# ENERGY SAVING POTENTIAL BY THE APPLICATION OF GREEN ROOFS IN GREEK SCHOOL BUILDINGS

G. Kotsiris<fnref n="1" />, A. Androutsopoulos<fnref n="2" />, E. Polychroni<fnref n="2" />, P.A. Nektarios<fnref n="1" />

Corresponding author email: geokotsis@aua.gr

## ABSTRACT

Green roof utilization is increasing in contemporary urban centers mainly due their numerous beneficial impacts that include microclimate improvement. However, their cooling and thermal insulating features have been studied mainly in either office buildings or hotels. The aim of the present study is to evaluate the contribution of green roof systems in the energy needs of school buildings, which constitute a special type of public buildings in Greece. Different types (extensive, semi-intensive) and construction options of green roof systems were studied on eight school buildings. The energy needs of the buildings were simulated with TRNSYS simulation software, before and after the installation of the green roofs, in order to quantify their potential energy savings. The results from the simulation confirmed previous research results that reported that all tested green roof types improved the energy savings for cooling. By contrast, the energy savings for heating were much less or even insignificant. It was also shown that the use of specialized materials, such as substrates with increased porosity or a drain layer with good thermal insulation characteristics are necessary to avoid heating penalties.

<figure><img src="image_397.png" /></figure>

<footnote n="1">¹ Lab. of Floriculture and Landscape Architecture, Dept. of Crop Science, Agricultural University of Athens, Iera odos 75, 118 55, Greece</footnote>

<footnote n="2">² Buildings Department, Division of Energy Efficiency, Centre for Renewable Energy Sources & Saving (CRES), 19th km Marathonos Av., Pikermi 19009, Greece</footnote>
# THE ADAPTIVE APPROACH AS A MEANS TO INCREASE GREEN ROOFING THE MEDITERRANEAN BASIN

**Nektarios P.A., Ntoulas N., Kotsiris G., Nydrioti E., Varela D., Kapsali T., Kokkinou I., Reisi E., Amountzias I.**

Lab. of Floriculture and Landscape Architecture, Dept. of Crop Science, Agricultural University of Athens, Athens, Greece

Corresponding author email: [pan@aua.gr](mailto:pan@aua.gr)

## ABSTRACT

Green roof construction has been steadily increasing worldwide due to the numerous environmental and aesthetic advantages that they provide within the often monotonous and hostile landscape of contemporary cities. In city centers, which are mostly occupied by older buildings that can withstand only minimal additional weight loads, extensive green roof types seem to be the only applicable solution for green roof construction. However, in the absence of government incentives, a financial investment is frequently substantiated from the residents only if the green roof is an accessible and functional space.

In order to achieve sustainable management solutions, it seems necessary to deviate from the existing formulaic approach in concern to green roof typology towards an adaptive approach. This deviation requires rigid data and knowledge of the most significant parameters for assuring green roof sustainability and accurately predicting resources inputs. These parameters include the type and the depth of the substrate, the necessary water inputs and the selected plant species.

Several substrate types will be presented along with their capacities as well as their effects on plant growth. The substrates commonly utilize coarse textured materials that are lightweight and possess an increased water holding capacity. In addition of significant importance is the source and percentage of the organic amendment of the substrate.

Substrate depth has been substantiated as one of the most important factors contributing to green roof sustainability. It is commonly accepted that as substrate depth increases the palette of plant selection as well as plant growth and their sustainability also increases. Substrate depth becomes of further importance in the challenging climatic conditions of the Mediterranean basin, especially in the swallow depths that are utilized in the extensive green roof types (2-15 cm).

However under an adaptive approach, a reduction of the substrate depth that otherwise would be intolerable for plant survivor, could be compensated by increasing water inputs. The exact prediction of the annual water inputs in an adaptive green roof system will depend on the selection of the plant species or plant phytocommunities.

**Acknowledgements:** This project was funded by the Hellenic General Secretariat of Research and Technology under the call of Bilateral Cooperation between Greece and China having the code number of 12CHN136.

<figure><img src="image_398.png" /></figure>
# VEGETATIVE COVER AS A SUSTAINABLE METHOD FOR REHABILITATING VARIOUS MINE TAILINGS THROUGH PHYTOSTABILIZATION

Jacobus M. Pretorius¹, Pieter W. van Deventer²

¹,² School of Geo- and Spatial Sciences / Soil Science, North-West University, Potchefstroom, South Africa
*Corresponding author email: 22186247@nwu.ac.za*

## ABSTRACT

Phytostabilization is a useful tool in the rehabilitation of tailings storage facilities (TSFs); it involves the reduction of trace element mobility in a growth medium by means of absorption. In some cases it also refers to the physical stabilization of any surface, i.e. side slopes of TSFs. Vegetative material increases soil cover (cover [C-] factor in soil erosion equation) and consequently decreases soil erosion and the mobilization of any contaminants, e.g. trace elements. Leaching of contaminants is also reduced due to the hydraulic control exercised by plant roots, and the addition of soil organic matter by plants decreases the solubility of trace elements in a growth medium.

The objective of this study was to assess the suitability of different plant species for the phytostabilization of various mine tailings. Twenty-eight plant species (17 tree species and 11 grass species) were established in nine completely different types of tailings materials as well as in a control soil. Physical and chemical properties of tailings which are significant for phytostabilization were analyzed. Detailed monitoring was conducted over a period of 24 months to identify successes and failures. Monitoring criteria included the assessment of plant performance (plant physiology, plant stress factors) and changes in tailings chemistry (pH, electrical conductivity).

Final results indicate that the correlation matrix is a useful tool to assist in the recommendation of specific species for different tailings materials. Various successful tailings and species combinations were identified, e.g. *Bambuseae* sp., when planted in coarse coal discard material; and *Chrysopogon zizanioides* (Vetiver), when planted in low-pyrite gold tailings. These combinations showed minimal plant stress factors, good growth, and good canopy cover, basal diameter growth and root development.

Concluding remarks: Vegetative covers have proven to be a valuable method in tailings rehabilitation where successful tailings-species combinations are used; various improvements in the physical and chemical properties of tailings can be observed. Determining the success of specific species in given tailings materials early on in the planning phase of the rehabilitation programme is therefore essential.

**Keywords:** vegetative cover, tailings storage facilities, trace elements, soil erosion, phytostabilization, monitoring, cover factor

**Acknowledgements:** Financial support from the Technology and Human Resources for Industry Programme (THRIP) of the National Research Foundation (NRF) [South Africa] as well as from Frazer Alexander Tailings is greatly appreciated.

<figure><img src="image_399.png" /></figure>
# SEDUMS ROLE IN METAL RETENTION AND EMISSION BY GREEN ROOFS

**Julie Schwager**<fnref n="1" />, **Amandine Ouros**<fnref n="1" />, **Jean-Claude Begin**<fnref n="2" />, **Véronique Ruban**<fnref n="3" />, **Françoise Watteau**<fnref n="2" />, **Jean-Louis Morel**<fnref n="2" />

*Corresponding author email: julie.schwager@cerema.fr*

## ABSTRACT

To reach the ambitious objectives of good chemical states for waters that have been introduced in Europe with the Water Framework Directive 2000/60/CE, it is important to control pollutant fluxes. Urban runoff being recognized as one of the major sources of pollutants in surface water and rainwater harvesting spreading in the meanwhile, upstream stormwater management seems to be necessary.

Vegetative roofs are constituted by two materials currently used in water depollution, i.e. a "soil" and plants. Therefore, this technique is often considered as a potential solution to obtain a good roof water quality.

Nevertheless, vegetative roof impacts on water quality have proven to be mixed. The main reason is probably because vegetative roofs are not designed in order to purify rainwater. The objective of this presentation is

to better understand plant impacts on metal storing and disposing in green roof to provide criteria for plant selection in order to improve roof water quality.

<figure><img src="image_400.png" /><figcaption>Fig. 1: Substrate section showing aggregates around *S. spurium* roots</figcaption></figure>

This study is mainly based on two complementary steps. The first one is the monitoring of green roofs built in 2010 near Nancy (France). Three plants associations have been used on eight different structures. The first one contains six species of *sedum* (*S. album*, *reflexum*, *spurium*, *floriferum*, *hispanicum*, *sexangulare*). This genus is widely used for green roofing for its ability to withstand harsh climatic conditions. The two other plant associations also contain some local plants such as *potentilla erecta*, *euphorbia polychroma* and *iris sibirica*. Plant development was studied every two months. Samplings were collected twice a year and metal content was analysed in root and aerial parts. Substrates were also sampled in 2010 and 2013 for total and extractible metal contents analyses and physico-chemical characterizations. Metal content in the fertilizer used when the plants were introduced was also evaluated. Atmospheric deposits and roof water were collected monthly and analysed to measure metal contents. Along with this in situ study, complementary experiments at meso- and microscopic scale were realized. Thin section (20 µm thickness) of substrates containing sedums roots were made in 2013 and observed thanks to a binocular magnifier. Roots and leaves ultra-thin sections (80 nm) were observed thanks to a transmission electron microscope equipped with an Energy-Dispersive X-ray microanalysis system used to detect metals in plant tissues.

The different plants showed comparable ability to retain metal, far from hyperaccumulator capacities. The global amount of metal stored in the plant compartment was mainly influenced by the biomass of the different species. It was also superior to the amount of metals brought by the atmospheric deposit, meaning that the use of hyperaccumulator may not be necessary. It was also noticed that metal contents in plants were highly correlated to metal contents in substrates, which are bigger than atmospheric deposits ones. This indicates that a part of available metals from the substrate is stored by the plants, which contributes to reduce metal lixiviation from the substrate by rain water and consequently to reduce metal concentration in green roof water. Cr content in fertiliser proved to be significant. A strong correlation was observed between Cr amounts brought by

<figure><img src="image_401.png" /></figure>

<footnote n="1">¹ Cerema, Direction Territoriale Est, Laboratoire de Nancy, Tomblaine, France</footnote>

<footnote n="2">² Laboratoire Sols et Environnement, Université de Lorraine - UMR INRA 1120, Vandoeuvre-lès-Nancy, France</footnote>

<footnote n="3">³ Ifsttar, Bouguenais, France</footnote>
fertiliser introduction and measured in plants. If this metal had not been retained by plants, fertilisation would have increased Cr content in roof water. Root plants seemed to foster aggregates formation (Figure 1), which can reduce small particles leaching. Organic matter content in substrate was also higher as plants were present than under bare surfaces. Both of these impacts indirectly improve water quality: the first one by reducing metal emission from the substrate and the second by improving the substrate ability to retain metals. Indeed, organic matter proved to have higher metal sorption ability than the mineral part of the substrate. However, plant roots also seemed to increase the availability of some metals in the substrate, such as Zn. This could have a negative impact on roof water quality if these elements are lixiviated by water before being absorbed by the roots. Ultrastructural observations of plant roots revealed that metals were mainly stored by cells containing condensed polyphenolic substances. Bacteria degrading these polyphenolic metabolites have also been noticed. This induces that storage of metal in green roof plants is only temporary. Consequently, if plants are introduced on green roof to improve water quality thanks to their ability to store metals, maintenance practice should be adapted to its characteristics. Seasonal mowing and collecting of aerial parts or periodical plants substitution could be considered.
# LAYER SOIL FORMATION ON TAILING HEAPS AFTER SOIL COVERS AND TREES AFORESTATION

Rogelio Carrillo González, Ma. del Carmen A. González Chávez

Edaphology, Colegio de Postgraduados Campus Montecillo, Texcoco, México

Corresponding author email: crogelio@colpos.mx

## ABSTRACT

As a result of mining activities and soft environmental regulations a sort of tailing heaps has been abandoned along the countryside in Mexico. Most of the treatments and managements of the tailing are too expensive to apply to millions of cubic meters of these residues. In addition, many of these materials are a cocktail of elements including toxic metals and metalloids. So, a proper disposal management such as electrochemical treatment, or dumping are not viable. One option to sort it out, followed by a mining company, was apply soil covers over the plateau of the damp and afforest with *Pennisetum clandestinum*, *Acacia farnesiana*, *Populous nigra*, *Casuarina equisetifolia* and *Eucalyptus camaldulensis* in different areas. Thirty years later a soil layer seems to be formed but some plant trees fall down or dry to died up right, no records about trace elements availability has been carried out on the soil layers formed during the evolution of the rhizospheric layer. Therefore the aim of this research was to follow the soil layer formation and trace elements availability depending of the plants set up.

We identified four layers in the soil profile: litter (organic matter), root development, transition between root and tailing, and tailing layer. *C. equisetifolia* formed a thick layer of organic matter (up to 10 cm), but did not allow the introduction of other pioneer plants, *P. nigra* formed a variable layer (2 - 20 cm) of organic matter but allowed the establishment of other plant species. *E. camaldulensis* behaved similarly to *P. nigra* in soil formation. *P. clandestinum* formed an organic matter top layer and a thick root layer. Trace elements concentration on the different layers formed varied depending on the predominant plant species established, however generally speaking we cannot say that plant covers reduce metal availability in the top layer. Copper concentration for instance was low in the first 5 cm of soil were *C. equisetifolia* was growing, and increased in deeper layers, but in the area of *C. equisetifolia* and *P. nigra* the concentration was higher in the top layer (0-10 cm), decreased in the second layer and decreased again. Some tree plants fall down because their roots avoided growing down the tailing and keep on a surface layer, so when wind blows the roots are broken. Soil formation and trace element distribution in the soil profile vary between the predominant plants species established, possible explanation are discussed in this study.

<figure><img src="image_402.png" /></figure>
# ADVANTAGES AND DISSADVANTAGES OF THE PRESENCE OF THE 1BL.1RS WHEAT-RYE TRANSLOCATION IN SUSTAINABLE WHEAT PRODUCTION

Ioannis N. Xynias<fnref n="1" />, Theano Lazaridou<fnref n="1" />, Chryssanthi Pankou<fnref n="2" />, Athanasios Mavromatis<fnref n="2" />, Demetrios G. Roupakias<fnref n="2" />

Corresponding author email: ioannis_xynias@hotmail.com; xynias@teikal.gr

## ABSTRACT

The 1BL.1RS wheat-rye translocation (i.e. a new B genome chromosome consisting of the long arm of chromosome 1B and the short arm of 1R replacing the normal 1B, Figure 1) is probably the most important one, used by many breeders. Although four sources of translocations and substitutions have been reported in wheat, the 1RS arm of the translocation, obtained by Riebesel in the 1920s, is originating from "Petkus" rye (Rabinovich, 1998). The first commercial cultivar (cv. "Salzmunder Bartweizen") caring the 1BL. 1RS translocation, was produced and released in Germany by Riebesel in 1957. However, in most current cultivars the translocation was transferred through the Russian cultivars Kavkaz and Aurora, which are progenies of the Riebesel lines. The translocated fragments of rye chromosome offer to the host cultivar a number of important agronomic traits, i. e. high yield potential, broad adaptation, resistance to drought and biotic stress conditions (diseases and insects) and increased regeneration capacity of the plants, contributing to a sustainable wheat production. More precisely, the short arm of 1R chromosome contains resistance genes against leaf rust (Lr26), stem rust (Sr31), stripe rust (Yr9) and powdery mildew (Pm8). The main disadvantages of the translocation are that the aforementioned genes are linked with decreased bread-making-quality, their action depends on the genetic background of the host cultivar and the prevailing environmental conditions, but most of all, they are no longer effective against new biotypes of the diseases. However, recent results demonstrate a considerable resistance of new Hellenic doubled haploid lines (lines obtained through anther- culture) not only to rusts and mildew but also to septoriasis under the conditions of Ukraine. It could be concluded from the results of various reports that the effects on yield and drought tolerance have been highly dependent on genetic background and environmental conditions of the host cultivar. For example, the 1BL.1RS translocation in spring cultivar 'Pavon' was correlated with the presence of a larger root biomass and higher grain yields especially under well watered conditions. For this, breeders should pay careful attention to the effects of genetic background and environment on drought tolerance of lines potentially conferred by rye translocations. New efforts (including introgression of new genes) are inevitable to broaden the narrow genetic background of cultivars carrying the translocation if its positive traits are to be exploited.

<figure><img src="image_403.png" /><figcaption>Figure 1. The normal 1B chromosome (left) and the translocated (right) and two corresponding storage protein genes.</figcaption></figure>

<figure><img src="image_404.png" /></figure>

<footnote n="1">¹ Technological Education Institution of Western Macedonia - Florina, School of Agriculture Technology, Food Technology and Nutrition, 53100 Florina, Hellas</footnote>

<footnote n="2">² Laboratory of Genetics and Plant breeding, School of Agriculture, Aristotle University of Thessaloniki, 54124 Thessaloniki, Hellas</footnote>
# EFFECT OF THE 1BL.1RS WHEAT - RYE TRANSLOCATION ON THE ANDROGENIC RESPONSE IN BREAD WHEAT

**Theano Lazaridou**<fnref n="1" />, **Chryssanthi Pankou**<fnref n="2" />, **Ioannis N. Xynias**<fnref n="1" />, **Demetrios G. Roupakias**<fnref n="2" />

Corresponding author e-mail: [thelazaridou@yahoo.gr](mailto:thelazaridou@yahoo.gr)

## ABSTRACT

Anther culture is the most convenient and effective technique to obtain doubled haploid (DH) plants and thus to accelerate the production of improved wheat cultivars. This androgenic response depends on culture medium, on cold or heat pretreatment and mainly on the genotype of the donor plants. In case of bread wheat (AABBDD) it was referred that the presence of the 1BL.1RS wheat-rye translocation could improve the embryoid production and green plant regeneration after anther culture. Thus, the present study was conducted to further investigate the effect of the aforementioned translocation on embryoid induction and green plant regeneration in bread wheat after anther culture. For this to be done, four bread wheat cultivars carrying the 1BL.1RS translocation (KVZ/Cgn, Acherontas, Elisavet, and Orpheus) and five bread wheat cultivars without the translocation (Acheloos, Vergina, Mykonos, Prelude and Chios) were used. Approximately 1500 anthers (derived from 30 spikes) from each cultivar, containing anthers in the mid (MU) to late uninucleate (LU) microspore developmental stage, were cultured after cold pre-treatment for 7 days at 4°C. The induction, regeneration, and rooting media used were the W14, 190-2 and basic MS medium respectively. The best anther culture response was recorded in two cultivars carrying the translocation (i. e. cvs Acherontas, 49.8 embryoids/100 anthers and KVZ/Cgn, 33.6 embryoids/100 anthers). Cultivar Orpheus (carrying the translocation) and Chios, (lucking the translocation) performed almost equally (23.8 embryoids/100 anthers and 22.1 embryoids/100 anthers respectively). More green plants were produced from the cultivars Acherontas (6.5 green plants/100 anthers) and KVZ/Cgn (5 green plants/100 anthers). No response was reported for the cultivars lucking the translocation 1BL.1RS except cultivars Acheloos and Chios. They produced 2.36 green plants/100 anthers and 2.9 green plants/100 anthers, respectively. It is concluded that the positive effect of the translocation 1BL.1RS observed on anther culture response of bread wheat cultivars cannot be attributed entirely to the presence of the translocation. This is because the genetic background of the cultivars carrying the translocation could be also important.

<figure><img src="image_405.png" /></figure>

<footnote n="1">¹ Technological Education Institution of Western Macedonia - Florina, School of Agriculture Technology, Food Technology and Nutrition, 53100 Florina, Hellas</footnote>

<footnote n="2">² Laboratory of Genetics and Plant breeding, School of Agriculture, Aristotle University of Thessaloniki, 54124 Thessaloniki, Hellas</footnote>
# EVALUATION OF PLANT SPECIES AND SOILLESS SUBSTRATES FOR MEDITERRANEAN SUSTAINABLE GREEN ROOF INSTALLATIONS

Damiano Biagiotti<fnref n="1" />,<fnref n="2" />, Germina Giagnacovo<fnref n="2" />, Rita Di Bonito<fnref n="2" />, Carlo Alberto Campiotti<fnref n="2" />

Corresponding author email: rita.dibonito@enea.it

## ABSTRACT

**Background:** The installation of green roofs and green facades on urban buildings has received a growing interest for the potential environmental benefits. One of the advantages of the green roofs is the reduction of the surface temperatures by the plant transpiration with mitigation of the "heat island" effect in the urban settlements [2]. In the continental regions the parameters of plant growth on green roofs have been object of study [1] but in the Mediterranean areas the development of the green roof technology still requires a research aimed at the evaluation of materials and plant species adapted to the seasonal climatic variations and the summer drought. The aim of this work is: 1) The selection of plant species and sustainable supporting substrates able to form a coverage for extensive green roofs in the climatic conditions of the Central Italy. 2) The collection of the environmental data on a green roof model for the prediction of the thermal insulation.

**Materials and methods:** 1) The species *Sedum acre* and *S. reflexum* were used for trials using a blend of soilless substrates (Agriterram™ and Agrilit 3, 50% each) and the addition of amendments or arbuscular mycorrhizal inoculum. The amendments were: Biochar from biomass and Compost from food waste, added at 10% Vol/Vol. The experiments were carried out on small containers (10 cm deep) under outdoor conditions during the four seasons. Periodically, the plant growth parameters were evaluated. 2) Containers of 7 m<fnref n="2" /> each were made with layers of substrates (10 cm deep) and installed on a building terrace. The plant species *Sedum* spp. and *Sempervivum* spp. were transplanted at the density of 16 plants/m<fnref n="2" />. The trial included two indigenous accessions collected from Italian regions and two species from collections. The addition of compost to the substrate was evaluated also. Periodically, the coverage of each species and the inter-species interaction were evaluated. In addition, the temperature on the surface and the bottom of the system plant-substrate, the radiation, the moisture content and the climatic parameters were collected by sensors connected to a data collection system. The data were analyzed by a specific software in relation to the climatic parameters, in order to estimate the mitigation effect related to the plant coverage.

**Results:** The addition of the Compost to the substrate produced a significant effect on the plant growth parameters, endurance and ability to form a dense canopy over the four seasons. The inoculum with arbuscular mycorrhizae promoted the plant growth and the root development. The experiment on containers simulating the green roof conditions indicated the ability of indigenous Italian accessions of *Sedum* and *Sempervivum* to compete and form a consistent coverage when growing on mixed culture with collection accessions. The evaluation of the physical parameters of the systems plant-substrates indicated a mitigation of the surface temperature in the summer season, related to the plant transpiration.

**Conclusions:** The data indicated the good performance of a blend of species of Crassulaceae from collections and some indigenous Italian accessions for the realization of extensive green roof coverage, under the climatic conditions of Central Italy. The addition of Compost or mycorrhizal inoculum to a soilless substrate blend of 10 cm deep have shown an improvement of the plant growth parameters, under a limited fertilization rate. The model plant-substrate system has shown a reduction of the surface temperature indicating a mitigation effect in the summer season. A long term experiment is necessary to confirm the results under the variability of the climatic factors.

## References:

1. Bates, J.P. Sadler, R. Mackay (2013) Urban Forestry and Urban Greening 12, 98-108

2. Sfakianaki, E. Pagalou, K. Pavlou, M. Santamouris and M. N. Assimakopoulos (2009) Int. J. Energy Res. 33, 1059-1069

**Acknowledgements:** The financial support of the Italian Ministry of Economic Development is greatly appreciated.

<figure><img src="image_406.png" /></figure>

<footnote n="1">¹,² Dept. DAFNE, Università degli Studi della Tuscia, Viterbo, Italy</footnote>

<footnote n="2">² Unit UTEE-Efficienza Energetica in Agricoltura, ENEA Casaccia, Roma, Italy</footnote>
# ANAEROBIC DIGESTATE AS A SUITABLE PLANT GROWTH PROMOTER IN SUB-ACID SOILS

Giacomo Lencioni<fnref n="1" />, Nicola Cavirani<fnref n="2" />, Davide Imperiale<fnref n="2" />, Marta Marmirolli<fnref n="2" />, Nelson Marmirolli<fnref n="1" />,<fnref n="2" />

Corresponding author email: giacomo.lencioni83@gmail.com

## ABSTRACT

Biogas production is a suitable renewable alternative to fossil fuels and it is increasingly becoming of deep interest. One of the by-products of this process is the anaerobic digestate (AD), which can be obtained with an anaerobic digestion of animal or plants biomasses or with municipal waste, with different chemical properties and composition. From August 2012, Italian legislation has changed the definition of AD, and the rules for its use. Currently AD is classified as a by-product, instead of a waste. Many studies have assessed that, under controlled supply conditions, it has fertilising properties. Digestate can be used as a replacement for mineral fertilisers, reducing costs to farmers and minimising greenhouse gas emissions from cultivation.

The aim of this work is determining AD (methanogenic digestate, from anaerobic fermentation of swine manure, in which on average 80% of the total nitrogen is NH₄) effect on horticultural model plants with different protocols. Germination index has been assessed in pea (Pisum sativum) and phytotoxicity in water cress (Lepidium sativum). Shoot elongation and leaf chlorophyll content have been measured in lettuce (Lactuca sativa) and endive (Cichorium endivia), with a special attention for the fresh/dry weights ratio and for the roots/shoots ratio. In barley (Hordeum vulgare) the roots elongation and the radical inhibition indexes were determined by processing the average weights and lengths of roots. Germination and phytotoxicity were assessed by *in vitro* experiment in agarized media, while shoots and roots elongation was assessed in pot experiments on soil.

From left to right the effect on growth of tomato plants (San Marzano cv) with increasing AD concentration is shown

<figure><img src="image_407.png" /></figure>

Additional tests were performed using other horticultural plants: tomato (Solanum lycopersicon, two cultivars), pepper (Capsicum annuum, two cultivars) and geranium (Pelargonium zonale, one cultivar). Four different concentrations of AD diluted in a nutrient solution were supplied in order to determine the effects on growth rate, total biomass, leaves chlorophyll content, number of flowers, fruit size, harvest index. Different types of soil have also been tested.

In the experiments with different types of soil, it was shown that the AD from swine manure, with pH 8.0-8.5, had the effect of soil liming. Therefore, it can also be used to adjust the pH of acid and sub-acid soils, besides providing a fertilising effect.

The results of the combined tests on model plants and horticultural plants revealed the optimum concentrations of AD to be applied in order to maximise plants growth and soil liming.

Future perspectives concern the application of AD to several different tomato cultivars in metal polluted soils, to estimate the effect of AD on metal plant uptake.

<figure><img src="image_408.png" /></figure>

<footnote n="1">¹ C.I.N.S.A. National Interuniversity Consortium for Environmental Sciences, San Marco 2847, 30124 Venezia, Italia</footnote>

<footnote n="2">² University of Parma, Department of Life Sciences, Parco Area delle Scienze 11/a, 43124 Parma, Italy</footnote>
<figure><img src="image_409.png" /></figure>

BIOFUELS & BIOMASS FOR ENERGY
<figure><img src="image_410.png" /></figure>
# TEMPORAL EVOLUTION OF SOIL QUALITY UNDER SHORT ROTATION COPPICE AS COMPARED TO FOREST, GRASSLAND AND ARABLE PLOTS

Marie Stauffer<fnref n="1" />,<fnref n="2" />, Corinne Leyval<fnref n="1" />, Jean-Jacques Brun<fnref n="3" />, Jacques Berthelin<fnref n="1" />

Corresponding author email: [marie.stauffer54@gmail.com](mailto:marie.stauffer54@gmail.com)

## ABSTRACT

Nowadays, agricultural soils are subjected to strong anthropic pressure, including their use for production of diversified energetic raw material competing with food production. Short Rotation Coppices (SRC) are an intensive crop of fast growing tree species intended to produce energy wood chips. Many studies have been led on SRC, but hardly any have as yet dealt with its impact on chemical/biological parameters and soil functions and publications on its impact on soil microbial communities are scarce.

We assessed the biological and physico-chemical properties of soils planted with SRC in situ for two years compared to agrosystem, forest and grassland soils. Samples were collected from field plots of the four land uses in the Aisne valley (France), in autumn and spring 2010 to 2012. Twenty-five different parameters involved in soil fertility, biological communities and activity were measured at four seasonal sampling times. Based on the parameters discriminating the four land uses, three soil quality indices were calculated to reflect fertility, biological activity and biological community.

For many parameters, SRC values were higher than agrosystem values, but lower than forest or grassland values. The calculated soil quality indices were relevant with the following gradient (arable field > SRC > grassland > forest). Six years after plantation, the SRC crop had increased the soil fertility and biological community indices compared to the agrosystem. However, biological activity indices were similar in the SRC and agrosystem plots although soil basal respiration and laccase activity were higher in the SRC than in the agrosystem. Our results demonstrate that Soil Quality Indices can be considered as valuable tools to improve agricultural management. Whether a balanced SRC ecosystem functioning will be maintained in the next 15-20 years will have to be verified.

Acknowledgements: We thank Philippe Lucas, the owner of the SRC plot. This project was supported by Kinomé company and ANRT (National Agency of Research and Technology) with a CIFRE (Industrial Convention of Research Formation) contract number 2009/13/07. ADEME (« Agence de l'Environnement et de la Maîtrise de l'Energie ») sponsored and supported the research (contract number 0975C0095).

<figure><img src="image_411.png" /></figure>

<footnote n="1">¹ Lorraine University, LIEC-CNRS, Faculty of Sciences and Technology, BP 70239, 54506 Vandoeuvre-les-Nancy Cedex, France</footnote>

<footnote n="2">² KINOMÉ, 8 Rue de Valmy, 93000 Montreuil sous Bois, France</footnote>

<footnote n="3">³ UR EMGR, IRSTEA, BP 76, 38402 Saint Martin D'Héres Cedex, France</footnote>
# ENERGY USE AND GREENHOUSE GAS EMISSIONS IN ORGANIC AND CONVENTIONAL PEACH AND KIWI ORCHARDS

**Marios Michos**<fnref n="1" />, **George C. Menexes**<fnref n="2" />, **Andreas P. Mamolos**<fnref n="1" />, **Constantinos A. Tsatsarelis**<fnref n="3" />, **Kiriaki L. Kalburtji**<fnref n="1" />

*Corresponding author email: mamolos@agro.auth.gr*

## ABSTRACT

Energy flow in peach and kiwi orchards can help to determine first the best management strategies for each species and second the possibility of using the one having best environmental advantages. Conventional and organic peach and kiwi orchards found in the Prefecture of Pella, northern Greece, were selected in order to (a) determine energy flow of the farming systems, (b) reveal groups of orchards on the basis of their similarities to energy inputs, (c) estimate gas emissions and carbon footprint.

Fifteen farms [four conventional and three organic kiwi orchards (KC1, KC2, KC3, KC4 and KO1, KO2, KO3); four conventional and four organic peach orchards (PC1, PC2, PC3, PC4 and PO1, PO2, PO3, PO4)] were selected during the years 2010-2013. The energy encapsulated by the crop was estimated using the duration of each operation, the number of machines and laborers, all field operation inputs (irrigation, fertilizer application, harvesting etc.) and production coefficients (plant protection products, fertilizers etc.). Non-renewable (fuels, chemical fertilizers and pesticides) and renewable energy inputs (human labour, animal manure) are the components of total energy inputs. Carbon dioxide, CH₄, and N₂O and CO₂-equivalent emissions were estimated for all agricultural practices, soils, fuel and fertilizers [1,2]. The Hierarchical Cluster Analysis method was applied using seven production coefficients' variables (fertilizers, diesel, herbicides, fungicides, insecticides, labor, machinery) in order to reveal groups of farming systems [3,4].

Hierarchical cluster analysis revealed three groups of the studied orchards (Fig. 1). Group 1 (PC1, PC2, PC3, PC4 and PO1, PO2, PO3, PO4) had the lowest total and non-renewable energy inputs. Group 2 (KC1, KC2, KC3, KC4) had intermediate total energy inputs and the highest non-renewable energy inputs, while the opposite was observed for Group 3 (KO1, KO2, KO3). The highest contributors in cluster formation were human labour, shoring of branches, herbicides, fungicides, and machinery.

The largest contributors for gas emissions were fuels and fertilizers. Organic kiwi orchards (Group 3) tended to have the lowest gas emissions. Local management practices mainly affected the grouping of the studied orchards revealing their importance for the development of agro-environmental plans. Finally agricultural policies should regulate the local agro-environmental balance.

<figure><img src="image_412.png" /></figure>

## References:

1. IPCC, (2006) Software for national gas inventories, Intergovernmental Panel on Climate Change. Available at: http://ipcc2006.air.sk/ (Accessed 24 February 2014).

2. EMEP/EEA, (2009) Air pollutant emission inventory guidebook, Technical report No 9. Copenhagen: European Environment Agency.

3. Michos, M.C., Mamolos, A.P., Menexes, G.C., Tsatsarelis, C.A., Tsirakoglou, V.M., Kalburtji, K.L., (2012) Energy inputs, outputs and greenhouse gas emissions in organic, integrated and conventional peach orchards. Ecol. Indicat. 13, 22–28.

4. Litskas, V.D., Karaolis, C.S., Menexes, G.C., Mamolos, A.P., Koutsos, T.M., Kalburtji, K.L., (2013) Variation of energy flow and greenhouse gas emissions in vineyards located in Natura 2000 sites. Ecol. Indicat. 27, 1–7.

**Acknowledgements:** The financial support by Research Committee of Aristotle University of Thessaloniki.

<figure><img src="image_413.png" /></figure>

<footnote n="1">¹ School of Agriculture, Laboratory of Ecology and Environmental Protection, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece</footnote>

<footnote n="2">² School of Agriculture, Laboratory of Agronomy, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece</footnote>

<footnote n="3">³ School of Agriculture, Laboratory of Agricultural Engineering, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece</footnote>
PHYTOREMEDIATION WITH MISCANTHUS PRODUCED FOR BIOENERGY

Lawrence C. Davis<fnref n="1" />ª, Larry E. Erickson<fnref n="1" />ᵇ, Ganga Hettiarachchi<fnref n="1" />ᶜ, Joni Mengarelli<fnref n="1" />ª, Valentina Pidlisnyuk<fnref n="2" />, Kraig Roozeboom<fnref n="1" />ᶜ, Tetyana Stefanovska<fnref n="3" />, Natalya Tatarina<fnref n="3" />

Corresponding author email: ldavis@ksu.edu

ABSTRACT

Optimal use of land for food, energy, and materials, and phytoremediation of degraded lands are both
prominent sustainability issues. Miscanthus, a C-4 perennial grass, has high biomass productivity with
good water use efficiency and low nutrient demands. It grows well on some contaminated and marginal
lands that are not being used for food crops. It has the potential to improve soil quality by adding organic
matter to soil, while as a perennial grass, it has advantages to reduce water and wind erosion. It can be
harvested and used directly for combustion heating in a pelleted form, or potentially be processed for
liquid biofuels and biochar. This system was recently reviewed [1].

There are many locations in the world where land is not being used for crop production because of past
degradation, contamination, and/or a failure to be farmed profitably. Lands contaminated by the
Chenobyl disaster, Ukraine, and brownfields in Slovakia are examples of where miscanthus has been used
for attempted recovery. In the US, mine spoils and depleted farmlands are being tested for large scale
production of miscanthus. Research in progress in Ukraine, Slovakia, and the United States by the authors
tries to address the need for phytoremediation of sites contaminated by past military activities. Heavy
metals, especially Pb, are present at the identified field sites, from gasoline spillage and combustion, small
arms firing ranges and diverse other activities. These are likely combined with diverse organic
contaminants. During 2013, environmental monitoring in Kamenetz-Podilsky, Ukraine, showed Co, Mn
and Pb as contaminants at one site and Cu, Zn and Cd at another. At Fort Riley, Kansas, Pb contamination
on former practice ranges covers hectares of land.

Current research includes testing the productivity of miscanthus on soils from the contaminated sites, as
well as the effect of the miscanthus on soil quality and contaminant fate. Pot studies in Ukraine show that
miscanthus grows on soils from the contaminated site and there is slow uptake of the heavy metals
observed during two seasons of growth. Potential insect pests and diseases are being monitored. Rates
and extent of contaminant uptake are critical factors in considering whether to use the biomass for
energy production. Pot studies in Kansas show that soil amendments, principally organic matter, are
important in establishment of miscanthus in poor soils where heavy metal contamination is present.
Examples of results with effective use of amendments will be included. Hydroponic growth tests have
been used to determine nitrogen requirements of miscanthus, and its tolerance of salts. Miscanthus has
been effectively established and monitored several seasons under natural climatic conditions in large field
plots at K-State, near Fort Riley, KS.

Safe and effective use of miscanthus in phytoremediation systems must examine how contaminants affect miscanthus growth, and also the extent to which miscanthus accumulates or excludes contaminants. Natural climatic conditions, diseases, and pests must also be considered during establishment, and for long-term maintenance.

References:

1. Valentina Pidlisnyuk, Tetyana Stefanovska, Edwin E. Lewis, Larry E. Erickson and Lawrence C. Davis (2014) Miscanthus as a Productive Biofuel Crop for Phytoremediation Critical Reviews in Plant Science 33:1-19.

Acknowledgements: Supported in part by: Kansas Agricultural Experiment Station; Kansas State Center for Hazardous Substance Research; Matej Bel University, Slovakia; a NATO Planning Grant "New Phytotechnology for Cleaning Contaminated Military Sites"; and National University of Life and Environmental Science, Ukraine

<figure><img src="image_414.png" /></figure>

<footnote n="1">¹Depts of: a; Biochemistry, b; Chemical Engineering, c; Agronomy, Kansas State University, Manhattan, KS USA</footnote>

<footnote n="2">²Faculty of Natural Science, Matej Bel University, 40 Tajoskeho, 94701, Banska Bystrica, Slovakia</footnote>

<footnote n="3">³National University of Life and Environmental Science, 13, Gerojv Oboronu, 03040, Ukraine</footnote>
# GROWING *LEMNA MINOR* IN HUMAN AND SYNTHETIC URINE FOR BIOMASS PRODUCTION, NUTRIENTS AND ANTIMICROBIALS REMOVAL

Evangelia I. Iatrou<fnref n="1" />, Aloupi Maria<fnref n="1" />, Athanasios S. Stasinakis<fnref n="1" />, Nikos S. Thomaidis<fnref n="2" />

Corresponding author email: iatrou@env.aegean.gr

## ABSTRACT

Constructed wetlands with aquatic plants such as the duckweed *Lemna minor* have been widely used for tertiary wastewater treatment. Moreover, in recent studies *Lemna minor* has been used as a feedstock for biofuel production as well as a good resource of proteins and starch. In domestic wastewater, more than half of nutrient content originates from urine. As a result, there are some recent studies using urine for microalgae cultivation. According to them, human urine has the potential to become a good medium for biomass production of microalgae with high content in nitrogen, phosphorous and protein. In recent studies, the cultivation of duckweeds in swine wastewater represent high amounts of biomass production and starch content depending on harvest time, nutrient content etc. To the best of our knowledge, no information is available for the cultivation of *Lemna minor* using urine.

Based on the above, the main objective of this research was to achieve *Lemna minor* cultivation using human and synthetic urine. For this reason, the effect of several parameters (urine dilution rate, temperature, addition of microelements) on duckweed growth rate was investigated. The ability of *Lemna minor* to remove selected nutrients (NH₄-N, P) and selected antimicrobials (Cefadroxil, Metronidazole, Trimethoprim, Sulfamethoxazole) from urine were also studied.

Duckweed *Lemna minor* was grown on diluted human (fresh, hydrolysed and stored for 1 day) and synthetic urine. Batch experiments were initially done using different dilution factors (1:2, 1:5, 1:10, 1:25, 1:50, 1:100, 1:150, 1:200, 1:250) of human and synthetic urine in order to calculate growth rate of *Lemna minor* under different conditions. Experiments were also done at different temperatures (12 °C, 18 °C and 24 °C), different initial mass of duckweed, addition of organic carbon sources and different concentrations of trace elements such as Cu, Fe, Mn, Mg etc. Afterwards, the best urine solution was selected and the crude protein, starch and lipids content of biomass were determined. Finally, the removal rates of selected antimicrobials compounds commonly found in urine were calculated.

According to the results, the combination of duckweed *Lemna minor* and human/synthetic urine can provide a valuable solution for removal of nutrients and antimicrobials compounds from urine. The production of duckweed biomass can also serve as a biofuel feedstock due to high starch and protein content.

**Acknowledgements:** This research has been co-financed by the European Union (European Social Fund – ESF) and Greek national funds through the Operational Program "Education and Lifelong Learning" of the National Strategic Reference Framework (NSRF) - Research Funding Program: **THALES**. Investing in knowledge society through the European Social Fund. (www.aegean.gr/environment/watermicropol).

<figure><img src="image_415.png" /></figure>

<footnote n="1">¹University of the Aegean, Department of Environment, Mytilene, Greece</footnote>

<footnote n="2">²University of Athens, Department of Chemistry, Athens, Greece</footnote>
# ANAEROBIC CO-DIGESTION OF SEWAGE SLUDGE WITH GRAPE RESIDUES FROM WINE PRODUCING PROCESS

A. E. Maragkaki<fnref n="1" />,<fnref n="2" />, M. Fountoulakis<fnref n="1" />, K. Lasaridi<fnref n="2" />, M. Kornaros<fnref n="3" />, T. Manios<fnref n="1" />

Corresponding author email: amaragkaki@staff.teicrete.gr

## ABSTRACT

The grape agro-industrial sector is of major economic importance in many countries worldwide. Wine producing process generates large amounts of by-products which are totally unexploited and in some cases dangerous for the environment. Greece has produced an annual yield of about 350 kt of must corresponding to 140 kt of wine-grape residues. Anaerobic digestion of biomass is a well-known biological process producing biogas and biofertiliser. A significant number of biogas plants have been built, and now the concept is spreading all over the world. Biogas plants treat various types of organic residues including sewage sludge, food industry residues and manure. Wineries have a specific, seasonal product and the amounts of grape residues generated may not be sufficient to make digestion cost-effective. However, the addition of grape residues in a centralized facility digesting sewage sludge would ensure the viability of the process.

Two type of influent feedstock were utilized: 100% sewage sludge (SS) and a 5% of grape residues and 95% sewage sludge in order to investigate the biogas production of the grape residues – sludge co-digestion. The continuous experiments were carried out in a 3L digester constructed from stainless steel. The digester was operated at 35°C with a total feeding volume of 125ml daily and hydraulic retention time of 24 days. Feedstock was added four times daily (every 6 hours). Biogas was collected by displacement of water. Initially, the reactor was inoculated with anaerobic sludge originating from the Municipal Sewage Treatment Plant of the city of Heraklion (MSTPH). Also sewage sludge was originated from MSTPH. Grape residues were obtained from a local winery (Alexakis SA) in Heraklion, Crete. The characteristics of the feedstock are summarized in Table 1.

<figure><img src="image_416.png" /></figure>

The initial feed was sewage sludge and the bioreactor was operated using this feed for 30 days. Grape residues were then added to the feed so that the reactor was fed continuously with 95% sewage sludge and 5% food waste. The reactor treating the sewage sludge produced 463 ± 109 ml/d biogas before the addition of grape residues 940.50 ± 53 ml/d biogas after the addition of food waste (5% v/v in the feed).

**Acknowledgements:** This research has been co-financed by the European Union (European Social Fund – ESF) and Greek national funds through the Operational Program "Education and Lifelong Learning" of the National Strategic Reference Framework (NSRF) - Research Funding Program: ARCHIMEDES III. Investing in knowledge society through the European Social Fund (Acronym: EXISTING).

<figure><img src="image_417.png" /></figure>

<footnote n="1">¹ Department of Agricultural Technology, Technological Educational Institute of Crete, Estavromenos, Heraklion, 71500, Greece</footnote>

<footnote n="2">²Department of Geography, Harokopio University, 70 El. Venizelou, 17671 Kallithea, Athens, Greece</footnote>

<footnote n="3">³ Department of Chemical Engineering, University of Patras, 1 Karatheodori, University Campus, 26500 Patras, Greece</footnote>
# HARVESTING OF MICROALGAE THROUGH MAGNETOPHORESIS: INHIBITORY EFFECT OF IONPS TOWARD BIOFUEL PRODUCTIVITY

Pey Yi Toh<fnref n="1" />, Joanna Wan Yii Tai<fnref n="1" />, Bee Wah Ng<fnref n="1" />,<fnref n="2" />, Derek Juinn Chieh Chan<fnref n="1" />

Corresponding author email: peyyi2003@yahoo.com; chderekchan@usm.my

## ABSTRACT

Microalgae are suggested as the potential third generation renewable fuel sources due to its high oil yield (Johnson, 1999). Magnetophoretic separation of microalgae has been proven as a feasible harvesting method to achieve up to 99% cell separation efficiency (Lim et al., 2012) which yield a biomass estimated at $180/ton (Toh et al., 2012). A previous study showed a highly effective attachment of the surface functionalized iron oxide nanoparticles (SF-IONPs) onto the *Chlorella* sp. cell surface via the electrostatic-mediated-attachment (Toh et al., 2014). In industrial application, the recycling of the supernatant after the microalgae harvesting is necessary to minimize the water utilization for cost cutting purpose and reutilize the remaining nutrients to avoid eutrophication after discharge to the environment. However there will be some SF-IONPs residuals in the supernatant due of its high colloidally stable property (Yeap et al., 2012). Even though there are a number of studies about the toxicity effect of iron oxide nanoparticles (IONPs) in the biomedical application (Singh et al., 2010; Buyukhatipoglu and Clyne, 2010; Hoskins et al., 2012), however, there is still limited study on the toxicity effect of the IONPs toward the cultivated microalgae. In this study, the effective attachment of SF-IONPs onto *Chlorella* sp. cells has inhibited the cells growth. The growth inhibition is directly proportional to the dosage of SF-IONPs in culture media. The 20 mg/L of SF-IONPs has retarded the growth of cells up to 37.02 ± 14.06 % with respect to the control group. Total lipid yield was observed to reduce significantly up to 43.70 % with respect to the control group. Therefore residual SF-IONPs of more than 10 mg/L present in the supernatant is highly advised not to be recycling since it will retard the oil yield and elevate the production cost of the biofuel. However, in the continuous culture of cells which contained 50 mg/L of SF-IONPs up to 26 days, the growth inhibition effect from the SF-IONPs was observed in the first 9 days. The microscopy viewing found that the cells attached with the SF-IONPs will finally die and become colourless cell debris. However, the cells that are free from the attachment of the SF-IONPs will growth continuously and performed cell division. The growth of new cells in the medium diluted the concentration of SF-IONPs and eliminates the growth inhibition effect toward the cells.

**Acknowledgements:** This work is supported by Fundamental Research Grant Scheme (FRGS) (Grant No. 203/PJKIMIA/6071271) and Research University Postgraduate Research Grant Scheme (USM-RU-PRGS) (Grant No. 1001/PJKIMIA/8045037) from Universiti Sains Malaysia, the Science Fund Research Grant (Grant No. 305/PJKIMIA/6013412) and the Membrane Science and Technology cluster USM. Toh, P.Y. was supported by the My PhD scholarship from Ministry of Higher Education of Malaysia.

<figure><img src="image_418.png" /></figure>

<figure><img src="image_419.png" /></figure>

<figure><img src="image_420.png" /></figure>

<figure><img src="image_421.png" /></figure>

<footnote n="1">¹School of Chemical Engineering, Universiti Sains Malaysia, Nibong Tebal, Penang 14300, Malaysia</footnote>

<footnote n="2">²School of Biological Sciences, Universiti Sains Malaysia, Minden, Penang 11800, Malaysia</footnote>
# ASSESSMENT OF JATROPHA CURCAS AS A POTENTIAL FEEDSTOCK FOR BIOFUEL PRODUCTION IN SOUTH MEDITERRANEAN COUNTRIES

**Eleni G. Papazoglou**

Department of Crop Science, Agricultural University of Athens, Greece

Corresponding author email: elpapazo@aua.gr

## ABSTRACT

Jatropha curcas L. is a perennial shrub or small tree of the Euphorbiaceae family, characterized by low input requirements and high adaptability in harsh environments. This plant species is naturally grown and cultivated in tropical and sub-tropical regions producing high quality bio-oil, which can be easily converted to biodiesel.

The EU funded JatroMed project (www.jatromed.aua.gr) is evaluating the ability of this crop to be cultivated in South Mediterranean countries, aiming at the reinforcement of the development of poor rural areas by covering the energy needs of local communities. In order to reach this goal, five countries, namely Greece (coordinator), Italy, Egypt, Morocco and Algeria have successfully planted eight worldwide selected genotypes of *Jatropha curcas* in three low income rural sites of Egypt (Borg El Arab in Alexandria governorate), Morocco (Hard Draa in Essaouira region) and Algeria (Adrar wilaya). On 2012, in each target country four genotypes were established (two common for comparison reasons and two different). Each genotype is cultivated in one hectare and is treated under different levels of fertilization, irrigation and pruning.

The plantations are well established, the first fruits are harvested and oil is produced and analyzed. A harvesting device is under development by modifying existing olive harvesters and validating the results with field tests. The economic analysis of the production chain (from sowing to bio-oil production) is performed by applying Discounted Cash Flow (DCF) and Activity Based Costing methods. Social aspects are considered, such as acceptability from the farmers, number of jobs created, implication of women, etc.

The results so far showed that the best performed genotype for Egypt is Michoacán (non-toxic from Mexico), for Morocco is JCLMax (toxic from India) and for Algeria is Veracruz (non-toxic from Mexico). In conclusion, *Jatropha curcas* seems to be a promising energy crop for bio oil production in South Mediterranean countries.

**Keywords:** *Jatropha curcas*, biofuels, energy crops, agronomic practices, harvest, socio-economics

**Acknowledgements:** The author would like to acknowledge the European Union (EuropeAid/128320/C/ACT/Multi) for financially supporting this work. The contents of this document are the sole responsibility of the author and can in no way be taken to reflect the views of the European Union.

<figure><img src="image_422.png" /></figure>
# PHYTOREMEDIATION POTENTIAL OF SELECTED ENERGETIC PLANTS (MISCANTHUS GIGANTEUS AND PHALARIS ARUNDINACEA) IN DEPENDENCE ON FERTILIZATION

Malgorzata Kacprzak, Karolina Rosikon, Anna Grobelak, January Bien

Institute of Environmental Engineering, Czestochowa University of Technology, Czestochowa, Poland
*Corresponding author email: mkacprzak@is.pcz.czest.pl*

## ABSTRACT

Phytoremediation is the process of using plants to clean soil contaminated with heavy metals and other hazardous compounds, and also allows to restore degraded soils utility and natural values. Degraded lands are usually excluded from the food production; however, may be suitable for the cultivation of energy crops such as *Miscanthus giganteus* (miscanthus) or *Phalaris arundinacea* (reed canary grass). These plants due to the ability to collect and accumulate heavy metals are excellent for phytoremediation. Depending on the plant species, metals may be absorbed in larger amounts in the roots or aerial parts. Both, miscanthus and reed canary grass, due to a very well developed root system, may meet another additional function, namely anti-erosion, which further supports the possibility of using them in the process of phytoremediation.

This paper presents the influence of fertilization in cultivation *Miscanthus giganteus* and *Phalaris arundinacea* in the process of phytoremediation of Cd, Ni and Zn. There were two types of fertilization – organic with the use of sewage sludge and non-organic with the use of mineral salts.

Research show that after first year of vegetation both Cd (0.344 mg kg⁻¹ d.m.) and Zn ions (29.47 mg kg⁻¹ d.m.) were the most accumulated by the biomass of miscanthus grown on soil fertilized with municipal sludge (OW). However, in the case of the reed canary grass, the correlation can be attributed to the Ni (3.76 mg kg⁻¹ d.m.) and Zn (122.6 mg kg⁻¹ d.m.). In turn, after the second year of vegetation all of the tested metals are characterized by a lower absorbtion as compared to the first year. In the case of reed canary grass Cd and Zn content in the biomass after the second year of vegetation was similar to the control, while the Ni concentration was lower in relation to the control. In the case of miscanthus, it was observed that the concentration of Ni and Zn in the biomass after the second year of vegetation was similar to the control, while the Cd content varied in dependence on the applied fertilizer.

Concluding, both miscanthus and reed canary grass are able to accumulation of heavy metals, although the quantity of absorbed ions were depended on fertilization and the time.

**Acknowledgements:** The research leading to these results has received funding from the Polish-Norwegian Research Programme operated by the National Centre for Research and Development under the Norwegian Financial Mechanism 2009-2014 in the frame of Project Contract No POL NOR/201734/76

<figure><img src="image_423.png" /></figure>
# USE OF TE CONTAMINATED SITES FOR THE PRODUCTION OF BIOMASS FOR BIOCHAR

Michael W.H. Evangelou<fnref n="1" />, Anette Brem<fnref n="2" />, Fabio Ugolini<fnref n="3" />, Samuel Abiven<fnref n="2" />, Rainer Schulin<fnref n="1" />

Corresponding author email: michaelEvangelou@env.ethz.ch

## ABSTRACT

Trace element (TE) contamination of soils is a worldwide problem. However, although not considered safe anymore for food production without clean-up, many of these soils may still be used to produce biomass for non-food purposes such as biochar. In a first step we investigated the suitability of the tree species willow (Salix sp.), poplar (Populus sp.), and birch (Betula pendula) which grew on a TE contaminated site in France. Birch revealed significantly lower leaf Cd concentrations (1.2 – 8.9 mg kg⁻<fnref n="1" />) than willow and poplar (5 – 80 mg kg⁻<fnref n="1" />), thus posing the lowest risk for TE contamination of surrounding areas. It also displayed lower bark Cd, Zn and Pb concentrations than poplar and willow making it suitable for biochar production. Thus, biochars were produced by pyrolysis at temperatures 450 and 700 °C from birch wood growing on the TE contaminated site in France and birch wood originating from non-contaminated soil. During the pyrolysis, in contrast to Cu, Fe, Mg, K, Mn and P, the elements Cd, Pb, S and Na volatilized. The produced biochars were applied as an amendment on a non-contaminated low-fertility soil, and its effect on the growth and metal accumulation of ryegrass (Lolium perenne, var. Calibra) as well as soil microbial abundance was investigated. The root biomass of the biochar treated plants was lower than that of the non-amended plants, while that of the shoot was higher. Plant shoot K and Zn concentrations were increased significantly by up to 7- and 3.3-fold respectively, owing to the application of TE-contaminated biochar. For P, Mg, Mn, Fe and Cu no significant increase in concentration could be detected. Neither the TE-contaminated biochar, nor the non-contaminated biochar had adverse effect on the bacterial community of the soil. The results show that if the contaminated sites are chosen according to their soil TE, e.g. Cd and Zn, the produced biochar could be used to biofortify crops which grow on soils with low Zn concentrations.

<figure><img src="image_424.png" /></figure>

<footnote n="1">¹Institute of Terrestrial Ecosystems, ETH Zürich, Universitätstrasse 16, CH-8092 Zürich, Switzerland</footnote>

<footnote n="2">²Soil Science and Biogeochemistry, Department of Geography, University of Zurich, Winterthurerstrasse 190, Zürich 8057, Switzerland</footnote>

<footnote n="3">³Institute of Biogeochemistry and Pollutant Dynamics, ETH Zürich, Universitätstrasse 16, CH-8092 Zürich, Switzerland</footnote>
# MICROALGAE FOR WASTEWATER TREATMENT, CO₂ MITIGATION AND BIOFUELS: DREAM OR SUSTAINABLE MAID FOR ALL WORK?

Jean-Paul Schwitzguébel<fnref n="1" />, Stephen Mackay<fnref n="1" />,<fnref n="2" />, Eduardo Gomes<fnref n="1" />, Rolene Bauer<fnref n="2" />, Christof Holliger<fnref n="1" />

Corresponding author email: [jean-paul.schwitzguebel@epfl.ch](mailto:jean-paul.schwitzguebel@epfl.ch)

## ABSTRACT

Over the last decade, intensive research has been carried out all over the world to explore the huge potential of microalgae for biofuel production, which could be coupled to the use of industrial flue gas to enhance CO₂ fixation as a greenhouse gas mitigation strategy and to remove pollutants from various wastewaters. However, large-scale microalgae production for such purposes is not yet technically and economically feasible. Challenges are thus to increase the efficiency of both algae production and conversion into biofuel. The following parameters are critical for sustainable microalgal biomass cultivation: CO₂ concentration; possible use of wastewater streams; light intensity and quality; photobioreactor design; batch, fed-batch or continuous growth conditions.

The objective of our research was thus to assess the influence of these parameters on the quantitative and qualitative production of biomass, lipids and added-value chemicals, taking into account the probable antagonism between biomass and lipid production, often depending on the nitrogen supply. Several microalgal species were grown at laboratory scale to compare the yield of biomass and lipid production. In addition to the potential use of oil for biodiesel production, we are currently working towards demonstrating the technical feasibility of an innovative process, for syngas production via hydrothermal processing of microalgae. The process is envisioned as a closed-cycle with respects to nutrients, water and CO₂, that are separated and reused for microalgae growth. The possible effect of increasing injection of CO₂ in the growth medium was also investigated, with the aim to recycle CO₂ from flue gas and enhance biomass production.

On the other hand, to achieve the economical and ecological sustainability of algal biofuel production, it will be necessary to integrate it with the extraction of high-value products in a biorefinery process. Carotenoids have been proposed as added-value compounds that could contribute to make microalgal biofuel production economically feasible. Therefore, the viability and sustainability of extracting carotenoids before the hydrothermal treatment of the remaining biomass to produce syngas were investigated.

Finally, harvesting and dewatering microalgae cells remains a technical challenge and typically contributes to 20-30% of biomass production costs and can represent up to half of the total cost of algal biofuels. The potential of co-culture of filamentous fungal species with microalgae in a lichenization process as a strategy to reduce the cost and energy consumption of harvesting and of the whole process seems promising and was thus investigated. In submerged cultures, filamentous microorganisms actually aggregate and grow as loosely packed pellets or compact granules. Microalgae cells were immobilized in these pellets and easily removed as an aggregate with the fungal cells. Pellet formation is strain specific and highly dependent on operational conditions during cultivation. This study was especially focused on lichen pellet formation during the co-culture of *Chlorella sorokiniana* and of an unidentified filamentous fungus, which was eventually characterized. While algae growth was optimal between pH 6 and 10, the highest pellet formation was observed in the pH range of 4-7, thus requiring a strict control of pH during the whole cultivation. The dense formed pellets did sediment in submerged cultures when the agitation was stopped and were easily harvested. The effect of such a co-cultivation on the production of added-value chemicals like carotenoids and on the biofuel potential of the remaining biomass is under evaluation, and will be compared to results obtained with cultivation of microalgae only.

The proposed strategies may significantly reduce energy demands with regards to harvesting and dewatering of microalgal biomass and improve the sustainability of the whole process, but should be assessed at larger scale.

**Acknowledgements:** The research was supported by the Swiss Secretariat for Education, Research and Innovation, in the framework of COST Action CM0903 (Utilisation of biomass for sustainable fuels and chemicals).

<figure><img src="image_425.png" /></figure>

<footnote n="1">¹ Laboratory for Environmental Biotechnology, EPFL, Lausanne, Switzerland</footnote>

<footnote n="2">² Department of Genetics, Stellenbosch University, Stellenbosch, South Africa</footnote>
# TECHNICAL, ECONOMIC, ENVIRONMENTAL AND SOCIAL SUSTAINABILITY OF GASIFICATION-BASED BIOENERGY SYSTEMS FOR THE MEDITERRANEAN AGRO-INDUSTRIAL SECTOR

P. Manara, A. Zabaniotou

Biomass Group, Department of Chemical Engineering, Aristotle University of Thessaloniki, Univ. Box 455, 54124 Thessaloniki, Greece

Corresponding author email: azampani@auth.gr

## ABSTRACT

Small-scale gasification-based bioenergy systems coupled could be innovative alternatives for CHP production when fueled with agro-residues, providing parallel benefits related to residues management and sustainable agri-food production and agriculture. In this study, an indicator-based estimation of sustainability, considering not only economic but also environmental and social issues was attempted for a gasification-based bioenergy system. The outcomes of the study indicate that small decentralized units could be used in a symbiotic way with industrial agro-companies or agricultural associations in an eco-efficient way to cover their needs in heating and electricity. However, commercial success depends on capital reduction instruments such as subsidies, electricity feed in tariff and biomass price. The use of agro residues and food wastes provides the benefit of zero cost feedstock and the mobile character of the unit the zero cost of biomass logistics.

<figure><img src="image_426.png" /></figure>
# SRWC POTENTIAL BIOENERGY MARKETS IN NORTH CAROLINA, USA

**Elizabeth Guthrie Nichols, Dennis W. Hazel**

Department of Forest and Environmental Resources, North Carolina State University, Raleigh, NC USA

Corresponding author email: egnichol@ncsu.edu hazeld@ncsu.edu

## ABSTRACT

Woody biomass will be a major feedstock for energy production, including biofuels, to secure the future of domestic energy production in the United States. Fast-growing, purpose-grown tree plantations, or short rotation woody crops, are one of the most promising methods to meet future woody biomass demands. Whereas considerable research has been done in the U.S. on SRWCs, relatively little has been done in North Carolina. Over the last two years, we have begun a research and extension program to develop an understanding of potential SRWC production in North Carolina in order to meet immediate woody biomass market demands for wood pellets and potential biofuel demand for ChemTex. Our programmatic objectives are to develop recommendations of species or clones for specific site conditions; to develop planting, management; and harvesting guidelines for site conditions; to estimate growth rates, harvest yields, and feedstock characteristics; to estimate production and harvesting costs on various lands; to understand potential pest and disease issues; and to evaluate wood chemistry of species and clones specific to energy wood use. We will report on performance results from randomized-block establishment trials at coastal to mountainous sites for select clones of *Populus*, native hardwoods, and eucalyptus. Site trials vary by land type from cropland, managed forests, degraded lands, and marginal lands.

**Acknowledgements:** The financial support by North Carolina Department of Agriculture, ArborGen, and GreenWood Resources are much appreciated.

<figure><img src="image_427.png" /></figure>
# DEVELOPING A BIOMASS TO ENERGY UTILIZATION MANAGEMENT SPATIAL DECISION SUPPORT SYSTEM - IT TOOL FOR THE ISLAND OF CRETE

M. Marneri, E. Dialynas, G. Daskalakis, V. Manios

EnvinHealth S.A, Environmental, Nutritional and Health Services SA, 3 A. Tsakalof, 71409, Heraklion, Crete, Greece

Corresponding author email: gdaskalakis@staff.teicrete.gr

## ABSTRACT

There are more than 1,000,000 tonnes of biomass that are produced in an annual base in Crete, the majority of which are currently disposed improperly (the least), where could be used in renewable energy production, either through anaerobic digestion or incineration systems. Support system will focus on biomass appropriate for incineration units. Their collection and utilization was (and still is) non-economical due to a number of reasons as for example their physiochemical characteristics (high moisture content), their seasonal production and their dispersion in a vast and mountainous terrain.

The main aim of the support system – IT TOOL is to collect, analyze and evaluate a series of information (new knowledge) that will result in the development of a scheme, that would allow the collection of biomass that is currently not collected or utilized in the island of Crete and its transformation into a fuel that could be used either in central heating systems of private households and public buildings or as fuel in renewable energy producing units, in an environmentally and economically profitable way.

IT system combines all geographical datasets collected and will be able to dynamically convey the information in order to support decisions of the biomass investors. IT tool use inherent capabilities of GIS which provides better analytic capabilities and facilitates decision-making to a greater extent for the stakeholders. IT can be extremely helpful in deriving the most meaningful information necessary for better decisions. IT tool allow users to interact in a comprehensive manner by changing parameters and making different scenarios.

IT tool that allows the easiest and more useful representation of the data collected meaning investments in environmentally friendly technologies among of which renewable energy production systems. It is estimated that more than 1.0 billion euros will be invested in biomass units, since until 2020 more than 350 MWe units must be installed and operate. Since more than 50 % of the will be incineration units their need will be well above 1,500,000 tn of dry biomass. In this case systems that would allow the economical and environmentally friendly collection, treatment and transportation of biomass will provide a huge benefit to the local economy.

**Acknowledgements:** This research has been co-financed by Greece and the European Union (European Social Funds) (NSRF) 2007-2013, through the Operational Program "Business Support for the employment of highly qualified personnel". Project title: "Developing a Biomass to Energy Utilization Management Scheme for the Island of Crete". Acronym: Cretenergy.

<figure><img src="image_428.png" /></figure>
# DEVELOPMENT OF A PRODUCTION AND UTILIZATION SYSTEM OF WOODY BIOMASS FOR HEATING MUNICIPAL BUILDINGS – BIOMASS

A. Maragkaki<fnref n="1" />,<fnref n="2" />, A. Papadaki<fnref n="1" />, M. Fountoulakis<fnref n="1" />, I. Petousi<fnref n="1" />,<fnref n="3" /> N. Markakis<fnref n="1" />, F. Galliou<fnref n="1" />, G. Daskalakis<fnref n="1" />, J. Sampathianakis<fnref n="1" />, T. Manios<fnref n="1" />

Corresponding author email: amaragkaki@staff.teicrete.gr

## ABSTRACT

Municipalities in both Greece and Cyprus pay tens of thousands per year for pruning and cleaning of gardens and parks. In most cases these materials led the funeral along with other municipal solid waste, with a significant economic burden on each municipality. The «BIOMASS» proposes to address all these plant residues not as a problem but as an opportunity. Lead partner in the project is the Municipality of Festos, while partners are the Technological Educational Institute of Crete, FODSA of Samos and Community Council of Voroklini.

The project «BIOMASS» has as its main objective: Utilization of woody debris produced by urban and agricultural activity and converts them into solid biofuel (pellets) for heating public buildings. Finally the specific objectives of the project are to develop a system for collecting the plant residues urban, develop a system for collecting the plant residues from agricultural crops, the development of a plant pellets for each operator, the development of a network of burners in public / municipal buildings will allow the use of pellets produced, environmental and economic assessment of the process, and technical education personnel to other Municipalities and individuals on how to develop and operate the related systems. The main expected results are reducing the amount of plant residues and petroleum consumption from stakeholders for heating public buildings and at the same time develop at least two to three new full-time jobs for the operation of each of these systems.

**Acknowledgements:** This project has been co-financed by the European Union (ERDG) and National Funds of Greece and Cyprus 20% through the Operational CBC program Greece-Cyprus 2007-2013. Project title: "Development of a Production and Utilization System of woody biomass for heating municipal buildings". Acronym: Biomass.

<figure><img src="image_429.png" /></figure>

<footnote n="1">¹ Department of Agricultural Technology, Technological Educational Institute of Crete, Estavromenos, Heraklion, 71500, Greece</footnote>

<footnote n="2">²Department of Geography, Harokopio University, 70 El. Venizelou, 17671 Kallithea, Athens, Greece</footnote>

<footnote n="3">³ Department of Civil Engineering, Leeds University, Leeds LS2 9JT, UK</footnote>
COMPARATIVE STUDY OF H₂-PRODUCTION UNDER SULFUR DEPLETION BY
THE GREEN ALGAE C. REINHARDTII AND S. OBLIQUUS

Aikaterini Papazi<fnref n="1" />, Aristidis Michoglou<fnref n="2" />, Antigoni Nikolaki<fnref n="2" />, Armida-Irine Gjindali<fnref n="1" />, Dimitrios Stfanakis<fnref n="2" />, Eleftherios Touloupakis<fnref n="2" />, Georgia Zaxariou<fnref n="3" />, Vassiliki Petroulea<fnref n="3" />, Nikolaos Chaniotakis<fnref n="2" />, Ioannis Sanakis<fnref n="3" />, Vassileios Petrouleas<fnref n="3" />, Georgios Tsiotis<fnref n="2" />, Anastasios Melis<fnref n="4" />, Kiriakos Kotzabasis<fnref n="1" />, Dimitrios Ghanotakis<fnref n="2" />

Corresponding author email: ghanotakis@chemistry.uoc.gr

ABSTRACT

Photosynthetic hydrogen production by green algae was first reported by Gaffron and Rubin [1]. Under anaerobic conditions, electrons are transferred through the photosynthetic electron transport chain to ferredoxin, and subsequently to hydrogenase, which converts protons to molecular hydrogen. Photosynthetic hydrogen production can take place either by a PSII dependent or a PSII independent pathway; in the first case water is the electron donor, whereas in the second case electrons originate from the catabolism of organic substrates through the NAD(P)H-dehydrogenase.

The expression and activity of hydrogenase are both very sensitive to the presence of oxygen. In the PSII-dependent pathway, the photolysis of water results to production of molecular oxygen, which inhibits hydrogenase activity and thus hydrogen production. An effective way for establishing oxygen depletion in *Chlamydomonas reinhardtii* cultures is the removal of sulfur from the culture medium [2]. Sulfur depletion results in partial deactivation of PSII; under these conditions consumption of oxygen in a closed system was found to be much higher than oxygen production. These growth conditions are appropriate for hydrogenase activation and the production of molecular hydrogen.

In the present contribution we compare the ability of two green algae (*Chlamydomonas reinhardtii* and *Scenedesmus obliquus*) to produce hydrogen under sulfur depleted conditions. Thermal conductivity hydrogen measurements using gas chromatography have clearly demonstrated that of the two species only *Chlamydomonas* cultures can produce high amounts of hydrogen under complete sulfur deficiency. Further experiments in *Scenedesmus* cultures using different sulfur concentrations in the growth medium clearly showed that low concentrations of sulfur are necessary for hydrogen production.

The above-mentioned different response of *Chlamydomonas* and *Scenedesmus* cultures to sulfur deficiency could be the key for better understanding photosynthetic hydrogen production. Comparative studies using the two algal species, by proteome analysis and EPR characterization of PSII, could allow us to explore the hydrogen production mechanism of *Chlamydomonas* under sulfur deficiency. In addition, optimization of the conditions for hydrogen production by *Chlamydomonas* cells under sulfur deprivation took place by changing growth parameters, such as the initial cell concentration, the initial culture volume in the hermitically closed bottles, the initial glucose concentration and the presence or not of shaking.

References:

1. Gaffron H., Rubin J. (1942) Fermentative and Photochemical Production of Hydrogen in Algae. *J Gen Physiol* 26, 219-240

2. Melis A., Zhang L., Forestier M., Ghirardi M.L., Seibert M. (2000) Sustained photobiological hydrogen gas production upon reversible inactivation of oxygen evolution in the green alga *Chlamydomonas reinhardtii*. *Plant Physiol* 122, 127-136

Acknowledgements: The financial support by THALES project Bio-hydrogen production by unicellular green algae (ALGAH2) contract No. MIS 377283 is greatly appreciated.

<figure><img src="image_430.png" /></figure>

<footnote n="1">¹Department of Biology, University of Crete, Greece</footnote>

<footnote n="2">²Department of Chemistry, University of Crete, Greece</footnote>

<footnote n="3">³Democritos, National Center for Scientific Research, Athens, Greece</footnote>

<footnote n="4">⁴Department of Plant and Microbial Biology, University of California, USA</footnote>
# PREDICTION OF FORAGE ENERGY POTENTIAL THROUGH THE REGIONAL DIFFERENCES IN GROWING CONDITIONS AND ENERGY POLICY

Pavel Tlustos<fnref n="1" />, Josef Hakl<fnref n="2" />, Pavel Fuksa<fnref n="2" />, Lukas Pacek<fnref n="1" />

Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, Kamycka 129, 165 21 Prague – Suchdol, Czech Republic

Corresponding author email: tlustos@af.cz.u.cz

## ABSTRACT

Recent development of renewable energy sources (RES) and production of biofuels in the Czech Republic was significantly modified by a system of quotas and subsidies, which does not sufficiently respond to all consequences of this development. A tension occurs between the renewable energy on the one hand and natural resources, biodiversity, food safety and protection of local climate on the other hand.

The only independent evaluation of all available sources and their need from local level can give valid answer which source of bioenergy is suitable with the respect of above mentioned limitation. Potential of RES (photo-, wind, water, biomass), existing energy production facilities including RES applications, energy utilizable wastes, wood harvesting residual potential are gathered into large databases within the LIFE+ project Regional Sustainable Energy Policy (ReStEP). The most complicated evaluation is in the case of biomass production and utilisation. There are several limitations affecting biomass production including natural ones, legislation and sustainable development.

Recently, the focus has been given on the most productive crops such as maize however these wide-row crops could have some negative impact on environment. Other forage crops including legume's monocultures, legume-grass mixtures and permanent grasslands should be used as a bioenergy source. Unlike to maize forage crops also provide some important benefits for environment such as nitrogen fixation, improvement of soil fertility and protection against soil erosion [2], as well as phytoremediation of contaminated land [3].

<table><thead><tr><th rowspan="2">Species</th><th>Yield</th><th>Energy content</th><th>Energy production</th></tr><tr><th>t.ha<sup>-1</sup></th><th>MJ.kg<sup>-1</sup></th><th>GJ.ha<sup>-1</sup></th></tr></thead><tbody><tr><td>Silage maize</td><td>12 – 20</td><td>17.0 – 18.7</td><td>210 – 370</td></tr><tr><td>Forage sorghum</td><td>10 – 18</td><td>17.1 – 18.6</td><td>170 – 330</td></tr><tr><td>Alfalfa</td><td>8 – 13</td><td>17.0 – 18.3</td><td>140 – 230</td></tr><tr><td>Red clover</td><td>7 – 12</td><td>16.9 – 18.1</td><td>120 – 210</td></tr><tr><td>Legume-grass mixture</td><td>5 – 14</td><td>16.9 – 18.5</td><td>85 – 250</td></tr><tr><td>Permanent grassland</td><td>3 – 11</td><td>16.7 – 18.6</td><td>50 – 200</td></tr></tbody></table>

The objective of this study is to show differences in the biomass and energy production of main forage crops with respect of climate, soil fertility, degradation and erosion, as well the limits for energy use of agricultural residues.

Productivity in terms of energy output is expressed as a calorific value determined by the adiabatic calorimeter IKA C 5000 according to ISO 1928 (2009). This value represents an energy potential for direct combustion of biomass and is used for calculation of energy balance, important for comparison of different species or management practices [1]. The simple comparison of forage crops and their groups by the potential yield, the calorific value and the energy output is given in the table. Wide margins of all parameters presented there show the highest energy production by maize at fertile soils under suitable conditions. Comparable energy content of all crops and environmental advantages of forage ones supports the strategy, that proper crop management, up to some extent of less productive grasslands can substitute maize as energy source in less favourable areas.

## References:

1. Fuksa, P., Hakl, J., Brant, V. (2013) Zemdirbyste, 100 (4), 355-362.

2. Hakl, J., Fuksa, P., Habart, J., Šantrůček, J. (2012) Plant Soil Environment, 58, (6), 288 - 293. ISO 1928: 2009.

3. Tlustoš P., et al. (2006) Plant Soil Environ. 52, 413 – 423

**Acknowledgements:** The support from European Commission LIFE+ program, project **LIFE10 ENV/CZ/000649 ReStEP** is highly appreciated.

<figure><img src="image_431.png" /></figure>

<footnote n="1">¹Department of Agro-Environmental Chemistry and Plant Nutrition,</footnote>

<footnote n="2">²Department of Forage Crops and Grassland Management</footnote>
<figure><img src="image_432.png" /></figure>

MONITORING
<figure><img src="image_433.png" /></figure>
# PHYTOMONITORING OF AIRBORNE PCB CONTAMINATION FROM TWO LOCAL POINT SOURCES IN THE CANADIAN ARCTIC

**Barbara A Zeeb<fnref n="1" />, Carol Luttmer<fnref n="2" />, Sarah Ficko<fnref n="3" />, Ken Reimer<fnref n="2" />, Allison Rutter<fnref n="4" />**

*Corresponding author email: zeeb-b@rmc.ca*

## ABSTRACT

Saglek Bay (LAB-2), located on the coast of Labrador is a former Polevault station operated from 1953 to 1971 when it was abandoned. In 1996, it was determined that the site was contaminated with polychlorinated biphenyls (PCBs) with concentrations in soils far exceeding the Canadian Environmental Protection Agency (CEPA) regulation of 50 µg/g. This led to remediation work carried out from 1999 to 2004 to remove and/or isolate all PCB-contaminated soil exceeding 50 µg/g and to further remediate parts of the site to <5 µg/g PCBs. In this study, spatial and temporal trends of PCB concentrations in the new-year growth of dwarf birch (*Betula glandulosa*) and willow (*Salix spp.*) were investigated over a period of fourteen (1997–2011) years in an effort to track ecosystem recovery following the removal of the point source of contamination.

Concentrations of PCBs in plants were measured before (2001–2002), during (2003–2004), and for six years after (2005–2010) the screening and containerization of PCB-contaminated soils. PCBs were measured as Aroclor 1260 equivalents (dry weight) by gas chromatography with an electron capture detector (GC/ECD) using U.S. EPA method 680/8270. During the remediation activities, ambient air PCB concentrations were measured using active air samplers for comparison to the passive plant samplers (i.e. the dwarf birch and willows). PCB concentrations measured by the active samplers reached a maximum of 0.037 µg/m<fnref n="3" /> which was below the project criteria of 0.15 µg/m<fnref n="3" /> indicating minimal source emissions. During the same time period, PCB concentrations in the passive samplers (i.e. plants) showed significant increases of 2–14 fold compared to the baseline data from previous years.

The birch data also showed significant changes between monitoring events within the 2003 and 2004 sampling seasons (June to September) and decreases when ambient air concentrations were low, indicating the sensitivity of new-year growth to reflect net accumulation and ambient conditions at a temporal scale of approximately two weeks. In the third year following remediation, concentrations decreased to below baseline levels reflecting the overall remediation and source removal at the site.

The BAF-5 radar site located on the summit of Resolution Island, Nunavut, tells a similar story. As part of the Distant Early Warning (DEW) Line, large quantities of PCBS were left when the site was abandoned in the 1970's. Approximately 4000 kg of pure PCBs were removed in 1999, while lower level PCBs were remediated and or disposed of using a variety of techniques between 1999 and 2005. Plant biomonitors accurately tracked airborne PCBs during and after remediation. Together, these data show that we can effectively use plants to phytomonitor terrestrial ecosystem recovery following removal of PCB point sources at remote Arctic sites.

**Acknowledgements:** The authors are grateful to the local wildlife monitors and contractors at the North Warning Systems Office LAB-2 site who provided assistance in the field, to AirZone Inc. for sharing meteorology and air monitoring data and to the Canadian Department of National Defence for funding.

<figure><img src="image_434.png" /></figure>

<footnote n="1">¹Dept. of Chemistry & Chemical Engineering, Royal Military College of Canada, Canada</footnote>

<footnote n="2">²Environmental Sciences Group, Royal Military College of Canada, Canada</footnote>

<footnote n="3">³Faculty of Agricultural, Life and Environmental Sciences, University of Alberta, Canada</footnote>

<footnote n="4">⁴School of Environmental Sciences, Queen's University, Canada</footnote>
# TERRESTRIAL PLANT BIOCONCENTRATION FACTORS FOR ORGANIC CHEMICALS: EXPERIMENTAL VALUES DATABASE AND SCREENING-LEVEL MODEL EVALUATION

Bill Doucette<fnref n="1" />, Erik Dettenmaier<fnref n="2" />, Chuba Shunthirasingham<fnref n="3" />, Jon Arnot<fnref n="3" />

Corresponding author email: [william.doucette@usu.edu](mailto:william.doucette@usu.edu)

## ABSTRACT

Quantifying and predicting the transfer of chemicals from the soil environment into terrestrial plants is important for assessing human and ecological risks, evaluating the use of plants as biomonitors of environmental contamination and predicting the effectiveness of phytoremediation. Empirical data describing chemical uptake by plants roots are generally expressed as ratios of chemical concentrations in the plant compartment of interest (e.g. shoots, roots, xylem sap) to that in the exposure medium, (soil, soil pore water, hydroponic solution) measured at the time the samples are collected. These ratios, that may or may not reflect equilibrium, are generally referred to as bioconcentration factors (BCFs) but have also been named for the specific plant tissue sampled such as root concentration factors, stem concentration factors or transpiration stream (xylem sap) concentrations factors. Plant-soil BCF values have been used directly as models to provide estimates of plant tissue concentrations from measured exposure concentrations and have also been used as input into mechanistic models and in risk assessments. However, the development of such models is dependent on quality experimental data that provides insights into the underlying processes of chemical uptake and biotransformation and physical losses within plants. We collected and reviewed measured data from over five hundred published papers to develop a terrestrial plant bioaccumulation database that was used to evaluate several screening-level plant uptake models. The database includes plant bioconcentration factors for 360 organic chemicals and 100 plant species. The review shows the lack of experimental data collected in a consistent, reproducible manner confounds comparisons between studies and limits model evaluation and development. Key parameters describing plant growth and health were often missing. Additional experimental data for ionic and ionogenic organic chemicals are needed along with information on the biotransformation of chemicals within plants. Until standard protocols for the measurement of plant uptake data are available and used to generate consistent, high quality data, models will be only able to provide order of magnitude estimates of root uptake and resulting tissue concentrations. Case studies of well-conducted plant studies are summarized to provide guidance for future standard protocol development.

**Acknowledgements:** The financial support by ExxonMobil Biomedical Sciences, the Graduate Assistance in Areas of National Need Program, and the Utah Water Research Laboratory is greatly appreciated.

<figure><img src="image_435.png" /></figure>

<footnote n="1">¹Utah State University, Utah Water Research Laboratory, Logan, UT USA</footnote>

<footnote n="2">²Chevron Energy Technology Company, Houston, TX, USA</footnote>

<footnote n="3">³ARC Arnot Research and Consulting Inc., Toronto, ON, Canada</footnote>
# PHYTOSCREENING FOR PERCHLORATE: RAPID ANALYSIS OF TREE SAP

Matt Limmer<fnref n="1" />, Danielle West<fnref n="1" />, Ruipu Mu<fnref n="1" />, Honglan Shi<fnref n="1" />, Joel Burken<fnref n="1" />, Kim Whitlock<fnref n="2" />

Corresponding author email: limmer.7@gmail.com (1401 N. Pine St, Rolla, MO 65409)

## ABSTRACT

Perchlorate is an emerging contaminant increasing encountered at contaminated sites, found in groundwaters of 49 states. The highly mobile pollutant can transport rapidly, making site assessment difficult and costly. A method to rapidly assess in-planta perchlorate concentrations was developed. Tree sap collected from tree cores via centrifugation at 30,000g was used to evaluate the perchlorate contamination. The sap sample was directly analyzed by an UFLC-MS/MS method after isotope labeled perchlorate internal standard addition and filtration. A hydroponic greenhouse study of willow trees was conducted to fully evaluate the analytic techniques and demonstrate that tree sap perchlorate concentrations were proportional to dosing solution perchlorate concentrations. The method was applied at the Longhorn Army Ammunition Plant in Karnack, Texas, USA to delineate a perchlorate plume that extends into a wooded area. At the field site, analysis of ~200 tree cores revealed plume boundaries similar to those obtained by traditional groundwater monitoring methods. Phytoscreening offered higher spatial resolution and detail than the limited groundwater sampling, for considerably less cost than the groundwater sampling efforts. This research demonstrates the potential for phytoscreening to extend beyond the traditionally studied organic compounds, providing a low-impact, rapid method for delineating perchlorate plumes in groundwater.

<figure><img src="image_436.png" /></figure>

<footnote n="1">¹ Missouri S&T, Rolla, MO USA</footnote>

<footnote n="2">² TechLaw, Inc.Chicago, IL USA</footnote>
COMPARISON OF TREE CORING AND SOIL GAS SAMPLING FOR SCREENING OF
CONTAMINATED SITES

Mette Algreen<fnref n="1" />, Marcel Stalder<fnref n="2" />, Charlotte E. Riis<fnref n="3" />, Jan Petersen<fnref n="4" />, Mariusz Kalisz<fnref n="5" />, Janusz Krupanek<fnref n="5" />, Stefan Trapp<fnref n="1" />, Mette Broholm<fnref n="1" />

Corresponding author email: mann@env.dtu.dk

ABSTRACT

Site characterization is often time consuming and a financial burden for the site owners, which raises
a demand for rapid and inexpensive (pre)screening methods.

Phytoscreening by tree coring has shown to be a useful tool to detect subsurface contamination, especially of chlorinated solvents. However the application and dissemination of the method is still limited. On the other hand, soil gas sampling for mapping of volatile organic compounds in the subsurface is a common and commercially applied method. Both methods are semi-quantitative, low-invasive and inexpensive, which makes them suitable as initial screening methods for site characterization.

The aim of this study is to compare tree coring and soil gas sampling to evaluate to which extent tree coring may supplement or substitute soil gas sampling as a site contaminant screening tool. And where both methods are feasible, evaluate when (with respect to compounds, soil properties, and locations) one method is preferred over the other.

Fields sampling was performed at European sites contaminated with fuel components or chlorinated solvents from former site activities (industrial production, gas stations, air base or gas plant) in fall 2012 and 2013. Samples from different tree species such as willow, asp, oak, birch and pine were collected and analyzed by headspace GC-MS.

The soil gas measurements were conducted by consulting engineering firms as part of the site characterization prior to the tree core sampling events. Results obtained both by tree coring and soil gas sampling are compared and held up against quantitative results obtained by groundwater-and/or soil sample analysis.

Significant correlation between the methods is not always the case. However, both methods can detect contamination in the shallow subsurface and then identify high risk areas. The uptake of BTEX into trees varies to a greater extent with the tree species and the site conditions than chlorinated solvents, which lead to greater uncertainty.

Both methods have their advantages and disadvantages. Hence, the methods supplement each other.

Based on results from these initial screening methods, other more advanced/quantitative and cost-
intensive methods can be focused, with the overall goal to make site characterization more complete
and/or efficient.

Acknowledgements: The financial support by FP-7 project TIMBRE contract No. 265364 is greatly appreciated.

<figure><img src="image_437.png" /></figure>

<footnote n="1">¹Technical University of Denmark, Kgs. Lyngby, Denmark</footnote>

<footnote n="2">²SolGeo AG, Solothurn, Schweiz</footnote>

<footnote n="3">³Niras, Allerød, Denmark</footnote>

<footnote n="4">⁴Region of Southern Denmark, Vejle, Denmark</footnote>

<footnote n="5">⁵Institute for Ecology of Industrial Areas, Katowice, Poland</footnote>
# PERSPECTIVES OF WOODY PLANT IMPLEMENTATION IN PHYTOREMEDIATION OF TOXIC ELEMENTS IN INDUSTRIAL CITIES

Svetlana Gorelova<fnref n="1" />, Marina Frontasyeva<fnref n="2" />, Sergey Lyapunov<fnref n="3" />, Anatoliy Gorbunov<fnref n="3" />, Olga Okina<fnref n="3" />

Corresponding author email: salix35@gmail.com

## ABSTRACT

Currently, more than 70% of the world population lives in large cities. Most Russian megalopolises are characterized as developed industrial complexes providing serious impact on the environment. Woody plants are the major representatives of the biota that are the biofilters of toxic compounds and fixators of carbon dioxide and sources of oxygen and organic matter in urban ecosystems. In our research as a model urban ecosystem was chosen a city of Tula, a large industrial capital of the region, bordering Moscow, Lipetsk, Orel, Kursk, and Kaluga regions. The city is known for the presence on its territory of a number of ferrous and nonferrous metallurgical enterprises producing pig iron, ferro-manganese, vanadium and chromium compounds, armory, machine building and defense enterprises. The results of our investigations showed that the soil of the city by 40% subject to polymetallic contamination with Fe, Cu, Zn, Pb, and As. In the air, an increased content of Fe and Cu is observed. Sampling was carried out in the buffer zone of metallurgical enterprises and in the sanitary protection plantations along the main city highways.

Analysis of biogeochemical activity of woody plants under high anthropogenic impact pollution conducted by means of two complementary analytical techniques – atomic absorption spectrometry (AAS) and instrumental neutron activation analysis (INAA) – revealed promising species of woody plants for phytoremediation of the environment:

Populus nigra - Mn, Fe, Ni, Zn, Cd, Pb

Betula pendula - Mn, Fe, Ni, Zn, Cd, Pb

Aesculus hippocastanum - Ni, Cu, As, Pb

Larix sibirica - Mn, Fe, Pb

Acer platanoides - Mn, Fe

Tilia cordata - Fe, Cu

Sorbus aucuparia - Fe

Cotoneaster lucidus - Mn, Fe, Pb, Cd, Cu, Ni

Crataegus monogyna - Fe, Ni

Caragana arborescens - Fe, Sb

Philadelphus coronarius - Pb, Sb

Spiraea japonica - Cd, Cu

Forsythia intermedia - Cd, Cu

Syringa josikaea - Cd, Sb

Syringa vulgaris - Fe

Cornus alba - Fe

Crataegus sanguinea - Ni

Data on biogeochemical activity of species and their adaptive capacity in terms of polymetallic pollution permits competently select woody plants for greenery planting of sanitary protection zones of the technologically polluted urban ecosystems and a combination of species will allow maximal remediation of the environment from toxic elements.

**Acknowledgements:** The financial support by the RFBR grant ‘The study of adaptive characteristics and buffer role of woody exotic species in the migration of toxic elements in the urban ecosystems” (project No. r_center_a 13-05-97508)

<figure><img src="image_438.png" /></figure>

<footnote n="1">¹ Department of Biology and technology of living systems, Faculty of Natural Sciences, L.N. Tolstoy TSPU, Tula, Russia</footnote>

<footnote n="2">² Department of Neutron Activation Analysis and Applied Research, Division of Nuclear Physics, Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research, Dubna, Russia</footnote>

<footnote n="3">³ Laboratory of Chemical and Analytical Researches, Geological Institute of RAS, Moscow, Russia</footnote>
# THE REPETITIVE-NON CODING DNA OF BRYOPHYTES, A DIFFERENT, SIMPLE, FAST, AND INEXPENSIVE KIND OF "EARLY GENETIC INDICATOR"

**Bassi Paola**<fnref n="1" />, **Ferraro Marina**<fnref n="2" />, **Basile Adriana**<fnref n="3" />

## ABSTRACT

Studies over the last years showed that the highly repetitive /non-coding DNA, which was once considered useless for the life of the organism, to the point of being called "Junk DNA" or "Selfish DNA", it is vice versa important for the adaptation of the organism to the environment. In fact, the external stimuli, such as temperature variations, pollutants, or even mechanical stimuli or viral infections, are able to induce quantitative changes just in this peculiar fraction of the genome. It is not about DNA modifications "harmful", but rather about a sort of "language" that the genome uses to respond to environmental stimuli: in fact the modifications of the non-coding DNA bring in turn, via a "conformational" effect, to an amendment of the higher structures of the double helix of DNA, thereby affecting, albeit indirectly, the transcription of the coding DNA.

Our idea is to propose the observation of the behavior of highly repetitive DNA as a tool to test the effect of the environmental pollution. In our opinion, in fact, such different kind of technological approach could provide a breakthrough in such field of research. Indeed, being the modulation of repetitive non-coding DNA one of the first responses of the genome to the environmental stresses, such a methodology allows us to test the effects of environmental pollution in a time earlier to that of other indicators. Also, given that the modifications of repetitive/non coding DNA are a response only "preparatory" to other, potential, genetic events, such different techniques could detect a potential risk of pollution also in the cases in which the natural defense reaction of the plant makes ineffective other kind of indicators.

In this context, we suggest to observe the behavior of repetitive DNA in answer to stresses in the genome of Bryophytes: in fact, the experimental data obtained by our group in both, aquatic and terrestrial species of these plants, experiments conducted either directly within the cell, by cytochemical analysis of the nuclear genome (characterization of DNA with A+ T and G+ C specific fluorochromes, *in situ* DNA hybridization, Electron Microscopy Analysis, etc.), or indirectly, through appropriate characterizations of the extracted DNA (kinetics of dissociation and recombination, *in vitro* hybridization, Southern Blotting, etc.), have shown that, differently to what happens for the other plants, the Bryophytes have a peculiar structure of the nucleus that consent accurate and immediate observation of the behavior of repetitive-non coding DNA, in a simple, fast and inexpensive way.

**Keywords:** environmental pollution, early genetic indicators, repetitive/non coding DNA, bryophytes

## References:

1. Zuckerlandl E. (1981) A general function of non-coding polynucleotide sequences. Mol Biol Reports 7, 149-158.

2. Bassi P. (1990) Quantitative variations of nuclear DNA during plant development: a critical analysis. Biological Reviews, 65, 185-225.

3. Bassi P. (1991) Repetitive/non coding DNA: a possible link between environment and gene expression in plants?. Biologisches Zent, 10, 1-13.

4. Cullis CA. (1999) The environment as an active generator of adaptive genomic variation. In Lerner H.R., Plant Responses to Environmental Stresses: from Phytohormones to Genome Reorganization. Dekker M., New York. Basel, 149-160.

5. Bassi P. (1999) Effects of Environmental stress on repetitive DNA behaviour in plants. In Lerner H.R., Plant Responses to Environmental Stresses: from Phytohormones to Genome Reorganization. Dekker M., New York. Basel, 161-170.

6. Basile, A., Cogoni, A.E., Bassi, P., Fabrizi, E., Sorbo, S., Giordano, S., Castaldo Cobianchi, R. (2001) Accumulation of Pb and Zn in gametophytes and sporophytes of the moss *Funaria hygrometrica* (Funariales). Annals of Botany 87(4), 537-543.

7. Bassi P., Basile A., Ferraro M., Masi M., Migliaccio D., Morelli G., Napolitano E., (2006) Plasticity of repetitive DNA in response to metal stress in Bryophytes. Plant Biosystems 140(1), 80-86.

8. Bassi P., Basile A., Sorbo S. (2014) Behaviour of metal-induced repetitive-non coding DNA in the protonemata of *Funaria hygrometrica*. Plant Biosystems, in press.

<figure><img src="image_439.png" /></figure>

<footnote n="1">¹ Dept. of Environmental Biology, Univ. "Sapienza", 00185 Rome, Italy</footnote>

<footnote n="2">² Dept. of Genetic and Mol. Biology, Univ. "Sapienza", 00185 Rome, Italy</footnote>

<footnote n="3">³ Dept. of Biol. Sciences, Univ. "Federico II", 80139 Naples, Italy</footnote>
# CHANGES IN THE LEVELS OF HSP70 IN RELATION TO HEAVY METALS EXPOSITION IN LEMNA MINOR

Lentini Marco<fnref n="1" />, De Lillo Alessia<fnref n="1" />, Castiglia Daniela<fnref n="2" />, Esposito Sergio<fnref n="1" />, Cardi Manuela<fnref n="1" />

Corresponding author email: marcolentini1985@gmail.com

## ABSTRACT

The heavy metal pollution is one of the most widespread pollution on Earth. In the last decade many attempts have been made to find suitable biological plant indicators and appropriate markers witnessing the levels of stress produced by heavy metals.

In our study we use *Lemna minor* as a bio-indicator, and HSP70 as a protein marker to evaluate the effects of heavy metals.

Specifically, we investigated the occurrence and abundance of HSP70 protein depending on different heavy metals, by changing both levels and time of exposure of specimens cultured *in vitro*.

We used different HSP70 antibodies able to recognize cytosolic, chloroplastic or mitochondrial isoforms, in order to recognize which of them is/are specifically involved in response to heavy metals stress, by conventional and 2-D western blotting.

The possible relationship(s) between the presence of different isoforms, their abundance, and of the changing heavy metals stress exposition and concentrations, are discussed in order to define this method as a useful tools to assess and monitor heavy metals' pollution in plants.

<figure><img src="image_440.png" /></figure>

<footnote n="1">¹ Dipartimento di Biologia – Università di Napoli Federico II – Via Cinthia - 80126 Napoli, Italy</footnote>

<footnote n="2">² CNR-IBBR (Istituto di Bioscienze e BioRisorse) - Via Università, 133 - I-80055 Portici (NA), Italy</footnote>
# AQUATIC MACROPHYTES AS POTENTIAL PHYTOREMEDIATORS AND BIOMONITORS OF POLLUTED WATER BODIES IN THE MIDDLE URAL

Maleva M.G., Chukina N.V., Borisova G.G.

Department of Plant Physiology and Biochemistry, Faculty of Biology, Institute of Natural Sciences, Ural Federal University named after the first President of Russia B. Yeltsin, Ekaterinburg, Russia

Corresponding author email: maria.maleva@mail.ru

## ABSTRACT

The Middle Ural is a highly urbanized region and the most water bodies are contaminated by a wide spectrum of pollutants, including heavy metals (HMs). Aquatic macrophytes are important components of freshwater ecosystems. Some species have expressive ability to bioconcentrate and bioaccumulate both nutrients and toxic elements in their green parts. Therefore they're actively used as biomonitors and phytoremediators [Borisova et al., 2014; Krems et al., 2013; Pratas et al., 2011]. Nevertheless the physiological and biochemical basis of metal hyperaccumulation and the mechanisms of tolerance in plants are not still studied enough.

In response to HMs stress in the plants leaves develop effective defense systems, including synthesis of thiols-enriched proteins and peptides – metallothioneins and phytochelatins [Prasad, 1999; Malec et al., 2009]. It is known, that the latter are not only able to bind the excess of HMs, but also help to neutralize toxic reactive oxigen species (ROS), generated under the HMs stress (Thornalley and Vasak, 1985). The aim of this study was to determine the accumulative ability of some macrophyte species and investigate the level of thiol-enriched compounds in comparison with HMs contamination of polluted water bodies in the Middle Ural.

The study objects were *Elodea canadensis* Michx., *Potamogeton perfoliatus* L., *Ceratophyllum demersum* L., *Batrachium trichophyllum* (Chaix.) Bosch. and *Lemna gibba* L. The studies were carried out on watercourses of Sverdlovsk region (Middle Ural, Russia) that differ in their extents of pollution: rivers Chusovaya (the Kama River tributary), Iset' and Pyshma (Tobol River tributaries), and other water bodies within the mentioned above river basins. Ikbulat Lake, Sysert' and Reshetka rivers have been taken as "conventionally clean" since these water bodies are subjected to the lowest anthropogenic impact. The plants were sampled in July during the period of blooming. The water samples were simultaneously collected. Integrated samples of leaves were collected at ≥10 plants. The HMs content in the leaves of aquatic macrophytes and in nonfiltered water was determined using atomic absorption spectrometry (AAS Vario 6, "Analytik Jena", Germany). The content of soluble and membrane-bound thiols was estimated according to Ellman (1959). The non-protein thiols (NPT) were estimated according to Nagalakshmi, Prasad (2001). Protein content was determined according to Bradford (1976), using bovine serum albumin (Sigma) as a standard. The parameters were determined in three replicates. The statistical significance of the results was determined using nonparametric Mann-Whitney U-test, *p*<0.05.

The most polluted with copper was Pyshma River (0.042 mg/l), nickel – Revda and Pyshma rivers (0.988 and 0.146 mg/l, respectively), zink – Iset' River (0.040 mg/l), manganese – Pyshma River (0.241 mg/l), iron – Chusovaya River (0.912 mg/l). *B. trichophyllum* and *C. demersum* showed the highest potential for accumulation of HMs (especially for manganese, zink, iron and copper). Others species exhibited more selective accumulative ability. A high ability to accumulate manganese and zink was also found in *E. canadensis* and *P. perfoliatus*, iron and zink – in *L. gibba*. The order of values of the biological accumulation of HMs in the most studied species was as follows: Ni < Cu < Fe < Zn < Mn. The metals content in the leaves of aquatic macrophytes growing in impact water environment could be several times higher then in unpolluted plants. For example, in the leaves of *L. gibba* from a watercourse subjected to increased anthropogenic load (the Chusovaya River), the content of Cu was 7.2 times higher and Zn was 4.3 times higher than with plants from "conventionally clean" water bodies (the Sysert' River).

The increased accumulation of HMs in plant tissues upon environmental pollution was accompanied by changes in the level of thiols. It was found, that the content of nonprotein thiols, soluble and membrane-bound thiols in macrophyte leaves depended on HMs impact. Plants that lived in waters with high anthropogenic pollution had a significant increase of thiols compared with the "conventionally clean" ones. Besides the content of soluble and membrane-bound proteins was also significantly raised. This indicated that the macrophytes could be used as a good bioindicators in continuous long period monitoring of contaminated water ecosystems.

Thus, it might be supposed that some species of aquatic plants can be more successful bioaccumulators and show high potential in possible use as phytoremediators. At the same time the plants tolerance to the stressful environmental conditions is determined by their ability to develop different protective mechanisms, among which is the most specific synthesis of nonprotein and protein thiols, that can neutralize the toxic effects of HMs.

<figure><img src="image_441.png" /></figure>
LONG-TERM MONITORING OF CHLORINATED SOLVENTS IN TREES

Matt Limmer, Amanda Holmes, Joel Burken

Missouri S&T, Rolla, MO USA

Corresponding author email: limmer.7@gmail.com (1401 N. Pine St, Rolla, MO 65409)

ABSTRACT

Phytoremediation, while an attractive low-impact technology for site remediation, poses difficulties regarding assessment of contaminant removal and long-term efficacy. Direct measurements and calculation of pollutant removal rates by trees onsite provide a clearer picture of phytoremediation efficacy in contrast to indirect measurements, such as groundwater monitoring wells. However, the removal rate is dependent on environmental conditions, resulting in variable *in planta* concentrations and contaminant removal rates. This project used *in planta* solid-phase microextraction (SPME) coupled with gas chromatography (GC) to measure chlorinated solvent concentrations in four trees at a contaminated site in Rolla, Missouri over a four year period. A single measurement was made in less than twenty minutes, with ten minutes of that time required for transportation to and from the GC. The sampling ports in each tree have provided more than 100 samples per tree over the four-year period. Concentrations in the tree exhibited a seasonal trend, with highest concentrations observed during the summer months, when translocation of groundwater was highest. Peak concentrations measured in the summer were approximately one order of magnitude higher than minimum concentrations measured in the winter and were correlated with calculated evapotranspiration (ET) based upon local weather. Contaminant removal rates were generated using calculated ET and *in planta* concentrations. This modeling approach and *in planta* measurements offer a new method to project contaminant mass removal rates for phytoremediation of organic contaminants.

<figure><img src="image_442.png" /></figure>
<figure><img src="image_443.png" /></figure>
<figure><img src="image_444.png" /></figure>

HALOPHYTES
<figure><img src="image_445.png" /></figure>
# PHYTOREMEDIATION OF HEAVY METALS FROM COASTAL WATERS AND THEIR MOLECULAR MECHANISMS

**Abid Ali Ansari**, Subrata Trivedi

Department of Biology, Faculty of Science, University of Tabuk, Kingdom of Saudi Arabia

Corresponding author email: aansari@ut.edu.sa

## ABSTRACT

Heavy metal pollution in coastal marine waters is a serious global problem that needs to be properly assessed and mitigated. It poses negative long-term implication on the health of human and also the ecosystem at large. Here we describe the causes and effects of heavy metal pollution on coastal waters. Phytoremediation of heavy metals is discussed with emphasis on Cadmium, Arsenic, Lead, Copper, Chromium, Manganese, Nickel, Vanadium and Zinc. Located between marine and terrestrial environments, mangroves are transitional coastal ecosystems which are found mostly in the tropical and subtropical regions. The phytoremediation potential of several mangrove species is discussed. Finally, the different molecular mechanism involved in phytoremediation of metals is described. There has been significant progress in determining the molecular basis for metal accumulation, which provides a strong scientific basis to outline several strategies for phytoremediation of metals from coastal waters. The metal transporter genes that are involved in hyperaccumulation of metals and biotechnological approaches including the transgenic plants are elucidated.

*Acknowledgements:* The authors would like to acknowledge financial support for this work, from the Deanship of Scientific Research (DSR), University of Tabuk, Tabuk, Saudi Arabia, under the no. S-0014-1435.

<figure><img src="image_446.png" /></figure>
# PRODUCTION OF THE HALOPHYTE FORAGE *ATRIPLEX AMNICOLA* FROM HEAVY METALS CONTAMINATED SOILS

## Mamdouh Alsayed Eissa

Department of Soils and Water, Faculty of Agriculture, Assiut University, Assiut, 71526, Egypt

Corresponding author email: emankamel500@gmail.com

## ABSTRACT

Most of the studies about *Atriplex* plants grown on contaminated soils focused on *Atriplex halimus* while the genus of *Atriplex* contains more than 100 species. There is little information available about river saltbush (*Atriplex amnicola*) grown on contaminated soils also its response to organic fertilization. This study aims to determine (i) heavy metals accumulation in the shoot tissues of river saltbush; (ii) toxicity of heavy metals in the stems and leaves of river saltbush and (iii) impact of compost on reduction of heavy metals uptake.

Field experiment was conducted out at Ellwan, Assiut Governorate, Egypt (located at Longitude and Latitude 27 11 43 N and 31 04 19 E respectively). Untreated sewage waste water was used to irrigate the soil for more than 50 years and that soil considered to be moderately heavy metals contaminated soil [1]. One month-old seedlings of river saltbush were transplanted in the field at 1×1 m spacing and the plot area was 2×5 m². Compost at rates of 0, 15 and 30 ton ha⁻¹ were applied and mixed with the soil before planting.

From our study we found that *Atriplex amnicola* plants were affected significantly by compost treatments. Increasing compost application rates to 30 ton ha⁻¹ increased the stems and leaves fresh weight by 9 and 14 %, dry weight by 14 and 9%, crude protein (CP) increased by 29 and 25% and ash content by 39 and 15% respectively. The high rate of compost reduced the Zn and Pb concentrations in stems by11% and 32%, while in leaves by 6 and 38% respectively. Despite the extremely high total heavy metals concentrations in the studied soil, plants of river saltbush were able to maintain shoot elements content below toxic level. Toxicity levels of Zn, Cu, Pb and Cd are 100-400, 20-100, 30-300 and 5-30 mg kg⁻¹ dry weight of plant respectively [2]. The above numbers about toxicity levels are belong to the plant health but metal concentrations are also important for animal health. Permissible limits of heavy metals in animal's forages are 10 mg kg⁻¹ for Cd, 10-100 mg kg⁻¹ for Pb, 15-500 mg kg⁻¹ for Cu and 250-1000 mg kg⁻¹ for Zn as reported by [3]. In examining the obtained data, it appears that river saltbush metal accumulation did not exceed these limits, and it is may be a source of forage for livestock production in contaminated soils. Crude protein (CP) in the leaves of *Atriplex amnicola* ranged between (20.1-25.2%). Na and K concentrations in the leaves of *Atiplex amnicola* were higher than those in traditional forage crops but this do not prevent using of them as forage [4]. The current study shows the ability of A. *amnicola* to reduce HMs accumulation in their aerial parts. This shrub contained more crude protein and ash content compared to the traditional forage crops and the produced plant material is suitable as safe animal forage.

## References:

1. Usman, A. and Mohamed, H.M. (2009) Effect of microbial inoculation and EDTA on the uptake and translocation of heavy metal by corn and sunflower. *Chemosphere* 76, 893-899

2. Kabata-Pendias, A. (2001) Trace elements in soils and plants (3rd ed.). Boca Raton, CRC.

3. National Research Council (2005) Mineral tolerance of animals. National Academies Press, Washington, DC

4. Masters, D., M. Tiong, P. Vercoeb and H. Norman (2010) The nutritive value of river saltbush (*Atriplex amnicola*) when grown in different concentrations of sodium chloride irrigation solution. *Small Ruminant Research* 91, 56-62.

<figure><img src="image_447.png" /></figure>
# USE OF HALOPHYTES FOR PHYTOREMEDIATION OF ROAD RUNOFF CONTAMINATED BY DEICING SALTS

Rémi Suaire<fnref n="1" />, Ivana Durickovic<fnref n="1" />, Marie-Odile Simonnot<fnref n="2" />

Corresponding author email: remi.suaire@cerema.fr

## ABSTRACT

In winter maintenance, between 200,000 and 2,000,000 tons of deicing salts are spread on the French road network every year. The main deicing salt used in France is sodium chloride (NaCl), and its impacts on water streams, soils and biota are reported in the literature of the countries using this product. Even though 95% of the applied road salt ends up in the environment, there is almost no specific treatment reported in the literature. The high loads of dissolved NaCl in road runoffs are only diluted by their passage through retention ponds used for road runoff treatment in France.

Halophytes have been successfully tested for phytoremediation of soils polluted by trace metals and for phytodesalination. The use of halophytes on a biofilter treating road runoff could therefore be a promising technique to reduce the environmental impacts of deicing salts.

The objective of this study is to demonstrate that halophytes are suitable for phytodesalination of road runoff. The bioaccumulation of NaCl and other road pollutants (such as trace metals) are therefore tested for several plants.

Samples from a retention pond in France were collected and analyzed in order to measure deicing salt and trace metal concentrations encountered in the field. The measurements performed on those samples showed that $Na^+$ concentrations ranged between 10 and 2900 mg/L in the retention pond water input under study, and $Cl^-$ concentrations ranged between 5 and 4000 mg/L. In the pond output water, $Na^+$ concentrations ranged between 10 and 1100 mg/L, and $Cl^-$ concentrations between 5 and 500 mg/L.

These data permitted the preparation of phytodesalination bioassays and the choice of halophytes suitable for our experimental site (*Armeria maritima* and *Atriplex hortensis*). Tests of the halophyte germination, growth and pollutant bioaccumulation have been led. The halophytes have been hydrated with brines of concentrations between 0 and 10 g/L (reflecting those measured in the field) under strictly controlled conditions (temperature, nutrition, hydration, light, substrate quality, etc.). Two types of sodium chloride were used for the preparation of brines: chemically pure NaCl and the deicing salt (containing, besides NaCl, 5% of impurities and additives). After two-month-long bioassays, the pollutant bioaccumulation will be measured: analysis of $Na^+$, $Cl^-$, and trace elements will be performed on the halophytes (in stems, leaves and roots), and on the substrate after plant harvest.

So far, the growth of the two species under study was analyzed and both *Armeria maritima* and *Atriplex hortensis* have shown a relatively low growth. Although *Atriplex hortensis* showed relatively higher fresh mass compared to *Armeria maritima*, dry mass/fresh mass ratio was significantly higher for *Armeria maritima* suggesting higher water uptake for *Atriplex hortensis*, which could be attributed to a better tolerance of salinity increase.

The analysis of $Na^+$, $Cl^-$ and trace metals accumulation in the plants is still in progress.

**Acknowledgements:** The financial support by CEREMA project DEFROUPHROU and by GEMCEA is greatly appreciated.

<figure><img src="image_448.png" /><figcaption>Figure 1. Growth of *Armeria maritima* (AM) and *Atriplex hortensis* (AH) treated with deionised water or brine with 500 mg/L of pure NaCl</figcaption></figure>

<figure><img src="image_449.png" /></figure>

<footnote n="1">¹CEREMA, DTer Est, Laboratory of Nancy, Tomblaine, France</footnote>

<footnote n="2">²LRGP, UMR CNRS 7274, Université de Lorraine, Nancy, France</footnote>
# MIXED HEAVY METAL TOLERANCE AND ACCUMULATION IN THE WETLAND HALOPHYTE *JUNCUS ACUTUS* L.

Evdokia Syranidou, Eleni Manousaki, Stavros Christofilopoulos, Nicolas Kalogerakis

School of Environmental Engineering, Technical University of Crete, Greece

Corresponding author email: nicolas.kalogerakis@enveng.tuc.gr

## ABSTRACT

Halophytic species are widespread in Mediterranean basin and are known to tolerate various environmental stresses including heavy metals. *Juncus acutus* L. which is a perennial wetland halophytic plant with a wide ecological range has proven before to be tolerance to various environmental stresses including heavy metal stress by zinc and hexavalent chromium. In this study, the mixed metal tolerance and accumulation ability of the plant is investigated in order to explore its potential for mixed heavy metal phytoremediation.

For that purpose, a 5 months (October to April) pot experiment was conducted (six pots per treatment) with *J. acutus* plants collected from the Souda Bay in Chania and planted to the same amount of a typical surface soil collected from an agricultural area in Chania. After a sufficient adaptation period, the experiment started with the addition of 10 ppm Cd, 150ppm Ni, 500ppm Zn and 500ppm Pb, in one dose as aqueous solutions. The experiment took place in an open air area under natural light conditions with temperatures ranging from 5.3 to 31.1 °C. The soil was always over-saturated, as the pots were irrigated with water to achieve a water layer above the soil surface in order to simulate wetland conditions. At the end of the experimental period, measures of metals concentrations in plant tissues and in water of the soil pores and measurements of plant biomass, chlorophyll content, shoot water content and shoot proteins were performed. Moreover, plants were, also, assayed for activities of antioxidant enzymes such as the guaiacol peroxidase (GPX) and catalase (CAT).

The results suggest that *J. acutus* is not a hyperaccumulator of the examined metals (Table); however, it showed high tolerance to mix heavy metal pollution since all plants showed no visible

<table><caption>Table. Metal concentration in *J. acutus* tissues (ppm). Values represent means ± standard error (n = 6).</caption><thead><tr><th></th><th></th><th>Cd</th><th>Pb</th><th>Ni</th><th>Zn</th></tr></thead><tbody><tr><td rowspan="2">Control</td><td>Shoots</td><td>n.d.</td><td>1,25 ± 0,08</td><td>n.d.</td><td>15,66 ± 2,3</td></tr><tr><td>Roots</td><td>n.d.</td><td>5,98 ± 0,9</td><td>12,78 ± 6,6</td><td>28,64 ± 5,6</td></tr><tr><td rowspan="2">Metal treatment</td><td>Shoots</td><td>1,59 ± 0,6</td><td>84,94 ± 30,7</td><td>100,22 ± 18,7</td><td>200,28 ± 39,5</td></tr><tr><td>Roots</td><td>12,77 ± 1,9</td><td>626,35 ± 106,5</td><td>259,65 ± 36,4</td><td>476,14 ± 55,9</td></tr></tbody></table>

toxicity symptoms such as chlorosis, necrosis, or root inhibition. Moreover, shoot proteins, chlorophyll content and shoot water content were not found to be statistically affected by the presence of metals and in addition, plant's biomass was increased in the presence of metals. Furthermore, GPX activity and CAT activity of exposed plants to metals was not statistically elevated in comparison with the controls.

All the above suggest that *Juncus acutus* L. is a heavy metal tolerant plant that could be used in phytoremediation strategies for revegetation of mixed heavy metal polluted areas.

*Acknowledgements:* This work was co-funded by the European Union in the Work Programme 2012 "COOPERATION" FP7-KBBE-2012-6 project - "Integrating biotreated wastewater reuse and valorization with enhanced water use efficiency to support the Green Economy in EU and India" (WATER4CROPS, Grant No. 311933).

<figure><img src="image_450.png" /></figure>
# ALTERNANTHERA BETTZICKIANA A POTENTIAL PHYTOREMEDIATOR: GROWTH, METAL UPTAKE AND OXIDATIVE STRESS

Shafaqat Ali<fnref n="1" />, Hafiz Muhammad Tauqeer<fnref n="1" />, Mujahid Farid<fnref n="1" />, Qasim Ali<fnref n="2" />

## ABSTRACT

Industrialization and urbanization have heavily contaminated irrigated soils by heavy metals which upset the ecosystems including surface and ground waters, human health, food and cause many drastic issues for food chain. Phytoremediation is a vital and inexpensive technique used to remediate heavy metal contaminated soils with textile effluents, tannery waste and wet lands. Plants that show high metal accumulation often show slow growth rate and little biomass production when grown on soils contaminated with metals. Exploration and cultivation of heavy metal tolerant plants with a better growth is a very effective method for the purpose of phytoremediation of different mediums contaminated with heavy metals.

In this investigation, varying physiological and biochemical attributes of ornamental plant *Alternanthera bettzickiana* due to its tolerance to high salinity levels as reported in literature. So, keeping in view the resistance of this plant against high salinity levels, the present experiment was performed to find out the potential of *A. bettzickiana* to different regimes of cadmium (Cd) in maintained in soil. Different levels of cadmium treatments (0 mM, 0.5 mM, 1 mM and 2 mM) were applied in solution form after the establishment of the cuttings. CdCl₂ was applied as a source of Cd. For the estimation of varying physiological and biochemical attributes the plants were harvested after 10 weeks of planting the plant stem cuttings. Potassium sulfate (K₂SO₄), urea (CH₄N₂O) and diammonium phosphate [(NH₄)₂HPO₄] were used as fertilizers @ 2.19:1.36:2.4 mg L⁻<fnref n="1" /> after every 15 days to maintain the NPK ratio in the soil. Plant height and biomass increased significantly with the application of Cd stress and the maximum gain in biomass was obtained when Cd was applied @ 2 mM concentration. No visual symptoms of metal toxicity were observed on plants leaves or roots at all Cd regimes. Of different enzymatic antioxidants, superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) activities were also affected significantly in Cd treated plants but the increasing or decreasing trend was plant part specific. SOD activity in roots decreased while increased in leaf due to Cd stress and the maximum increase in leaf SOD was recorded at 2 mM Cd level. However, the opposite trend was true for CAT activity in root and leaf. While the POD activity increased in both root and leaf due to Cd stress. Leaf and root MDA content as indicator of lipid peroxidation also increased due to Cd application in both root and leaf and more increase was at 2 mM Cd level. Furthermore, soil Cd application increased its internal content in both the roots and leaves and the maximum Cd content was in plants treated with 2mM Cd level. From the results it was concluded that *A. bettzickiana* plants accumulated high concentrations of Cd when grown under cadmium stress and showed tolerance these levels. Thus we can conclude that *A. bettzickiana* seems to be valuable for phytostabilisation strategies.

<figure><img src="image_451.png" /></figure>

<figure><img src="image_452.png" /></figure>

<footnote n="1">¹Department of Environmental Sciences, Government College University, Allama Iqbal Road Faisalabad, 38000, Pakistan</footnote>

<footnote n="2">²Department of Botany, Government College University, Allama Iqbal Road Faisalabad, 38000, Pakistan
Corresponding Author email: shafaqataligill@yahoo.com</footnote>
# PHYTOREMEDIATION OF A HYPERSALINE SOIL IN KUWAIT BY RHIZOSPHERIC AND PHYLLOSPHERIC MICROORGANISMS ASSOCIATED WITH ORGANS OF A HALOPHILIC WILD PLANT

Dina M. Al-Mailem, Mais Marafie, Mohamed Eliyas, Samir Radwan

Microbiology Program, Department of Biological Sciences, Faculty of Science, Kuwait University, Kuwait

Corresponding author email: dina.almailem@ku.edu.kw

## ABSTRACT

Hydrocarbonoclastic microorganisms associated with roots and aerial organs of *Halocnemum strobilaceum*, a wild, halophytic plant growing in a hypersaline coastal area in Kuwait, were counted and isolated. The NaCl concentration of this coastal area ranged between 2 M and 4 M. Up to 8 x 10⁴ colony forming units (CFU) g⁻¹ of extremely halophilic, hydrocarbonoclastic microorganisms were counted in the rhizosphere, and several hundreds of CFU g⁻¹ in the phyllosphere of this plant. According to their 16S rRNA-gene sequences, the rhizospheric microorganisms were affiliated with the two archaea, *Halobacterium* sp. and *Halococcus* sp., and with the five extreme halophilic bacteria, *Brevibacillus borstenlensis*, *Pseudoalteromonas rutherila*, *Halomonas sinaensis*, *Pseudomonas stutzeri*, *Mycobacterium bolletii* and *Panibacillus polymyxa*. The phyllospheric microorganisms were affiliated with the extreme halophilic bacterium *Ochrobactrum* sp. and the halophilic dimorphic yeast *Candida utilis*. All individual organisms, when used as inocula could consume, in batch culture, crude oil and pure aliphatic and aromatic hydrocarbons in mineral media, with 2 M salinity. Also the total rhizospheric and phyllospheric microflora associated with the plant when used as inocula were effective in hydrocarbon removal at high salinity. It was concluded that this wild, halophytic plant species is a suitable agent for the phytoremediation of hypersaline environments polluted with oil.

**Acknowledgements:** This work was funded and supported by the University of Kuwait, Research project (RS 01/12).

<figure><img src="image_453.png" /></figure>
CICHORIUM SPINOSUM AS A PHYTOREMEDIATION SPECIES

Mendoni Eleni<fnref n="1" />, Salonikioti Anastasia<fnref n="1" />, Petropoulos Spyridon<fnref n="1" />, Antoniadis Vasilios<fnref n="2" />, Efthimia Levizou<fnref n="3" />

Corresponding author e-mail: fangio57gr@gmail.com

ABSTRACT

Increased salinity is an ever-growing environmental problem that severely affects agriculture by limiting crop production and by causing loss of agricultural land. According to the United Nations Food and Agriculture Organization approximately 20-30% of all agricultural lands are becoming saline, whereas Global Assessment of Soil Degradation (GLASOD) reports that about 8% of all salt-affected soils are human-induced.

In the present study, *Cichorium spinosum* plants, which are native to the island of Crete, were grown under medium to high salinity levels (4-8 dS/m) by adding NaCl to irrigation water, in order to evaluate their productivity and mineral composition, and were compared to control treatment (1.8 dS/m). From the results it is observed that *Cichorium spinosum* is tolerant to medium salinity levels (4 dS/m), since the number of leaves, fresh weight of leaves and total leaf area is significantly higher than that at high salinity levels (8 dS/m), without differ significantly from the control treatment (1.8 dS/m). In addition, chlorophyll content was not affected by salinity level, whereas leaf thickness increased significantly for plants grown under medium salinity levels (4 dS/m). Regarding mineral composition, Na⁺ content increased significantly whereas Mg<fnref n="2" />⁺, Mn and K⁺ decreased with increasing salinity. Ca<fnref n="2" />⁺ and Fe<fnref n="3" />⁺ content was not affected.

Therefore *Cichorium spinosum* could be considered a species with tendency to accumulate Na⁺ without its productivity being affected. In this context, the cultivation of wild native species such as *Cichorium spinosum* could be a useful means to alleviate problems of saline soils by cultivating it for phytoremediation purposes, whereas the final products could be marketed as edible herbs. In addition, this species could be cultivated in areas where irrigation water is of low quality due to high content of NaCl (coastal areas or areas where ground water is saline), allowing farmers to exploit these areas and gradually improve their soils.

<figure><img src="image_454.png" /></figure>

<footnote n="1">¹ Laboratory of Vegetable Production, University of Thessaly, Fytokou Street, N. Ionia, Magnissia, Greece</footnote>

<footnote n="2">² Laboratory of Soil Science, University of Thessaly, Fytokou Street, N. Ionia, Magnissia, Greece</footnote>

<footnote n="3">³ Laboratory of Plant Physiology, University of Thessaly, Fytokou Street, N. Ionia, Magnissia, Greece</footnote>
<figure><img src="image_455.png" /></figure>
<figure><img src="image_456.png" /></figure>

EMERGING ISSUES
<figure><img src="image_457.png" /></figure>
# SUSTAINABLE REUSE OF NITROGEN-LADEN PROCESS WATER IN POPLAR TREE VADOSE ZONES AT VARIOUS SCALES

Hayden Ausland<fnref n="1" />, Lou Licht<fnref n="2" />, Craig Just<fnref n="1" />

Corresponding author email: craig-just@uiowa.edu

## ABSTRACT

Process water from industrial-scale preparation of food products from potatoes, onions and similar produce contains relatively high levels of organic carbon and nitrogen. Large processing facilities generate over 5 million gallons of process water daily with land irrigation being one of the disposal methods of choice. Irrigation to produce a commodity crop, such as alfalfa or sugar beets, represents a beneficial reuse of this process water. But, over application of process water, particularly in sandy soils, can cause nitrate contamination of shallow aquifers at levels approaching 10 times the Federal standard for drinking water. The critical question then is:

*How can the reuse benefits of this nitrogen-laden process water be maximized in a way that protects groundwater while maintaining the economic viability of the food processor?*

Our central hypothesis was that deeply rooted poplar trees increase the vadose zone dwell time, resulting in enhanced nitrogen removal. A pilot-scale system (Figure 1) was utilized to test the impact of poplar tree root zones on dwell time and on reactive transport of organic carbon and nitrogen that was applied via an intermittently-dosed synthetic wastewater. The results from the pilot-scale system were used to calibrate a 2-dimensional conservative and reactive transport model (Hydrus 2D/3D) configured with the Constructed Wetlands 2D biological processes module. The Hydrus model was then used to predict the impact of nitrate on groundwater using various year-round irrigation scenarios with consideration of spatial and temporal effects inherent to the full-scale application of this enhanced vadose zone nitrogen removal approach.

Pilot-scale data indicated a significant increase in dwell time attributable to poplar tree roots in sandy soils. This dwell time enhancement led to a commensurate increase in denitrification during dormancy (4.5 ± 2°C). While significantly less nitrate was measured in the effluent of rooted test cells, the nitrate concentration in all test cell effluents increased over time to levels exceeding 70 mg-N L⁻<fnref n="1" />. The synthetic wastewater contained less than 1 mg-N L⁻<fnref n="1" /> nitrate; therefore, the steady increase of effluent nitrate was attributed to hydrolysis of accumulated, slowly degradable organic nitrogen followed by rapid nitrification. A nitrogen mass balance analysis was performed to estimate the dwell-time specific, first-order rate coefficients for hydrolysis and denitrification at the pilot-scale.

The Hydrus model was able to accurately fit (R<fnref n="2" /> > 0.94; RMSE < 0.052) van Genuchten shape, saturated hydraulic conductivity, dispersivity, and immobile water content parameters from pilot-scale bromide tracer studies. The biochemical parameters considered were dissolved oxygen, organic matter (readily and slowly degradable and inert), ammonium, nitrite, nitrate, nitrogen gas, inorganic phosphorus, and heterotrophic and autotrophic organisms. Heterotrophic bacteria were assumed responsible for hydrolysis, OM mineralization, and denitrification. Autotrophic bacteria were assumed responsible for a two-step nitrification process (NH₄⁺ to NO₂⁻ and NO₂⁻ to NO³⁻). Initial values for these parameters were obtained from [Langergraber & Šimunek, 2005] and over 50 model iterations were performed to determine optimal values. Once optimized, simulations of 720 hours of intermittent dosing were run for both the rooted and non-rooted scenarios. The effluent nitrate concentration in the rooted simulation was 6.4 mg/L compared to 12.0 mg/L in the non-rooted system. The simulated impact of roots on nitrate was consistent with the effluent nitrate concentrations measured in early pilot-scale testing cells with roots (1.0 mg/L) and without roots (5.1 mg/L). The study provides strong evidence that less nitrate escapes a deeply rooted vadose zone than a non-rooted, or shallow rooted, system even during dormancy (4.5°C). Results from Hydrus simulations at larger spatial and temporal scales are forthcoming.

**Reference:** Langergraber, G., & Šimunek, J. (2005) Modeling variably saturated water flow and multicomponent reactive transport in constructed wetlands. *Vadose Zone Journal, 4*(4), 924-938. doi:10.2136/vzj2004.0166

**Acknowledgements:** Funding provided by the Port of Morrow, Boardman, Oregon under UI Project #1186790.

<figure><img src="image_458.png" /><figcaption>Figure 1: Pilot-scale testing system with dosing tanks, irrigation system and 10 cells.</figcaption></figure>

<figure><img src="image_459.png" /></figure>

<footnote n="1">¹ Civil and Environmental Engineering, University of Iowa, Iowa City, Iowa, USA</footnote>

<footnote n="2">² Ecolotree, Inc., North Liberty, Iowa, USA</footnote>
# REMOVAL OF CHROMIUM (VI) FROM THE AQUEOUS SOLUTION USING PERSIMMON TANNIN GEL

## Tomonobu Hatano, Takehiko Tsuruta

Hachinohe Institute of Technology, Japan

Corresponding author email: tsuruta@hi-tech.ac.jp

## ABSTRACT

Chromium is used in many items, such as textile, leather tanning, electroplating, metal finishing industries, wood treatment, corrosion control, oxidation, and anodizing industries. In the case of absorption was done at high levels, chromium can generate serious problems and, when concentration reaches 100 µg/g body wt., it can ultimately become lethal. Recently, the main processes for elimination chromium are adsorption, reverse osmosis and chemical reactions that contain reduction and precipitation. Among them adsorption has been shown as a feasible alternative method for removing traces of chromium from wastewater. Many different adsorbents were tried to remove chromium from wastewaters.

We have reported that microorganisms could remove many kinds of toxic and useful metals, such as lithium, uranium, thorium, rare earths, and gold from the aqueous solution. Additionally, immobilized persimmon gel removed gold (III) from hydrogen tetrachloroaurate(III) solution. However, microorganisms could remove little amount of chromium from the chromium (VI) solution. Therefore, the removal of chromium from aqueous chromium (VI) solution using immobilized persimmon gel in this study was examined.

The removal of chromium using persimmon tannin adsorbent was strongly affected by the pH of the solution. The amounts of chromium (VI) removed were quantitatively under pH 2, however, those were decreased with increasing the pH of the solution. The amount of total chromium removed was maximal at pH 2, however, those were decreased with increasing or decreasing the pH of the solution. It seems that the positively charged protonated hydroxy groupes of persimmon tannin and negatively charged hydrogen chromate ions are bonded at low pH region. Additionally, the amount of reduced chromium was increased with decreasing the pH of the solution.

<figure><img src="image_460.png" /><figcaption>Fig. 1. The effect of pH on the removal of Cr by persimmon tannin gel.</figcaption></figure>

15 mg (dry wr. basis) of the gel were suspended in 100 ml of the solution containing 2.5 ppm of chrome for 1h at room temperature.

The removal of chromium using immobilized persimmon tannin adsorbent was strongly affected by the chromium concentration of the solution. Immobilized persimmon tannin adsorbent can remove chromium quantitatively from the solution containing lower 5 ppm chromium (VI), however, the ratio of the amount of chromium (VI) removed toward that of initial chromium (VI) was decreased with increasing the initial chromium (VI) concentration.

The amount of chromium (µmol/g dry wt. adsorbents) was increased with increasing the initial chromium (VI) concentration of the solution. When the initial chromium (VI) concentration was 1700 µM, 4860 µmol of chromium (VI) was removed. The amount of chromium removed was fitted with *Langmuir's* isotherm. The amount of reduced chromium was maximal from the solution containing 400-800 µM chromium (VI).

The removal of chromium using immobilized persimmon tannin absorbents was strongly affected by the adsorbent amounts. The ratio of the amount of removed chromium (VI) per that of initial chromium (VI) was increased with increasing the amount of adsorbent, however, the amount of chromium removed (µmol/g dry wt. adsorbent) was decreased with increasing the amount of adsorbent. The amount of total reduced chromium was increased with increasing the amount of adsorbent, however, the amounts of reduced chromium (µmol/g dry wt. adsorbent) was maximal using 10-15 mg of adsorbent (dry wt. basis).

Adsorption of chromium (VI) was very rapidly and reached equilibrium within two hours. Reduction of chromium (VI) was a little slower than adsorption. The time course of chromium removal was fitted by an adsorption model using time-dependent *Langmuir equation*.

<figure><img src="image_461.png" /></figure>
# THE IMPORTANCE OF PLANTS WHEN USING *IN SITU* CARBON SORBENTS FOR THE REMEDIATION OF ORGANIC CONTAMINANTS

Mackenzie J. Denyes<fnref n="1" />, Allison Rutter<fnref n="2" />, Barbara A. Zeeb<fnref n="1" />

## ABSTRACT

Risk based remediation technologies for organic contaminants are gaining in popularity. An example of such a technology is the use of carbon sorbents, specifically biochar and activated carbon (AC), which immobilize the contaminant *in situ* and thereby reduce contaminant uptake in plants and invertebrates. To date, most studies have focused on AC and in sediment systems and there is less data available for biochar, which is a greener more cost-effective material capable of improving plant growth and sustainably sequestering carbon. Our previous work demonstrates that the addition of 2.8% biochar and AC to highly PCB-contaminated soil (>50 µg/g, Aroclor 1254/1260) reduces the phytoavailability of PCBs by ~70%. These carbon amendments also reduced the bioavailability of PCBs to *Eisenia fetida* (worm) (average 57% reduction) and restored the microbial community structure and function to that of a remediated site ([PCB]~soil~ < 3 µg/g). However, when applying this technology to a different organic contaminant, DDT, neither AC nor biochar were successful in minimizing DDT phytoavailability. These findings are in contrast to the existing literature which reports that AC reduces DDT bioavailability by greater than 90%. Many of these studies originate from researchers studying sediment systems. Generally, these studies have utilized equilibrium passive sampling devices based on polyoxymethylene (POM) (a polymer) to estimate the effectiveness of carbon sorbents, and have largely neglected the role that plants play in contaminant mobility. For this reason, we included both passive POM sampling and worm bioavailability studies in our two-year plant trials with carbon amendments. We found that carbon amendments significantly reduced the uptake of contaminants into the POM samplers by 32% and into worms by 50%, while no significant reductions in plant uptake were achieved. Thus, although passive samplers may be adequate for predicting contaminant availability to invertebrates such as worms, further research is required using plants to determine the actual potential of carbon amendments in soil systems as a remediation technology.

<figure><img src="image_462.png" /></figure>

<footnote n="1">¹ Department of Chemistry and Chemical Engineering, Royal Military College of Canada, Kingston, ON, K7K 7B4 Canada</footnote>

<footnote n="2">² School of Environmental Studies, Biosciences Complex, Queen's University, Kingston, ON, K7L 3N6 Canada
*Corresponding author email: mackenzie.denyes@rmc.ca*</footnote>
# ASSESSING THE AMENDING POTENTIAL OF LOW CARBON FOOTPRINT MATERIALS IN Zn MINING AREAS

Gabriel Caixeta Martins<fnref n="1" />, Ingrid Fernanda Santana Alvarenga<fnref n="2" />, Luiz Roberto Guimarães Guilherme<fnref n="1" />

Corresponding author email: guilherm@dcs.ufla.br

## ABSTRACT

Mining activities are essential for society's development. Yet, they cause environmental impacts, which must be minimized so that the benefits of this activity can overlap the likely risks it might pose. Open-pit mining often involves the removal of the soil overlaying the ore body. At the end of the exploration process, the areas contain materials with heterogeneous soil physicochemical characteristics, often unsuitable for plant growth. Improving soil chemical, biological, and physical conditions is a requirement for recovering most of these areas. For that, it is often necessary to apply suitable soil amendments in order to alleviate the potential phytotoxicity and/or provide plant nutrients. Such amendments must be chosen not only based on their efficiency and cost, but also according to their carbon footprint, which is currently a trend for any "green remediation" proposal. This study aims at evaluating the amending potential of biocompounds, limestone, sewage sludge, and biochar in a substrate collected from Zn-mine degraded areas. Our main goal is to select the best soil amendment for the recovery of these areas – with a focus on low-cost, locally produced materials –, using *Andropogon gayanus* as cover crop for a phytotechnology project.

The experiment was conducted under greenhouse conditions with soils samples collected from a zinc mining area located in the State of Minas Gerais, Brazil (clay 560 g kg⁻<fnref n="1" />; sand 145 g kg⁻<fnref n="1" />; pH 5.9; P 3,5 mg dm⁻³; K 14.8 mg dm⁻³; Ca 0.9 cmol_c dm⁻³; Mg 0.7 cmol_c dm⁻³; Al 0.4 cmol_c dm⁻³; effective CEC 3.8 cmol_c dm⁻³; Zn_total 530 mg L⁻³; Cd_total 1.6 mg L⁻³; Pb_total 1,253 mg L⁻³). We tested four amendments and five increasing rates for each material, with three replicates: a wood-based biochar residue (0; 2.5; 7.5; 15; 30 g kg⁻<fnref n="1" />), a sewage sludge (0; 1.25; 2.5; 5; 10 g kg⁻<fnref n="1" />), a by-product limestone (0; 0.5; 1; 2; 4 g kg⁻<fnref n="1" />), and a biocompound (0; 1.25; 2.5; 5; 10 g kg⁻<fnref n="1" />). All amendments were collected at sites nearby the degraded area, being locally produced or occurring as by-products of neighboring industrial activities. The materials were added to the soil and incubated for 60 days with soil humidity at 60% of field capacity. After incubation, soils were tested for pH (in water) and phytoavailable levels of Cd, Pb, and Zn and subsequently sowed with 15 *Andropogon gayanus* seeds, from which we evaluated germinability, plant survival and shoot height.

Increasing limestone and biochar rates caused soil pH to increase up to 2.5 units, hence reducing phytoavailable Cd (~32% limestone and ~57% biochar) and Zn (~28% limestone and ~41% biochar), with little effect upon the availability of Pb (~5% biochar and ~3% limestone). Adding sewage sludge and biocompound had little effect on soil pH, yet they still reduced the availability of Cd (~7% sewage sludge and ~14% biocompound), while causing Zn phytoavailability to decrease (biocompound ~24%) or increase (~14% sewage sludge). Phytoavailable Pb increased slightly following the application of sewage sludge (~8%) and biocompound (~10%).

Germinability was affected differently due to the treatments, with little effect caused by biochar and a trend of reduction caused by adding sewage sludge, limestone, and biocompound. Nevertheless, despite the observed reductions in the germination of *Andropogon gayanus* seeds, all treatments increased plant survival, with the largest increases in *A. gayanus* survival being observed with the addition of 5 g kg⁻<fnref n="1" /> of sewage sludge (~122%) and 10 g kg⁻<fnref n="1" /> of biocompound (~138%). Similarly, shoot height was also positively affected by the addition of all soil amendments, with best results observed following the application of 1 g kg⁻<fnref n="1" /> limestone (~90%), 10 g kg⁻<fnref n="1" /> of biocompound (~70%), 15 g kg⁻<fnref n="1" /> biochar (~54%), and 5 g kg⁻<fnref n="1" /> of sewage sludge (~51%). This positive effect is possibly a result of improvements in nutritional status.

Considering their local availability and low cost and due to the improvements caused in soil chemical attributes, as well as on plant survival and shoot height, we conclude that all evaluated materials could be used as soil amendments for the studied Zn mine area, being suitable for further testing under field conditions. Additional research is underway to assess if the combined use of the tested soil amendments could result in enhancement of the beneficial effects.

**Acknowledgements:** The financial support provided by FAPEMIG, CAPES and CNPq.

<figure><img src="image_463.png" /></figure>

<footnote n="1">¹Soil Science Department, Federal University of Lavras, Brazil</footnote>

<footnote n="2">²Biology Department/Vegetal Physiology Sector Federal University of Lavras, Brazil</footnote>
# LED LIGHT QUALITIES AS AN ARTIFICIAL STIMULUS IMPROVE THE GROWTH OF TWO MEDITERRANEAN SPECIES CULTIVATED INTO A CONTROLLED ENVIRONMENT

Sonia Smirnakou<fnref n="1" />, Kalliopi Radoglou<fnref n="2" />

Corresponding author email: kradoglo@fmenr.duth.gr

## ABSTRACT

Plant illumination is particularly important in a controlled environment. Light-emitting diodes (LED) is a promising light source used in plant physiology research. However there have been very few investigations on the application of LEDs in plant growth chambers. In the present study we evaluate the effects of different LED or fluorescent lights on the growth traits of two Mediterranean species, *Arbutus unedo* L. and *Myrtus communis* L. seedlings cultivated in mini-plug containers fulfilled with stabilized medium soil substrate. Seedlings were uniformly grown into two growth chambers for seven weeks under light-emitting diodes (LEDs) (L20AP67, AP673L,G2, AP67, NS1 -Valoya) or Fluorescent light (FL).LEDs used emitted a continuous spectrum thanks to a mixture of ultraviolet, blue, green, red, far-red and infra-red in various percentages. Furthermore root growth potential (RGP) was used as a performance attribute for seedling quality evaluation.

During the seven week experimental period the effect of different light treatments was significant and species dependent by means of height growth rate, the leaf colour rating and the leaf number. L20AP67 LED and FL light induced significantly taller seedlings but showed less beneficial effect on the root development for both species compared to the rest of LEDs. *Arbutus unedo* seedlings showed significantly lower dry weight mass of leaves, shoots and roots under the illumination of FL and L20AP67. While *Myrtus* seedlings better adapted under

the AP67 LED illumination for the same parameters tested. Root-to-shoot ratio for both species obtained significantly lower under the FL light than LEDs. After 31 days into the RGP bath, *Arbutus* seedlings pre-cultivated under the G2 LED better predicted the new root length and dry weight thus almost doubled. For *Myrtus* seedlings that pre-cultivated under the FL no significant effect found for the new root length, while G2 LED obtained the heaviest roots among all treatments.

<figure><img src="image_464.png" /></figure>

<figure><img src="image_465.png" /></figure>

These results confirm and extend the results of other studies indicating that light quantity and quality interact to determine plant morphology. A thorough understanding of this interaction is essential to the development of light sources for optimal plant growth and development.

**Acknowledgements:** The financial support by FP-7 project REGENFOREST contract No. 286067 is greatly appreciated.

<figure><img src="image_466.png" /></figure>

<footnote n="1">¹ Forest Research Institute, Hellenic Agricultural Organization "DEMETER", Thessaloniki, Greece, e-mail: sosmirnakou@gmail.com</footnote>

<footnote n="2">² Department of Forestry and Management of the Environment and Natural Resources Democritus University of Thrace (DUTH), Greece</footnote>
# PHYTOEXTRACTION AND ECOLOGICAL CATALYSIS: SYMBIOSIS FOR FUTURE

## Pr. Claude Grison

CNRS-STRATOZ - Cap Alpha - Avenue de l'Europe- 34830 Clapiers, France

Corresponding author email: claudegrison@cefe.cnrs.fr

## ABSTRACT

Phytotechnologies such as metals phytoextraction are very attractive solutions for the reclamation of contaminated brownfields. However, the economic potential of these technologies remains poor, due to a lack of credible outlets for metal-contaminated biomass. Strong efforts are still necessary to recycle metals from contaminated-biomass, so as to develop the full economic potential of phytoextraction. This is particularly true in a context of resource depletion, with increasing metals prices and market volatility.

In this communication, we aim to present a new outlet for metal contaminated-biomass. Phytoextraction generates plant wastes, which are valued through an innovative concept of ecological recycling. Because certain plants are able to hyperaccumulate primary or strategic metals as Ni2+, Mn2+, Cu2+, Co2+, Zn2+ and Pd2+ can be directly used as catalysts in organic fine chemical reactions. Ecocatalysts derived from hyperaccumulators are useful for the synthesis of molecules with high added value in fine and industrial chemistry (aroma and cosmetics with the «natural» label, drugs and oligomers with biological interests, highly functionnalized aromatic heterocycles, chiral structures, key intermediate in various industrial chemical process and biopecticides). They allow the development of substitutive reagents to oxidants forbidden by REACH too. The new catalysts represent very efficient alternative to catalysts derived from metallurgy, with a three dimension domain of chemistry/ecology/environment.

This original approach brings the first perspective of enhancing the unique biomass and establishes a new field of Green Chemistry: Ecocatalysis. It is already a Green Revolution in Chemistry. Phytoextraction are source of innovation in organic synthesis and organic synthesis is the driving force of developed phytotechnologies.

These results are the first use of metal-rich biomass in organic synthesis and constitute a strong encouragement for the economic development of phytoextraction programs for metal-bearing soils.

## Selected References:

1. Design and performance of supported Lewis acid catalysts derived from metal contaminated biomass for Friedel-Craft's alkylation and acylations, Catalysis Today 189 (2012) 111-116, G. Losfeld, V. Escande, P. Vidal De La Blache, L. L'Huillier and C. Grison

2. The chemical exploitation of nickel phytoextraction: An environmental, ecologic and economic opportunity for New Caledonia, Chemosphere 89 (2012) 907-910, G. Losfeld, V. Escande, T. Jaffré, L. L'Huillier and C. Grison

3. Lewis-acid catalysts: a potential exploitation for zinc and nickel phytoextraction, Environmental Impact 2012, WIT Transactions on ecology and the environment vol. 186 G. Losfeld, P. Vidal De La Blache, V. Escande and C. Grison

4. Metallophyte wastes and polymetallic catalysis: a promising combination in green chemistry. The illustrative synthesis of 5'-capped RNA, RSC Advances 15 (2013) 5204-5212, Y. Thillier, G. Losfeld, V. Escande, C. Dupouy, J.-J. Vasseur, F. Debart and C. Grison

5. Ecological catalysis and phytoextraction: symbiosis for future, Applied Catalysis B: Environmental in Press 2013, V. Escande, L. Garoux, C. Grison, Y. Thillier, F. Debart, J.-J. Vasseur, C. Boulanger and C. Grison

6. Psychotria douarrei and Geissois pruinosa, novel resources for the plant-based catalytic chemistry, RSC Advances in Press 2013, C. Grison, C. Grison, V. Escande, C. Boulanger, L. Garoux and E. Petit

<figure><img src="image_467.png" /></figure>
# SOCIAL AND ENVIRONMENTAL ASPECTS FROM THE IMPLEMENTED RENEWABLE ENERGY PARKS (SOLAR, AIR, BIOMASS) IN CYPRUS

Michalis Tsangas, Antonis A. Zorpas

Cyprus Open University, Faculty of Pure and Applied Science, Environmental Conservation and Management, P.O.Box 12794, 2252, Latsia, Nicosia, Cyprus

Corresponding author email: antonis.zopras@ouc.ac.cy; antoniszorpas@yahoo.com

## ABSTRACT

Renewable energy sources are, nowadays, not only a necessity but also a choice. Many technologies to exploit them have been established and operate all over the world, producing energy and offering multiple benefits. However the renewable energy projects are not always without negative impacts to the environment and the society, which must be effectively evaluated and confronted, in order only the positive impact to remain. However the research question is does this happen in all cases? The aim of this paper is to evaluate from a social and an environmental aspect, renewable energy projects which has been established in Cyprus previous year, in order to find out if they are licensed and constructed effectively, if the necessary preventive measures are implemented against social and environmental impacts during the projects operation and which kind of renewable energy sources is implemented more efficiently, socially and environmentally. The object is also to infer conclusions for the renewable energy sources sector in Cyprus. A number of techno economic, institutional, environmental and social criteria have been appointed, according to which the evaluation of renewable energy projects is performed. Data from three different established renewable energy projects (solar park, air park, Biomass Park) have been collected. Firstly it is tested if their social and environmental performance presents variation. Afterwards the projects are compared according to the above criteria, in order to identified, which project is more efficiently. Based on the results and of literature review, a SWOT analysis was prepared for the renewable energy sector in Cyprus. The results indicated that the projects do not differ from each other and that they run efficiently, environmentally and socially. Based on the multicriteria analysis the photovoltaic park emerges as the more socially and environmentally efficient, with the wind parks coming second and the biogas unit third. Finally a significant number of strengths and opportunities, and a comparatively smaller number of weaknesses and threats are concluded for the renewable energy sector in Cyprus.

<figure><img src="image_468.png" /></figure>
# DEVELOPING A PRACTICAL DECISION SUPPORT TOOL (DST) FOR THE APPLICATION OF PHYTOTECHNOLOGIES

Andrew Cundy<fnref n="1" />, Kenechukwu Onwubuya<fnref n="1" />, Paul Bardos<fnref n="1" />,<fnref n="2" />, Markus Puschenreiter<fnref n="3" />, Nele Witters<fnref n="4" />, Jaco Vangronsveld<fnref n="4" />, Michel Mench<fnref n="5" />, Ingo Mueller<fnref n="6" />

Corresponding author email: A.Cundy@brighton.ac.uk

## ABSTRACT

A range of systems and tools have been proposed to support decision making within the contaminated land arena. A critical review, coupled with stakeholder feedback collected during the European Commission FP6 ERANET project SUMATECS and presented in Onwubuya et al (2009), has highlighted a general lack of stakeholder knowledge of these systems and tools, and specifically of those which can be used to support the selection and application of phytotechnologies. Based on stakeholder feedback, existing tools were regarded as too general (containing insufficient detail on the range of remediation options available), or alternatively were too complicated for regular or widespread use by decision makers in selecting and applying phyto-based (and indeed other) remediation technologies. Following the SUMATECS project, the European Commission FP7 GREENLAND project was established to overcome a range of barriers to the practical application of phytotechnologies and other gentle remediation options (GRO). A key component of this project was the development, using data from long-term (> 5 year) phytotechnology site trials across Europe, of a practical and simple decision support tool (DST) focused on the wider contaminated land community. This DST is presented here.

The GREENLAND DST is a simple Microsoft Excel-based tool targeted on non-specialists which follows (in line with existing national decision support guidelines) a tiered approach (below, right). The tool is designed to interface with existing national guidance at the options appraisal stage, although the DST also has applicability at earlier (site planning) stages. The tiered approach used consists of three phases: phase 1 (feasibility), phase 2 (semi-quantitative assessment) and phase 3 (technical assessment).

Each phase terminates in a decision point (Yes = proceed to next phase; No = return to options appraisal), with increasing complexity and time investment requirements from phase 1 to 3. Phase 1 includes: (a) a (qualitative) contaminant matrix, in the format of the UK's CLR-11 remediation option applicability matrix, which forms a decision checklist for the site to be remediated, and consists of a list of phytotechnologies which have been checked against a list of Contaminants of Concern (CoC) through detailed literature review in order to provide evidence that the methods have previously been used in the remediation of the specified contaminants on (at least) a research scale; and (b) examples of practical, large-scale phytotechnology applications.

Phase 2 contains modules on: stakeholder engagement (including guidelines for stakeholder engagement and criteria for the

identification of different stakeholder profiles/ categories); sustainability assessment (drawing on SURF-UK sustainability indicators, and tables showing potential economic, environmental and social benefits); and economic assessment (using a simplified “cost calculator” for quantifying (within limits) site costs and local and wider economic benefit). Phase 3 contains detailed technical information for practical remediation project design and implementation, and outlines “operating windows” for phytotechnology application. The DST has been populated and “calibrated” using data from long-term European site trials within the GREENLAND site network, and a major redevelopment project in east London, UK. Ongoing work is aimed at finalising the DST using an iterative approach involving validation by a range of experts and potential end-users.

**Acknowledgements:** The authors are grateful for financial support from the European Commission under the Seventh Framework Programme for Research (FP7-KBBE-266124, GREENLAND).

<figure><img src="image_469.png" /></figure>

*Increasing complexity and time investment*

*"N" and red arrow indicates return to Options Appraisal in national guidance*

<figure><img src="image_470.png" /></figure>

<footnote n="1">¹School of Environment and Technology, University of Brighton, UK</footnote>

<footnote n="2">²r3 Environmental Technology Ltd., Reading, UK</footnote>

<footnote n="3">³University of Natural Resources and Life Sciences (BOKU), A-3430 Tulln, Austria</footnote>

<footnote n="4">⁴Centre for Environmental Sciences (CMK), Hasselt University, 3590 Diepenbeek, Belgium</footnote>

<footnote n="5">⁵UMR BIOGECO INRA 1202, University of Bordeaux, F-33405 Talence, France</footnote>

<footnote n="6">⁶Saxon State Agency for Environment, Agriculture and Geology, D-01109 Dresden, Germany</footnote>
# EFFECT OF ANTIOXIDANT RICH SPICES, CLOVE AND CARDAMOM EXTRACTS ON THE METABOLIC ENZYME ACTIVITY OF LABEO ROHITA

**Oyas Ahmed Asimi<fnref n="1" />, Norattam Puri Sahu<fnref n="2" />, Asim Kumar Pal<fnref n="3" />**

*Corresponding author email: asimioa@gmail.com*

## ABSTRACT

Backgrout: Herbs, spices, and various plant extracts have received increased attention as possible replacement of antibiotics. In this view, aromatic substances and essential oils extracted from various plants became interesting due to their antimicrobial [Guler, et al., 2005], antioxidant [Dragland, et al., 2003] effects and their stimulating effects on animal performance [Ciftci, et al., 2005, Guler, et al., 2005b] and digestive enzymes [Lee, et al., 2003]. These plant extracts are usefull in reducing stress.

Objectives: Studies indicate that clove and cardamom have a number of medicinal properties and is a potent antioxidant, the present study was undertaken to evaluate their anti-stress effect in *Labeo rohita*.

Methods: Spice extracts, Experimental design and diets, Enzyme analysis, Statistical analysis.

Results: The present study was conducted to evaluate the protective effect of clove (Clo) and cardamom (Cdm) extracts as natural antioxidants in the diet of *Labeo rohita* fingerlings. Both the spice extracts were mixed separately or in combination at the level of 0.5 and 1.0% in the fish feed. Thus, seven experimental diets were the following Clo-0.5, Clol-1.0, Cdm-0.5, Cdm-1.0, CC-0.5 and CC-1.0 and control with no extract. The results revealed that, SOD and CAT activities were significantly (p<0.05) higher in Clo-0.5 compared to the control. AST, ALT, LDH, MDH and G-6-Pase activities in liver and muscle decreased significantly (p<0.05) in all treated groups as compared to the control. Significantly (p<0.01) lowest activities was observed in treatment group Clo.-0.5%.

Conclusion: The results suggest that the dietary administration of clove extract supplementation at a concentration of 0.5% level possess good anti-stress activity.

Acknowledgements: The first author is highly thankful to the Director/Vice-chancellor, Central Institute of Fisheries Education, Mumbai, India for providing facilities for carrying out the work.

<figure><img src="image_471.png" /></figure>

<footnote n="1">¹ Fish Nutrition & Biochemistry, Faculty of Fisheries, Sher-e-Kashmir University of Agricultural Sciences and Technology-Kashmir, India</footnote>

<footnote n="2">² Fish Nutrition, Biochemistry & Physiology, Central Institute of Fisheries Education, Mumbai, India</footnote>

<footnote n="3">³ Joint Director, Central Institute of Fisheries Education, Mumbai, India</footnote>
# PROCESSING OF PLANT BIOMASS HARVESTED AT TRACE ELEMENT-CONTAMINATED SITES MANAGED BY GENTLE (PHYTO) REMEDIATION OPTIONS

Valérie Bert<fnref n="1" />, Serge Collet<fnref n="1" />, Florence Delgratta<fnref n="1" />, Jannis Dimitriou<fnref n="2" />, Rolf Herzig<fnref n="3" />, Jolien Olga Janssen<fnref n="4" />, Jaco Vangronsveld<fnref n="4" />, Silke Neu<fnref n="5" />, Michel Mench<fnref n="6" />, Anne Loppinet-Serani<fnref n="7" />, Patrick Lemaître<fnref n="8" />, Mathieu Chaillou<fnref n="8" />, Carine Richer<fnref n="8" />, Markus Puschenreiter<fnref n="9" />

Corresponding author email: valerie.bert@ineris.fr

## ABSTRACT

Depending on the GRO (phytoextraction, in situ immobilization and aided phytostabilization) set up on a polluted site and the type of plant used, harvested plant parts may contain concentrations of trace elements (TE) that may be higher than those found in similar vegetation grown on uncontaminated soils (background). To render such technologies economically attractive and feasible, harvested biomass should be valuable and enter valuation pathways. On contaminated lands, plants may serve to provide feedstocks for bioenergy, non-food products and biofuels and, thus, may contribute to achieve the EU aim by 2020, i.e. to get 20% of its energy from renewable sources. As a potential advantage, these plants will not compete with plants grown on uncontaminated lands as contaminated lands are not suitable for food production. In GREENLAND (http://www.greenland-project.eu/), one task aimed at testing various conventional and innovative technologies of biomass valorisation, such as combustion, anaerobic digestion, pyrolysis and solvolysis, and determining the fate of the TE in the resulting products of each conversion process. Assays were carried out on a wide range of plant species cultivated at the field trials of the GREENLAND partners.

Figure 1 shows the distribution of Zn in the emissions, i.e. bottom ash, particulate fraction (fly ash) and gaseous fraction of the flue gas, as a result of combustion assays performed in a biomass boiler of 40kW on willows and poplars cultivated for phytoextraction purposes and the comparison with corresponding virgin wood (control).

For all assays, Zn occurred mainly (> 50%) in the fly ash. The bottom ash represents the second compartment for the occurrence of Zn whereas the gaseous fraction of the flue gas represents a minor compartment for Zn emissions. The distribution is not depending on the initial burnt wood, i.e. virgin wood (control) or Zn enriched wood (phytoextraction). The use of TE enriched wood in biomass boilers seems possible if they are equipped with efficient filters to avoid air pollution. Results are in accordance with [1,2]. Anaerobic digestion, pyrolysis and solvolysis assays were performed in batch reactors. Results from all these conversion technologies will be presented and discussed as well as the possibility to use end products of the processes.

<figure><img src="image_472.png" /></figure>

The valuation of plants produced by GRO seems possible if TE do not disturb the functioning and the performance of the process and if such plant use complies with current regulation.

## References:

1. Delplanque, M., et al. (2013) *Biomass and Bioenergy*, 49, 160-170.

2. Chalot, M., et al. (2012) *Environnemental Science and Technology*, 46(24), 13361-9.

**Acknowledgements:** This work was financially supported by European funds through GREENLAND (FP7, KBBE-2010-4, 26124).

<figure><img src="image_473.png" /></figure>

<footnote n="1">¹INERIS, Technologies and Sustainable and Clean Processes, Parc Technologique Alata, BP2, 60550 Verneuil en Halatte, France</footnote>

<footnote n="2">²Department of Crop Production Ecology, Swedish University of Agriculture Sciences, SE-750 07 Uppsala, Sweden</footnote>

<footnote n="3">³Phytotech Foundation, Umweltbeobachtung und oekologische Planung Quartiergasse 12, 3013 Bern, Switzerland</footnote>

<footnote n="4">⁴Hasselt University, Centre for Environmental Sciences, Agoralaan building D 3590 Diepenbeek, Belgium</footnote>

<footnote n="5">⁵Saxon state office for environment, Agriculture and Géologie, Referat 42, Boden, Altlasten, Department 42, Soil, Contaminated Sites, Halsbrücker Straße 31a, 09599 Freiberg, Germany</footnote>

<footnote n="6">⁶INRA, UMR BIOGECO INRA 1202, University of Bordeaux 1, 33405 Talence, France</footnote>

<footnote n="7">⁷CNRS, Université de Bordeaux, ICMCB, 87 avenue du Dr. A. Schweitzer, Pessac, F-33608, France</footnote>

<footnote n="8">⁸INNOVEOX SAS, 18 Rue D'Aguesseau, 75008 Paris, France</footnote>

<footnote n="9">⁹University of Natural Resources and Life Sciences, Department of Forest and Soil Sciences, Konrad Lorenz Strasse 24, 3430 Tulln, Austria</footnote>
# SEMI-CONTINUOUS TREATMENT OF PENTACHLOROFENOL BY BIOLOGICAL FENTON REACTION

Yoshihiko Inagaki<fnref n="1" />, Shoichi Nagahashi<fnref n="1" />, Ami Fujiwara<fnref n="1" />, Andre Rodrigues dos Reis<fnref n="2" />, Yutaka Sakakibara<fnref n="1" />

Corresponding author email: sakaki@waseda.jp

## ABSTRACT

**Introduction:** Environmental pollutants such as pharmaceutical and personal care products (PPCPs) and endocrine disrupting chemicals (EDCs) have been detected in aquatic environment. Recent studies demonstrated EDCs can be treated by plants, plant tissues, and/or plant enzymes. We proposed a biological Fenton reaction (BFR), by which organic pollutants are oxidized with hydroxyl radicals produced from endogenous hydrogen peroxide ($H_2O_2$) in the presence of ferrous ions ($Fe^{2+}$). However, BFR was investigated only in batch treatments, and continuous or semi-continuous performance has not been known. Here we applied BFR to semi-continuous treatment of a recalcitrant pollutant, pentachlorophenol (PCP). In addition, in order to verify the oxidation performance, electron spin resonance (ESR) technique was employed to detect hydroxyl radicals produced by BFR.

**Methods used:** Aquatic plants used in this study were floating plants, Amazon frogbits (*Limnobium laevigatum*). Semi-continuous experiments were designed to repeat a fill and draw treatment 5 times, where solution in vessel was changed to feed solution every 3 days. Plants (20g fresh weight (FW)) were cultivated in a 2.5 L glass vessel and were maintained under intermittent illumination at 3000lx provided by white fluorescent lamps with light periods of 16 hours and dark periods of 8 hours. Concentrations of PCP and $Fe^{2+}$ in the feed solution were set at 0.38 in first two cycles and 0.76µM in the rest cycles, and 0.18mM, respectively, and measurements were made for PCP and endogenous $H_2O_2$ concentrations. Control tests using no aquatic plant with iron compounds and aquatic plants with no iron compound were conducted. ESR analyses were made using DEPMPO (5-(diethoxyphosphoryl)-5-methyl-pyrroline-N-oxide) as a spin trapping agent in the presence and absence of iron compounds; and observed results were compared with that of Fenton reagents (0.05mM ferrous sulfate heptahydrate and 0.2mM $H_2O_2$). Aquatic plant samples (0.3g-FW) were ground using a mortar and pestle with 0.05mM $FeSO_4 \cdot 7H_2O$ and 0.2mM DEPMPO. The mixture was centrifuged at 12,000×g and 4°C for 5 minutes and the supernatant was used for the measurement of ESR.

**Results and discussion:** Figure 1 shows the result of semi-continuous experiments. Significant difference of PCP removals was observed repeatedly in the different conditions. Moreover, addition of iron compounds facilitated the removal of PCP in comparison with the other two conditions. Figure 2 compares three ESR spectrums for (1) plant with $Fe^{2+}$, (2) plant without $Fe^{2+}$, and (3) Fenton reagent. Six ESR spectrums peaks specific to DEPMPO-OH adduct, which are shown by the inverted triangle marks in the result for Fenton reagents, were detected in BFR, while no clear peaks were observed in the absence of iron compounds. Thus, hydroxyl radicals can be produced by the addition of iron compounds to aquatic plants. Measurement of endogenous $H_2O_2$ showed that hydrogen peroxide tended to decrease in the presence of $Fe^{2+}$, but was kept at least around 0.1mM in plants during semi-continuous experiment.

<figure><img src="image_474.png" /><figcaption>Figure 1: Time course change of PCP concentration under different conditions</figcaption></figure>

**Conclusions:** The possibility of a biological Fenton reaction (BFR) was demonstrated through a semi-continuous experiments and ESR analysis. A further study on operating conditions and long-term experiments will be needed to evaluate precise performance.

**Acknowledgements:** This study was supported in part by Grant-in-Aid for Challenging Exploratory Research (#23656334) and for JSPS Fellows (#25767).

<figure><img src="image_475.png" /><figcaption>Figure 2: ESR spectrums for three conditions</figcaption></figure>

<figure><img src="image_476.png" /></figure>

<footnote n="1">¹Dept of Civil and Environmental Engineering, Waseda University, Tokyo, Japan</footnote>

<footnote n="2">²Dept of Soil Science, Federal University of Lavras, Lavras, Brazil</footnote>
# THE USE OF POLYPHENOLIC COMPOUNDS OBTAINED FROM OLIVE OIL MILL WASTE (OMWW) AS ECOLOGICAL PLANT PROTECTION AGENT AGAINST SEVERAL FUNGAL PATHOGENS ON TOMATO PLANTS. STUDIES IN VITRO AND IN VIVO

S.V. Leontopoulos<fnref n="1" />, I. Giavasis<fnref n="2" />, K. Petrotos<fnref n="1" />, Ch., Makridis<fnref n="3" />

Corresponding author email: s_leontopoulos@yahoo.com

## ABSTRACT

The assessment of different concentrations and forms of polyphenols (free and encapsulated) from olive oil mill waste (OMWW) as plant protection materials against economically important plant pathogenic fungi in *in vitro* and *in vivo* tests is the main research area of this work. In the first stage of the experimental process, it was assessed the zone of inhibition in mm of several fungal pathogens using the methods of disk diffusion assay, and well diffusion assay. In the second stage of the evaluation, the effect of polyphenols against 14 fungal microorganisms was examined in order to determine the MIC / microbicidal concentration (MIC / MFC). In a later stage *in vivo* evaluation of liquid polyphenols (LFP) obtained from OMWW as natural bio-chemicals against several fungal pathogens on tomato plants was carried. The evaluation of the results obtained by determining the MIC and MLC, demonstrated that the fungus *Aspergillus flavus* appeared highly resistant to the LFP concentration required minimum sample rate > 35-40 % for inhibition and killing effects respectively. In order of major protection resulting from the use of polyphenolic compound against major diseases, *Botrytis cinerea*, *Sclerotinia sclerotiorum* and *Ascochyta lentis* gave the most promising results. Moreover, the use of low concentration of LFP at 5 and 10 % could control in some cases fungal pathogens. However, higher concentration of LFP (20 and 30%) appeared possible phytotoxic effects.

<figure><img src="image_477.png" /></figure>

<footnote n="1">¹ Department of Biosystems Engineering, Technological Educational Institute of Thessaly (T.E.I.), 41110, Greece</footnote>

<footnote n="2">² Department of Food Science, Technological Educational Institute of Thessaly (T.E.I.), 41110, Greece</footnote>

<footnote n="3">³ Department of Animal Production, Technological Educational Institute of Thessaly (T.E.I.), 41110, Greece</footnote>
# BIOMANAGEMENT OF PESTICIDE CONTAMINATED SOILS: MICROBIOLOGICAL METHODS FOR FEASIBILITY ASSESSMENT AND MONITORING

Nejla Hechmi<fnref n="1" />,<fnref n="2" />, Nadhira Ben Aissa<fnref n="2" />, Hassen Abdenaceur<fnref n="1" />, Naceur Jedidi<fnref n="1" />

Corresponding author email: nejlahechminet@yahoo.fr

## ABSTRACT

There is rising public concern as a wide variety of toxic organic chemicals are being introduced deliberately into the environment. Pesticides are one common example of these chemicals, which enter the soils frequently in recent years. Chlorinated phenols (CPs) are a group of ionisable organic compounds of major environmental concern. Chlorophenols, particular those with three or more chlorine atoms, have gained an increasing use as fungicides, herbicides and insecticides [1]. Pentachlorophenol (PCP) from the category of CPs generated harmful effects in aquaculture or soil systems [2]. Thus, the cleanup PCP contaminated soils is imperative and it is critical to develop efficient approaches to remove multiple contaminants from soils.

The need to remediate these natural resources has led to the development of new technologies that emphasize the destruction of the pollutants rather than the usual approach of disposal. Bioremediation is the use of biological interventions of biodiversity for mitigation of the noxious effects caused by environmental pollutants in a given site [3]. Bioremediation with divers' aspects more recently, phytoremediation has emerged as one of the alternative technologies for removing contaminants from the soil. Furthermore, monitoring entire microbial communities in soil has traditionally been a very time-consuming task.

In this contribution we used phospholipid fatty acid (PLFA) profiles, one based on the extraction of lipids directly from soil, indicating the effects of Pentachlorophenol on soil microbial community structure. Four native plant species, white clover, ryegrass, alfalfa, and rapeseed, were studied, in order to confirm which species of plants improved the biodegradation ability for chlorinated pollutant.

After 60 days cultivation, white clover, raygrass, alfalfa and rapseed all significantly enhanced the degradation of PCP in soils. The content of the biomarker PLFAs was significantly higher in the rhizosphere samples than the unplanted samples (p<0.05). Significant difference of the microbial community structure was observed in samples of alfalfa from other rhizosphere and unplanted samples, indicating the rhizosphere of alfalfa enhanced the tolerance of soil microorganisms to pentachlorophenol significantly.

Phytoremediation appeared to have great potential for the remediation of PCP contaminated soil. The presence of vegetation significantly promoted the dissipation of PCP in the soil environment. Remediation capacity varied greatly among plant species. The difference was attributed not just to the effect of higher overall biomass yields but to different rhizosphere microbial community structure.

**Keywords:** soil, pentachlorophenol, phytoremediation, phospholipid fatty acids, endoenzyme activity

## References:

1. Roy, S. and Hänninen, O. (1994), Pentachlorophenol: Uptake/elimination kinetics and metabolism in an aquatic plant, *Eichhornia Crassipes*, Environ Toxicol Chem 13, 763–773.

2. Hechmi, N., Ben Aissa, N., Abdenaceur, H. and Jedidi, N. (2013), Phytoremediation Potential of Maize (Zea mays L.) in Co-contaminated Soils with Pentachlorophenol and Cadmium. Int J Phytorem 15, 703–713.

3. Asha Latha, A., Reddy, S. (2013), Review On Bioremediation – Potential Tool For Removing Environmental Pollution, Int J Basic Appl Chem Sci 3(3), 21-33.

<figure><img src="image_478.png" /></figure>

<footnote n="1">¹ Laboratory of Wastewater Treatment, Water Research and Technologies Centre (CERTE), Technopole Borj Cedria BP 273, Soliman 8020, Tunisia</footnote>

<footnote n="2">² National Agronomic Institute of Tunisia, 43 Avenue Charles Nicolle City of Mahrajene 1082, Tunisia</footnote>
# OPTIMIZATION OF CADMIUM (Cd<fnref n="2" />⁺) REMOVAL FROM AQUEOUS SOLUTIONS BY NOVEL BIOSORBENT

Alaa El Din Mahmoud<fnref n="1" />, Manal Fawzy<fnref n="1" />, Ahmed Radwan<fnref n="2" />

Corresponding author email: alaa-mahmoud@alexu.edu.eg

## ABSTRACT

In this research, Dead leaves of a common ornamental plant, *Dracaena draca* known also as dragon tree was used as a biosorbent for the removal of Cadmium (Cd<fnref n="2" />⁺) from aqueous solutions using a full 2³ factorial experimental design. Three factors were investigated at two different levels, metal ion concentration (X=10 and 100ppm), hydrogen ion concentration (pH=2 and 7) and biomass dose (BD=0.1 and 0.5g). Experiments were carried out in duplicates with 50 ml of Cd<fnref n="2" />⁺ solutions at room temperature.

Cd<fnref n="2" />⁺ concentrations in the solutions were determined using Varian Inductively Coupled Plasma (ICP-AES). When comparing observed values (experimental) with calculated values (model), they were set closely together that allowed suggesting a normal distribution where (R<fnref n="2" />=0.9938). Characterizations of the biosorbent were done by FT-IR and SEM. The FT-IR analysis of biomass before and after biosorption indicated the involvement of hydroxyl, carboxylic acids, alcohol and carboxylate groups in biosorption of Cd<fnref n="2" />⁺.

Results also showed that the most significant effect for Cd<fnref n="2" />⁺ biosorption was ascribed to (X). The interaction effects of (pH BD) and (X pH) were found to have significant influence on Cd<fnref n="2" />⁺ removal efficiency. The highest Cd<fnref n="2" />⁺ removal percentage attained by 79.60% at X=10 ppm, pH=7 and BD=0.5g. The reusability of the biosorbent was tested in three desorption cycles and the regeneration efficiency was above 99.7%.

**Acknowledgements:** The financial support by Young Scientist Award, National Man and Biosphere Egyptian Committee (UNESCO) is greatly appreciated.

<figure><img src="image_479.png" /></figure>

<footnote n="1">¹Environmental Sciences Department, Faculty of Science, Alexandria University, Alexandria, Egypt</footnote>

<footnote n="2">²Physical Oceanography Department, National Institute of Oceanography and Fisheries, Alexandria, Egypt</footnote>
CONCENTRATIONS OF PERFLUOROOCTANOATE AND PERFLUOROOCTANE SULFONATE IN SEDIMENT OF WESTERN CAPE RIVERS, SOUTH AFRICA

John Baptist Nzukizi Mudumbi<fnref n="1" />, Seteno Karabo Obed Ntwampe<fnref n="2" />, Munyololo Muganza<fnref n="1" />, Andrew Rand<fnref n="1" />, Okechukwu Jonathan Okonkwo<fnref n="3" />

Corresponding author email: Ntwampes@cput.ac.za

ABSTRACT

In this study, we report for the first time, concentrations of perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) in sediments from a South African environment, particularly for the largest catchment areas of the Western Cape, South Africa. This area is associated to the largest agricultural sector, of which the produce and end-products are intended for the international markets. Therefore, worldwide these compounds have been investigated and documented, but in South Africa in general, and in the Western Cape Province in particular, this still yet to be done. Thus, the aim of this study was to determine the concentration of PFOA and PFOS in riparian sediments of the three rivers of the Western Cape; and the objectives were: a) to analysis and quantification of PFOS and PFOA contamination levels in sediments from the three rivers; b) to determine sediment's physico-chemical characteristics that may facilitate PFOA and PFOS sorption on sediments, c) to correlate PFOA/PFOS distribution in the three rivers to domestic, industrial and agricultural areas coverage of the three rivers. Sample preparation and analysis were based on the ISO 25101: 2009(E) method, using solid phase extraction (SPE) followed by liquid chromatography combined with electrospray tandem mass spectrometry (LC-MS/MS). Sediment granulometry core water characteristics and percent total organic carbon were also quantified.

From the results, PFOS and PFOA were observed in all the river sediment samples and were found in concentrations up to 19 ng/g and 187 ng/g for Salt River, 121 ng/g and 772 ng/g for Diep River, and 75 ng/g and 193 ng/g for Eerste River, for both PFOS and PFOA, respectively. Additionally, it was observed in this study that PFOA sorption onto the sediment was high at a higher pH (> 8), high %TOC in smaller grain size (< 0.1mm), and high salinity, instead of low salinity including pH (< 8) as observed for the Diep River, while the prevalence of PFOS concentration was lower under similar conditions. For the Eerste River, PFOS concentration in the sediment samples was lower at a lower pH (> 7.95), %TOC (> 16% and 20% w/w) for sediment grains < 0.1mm and > 0.5mm, respectively, with low nitrate, phosphates and ammonium concentrations. For the Salt River, although high salinity was measured as 418 mg/L with a high %TOC in grain sediment of size > 0.5 mm, and a high phosphate content, PFOS were below the detection limit in several samples with PFOA and PFOS concentrations being high in other sample as ionic organic chemicals, such as PFOA and PFOS, salinity and pH largely increase their extent of sorption onto solid matrices.

Some of the PFOA and PFOS concentrations are higher than those previously reported in similar studies in various countries; this suggests there is cause for concern. Although sediment has a poor sorption capacity for both PFOS and PFOA in comparison with plants, the prevalence of PFOS and PFOA in sediment samples from rivers from which irrigation water is sourced for agricultural purposes, indicates a risk of agricultural produce contamination.

With such an insight, it is obvious that PFOA and PFOS in South African rivers pose a significant threat to communities using river water in riparian areas.

Acknowledgements: The authors are grateful to the National Research Foundation (NRF) of South Africa for financial support (Thuthuka Grant), Prof. J. Yu (National University of Singapore) for her valuable assistance, and Ms L. Muedi (NRF intern), Ms L. Marshall and Mr D. Roman (Cape Peninsula University of Technology) for their support.

<figure><img src="image_480.png" /></figure>

<footnote n="1">¹ Faculty of Applied Sciences, Department of Environmental and Occupational Studies, Cape Peninsula University of Technology, Cape Town, South Africa</footnote>

<footnote n="2">² Faculty of Applied Sciences, Department of Agriculture and Food Sciences, Cape Peninsula University of Technology, Cape Town, South Africa</footnote>

<footnote n="3">³ Faculty of Science, Department of Environmental, Water and Earth Sciences, Tshwane University of Technology, Pretoria, South Africa</footnote>
# TRANSPRATION TO THE *PTERIS VITTATA* L. ARSENIC ENRICHMENT BY SEVERAL DIFFERENT TEST METHODS

Han zhou Hao<fnref n="1" />,<fnref n="2" />, Zheng yi Fu<fnref n="3" />, Xue bin Zhong<fnref n="1" />,<fnref n="2" />, Hua Wei<fnref n="1" />,<fnref n="2" />

Corresponding author email: haohz110@163.com

## ABSTRACT

Biological factors (leaf area, stomatal function, root quantity and distribution) and abiotic factors (climate, soil moisture, etc.) joint action in plant transpiration. Transpiration rate closely relationship with stomatal opening and closing about, when fully open stomata, transpiration rate can be increased by 20% to 40%.

Generally, the regulation of transpiration based on different methods: (1) by adjusting the relative humidity in artificial climate chamber to regulate different transpiration rates. ZHANG Yong et al found that using this method, high transpiration of tomato seedlings under Cd and Pb content than low transpiration increased by 1.47-1.73 times and 1.25-1.75 times. Sharif Ali Salah showed that with this method, *Triticum aestivum* wheat varieties under high transpiration rate of Cd, Zn accumulation increased significantly lower than the low transpiration rate. (2) use of alternate root osmotic stress, this method reduces transpiration under the premise of maintaining normal growth of plants, half of the roots into the nutrient solution, and the other half roots in nutrient solution + PEG6000 (polyethylene glycol, penetrant manufacturing water Stress) to a cycle of alternating 12h. (3) Some of exogenous hormones. As cytokinins (cytokinin, CK) can induce stomatal opening. (4) shading.

Studies have shown that plant transpiration process of absorption of heavy metals play an important role. Plants absorb pollutants to the performance of the water soluble contaminants can reach the root showed that diffusion through channels and freedom apoplast, after being absorbed by roots, part of the root store down the rest of the transpiration stream through the xylem vessels with rising together from the root transport in the blade.

This experiment by *Pteris vittata* L. as object, studies the high transpiration rate (artificial climate box regulation), low transpiration rate (artificial climate box regulation) and dry land treasure of *Pteris vittata* L. arsenic enrichment regulation function. Research results show that: although HR and LR has influence to the *Pteris vittata* L. biomass, but under the condition of different arsenic treatment is different, Under the condition of without arsenic, high transpiration rate ( HR ) and low transpiration rate (LR ) compared to the total biomass, aboveground biomass, underground biomass increased obviously, under the condition of arsenic, HR and LR treatment compared to the total biomass increases, the difference is not big, especially the underground biomass change is not obvious. The difference of transpiration conditions lead to the *Pteris vittata* L. arsenic concentration difference is obvious, Arsenic and without arsenic, HR are better than arsenic concentration under the condition of LR have obvious growth, under As100 treatment HR is increased by 22.7% than LR conditions, under As200 treatment HR is increased by 27.1% than LR conditions. Transpiration of *Pteris vittata* L. arsenic accumulation contribution rate is extremely low; the contribution rate of different transpiration or different arsenic treatment is small differences. Dry land treasure, can obviously inhibit the *Pteris vittata* L. transpiration, especially the T2 processing, the effect is much better. Don't add dry land treasure but arsenic treatment, the *Pteris vittata* L. also received obviously inhibits transpiration, when arsenic treatment, the transpiration of the *Pteris vittata* L. is restrained. Without arsenic, biological enrichment factor has certain change, Plus arsenic, transfer coefficient and biological enrichment factor change is big, Especially under the treatment of the T2 and arsenic is very significant, transfer coefficient and the concentration coefficient is decreased by 27.8% and 36.4% respectively, it proves that combination of dry land treasure and arsenic has an important effect of *Pteris vittata* L. transshipment and concentration of arsenic.

Acknowledgements: The financial support by Health Department of Hubei Province of P.R. China (No. JX6B29).

<figure><img src="image_481.png" /></figure>

<footnote n="1">¹ Resources and Environment College, Hubei University of Science and Technology, Xianning, Hubei 437100, China</footnote>

<footnote n="2">² Land and Water Resources Research Center of the Middle Reaches of Yangtze River, Xianning, Hubei 437100, China</footnote>

<footnote n="3">³ Dept of Analytical Chemistry, Connecticut Agricultural Experiment Station, New Haven, USA</footnote>