title: Phytochemical investigations and antiproliferative secondary metabolites from Thymus alternans growing in Slovakia.
abstract: BACKGROUND
  Thymus alternans Klokov (Lamiaceae) is a neglected species of the genus Thymus (Sect. Serpyllum) endemic to Carpathian area, where it is used as a flavouring agent and for medicinal purposes.
  
  
  OBJECTIVE
  The aim of the work was to identify antiproliferative constituents from the flowering aerial parts of this plant.
  
  
  METHODS
  Thymus alternans extracts were analyzed by HPLC-MSnand subjected to extensive chromatographic separations. The isolated compounds (phenolics and triterpenes) were structurally elucidated by MS and 1D and 2D NMR experiments. Essential oil (EO) composition was determined by GC-FID and GC-MS. Six purified triterpenes and EO were assayed for in vitro antiproliferative activity against a panel of human cancer cells, namely, breast (MDA-MB 231), colon (HCT-15 and HCT116), lung (U1810), pancreatic (BxPC3), melanoma (A375) and cervical carcinoma (A431) cells.
  
  
  RESULTS
  The structures of the isolated compounds were achieved on the basis of H-NMR and MS experiments. Luteolin-4'-O-β-d-glucopyranoside (P1), chrysoeriol-7-O-β-d-glucopyranoside (P2), chrysoeriol-5-O-β-d-glucopyranoside (P3), apigenin-7-O-β-d-glucopyranoside (P4), rosmarinic acid (P5), rosmarinic acid-3'-O-β-d-glucopyranoside (P6), caffeic acid-3-O-β-d-glucopyranoside (P7), 3α-hydroxy-urs-12,15-diene (T1), α-amyrin (T2), β-amyrin (T3), isoursenol (T4), epitaraxerol (T5), and oleanolic acid (T6). GC-MS analysis revealed that the EO of T. alternans was devoid of phenols and belonged to the nerolidol-chemotype, that is typical of the Sect. Serpyllum. The six purified triterpenes (T1-T6) were active with IC50ranging from 0.5 to 5 μM being comparable or better than those of reference compounds betulinic acid and cisplatin. The EO exhibited significant effects on A375, MDA-MB 231 and HCT116 cell lines with IC50in the range of 5-8 μg/mL.
  
  
  CONCLUSIONS
  The reported results suggest that T. alternans can be considered as a good source of phytoconstituents with possible importance in the pharmaceutical field.
authors.affiliation: a Department of Pharmaceutical and Pharmacological Sciences , University of Padova , Padova , Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Dall'Acqua
authors.affiliation: a Department of Pharmaceutical and Pharmacological Sciences , University of Padova , Padova , Italy.
authors.forename: Gregorio
authors.initials: G
authors.lastname: Peron
authors.affiliation: a Department of Pharmaceutical and Pharmacological Sciences , University of Padova , Padova , Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Ferrari
authors.affiliation: a Department of Pharmaceutical and Pharmacological Sciences , University of Padova , Padova , Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Gandin
authors.affiliation: b School of Pharmacy , University of Camerino , Camerino , Italy.
authors.forename: Massimo
authors.initials: M
authors.lastname: Bramucci
authors.affiliation: b School of Pharmacy , University of Camerino , Camerino , Italy.
authors.forename: Luana
authors.initials: L
authors.lastname: Quassinti
authors.affiliation: c Department of Botany, Institute of Biology and Ecology , P. J. Šafárik University , Košice , Slovakia.
authors.forename: Pavol
authors.initials: P
authors.lastname: Mártonfi
authors.affiliation: b School of Pharmacy , University of Camerino , Camerino , Italy.
authors.forename: Filippo
authors.initials: F
authors.lastname: Maggi
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000302:isolation & purification / Q000494:pharmacology*; D049109:Cell Proliferation / Q000187:drug effects*; D002851:Chromatography, High Pressure Liquid; D002945:Cisplatin / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D008401:Gas Chromatography-Mass Spectrometry; D045325:HCT116 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D009369:Neoplasms / Q000188:drug therapy* / Q000473:pathology; D009822:Oils, Volatile / Q000302:isolation & purification / Q000494:pharmacology*; D053978:Pentacyclic Triterpenes; D064209:Phytochemicals / Q000302:isolation & purification / Q000494:pharmacology*; D008517:Phytotherapy; D035261:Plant Components, Aerial; D010936:Plant Extracts / Q000302:isolation & purification / Q000494:pharmacology*; D010938:Plant Oils / Q000302:isolation & purification / Q000494:pharmacology*; D010946:Plants, Medicinal; D018154:Slovakia; D046930:Thymus Plant / Q000737:chemistry* / Q000254:growth & development; D014315:Triterpenes / Q000494:pharmacology; D000094062:Betulinic Acid
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D009822:Oils, Volatile; D053978:Pentacyclic Triterpenes; D064209:Phytochemicals; D010936:Plant Extracts; D010938:Plant Oils; D014315:Triterpenes; D002945:Cisplatin; D000094062:Betulinic Acid
keywords: GC-MS; HPLC-MS; Triterpene; essential oil; flavonoids