Supporting Information for. revealed defense and detoxification mechanism of. cucumber plant under nano-cu stress

Transcription

1 Supporting Information for 1 H NMR and GC-MS based metabolomics revealed defense and detoxification mechanism of cucumber plant under nano-cu stress Lijuan Zhao, Yuxiong Huang, Jerry Hu, ǂ Hongjun Zhou, Adeyemi S. Adeleye, Arturo A. Keller * Bren School of Environmental Science & Management, University of California, Santa Barbara, California , United States ǂ Materials Research Laboratory, University of California, Santa Barbara, California , United States Neuroscience Research Institute and Molecular, Cellular and Developmental Biology, University of California Santa Barbara, Santa Barbara, CA 93106, USA University of California, Center for Environmental Implications of Nanotechnology, Santa Barbara, California United States *Corresponding author. Tel: ; fax: address: keller@bren.ucsb.edu

2 Additional experiment: In order to ecucidate the role of amino acid and citric acid on the uptake of nano-cu by cucumber plant, additional experiments were conducted. In this assay, 2-week-old cucumber plants were exposed to 20 ppm nano-cu (Control), 20 ppm nano-cu + serine-l (6.25 mm), 20 ppm nano-cu + serine-m (12.5 mm), 20 ppm nano-cu + serine-h (25 mm), 20 ppm nano-cu + citric acid-l (6.25 mm), 20 ppm nano-cu + citric acid-m (12.5 mm), 20 ppm nano-cu + citric acid-h (25 mm) for one week. There were a total of 7 treatments and each treatment had three replicates. At harvest, the ph of nutrient solution, the total Cu concentration in nutrient solution, ph and the Cu concentration in different tissues were analyzed by ph meter and ICP-MS (Agilent 6900), respectively.

3 Figure S1. Fresh biomass of different tissues from cucumber plants treated with nano-cu at 0, 10 and 20 mg/l. All data show the mean of a total of four replicates (nine plants per replicate ). Error bars represent standard deviation. * indicates a significant difference (Tukey s HSD multiple comparison at p 0.05).

4 100% 98% 96% 94% 92% 90% 88% 86% 84% 82% Control 10 ppm Nano- Cu 20 ppm Nano- Cu Leaf Stem Root Figure S2. Cu distribution in different plant tissues.

5 Figure S3. Principal component analysis (PCA) scores plot (PC1 vs PC2) from 1 H NMR spectra of leaf extracts performed by MestreNova for leaves of cucumber plants exposed to 0, 10 mg/l and 20 mg/l nano-cu. Green = control, blue = 10 mg/l, and red = 20 mg/l.

6 Figure S4. Principal component analysis (PCA) scores plot (PC1 vs PC2) from 1 H NMR spectra of root exudates extracts performed by MestreNova for cucumber plants exposed to 0, 10 mg/l and 20 mg/l nano-cu. Green = control, blue = 10 mg/l, and red = 20 mg/l.

7 Figure S5-A. One-dimensional 1 H NMR spectra of cucumber root extracts. Cucumber plants were exposed to 0, 10 ppm, and 20 ppm of nano-cu for one week. Green lines represent Control, Blue lines represent 10 ppm, and Red lines represent 20 ppm. Each treatment has four replicates. The PCA loadings plots identify the regions of 1 H NMR spectra that contribute to the differences in PC1: the region of amino acids and organic acids (δ )

8 Figure S5-B. One-dimensional 1 H NMR spectra of cucumber root extracts. Cucumber plants were exposed to 0, 10 ppm, and 20 ppm of nano-cu for one week. Green lines represent Control, Blue lines represent 10 ppm, and Red lines represent 20 ppm. Each treatment has four replicates. The PCA loadings plots identify the regions of 1 H NMR spectra that contribute to the differences in PC1: sugar region (δ )

9 Figure S5-C. One-dimensional 1 H NMR spectra of cucumber root extracts. Cucumber plants were exposed to 0, 10 ppm, and 20 ppm of nano-cu for one week. Green lines represent Control, Blue lines represent 10 ppm, and Red lines represent 20 ppm. Each treatment has four replicates. The PCA loadings plots identify the regions of 1 H NMR spectra that contribute to the differences in PC1: aromatic region (δ )

10 Figure S6. GC-MS quantification data of 11 up-regulated amino acids in root exudates in response to exposure at 10 mg/l and 20 mg/l nano-cu.

11 PH in nutrient solution ph of nutrient solution nano-cu nano-cu with 6.25 nano-cu with 6.25 mm Serine mm Citrate A Copper concentration in nutrient solution (mg/l) Cu in nutrient solution nano-cu nano-cu with nano-cu with 6.25 mm 6.25 mm Serine Citrate B Copper concentration in root (mg/kg dry weight) nano-cu Cu in root tissues nano-cu with 6.25 mm Serine nano-cu with 6.25 mm Citrate C Figure S7. ph in nutrient solution (A) free Cu in nutrient solution (B) and Cu concentration in cucumber root tissues (C).Two-week-old cucumber seedlings were cultivated for 48 hours in half strength Hogland nutrient solution containing 20 mg/l nano-cu with 6.25 mm of serine or citric acids.