Tracing intracellular localization and chemical forms of copper in Elsholtzia splendens with cluster analysis

Copper (Cu) accumulation in soils is becoming a major environmental problem. Elsholtzia splendens, a Cu-tolerant plant growing in Cu mine areas, can accumulate high levels of Cu. This study develops a systematic method of detecting the intracellular localization and different chemical forms of Cu to comprehensively understand the mechanisms involved in Cu tolerance and detoxification of E. splendens. Various chemical forms of Cu were absorbed by E. splendens, the highest proportion of which was stored in its cell walls (68%) and vacuoles (42%). Most of the Cu in the roots, stems and leaves were extracted by 2% hydrate acetic acid (HAc) in controls. The distribution characteristics of Cu, particularly in the roots, were closely related to the Cu tolerance of E. splendens. In addition, the binding capacity of Cu to soluble salts or proteins decreased and its binding capacity to insoluble oxalate increased with the increasing Cu supply. These results indicated that Cu ions were integrated with compounds of low bioavailable, such as undissolved phosphate or oxalate, which contributed to the metal tolerance and detoxification of E. splendens. Cluster analysis, based on Cu abundance levels, reveals that correlations exist between the chemical forms of Cu and their intracellular locations, which in turn indicates a relationship between their migration activity and toxicity.