Natural mycelial exopolysaccharide (EPS) and endopolysaccharide (ENS) extracted from bioreactor-cultivated European Ganoderma applanatum mushrooms are of potential high commercial value for both food and adjacent biopharmaceutical industries. In order to evaluate their potential toxicity for aquaculture application, both EPS (0.01-10 mg/mL) and ENS (0.01-10 mg/mL) extracts were tested for Zebrafish Embryo Toxicity (ZFET); early development effects on Zebrafish Embryos (ZE) were also analyzed between 24 and 120 h post-fertilization (HPF). Both EPS and ENS are considered non-toxic with LC50 of 1.41 mg/mL and 0.87 mg/mL respectively. Both EPS and ENS did not delay hatching and teratogenic defect towards ZE with <1.0 mg/mL, respectively. No significant changes in the ZE heart rate were detected following treatment with the two compounds tested (EPS: 0.01-10 mg/mL: 176.44 ± 0.77 beats/min and ENS: 0.01-10 mg/mL: 148.44 ± 17.75 beats/min) compared to normal ZE (120-180 beats/min). These initial findings support future pre-clinical trials in adult fish models with view to safely using EPS and ENS as potential feed supplements for supplements for development of the aquaculture industry.
This study focuses on the evaluation of the performance of Chlorella sp. in removing nutrient in aquaculture wastewater and its correlation with the kinetic growth of Chlorella sp. The treatment was applied with various Chlorella sp. inoculation dosage ranging from 0% to 60% (v/v) of wastewater. The optimum inoculation dosage was recorded at 30% (v/v) with effluent concentration of ammonia and orthophosphate recording at 0.012mgL(-1) and 0.647mgL(-1), respectively on Day 11. The optimum dosage for bio-flocculation process was obtained at 30mgL(-1) of Aspergillus niger with a harvesting efficiency of 97%. This type of development of phytoremediation with continuous bio-harvesting could promote the use of sustainable green technology for effective wastewater treatment.
Illumination factors such as length of photoperiod and intensity can affect growth of microalgae and lipid content. In order to optimize microalgal growth in mass culture system and lipid content, the effects of light intensity and photoperiod cycle on the growth of the marine microalgae, Nannochloropsis sp. were studied in batch culture. Nannochloropsis sp. was grown aseptically for 9 days at three different light intensities (50, 100 and 200 μmol m(-2) s(-1)) and three different photoperiod cycles (24:0, 18:06 and 12:12 h light:dark) at 23 °C cultivation temperature. Under the light intensity of 100 μmol m(-2) s(-1) and photoperiod of 18 h light: 6 h dark cycle, Nannochloropsis sp. was found to grow favorably with a maximum cell concentration of 6.5×10(7) cells mL(-1), which corresponds to the growth rate of 0.339 d(-1) after 8 day cultivation and the lipid content was found to be 31.3%.
Exploring novel biological anti-quorum sensing (QS) agents to control membrane biofouling is of great worth in order to allow sustainable performance of membrane bioreactors (MBRs) for wastewater treatment. In recent studies, QS inhibitors have provided evidence of alternative route to control membrane biofouling. This study investigated the role of Piper betle extract (PBE) as an anti-QS agent to mitigate membrane biofouling. Results demonstrated the occurrence of the N-acyl-homoserine-lactone (AHL) autoinducers (AIs), correlate QS activity and membrane biofouling mitigation. The AIs production in bioreactor was confirmed using an indicator strain Agrobacterium tumefaciens (NTL4) harboring plasmid pZLR4. Moreover, three different AHLs were found in biocake using thin layer chromatographic analysis. An increase in extracellular polymeric substances (EPS) and transmembrane pressure (TMP) was observed with AHL activity of the biocake during continuous MBR operation, which shows that membrane biofouling was in close relationship with QS activity. PBE was verified to mitigate membrane biofouling via inhibiting AIs production. SEM analysis further confirmed the effect of PBE on EPS and biofilm formation. These results exhibited that PBE could be a novel agent to target AIs for mitigation of membrane biofouling. Further work can be carried out to purify the active compound of Piper betle extract to target the QS to mitigate membrane biofouling.
Poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] copolymer is noted for its high biocompatibility, which makes it an excellent candidate for biopharmaceutical applications. The wild-type Cupriavidus sp. USMAA1020 strain is able to synthesize P(3HB-co-4HB) copolymers with different 4HB monomer compositions (up to 70mol%) in shaken flask cultures. Combinations of 4HB carbon precursors consisting of 1,6-hexanediol and γ-butyrolactone were applied for the production of P(3HB-co-4HB) with different 4HB molar fraction. A sharp increase in 4HB monomer composition was attained by introducing additional copies of PHA synthase gene (phaC), responsible for P(3HB-co-4HB) polymerization. The phaC of Cupriavidus sp. USMAA1020 and Cupriavidus sp. USMAA2-4 were cloned and heterologously introduced into host, wild-type Cupriavidus sp. USMAA1020. The gene dosage treatment resulted in the accumulation of 93mol% 4HB by the transformant strains when grown in similar conditions as the wild-type USMAA1020. The PHA synthase activities for both transformants were almost two-fold higher than the wild-type. The ability of the transformants to produce copolymers with high 4HB monomer composition was also tested in large scale production system using 5L and 30L bioreactors with a constant oxygen mass transfer rate. The 4HB monomer composition could be maintained at a range of 83-89mol%. The mechanical and thermal properties of copolymers improved with increasing 4HB monomer composition. The copolymers produced could be tailored for specific biopharmaceutical applications based on their properties.
Microbial electrolysis cells (MECs) are perceived as a potential and promising innovative biotechnological tool that can convert carbon-rich waste biomass or wastewater into hydrogen (H2) or other value-added chemicals. Undesired methane (CH4) producing H2 sinks, including methanogens, is a serious challenge faced by MECs to achieve high-rate H2 production. Methanogens can consume H2 to produce CH4 in MECs, which has led to a drop of H2 production efficiency, H2 production rate (HPR) and also a low percentage of H2 in the produced biogas. Organized inference related to the interactions of microbes and potential processes has assisted in understanding approaches and concepts for inhibiting the growth of methanogens and profitable scale up design. Thus, here in we review the current developments and also the improvements constituted for the reduction of microbial H2 losses to methanogens. Firstly, the greatest challenge in achieving practical applications of MECs; undesirable microorganisms (methanogens) growth and various studied techniques for eliminating and reducing methanogens activities in MECs were discussed. Additionally, this extensive review also considers prospects for stimulating future research that could help to achieve more information and would provide the focus and path towards MECs as well as their possibilities for simultaneously generating H2 and waste remediation.
Aerobic granular sludge has a number of advantages over conventional activated sludge flocs, such as cohesive and strong matrix, fast settling characteristic, high biomass retention and ability to withstand high organic loadings, all aspects leading towards a compact reactor system. Still there are very few studies on the strength of aerobic granules. A procedure that has been used previously for anaerobic granular sludge strength analysis was adapted and used in this study. A new coefficient was introduced, called a stability coefficient (S), to quantify the strength of the aerobic granules. Indicators were also developed based on the strength analysis results, in order to categorize aerobic granules into three levels of strength, i.e. very strong (very stable), strong (stable) and not strong (not stable). The results indicated that aerobic granules grown on acetate were stronger (high density: >150 g T SSL(-1) and low S value: 5%) than granules developed on sewage as influent. A lower value of S indicates a higher stability of the granules.
Thermophilic treatment of palm oil mill effluent (POME) was studied in a novel integrated anaerobic-aerobic bioreactor (IAAB). The IAAB was subjected to a program of steady-state operation over a range of organic loading rate (OLR)s, up to 30 g COD/L day in order to evaluate its treatment capacity. The thermophilic IAAB achieved high chemical oxygen demand (COD), biochemical oxygen demand (BOD) and total suspended solids (TSS) removal efficiencies of more than 99% for OLR up to 18.5 g COD/L day. High methane yield of 0.32 LCH(4) (STP)/g COD(removed) with compliance of the final treated effluent to the discharge limit were achieved. This is higher than that of the mesophilic system due to the higher maximum specific growth rate (μ(max)) of the thermophilic microorganisms. Besides, coupling the model of Grau second order model (anaerobic system) with the model of Monod (aerobic system) will completely define the IAAB system.
The performance of a compost biofilter inoculated with mixed microbial consortium was optimized for treating a gas-phase mixture of benzene and toluene. The biofilter was acclimated to these VOCs for a period of ∼18d. The effects of concentration and flow rate on the removal efficiency (RE) and elimination capacity (EC) were investigated by varying the inlet concentration of benzene (0.12-0.95g/m(3)), toluene (0.14-1.48g/m(3)) and gas-flow rate (0.024-0.072m(3)/h). At comparable loading rates, benzene removal in the mixture was reduced in the range of 6.6-41% in comparison with the individual benzene degradation. Toluene removal in mixture was even more affected as observed from the reductions in REs, ranging from 18.4% to 76%. The results were statistically interpreted by performing an analysis of variance (ANOVA) to elucidate the main and interaction effects.
Solid-state fermentation (SSF) was employed to enhance the nutritive values of palm kernel cake (PKC) for poultry feeding. Aspergillus flavus was isolated from local PKC and utilized to increase the mannose content of PKC via the degradation of β-mannan in PKC; evaluation was done for batch SSF in Erlenmeyer flasks and in a novel laterally aerated moving bed (LAMB) bioreactor. The optimum condition for batch SSF in flasks was 110% initial moisture content, initial pH 6.0, 30 °C, 855 μm particle size, and 120 h of fermentation, yielding 90.91 mg mannose g⁻¹ dry PKC (5.9-fold increase). Batch SSF in the LAMB at the optimum condition yielded 79.61 mg mannose g⁻¹ dry PKC (5.5-fold increase) within just 96 h due to better heat and mass transfer when humidified air flowed radially across the PKC bed. In spite of a compromise of 12% reduction in mannose content when compared with the flasks, the LAMB facilitated good heat and mass transfer, and improved the mannose content of PKC in a shorter fermentation period. These attributes are useful for batch production of fermented PKC feed in an industrial scale.
Biological removal of carbon, nitrogen and sulfur is drawing increasing research interest in search for an efficient and cost-effective wastewater treatment. While extensive work on separate removal of nitrogen and sulfur is well documented, investigation on simultaneous denitrifying sulfide removal has only been reported recently. Most of the work on denitrifying sulfide removal has been focusing on bioreactor performance, loading and operating conditions. Nonetheless, underlying principles elucidating the biochemical reactions and the mechanisms of the microbial degradation are yet to be established. In addition, unstable denitrifying sulfide removal which is a major operating problem that hinders practical application of the process, is yet to be resolved. This paper provides a review on the state-of-the-art development of simultaneous biological removal of sulfur, nitrogen and carbon. Research on bioreactor operation and performance, reactor configurations, mechanisms and modeling work including the use of mass balance analysis and artificial neural networks is delineated. An in-depth discussion on the microbial community and functional consortium is also provided. Challenges and future work on simultaneous biological removal of nitrogen-sulfur-carbon are also outlined.
Sulfide inhibition to nitrifying bacteria has prevented the integration of digestate nitrification and biogas desulfurization to simplify anaerobic digestion systems. In this study, liquid digestate with NaHS solution was treated using nitrifying sludge in a sequential-batch reactor with a long fill period, with an ammonium loading rate of 293 mg-N L-1 d-1 and a stepwise increase in the sulfide loading rate from 0 to 32, 64, 128, and 256 mg-S L-1 d-1. Batch bioassays and microbial community analysis were also conducted with reactor sludge under each sulfide loading rate to quantify the microbial acclimatization to sulfide. In the reactor, sulfide was completely removed. Complete nitrification was maintained up to a sulfide load of 128 mg-S L-1 d-1, which is higher than that in previous reports and sufficient for biogas treatment. In the batch bioassays, the sulfide tolerance of NH4+ oxidizing activity (the 50% inhibitory sulfide concentration) increased fourfold over time with the compositional shift of nitrifying bacteria to Nitrosomonas nitrosa and Nitrobacter spp. However, the sulfur removal rate of the sludge slightly decreased, although the abundance of the sulfur-oxidizing bacteria Hyphomicrobium increased by 30%. Therefore, nitrifying sludge was probably acclimatized to sulfide not by the increasing sulfide removal rate but rather by the increasing nitrifying bacteria, which have high sulfide tolerance. Successful simultaneous nitrification and desulfurization were achieved using a sequential-batch reactor with a long fill period, which was effective in facilitating the present acclimatization.
Moving bed sequencing batch reactors (MBSBRs) packed with 8% (v/v) of 8-, 27- and 64-mL polyurethane (PU) foam cubes, respectively, were investigated for simultaneous 4-chlorophenol (4-CP) and nitrogen removal at increasing 4-CP concentration. When the 4-CP concentration exceeded 300 mg L(-1), the MBSBR with 27-mL foam cubes was observed to outperform the other MBSBRs in removing 4-CP and nitrogen. The reasons were: (1) there were more biomass in inner layer of the 27-mL cubes, compared to that of the 8-mL cubes, which was more shielded from the inhibitory effect of 4-CP and (2) the 27-mL cubes were more mobile than the 64-mL cubes. Although increasing 4-CP concentration to 600 mg L(-1) resulted in incomplete removal of 4-CP in the MBSBRs, results of the batch reactor with 27-mL foam cubes showed that complete 4-CP removal within the REACT period could be achieved by increasing the packing volume to 20%.
Central composite design (CCD) and response surface methodology (RSM) were employed to optimize four important variables, i.e. amounts of oil, bacterial inoculum, nitrogen and phosphorus, for the removal of selected n-alkanes during bioremediation of weathered crude oil in coastal sediments using laboratory bioreactors over a 60 day experimentation period. The reactors contained 1 kg soil with different oil, microorganisms and nutrients concentrations. The F Value of 26.89 and the probability value (P < 0.0001) demonstrated significance of the regression model. For crude oil concentration of 2, 16 and 30 g per kg sediments and under optimized conditions, n-alkanes removal was 97.38, 93.14 and 90.21% respectively. Natural attenuation removed 30.07, 25.92 and 23.09% n-alkanes from 2, 16 and 30 g oil/kg sediments respectively. Excessive nutrients addition was found to inhibit bioremediation.
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).
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.
The purpose of this study was to evaluate the feasibility of producing bioethanol from palm-oil mill effluent generated by the oil-palm industries through direct bioconversion process. The bioethanol production was carried out through the treatment of compatible mixed cultures such as Thrichoderma harzianum, Phanerochaete chrysosporium, Mucor hiemalis, and yeast, Saccharomyces cerevisiae. Simultaneous inoculation of T. harzianum and S. cerevisiae was found to be the mixed culture that yielded the highest ethanol production (4% v/v or 31.6 g/l). Statistical optimization was carried out to determine the operating conditions of the stirred-tank bioreactor for maximum bioethanol production by a two-level fractional factorial design with a single central point. The factors involved were oxygen saturation level (pO(2)%), temperature, and pH. A polynomial regression model was developed using the experimental data including the linear, quadratic, and interaction effects. Statistical analysis showed that the maximum ethanol production of 4.6% (v/v) or 36.3 g/l was achieved at a temperature of 32 degrees C, pH of 6, and pO(2) of 30%. The results of the model validation test under the developed optimum process conditions indicated that the maximum production was increased from 4.6% (v/v) to 6.5% (v/v) or 51.3 g/l with 89.1% chemical-oxygen-demand removal.
Bio-hydrogen production from wastewater using sludge as inoculum is a sustainable approach for energy production. This study investigated the influence of initial pH and temperature on bio-hydrogen production from dairy wastewater using pretreated landfill leachate sludge (LLS) as an inoculum. The maximum yield of 113.2 ± 2.9 mmol H2/g chemical oxygen demand (COD) (12.8 ± 0.3 mmol H2/g carbohydrates) was obtained at initial pH 6 and 37 °C. The main products of volatile fatty acids were acetate and butyrate with the ratio of acetate:butyrate was 0.4. At optimum condition, Gibb's free energy was estimated at -40 kJ/mol, whereas the activation enthalpy and entropy were 65 kJ/mol and 0.128 kJ/mol/l, respectively. These thermodynamic quantities suggest that bio-hydrogen production from dairy wastewater using pretreated LLS as inoculum was effective and efficient. In addition, genomic and bioinformatics analyses were performed in this study.
In this study, endoglucanase was produced from oil palm empty fruit bunch (OPEFB) by a locally isolated aerobic bacterium, Bacillus pumilus EB3. The effects of the fermentation parameters such as initial pH, temperature, and nitrogen source on the endoglucanase production were studied using carboxymethyl cellulose (CMC) as the carbon source. Endoglucanase from B. pumilus EB3 was maximally secreted at 37 degrees C, initial pH 7.0 with 10 g/l of CMC as carbon source, and 2 g/l of yeast extract as organic nitrogen source. The activity recorded during the fermentation was 0.076 U/ml. The productivity of the enzyme increased twofold when 2 g/l of yeast extract was used as the organic nitrogen supplement as compared to the non-supplemented medium. An interesting finding from this study is that pretreated OPEFB medium showed comparable results to CMC medium in terms of enzyme production with an activity of 0.063 U/ml. As OPEFB is an abundant solid waste at palm oil mills, it has the potential of acting as a substrate in cellulase production.