High Pressure Liquid Chromatographic Analysis of Polyphenols in Leaves of Eucalyptus camaldulensis, E. globulus and E. rudis: Proanthocyanidins, Ellagitannins and Flavonol Glycosides

Abstract

Polyphenols of adult leaves from Eucalyptus globulus, E. camaldulensis and E. rudis grown in Spain, in two areas with different edaphoclimatical characteristics, were estimated by chemical methods and by high pressure liquid chromatography. Leaf extracts consisted mainly of proanthocyanidins, ellagitannins and flavonol glycosides. Knowledge of the ellagitannins allows the discrimination of the three species, with E. globulus being well differentiated from the other two. However, the pattern of flavonol glycosides only permitted the discrimination of E. rudis with respect to E. camaldulensis and E. globulus. EXPERIMENTALSamples. Adult leaves of E. camaldulensis, E. globulus and E. rudis were collected in two Spanish arboreta. The climate at one of the sites, located in the South Western province of Huelva, is warmer and drier than that of the other, which is in the North Eastern province of Pontevedra. Five samples, each from a different tree, were collected for each species at each geographical origin, with the exception of E. camaldulensis from Pontevedra where only two trees were sampled.Extraction. Tannins were extracted from dried and ground leaf samples (10 g) with 300 mL methanol:water (80:20), at room temperature in the dark for 24 h (Conde et al., 1995). The suspensions were filtered, the methanol was removed by vacuum distillation and the aqueous solution was extracted with ethyl ether and freeze dried. The aqueous solution was used for the quantitative analysis of total phenols, proanthocyanidins and ellagitannins, whilst the lyophilized material was used for HPLC analysis after redissolving in methanol:water (1:1).Total phenol content. This was determined by the Folin-Ciocalteu method (Singleton and Rossi, 1965) using aqueous solutions of gallic acid (2-40 g/mL) as standard. The molar absorbance obtained in our analysis conditions was 22.3ϫ 10 3 /mol/cm. Proanthocyanidin content. The evaluation of proanthocyanidins was carried out using the vanillin method (Swain and Hillis, 1959). Calibration was achieved with aqueous solutions of catechin (2-40 g/mL) and a molar absorbance of 34.3ϫ 10 3 /mol/cm was obtained under our conditions. Ellagitannin content. Ellagitannins were estimated by the HPLC evaluation of the ellagic acid obtained following acid hydrolysis.

Antioxidant effects and phytochemical analysis of crude and chromatographic fractions obtained from Eucalyptus globulus bark

“…For the three assays, crude extract displayed the strongest antioxidative activity that was significantly (p<0.05) different from those of its fractions and the tested standards. Activities of all extracts are probably due to the presence of polyphenols especially hydrolyzable tannins which have been previously reported in E. globulus (Cadahia et al, 1997;Hou et al, 2000;Amakura et al, 2009). Eyles et al (2004) reported that antioxidative activity of wounded wood of E. globulus extracts is due to hydrolyzable tannins including pedunculagin, di-, tri-and tetragalloylglucose.…”

“…Phenolic compounds extraction in plant materials is influenced by their chemical nature, the extraction method employed, sample particle size and storage time (Naczk and Shahidi, 2004). As reported in the literature, major compounds of E. globulus plant are hydrolyzable tannins and flavonol derivatives (Cadahia et al, 1997;Conde et al, 1997;Atoui et al, 2005). So, we used aqueous acetone (70%) as extraction solvent which is found to be better for the extraction of tannins (Cork and Krockenberger, 1991) and flavonoids (Cowan, 1999).…”

Qualitative and Semi‐quantitative Analysis of Phenolics in Eucalyptus globulus Leaves by High‐performance Liquid Chromatography Coupled with Diode Array Detection and Electrospray Ionisation Mass Spectrometry

“…, 6 ). These flavonols derivatives have already been described in Eucalyptus species (Cadahia et al ., ; Conde et al ., ; Santos and Waterman, , ; Atoui et al ., ; Amakura et al ., ; Boulekbache‐Makhlouf et al ., ).…”

Intraspecific variation in the growth and survival of juvenile fish exposed to Eucalyptus leachate

“…Red gum leachate contains over 90 chemical compounds, some of which are toxic to aquatic organisms (Hillis ; Cadahía et al. ; Conde et al. ; Farah et al.…”