This study reports an efficient fed-batch strategy to improve poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-4-hydroxybutyrate) [P(3HB-co-3HV-co-4HB)] terpolymer production by Cupriavidus sp. USMAA2-4 with enhanced mechanical properties in bioreactor. The cultivations have been performed by combining oleic acid with γ-butyrolactone at different concentration ratios with 1-pentanol at a fixed concentration. The batch and fed-batch fermentations have resulted in P(3HB-co-3HV-co-4HB) with compositions of 9-35 mol% 3HV and 4-24 mol% 4HB monomers. The DO-stat fed-batch fermentation strategies have significantly improved the production with a maximum 4.4-fold increment of cell dry weight (CDW). Besides, appropriate feeding of the substrates has resulted in an increment of terpolymer productivity from 0.086-0.347 g/L/h, with a significantly shortened cultivation time. The bacterial growth and terpolymer formation have been found to be affected by the concentration of carbon sources supplied. Characterization of P(3HB-co-3HV-co-4HB) has demonstrated that incorporation of 3HV and 4HB monomer has significantly improved the physical and thermodynamic properties of the polymers, by reducing the polymer's crystallinity. The tensile strength, Young's modulus of the terpolymer has been discovered to increase with the increase of M w. The fed-batch fermentation strategies employed in this study have resulted in terpolymers with a range of flexible materials having improved tensile strength and Young's modulus as compared to the terpolymer produced from batch fermentation. Possession of lower melting temperature indicates an enhanced thermal stability which broadens the polymer processing window.
Microbial strain optimization focuses on improving technological properties of the strain of microorganisms. However, the complexities of the metabolic networks, which lead to data ambiguity, often cause genetic modification on the desirable phenotypes difficult to predict. Furthermore, vast number of reactions in cellular metabolism lead to the combinatorial problem in obtaining optimal gene deletion strategy. Consequently, the computation time increases exponentially with the increase in the size of the problem. Hence, we propose an extension of a hybrid of Bees Algorithm and Flux Balance Analysis (BAFBA) by integrating OptKnock into BAFBA to validate the result. This paper presents a number of computational experiments to test on the performance and capability of BAFBA. Escherichia coli, Bacillus subtilis and Clostridium thermocellum are the model organisms in this paper. Also included is the identification of potential reactions to improve the production of succinic acid, lactic acid and ethanol, plus the discussion on the changes in the flux distribution of the predicted mutants. BAFBA shows potential in suggesting the non-intuitive gene knockout strategies and a low variability among the several runs. The results show that BAFBA is suitable, reliable and applicable in predicting optimal gene knockout strategy.
Green procedure for synthesizing silver nanoparticles (AgNPs) is currently considered due to its economy and toxic-free effects. Several existing works on synthesizing AgNPs using leaves extract still involve the use of physical or mechanical treatment such as heating or stirring, which consume a lot of energy. To extend and explore the green extraction philosophy, we report here the synthesis and antibacterial evaluations of a purely green procedure to synthesize AgNPs using Carica papaya, Manihot esculenta, and Morinda citrifolia leaves extract without the aforementioned additional treatment. The produced AgNPs were characterized using the ultraviolet-visible spectroscopy, field emission scanning electron microscopy, energy-dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, and antibacterial investigations. For antibacterial tests, two bacteria namely Escherichia coli and Bacillus cereus were selected. The presently employed method has successfully produced spherical AgNPs having sizes ranging from 9 to 69 nm, with plasmonic characteristics ranging from 356 to 485 nm, and energy-dispersive X-ray peak at approximately 3 keV. In addition, the smallest particles can be produced when Manihot esculenta leaves extract was applied. Moreover, this study also confirmed that both the leaves and synthesized AgNPs exhibit the antibacterial capability, depending on their concentration and the bacteria type.
Oil palm fronds are the most abundant lignocellulosic biomass in Malaysia. In this study, fronds were tested as the potential renewable biomass for ethanol production. The soaking in aqueous ammonia pretreatment was applied, and the fermentability of pretreated fronds was evaluated using simultaneous saccharification and fermentation. The optimal pretreatment conditions were 7 % (w/w) ammonia, 80 °C, 20 h of pretreatment, and 1:12 S/L ratio, where the enzymatic digestibility was 41.4 % with cellulase of 60 FPU/g-glucan. When increasing the cellulase loading in the hydrolysis of pretreated fronds, the enzymatic digestibility increased until the enzyme loading reached 60 FPU/g-glucan. With 3 % glucan loading in the SSF of pretreated fronds, the ethanol concentration and yield based on the theoretical maximum after 12 and 48 h of the SSF were 7.5 and 9.7 g/L and 43.8 and 56.8 %, respectively. The ethanol productivities found at 12 and 24 h from pretreated fronds were 0.62 and 0.36 g/L/h, respectively.
Rice bran (RB) and de-oiled rice bran (DRB) have been treated and used as the carbon source in acetone-butanol-ethanol (ABE) production using Clostridium saccharoperbutylacetonicum N1-4. The results showed that pretreated DRB produced more ABE than pretreated RB. Dilute sulfuric acid was the most suitable treatment method among the various pretreatment methods that were applied. The highest ABE obtained was 12.13 g/L, including 7.72 g/L of biobutanol, from sulfuric acid. The enzymatic hydrolysate of DRB (ESADRB), when treated with XAD-4 resin, resulted in an ABE productivity and yield of 0.1 g/L h and 0.44 g/g, respectively. The results also showed that the choice of pretreatment method for RB and DRB is an important factor in butanol production.
Armillaria sp. F022 is a white-rot fungus isolated from a tropical rain forest in Indonesia that is capable of utilizing pyrene as a source of carbon and energy. Enzymes production during the degradation process by Armillaria sp. F022 was certainly related to the increase in biomass. In the first week after incubation, the growth rate rapidly increased, but enzyme production decreased. After 7 days of incubation, rapid growth was observed, whereas, the enzymes were produced only after a good amount of biomass was generated. About 63 % of pyrene underwent biodegradation when incubated with this fungus in a liquid medium on a rotary shaker (120 rpm, 25 °C) for 30 days; during this period, pyrene was transformed to five stable metabolic products. These metabolites were extracted in ethyl acetate, isolated by column chromatography, and then identified using thin layer chromatography (TLC) and gas chromatography-mass spectrometry (GC-MS). 1-Hydroxypyrene was directly identified by GC-MS, while 4-phenanthroic acid, 1-hydroxy-2-naphthoic acid, phthalic acid, and protocatechuic acid were identified to be present in their derivatized forms (methylated forms and silylated forms). Protocatechuic acid was the end product of pyrene degradation by Armillaria sp. F022. Dynamic profiles of two key enzymes, namely laccase and 1,2-dioxygenase, were revealed during the degradation process, and the results indicated the presence of a complicated mechanism in the regulation of pyrene-degrading enzymes. In conclusion, Armillaria sp. F022 is a white-rot fungus with potential for application in the degradation of polycyclic aromatic hydrocarbons such as pyrene in the environment.
Herein, we systematically reported the capability of T. harzianum RY44 for decolorization of Mordant orange-1. The fungi strains were isolated from the Universiti Teknologi Malaysia tropical rain forest. For initial screening, the decolorization was conducted using 50 strains of the fungi for 20 days incubation time and the best performance was selected. Then, the decolorization capability and fungal biomass were evaluated using different dye concentrations, namely, 0, 50, 75 and 100 ppm. Effects of the carbon sources (fructose, glucose, and galactose), nitrogen sources (ammonium nitrate, ammonium sulfate and yeast extract), surfactant (tween 80), aromatic compounds (benzoic acid, catechol and salicylic acid), and pH on the decolorization efficiency were examined. This study has found that the employed carbon sources, nitrogen sources, and aromatic compounds strongly enhance the decolorization efficiency. In addition, increasing the surfactant volume and pH generally decreased the decolorization efficiencies from 19.5 to 9.0% and 81.7 to 60.5%, respectively. In the mechanism philosophy, the present work has found that Mordant orange-1 were initially degraded by T. harzianum RY44 to benzoic acid and finally transformed into salicylic acid.
Polycyclic aromatic hydrocarbon is a toxic recalcitrant environmental pollutant and its removal from the environment is very essential. In this study, a novel S1 strain isolated from the tropical rain forest was identified as Candida species based on 18S rRNA. The pyrene biodegradation was performed by Candida sp. S1. Pyrene was 35% degraded in 15 days. The percentage of pyrene biodegradation increased up to 75% with 24 g L-1of sodium chloride and decreased along with increasing salinity. Under the acidic condition, the biodegradation was increased up to 60% at pH 5. It was also found that the increasing glucose concentration of more than 10 g L-1had no significant effect on pyrene biodegradation, while agitation proved to have greater influence. There was a positive relationship between biomass growth and biodegradation rate of pyrene. One pyrene metabolite was identified from the extract solution and analyzed by a thin-layer chromatography, UV-visible absorption and gas chromatography-mass spectrometry. The metabolite found in the pyrene degradation was benzoic acid. Suitable conditions must be found to promote a successful microbial augmentation in liquid culture.
Anaerobic treatment processes to remove organic matter from palm oil mill effluent (POME) have been used widely in Malaysia. Still the amounts of total organic and total mineral released from POME that may cause degradation of the receiving environment need to be verified. This paper proposes the use of the hydrodynamic equations to estimate performance of the cascaded anaerobic ponds (CAP) and to calculate amounts of total organic matter and total mineral released from POME. The CAP efficiencies to remove biochemical oxygen demands, chemical oxygen demands, total solids and volatile solids (VS) as high as 94.5, 93.6, 96.3 and 98.2 %, respectively, are estimated. The amounts of total organic matter and total mineral as high as 538 kg VS/day and 895 kg FS/day, respectively, released from POME to the receiving water are calculated. The implication of the proposed hydrodynamic equations contributes to more versatile environmental assessment techniques, sometimes replacing laboratory analysis.
Polycyclic aromatics hydrocarbons (PAHs) are ubiquitous and toxic pollutants that are dangerous to humans and living organism in aquatic environment. Normally, PAHs has lower molecular weight such as phenanthrene and naphthalene that are easy and efficient to degrade, but high-molecular-weight PAHs such as chrysene and pyrene are difficult to be biodegraded by common microorganism. This study investigated the isolation and characterization of a potential halophilic bacterium capable of utilizing two high-molecular-weight PAHs. At the end of the experiment (25-30 days of incubation), bacterial counts have reached a maximum level (over 40 × 1016 CFU/mL). The highest biodegradation rate of 77% of chrysene in 20 days and 92% of pyrene in 25 days was obtained at pH 7, temperature 25 °C, agitation of 150 rpm and Tween 80 surfactant showing to be the most impressive parameters for HMWPAHs biodegradation in this research. The metabolism of initial compounds revealed that Hortaea sp. B15 utilized pyrene to form phthalic acid while chrysene was metabolized to form 1-hydroxy-2-naphthoic acid. The result showed that Hortaea sp. B15 can be promoted for the study of in situ biodegradation of high molecular weight PAH.
Due to environmental concern, the research to date has tended to focus on how textile dye removal can be carried out in a greener manner. Therefore, this study aims to evaluate the decolorization and biotransformation pathway of Mordant Orange-1 (MO-1) by Cylindrocephalum aurelium RY06 (C. aurelium RY06). Decolorization study was conducted in a batch experiment including the investigation of the effects of physio-chemical parameters. Enzymatic activity of C. aurelium RY06 during the decolorization was also investigated. Moreover, transformation and biodegradation of MO-1 by C. aurelium RY06 were observed using the gas chromatography-mass spectrometry. Manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase, and 2,3-dioxygenase enzymes were detected during the decolorization. In general, the present work concluded that the MO-1 was successfully degraded by C. aurelium RY06 and transformed to be maleic acid and to be isophtalic acid.
Polyporus sp. S133 decolorized the Amaranth in 72 h (30 mg L(-1)) under static and shaking conditions. Liquid medium containing glucose has shown the highest decolorization of Amaranth by Polyporus sp. S133. When the effect of increasing inoculum concentration on decolorization of Amaranth was studied, maximum decolorization was observed with 15 % inoculum concentration. Significant increase in the enzyme production of laccase (102.2 U L(-1)) was observed over the period of Amaranth decolorization compared to lignin peroxidase and manganese peroxidase. Germination rate of Sorghum vulgare and Triticum aestivum was less with Amaranth treatment as compared to metabolites obtained after its decolorization. Based on the metabolites detected by GC-MS, it was proposed that Amaranth was bio-transformed into two intermediates, 1-hydroxy-2-naphthoic acid and 1,4-naphthaquinone. Overall findings suggested the ability of Polyporus sp. S133 for the decolorization of azo dye and ensured the ecofriendly degradation of Amaranth.
A lipase catalysed enantioselective hydrolysis process under in situ racemization of the remaining (R)-ibuprofen ester substrate with sodium hydroxide as the catalyst was developed for the production of S-ibuprofen from (R,S)-ibuprofen ester in isooctane. Detailed investigations on parameters study indicated that 0.5 M NaOH, addition of 20% (v/v) co-solvent (dimethyl sulphoxide), operating temperature of 45 degrees C, and 40 mmol/L substrate gave 86% conversion and 99.4% optical purity of S-ibuprofen in dynamic kinetic resolution. Meanwhile, in common enzymatic kinetic resolution process, only 42% conversion of the racemate and 93% enantiomeric excess of the product was obtained which are of lower values as compared to dynamic kinetic resolution. The S-ibuprofen produced during each process was evaluated and approximately 50% increment in concentration of S-acid product was produced when dynamic kinetic resolution was applied into the process.
Anode biofilm is a crucial component in microbial fuel cells (MFCs) for electrogenesis. Better knowledge about the biofilm development process on electrode surface is believed to improve MFC performance. In this study, double-chamber microbial fuel cell was operated with diluted POME (initial COD = 1,000 mg L(-1)) and polyacrylonitrile carbon felt was used as electrode. The maximum power density, COD removal efficiency and Coulombic efficiency were found as 22 mW m(-2), 70 and 24 %, respectively. FTIR and TGA analysis confirmed the formation of biofilm on the electrode surface during MFC operation. The impact of anode biofilm on anodic polarization resistance was investigated using electrochemical impedance spectroscopy (EIS) and microbial community changes during MFC operation using denaturing gradient gel electrophoresis (DGGE). The EIS-simulated results showed the reduction of charge transfer resistance (R ct) by 16.9 % after 14 days of operation of the cell, which confirms that the development of the microbial biofilm on the anode decreases the R ct and therefore improves power generation. DGGE analysis showed the variation in the biofilm composition during the biofilm growth until it forms an initial stable microbial community, thereafter the change in the diversity would be less. The power density showed was directly dependent on the biofilm development and increased significantly during the initial biofilm development period. Furthermore, DGGE patterns obtained from 7th and 14th day suggest the presence of less diversity and probable functional redundancy within the anodic communities possibly responsible for the stable MFC performance in changing environmental conditions.
The effects of structure and concentration of surfactants on the biodegradation of fluoranthene, a three rings polycyclic aromatic hydrocarbon in the aqueous phase, as well as their effects on the biodegradation and enzyme activity were investigated. The toxicity ranking of studied surfactants is: non-ionic Tween 80 4,500 mg/L) was showed by Tween 80 (10 mg/L) culture, manifesting that the non-ionic surfactant present in the culture were beneficial to the fungal growth. Laccase showed the highest enzymes activity in all surfactants culture. Non-ionic Tween 80 showed a significant result for laccase activity (1,902 U/L) in the Armillaria sp. F022 culture. The increased enzymes cumulative activity may stem directly from the rising fluoranthene biodegradability as addition of appropriate surfactants. The biotransformation of fluoranthene was greatly improved by Tween 80, and totally fluoranthene degradation was obtained as Tween 80 was 10 mg/L. Two fluoranthene metabolites were isolated from the culture medium and analyzed by a thin layer chromatography, UV visible spectrometer and gas chromatography-mass spectrometry (GC-MS). The oxidation of fluoranthene is initiated by oxygenation at the C-2,3 positions resulting 9-fluorenone. At the end of experiment, one metabolite was detected in the culture extract and identified as phthalic acid. Evidently, Armillaria sp. F022 seems efficient, high effective and deserves further application on the enhanced bioremediation technologies for the treatment of fluoranthene-contaminated soil.
This paper demonstrates Pseudomonas cepacia lipase catalyzed hydrolysis of p-nitrophenyl palmitate under irradiation of light with wavelengths of 250-750 nm. The reaction follows Michaelis-Menten Kinetics and the light irradiation increases the overall rate of hydrolysis. Using Lineweaver-Burk plot K M and V max values for the reaction in presence of light are found to be 39.07 and 66.67 mM/min/g, respectively; while for the same reaction under dark condition, the values are 7.08 and 10.21 mM/min/g. The linear form of enzyme dependent rate of reaction confirms that no mass-transfer limitations are present and the reaction is a kinetically controlled enzymatic reaction.
Microalgal lipid production by Chlorella protothecoides using sugarcane bagasse hydrolysate was investigated in this study. First, maximum glucose and reducing sugar concentrations of 15.2 and 27.0 g/L were obtained in sugarcane bagasse hydrolysate (SCBH), and the effects of different percentages of glucose and xylose on algal cultivation were investigated. Afterwards, SCBH was used as a carbon source for the cultivation of C. protothecoides and higher biomass concentration of 10.7 g/L was achieved. Additionally, a large amount of fatty acids, accounting up to 16.8% of dry weight, were accumulated in C. protothecoides in the nitrogen-limited (0.1-1 mmol/L) culture. Although SCBH inhibited fatty acid accumulation to a certain degree and the inhibition was aggravated by nitrogen starvation, SCBH favored microalgal cell growth and fatty acid production. The present study is of significance for the integration of cost-effective feedstocks production for biodiesel with low-cost SCBH as well as environmentally friendly disposal of lignocellulosic wastes.
A functionalized polystyrene nanofiber (PSNF) immobilized β-galactosidase assembly (PSNF-Gal) was synthesized as a nanobiocatalyst aiming to enhance the biocatalyst stability and functional ability. The PSNF fabricated by electrospinning was functionalized through a chemical oxidation method for enzyme binding. The bioengineering performance of the enzyme carriers was further evaluated for bioconversion of lactose to galacto-oligosaccharides (GOS). The modified PSNF-Gal demonstrated distinguished performances to preserve the same activity as the free β-galactosidase at the optimum pH of 7.0, and to enhance the enzyme stability of PSNF-Gal in an alkaline condition up to pH 10. The PSNF assembly demonstrated improved thermal stability from 37 to 60 °C. The nanobiocatalyst was able to retain 30 % of its initial activity after ninth operation cycles comparing to four cycles with the unmodified counterpart. In contrast with free β-galactosidase, the modified PSNF-Gal enhanced the GOS yield from 14 to 28 %. These findings show the chemically modified PSNF-based nanobiocatalyst may be pertinent for various enzyme-catalysed bioprocessing applications.
The influence of water activity and water content was investigated with farnesyl laurate synthesis catalyzed by Lipozyme RM IM. Lipozyme RM IM activity depended strongly on initial water activity value. The best results were achieved for a reaction medium with an initial water activity of 0.11 since it gives the best conversion value of 96.80%. The rate constants obtained in the kinetics study using Ping-Pong-Bi-Bi and Ordered-Bi-Bi mechanisms with dead-end complex inhibition of lauric acid were compared. The corresponding parameters were found to obey the Ordered-Bi-Bi mechanism with dead-end complex inhibition of lauric acid. Kinetic parameters were calculated based on this model as follows: V (max) = 5.80 mmol l(-1) min(-1) g enzyme(-1), K (m,A) = 0.70 mmol l(-1) g enzyme(-1), K (m,B) = 115.48 mmol l(-1) g enzyme(-1), K (i) = 11.25 mmol l(-1) g enzyme(-1). The optimum conditions for the esterification of farnesol with lauric acid in a continuous packed bed reactor were found as the following: 18.18 cm packed bed height and 0.9 ml/min substrate flow rate. The optimum molar conversion of lauric acid to farnesyl laurate was 98.07 ± 0.82%. The effect of mass transfer in the packed bed reactor has also been studied using two models for cases of reaction limited and mass transfer limited. A very good agreement between the mass transfer limited model and the experimental data obtained indicating that the esterification in a packed bed reactor was mass transfer limited.
In this study, laccase was immobilized on nylon 6,6/Fe(3+) composite (NFC) nanofibrous membrane and used for the detoxification of 3,3'-dimethoxybenzidine (DMOB). The average size and tensile strength of the NFC membrane were found to be 60-80 nm (diameter) and 2.70 MPa, respectively. The FTIR results confirm that the amine (N-H) group of laccase was attached with Fe(3+) particles and the carbonyl (C=O) group of NFC membrane via hydrogen bonding. The half-life of the laccase-NFC membrane storage stability was increased from 6 to 11 weeks and the reusability was significantly extended up to 43 cycles against ABTS oxidation. Enhanced electro-oxidation of DMOB by laccase was observed at 0.33 V and the catalytic current was found to be 30 µA. The DMOB-treated mouse fibroblast 3T3-L1 preadipocytes showed maximum (97 %) cell inhibition at 75 µM L(-1) within 24 h. The cytotoxicity of DMOB was significantly decreased to 78 % after laccase treatment. This study suggests that laccase-NFC membrane might be a good candidate for emerging pollutant detoxification.