| Lane | Athlete | Team | Seed | Note |
|---|---|---|---|---|
| 3 | MOREIRA DE SOUZA, Isabella | Lake Nona |
NT | |
| 4 | CHAN, Taylor | Dalton School | NT | |
| 5 | HICKS, Aaliyah | Lake Nona |
NT |
| Lane | Athlete | Team | Seed | Note |
|---|---|---|---|---|
| 1 | DASSORI, Lillian | Dalton School | 1:07.00 | |
| 2 | GUITTON, Esme | Winter Park |
1:04.53 | Timber Creek Elite Classic 2025 |
| 3 | BROWN, Acasia | Ocoee |
59.23 | |
| 4 | VALLE, Alexie | Timber Creek |
55.87 | Timber Creek Elite Classic 2025 |
| 5 | THOMPSON, Tailin | Timber Creek |
57.80 | 2025 The Ellis Elite 16 Track and Field Invitational |
| 6 | CAMPOS, Ana | Winter Park |
1:04.38 | Timber Creek Elite Classic 2025 |
| 7 | RICHEMOND, Rickny | Ocoee |
1:06.93 | Ocoee Invitational (Meet is Full) |
| 8 | NEMO, Avery | Riverdale Country School | 1:15.00 |