Polydopamine functionalized dendritic fibrous silica nanoparticles as a generic platform for nucleic acid-based biosensing.

Accurate and rapid detection of nucleic acid sequences is of utmost importance in various fields, including disease monitoring, clinical treatment, gene analysis and drug discovery. In this study, we developed a "turn-on" fluorescence biosensor that enables simple and highly efficient detection of nucleic acid biomarkers. Our approach involves the utilization of 6-carboxyfluorescein modified single-stranded DNA (FAM-ssDNA) as molecular recognition element, along with polydopamine-functionalized dendritic fibrous nanosilica (DFNS). FAM-ssDNA serves as both specific molecular recognition element for the target analyte and reporter capable of transducing a detectable signal through Watson-Crick base pairing. The polydopamine-functionalized DFNS (DFNS@DA) exhibits strong binding to FAM-ssDNA via polyvalent metal mediated coordination leading to effective quenching by fluorescence resonance energy transfer. In the presence of a complementary target sequence, FAM-ssDNA forms hybridized structure and detaches from DFNS@DA, which causes an increased fluorescence emission. The analytical system based on FAM-ssDNA and DFNS@DA demonstrates exceptional sensitivity, selectivity, and rapid response for the detection of nucleic acid sequences, leveraging the high adsorption and quenching properties of DFNS@DA. For the first proof of concept, we demonstrated the successful detection of microRNA (miR-21) in cancer cells using the FAM-ssDNA/DFNS@DA system. Our results highlight the promising capabilities of DFNS@DA and nucleic acid-based biosensors, offering a generic and cost-effective solution for the detection of nucleic acid-related biomarkers.

Dual-amplification system based on CRISPR-Cas12a and horseradish peroxidase-tethered magnetic microspheres for colorimetric detection of microcystin-LR.

A novel dual-amplification system based on CRISPR-Cas12a and horseradish peroxidase (HRP) was developed for colorimetric determination of MC-LR. This dual-amplification was accomplished by combining the nuclease activity of CRISPR-Cas12a with the redox activity of HRP. HRP linked to magnetic beads through an ssDNA (MB-ssDNA-HRP) was used to induce a color change of the 3,3',5,5'-tetramethylbenzidine (TMB)-H2O2 chromogenic substrate solution. Specific binding of MC-LR with its aptamer initiated the release of a complementary DNA (cDNA), which was designed to activate the trans-cleavage activity of CRISPR-Cas12a. Upon activation, Cas12a cut the ssDNA linker in MB-ssDNA-HRP, causing a reduction of HRP on the magnetic beads. Consequently, the UV-Vis absorbance of the HRP-catalyzed reaction was decreased. The dual-signal amplification facilitated by CRISPR-Cas12a and HRP enabled the colorimetric detection of MC-LR in the range 0.01 to 50 ng·mL-1 with a limit of detection (LOD) of 4.53 pg·mL-1. The practicability of the developed colorimetric method was demonstrated by detecting different levels of MC-LR in spiked real water samples. The recoveries ranged from 86.2 to 118.5% and the relative standard deviation (RSD) was 8.4 to 17.6%. This work provides new inspiration for the construction of effective signal amplification platforms and demonstrates a simple and user-friendly colorimetric method for determination of trace MC-LR.

Recyclable nanoparticles based on a boronic acid-diol complex for the real-time monitoring of imprinting, molecular recognition and copper ion detection.

Molecularly imprinted polymers (MIPs) have now become one of the most remarkable materials in the field of molecular recognition. Although many efforts have been made to study the process and mechanism of molecular imprinting, it has not been possible to monitor the interactions between the template and the growing polymer chains under real-time experimental conditions. The behavior of the template-monomer complex during the whole polymerization process has remained largely unknown. In this work, we introduce a fluorescence technique that allows monitoring of the template-functional monomer complex during an actual imprinting process, as well as the real-time signaling of template binding and dissociation from the imprinted polymer. For the first proof-of-principle, we select Alizarin Red S (ARS) and 4-vinylphenylboronic acid as the template and functional monomer, respectively, to synthesize MIP particles via precipitation polymerization. As the formation of the template-functional monomer complex leads to strong fluorescence emission, it allows the status of the template binding to be monitored throughout the whole reaction process in real time. Using the same fluorescence technique, the kinetics of template binding and dissociation can be studied directly without particle separation. The hydrophilic MIP particles can be used as a scavenger to remove ARS from water. In addition, the MIP particles can be used as a recyclable sensor to detect Cu ions. As the Cu ion forms a stable complex with ARS, it causes ARS to dissociate from the MIP nanoparticles, leading to effective fluorescence quenching. The non-separation analytical method based on fluorescence measurement provides a convenient means to study molecular imprinting reactions and the kinetics of molecular recognition using imprinted polymers. The recyclable nanoparticle sensor allows toxic Cu ions to be detected directly in water in the range of 0.1-100 µM with a recovery of 84-95%.

Boronic Acid Functionalized Nanosilica for Binding Guest Molecules.

Dendritic fibrous nanosilica (DFNS) has very high surface area and well-defined nanochannels; therefore, it is very useful as supporting material for numerous applications including catalysis, sensing, and bioseparation. Due to the highly restricted space, addition of molecular ligands to DFNS is very challenging. This work studies how ligand conjugation in nanoscale pores in DFNS can be achieved through copper-catalyzed click reaction, using an optional, in situ synthesized, temperature-responsive polymer intermediate. A clickable boronic acid is used as a model to investigate the ligand immobilization and the molecular binding characteristics of the functionalized DFNS. The morphology, composition, nanoscale pores, and specific surface area of the boronic acid functionalized nanosilica were characterized by electron microscopy, thermogravimetric and elemental analysis, Fourier transform infrared spectroscopy, and nitrogen adsorption-desorption measurements. The numbers of boronic acid molecules on the modified DFNS with and without the polymer were determined to be 0.08 and 0.68 mmol of ligand/g of DFNS, respectively. We also studied the binding of small cis-diol molecules in the nanoscale pores of DFNS. The boronic acid modified DFNS with the polymer intermediate exhibits higher binding capacity for Alizarin Red S and nicotinamide adenine dinucleotide than the polymer-free DFNS. The two types of boronic acid modified DFNS can bind small cis-diol molecules in the presence of large glycoproteins, due in large part to the effect of size exclusion provided by the nanochannels in the DFNS.

Hypolipidemic, anti-inflammatory, and anti-atherosclerotic effects of tea before and after microbial fermentation.

BACKGROUND: Microbial fermentation significantly affects the flavor and efficacy of tea. It is generally believed that fermented tea is more effective in lowering lipids, while unfermented tea can more effectively inhibit inflammation. However, there is not sufficient evidence to support this claim. To systematically compare the hypolipidemic, anti-inflammatory, and anti-atherosclerotic effects of tea before and after microbial fermentation, hyperlipidemic rats and inflammatory injury cells were treated with Monascus purpureus-fermented pu-erh tea water extract (MPT) and sun-dried green tea water extract (SGT), respectively. RESULTS: MPT, with higher levels of theabrownins, flavonoids, gallic acid (GA), and lovastatin, was more effective in reducing serum triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), and inflammatory cytokines (TNF-α, IL-1ß, and IL-6), while SGT, with higher levels of tea polyphenols, amino acids, (-)-epigallocatechin gallate (EGCG), and theaflavins, was more effective in increasing serum high-density lipoprotein cholesterol (HDL-C) in hyperlipidemic rats. The foam cells on the arterial wall of the rats in the MPT group were visibly less, and the thrombosis time was longer than that in the SGT group. Cell experiments showed that MPT was more effective in protecting endothelial cells from damage than SGT. CONCLUSION: Surprisingly, Monascus purpureus-fermented pu-erh tea not only had better hypolipidemic and anti-atherosclerotic effects than its raw material (sun-dried green tea), but also was superior in anti-inflammatory effects to the latter, which was possibly attributable to the great changes in functional ingredients during microbial fermentation.

A facile and general approach for the preparation of boronic acid-functionalized magnetic nanoparticles for the selective enrichment of glycoproteins.

Biomedical applications and biomarkers for early clinical diagnostics and the treatment of diseases demand efficient and selective enrichment platforms for glycoproteins. Thus, we herein report a facile and general approach for the preparation of boronic acid-functionalized magnetic nanoparticles for the selective enrichment of glycoproteins. More specifically, Fe3O4 magnetic nanoparticles were initially prepared via a solvothermal reaction, and core-shell-structured Fe3O4@SiO2 nanoparticles were obtained according to a sol-gel process. Subsequently, the Fe3O4@SiO2 surfaces were modified using 4-formylphenylboronic acid to allow the formation of strong yet reversible covalent bonds between boronic acid (BA) and the cis-1,2-diol groups of glycoproteins. The morphology and structure of the Fe3O4, Fe3O4@SiO2, and Fe3O4@SiO2-BA nanoparticles were characterized by scanning electron microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and vibrating sample magnetometry, thereby confirming their successful preparation. The obtained BA-modified Fe3O4@SiO2 magnetic nanoparticles were applied in the attempted enrichment of two glycoproteins (ovalbumin (OVA) and transferrin (TRF)) and two non-glycoproteins (bovine serum albumin (BSA) and cytochrome c (Cyt C)). The results confirmed a significant difference in affinity between glycoproteins and non-glycoproteins. In addition, the recognition capability of the Fe3O4@SiO2-BA nanoparticles was demonstrated by the selective enrichment of glycoproteins from a complex system containing both glycoproteins (i.e., TRF) and non-glycoproteins (i.e., Cyt C).

Molecularly imprinted electrospun nanofibers for adsorption of 2,4-dinitrotoluene in water.

The detection and adsorption of nitroaromatic compounds such as 2,4-dinitrotoluene (DNT) is of great importance, but the selective detection of trace DNT in water remains a challenge. Here, we report molecularly imprinted polymer (MIP) nanofibers fabricated by electrospinning using DNT as a template molecule and poly(allylamine) as a functional macromer. The physical and chemical properties of the nanofibers were characterized by Fourier transform infrared (FTIR) spectroscopy, X-ray photoelectron spectroscopy (XPS), and scanning electron microscopy (SEM), and the DNT adsorption and selectivity properties were also studied by high performance liquid chromatography (HPLC). The results show that the nanofibers prepared using 20 wt% poly(ethylene terephthalate) (PET) in solvent had a good morphology and an optimal ratio of poly(allylamine) and DNT, and the imprinted nanofibers showed specific adsorption of DNT, in accordance with the Freundlich isotherm model and Scatchard analysis. Furthermore, the imprinted nanofibers showed remarkable stability and reusability, losing only 3% of their performance after eight cycles. Thus, the imprinted nanofibers fabricated by this strategy could be used for trace DNT extraction, separation, and further construction of sensors in the near future.

Determination and comparison of γ-aminobutyric acid (GABA) content in pu-erh and other types of Chinese tea.

Two previous studies have reported that pu-erh tea contains a high level of γ-aminobutyric acid (GABA), which is the major inhibitory neurotransmitter in the central nervous system and has several physiological functions. However, two other researchers have demonstrated that the GABA content of several pu-erh teas was low. Due to the high value and health benefits of GABA, analysis of mass-produced pu-erh tea is necessary to determine whether it is actually enriched with GABA. A high-performance liquid chromatography (HPLC) method was developed for the determination of GABA in tea, the results of which were verified by amino acid analysis using an Amino Acid Analyzer (AAA). A total of 114 samples of various types of Chinese tea, including 62 pu-erh teas, 13 green teas, 8 oolong teas, 8 black teas, 3 white teas, 4 GABA teas, and 16 process samples from two industrial fermentations of pu-erh tea (including the raw material and the first to seventh turnings), were analyzed using HPLC. Statistical analysis demonstrated that the GABA content in pu-erh tea was significantly lower than that in other types of tea (p < 0.05) and that the GABA content decreased during industrial fermentation of pu-erh tea (p < 0.05). This mass analysis and comparison suggested GABA was not a major bioactive constituent and resolved the disagreement GABA content in pu-erh tea. In addition, the GABA content in white tea was found to be significantly higher than that in the other types of tea (p < 0.05), leading to the possibility of producing GABA-enriched white tea.