Research indicates possible green solution for manganese-contaminated soils

Study suggests eucalyptus could be the key to a sustainable and cost-effective solution for recovering manganese-contaminated soils

 

Manganese (Mn) is an essential micronutrient for plants, but at high concentrations, it can become toxic. However, Eucalyptus tereticornis appears to be remarkably tolerant to Mn, even at levels well above those that would cause harm to other plant species. The mechanism(s) underlying this ability were not understood based on scientific literature. From a study that monitored the Mn absorption in these plants, published in the Journal of Hazardous Materials, researchers from the Department of Plant Biology at the State University of Campinas (Unicamp) and from The Brazilian Synchrotron Light Laboratory (LNLS), from the Brazilian Center for Research in Energy and Materials (CNPEM), demonstrated how E. tereticornis can tolerate and detoxify high levels of Mn in its environment.  

The article, entitled “Tissue-level distribution and speciation of foliar manganese in Eucalyptus tereticornis by µ-SXRF and µ-XANES shed light on its detoxification mechanisms” led by Vinicius H. De Oliveira at Unicamp, presents the locations in the plant organism where Mn is accumulated, in what forms this element is assimilated, and even elucidates some of the mechanisms responsible for this ability to tolerate high concentrations of the metal used by E. tereticornis. This characteristic could be explored for environmental remediation purposes, particularly in contaminated soils.  

According to LNLS Soil Science Advisor, Dr. Dean Hesterberg, one of the article’s authors, it is not just the total Mn concentration that is important for understanding contaminated soils. “In acidic soils and especially under reducing redox conditions, manganese minerals are more soluble, which generally increases Mn availability for plant uptake. This can impact plants, which mainly absorb dissolved Mn. And, in Brazil, there are many acidic soils”, says Hesterberg.  

Synchrotron radiation imaging techniques

To gain evidence of how eucalyptus tolerates Mn-rich soils, researchers mapped the Mn distribution within Eucalyptus tereticornis leaves over time. This was possible through advanced techniques available at the Carnaúba beamline of the electron accelerator and synchrotron light generator, Sirius. The techniques used in the work included synchrotron micro scanning X-ray fluorescence imaging (µ-SXRF) and micro X-ray Absorption Near-Edge Structure (µ-XANES) spectroscopy.

Both use synchrotron radiation, a type of light released when electrons are accelerated to speeds very close to that of light. This usually happens by making them travel in a circular path, through strong magnetic fields, as is the case with the Sirius machine. Synchrotron light is incredibly bright and tunable over a wide range of wavelengths. In this way, the Carnaúba beamline uses light at X-ray wavelengths produced by the Sirius accelerator.

µ-SXRF

The synchrotron micro scanning X-ray fluorescence imaging (µ-SXRF) technique is used to investigate the elemental distribution and composition of materials on a microscopic scale. Fluorescence occurs because when materials are exposed to X-rays, atoms in the sample are excited and emit secondary (or fluorescent) X-rays when de-excited. The energy of these emissions serves as a fingerprint of each chemical element. This allows scientists to identify and quantify the composition of the studied material.  

LNLS/CNPEM researcher Dr. Carlos Alberto Pérez, one of the study’s authors, explains a little about how the technique works. “The µ-SXRF works based on X-ray optical equipment. The equipment has a monochromator, a crystal that defines a specific energy for the sample excitation. Another part of the equipment is the nanofocusing of this monochromatic light. This way, an X-ray beam that is about 100 times smaller than a human hair is created”.  

Through this beam of light, researchers are able to scan the sample, point by point, which generates an image with thousands of pixels. X-ray fluorescence is emitted as the beam hits each of these points. At the end, the pixels are computed using a program to generate an image, called elemental map. 

Elemental maps can be constructed for several specific chemical elements. In the case of the research published in the Journal of Hazardous Materials, the group of scientists assembled the elemental map of Mn in eucalyptus tissues. Thus, they were able to compare the presence of Mn in the plant’s leaf tissues, when it grew with an abundance of Mn and when it grew with normal amounts of the metal.  

μ-XANES

Micro-X-Ray Absorption Near Edge Structure (μ-XANES) spectroscopy, in turn, is used to probe the chemical state and electronic structure of specific elements in a sample. It is a sub-technique of X-ray Absorption Spectroscopy (analysis of how a sample absorbs X-rays), focusing on the energy band near the absorption edge of the element being studied. That is why the technique’s name brings the term ‘near the edge’.  

Hesterberg says that “unlike µ-SXRF, which is a fixed energy and scanning technique, µ-XANES is a variable energy technique. The absorption edge region is where there is a large increase in X-ray absorption by the sample”.  

Analysis of the edge region made it possible to discover the manganese oxidation state, that is, whether the element was in the form Mn²⁺, Mn³⁺ or Mn⁴⁺. Therefore, the technique allowed the researchers to understand if the manganese was in an oxidized form, or in a mineral state, and what coordinating atoms are likely around it. This means understanding what strategies eucalyptus uses to detoxify itself from the metal.  

Tolerance to high manganese concentrations

The study demonstrated that E. tereticornis tolerates high Mn concentrations without visible symptoms of toxicity or decrease in leaf biomass. Besides that, the two mentioned synchrotron techniques were used at three different points on the plant specimen leaves: the leaf vein, secretory cavity and leaf mesophyll regions.  

“Analysis of the µ-XANES spectra indicated that Mn was more closely bound to organic chelators, as opposed to being in a mineral state. Knowing these differences in the manganese chemical form or speciation helps us identify how the plant prevents toxicity by from this metal”, says Hesterberg.  

The results suggest two strategies used by Eucalyptus tereticornis to control Mn excess in its tissues. The first of these is through vacuolar sequestration, i.e., storage takes place within a region of leaf tissue called the vacuole, an organelle that stores water, food, waste, and other substances. This prevents the metal from interfering with essential processes, allowing the plant to thrive even in Mn-rich environments.  

This conclusion resulted from μ-SXRF measurements, which verified an accumulation of manganese in the leaf mesophyll region, indicating the vacuolar sequestration strategy. “The mesophyll has a large proportion of vacuoles, so this would be a logical place for manganese to go, as they absorb toxins or surplus components from the tissues around them”, tells Pérez.  

The second is through binding with organic acids (OAs). μ-XANES measurements revealed that Mn in leaf veins was mainly bound to OAs, for example, in the forms of Mn(II)-citrate and Mn(II)-malate. This binding process is called complexation – binding metals with OAs, which is similar to the mechanism of chelation used to remove metals from the human body. These compounds help to inhibit Mn toxicity. 

Interestingly, Mn was not found in high concentrations in calcium oxalate crystals (CaOx), a strategy used by other plants as a Mn sink.

A solution for contaminated soils?

The study’s findings have important implications beyond understanding how plant biology works. Since Eucalyptus tereticornis can tolerate and detoxify high levels of Mn, it could potentially be used for phytoremediation – when we use plants to remediate or clean environments contaminated by substances, such as metals for example. In particular, regions with acidic soils could benefit as they are usually manganese-rich. 

For Hesterberg, “perhaps eucalyptus could be used to carry out this phytoremediation, and develop a useful product, having more benefits from eucalyptus production than treating the soil chemically. We can calculate how many years it would take to clean an area in this way, which could be decades. However, this can be an inexpensive and very low-tech strategy”. Furthermore, this use of eucalyptus “may be more suitable for areas that have not been used, or even abandoned for many years”, Pérez added.  

Given the economic value of eucalyptus species, through the production of cellulose, wood or essential oils, E. tereticornis can offer a double benefit: environmental cleaning combined with an economic return.  

However, more research is still needed to explore the underlying molecular mechanisms in more depth to enhance detoxification strategies using plant specimens.  

According to Pérez, other elements, such as iron and zinc, which are essential for plants, can be researched using the Carnaúba beamline techniques. Hesterberg also gave ideas on possible issues to be investigated at CNPEM: “For example, are there other species that can accumulate more manganese? Can these detoxification mechanisms be generalized to other plants? Would it be possible to genetically modify these plants to increase their phytoremediation capacity? You could also look into how other nutrients affect manganese accumulation”.

About CNPEM

The Brazilian Center for Research in Energy and Materials (CNPEM) is home to a state-of-the-art, multi-user and multidisciplinary scientific environment and works on different fronts within the Brazilian National System for Science, Technology and Innovation. A social organization overseen by the Ministry of Science, Technology and Innovation (MCTI), CNPEM is driven by research that impacts the areas of health, energy, renewable materials, and sustainability. It is responsible for Sirius, the largest assembly of scientific equipment constructed in the country, and is currently constructing Project Orion, a laboratory complex for advanced pathogen research. Highly specialized science and engineering teams, sophisticated infrastructure open to the scientific community, strategic lines of investigation, innovative projects involving the productive sector, and training for researchers and students are the pillars of this institution that is unique in Brazil and able to serve as a bridge between knowledge and innovation. It is responsible for operating the Brazilian Synchrotron Light (LNLS), Biosciences (LNBio), Nanotechnology (LNNano), and Biorenewables (LNBR) National Laboratories, as well as the Ilum School of Science, which offers a bachelor’s degree program in science and technology with support from the Ministry of Education (MEC).