Biodegradable polymers as solid substrate and
biofilm carrier for denitrification in recirculated
aquaculture systems
A. Boley *, W.-R. Mu¨ller, G. Haider
Uni6ersita¨t Stuttgart,Institut fu¨r Siedlungswasserbau,Wassergu¨te- und Abfallwirtschaft, Arbeitsbereich Biologie,Bandta¨le 2,D-70569 Stuttgart, Germany
Abstract
A simple process for nitrate removal is proposed for its application in aquaculture. Biodegradable polymer pellets are acting as solid substrate and biofilm carrier for denitrifica-tion. Laboratory experiments with conventional aquaria and fish were used to examine the feasibility and a first evaluation of the process performance in a recirculated aquaculture system. All over the test-period the fish were in a good condition. Nitrate concentrations in the aquaria with treatment were low compared to the untreated reference system. A further advantage was the stability of the pH in the units with denitrification whereas pH of the untreated water decreased due to nitrification. © 2000 Elsevier Science B.V. All rights reserved.
Keywords:Water treatment; Recirculating systems in aquaculture; Denitrification; Biodegradable poly-mers; Solid substrates
www.elsevier.nl/locate/aqua-online
1. Introduction
In aquaculture systems nitrate removal is a problem which has not always found satisfactory solutions in practice. Modern technology of water treatment in recircu-lating systems consists of solid waste removal, carbon-removal and nitrification, pH and CO2control (Fig. 1). Consumption of energy and water in those systems can be lowered if the nitrate produced in the aerobic biofilter unit is reduced by a denitrification step. This diminishes the fresh water addition and the amount and impact of the wastewater.
* Corresponding author. Tel.: +49-711-6855441; fax:+49-711-6853729.
E-mail address:[email protected] (A. Boley)
76 A.Boley et al./Aquacultural Engineering22 (2000) 75 – 85
Denitrification is defined as the biological nitrate reduction sequence NO3
−
NO2
−N
2ON2. We restrict the discussion to the heterotrophic biological process where organisms gain energy and carbon from organic compounds. A conventional technique is to add an organic carbon source (e.g. ethanol, acetic acid) to a denitrification reactor (Frick and Richard, 1985; Sto¨ver and Roennefahrt, 1990). The disadvantage of this treatment process is the need of a close, rather sophisti-cated and costly process control, the risk of overdosing and a deepened knowledge about the operation of this biological system.
In contrast to conventional treatment units, denitrification with biodegradable polymers presented here is a simple process. Microorganisms use the biopolymer in form of pellets as biofilm carrier and simultaneously as water insoluble carbon source for denitrification, which is accessible only by enzymatic attack (Mu¨ller et al., 1992; Wurmthaler, 1995).
The scheme in Fig. 2 elucidates the difference between conventional denitrifica-tion and the new process presented here. In convendenitrifica-tional denitrificadenitrifica-tion with a fixed bed reactor a biofilm will grow on the inert carrier and denitrification takes place whenever the water contains NO3
−, soluble organic substrate and trace elements.
End-products are N2, H2O, CO2 and biomass. The new system with biodegradable
Fig. 1. Scheme of a modern recirculated aquaculture system.
polymers does not require an external dosing of soluble organic substrate as the polymer itself acts as biofilm carrier and organic carbon source.
Heterotrophic denitrification positively influences the pH of the water. If proteins are metabolized by fish, the end-products of respiration after hydrolysis to amino acids (e.g. glycine) are NH4+ and HCO3−, which are excreted via gills (Eq. (1);
Forster and Goldstein, 1969):
NH2CH2COOH+1.5 O2NH4++HCO3−+CO2 (1)
The nitrification equation with biomass formation (Wheaton et al., 1994, Eq. (2)) indicates the production of protons (catalyzed by enzymes of, e.g. Nitrosomonas and Nitrobacter species):
Decreasing pH values have to be coped with by adding, e.g. NaHCO3
−.
The use of a biodegradable polymer as organic carbon substrate, e.g. PHB, leads to biomass, carbon dioxide and simultaneous reduction of nitrate to elementary nitrogen. With a yield coefficient of 0.45 g biomass/g PHB assumed (Heinemann, 1995), the summarized denitrification equation including biomass formation can be given as:
0.494 C4H6O2+NO3−
0.130 CO2+HCO3−+0.415 N2+0.169 C5H7O2N+0.390 H2O (3)
The summary equation (nitrification and denitrification) results in:
1.021 NH4+
+1.021 HCO3−
+1.895 O2+0.494 C4H6O2
3.369 H2O+2.044 CO2+0.415 N2+0.190 C5H7O2N (4)
CO2 produced can be stripped by aeration. If all the nitrate produced is denitrified, the pH remains constant.
2. Materials and methods
The examination of solid substrates in form of biodegradable polymers for denitrification purposes in aquaculture has been carried out in simple laboratory-scale test systems (Fig. 3). We used four commercially available 100-l aquaria operated in parallel. Each aquarium was equipped with an aerobic biofilter-unit filled with SIPORAX-packing (Schott, 0.75 l). The total volume (aquarium+bi-ofilter) was 82.5 l. It was filled with tap water and the temperature was adjusted to :25°C. The tank was illuminated 10 h/day. Each aquarium contained 14 fish
(Leuciscus idus) with an initial total biomass of 80 g. The feeding rate of one tank
78 A.Boley et al./Aquacultural Engineering22 (2000) 75 – 85
Fig. 3. Aquarium system for testing denitrification. Period 1: Carbon removal+nitrification only; period 2: Carbon removal+nitrification+denitrification.
Table 1
Material characterization
Bionolle
Short name PHB PCL
Bionolle c6010 BIOPOL D400 GN
Trade name, type TONE P 787
[C6H8O4]n
Chemical formula [C4H6O2]n [C6H10O2]n
294 264
Mass (g) per unit 265
0.52 0.39 0.46
Total surface (m2)
Showa Denko Union Carbide
Manufacturer Monsanto
Before starting the experiments the biofilters for carbon removal and nitrification were subjected to a conditioning phase with a medium containing ammonia to secure a good nitrification performance.
The first period of the experiments was confined to nitrification via biofilter. In period 2 the denitrification units were connected. They consisted of small fixed bed reactors (‘denireactors’) with a volume of 0.375 l. As subsequent aerobic treatment, small aerated fixed bed units (volume 420 ml), with SIPORAX-packing, were installed for polishing to avoid possibly occurring byproducts (e.g. NO2−). Different
biodegradable polymers pellets (Table 1) were used as packing for the denireactors and, as reference, one was operated with glass beads. The polymers to be tested were filled into the closed denireactors without pretreatment and enclosed in the system by a plastic foam bottom and a cover above.
Relevant water-parameters in the tanks were examined. Weekly measurements of temperature, pH, oxygen, conductivity, NH4
+, NO
Occasionally the dissolved organic carbon (DOC) concentration was determined. The water volume added for compensation of evaporation was taken into account. A simple model for evaluating the performance of the denireactors with different polymer packing has been used after the start-up and beyond the lag-time periods of these units. The influence of the oxygen has not yet been taken into account as well as NO2− was not included into the model.
The NO3−concentration in the aquarium, considered as complete mixed tank, as
function of time can be described as follows:
dcA/dt=(QD*(cE−cA)+mNO3)/VA (5)
The lowest concentration in the aquariumcA0 to be achieved under steady state conditions, i.e. with an effluent concentration of the denireactor cE=0 is deter-mined by the relation:
cA0=mNO3/QD (6)
wherecAis thecA0NO3
− conc., aquarium tank (mg/l N-NO
3
−);c
Eis the NO3
−conc.,
effluent denireactor (mg/l N-NO3−);m
NO3is the daily production of NO3− in system (mg/day N-NO3−);Q
Dis the recirculation rate=throughput denireactor (l/h); and VA is the water volume of aquarium tank (l).
The overall volumetric denitrification performancerDVin mg/(Lh) N-NO3
− of a
denireactor is given by Eq. (7).
rDV=QD*(cE−cA)/VD (7)
rDVis the overall volumetric denitrification rate of a denireactor (mg/(lh) N-NO3−);
andVD is the denireactor volume (l).
3. Results
Due to a preconditioning of the biofilters, ammonium and nitrite concentrations were low during the whole test-periods 1 and 2. NH4+ did not exceed 0.1 mg/l
(N-NH4+), NO2−was below 0.05 mg/l N-NO2−after the first day. Temperature was
stable in a range of 25.1 – 26.1°C. DOC values increased slowly during the tests, beginning with 3 – 4 mg/l they did not exceed 5 – 7 mg/l at the end of the tests.
In period 1 NO3− concentrations increased in all four aquaria in a very similar
way (Fig. 4). In this period and from the reference aquarium system, the daily production of nitrate could be calculated to 56.1 (95) mg/day N-NO3−.
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Table 2
Estimated maximal denitrification velocities of tested materials
Specific surface Temp. (°C) Concentration range 5–40 mg/l N-NO3−
Solid substrate Flowrate (l/h)
(m2/l)
Surface related rates (mg N-NO3−/(m2h)) Volumetric rates (mg N-NO3−/
(lh))
0.4–0.6
1.49 20–25
PHB 7–41 5–28
0.87 20–25 0.2–0.3 20–160
PCL 21–166
0.3–0.6 1.5–10; 12–77 1.3–9; 10.5–67 -Bionolle, period 1 1.22 20–25
As Fig. 4 shows the theoretical concentration limits (about 5 mg/l, Eq. (6)) have approximately been achieved with PCL and Bionolle at the end of test. Nitrate concentrations in the effluent of these denireactors were below the detection limit (0.23 mg/l N-NO3
−). This confirmed our assumptions.
In contrast to these results the aquarium with the PHB denireactor reached the equilibrium already at a concentration of 18 mg/l N-NO3−. This decrease of
performance (=decrease of denitrification velocity) can probably be explained by clogging and short-circuiting of the denireactor due to excess biomass production, which has been observed after the end of period 2.
As the acid neutralizing capacity of the tap-water was low (ANC=1 mmol/l), pH values decreased with time, due to nitrification (Fig. 5). To prevent extensive decrease of pH, it was adjusted twice with NaHCO3, which was added to the reference aquarium (packing with glass beads) at days 71 and 100. For the aquarium with the PHB denireactor NaHCO3 addition was not necessary because at day 71 denitrification had already started. The start of denitrification
immedi-Fig. 4. Nitrate concentrations in testsystems. Temperature: 25 – 26°C.
Fig. 5. pH in testsystems. Temperature 25 – 26°C. Arrows indicate pH adjustment with NaHCO3. (After
82
Denitrification velocities in fixed bed reactors with different substrates
Temp. (°C) Volumetric rate (mg
Carrier-material Spec. surface Substrate Surface related rate (mg N- Type of water and in-stallation Burned clay 1.3 Acetic Acid
0.9 Ethanol 12–13 54–66 Drinking water plantc
Burned clay 49–59
11
1.55 PHB 10 16 Tap water,
labora-PHB
1.2 PCL 15 10 Tap water,
labora-PCL 13
tory-scalee
Fluidized bed, aqua-(d=0.3–0.9 mm)
Sand Dissolved organic 22.5–27 36
substrates culture systemf
Table 4
Estimated costs of substrates for nitrate removal
Price of substrate
Substrate Consumption of substrate (kg Costs of denitrification (€/kg substrate) substrate/kg N-NO−3) (€/kg N-NO−3)
Bionollec6010 Commercially not available (C6H4O2)n
ately may lead to an increase of pH. For the PCL and Bionolle denireactor NaHCO3 was also added at day 71, because denitrification had not yet started. Later pH increased too, therefore an adjustment was no more required. These results are compatible with Eq. (4).
After both test-periods the fish were in a good condition and no fish died. They almost doubled their initial body weight all together up to 145 g (95%) per aquarium.
4. Discussion
Denitrification systems are not yet common practice in aquaculture and until now they were mostly installed for research purposes. The reason is that toxicity of nitrate is low, compared with nitrite and ammonia.
A comparison of the polymer based denitrification presented here with conven-tional denitrification processes is shown in Table 3. The volumetric and surface related denitrification rates with PHB and PCL as substrates are lower than the respective rates with methanol and ethanol. However the same order of magnitude as with acetic acid as substrate could be observed.
84 A.Boley et al./Aquacultural Engineering22 (2000) 75 – 85
5. Conclusions
The denitrification process based on the use of solid substrates (biodegradable polymers) can not yet compete in its performance with the classical treatment units for biological nitrate removal with liquid substrates. Preliminary deliberations for this new denitrification process in aquaculture suggest that this is not a low-cost process at present. A cost-benefit analysis could not yet be carried out as data close to reality are lacking. However when extrapolating these laboratory-scale results and weighing the advantages, which are the user-friendly simplicity and safety of this process in relation to the disadvantages as the high costs of the solid substrates, we remain optimistic. A positive expectation is: reduction of clean water require-ment, reduction of waste water production, reduction of energy consumption which will contribute to favor an application in the future.
Acknowledgements
This work was supported by Deutsche Bundesstiftung Umwelt and European Communities, INCO-DC.
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