173
Analysis of formation and characterization of hydrogels originated from structural extracellular polymeric substances (sEPS) extracted from aerobic granular sludge (AGS)
R. Campo*, B. Pagliaccia*, E. Carretti**, D. Berti**, S. Caffaz***, C. Lubello*, T. Lotti*
* Università degli Studi di Firenze, Dipartimento di Ingegneria Civile e Ambientale (DICEA), Via di S.Marta 3, 50139, Firenze.
** Università degli Studi di Firenze, Dipartimento di Chimica “Ugo Schiff”, Via della Lastruccia, 13 - 50019 Sesto Fiorentino (FI).
*** Publiacqua s.p.a., Impianto di depurazione San Colombano, Via di Romania snc, 50055, Lastra a Signa (FI).
Keywords: sEPS; hydrogel; resource recovery; rheological study; water binding capacity Summary of key findings
This study reports on hydrogel formation and characterization from structural extracellular polymeric substances (sEPS) extracted from aerobic granular sludge (AGS). The following key-findings can be drawn: i) hydrogel can be formed starting from 3.3%wt sEPS on; ii) hydrogels maintain a constant molar ratio between Ca and C with the increase of the %wt sEPS; iii) the thermal behaviour of sEPS shows that the cross-linking reaction mainly involves the polysaccharidic fraction of biopolymers; iv) water- binding capacity of sEPS was confirmed by the increase of bound water fraction of hydrogel; v) rheological results highlighted a discontinuity of the solid-like mechanical properties of hydrogel at increasing sEPS%wt at approximately 5%wt. These findings reveal the peculiar physico-chemical characteristics of sEPS which nowadays are increasingly gaining interest in the field of resource recovery.
Background and relevance
AGS is a promising technology for wastewater treatment able to simultaneously remove carbon (C), nitrogen (N) and phosphorus (P). Recently, the possibility to extract sEPS from AGS having hydrogel formation abilities, has opened new scenarios on biopolymers recovery from AGS (Felz et al., 2016) and it occupies a prominent position in the general field of resources recovery (Kehrein et al., 2020). In this context, the aim and the relevance of this study are to investigate the hydrogel formation capability upon cross-linking with calcium for a wide range of concentration of acqueous sEPS solutions (from 2%wt up to about 10%wt) and to characterize the hydrogels in terms of elemental composition, thermogravimetric behaviour, water-binding capacity and reological properties. Among the sEPS features, the water-binding capacity can be applied in agriculture to increase water retention of soils (Milani et al., 2017), while the mechanical properties characterization is a pre-requisite for the evaluation of potential applications in several industrial sectors where biopolymers-based hydrogels are getting increasing attention due to their unique properties such as high water content, softness, flexibility and biocompatibility.
Results and Discussion
Structural EPS (sEPS) were extracted according to Felz et al. (2016), while hydrogels formation was performed by diffusion of Ca2+ from a 2.5%wt CaCl2 aqueous cross-linking solution into the sEPS matrix through a dyalisis membrane (3.5 kDa MWCO). Nine sEPS solutions at increasing biopolymer concentration (~2-10%wt) were prepared and placed in a 0.95 mL cylinder (Height/Diameter = 1.7) sealed at the bottom and top by the dyalisis membrane. Each cylinder was then put into a 250 mL becher filled with 200 mL of 2.5%wt CaCl2 solution for 24h. Fig.1 shows a conceptual diagram phase ofhydrogel formation from different sEPS concentrations. Experimental results revealed that hydrogel formation was possible starting from a 3.3%wt as gVSsEPS/gWet-WeightsEPS, upon controlled cross- linking with Ca2+. A yellow-brown bulk coloration was observed with the increase of sEPS
174
concentration, likely due to humic-fulvic acids release from hydrogel. This experimental evidence was supported by a sulphur decrease in the hydrogel (from 1.39 of sEPS to 0.89%wt of hydrogel dried samples) (Morra et al., 1997). Table 1 reports the sEPS characterization and elemental composition of both sEPS and the hydrogel. sEPS yield extraction was 0.23 gVSsEPS/gVSAGS, similar to literature result (Lin et al., 2010; Felz et al., 2016). Proteins (PN) of sEPS were determined with the BCA assay (Smith et al., 1985) whereas polysaccharides (PS) of sEPS were determined with the anthrone sulphuric acid method (Dreywood, 1946) using both d-glucose and sodium alginate as standards. PSGlu analysed with d-glucose as standard was 128.5 mgPSGlu /gVSsEPS, similar to literature results, whereas both PSAlg and PNBSA were noticeable higher than literature data (942.7 vs. 486.2 mgPSAlg/gVSsEPS by Lin et al. 2013, and 617.8 vs. 381 mgPNBSA/gVSsEPS by Felz et al. 2019, respectively). PSAlg was 7.3 times greater than PSGlu, thus suggesting an alginate-like composition of sEPS from AGS as reported by (Lin et al., 2010).
The C-H-N elemental composition and P content in sEPS and hydrogel were measured according to a previous study of the same group (Lotti et al., 2019). The Ca and S in sEPS and hydrogel were measured by ICP-OES (PerkinElmer OPTIMA 2000™ ICP). Comparative elemental analysis of sEPS and hydrogel revealed an almost constant nitrogen molar ratio (0.116 vs. 0.110 mol N/C-mol) and a value close to literature data (0.139 mol N/C-mol by Felz, 2019). An important Ca/C molar ratio increase from sEPS to hydrogel composition (0.003 vs. 0.071 mol Ca/C-mol) confirmed the inclusion of Ca2+ ions inside the three-dimensional network of sEPS matrix during the cross-linking reaction (hydrogel formation). Interestingly, the Ca/C molar ratio holds constant for all hydrogels formed independently from the increasing %wt sEPS. Thermogravimetric (TG) and differential thermogravimetric (DTG) analysis of dried sEPS and hydrogel were performed with a thermogravimetric analyser (Q5000 TA Instruments) heating the samples from room temperature to 800°C at 10°C/min under a nitrogen flow rate of 100 ml·min-1. TG and DTG curves of sEPS (Fig.1.2 a) revealed three main stages during samples thermal decomposition: i) dehydration referred to the elimination of residual moisture below 200°C that could not be removed during drying in oven at 105°C for 48 hours (weight loss next to 5.6%wt); ii) volatilization of the organic matter (about 200-535°C); iii) decomposition of the mineral fraction contained in sEPS. From the deconvolution of the DTG curve of sEPS (Fig.1.2 b) it is possible to identify at least three peaks (Bach and Chen, 2017): i) a first peak close to 270°C likely due to low thermal resistant proteins (PN I); ii) a second peak close to 290°C likely due to high thermal resistant polysaccharides (PS) such as sodium alginate (Li et al., 2019); iii) a third peak close to 300°C likely due to high thermal resistant proteins (PN II). Also by observing the deconvolution of the DTG curve of hydrogel in the temperature range 200-500°C (Fig.1.2 b), it is possible to note at least three distinct peaks : i) a first peak close to 270°C that is attributable to the low thermal resistant proteins (PN I) that are not involved in the cross-linking reaction, similar and close to the first peak in the DTG curve of sEPS sample; ii) a second peak close to 300°C that might be attributed to the high thermal resistant proteins (PN II) that are not involved in the cross-linking reaction. This last peak is similar and close to the third peak in the DTG curve of sEPS sample. Considering this experimental evidence, it is reasonable to assert that both PN I and PN II do not undergo substantial structural modifications during the cross- linking reaction; iii) a third peak close to 350°C, which is shifted of almost 60°C with respect to the second peak of sEPS DTG curve, relative to the high thermal resistant polysaccharides like alginates.
This shift is likely due to the higher energy needed to thermally decompose the polysaccharides that are strongly cross-linked with Ca2+ in the three-dimensional network of the hydrogel. Additional peaks around 400-500°C for both sEPS and sEPS-based hydrogel DTG curves are attributable to lipids thermal decomposition (Bach and Chen, 2017). These speculations are corroborated by FT-IR analysis.
The bound/bulk water measurement in hydrogels was assessed by differential scanning calorimetry (DSC, Q1000 TA Instruments). DSC analysis showed a sEPS%wt dependent bound water fraction (up to 15%) with a net prevalence of non-freezable population and bulk water fraction. Oscillatory shear measurements were carried out with a plate-plate geometry (20 mm diameter, 400 µm gap) on a Discovery Hybrid Rheometer (Disc.HR-3, TA Instruments) working in controlled shear stress.
Preliminary rheological tests revealed a discontinuity of the solid-like mechanical properties at increasing sEPS%wt at approximately 5%wt sEPS. Unfortunately rheological data analysis were not completed due to COVID-19 work limitations and will be included in the presentation at the Biofilms 2020 virtual conference.
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Figure 1.1 Conceptual diagram phase of hydrogel formation from different sEPS concentrations (%
[gVS/gWetWeight]). The cross-linking reaction was promoted by Ca2+ diffusion from a CaCl2 2.5%wt solution into the sEPS matrix through a dyalisis membrane (3.5 kDa MWCO).
Table 1.1 sEPS characterization and elemental composition of sEPS and the hydrogel. Reference studies are *(Felz et al., 2016), **(Lin et al., 2010), ***(Felz et al., 2019),****(Felz, 2019).
Physico-chemical
characterization Unit This study Reference studies
VS/TS, sEPS gVSsEPS/gTSsEPS 0.87 ± 0.01 0.87* -
Yield, sEPS gVSsEPS/gVSAGS 0.230 ± 0.010 0.200 ± 0.010* 0.160 ± 0.004**
PN, as BSA equivalent mgPNBSA/gVSsEPS 617.8 ± 0.7 381.0 ± 5.0*** < detection limit of
Bradford Standard Assay**
PS, as glucose equivalent mgPSGlu/gVSsEPS 128.5 ± 3.1 138.0 ± 2.5*** 122.5 **
PS, as alginate equivalent mgPSAlg/gVSsEPS 942.7 ± 15.1 - 486.2 ± 22.3**
PN/PSGlu gPNBSA/gPSGlu 4.81 ± 0.05 2.76 ± 0.08 -
PN/PSAlg gPNBSA/gPSAlg 0.66 ± 0.07 - -
TN mgN/gVSsEPS 64.46 ± 0.49 - -
TP mgP/VSsEPS 5.46 ± 0.01 - -
sEPS Elemental composition This study Reference study****
C mol C/C-mol 1 1
H mol H/C-mol 1.678 1.807
N mol N/C-mol 0.116 0.139
P mol P/C-mol 0.003 0.024
S mol S/C-mol 0.010 0.013
Ca mol Ca/C-mol 0.003 -
O mol O/C-mol 0.453 -
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Figure 1.2 (a) Thermogravimetric (TG – continuous line) and differential thermogravimetric (DTG – dashed line) curves of dried sEPS. (b) comparison between DTG curves of dried sEPS (red) and hydrogel (black) formed by a cross-linking reaction promoted by Ca2+ diffusion from a CaCl2 2.5%wt solution into the sEPS matrix through a dyalisis membrane (3.5 kDa MWCO).
References
Bach, Q.V., Chen, W.H., 2017. A comprehensive study on pyrolysis kinetics of microalgal biomass. Energy Convers. Manag.
https://doi.org/10.1016/j.enconman.2016.10.077
Dreywood, R., 1946. Qualitative Test for Carbohydrate Material. Ind. Eng. Chem. - Anal. Ed.
https://doi.org/10.1021/i560156a015
Felz, S., 2019. Structural Extracellular Polymeric Substances from Aerobic Granular Sludge.
Felz, S., Al-Zuhairy, S., Aarstad, O.A., van Loosdrecht, M.C.M., Lin, Y.M., 2016. Extraction of Structural Extracellular Polymeric Substances from Aerobic Granular Sludge. J. Vis. Exp. https://doi.org/10.3791/54534
Felz, S., Vermeulen, P., van Loosdrecht, M.C.M., Lin, Y.M., 2019. Chemical characterization methods for the analysis of structural extracellular polymeric substances (EPS). Water Res. https://doi.org/10.1016/j.watres.2019.03.068
Kehrein, P., Van Loosdrecht, M., Osseweijer, P., Garfí, M., Dewulf, J., Posada, J., 2020. A critical review of resource recovery from municipal wastewater treatment plants-market supply potentials, technologies and bottlenecks. Environ. Sci. Water Res. Technol. https://doi.org/10.1039/c9ew00905a
Li, X., Lin, S., Hao, T., Khanal, S.K., Chen, G., 2019. Elucidating pyrolysis behaviour of activated sludge in granular and flocculent form: Reaction kinetics and mechanism. Water Res. https://doi.org/10.1016/j.watres.2019.06.074
Lin, Y., de Kreuk, M., van Loosdrecht, M.C.M., Adin, A., 2010. Characterization of alginate-like exopolysaccharides isolated from aerobic granular sludge in pilot-plant. Water Res. 44, 3355–3364.
Lotti, T., Carretti, E., Berti, D., Martina, M.R., Lubello, C., Malpei, F., 2019. Extraction, recovery and characterization of structural extracellular polymeric substances from anammox granular sludge. J. Environ. Manage.
https://doi.org/10.1016/j.jenvman.2019.01.054
Milani, P., França, D., Balieiro, A.G., Faez, R., 2017. Polymers and its applications in agriculture. Polímeros 27, 256–266.
Morra, M.J., Fendorf, S.E., Brown, P.D., 1997. Speciation of sulfur in humic and fulvic acids using X-ray absorption near-edge structure (XANES) spectroscopy. Geochim. Cosmochim. Acta. https://doi.org/10.1016/S0016-7037(97)00003-3 Smith, P.K., Krohn, R.I., Hermanson, G.T., Mallia, A.K., Gartner, F.H., Provenzano, M.D., Fujimoto, E.K., Goeke, N.M.,
Olson, B.J., Klenk, D.C., 1985. Measurement of protein using bicinchoninic acid. Anal. Biochem.
https://doi.org/10.1016/0003-2697(85)90442-7
177 Presenting Author
Dr. Campo Riccardo Post Doctoral researcher
Università degli Studi di Firenze, Dipartimento di Ingegneria Civile e Ambientale (DICEA), Via di S.Marta 3, 50139, Firenze.
Is the presenting author an IWA Young Water Professional? N (i.e. an IWA member under 35 years of age)
Bio: Riccardo Campo is currently a post-doctoral researcher at the University of Florence (Italy) working in the research group of environmental and sanitary engineering. Recently, he is dealing with three main research topics: i) aerobic granular sludge (AGS) technology for low C/N wastewater treatment; ii) extracellular polymeric substances (EPS) extraction from AGS, characterization and recovery; iii) phosphorous recovery from AGS. In the recent past, his research was focused on AGS technology for the treatment of industrial wastewater (oily wastewater, fish-canning wastewater), membrane bioreactors (MBRs) and moving bed biofilm reactors (MBBRs) for the treatment of civil and industrial wastewater.
https://www.scopus.com/authid/detail.uri?authorId=56870931700
178
Extraction conditions govern the quantity and properties of gel-forming EPS recovered from aerobic granules.
Bou Sarkis, A.*,***, Pagliaccia, B. **,*** Durieux, S. *** Bounouba, M. *** Bessiere, Y. *** Derlon, N. ****
Paul, E. ***and Girbal-Neuhauser, E.*
* LBAE, Université de Toulouse, UPS, Auch, France
** DICEA, University of Florence, Firenze, Italy
***TBI, Université de Toulouse, CNRS, INRAE, INSA, Toulouse, France
**** EAWAG, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf, Switzerland Keywords: Aerobic granules; extraction selectivity; extracellular polymeric substances.
Summary of key findings
Structural EPS extracted from aerobic granules constitute valuable raw biomaterials for industrial applications but their properties differ greatly with the chemicals used for granule solubilization. Four solvents were evaluated and performances in terms of yield and selectivity were investigated. Alkaline solvents appear as the most efficient extractant but it was shown that NaOH is two times more efficient than Na2CO3 for granule solubilization. However, considering the additional precipitation step needed for enrichment in gel-forming EPS, it was noted that the polymers extracted with Na2CO3 are precipitated to a larger extent than those extracted in NaOH (25% precipitated against 6,6%). This can be explained by a better precipitation of molecules present in Na2CO3 since the size profiles reveal that the Na2CO3
protocol was able to extract EPS with higher molecular weights compared to the EPS extracted with the NaOH protocol. The findings of this study can help to set strategies for the extraction of gel-forming EPS from densified aggregates for their valorization as innovative biomaterials.
Background and relevance
The extracellular polymeric substances (EPS) secreted by aerobic granular sludge are mainly associated with the physical structure, rheological behavior and stability of the granules. EPS can form hydrogels that have many industrial applications, such as coating materials or biosorbents with interesting swelling or de-swelling characteristics, etc. (Lin et al., 2015).
The EPS components are far complex and to be able to benefit from their interesting properties, the EPS extraction methods need to be optimized according to their efficiency and selectivity for EPS recovery for targeted industrial applications. In particular, the effect of solvent has to be addressed.
Indeed, harsh extraction techniques might be required to solubilize the extracellular matrix and recover the structural EPS from dense heterogeneous aggregates such as biofilms and aerobic granules. Alkaline extraction is widely used for the extraction of functional gel-forming constituents in aerobic granules and relies in general on the use of NaOH or Na2CO3 ( Lin et al., 2010, Seviour et al., 2010, Felz et al., 2016) but it is not clear how the base used during extraction influences the quantity and composition of EPS that are recovered. Alkaline conditions induce many charged groups, such as carboxylic groups in protein and polysaccharides, to be ionized (Sheng et al., 2010). Another possible extraction technique is the use of urea, which is a small molecule that can have a direct (cleavage of hydrogen bonds in the EPS matrix) and an indirect (by altering the solvent environment and therefore influencing the hydrophobicity) effect on molecules present in the EPS matrix (Bennion & Daggett, 2003).
The aim of this study is thus to explore the performances of harsh extraction protocols coupling chemical and thermal methods on EPS recovery in terms of yield and selectivity for the extraction of gel-forming polymers. A specific focus will be held on the size of the EPS extracted. Indeed, the size of the molecules influence the gelation capacity, and in the case of alginates a higher molecular weight gives the compound a better gel formation kinetics and enhanced viscosities (Fernández Farrés & Norton, 2014).
Thus, selectivity towards high molecular weights can be a criterion to select an extraction protocol.
179 Methodology and Results
Methodology
The extraction protocol was divided into two steps: a solubilization of polymers (step 1) and a precipitation of the solubilized polymers (step 2). For EPS solubilization, 0.4g of dried granules (produced in Eawag, according to Layer et al., 2019) were rehydrated in the chosen solvent (NaCl, urea, Na2CO3 or NaOH, all at 0,2 mol. L-1) and the granule suspension was homogenized using ultraturrax T25. A heating phase at 80°C for 60 minutes was added in order to enhance granules disintegration and EPS solubilization. After centrifugation the supernatant was recovered (named S1) and the pellet (named P1) was analyzed for its content in volatile solids (VS). As previously described for the precipitation of alginate like polymers (Lin et al 2010), the pH of the S1 extract was adjusted to pH 2±0.02 with 1 M HCl, and after incubation for 30 minutes at 4°C, the precipitate was collected by centrifugation. The precipitate was solubilized in 0.1 M NaOH, dialyzed for 36 hours against distilled water and lyophilized (named P2). The lyophilizate was resuspended in 2 mL of NaOH 0.1M (named S2).
Results
Table 1 shows the efficiency of 4 protocols based on different chemical solvents for the recovery of organic matter from aerobic granules.
Alkaline extraction, i.e. Na2CO3 or NaOH, based on the increase of repulsive forces in the EPS matrix is more efficient for solubilizing the granule matrix (S1 samples) in comparison with urea which is able to solubilize hydrophobic polymers and NaCl that can be considered here as a control for high temperature alone. NaOH was the most efficient for solubilizing granules (661.96±47.2 mgVS solubilized/gVS) followed by Na2CO3 with a solubilizing power diminished by half approximately (331.8±12.93 mgVS solubilized/gVS). After acidic precipitation, dialysis and lyophilization, we can see an inversion in the final Y2 yield leading to 43,93 ± 11,84 mgVS precipitated/gVS for NaOH and 83,16
± 9,82 for Na2CO3 in S2 final samples. Thus, only 6,6% of the NaOH solubilized polymers were precipitated while the precipitation has concerned 25% of the polymers solubilized in Na2CO3. Therefore, the solubilization should not be the only parameter considered to choose the solvent, because the recovery of EPS after the precipitation step might be different to what is observed during solubilization and the yields can be inversed.
Table 1: Effect of solvents on the yield of solubilization (Y1) and precipitation (Y2) of VS in granules Extraction method Y1 a : mg VS solubilized/ gVS granule Y2 b : mg VS precipitated/gVS granule
NaCl 0,2 M 80°C 146,76 ± 42,79 4,48± 0,54
NaOH 0,2 M 80°C 661,96 ± 47,2 43,93 ± 11,84
Na2CO3 0,2 M 80°C 331,8 ± 12,93 83,16 ± 9,82
Urea 0,2 M 80°C 85,16 ± 29,73 13,07 ± 2,47
a VS recovered in S1 after chemical and thermal solubilization of initial VS in granules
b VS recovered in S2 after acidic precipitation of solubilized VS in S1
For the extracts obtained after NaOH / 80°C solubilization (Figure 1A) it can be observed that a major part of the extract consists of low molecular weight (89.7 % surface area). For Na2CO3 this fraction is less represented but remains a major part of the extract (60.9% surface area) and an individualized peak of medium molecular weight (24.7% surface area) can be noted. The chromatograms obtained after resolubilization of the lyophilized EPS (Figure1B) show us that Na2CO3 has an additional peak of high molecular weight eluted at 8 mL in comparison with NaOH and the medium molecular weight peak is more significant in Na2CO3. Therefore, the extraction protocol has an effect on the selectivity of molecules in terms of their molecular weight. NaOH seems to be more specific for the solubilization of low molecular weights that are mostly lost during precipitation and the dialysis step whereas Na2CO3 is more convenient to recover high and medium molecular weight molecules.
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Figure 1: Selectivity of NaOH (dotted line) and Na2CO3 (solid line) on the size profiles of solubilized polymers S1 (A) and acid precipitated polymers S2 (B).
Size Exclusion Chromatography was performed on Superdex 200 column with Tris HCl 50 mM; NaCl 0,1M, pH 9±0.02 as the mobile phase circulating at 0.5 mL/min. The detection of molecules was done at 280 nm. The column separates effectively molecules between 600 kDa and 10 kDa. The calibration curve was done using proteins with sizes varying from 669 kDa (thyroglobulin) to 6.5 kDa (aprotinin). LMW=low molecular weight (<20 kDa), MMW= medium molecular weight (20 to 200 kDa), HMW= high molecular weight (>200 kDa).
Discussion
The results presented here indicate that the use of alkaline extraction methods is efficient for the extraction of EPS giving a glimpse of the possibilities of upgrading the constituents of aerobic granular sludge. This is consistent with literature since after comparison of physical or chemical methods, Feng et al (2019) showed that alkaline extraction using sodium carbonate and sodium hydroxide are the most efficient for EPS recovery from aerobic granules. Lin et al (2010) were successful at producing hydrogels with calcium chloride with extracted EPS by Na2CO3/ 80°C that they called ALE for alginate like exopolymers while a complex heteropolysaccharide called Granulan was extracted from granules by the use of sodium hydroxide (Seviour et al., 2012).
However, besides the interesting yields offered by these harsh alkaline methods for recovery of gel- forming polymers, our results suggest that depolymerization of molecules due to the alkaline conditions can occur. Temperature and pH govern the depolymerization rate of Alginate, which undergoes β- elimination and oxidative-reductive depolymerization (ORD) (Holme et al., 2008). The results obtained suggest a higher depolymerization with NaOH than in Na2CO3 that may significantly reduce the gelling capacity of the recovered polymers.
In conclusion, the extraction protocol influences to a large extent the amount of recovered EPS but also the type of molecules extracted. With sodium hydroxide mostly small molecules are selected which might be due to lysis or degradation of high molecular weight molecules, whereas sodium carbonate has a good selectivity for high molecular weights and selects less of the low molecular weights. The type of the extractant used should also be selected based on the efficiency after precipitation and not only the solubilization step. Further investigations will be presented to explore the influence of parameters such as temperature, and solvent concentration to be able to select a unified protocol for the extraction of EPS that have gelifying capacities.
181
References:
Bennion, B. J., & Daggett, V. (2003). The molecular basis for the chemical denaturation of proteins by urea.
Proceedings of the National Academy of Sciences of the United States of America, 100(9), 5142–5147.
https://doi.org/10.1073/pnas.0930122100
Felz, S., Al-Zuhairy, S., Aarstad, O. A., van Loosdrecht, M. C. M., & Lin, Y. M. (2016). Extraction of structural extracellular polymeric substances from aerobic granular sludge. Journal of Visualized Experiments, 2016(115), 1–8. https://doi.org/10.3791/54534
Fernández Farrés, I., & Norton, I. T. (2014). Formation kinetics and rheology of alginate fluid gels produced by in- situ calcium release. Food Hydrocolloids, 40, 76–84. https://doi.org/10.1016/j.foodhyd.2014.02.005
Holme, H. K., Davidsen, L., Kristiansen, A., & Smidsrød, O. (2008). Kinetics and mechanisms of depolymerization of alginate and chitosan in aqueous solution. Carbohydrate Polymers, 73(4), 656–664.
https://doi.org/10.1016/j.carbpol.2008.01.007
Layer, M., Adler, A., Reynaert, E., Hernandez, A., Pagni, M., Morgenroth, E., Holliger, C., & Derlon, N. (2019).
Organic substrate diffusibility governs microbial community composition, nutrient removal performance and kinetics of granulation of aerobic granular sludge. Water Research X, 4, 100033.
https://doi.org/10.1016/j.wroa.2019.100033
Lin, Y., de Kreuk, M., van Loosdrecht, M. C. M., & Adin, A. (2010). Characterization of alginate-like
exopolysaccharides isolated from aerobic granular sludge in pilot-plant. Water Research, 44(11), 3355–
3364. https://doi.org/10.1016/j.watres.2010.03.019
Lin, Y. M., Nierop, K. G. J., Girbal-Neuhauser, E., Adriaanse, M., & van Loosdrecht, M. C. M. (2015). Sustainable polysaccharide-based biomaterial recovered from waste aerobic granular sludge as a surface coating material. Sustainable Materials and Technologies, 4, 24–29. https://doi.org/10.1016/j.susmat.2015.06.002 Seviour, T., Donose, B. C., Pijuan, M., & Yuan, Z. (2010). Purification and conformational analysis of a key
exopolysaccharide component of mixed culture aerobic sludge granules. Environmental Science and Technology, 44(12), 4729–4734. https://doi.org/10.1021/es100362b
Seviour, T., Yuan, Z., van Loosdrecht, M. C. M., & Lin, Y. (2012). Aerobic sludge granulation: A tale of two polysaccharides? Water Research, 46(15), 4803–4813. https://doi.org/10.1016/j.watres.2012.06.018 Sheng, G. P., Yu, H. Q., & Li, X. Y. (2010). Extracellular polymeric substances (EPS) of microbial aggregates in
biological wastewater treatment systems: A review. Biotechnology Advances, 28(6), 882–894.
https://doi.org/10.1016/j.biotechadv.2010.08.001
Presenting Author
Bou Sarkis Abdo PhD student
LBAE, Université de Toulouse, UPS, Auch, France
TBI, Université de Toulouse, CNRS, INRAE, INSA, Toulouse, France Is the presenting author an IWA Young Water Professional? Y/N N
Bio: Bou Sarkis Abdo is a second year PhD student at the Paul Sabatier University.
He obtained a bachelor’s in biology and a food engineering degree from the Lebanese University. His current research interests are oriented towards the environmental sciences, specifically towards the use of microbial aggregates for the purification of water and their valorization.
182
Effect of mass transfer on the S0 and N2O accumulation in sulfide-oxidizing autotrophic denitrification process: batch experiments and modeling
evaluation
Zou, X.*, Chen, G.H.*, Wu, D.*
*Department of Civil and Environmental Engineering, The Hong Kong University of Science and Technology, Hong Kong SAR
*
Keywords: Sulfur-based autotrophic denitrification; SOB biofilms; reaction intermediates Summary of key findings
Through experimental and modeling observation, we found:
(1) High-rate sulfide-oxidizing bacteria biofilms (SOBiflms) could be developed in moving-bed biofilm reactor (MBBR) through long term reactor operation.
(2) The occurrence of mass transfer limitation in SOBiflms attribute to the production of of intermediates i.e. elemental sulfur and nitrous oxide.
Background and relevance
Sulfide-oxidizing autotrophic denitrification is a promising alternative to conventional heterotrophic denitrification in wastewater biological nitrogen removal. The sulfur-oxidation and autotrophic denitrification intermediates – elemental sulfur (S0) and nitrous oxide (N2O) is crucial. But their roles in the sulfur-nitrogen transformations are yet to be studied in-depth.
Our team has developed a bench-scale SOAD MBBR and continuously operate this reactor over 700 days. We taken mature biofilm from this bioreactor to further investigate the effect of external mass transfer on the SOAD process (for SOAD biofilm system). In detail, the MBBR biofilm samples were separated into two groups, one group mainted its original form; the other group of biofilm biomass was scrapped off and resuspend in batch reactor with synthetic sewage mimicking no mass transfer limitation. We used the two biofilm/biomass sample groups to conduct comparative batch
expeirments. The obtained experimental results were used to determine SOB biofilm model parameter.
Key Results
• Compared to batch test with MBBR, batch test with detached MBBR sludge results in 34% less N2O production and 49% less S0 production. Which means, mass transfer on the biofilm has an signficant effect to the accumulation of S0 and N2O (Fig.1A and B).
• Meanwhile, the sludge from detached MBBR and biofilm were both well described by the sulfide-oxidizing autotrophic denitrification model conducted by Aquasim (Fig.1C and D).
• This study gives the first intuition about how the mass transfer effect influences the N2O emission in SOAD biofilm system through experimental investigation and model verification.(the matrix of biofilm model is referred to Table 1, according to Stijin et al., (accepted)).
183
a b
Figure 1.1 Nitrous oxide accumulation in batch tests (a); Element sulphur accumulation in batch tests (b);
Kinetic simulation of element sulphur accumulation (c and d) (Note: with and without MBBR stand for SOB biofilm and SOB biomass samples respectively)
Table 1.1 Stoichiometric matrix for the bioconversion model.
Component → SH2S SS0 SSO42- SNO3- SNO2- SN2O XSOB XI
Process ↓ g S·m-3 g S·m-3 g S·m-3 g N·m-3 g N·m-3 g N·m-3 g COD·m- g COD·m
-3 3
1a. H2S -1 1 -(0.5-YH2S) (0.5-YH2S)
denitratation
1b. H2S YH2S
-1 YH2S
1 1.14YH2S 1.14YH2S
-(0.5-YH2S) (0.5-YH2S) 1 denitritation 1
2a. S0 YH2S YH2S
-1 1 -(1.5-YS0) 1.72YH2S
(1.5-YS0) 1.72YH2S
denitratation 1
2b. S0 YS0
-1 Y1 S0
1.14YS0 1.14YS0
-(1.5-YS0) (1.5-YS0) denitritation
3. Decay of XSOB YS0 Y S0 1.72YS0 1.72YS0 1
-1 fI
References (not incluced in page count, but please keep to a reasonable length)
Cui, Y. X.; Wu, D.; Mackey, H. R.; Chui, H. K.; Chen, G. H. Application of a Moving-Bed Biofilm Reactor for Sulfur- Oxidizing Autotrophic Denitrification. Water Science and Technology. 2017, 77(4), 1027-1034.
S.O. Decru, J.E. Baeten, Y.X. Cui, D. Wu, G.H. Chen, E.I.P. Volcke. Model-based analysis of sulfur-based denitrification in a biofilm reactor. Water Science and Technology. 2020.
C d
184 Presenting Author
Mr. Zou Mphil student
My affiliation: The Hong Kong University of Science and Technology Is the presenting author an IWA Young Water Professional? N (i.e. an IWA member under 35 years of age)
Bio: Xu Zou is a wastwater process simulation specialist with two years successful experience in novel wastewater process kinetic developing and modeling.
185