EXCHANGE OF PAH BETWEEN AQUEOUS PHASE AND BIOFILMS OF SOIL BACTERIA
Daniel Wickea, Uta Böckelmannb, Thorsten Reemtsmac
a Dept. of Water Quality Control, Technical University of Berlin, KF 4, Strasse des 17 Juni 135, 10623
Berlin, Germany, email daniel.wicke@tu-berlin.de
b Dept. of Environmental Microbiology, Technical University of Berlin, FR1-2, Franklinstrasse 29,
10587 Berlin, Germany, email uta.boeckelmann@tu-berlin.de
c Dept. of Water Quality Control, Technical University of Berlin, KF 4, Strasse des 17 Juni 135, 10623
Berlin, Germany, email thorsten.reemtsma@tu-berlin.de
Introduction
Urban soils accumulate considerable amounts of organic contaminants, such as PAHs and PCBs. For
example, our measurements of samples from a public park (Tiergarten) in the centre of Berlin revealed
PAH concentrations of more than 10 µg/g dry soil (Chrysene and Benzo[a]-anthracene). Identifying the
mechanisms responsible for controlling the sorption and desorption behaviour of organic contaminants
in soils is critical to understand and to predict the fate and transport of these compounds in the
subsurface environment.
Biofilms – forming the boundary layer between soil particles and the surrounding water, are suspected
to play an important role in mass transfer processes between these two phases. Important processes
are sorption / desorption to biofilms as well as mass transport and transformation reactions within
biofilms. Especially quantitative aspects of the exchange processes (diffusion coefficients, sorption
rates) are of particular interest. Due to the complexity and heterogeneity of the soil system, it is difficult
to study biofilm sorption directly in soils. As a consequence, we developed an experimental setup that
allows us to study these processes under controlled laboratory conditions with biofilms attached to a
solid support in small reactors.
Experimental approach
In order to investigate thermodynamics and kinetic aspects of sorption and desorption processes of
organic contaminants (such as PAH) to biofilms, the concept of a differential loop reactor has been
chosen. High flow rates are used to ensure that diffusion processes occurring outside of the biofilm
layer are minimized, and to achieve a differential change of contaminant concentration in the reactor.
The reactors (empty volume of approximately 80 mL) were filled with glass beads with a diameter of 3
mm as adhesion material for the selected microorganisms (surface area of around 850 cm²). The
sterilized reactors were inoculated with different bacterial strains, isolated from soil samples.
Appropriate strains of soil microorganisms were selected that meet the requirements of the
experimental conditions. Especially the formation of firmly attached biofilms that resist high flow rates
was required. With the selected soil bacteria, sufficient biofilm mass has been established within 7
days of growth using TSB (Tryptic Soy Broth) medium.
After biofilm establishment, reactors were immediately used for sorption experiments. Due to the
hydrophobicity of the target analytes (phenanthrene, fluoranthene, pyrene), all components of the
reactor system had to be carefully selected to minimize sorption (glass, teflon). Samples were taken
up to every minute to be able to observe fast sorption rates. Samples (800 µl) were filled in vials
containing 200 µl acetonitrile to prevent sorption of the selected PAH to the glass walls, and measured
directly using high-pressure liquid chromatography with programmed fluorescence detection.
With this experimental setup, the influence of external effects on the exchange processes, such as
drying of the biofilms, temperature, the effects of
salts and the addition of soil humic acids to the
Experimental results - pyrene
solution phase has also been investigated.
1,2
1,0
0,8
c/c 0
Results
Figure 1 shows exemplarily the normalized
concentrations of one of the three selected PAH
(pyrene) over time. It can be seen that
equilibrium was almost established within two
hours.
Regarding thermodynamics of the sorption
process, distribution coefficients have been
calculated and related to the organic carbon
content of the biofilm. An average carbon content
of approximately 50 % for bacterial cells has been
log k ow = 4,9
0,6
0,4
0,2
0,0
0
20
40
60
80
100
120
140
time [min]
Figure 1: Sorption of pyrene to a biofilm formed
by strain 30A (Rhizobiaceae)
c/co
used for calculation. The derived log KOC values Table 1: log KOC and Ds values for sorption/ diffuare listed in Table 1. KOC-values in the literature sion of selected PAH in biofilm of a soil bacteria
for sorption of PAH to other organic sorbents are
Ds [10-6 cm²/s]
compound
log KOC
in the same range. For example, Jonassen et al.
(Jonassen 2003) are reporting log KOC values for
0,3
Phenanthrene
4,1
phenanthrene, anthracene, and pyrene of 4.2,
0,2
Fluoranthene
4,3
4.3, and 5.0, respectively using Aldrich humic
acid as organic carbon.
Pyrene
4,4
0,2
Regarding the kinetics of the sorption process it
can first be seen that sorption of PAH to biofilm is
a comparatively fast process. Equilibrium concentration is mostly reached within the 120 minutes of
the experiment. Furthermore, most profiles are showing a little discontinuity after 10 to 15 minutes of
the experiment. This could indicate a 2-stage process with a rapid sorption to the unloaded biomass
within the first 10 to 15 minutes, and a second phase of slower diffusion when the concentration at the
biofilm surface is raised to a certain level. Slower diffusion into the biofilm could then be responsible
for further sorption until equilibrium after two hours. Occurrence of diffusion in the boundary layer (if
not totally repressed) would
also influence the kinetics of
Pyrene - tw o-stage sorption
the sorption process. For the
determination of intraparticle/
1,1
measured
calculated
intrabiofilm diffusion coeffi1,0
cients, the KIN 1.3 software by
0,9
E. Worch has been used
0,8
(Worch 2001). With this soft0,7
ware experimentally obtained
0,6
curves can be simulated using
0,5
the linear driving force (LDF)
0,4
model. An example of a
0
20
40
60
80
100
120
modelled curve assuming a
tim e [m in]
two-stage sorption process is
Figure 2: Modelled 2-stage sorption curve of pyrene to soil biofilm shown in Figure 2. Resulting
values for the diffusion coefficient (Ds) are also listed in Table 1. In comparison with water (D s ~ 6∙10-6 cm²/s) the diffusion of the
considered PAH in biofilms of soil bacteria is approximately one order of magnitude slower. Drying of
the biofilm prior the sorption experiment as well as incorporation of soil humic acids (purchased from
IHSS) did not significantly change the extent and kinetics of the sorption process.
It could be shown that the described experimental system and model is useful to investigate the
kinetics and extent of sorption of hydrophobic pollutants such as PAH to biofilm of soil bacteria. This
data can be used for modelling the fate and transport of hydrophobic compounds in soil systems.
Perspectives
Further investigations will include the influence of biofilm hydrophobicity on the sorption behaviour of
PAH. Desorption of PAH from biofilms will also be investigated as the second important aspect of the
exchange process. In addition, other soil bacteria will be employed, especially strains that are able to
degrade certain PAH (e.g. phenanthrene-dregrading strains) to study the formation of metabolites that
might be more soluble and therefore more mobile than the PAH itself.
Keywords: sorption kinetics, biofilm, urban soils, PAH
Jonassen, K.; Nielsen, T.; Hansen, P. E.: The application of high-performance liquid chromatography
humic acid columns in determination of KOC of polycyclic aromatic compounds. Environmental
Toxicology and Chemistry (2003), 22, 741-745
Worch, E: KIN 1.3. Software, 2001. Program for determination of surface diffusion coefficients from
experimental data, Department of Water Chemistry, Technical University of Dresden,
Germany