| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 3 | East Ridge | A | NT | |
| 4 | Windermere Prep | A | DNS | |
| 5 | Cornerstone Charter | A | NT |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 2 | Lake Buena Vista | A | 44.95 | FHSAA 2A Region 2 |
| 3 | Freedom (Orlando) | A | 44.35 | 44.35 Metro East Conference Championships |
| 4 | Foundation Academy | A | 43.48 | FHSAA Class 1A Track & Field State Championships |
| 5 | Orlando Christian Prep | A | 43.76 | 43.76 Highlander Classic 2022 |
| 6 | West Oaks | A | 44.83 | |
| 7 | Mount Dora Christian | A | 45.00 |