System for bioremediation of phenol-polluted waters based on the application of the common duckweed (Lemna minor L.) and the bacteria of its rhizosphere

U ovoj doktorskoj disertaciji ispitivano je uklanjanje fenola iz tečne hranljive podloge pomoću sočivice (Lemna minor L.) i bakterija njene rizosfere. Od ukupnog broja bakterijskih izolata rizosfere, izolovano je 6 sojeva kao najbolji kandidati za uklanjanje fenola: Lelliottia sp. 11, Klebsiella oxytoca 14, Serratia marcescens 27, Hafnia alvei 37, H. paralvei 43, i S. nematodiphila 51. Sočivice inokulisane bilo kojim od ovih sojeva efikasnije u uklanjanju fenol od površinski sterilisanih sočivica kao i od bilo kog od ovih sojeva u obliku slobodne suspenzije (bez sočivica). Soj H. paralvei 43, u kombinaciji sa sočivicom, bio je najefikasniji u uklanjanju fenola. Ovaj bakterijski soj utiče i na antioksidativni odgovor sočivice što se ogleda u povećanim enzimskim (GPX) i neenzimskim (ukupni solubilni proteini biljnog tkiva, MDA i H2O2) parametrima oksidativnog stresa.Zimogramskom detekcijom GPX utvrđeno je prisustvo pet različitih izoformi u tkivu sočivice gajenoj u prisustvu fenola, dok su četiri izoforme detektovane kada su na korenu bile prisutne bakterije bez fenola Fluorescentom mikroskopijom pokazano je da je K. oxytoca 14 endofit sa najizraženijim efektom na multiplikaciju sočivica sa ili bez fenola. Iz hranljive podloge bez fenola u kojoj su gajene sočivice i K. oxytoca 14 izolovano je 6 fenolnih jedinjenja: luteolin 6,8-di-C-heksozid, p-hidroksibenzoeva kiselina, kafeinska kiselina, apigenin 6-C-(2"-pentozil)heksozid i p-kumarinska kiselina. Rezultati ove doktorske disertacije pokazuju da su odabrani bakterijski sojevi u kombinaciji sa biljkama sočivice potencijalno rešenje za bioremedijaciju vode zagađene fenolom.In this dissertation, the possibility of phenol removal by the common duckweed (Lemna minor L.) and by the bacteria of its rhizosphere, was investigated. Six bacterial strains, selected according to their ability of growth on the minimal nutrient medium with phenol, high phenol-resistance, sensitivity to typical antibiotics, biofilm formation, positive effect on the multiplication rates of the plants and bacteria-plant interactions on the root surface were: Lelliottia sp. 11, Klebsiella oxytoca 14, Serratia marcescens 27, Hafnia alvei 37, H. paralvei 43, and S. nematodiphila 51. Duckweed inoculated with any of the aforementioned strains were more efficient in phenol removal than the surface-sterilized duckweed or the bacterial suspensions. The most efficient bacterial strain/duckweed system is the one based on H. paralvei 43 and plants. This bacterial strain also modulates the antioxidant response of the plant, indicated by the non-enzymatic (total AsA, soluble proteins, MDA and H2O2) and the enzymatic parameters. The zymogram detection of GPX in the plants showed 5 phenol-induced GPX isoforms, whereas the bacteria alone induced 4 different GPX isoforms. Bacteria showed a tendency to lower the intensity of GPX expression in plants. K. oxytoca 14 is an endophyte which induced a rapid multiplication of the duckweed. Six phenolic compounds were identified in the nutrient medium used to co-culture K. oxytoca 14 and the duckweed: lutheolin 6,8-di-C-hexoside, p-hidroxybenzoic acid, caffeic acid, apigenin 6-C-(2"-pentosyl) hexoside and p-coumaric acid. The results of this dissertation show that the selected bacteria /duckweed systems present an efficient solution for the bioremediation of the phenol-polluted wastewaters

Phenol Removal Capacity of the Common Duckweed (Lemna minor L.) and Six Phenol-Resistant Bacterial Strains From Its Rhizosphere: In Vitro Evaluation at High Phenol Concentrations

The main topic of this study is the bioremediation potential of the common duckweed, Lemna minor L., and selected rhizospheric bacterial strains in removing phenol from aqueous environments at extremely high initial phenol concentrations. To that end, fluorescence microscopy, MIC tests, biofilm formation, the phenol removal test (4-AAP method), the Salkowski essay, and studies of multiplication rates of sterile and inoculated duckweed in MS medium with phenol (200, 500, 750, and 1000 mg L−1) were conducted. Out of seven bacterial strains, six were identified as epiphytes or endophytes that efficiently removed phenol. The phenol removal experiment showed that the bacteria/duckweed system was more efficient during the first 24 h compared to the sterile duckweed control group. At the end of this experiment, almost 90% of the initial phenol concentration was removed by both groups, respectively. The bacteria stimulated the duckweed multiplication even at a high bacterial population density (>105 CFU mL−1) over a prolonged period of time (14 days). All bacterial strains were sensitive to all the applied antibiotics and formed biofilms in vitro. The dual bacteria/duckweed system, especially the one containing strain 43-Hafnia paralvei C32-106/3, Accession No. MF526939, had a number of characteristics that are advantageous in bioremediation, such as high phenol removal efficiency, biofilm formation, safety (antibiotic sensitivity), and stimulation of duckweed multiplication

Selectivity issues in affinity-based biochemical sensors: Determining the ratio of similar biomolecules in binary mixtures

We consider selectivity of affinity-based nanosensors utilizing resonance shift due to the presence of adsorbed analyte. Among such devices we analyze mass-based sensors utilizing mechanical resonance in e.g. micro or nanocantilevers and all-optical refractometric sensors utilizing surface plasmon polariton resonance. The sensitivity of such devices can be extremely high, reaching single-molecule level, however their selectivity is limited by the differences in mass in the first case or in refractive index values between different analytes in the second. The typical approach is to use some kind of receptors on the sensor surface with highly specific binding of a targeted analyte. The properties of a given biomolecule, for instance protein, will vary between its different conformations due to different arrangements of their atoms in space. Since the conformation of a molecule is critically important for its function, it is of interest to determine the ratio between different conformational isomers in a given sample. In general, different conformations of a biomolecule may have different affinity toward binding sites on the surface of an affinity-based nanosensor, as well as different surface-volume ratios. We argue that the analysis of adsorption kinetics ensures sufficient data to discriminate between different conformations in a mixture

The time response of plasmonic sensors due to binary adsorption: analytical versus numerical modeling

In order to allow for multiscale modeling of complex systems we focus on various approaches to modeling binary adsorption. We consider multiple methods of modeling the temporal response of general plasmonic sensors. We start from the analytical approach. The kinetics of adsorption and desorption is modeled both as a first order reaction and as a second order reaction. The criteria for their validity and the choice between them in the case of two-component adsorption are established. Due to the nonlinearities of the second order reactions and the lack of their analytical solutions, computer aided modeling is considered next, also in multiple ways: the employment of numerical solvers, fitting of experimental results, the stochastic simulation algorithms and the employment of artificial neural networks (ANN). The examples we present illustrate the advantages and disadvantages of the particular approaches. The goal is to aid the concurrent multiscale modeling of adsorption-based devices. Machine learning in ANN performed here is used to estimate the equilibrium values of adsorbed quantities. The obtained results show that to train an ANN for the estimation of the equilibrium adsorption quantities the Levenberg-Marquardt and the Bayesian regularization algorithms are less efficient than the quasi-Newton BFGS (Broyden-Fletcher-Goldfarb-Shanno) algorithm.This is a post-peer-review, pre-copyedit version of an article (Jakšić, O., Jokić, I., Jakšić, Z. et al. The time response of plasmonic sensors due to binary adsorption: analytical versus numerical modeling) published in Applied Physics A: Materials Science and Processing (Appl. Phys. A 126, 342, 2020) The final authenticated version is available online at: [https://doi.org/10.1007/s00339-020-03524-3]The published version: [http://cer.ihtm.bg.ac.rs/handle/123456789/3494

Nanoaperture array-based plasmonic sensors of dangerous substances using transparent conductive oxides

Detection of dangerous substances like explosives, pathogenic microorganisms and toxic chemical is of utmost interest for homeland defense and anti-terrorist actions. Among the devices of choice for that purpose are surface plasmon resonance (SPR) chemical-biological sensors since they are ultra-fast, highly sensitive and label-free. We consider one of the advanced types of SPR sensors, those with ordered arrays of subwavelength apertures in plasmonic material. Such sensors show high sensitivity in transmission readout mode, are useful for both liquid and gaseous analytes and can be used as a building block for complex microfluidic systems for multianalyte detection. Simultaneously, high electromagnetic field concentrations in nanoapertures enhance nonlinear effects, thus further facilitating detection of complex molecules of dangerous substances. We performed finite element simulation of the performance of such devices for various materials and nanoaperture sizes and shapes. We dedicated special attention to the case when the subwavelength array is formed in transparent conductive oxide (TCO) host. An example of TCO is tin oxide, routinely used in thin layers for window defrosting and defogging elements in armored vehicles. We show that compared to conventional nanoaperture array sensors based on metal hosts the TCO-based devices simultaneously offer enhanced selectivity and retain high sensitivity

Electrodeposition of Nb and Al from chloroaluminate melt on vitreous carbon

Niobium and aluminium were electrodeposited at 200 °C under argon atmosphere onto vitreous carbon from inorganic chloroaluminate melts (AlCl3+NaCl) with added niobium. Niobium was introduced into the electrolyte by anodic dissolution of metallic niobium or by chemical dissolution of Nb2O5 in a melt of equimolar AlCl3+NaCl mixture. The processes of deposition/dissolution onto/from vitreous carbon were investigated by cyclic voltammetry and chronoamperometry. Characterization of the obtained deposits was done by Scanning Electron Microscopy (SEM) and Energy Dispersive Spectroscopy (EDS). The only observed reduction processes on the working electrode in the potential window from 1.000 V to – 1.000 V vs. Al, were individual niobium deposition and codeposition of niobium and aluminium with Al-Nb alloys formation. Electrodeposition of niobium from the chloroaluminate melt with added niobium (V) oxide seems to start at around – 0.100 V vs. Al and at about – 0.200 V vs. Al aluminium starts codepositing. During the codeposition Nb-Al alloys were formed. Niobium deposition starting potential from the electrolyte with niobium added by anodic dissolution starts at 0.100 V vs. Al, and aluminium codeposition starting potential was at around – 0.025 V vs. Al, followed by Nb/Al alloy formation

Poređenje biosorpcije sa Aureobasidium pullulans soj CH-1 i polimer sorpcije na Poli(GMA-co-EGMA)teških metala iz otpadnih voda

The intact microbial cells as well as derived microbial products can be highly efficient bioacummulators of both soluble and participate forms of metals, especially from dilute external concentrations (1,2). The reason for testing Aureobasidium pullulans as a biosorbent is the fact that this fungus presents a ubiquitous saprophite that occurs commonly in the phytoshere (3). Therefore, it has enabled the use of carbohydrate wastes as substrates for the production of pullulan such as the peat hydrolysate. The metal removal and recovery processes include: precipitation, adsorption and biosorption, electrowinning and electocoagulation, cementation, membrane separation, solvent extraction and ion exchange (4). Galvanic waste effluent contain heavy metals ions, such as Cu2f, Cr1+, Cr6+, Cd2+, Ni2+, etc., which present environmental hazard. Having all this in mind, the aim of this paper is to examine the possibility of applying Aureobasidium pullulam CH-1 (Collection of Microorganisms, IChTM) as a biosorbent of metals and biosynthesis of pullulan at the same time on peat hydrolisate, distribution of sorbed metals between biomass and pullulan, as well as the correlation between biosorption Aureobasidium pullulam CH-1 and polymer sorption on poly(GMA-co-EGDMA)-en of heavy metals from galvanic waste effluent.Ispitivanje Aureobasidium pullulans, soj CH-1 kao biosorbenta je urađeno na kiselom hidrolizatu treseta sa lokaliteta Vlasinskog jezera. Kiseli hidrolizat treseta je dobijen na isti način kao i otpadne vode pri industrijskoj obradi treseta. Rezultati su pokazali da Aureohusidium pullulans, soj CH-1 može istovremeno da vrši biosorpciju metala i biosintezu pululana pri fermentacijama na kiselom hidrolizatu treseta. U cilju poređenja, za sorpciju teških metala iz galvanskih otpadnih voda, osim Aureobasidium pullulans soj CH-1 korišćen je i makroporozni kopolimer, poli(gicidilmetakrilat-co-etilenglikoldimetakrilat) modifikovan sa etilendiaminom, poli(GMA-co-EGDMA)-en. Rezultati uporednih eksperimenata biosorpcije na Aureobasidium pullulans soj CH-1 i sorpcije na polimernom sorbentu su pokazali da se oba postupka mogu uspešno koristiti za uklanjanje teških metala iz galvanskih otpadnih voda. Pokazalo se da je pri datim eksperimentalnim uslovima, za uklanjanje cinka uspešniji postupak biosorpcija, dok je poli(GMA-co-F.GDMA)-en znatno efikasniji za uklanjanje bakra iz galvanskih otpadnih voda

Poređenje biosorpcije sa Aureobasidium pullulans soj CH-1 i polimer sorpcije na Poli(GMA-co-EGMA)teških metala iz otpadnih voda

The intact microbial cells as well as derived microbial products can be highly efficient bioacummulators of both soluble and participate forms of metals, especially from dilute external concentrations (1,2). The reason for testing Aureobasidium pullulans as a biosorbent is the fact that this fungus presents a ubiquitous saprophite that occurs commonly in the phytoshere (3). Therefore, it has enabled the use of carbohydrate wastes as substrates for the production of pullulan such as the peat hydrolysate. The metal removal and recovery processes include: precipitation, adsorption and biosorption, electrowinning and electocoagulation, cementation, membrane separation, solvent extraction and ion exchange (4). Galvanic waste effluent contain heavy metals ions, such as Cu2f, Cr1+, Cr6+, Cd2+, Ni2+, etc., which present environmental hazard. Having all this in mind, the aim of this paper is to examine the possibility of applying Aureobasidium pullulam CH-1 (Collection of Microorganisms, IChTM) as a biosorbent of metals and biosynthesis of pullulan at the same time on peat hydrolisate, distribution of sorbed metals between biomass and pullulan, as well as the correlation between biosorption Aureobasidium pullulam CH-1 and polymer sorption on poly(GMA-co-EGDMA)-en of heavy metals from galvanic waste effluent.Ispitivanje Aureobasidium pullulans, soj CH-1 kao biosorbenta je urađeno na kiselom hidrolizatu treseta sa lokaliteta Vlasinskog jezera. Kiseli hidrolizat treseta je dobijen na isti način kao i otpadne vode pri industrijskoj obradi treseta. Rezultati su pokazali da Aureohusidium pullulans, soj CH-1 može istovremeno da vrši biosorpciju metala i biosintezu pululana pri fermentacijama na kiselom hidrolizatu treseta. U cilju poređenja, za sorpciju teških metala iz galvanskih otpadnih voda, osim Aureobasidium pullulans soj CH-1 korišćen je i makroporozni kopolimer, poli(gicidilmetakrilat-co-etilenglikoldimetakrilat) modifikovan sa etilendiaminom, poli(GMA-co-EGDMA)-en. Rezultati uporednih eksperimenata biosorpcije na Aureobasidium pullulans soj CH-1 i sorpcije na polimernom sorbentu su pokazali da se oba postupka mogu uspešno koristiti za uklanjanje teških metala iz galvanskih otpadnih voda. Pokazalo se da je pri datim eksperimentalnim uslovima, za uklanjanje cinka uspešniji postupak biosorpcija, dok je poli(GMA-co-F.GDMA)-en znatno efikasniji za uklanjanje bakra iz galvanskih otpadnih voda

Antioxidative Responses of Duckweed (Lemna minor L.) to Phenol and Rhizosphere-Associated Bacterial Strain Hafnia paralvei C32-106/3

Duckweed (L. minor) is a cosmopolitan aquatic plant of simplified morphology and rapid vegetative reproduction. In this study, an H. paralvei bacterial strain and its influence on the antioxidative response of the duckweeds to phenol, a recalcitrant environmental pollutant, were investigated. Sterile duckweed cultures were inoculated with H. paralvei in vitro and cultivated in the presence or absence of phenol (500 mg L−1), in order to investigate bacterial effects on plant oxidative stress during 5 days. Total soluble proteins, guaiacol peroxidase expression, concentration of hydrogen peroxide and malondialdehyde as well as the total ascorbic acid of the plants were monitored. Moreover, bacterial production of indole-3-acetic acid (IAA) was measured in order to investigate H. paralvei’s influence on plant growth. In general, the addition of phenol elevated all biochemical parameters in L. minor except AsA and total soluble proteins. Phenol as well as bacteria influenced the expression of guaiacol peroxidase. Different isoforms were associated with phenol compared to isoforms expressed in phenol-free medium. Considering that duckweeds showed increased antioxidative parameters in the presence of phenol, it can be assumed that the measured parameters might be involved in the plant’s defense system. H. paralvei is an IAA producer and its presence in the rhizosphere of duckweeds decreased the oxidative stress of the plants, which can be taken as evidence that this bacterial strain acts protectively on the plants during phenol exposure

Electrodeposition of Nb and Al from chloroaluminate melt on vitreous carbon

Niobium and aluminium were electrodeposited at 200 °C under argon atmosphere onto vitreous carbon from inorganic chloroaluminate melts (AlCl3+NaCl) with added niobium. Niobium was introduced into the electrolyte by anodic dissolution of metallic niobium or by chemical dissolution of Nb2O5 in a melt of equimolar AlCl3+NaCl mixture. The processes of deposition/dissolution onto/from vitreous carbon were investigated by cyclic voltammetry and chronoamperometry. Characterization of the obtained deposits was done by Scanning Electron Microscopy (SEM) and Energy Dispersive Spectroscopy (EDS). The only observed reduction processes on the working electrode in the potential window from 1.000 V to – 1.000 V vs. Al, were individual niobium deposition and codeposition of niobium and aluminium with Al-Nb alloys formation. Electrodeposition of niobium from the chloroaluminate melt with added niobium (V) oxide seems to start at around – 0.100 V vs. Al and at about – 0.200 V vs. Al aluminium starts codepositing. During the codeposition Nb-Al alloys were formed. Niobium deposition starting potential from the electrolyte with niobium added by anodic dissolution starts at 0.100 V vs. Al, and aluminium codeposition starting potential was at around – 0.025 V vs. Al, followed by Nb/Al alloy formation