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.
A recently reported stable and efficient EBPR system at high temperatures around 30 °C has led to characterization of kinetic and stoichiometric parameters of the Activated Sludge Model no. 2d (ASM2d). Firstly, suitable model parameters were selected by identifiability analysis. Next, the model was calibrated and validated. ASM2d was found to represent the processes well at 28 and 32 °C except in polyhyroxyalkanoate (PHA) accumulation of the latter. The values of the kinetic parameters for PHA storage (q PHA), polyphosphate storage (q PP) and growth (μ PAO) of polyphosphate-accumulating organisms (PAOs) at 28 and 32 °C were found to be much higher than those reported by previous studies. Besides, the value of the stoichiometric parameter for the requirement of polyphosphate for PHA storage (Y PO4) was found to decrease as temperature rose from 28 to 32 °C. Values of two other stoichiometric parameters, i.e. the growth yield of heterotrophic organisms (Y H) and PAOs (Y PAO), were high at both temperatures. These calibrated parameters imply that the extremely active PAOs of the study were able to store PHA, store polyphosphate and even utilize PHA for cell growth. Besides, the parameters do not follow the Arrhenius correlation due to the previously reported unique microbial clade at 28 and 32 °C, which actively performs EBPR at high temperatures.
The use of biomaterials or microorganisms in PAHs degradation had presented an eye-catching performance. Pleurotus eryngii is a white rot fungus, which is easily isolated from the decayed woods in the tropical rain forest, used to determine the capability to utilize naphthalene, a two-ring polycyclic aromatic hydrocarbon as source of carbon and energy. In the meantime, biotransformation of naphthalene to intermediates and other by-products during degradation was investigated in this study. Pleurotus eryngii had been incubated in liquid medium formulated with naphthalene for 14 days. The presence of metabolites of naphthalene suggests that Pleurotus eryngii begin the ring cleavage by dioxygenation on C1 and C4 position to give 1,4-naphthaquinone. 1,4-Naphthaquinone was further degraded to benzoic acid, where the proposed terepthalic acid is absent in the cultured extract. Further degradation of benzoic acid by Pleurotus eryngii shows the existence of catechol as a result of the combination of decarboxylation and hydroxylation process. Unfortunately, phthalic acid was not detected in this study. Several enzymes, including manganese peroxidase, lignin peroxidase, laccase, 1,2-dioxygenase and 2,3-dioxygenase are enzymes responsible for naphthalene degradation. Reduction of naphthalene and the presence of metabolites in liquid medium showed the ability of Pleurotus eryngii to utilize naphthalene as carbon source instead of a limited glucose amount.
The introduction of plastic sectors has resulted in the presence of microplastics (MPs) in water systems, which has become a global issue that has attracted scientific and community awareness. MPs can be detected in a variety of sources such as beauty products, manufacturing effluent, or fishing activities. This study examined the repercussions posed by MPs' prevalence on land and marine environments and human health issues. Henceforth, remediation technologies must be introduced to shift out MPs from the water supplies in order to sustain the environmental quality for future generations, the benefits and drawbacks of the technology applied. This study also portrays difficulties encountered in MP research as the hurdles must be mastered in order to properly comprehend the MPs. The cooperation between nations is the most critical aspect in fully tackling MP issues as it can be easily carried by wind or water and its damage can be larger than predicted.
Insufficient power generation from a microbial fuel cell (MFC) hampers its progress towards utility-scale development. Electrode modification with biopolymeric materials could potentially address this issue. In this study, medium-chain-length poly-3-hydroxyalkanoates (PHA)/carbon nanotubes (C) composite (CPHA) was successfully applied to modify the surface of carbon cloth (CC) anode in MFC. Characterization of the functional groups on the anodic surface and its morphology was carried out. The CC-CPHA composite anode recorded maximum power density of 254 mW/m2, which was 15-53% higher than the MFC operated with CC-C (214 mW/m2) and pristine CC (119 mW/m2) as the anode in a double-chambered MFC operated with Escherichia coli as the biocatalyst. Electrochemical impedance spectroscopy and cyclic voltammetry showed that power enhancement was attributed to better electron transfer capability by the bacteria for the MFC setup with CC-CPHA anode.
In the current scenario, many synthetic chemicals have used long-term to control pests and mosquitoes, leading to the resistance of strains and toxicity effect on human beings. To overcome the adverse problem in recent advances, the scientific community is looking into nanofabricated pesticides and mosquitoes. This study aims to synthesize the recyclable chitosan-coated cadmium nanoparticles (Ch-CdNps) using Plumeria alba flower extract, which was further applied for insecticidal and mosquitocidal activities. The synthesized Ch-CdNps were confirmed by UV spectroscopy and FTIR analysis. The XRD, TEM, and DLS results confirmed the crystallinity with a spherical shape at 80-100 nm. The insecticidal activity proves that Ch-CdNps inhibited Helicoverpa armigera and Spodoptera litura at 100 ppm. In mosquitocidal, LC50 values of larvicidal of 1st instar were 4.116, 4.33, and 4.564 µg/mL, and the remaining three stages of instars, pupicidal, adulticidal, longevity, fecundity, and ovicidal assays inhibit the Anopheles stephensi followed by Aedes aegypti and Culex quinquefasciatus. Further, the first-order kinetics of photocatalytic degradation of methylene blue and methyl orange was confirmed. Based on the obtained results, Ch-CdNps can inhibit the pest, mosquitoes, and photocatalytic degradation.
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.
Cresol Red belongs to the triphenylmethane (TPM) class of dyes which are potentially carcinogenic or mutagenic. However, very few studies on biodegradation of Cresol Red were investigated as compared to other type dyes such as azo and anthraquinone dye. The aim of this work is to evaluate triphenylmethane dye Cresol Red degradation by fungal strain isolated from the decayed wood in Johor Bahru, Malaysia. Detailed taxonomic studies identified the organisms as Trichoderma species and designated as strain Trichoderma harzianum M06. In this study, Cresol Red was decolorized up to 88% within 30 days under agitation condition by Trichoderma harzianum M06. Data analysis revealed that a pH value of 3 yielded a highest degradation rate among pH concentrations (73%), salinity concentrations of 100 g/L (73%), and a volume of 0.1 mL of Tween 80 (79%). Induction in the enzyme activities of manganese peroxidase, lignin peroxidase, laccase, 1,2- and 2,3-dioxygenase indicates their involvement in Cresol Red removal. Various analytical studies such as Thin-Layer Chromatography (TLC), UV-Vis spectrophotometer, and Gas chromatography mass spectrometry (GC-MS) confirmed the biotransformation of Cresol Red by the fungus. Two metabolites were identified in the treated medium: 2,4-dihydroxybenzoic acid (t R 7.3 min and m/z 355) and 2-hydroxybenzoic acid (t R 8.6 min and m/z 267). Based on these products, a probable pathway has been proposed for the degradation of Cresol Red by Trichoderma harzianum M06.
The under-treated wastewater, especially remaining carcinogenic aromatic compounds in wastewater discharge has been expansively reported, wherein the efficiency of conventional wastewater treatment is identified as the primary contributor source. Herein, the advancement of wastewater treatments has drawn much attention in recent years. In the current study, combined sequential and hybridized treatment of thermolysis and coagulation-flocculation provides a novel advancement for environmental emerging pollutant (EP) prescription. This research is mainly demonstrating the mitigation efficiency and degradation pathway of pararosaniline (PRA) hybridized and combined sequential wastewater treatment. Notably, PRA degradation dominantly via a linkage of reaction: thermal cleavage, deamination, silication and diazene reduction. Thermolysis acts as an initiator for the PRA decomposition through thermally induced bond dissociation energy (BDE) for molecular fragmentation whilst coagulation-flocculation facilitates the formation of organo-bridged silsesquioxane as the final degradation product. Different from conventional treatment, the hybridized treatment showed excellent synergistic degradability by removing 99% PRA and its EPs, followed by combined sequential treatment method with 86% reduction. Comprehensive degradation pathway breakdown of carcinogenic and hardly degradable aromatic compounds provides a new insight for wastewater treatment whereby aniline and benzene are entirely undetectable in effluent. The degradation intermediates, reaction derivatives and end products were affirmed by gas chromatography-mass spectrometry, Fourier transform infrared spectroscopy and ultraviolet-visible spectrophotometry (GC-MS, FTIR and UV-Vis). This finding provides valuable guidance in establishing efficient integrated multiple-step wastewater treatments.
This study showed that bio-functional silver nanoparticles (AgNPs) and zinc oxide nanoparticles (ZnONPs) were synthesized in aqueous extracts of Gymnema sylvestre leaves and tested for toxicity assessment against triple-negative breast cancer cells (TNBC). Biofunctional nanoparticle (NPs) samples were characterized using UV-Vis spectroscopy, FT-IR, XRD, SEM, and TEM. The results showed that the phytofabrication of AgNPs resulted in a dark brown, UV-vis maximum absorbance peak at 413 nm. The AgNPs were crystalline and spherical, with sizes ranging from 20 to 60 nm, as confirmed by the XRD pattern and TEM images. Another phytofabrication of ZnONPs exhibited a white precipitate corresponding to a UV-Vis maximum absorption peak at 377 nm and a fine micro flower morphology with a particle-sized tribution between 100 and 200 nm. In addition, FT-IR spectra showed that bioorganic compounds are associated with NPs that respond to reduced Ag+ ions and AgNPs tabilizers. Invitro cytotoxicity studies revealed the potent anti-cancer effects of phytofabricated AgNPs and ZnONPs on TNBC cells. Furthermore, the AO/EB double staining assay results proved that apoptotic cells are distinguished by greenish-yellow fluorescence of the cell nuclei with IC50 concentrations of 44 ± 0.8 µg/mL for AgNPs and 26.2 ± 0.5 µg/mL for ZnONPs, respectively. Based on our results, we expect that the anticancer function of the biofunctional NPs is due to the apoptotic activation of TNBC cells by increased ROS. Therefore, the presented study demonstrated that biofunctional AgNPs and ZnONPs have excellent prospects for the anti-cancer activity that can be used in pharmaceutical and medical fields.
Employing aerobic fermentation, Gram-negative bacteria belonging to the genus Xanthomonas produce the high molecular weight natural heteropolysaccharide known as xanthan. It has various amounts of O-acetyl and pyruvyl residues together with D-glucosyl, D-mannosyl, and D-glucuronyl acid residues in a molar ratio of 2:2:1. The unique structure of xanthan allowed its various applications in a wide range of industries such as the food industry, pharmacology, cosmetics and enhanced oil recovery primarily in petroleum. The cultivation medium used in the manufacture of this biopolymer is critical. Many attempts have been undertaken to generate xanthan gum from agro-based and food industry wastes since producing xanthan gum from synthetic media is expensive. Optimal composition and processing parameters must also be considered to achieve an economically viable manufacturing process. There have been several attempts to adjust the nutrient content and feeding method, temperature, pH, agitation and the use of antifoam in xanthan fermentations. Various modifications in technological approaches have been applied to enhance its physicochemical properties which showed significant improvement in the area studied. This review describes the biosynthesis production of xanthan with an emphasis on the importance of the upstream processes involving medium, processing parameters, and other factors that significantly contributed to the final application of this precious polysaccharide.
Armillaria sp. F022, a white-rot fungus isolated from decayed wood in tropical rain forest was used to biodegrade anthracene in cultured medium. The percentage of anthracene removal by Armillaria sp. F022 reached 13 % after 7 days and at the end of the experiment, anthracene removal level was at 87 %. The anthracene removal through sorption and transformation was investigated. 69 % of eliminated anthracene was transformed by Armillaria sp. F022 to form other organic structure, while only 18 % was absorbed in the mycelia. In the kinetic experiment, anthracene dissipation will not stop even though the biomass had stopped growing. Anthracene removal by Armillaria sp. F022 was correlated with protein concentration (whole biomass) in the culture. The production of enzyme was affected by biomass production. Anthracene was transformed to two stable metabolic products. The metabolites were extracted in ethyl-acetate, isolated by column chromatography, and then identified using gas chromatography-mass spectrometry (GC-MS).
Production of extracellular laccase by the white-rot fungus Pycnoporus sanguineus was examined in batch submerged cultures in shake flasks, baffled shake flasks and a stirred tank bioreactor. The biomass growth in the various culture systems closely followed a logistic growth model. The production of laccase followed a Luedeking-Piret model. A modified Luedeking-Piret model incorporating logistic growth effectively described the consumption of glucose. Biomass productivity, enzyme productivity and substrate consumption were enhanced in baffled shake flasks relative to the cases for the conventional shake flasks. This was associated with improved oxygen transfer in the presence of the baffles. The best results were obtained in the stirred tank bioreactor. At 28 °C, pH 4.5, an agitation speed of 600 rpm and a dissolved oxygen concentration of ~25 % of air saturation, the laccase productivity in the bioreactor exceeded 19 U L(-1 )days(-1), or 1.5-fold better than the best case for the baffled shake flask. The final concentration of the enzyme was about 325 U L(-1).
This review is focused on the production of microbial lipases by high cell density fermentation. Lipases are among the most widely used of the enzyme catalysts. Although lipases are produced by animals and plants, industrial lipases are sourced almost exclusively from microorganisms. Many of the commercial lipases are produced using recombinant species. Microbial lipases are mostly produced by batch and fed-batch fermentation. Lipases are generally secreted by the cell into the extracellular environment. Thus, a crude preparation of lipases can be obtained by removing the microbial cells from the fermentation broth. This crude cell-free broth may be further concentrated and used as is, or lipases may be purified from it to various levels. For many large volume applications, lipases must be produced at extremely low cost. High cell density fermentation is a promising method for low-cost production: it allows a high concentration of the biomass and the enzyme to be attained rapidly and this eases the downstream recovery of the enzyme. High density fermentation enhances enzyme productivity compared with the traditional submerged culture batch fermentation. In production of enzymes, a high cell density is generally achieved through fed-batch operation, not through perfusion culture which is cumbersome. The feeding strategies used in fed-batch fermentations for producing lipases and the implications of these strategies are discussed. Most lipase-producing microbial fermentations require oxygen. Oxygen transfer in such fermentations is discussed.
Microplastics (MPs) in environmental studies have revealed that public sewage treatment plants are a common pathway for microplastics to reach local surroundings. Microplastics are becoming more of a worry, posing a danger to both marine wildlife and humans. These plastic items not only contribute to the macrocosmic proliferation of plastics but also the scattering of microplastics and the concentration of other micropollutant-containing objects, increasing the number of pollutants identified. Microplastics' behavior, movement, transformation, and persistence mechanisms, as well as their mode of action in various wastewater effluent treatment procedures, are still unknown. They are making microplastics made from wastewater a big deal. We know that microplastics enter wastewater treatment facilities (WWTPs), that wastewater is released into the atmosphere, and that this wastewater has been considered to represent a threat to habitats and ground character based on our literature assessment. The basic methods of wastewater and sewage sludge, as well as the treatment procedure and early characterization, are covered throughout the dissection of the problematic scientific conceptualization.
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.
The present study reports the effects of three commercial immobilized lipases namely Novozyme 435 from Candida antarctica lipase B (CALB), Lipozyme TL IM from Thermomyces lanuginosus and Lipozyme RM IM from Rhizomucor miehei on the production of trimethylolpropane (TMP) ester from high oleic palm methyl ester (HO-PME) and TMP. The TMP ester is a promising base oil for biolubricants that are easily biodegradable and non-toxic to humans and the environment. Enzymatic catalysts are insensitive to free fatty acid (FFA) content, hence able to mitigate the side reactions and consequently reduce product separation cost. The potential of these enzymes to produce TMP ester in a solvent-free medium was screened at various reaction time (8, 23, 30 and 48 h), operating pressure (0.1, 0.3 and 1.0 mbar) and enzyme dosage (1, 3, 5 and 10% w/w). The reaction was conducted at a constant temperature of 70 °C and a molar ratio of 3.9:1 (HO-PME: TMP). Novozyme 435 produced the highest yield of TMP ester of 95.68 ± 3.60% under the following conditions: 23 h reaction time, 0.1 mbar operating pressure and 5% w/w of enzyme dosage. The key lubrication properties of the produced TMP ester are viscosity index (208 ± 2), pour point (- 30 ± - 2 °C), cloud point (- 15 ± - 2 °C), onset thermal degradation temperature (427.8 °C), and oxidation stability, RPVOT (42 ± 4 min). The properties of the TMP ester produced from the enzymatic transesterification are comparable to other vegetable oil-based biolubricants produced by chemical transesterification.
Pasteurella multocida serotype B:2 is the causative agent of haemorrhagic septicaemia, a fatal disease in cattle and buffaloes. For use as a vaccine in the treatment of HS disease, an efficient cultivation of attenuated gdhA derivative P. multocida B:2 (mutant) for mass production of viable cells is required. In this study, the role of amino acids and vitamins on the growth of this particular bacterium was investigated. Initially, three basal media (Brain-heart infusion, Terrific broth, and defined medium YDB) were assessed in terms of growth performance of P. multocida B:2. YDB medium was selected and redesigned to take into account the effects of amino acids (glutamic acid, cysteine, glycine, methionine, lysine, tyrosine, and histidine) and vitamins (vitamin B1, nicotinic acid, riboflavin, pyridoxine, pantothenic acid, and biotin). High viable cell number was largely affected by the availability of micronutrient components and macronutrients. Histidine was essential for the growth whereby a traceable amount (20 mM) was found to greatly enhance the growth of gdhA derivative P. multocida B:2 mutant (6.6 × 109 cfu/mL) by about 19 times as compared to control culture (3.5 × 108 cfu/mL). In addition, amongst the vitamins added, riboflavin exhibited the highest impact on the viability of gdhA derivative P. multocida B:2 mutant (5.3 × 109 cfu/mL). Though the combined histidine and riboflavin in the culture eventually did not promote the stacking impact on cell growth and cell viability, nonetheless, they were still essential and important in either growth medium or production medium.
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.
In this study, a newly isolated ascomycete fungus Trichoderma lixii F21 was explored to bioremediate the polar [Alizarin Red S (ARS)] and non-polar [Quinizarine Green SS (QGSS)] anthraquinone dyes. The bioremediation of ARS and QGSS by T. lixii F21 was found to be 77.78 and 98.31 %, respectively, via biosorption and enzymatic processes within 7 days of incubation. The maximum biosorption (ARS = 33.7 % and QGSS = 74.7 %) and enzymatic biodegradation (ARS = 44.1 % and QGSS = 23.6 %) were observed at pH 4 and 27 °C in the presence of glucose and yeast extract. The laccase and catechol 1,2-dioxygenase produced by T. lixii F21 were involved in the molecular conversions of ARS and QGSS to phenolic and carboxylic acid compounds, without the formation of toxic aromatic amines. This study suggests that T. lixii F21 may be a good candidate for the bioremediation of industrial effluents contaminated with anthraquinone dyes.