Search Results (495)

Since the introduction of the DNA origami technology by Seeman and Rothemund, the integration of functional entities (nanoparticles, quantum dots, antibodies, etc.) has been of huge interest to broaden the area of applications for this technology. The possibility of precise functionalization of the DNA origami technology gives opportunity to build up complex novel structures, opening up endless opportunities in medicine, nanotechnology, photonics and many more. The main advantage of the DNA origami technology, namely the self-assembly mechanism, can represent a challenge in the construction of complex mixed-material structures. Commonly, DNA origami structures are purified post-assembly by filtration (either spin columns or membranes) to wash away excess staple strands. However, this purification step can be critical since these functionalized DNA origami structures tend to agglomerate during purification. Therefore, custom production and purification procedures need to be applied to produce purified functionalized DNA origami structures. In this paper, we present a workflow to produce functionalized DNA origami structures, as well as a method to qualify the successful hybridization of a quantum dot to a square frame DNA origami structure. Through the utilization of a FRET fluorophore–quencher pair as well as a subsequent assembly, successful hybridization can be performed and confirmed using photoluminescence measurements. Full article

A method combining magnetic solid-phase extraction (MSPE) with ultra-high performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) was developed for the determination of diazepam residues in aquatic products. A novel magnetic nanoparticle material, Fe₃O₄@SiO₂@DVB-NVP, was synthesized and applied as an adsorbent for sample cleanup. The sample preparation procedure involved extraction with 1% ammonia–acetonitrile, followed by purification using the MSPE technique to efficiently remove matrix interferents. Chromatographic separation was achieved on an ACQUITY UPLC BEH C₁₈ column with a gradient elution program using a mobile phase composed of 0.1% formic acid–2 mM ammonium acetate solution and methanol. Detection was performed under multiple-reaction monitoring (MRM) mode with positive electrospray ionization (ESI⁺). Quantification was carried out using the external standard method. The synthesized magnetic material was characterized using SEM, TEM, FTIR, XRD, BET, and VSM, confirming its mesoporous structure, strong adsorption capacity, and excellent magnetic responsiveness. The method demonstrated good linearity over the concentration range of 0.25–50 μg/L (r² = 0.997). The limits of detection and quantification were 0.20 μg/kg and 0.50 μg/kg, respectively. Average recoveries from spiked blank matrices at three levels (0.5, 2.5, and 5.0 μg/kg) ranged from 89.3% to 119.7%, with relative standard deviations (RSDs) between 0.8% and 10.2%. The proposed method is highly selective, exhibits minimal matrix interference, and provides reliable quantitative performance, making it suitable for the qualitative and quantitative analysis of diazepam residues in aquatic products. Full article

The saliva of the medicinal leech Hirudo medicinalis contains a wide range of biologically active compounds, including multiple anticoagulants. Previously, we identified a novel cysteine-rich anticoagulant protein (CRA) from leech saliva and produced it recombinantly in Escherichia coli, demonstrating its potential as a basis for new anticoagulant drugs. In this study, we developed an optimized procedure for scalable production and purification of recombinant CRA. The purified protein was analyzed for common contaminants originating from E. coli, such as endotoxins, bacterial proteins, and DNA, and its anticoagulant properties were evaluated using standard clotting assays. Across three independent experiments, the yield of purified CRA ranged from 3.7 to 5.5 mg per liter of bacterial culture, with impurity levels per milligram of protein ranging from 7.1–31.2 ng of bacterial proteins, 1.2–15.1 ng of DNA, and 60–1445 EU of endotoxins. The purified CRA displayed electrophoretic and chromatographic homogeneity and retained strong anticoagulant activity. Additionally, a truncated form of CRA lacking the C-terminal region was produced and characterized. This variant lost membrane affinity and showed altered activity profiles, with higher thrombin time activity but reduced prothrombin time and activated partial thromboplastin time activities compared with the full-length protein. Full article

Introduction. Over the past two decades, the αGal (Galα1–3Galβ1–4GlcNAc–R) epitope, a carbohydrate found in many non-primate mammals, has gained significant relevance in medicine due to its association with an increasing number of allergic reactions to animal-derived foods, drugs, and medical devices. Due to a mutated gene coding for α1,3-galactosyltransferase (α1–3GT), humans lack αGal and, therefore, naturally produce anti-α-Gal antibodies (IgM, IgA, and IgG), especially in the context of a xenotransplantation, which can lead to extreme immunological reactivity, including hyperacute rejection of the transplant. Recently, these uncontrollable immune reactions have driven demand for more accurate procedures to better detect αGal in animal-derived foods or bioprosthetics. The currently most widely used α-Gal-specific monoclonal antibody is an IgM antibody (clone M86), developed in Ggta1 KO mice and isolated from hybridoma tissue culture. As the IgM isotype has limited purification properties, specificity, and sensitivity, we aimed to produce a novel IgG antibody with high affinity and extensive applicability. Methods. An experimental murine IgG1 anti-αGal antibody (IgG-αGalomab) was developed by immunization of Ggta1 knockout (KO) mice, and its affinity was evaluated using ELISA, Western blot, flow cytometry, and immunohistochemistry/immunofluorescence. Results. Compared to IgM-M86, IgG-αGalomab demonstrated ~1200-fold higher binding potency and lower cross-reactivity with competitive molecules, i.e., bovine serum albumin, galactobiose, and lactose. Unlike IgM-M86, IgG-αGalomab showed an increasing affinity over time in the binding tests performed on xenogeneic tissues. Notably, high-affinity for αGal was detected by Western blot at high dilution [1:200,000] of IgG-αGalomab compared to IgM-M86 [1:1000]. By flow cytometry, specificity and dose-dependent response were confirmed using in vitro cultures of porcine and human fibroblasts. Finally, in immunofluorescence and immunohistochemistry analysis, αGal was demonstrated to be detectable by IgG-αGalomab at a dilution of [1:1000], while IgM-M86 was demonstrated to be detectable at [1:100]. Conclusions. Altogether, our newly developed antibody showed high sensitivity and specificity for α-Gal in various applications. Based on its potential binding capacity, IgG-αGalomab could have important applications in precision medicine for predicting, treating, and preventing immune-mediated phenomena of patients in different medical areas. Full article

Two HPLC-based analytical methods, one with DAD and the other with an FLD detector, were developed and validated for the simultaneous analysis of B₁, B₂, and B₆ vitamins, both in pharmaceutical gummies and in gastric and intestinal fluids (with water or milk or orange juice). For the detection of B₁ by fluorometry, a pre-column oxidation/derivatization process was accomplished in the presence of B₂ and B₆ vitamins. The methods were performed on an Aqua column (250 mm × 4.6 mm, 5 mm) at 40 °C, with isocratic elution (70% NaH₂PO₄ buffer pH 4.95 and 30% methanol) and a flow rate of 0.9 mL/min. Both were validated according to ICH specifications in terms of linearity (R² > 0.999), accuracy (% Mean Recovery 100 ± 3%) and precision (%RSD < 3.23). For the analysis of the samples, a stability study (in diluents, pH and fluids) was conducted, while for their purification two different extraction procedures, a liquid/solid for the gummies (%Recovery > 99.8%) and a Solid Phase Extraction (SPE) for the Gastrointestinal (G.I.) fluids, (%Recovery 100 ± 5%) were developed. Finally, to investigate whether the co-administration of B-complex with water, orange juice or milk plays a significant role in their release from gummies, a three-phase in vitro digestion protocol was applied. The results did not show significant differences with a slight superiority in the release of B₂ and B₆ with water, while B₁ with orange juice. Full article

This innovative study introduces an eco-conscious and cost-effective approach to synthesizing gelatin-based carbon dots (CDs) via two distinctive methods: hydrothermal processing in a muffle furnace (CDs-MF) and domestic microwave (CDs-MW). Both strategies harness natural, low-cost materials and prioritize simplicity, sustainability, and environmental friendliness, culminating in effective fluorescent sensing of the pesticide thiamethoxam (TMX). For the hydrothermal route, the investigation explores two purification approaches—ultracentrifugation (CDs-MF-C) and 0.22 µm syringe filtration (CDs-MF-F)—while the microwave-derived CDs (CDs-MW) undergo dialysis alone. This study aims to investigate how synthesis and purification impact the CDs structural, morphological, and photophysical characteristics. The difference in size was obtained from transmission electron microscopy (TEM): 30–40 nm for CDs-MF-C, 12–15 nm for CDs-MF-F, and 3–6 nm for CDs-MW. Fluorescence emission performance reveals that CDs-MF-F performs a fluorescence quantum yield of 27%, CDs-MF-C at 23%, and CDs-MW at a modest 3%. All variants exhibit TMX detection via fluorescence quenching through the inner filter effect (IFE). Analytically, CDs-MF-C stands out with the lowest detection limit (LOD = 0.396 ppm) and quantification limit (LOQ = 1.317 ppm), followed by CDs-MF-F (LOD = 0.475 ppm; LOQ = 1.585 ppm) and CDs-MW (LOD = 0.549 ppm; LOQ = 1.831 ppm). These findings emphasize the unique interplay between the synthesis pathway, purification strategy, and functional performance, demonstrating the critical importance of tuning structural properties for optimizing carbon-dot sensors. Full article

Sweet potato stems and leaves (SPSL) are rich in bioactive polyphenols, yet their utilization remains underexplored. This study established an efficient method for SPSL polyphenol enrichment using macroporous resins, with UHPLC-QE-MS/MS characterization of the purified polyphenols (PP) and subsequent evaluation of anti-inflammatory activity. The results showed that NKA-II resin demonstrated the best purification effect on SPSL polyphenols among the six tested resins. The optimal enrichment procedure of NKA-II resin was as follows: loading sample pH 3.0, 4.48 mg CAE/mL concentration, and 80% ethanol (v/v) eluent. A total of 19 major compounds were characterized in PP, including 12 phenolic acids and seven flavonoids, with a polyphenol purity of 75.70%. PP pretreatment (100 and 500 μg/mL) significantly inhibited LPS-induced release of NO (by 40.62% and 68.61%), IL-1β (by 40.07% and 68.34%), IL-6 (by 40.63% and 52.41%), and TNF-α (by 52.29% and 73.76%) compared to the LPS group (p < 0.05), demonstrating potent anti-inflammatory effects. Western blot analysis revealed that PP exerted anti-inflammatory effects by inhibiting the NF-κB (via suppression of IκBα phosphorylation/degradation and blockade of p65 nuclear translocation) and MAPK (via inhibition of p38, ERK, and JNK phosphorylation) signaling pathways. These findings support the utilization of this agricultural by-product in functional food development, particularly as a source of natural anti-inflammatory compounds for dietary supplements or fortified beverages. Full article

Nanofiltration (NF) is considered a competitive purification method for acidic stream treatments. However, conventional thin-film composite NF membranes degrade under acid exposures, limiting their applications in industrial acid treatment. For example, wet-process phosphoric acid contains impurities of multivalent metal ions, but NF membrane technologies for impurity removal under harsh conditions are still immature. In this work, we develop a novel strategy of acid-resistant nanofiltration membranes based on interfacial polymerization (IP) of polyethyleneimine (PEI) and cyanuric chloride (CC) with the s-triazine ring. The IP process was optimized by orthogonal experiments to obtain positively charged PEI-CC membranes with a molecular weight cut-off (MWCO) of 337 Da. We further applied it to the approximate industrial phosphoric acid purification condition. In the tests using a mixed solution containing 20 wt% P₂O₅, 2 g/L Fe³⁺, 2 g/L Al³⁺, and 2 g/L Mg²⁺ at 0.7 MPa and 25 °C, the NF membrane achieved 56% rejection of Fe, Al, and Mg and over 97% permeation of phosphorus. In addition, the PEI-CC membrane exhibited excellent acid resistance in the 48 h dynamic acid permeation experiment. The simple fabrication procedure of PEI-CC membrane has excellent acid resistance and great potential for industrial applications. Full article

Peptides play a crucial role in the development of pharmaceuticals and functional foods. Multiple studies have shown that natural bioactive peptides possess antioxidant, antihypertensive, anti-tumor, and anti-inflammatory activities. Marine bioactive peptides, especially those sourced from fish, constitute a substantial reservoir of these molecules. Although considerable research has been undertaken on fish-derived peptides, studies specifically concerning those from tuna are limited. Tuna, a marine fish of high nutritional value, generates substantial by-product waste during fishing and processing. Therefore, it is essential to conduct an evaluation of the advancements in study on tuna-derived active peptides and to offer a perspective on the direction of future investigations. This review integrates prospective bioactive peptides derived from tuna and reports contemporary strategies for their investigation, including extraction, purification, screening, identification, and activity evaluation procedures, including Yeast Surface Display (YSD) and molecular docking. This review seeks to promote the continued investigation and application of bioactive peptides derived from tuna. Full article

One of the key elements in the analysis of gene expression and its post-translational regulation is miRNAs. Degradome-seq analyses are performed to analyze the cleavage of target RNAs in the transcriptome. This work presents the first degradome-seq library preparation protocol that enables successful construction of libraries, even from highly degraded RNA samples with RIN below 3, thus significantly expanding the possibilities for research when working with low-quality material. The developed protocol improves the efficiency of library preparation in degradome-seq analysis used to identify miRNA targets, reduces library preparation time, and lowers the cost of purchasing reagents by using reagents from the RNA-seq library preparation kit and proprietary-designed primers. A crucial feature of this new protocol is optimizing the purification step for short library fragments, which increases the yield of correctly sized fragments compared to previously used methods. This is achieved by implementing an original method involving tube-spin purification with gauze and precipitation using sodium acetate with glycogen, greatly enhancing recovery efficiency—a factor especially critical when working with degraded RNA. Cloning to a plasmid and sequencing of the inserted fragment verified the correctness of the library preparation using the developed protocol. This protocol represents a groundbreaking tool for degradome research, enabling the construction and sequencing of degradome libraries, even from degraded samples previously considered unsuitable for such analyses. This is due to the use of residues from the sRNA-seq library kit. It noticeably reduces the cost of library construction. The precision of the excised fragment after electrophoresis was performed during the procedure to isolate fragments of the correct length, which was improved using additional size markers. Compared to previously used methods, optimizing the purification method of degradome-seq libraries allowed an increase in the yield of fragments obtained. Full article

Nanomaterials possess unique physicochemical properties that position them as promising candidates for environmental remediation, particularly in the removal of persistent organic pollutants (POPs) from aqueous systems. Their high surface area, tunable functionality, and strong adsorption capabilities have attracted significant attention. In this context, this paper reviews the mechanisms of nanomaterial-based POP decontamination, also providing a critical overview of the limitations and challenges in applying these methods. Specifically, issues of stability, reusability, and aggregation are discussed, which can lead to performance decay during repeated use. In addition, the practical application requires nanocomposites to enable efficient separation and mitigate agglomeration. Environmental concerns also arise from nanomaterials’ fate, transport, and potential toxicity, which may impact aquatic ecosystems and non-target organisms. When checking for large-scale application feasibility, impurities typically add to production costs, recovery problems, and general infrastructure limitations. In addition to these points, there are no standard guidelines or clear risk assessment procedures for registering a product. Unprecedented cross-disciplinary research between natural, human, and technological studies and outreach programs is needed to facilitate the development and diffusion of the results. The barriers will eventually be breached to move from laboratory success in developing the desperately needed new water purification technologies to field-ready water treatment solutions that can address the global POP contamination problem. Full article

Encouraging discoveries and excellent advances in the fight against cancer have led to innovative therapies such as photothermal therapy (PTT), photodynamic therapy (PDT), drug targeting (DT), gene therapy (GT), immunotherapy (IT), and therapies that combine these treatments with conventional chemotherapy (CT). Furthermore, 2,041,910 new cancer cases and 618,120 cancer deaths have been estimated in the United States for the year 2025. The low survival rate (<50%) and poor prognosis of several cancers, despite aggressive treatments, are due to therapy-induced secondary tumorigenesis and the emergence of drug resistance. Moreover, serious adverse effects and/or great pain usually arise during treatments and/or in survivors, thus lowering the overall effectiveness of these cures. Although prevention is of paramount importance, novel anticancer approaches are urgently needed to address these issues. In the field of anticancer nanomedicine, carbon nanotubes (CNTs) could be of exceptional help due to their intrinsic, unprecedented features, easy functionalization, and large surface area, allowing excellent drug loading. CNTs can serve as drug carriers and as ingredients to engineer multifunctional platforms associated with diverse treatments for both anticancer therapy and diagnosis. The present review debates the most relevant advancements about the adjuvant role that CNTs could have in cancer diagnosis and therapy if associated with PTT, PDT, DT, GT, CT, and IT. Numerous sensing strategies utilising various CNT-based sensors for cancer diagnosis have been discussed in detail, never forgetting the still not fully clarified toxicological aspects that may derive from their extensive use. The unsolved challenges that still hamper the possible translation of CNT-based material in clinics, including regulatory hurdles, have been discussed to push scientists to focus on the development of advanced synthetic and purification work-up procedures, thus achieving more perfect CNTs for their safer real-life clinical use. Full article

Selenium sulfide, the active ingredient of traditional antidandruff shampoos, is industrially produced from selenium dioxide (SeO₂) and hydrogen sulfide (H₂S) under acidic conditions. This reaction can also be carried out with natural H₂S and H₂S generated by sulfate-reducing bacteria (SRB). These bacteria are robust and, by relying on their conventional growth medium, also thrive in “waste” materials, such as a mixture of cabbage juice and compost on the one side, and a mixture of spoiled milk and mineral water on the other. In these mixtures, SRB are able to utilize the DL-lactate and sulfate (SO₄²⁻) present naturally and produce up to 4.1 mM concentrations of H₂S in the gas phase above a standard culture medium. This gas subsequently escapes the fermentation vessel and can be collected and reacted with SeO₂ in a separate compartment, where it yields, for instance, pure selenium sulfide, therefore avoiding the need for any cumbersome workup or purification procedures. Thus “harvesting” H₂S and similar (bio-)gases produced by the fermentation of organic waste materials by suitable microorganisms provides an elegant avenue to turn dirty waste into valuable clean chemical products of considerable industrial and pharmaceutical interest. Full article

Microbial lipids are substances of high added value produced by single-cell organisms grown on simple substrates. These lipids, depending on the producing organism, may contain rare fatty acids, whose isolation and purification from non-microbial sources usually is an inefficient and costly procedure. Such fatty acids mostly include members of the omega-3 and omega-6 families of polyunsaturated fatty acids, which are credited with potential anticancer, anti-inflammatory, cardioprotective, and neuroprotective actions. However, their poor solubility in aqueous solutions often restricts their potential applications, as routes other than dietary consumption are unavailable. A promising approach for administering them is their conversion into alkali salts, mostly with lithium or potassium, which are water-soluble and bio-assimilable. In this article, all studies investigating the potential anticancer effects of alkali salts of fatty acids isolated from microorganisms were reviewed in an attempt to sum up existing knowledge and encourage further research. Full article

Bioprotection in winemaking refers to the use of naturally occurring microorganisms—mainly non-Saccharomyces yeasts—to inhibit the growth of spoilage microbes and reduce the need for chemical preservatives like sulfur dioxide (SO₂). Numerous studies have demonstrated the benefits of non-Saccharomyces as bioprotectants. However, the use of Saccharomyces cerevisiae as a bioprotectant has been studied very little. Furthermore, it can offer many advantages for the production of sulfite-free wines. To test if S. cerevisiae could be used in bioprotection, we compared the ability of different strains to inhibit the growth of Brettanomyces bruxellensis and Hanseniaspora uvarum. Among the strains tested, the S. cerevisiae Sc54 strain isolated from the vineyard of the Bekaa plain was selected. To investigate its mechanisms of action, we analyzed its metabolite production, including acetic acid and ethanol. Taking into account the low levels of these metabolites and the lack of similar inhibition patterns in media supplemented with acetic acid and ethanol, it appears that other factors contribute to its antagonistic properties. Nutrient competition was ruled out as a factor, as the growth inhibition of B. bruxellensis and H. uvarum occurred rapidly within the first 24 h of co-culture. In this study, we explored the role of the S. cerevisiae killer toxin (Sc54Kt) as a bioprotective agent against H. uvarum and B. bruxellensis spoilage yeasts. Purification procedures with ethanol allowed the extraction of Sc54Kt, yielding two concentrations (0.185 and 0.5 mg/mL). Remarkably, semi-purified Sc54Kt exhibited inhibitory effects at both concentrations under winemaking conditions, effectively controlling the growth and metabolic activity of the target spoilage yeasts. Overall, these findings demonstrate that S. cerevisiae Sc54 not only exerts a strong bioprotective effect but also contributes to improving the quality of wine. The results suggest that S. cerevisiae Sc54 is a promising bioprotective agent for mitigating spoilage yeasts in winemaking, offering a natural and effective alternative to conventional antimicrobial strategies. Full article