| Lane | Athlete | Yr. | Team | Seed | Note |
|---|---|---|---|---|---|
| 2 | Austin, Leiana | FR | East River | 24.42 | St. Cloud Invitational 2020 |
| 3 | Perry, Rodnay | FR | East River | 21.67 | St. Cloud Invitational 2020 |
| 4 | Decker, Klarissa | SO | Freedom (Orlando) | 19.98 | Toho Invitational 2020 |
| 5 | Harris, Dayna | FR | East River | 21.34 | St. Cloud Invitational 2020 |
| 6 | Ugwuoke, Hope | SO | Timber Creek | 22.08 | TC Time Trial |
| 7 | Wilson, Hanna | SO | Freedom (Orlando) | NT |