Variation in the contents of pseudohypericin and hypericin in Hypericum perforatum from Lithuania Edita Bagdonait _ e a , Pavol Mártonfi b , Miroslav Rep  cák b , Juozas Labokas a, * a Institute of Botany, Nature Research Centre,  Zaliu ˛ ju ˛ E zeru ˛ g. 49, Vilnius LT-08406, Lithuania b P. J.  Safárik University, Faculty of Science, Institute of Biology and Ecology, Department of Botany, Mánesova 23, SK-04154 Ko sice, Slovakia article info Article history: Received 17 May 2010 Accepted 8 August 2010 Keywords: HPLC Hypericum perforatum Hypericin Pseudohypericin Total hypericins abstract Hypericin and hypericin-like substances are considered the main active compounds in Hypericum perforatum L. (Hypericaceae). In this work pseudohypericin and hypericin of H. perforatum collected in Lithuania were quantified. Studies on accumulation dynamics and between-accession variation of the contents of these secondary metabolites were carried out by high performance liquid chromatography (HPLC). The data were statistically pro- cessed with ANOVA and PCA. Significant difference between pseudohypericin and hypericin content in floral budding and full flowering stages was detected. The highest amounts of the secondary metabolites were observed in the flowering stage. The study revealed evident within population variations in H. perforatum. Mean concentrations of pseudohypericin and hypericin among accessions varied from 3.45 to 6.82 mg/g and from 1.17 to 2.59 mg/g, respectively. Accessions of H. perforatum showed remarkable differences in chemical composition depending on the provenance of plants. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction St. John’s wort (Hypericum perforatum L., Hypericaceae), a perennial herbaceous plant native to Europe, has been the focus of many studies due to its use as a natural medicine. The crude drug Hyperici herba consists of the aerial parts of H. perforatum. Extracts of this drug used in therapy are prepared by extracting the upper aerial parts of the plant using ethanol:water or methanol:water mixtures (Avato and Guglielmi, 2004). The plant and the extract of H. perforatum contain a wide range of compounds (Poutaroud et al., 2001), but are evaluated mainly by the contents of hypericin and pseudohypericin. Their hydrated proto-forms, protohypericin and protopseudohypericin, occur as the biosynthetic precursors which are subse- quently transformed into the cyclic compounds by exposure to light. The protopseudohypericin, pseudohypericin, proto- hypericin and hypericin, altogether called naphthodianthrones, are active ingredients of H. perforatum and are commonly calculated as total hypericins, remain as popular marker substances and are used as a quality control measure in standardized extracts and phytopharmaceuticals (Schmidt, 2003). The quality and quantity of hypericins in H. perforatum depend on various factors. The contents of hypericin vary significantly in plants collected from different locations and depend on plant development stage too. It has been reported that H. perforatum grown in different regions of Bulgaria (Kitanov, 2000), Italy (Pietta et al., 2001), India (Verma et al., 2008), Greece (Gioti et al., 2009) and Tunisia (Hosni et al., 2010) showed observable variations in hypericin concentrations. Pop- ulations of H. perforatum in geographically separated regions synthesize different amounts of the major bioactive compounds, probably, due to the specific environments and/or genetic variations of the plants. Southwell and Bourke (2001) observed the variation with the season in the concentration of total hypericins of H. perforatum. They reported that the concentrations of * Corresponding author. Tel.: þ370 52729930; fax: þ370 52729950. E-mail address: juozas.labokas@botanika.lt (J. Labokas). Contents lists available at ScienceDirect Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco 0305-1978/$ – see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2010.08.005 Biochemical Systematics and Ecology 38 (2010) 634–640