Supplementary Methods 2: Chemical analysis of total phenolics and total triterpenoid
Leaves were flash frozen in liquid nitrogen and stored at -80°C for analysis. Total phenolic concentration was calculated with the modified Folin-Ciocalteau method (Singleton & Rossi, 1964). Briefly, an aliquot of 1 mL of free phenolic acid extract was added to 5 mL of water, 1.0 mL Folin-Ciocalteau reagent, and 3 ml of saturated sodium carbonate solution (7.5%) and mixed in a screw-top test tube. The absorbance was measured at 765 nm with a spectrophotometer, after incubation for 2 h at room temperature. Quantification was based on the standard curve, established with 0, 50, 100, 150, 200 and 250 mg/L of gallic acid, and the results were expressed as gallic acid equivalent in milligrams per gram dry weight (mg GAE/g DW). Total triterpenoid concentration was determined via vanillin-glacial acetic acid-perchloric acid spectrophotometry using oleanolic acid as the standard (Fan & He, 2006). Briefly, total triterpenoids were extracted by ultrasonic extraction using anhydrous ethanol as a solvent. Oleanolic acid was used as the reference, 5% vanillin-acetic acid glacial solution and perchlorate acid were used as the chromogenic agent, followed by water bath at 70°C for 15min, and the absorbance was measured at 560 nm. Because the amount of one leaf was not sufficient for each chemical analysis, we gathered leaves from six plants as a biological replication. There were 12 replicates for each combination of health plant and herbivory treatment for each non-native species or their native congeners.