Ashton demanded an immediate release of those persons "being illegally detained by armed groups," Xinhua reported.
Ashton also asked Russia to "take concrete action in support of the Geneva accord," which set out key steps to de-escalate tensions and restore security.
"I notably condemn the shooting of Kharkiv's mayor Henadiy Kernes Monday and the detention of military observers from the Organization for Security and Cooperation in Europe (OSCE) participating states in Sloviansk by armed separatists since last week," Ashton added.
Ashton made the remarks after the EU imposed travel bans and assets freeze on 15 Russian officials ealr Tuesday.
Russia criticised the new EU sanctions, saying the 28-nation bloc should be ashamed of itself.