Gadolinium Accumulation and Toxicity on In Vitro Grown Stevia rebaudiana: A Case-Study on Gadobutrol.

Gadolinium-based contrast agents are molecular complexes which are extensively used for diagnostic purposes. Apart from their tremendous contribution to disease diagnostics, there are several issues related to their use. They are extremely stable complexes and potential contaminants of surface and ground waters, an issue which is documented worldwide. The irrigation of fields with contaminated surface waters or their fertilization with sludge from wastewater treatment plants can lead to the introduction of Gd into the human food supply chain. Thus, this study focused on the potential toxicity of Gd on plants. For this purpose, we have studied the molecular effects of gadobutrol (a well-known MRI contrast agent) exposure on in vitro-grown Stevia rebaudiana. The effects of gadobutrol on plant morphology, on relevant plant metabolites such as chlorophylls, carotenoids, ascorbic acids (HPLC), minerals (ICP-OES), and on the generation of free radical species (MDA assay and EPR) were assessed. Exposures of 0.01, 0.05, 0.1, 1, and 3 mM gadobutrol were used. We found a correlation between the gadobutrol dose and the plant growth and concentration of metabolites. Above the 0.1. mM dose of gadobutrol, the toxic effects of Gd+3 ions became significant.

Cellular Internalization of Beta-Carotene Loaded Polyelectrolyte Multilayer Capsules by Raman Mapping.

Raman mapping is becoming a very useful tool in investigating cells and cellular components, as well as bioactive molecules intracellularly. In this study, we have encapsulated beta-carotene using a layer-by-layer technique, as a way to enhance its stability and bioavailability. Further, we have used Raman mapping to characterize the as-obtained capsules and monitor their uptake by the human retinal epithelial D407 cells. We were able to successfully map the beta-carotene distribution inside the capsules, to localize the capsules intracellularly, and distinguish between capsules and other cellular components.

Knee osteoarthritis grading by resonant Raman and surface-enhanced Raman scattering (SERS) analysis of synovial fluid.

In this preliminary study on synovial fluid (SF), knee osteoarthritis (OA) grading of n = 23 patients was accomplished by combining two methods: resonant Raman spectroscopy, and surface-enhanced Raman scattering (SERS) of native proteins acquired with iodide-modified silver nanoparticles and a laser emitting at 633 nm. Based on principal component analysis-linear discriminant analysis (PCA-LDA), the SERS spectra of proteins enabled the classification of low-grade and high-grade OA groups with an accuracy of 91%. Resonant Raman spectra of SF, recorded with laser excitation at 532 nm, exhibited carotenoid-associated bands that were less intense in the case of high-grade knee OA patients. Based on the resonant Raman spectra, the grading of OA patients was accomplished with an accuracy of 74%. Concatenating SERS and Raman spectral information increased the classification accuracy between the two groups to 100%. These results demonstrate the potential of Raman and SERS as a point-of-care method for aiding OA grading.

Raman spectroscopy applications in rheumatology.

Raman spectroscopy is a type of vibrational spectroscopy based on the inelastic scattering of photons, which has attracted much attention due to its potential clinical application in rheumatology. In this review, we discuss the typical spectral features of cartilage, bone, synovial fluid, and pathologic crystal deposits, as well as methods of amplifying the Raman signal of biofluids such as drop-coating deposition Raman spectroscopy. Further, applications of Raman and drop-coating deposition Raman spectroscopy in osteoarthritis are described, highlighting the clinical potential of these methods. We also discuss the role of Raman and related techniques in analyzing pathologic crystals such as monosodium urate, calcium pyrophosphate dihydrate, and hydroxyapatite. The results presented in this review demonstrate that Raman spectroscopy has grown past the stage of proof-of-concept, especially in the case of pathologies involving crystal depositions such as gout and calcium pyrophosphate deposition disease , for which the method has been validated on large number of samples. As the medical community becomes more and more aware of Raman spectroscopy, it is envisioned that it will become a standard technique in the near future.

Phytochemical Characterization of Five Edible Purple-Reddish Vegetables: Anthocyanins, Flavonoids, and Phenolic Acid Derivatives.

Vegetables comprise a significant portion of our daily diet with their high content in nutrients including fiber, vitamins, minerals, as well as phenolic compounds. Vegetable consumption has been shown to be positively associated with the prevention of several degenerative diseases thanks to their bioactive compounds. Accordingly, five selected vegetables, namely, red chicory, red onion, eggplant, purple sweet potato, and black carrot were thoroughly assessed for their phenolic content in this study. For this purpose, the total phenolic and flavonoid content of these five vegetables and their antioxidant activities that are based on three common methods ABTS radical cation decolorization assay (ABTS), Cupric Ion Reducing Antioxidant Capacity (CUPRAC), and DPPH scavenging activity assay were determined. Additionally, HPLC-PDA/Electrospray ionization coupled with mass spectrometry (HPLC-PDA/-ESI+-MS)-based identification and quantification of the members belonging to polyphenols present in each vegetable were determined. Statistical correlations between antioxidant activities and the specific type of phenolic compounds, such as anthocyanins, flavonoids, anthocyanins, and phenolic acids were further elucidated. Phenolic acids (chlorogenic and syringic acids) were found to be the most abundant compounds that are present in all samples. Among the anthocyanins, cyaniding derivatives were present in all vegetables. In terms of their antioxidant activities, the analyzed vegetables were ranked as red chicory > purple sweet potato > black carrot > eggplant > red onion, in descending order. Superior antioxidant activities exhibited by red chicory and purple sweet potato were attributed to the high content of phenolic compounds, especially flavonols (quercetin-3,4-O-diglucoside) in red chicory and anthocyanins (peonidin-3-caffeoyl p-hydroxybenzoylsophoroside-5-glucoside) in purple sweet potato.

Warfarin-Capped Gold Nanoparticles: Synthesis, Cytotoxicity, and Cellular Uptake.

Currently, research studies on nanoparticle cytotoxicity, uptake or internalization into the body's cells are of great interest for the improvement of diagnostic and therapeutic applications. We report here the synthesis and characterization of very stable novel warfarin-capped gold nanoparticles with an average diameter of 54 ± 10 nm which were prepared using sodium warfarin as a reducing agent. The nanoparticles were tested in terms of cytotoxicity and cellular internalization in vitro on two cell lines: normal lung fibroblast HFL-1 and human retinal pigment epithelial D407 cells. Our results showed that the normal lung fibroblast HFL-1 cells were more sensitive to the nanoparticle treatment compared to the human retinal pigment epithelial D407 cells. Moreover, any signs of potential cytotoxicity occurred during the first 24 h of treatment, the cellular viability remaining largely unchanged for longer exposure times. Transmission electron microscopy and dark field hyperspectral imaging revealed that the nanoparticles were effectively delivered and released to the HFL-1 and D407 cells' cytoplasm. Our results provide valuable information to further investigate sodium warfarin-capped gold nanoparticles for possible biological applications.

Valorification of crude glycerol for pure fractions of docosahexaenoic acid and ß-carotene production by using Schizochytrium limacinum and Blakeslea trispora.

The goal of this research is the investigation of a way to maximize the production of docosahexaenoic acid (DHA) and ß-carotene by optimizing the culture conditions of their sources, microalgae Schizochytrium limacinum and fungus Blakeslea trispora respectively, in a fermentation medium. The influencing factors in the fermentation process for producing DHA and ß-carotene have proven to be: the concentration of carbon source (different glycerol crude and pure concentrations) for both of them, and in particular temperature for DHA and pH for ß-carotene. Testing the effect of these parameters was determined: biomass, DHA and ß-carotene concentration. The highest production by S. limacinum was obtained at 25 °C, while using a quantity of 90 g/L of glycerol (crude or pure) as a carbon source. Temperature was the main factor that influenced the biosynthesis of DHA. The quantification of DHA was made by GC-MS chromatography, followed by a purification process, with the end result of DHA in pure phase. The maximum quantities for ß-carotene production were obtained with pH 7 and 60 g/L of crude glycerol. The results highlight the possibility of using crude glycerol as a low-cost substrates for growth of microalgae S. limacinum and of fungus B. trispora in order to obtain the crucial molecules: docosahexaenoic acid and ß-carotene.

Antiproliferative and Apoptotic Potential of Cyanidin-Based Anthocyanins on Melanoma Cells.

Elderberries are known for their high anthocyanins content, which have been shown to possess anti-proliferative and anti-cancer effects. Anthocyanins enriched extract (AEE) was obtained from elderberries and was characterized by LC/DAD/ESI-MS analysis. Five cyanidin-based anthocyanins were identified, among which Cy-3-O-samb was the major compound (51%). The total anthocyanins content of AEE was 495 mg Cy-3-O-samb/100 g FW. AEE inhibited proliferation of metastatic B16-F10 murine melanoma cells, in a concentration-dependent manner, with an IC50 of 264.3 µg/mL. LDH (lactate dehydrogenase), as a marker of membrane integrity, increased 74% in B16-F10 cells treated with 250 µg/mL AEE, compared to control. It was observed that apoptosis is the mechanism of melanoma cell death after AEE treatment, confirmed morphologically by acridine orange/ethidium bromide double staining and TUNEL analysis. These results indicate that elderberry-derived anthocyanins might be utilized in future applications as topical adjuvant in skin cancer therapy.

Melanoma Inhibition by Anthocyanins Is Associated with the Reduction of Oxidative Stress Biomarkers and Changes in Mitochondrial Membrane Potential.

Anthocyanins are water soluble pigments which have been proved to exhibit health benefits. Several studies have investigated their effects on several types of cancer, but little attention has been given to melanoma. The phytochemical content of nine different berry samples was assessed by liquid chromatography followed by electrospray ionization mass spectrometry (LC-ESI+-MS). Twenty-six anthocyanins were identified, after a previous C18 Sep-pak clean-up procedure. Chokeberry and red grape anthocyanins rich extracts (C-ARE and RG-ARE) were selected to be tested on normal and melanoma cell lines, due to their different chemical pattern. C-ARE composition consists of cyanidin aglycone glycosylated with different sugars; while RG-ARE contains glucosylated derivatives of five different aglycones. Both C-ARE and RG-ARE anthocyanins reduced proliferation, increased oxidative stress biomarkers and diminished mitochondrial membrane potential in melanoma cells, having no negative influence on normal cells. A synergistic response may be attributed to the five different aglycones present in RG-ARE, which proved to exert greater effects on melanoma cells than the mixture of cyanidin derivatives with different sugars (C-ARE). In conclusion, C-ARE and RG-ARE anthocyanins may inhibit melanoma cell proliferation and increase the level of oxidative stress, with opposite effect on normal cells. Therefore, anthocyanins might be recommended as active ingredients for cosmetic and nutraceutical industry. Graphical Abstract ᅟ.

Antiproliferative and antioxidant properties of anthocyanin rich extracts from blueberry and blackcurrant juice.

The present study was aimed at evaluating the antiproliferative potential of anthocyanin-rich fractions (ARFs) obtained from two commercially available juices (blueberry and blackcurrant juices) on three tumor cell lines; B16F10 (murine melanoma), A2780 (ovarian cancer) and HeLa (cervical cancer). Individual anthocyanin determination, identification and quantification were done using HPLC-MS. Antioxidant activity of the juices was determined through different mechanism methods such as DPPH and ORAC. For biological testing, the juices were purified through C18 cartridges in order to obtain fractions rich in anthocyanins. The major anthocyanins identified were glycosylated cyanidin derivatives. The antiproliferative activity of the fractions was tested using the MTT assay. The antiproliferative potential of ARF was found to be associated with those bioactive molecules, anthocyanins due to their antioxidant potential. The results obtained indicated that both blueberry and blackcurrants are rich sources of antioxidants including anthocyanins and therefore these fruits are highly recommended for daily consumption to prevent numerous degenerative diseases.

Effects of polystyrene nanoplastics exposure on in vitro-grown Stevia rebaudiana plants.

Nanoplastics (NPs) as environmental contaminants have received increased attention in recent years. Numerous studies have suggested possible negative effects of plants exposure to NPs, but more data are needed with various plants under different exposure conditions to clarify the underlying phytotoxicity mechanisms. In this study, we investigated the effect of polystyrene nanoplastics (PSNPs; 28.65 nm average diameter) exposure (10, 100 and 250 mg/L) on plant morphology and production of relevant metabolites (steviol glycosides, chlorophylls, carotenoids, and vitamins) of in vitro-grown Stevia rebaudiana plantlets. Additionally, we used dark field microscopy combined with fluorescence hyperspectral imaging for the visualization of internalized PSNPs inside plant tissues. At higher concentrations (>100 mg/L), PSNPs were shown to aggregate in roots and to be transported to leaves, having a significantly negative impact on plant growth (reduced size and biomass), while increasing the production of metabolites compared to controls, most probably because of response to stress. The production of steviol glycosides presented a biphasic dose-response suggestive of hormesis, with the highest values at 10 mg/L PSNPs (1.5-2.2-fold increase compared to controls), followed by a decline in production at higher concentrations (100 and 250 mg/L), but with values comparable to controls. These results are promising for future in vivo studies evaluating the effect of NP exposure on the production of steviol glycosides, the natural sweeteners from stevia.

Citrus essential oils - Based nano-emulsions: Functional properties and potential applications.

Citrus essential oils are natural products with various bioactive properties (e.g., antimicrobial, antioxidant, and antimutagenic activities), that are generally recognized as safe (GRAS) by Food and Drug Administration (FDA) to be used as flavorings and food additives. Nonetheless, due to their high volatility, low solubility in water, low thermal stability, susceptibility to oxidation, and strong flavor, their applications in the food industry are limited. Nanotechnology allows the incorporation of citrus essential oils into nano-emulsion systems, thus protecting them from the deterioration caused by external factors and maintaining or even improving their functional properties. This study aims to summarize the antioxidant, antimicrobial, and antimutagenic effects of the nano-emulsions based on essential oils from citrus peels with emphasis on their mechanisms of action and potential applications in, e.g., foods, pharmaceuticals, and cosmetics.

Nano-Encapsulation of Citrus Essential Oils: Methods and Applications of Interest for the Food Sector.

Citrus essential oils possess many health-promoting benefits and properties of high interest in the food and agri-food sector. However, their large-scale application is limited by their sensitivity to environmental factors. Nanostructures containing citrus essential oils have been developed to overcome the high volatility and instability of essential oils with respect to temperature, pH, UV light, etc. Nanostructures could provide protection for essential oils and enhancement of their bioavailability and biocompatibility, as well as their biological properties. Nano-encapsulation is a promising method. The present review is mainly focused on methods developed so far for the nano-encapsulation of citrus essential oils, with emphasis on lipid-based (including liposomes, solid lipid nanoparticles, nanostructured lipid particles, and nano- and micro-emulsions) and polymer-based nanostructures. The physico-chemical characteristics of the obtained structures, as well as promising properties reported, with relevance for the food sector are also discussed.

SERS liquid biopsy in breast cancer. What can we learn from SERS on serum and urine?

SERS analysis of biofluids, coupled with classification algorithms, has recently emerged as a candidate for point-of-care medical diagnosis. Nonetheless, despite the impressive results reported in the literature, there are still gaps in our knowledge of the biochemical information provided by the SERS analysis of biofluids. Therefore, by a critical assignment of the SERS bands, our work aims to provide a systematic analysis of the molecular information that can be achieved from the SERS analysis of serum and urine obtained from breast cancer patients and controls. Further, we compared the relative performance of five different machine learning algorithms for breast cancer and control samples classification based on the serum and urine SERS datasets, and found comparable classification accuracies in the range of 61-89%. This result is not surprising since both biofluids show striking similarities in their SERS spectra providing similar metabolic information, related to purine metabolites. Lastly, by carefully comparing the two datasets (i.e., serum and urine) we show that it is possible to link the misclassified samples to specific metabolic imbalances, such as carotenoid levels, or variations in the creatinine concentration.

The effect of 100-200 nm ZnO and TiO2 nanoparticles on the in vitro-grown soybean plants.

Engineered nanomaterials are increasingly used in everyday life applications and, in consequence, significant amounts are being released into the environment. From soil, water, and air they can reach the organelles of edible plants, potentially impacting the food chain and human health. The potential environmental and health impact of these nanoscale materials is of public concern. TiO2 and ZnO are among the most significant nanomaterials in terms of production amounts. Our study aimed at evaluating the effects of large-scale TiO2 (~100 nm) and ZnO (~200 nm) nanoparticles on soybean plants grown in vitro. The effect of different concentrations of nanoparticles (10, 100, 1000 mg/L) was evaluated regarding plant morphology and metabolic changes. ZnO nanoparticles showed higher toxicity compared to TiO2 in the experimental set-up. Overall, elevated levels of chlorophylls and proteins were observed, as well as increased concentrations of ascorbic and dehydroascorbic acids. Also, the decreasing stomatal conductance to water vapor and net CO2 assimilation rate show higher plant stress levels. In addition, ZnO nanoparticle treatments severely affected plant growth, while TEM analysis revealed ultrastructural changes in chloroplasts and rupture of leaf cell walls. By combining ICP-OES and TEM results, we were able to show that the nanoparticles were metabolized, and their internalization in the soybean plant tissues occurred in ionic forms. This behavior most likely is the main driving force of nanoparticle toxicity.

In situ laser-induced photochemical silver substrate synthesis and sequential SERS detection in a flow cell.

A new, simple, and effective approach for multianalyte sequential surface-enhanced Raman scattering (SERS) detection in a flow cell is reported. The silver substrate was prepared in situ by laser-induced photochemical synthesis. By focusing the laser on the 320 µm inner diameter glass capillary at 0.5 ml/min continuous flow of 1 mM silver nitrate and 10 mM sodium citrate mixture, a SERS active silver spot on the inner wall of the glass capillary was prepared in a few seconds. The test analytes, dacarbazine, 4-(2-pyridylazo)resorcinol (PAR) complex with Cu(II), and amoxicillin, were sequentially injected into the flow cell. Each analyte was adsorbed to the silver surface, enabling the recording of high intensity SERS spectra even at 2 s integration times, followed by desorption from the silver surface and being washed away from the capillary. Before and after each analyte passed the detection window, citrate background spectra were recorded, and thus, no "memory effects" perturbed the SERS detection. A good reproducibility of the SERS spectra obtained under flow conditions was observed. The laser-induced photochemically synthesized silver substrate enables high Raman enhancement, is characterized by fast preparation with a high success rate, and represents a valuable alternative for silver colloids as SERS substrate in flow approaches.

SERS liquid biopsy: An emerging tool for medical diagnosis.

Surface-enhanced Raman scattering (SERS) is emerging as a novel strategy for biofluid analysis. In this review, we delineate four experimental SERS protocols that are frequently used for the profiling of biofluids: 1) liquid SERS for the detection of purine metabolites; 2) iodide-modified liquid SERS for the detection of proteins; 3) dried SERS for the detection of both purine metabolites and proteins; 4) resonant Raman for the detection of carotenoids. To explain the selectivity of each experimental SERS protocol, we introduce a heuristic model for the chemisorption of analytes mediated by adsorbed ions (adions) onto the SERS substrate. Next, we show that the promising results of SERS liquid biopsy stem from the fact that the concentration levels of purine metabolites, proteins and carotenoids are informative of the cellular turnover rate, inflammation, and oxidative stress, respectively. These processes are perturbed in virtually every disease, from cancer to autoimmune maladies. Finally, we review recent SERS liquid biopsy studies and discuss future steps that are required for translating SERS in the clinical setting.

Separation and Purification of Biogenic 1,3-Propanediol from Fermented Glycerol through Flocculation and Strong Acidic Ion-Exchange Resin.

In the present work, was investigated the separation and purification procedure of the biogenic 1,3-propanediol (1,3-PD), which is a well-known valuable compound in terms of bio-based plastic materials development. The biogenic 1,3-PD was obtained as a major metabolite through the glycerol fermentation by Klebsiella pneumoniae DSMZ 2026 and was subjected to separation and purification processes. A strong acidic ion exchange resin in H+ form was used for 1,3-PD purification from the aqueous solution previously obtained by broth flocculation. The eluent volume was investigated considering the removal of the secondary metabolites such as organic acids (acetic, citric, lactic, and succinic acids) and 2,3-butanediol (2,3-BD), and unconsumed glycerol. It was observed that a volume of 84 mL of ethanol 75% loaded with a flow rate of 7 mL/min completely remove the secondary metabolites from 10 mL of concentrated fermented broth, and pure biogenic 1,3-PD was recovered in 128 mL of the eluent.

Soybean Interaction with Engineered Nanomaterials: A Literature Review of Recent Data.

With a continuous increase in the production and use in everyday life applications of engineered nanomaterials, concerns have appeared in the past decades related to their possible environmental toxicity and impact on edible plants (and therefore, upon human health). Soybean is one of the most commercially-important crop plants, and a perfect model for nanomaterials accumulation studies, due to its high biomass production and ease of cultivation. In this review, we aim to summarize the most recent research data concerning the impact of engineered nanomaterials on the soya bean, covering both inorganic (metal and metal-oxide nanoparticles) and organic (carbon-based) nanomaterials. The interactions between soybean plants and engineered nanomaterials are discussed in terms of positive and negative impacts on growth and production, metabolism and influences on the root-associated microbiota. Current data clearly suggests that under specific conditions, nanomaterials can negatively influence the development and metabolism of soybean plants. Moreover, in some cases, a possible risk of trophic transfer and transgenerational impact of engineered nanomaterials are suggested. Therefore, comprehensive risk-assessment studies should be carried out prior to any mass productions of potentially hazardous materials.

The role of Ag+, Ca2+, Pb2+ and Al3+ adions in the SERS turn-on effect of anionic analytes.

In our recent studies we highlighted the role of adsorbed ions (adions) in turning on the surface-enhanced Raman scattering (SERS) effect in a specific mode for anionic and cationic analytes. In this work, we emphasize the role of Ag+, Ca2+, Pb2+ and Al3+ adions in the specific adsorption of anionic analytes such as the citrate capping agent and three organic acids. Our results suggest an adion-specific adsorption mechanism: the adsorption of anionic analytes is facilitated by positively charged adions such as Ag+, Ca2+, Pb2+ or Al3+, which provide adsorption sites specific for the anionic analytes. The turn-on of the SERS effect is explained in the context of the chemical mechanism of SERS. The adions form SERS-active sites on the silver surface enabling a charge transfer between the adsorbate and the silver surface. High-intensity SERS spectra of uric acid, salicylic acid and fumaric acid could be recorded at a concentration of 50 µM only after activation of the colloidal silver nanoparticles by Ca2+, Pb2+ or Al3+ (50 µM). The chemisorption of the three anionic species to the silver surface occurs competitively and is enhanced with the anions of higher affinities to the silver surface as indicated by the SERS spectra of corresponding mixed solutions.