Master’s dissertation submitted in partial fulfilment of the ......Prof. Filip Tack for guiding me during all the stages of the work on this master thesis and for the opportunity

Master’s dissertation submitted in partial fulfilment of the requirements for the joint degree of

International Master of Science in Environmental Technology and Engineering

an Erasmus+: Erasmus Mundus Master Course jointly organized by

Ghent University, Belgium University of Chemistry and Technology, Prague, Czech Republic UNESCO-IHE Institute for Water Education, Delft, the Netherlands

Academic year 2016 – 2017

Heavy metal behaviour in contaminated soil amended with biochar

Host University:

Ghent University, Belgium

Korea Biochar Research Center, Kangwon National University, South Korea

Anna Tsibart Promotor: Professor Filip M.G. Tack Co-promotor: Professor Yong Sik Ok

This thesis was elaborated at Ghent University and defended at Ghent University within the framework of the European Erasmus Mundus Programme “Erasmus Mundus International Master of Science in

Environmental Technology and Engineering " (Course N° 2011-0172)

© [2017] [Ghent], [Anna Tsibart], Ghent University, all rights

reserved.

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Acknowledgement

I am deeply grateful for the help and kind support to the following people:

Prof. Filip Tack for guiding me during all the stages of the work on this master thesis and

for the opportunity to visit Korea

Prof. Yong Sik Ok for accepting me as a visiting student in Korea Biochar Research Center

and sharing his deep knowledge in biochar

Xiao Yang for guiding me in the laboratory work in Korea and sharing knowledge in biochar

characterization

Reinhart van Poucke for the help in the laboratory work in Belgium and feedback for the

thesis

Ali El-Naggar for his kind help and valuable advice during my stay in Korea

Kumuduni Panlansooriya for her support and kindness in Korea

Yasser Awad for the assistance during laboratory experiments in Korea

Melgü Kizilmese for her help in heavy metal analysis in Ghent laboratory

3

Abstract

Soil contamination with heavy metals is increasing over the last decades worldwide, so

effective remediation approaches are required. Biochar is widely studied as a soil amendment

capable of immobilizing heavy metals in soils. The aim of this study is to assess the efficiency of

biochar application for the remediation of soils contaminated with the mixture of heavy metals. For

this purpose, a 21-day incubation experiment was conducted with two soils with different

contamination levels and having various soil properties. The tested soils were collected from two

sites. The soil from Lommel (Belgium) had high concentrations of Cd, Pb, Zn because of Zn smelter.

Another soil was taken from an agricultural site nearby mining activities (Gongju, South Korea).

Seven biochars were screened to evaluate their efficacy of metal immobilization. Metal behavior was

assessed by the extraction of 0.01 M CaCl2 and 0.05 EDTA. The CaCl2-extractable Cd and Zn forms

were reduced after biochar application, however showing no significant decrease in Pb and As

concentrations. The most effective biochar reducing Cd and Zn mobility was FW+wood. This

amendment also reduced the amount of released metals under the conditions of pH change.

Electrostatic attraction and complexation with O-containing functional groups were suggested as

the most important mechanisms affecting Cd and Zn sorption. The important properties of biochars

affecting their immobilization properties were high pH, high O-content, high surface area and the

presence of C-O-C functional groups. The concentration of EDTA-extractable metals did not change

significantly after biochar application. It indicates the environmental risk related to heavy metals at

studied sites, especially in case of sandy soil. Moreover, studied amendments induced pH, EC and

organic matter increase at both studies sites.

4

Contents 1. Introduction .................................................................................................................................................... 5

2. Literature review ............................................................................................................................................ 6

2.1. Heavy metals sources and behaviour in soil ................................................................................................ 6

2.2. Remediation approaches for soils contaminated with heavy metals .......................................................... 8

2.3. Biochar as a soil amendment ....................................................................................................................... 9

2.3.1. Biochar properties ................................................................................................................................ 9

2.3.2. Environmental benefits of biochar application to the soils................................................................ 10

2.3.3. Biochar effect on soil properties ........................................................................................................ 11

2.3.4. Biochar application for heavy metal immobilization .......................................................................... 12

3. Materials and methods ................................................................................................................................ 14

3.1. Soil collection and characterization ........................................................................................................... 14

3.2. Preliminary incubation experiment ........................................................................................................... 14

3.3. Main Incubation experiment ..................................................................................................................... 15

3.4. Biochar production .................................................................................................................................... 15

3.5. Biochar characterization ............................................................................................................................ 16

3.6. Soil characterization .................................................................................................................................. 17

3.7. Data and statistical analysis ....................................................................................................................... 18

4. Results .......................................................................................................................................................... 19

4.1. Preliminary 1-day incubation experiment ................................................................................................. 19

4.2. Main 21-day incubation experiment ......................................................................................................... 20

4.2.1. Soil characteristics .............................................................................................................................. 20

4.2.2. Biochar characteristics ....................................................................................................................... 20

4.3. Changes in soil properties .......................................................................................................................... 25

4.4. Metal immobilization performance ........................................................................................................... 26

4.4.1. Consecutive extraction with 0.01 M CaCl2 ......................................................................................... 26

4.4.2. Consecutive extraction with 0.05 M EDTA ......................................................................................... 30

4.4.3. pH stat test results ............................................................................................................................. 35

5. Discussion .......................................................................................................................................................... 39

5.1. Effect of biochar on soil physico-chemical properties ............................................................................... 39

5.2. Effect of biochar on the CaCl2-extractable forms of heavy metals ............................................................ 40

5.3. Effect of biochar on EDTA- extractable forms of heavy metals ................................................................. 43

5.4. pH-dependent release of heavy metals from biochar amended soils ....................................................... 44

6. Conclusion and recommendation ..................................................................................................................... 46

6.1. Conclusion.................................................................................................................................................. 46

6.2. Recommendation ...................................................................................................................................... 46

References ............................................................................................................................................................ 48

5

1. Introduction

Soil contamination with heavy metals has attracted a lot of attention over the last years, as

these elements are toxic and mobile in the environment. Due to their potential for entering to the

food chain, they pose a significant risk for human health. Heavy metals are persistent in the

environment and cannot be biologically degraded. For these reasons, various methods for soil clean-

up were developed recently. Many of the developed methods are efficient, but at the same time,

they have certain limitations because they are expensive or time-consuming. Nowadays, there is

much interest for chemical soil amendments, that are able to immobilize heavy metals. These

amendments include various clay minerals, bentonite, lime, limestone, zeolites, phosphates, organic

composts and others (Khalid et al., 2017; Xu et al., 2017).

A number of studies showed the efficiency of biochar to reduce the mobility of inorganic

contaminants because of its large surface area, high porosity and presence of surface functional

groups. Some studies confirmed the reduction of heavy metal bioavailability for the plants (Ahmad

et al., 2014; Xu et al., 2012, Al-Wabel et al., 2015).

However, much research focuses on soils freshly spiked with heavy metals, but for soils taken

from polluted sites, these require more detailed investigation. Moreover, the mechanisms reducing

metal mobility remain unclear for each heavy metal, especially in cases where different metals are

involved. The effect of biochar properties on heavy metals stabilization needs to be better

understood. The objective of this study is to assess the efficiency of biochar application for the

remediation of soils contaminated with a mixture of heavy metals.

The objectives of this study are:

1. To screen various biochars for their capability to immobilize heavy metals in

contaminated soils;

2. To reveal the link between the properties of biochar and metal immobilization

performance;

3. To assess the mechanisms of biochar effects on the mobility and bioavailability of

different metals in the amended soils;

4. To compare biochar effect on soils having different properties and contamination level;

5. To study the change of chemical soil properties after biochar application.

6

2. Literature review

2.1. Heavy metals sources and behaviour in soil

Heavy metals and metalloids are elements, which are toxic for humans, animals, plants and

affect environmental quality. Nowadays, the contamination with heavy metals is increasing because

of their usage in various technologies (Khalid et al., 2017).

Pb, Cd, Hg and As are often considered as priority pollutants as they are very toxic, are

released in a large concentrations and are persistent in the environment (Wang, Chen, 2014). They

are part of the list of top 20 Hazardous Substances of the Agency for Toxic Substances and Disease

Registry (ATSDR, 2012) and the United States Environmental Protection Agency (US EPA) (Khalid et

al., 2017). Zn and Cu are essential elements for organisms. For instance, immune system functioning,

cognitive abilities are dependent on zinc in living organisms. However, these elements may be toxic

in high concentrations (Wani et al., 2017). Most heavy metals exist in the cationic form, but some of

them (Cr(VI), Mo(VI), Au(III), Se(V), V(V), As(V) can be found in the anionic form, which influences

their properties and behaviour in the environment (Wang, Chen, 2014).

Sources of heavy metals in soils are both natural (pedogenic, geogenic) and related to human

activity (anthropogenic) (Khalid et al., 2017, Bolan et al., 2014). On a large scale heavy metal

concentrations are mostly dependant on geogenic sources (Toth et al., 2016). Soils with elevated

concentrations of heavy metal could receive them from the weathering of parent materials,

sedimentary rocks and coals, volcanic eruption and erosion (Toth et al., 2016; Khalid et al., 2017,

Thangarajan et al., 2017). For instance, coal is considered to be major source for As (Ferguson, Gavis,

1972). Hg is emitted to the atmosphere, and further accumulated in the soils from volcanic activity

(Varecamp, Buseck, 1986). Generally, heavy metal arrived from parent material are not bioavailable

for plants in most cases (Bolan et al., 2014).

Anthropogenic sources of heavy metals include application contaminated sewage sludge and

fertilizers, pesticides to the soils and irrigation with wastewaters, disposal of waste (Pourrut et al.,

2011, Khalid et al., 2017; Bolan et al., 2014). For example, common source of Cd is application of

Cd-containing P-fertilizers where Cd arrive from phosphate rocks (Bolan et al., 2014). Both organic

fertilizers (manure, sewage sludge) and mineral fertilizers can serve as a source of Zn (Jensen et al.,

2017). Other sources are mining and refining of ores (Ouyang et al., 2017). Some elements are

released in pyro- metallurgical process after mining. Cd, for instance could be emitted as by-product

of Zn-refining process (Pourrut et al., 2011). Metals also arrive from the exhausts of vehicles,

combustion of gasoline containing lead, smelting (Khalid et al., 2017, Yan et al., 2017). Heavy metals

from anthropogenic sources are mostly highly available by plants (Bolan et al., 2014).

7

According to Ross et al. (1994) different sources of metals correspond to different set of

contaminants in soils. For instance, agricultural soils often have elevated concentrations of Zn, As,

Pb, Cd, Cu, Se and U. The group of Cd, Pb, As and Hg arrive from metalliferous mining and smelting.

Industry is the source of Cd, Hg, As, Cr, Cu, Co, Ni and Zn in soils. Waste disposal causes high

contamination with As, Pb, Cu, Cd, Cr, Zn and Hg. The group of As, Pb, Cr, Hg, Cu, Cd and U arrive

from atmospheric deposition.

Soils are the most important sink for the metals in the environment. It was estimated that >

10 million contaminated sites existed in the world, and more than a half of them were polluted with

heavy metals (Khalid, 2017). Soil contamination with heavy metals pose environmental risks. It can

affect the human health via food chains (Dai et al., 2015, Yuan et al., 2017). Moreover, there is a

possibility of accumulation in ground water, water bodies and sediments (Antoniadis et al., 2017).

The accumulation rate of heavy metal in the food chain is determined by properties of the

soil, amount of heavy metal and the uptake rate by plants (Bolan et al., 2014; Yuan et al., 2017).

After arrival to the soil, heavy metals and metalloids are subjected to various processes. These

processes include sorption, precipitation, complexation reaction, plant uptake, leaching and

volatilization.

The sorption of heavy metals in soils depends on the soil properties, especially pH, clay and

organic carbon content. During this process, charged metal ions are attracted by soil particles by

electrostatic attraction and chemical bond formation (Xiao et al., 2015, Bolan et al., 2014).

Precipitation process is important at higher pH values, high amount of metal ions and high

concentration of SO42−, CO3

2−, OH−, and HPO42− groups. Cation complexation with inorganic and

organic ligand ions is another important mechanism of metal retention in soil. These interactions

are controlled by pH and the amount of ligands available. The processes of microbial

oxidation/reduction is significant for some metalloids (As, Cr, Hg, and Se). For instance, microbial

oxidation decreases As mobility. As(III) presents in soil in reduced environment, and less mobile form

As(V) - in oxidized (Bolan et al., 2014). Volatilization takes place only if the element is able to form

gaseous compounds, for instance As, Hg, and Se (Bolan et al., 2014).

Soil chemical properties (pH, redox conditions, cation-exchange capacity, soil mineralogy,

biological and microbial conditions, cation levels) could have an effect on metal availability for plants

in soils and define related environmental risks (Pourrut et al., 2011; Bolan et al., 2014). For instance,

cationic heavy metals are more mobile under acidic conditions as their sorption increases under

higher pH, and anionic elements – under alkaline conditions (Antoniadis et al., 2017). Also, typically,

soils rich in clay are able to retain higher amount of metals. So, metals are more mobile in light-

textured soil. The amount of adsorbed metal depends on CEC of clay minerals. Soils with high CEC

retain both cations and anions (Antoniadis et al., 2017). Soil organic matter favours metal retention

8

as it adds the CEC to soil. Especially, carboxylic and hydroxyl functional groups play an important

role in metal sorption (Antoniadis et al., 2017). Some metals are adsorbed to soil particles; for

instance, Pb mobility in the environment and in soils depends on the solubility of solid particles

bearing Pb (Yan et al., 2017).

The mobility of elements in soils depends also on the nature and quantity of these elements.

Metals which can form strong covalent bonds are more strongly adsorbed to soil

(Hg>Pb>Cd>Co>Ni>Zn>Cu>Cr) (Antoniadis et al., 2017).

2.2. Remediation approaches for soils contaminated with heavy metals

The group of heavy metals and metalloids cannot be degraded biologically or chemically

(Khalid et al., 2017, Bolan et al., 2014; Ouyang et al., 2017). In addition even though law restrictions

favoured the reduction of emissions of metals in developed countries, there are still many historically

contaminated sites. Many developing countries still currently have high emission levels of heavy

metals (Kostarelos et al., 2015), Therefore, there is a need for remediation of contaminated soils

(Bolan et al., 2014; Yuan et al., 2017).

The exciting approaches to soil clean-up include physical, chemical and biological methods,

which could be implemented separately or in the combination (Khalid at al., 2017).

Physical methods are divided into soil replacement, soil isolation, vitrification, electrokinetic

method. Biological methods include phytostabilization, phytoevaporation and phytoextraction.

Immobilization and soil washing are classified as chemical methods (Khalid et al., 2017; Xu et al.,

2017). All these strategies could be divided to one of two approaches: 1) heavy metal removal; 2)

mobility and bioavailability decrease (Floris et al., 2017). For instance, soil extraction and

bioremediation refer to the first approach and metal immobilization with different amendments

represents the second approach.

The remediation technologies should depend on the specific site to make the remediation

more feasible and efficient. However, in many cases the soil remediation task could be very

challenging as most of the techniques are very costly, time-consuming, have some limitations and

could destroy the environment (Khalid at al., 2017).

For instance, physical methods like soil replacement or soil washing, could be used only within

small areas (Yao et al., 2012, Xu et al., 2017). These methods are very expensive, and suited for high

level of contamination. Also the thermal decomposition method, which uses steam or microwave to

convert a contaminant to volatile form, takes a relatively long time (Yao et al., 2012).

Bioremediation is the use of plants and associated microoganisms to reduce the concentration

of heavy metal (Yao et al., 2012). Generally, bioremediation is considered to be a cost-effective and

environmentally friendly approach (Yao et al., 2012) but the method is not applicable for highly

9

contaminated sites (Khalid et al., 2017). Phytoextraction technique based on the plant uptake of the

pollutants and further harvest of aboveground parts is economically feasible and applicable over a

large scale. However, the main disadvantage is a long period of a remediation and dependence on

growing conditions of specific plants (Khalid et al., 2017, Yao et al., 2012).

Chemical methods of heavy metal remediation include application of clay minerals, natural

sepiolite, palygorskite, bentonite, lime and limestone, zeolites, phosphates, and organic composts

(Khalid et al., 2017; Xu et al., 2017). Metal immobilization can be performed due to adsorption,

precipitation, and complexation processes. As a results heavy metals are transferred to the solid

phase and their mobility and bioavailability is reduced (Bolan et al., 2014; Xu et al., 2017). Chemical

method are rapid and effective (Khalid et al., 2017), but some of these amendments can have

undesired side-effects, for instance, solubility of some essential trace elements can decrease and

plant growth can slow down (Floris et al., 2017). Therefore, the amendments should be safe for soil

and not have negative effects on soil properties and fertility (Al-Wabel et al., 2015).

2.3. Biochar as a soil amendment

2.3.1. Biochar properties

According to numerous studies, conducted over the last few years, biochar demonstrated

good potential as a soil amendment favoring heavy metal immobilization (Bashir et al., 2017, Houben

et al., 2013, Lu et al., 2014, Yadav et al., 2017, Wang et al., 2017, Melo et al., 2013, Cao et al., 2009,

Li et al., 2009).

Biochar is a product made after thermal decomposition of organic material under limited

supply of oxygen and a temperature below 900°C (Lehmann, Joseph, 2009). Biochar can be obtained

from pyrolysis of plant-derived biomass (wood bark, rice husk, pine wood etc.) or non-plant derived

biomass (dairy and chicken manure) (Godlewska et al., 2017). Traditional source of biochar includes

lump charcoal from primitive and modern kilns. Also it can be produced in gasifiers, be co-product

or by-product in retorts and bio-gas and bio-oil technologies (McLaughlin et al., 2009).

Biochar is produced at various pyrolysis characteristics (heating rate, highest treatment

temperature, pressure, reaction residence time, reaction vessel, pre-treatment, post-treatment).

Temperature and feedstock are considered the most important factors affecting biochar properties

(Downie et al., 2009).

Biochar surface area have numerous micropores with < 2 nm diameter, which gives adsorptive

properties to biochar (Downie et al, 2009; Mendonça et al., 2017). The surface area and pore size

grow with the temperature because of functional groups destruction (Angin, Sensöz, 2014). However,

at certain a temperature point, some deformation occurs and surface area starts reducing (Downie

et al, 2009). Uchimiya et al., 2011 detected these changes at temperature above 700 °C. It was

http://www.sciencedirect.com/science/article/pii/S0045653513015051#b0495

10

confirmed by the findings of Chun et al. (2004), at 700 °C surface area decreased. Pore size can

influence metal sorption as metals cannot be adsorbed by very small pores (Ahmedna et al., 2004).

Ahmad et al., (2014) suggested that plant-derived biomass generally has higher surface area than

biochars from manure or biosolids.

Elemental composition of biochar generally depends on pyrolysis temperature. Generally,

biochar has a high content of C with high amount of aromatic structures (Lehmann, Joseph, 2009).

At higher temperatures C content normally increases and structure becomes more condensed

(Angin, Sensöz, 2014). Moreover, process of C graphitization, dehydrotation, and deoxygenation of

biomass take place (Ahmad et al., 2014; Mendonça et al., 2017). For this reason with H/C and O/C

ratios decrease under the high temperature. The amount of surface functional groups (carboxylic,

amino, and hydroxyl) generally decreases (Li et al., 2017). Oxygen content decreases with the

temperature because of the decomposition of oxygen surface groups (Mendonça et al., 2017). The

amount of volatile matter content also decreases with high temperature (Hagner et al., 2016).

Biochar pH depends on temperature and feedstock properties (Li et al., 2017). Generally pH

of biochar has alkaline values and rises with the pyrolysis temperature (Angin, Sensöz, 2014) because

of the formation of ash. Moreover, the amount of base cations is higher at greater temperature,

favoring pH increase (Yuan et al., 2011).

High temperatures favor depolymerization of biomass (Keiluweit et al., 2010), but this process

is nor observed in the biochars from non-plant feedstocks because they do not have lignocellulosic

molecules (Ahmad et al., 2014).

After being added to the soil, biochar interacts and aggregates with mineral and organic

matter. Possible degradation may occur due to biotic degradation of a labile biochar fraction,

erosion, leaching, pedoturbation (Lehmann et al., 2009). However, biochar is very stable in the

environment because of its organo-chemical and physical structure. For example, biochar from forest

fires could be more than 10000 years old (Lehmann et al., 2009). Kuzyakov et al. (2009) suggested

that biochar residence time in soils of temperate climates is about 2000 years.

2.3.2. Environmental benefits of biochar application to the soils

Biochar application can contribute in solving various environmental problems – greenhouse

gases emissions, high CO2 concentration in the atmosphere, managing organic waste (Godlewska

et al., 2017, Ahmad et al., 2014). Biochar is also applied for soil improvement and energy production

(Lehmann, Joseph, 2009, Ahmad et al., 2014, Sohi et al., 2012). Biochar is considered as

environmentally friendly ameliorant as local and renewable resources are used for its production

(Lehmann, Joseph, 2009).

http://www.sciencedirect.com/science/article/pii/S0045653517304356#bib14

http://www.sciencedirect.com/science/article/pii/S0167198716301040#!

11

Various waste biomass including crop residue, manure and sludge are used for biochar

production (Lu et al., 2017, Xu et al., 2013, Cao et al., 2009, Melo et al., 2013). The usage of biochar

favors managing this waste, and reduces pollution loading to the environment. During the process

of charring the volume and weight of the waste is reduced. another benefit is decreasing methane

emissions from landfill (Lehmann, Joseph, 2009, Ahmad et al., 2014). Moreover, pyrolysis processes

reduces pathogenic microorganisms from sludge or manure biochar (Lehmann, Joseph, 2009, Ahmad

et al., 2014). It also has the benefit of decreasing energy used in the long-distance transport of

waste (Lehmann, Joseph, 2009, Ahmad et al., 2014).

Biochar application to agricultural lands is able to sequester atmospheric carbon dioxide and

mitigate climate change (Lehmann, Joseph, 2009, Godlewska et al., 2017). Biochar is very stable in

soils due to its physical structure (Kuzyakov, 2009, Lehmann, Joseph, 2009). This long-term stability

plays an important role in reducing CO2 emissions as it decreases the rate at which

photosynthetically fixed carbon is transmitted to the atmosphere (Woolf et al., 2010). Biochar was

suggested as a sink for atmospheric CO2 by Glaser et al., (2002). Zweiten et al., (2009) also reported

reduction of CH4 emissions by biochar application (Zweiten et al., 2009). Globally, biochar

implementation can reduce total greenhouse emissions by 12% annually (Woolf et al., 2010).

2.3.3. Biochar effect on soil properties

Biochar can positively affect soils physical and chemical properties (ion exchange capacity,

porosity, water holding capacity, retention of nutrients or microbial activity) (Godlewska et al., 2017;

Hussian et al., 2016, Glaser et al., 2002).

According to Glaser et al. (2002) and Verheijen et al., (2010), biochar addition to soil increases

its cation exchange capacity and improves higher nutrient retention in comparison to untreated soil.

Moreover, biochar has a low bulk density, so its additions can reduce bulk density of soil and

improve soil structure. Also, biogeochemical processes in soils are altered after biochar addition due

to changes in microbial communities and changes in enzyme activities (Ahmad et al., 2014). All

these processes improve soil fertility and provide better crop productivity after biochar addition

Glaser et al., (2002). The study of Al-Wabel et al. (2015) showed that biochar application improved

maize growth (which plant?).

Water retention of soil increases after biochar application (Downie et al., 2009) because net

soil surface area increases (Verheijen et al., 2010). Glaser et al., (2002) suggested the increase of

water holding capacity after biochar additions was because of high organic matter content in biochar

(Glaser et al., 2002). This effect is long-term because of biochar stability and recalcitrance (Verheijen

et al., 2010). Due to additional water and nutrients in the micropores, biochar may improve plant

12

water availability, especially for sandy soils during dry periods. However, in case of small pores,

biochar particles can block soil pores, reducing the infiltration ability of soil (Verheijen et al., 2010).

Soils organic matter is significantly altered by the addition of biochar (Ahmad et al., 2014).

For instance, it can cause the positive priming effect (Zimmerman et al., 2011) by faster

decomposition of soil native C by changing microbiological conditions (Kuzyakov at al. (2009). In

other cases, the negative priming effect was observed due to the adsorption of dissolved organic C

and its slower decomposition. Generally, the amount of mobile and resident organic matter, its

sorption capacity affects the positive or negative direction of priming (Ahmad et al., 2014). In

addition, positive priming effect is more pronounced in the case of application of biochars produced

at lower temperature. Addition of biochars produced at higher temperature causes mostly negative

priming effect (Zimmerman et al., 2011). However, over the long term organic matter is strongly

adsorbed to biochar causing C storage in soils (Zimmerman et al., 2011).

Because of its alkaline nature biochar has a liming effect on soil. The extent of this effect

varies depending on biochar properties, its feedstock and production temperature. (Ahmad et al.,

2014).

2.3.4. Biochar application for heavy metal immobilization

Biochar is considered as a soil ameliorant having a great potential for immobilizing heavy

metals (Ahmad et al., 2014; Xu et al., 2012, Al-Wabel et al., 2015). However, the ability of biochar to

adsorb contaminants varies depending on biochar physico-chemical properties and target pollutant

(Ahmad et al., 2014). The most important properties are feedstock and pyrolysis temperature.

The main mechanisms of reducing metal mobility include processes of complexation with

functional groups, cation exchange with biochar surface, precipitation and formation of insoluble

species, electronic attraction to biochar surface, reduction and further sorption of reduced

compounds (Li et al., 2017; Ahmad et al., 2014). Also often, these mechanisms can act at the same

time.

Cation exchange capacity can predominate in cases of biochar having relatively high CEC and

high amount of Ca, K, Mg, Na (Harvey et al., 2011, Li et al., 2017). In the case of biochars with low

CEC other mechanisms play an important role in Cd sorption. They include complexation with

carboxylic surface functional group and precipitation. Uchimiya et al., (2011) observed the important

role of carboxyl, hydroxyl, and phenolic functional groups for metal binding. Biochars derived from

various feedstocks can have different mechanism of metal sorption. For instance, manure biochar

can contain high amounts of carbonate and phosphate. For this reason, precipitation is the main

mechanism of Cd immobilization by manure biochar (Xu et al., 2013).

13

Cadmium and lead are divalent cations and their sorption behaviour is similar. Pb sorption is

defined by the same mechanisms as sorption of Cd and depends on feedstock and pyrolysis

temperature of biochar (Li et al., 2017). Non-electrostatic mechanisms are considered as dominating

for Pb (Li et al., 2016; Clemente et al., 2017; Tang et al., 2013; Uchimiya et al., 2011). But Cao et al.

(2011) reported immobilization of Pb by forming Pb5(PO4)3(OH) in soils amended with dairy manure

biochar. Uchimiya et al. (2012) showed higher Pb immobilization performance in case of low pyrolysis

temperature biochars. These biochars contained O-containing functional groups playing an

important role in altering of Pb mobility.

Zn immobilisation by biochars is controlled by complexation, electrostatic attraction, and

precipitation. Moreover, Ca-oxalate (CaC2O4) crystals may be responsible for the increased ability

of some biochars to remove Cd and Zn (Clemente et al., 2017). Lu et al. (2014) reported that

reduction in biochar size can enhance the effectiveness of Zn immobilization illustrating biochar size

also should be taken into consideration when in use as soil amendment.

Arsenic behaviour is a bit different in comparison to Pb and Cd. As sorption is governed by

complexation with hydroxyl, carboxyl, and C-O ester of alcohols functional groups and electrostatic

interactions. Biochar produced at lower temperature is potentially more efficient in As removal (Li

et al., 2017). However, in some cases biochar application does not induce As immobilization. As

(AsO43-, AsO3

3-) is attracted by positively charged sites in soil, but after the biochar application pH

increases and amount of these sites becomes lower. Another possible mechanism increasing its

mobility in the amended soils is electrostatic repulsion with biochar surface (Igalavithana et al., 2017).

Moreover, cation exchange capacity in soil increases after adding biochar amendment and

soil pH range shifts towards neutral and alkaline range. Under these conditions metal mobility

decreases and the mobilization of oxyanions increases (Al-Wabel et al., 2015.; Lu et al., 2014).

Ahmad et al. (2014) suggested that low temperature pyrolysed biochar with the high amount

of O-containing functional groups generally show good efficiency for heavy metal stabilization. High

surface area biochars with a high amount of pores produced under high temperature are more

effective for organic contaminants (Ahmad et al., 2014). However, in the studied soils both factors

were important for Cd and Zn stabilization.

So, biochar has received a lot of attention in the last years. However, there are still some

uncertainties in this field. Many studies focused on soils, which were spiked with heavy metals under

laboratory conditions. There is still lack of research on aged field contaminated soils (Lu et al., 2014).

Moreover, biochars vary greatly in their properties and ability to adsorb the contaminants. More

field studies and in-situ experiments are required for interpreting the mechanisms of various biochars

and long-term effect on the contaminated soils.

14

3.Materials and methods

3.1. Soil collection and characterization For this study soils were collected from two sites with different contamination level. One soil

was sampled in Lommel (Limburg, North-East Belgium, 51°13′ N, 5°15′ E) in a site nearby two zinc

smelters. The zinc smelter of Lommel was operated in the period of 1904-1974. The zinc smelter of

Balen was built in 1889 and continues its operation until now. So the site was historically polluted

with Cd, Zn and Pb because of atmospheric deposition and zinc ashes application in the road

construction (Van Nevel et al., 2011). Cadmium was emitted into the ecosystem as a as by-product

of the Zn refining process. Nowadays, an area of approximately 700 km2 is diffusively polluted by

Cd, Zn and Pb (Meers et al., 2010). Lower concentrations of Cu, As, and Hg were also found at this

site but they did not exceed the regional safety levels (Van Nevel et al., 2011). According to the

FAO/UNESCO soil classification the soil could be classified as a Humic Podzol, which is characterized

by sandy texture and high acidity. These factors favor high mobility of heavy metals, so there is a

risk of human exposure in the area.

Another soil was sampled near the Tancheon historical mine at Gongju-si (36.44° N, 127.12°E,

Chungcheongnam-do Province, Korea). Vegetable crops were cultivated at the site, but because of

the high contamination with As and Pb, during last few years agriculture was prohibited

(Igalavithana et al. 2017). The soils could be classified as Inceptisols. These soils had slightly acidic

pH, which can cause mobilization of some metals and a high risk of human exposure.

The air-dried soils were passed through a 2 mm sieve and homogenized. pH, and EC were

determined in the mixture of soil and water with the weight ratio 1:5 soil: deionized water (Thomas,

1996; Rhoades, 1996). The measurement was conducted using a pH electrode (Orion Virsa Star,

Thermo Scientific, USA). For the EC determination, the suspension was filtered using highly retentive

filter paper (Whatman 42, diameter 110 mm, 100 circles). The measurement was performed by an

EC electrode (Orion Virsa Star, Thermo Scientific, USA). Total metal concentrations were analyzed in

a microwave-digested sample (USEPA method 3051, MARS, HP-500 plus, CEM Corp., NC, USA) by

ICP-OES (Ahmad et al., 2016; Smith and Mullins, 1991; USEPA, 2007). Soil texture was analyzed by

pipette method (Gee et al., 1986). Total N was defined by mineralization and followed by steam

distillation followed by steam distillation and quantitative analysis via titration (Van Ranst et al.,

1999).

3.2. Preliminary incubation experiment

In order to select the most effective biochars for metal immobilization a preliminary

experiment with various biochars was conducted. These biochars had different feedstocks, but all of

them could be potentially found in Belgium and in Korea.

15

Twenty grams of Lommel soil was placed in each incubation bottle for 24 hours into the

incubator MIR-554, SANYO (Japan). The biochar samples (0.6 g = 3% application rate, 45 t/ha) and

the water (1.5 mL = 70% of water holding capacity) were added to each bottle with the Belgian soil.

48 experimental units (15 biochar types + 1 control, in three replicates) were prepared in total. After

the incubation the samples were dried at 30℃ for 2 hours and the pH and EC were measured as

described above.

Based on the results of the preliminary experiment the 7 most effective biochars were selected

for the incubation experiment. These were: 40% food waste+60% wood (NUS); Wood 100% (NUS);

Vermont Biochar; 30% Chicken Manure+70% Wood (NUS); AD residue+wood (NUS); Sorghum 600℃;

Diary Manure.

3.3. Main Incubation experiment

Assuming a soil depth of 10 cm and a bulk density of 1500 kg/m3 the biochars were applied

to the 95 g of soil at a rate of 5%, which was equal to the application rate 75 t biochar/ha. The

experiment was conducted in closed 600 mL high-density polyethylene bottles. The water content

in the samples was maintained at 70% of the water holding capacity. Control soils were incubated

without amendments. All treated and control soils were triplicated. Biochar amended soils and

control samples were incubated at 25 °C for 21 days under dark conditions in the incubator (MIR-

554, SANYO, Japan).

3.4. Biochar production

Most of selected biochars are by-product of the gasification technology. Four biochars were

obtained as a by-product of a 10 kWt fixed bed downdraft gasifier (All Power Labs, Berkeley, US,

http://www.allpowerlabs.com/).

The biochar food waste (40%) and wood waste (60%) was obtained from the National

University of Singapore (NUS). The feedstock for gasification included the mesquite wood chips

purchased from KingsFord (USA) and a dried food waste (rice, noodles, pasta, mat, eggs, vegetable

matter). The feedstock was gasified at a temperature 25 – 800 ℃ and a heating rate of 20 ℃/min.

(Yang et al., 2016). The food waste could be gasified only in a mixture as it has a high moisture

content which affects the quality of syngas produced. However, the increasing of the food waste

proportion favoured the high heating value of a syngas (Yang et al., 2016). The biochar chicken

manure (30%)+wood waste (70%) was also obtained from NUS. Chicken manure was used for

obtaining CaO catalysts for the production of biodiesel. The dried chicken manure was calcined

under air for 4 hours at temperature 550-950 °C (Maneerung et al., 2016). Another feedstock from

this gasifier included only mesquite wood chips (100 % wood waste) (Yang et al., 2016) and the

mixture of anaerobic digestion residue and wood waste.

http://www.allpowerlabs.com/

16

The other two biochars (dairy manure and sorghum) were obtained in a pilot scale bubbling

fluidized bed biomass gasifier developed by the Texas A&M University (TAMU) at College Station

(Maglinao et al., 2015). Biochar preparation conditions are shown in Table 1.

Table 1. Preparation conditions of studied biochars

Feedstock Wood

100%

Food

waste

40%+

wood

waste 60%

Chicken

manure

30%+ wood

waste 70%

AD

residue+

wood

waste

Vermont

biochar

(pulp

wood,

hardwood

tops)

Dairy

manure

Sorghum

Process Gasification Gasification Gasification Gasification Pyrolysis Gasification Pyrolysis

for energy

production

Reactor APL Gasification Retort kiln Fluidized bed biomass

gasifier

Dairy manure was collected in an open pit at the Southwest Regional Dairy Center (University

in Stephenville, Texas). After this it was air-dried under ambient conditions and further dried at 85°C.

Finally the moisture content reached 1%. Particle size of the gasification residue was between 16

and 40 meshes (Nam et al., 2016).

High tonnage sorghum was planted at the Texas Agrilife Research farm in Burleson county,

Texas. This feedstock had a low lignin content. Prior to the gasification procedure sorghum was

grained to 10 mm size particles and dried to 10% moisture content (Maglinao et al., 2015).

Vermont biochar is commercially available on the market. This biochar was obtained from

pyrolysis process (organic material decomposition by heating with no oxygen supply). Biochar was

produced in a retort kiln. The feedstock was presented by the “waste wood” (pulp wood, sawmills

slabs, hardwood tops). Also microorganisms (bacteria, fungi) were added to the biochar

(http://www.vermontbiochar.com/).

3.5. Biochar characterization For the determination of moisture, ash, mobile and resident matter content in the biochar

samples the proximate analysis was performed (McLaughlin et al., 2009). Moisture was analyzed in

the samples as a difference of weight of air-dried sample and a sample heated at 105 ℃ for 24

hours to a constant weight. Mobile matter (volatile or labile matter), representing the non-carbonized

portion of biochar, was determined as a weight difference between the initial sample and sample

heated in a covered crucible in a muffle (WiseTherm, Germany) at 450 ℃ for 1 h. Ash was measured

17

after heating the sample at 750 ℃ for 1 hour in an open-top crucible. Resident matter (fixed carbon)

was found as a difference between the initial sample and moisture, ash and mobile matter (Ahmad

et al., 2012). Biochars were analyzed for elemental composition (C, H, O, N, S) by elemental analyzer

EuroEA 3000 (EuroVector, Pavia, Italy), and after this the ratio O/C were calculated.

For the pH determination in biochar the extraction was performed at a 1 g : 20 mL soil-to-

deionized water ratio (Thomas et al., 1996). The extraction of the mixture was followed by shaking

for 1.5 h. After this the mixture was left for 10 min, than stirred again, and the electrode was inserted

into the suspension. After pH measurement the suspension was filtered using highly retentive filter

paper, and electrical conductivity was determined in the filtered supernatant. Orion Versa Star Pro

pH/Conductivity Multiparameter Benchtop Meter (Thermo Scientific, Waltham, MA, USA) was used

for pH and EC determination. Each solution was measured in triplicate (Thomas et al., 1996).

For CEC analysis percolation tubes were filled with 0.5 cm of white sand. After this a mixture

of 1g of biochar and 12.5 g of sand was added to the tubes. The last layer included 0.5 cm of pure

white sand. The tubes were percolated with 75 mL of ammonium acetate, 150 mL of denaturated

ethanol, 250 mL of 1 M KCl. Fifty mL of the last solution was placed in a distillation flask and then

a spoon of MnO was added. In the distillation process NH4+ was recovered in the boric acid flask.

The distillate was titrated with Titrino (Metrohm, Switzerland) with 0.01 M HCl until the solution

changed back into a red colour. The amount of HCl added was used to determine cation exchange

capacity (Van Ranst et al., 1999).

Analysis of surface area was performed using adsorption apparatus Gemini VII 3.03

(Micromeritics, Norcross, GA, USA). Biochar samples were pretreated at 473 K for 6 hours and were

adsorbed by N2 at 77 K. For the determination of biochar surface area the Brunauer–Emmett–Teller

(BTE) method was applied. Average pore volume and diameter were obtained with Barret–Joyner–

Halender (BJH) method.

Biochar surface functional groups were characterized by Fourier transform infrared

spectroscopy (FT-IR). The potassium bromide pellet method with wavenumber range of 600-4000

cm-1 was used. The resolution 4 cm-1 was used to scan the pellets. Surface functional groups were

determined after the baseline correction using the software correction Thermo Scientific

Omnic.

Biochar samples were analysed by scanning with an electron microscope with energy-

dispersive X-ray spectroscopy (SEM-EDX; Hitachi S-4800 with ISIS 310, Japan) to obtain

morphological images of biochar surface and elemental distribution on the surface.

3.6. Soil characterization

For the pH determination in the soils the extraction ratio 1:5 soil: deionized water with 5 g of

soil was used (Thomas, 1996). The duration of shaking was 1 h. After filtering the suspension EC was

18

determined (Rhoades, 1996). Each solution was measured in triplicate. Total N was defined by

mineralization and quantitative analysis by steam distillation followed by steam distillation (Van

Ranst et al., 1999). Organic C was analyzed by loss of ignition method at 550 °C (Van Ranst et al.,

1999).

Consecutive CaCl2 and EDTA extractions were conducted for assessing mobile and bioavailable

forms of metals. 1g of soil was used for extraction with 10 mL of 0.01 M CaCl2 solution. After 24 h

shaking at 550 rpm the samples were centrifuged and pure extract without solid particles was taken

out with a pipette, filtered and analysed for heavy metals. Extractions were repeated 4 times. The

same procedures were performed with 0.05 M EDTA solution. The concentration of heavy metals in

the obtained extracts were analyzed using ICP-OES (Optima 7300 DV, Perkin-Elmer, USA).

For pHstat leaching tests 10 g of soil from each treatment was mixed with 50 mL of distilled

water with a magnetic stirrer at 550 rpm. The pH-meter was used for monitoring acidity changes.

A 10 mL initial sample was collected from the solution. 0.6 M HCl acid was added to lower the pH.

At each 0.5 pH step 5 mL of sample was taken until the final pH=4.0. The collected soil suspensions

were filtered and measured in the ISP-OES machine for Zn, Pb, Cd and As (Optima 7300 DV, Perkin-

Elmer, USA). Finally, an adsorption capacity of each treatment was estimated at different pH levels.

3.7. Data and statistical analysis A one-way ANOVA followed by a Tukey's test were conducted to assess significant differences

between means in each treatment for heavy metal concentration. All statistical analyses were carried

out using SPSS v.14.

19

4.Results

4.1. Preliminary 1-day incubation experiment

After the preliminary 1-day incubation pH increased in all amended soils in comparison to

control soil (Table 2). pH values exceeded pH=7 after adding 3% of biochar (40% food waste+60%

wood (NUS); Wood 100% (NUS); Vermont Biochar; 30% Chicken Manure+70% Wood (NUS);

AD+wood (NUS); Sorghum 600℃; Diary Manure). In case of other biochars the soils remained below

pH=7.

Table 2. The pH and EC (mean ± standard deviation) in control and biochar amended soil

incubated for 24 hours (n=3).

Moreover, some tested biochars caused an increase in soil EC, others induced a decrease. EC

of the biochar amended soils did not exceed the saline limits of the soils (0.4 S/m) (Scianna, 2002).

Thus, all soil samples after incubation could be classified as nonsaline. For this reason, EC change in

the soil could not give a negative effect on the plant growth after biochar application.

Biochar pH EC, S/m

40% food waste+ wood 7.68±0.07 0.0666±0.0028

Black Carbon (China) 7.13±0.04 0.0593±0.0032

Wood Japan 6.65±0.04 0.0126±0.0051

Wood 100% 7.29±0.07 0.0176±0.0004

Wood charcoal (Korea) 6.59±0.04 0.0115±0.0002

Sewage Sludge 6.39±0.06 0.0074±0.0049

Vermont Biochar 7.35±0.11 0.0192±0.0034

Wakefield agricultural carbon 6.74±0.11 0.0108±0.0088

Chicken Manure+ wood waste 7.35±0.05 0.0763±0.0017

AD char from NUS+wood waste 7.06±0.06 0.0265±0.0022

Sorghum 500 ℃ 6.78±0.04 0.0945±0.0026

Hoffman Horticultural Charcoal 6.78±0.03 0.0121±0.002

Sorghum 600℃ 7.10±0.03 0.0666±0.0033

Diary Manure 7.15±0.06 0.0591±0.0034

Algae 6.93±0.07 0.0125±0.0052

Control 6.35±0.08 0.0176±0.0063

20

In addition, the mobility and behavior of target heavy metals for the Lommel soil (Cd, Zn,

Pb) mostly depends on the pH (Liang et al., 2017). For this reason, biochars with higher ability to

change the soil pH were selected for the main incubation test.

4.2. Main 21-day incubation experiment

4.2.1. Soil characteristics

Tested soils had different texture (Table 3). Gongju soil had a sandy loam texture and Lommel

soil was sandy. Other properties were similar. The pH in both soils was slightly acidic, in the Gongju

soil it was 6.29, and in the Lommel soil – 5.56. Both studied soils had low total nitrogen - 2.0 g/kg

and 1.1 g/kg in the Gongju and Lommel soils respectively.

The level of contamination with heavy metals was higher in Gonju soil, and the main pollutants

were As and Pb. The main contaminants for Lommel site were Cd, Pb, Zn. In both soils the

concentrations of heavy metals exceeded the regional contamination warning limit (De Temmerman

et al., 2003, Ministry of Environment, 2016).

Table 3. The properties of the studied soils

Soil type Soil

texture

Sand,

%

Silt,

%

Clay,

%

pH Total N,

g/kg

EC,

S/m

As,

mg/kg

Pb,

mg/kg

Cd,

mg/kg

Zn,

mg/kg

Gongju soil

(Korea)

Sandy

loam

68.8 21.7 9.4 6.3 2.0 0.0481 448.6

593.9

5.8

178

Contamination warning limit (Korea) 25 200 4 300

Lommel soil

(Belgium)

Sand 92.0 2.4 5.6 5.5 1.1 0.0213 9.6

223.6

10

395.9

Contamination warning limit (Belgium) 1.2 200

4.2.2. Biochar characteristics

The amount of mobile matter varied between the biochars depending on the feedstock and

temperature (Table 4). Woody biochars (wood waste 100% and Vermont) had the highest values of

mobile matter. The mobile matter decreased at lower proportions of wood and it was the lowest in

the case of dairy manure biochar. Resident matter had higher values in biochars produced at the

higher temperatures – wood and AD+wood.

Carbon content was generally higher at the high temperature, due to the removal of functional

groups and greater carbonization. O and H levels were relatively low in all the samples. The degree

of polarity (O/C) was the highest in dairy manure and sorghum biochars.

A higher pH level was also observed in biochars produced at 800°C due to removal of acidic

functional groups. Low EC was detected in woody biochars (wood 100% and Vermont biochars) and

dairy manure biochars, which can potentially influence low ion exchange and high HM mobility.

21

Table 4. Physico-chemical properties of studied biochars

Biochar Mobile

matter, % Ash, %

Resident

matter, %

C, % H, % O,

%

O/C CEC,

cmol(+)/

100g

pH EC, S/m

Diary

manure

11.6±1.0 85.0±0.6 1.7±0.9 9.8 - 5.0 0.5 1.48±0.30 9.72±0.06 0.082 ±0.0018

Vermont

biochar

33.2±5.6 25.1±3.6 32.2±8.2 78.5 1.77 19.7 0.19 2.66±0.02

10.09±0.05 0.1153±

0.0036

Sorghum 25.8±2.1 43.6±1.5 24.9±1.7 41.6 2.7 10.8 0.3 3.02±0.04 9.17±0.06 0.7293±0.022

Chicken

manure30%

+ wood

waste 70%

26.4±2.9 12.7±2.3 43.4±1.5 65.2 1.7 10.0 0.2 4.64±1.15 10.32±0.02 0.6940±0.0208

Food waste

40% +

wood waste

60%

27.3±1.6 29.5±0.8 36.3±1.4 45.4 - 11.0 0.2 4.01±1.00 10.50±0.03 0.6650±0.0022

AD residue

+wood

waste

28.3±4.7 12.7±2.3 54.3±6.2 49.4 1.2 6.7 0.1 4.06±0.17 9.94 ±0.10 0.1671±0.0016

Wood 100% 37.0±3.3 8.8±1.0 47.0±4.2 69.4 - 8.4 0.1 11.17±0.37 9.96±0.01 0.0826±0.0016

All the biochars had relatively low CEC (1.5 - 4.6 cmol(+)/100g). The highest value was observed

in case of wood waste biochar (11.17 cmol(+)/100 g).

Surface area and pore characteristic of the studied biochars are shown in the Table 5.

Table 5. Surface area and pore properties of biochars.

Biochar BET Surface area,

m2/kg

Pore volume,

cm3/kg

Pore size,

nm

Dairy manure 3860 0.07 9.14

Vermont 246500 0.08 5.08

Sorghum 4050 0.01 12.28

30% Chicken manure + wood 347800 0.09 3.11

40% food waste + wood 159250 0.07 2.96

AD + wood 23320 0.02 2.74

Wood 100% 112910 0.03 3.14

22

The surface area increased in the biochars produced at higher temperatures (Chicken

manure+wood, food waste+wood, Vermont, wood biochars). Micropore area was higher in the same

samples but pore size was significantly reduced at higher temperatures.

SEM at 1000× magnification showed that the studied biochars had a porous structure (Fig. 1),

but the size and amount of pores differed. It could indicate the different ability of biochar to adsorb

contaminants.

FTIR analysis revealed the presence of various functional groups (Fig. 2). The peak at the

wavelength 3200-3500 is assigned to the bonded hydroxyl O-H groups (Igalavithana et al., 2017;

Bekiaris et al., 2016). It was observed in Wood, wood+AD, wood+ chicken manure biochars, which

were produced in the gasifier at the temperature range 25-800°C. Probably their heating up to 800

°C was very short, as above 600°C breakdown of O-H groups and water removal takes place.

Carboxylic acids (C=O stretching) groups were detected in Wood, Wood+Chicken manure, dairy

manure, Wood+food waste biochars at 1665-1760 cm-1. Aliphatic C-O-C finctional groups were

found in Vermont, DM, Sorghum, Wood+FW in the wavelength region 1000-1320 cm-1. The peak

at 885-760 cm-1 indicated aromatic C-H bond, this peak was found in Vermont, dairy manure,

sorghum biochars (600°C), and food waste+wood biochars. Other biochars formed at temperatures

up to 800 °C did not show the peak at these wavelength.

XRD analysis indicated the presence of SiO2 mineral in the sorghum dairy manure, wood and

Vermont biochars (Fig. 3). CaSO2 was detected in Sorghum, Wood +FW, Wood+CM, Wood+AD

amendements. The same biochars demonstrated the presence of calcite CaCO3. Peaks corresponding

to KCl (Sylvite) were observed for FW+Wood, CM+Wood, AD+Wood and Vermont biochars.

The carbon structure of biochars was investigated using Raman spectroscopy (Fig. 4). Two

peaks could be distinguished in all the samples at 1354 cm-1 (ID, defect), showing the presence of

defects in carbon structure and at 1595 cm-1 (IG, graphite), showing more organized graphene

structures. The ID peak corresponds to polyaromatic carbons, which do not have special order

(Mendonça et al., 2017). More pronounced and higher IG bands were observed in sorghum,

wood+AD, DM-600 biochars, so these amendments had a more organized structure.

Also, in case of high total aromaticity of the biochar, the light absorptivity of char increases,

and Raman intensity is low (Keown et al., 2007). Thus, Vermont biochar, wood, wood+chicken

manure, food waste+wood biochar had the lowest Raman intensity, indicating a higher degree of

aromaticity in comparison to other biochars. The same biochars had a high BET surface area.



23

Fig. 1. SEM images at 1000 magnification images of studied biochars. a - Dairy manure; b- Vermont biochar;

c- Sorghum; d- 30% Chicken manure+wood; e- 40% food waste+wood; f - AD+wood; g- 100% wood.

b

c

a

a b

d

e f

g

24

Fig 2. Surface functional group analysis of biochars using Fourier transform infrared spectroscopy

(FT-IR).

Fig. 3. XRD of studied biochars.

25

Fig. 4. Raman spectra of the biochar samples.

4.3. Changes in soil properties Soil pH increased by 1-2 units after application of biochars (Table 6). For the Lommel soil the

most effective biochars were Vermont, Food waste+wood, chicken manure+wood and 100% wood.

For the Gongju soil Vermont and food waste biochars caused the most significant pH changes due

to their highly alkaline nature.

The soil EC increased to the highest values of 0.1709 S/m in the Sorghum amended Gongju

soil. Chicken manure and food waste amendments also caused significant increase of EC values in

both soils.

26

Table 6. The pH and EC (mean ± standard deviation) in control and biochar amended soil

incubated for 21-day (n=3).

Sample pH EC, S/m LOI (550°C), % DW

Lommel Dairy manure 7.19±0.07 0.0450±0.0013 0.16±0.03

Lommel Vermont 8.14±0.06 0.04243±0.0031 0.25±0.01

Lommel Sorghum 7.45±0.03 0.1557±0.0064 0.21±0.01

Lommel CM + wood 7.64±0.09 0.1250±0.0015 0.22±0.03

Lommel FW + wood 8.11±0.04 0.1369±0.0086 0.20±0.03

Lommel AD + wood 7.53±0.08 0.0471±0.0079 0.24±0.02

Lommel wood 7.67±0.06 0.0245±0.0033 0.24±0.04

Lommel control 6.29±0.15 0.0213±0.0092 0.12±0.01

Gongju Dairy manure 6.66±0.07 0.0671±0.0062 0.19±0.02

Gongju Vermont 7.17±0.08 0.0583±0.0018 0.27±0.03

Gongju Sorghum 6.84±0.07 0.1709 ±0.0092 0.22±0.02

Gongju CM + wood 6.91±0.08 0.1488±0.0063 0.29±0.06

Gongju FW + wood 7.27±0.02 0.1569±0.0062 0.34±0.02

Gongju AD + wood 6.70±0.09 0.0571±0.0081 0.34±0.05

Gongju wood 6.58±0.08 0.044±0.0032 0.29±0.04

Gongju control 5.50±0.08 0.0481±0.0076 0.20±0.02

The content of soil organic matter (soil on ignition) increased in soils after biochar application.

The control soil sample collected from the Lommel site had 0.12% of organic matter. The ANOVA

was significant F (7, 16)=7.315, p=0.01. Tuckey HSD tests revealed significant pairwise differences

between the means of control soil and soils with amendments - Vermont, sorghum, chicken

manure+wood, AD+wood, Wood biochars, p<0.05. Dairy manure and food waste biochar amended

soils, do not significantly differ from control soil, p>0.05. The highest values of LOI % were observed

in soils treated with Vermont, AD+wood and wood biochar (0.25, 0.23 and 0.24% respectively). These

biochars had the highest carbon content according to proximate analysis.

In the control Gongju soil the content of matter was higher - 0.20%. The statistically significant

increase in total organic carbon was observed in samples treated with FW+wood, AD+wood p<0.05.

The ANOVA was significant F (7, 16)=5.605, p=0.01. Others treated soils, do not significantly differ

from control soil, p>0.05.

4.4. Metal immobilization performance

4.4.1. Consecutive extraction with 0.01 M CaCl2

CaCl2 extractions were performed in order to estimate available heavy metals in soil (Kabata-

Pendias, 2004). Consecutive extraction gave information about the environmental behavior of heavy

27

metals and their release if soils were continuously washed with the solution. The concentration of

Cd and Zn extracted with 0.01 M CaCl2 in the Lommel soil are presented in Fig. 7 and 8 respectively.

Cd was released in higher amounts from the control soil in all the four consecutive extractions

(Fig. 5).

Fig. 5. 0.01 M CaCl2 extractable Cd concentration in Lommel soil after 21-day incubation. The error

bar indicate standard deviation (n=3).

All biochar amended soils had lower total CaCl2-extractable Cd concentrations (n=3, p<0.05).

The ANOVA test for the total extractable Cd concentration gave significant F values, (F (7, 16)=56.296,

p=0.001). Treatment with Vermont, Sorghum, and FW+wood biochars was especially effective. Cd

was not detected at the first extraction in these cases, but further extractions caused Cd

immobilization. The least effective biochar was AD + wood. In this case, 0.35 mg/kg Cd was released

at the first extraction, and next extraction steps caused the release of a higher Cd amount. The total

amount of Cd released from the soil was the lowest in soil treated with food waste+wood biochar.

Concentration of CaCl2-extractale Zn was high in the control soil, at the first extraction the

values reached 59 mg/kg, and further decrease to 26 mg/kg at the forth extraction (Fig. 6). According

to Tuckey HSD tests, meanscores of total CaCl2-extractable Zn concentration in control soil and all

the treatments were significantly different (n=3, p<0.05).

The best results were shown by food waste (40%)+ wood (60%) and sorghum biochars. At

the first Zn concentration was below the detection limit. Further extraction steps caused the release

of Zn. Soils amended with dairy manure, chicken manure+wood, AD residue+wood, and wood 100%

had higher concentration of Zn. The first extraction released 8, 6, 11, 12 mg/kg Zn respectively. The

second extraction released even higher amounts of Zn 10, 9, 14, 17 mg/kg Zn. Dairy manure and

0

0.2

0.4

0.6

0.8

1

1.2

Dairy manure Vermontbiochar

Soghum 600C 30% chickenmanure+wood

40% foodwaste+wood

ADresidue+wood

Wood 100% Control

0.0

1 M

CaC

l 2ex

trac

tab

le C

d (

mg

/kg)

1 extraction 2 extraction 3 extraction 4 extraction

28

chicken manure (30%) +wood (70%) biochar demonstrated low Cd release in the first extraction, but

the amount of mobile Zn increased in the second extraction and remained high in the third and

fourth extractions. On the contrary, soil amended with Vermont biochar released the highest amount

of Zn at the first extraction step– 11 mg/g, and the following extractions provided lower amounts

of extractable Zn – 4, 4 and 2 mg/kg respectively.

Fig. 6. 0.01 M CaCl2 extractable Zn concentration in Lommel soil after 21-day incubation. The error


The control Gongju soil had in total 3.7 CaCl2-extractable mg/kg of As (Fig. 7). The first

extraction released 1.3 mg/kg As from the soil, the next extraction steps gave lower As

concentrations. At the first extraction 1.3, 1.2, 1.2 and 1.2 mg/kg were mobilized from the samples

amended with sorghum, dairy manure, food waste and wood biochars respectively. But the

difference in As mobility was not statistically confirmed, the ANOVA was not significant (F (7,

16)=1.95, p=0.359).

Total Pb concentration released throughout four consecutive CaCl2 extractions was 1.02 mg/kg

at the control soil (Fig. 8). The application of AD+wood, chicken manure+wood and sorghum

biochars caused a slight reduction in Pb mobility but this trend was statistically not significant (F (7,

16)=0.151, p=0.991). Very low amounts of Pb were released in the first and second extractions in

control and biochar amended soils. The third and fourth stages of extraction mobilized higher

amount of Pb.

0

10

20

30

40

50

60

70

Dairy manure Vermontbiochar

Soghum 600C 30% chickenmanure+wood

40% foodwaste+wood

ADresidue+wood

Wood 100% Control

0.0

1 M

CaC

l 2ex

trac

tab

le Z

n (

mg

/kg)


29

Fig. 7. 0.01 M CaCl2 extractable As concentration in Gonju soil after 21-day incubation. The error


Fig. 8. 0.01 M CaCl2 extractable Pb concentration in Gongju soil after 21-day incubation. The error


The Gongju soil had much less Zn than the Lommel soil (10 versus 161 mg/kg) (Fig. 9). In the

case of Gonju soil biochar treatment favored significant Zn immobilization in all amended soils (F

(7, 16)=16.154, p=0.001), especially in the case of Chicken manure and Vermont biochar. After the

first extraction, only 0.2-0.6 mg/g of Zn was released in the treated soils. However, this amount was

much higher after the third and fourth extractions (1.7-2.8 mg/kg).

0

0.4

0.8

1.2

1.6

Dairymanure

Vermontbiochar

Sorghum Chickenmanure +

wood

Foodwaste+wood

AD+wood Wood Contol

0.0

1 M

Cac

l 2ex

trac

tab

le A

s (m

g/k

g)


0

0.2

0.4

0.6

0.8

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

1 M

Cac

l 2ex

trac

tab

le P

b (

mg

/kg)


30

Fig. 9. 0.01 M CaCl2 extractable Zn concentration in Gongju soil after 21-day incubation. The error


4.4.2. Consecutive extraction with 0.05 M EDTA

0.05M EDTA extraction was performed in order to assess the potentially mobile forms or plant

available forms of metals (Takáč et al., 2009). This pool includes dissolved and some exchangeable

fractions of metals. Chelating properties of EDTA favour the formation of soluble metal complexes

which cannot be reabsorbed or precipitated. Single extraction can underestimate the heavy metal

bioavailability (Mendoza et al., 2006), for this reason, consecutive four-stage extraction with 0.05

EDTA was undertaken.

In general, in both studied soils, the bioavailable concentration of all heavy metals except As

was very high at first extraction. Much less was released at further steps of extraction.

Control soil in total released 7.9 mg/kg of Cd 0.05 EDTA extractable Cd (Fig. 10). At the first

extraction 7.2 mg/kg was released. Most biochar amended samples released 7.3-8.5 mg/kg Cd. The

slight reduction of total EDTA extractable Cd concentration in FW+wood and AD+wood samples

was 7.0 and 7.1 mg/kg. But post hoc comparisons did not revealed significant pairwise differences

between the mean scores of the control soil and all the treated soils (p>0.05). Some statistically

significant differences were observed between Dairy manure and FW+wood, and Dairy manure and

AD+wood treatments.

0

1

2

3

4

5

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

1 M

caC

l 2ex

trac

tab

le Z

n (

mg

/kg)


31

Fig. 10. 0.05 M EDTA extractable Cd concentration in Lommel soil after 21-day incubation.

The error bar indicate standard deviation (n=3).

0.05 M EDTA-extractable Pb concentrations are much higher at the first extraction (Fig. 11).

Relatively lower Pb concentrations were detected at further extraction steps, as the amount of

bioavailable Pb was depleted. The total Pb concentration of the control soil was 193 mg/kg.

Soils treated with food waste+wood biochar and Ad+ wood biochar demonstrated slight Pb

immobilization. The total EDTA extractable Pb concentration was 177 and 173 mg/kg respectively.

However, according to one-way ANOVA and post hoc tests these differences were not statistically

significant.

Fig. 11. 0.05 M EDTA extractable Pb concentration in Lommel soil after 21-day incubation. The

error bar indicate standard deviation (n=3).

0

2

4

6

8

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le C

d (

mg

/kg)


0

50

100

150

200

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le P

b

(mg

/kg)


32

Total Zn concentration in the control Lommel sample was 353 mg/kg. (Fig. 12) After treatment

with Dairy manure, Vermont, Sorghum and Chicken manure mixture with wood, the amount of

extractable Zn increased and reached 396, 378, 379, 365 mg/kg. However, these differences were

not statistically significant (p>0.05).

Fig. 12. 0.05 M EDTA extractable Zn concentration in Lommel soil after 21-day incubation.


The lowest total amount of bioavailable As (17.4 mg/kg) was detected in control soil (Fig. 13).

In all biochar amended soils the concentration of bioavailable As increased to 17.6 – 21.3 mg/kg.

The highest values of As concentration were detected for dairy manure and Vermont biochar (21.3

and 21.2 mg/kg respectively). In all these cases the ANOVA was not significant (F (7, 16)=1.470,

p=0.246). The second extraction stage and further extraction stages released relatively high amount

of As in comparison to other heavy metals.

Total concentration of EDTA-extractable Cd in the Gongju soil was 0.39 mg/kg in the control

soil (Fig. 14). After the incubation, the amount of bioavailable Cd increased to 0.41-0.56 mg/kg in

all biochar amended soils. However, according to the Tuckey test, no difference between control

and treated samples was revealed (p>0.05). Statistically significant differences were observed

between Vermont and wood biochars.

0

100

200

300

400

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le Z

n (

mg

/kg)


33

Fig. 13. 0.05 M EDTA extractable As concentration in Gongju soil after 21-day incubation.


Fig. 14. 0.05 M EDTA extractable Cd concentration in Gonju soil after 21-day incubation.


The EDTA-extractable concentration of Pb in control Gongju soil was 129.8 mg/kg Pb (Fig.

15). In all soils treated with biochars the concentration of Pb available for plants changed to 132.4-

162.8 mg/kg. However, all the differences were not significant (p>0.05). The major part of

bioavailable Pb was extracted at the first and second extraction stages.

0

3

6

9

12

15

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le A

s, m

g/k

g


0

0.1

0.2

0.3

0.4

0.5

0.6

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le C

d (

mg

/kg)


34

Fig. 15. 0.05 M EDTA extractable Pb concentration in Gongju soil after 21-day incubation. The

error bar indicate standard deviation (n=3).

Zn concentration throughout the consecutive extraction is presented in Fig. 16. Dairy manure,

Vermont and Sorghum biochars released significantly higher amounts (42.0-58.8 mg/kg) of EDTA-

extractable Zn in comparison to the control soil (38.7 mg/kg).

Fig. 16. 0.05 M EDTA extractable Zn concentration in Gongju soil after 21-day incubation.


0

40

80

120

160

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le P

b (

mg

/kg)


0

10

20

30

40

50

Dairymanure

Vermontbiochar


wood

Foodwaste+wood

AD+wood Wood Contol

0.0

5 M

ED

TA e

xtra

ctab

le Z

n (

mg

/kg)


35

4.4.3. pH stat test results

The pH stat leaching test was aimed to investigate metal release depending on the pH.

The behavior of all studied heavy metals demonstrated the dependence on the pH values (Fig. 17-

22). Leaching curves had similar patterns for Cd, Zn, As. Generally, at lower pH values higher metal

concentrations were detected.

The biochar amended soils demonstrated the same curve patterns, but the amount of released

metals were slightly different. For instance, at acidic pH values Cd was extracted in lower amounts

in treated soils (Venegas et al, 2016). However, dairy manure and Vermont biochars gave a higher

Cd extraction yield. Zn demonstrated the same behavior in Lommel soil. In Gongju soil all treated

soils showed higher concentration of extractable Zn, it was especially high in the case of AD residue

+ wood biochar. No clear trend for Pb in both soils was observed. The maximal Pb yield was

observed at different pH values in case of different biochars, but the amount of released Pb in

biochar amended soils was generally lower in comparison to control soil. In Gongju soil at low pH

Pb was extracted in higher amounts from biochar treated soils, especially in the case of AD +wood

biochar in comparison to the control soils. A different trend was observed in the case of As. After

all biochar application except wood biochar As was extracted from the soil in higher amounts.

36

Fig. 17. Leached amount (mg/kg) of Cd as a function of pH during the pHstat test in samples control

and biochar amended Lommel soil.

Fig. 18. Leached amount (mg/kg) of Pb as a function of pH during the pHstat test in

samples control and biochar amended Lommel soil.

0

1

2

3

4

0 2 4 6 8 10

Extr

acta

ble

Pb

(m

g/k

g)

pH

30%chicken manure+70%wood40%food waste+60%woodAD residue+woodwood 100%

0

0.2

0.4

0.6

0.8

0 2 4 6 8 10

Extr

acta

ble

Cd

(m

g/k

g)

pH

dairy manurevermont biocharsorghumcontrol

0

0.2

0.4

0.6

0.8

0 2 4 6 8 10

Extr

acta

ble

Cd

(m

g/k

g)

pH30%chicken manure+70%wood40%food waste+60%woodAD residue+woodwood 100%

0

1

2

3

4

0 2 4 6 8 10

Extr

acta

ble

Pb

(m

g/k

g)

pHdairy manurevermont biocharsorghumcontrol

37


samples control and biochar amended Lommel soil.

Fig. 20. Leached amount (mg/kg) of As as a function of pH during the pHstat test in

samples control and biochar amended Gonju soil.

0

10

20

30

40

50

0 2 4 6 8 10Extr

acta

ble

Pb

(m

g/k

g)


0

10

20

30

40

50

0 2 4 6 8 10Extr

acta

ble

Zn

(m

g/k

g)


0

0.2

0.4

0.6

0.8

1

1.2

0 2 4 6 8

Extr

acta

ble

As

(mg

/kg)


0

0.2

0.4

0.6

0.8

1

1.2

0 5 10

Extr

acta

ble

As

(mg

/kg)


38


samples control and biochar amended Gongju soil.

Fig. 22. Leached amount (mg/kg) of Zn as a function of pH during the pHstat test in

samples control and biochar amended Gonju soil.

0

1

2

3

4

5

6

7

0 2 4 6 8

Extr

acta

ble

Zn

(m

g/k

g)


0

1

2

3

4

5

6

7

0 2 4 6 8

Extr

acta

ble

As

(mg

/kg)


0

1

2

3

4

5

0 2 4 6 8

Extr

acta

ble

Pb

(m

g/k

g)

pHdairy manure40%food waste+60%woodAD residue+woodwood 100%

0

1

2

3

4

5

0 2 4 6 8

Extr

acta

ble

Pb

(m

g/k

g)


39

5. Discussion

5.1. Effect of biochar on soil physico-chemical properties The significant increase in soil pH in biochar amended soils is widely reported in the literature

and is in agreement with other studies (Bashir et al., 2017, Houben et al. 2013, Abujabhah et al,

2016). The Vermont and Chicken manure+wood biochars had the highest pH values and they

induced the most significant pH change in Lommel and Gongju soils. According to Bashir et al.

(2017) and Yuan and Xu (2011) the biochars with the highest ash content and rich and in oxygen

containing functional groups cause pH to increase. Vermont and CM+wood biochars did not have

the highest ash content among the tested biochars. These amendments were produced mostly from

wood feedstock, which generally have a high percentage of base cations (Ca2+, Mg2+, K+, Na+) (Novak

et al., 2009). These cations can form carbonates, oxides, and hydroxides during the pyrolysis process.

The presence of CaCO3 was confirmed by XRD analysis in chicken manure+wood biochar and sylvite

(KCl) in Vermont biochar. Applied to the soils these substances dissolute, increasing soil pH (Houben

et al., 2013, Novak et al., 2009).

The increase in soil EC was in agreement with previous published results (Fellet et al. 2011;

Mohamed et al., 2015; Lu et al., 2014). EC indicates an increase in salinity of incubated soils because

of the presence of Na, Mg, Ca and carbonates in biochars (Yadav et al., 2016). Sorghum, CM+wood

and FW+wood biochars demonstrated higher effects on soil EC. The presence of CaCO3 and KCl,

was confirmed by XRD. However, in all biochar treated soils EC values did not exceed the saline limit

(4 dS m−1) (Scianna, 2002) and cannot cause salt stress to plant growth, so biochar amendment at

5% application rate cannot affect the plant growth. For the biochar addition at higher application

rates EC values should be carefully calculated in order to avoid the risk of salinization (Yadav et al.,

2016). According to previous findings, the EC in biochar can increase at higher pyrolysis temperature

(Yadav et al., 2016). Although, in the tested biochars this trend was not observed. Probably the

feedstock properties had more prominent effect on the EC values. According to Lu et al. (2014), the

EC increase after biochar application maybe important for metal mobility depending on the

dominant ion interactions in soil solution (Lu et al., 2014).

The increase of soil organic matter content (measured via LOI) was observed in biochar

amended soils of both the Lommel and Gongju sites. These results (Table 6) are in agreement with

previous findings (Raya-Moreno et al., 2017). According to Heiri and Lotter (2001), LOI 550 °C can

overestimate the amount of organic matter in soils because of dehydration processes in clays, and

hydroxides. However, other findings showed that LOI could be accepted as a suitable method for

biochar analysis in the field (Koide et al., 2011). In the current study, FW+wood and AD+wood

biochars induced a soil organic matter change in Gongju soil. This was probably due to a relatively

https://link.springer.com/article/10.1007/s11368-017-1796-z#CR16

https://link.springer.com/article/10.1007/s11356-017-9521-4#CR26








http://www.sciencedirect.com/science/article/pii/S0048969717306897#bb0055

40

high portion of resident matter in these biochars (63 % and 54.3 % respectively). However, for the

Lommel soil organic matter increase was detected in all the treated samples, excluding FW+wood

and dairy manure biochars. Negligible effects of dairy manure biochar could be explained by the

high ash content (85%) and low amount of carbon (9.8%). Generally biochar application induces an

increase of recalcitrant fraction of soil organic carbon and further C sequestration in soils, improving

productivity of crops and soil quality (Lehmann, Joseph, 2009).

5.2. Effect of biochar on the CaCl2-extractable forms of heavy metals

Total metal content is taken into account while considering contamination warning limits in

different regions, but environmental risks mostly depend on mobile and bioavailable fractions of

metals (Houben et al., 2013). According to Pueyo et al. (2004), extraction with 0.01 mol/L CaCl2 is

supposed to be an appropriate method for accessing the mobility of Cd, Cu, Pb, and Zn. Lu et al.,

(2017) suggested that CaCl2 extraction represents metal concentration in pore water. Bergknut et al.

(2007) reported that Cd availability for plants and toxicity for soils could be assessed with CaCl2

extraction.

According to the obtained results, the concentration of CaCl2-extractable Cd and Zn decreased

in all biochar amended soil samples. CaCl2-extracrable Cd concentration in Lommel soil decreased

to 50-90%. The most efficient biochars FW+wood adsorbed the 92% of Cd and Vermont biochar

87% of biochar. Less effect was detected at AD+wood biochar. It decreased Cd concentration to

50%. Similar trends were detected for CaCl2-extractable Zn in Lommel soil. The application of food

waste and Vermont biochars reduced the amount of Zn at 94% and 87% respectively. Lower

reduction rates were observed for AD+wood and wood biochars (64 and 65% respectively). These

results are in accordance with the findings of Houben et al (2013), who observed a reduction in the

mobility of Cd and Zn after biochar application. Cd immobilization in biochar treated soil was also

reported by Wang et al. (2017).

In our study Cd and Zn concentration showed high pH dependency. Both Vermont and

FW+wood biochars had the highest pH values (10.09 and 10.5 respectively) and induced the most

significant pH changes in Lommel and Gongju soils. Under higher pH most heavy metals are

converted to insoluble forms (carbonates, phosphates, oxides and hydroxides) and decrease their

mobility, and heavy metal solubility is reduced (Wang et al., 2017). High pH dependence of CaCl2 -

extractable Cd and Zn concentrations on pH of applied biochars was also confirmed in the studies

of Houben et al (2013) and Rieuwerts et al. (2006).

Another process defining the high performance of Vermont and FW biochars for Cd

immobilization could be the formation of CdCO3 and its further precipitation. In our study, X-ray

diffraction analysis confirmed the presence of CaCO3 mineral in FW+wood biochar, but not in

Vermont biochar. CM+wood, AD+wood and sorghum biochars also showed the presence of CaCO3.

For this reason, potentially CdCO3 could form in these samples and further precipitate. Xu et al.

(2013) suggested that dairy manure biochar often contain carbonate and phosphate, so precipitation

41

plays an important role in Cd stabilization by the biochars of this type. However, in tested dairy

manure sample, this substance was not found, suggesting that in this case other mechanisms are

responsible for metal sorption.

The immobilization performance of the most efficient biochars correlated with surface area

and highest pore volume. Vermont, CM+wood, FW+wood had the highest surface area (246500,

347800, 159200 m2/kg respectively) and highest pore volume (0.7, 0.8, 0.9 cm3/g respectively)

among the studied amendments. A higher degree of aromaticity in these biochars was confirmed

by Raman spectra. Therefore, electrostatic interactions with negatively charged biochar surface could

be suggested as another factor defining the cation sorption (Li et al., 2017). Same findings were

presented by the field study of Bian et al. (2014), studying Cd immobilization in biochar amended

soils.

Oxygen content in the tested biochars had the positive correlation with concentrations of

CaCl2-extractable Zn and Cd. The amount of oxygen was maximal in the most efficient biochars.

Vermont, FW+wood and CM+wood biochars, had 19, 11 and 10% of oxygen respectively. These

findings indicate the possible complexation of Cd and Zn on the surface of oxygen-containing

functional groups (Uchimiya et al., 2011; Ahmad et al., 2014). Moreover, these biochars had the most

pronounced peaks in aliphatic C-O-C functional groups at FTIR spectra, so this group potentially

participates in the sorption of Cd and Zn (Ahmad et al., 2014). Carboxylic and hydroxyl groups were

detected in the wood and wood+CM biochars. Carboxylic groups were also found in dairy manure

and wood+FW biochars. All these biochars induced metal stabilization, but did not provide the

maximal effect on heavy metal immobilization. This finding was in agreement with Trackal et al.

(2014), who reported no shift in carboxyl group peaks at the FTIR spectra before and after biochar

application. For this reason, formation of complexes with carboxyl groups is considered as not the

most important mechanism for Cd stabilization Trackal et al. (2014).

CEC of biochars did not affect the reduction of metal mobility. CEC was relatively high only in

case of wood biochar – it reached 11 cmol(+)/100g. Other biochars had significantly lower CEC

values. Wood biochar demonstrated lower efficiency of Cd. It indicates that cation exchange was a

relatively less important mechanism of metal immobilization. However, according to Harvey et al.

(2011) cation exchange can be the most important mechanism for Cd immobilization in the case

that biochar has high CEC. Moreover, Wang et al., (2014) observed the decrease of CaCl2-extractable

concentration metals decreases at higher CEC (Wang et al., 2014), It could indicate that other

sorption mechanisms were stronger in the studied soils.

In the control soil the amount of CaCl2-extractable Cd and Zn was relatively low, and in the

control Lommel soil the amount of Cd and Zn decreased in the second and further leaching events.

It indicates the depletion of the pool of mobile metals. In Gongju soil Zn concentration was high at

each extraction stage, indicating that sorption was stronger in the loamy soil. After adding the

biochars the total concentration of mobile Cd and Zn generally reduced. However, the amount



42

released at the first step of extraction was the same as at the further extraction stagesm it did not

decrease significantly. The probable reason is that the mobile forms of heavy metals are not strongly

retained by biochars or more time is required for stronger sorption.

According to obtained results, the properties of biochar influencing Cd and Zn behaviour in

soils were pH, BET surface area, oxygen content and amount of oxygen-containing functional groups.

So, for selecting the biochars for field application these properties should be taken into account.

In the studied soil none of the biochar amendments induced Pb immobilization. Generally, Pb

was extracted with 0.01 M CaCl2 in a very low amount. In the control Gongju soil CaCl2-extractable

Pb concentration was 1 mg/kg, while the total concentration was 593 mg/kg. It indicates relatively

low mobility of this element and shows relatively low toxicological risk of leaching to the

groundwater.

The possible explanation for the lack of a pronounced effect of biochar on Pb sorption in

tested soils is absence of P-bearing minerals, which were not detected with X-ray diffraction analysis.

According to Cao et al. (2011), the important mechanism for reducing Pb-concentrations was the

reaction with P-bearing material, further formation and precipitation of hydroxypyromorphite

Pb5(PO4)3OH. Authors consider sorption to biochar surface as a less important immobilization

mechanism (Cao et al., 2011). Another reason could be competitive effect of Cd and Al with Pb

adsorption. Han et al., (2017) observed reduction in Pb adsorption in the presence of Cd up to 19.3-

42.2% and in the presence of Al - up to 80.6-95.8%. The competition by these elements reduced

the available sorption sites on biochars surface (Han et al., 2017).

CaCl2-extractable concentration of As in Gongju soil was very low. With the control soil 3.7

mg/kg was extracted while its total concentration was 448.6 mg/kg. This finding also indicates low

mobility of the element in the soil and relatively low environmental risk. The biochar application did

not induce any significant change in CaCl2-extractable As concentration. Generally, high pH values

after biochar application facilitate As mobility and availability for the plants (Joseph et al., 2010). In

Gongju soil pH remained slightly acidic or neutral, it could be a possible reason that As did not

increase its mobility in soil. The study of Igalavithana et al., (2017) suggested that high As mobility

in treated samples maybe related to the presence of PO43- ion competing with As(V) for the

sorption sites on biochar surface. Moreover PO43- ion were not detected, so the biochar application

did not cause significant mobilization of As ions. It is possible longer field experiments are needed

to predict As behaviour in treated soils.

The efficiency of biochars was slightly different in two studied soils. For instance, changes in

CaCl2-extractable Zn concentration were more pronounced in the Lommel soil, which is more sandy

and poor in organic matter. These findings were confirmed by the study of Melo et al. (2013). The

authors suggest that clay or loamy soils are more able to retain pollutants compared to sandy soils.

43

5.3. Effect of biochar on EDTA- extractable forms of heavy metals

EDTA has strong buffering capacity and ability to chelate the metal cations (Pb, Cd, Zn),

forming soluble complexes with these metals (Naghipour et al., 2016). EDTA extractable

concentration of heavy metals can represent a pool of metals available for plants (Li et al., 2017).

The concentrations of EDTA-extractable metals were very high in the Lommel soil. From the

control soil Cd, Pb and Zn were extracted at 7.9, 194.1 and 355.2 mg/kg, while their total

concentrations were 10.0, 224 and 396 mg/kg respectively. It indicates high mobility in sandy soils

and a high toxicological risk. However, the Gongju soil had relatively low EDTA-extractable

concentrations of elements with relatively low environmental risk introducing the contaminants to

the plants. The amount of As, Cd, Pb and Zn released was 17.2, 0.3, 13.3 and 39.3 mg/kg respectively,

while total concentrations reached 448.6, 5.8, 593.9 and 178.0 mg/kg respectively. Generally, EDTA-

extractable concentrations of Cd and Zn were higher in Lommel soil in comparison to Gongju soil,

following the trends of their total concentration at these sites. It could be suggested that metals are

better retained by the loamy and the richer in organic carbon Gongju soil in comparison to the

sandy Lommel soil. The influence of physico-chemical properties for the amount of bioavailable Cd

was also shown by Li et al. (2017). Their study reported positive correlations of bioavailable

concentration of Cd to soils general physico-chemical properties including pH, OC, CEC and EC.

According to the obtained results, amendments did not cause any significant reduction in

mobility of all studied pollutants in both soils. The possible reason of the minor change in the EDTA-

extractable amount of trace elements is a relatively short 21-day incubation period. It was in

accordance with the study of Almaroai et al. (2014) 21-day experiment was performed in order to

assess Pb bioavailability for plants in biochar amended soils. The irrigation with the deionized water

did not change the amount of Pb available for the plants. Most of the studies report 30-day or 45-

day incubation experiments. For instance, in the 45-days incubation experiment of Igalavithana et

al. (2017), Pb was immobilized by crop residue biochars from the crop residues, but the concentration

of As did not change. Abbas et al. (2017) observed the decrease of EDTA-extractable Cd in

agricultural soil after application of rice straw biochar. According to Li et al. (2017), bamboo and rice

straw biochars were efficient in reducing bioavailability of Zn, Pb, Cd, Cu. Fellet et al. (2014) detected

a good performance of manure biochar for Pb and Cd immobilization.

The possible reason was the lack of pronounced changes in organic carbon in incubated

soils, because organic matter plays an important role in sorption of some metals. Also low change

in pH in the neutral Gongju soil could be another reason for negligible effect of biochar application.

However, some studies also observe a lack of significant effect on the concentration of bioavailable

heavy metals. For instance, Soja et al. (2017) reported mobilizing of EDTA-extractable Cu in neutral

soil and no significant effect compared to biochar amended acidic soil. In the study of Xu et al

44

(2016) little effect of bamboo biochar on Pb and Cd bioavailability was indicated, but at the same

time pH values increased significantly in the treated soils. In addition, findings of Li et al. (2017)

showed high efficiency of rice straw biochar in reducing Cd in lightly contaminated soils, but no

significant result in the soil with high level of contamination. The reason was that capacity of biochar

to adsorb pollutants at the application rate 20 t/ha was not enough to reduce their amount in soils.

In this study, the application rate was 75 t/ha, (5%) so even in case of very high contamination level

at Gongju site the adsorption capacity of biochars should not be exceeded. So a possible reason for

the little reduction in the bioavailable metals in tested soils was lack of change in organic carbon

content. Also, probably the biochar application in the Gongju soil did not cause the significant

change in pH values, and did not reduced metal mobility.

In the studied soils the concentration of CaCl2-extractable Zn and Cd was reduced by biochar

application, indicating lower risk of their leaching to groundwater. Pb and As were less mobile and

did not pose toxicological risk. However, EDTA-extractable fraction, related to plant availability did

not change after biochar application. It means that there is still a risk of metal introduction to the

food chains in agricultural area, especially in case of sandy soil.

5.4. pH-dependent release of heavy metals from biochar amended soils

pHstat leaching tests were conducted to estimate heavy metal behavior in soils and potential

immobilization in situ at changing pH from contaminated soils. In addition, it allows a way to assess

the buffering capacity of various amendments, including biochar (Cappuyns, Swennen, 2008) and

predicting maximal contaminant availability and mobility in soils (Rigol et al., 2009). This approach

indicates the end points for the remediation of contaminated soils and can be useful in assessing

the environmental risk of heavy metal (González-Núñez et al., 2012, Kosson et al., 2002).

Tests demonstrated the dependence of metal leachability on pH change. For instance, in

both studied soils Cd and Zn displayed an increased release at higher pH values. It was in agreement

with previous findings. According to Cappuyns, Swennen (2008) Cd and Zn generally are rapidly

released following slower release at the end of the experiment. These elements could be desorbed

easily at low pH or they can be released from nonstable carbonates. The reason for their increased

release under acidic conditions was competition of protons with metal cations for sorption sites and

higher amount of protons at lower pH (Venegas et al., 2016, González-Núñez et al., 2012).

Biochar application slightly changed the amount of released metals. At acidic pH values Cd

and Zn were extracted in lower amounts in treated soils. It can be possibly explained by the

increasing of the amount sorption sites with addition of biochars (Venegas et al, 2016). However,

the addition of Vermont and dairy manure biochars caused the reduction of Cd and Zn extraction

http://www.sciencedirect.com/science/article/pii/S001670611630249X#!

45

at pH=4. These two biochars had lower CEC in comparison to other biochars (1,48 and 1.66

cmol(+)/100 g respectively). For this reason, they probably did not have a high amount of sorption

sites to retain metals. Therefore, Vermont and Food waste biochar showed a promising sorption

capacity for CaCl2-extractable Cd and Zn. However, only the FW+wood biochar was efficient for

decrease in Cd and Zn leaching in both soils under low pH.

In both tested soils, Pb was leached in very small amounts and no clear trend was observed

in Pb behavior. In the studied soils tests were conducted between the pH values 4 and 7,5, but

changes in its leaching behavior generally start at lower pH (Cappuyns and Swennen, 2008). Thus in

the environment at lower pH the risk of Pb mobilization remains low. For Pb in the Gongju soil,

AD+wood and Vermont biochar showed slightly higher concentration than in the control soil. The

reason could be the same as in case of Zn and Cd. Other treatments did not display any effect on

Pb leaching rates.

Arsenic was also released in a very low amount at various pH-values in comparison to its

total concentration, which indicates its low mobility. Cappuyns et Swennen (2008) reported that As

exists is an anion and demonstrates reabsorption behavior in pH leaching test. However, these

changes occur from studies with pH values 4-7. For As the application of all biochars demonstrated

slightly higher concentrations of the pollutant at acidic pH in comparison to the control soil.

However, the general yield of As was maintained low and its leaching rates were almost negligible.

Generally, according to the obtained results, Gongju soil had generally smaller

concentrations of all released metals in comparison to Lommel soil. The reason could be stronger

sorption of metals by clay minerals and organic matter in this soil.

Results of this study are similar to the findings of Houben et al. (2013). The authors did not

observed significant change in metal release in biochar treated soils. However, biochar application

caused an increase of acid neutralizing capacity, so a longer period will be required to reach

hazardous pH levels. According to Houben et al. (2013), biochar application is useful if the soil pH

is a problematic factor.

Taking into the account the results of both CaCl2 consecutive extraction and pHstat leaching

test, the most effective biochar was FW+wood biochar. Its application caused the decrease of CaCl2-

extractable Cd and Zn. Moreover, under the condition of decreasing pH, the metal release will be

lower than in untreated soils. Application of the Vermont biochar can reduce Cd and Zn

concentration at neutral or base pH. In case of acidic pH, the metal release increases.

46

6. Conclusion and recommendation

6.1. Conclusion According to the obtained results, the most effective soil amendment was FW+wood biochar.

The application of this amendment caused reduction of CaCl2-extractable Zn and Cd in both studied

soil. Moreover, at low pH the amount of leached metals decreased after application of this biochar.

Vermont biochar showed good remediation potential, but at pH=4 it caused metal mobilization in

comparison to the control soils.

The most important properties of biochars causing their high sorption capacity were high pH,

high O-content, high surface area and the presence of C-O-C functional groups. Therefore, it could

be suggested that the most important mechanisms of Cd and Zn removal were electrostatic

attraction and complexation with oxygen-containing functional group. Moreover, CEC is important

for the performance of biochars under changing pH conditions.

The behavior of CaCl2-extractable As and Pb was not affected by biochar addition. However,

their extraction rate and mobility in the studied soils was relatively low and they did not pose high

risk of leaching to groundwater at studied sites. Loamy soil demonstrated higher retention of heavy

metals in comparison to sandy and soils poor in organic matter. For this reason, biochar application

was more effective in sandy soils.

The EDTA- extractable concentration of metals did not reduce after biochar addition.

Therefore, in sandy soil there is still an environmental risk of plant uptake and transfer to the food

chains.

Moreover, biochar application induced changes in soil chemical properties. For instance, it

had significant liming effect on both studied soils. Vermont and CM+wood biochars showed the

highest pH increase due to the higher amount of base cations. Soil electrical conductivity was higher

in Lommel and Gongju soils after adding the biochar, but did not exceed saline limit. It indicates

that 5% application rate of tested biochars does not bring the risk of soil salinization. The increase

of soil organic matter content was detected when biochars with a higher portion of volatile matter

were added to the soils (FW+wood and AD+wood).

6.2. Recommendation

Seven biochars were screened in this study for their capability to remediate contaminated

soils. Food waste+wood biochar showed best potential for metal immobilization. Further research

could be focused on more detailed study of this biochar. Biochars produced under different

conditions and with different proportion of food waste and wood waste in a feedstock have to be

systematically studied. Moreover, various soil types with different set of contaminants and different

47

concentrations should be tested as the conditions of Gongju and Lommel soil could not be

extrapolated to many sites. Also, adding of food waste+wood biochar at different application rates

should be tested in order to make remediation effective and economically feasible. In addition,

longer field experiments are required to assess the aging processes and long-term sorption behavior

of metals.

This study showed that the most important properties of biochar affecting the sorption of Cd

and Zn are pH, surface area, oxygen content and CEC. Searching more biochar with similar properties

could be useful for developing practical recommendation for agriculture.

48

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Master’s dissertation submitted in partial fulfilment of the ......Prof. Filip Tack for guiding me during all the stages of the work on this master thesis and for the opportunity