| 24 May 25 | r01    | Mito HollyHockMito HollyHoc | 0 - 1 | SagamiharaSagamihara |
| 24 May 25 | r01 | Nara ClubNara Club | 3 - 1 | Niigata University H and Niigata Unive |
| 24 May 25 | r01 | FC GifuFC Gifu | 3 - 1 | Toyama ShinjoToyama Shinjo |
| 24 May 25 | r01 | Tochigi SCTochigi SC | 5 - 0 | Oyama SCOyama SC |
| 24 May 25 | r01 | Sendai UniversitySendai Univer | 2 - 4 | Toyo UniversityToyo Universi |
| 24 May 25 | r01 | Kyoto Sangyo UniversityKyoto Sangyo | 3 - 1 | Moriyama Samurai 2000Moriyama Samu |
| 24 May 25 | r01 | Brew SagaBrew Saga | 2 - 1 | Hirondelle KumamotoHirondelle Ku |
| 24 May 25 | r01 | Veroskronos TsunoVeroskronos T | 2 - 1 | Gainare TottoriGainare Totto |
| 24 May 25 | r01 | Kamatamare SanukiKamatamare Sa | 1 - 1 | Kochi UnitedKochi United |
| 25 May 25 | r01 | Matsumoto YamagaMatsumoto Yam | 2 - 1 | FC OsakaFC Osaka |
| 25 May 25 | r01 | Okinawa SVOkinawa SV | 4 - 1 | Verspah OitaVerspah Oita |
| 25 May 25 | r01 | ImabariImabari | 0 - 2 | Kagoshima UnitedKagoshima Uni |
| 25 May 25 | r01 | Zweigen KanazawaZweigen Kanaz | 6 - 0 | Chukyo UniversityChukyo Univer |
| 25 May 25 | r01 | Arterivo WakayamaArterivo Waka | 4 - 3 | Basara HyogoBasara Hyogo |
| 25 May 25 | r01 | Thespa GunmaThespa Gunma | 1 - 0 | Hosei UniversityHosei Univers |
| 25 May 25 | r01 | ReinMeer AomoriReinMeer Aomo | 5 - 0 | BTOP HokkaidoBTOP Hokkaido |
| 25 May 25 | r01 | Fukui UnitedFukui United | 1 - 0 | Honda FCHonda FC |
| 25 May 25 | r01 | Veertien MieVeertien Mie | 1 - 0 | Yamanashi Gakuin UniversiYamanashi Gak |
| 25 May 25 | r01 | Fukushima UnitedFukushima Uni | 9 - 1 | Tokyo International UniveTokyo Interna |
| 25 May 25 | r01 | Iwate Grulla MoriokaIwate Grulla | 6 - 0 | North Asia UniversityNorth Asia Un |
| 25 May 25 | r01 | Belugarosso IwamiBelugarosso I | 0 - 1 | Giravanz KitakyushuGiravanz Kita |
| 25 May 25 | r01 | Kataller ToyamaKataller Toya | 2 - 1 | Juntendo UniversityJuntendo Univ |
| 25 May 25 | r01 | Fukuyama CityFukuyama City | 2 - 1 | FC TokushimaFC Tokushima |
| 25 May 25 | r01 | Oita TrinitaOita Trinita | 2 - 0 | Lvnirosso NCLvnirosso NC |
| 25 May 25 | r01 | Baleine ShimonosekiBaleine Shimo | 2 - 1 | International Pacific UniInternational |
| 25 May 25 | r01 | RB Omiya ArdijaRB Omiya Ardi | 0 - 1 | University of TsukubaUniversity of |
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