Assessment of mobility and bioavailability of mercury compounds in sewage sludge and composts

Environmental Research 156 (2017) 394–403

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Assessment of mobility and bioavailability of mercury compounds in sewage sludge and composts☆

MARK

⁎

Beata Janowska , Kazimierz Szymański, Robert Sidełko, Izabela Siebielska, Bartosz Walendzik Koszalin University of Technology, Faculty of Civil Engineering, Environmental and Geodetic Sciences, Department of Waste Management, ul. Śniadeckich, 75-453 Koszalin, Poland

A R T I C L E I N F O

A B S T R A C T

Keywords: Mercury Fractionation Bioavailability Sewage sludge Compost Mobility factor

Content of heavy metals, including mercury, determines the method of management and disposal of sewage sludge. Excessive concentration of mercury in composts used as organic fertilizer may lead to accumulation of this element in soil and plant material. Fractionation of mercury in sewage sludge and composts provides a better understanding of the extent of mobility and bioavailability of the different mercury species and helps in more informed decision making on the application of sludge for agricultural purposes. The experimental setup comprises the composing process of the sewage sludge containing 13.1 mg kg−1 of the total mercury, performed in static reactors with forced aeration. In order to evaluate the bioavailability of mercury, its fractionation was performed in sewage sludge and composts during the process. An analytical procedure based on four-stage sequential extraction was applied to determine the mercury content in the ion exchange (water soluble and exchangeable Hg), base soluble (Hg bound to humic and fulvic acid), acid soluble (Hg bound to Fe/Mn oxides and carbonates) and oxidizable (Hg bound to organic matter and sulphide) fractions. The results showed that from 50.09% to 64.55% of the total mercury was strongly bound to organo-sulphur and inorganic sulphide; that during composting, increase of concentrations of mercury compounds strongly bound with organic matter and sulphides; and that mercury content in the base soluble and oxidizable fractions was strongly correlated with concentration of dissolved organic carbon in those fractions.

1. Introduction Management and disposal of sewage sludge being the product of sewage treatment is in highly industrialised countries a logistic, economic, and primarily, environmental problem. Sewage sludge is considered as dangerous waste as it contains high organic load, chemical pollutants including heavy metals, pesticides and other dangerous organic compounds (Białobrzewski et al., 2015; Dong et al., 2013). Sewage sludge also makes sanitary hazard due to the occurrence of pathogenic bacteria, viruses and other pathogenic organisms (Bień, 2002; Iacovidou et al., 2012). The main disposal processes of sewage sludge is its use in agriculture and degraded land reclamation. The disposal processes comprise composting, incineration and deposition (Ramdani et al., 2015; Samolada and Zabaniotou, 2014; Fonts et al., 2012). Sewage sludge intended for agriculture application and soils

reclamation should comply with the requirements pertaining to its chemical composition, including heavy metals content and sanitary conditions. Excessive heavy metals concentration limits the use of sewage sludge for fertilization due to its toxic action and ability to bioaccumulate in plant material (Dong et al., 2013; Ingelmo et al., 2012; Ramdani et al., 2015). The process of sewage sludge composting could be used for producing organic fertilizer, biofuel and other forms of renewable energy sources (Fonts et al., 2012; Iacovidou et al., 2012). However, heavy metals content, including mercury, in composts and biofuel limits the possibility of the use of this material in agriculture and energy generation. One criteria for assessing the quality of compost is the content of heavy metals, including mercury compounds in such material (Hseu et al., 2010; Janowska and Szymański, 2009; Confesor et al., 2008; Szymański et al., 2005). Mercury is one of the most dangerous environmental pollutants, featuring high chemical and biological activities that create compounds

Abbreviations: EX, exchangeable fraction; BS, base soluble fraction; AS, acid soluble fraction; OX, oxidizable fraction; PE, population equivalent; HgT, total mercury concentration; TOC, total organic carbon; DOC, dissolved organic carbon; Mf, mobility factor; r, Pearson's correlation coefficient ☆ This research work was financed by the Polish State Committee for Scientific Research within the project No. N N523 743840. ⁎ Corresponding author. E-mail addresses: [email protected] (B. Janowska), [email protected] (K. Szymański), [email protected] (R. Sidełko), [email protected] (I. Siebielska), [email protected] (B. Walendzik). http://dx.doi.org/10.1016/j.envres.2017.04.005 Received 30 July 2016; Received in revised form 28 February 2017; Accepted 4 April 2017 0013-9351/ © 2017 Elsevier Inc. All rights reserved.

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2. Methods

of diversified properties (AMAP, 2013, Syversen and Kaur, 2012). Biogeochemical mercury circulation depends not only on its concentration but, primarily, on its forms of occurrence. Mercury may occur in nature in the form of volatile alkylated compounds, particularly as methyl mercury and ethyl mercury, or in the elemental form Hg0 (Kabata-Pendias, 2010; Mao et al., 2016). It can also form water-soluble complex compounds where ligands can be chlorides or sparingly soluble organic complexes (Hutchison and Atwood, 2003; Kabata-Pendias, 2010; Han et al., 2003; Wang et al., 2012; Xu et al., 2015). Mercury shows strong affinity to compounds containing sulphur and chlorine as well as to organic compounds and organic matter (Hutchison and Atwood, 2003; Kabata-Pendias, 2010; Xu et al., 2015). Organomercury species such as methyl mercury are more mobile than inorganic mercury forms, and thus more easily bioaccumulated and considered as more toxic to humans than either elemental Hg0 or Hg salts. Mercury readily chelates with organic matter in the form of humic and fulvic acids, forming stable complexes (Hutchison and Atwood, 2003; Kabata-Pendias, 2010; Xu et al., 2015). The quantity of this element in sewage sludge generated in wastewater treatment plants depends on the wastewater treatment technology and on the wastewater properties (Balogh and Nollet, 2008; Mao et al., 2016). In the studies by Mukherjee et al., (2004) the average values of the total mercury concentration in sewage sludge were all in the range from 0.6 to 3 mg kg−1 (DM). The environmental mobility and toxicological effects of mercury are strongly dependent on the chemical species present (Kabata-Pendias, 2010). Mercury in sewage sludge and compost could be extracted by means of various chemical reagents in order to determine the different mercury species and partitions, providing useful information about their toxicology, bioavailability and biogeochemical reactivity. Sequential extraction allows for determination of bond strengths and differentiation of chemical species of mercury: elemental mercury Hg°, water-soluble compounds (HgCl2), compounds associated with carbonate, associated with hydrous oxides of Fe and Mn, bound to organic substances and mercury (II) sulphide (Bloom et al., 2003; Boszke et al., 2008; Fernandez–Martinez et al.al., 2006; Fernandez-Martinez and Rucandio, 2013; Han et al., 2003; Han et al., 2006; Issaro et al., 2009; Ram et al., 2009; Reis et al., 2010). Many research papers were dedicated to mercury speciation in environmental and biological samples. The literature survey presented in Ibanez-Palomino et al. has shown that approximately 80% of the papers pertain to definition of forms of mercury occurring in water, bottom sediments and in aquatic organisms; approximately 12% of the papers refer to determination of mercury forms in biological samples, and 5% of the papers pertain to soil (Ibáñez-Palomino et al., 2012). Information describing fractionation of mercury in samples of compost and sewage sludge can be found in few publications (Cappon, 1987; Shoham-Frider et al., 2007; Lomonte et al., 2010), although the fractionation of mercury based on sequential extraction allows for definition of mobility and bioavailability of this element. There is a need for further research leading to the definition of the qualitative and quantitative forms of mercury, which findings could help evaluate the fertilizing properties of composts. In line with this motivation, the objective of the study documented in this article was to examine the relationships between the changes in the organic matter and bioavailability of Hg. The study applied sequential chemical extractions to determine Hg partitioning in sewage sludge and composts. The results were tested on the mixture of sewage sludge and timber shavings during its aerobic bath composting and with an initial C/N ratio of 26.

2.1. Test material The used sewage sludge was obtained from the wastewater treatment plant of Sianów, Poland, featuring the capacity of 15000 PE (population equivalent). Samples of the sewage sludge were taken from the open air sludge drying bed, in autumn and spring (2011–2012), where it was stored for approximately one year (it was dependent on the weather conditions). This was a mechanical and biological wastewater treatment plant using trickling filter, which was built and put into operation at the beginning of the 1990s. The mechanical system comprised a manually cleaned screen, two-chambers and a trap featuring a trapezoid-shape cross-section. The biological system comprised a trickling filter with sprinkler system. The sediment part of the wastewater treatment plant included the secondary sedimentation tank with sludge scraper, open sludge chambers where the sludge was concentrated, and drying beds. In 2015, the said wastewater treatment plant was closed. 2.2. Composting of sewage sludge The composting of the sewage sludge was performed in two static bioreactors (KI and KII) with forced aeration, featuring the capacity of 60 L. The bioreactors were enclosed, insulated containers with controlled flow of air and temperature (Siebielska, 2014). The bioreactor feedstock was approximately 50 L. Intense aeration was performed during the first 21 days of the process. During this stage, the volume of air supplied to bioreactors was 2.5 L min−1 (Siebielska and Sidełko, 2015). The material intended for composting was a mixture of sewage sludge with some amount of structural material such as timber shavings. The weight proportion of the sewage sludge to the structural material was 2:1. In order to ensure uniform distribution of air across the entire volume, polypropylene rings were added to the composted biomass; the proportion of weight of the sewage sludge to the polypropylene rings was 10:1. Each composting cycle lasted for 182 days. During the first composting cycle, composted biomass samples were taken for testing after 3, 7, 10 and 14 days, and subsequently every 7 days, i.e. after 21 and 28 days. For subsequent 6 weeks, the samples were taken every 14 days, every 21 days during the following 12 weeks, and the final sample was taken after 28 days. 2.3. Physico-chemical analyses Moisture, organic matter content, pH, total organic carbon (TOC), total nitrogen and total mercury content (HgT) were determined in samples of sludge waste intended for composting and taken from the bioreactors. The organic matter content was determined by the loss on ignition of the dry mass at 550 °C. The content of total organic carbon and total nitrogen were determined using the Vario Max CN macro analyser. The total mercury (HgT) content was determined in air-dry samples of the tested material, which was ground and sifted through 0.75 mm mesh sieve. In such prepared samples, the content of HgT was determined using spectrophotometer LECO AMA-245 dedicated to mercury determination. Mercury fractionation was performed based on the sequential extraction regime presented in Ram's paper, which recommended this method as highly repeatable; extraction solutions and extraction conditions applied prevented mercury loss (Ram et al., 2009). Mercury compounds contents were determined in the ion exchange (EX), base soluble (BS), acid soluble (AS) and oxidizable (OX) fractions. Air-dried, ground and sifted (0.75 mm) 1 g sample was subjected to sequential extraction, the regime of which is presented in Table 1. After each extraction stage, the samples were centrifuged (MPW – 350 395

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Table 1 Sequential extraction procedure (Ram et al., 2009). Step

FI FII FIII FIV

Fraction

Exchangeable (EX) Base soluble (BS) (HA & FA) Acid soluble (AS) (Fe and Mn oxides) Oxidizable (OX)

(organics and sulphides)

Extraction solution

Table 2 Physico-chemical properties of the sewage sludge intended for composting and of biomass constituting bioreactor feedstock. Extraction conditions Temperature

Duration

15 mL 10% KCl 10 mL 0.1 M NaOH

20 °C 20 °C

1h 30 h

10 mL1.0 M HCl

20 °C

6h

50 °C

5h

7 mL30% H2O2 (pH=2) +2 mL 0.02 M HNO3 4 mL 2.0 M NH4Cl w 20% HNO3 topped up with water to 25 mL

20 °C

Unit of measure

Sewage sludge

Bioreactor feedstock

Moisture Organic matter pH TOC Total nitrogen HgT C/N

% % – % % mg kg−1 DM –

78.0 ± 0.8 68.0 ± 0.3 6.58 ± 0.02 35.62 ± 0.17 2.97 ± 0.01 13.1 ± 1.6 13:1

65.0 ± 1.4 83.5 ± 2.12 6.19 ± 0.03 39.96 ± 0.64 1.67 ± 0.08 7.07 ± 0.57 26:1

xides, and with carbonates (Boszke et al., 2008). Following the extraction with 1 M HCl, dissolution of carbonates can occur with formation of mercury II chloride, which is a readily soluble compound (Ram et al., 2009). The statistical analysis of the results obtained was performed using STATISTICA PL 12 package (StatSoft, Poland).

1h

centrifuge) for 10 min at 5000 rpm, then washed with 5 mL of deionised water and centrifuged again. Both eluates were combined and the mercury content was determined in such prepared extracts through the LECO AMA-254 mercury analyser. Three parallel determinations were performed for each tested sample. In specific eluates obtained after each extraction stage, dissolved organic carbon (DOC) content was determined using the Vario Max CN macro analyser in accordance with the manufacturer's application note.

3. Results and discussion 3.1. Composting The composting of the sewage sludge was performed in model conditions in static bioreactors. Increase of temperature during the first stage of composting and the loss of organic matter is an important indicator to evaluate the intensity of the processes performed (Bernal et al., 2009; Białobrzewski et al., 2015). The composted sewage sludge reached the maximum temperature of 48 °C but the period of hightemperature composting did not exceed 10 days. This could be caused by the small volume of the composted material (50 L). The cooling of the composted feedstock could also be caused by the heat exchange with the surroundings, originating from insufficient system insulation. Table 2 shows the average values of the selected physico-chemical parameters describing properties of the sewage sludge and the material that makes the bioreactor feedstock. The concentration of HgT was 13.1 ± 1.6 mg kg−1 DM in the composted sewage sludge and 7.07 ± 0.57 mg kg−1 (DM) in the bioreactor feedstock. Such sewage sludge can be used for agricultural purposes and for land reclamation. According to the regulation by the Polish Ministry of Natural Environment of 6 February 2015 (O.J.L. 2015, Section 257) the admissible content is 16 mg kg−1 (DM). Such high mercury concentration in the tested sewage sludge could be caused by infiltration of metallic mercury that makes the sealing of the trickling filter sprinkler bundage. Total mercury concentration (HgT) was increasing during composting in subsequent days. The average HgT content in the samples taken from the bioreactor after 182 days was 10.8 ± 0.4 mg kg−1 DM (Fig. 1). Due to the mercury content, such produced composts cannot be used as organic fertilizers. According to the Regulation by the Polish Ministry of Agriculture and Rural Development of 18 June 2008 concerning the execution of some regulations of the Fertilizers and Fertilization Act (O.J.L. No. 119, Section 765), the mercury content in organic fertilizers that support plant growth must not exceed 2 mg kg−1 DM. In the sewage sludge, the total organic carbon (TOC) content was 35.62% and the total nitrogen concentration was 2.97%. The average TOC content in the samples taken from the bioreactor during composting was kept within the limits of 39.96% (bioreactor feedstock) to 34.06% (after 182 days). The TOC content deficit during composting was 14.7% compared with the content in the compost feedstock. Therefore, it can be assumed that the decomposition of organic matter and the formation of humic compounds had indeed happened (Epstein, 1997; Bernal et al., 2009). The C/N ratio in the sewage sludge samples was 13:1, increasing to 26:1 after addition of the timber shavings. The C/N ratio in the samples

2.3.1. UV–Vis spectroscopy of compost samples In NaOH supernatants obtained after the second extraction stage, absorbance (A) was measured at the λ=280 nm, λ=472 nm and λ=664 nm wavelengths (Zbytniewski and Buszewski, 2005) (UV–VIS – 6715UV/VIS JENWAY spectrometer). Then the absorbance ratios of Q2/6=A280/664; Q4/6=A472/664; Q2/4=A280/472 were determined to indicate the degree of humification. The Q2/4 ratio reflects the proportion between the lignins and other materials at the beginning of humification, and the content of the materials at the beginning of transformation. The Q2/6 ratio denotes the relationship between the non-humified and strongly humified material. The Q4/6 ratio, often called the humification index, is the most often calculated ratio. 2.4. Mobility factor (Mf) The mobility of elements was defined as their ability to transfer from the sample solid phase where the element in a given form is poorly bound and can be freed in natural conditions into ionic form (Boszke et al., 2008; Zhu et al., 2014). To determine the mobility of metals in environmental samples, the following formula (Dong et al., 2013) was used to calculate the mobility factor Mf:

Mf =

Parameter

FI + FII + FIII ∙ 100% HgT

where FI, FII and FIII represent the concentration of Hg extracted from the EX, BS and AS fractions; and HgT represents the total concentration of mercury in mg kg−1 of dry sludge. The ion exchange (EX), base soluble (BS) and acid soluble (AS) fractions were considered as mobile fractions. In the base soluble fraction, apart from the humic acids generated in the compost ripening stage, organomercury mobile compounds can occur. Mercury forms, with humic and fulvic acids, constitute stable complex compounds but those compounds can decompose, which can lead to the release of mercury into the natural environment (Fernandez-Martínez and Rucandio, 2013; Lomonte et al., 2010; Shoham-Frider et al., 2007). The acid soluble fraction has been operatively defined as “reactive mercury compounds” or “bioavailable inorganic mercury”. Mercury can also form bonds with iron monosulphides, iron and manganese hydro396

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Fig. 1. Change of HgT, total organic carbon contents and C/N values during composting.

taken after 182 days was 16:1. The changes of the C/N ratio is illustrated in Fig. 1. The obtained compost had dark colour and earthy smell.

compost samples taken from the bioreactor on the 182nd day was 5.94. The values of the Q4/6 absorption ratio exceeding 5 may indicate that the humification process was incomplete.

3.1.1. UV–VIS spectroscopic properties of the compost organic matter UV–VIS spectrometry is a broadly applied technique used for determination of molecular properties of humic substances, i.e. molecule sizes and the degree of condensation and polymerization of aromatic constituents (Zbytniewski and Buszewski, 2005; Sellami et al., 2008; Albrecht et al., 2011). Based on the analysis of absorption spectra of humic substances in the alkaline extracts within the UV–VIS range, areas specific to the composting process can be determined. There are three important areas in a spectrum that are related to absorbance measured at the λ=280 nm, λ=472 nm and λ=664 nm wavelengths. The maximum absorbance at λ=280 nm is caused by the occurrence of chinone and phenolic structures originating from the decomposition of lignin and aliphatic structures that are characteristic to the initial composting stage. Absorbance at the λ=472 nm wavelength originates from the occurrence of organic compounds being products of the depolymerization of macromolecules by microorganisms, and reflects chemical composition at the beginning of the humification process. The presence of aromatic compounds containing many oxygen atoms causes absorbance of the radiation at the λ=664 nm wavelength. The presence of such structures indicates compost maturity. The determination of the Q2/4=A280/479; Q2/ 6=A280/664; Q4/6=A472/664 absorbance ratios allows for definition of the degree of compost maturity and transformations of organic matter during composting (Zbytniewski and Buszewski, 2005; Albrecht et al., 2011). Changes of the Q2/4, Q2/6 and Q4/6 ratios that take place during composting are illustrated in Fig. 2. The decrease in values of those ratios was noted in the samples of composted sewage sludge after three days. At that time, rapid microbiological decomposition of sugars and proteins could happen (Shao et al., 2013). Then, before the 21st day, considerable increase of values of those ratios had happened. As of the 28th to 70th day of composting, insignificant increase of values of those ratios was observed. The resynthesis processes started to prevail at that stage of composting. Small increases of the Q2/6 and Q4/6 ratios indicate the presence of the compounds containing phenolic and benzenecarboxylic groups in their structures (Sellami et al., 2008). After the 70th day of the composting process, the determined values of the ratios stabilised (Fig. 2). In particular, the value of the Q4/6 ratio of the

3.2. Sequential extraction 3.2.1. Mercury fractionation Mercury fractionation in the sewage sludge and composts was performed using the analytical procedure regime presented by Ram et al. (2009). The highest values of the Hg concentration during the process were determined in the OX fraction, whereas the EX and AS fractions featured the lowest Hg concentrations. Mercury in the sewage sludge occurred mainly in the form of compounds with organic matter and sulphides. The average percentage of Hg in the OX fraction was 64.95% ± 4.50% of the total Hg content. The percentage of mercury compounds bound with the fulvic and humic acids was 7.68% ± 1.26%, in the acid soluble fraction was 0.19% ± 0.02%., and in the ion exchange fraction was 0.14% ± 0.07%. Similar distribution of mercury compounds was determined by Ram et al. in bottom sediments using the same sequential extraction regime (Ram et al., 2009). Mercury occurred in the tested samples, mostly in the oxidizable fraction, in the range from 65% to 95%. Mercury compounds in the base soluble fraction ranged between 5% and 11% of the total mercury content, whereas those soluble in acids constituted approximately 1.5% of the content. The concentration of mercury ionic forms was below the limit of determination (Ram et al., 2009). Fractionation of mercury in the activated sewage sludge obtained from the certified reference material NIST-2781 was described in Shoham-Frider's paper (Shoham-Frider et al., 2007). The authors applied a five-stage sequential extraction based on the analytical regime developed by Bloom et al. (2003) and Shoham-Frider et al. (2007). Total Hg content and its concentration in particular eluates obtained in subsequent extraction stages was determined by the application of the CV-AFS technique. In the tested samples, mercury occurred mainly in the fraction IV extracted with 12 M HNO3, with Hg percentage constituting 70% of the total content of this element. This fraction was defined as strongly complexed, i.e. containing elemental Hg°, mercury in the form of sulphur-organic compounds, Hg-Ag amalgamates and Hg bound with crystalline Fe/Mn oxides. Mercury content in the fraction obtained from the extraction with 1 M KOH was 10% (FIII – organic Hg chelate complexes). Mercury in the mercury sulphide form – fraction V (aqua regia), made approximately 7% of the total content. 397

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Fig. 2. Change of absorbance ratios: Q2/4, Q2/6 and Q4/6 in 0.1 M NaOH extract during composting.

Average percentage of the organomercury compounds soluble in NaOH in the bioreactor feedstock made 10.16% of the total Hg content. As of the beginning of the process, the decrease of the percentage of the base soluble Hg compounds was noted. The lowest value was noted in the samples taken from the bioreactor on the 91st day; the value made 4.00% of the Hg content (Fig. 4). At this stage of the process, organic matter decomposition occurred. At the final composting stage, i.e. from the 112th to the 182nd day, the percentage of Hg in the BS fraction slowly increased to reach 5.14% at the last day of the process. At this stage, re-synthesis of the organic compounds and binding of Hg with humic acids being generated at that time was probably going on. The chemical composition of this fraction depended on the speed of the biochemical transformations associated with the decomposition of organic matters and with resynthesis of the humic substance compounds. Mercury present in the fraction bound with organic matter and sulphides (OX) in the bioreactor feedstock made 53.26% of the total mercury content. Change of the mercury percentage in the OX fraction during composting is illustrated in Fig. 5. During the first three days of the composting process, this percentage decreased down to 50.09%, maintained until the 10th day. In the 14th day of the process, the percentage increased to 58.23% and stayed at that level until the 91st day. At the final stage of composting, the Hg percentage in the OX fraction increased again, reaching 64.55% HgT on the 182nd day.

The percentage of the water soluble (FI) mercury compounds was 0.03 ± 0.05%. However, compounds soluble in 0.1 M acetic acid and in 0.01 M HCl (FII) made between 1.0% and 1.3% of the total content of the researched element (Shoham-Frider et al., 2007). Similar distribution of mercury compounds in particular fractions was obtained for the certified reference material. The tested sewage sludge contained approximately 31% of total organic carbon. The same sequential extraction regime was used by Lomonte et al. for evaluation of mobility and bioavailability of mercury occurring in sewage sludge (Lomonte et al., 2010). The authors performed fractionation of mercury in sewage sludge samples taken from the sludge lagoons. The concentration of total organic carbon in those samples varied between 36% and 44%. Unlike the results obtained by ShohamFrider et al., mercury in those samples occurred mainly in the fraction III – 59% (base soluble) (Shoham-Frider et al., 2007). Mercury compounds present in the fractions FIV and FV made approximately 40% of the total content of this element. However, the percentages of water soluble mercury compounds – FI and those occurring in fraction II were below 2% (Lomonte et al., 2010). Mercury percentage in the EX fraction of the bioreactor feedstock made 0.37% of the total Hg content. After three days of composting, it decreased to 0.22%. Then, small increase of the mercury compounds percentage in the determined fraction was noted before the day 14. In subsequent days of the process, percentages changed in small amounts ranging between 0.24% and 0.34%. The content of the ion exchangeable mercury compounds in the samples taken on the 182nd day of composting made 0.31% of the total Hg content. Changes to the Hg percentages in the ion exchangeable fraction are illustrated in Fig. 3. Mercury compounds present in the AS fraction in the mass of the bioreactor feedstock made 0.32% of the total Hg content. However, average percentage of the acid soluble compounds remained within the interval of 0.21–0.43% (Fig. 3). Until approximately the 28th day, the percentage was slightly increasing whereas it decreased after the 42nd day. Mercury compounds soluble in acids in compost samples taken at the end of the composting process made 0.21% of the total Hg concentration, while mercury compounds soluble in acids made small percentage of the total mercury content in the composted sludge.

3.2.2. DOC content The average percentage of DOC in the tested sewage sludge was 1.74% in the EX fraction, 14.05% in the BS fraction, 2.06% in the AS fraction and 6.17% in the OX fraction. The highest DOC concentration values that occurred during the composting process were found in the BS and OX fractions whereas the lowest values in the EX fraction. The DOC percentage in the EX fraction in the compost feedstock was 1.22%. After 7 days of the process, the percentage increased up to 1.80%. This stage shown rapid increase of the temperature originating from the intense microbiological decomposition of such organic compounds as monosaccharides, proteins, peptides and aminoacids (Epstein, 1997; Shao et al., 2013). During the following days, the DOC 398

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Fig. 3. Change of the mercury compound percentages in the EX and AS fractions occurring during the composting process.

ing, the percentage of the tested analyte increased up to 2.04% of the TOC. Then, as of the 21st day, it decreased (1.29%) and stabilised during subsequent days at the level not exceeding 1.5%. During the last weeks of composting, i.e. from the 154th day, a small increase up to 1.75% was noted (Fig. 6). The applied sequential extraction regime has defined fraction BS as a fraction in which mercury occurred in a form bound with humic and fulvic acids (Ram et al., 2009). However, the chemical composition of these solutions depends on the composting stage. The technique of separating humic substances consisting in their extraction with sodium hydroxide is non-specific because, in such condition, also those compounds that are not of humic compounds type can migrate to the solution (Sellami et al., 2008; Zbytniewski and Buszewski, 2005).

percentage decreased, reaching its lowest value of 0.81% during the period from the 21st to the 42nd day. Subsequently, the DOC percentage increased to approximately 1.00% and remained at this level until the 133rd day. In the end stage of the composting process, the DOC percentage decreased in the EX fraction. The percentage was, in the samples taken on the 182nd day, 0.87% of the total TOC content (Fig. 6). Content decrease of the dissolved organic carbon in the EX fraction was observed during composting. Another change in analyte was noted in the paper published by Zbytniewski and Buszewski: increase of water-soluble DOC concentration before the 19th day of composting (Zbytniewski and Buszewski, 2005). The percentage of DOC in the AS (acid soluble) fraction in the bioreactor feedstock was 1.28% TOC. Before the 14th day of compost-

Fig. 4. Percentage changes of the mercury compounds in the BS fraction and the Mf factor during composting.

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Fig. 5. Percentage changes of the mercury compounds in the OX fraction during composting.

Fig. 6. Change of percentage of DOC in the EX and AS fractions during composting.

decomposition was progressing, fatty acids, methyl esters of those acids and products of lignin decomposition, i.e. hydroquinone groups, appeared. The content of aromatic organic compounds increased whereas the content of aliphatic groups decreased. During compost ripening stage oxygen content increased as this originated from the presence of carbonyl and phenol groups (Amir et al., 2006a, 2006b; Veeken et al., 2000). The DOC concentration in the oxidizable fraction corresponds to the content of organic compounds, which did not decompose to carbon dioxide in the extraction conditions. The DOC percentage in the OX fraction, in biomass intended for composting, was 4.87% of the TOC. During the first two weeks of composting, the DOC percentage increased up to 5.96% on 14th day (Fig. 7). In subsequent days, the

According to Fernandez-Martinez and Rucandio, a more selective reagent for mercury compounds bound with fulvic and humic acids is Na4P2O7 (Fernandez-Martinez and Rucandio, 2013). The percentage of DOC in the BS fraction in the biomass intended for composting was 10.53% of the TOC. During the first stage of composting, the percentage was decreasing and on the 56th day, it reached the lowest value of 7.20%. During the first days of the process, this fraction contained sugars, lignin, nitrogen-containing organic compounds, and organic compounds of low molecular weight. Those compounds are easily biodegradable (Jouraiphy et al., 2005; Shao et al., 2013). During subsequent days, this value increased slightly and amounted to, in samples taken during the last day of composting, 8.81% of the total organic carbon content (Fig. 7). As microbiological 400

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Fig. 7. Change of percentage of DOC in the BS and OX fractions during composting.

in that fraction was increasing. Concentration of mercury compounds in the EX fraction was positively correlated with absorbance ratios: Q2/4 (r=0.71; P < 0.05), Q2/6 (r=0.77; P < 0.05) and Q4/6 (r=0.80; P < 0.05). The percentage of acid soluble mercury compounds did not exceed 0.5% of the HgT total content. The analysis did not indicate statistically significant relationships between the Hg concentration and the contents of other determined indicators like TOC or C/N. Mercury content in acid soluble fraction did not depend on the concentration of dissolved organic carbon determined in eluates of this fraction. By definition, in this fraction, mercury compounds bound with Fe/Mn oxides are being determined (Ram et al., 2009). In the acid environment, decomposition of carbonates and phosphates, with which mercury can combine, may also happen (Boszke et al., 2008). Percentage of ion exchangeable and water soluble mercury compounds in environmental samples (bottom sediments, soil) is small due to the fact that this element shows strong affinity to organic matter and those compounds are sparingly soluble in water (Kabata-Pendias, 2010; Lomonte et al., 2010; Moreno et al., 2005; Shoham-Frider et al., 2007; Xu et al., 2015; Zhang et al., 2009). In the acid-soluble fraction, the DOC percentage did not exceed, in the majority of samples, 2.0% of the total organic carbon content. Just like in the case of mercury compounds, explicit statistical analysis results were not obtained. High mercury affinity to organic matter containing sulfhydryl groups limits its sorption by argillaceous materials. Mercury compounds can also easily transform into volatile forms – elemental mercury or alkyl derivatives (Hutchison and Atwood, 2003; Kabata-Pendias, 2010; Moreno et al., 2005; Wang et al., 2012; Xu et al., 2015). Such processes can explain low mercury compounds content in the ion exchangeable fraction and in the acid soluble fraction. The concentration of mercury compounds in the base soluble fraction was influenced by the content of dissolved organic carbon present in that fraction. This was indicated by positive values of the correlation coefficients r=0.76 (P < 0.05). Changes of the Hg concentrations in the BS fraction were strongly positively correlated with the changes of the C/N ratio. This can prove that during the initial composting stage, in which organic matter decomposition proceeded, concentration of base soluble mercury compounds was higher than during the humification stage, i.e. during the formation of substances featuring humic compounds. Negative correlations with absorbance

decrease of the DOC percentage was noted. Samples of the composted sewage sludge taken on the 21st and 28th days of the process featured the lowest percentage of DOC of approximately 5.55%. As of the 42nd day, the DOC percentage was increasing again: the DOC percentage in the samples taken on the 182nd day was 7.94% of the TOC. The higher DOC percentage than in the oxidizable fraction was determined in the NaOH extracts before the 56th day of the process. In the final stage of composting, the DOC percentage in both fractions was at a similar level (Fig. 7). 3.3. Mobility factor Fig. 4 shows how the mobility factor changed during composting. The highest Mf values were determined for those biomass samples that were intended for composting (Mf=11.19) and for the samples taken from the bioreactor at the initial stage. During first four weeks of composting, the Mf value was within the 8.13–10.01 interval. During consecutive days of composting, the Mf values decreased, the lowest value of 4.54 determined for the samples taken from the bioreactor on the 91st day of the process. Then, insignificant increase of the mobile Hg forms concentrations was observed: the Mf value in the samples taken from the bioreactor on the 182nd day was 5.54. Low Mf values indicate that during composting, mercury occurs in the form that is strongly bound with the sample solid phase. Therefore, immobilisation of mercury compounds occurs during the composting process. 3.4. Statistical analysis Total mercury (HgT) content was strongly negatively correlated with the total organic carbon (TOC) concentration (r=−0.79; P < 0.05) and the C/N ratio (r=−0.92; P < 0.05). Due to the decomposition of the organic matter, mercury concentration in the composted biomass was increasing. The percentage of mercury compounds in compost samples occurring in ion exchangeable form did not exceed 0.4% of the total content of this element. Concentration of ion exchangeable compounds of mercury was negatively correlated with the DOC content in the EX fraction (r=−0.53; P < 0.05) and with the C/N value (r=−0.55; P < 0.05). As the organic matter content decreased, Hg concentration 401

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concentrations in the EX and AS fractions. Increase of dissolved organic carbon content in the BS and OX fractions took place as well. Those changes proceeded in the final composting stage on approximately the 133rd day.

ratios prove that the base soluble mercury compounds were formed during the first composting stage, i.e. during the course of organic matter decomposition – Q2/4 (r=−0.76; P < 0.05), Q2/6 (r=−0.73; P < 0.05), Q4/6 (r=−070; P < 0.05). During the final stage of composting, slight increase of the Hg base soluble compounds percentage occurred, which could be influenced by the formation of the humification products with high content of oxygen atoms and aromatic compounds. The DOC content in this fraction was positively correlated with the C/N and TOC content. Increase of concentrations of the mercury compounds present in the OX fraction was observed during the composting process. Percentage of mercury bound with organic matter varied during the entire composting cycle, from 50.09% to 64.55% of the total Hg content. Mercury content in the OX fraction was strongly negatively correlated with the total organic carbon concentration (r=−0.83; P < 0.05) and with the C/N value (r=−0.93; P < 0.05). Percentage of mercury compounds in the OX fraction was increasing with decomposition of organic matter during composting process. Determined correlation coefficients indicate strong, statistically significant relationship between the Hg compounds content in the oxidizable fraction and the DOC concentration in the OX (r=0.90; P < 0.05) as well as the BS fractions (r=−0.67; P < 0.05). Increase of the DOC percentage in the OX fraction was associated with the compost stabilisation stage and the formation of humic substances. Resynthesis of organic compounds, which were generated from the decomposition of organic matter, may have led to the generation of humic and fulvic acids. Those substances are capable of complexing mercury ions due to the presence of carbonyl, hydroxyl and thiol groups (Wallschläger et al., 1998; Zhang et al., 2009). Hg can probably make covalent bonds with thiol functional groups (Ravichandran, 2004). These compounds show limited bioavailability. However, the processes of microbiological or chemical methylation of mercury proceeding in soil can lead to the origination of mobile forms of this element (Kabata-Pendias, 2010). Similar conclusions were presented by Shoham-Frider et al. (2007). They concluded that in the samples rich in organic matter, e.g. sewage sludge and polluted bottom sediments mercury is strongly bound with sulphide ligands. The oxidizable fraction is obtained through the application of a mixture of 30% H2O2 and 0.02 M HNO3 at the temperature of 50 °C (Ram et al., 2009). In such conditions, total decomposition of organic matter does not occur. It appears from the research work performed by Bloom et al. that decomposition of sparingly soluble mercury compounds (HgS) occurs due to the action of aqua regia (Bloom et al., 2003).

4. Conclusions Sequential extraction of Hg indicates that from 50% to 64% of HgT is organic and sulphide-bound. Percentage of mercury in the BS fraction was, in the final composting stage, approximately 5% HgT. Percentage of ion-exchangeable and acid-soluble mercury compounds did not exceed 0.4% of the total content of this element. The study shows that the distribution of mercury compounds in particular fractions depends on the changes in the organic matter during composting process. Though the total Hg concentrations are high in the tested samples, but the partition chemistry data indicates that the Hg in sewage sludge and composts is not bioavailable. Ethical statement the authors declare that they have no conflict of interest with the research process which outcomes are presented in this article. Acknowledgements This research was financed by the Polish State Committee for Scientific Research within the project No. N N523 743840. References Albrecht, R., Le Petit, J., Terrom, G., Perissol, C., 2011. Comparison between UV spectroscopy and nirs to assess humification process during sewage sludge and green wastes co-composting. Bioresour. Technol. 102 (6), 4495–4500. AMAP/UNEP, 2013. Technical background report for the global mercury assessment 2013. Arctic Monitoring and Assessment Programme, Oslo, Norway/UNEP Chemicals Branch, Geneva, Switzerland. Amir, H., Hafidi, M., Lemee, L., Baily, J.R., Merlina, G., Kaemmeer, M., Revel, J.R., Ambles, A., 2006a. Structural characterization of fulvic acids, extracted from sewage sludge during composting, by thermochemolysis-gas chromatography-mass spectrometry. J. Anal. Appl. Pyrolysis 77, 149–158. Amir, H., Hafidi, M., Lemee, L., Merlina, G., Guiresse, M., Pinelli, E., Revel, J.C., Baily, J.R., Ambles, A., 2006b. Structural characterization of humic acids, extracted from sewage sludge during composting, by thermochemolysis-gas chromatography-mass spectrometry. Process Biochem. 41, 410–422. Balogh, S.J., Nollet, Y.H., 2008. Mercury mass balance at a wastewater treatment plant employing sludge incineration with offgas mercury control. Sci. Total Environ. 389, 125–131. Bernal, M.P., Alburquerque, J.A., Moral, R., 2009. Composting of animal manures and chemical criteria for compost maturity assessment. A review. Bioresour. Technol. 100, 5444–5453. Białobrzewski, I., Mikš-Krajnik, M., Dach, J., Markowski, M., Czekała, W., Głuchowska, K., 2015. Model of the sewage sludge-straw composting process integrating different heat generation capacities of mesophilic and thermophilic microorganisms. Waste Manag. 43, 72–83. Bień J.B., 2002. Sewage sludge. Theory and Practise. 1st ed. Wydawnictwo Politechniki Częstochowskiej, Częstochowa (in polish). Bloom, N.S., Preus, E., Katon, J., Hiltner, M., 2003. Selective extractions to assess the biogeochemically relevant fractionation of inorganic mercury in sediments and soil. Anal. Chim. Acta 479, 233–248. Boszke, L., Kowalski, A., Astel, A., Barański, A., Gworek, B., Siepak, J., 2008. Mercury mobility and bioavailability in soil from contaminated area. Environ. Geol. 55, 1075–1087. Cappon, C.J., 1987. Uptake and speciation of mercury and selenium in vegetable crops grown on compost-treated soil. Water Air Soil Pollut. 34 (4), 353–361. Confesor, R.B., Hamlett, J.M., Shannon, R.D., Graves, R.E., 2008. Potential pollutants from farm, food and yard waste compost at differing ages: Part I. Physical and chemical properties. Compost Sci. Util. 16 (4), 228–238. Dong, B., Liu, X., Dai, L., Dai, X., 2013. Changes of heavy metal speciation during highsolid anaerobic digestion of sewage sludge. Bioresour. Technol. 131, 152–158. Epstein, E., 1997. The Science of Composting. Technomic publishing Company Inc., Lancaster, PA. Fernandez-Martinez, R., Loredo, J., Ordonez, A., Rucandio, M.I., 2006. Physicochemical characterization and mercury speciation of particle-size soil fractions from an abandoned mining area in Mieres, Asturias (Spain). Environ. Pollut. 142, 217–226. Fernandez-Martínez, R., Rucandio, I., 2013. Assessment of a sequential extraction method to evaluate mercury mobility and geochemistry in solid environmental samples.

3.5. Transformations of the Hg compounds in the composting stage Biochemical transformations associated with mineralisation of organic matter and humification processes affected the distribution of mercury compounds in particular fractions. The percentage of basesoluble mercury compounds at the initial composting stage lasting up to approximately the 28th day was decreasing. At that time, concentration of ion-exchangeable mercury compounds was also decreasing. The changes of mercury concentrations in the base soluble fraction depended on the rate of transformation of organic matter. Mercury ions generated due to the biodegradation of organic compounds could be bound by sulphide ligands or form sparingly soluble sulphides (Ravichandran, 2004; Zhang et al., 2009). They could also occur in the forms of other compounds insoluble in NaOH. During the humification stage, concentrations of mercury compounds occurring in the base soluble fraction and in the oxidiziable fraction were increasing. This process started on approximately the 56th day and lasted until approximately the 112nd day of the composting process. At that time, content increase of dissolved organic carbon in the base-soluble fraction also proceeded. The organic matter stabilisation stage indicated increase of the Hg percentage in the OX and BS fractions as well as decrease of 402

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