Molecules Of Interest Research Articles

Determination of Oxygen Heterocyclic Compounds in Foods Using Supercritical Fluid Chromatography-Tandem Mass Spectrometry.

The aim of this research was to determine oxygen heterocyclic compounds in twenty-six Citrus- and cinnamon-flavoured foods using supercritical fluid chromatography in combination with triple-quadrupole mass spectrometry (SFC-QqQ-MS). According to the authors' knowledge, this is the first report on the determination of these molecules in foods by means of the SFC-QqQ-MS technique. The analytical technique normally used for their determination in foods is liquid chromatography coupled with a photodiode array detector. However, supercritical fluid chromatography is proving to be a valid alternative approach to investigating coumarins, furocoumarins and polymethoxyflavones. According to the results presented herein, each sample analysed showed the presence of molecules of interest. Coumarin was found in all the cinnamon-flavoured samples analysed in a low concentration. The presence of oxygen heterocyclic compounds in all the Citrus-flavoured samples, according to the label, comfirmed that the foods selected for this research article were prepared with Citrus fruits. Among the samples analysed, mandarin juice was the richest in bioactive compounds, representing a good source of polymethoxyflavones in a diet.

A Review on the Antimutagenic and Anticancer Effects of Cysteamine.

Cancer is one of the leading causes of death worldwide. First-line treatments usually include surgery, radiotherapy, and/or systemic therapy. These methods can be associated with serious adverse events and can be toxic to healthy cells. Despite the new advances in cancer therapies, there is still a continuous need for safe and effective therapeutic agents. Cysteamine is an aminothiol endogenously synthetized by human cells during the degradation of coenzyme-A. It has been safely used in humans for the treatment of several pathologies including cystinosis and neurodegenerative diseases. Cysteamine has been shown to be a potent antimutagenic, anticarcinogenic, and antimelanoma in various in vitro and in vivo studies, but a review on these aspects of cysteamine's use in medicine is lacking in the current literature. The efficacy of cysteamine has been shown in vitro and in vivo for the treatment of different types of cancer, such as gastrointestinal cancer, pancreatic cancer, sarcomas, hepatocellular carcinoma, and melanoma, leading to the significant reduction of lesions and/or the increase of survival time. Although the mechanisms of action are not fully understood, possible explanations are (i) free radical scavenging, (ii) alteration of the tumor cell proliferation by affecting nucleic acid and protein synthesis or inhibition of DNA synthesis, and (iii) hormone regulation. In conclusion, regarding the high safety profile of cysteamine and the current literature data presented in this article, cysteamine might be considered as an interesting molecule for the prevention and the treatment of cancer. Further clinical studies should be performed to support these data in humans.

The fungal quorum-sensing molecule, farnesol, regulates the immune response of vaginal epithelial cells against Candida albicans

BackgroundFarnesol is a Candida-secreted quorum-sensing molecule of great interest as a potential antifungal agent for serious and hardly curable infections—candidiasis, especially vulvovaginal candidiasis (VVC).MethodsThe effect of farnesol on cellular morphology and viability and evaluated the production of Th1 (IL-2), Th2 (IL-4), proinflammatory (IL-6), chemotactic (IL-8), and Th17 (IL-17) cytokines in the culture supernatants of vaginal epithelial cell line (VK2) were evaluated. Moreover, we tested the inhibitory effect of farnesol on C. albicans adhesion. Scanning electron microscopy was conducted to observe any VK2 cell ultrastructural changes.ResultsOnly low concentrations (≤ 50 µmol/L) of farnesol did not affect the morphology and viability of the VK2 cells (P > 0.05). Farnesol reduced the adhesion of C. albicans to the VK2 cells. When treated with farnesol, statistical elevated levels of both IL-4 and IL-17 secreted by the infected VK2 cells were present in the culture supernatants (P < 0.05).ConclusionsFarnesol acts as a stimulator to up-regulate the Th17-type innate immune response, as well as Th2-type humoral immunity following C. albicans infection. Further research is required to select the optimal therapeutic dose to develop efficacious and safe mucosal immune adjuvant for treating VVCs.

4a′-Hydroxy-3′,3′,5,6′,6′,7-hexamethyl-3′,4′,4a′,6′,7′,9a′-hexahydrospiro[indole-3,9′-xanthene]-1′,2,8′(1H,2′H,5′H)-trione

Pseudo-multicomponent reactions (Pseudo-MCRs) have led to a variety of compounds with interesting biological properties, especially desirable in the pharmaceutical industry. The isatin nucleus could be considered a privileged scaffold for the design of biologically active substances. Dimedone is an interesting and versatile molecule for most organic transformations, especially one-pot and multicomponent reactions. Xanthene derivatives are still an attractive research field for both academia investigations and industry. In this investigation, a simple and efficient tandem Knoevenagel–Michael protocol with subsequent cyclization for the synthesis of the previously unknown 4a′-hydroxy-3′,3′,5,6′,6′,7-hexamethyl-3′,4′,4a′,6′,7′,9a′-hexahydrospiro[indole-3,9′-xanthene]-1′,2,8′(1H,2′H,5′H)-trione was elaborated. The suggested method is based on the pseudo-MCR of 5,7-dimethylisatin and dimedone. The structure of the earlier unknown compound was proven using 1H, 13C-NMR, and IR spectroscopy, mass spectrometry, and elemental analysis. To compare the developed protocol with the existing ones, unsubstituted spiro[indole-3,9′-xanthene] was synthesized. Its structure has been proven using two-dimensional (2D) NMR spectroscopy techniques.

Non-canonical two-step biosynthesis of anti-oomycete indole alkaloids in Kickxellales

BackgroundFungi are prolific producers of bioactive small molecules of pharmaceutical or agricultural interest. The secondary metabolism of higher fungi (Dikarya) has been well-investigated which led to > 39,000 described compounds. However, natural product researchers scarcely drew attention to early-diverging fungi (Mucoro- and Zoopagomycota) as they are considered to rarely produce secondary metabolites. Indeed, only 15 compounds have as yet been isolated from the entire phylum of the Zoopagomycota.ResultsHere, we showcase eight species of the order Kickxellales (phylum Zoopagomycota) as potent producers of the indole-3-acetic acid (IAA)-derived compounds lindolins A and B. The compounds are produced both under laboratory conditions and in the natural soil habitat suggesting a specialized ecological function. Indeed, lindolin A is a selective agent against plant-pathogenic oomycetes such as Phytophthora sp. Lindolin biosynthesis was reconstituted in vitro and relies on the activity of two enzymes of dissimilar evolutionary origin: Whilst the IAA–CoA ligase LinA has evolved from fungal 4-coumaryl-CoA synthetases, the subsequently acting IAA-CoA:anthranilate N-indole-3-acetyltransferase LinB is a unique enzyme across all kingdoms of life.ConclusionsThis is the first report on bioactive secondary metabolites in the subphylum Kickxellomycotina and the first evidence for a non-clustered, two-step biosynthetic route of secondary metabolites in early-diverging fungi. Thus, the generally accepted “gene cluster hypothesis” for natural products needs to be reconsidered for early diverging fungi.

Molecular Properties of 3d and 4f Coordination Compounds Deciphered by Raman Optical Activity Spectroscopy.

Molecular properties of coordination compounds can be efficiently studied by vibrational spectroscopy. The scope of Raman spectroscopy has been greatly enhanced by the introduction of Raman optical activity (ROA) sensitive to chirality. The present review describes some of its recent applications to study the coordination compounds. 3d and 4f metal complexes often absorb the excitation light, or exhibit luminescence. Therefore, effects caused in ROA spectra by electronic circular dichroism (ECD) and circularly polarized luminescence (CPL) must be taken into consideration.In 3d metal complexes ECD and circularly-polarized Raman scattering compete with the resonance ROA (RROA) signal. Pure RROA spectrum can thus be obtained by subtracting the so-called ECD-Raman component. CPL is frequently encountered in 4f systems. While it can mask the ROA spectra, it is useful to study molecular structure. These electronic effects can be reduced by using near-infrared excitation although vibrational ROA signal is much weaker compared to the usual green laser excitation scenario. The ROA methodology is thus complex, but capable of providing unique information about the molecules of interests and their interaction with light.

C-Phycocyanin: A Phycobiliprotein from Spirulina with Metabolic Syndrome and Oxidative Stress Effects.

Spirulina maxima is a cyanobacterium considered a "superfood" due to its metabolites and nutrient content. These include a complex mixture of minerals, vitamins, fatty acids, proteins, and accessory pigments. In recent years, it has positioned itself as a promising source of bioactive molecules for the treatment of several diseases, including metabolic syndrome, coronary diseases, cancer, and the improvement of health modulating oxidative stress. C-Phycocyanin (C-PC) is a photosynthetic pigment from green-blue cyanobacterium and the most abundant phycobiliprotein in the Spirulina genus with various pharmacological properties attributed due to its antioxidant capacity but has no specific cellular target. This has made it a molecule of great interest in biomedical research. This review focuses on the pharmacological effects and the benefits on metabolic syndrome and oxidative stress of C-PC.

Sampling Cerebrospinal Fluid and Blood from Lateral Tail Vein in Rats during EEG Recordings.

Because the composition of body fluids reflects many physiological and pathological dynamics, biological liquid samples are commonly obtained in many experimental contexts to measure molecules of interest, such as hormones, growth factors, proteins, or small non-coding RNAs. A specific example is the sampling of biological liquids in the research of biomarkers for epilepsy. In these studies, it is desirable to compare the levels of molecules in cerebrospinal fluid (CSF) and in plasma, by withdrawing CSF and plasma in parallel and considering the time distance of the sampling from and to seizures. The combined CSF and plasma sampling, coupled with video-EEG monitoring in epileptic animals, is a promising approach for the validation of putative diagnostic and prognostic biomarkers. Here, a procedure of combined CSF withdrawal from cisterna magna and blood sampling from the lateral tail vein in epileptic rats that are continuously video-EEG monitored is described. This procedure offers significant advantages over other commonly used techniques. It permits rapid sampling with minimal pain or invasiveness, and reduced time of anesthesia. Additionally, it can be used to obtain CSF and plasma samples in both tethered and telemetry EEG recorded rats, and it may be used repeatedly across multiple days of experiment. By minimizing the stress due to sampling by shortening isoflurane anesthesia, measures are expected to reflect more accurately the true levels of investigated molecules in biofluids. Depending on the availability of an appropriate analytical assay, this technique may be used to measure the levels of multiple, different molecules while performing EEG recording at the same time.

Electrochemical Properties of Derivatives of 1,3-Diphenylisobenzofuran – Chromophores for Singlet Fission

Nowadays, there are efforts to apply alternative sources of renewable energy. One of these sources is solar energy. Solar energy represents a renewable free source of energy that is sustainable and totally inexhaustible. The standard efficiency of solar cells is now below 30 %. Singlet fission has attracted rising attention because it promises to increase the maximum of theoretical efficiency of single-junction solar cells. While solar cells after absorption one photon generates one electron, singlet fission is a photophysical process, whereby a singlet exciton generated by irradiation splits into two triplet excitons [1,2]. In this way, one photon can generate theoretically two electrons which should increase the efficiency of inexpensive single-junction solar cells. Therefore, recently substantial attention has been paid to search for suitable chromophores for singlet fission.When looking for new chromophores for efficient SF, their crystalline form and molecular packing, the relative energies of the S0 (ground state), T1 (lowest triplet), and S1 (first excited singlet) states, where energy of the triplet state should be lower than a half of the first excited singlet energy, and the potentials of one-electron reduction and one-electron oxidation are crucial. Redox properties of molecules for SF are thus critical for their use, therefore electrochemical approach is necessary.Molecules on the base of 1,3-diphenylisobenzofuran (DPIBF) attract an interest because of their possible efficiency for singlet fission. In the last few years we have started to investigate various types of derivatives of DPIBF. One of our first studies [3] was focused on a series of fluorinated derivatives of DPIBF where the influence of number and position of fluorine atoms in the molecule on the redox potentials and mechanism is followed. Next to that we studied series of dimer derivatives of DPIBF [4]. The location and the nature of the bridging unit between two DPIBF redox centres attracted our attention because in this way the shape of the molecule, its extent of electron delocalization and, thus, possible ability of formation of biradicaloid can be changed.The most interesting dimeric molecule appeared to be that one with the elongated π-conjugated system, the “quasi dimer” of DPIBF (2). Because of that, quasi dimer was investigated in more details in comparison with other three molecules by standard electrochemical techniques, the in-situ UV-vis and EPR spectroelectrochemical techniques. The three other derivatives for comparison were parent diphenylisobenzofuran (1), direct dimer (3) and methylene bridged dimer (4). The main goal of this deeper study was to characterize properly not only electrochemical properties of the molecules, but also their absorbance and fluorescence properties and stability in solvents. Acknowledgement This work was supported by the grant 21-23261S (Grant Agency of the Czech Republic) and by the institutional support RVO: 61388955.

Valorization of bio-colorants extracted from Hypercium scabrum L. plant for sustainable and ecological coloration of wool yarns

Recently, natural dyes are being explored all over the world as safer and highly sustainable bio-based alternatives to synthetic dyes. Agricultural wastes and plant by-products are the most commonly explored alternatives with dual benefits of waste reclamation and sustainable dye production with extra value-adds. Hypercium scabrum plant contains interesting bio-dye molecules with high flavonoids and tannin contents. The present study aims at exploring the potential of H. scabrum plant extract to color wool textiles with a focus on sustainable bio-dye production and fastness properties. The extracted bio-dye was quantitatively (for total phenolic (2.733 mg per CE/g) and total flavonoid (1.140 mg per GAE/g) content using the Folin–Ciocalteu method) and qualitatively (UV–Vis, FT-IR, and EDX) characterized. The effect of dyeing parameters like pH (2–8), temperature (60–90 °C), dry-weight content of plant material as a dye (25–150% o.w.f.), and dyeing time (15–120 min) on color strength (K/S) values were assessed. Color fastness assays showed good resistance to light, washing, and rubbing. The effect of artificial aging (Xenon arc lamp) on the color strength of dyed wool yarns under different exposure times (0–48 h) was explored. The highest color fading occurred in control dyed samples with a first-order rate constant of 131.57 h−1 and a half-life period of 5.26 x 10−3 h. Color difference (ΔE) values suggested that mordanted samples showed less fading compared to control dyed samples at equal times of Xenon exposure. Additionally, the dyed samples were washed in double distilled water, tap water, and 4 g/L NaCl solution to check their effects on the corresponding K/S values while 4 g/L NaCl solution mimics the real conditions of perspiration. Maximum color leaching occurred in 4 g/L NaCl washing with a first-order rate constant of 11.57 min−1. Cost analysis of the dye extraction and dyeing procedure revealed that the process is sustainable and economical. Thus, the use of H. scabrum whole plant can provide a clean, economical, and sustainable source of alternative natural dyes that can be used to substitute synthetic analogs.

Impedance Spectroscopy: A New Tool for Dairy Analysis

In this work, an impedimetric bioelectronic tongue (bioET) formed by an array of interdigitated microelectrodes sensors was developed and applied in the discrimination of chemical components usually found in milk samples. Few studies of impedimetric bioETs applied in the dairy industry can be found in the bibliography due to the complexity of the characterization of milk matrixes. Therefore, the development and study of the capabilities of multisensory systems containing different advanced materials such as nanomaterials combined with biological molecules is a new field of research.In order to develop an array of sensors with enhanced sensitivity and selectivity, sensors were modified with silver nanoparticles (AgNPs) and enzymes including galactose oxidase (GaOx), glucose oxidase (GOx), beta-galactosidase (β-gal), urease (Ure) and lipase (Lip) were immobilized on the sensors surface. Silver nanoparticles have proved to be an excellent option to enhance the performance of biosensors due to their electrochemical properties, their high electrical conductivity, and their ability to amplify bio electrochemical signals.Atomic force microscopy (AFM) was used to study the distribution of AgNPs and enzymes in the sensor surface. Silver nanoparticles showed a homogeneous arrangement along the entire surface, moreover after the enzyme immobilisation a pronounced increase in the sensors surface roughness was observed ensuring the process of the biological molecule (Figure 1.A and Figure 1.B).The impedance analysis of standard solutions of interest in milk (glucose, galactose, lactose, urea and triglycerides) demonstrated that sensors with a higher concentration of silver nanoparticles (AgNPs) in their structure reported higher sensitivities towards compounds found in milk with a reduction in the impedance module, indicating that there has been an improvement in electronic transfer due to the interaction of the nanomaterial and the electrode surface an example of the sensor behaviour is shown in Figure 1.C. Moreover, sensors that combined the nanomaterial with an enzyme showed a greater ability to differentiate between increasing concentrations of the enzymes target molecules (Figure 1.D). Taking as an example the case of glucose the sensor modified with AgNPs and glucose oxidase achieved a lower impedance module compared with the other sensors thanks to its lower resistance to electronic transfer in the sensor due to the enzymatic activity enhanced by the silver nanoparticles.Once all the standard solutions had been analysed, the chemometric analysis of their response was carried out applying a principal components analysis (PCA) approach which showed that the bioET developed was able to discriminate between the standard solutions (glucose, galactose, lactose, urea and triglycerides), resulting in five clusters according to their nature with a 79% of the variance of the original data obtained by the first two components of the system. Sugars appear in the positive region of the first component divided into three groups (lactose, galactose and glucose). In the negative region of the first component appears urea and the suspension of triglycerides that can be differentiated over the second component.Lastly, a first approach was made to the analysis of milk samples with different nutritional characteristics such as their fat, lactose and urea content. The utilization of the bioelectronic tongue showed how the different contents of molecules of interest was directly related with the response of the impedimetric sensors due to changes in the interaction of the sensors surface and the milk matrix. As it was expected, higher concentrations of fat caused an increase in the impedance module due to their interference in the electronic transfer process. Furthermore, it was confirmed that the biosensors developed showed specific responses towards milk samples with higher values of sugar and proteins contents ensuring a high degree of cross-selectivity in the system.Figure 1: A) Characterization by AFM of the sensor modified with AgNPs. B) Characterization by AFM of the biosensor with AgNPs and GOx. C) Nyquist diagram of sensors with 20 (orange), 30 (black) and 40 μl (blue) of AgNPs in 0.1M glucose. D) Nyquist diagram of the response of a sensor with AgNPs and GOx in glucose 0.1M (red), 0.01M (green) and 0.001M (grey). Figure 1

Direct early growth response-1 knockdown decreases melanoma viability independent of mitogen-activated extracellular signal-related kinase inhibition.

To investigate downstream molecular changes caused by mitogen-activated protein kinase (MEK) inhibitor treatment and further explore the impact of direct knockdown of early growth response-1 ( EGR1 ) in melanoma cell culture. RNA-sequencing (RNA-Seq) was performed to determine gene expression changes with MEK inhibitor treatment. Treatment with MEK inhibitor (trametinib) was then assessed in two cutaneous (MEL888, MEL624) and one conjunctival (YUARGE 13-3064) melanoma cell line. Direct knockdown of EGR1 was accomplished using lentiviral vectors containing shRNA. Cell viability was measured using PrestoBlueHS Cell Viability Reagent. Total RNA and protein were assessed by qPCR and SimpleWestern. RNA-Seq demonstrated a profound reduction in EGR1 with MEK inhibitor treatment, prompting further study of melanoma cell lines. Following trametinib treatment of melanoma cells, viability was reduced in both cutaneous (MEL888 26%, P < 0.01; MEL624 27%, P < 0.001) and conjunctival (YUARGE 13-3064 33%, P < 0.01) melanoma compared with DMSO control, with confirmed EGR1 knockdown to 0.04-, 0.01-, and 0.16-fold DMSO-treated levels (all P < 0.05) in MEL888, MEL624, and YUARGE 13-3064, respectively. Targeted EGR1 knockdown using shRNA reduced viability in both cutaneous (MEL624 78%, P = 0.05) and conjunctival melanoma (YUARGE-13-3064 67%, P = 0.02). RNA-Sequencing in MEK inhibitor-treated cells identified EGR1 as a candidate effector molecule of interest. In a malignant melanoma cell population, MEK inhibition reduced viability in both cutaneous and conjunctival melanoma with a profound downstream reduction in EGR1 expression. Targeted knockdown of EGR1 reduced both cutaneous and conjunctival melanoma cell viability independent of MEK inhibition, suggesting a key role for EGR1 in melanoma pathobiology.

PCS/Bonds and PCS0: Pick your molecule and get its accurate structure and ground state rotational constants at DFT cost.

An unsupervised computational protocol is proposed with the aim of obtaining accurate structures of large molecules in the gas phase at the cost of standard density functional theory (DFT) computations. The whole workflow is fully automated and provides optimized equilibrium geometries and ground state rotational constants to be directly compared with experiments. The results for a panel of molecules of biological or medicinal interest show that the accuracy of the results delivered by the new tool at the cost of a single DFT geometry optimization is close to that delivered by state-of-the-art composite wavefunction methods for small semi-rigid molecules.

Nuclear receptor superfamily structural diversity in pacific oyster: In silico identification of estradiol binding candidates

The increasing presence of anthropogenic contaminants in aquatic environments poses challenges for species inhabiting contaminated sites. Due to their structural binding characteristics to ligands that inhibit or activate gene transcription, these xenobiotic compounds frequently target the nuclear receptor superfamily. The present work aims to understand the potential interaction between the hormone 17-β-estradiol, an environmental contaminant, and the nuclear receptors of Crassostrea gigas, the Pacific oyster. This filter-feeding, sessile oyster species is subject to environmental changes and exposure to contaminants. In the Pacific oyster, the estrogen-binding nuclear receptor is not able to bind this hormone as it does in vertebrates. However, another receptor may exhibit responsiveness to estrogen-like molecules and derivatives. We employed high-performance in silico methodologies, including three-dimensional modeling, molecular docking and atomistic molecular dynamics to identify likely binding candidates with the target moecule. Our approach revealed that among the C. gigas nuclear receptor superfamily, candidates with the most favorable interaction with the molecule of interest belonged to the NR1D, NR1H, NR1P, NR2E, NHR42, and NR0B groups. Interestingly, NR1H and NR0B were associated with planktonic/larval life cycle stages, while NR1P, NR2E, and NR0B were associated with sessile/adult life stages. The application of this computational methodological strategy demonstrated high performance in the virtual screening of candidates for binding with the target xenobiotic molecule and can be employed in other studies in the field of ecotoxicology in non-model organisms.

Controllable Photocleavage of Blebbistatin Derivatives as Photoremovable Protecting Groups.

Blebbistatin was demonstrated as a promising two-photon near-infrared activated photoremovable protecting group of hydroxyl radicals with various potential applications. However, the photocleavage mechanism of the blebbistatin derivatives remains ambiguous. Herein, blebbistatin derivatives with various electronic characteristic leaving groups were synthesized and studied, and the photocleavage mechanism(s) and the tunable effect of the leaving groups were unveiled by combining photoproduct analysis, reactive oxygen radical species detection, femtosecond transient absorption spectroscopy, and density functional theory calculation. More substantial electron-withdrawing leaving groups facilitate heterolysis of the C-O bond, which results in a cationic intermediate and a corresponding remnant. Weaker electron-withdrawing groups lead to a higher proportion of homolysis of the C-O bond, accompanied by the generation of the reactive oxygen radical species. With this structure-property relationship, the protected groups of the molecules of interest can be rationally chosen to satisfy the different requirements needed for specific applications.

Bigels as Delivery Systems: Potential Uses and Applicability in Food.

Bigels have been mainly applied in the pharmaceutical sector for the controlled release of drugs or therapeutics. However, these systems, with their intricate structures, hold great promise for wider application in food products. Besides their classical role as carrier and target delivery vehicles for molecules of interest, bigels may also be valuable tools for building complex food structures. In the context of reducing or even eliminating undesirable (but often highly functional) food components, current strategies often critically affect food structure and palatability. The production of solid fat systems that are trans-fat-free and have high levels of unsaturated fatty acids is one of the challenges the food industry currently faces. According to recent studies, bigels can be successfully used as ingredients for total or partial solid fat replacement in complex food matrices. This review aims to critically assess current research on bigels in food and pharmaceutical applications, discuss the role of bigel composition and production parameters on the characteristics of bigels and further expand the use of bigels as solid fat replacers and functional food ingredients. The hydrogel:oleogel ratio, selected gelators, inclusion of surfactants and encapsulation of molecules of interest, and process parameters (e.g., temperature, shear rate) during bigel production play a crucial role in the bigel's rheological and textural properties, microstructure, release characteristics, biocompatibility, and stability. Besides exploring the role of these parameters in bigel production, future research directions for bigels in a food context are explored.

Exploring the therapeutic potential of an antinociceptive and anti-inflammatory peptide from wasp venom

Animal venoms are rich sources of neuroactive compounds, including anti-inflammatory, antiepileptic, and antinociceptive molecules. Our study identified a protonectin peptide from the wasp Parachartergus fraternus' venom using mass spectrometry and cDNA library construction. Using this peptide as a template, we designed a new peptide, protonectin-F, which exhibited higher antinociceptive activity and less motor impairment compared to protonectin. In drug interaction experiments with naloxone and AM251, Protonectin-F's activity was decreased by opioid and cannabinoid antagonism, two critical antinociception pathways. Further experiments revealed that this effect is most likely not induced by direct action on receptors but by activation of the descending pain control pathway. We noted that protonectin-F induced less tolerance in mice after repeated administration than morphine. Protonectin-F was also able to decrease TNF-α production in vitro and modulate the inflammatory response, which can further contribute to its antinociceptive activity. These findings suggest that protonectin-F may be a potential molecule for developing drugs to treat pain disorders with fewer adverse effects. Our results reinforce the biotechnological importance of animal venom for developing new molecules of clinical interest.

Spatial-temporal tracking of molecular species in Geant4-DNA track structure simulations

Monte Carlo particle transport codes have been used to simulate the radiolysis of water and subsequent evolution of molecular species during the chemical stage. In Geant4, accounting of molecules is performed using a molecule counter which provides only a global molecule count. We developed a modular counter implementation that was used to perform spatial-temporal tracking of molecule counts. This custom counter was validated in homogeneous and simplistic split geometries for 1 MeV electrons and 50 MeV protons.Using a spherical cell with one nucleus and one hundred mitochondria as an example, the radiolytic yields in the cytoplasm, nucleus, and mitochondria were compared between targeted (electrons originating from within mitochondria) and unspecific (electrons originating from within cytoplasm) for (very) low energy electrons with kinetic energies of 100 eV to 1 keV.At short times radiolytic yields in source volumes were consistent with reference yields obtained in a large homogeneous volume. Past 1 ns diffusion out of the source volumes led to yield reductions. For volumes abutting the source volumes radiolytic yields at short times strongly depended on track end effects, most pronounced for 100 eV electrons. Similarly, diffusion out of the source volumes greatly increased yields in abutting volumes past 1 ns with magnitude decreasing and time of increase increasing with increasing electron energy. For higher energy electron beams (100 keV and 200 keV) radiolytic yields in all volumes were consistent with reference yields for large statistics.Thus, in cases where energy deposit sites are uniformly distributed in the volume of interest and energies of the charged particles are high to avoid track end effects reference yields may be used in place of resource intensive chemical stage simulations. Spatially heterogeneous energy deposits, as observed in targeted ɑ or β radiation therapy or with targeted nanoparticles, using a spatially sensitive molecule counter is suggested when estimating radiolytic yields outside of the source volumes or at distances that are within the diffusion range of the molecules of interest.

Immunofluorescence Microscopy.

Visualizing fluorescence-tagged molecules is a powerful strategy that can reveal the complex dynamics of the cell. One robust and broadly applicable method is immunofluorescence microscopy, in which a fluorescence-labeled antibody binds the molecule of interest and then the location of the antibody is determined by fluorescence microscopy. The effective application of this technique includes several considerations, such as the nature of the antigen, specificity of the antibody, permeabilization and fixation of the specimen, and fluorescence imaging of the cell. Although each protocol will require fine-tuning depending on the cell type, antibody, and antigen, there are steps common to nearly all applications. This article provides protocols for staining the cytoskeleton and organelles in two very different kinds of cells: flat, adherent fibroblasts and thick, free-swimming Tetrahymena cells. Additional protocols enable visualization with widefield, laser scanning confocal, and eSRRF super-resolution fluorescence microscopy. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Immunofluorescence staining of adherent cells such as fibroblasts Basic Protocol 2: Immunofluorescence of suspension cells such as Tetrahymena Basic Protocol 3: Visualizing samples with a widefield fluorescence microscope Alternate Protocol 1: Staining suspension cells adhered to poly-l-lysine-coated coverslips Alternate Protocol 2: Visualizing samples with a laser scanning confocal microscope Alternate Protocol 3: Generating super-resolution images with SRRF microscopy.

Kinetic Extraction of Fucoxanthin from Undaria pinnatifida Using Ethanol as a Solvent.

Fucoxanthin (Fx) has been proven to exert numerous biological properties, which makes it an interesting molecule with diverse industrial applications. In this study, the kinetic behavior of Fx was studied to optimize three variables: time (t-3 min to 7 days), temperature (T-5 to 85 °C), and concentration of ethanol in water (S-50 to 100%, v/v), in order to obtain the best Fx yield from Undaria pinnatifida using conventional heat extraction. The Fx content (Y1) was found through HPLC-DAD and expressed in µg Fx/g of algae sample dry weight (AS dw). Furthermore, extraction yield (Y2) was also found through dry weight analysis and was expressed in mg extract (E)/g AS dw. The purity of the extracts (Y3) was found and expressed in mg Fx/g E dw. The optimal conditions selected for Y1 were T = 45 °C, S = 70%, and t = 66 min, obtaining ~5.24 mg Fx/g AS; for Y2 were T = 65 °C, S = 60%, and t = ~10 min, obtaining ~450 mg E/g AS; and for Y3 were T = 45 °C, S = 70%, and t = 45 min, obtaining ~12.3 mg Fx/g E. In addition, for the selected optimums, a full screening of pigments was performed by HPLC-DAD, while phenolics and flavonoids were quantified by spectrophotometric techniques and several biological properties were evaluated (namely, antioxidant, antimicrobial, and cholinesterase inhibitory activity). These results could be of interest for future applications in the food, cosmetic, or pharmaceutical industries, as they show the Fx kinetic behavior and could help reduce costs associated with energy and solvent consumption while maximizing the extraction yields.