Synthesis of Cycloaliphatic Polyamides via Palladium-Catalyzed Hydroaminocarbonylative Polymerization.

Polyamides represent one class of materials that is important in modern society. Because of the numerous potential applications of polyamides in various fields, there is a high demand for new polyamide structures, which necessitates the development of new polymerization methods. Herein, we report a novel and efficient palladium-catalyzed hydroaminocarbonylative polymerization of dienes and diamines for the synthesis of cycloaliphatic polyamides. The method employs readily available starting materials, proceeds in an atom-economic manner, and creates a series of new functional polyamides in high yields and high molecular weights. In contrast with the traditional polyamides based on adipic acid, the cycloaliphatic polyamides have superior thermal resistance, higher glass-transition temperature, and better solubility in common organic solvents, thus probably featuring the merits of high-performance and good processability.

Hollow versatile Ag@Pt alloy nanoparticles with nanozyme activity for detection and photothermal sterilization of Helicobacter pylori.

In view of a large number of people infected with Helicobacter pylori (H. pylori) with great harm followed, there is an urgent need to develop a non-invasive, easy-to-operate, and rapid detection method, and to identify effective sterilization strategies. In this study, highly specific nanoprobes with nanozyme activity, Ag@Pt nanoparticles (NPs) with the antibody, were utilized as a novel lateral flow immunoassay (LFIA). The optical label (Ag@Pt NPs) was enhanced by the introduction of the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) and compared with a gold nanoparticles (Au NPs) optical label. Under the optimal condition, Ag@Pt-LFIA and TMB-enhanced Ag@Pt-LFIA for H. pylori were successfully established, two of which were over twofold and 100-fold more sensitive than conventional visual Au NP-based LFIA, respectively. Furthermore, Ag@Pt NPs with the antibody irradiated with NIR laser (808 nm) at a power intensity of 550 mW/cm2 for 5 min exhibited a remarkable antibacterial effect. The nanoprobes could close to bacteria through effective interactions between antibodies and bacteria, thereby benefiting photothermal sterilization. Overall, Ag@Pt NPs provide promising applications in pathogen detection and therapeutic applications.

Palladium-Catalyzed Inward Isomerization Hydroaminocarbonylation of Alkenes.

In contrast to the kinetically favored outward isomerization-hydrocarbonylation of alkenes, the disfavored inward isomerization-hydrocarbonylation of alkenes remains an important challenge. Herein, we have developed a novel and effective palladium-catalyzed inward isomerization-hydroaminocarbonylation of unactivated alkenes and aniline hydrochlorides for the formation of synthetically valuable α-aryl carboxylic amides in high yields and high site-selectivities. The high efficiency of the reaction is attributed to a relay catalysis strategy, in which the Markovnikov-favored [PdH]-PtBu3 catalyst is responsible for inward isomerization, while the [PdH]-Ruphos catalyst is responsible for hydroaminocarbonylation of the resulting conjugated aryl alkenes. The reaction exhibits highly functional group tolerance and provides a new method for formal carbonylation of remote C(sp3)-H bond.

Mix-and-Read Nanobody-Based Sandwich Homogeneous Split-Luciferase Assay for the Rapid Detection of Human Soluble Epoxide Hydrolase.

The soluble epoxide hydrolase (sEH) is possibly both a marker for and target of numerous diseases. Herein, we describe a homogeneous mix-and-read assay for the detection of human sEH based on using split-luciferase detection coupled with anti-sEH nanobodies. Selective anti-sEH nanobodies were individually fused with NanoLuc Binary Technology (NanoBiT), which consists of a large and small portion of NanoLuc (LgBiT and SmBiT, respectively). Different orientations of the LgBiT and SmBiT-nanobody fusions were expressed and investigated for their ability to reform the active NanoLuc in the presence of the sEH. After optimization, the linear range of the assay could reach 3 orders of magnitude with a limit of detection (LOD) of 1.4 ng/mL. The assay has a high sensitivity to human sEH and reached a similar detection limit to our previously reported conventional nanobody-based ELISA. The procedure of the assay was faster (30 min total) and easy to operate, providing a more flexible and simple way to monitor human sEH levels in biological samples. In general, the immunoassay proposed here offers a more efficient detection and quantification approach that can be easily adapted to numerous macromolecules.

Development of a sandwich ELISA for the specific quantitation of hemagglutinin (HA)-tagged proteins during their inducible expression in Escherichia coli.

Heavy single-chain antibodies (VHH or nanobodies) are popular in the medical and analytical fields due to its small size, high solubility, stability, and other advantageous features. However, the usage of VHHs is limited by the low yield of its production and purification. In order to determine the optimal purification strategy for VHH to improve the yield, a method to monitor purification at the intermediate steps is needed. In this study, a simple, sensitive, low-cost sandwich enzyme-linked immunosorbent assay (ELISA) was developed to quantitate VHHs throughout the purification steps. Under optimized conditions, the assay has a sensitivity of 0.149 OD·mL/ng and a limit of detection (LOD) of 0.029 ng/mL. The average recoveries of the assay against the spiked samples were 101.9-106.0% and 100.7-108.0%. The method was applied to a variety of real samples for the detection of different VHHs in bacterial cell media. High amount of VHHs (up to 41.3 mg/mL), which are comparable to the average yield of VHH in standard production protocols, were detected in the media. This study raises attention to the problem of protein losses in cell culture supernatants and provides a method for the continuous detection of the protein abundance to optimize the expression and purification protocols especially for nanobodies.

The Generation of a Nanobody-Based ELISA for Human Microsomal Epoxide Hydrolase.

A microsomal epoxide hydrolase (mEH) metabolizes in vivo in both xenobiotic and endogenous epoxides associated with signaling function. Findings in patients suggest that mEH might be a biomarker for several diseases, including metastatic cancer and viral hepatitis. To easily quantify mEH, nanobodies specific to the human mEH were isolated from a phage library of llama VHHs. Four unique clones were obtained and used for developing ELISAs. Three formats of double antibody sandwich assays were investigated using different detection strategies. Using PolyHRP, the signal was strongly amplified, yielding a 22-fold lower LOD (12 pg mL-1) than the 'conventional'. To further validate the performance of the immunoassays, human tissue samples were analyzed by nanobody-based ELISAs and compared to the enzyme activities (R2 > 0.95). The results demonstrate that these nanobodies are powerful tools for the quantification of human mEH and could eventually result in a bedside assay.

Generation of bioluminescent enzyme immunoassay for ferritin by single-chain variable fragment and its NanoLuc luciferase fusion.

Ferritin, widely present in liver and spleen tissue, is considered as a serological biomarker for liver diseases and cancers. The detection of ferritin may be an important tool in health diagnosis. In this study, 14 non-immunized chicken spleens were utilized to construct a single-chain fragment (scFv) phage library. After 4 rounds of panning, 7 unique clones were obtained. The optimal clone was further screened and combined with NanoLuc luciferase (Nluc) as a dual functional immunoprobe to bioluminescent enzyme immunoassay (BLEIA), which was twice as sensitive as its parental scFv-based double-sandwich enzyme-linked immunoassay (ds-ELISA). The cross-reactivity analysis revealed that the proposed methods were highly selective and suitable for clinical detection. To further verify the performance of the immunoassays, serum samples were tested by the proposed methods and a commercial ELISA kit, and there was a good correlation between the results. These results suggested that scFv fused with Nluc might be a powerful dual functional tool for rapid, practically reliable, and highly sensitive ferritin detection.

Asymmetric Markovnikov Hydroaminocarbonylation of Alkenes Enabled by Palladium-Monodentate Phosphoramidite Catalysis.

A palladium-catalyzed asymmetric Markovnikov hydroaminocarbonylation of alkenes with anilines has been developed for the atom-economical synthesis of 2-substituted propanamides bearing an α-stereocenter. A novel phosphoramidite ligand L16 was discovered which exhibited very high reactivity and selectivity in the reaction. This asymmetric Markovnikov hydroaminocarbonylation employs readily available starting materials and tolerates a wide range of functional groups, thus providing a facile and straightforward method for the regio- and enantioselective synthesis of 2-substituted propanamides under ambient conditions. Mechanistic studies revealed that the reaction proceeds through a palladium hydride pathway.

Palladium-Catalyzed Markovnikov Hydroaminocarbonylation of 1,1-Disubstituted and 1,1,2-Trisubstituted Alkenes for Formation of Amides with Quaternary Carbon.

Hydroaminocarbonylation of alkenes is one of the most promising yet challenging methods for the synthesis of amides. Herein, we reported the development of a novel and effective Pd-catalyzed Markovnikov hydroaminocarbonylation of 1,1-disubstituted or 1,1,2-trisubstituted alkenes with aniline hydrochloride salts to afford amides bearing an α quaternary carbon. The reaction makes use of readily available starting materials, tolerates a wide range of functional groups, and provides a facile and straightforward approach to a diverse array of amides bearing an α quaternary carbon. Mechanistic investigations suggested that the reaction proceeded through a palladium hydride pathway. The hydropalladation and CO insertion are reversible, and the aminolysis is probably the rate-limiting step.

A Pt-Ir nanocube amplified lateral flow immunoassay for dehydroepiandrosterone.

The traditional gold-nanoparticle-based lateral flow immunoassay (LFIA) cannot satisfy the requirements for the sensitive detection of dehydroepiandrosterone (DHEA) in human urine. To enhance the sensitivity of the LFIA, platinum-iridium nanocubes (Pt-Ir NCs) with high catalytic efficiency and stability were synthesized and labelled with polyclonal antibody (pAb) to form a pAb-Pt-Ir probe. For the detection of DHEA, a novel LFIA with Pt-Ir NCs as an optical label and an enhanced LFIA in which the peroxidase-like activity of the Pt-Ir NCs was triggered by the introduction of the chromogenic substrate 3-amino-9-ethyl-carbazole (AEC) were developed and compared with a LFIA with platinum nanocubes (PtNCs) as an optical label. The visual limit of detection was 0.5 ng mL-1 for Pt-Ir-LFIA and 0.05 ng mL-1 for AEC-enhanced Pt-Ir-LFIA, in comparison to 100 ng mL-1 for PtNCs-LFIA and 50 ng mL-1 for AEC-enhanced PtNCs-LFIA. The average recoveries from spiked urine samples ranged from 90.8% to 110.4%, with a coefficient of variation below 12.6%, suggesting the accuracy and reliability of our developed immunoassay. Achieving excellent sensitivity, specificity, and reproducibility, Pt-Ir-LFIA provided a promising platform for monitoring DHEA.

Fluorescence polarization immunoassay for rapid determination of dehydroepiandrosterone in human urine.

In this paper, five fluorescein-labeled dehydroepiandrosterone (DHEA) derivatives (tracers) with different chain lengths between the fluorescein and hapten were synthesized and featured so as to establish a fluorescence polarization immunoassay (FPIA) for DHEA detection in human urine samples with previously prepared polyclonal antibody against DHEA. The outcomes of the structure of tracer on FPIA sensitivity were investigated. Under the optimal condition, the fluorescence polarization value (FP) decreases linearly in DHEA concentration, ranging from 1.6 to 243.3 ng mL-1, with the limit of detection of 1.1 ng mL-1 and IC50 value of 25.1 ng mL-1. Moreover, the developed FPIA was time-saving as it could complete the detection within 3 min. FPIA and commercial enzyme-linked immunosorbent assay kit were both applied to analyze the spiked human urine samples with DHEA. Excellent recoveries (92.1-108.0%) and satisfactory correlation coefficient (R2 = 0.98) were acquired with the two methods, indicating that the developed FPIA was a fast and efficient screening immunoassay with accuracy and sensitivity for DHEA detection in human urine samples. Graphical abstract.

Development of enzyme-free single-step immunoassays for glycocholic acid based on palladium nanoparticle-mediated signal generation.

Palladium nanoparticles (PdNPs) are composed mainly of inert noble metals, and their outstanding properties have attracted wide attention. PdNPs are not only capable of mimicking the oxidase-like characteristics of natural bio-enzymes, but they also present a clear black band in the test zone. In this work, the synthesized PdNPs promoted a transformation of colorless tetramethylbenzidine (TMB) to a blue oxidation product of TMB, providing a Km value of 0.09 mM for TMB, and revealing the good catalytic performance of the synthesized PdNPs. For both signal generation and amplification, PdNPs effectively replaced natural bio-enzymes as a new labeling tag. Thus, the PdNP-based enzyme-free single-step immunoassays were successfully developed for efficient and sensitive detection of glycocholic acid (GCA). Under optimal conditions, a noticeable linear relationship was identified by the enzyme-linked immunosorbent assay (ELISA) over a range of 8-2390 ng/mL, while the visual limit of detection (vLOD) in the constructed lateral flow immunoassay (LFA) was 10 ng/mL for GCA. The recovery rate in spiked urine samples obtained by the ELISA ranged from 84.2 to 117.9%, which was consistent with the results in LFA. The present work demonstrates the potential of PdNPs as labeling matrices in enzyme-free single-step immunoassays.

Palladium-Catalyzed Asymmetric Markovnikov Hydroxycarbonylation and Hydroalkoxycarbonylation of Vinyl Arenes: Synthesis of 2-Arylpropanoic Acids.

Asymmetric hydroxycarbonylation is one of the most fundamental yet challenging methods for the synthesis of carboxylic acids. Herein, we reported the development of a palladium-catalyzed highly enantioselective Markovnikov hydroxycarbonylation of vinyl arenes with CO and water. A monodentate phosphoramidite ligand L6 plays vital role in the reaction. The reaction tolerates a range of functional groups, and provides a facile and atom-economical approach to an array of 2-arylpropanoic acids including several commonly used non-steroidal anti-inflammatory drugs. The catalytic system has also enabled an asymmetric Markovnikov hydroalkoxycarbonylation of vinyl arenes with alcohols to afford 2-arylpropanates. Mechanistic investigations suggested that the hydropalladation is irreversible and is the regio- and enantiodetermining step, while hydrolysis/alcoholysis is probably the rate-limiting step.

Platinum nanoflowers with peroxidase-like property in a dual immunoassay for dehydroepiandrosterone.

Platinum nanoflowers (PtNFs) were utilized in a competitive enzyme-linked immunosorbent assay (ELISA) and in a lateral flow immunoassay (LFIA) for superior peroxidase-like activity and intense brown color, respectively. PtNFs were linked to the polyclonal antibody (pAb) to form the pAb-PtNFs probes for the dual immunoassay. Based on optimized pAb-PtNF probes, both enzyme-linked immunosorbent assay (PtNFs-ELISA) and lateral flow immunoassay (PtNFs-LFIA) perform very well. The absorbance at 450 nm decreases linearly in the DHEA concentration range 2.1 to 118.1 ng mL-1, and the limit of detection is 1.3 ng mL-1 and the IC50 value is 15.7 ng mL-1 of PtNFs-ELISA. The visual cut-off value of PtNFs-LFIA is 10.0 ng mL-1. The average recoveries from spiked samples range from 95.0 to 108.9% with a coefficient of variation below 12.2%. Excellent recoveries and correlation between the two methods were observed. Furthermore, the designed immunosensors exhibited good selectivity, confirming a broad development prospect in DHEA monitoring. Graphical Abstract.

A colorimetric sensing strategy based on enzyme@metal-organic framework and oxidase-like IrO2/MnO2 nanocomposite for α-glucosidase inhibitor screening.

A highly sensitive colorimetric sensing strategy based on enzyme@metal-organic framework (GAA@Cu-MOF) and IrO2/MnO2 nanocomposite was exploited innovatively for screening of α-glucosidase (GAA) inhibitors. IrO2/MnO2 nanocomposite exhibits excellent oxidase-mimicking activity which can directly catalyze the oxidation of 3,3,5,5,-tetramethylbenzidine (TMB) into a blue product with an absorption maximum at 652 nm. And GAA@Cu-MOF can decompose L-ascorbic acid-2-O-α-D-glucopyranosyl (AAG) to ascorbic acid (AA). The produced AA can destroy the IrO2/MnO2 nanocomposite and reduce its oxidase-like activity. However, the generation of AA is restricted when GAA inhibitors are added to the system, which allows the oxidase-like activity of the IrO2/MnO2 nanocomposite to be maintained. In view of this, a method for screening of GAA inhibitors was developed. In addition to enhancing the stability of GAA, the method can also effectively avoid the potential interference of H2O2 in the screening process of GAA inhibitors, which helps to improve the sensitivity of the method. Therefore, highly sensitive determination for acarbose and ascorbic acid are achieved with detection limits of 6.27 nM and 1.23 µM, respectively. The proposed method was successfully applied to screen potential GAA inhibitors from oleanolic acid derivatives. Graphical abstract.

Differential responses of carbon-degrading enzyme activities to warming: Implications for soil respiration.

Extracellular enzymes catalyze rate-limiting steps in soil organic matter decomposition, and their activities (EEAs) play a key role in determining soil respiration (SR). Both EEAs and SR are highly sensitive to temperature, but their responses to climate warming remain poorly understood. Here, we present a meta-analysis on the response of soil cellulase and ligninase activities and SR to warming, synthesizing data from 56 studies. We found that warming significantly enhanced ligninase activity by 21.4% but had no effect on cellulase activity. Increases in ligninase activity were positively correlated with changes in SR, while no such relationship was found for cellulase. The warming response of ligninase activity was more closely related to the responses of SR than a wide range of environmental and experimental methodological factors. Furthermore, warming effects on ligninase activity increased with experiment duration. These results suggest that soil microorganisms sustain long-term increases in SR with warming by gradually increasing the degradation of the recalcitrant carbon pool.

Potential negative consequences of geoengineering on crop production: A study of Indian groundnut.

Geoengineering has been proposed to stabilize global temperature, but its impacts on crop production and stability are not fully understood. A few case studies suggest that certain crops are likely to benefit from solar dimming geoengineering, yet we show that geoengineering is projected to have detrimental effects for groundnut. Using an ensemble of crop-climate model simulations, we illustrate that groundnut yields in India undergo a statistically significant decrease of up to 20% as a result of solar dimming geoengineering relative to RCP4.5. It is somewhat reassuring, however, to find that after a sustained period of 50 years of geoengineering crop yields return to the nongeoengineered values within a few years once the intervention is ceased.

Dual lateral flow assay based on PdRu nanocages for human Papillomavirus detection.

Cervical cancer is one of the most common gynecological malignancies, with the vast majority of which being caused by persistent infection with Human Papillomavirus (HPV) 16 and 18. The current available HPV detection methods are sensitive and genotyped but are restricted by expensive instruments and skilled personnel. The development of an easy-to-use, rapid, and cost-friendly analysis method for HPV is of great need. Herein, hollow palladium-ruthenium nanocages modified with two oligonucleotides (PdRu capture probes) were constructed for genotyping and simultaneous detection of target nucleic acids HPV16 and HPV18 by dual lateral flow assay (DLFA). PdRu capture probes were endowed with bi-functions for the first time, which could be used to output signals and hybridize target nucleic acids. Under optimized conditions, the PdRu based-DLFA with detection limits of 0.93 nM and 0.19 nM, respectively, exhibited convenient operation, and high sensitivity. Meanwhile, the DLFA achieved excellent rapid detection within 20 min, which was attributed to capture probes that can be directly bound to amplification-free target nucleic acids. Therefore, the development of PdRu-based DLFA can be utilized for rapid, sensitive, and simultaneous genotyping detection of HPV16 and HPV18, showing great application for nucleic acid detection.

Integrated physiological, biochemical, and transcriptomics analyses reveal the underlying mechanisms of high nitrogen use efficiency of black sesame.

Cultivating high nitrogen use efficient varieties is a sustainable solution to mitigating adverse effects on the environment caused by excessive nitrogen fertilizer application. However, in sesame, although immoderate nitrogen fertilizers are used to promote yield, the molecular basis of high nitrogen use efficiency (NUE) is largely unknown. Hence, this study aimed to identify high NUE black sesame variety and dissect the underlying physiological and molecular mechanisms. To achieve this, seventeen seedling traits of 30 black sesame varieties were evaluated under low nitrogen (LN) and high nitrogen (HN) conditions. Dry matter accumulation, root parameters, shoot nitrogen accumulation, and chlorophyll content are important factors for evaluating the NUE of sesame genotypes. The variety 17-156 was identified as the most efficient for N utilization. Comparative physiological and transcriptomics analyses revealed that 17-156 possesses a sophisticated nitrogen metabolizing machinery to uptake and assimilate higher quantities of inorganic nitrogen into amino acids and proteins, and simultaneously improving carbon metabolism and growth. Specifically, the total nitrogen and soluble protein contents significantly increased with the increase in nitrogen concentrations. Many important genes, including nitrate transporters (NPFs), amino acid metabolism-related (GS, GOGAT, GDH, etc.), phytohormone-related, and transcription factors, were significantly up-regulated in 17-156 under HN condition. In addition, 38 potential candidate genes were identified for future studies toward improving sesame's NUE. These findings offer valuable resources for deciphering the regulatory network of nitrogen metabolism and developing sesame cultivars with improved NUE.

Application of Bioactive Materials for Osteogenic Function in Bone Tissue Engineering.

Bone tissue defects present a major challenge in orthopedic surgery. Bone tissue engineering using multiple versatile bioactive materials is a potential strategy for bone-defect repair and regeneration. Due to their unique physicochemical and mechanical properties, biofunctional materials can enhance cellular adhesion, proliferation, and osteogenic differentiation, thereby supporting and stimulating the formation of new bone tissue. 3D bioprinting and physical stimuli-responsive strategies have been employed in various studies on bone regeneration for the fabrication of desired multifunctional biomaterials with integrated bone tissue repair and regeneration properties. In this review, biomaterials applied to bone tissue engineering, emerging 3D bioprinting techniques, and physical stimuli-responsive strategies for the rational manufacturing of novel biomaterials with bone therapeutic and regenerative functions are summarized. Furthermore, the impact of biomaterials on the osteogenic differentiation of stem cells and the potential pathways associated with biomaterial-induced osteogenesis are discussed.