The content of GLC and FRU in leaves was evaluated by measuring the NADPH absorption after successive additions of the coupling enzymes glucose-6-P-dehydrogenase, hexokinase, phosphoglucose-isomerase and invertase  using a UV/AZD6738 solubility dmso visible spectrophotometer (Tecan GENios Microplate Reader, Männedorf, Switzerland) at 340 nm. AA was estimated by a colorimetric Alvespimycin cost 2.6-dichlorophenol-indophenol (DIP) method . The AA content was estimated using a UV/visible spectrophotometer (Novaspec II, Pharmacia Biotech AB, Uppsala, Sweden) at 520 nm. CA content was determined by measuring the NADH oxidation after addition of l-malate dehydrogenase, l-lactate dehydrogenase, oxaloacetate and pyruvate 
using a UV/visible HDAC inhibitor spectrophotometer (Novaspec II, Pharmacia Biotech AB, Uppsala, Sweden) at 340 nm. Finally, according to Marinova et al. , PP leaf content was determined following a modified Folin-Ciocalteu method . After incubation, the absorbance of the leaf extracts was determined using a UV/visible spectrophotometer (Novaspec
II, Pharmacia Biotech AB, Uppsala, Sweden) at 750 nm. The enzymatic test kit was purchased from R-Biopharm AG (Darmstadt, Germany). Data analysis Plants were arranged in a randomized design (nine plants per species per treatment, one plant per pot). One-way analysis of variance (ANOVA) was carried out to test the differences in the plants’ behaviour. The statistical significance of differences between mean values was determined using Bonferroni’s test (p < 0.05). Different letters in Tables 1 and 2 are used to indicate means that were statistically different at p < 0.05. Statistical analysis was performed using the SPSS program (ver. 17, SPSS Inc.,
Chicago, IL, USA). Table 1 Concentration of Ag in the roots, stems and leaves of the plants and Ag TF Species Ag roots Ag stem Ag leaves Translocation factor Inositol monophosphatase 1 (mg kg−1 DW) (mg kg−1 DW) (mg kg−1 DW) (× 100) Brassica juncea 82,292 a 57,729 a 6,156 a 7.48 a (5,394) (598) (516) (0.92) Festuca rubra 62,365 b 2,777 c 2,459 b 3.94 b (1,990) (2,738) (258) (0.36) Medicago sativa 19,715 c 25,241 b 4.31 c 0.022 c (2,369) (5,004) (0.84) (0.003) The means (n = 3) with the same letter were not significantly different (Bonferroni’s test; p < 0.05). The mean standard error (n = 3) is in brackets. TF, translocation factor; DW, dry weight. Table 2 Content of GLC, FRU, AA, CA and PP in the leaves of the plants Species GLC FRU AA CA PP (mmol kg−1 FW) (mmol kg−1 FW) (mg kg−1 DW) (mg kg−1 DW) (mg GA Eq. 100 g−1 DW) Brassica juncea 1.61 b 2.17 b 3,878 a 10.2 a 711 a (0.64) (1.07) (548) (0.48) (48.6) Festuca rubra 70.4 a 57.8 a 119 c 11.2 a 580 b (12.9) (14.7) (92.4) (2.59) (37) Medicago sativa 8.17 b 7.37 b 1459 b 5.12 a 528 b (0.58) (0.57) (359) (1.68) (18.9) The means (n = 3) with the same letter were not significantly different (Bonferroni’s test; p < 0.05). The mean standard error (n = 3) is in brackets.