The women's 400 metres event at the 2000 World Junior Championships in Athletics was held in Santiago, Chile, at Estadio Nacional Julio Martínez Prádanos on 17, 18 and 20 October.
| Gold | Jana Pittman | |
| Silver | Aneta Lemiesz | |
| Bronze | Norma González |
20 October
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| align=left | align=left | | 52.45 | ||||
| align=left | align=left | | 52.78 | ||||
| align=left | align=left | | 53.30 | ||||
| 4 | align=left | align=left |  | 53.46 | |||
| 5 | align=left | align=left |  | 53.58 | |||
| 6 | align=left | align=left |  | 53.69 | |||
| 7 | align=left | align=left |  | 53.95 | |||
| 8 | align=left | align=left |  | 54.32 |
18 October
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | | 53.00 | Q | ||
| 2 | align=left | align=left | | 53.09 | Q | ||
| 3 | align=left | align=left | | 53.38 | Q | ||
| 4 | align=left | align=left | | 53.61 | Q | ||
| 5 | align=left | align=left | | 53.64 | |||
| 6 | align=left | align=left | 53.86 | ||||
| 7 | align=left | align=left | | 54.29 | |||
| 8 | align=left | align=left | | 56.09 |
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | | 53.42 | Q | ||
| 2 | align=left | align=left | | 53.69 | Q | ||
| 3 | align=left | align=left | | 53.79 | Q | ||
| 4 | align=left | align=left | | 54.02 | Q | ||
| 5 | align=left | align=left | 54.27 | ||||
| 6 | align=left | align=left | | 54.73 | |||
| 7 | align=left | align=left | | 54.78 | |||
| 8 | align=left | align=left | 55.07 |
17 October
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | | 54.01 | Q | ||
| 2 | align=left | align=left | | 54.14 | Q | ||
| 3 | align=left | align=left | | 54.38 | Q | ||
| 4 | align=left | align=left | 54.72 | q | |||
| 5 | align=left | align=left |  | 56.52 | |||
| 6 | align=left | align=left | 57.09 |
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | | 53.79 | Q | ||
| 2 | align=left | align=left | | 53.82 | Q | ||
| 3 | align=left | align=left | 53.85 | Q | |||
| 4 | align=left | align=left | | 53.88 | q | ||
| 5 | align=left | align=left |  | 55.12 | |||
| 6 | align=left | align=left |  | 55.40 | |||
| 7 | align=left | align=left |  | 61.21 | |||
| align=left | align=left | | DQ |
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | 53.91 | Q | |||
| 2 | align=left | align=left | | 54.02 | Q | ||
| 3 | align=left | align=left | | 54.20 | Q | ||
| 4 | align=left | align=left | | 54.25 | q | ||
| 5 | align=left | align=left | 56.33 |
| Rank | Name | Nationality | Time | Notes | |||
|---|---|---|---|---|---|---|---|
| 1 | align=left | align=left | | 54.27 | Q | ||
| 2 | align=left | align=left | | 54.36 | Q | ||
| 3 | align=left | align=left | | 54.63 | Q | ||
| 4 | align=left | align=left | | 54.82 | q | ||
| 5 | align=left | align=left | 55.49 | ||||
| 6 | align=left | align=left | 56.51 | ||||
| 7 | align=left | align=left | 57.73 |
According to an unofficial count, 26 athletes from 18 countries participated in the event.