T. grandiflorum favors well-drained, neutral to slightly acid soils, usually in second- or young-growth forests. In the Northern parts of its range it shows an affinity for maple or beech forests, but has also been known to spread into nearby open areas. Depending on geographical factors, it flowers from late April to early June, just after T. erectum. Like many forest perennials, T. grandiflorum is a slow growing plant. Its seeds require double dormancy, meaning they normally take at least two years to fully germinate. Like most species of Trillium, flowering age is determined largely by the surface of the leaf and volume of the rhizome the plant has reached instead of age alone. Because growth is very slow in nature, T. grandiflorum typically requires seven to ten years in optimal conditions to reach flowering size, which corresponds to a minimum of 36 cm2 (5.6 sq in) of leaf surface area and 2.5 cm3 (0.15 cu in) of rhizome volume. In cultivation, however, wide disparity of flowering ages are observed.
Due to the popularity of Trillium grandiflorum as a garden specimen, conservation concerns have been raised as the vast majority of plants sold in commercial nurseries are believed to be collected from the wild. Indeed, there is little indication of any commercial nursery growth. Frederick and Roberta Case, botanists who specialize in trilliums, wrote in 1997, “to our knowledge, no true commercial quantity 'propagation' takes place at the present time.” Such heavy collecting, combined with other pressures such as habitat destruction and grazing, may effectively endanger the plants in some areas. Trillium grandiflorum is legally listed as vulnerable in Quebec, primarily due to habitat destruction as the plant is found in forests neighboring the province's most populous regions. It is listed as exploitably vulnerable in the state of New York, meaning that it might become threatened should certain factors not be mitigated. In Maine, it is listed as endangered, and might have been extirpated entirely.