Aerobic dynamic feeding (ADF) strategy was applied in sequencing batch reactor (SBR) to accumulate polyhydroxyalkanoate (PHA) in aerobic granules. The aerobic granules were able to remove 90% of the COD from palm oil mill effluent (POME). The volatile fatty acids (VFAs) in the POME are the sole source of the PHA accumulation. In this work, 100% removal of propionic and butyric acids in the POME were observed. The highest amount of PHA produced in aerobic granules was 0.6833mgPHA/mgbiomass. The PHA formed was identified as a P (hydroxybutyrate-co-hydroxyvalerate) P (HB-co-HV).
The influence of hydraulic retention time (HRT, 24, 12, and 6h) on the physical characteristics of granules and performance of a sequencing batch reactor (SBR) treating rubber wastewater was investigated. Results showed larger granular sludge formation at HRT of 6h with a mean size of 2.0±0.1mm, sludge volume index of 20.1mLg(-1), settling velocity of 61mh(-1), density of 78.2gL(-1) and integrity coefficient of 9.54. Scanning electron microscope analyses revealed different morphology of microorganisms and structural features of granules when operated at various HRT. The results also demonstrated that up to 98.4% COD reduction was achieved when the reactor was operated at low HRT (6h). Around 92.7% and 89.5% removal efficiency was noted for ammonia and total nitrogen in the granular SBR system during the treatment of rubber wastewater.
A fed-batch strategy was established based on the maximum substrate uptake rate (MSUR) of Pseudomonas aeruginosa USM-AR2 grown in diesel to produce rhamnolipid. This strategy matches the substrate feed rates with the substrate demand based on the real-time measurements of dissolved oxygen (DO). The MSUR was estimated by determining the time required for consumption of a known amount of diesel. The MSUR trend paralleled the biomass profile of Ps. aeruginosa USM-AR2, where the MSUR increased throughout the exponential phase indicating active substrate utilization and then decreased when cells entered stationary phase. Rhamnolipid yield on diesel was enhanced from 0·047 (g/g) in batch to 0·110 (g/g) in pulse-pause fed-batch and 0·123 (g/g) in MSUR fed-batch. Rhamnolipid yield on biomass was also improved from 0·421 (g/g) in batch, 3·098 (g/g) in pulse-pause fed-batch to 3·471 (g/g) using MSUR-based strategy. Volumetric productivity increased from 0·029 g l(-1) h(-1) in batch, 0·054 g l(-1) h(-1) in pulse-pause fed-batch to 0·076 g l(-1) h(-1) in MSUR fed-batch.
Sequencing batch reactor (SBR) is one of the various methods of biological treatments used for treating wastewater and landfill leachate. This study investigated the treatment of landfill leachate and domestic wastewater by adding a new adsorbent (powdered ZELIAC; PZ) to the SBR technique. ZELIAC consists of zeolite, activated carbon, lime stone, rice husk ash, and Portland cement. The response surface methodology and central composite design were used to elucidate the nature of the response surface in the experimental design and describe the optimum conditions of the independent variables, including aeration rate (L/min), contact time (h), and ratio of leachate to wastewater mixture (%; v/v), as well as their responses (dependent variables). Appropriate conditions of operating variables were also optimized to predict the best value of responses. To perform an adequate analysis of the aerobic process, four dependent parameters, namely, chemical oxygen demand (COD), color, ammonia-nitrogen (NH3-N), and phenols, were measured as responses. The results indicated that the PZ-SBR showed higher performance in removing certain pollutants compared with SBR. Given the optimal conditions of aeration rate (1.74 L/min), leachate to wastewater ratio (20%), and contact time (10.31 h) for the PZ-SBR, the removal efficiencies for color, NH3-N, COD, and phenols were 84.11%, 99.01%, 72.84%, and 61.32%, respectively.
A Pichia pastoris transformant carrying the cutinase cDNA of Glomerella cingulata was over-expressed in a 5L bioreactor (2.0L working volume) under fed-batch conditions. Bioreactor experiments rely on varying selected parameters in repeated rounds of optimisation: here these included duration of induction, pH and temperature. Highest cell densities (320gL(-1) wet cell weight) with a cutinase production of 3800mgL(-1) and an activity of 434UmL(-1) were achieved 24h after induction with methanol in basal salt medium (at pH 5 and 28°C). Characterisation of the cutinase showed that it was stable between pH 6 and pH 11, had an optimum pH of 8.0 and retained activity for 30min at 50°C (optimum temperature 25°C).The preferred substrates of G. cingulata cutinase were the medium- to long-chain ρ-nitrophenyl esters of ρ-nitrophenylcaprylate (C8), ρ-nitrophenyllaurate (C12) and ρ-nitrophenylmyristate (C14), with the highest catalytic efficiency, kcat/Km of 7.7±0.7mM(-1)s(-1) for ρ-nitrophenylcaprylate. Microscopic analyses showed that the G. cingulata cutinase was also capable of depolymerising the high molecular weight synthetic polyester, polyethylene terephthalate.
A xylanase gene (xyn2) from Trichoderma reesei ATCC 58350 was previously cloned and expressed in Kluyveromyces lactis GG799. The production of the recombinant xylanase was conducted in a developed medium with an optimised batch and with fed-batches that were processed with glucose. The glucose served as a carbon source for cell growth and as an inducer for xylanase production. In a 1-L batch system, a glucose concentration of 20 g L(-1) and 80 % dissolved oxygen were found to provide the best conditions for the tested ranges. A xylanase activity of 75.53 U mL(-1) was obtained. However, in the batch mode, glucose depletions reduced the synthesis of recombinant xylanase by K. lactis GG799. To maximise the production of xylanase, further optimisation was performed using exponential feeding. We investigated the effects of various nitrogen sources combined with the carbon to nitrogen (C/N) molar ratio on the production of xylanase. Of the various nitrogen sources, yeast extract was found to be the most useful for recombinant xylanase production. The highest xylanase production (110.13 U mL(-1)) was measured at a C/N ratio of 50.08. These conditions led to a 45.8 % increase in xylanase activity compared with the batch cultures. Interestingly, the further addition of 500 g L(-1) glucose led to a 6.2-fold increase (465.07 U mL(-1)) in recombinant xylanase activity. These findings, together with those of the exponential feeding strategy, indicate that the composition of the C/N molar ratio has a substantial impact on recombinant protein production in K. lactis.
The effects of azide on electron transport of exoelectrogens were investigated using air-cathode MFCs. These MFCs enriched with azide at the concentration higher than 0.5mM generated lower current and coulomb efficiency (CE) than the control reactors, but at the concentration lower than 0.2mM MFCs generated higher current and CE. Power density curves showed overshoot at higher azide concentrations, with power and current density decreasing simultaneously. Electrochemical impedance spectroscopy (EIS) showed that azide at high concentration increased the charge transfer resistance. These analyses might reflect that a part of electrons were consumed by the anode microbial population rather than transferred to the anode. Bacterial population analyses showed azide-enriched anodes were dominated by Deltaproteobacteria compared with the controls. Based on these results it is hypothesized that azide can eliminate the growth of aerobic respiratory bacteria, and at the same time is used as an electron acceptor/sink.
A novel wastewater treatment system consisting of an up-flow anaerobic sludge blanket (UASB) reactor and a down-flow hanging sponge (DHS) reactor with sulfur-redox reaction was developed for treatment of municipal sewage under low-temperature conditions. In the UASB reactor, a novel phenomenon of anaerobic sulfur oxidation occurred in the absence of oxygen, nitrite and nitrate as electron acceptors. The microorganisms involved in anaerobic sulfur oxidation have not been elucidated. Therefore, in this study, we studied the microbial communities existing in the UASB reactor that probably enhanced anaerobic sulfur oxidation. Sludge samples collected from the UASB reactor before and after sulfur oxidation were used for cloning and terminal restriction fragment length polymorphism (T-RFLP) analysis of the 16S rRNA genes of the bacterial and archaeal domains. The microbial community structures of bacteria and archaea indicated that the genus Smithella and uncultured bacteria within the phylum Caldiserica were the dominant bacteria groups. Methanosaeta spp. was the dominant group of the domain archaea. The T-RFLP analysis, which was consistent with the cloning results, also yielded characteristic fingerprints for bacterial communities, whereas the archaeal community structure yielded stable microbial community. From these results, it can be presumed that these major bacteria groups, genus Smithella and uncultured bacteria within the phylum Caldiserica, probably play an important role in sulfur oxidation in UASB reactors.
The applicability of the enhanced biological phosphorus removal (EBPR) process for the removal of phosphorus in warm climates is uncertain due to frequent reports of EBPR deterioration at temperature higher than 25 °C. Nevertheless, a recent report on a stable and efficient EBPR process at 28 °C has inspired the present study to examine the performance of EBPR at 24 °C-32 °C, as well as the PAOs and GAOs involved, in greater detail. Two sequencing batch reactors (SBRs) were operated for EBPR in parallel at different temperatures, i.e., SBR-1 at 28 °C and SBR-2 first at 24 °C and subsequently at 32 °C. Both SBRs exhibited high phosphorus removal efficiencies at all three temperatures and produced effluents with phosphorus concentrations less than 1.0 mg/L during the steady state of reactor operation. Real-time quantitative polymerase chain reaction (qPCR) revealed Accumulibacter-PAOs comprised 64% of the total bacterial population at 24 °C, 43% at 28 °C and 19% at 32 °C. Based on fluorescent in situ hybridisation (FISH), the abundance of Competibacter-GAOs at both 24 °C and 28 °C was rather low (<10%), while it accounted for 40% of the total bacterial population at 32 °C. However, the smaller Accumulibacter population and larger population of Competibacter at 32 °C did not deteriorate the phosphorus removal performance. A polyphosphate kinase 1 (ppk1)-based qPCR analysis on all studied EBPR processes detected only Accumulibacter clade IIF. The Accumulibacter population shown by 16S rRNA and ppk1 was not significantly different. This finding confirmed the existence of single clade IIF in the processes and the specificity of the clade IIF primer sets designed in this study. Habitat filtering related to temperature could have contributed to the presence of a unique clade. The clade IIF was hypothesised to be able to perform the EBPR activity at high temperatures. The clade's robustness most likely helps it to fit the high-temperature EBPR sludge best and allows it not only to outcompete other Accumulibacter clades but coexist with GAOs without compromising EBPR activity.
Treatment of industrial waste water (e.g. textile waste water, phenol waste water, pharmaceutical etc) faces limitation in conventional treatment procedures. Advanced oxidation processes (AOPs) do not suffer from the limits of conventional treatment processes and consequently degrade toxic pollutants more efficiently. Complexity is faced in eradicating the restrictions of AOPs such as sludge formation, toxic intermediates formation and high requirement for oxidants. Increased mass-transfer in AOPs is an alternate solution to this problem. AOPs combined with Fluidized bed reactor (FBR) can be a potential choice compared to fixed bed or moving bed reactor, as AOP catalysts life-span last for only maximum of 5-10 cycles. Hence, FBR-AOPs require lesser operational and maintenance cost by reducing material resources. The time required for AOP can be minimized using FBR and also treatable working volume can be increased. FBR-AOP can process from 1 to 10 L of volume which is 10 times more than simple batch reaction. The mass transfer is higher thus the reaction time is lesser. For having increased mass transfer sludge production can be successfully avoided. The review study suggests that, optimum particle size, catalyst to reactor volume ratio, catalyst diameter and liquid or gas velocity is required for efficient FBR-AOP systems. However, FBR-AOPs are still under lab-scale investigation and for industrial application cost study is needed. Cost of FBR-AOPs highly depends on energy density needed and the mechanism of degradation of the pollutant. The cost of waste water treatment containing azo dyes was found to be US$ 50 to US$ 500 per 1000 gallons where, the cost for treating phenol water was US$ 50 to US$ 800 per 1000 gallons. The analysis for FBR-AOP costs has been found to depend on the targeted pollutant, degradation mechanism (zero order, 1st order and 2nd order) and energy consumptions by the AOPs.
A sequencing batch reactor (SBR) with a working volume of 8 L and an exchange ratio of 25% was used to enrich biomass for the treatment of the anaerobically treated low pH palm oil mill effluent (POME). The influent concentration was stepwise increased from 5000 ± 500 mg COD/L to 11,500 ± 500 mg COD/L. The performance of the reactor was monitored at different organic loading rates (OLRs). It was found that approximately 90% of the COD content of the POME wastewater was successfully removed regardless of the OLR applied to the SBR. Cycle studies of the SBR show that the oxygen uptake by the biomass while there is no COD reduction may be due to the oxidation of the storage product by the biomass. Further, the growth kinetic parameters of the biomass were determined in batch experiments using respirometer. The maximum specific growth rate (μmax) was estimated to be 1.143 day(-1) while the half saturation constant (Ks) with respect to COD was determined to be 0.429 g COD/L. The decay coefficient (bD) and biomass yield (Y) were found to be 0.131 day(-1) and 0.272 mg biomass/mg COD consumed, respectively.
The locally isolated filamentous fungus Cunninghamella bainieri 2A1 was cultivated in a 5 L bioreactor to produce lipid and gamma-linolenic acid (GLA). The optimization was carried out using response surface methodology based on a central composite design. A statistical model, second-order polynomial model, was adjusted to the experimental data to evaluate the effect of key operating variables, including aeration rate and agitation speed on lipid production. Process analysis showed that linear and quadratic effect of agitation intensity significantly influenced lipid production process (P < 0.01). The quadratic model also indicated that the interaction between aeration rate and agitation speed had a highly significant effect on lipid production (P < 0.01). Experimental results showed that a lipid content of 38.71% was produced in optimum conditions using an airflow rate and agitation speed of 0.32 vvm and 599 rpm, respectively. Similar results revealed that 0.058(g/g) gamma-linolenic acid was produced in optimum conditions where 1.0 vvm aeration rate and 441.45 rpm agitation rate were used. The regression model confirmed that aeration and agitation were of prime importance for optimum production of lipid in the bioreactor.
An innovative approach using soybean residues for the production of bioflocculants through solid-state fermentation was carried out in 4.5 L near-to-adiabatic bioreactors at pilot-scale level. An added inoculum of the strain Bacillus subtilis UPMB13 was tested in comparison with control reactors without any inoculation after the thermophilic phase of the fermentation. The flocculating performances of the extracted bioflocculants were tested on kaolin suspensions, and crude bioflocculants were obtained from 20 g of fermented substrate through ethanol precipitation. The production of bioflocculants was observed to be higher during the death phase of microbial growth. The bioflocculants were observed to be granular in nature and consisted of hydroxyl, carboxyl and methoxyl groups that aid in their flocculating performance. The results show the vast potential of the idea of using wastes to produce bioactive materials that can replace the current dependence on chemicals, for future prospect in water treatment applications.
The biosynthesis of biomedical products including lipid and gamma-linolenic acid (GLA) by Cunninghamella bainieri 2A1 was studied in repeated batch fermentation. Three key process variables, namely, glucose concentration, ammonium tartrate concentration, and harvesting time, were optimized using response surface methodology. Repeated batch fermentation was carried out by the cultivation of Cunninghamella bainieri 2A1 in nitrogen-limited medium with various nitrogen concentration (1-4 g/L) and glucose concentration (20-40 g/L) at three time intervals (12 h, 24 h, and 48 h). Experimental results showed that the highest lipid concentration of 6.2 g/L and the highest GLA concentration of 0.4 g/L were obtained in optimum conditions, where 20.2 g/L glucose, 2.12 g/L ammonium tartrate, and 48 h harvesting time were utilized. Statistical results showed that the interaction between glucose and ammonium tartrate concentration had highly significant effects on lipid and GLA biosynthesis (P < 0.01). Moreover, harvesting time had a significant interaction effect with glucose and ammonium tartrate concentration on lipid production (P < 0.05).
A pilot-scale production of lipase using palm oil mill effluent (POME) as a fermentation basal medium was carried out, and parameters for immobilization of the produced lipase were optimized. Lipase production in a 300-L bioreactor was performed using two proposed strategies, constant power per volume (P/V) and constant tip speed. Moreover, lipase immobilization on different materials was also investigated. Lipase production was performed using liquid-state bioconversion of POME as the medium and Candida cylindracea as the inoculum. The fermentation medium was composed of 1% total suspended solids (TSS) of POME, 0.5% (w/v) peptone, 0.7% (v/v) Tween-80, and 2.2% inoculum. The medium composition was decided on the basis of the medium optimization results of a previous study. The fermentation was carried out for 48 h at 30°C and pH 6. The maximum lipase production was 5.72U/mL and 21.34 U/mL, obtained from the scale-up strategies of constant tip speed and P/V, respectively. Four accessible support materials were screened for their potential use in immobilization. The most suitable support material was found to be activated carbon, with a maximum immobilization of 94%.
In this work, lipase from Candida rugosa was immobilized onto chitosan/graphene oxide beads. This was to provide an enzyme-immobilizing carrier with excellent enzyme immobilization activity for an enzyme group requiring hydrophilicity on the immobilizing carrier. In addition, this work involved a process for the preparation of an enzymatically active product insoluble in a reaction medium consisting of lauric acid and oleyl alcohol as reactants and hexane as a solvent. This product enabled the stability of the enzyme under the working conditions and allowed the enzyme to be readily isolated from the support. In particular, this meant that an enzymatic reaction could be stopped by the simple mechanical separation of the "insoluble" enzyme from the reaction medium. Chitosan was incorporated with graphene oxide because the latter was able to enhance the physical strength of the chitosan beads by its superior mechanical integrity and low thermal conductivity. The X-ray diffraction pattern showed that the graphene oxide was successfully embedded within the structure of the chitosan. Further, the lipase incorporation on the beads was confirmed by a thermo-gravimetric analysis. The lipase immobilization on the beads involved the functionalization with coupling agents, N-hydroxysulfosuccinimide sodium (NHS) and 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC), and it possessed a high enzyme activity of 64 U. The overall esterification conversion of the prepared product was 78% at 60 °C, and it attained conversions of 98% and 88% with commercially available lipozyme and novozyme, respectively, under similar experimental conditions.
Clostridium sp. strain Ade.TY is potentially a new biohydrogen-producing species isolated from landfill leachate sludge. Here we present the assembly and annotation of its genome, which may provide further insights into its gene interactions for efficient biohydrogen production.
Start-up period is considered to be the most unstable and difficult stage in anaerobic process and usually takes a long time due to slow-degree adaptation of anaerobic microorganisms. In order to achieve a shorter start-up period, a novel modified anaerobic baffled reactor (MABR) has been developed in this study, where each modified baffle has its own characteristics (form/shape) to facilitate a treatment ofrecycled paper mill effluent (RPME). The results ofphysico-chemical characteristics showed that effluent from recycled paper mill consisted of 4328mgL-1 chemical oxygen demand (COD), 669mg L-1 biochemical oxygen demand and 501mg L-1 volatile fatty acid. It also consisted of variety of heavy metals such as zinc, magnesium, iron and nickel at concentrations of 1.39, 12.19, 2.39 and 0.72 mgL-1, respectively. Performance of MABR during the start-up period showed that methane production reached 34.7% with COD removal of 85% at steady state. The result indicates that MABR was successfully operated during the start-up period in treating RPME within a period of less than 30 days.
Clostridium bifermentans strain WYM is an effective biohydrogen producer isolated from landfill leachate sludge. Here, we present the assembly and annotation of its genome, which may provide further insights into the metabolic pathways involved in efficient biohydrogen production.