The Effect of Cellulose Nanocrystal Reassembly on Latex-Based Pressure-Sensitive Adhesive Performance.

Different cellulose nanocrystal (CNC) forms (dried vs never-dried) can lead to different degrees of CNC reassembly, the formation of nanofibril-like structures, in nanocomposite latex-based pressure-sensitive adhesive (PSA) formulations. CNC reassembly is also affected by CNC sonication and loading as well as the protocol used for CNC addition to the polymerization. In this study, carboxylated CNCs (cCNCs) were incorporated into a seeded, semibatch, 2-ethylhexyl acrylate/methyl methacrylate/styrene emulsion polymerization and cast as pressure-sensitive adhesive (PSA) films. The addition of CNCs led to a simultaneous increase in tack strength, peel strength, and shear adhesion, avoiding the typical trade-off between the adhesive and cohesive strength. Increased CNC reassembly resulted from the use of dried, redispersed, and sonicated cCNCs, along with increased cCNC loading and addition of the cCNCs at the seed stage of the polymerization. The increased degree of CNC reassembly was shown to significantly increase the shear adhesion by enhancing the elastic modulus of the PSA films.

Mammal hyaluronidase activity on chondroitin sulfate and dermatan sulfate: Mass spectrometry analysis of oligosaccharide products.

Mammalian hyaluronidases are endo-N-acetyl-D-hexosaminidases involved in the catabolism of hyaluronic acid (HA) but their role in the catabolism of chondroitin sulfate (CS) is also examined. HA and CS are glycosaminoglycans implicated in several physiological and pathological processes, and understanding their metabolism is of significant importance. Data have been previously reported on the degradation of CS under the action of hyaluronidase, yet a detailed structural investigation of CS depolymerization products remains necessary to improve our knowledge of the CS depolymerizing activity of hyaluronidase. For that purpose, the fine structural characterization of CS oligosaccharides formed upon the enzymatic depolymerization of various CS subtypes by hyaluronidase has been carried out by high-resolution Orbitrap mass spectrometry (MS) and extreme UV (XUV) photodissociation tandem MS. The exact mass measurements show the formation of wide size range of even oligosaccharides upon digestion of CS-A and CS-C comprising hexa- and octa-saccharides among the main digestion products, as well as formation of small quantities of odd-numbered oligosaccharides, while no hyaluronidase activity was detected on CS-B. In addition, slight differences have been observed in the distribution of oligosaccharides in the digestion mixture of CS-A and CS-C, the contribution of longer oligosaccharides being significantly higher for CS-C. The sequence of CS oligosaccharide products determined XUV photodissociation experiments verifies the selective ß(1 â†’ 4) glycosidic bond cleavage catalyzed by mammal hyaluronidase. The ability of the mammal hyaluronidase to produce hexa- and higher oligosaccharides supports its role in the catabolism of CS anchored to membrane proteoglycans and in extra-cellular matrix.

TUTORIAL: ION ACTIVATION IN TANDEM MASS SPECTROMETRY USING ULTRA-HIGH RESOLUTION INSTRUMENTATION.

Tandem mass spectrometry involves isolation of specific precursor ions and their subsequent excitation through collision-, photon-, or electron-mediated activation techniques in order to induce unimolecular dissociation leading to formation of fragment ions. These powerful ion activation techniques, typically used in between mass selection and mass analysis steps for structural elucidation, have not only found a wide variety of analytical applications in chemistry and biology, but they have also been used to study the fundamental properties of ions in the gas phase. In this tutorial paper, a brief overview is presented of the theories that have been used to describe the activation of ions and their subsequent unimolecular dissociation. Acronyms of the presented techniques include CID, PQD, HCD, SORI, SID, BIRD, IRMPD, UVPD, EPD, ECD, EDD, ETD, and EID. The fundamental principles of these techniques are discussed in the context of their implementation on ultra-high resolution tandem mass spectrometers. © 2020 John Wiley & Sons Ltd. Mass Spec Rev.

Interactive role of miR-29, miR-93, miR-205, and VEGF in salivary adenoid cystic carcinoma.

OBJECTIVES: Salivary adenoid cystic carcinoma (SACC) is one of the most common salivary gland tumors in which patients encounter local recurrence and lung metastases. Understanding prognostic biomarkers in SACC is essential for future development in prognosis and treatment. This study aimed to assess the expression level of vascular endothelial growth factor (VEGF) and its potential regulatory microRNAs in SACC for prognostic determination. MATERIAL AND METHODS: The expression of VEGF in SACC samples was assessed using immunohistochemistry. Potential regulatory microRNAs were evaluated using quantitative reverse transcription-polymerase chain reaction. Associations between VEGF and microRNAs expression and clinicopathological parameters were investigated. RESULTS: VEGF expression levels positively correlated with histologic grade (p = .004) and treatment modality (p = .04). Decreased expression of miR-29a (p = .01) and increased expression of miR-93-5p and miR-205 (both p < .0001) were observed in SACC compared to normal salivary gland tissue. MiR-93-5p showed a positive association (p = .02) with VEGF overexpression. CONCLUSIONS: Our results showed the downregulation of miR-29 and overexpression of miR-93 and miR-205 in the SACC group, and the correlation between miR-93 and VEGF suggests these biomarkers as potential prognostic markers in the future.

Benchmarking higher energy collision dissociation (HCD) by investigation of binding energies of gas-phase host-guest complexes of hemicryptophane cages.

Synthesis of host molecules that feature well-defined characteristics for molecular recognition of guest molecules is often a major aim of synthetic host-guest (H-G) chemistry. A key consideration in evaluating the selectivity of hosts and the affinities of guests is the measurement of binding energies of obtained H-G complexes. In contrast to nuclear magnetic resonance (NMR) or fluorescence measurements that are capable of measuring binding strengths in solution, mass spectrometry offers the opportunity to measure gas-phase binding energies. Presented in this article is a higher energy collision dissociation (HCD) approach for determining critical energies of dissociation of H-G complexes. Experiments were performed on electrospray ionization (ESI)-generated H-G pairs in an LTQ-XL/Orbitrap hybrid instrument. The presented HCD approach requires preliminary calibration of the internal energy distribution of generated ions that was achieved by the use of activation parameters that were known from previous low-energy collision-induced dissociation (low-energy CID) experiments. Internal energy deposition was modeled based on a truncated Maxwell-Boltzmann distribution and characteristic temperature (Tchar ). Using this method, critical energies of dissociation were determined for 10 H-G biologically relevant complexes of the heteroditopic hemicryptophane cage host (Host). Obtained results are compared with those found previously by low-energy CID. The use of this HCD technique is relatively straightforward, although its implementation does require knowledge (or a presumption) about the Arrhenius pre-exponential factor of the complexes to obtain their critical energies of dissociation.

Comprehensive structural assignment of glycosaminoglycan oligo- and polysaccharides by protein nanopore.

Glycosaminoglycans are highly anionic functional polysaccharides with information content in their structure that plays a major role in the communication between the cell and the extracellular environment. The study presented here reports the label-free detection and analysis of glycosaminoglycan molecules at the single molecule level using sensing by biological nanopore, thus addressing the need to decipher structural information in oligo- and polysaccharide sequences, which remains a major challenge for glycoscience. We demonstrate that a wild-type aerolysin nanopore can detect and characterize glycosaminoglycan oligosaccharides with various sulfate patterns, osidic bonds and epimers of uronic acid residues. Size discrimination of tetra- to icosasaccharides from heparin, chondroitin sulfate and dermatan sulfate was investigated and we show that different contents and distributions of sulfate groups can be detected. Remarkably, differences in α/ß anomerization and 1,4/1,3 osidic linkages can also be detected in heparosan and hyaluronic acid, as well as the subtle difference between the glucuronic/iduronic epimers in chondroitin and dermatan sulfate. Although, at this stage, discrimination of each of the constituent units of GAGs is not yet achieved at the single-molecule level, the resolution reached in this study is an essential step toward this ultimate goal.

Investigation of activation energies for dissociation of host-guest complexes in the gas phase using low-energy collision induced dissociation.

A low-energy collision induced dissociation (CID) (low-energy CID) approach that can determine both activation energy and activation entropy has been used to evaluate gas-phase binding energies of host-guest (H-G) complexes of a heteroditopic hemicryptophane cage host (Zn (II)@1) with a series of biologically relevant guests. In order to use this approach, preliminary calibration of the effective temperature of ions undergoing resonance excitation is required. This was accomplished by employing blackbody infrared radiative dissociation (BIRD) which allows direct measurement of activation parameters. Activation energies and pre-exponential factors were evaluated for more than 10 H-G complexes via the use of low-energy CID. The relatively long residence time of the ions inside the linear ion trap (maximum of 60 s) allowed the study of dissociations with rates below 1 s-1 . This possibility, along with the large size of the investigated ions, ensures the fulfilment of rapid energy exchange (REX) conditions and, as a consequence, accurate application of the Arrhenius equation. Compared with the BIRD technique, low-energy CID allows access to higher effective temperatures, thereby permitting one to probe more endothermic decomposition pathways. Based on the measured activation parameters, guests bearing a phosphate (-OPO3 2- ) functional group were found to bind more strongly with the encapsulating cage than those having a sulfonate (-SO3 - ) group; however, the latter ones make stronger bonds than those with a carboxylate (-CO2 - ) group. In addition, it was observed that the presence of trimethylammonium (-N(CH3 )3 + ) or phenyl groups in the guest's structure improves the strength of H-G interactions. The use of this technique is very straightforward, and it does not require any instrumental modifications. Thus, it can be applied to other H-G chemistry studies where comparison of bond dissociation energies is of paramount importance.

Investigation of Hemicryptophane Host-Guest Binding Energies Using High-Pressure Collision-Induced Dissociation in Combination with RRKM Modeling.

In advancing host-guest (H-G) chemistry, considerable effort has been spent to synthesize host molecules with specific and well-defined molecular recognition characteristics including selectivity and adjustable affinity. An important step in the process is the characterization of binding strengths of the H-G complexes that is typically performed in solution using NMR or fluorescence. Here, we present a mass spectrometry-based multimodal approach to obtain critical energies of dissociation for two hemicryptophane cages with three biologically relevant guest molecules. A combination of blackbody infrared radiative dissociation (BIRD) and high-pressure collision-induced dissociation (high-pressure CID), along with RRKM modeling, was employed for this purpose. For the two tested hemicryptophane hosts, the cage containing naphthyl linkages exhibited stronger interactions than the cage bearing phenyl linkages. For both cages, the order of guest stability is choline > acetylcholine > betaine. The information obtained by these types of mass spectrometric studies can provide new insight into the structural features that most influence the stability of H-G pairs, thereby providing guidance for future syntheses. Graphical Abstract.

Microscaffolds by Direct Laser Writing for Neurite Guidance Leading to Tailor-Made Neuronal Networks.

While modern day integrated electronic circuits are essentially designed in a 2D fashion, the brain can be regarded as a 3D circuit. The thus enhanced connectivity enables much more complex signal processing as compared to conventional 2D circuits. Recent technological advances in the development of nano/microscale 3D structuring have led to the development of artificial neuron culturing platforms, which surpass the possibilities of classical 2D cultures. In this work, in vitro culturing of neuronal networks is demonstrated by determining predefined pathways through topological and chemical neurite guiding. Tailor-made culturing substrates of microtowers and freestanding microtubes are fabricated using direct laser writing by two-photon polymerization. The first scaffold design that allows for site-specific cell attachment and directed outgrowth of single neurites along defined paths that can be arranged freely in all dimensions, to build neuronal networks with low cell density, is presented. The neurons cultured in the scaffolds show characteristic electrophysiological properties of vital cells after 10 d in vitro. The introduced scaffold design offers a promising concept for future complex neuronal network studies on defined neuronal circuits with tailor-made design specific neurite connections beyond 2D.

Sculpturing wafer-scale nanofluidic devices for DNA single molecule analysis.

We present micro- and nanofluidic devices with 3D structures and nanochannels with multiple depths for the analysis of single molecules of DNA. Interfacing the nanochannels with graded and 3D inlets allows the improvement of the flow and controls not only the translocation speed of the DNA but also its conformation inside the nanochannels. The complex, multilevel, multiscale fluidic circuits are patterned in a simple, two-minute imprinting step. The stamp, the key of the technology, is directly milled by focused ion beam, which allows patterning nanochannels with different cross sections and depths, together with 3D transient inlets, all at once. Having such a variety of structures integrated in the same sample allows studying, optimizing and directly comparing their effect on the DNA flow. Here, DNA translocation is studied in long (160 µm) and short (5-40 µm) nanochannels. We study the homogeneity of the stretched molecules in long, meander nanochannels made with this technology. In addition, we analyze the effect of the different types of inlet structures interfacing short nanochannels. We observe pre-stretching and an optimal flow, and no hairpin formation, when the inlets have gradually decreasing widths and depths. In contrast, when the nanochannels are faced with an abrupt transition, we observe clogging and hairpin formation. In addition, 3D inlets strongly decrease the DNA molecules' speed before they enter the nanochannels, and help capturing more DNA molecules. The robustness and versatility of this technology and DNA testing results evidence the potential of imprinted devices in biomedical applications as low cost, disposable lab-on-a-chip devices.

Culturing and patch clamping of Jurkat T cells and neurons on Al2O3 coated nanowire arrays of altered morphology.

Nanowire substrates play an increasingly important role for cell cultures as an approach for hybrid bio-semiconductor junctions. We investigate Jurkat T cells and neurons from mice cultured on Al2O3 coated ordered and randomly distributed nanowires. Cell viability was examined by life/membrane staining reporting comparable viability on planar and nanowire substrates. Imaging the hybrid interface reveals a wrapping of the cell membrane around the very nanowire tip. Patch clamp recordings show similar electrophysiological responses on each type of nanowires compared to planar control substrates. We demonstrate that the morphological characteristic of the nanowire substrate plays a subordinate role which opens up the arena for a large range of nanowire substrates in a functionalized application such as stimulation or sensing.

Correction: Culturing and patch clamping of Jurkat T cells and neurons on Al2O3 coated nanowire arrays of altered morphology.

[This corrects the article DOI: 10.1039/C8RA05320K.].

Low-energy collision-induced dissociation (low-energy CID), collision-induced dissociation (CID), and higher energy collision dissociation (HCD) mass spectrometry for structural elucidation of saccharides and clarification of their dissolution mechanism in DMAc/LiCl.

The dissolution mechanism of oligosaccharides in N,N-dimethylacetamide/lithium chloride (DMAc/LiCl), a solvent used for cellulose dissolution, and the capabilities of low-energy collision-induced dissociation (low-energy CID), collision-induced dissociation (CID), and higher energy collision dissociation (HCD) for structural analysis of carbohydrates were investigated. Comparing the spectra obtained using 3 techniques shows that, generally, when working with monolithiated sugars, CID spectra provide more structurally informative fragments, and glycosidic bond cleavage is the main pathway. However, when working with dilithiated sugars, HCD spectra can be more informative providing predominately cross-ring cleavage fragments. This is because HCD is a nonresonant activation technique, and it allows a higher amount of energy to be deposited in a short time, giving access to more endothermic decomposition pathways as well as consecutive fragmentations. The difference in preferred dissociation pathways of monolithiated and dilithiated sugars indicates that the presence of the second lithium strongly influences the relative rate constants for cross-ring cleavages vs glycosidic bond cleavages, and disfavors the latter. Regarding the dissolution mechanism of sugars in DMAc/LiCl, CID and HCD experiments on dilithiated and trilithiated sugars reveal that intensities of product ions containing 2 Li+ or 3 Li+ , respectively, are higher than those bearing only 1 Li+ . In addition, comparing the fragmentation spectra (both HCD and CID) of LiCl-adducted lithiated sugar and NaCl-adducted sodiated sugar shows that while, in the latter case, loss of NaCl is dominant, in the former case, loss of HCl occurs preferentially. The compiled evidence implies that there is a strong and direct interaction between lithium and the saccharide during the dissolution process in the DMAc/LiCl solvent system.

Grafting the sol-gel based sorbents by diazonium salts: a novel approach toward unbreakable capillary microextraction.

The present work deals with a novel approach for grafting a sol-gel based sorbent, using diazonium salts for preparation of an unbreakable capillary microextraction (CME) device in on-line combination with high performance liquid chromatography (HPLC). The use of diazonium salts modifier allowed all types of metallic and non-metallic substrates to be used without any limitation. Substrates including copper, brass, stainless steel and polytetrafluoroethylene (PTFE) were chosen to be functionalized by chemical or electrochemical reduction of 4-amino phenyl acetic acid. Then, 3-(trimethoxysilyl)propylamine (3TMSPA) was selected as the precursor and the only reagent for preparation of the desired surface chemical bonded sorbent. The presence of chemical bond between substrate, diazonium salts and 3TMSPA is more probably responsible for thermal and solvent stability and long lifetime of the prepared sorbent. Characterization of the aryl group formation on the various substrates along with the prepared sorbents was thoroughly investigated by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and thermogravimetry analysis (TGA). Typically, one of the prepared sorbents, deposited on the inner surface of the copper tube, was selected for assessing the developed method. The CME device was used for on-line extraction of atrazine, ametryn and terbutryn, as model compounds, from the aquatic media. After extraction, the HPLC mobile phase was used for on-line desorption and elution of the extracted analytes from the CME loop, containing the grafted sol-gel based sorbent, through the HPLC column. Figures of merit of the developed method were also obtained in which the linearity for the analytes was in the range of 30-1000µgL(-1). The value of LOD (S/N=3) for all analytes was 10µgL(-1) and the RSD% values (n=5) were all below 9.4% at the 500µgL(-1) level. Applicability of the developed method was examined by analyzing some real water samples in which the relative recovery percentage ranged from 75 to 95%.