with the app. This is consistent with findings from other studies that have hypothesized that the strength of existing morning routines and circadian rhythms may make the morning an ideal time to build new habits. In the long term, less temporal consistency in meditation sessions was associated with more persistent app use, suggesting there are benefits from maintaining flexibility in behavior performance. These findings improve our understanding of how to promote enduring healthy lifestyles and can inform the design of mHealth strategies for maintaining behavior changes., (©Vincent Berardi, Rylan Fowers, Gavriella Rubin, Chad Stecher. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 10.04.2023.)