| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 1 | East River | A | Tyghter, Mykaela | Jackson, Jada | Feliciano, Alexcia | Sanders, Enijah | DNS | |
| 2 | Cypress Creek | A | Howard, Dezmi | Rivera, Nyasia | Mlodzinski, Karissa | Frazer, Ariana | 51.56 | |
| 3 | First Academy (Orlando) | A | Cook, Nia | Martin, Oliva | Wilson, Jayde | Are, Nathalie | 50.00 | |
| 4 | Boone | A | Outler, Reathasia | Jordan, Sophia | Strange, Sarah | Lee, Javia | 48.31 | FHSAA 4A Region 1 |
| 5 | Lake Mary | A | Powell, Jessiah | Faulkner, Kayla | Cole, Courtney | Bradley, Cymiah | 48.79 | FHSAA 4A Region 1 |
| 6 | Winter Park | A | Bernbaum, Georgia | Day, Jani | Parrish, Emily | Devlin, Alexis | 50.50 | |
| 7 | Lake Nona | A | Slater, Arianna | Ellis, Kathleen | Duplan, Brianna | Jacobs, Brianna | 53.73 | Tohopekaliga Tigers Invitational |
| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 3 | Timber Creek | A | Archibald-Bowers, Giselle | Estrella, Emily | Henderson, Savannah | Estrella, Kiara | NT | |
| 4 | Lake Highland Prep | A | Johnson, Macy | Wheeler, Lana | Brown, Caroline | Bines, Carly | NT | |
| 5 | Master's Academy (Oviedo) | A | Falco, Ana | Fyock, Ashley | Wilson, Ryleigh | Wilson, Alexa | DQ |