2.5.1 Total Cd and Cu content in moss and soil
We dried moss tissue from all populations and treatments at 50 ºC,
including field samples, and ground them to a fine powder in a tissue
lyser (n=3 replicates per sample). Then, we digested between 1-17 mg of
plant tissue in small Teflon vessels with 2 ml of HNO3(100%, TraceMetal Grade, Fisher Chemical) in a hot plate at 175ºC.
After 2 h, we added 0.8 ml of H2O2 (ACS
grade) to each sample and incubated them again for 2h at 175ºC. We also
prepared one sample of certified reference material (M2,Pleurozium schreberi moss tissue, Steinnes, Rühling, Lippo, &
Makinen, 1997) in the same way, and analyzed it along with analytical
blanks after every 12 samples to control for instrumental precision and
contamination.
Similarly, we dried soil samples from the field (n=3 replicates per
sample), sieved them to separate the 2 mm fraction, and ground them to a
fine powder before analysis. Between 9-30 mg of dried soil were digested
in small Teflon vessels with 2 ml of HNO3 (100%) for 8h
at 175ºC. This acid dissolves only a very small fraction of structural
minerals in the soil recovering mainly the most labile/reactive fraction
of heavy metals (Melo, Batista, Gilkes, & Rate, 2016). We used one
certified reference material prepared in the same way as the samples,
once every 12 samples, to control for instrumental precision (marine
sediment PACS-2, National Research Council of Canada).
Total contents of Cd and Cu were determined in both matrices at the
College of Marine Sciences (University of South Florida) by Inductively
Coupled Plasma Mass Spectrometry (ICP-MS, Agilent 7500cx). The
percentage recovery averaged 82% and 86% for Cu in plant and soil
respectively, and 102% and 131% for Cd in plant and soil respectively.