Application of biomacromolecules encapsulation systems for the long-term storage of Lactobacillus plantarum F1 and Lactobacillus reuteri 182.

The aim of this study was to improve the viability of lactic acid bacteria (LAB) during extended storage of 1 year and mechanical characteristics of the calcium alginate beads with co-encapsulation of prebiotics and chitosan coating and subsequent freeze drying. The results revealed that the addition of trehalose to alginate matrix effectively protects the LAB cells during freeze drying, i.e., the survival rate has increased up to more than 92.5 %. Chitosan coating reinforced Ca-alginate beads, therefore the sphericity and mechanical strength of the beads improved. The findings also showed that bacteria encapsulation with the prebiotics resulted in more cells stability during the prolonged storage of 1 year and were 4.82 ± 0.06 log CFU g-1 in the lyophilized alginate-trehalose beads for Lactobacillus plantarum and 5.64 ± 0.08 log CFU g-1 in the lyophilized alginate-trehalose-inulin beads for Lactobacillus reuteri. No survival, however, was noted for the LAB cells in wet capsules after the same period. This study demonstrated that prebiotics had a significant impact on the viability of cells during freeze drying and storage. What is more, physical properties of the alginate beads were enhanced by coating beads with the chitosan.

Microbial Biofuel Cells: Fundamental Principles, Development and Recent Obstacles.

This review focuses on the development of microbial biofuel cells to demonstrate how similar principles apply to the development of bioelectronic devices. The low specificity of microorganism-based amperometric biosensors can be exploited in designing microbial biofuel cells, enabling them to consume a broader range of chemical fuels. Charge transfer efficiency is among the most challenging and critical issues while developing biofuel cells. Nanomaterials and particular redox mediators are exploited to facilitate charge transfer between biomaterials and biofuel cell electrodes. The application of conductive polymers (CPs) can improve the efficiency of biofuel cells while CPs are well-suitable for the immobilization of enzymes, and in some specific circumstances, CPs can facilitate charge transfer. Moreover, biocompatibility is an important issue during the development of implantable biofuel cells. Therefore, biocompatibility-related aspects of conducting polymers with microorganisms are discussed in this review. Ways to modify cell-wall/membrane and to improve charge transfer efficiency and suitability for biofuel cell design are outlined.

Influence of TiO2 and ZnO Nanoparticles on α-Synuclein and ß-Amyloid Aggregation and Formation of Protein Fibrils.

The most common neurological disorders, i.e., Parkinson's disease (PD) and Alzheimer's disease (AD), are characterized by degeneration of cognitive functions due to the loss of neurons in the central nervous system. The aggregation of amyloid proteins is an important pathological feature of neurological disorders.The aggregation process involves a series of complex structural transitions from monomeric to the formation of fibrils. Despite its potential importance in understanding the pathobiology of PD and AD diseases, the details of the aggregation process are still unclear. Nanoparticles (NPs) absorbed by the human circulatory system can interact with amyloid proteins in the human brain and cause PD. In this work, we report the study of the interaction between TiO2 nanoparticles (TiO2-NPs) and ZnO nanoparticles (ZnO-NPs) on the aggregation kinetics of ß-amyloid fragment 1-40 (ßA) and α-synuclein protein using surface-enhanced Raman spectroscopy (SERS) and tip-enhanced Raman spectroscopy (TERS). The characterizations of ZnO-NPs and TiO2-NPs were evaluated by X-ray diffraction (XRD) spectrum, atomic force microscopy (AFM), and UV-Vis spectroscopy. The interaction of nanoparticles with amyloid proteins was investigated by SERS. Our study showed that exposure of amyloid protein molecules to TiO2-NPs and ZnO-NPs after incubation at 37 °C caused morphological changes and stimulated aggregation and fibrillation. In addition, significant differences in the intensity and location of active Raman frequencies in the amide I domain were found. The principal component analysis (PCA) results show that the effect of NPs after incubation at 4 °C does not cause changes in ßA structure.

Assessment of TiO2 Nanoparticle Impact on Surface Morphology of Chinese Hamster Ovary Cells.

The process of nanoparticles entering the cells of living organisms is an important step in understanding the influence of nanoparticles on biological processes. The interaction of nanoparticles with the cell membrane is the first step in the penetration of nanoparticles into cells; however, the penetration mechanism is not yet fully understood. This work reported the study of the interaction between TiO2 nanoparticles (TiO2-NPs) and Chinese hamster ovary (CHO) cells using an in vitro model. The characterization of crystalline phases of TiO2 NPs was evaluated by transmission electron microscopy (TEM), X-ray diffraction (XRD) spectrum, and atomic force microscopy (AFM). Interaction of these TiO2 nanoparticles (TiO2- NPs) with the CHO cell membrane was investigated using atomic force microscopy (AFM) and Raman spectroscopy. The XRD analysis result showed that the structure of the TiO2 particles was in the rutile phase with a crystallite size of 60 nm, while the AFM result showed that the particle size distribution had two peaks with 12.1 nm and 60.5 nm. The TEM analysis confirmed the rutile phase of TiO2 powder. Our study showed that exposure of CHO cells to TiO2-NPs caused morphological changes in the cell membranes and influenced the viability of cells. The TiO2-NPs impacted the cell membrane surface; images obtained by AFM revealed an 'ultra structure' with increased roughness and pits on the surface of the membrane. The depth of the pits varied in the range of 40-80 nm. The maximal depth of the pits after the treatment with TiO2-NPs was 100% higher than the control values. It is assumed that these pits were caveolae participating in the endocytosis of TiO2-NPs. The research results suggest that the higher maximal depth of the pits after the exposure of TiO2-NPs was determined by the interaction of these TiO2-NPs with the cell's plasma membrane. Moreover, some of pits may have been due to plasma membrane damage (hole) caused by the interaction of TiO2-NPs with membrane constituents. The analysis of AFM images demonstrated that the membrane roughness was increased with exposure time of the cells to TiO2-NPs dose. The average roughness after the treatment for 60 min with TiO2-NPs increased from 40 nm to 78 nm. The investigation of the membrane by Raman spectroscopy enabled us to conclude that TiO2-NPs interacted with cell proteins, modified their conformation, and potentially influenced the structural damage of the plasma membrane.

Endophytes from blueberry (Vaccinium sp.) fruit: Characterization of yeast and bacteria via label-free surface-enhanced Raman spectroscopy (SERS).

Blueberries (Vaccinium sp.) are consumed all around the globe, however, their endophytic community has not been thoroughly researched, specifically their fruit endophytes. We aimed to isolate and analyze easily cultivable blueberry fruit endophytes to help in future research, concerning probiotic microorganisms. Twelve strains were isolated in this pilot study, genetically homologous with Staphylococcus hominis, Staphylococcus cohnii, Salmonella enterica, Leuconostoc mesenteroides, and [Candida] santamariae. To determine the molecular composition of these isolates we used label-free surface-enhanced Raman spectroscopy (SERS). To our knowledge, this is the first time that SERS spectra for L. mesenteroides and C. santamariae are presented, as well as the first report of Candida yeast, isolated specifically from blueberry fruits. Our findings suggest that the differences in tested yeast and bacteria SERS spectra and subsequent differentiation are facilitated by minor shifts in spectral peak positions as well as their intensities. Moreover, we used principal component and discriminant function analyses to differentiate chemotypes within our isolate group, proving the sensitivity of the technique and its usefulness to recognize different strains in plant-associated microbe samples, which will aid to streamline future studies in biofertilizers and biocontrol agents.

Evaluation of a Yeast-Polypyrrole Biocomposite Used in Microbial Fuel Cells.

Electrically conductive polymers are promising materials for charge transfer from living cells to the anodes of electrochemical biosensors and biofuel cells. The modification of living cells by polypyrrole (PPy) causes shortened cell lifespan, burdens the replication process, and diminishes renewability in the long term. In this paper, the viability and morphology non-modified, inactivated, and PPy-modified yeasts were evaluated. The results displayed a reduction in cell size, an incremental increase in roughness parameters, and the formation of small structural clusters of polymers on the yeast cells with the increase in the pyrrole concentration used for modification. Yeast modified with the lowest pyrrole concentration showed minimal change; thus, a microbial fuel cell (MFC) was designed using yeast modified by a solution containing 0.05 M pyrrole and compared with the characteristics of an MFC based on non-modified yeast. The maximal generated power of the modified system was 47.12 mW/m2, which is 8.32 mW/m2 higher than that of the system based on non-modified yeast. The open-circuit potentials of the non-modified and PPy-modified yeast-based cells were 335 mV and 390 mV, respectively. Even though applying a PPy layer to yeast increases the charge-transfer efficiency towards the electrode, the damage done to the cells due to modification with a higher concentration of PPy diminishes the amount of charge transferred, as the current density drops by 846 µA/cm2. This decrease suggests that modification by PPy may have a cytotoxic effect that greatly hinders the metabolic activity of yeast.

Surface-enhanced Raman scattering sensors for biomedical and molecular detection applications in space.

The detection of molecular traces in the environment is a technical problem that is critical in pollutant control procedures at all stages of spacecraft assembly, in space flight, as well as in other technological processes such as food production, medical diagnostics, environmental control, warfare. However, in the aerospace industry, it is necessary to detect molecular traces of contaminants with extreme sensitivity, as even concentrations as low as part-per-billion (ppb) can be critical during long missions. The high sensitivity of the Volatile Organic Compounds (VOCs) detection within the air can be a challenge because of the poor affinity of VOC's to the metal surface of the sensor substrate. In this work, we present a surface-enhanced Raman scattering (SERS) spectroscopy technique as a highly sensitive and selective molecular sensor for gas trace detection not sensitive to molecules adsorbtion on sensing element. The developed hybrid SERS platform for molecular trace detection is supported by the hybrid nanoplasmonic porous silicon membrane in conjunction with micropump to achieve the trace level detection of VOCs in the environment. The combination of silicon membrane, made by electrochemical etching of the microchannels in the silicon chip, with chemical deposition of the silver nanoparticles inside the channels, produce a porous Ag nanoparticles membrane with a high density of plasmonic nanostructures ("hot spots"). The micropump integrated with the SERS sensor, pump the air with VOC's molecules through the plasmonic membrane "hot spots" to increase the probability of interaction of VOC's molecules with SERS substrate and to increase the enhancement factor. The sensor chip structure was designed, gas flow in the sensor was simulated, and the sensor was fabricated using 3D printing. The limit of detection of hydrazine with concentration level 10-12 M from solution and the vapor phase 0.1 ppm was demonstrated. The anisole vapors with concentration 0.5 ppb spectra in the air were recorded. Our results demonstrate that plasmonic membrane can be used as a high enhancement factor SERS sensor for many pollutants molecules detection with the nanomolar sensitivity and can be applied in the design of sensors for space applications, environment control, biomedical diagnostic. Supplementary Information: The online version contains supplementary material available at 10.1007/s12567-021-00356-6.

Baker's Yeast-Based Microbial Fuel Cell Mediated by 2-Methyl-1,4-Naphthoquinone.

Microbial fuel cell (MFC) efficiency depends on charge transfer capability from microbe to anode, and the application of suitable redox mediators is important in this area. In this study, yeast viability experiments were performed to determine the 2-methyl-1,4-naphthoquinone (menadione (MD)) influence on different yeast cell species (baker's yeast and Saccharomyces cerevisiae yeast cells). In addition, electrochemical measurements to investigate MFC performance and efficiency were carried out. This research revealed that baker's yeast cells were more resistant to dissolved MD, but the current density decreased when yeast solution concentration was incrementally increased in the same cell. The maximal calculated power of a designed baker's yeast-based MFC cell anode was 0.408 mW/m2 and this power output was registered at 24 mV. Simultaneously, the cell generated a 62-mV open circuit potential in the presence of 23 mM potassium ferricyanide and the absence of glucose and immobilized MD. The results only confirm that MD has strong potential to be applied to microbial fuel cells and that a two-redox-mediator-based system is suitable for application in microbial fuel cells.

Towards Microorganism-Based Biofuel Cells: The Viability of Saccharomyces cerevisiae Modified by Multiwalled Carbon Nanotubes.

This research aimed to evaluate the toxic effect of multi-walled carbon nanotubes (MW-CNTs) on yeast cells in order to apply MW-CNTs for possible improvement of the efficiency of microbial biofuel cells. The SEM and XRD analysis suggested that here used MW-CNTs are in the range of 10-25 nm in diameter and their structure was confirmed by Raman spectroscopy. In this study, we evaluated the viability of the yeast Saccharomyces cerevisiae cells, affected by MW-CNTs, by cell count, culture optical density and atomic force microscopy. The yeast cells were exposed towards MW-CNTs (of 2, 50, 100 µg/mL concentrations in water-based solution) for 24 h. A mathematical model was applied for the evaluation of relative growth and relative death rates of yeast cells. We calculated that both of the rates are two times higher in the case if yeasts were treated by 50, 100 µg/mL of MW-CNTs containing solution, comparing to that treated by 0 and 2 µg/mL c of MW-CNTs containing solution. It was determined that the MW-CNTs have some observable effect upon the incubation of the yeast cells. The viability of yeast has decreased together with MW-CNTs concentration only after 5 h of the treatment. Therefore, we predict that the MW-CNTs can be applied for the modification of yeast cells in order to improve electrical charge transfer through the yeast cell membrane and/or the cell wall.

Evanescent-field-induced Raman scattering for bio-friendly fingerprinting at sub-cellular dimension.

Evanescent field induced chemical imaging concept has been realized in analytical platform based on the µ-tip-enhanced Raman scattering spectroscopy (µ-TERS). The technique aimed to minimize thermal decomposition of dried biological sample as the result of huge concentration of optical field near the tip by increasing the size of an aperture-less "excitation source". µ-TERS technique is similar to classical biosensor systems based on propagating surface plasmon resonance phenomenon but with sensitive elements a few micrometers in size that can be targeted to the area of interest. The utility of the concept is exemplified by the analysis of dried single cell envelope of genetically modified Saccharomyces cerevisiae yeast cells, which do not have any heat-removing pathways, by water as in the case of the living cell. Practical excitation conditions effective for µ-TERS Raman observation of single layer dried biological samples without photodamage-related spectral distortion have been determined - the allowable limit is above 30s at 13 µW/µm(2). Finally, potential of µ-TERS spectroscopy as new bio-friendly instrumental platform for chemical fingerprinting and analytical characterization of buried nanoscale features is discussed.

In vivo characterization of protein uptake by yeast cell envelope: single cell AFM imaging and µ-tip-enhanced Raman scattering study.

Direct detection of biological transformations of single living cells in vivo has been performed by the advanced combination of local topographic imaging by Atomic Force Microscopy (AFM) and label-free sub-surface chemical characterization using new µ-Tip-Enhanced Raman Spectroscopy (µ-TERS). The enhancing mechanism for µ-TERS tips with micrometre range radius differs significantly to that of the conventional tapered structures terminated by a sharp apex and conditioned by the effects of propagating instead of localizing surface plasmon resonance phenomena. Sub-wavelength light confinement in the form of a nonradiative evanescent wave near the tip surface with penetration depth in the sub-micrometre range opens the way for monitoring of subsurface processes near or within the cell wall, inaccessible by other methods. The efficiency of the approach has been demonstrated by the analysis of the cell envelope of genetically modified (by glucose dehydrogenase (GDH) gene bearing Kluyveromyces lactis toxin signal sequence) yeast cells enriched by GDH protein. The presence of trans-membrane fragments in GDH together with the tendency to form active dimers and tetramers causes the accumulation of the proteins within the periplasmic space. These results demonstrate that the advanced combination of AFM imaging and subsurface chemical characterization by the novel µ-TERS technique provides a new analytical tool for the investigation of single living cells in vivo.