Methodology and experimental approach

1. Production of isotopically labelled (13C & 15N) sewage sludge and biochar for incubation and tracing experiments

Labelled (13C and 15N) and un-labelled sewage sludges were obtained at CENTA station. The labelling of the sludge was carried out by incubating the water sludge in a biomass reactor tank during 24-48 h, which were previously conditioned and feed with 13C-glucose and 15N-NO3.

The sludge produced was used as feedstock to produce hydrochars and biochars under different conditions (different temperature and pyrolysis time) at ATB-Potsdam in 2013 and 2014. Biochars (at 600ºC) and hydrochars (200ºC and 260ºC; 0,5, 1, 2 and 3 hours). The success of the labelling of the chars was assessed by elemental (C, N) and isotopic analysis (δ13C and δ15N). The abundance of 13C and 15N of the chars was aprox. two orders of magnitude greater than the natural abundance of 13C and 15N. Nevertheless, due to the slow production of 13C-15N labelled sludge, which are strictly needed as feedstock, the amount of labelled biochars is limited. Thus, experiments with labelled and un-labelled biochars are being carried out at the same time in order to achieve the goals of Biocharisma project.

During the development of biocharisma project further biochar samples from different feedstock and pyrolytic process have been analysed and tested. In this sense, it should be remarked the participation of Prof. Knicker and JMR in the European network devoted to the sustainable use of biochar (TD1107 Biochar COST action), this network facilitated the availability of 3 additional biochar samples, produced from wood, paper sludge and sewage sludge respectively by fast pyrolysis at 620 ºC. They were incorporated to the characterization and experiments of biocharisma project. In addition, Torres wine-yard Company sent 5 kg of a biochar sample produced by the traditional kiln methodology from old wineyard wood which has been also used in this project. In a second step, biochars were produced from labelled and un-labelled organic residues in a small scale pyrolysis reactor at ATB-Potsdam (Germany). Hydorchars were produced by hydrothermal carbonization HTC at 200°C, biochars were produced by dry pyrolysis at 600°C.


2. Characterization of biochars produced from sewage sludge and vegetation residues to be applied as soil amendment

-Bulk characterization

All the biochars and hydrochars produced as well as the biochars sent by COST action network and companies were characterized. Characterization was performed by elemental analysis (C,H,N;S), and by the determination of chemical and physical properties such as, pH, electrical conductivity (EC), ash content, specific surface area and water holding capacity (WHC). Fourier-Transform Infrared Spectroscopy (FT-IR) and 13C solid-state NMR techniques were also applied to elucidate the molecular structure and main chemical groups, whereas field emission scanning electron microscopy (FE-SEM) was used to investigate the surface topography and chemical composition of bio/hydrochars.

Biochars from wood and paper sludge revealed comparable elemental composition, pH, water holding capacity and ash content. The H/C and O/C atomic ratios suggested high aromaticity of all biochars, which was confirmed by 13C solid-state NMR spectroscopy. The FT-IR spectra confirmed the aromaticity of all the biochars as well as several specific differences in their composition. The SEM-EDS distinguished compositional and structural differences of the studied biochars such as macropores on the surface of wood-pyrolysis biochars, collapsed structures in paper sludge biochar, high amount of mineral deposits (rich in Al, Si, Ca and Fe) and organic phases in sewage sludge biochars and vessel structures for kiln wood biochar. Hydrochars from sludge presented a much lower C content (usually below 20%) and lower aromaticity than pyrochars. The latter was confirmed by the high H/C ratio and by the spectra of 13C NMR spectroscopy. The organic fraction of hydrochars was enriched in alkyl compounds. In addition, they were abundant in minerals and their pH values were closer to 7, which pointed to their appropriateness as soil ameliorants instead of C stabilization.

-Solid state 13C-NMR spectroscopy

Solid-state NMR, in particular solid-state 13C and 15N NMR is a well-recognized and broadly used analytical tool for the study of formation and stabilization of OM in soils and sediments (Wilson, 1987). Biocharisma takes advantage of the deep knowledge of Prof. Knicker in the field of NMR spectroscopy. The necessary equipment is available through by the Bruker Avance III instrument of the host research group.

Solid state NMR Spect_Bruker

-Analytical Pyrolysis

Pyrolysis-GC/MS have been performed in a double shot pyrolyzer (model 2020, Frontier Laboratories) directly connected to an Agilent 6890 GC-MS system, available at the MOSS group.


3. C-sequestration potential and fate of biochar in typical Mediterranean agricultural soils

The input of charred residues from biochar is expected to change the quantity and quality of soil OM but also its role as long-term C sink within the global C cycle. In order to achieve this objective, field and incubation based experiments will be studied by using different methodologies:

Determination of the respiration (Respicond)

The humification efficiency and the C-sequestration potential of biochars are a result of complex interactions between substrate, microbial population and environmental factors and thus differ for different treatments. Monitoring the potential CO2 release from the 13C-labelled biochars and of the respective amended soils in laboratory incubation experiments using a Respicond apparatus, will enhance to the understanding of the potential of the product to contribute or to decrease CO2 emission and thus contribute to C sequestration. The Respicond apparatus is a sophisticated computerized respirometer available at the HRI based on the principle of using the change of conductivity of an electrolyte (KOH) for detecting CO2. The obtained data will be used as possible means of quality control of the developed products.

Imagen110 mg of dry sample material of biochar and selected soils will be incubated in a Respicond Apparatus IV at 60% of the water holding capacity after inoculation with a microbial suspension. The CO2-production will be monitored every 2h for a time frame of two months. In addition, CO2 evolved in the experimental field chambers will be determined by the alkali trapping method (KOH). The obtained CO2 accumulation curve will be fitted using a two exponential decay model which allows the calculation of the mean residence times of the respective material.

Field experimental plots

The field experiments are being carried out on experimental micro-plots installed in the farm ‘La Hampa’ of the CSIC. This experimental farm of 45 ha is located 13 km from Seville city (37°16′59″ N, 06°04′03″ W).

4. Risk assessment of release of Polycyclic aromatic hydrocarbon (PAHs) due to biochar addition and strategies for their avoidance

The procedure applied for assessing the Polycyclic aromatic hydrocarbons was based on that reported by Bucheli et al., (2004). Nevertheless, the analytical procedure for the extraction and assessment of PAHs in biochar amended soils have been refined for this issue. Results of those analyses have been already submitted to an international scientific journal. The obtained concentrations were compared to those given as a threshold by European laws (CEN 15527: 2008 for biochar and ISO 18287: 2006 for water samples.

-Green house pot experiments and field experiments

Green house experiments

Pot experiments were carried out at green house facilities at IRNAS-CSIC. Germination rates and soil fertility of Lolium perenne at a Mediterranean calcic Cambisol were tested under different biochar doses (0, 10, 20 and 40 t/ha). Changes were related to the physical and chemical properties of the biochars tested. Biochar amendment improved germination rates and soil fertility (excepting for kiln wood biochar), and had no negative pH impact on the already alkaline soil. Application of biochars and hydrochars from sewage sludge, the richest in minerals and nitrogen, resulted in the highest soil fertility. In this case, increase of the dose went along with an enhancement of plant production. Considering costs due to production and transport of biochar, for all used chars with the exception of sludge hidrochars, the application of 10 t/ha turned out as the most efficient for the crop and soil used in the present incubation.


Field experiments

The field studies are being performed at the experimental station “La Hampa” from IRNAS-CSIC, located in the Guadalquivir river valley (SW Spain; 37º 21.32’ N, 6º 4.07’ W), Seville.

The first field experiment was carried out from January 2013 to August 2013 by seeding with Helianthus annuus the same calcic Cambisol also used as matrix for the pot experiment. Soil was amended with doses equivalent to 0 (control), 1.5 and 10 t ha-1 of five biochars, making a total of 12 different treatments. Soil properties and composition were monitored during the growing time including elemental composition, pH, water holding capacity and soil microbial biomass. After 7 months of growth, sunflower plants were harvested. Plant height, chlorophyll content and sunflower seeds production were recorded. Results of this field experiment are still being processed. Nevertheless, preliminary results indicated that addition of biochar did not alter negatively physical properties (pH, EC) or composition of this alkaline soil, on the contrary biochar addition caused a slight improvement of the WHC and soil porosity. Those changes produced a faster development of plant shoots. However, at the end of the experiment, biochar amendment caused no significant increase on the agronomic production for any of the tested biochars. From these preliminary results, we conclude that biochar amendment improved physical-surficial characteristics of the calcic Cambisol from an agronomic point of view, but under the typical Mediterranean climate those changes seem to leave the harvested seed yields unaffected.

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