Different in vitro cultures of Valerianaceae were analysed for valepotriate content [(iso)valtrate, acevaltrate, didrovaltrate] in a study on properties of production in vitro (plant species, growth conditions, differentiation level, valepotriate content of the medium after growth). The in vitro cultures were: callus cultures of Valeriana officinalis L., Valerianella locusta L. and Centranthus ruber L.DC.; a suspension culture of Valeriana officinalis L. and a root organ culture of Centranthus ruber L.DC. All of the cultures produced valepotriates in vitro in different amounts. None of the media that had served for growth contained any valepotriates. In order to characterize the in vitro growth more precisely different parameters were analysed at different time intervals during a growth period in one of the cultures (callus culture of Valeriana officinalis L.). These different parameters were: fresh and dry weight, lipid and nitrogen content and (iso)valtrate content. This study during a growth period was performed on two media differing in plant hormone content.