A Synergetic Pore Compartmentalization and Hydrophobization Strategy for Synchronously Boosting the Stability and Activity of Enzyme.

Spatial immobilization of fragile enzymes using a nanocarrier is an efficient means to design heterogeneous biocatalysts, presenting superior stability and recyclability to pristine enzymes. An immobilized enzyme, however, usually compromises its catalytic activity because of inevasible mass transfer issues and the unfavorable conformation changes in a confined environment. Here, we describe a synergetic metal-organic framework pore-engineering strategy to trap lipase (an important hydrolase), which confers lipase-boosted stability and activity simultaneously. The hierarchically porous NU-1003, featuring interconnected mesopore and micropore channels, is precisely modified by chain-adjustable fatty acids on its mesopore channel, into which lipase is trapped. The interconnected pore structure ensures efficient communication between trapped lipase and exterior media, while the fatty acid-mediated hydrophobic pore can activate the opening conformation of lipase by interfacial interaction. Such dual pore compartmentalization and hydrophobization activation effects render the catalytic center of trapped lipase highly accessible, resulting in 1.57-fold and 2.46-fold activities as native lipase on ester hydrolysis and enantioselective catalysis. In addition, the feasibility of these heterogeneous biocatalysts for kinetic resolution of enantiomer is also validated, showing much higher efficiency than native lipase.

Facile Synthesis of Multifunctional Mesoporous Starch-Based Microparticle for Effective Hemostasis and Wound Healing.

Uncontrolled hemorrhage and infection are the principal causes of mortality associated with trauma in both military and civilian medical settings. Modified starch granules have emerged as a safe hemostatic agent for irregular and noncompressible wounds, but their performance is constrained by limited hemostasis efficiency and modest antibacterial activity. This study reported a directed self-assembly approach for a multifunctional mesoporous starch-based microparticle loaded with chitosan and calcium ions (Ca@MSMP) used for rapid hemostasis and wound healing. Directed self-assembly of uniform Ca@MSMP with a hierarchical hollow structure in the presence of chitosan was confirmed by scanning electron microscopy (SEM) analysis and pore structure analysis. The resulting Ca@MSMP exhibited a well-defined spherical shape and uniform size of 1 µm and demonstrated excellent antibacterial activity (>95%) without hemolytic activity. Importantly, Ca@MSMP enhanced blood coagulation and platelet aggregation via the synergistic effect of rapid calcium release and chitosan-mediated electrostatic interactions, leading to a significant decrease in blood loss and reduction in hemostasis time in rat tail amputation and liver injury models. In comparative analyses, Ca@MSMP significantly outperformed the commercial hemostatic agent Quickclean, notably enhancing the healing of full-thickness skin wounds in vivo by effectively preventing infection. These results underscore the potential of this innovative hemostatic material in diverse clinical scenarios, offering effective solutions for the management of bleeding in wounds that are irregularly shaped and noncompressible.

Hydrogen-Bonded Supramolecular Nanotrap Enabling the Interfacial Activation of Hosted Enzymes.

Engineering nanotraps to immobilize fragile enzymes provides new insights into designing stable and sustainable biocatalysts. However, the trade-off between activity and stability remains a long-standing challenge due to the inevitable diffusion barrier set up by nanocarriers. Herein, we report a synergetic interfacial activation strategy by virtue of hydrogen-bonded supramolecular encapsulation. The pore wall of the nanotrap, in which the enzyme is encapsulated, is modified with methyl struts in an atomically precise position. This well-designed supramolecular pore results in a synergism of hydrogen-bonded and hydrophobic interactions with the hosted enzyme, and it can modulate the catalytic center of the enzyme into a favorable configuration with high substrate accessibility and binding capability, which shows up to a 4.4-fold reaction rate and 4.9-fold conversion enhancements compared to free enzymes. This work sheds new light on the interfacial activation of enzymes using supramolecular engineering and also showcases the feasibility of interfacial assembly to access hierarchical biocatalysts featuring high activity and stability simultaneously.

Green synthesis of stable hybrid biocatalyst using a hydrogen-bonded, π-π-stacking supramolecular assembly for electrochemical immunosensor.

Rational integration of native enzymes and nanoscaffold is an efficient means to access robust biocatalyst, yet remains on-going challenges due to the trade-off between fragile enzymes and harsh assembling conditions. Here, we report a supramolecular strategy enabling the in situ fusion of fragile enzymes into a robust porous crystal. A c2-symmetric pyrene tecton with four formic acid arms is utilized as the building block to engineer this hybrid biocatalyst. The decorated formic acid arms afford the pyrene tectons high dispersibility in minute amount of organic solvent, and permit the hydrogen-bonded linkage of discrete pyrene tectons to an extended supramolecular network around an enzyme in almost organic solvent-free aqueous solution. This hybrid biocatalyst is covered by long-range ordered pore channels, which can serve as the gating to sieve the catalytic substrate and thus enhance the biocatalytic selectivity. Given the structural integration, a supramolecular biocatalyst-based electrochemical immunosensor is developed, enabling the pg/mL detection of cancer biomarker.

Impact of solvent polarity on the morphology, physicochemical properties, and digestibility of A-type resistant starch particles.

Resistant starch particles (RSP) formed by antisolvent precipitation method has attracted much attention as a functional food ingredient having beneficial effects on obesity and diabetes. However, the effect of solvent polarity on the physicochemical properties and digestibility of RSP remains unclear. Here, n-propanol, isopropanol, acetone, and ethanol were employed as antisolvents to prepare RSP. The width and length of the resulting RSP decreased from 0.87 µm to 0.59 µm and from 2.56 µm to 1.31 µm, respectively, upon increasing the solvent polarity, while dramatically decreasing their crystallinity and the gelatinization enthalpy from 80.5% to 62.3% and from 67.9 ± 14.4 J/g to 41.5 ± 8.3 J/g, respectively, suggesting that solvent polarity is critical factor determining morphology, crystallinity, and thermostability of RSP. Furthermore, the level of resistant starch in RSP was found to be inversely proportional to the degree of solvent polarity, which would provide a useful means for modulating the digestibility of RSP.

Detection of bile acids in small volume human bile samples via an amino metal-organic framework composite based solid-phase microextraction probe.

In recent years, bile acids (BAs), the important component of bile, were found closely related to the occurrence and development of diseases, therefore, determination of BAs in bile samples is of great significance. However, biological matrix complexity and low concentrations of BAs were still challenging for BA detection in small amount of bile samples. In this work, a core-shell NH2-MIL101@mSiO2 was designed to improve the capture ability of BAs in biological samples, as well as possess good biocompatibility. Subsequently, solid-phase microextraction (SPME) probe of the NH2-MIL101@mSiO2 was coupled with HPLC-MS/MS to establish the analysis method for detecting eight BAs in bile samples. The established method received extraction efficiencies of (30-2143)-fold higher than those of the commercial probes and low limit of detection (LOD ≤ 0.21 ng mL-1). The miniaturization of SPME sampling devices, as well as the low LOD of this work, endowed this method advantage of low consumption of bile samples (30 µL). Based on the proposed method, eight BAs in bile samples of pancreatic cancer patients and cholelithiasis patients were detected successfully. A distinct difference was found in the concentrations of four targeted BAs in bile samples from pancreatic cancer patients and cholelithiasis patients. This work provided a method for quantification of eight BAs in small volume human bile samples, and it could open up a perspective regarding the relationship between BA metabolism and the occurrence of diseases.

Hydrogen-bonded organic framework biomimetic entrapment allowing non-native biocatalytic activity in enzyme.

Nature programs the structural folding of an enzyme that allows its on-demand biofunctionality; however, it is still a long-standing challenge to manually modulate an enzyme's conformation. Here, we design an exogenous hydrogen-bonded organic framework to modulate the conformation of cytochrome c, and hence allow non-native bioactivity for the enzyme. The rigid hydrogen-bonded organic framework, with net-arranged carboxylate inner cage, is in situ installed onto the native cytochrome c. The resultant hydrogen-bonded nano-biointerface changes the conformation to a previously not achieved catalase-like species within the reported cytochrome c-porous organic framework systems. In addition, the preserved hydrogen-bonded organic framework can stabilize the encapsulated enzyme and its channel-like pores also guarantee the free entrance of catalytic substrates. This work describes a conceptual nanotechnology for manoeuvring the flexible conformations of an enzyme, and also highlights the advantages of artificial hydrogen-bonded scaffolds to modulate enzyme activity.

Fabrication of starch/zein-based microcapsules for encapsulation and delivery of fucoxanthin.

As a drug carrier, starch-based microparticle (SMP) has attracted widespread attention. However, because SMP is commonly formed in aqueous media, it is facing the challenge of encapsulation of hydrophobic bioactive. Here, we present an effective method for encapsulating fucoxanthin (Fx), a model hydrophobic bioactive, within SMP formed by self-assembly of short-chain glucans (SCG), using zein nanoparticles as intermediate vectors. SMP exhibited higher encapsulation efficiency to chitosan-coated Fx-zein nanoparticles (∼91%) at a given concentration of 50 µg/mL compared to soybean polysaccharide-coated ones. Fx in SMP was found to be more stable against UV radiation-induced degradation and FeCl3-catalyzed oxidative stresses than free Fx. Furthermore, SMP conferred controlled release of Fx in the duodenum (6%), jejunum (13%), ileum (32%), and colon (42%), implying that this approach could be useful in designing an effective drug carrier for delivering several hydrophobic bioactives to different parts of the intestine.

Facile Synthesis of a Fluorinated-Squaramide Covalent Organic Framework for the Highly Efficient and Broad-Spectrum Removal of Per- and Polyfluoroalkyl Pollutants.

The escalating contamination by per- and polyfluoroalkyl substances (PFAS) has become an urgent issue in recent years, and the structural diversity of PFAS is the major challenge for effective pollution control. Herein, we take the intrinsic advantages of squaramide and prepare a new two-dimensional covalent organic framework (FSQ-1) that exhibits broad-spectrum PFAS affinity. The tailor-made linker forges hydrogen-bond donors, hydrogen-bond acceptors, and fluorophilic segments into one framework. The obtained material exhibits multipoint and multitype affinity to PFAS with different structures, by which high-efficient and broad-spectrum removal of various PFAS can be simultaneously achieved. Notably, the thermodynamic profiles provided by isothermal titration calorimetry (ITC) experiments further illustrate the underlying mechanism of the broad-spectrum affinity. FSQ-1 can also be applied for efficient PFAS extraction in trace-level PFAS analysis.

Noncovalently Tagged Gas Phase Complex Ions for Screening Unknown Contaminant Metabolites in Plants.

Screening the metabolites of emerging organic contaminants (EOCs) from complicated biological matrices is an important but challenging task. Although stable isotope labeling (SIL) is frequently used to facilitate the identification of contaminant metabolites from redundant interfering components, the isotopically labeled reagents are expensive and difficult to synthesize, which greatly constrains the application of the SIL method. Herein, a new online noncovalent tagging method was developed for screening the metabolites of 1H-benzotriazol (BT) based on the characteristic structural moieties reserved in the metabolites. By selecting ß-cyclodextrin (ß-CD) as a macrocyclic tagging reagent, metabolites with the reserved moiety were expected to exhibit a characteristic shift of the mass-to-charge ratio (Δm/z = 1134.3698) after being noncovalently tagged by ß-CD. Based on the characteristic mass shift, the suspected features were reduced by 1 order of magnitude, as numerous interfering species that could not be effectively tagged by ß-CD were excluded. From these suspected features, two metabolites of BT that have not been reported before were successfully screened out. The significant characteristic mass shift caused by the noncovalent tagging method is easier to identify with more confidence than the previously reported SIL method. Besides, noncovalent tagging reagents can be much more accessible and less expensive than isotopically labeled reagents. Hence, this online noncovalent tagging method can be an intriguing alternative to the conventional SIL method.

Facile fabrication of composited solid phase microextraction thin membranes for sensitive detections of trace hydroxylated polycyclic aromatic hydrocarbons in human urine.

Solid phase microextraction (SPME) has potential to be used for the high-performance enrichments of hydroxylated polycyclic aromatic hydrocarbons (OH-PAHs), which are important biomarkers of PAH exposure. By choosing suitable adsorbent, it is conducive to fabricate new-type of SPME device for improved extraction efficiencies towards OH-PAHs. In this study, a novel method of surface solvent evaporation has been proposed to fabricate SPME thin membrane, integrating the advantages of polydimethylsiloxane (PDMS) and different porous adsorbents. The powdery metal organic framework (Uio66-NH2), porous polymer (powdery polymer aerogel, PPA) and ordered mesoporous carbon (OMC) have been chosen as typical adsorbents and fabricated as thin membranes successfully, indicating the universality of the proposed method for membrane fabrication. Comparing the extraction efficiencies of three prepared membranes towards OH-PAHs, the OMC-PDMS membrane has demonstrated best enrichment efficiencies. The OMC-PDMS membrane was used for the enrichments of trace OH-PAHs in human urine of both smokers and nonsmokers, combining with liquid chromatographic tandem mass spectrometry (LC-MS/MS). The detection limits were in the range of 0.15-0.39 ng L-1, and satisfactory recoveries were found to be 82.1%-115%. It can be seen that the universal strategy to fabricate SPME membrane is helpful to achieve broad-spectrum or selective enrichments of target analytes from complex matrix by simple modulation of membrane components.

In-situ layer-by-layer synthesized TpPa-1 COF solid-phase microextraction fiber for detecting sex hormones in serum.

The secretion disorder of sex hormones is the source that leads to the occurrence of many diseases such as polycystic ovarian syndrome (PCOS), hyperandrogenism and so on. There exist physiological changes in human body when slight fluctuations in concentrations of sex hormones happen. Therefore, it's of great significance for accurate detection of sex hormones in human body. In this work, TpPa-1 COF solid-phase microextraction (SPME) fiber was prepared using high-efficient in-situ synthesis strategy and coupled with HPLC-MS/MS to detect three sex hormones, including Progesterone (P), testosterone (T) and dehydroepiandrosterone (DHEA) in human serum. The thickness of the coating reached 7 µm within 2 h. Under the optimal conditions, the established method presented low limit of detections (LODs, ≤ 0.75 ng/mL), wide linear ranges (0.100-100 ng mL-1) and good reproducibility, and three sex hormones (T, P, DHEA) were successfully detected and quantified in human serum. In conclusion, the established SPME method presented high-efficient fiber preparation and good analytical performances of sex hormone detection, therefore was in great potential for application in clinical.

Trends in sensitive detection and rapid removal of sulfonamides: A review.

Sulfonamides in environmental water, food, and feed are a major concern for both aquatic ecosystems and public health, because they may lead to the health risk of drug resistance. Thus, numerous sensitive detection and rapid removal methodologies have been established. This review summarizes the sample preparation techniques and instrumental methods used for sensitive detection of sulfonamides. Additionally, adsorption and photocatalysis for the rapid removal of sulfonamides are also discussed. This review provides a comprehensive perspective on future sulfonamide analyses that have good performance, and on the basic methods for the rapid removal of sulfonamides.

Polydopamine decorated ordered mesoporous carbon for efficient removal of bilirubin under albumin-rich conditions.

Excess bilirubin in the body will lead to serious health problems; however, its efficient removal remains a challenge in the clinical field because the available sorbent materials still suffer from serious performance issues, performance declining in a high-content albumin environment. Herein, we prepared a novel polydopamine (PDA) decorated ordered mesoporous carbon (OMC) material for the efficient removal of bilirubin in albumin-rich conditions. OMC was used as the supporting material due to its high specific surface area and its good affinity to hydrophobic analytes. PDA was then decorated on the OMC material through a facile self-assembly process to form a surface-imprinted layer. The obtained PDA-coated OMC material (OMC@PDA) exhibited excellent adsorption performance towards bilirubin in albumin-free conditions, in which its theoretical maximum adsorption amount was calculated to be 513.54 mg g-1. The imprinted PDA layer, for which the association constant towards bilirubin reached 4.51 × 104 M-1, endowed OMC@PDA with a competitive affinity compared to albumin. Therefore the materials showed good adsorption capacity and efficiency even in an albumin-rich environment (the adsorption equilibrated at 122.7 mg g-1 in 30 min). In addition, the good biocompatibility of OMC@PDA was demonstrated by hemolysis assay and protein fouling evaluation, which indicated the feasibility of applying this material in clinical situations.

A Convenient and Versatile Amino-Acid-Boosted Biomimetic Strategy for the Nondestructive Encapsulation of Biomacromolecules within Metal-Organic Frameworks.

Herein, an amino-acid-boosted biomimetic strategy is reported that enabled the rapid encapsulation, or co-encapsulation, of a broad range of proteins into microporous metal-organic frameworks (MOFs), with an ultrahigh loading efficiency. It relies on the accelerated formation of prenucleation clusters around proteins via a metallothionein-like self-assembly. The encapsulated proteins maintained their native conformations, and the structural confinement within porous MOFs endowed enzymes with excellent bioactivity, even in harsh conditions (e.g. in the presence of proteolytic or chemical agents or at high temperature). Furthermore, owing to the merits of nondestructive and protein surface charge-independent encapsulation, the feasibility of this biomimetic strategy for biostorage, enzyme cascades, and biosensing was also verified. It is believed that this convenient and versatile encapsulation strategy has great promise in the important fields of biomedicine, catalysis, and biosensing.

High-Efficiency, Matrix Interference-Free, General Applicable Probes for Bile Acids Extraction and Detection.

Although bile acids (BAs) have been suggested as important biomarkers for endocrine diseases, the identification and quantification of different BAs are still challenges due to their enormous species and wide range concentrations. Herein, a copolymer probe based on ß-cyclodextrin (ß-CD) is fabricated through a simple in-mold photopolymerization for the selective extraction of BAs. Through the unique stereochemical affinity between BAs and the cavity of ß-CD, the custom probe shows superior enriching capacities to series BAs. Moreover, the outstanding extraction ability is proved to be consistent in various interfering conditions, including pH changing and the addition of complex matrix. Further comparison shows that the stereostructure of the nucleus of BAs plays a vital role during the formation of the ß-CD/BA complex, indicating the potential for efficient extraction of other BAs, including their structural analogues or some unknown ones. The developed probe is used for solid phase microextraction, and the limits of detection are lower than 0.075 ng mL-1 by coupling to high performance liquid chromatography-tandem mass analysis. The results in this study highlight the potential for effective improvement of immediate detection and profiling of BAs in real samples, which will make a tremendous impact in the analytical field or clinical diagnosis.

Synthesis and application of magnetic molecularly imprinted polymers in sample preparation.

Magnetic molecularly imprinted polymers (MMIPs) have superior advantages in sample pretreatment because of their high selectivity for target analytes and the fast and easy isolation from samples. To meet the demand of both good magnetic property and good extraction performance, MMIPs with various structures, from traditional core-shell structures to novel composite structures with a larger specific surface area and more accessible binding sites, are fabricated by different preparation technologies. Moreover, as the molecularly imprinted polymer (MIP) layers determine the affinity, selectivity, and saturated adsorption amount of MMIPs, the development and innovation of the MIP layer are attracting attention and are reviewed here. Many studies that used MMIPs as sorbents in dispersive solid-phase extraction of complex samples, including environmental, food, and biofluid samples, are summarized. Graphical abstract The application of magnetic molecularly imprinted polymers (MIPs) in the sample preparation procedure improves the analytical performances for complex samples. MITs molecular imprinting technologies.

Improving the Sensitivity of Solid-Phase Microextraction by Reducing the Volume of Off-Line Elution Solvent.

Solid-phase microextraction (SPME) coupled with liquid chromatograph (LC) is widely used to detect polar and ionic organic compounds, including various pharmaceuticals and endogenous bioactive compounds. In this study, a small-sized insert tube for use in the commercial autosampler vial was designed for eluting the extracted analytes from SPME fibers for LC analysis. By using this custom-made insert tube as an alternative to the commercial insert tube, the volume of the elution solvent was reduced by four-fifths. Even though smaller fractions of the analytes were eluted from the fiber coatings, the analyte concentrations in the elution solutions were substantially increased by using the custom-made insert tube. Therefore, larger amounts of the analytes could be injected to LC and higher signal-to-noise ratios could be achieved, even at smaller injection volumes. Since the elution in the custom-made insert tube was nonexhaustive, four strategies were developed to figure out the extracted amounts in the fiber coatings. In combination with the sampling-rate calibration method, these strategies were successfully used to determine the concentrations of fluoxetine in living tilapias. This study provides a simple but effect way for improving the analytical sensitivity when coupling SPME with LC.

Rapid in vivo determination of tetrodotoxin in pufferfish (Fugu) muscle by solid-phase microextraction coupled to high-performance liquid chromatography tandem mass spectrometry.

Tetrodotoxin (TTX) is one of the most toxic substances of non-protein in nature. In present study, a rapid and sensitive method based on in vivo solid-phase microextraction (SPME) coupled to liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed to detect the endogenous TTX in pufferfish (Fugu). Fiber evaluation experiments demonstrated that, compared with the commercial PDMS and PA fibers, the home-made electrospun PS@PDA-GA fibers exhibited much better extraction performance towards to TTX in water (120 times and 20 times, respectively), under the optimized conditions. Then, the home-made SPME fibers were employed to extract TTX in spiked homogeneous fish muscle samples, and a LC-MS-MS was used for the analysis. The reproducibilities (RSDs of inter and intra fiber were 12.1%, and 7.9% respectively), linear range (10-1000ngg-1, R2=0.9963) and sensitivity (the LOD was 2.3ngg-1) of the method were found to be excellent and satisfactory for further in vivo experiments. Especially the LOD of the established method is lower than the National Standard Method of China (GB/T 23217-2008, LOD 50ngg-1). Subsequently, the method was successfully applied to detect the TTX in the dorsal muscle of living pufferfish, and the accuracy was verified with traditional liquid extraction (LE) method. In general, this is the first study to detect TTX in pufferfish by in vivo sampling method, which provided a promising alternative method for the studies of TTX, and also advanced the implementation of SPME for more in vivo studies.