An innovative nutritional slow-release packing material with functional microorganisms for biofiltration: Characterization and performance evaluation

Journal of Hazardous Materials 366 (2019) 16–26

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An innovative nutritional slow-release packing material with functional microorganisms for biofiltration: Characterization and performance evaluation

T

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Zhuowei Chenga, Ke Fenga, Danhua Xua, Christian Kennesb, Jianmeng Chena, , Dongzhi Chena, ⁎ Shihan Zhanga, Jiexu Yea, Dionysios D. Dionysiouc, a

College of Environment, Zhejiang University of Technology, Hangzhou, 310009, China Chemical Engineering Laboratory, Faculty of Science, University of La Coruna, 15001, Spain Environmental Engineering and Science Program, Department of Chemical and Environmental Engineering (ChEE), University of Cincinnati, Cincinnati, OH 45221-0012, USA b c

G R A P H I C A L A B S T R A C T

A R T I C LE I N FO

A B S T R A C T

Keywords: Nutritional slow-release pH buffer Function microorganisms Biotrickling filter Removal performance

The type of packing material for biofiltration has a great impact on microbial growth and pollutant removal. This study evaluated the feasibility of a nutritional slow-release packing material with functional microorganisms (NSRP-FM) in a biofilter for the removal of gaseous n-butyl acetate. Through the emulsification-cross linked process and microbial immobilization, an innovative packing material was obtained, with a specific surface area of 2.45 m2 g−1 and a bulk density of 40.75 kg m-3. The cumulative release rates of total phosphorus and total nitrogen were 90.6% and 75.6%, respectively, as measured while continuously spraying deionized water. To evaluate the performance of biofiltration, NSRP-FM was compared with the commercial polyurethane foam (PUfoam), in two identical biotrickling filters (BTFs). The BTF packed with the prepared NSRP-FM maintained a consistent removal efficiency (over 95%) without nutrients addition and pH adjustment. The other BTF had poor removal performance, and the removal efficiency declined to 65% when there was no pH adjustment. Energy dispersive X-ray spectroscopy (EDS) analysis of NSRP-FM showed that inorganic elements were released during the operation of BTF. The abundance of functional microorganisms suggested that the prepared NSRP-FM provided a better environment for microbial growth, despite changes in the operating conditions.

⁎

Corresponding authors. E-mail addresses: [email protected] (J. Chen), [email protected] (D.D. Dionysiou).

https://doi.org/10.1016/j.jhazmat.2018.11.070 Received 13 May 2018; Received in revised form 15 November 2018; Accepted 16 November 2018 Available online 17 November 2018 0304-3894/ © 2018 Published by Elsevier B.V.

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1. Introduction

containing starch as the carbon source reduced the start-up period of a biofilter [11]. As mentioned previously, the inoculated microorganisms play key roles for the performance of the biofilter. Since specific microorganisms are necessary for the removal of most VOCs and odorants, some researchers suggested that the functional microbial agents should be prepared, not only for optimal microbial activity but also for the quick biofilm formation [21,22]. If the functional microorganisms were fixed on the packing material, the inoculation process was much simpler, and no inoculum was needed during start-up. However, only few studies prepared and evaluated the performance of packing materials embedded with microorganisms. Most used mixed microbial cultures originating from acclimated activated sludge, rather than specific VOCdegrading strains [20,23]. This study aimed to synthesize a novel packing material with slowrelease nutrients, pH buffer capacity and two functional microbial species for the removal of gaseous n-butyl acetate. Some physicochemical properties (bulk density, specific surface area, bulking capacity etc.) of the prepared packing material were evaluated, along with the stability of microbial activity for long-term storage. A comparison was carried out using two identical biotrickling filters (BTFs) packed with the prepared material and commercial PU-foam. Evaluation of the start-up time, elimination capacity, microbial structure, pH and electric conductivity of the recirculation liquid was conducted.

There are numerous techniques designed for the treatment of gaseous organic pollutants (mainly volatile organic compounds, VOCs) and odorous compounds (mainly those containing sulfur) from industrial production and waste treatments. Of these methods, biopurification is considered one of the most effective, especially for larger volumes of gaseous pollutants at low concentrations [1,2]. Biopurification has been widely studied and implemented in real world applications due to its numerous advantages, including no secondary pollution, lower cost, convenient maintenance and operation [3]. Recently, bioaerosol emissions from the biofilters have become a considerable and key issue, since it would seriously affect the comfort of surroundings and the health of nearby people. The emissions of bioaerosol could be related to several factors, such as the type of carrier used, bioreactor configuration, and any other conditions that affects reactor performance [4,5]. Also, the utilization of some other technologies (e.g. UV photodegradation) could decrease the emission of bioaerosol [6]. During biopurification, gaseous pollutants are transferred to a biofilm, which was developed on the packing material. The microbial activity within the biofilm is considered to be the major factor that influences purification efficiency; however, the packing material, which provides the microorganisms with favorable environmental moisture, pH, nutrients and oxygen supply, is also an important factor for efficiency and stability of the biofilm [7,8]. According to previous studies, the ideal packing material should have specific physicochemical properties, such as suitable particle size, large surface area, high porosity, and so on [9–11]. To date, both organic and inert materials have been used as packing materials in industrial treatments. Malhautier et al. [12] reported that approximately 87% of full-scale operation biofilters were equipped with organic packing material, among which the most popular were wood chips and compost. However, the life span of organic materials is often shorter than that of inert materials (e.g., polyurethane foam or lava rock). Thus, the organic packing materials must be replaced frequently, accounting for nearly more than 45% of the total operating costs [13]. Meanwhile, the inert materials tend to be more expensive and usually require periodic nutrient supply [14]. As a result, the cost related to the inert packing materials accounts for half of the total costs (investment plus operating costs). Over the last few years, most researchers directed their attention to the development of new packing materials [15,16]. Some modified the original packing materials by physicochemical methods, in order to increase specific surface area, reduce the resistance and so on. Li et al. [17] prepared a novel packing material consisting of iron oxide-coated porous ceramsite through surface coating method for NO removal, and it took only 8 d for biofilm formation since the surface of new packing was more suitable for microbial growth. Others mixed the inert and organic material to obtain a novel packing material with their respective advantages. Brandt et al. [18] mixed composted leaves with expanded vermiculite, and results showed the mixtures provided an attractive environment for microbial growth and better diffusion for CH4. Recently, some nutrient sources necessary for microbial growth are artificially added into the packing material during the preparation [19]. In addition, because some deleterious acidic substances are formed during the microbial degradation, some compounds with high buffer capacities, such as CaCO3 and phosphate, are also incorporated into the packing materials [20]. The development of packing material has led to an improvement in the stability and performance of biofilters, including fast startup, high removal capacity, and short recovery time under unfavorable conditions (e.g. a lack of nutrient supply, frequent pH fluctuations). Dumonta et al. [10] developed a new cylindricalshaped extrudate called UP20 for the treatment of H2S, and the results showed that this packing material offered an advantage to the performance at high pollution load via self-regulating of pH conditions and nutrients. Compared to traditional PU-foam, a modified foam

2. Material and method 2.1. Microorganisms and culture medium One fungus Aspergillus fumigates HD-2 (NCBI accession NO. KJ764711) and one bacterium Ralstonia pickettii L2 (NCBI accession NO. GQ906999), both having specific biodegradability to n-butyl acetate, were chosen as the functional microorganisms. These two strains were isolated by our group and stored in China Center for Type Culture Collection under CCTCC No. M2014531 and M209250. More details about these two strains could be found in our previous studies [24,25]. These two strains were inhabited on the potato-dextrose (PDA)-agar and Luria-Bertani (LB)-agar, respectively, and both stored under 4℃. R. pickettii L2 and A. fumigates HD-2 were inoculated in LB liquid medium (pH = 7.0–7.2) and PDA liquid medium (pH = 7.0–7.2) for enrichment at 30℃ and 160 rpm, respectively [22]. 2.2. Packing material preparation Sodium alginate (SA) solution (1.5 g·L−1) was prepared using mineral inorganic medium (K2HPO4∙3H2O 9.42 × 10−1 g∙L−1, KH2PO4 2.34 × 10−1 g∙L−1, NaNO3 1.70 g L−1, NH4Cl 9.80 × 10-1 g∙L−1) and then stored at -4℃ for 12 h. Calcium chloride (CaCl2) solution (22.2 g·L−1) was added to the SA solution in two steps. First, a small amount of CaCl2 solution was sprayed on the surface of the SA solution. After a thin film formed on the surface of SA solution, CaCl2 solution was slowly added to the SA solution using a glass rod. The mixture was kept at -4℃ for the reaction to proceed, and the alginate gel formed after 72 h. To remove residual water, the alginate gel was dried at 4850℃. Finally, the obtained porous alginate gel was cut into several cubes with length of 15 mm. R. pickettii L2 and A. fumigates HD-2 were cultured to their logarithmic growth period (after 3 h and 32 h, respectively, from Supplementary Information (SI) Fig. S1) under aseptic condition, respectively. The obtained strains were mixed with each other at equal amounts of protein (containing by R. pickettii L2 or A. fumigates HD-2) and suspended in the sterilized deionized water. Then, 10 g alginate-gel cubes were immerged into 100 mL strain suspension (2 mg protein∙mL−1) for adsorption (60 min, optimal) and then washed by mineral inorganic medium to remove any microorganisms not firmly 17

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adsorbed on the surface. This process was repeated 3 times and finally all the cubes were dried at 30℃ until the water in the alginate-gel leaving off. Finally, the nutritional slow-release packing material with functional microorganisms (NSRP-FM) was obtained. The cells on NSRP-FM were in the dormancy period and the activity would recover when there was water and carbon source from outside. For the comparison, SA-gel without any mineral inorganic medium and SA-gel without any functional microorganisms were prepared as the control sample.

stream into the pure n-butyl acetate liquid and mixing with another air stream to get different inlet concentrations. The mixed gas was introduced into the biofilters from the bottom and the recirculation liquid was sprayed from the top. The component of the recirculation liquid could be found in Supplementary Information (SI) Table S1. The BTFs were operated for more than 2 months under continuous mode, and it was divided into four stages, Stage I from day 0–11, Stage II from day 11–31, Stage III from day 31–44, and Stage IV from day 4560. The different operational parameters are provided in Table S1. During the whole operation, the recirculation flow was set at a constant flow rate of 150 mL∙min−1, sustaining enough moisture for the microbial growth. Stage I was for the start-up of both biofilters. n-Butyl acetate-acclimated activated sludge, mixed with the strain suspension (containing R. pickettii L2 and A. fumigates HD-2 at their logarithmic growth period), was used as the inoculum for BTF2. Since the prepared NSRP-FM contained these two strains, only the acclimated activated sludge was used for the inoculation of BTF1. To make sure the conditions were identical for both biofilters, the biomass (expressed as the protein) of the strain suspension and the NSRP-FM were nearly the same. The utilization of acclimated activated sludge could enhance the formation of biofilm quickly and sustain the stable operation since it contained many other microorganisms [28,29]. From Stage II to IV, no nutrient solution was artificially introduced. Moreover, in Stage III, the pH value of the recirculation liquid was also not adjusted by NaOH.

2.3. Analysis of physical and chemical properties Dimension, density (apparent and bulk) and water-holding capacity were analyzed in order to validate the prepared materials as potential packing material for biofilters, according to the measurements proposed by Dorado et al. [26]. Specific surface area and porosity were measured by a volumetric sorption analyzer (ASAP 2020, Micromeritics Co., USA). The morphology of NSRP-FM was determined by field emission scanning electron microscopy (SEM) (Philips XL-30-ESEM, Holland). The surface chemical group and elemental analyses were carried out by Fourier transform infrared spectroscopy (FT-IR) (Thermo Nicolet6700, USA) and energy dispersive X-ray spectroscopy (EDS) (Oxford AZtecX-Max80, England), respectively. The release rate of total nitrogen (TN) and total phosphorus (TP) for NSRP-FM were measured as follows: randomly selected packing materials (10 g) were packed in a column which was made of plexiglass, and deionized water was sprayed from the top. When the filler layer began to drop liquid, a certain amount of leaching liquid was collected every 24 h. TN and TP were measured according to the National Standard Methods HJ 636–2012 and GB 11893-89 [27]. pH buffer capacity was evaluated under weak acidic (pH 5.5) conditions. Simply, 2.4 g of randomly selected packing materials were immersed in 800 mL distilled water at pH 5.5. An automatic pH monitor was used to measure the pH value of the leaching solution. When the pH value increased to 7.2, HCl was used to adjust the pH to its initial value (5.5).

2.6. Analytical methods Gaseous n-butyl acetate was measured by an Agilent 6890 gas chromatograph (Agilent Technologies, Palo Alto, CA, USA) using a HPInnowax column (30 m × 0.32 mm × 0.5 μm) with injector 200℃ oven 100℃ and detector 180℃. The intermediates from the microbial degradation of n-butyl acetate were determined by GC–MS and ion chromatography, using the n-hexane as the extracting agent. More details could be found in Supplementary Information S1. For microbial analysis, 2 g packing materials were collected periodically from each biofilter. Genomic DNA was extracted using the E.Z.N.A.® soil DNA Kit (Omega Bio-tek, Norcross, GA, U.S.) according to the procedures. Amplicon and pyrosequencing were performed by MajorBio Pharma-ceutical Technology Co., Ltd. (Shanghai, China) using a 454/RocheGS-FLX Titanium instrument (Roche, Nutley, NJ). Pyrosequence reads were analyzed using Amplicon Noise 1.25. More details could be found in S2. All values presented in the figures were the averages of a minimum of three independent experiments, and each experiment was conducted in triplicate. The error bars represented the standard deviations of triplicates.

2.4. Biomass and biodegradation activity analysis Two grams of packing materials were taken randomly for microbial analysis after storage time of 0, 7, 15, 30, and 60 d at room temperature. After shaking with sterilized phosphate buffer solution (pH = 7) for 3 h (280 rpm), the packing materials were partially broken, and the microorganisms were released into the solution. Then, the obtained microorganisms were disrupted by an ultrasonic cell disruption device (JY 92-II, Scientz Biotechnology Co., Ltd, Ningbo, China), the ultrasonic frequency of which was 15 kHz for R. pickettii L2 and 20 kHz for A. fumigates HD-2. The whole process was divided into 99 cycles, and every cycle contained 5 s working time and 10 s intermittent time. Finally, the obtained protein contents were determined by Folin-phenol method. Determination of the microbial biodegradation activity was processed as follows: 2 g packings were put into a series of flasks containing 50 mL of deionized water, with n-butyl acetate as the sole carbon source (180 mg L−1). The flasks were shaken at 30℃ with a speed of 160 rpm. The concentration of n-butyl acetate was measured every 12 h.

3. Results and discussion 3.1. Physical properties The physical properties of the NSRP-FM and other packing materials are listed in Table 1. The bulk density of the NSRP-FM was lower than other packing materials, except the PU-foam. Such a low bulk density is necessary to avoid cracking of the packing bed and the heavy weight on packing-supports [19]. The specific surface area of NSRP-FM was lower than PU-foam but higher than other packing materials, indicating that the NSRP-FM provided more surface area for microorganism attachment as well as gas-liquid contact. The large specific surface area was mainly attributed to the addition of CaCl2 into the sodium alginate solution to form porous structure. PU-foam is a good packing material and has been widely used in most biopurification systems for gaseous pollutants [30,31]. Therefore, further analysis was carried out between PU-foam and NSRP-FM. Based on the N2 adsorption-desorption curve, the pore size distributions were plotted according to the Barrett-Joyner-Halenda (BJH) equation, and

2.5. Biotrickling filter set-up and operational parameters Two identical BTFs are depicted in Fig. 1a, in which one was packed with NSRP-FM (marked as BTF1) and the other with commercial PU foam (marked as BTF2). Fig. 1b gives the schematic diagram of the BTF system. Gaseous n-butyl acetate was supplied by introducing a dry air 18

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Fig. 1. (a) Experimental pilot-scale biofilters. BTF1: NSRP-FM; BTF2: PU-foam. (b) Schematic diagram of the BTF system for n-butyl acetate purification.

elements. From the SEM images of NSRP-FM in Fig. 2 a and b, many holes and channels are distributed along the cross and longitudinal sections, respectively. Also, it could be seen (Fig.2c) that some holes existed on the PU-foam. The FT-IR spectrum (Fig. S3) of NSRP-FM shows the absorption peaks at 818.6 cm−1, 1594.0 cm−1 and 3352.6 cm−1, which were attributed to the bending vibration of surface CeC, C]O and OeH groups. These polar functional groups could enhance the surface hydrophilicity and promote the adhesion of microorganisms on packing material surface [34]. Gel hydration can form micropores, so that the nutrient elements can be released via concentration difference and vapor difference [35]. Fig. 3 a and b show the release rate and cumulative release amount of TN and TP from continuous water spray for about 25 d. Clearly, the release rate of TN was much faster than that of TP. Especially in the later phase, the release rates of TN and TP were 23 and 3 mg L−1·d−1, respectively. Through the calculation, the total released amount for TN and TP were 405.5 and 200.5 mg L−1; while the amount of TN and TP contained by NSRP-FM was 536.5 and 221.2 mg L-1, respectively. Therefore, 75.6% of TN and 90.6% of TP were released during the process. Besides P and N, NSRP-FM also contained other elements. Since there was a positive correlation between the electrical conductivity (a comprehensive indicator) and the ion concentration in solution, electrical conductivity was used to indirectly reflect the nutrient level. The initial conductivity of spraying water was nearly 0 μs·cm-1 (similar to that of the self-made deionized water). After spraying for 12 d, the conductivity of the circulating water rose to 1515 μs·cm-1, with an average release rate of 70∼80 μs·cm-1·d-1. Some nutrients (such as N, P and metal ions) contained by NSRP-FM were slowly released into the external deionized water and formed some inorganic salts, thus causing the electrical conductivity increasing. For the native sample SA-gel without any mineral inorganic medium, the final conductivity of the leaching liquid was stabilized at nearly 400 μs·cm-1, much less than the one for the prepared NSRP-FM (See Fig.

the results are shown in Fig. S2. The pores were mainly in the range of 0.5–3 nm for these two packing materials. These pores were mesoporous (2–50 nm) and micropores (< 2 nm), which are smaller than the mean length of most microorganism cells. This suggests that biomass attachment could easily take place on the surfaces of these materials [32]. According to Table 1, both PU-foam and NSRP-FM had higher porosity and lighter bulk density compared with other packing materials, implying a lower pressure drop across the bed during the operation. In addition, adsorption tests were performed at the filter (figure not shown) with an inlet n-butyl acetate concentration of 200 mg m−3. The results showed that the packing bed was saturated after 40 min and 45 min for NSRP-FM and PU-foam, respectively. Based on these basic physical properties, PU-foam was selected as the control one for the comparison of performance. 3.2. Chemical and slow-release properties EDS analysis was employed to determine the surface elemental compositions of the packing materials, and the results are shown in Table 2. The high percentage of some elements (N, P, Na, Cl, Ca) on NSRP-FM indicated that the dispersion of those elements, required by microbial growths, was achieved through the preparation process. However, PU-foam did not contain these elements, with the exception of C and O, which were the basic components of polyurethane. From Table 2, Na and Ca were distributed at different positions; that is, on the cross section, the content of Na (24.41%) was higher than that of Ca (17.85%); while on the longitudinal section, the content of Ca (32.34%) was higher than that of Na (6.86%). This might be explained by the “egg-box model" proposed by Sikorski et al. [33], which describes the reaction process between sodium alginate and calcium ion. When Ca2+ reacted with alginate to form a gel, a vertical channel was generated. As a result, more Ca2+ would distribute in this direction. Meanwhile, these vertical channels could be used as the release channels for nutrient

Table 1 Basic properties of packing materials prepared in this study and reported in the literature. Characteristic

Ceramsite

PU-foam

UP20

CM-5

NSRP-FM

Reference Type Dimension (mm) Bulk density (kg m−3) Specific surface area(m2 g−1) Porosity(%)

Liang et al., 2007 Organic 10-13 420-460 1.5-5 45-48

Cheng et al., 2011 Inorganic Φ14-18 35 4.43 90.8

Dumont et al., 2010 Release Φ7*15 920 <1 –

Zhu et al., 2016 Release Φ12*20 470 3.91 39

This study Release Φ15 40.75 2.45 92.6

19

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Table 2 Surface elemental analysis of packings before and after operating in BTFs. Element

NSRP-FM

Element

Cross section Before

N Na P Cl K Ca C O Total

Longitudinal section After

Before

Before After

wt.%

wt.% Sigma

wt.%

wt.% Sigma

wt.%

wt.% Sigma

wt.%

wt.% Sigma

18.19 24.41 4.98 33.27 1.30 17.85

0.72 0.26 0.10 0.33 0.07 0.21

0.32 0.40 0.27 0.28 0.28 7.66 48.68 42.11

0.73 0.03 0.03 0.02 0.02 0.09 0.43 0.39

17.35 6.86 2.01 38.51 2.93 32.34

1.03 0.16 0.12 0.54 0.14 0.48

0.92 0.27 0.25 0.26 0.32 22.57 40.82 34.58 100

1.28 0.05 0.06 0.05 0.06 0.37 0.66 0.59

100

PU-foam

100

C O Al Total C O Al Total

wt.%

wt.% Sigma

67.59 29.29 3.13 100 After 72.61 27.38 0.01 100

0.91 0.92 0.18

0.20 0.20 0.02

Fig. 2. EDX spectra and SEM images of different packing materials (a) Cross section of NSRP-FM; (b) Longitudinal section of NSRP-FM; (c) Section of PU-foam.

the leaching liquid of SA-gel without medium, the same strains could not grow well and the degradation rate of n-butyl acetate was only 3.97 mg L-1 h−1, much smaller than that of the one for NSRP-FM (8.08 mg L-1 h−1). Such results suggested that the prepared NSRP-FM contained more inorganic nutrients, which could release into the leaching liquid and thus maintained the microbial growth. During the microbial biodegradation of organic pollutants, some acidic intermediates are formed and, thus, lower the pH of the surrounding environment [36]. From Fig. 3c, the prepared NSRP-FM could adjust pH value from 5.5 to 7.5 after 1 d for several cycles. The reason might be some compounds (calcium salt) or ions (HPO42−, H2PO4-) contained by NSRP-FM had pH buffering capacities. When they slowly entered into the external solution, they could regulate pH value. Such

S4).Through the comparison of SEM images before and after water spraying, the roughness of the NSRP-FM surface increased and some new holes appeared, implying that it was more conducive to microbial attachment and growth than PU-foam. From the SEM images of NSRPFM and SA-gel without any microorganisms (Fig. S4 b and c), it could not be found any obvious microorganisms covering on the surface of the latter, suggesting the selected strains R. pickettii L2 and A. fumigates HD2 could effectively immobilized on the packing NSRP. In addition to conductivity, the leaching liquid of SA-gel without medium and NSRP-FM were used for the evaluation of nutrient level for microbial growth. From Fig. S5, the inoculated strains grew very well in the leaching liquid of the prepared NSRP-FM, and 200 mg L−1 n-butyl acetate could be completely removed in nearly 24 h. But in the case of 20

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Fig. 3. The release rate and total released amount for P (a) and N (b) and the pH buffer capacity under weak acid conditions (c) of the prepared NSRP-FM.

microorganisms, the storage stability of the microbial activity is much more important. Fig. 4b shows the variation of biomass and degradation ability of NSRP-FM, which was stored for 0, 7, 30 and 60 d. After being preserved at room temperature for 7 d, the biomass contained by the dry NSRP-FM decreased slightly, compared to the freshly prepared material, from 14.61 to 12.51 mg protein·g−1 dry packing material. Meanwhile, the microorganisms fixed were able to effectively degrade n-butyl acetate, with 96% removal efficiency in 36 h. When the storage time was extended to either 30 d or 60 d, the degradation ability remained stable, while the biomass decreased a little (from 11.57 to 11.21 mgprotein g-1 packings).

results suggested that this prepared material had better pH buffer capacity and maintained an optimal pH for microbial growth. 3.3. Microbial properties The effect of packing material on the activity and biodegrading ability of microorganisms is another important consideration [32]. The effects of these two packing materials on the degradation performance of R. pickettii L2 and A. fumigates HD-2 were investigated. From Fig. 4a, adsorption of n-butyl acetate by NSRP-FM or PU-foam was low, about 20 mg L−1 in nearly 36 h. The removal efficiency by A. fumigates HD-2 combined with packing material was much higher than that by A. fumigates HD-2 alone (almost 4 times higher). In the case of bacterium R. pickettii L2, the enhancement by the packing material was not so obvious, with slightly higher removal efficiency (nearly 1.2 times). Bacteria are more inclined to liquid phase growth media, while fungi prefer solid surfaces [28,37]. As a result, the growth and biodegradation activity of R. pickettii L2 in combination with packing material was not as advantageous as that of A. fumigates HD-2. For the packing material with embedded functional

3.4. Biofilter evaluation 3.4.1. Operation performance In order to assess the physicochemical and microbial properties of the prepared NSRP-FM on the removal of a synthetic waste gas, two BTFs packed with NSRP-FM and PU-foam were set up. Fig. 5a displays the performances of these BTFs throughout the operation. Stage I corresponds to the start-up phase. By day 5, both biofilters

Fig. 4. The effects of prepared NSRP on the selected strains’ biodegradable activity to n-butyl acetate (a) and the storage stability of the prepared NSRP-FM (b). 21

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Fig. 5. Evolution of the removal of gaseous n-butyl acetate throughout the operation of BTFs(a), conductivity of the recirculation liquid in stage II (from day 11–31) (b), pH of the recirculation liquid in stage III (from day 31–44) (c), and conductivity and pH measurements of the recirculation liquid in stage IV (from day 45–60) for BTF1 (using NSRP-FM as packings) and BTF2 (using PU-foam as packings) (d).

nutrients, and, therefore, both the microbial activity and RE were influenced to a certain degree. In order to further verify whether the lack of nutrients affected the removal efficiency, the conductivity of recirculation liquid was measured throughout this stage. From Fig. 5b, the conductivity of recirculation liquid in BTF1 decreased from 4000 to 3000 μS cm−1. In BTF2, the conductivity decreased sharply and stabilized at approximately 500 μS cm−1 on day 22. Interestingly, the RE decreased significantly at that time. The biomass in the packing materials was estimated by measuring the protein amount, and the results were 2.98 ± 0.08 and 2.28 ± 0.06 mg protein·g-1 packing for BTF1 and BTF2, respectively. The former was 30% higher than the later in the case of biomass. The pH buffering capacity of the prepared packing material was investigated in Stage III, in which the pH of the recirculation liquid was not adjusted. From Fig. 5a, the RE of BTF1 was much more stabilized than that of BTF2, and the average value was 94%. However, the RE of

removed 90% n-butyl acetate, but the efficiencies were not stabilized. The removal efficiency (RE) of BTF1 was finally stabilized at 98% on day 9, while BTF2 required an additional 3 days to achieve 98%. These results show the starting speed of the biofilter packed with NSRP-FM was slightly faster than that of the one packed with PU-foam. Since the biodegradability for n-butyl acetate was much better, along with the nbutyl acetate-degraders R. pickettii L2 and A. fumigates HD-2, the REs for both BTFs were much higher. In Stage II, the recirculation liquid was replaced with deionized water, and other process parameters were not changed. Both the REs decreased to different extents. At the end of this stage (30 d), quasisteady RE was 94% for BTF1 and 89% for BTF2, respectively. The low RE for BTF2, compared to BTF1, was likely caused by the lack of nutrients. Since the prepared NSRP-FM could release nutrients for microbial growth during operation, the RE was higher and more stable than the BTF with PU-foam. The PU-foam itself did not contain any 22

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Fig. 6. Relative abundances at order level (a) and relative abundances at the genus level (b) for bacteria for different stages of the two BTFs. Stage I from day 0–11, Stage II from day 11–31, Stage III from day 31-44.

BTF2 gradually decreased and finally declined to 65%. At the same time, the biomass amounts on the packing materials were 2.84 ± 0.03 and 1.91 ± 0.04 mg protein·g−1 packing for BTF1 and BTF2, respectively. The sharp decrease in removal efficiency for BTF2 was attributed to two factors: (1) the lack of nutrients and (2) the acidified environment. The prepared NSRP-FM could slowly release some nutrients, some of which also had the pH buffering capacity. As a result, the microorganisms in BTF1 could be able to grow well owing to enough inorganic components and neutral environment, thus not affecting the removal of n-butyl acetate. Starting from day 31, the pH of the recirculation liquid in both biofilters was measured, and the results are shown in Fig. 5c. The pH of the recirculation liquid of BTF2 decreased with time, and an acidified environment (pH 6.5) was formed, which could influence the microbial activity. Conversely, the environment in BTF1 remained neutral. Stage IV was designed to investigate the responses of both biofilters to inlet concentration changes of n-butyl acetate. In this stage, deionized water was used as the recirculation liquid for both BTFs, and pH adjustment was only carried out for BTF2. For BTF1, regardless of concentration, the RE was high (95–99%). However, the RE of BTF2 fluctuated greatly. The initial rise in RE (from 64% to 80%) was attributed to the change in acidified environment. Upon pH adjustment, the environment in BTF2 gradually reached near neutral (50 d, pH 6.9), and thus, some microbial activity recovered with an increase in RE. Subsequent decline in RE was caused by the inlet concentration changes, and the microorganisms were unable to adapt to the substrate variation in a short time. These differences suggested that BTF1 was more stable than BTF2 when the process parameters were changed. Fig. 5d describes the variation of conductivity and pH of recirculation liquid in this stage. The prepared NSRP-FM maintained better pH buffering capacity and nutrient release capacity in BTF1. However, the nutrient level in BTF2 was relative low level (conductivity < 200 μS cm−1). Other studies about the treatment of n-butyl acetate were mostly focused on the mixed waste gas. Ondarts et al. [38] used the biofilter packed with a mixture of compost and activated carbon for the treatment of mixed waste gas containing ∼93 μg m−3 n-butyl acetate, and more than 90% REs were achieved. Wu et al. [39] used a three-segment biofilter packed with the mixture of mature pig compost, forest soil and polyethylene (PE) for the treatment of the mixtures of n-butyl acetate, p-xylene and ammonia gas (NH3). When the concentration was 100–300 mg m−3 at EBRT of 90 s, 100% of n-butyl acetate was

removed, a slight higher than our study (94–96% under the same concentration and EBRT). Although the removal performances were similar among these studies, the biofilter packed with the prepared NSRP-FM did not need nutrients supply or pH adjustment from outside, suggesting the operation and maintenance of such bioreactor would be more convenient. From the SEM images in Fig.S6a, many mycelia are observed, completely covering the surface of NSRP-FM, and behind them some bacteria are seen. Compared with Fig.S6c, there are less mycelia and bacteria covering the PU-foam. These SEM images were consistent with the RE of BTF1, suggesting that there was adequate biomass growing in BTF1 since the biofilm was well structured and not too dense. This allowed better diffusion of nutrients and carbon sources [11,40]. Since the prepared NSRP-FM could release some nutrients during operation, the fungi and bacteria on the packing surface grew very well, even without the supply of external nutrients. However, in BTF2, there was not a continuous supply of nutrients, and thus, the microorganisms could not grow normally. Furthermore, since the pH of circulating liquid was not adjusted artificially in some stages, the growth of bacteria, which preferred a neutral environment [28], was inhibited under acidified conditions. Through the amount difference of elements distributing on the NSRP-FM’s surface, it was testified that some inorganic elements released during the utilization of the prepared packing material (Table 2). The average release for TP and TN were 92% and 97%, respectively. Other elements, especially some metals, were also released. The increase in the abundance of elements C and O might originate from the biomass, since the chemical formula for bacterium and fungus could be expressed as C5H7NO2 [41]. For PU-foam, the types of elements present did not change, and only the abundances varied, which were related to the microbial growth.

3.4.2. Microbial diversity analysis The abundance and diversity of microorganisms in the biofilters from stage I to III were analyzed by pyrosequencing. Supplementary Information S3 gives the raw findings of pyrosequencing. A total of 15 bacterial orders were detected in all samples, with abundances greater than 1%; while the abundance smaller than 1% was classified as “others” (Fig.6a). The order Burkholderiales, which was the upper level for the inoculated genus Ralstonia, was one of the dominant orders during the operation. Although there was no continuous supply of inorganic nutrients in stage II, the remaining inorganic nutrients, which 23

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Table 3 The predominant bacteria at the genus level and their roles in both BTFs. Order

Average relative abundance (%) BTF1

BTF2

Burkholderiales

16.275

13.445

Sphingobacteriales Xanthomonadales

11.725 11.219

5.315 6.491

Rhizobiales Corynebacteriales Flavobacteriales

13.594 12.625 8.717

17.831 11.009 9.984

First or second Genus

Roles

Reference

Ralstonia Acidovorax Ferruginibacter Xanthomonas Rhodanobacter Ochrobactrum Rhodococcus Flavobacterium

inoculated strain acrylic acid, CB Organic water degradation phenol and ammonia BTEX phenol toluene, formaldehyde possesses monooxygenases

– [43–45] [46] [47] [48] [49] [50] [51]

Fig. 7. Relative abundances at order level (a) and relative abundances at the genus level (b) for fungi for different stages of the two BTFs. Stage I from day 0–11, Stage II from day 11–31, Stage III from day 31-44.

growths. Table 3 shows the first or second bacteria types according to their abundances at the genus level, along with their average relative abundances and roles. Acidovorax species are commonly found in organic compound treatment reactors, such as ethanol, methanol, and glucose [43,44]. Wang et al. [45] reported that Acidovorax species grew in weak acid environments and used some acidic substances (e.g. acrylic acid) as their carbon sources. Therefore, the abundance of Acidovorax species in the Stage III sample from BTF2 was much higher than others. Xanthomonas species [47] and Ochrobactrum species [48] are known to degrade phenols and other aromatic compounds, while some Rhodanobacter species [49] and Rhodococcus species [50] have been used to degrade BTEX compounds (toluene, etc.). Moreover, Flavobacterium species possess an important enzyme called monooxygenase, which catalytically oxidizes various organic pollutants [51]. Compared with the bacteria, the number of microorganisms of fungal orders was relatively less (about 6 types) (Fig. 7a). According to their abundances, the first order was Chaetothyriales, with abundances ranging from 64.7% to 85.8% and 41.8% to 70.2% for BTF1 and BTF2, respectively. Some Chaetothyriales species are able to use toluene and other hydrocarbons as the sole carbon source in some growth-limiting surroundings [52]. As a result, in the present study, the abundance of Chaetothyriales species was relatively high regardless of the operating conditions. The second order in BTF1 was Eurotiales. It was the upper level of genus Aspergillus, which suggested that the inoculated fungal species A. fumigates HD2 existed, and its amount decreased to 6.6% and 8.0% for Stage II and III, respectively. The variation in abundances suggests that the growth demand of the inoculated A. fumigates HD2 was strict, and its growth was influenced much more by external

were indirectly reflected by the conductivity of the recirculation liquid, maintained the growth of Ralstonia pickettii L2 and other bacteria. As a result, the removal of n-butyl acetate did not change much more (RE from 94% to 90%) and a higher abundance of Burkholderiales (20.2%) was observed in BTF2. In Stage III, the pH of the recirculation liquid declined gradually to approximately 6.5. Because most bacteria thrive in neutral pH conditions, such as the strain R. pickettii L2 [42], the gradually acidified environment hinders the bacterial growth. Therefore, the abundance of Burkholderiales decreased to 7.7% (which implied the genus belonged to this order also decreased in abundance), and the RE of n-butyl acetate decreased to 66%. But in BTF1, the abundance of Burkholderiales increased continuously from Stage I to Stage III, with no supply of inorganic nutrients and no adjustment of pH. Because the slow-releasing nutrients and pH buffering capacity of NSRP-FM, a more suitable environment for R. pickettii L2 and other functional bacteria was created, despite changes in the external environment. As a result, the RE of n-butyl acetate was high (> 95%). A detailed description of the genus distribution of the order Burkholderiales is provided in Fig. 6b. It was found the Ralstonia, which was the upper level of the inoculated species R. pickettii L2, was ranked the first, according to the greatest abundance in BTF1. However, its abundance decreased sharply and was not the first genus in BTF2 in Stage III. These differences indicate that the pH variation had an obvious effect on the distribution and growth of R. pickettii L2. Since n-butyl acetate could be biodegraded by the strain R. pickettii L2 and A. fumigates HD-2 to acetaldehyde, butanol and some organic acids (determined by GCMS and IC which are supplemented in Fig. S7), all of them would be utilized as the carbon sources for other microbial 24

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environment changes. Compared to the NSRP-FM, the common PUfoam did not have slow-releasing nutrients and pH buffering capacity, and it was unable to resist changes to the environment. As a result, the abundance of Eurotiales in BTF2 was 2% and nearly 0.2% in Stage II and Stage III, respectively. According to the genus distribution bar plot (Fig. 7b), the species A. fumigates was the most abundant among all the samples.

[12] [13] [14]

[15]

4. Conclusions

[16]

A new packing material NSRP-FM was developed for the treatment of VOCs. The NSRP-FM showed appropriate physicochemical properties for packing material, such as density, specific surface area. NSRP-FM exhibited a suitable slow-release rate for nutrients (N, P and others), as well as pH buffering capability, which could provide a better environment for microbial growths and reproduce. In comparison with PUfoam, BTF packed with the NSRP-FM showed higher and more stable removal performance for n-butyl acetate, even when there were no nutrients supplement and no pH adjustment. Microbial diversity analysis showed the inoculated strains in NSRP-FM could grow well during the whole operation. These results suggest that NSRP-FM is an innovative and applicable packing material for waste gas biopurification. The effect of NSRP-FM on the bioaerosols emissions should be evaluated in the future.

[17]

[18]

[19] [20]

[21]

[22]

Acknowledgments [23]

The authors would like to thank the Key Project of Zhejiang Province Natural Science Foundation (LZ17E080001), the National Natural Science Foundation of China (51678528) and Young Talent Cultivation Project of Zhejiang Association for Science and Technology (grant number 2016YCGC014) provided financial supports to the research.

[24]

[25] [26]

Appendix A. Supplementary data

[27]

Supplementary material related to this article can be found, in the online version, at doi:https://doi.org/10.1016/j.jhazmat.2018.11.070.

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