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Delayed wound healing is one of the major diabetes complications that occur in 25% of diabetic patients. Specific wound management and combination treatment are required to repair the wound, which still remains a challenge with few effective therapies available currently. In this work, a new H 2 S donor PRO-F , which is characterized by the capability to promote wound healing in diabetes, was designed. PRO-F can be activated by light without consuming endogenous substances and the accompanying fluorescent signal makes the real-time tracking of released H 2 S possible. PRO-F is able to deliver H 2 S in an intracellular environment with moderate release efficiency (50%), which presents cytoprotective effects against excessive reactive oxygen species (ROS) induced damage. Furthermore, the potential of PRO-F to enhance chronic wound healing was validated by employing diabetic models. This work provides new insights into the therapeutic role of H 2 S donors in complex wound conditions, which should advance the pathophysiological research associated with H 2 S.

Early diagnosis of rheumatoid arthritis (RA) is crucial to prevent deterioration and improve the prognosis of disease outcome. However, current clinical diagnostic methods are unable to achieve accurate and early detection of RA. In this work, we designed an activatable organic nanoprobe ( ONP-CySe ) capable of specific and real-time imaging of ClO - in early RA. ONP-CySe comprises a near-infrared fluorescent selenomorpholine-caged cyanine dye as the sensing component and an amphiphilic triblock copolymer triphenyl phosphine derivative for mitochondria targeting. Our results showed that ONP-CySe successfully detected elevated levels of ClO - in the mitochondria of macrophages with high selectivity, low limit of detection (31.5 nM), excellent photostability, and good biocompatibility. Furthermore, ONP-CySe can also be used to monitor anti-inflammatory responses and efficacies of RA therapeutics, such as selenocysteine and methotrexate, in BALB/c mouse models. Therefore, our research proposes a universal molecular design strategy for the detection of ClO - , which holds potential for early diagnosis and drug screening for RA.

Peroxynitrite (ONOO - ), a kind of reactive oxygen species, plays an indispensable role in many physiological processes. The stability and reactivity of ONOO - are significantly affected by the pH of the environment. A novel fluorescent probe RN-NA that can simultaneously respond to ONOO - and pH was proposed and constructed based on a rational-designed multifunctional fluorescence resonance energy transfer (FRET) platform. The RN-NA probe exhibited a remarkably different fluorescence change in response to ONOO - and pH. The fluorescence signals at 525 and 710 nm increased about 4-fold with a pH change from 8.0 to 3.0. The changes in fluorescence at 525 nm are mainly attributed to photo-induced electron transfer, and the fluorescence enhancement at 710 nm was mainly due to acid-induced open-closed circulation. In the presence of ONOO - , the fluorescence at 525 nm increased 5-fold, while the fluorescence at 710 nm was almost completely diminished. Up to 70-fold fluorescence enhancement was observed in the ratiometric channel F 525 / F 710 . In the cell imaging experiment, the intracellular pH was adjusted using H + /K + ionophore and nigericin, and the endogenous ONOO - was generated by lipopolysaccharide (LPS) and γ-interferon (IFN-γ). The RN-NA probe can respond to cellular pH and endogenous ONOO - with remarkable fluorescence changes in both red/green and ratiometric channels.

Hydrogen sulfide (H 2 S) is a vital endogenous signal molecule that exerts critical physiological functions such as biological regulation and cytoprotection. Despite significant progress in developing H 2 S donors, site-specific delivery and controllable release of H 2 S in biological systems remain a key challenge. Herein, we develop new Cys-triggered fluorescent H 2 S donor Pro-S that is composed of a dicyanoisophorone-based near-infrared (NIR) fluorescent dye and a thiocarbamate moiety. The H 2 S donor releases H 2 S under the attack of Cys, accompanied by the release of a fluorescent reporter, which enables the real-time capturing of H 2 S by fluorescence spectroscopy or microscopy. Pro-S exhibits strong NIR fluorescence enhancement (70-fold), excellent controllable H 2 S release (30 min), high H 2 S release efficiency (62%), and well live-cell compatibility, allowing for visualization of H 2 S release in cells and zebrafish. Moreover, Pro-S presents a good effect of anti-inflammation in RAW 264.7 cells. This work provides a new idea for the design of H 2 S donors, which may be beneficial to the comprehension of the potential mechanism of inflammation and optimization of treatment strategies.

Multilayered and stacked cellulose paper has emerged as a promising platform for construction of three-dimensional (3D) cell culture because of its low cost, good biocompatibility, and high porosity. However, its poor light transmission makes it challenging to directly and clearly monitor cell behaviors (e.g., growth and proliferation) on the paper-based platform using an optical microscope. In this work, we developed a transparent microcrystalline cellulose/polyvinyl alcohol (MCC/PVA) paper with irregular pores through dissolution and regeneration of microcrystalline nanocellulose, addition of a porogen reagent (NaCl), and subsequently dipping in PVA solutions. The transparent MCC paper displays high porosity (up to 90%), adjustable pore size (between 23 and 46 μm), large thickness (from 315 to 436 μm), and high light transmission under water (>95%). Through further modification of the transparent MCC paper with PVA, the obtained transparent MCC/PVA paper shows enhanced mechanical properties (dry and wet strengths), good hydrophilicity (with a contact angle of 70.8°), and improved biocompatibility (cell viability up to 90%). By stacking and destacking multiple layers of the transparent MCC/PVA paper, it has been used for both two-dimensional and three-dimensional cell culture platforms. The transparent MCC/PVA paper under water enables both direct observation of cell morphology by an optical microscope via naked eyes and fluorescence microscope after staining. We envision that the developed transparent MCC/PVA paper holds great potential for future applications in various bioanalytical and biomedical fields, such as drug screening, tissue engineering, and organ-on-chips.

Lignocellulosic biomass is mainly composed of polysaccharides and lignin. The complexity and diversity of the plant cell wall polymers makes it difficult to isolate the components in pure form for characterization. Many current approaches to analyzing the structure of lignocellulose, which involve sequential extraction and characterization of the resulting fractions, are time-consuming and labor-intensive. The present study describes a new and facile system for rationally derivatizing and dissolving coarsely ground plant cell wall materials. Using ionic liquids (EmimAc) and dichloroacetyl chloride as a solvent/reagent produced mildly acetylated whole cell walls without significant degradation. The acetylated products were soluble in DMSO- d 6 from which they can be characterized by solution-state two-dimensional nuclear magnetic resonance (2D NMR) spectrometry. A distinct advantage of the procedure is that it realizes the dissolution of whole lignocellulosic materials without requiring harsh ball milling, thereby allowing the acquisition of high-resolution 2D NMR spectra to revealing structural details of the main components (lignin and polysaccharides). The method is therefore beneficial to understanding the composition and structure of biomass aimed at its improved utilization.

Small-molecular fluorescence sensors have become promising detection tools in many fields attributing to their high sensitivity, excellent temporal and spatial resolution, and low cytotoxicity. However, high concentration or aggregation-induced fluorescence quenching effect has usually hindered the development of traditional fluorescence dyes. Herein, a new fluorophore cyanovinylene dye BMZ with excimer emission property has been constructed. It shows an obvious enhanced and red-shift emission upon aggregation in aqueous solution, which overmatches the conventional pyrene with longer absorption and emission wavelengths. Using this unique optical property, a new fluorescence probe BMZ-Gal has been developed for trapping of β-galactosidase (β-Gal) activity with high selectivity, low limit of detection of 0.17 U, and rapid recognition, which is based on the β-Gal-induced formation of red-shift emitting excimer. β-Gal has a strong affinity for BMZ-Gal , which is verified through the Michaelis-Menten constants ( K m , 1.87 μM) and the hydrolysis efficiencies ( K cat / K m , 1.78 × 10 3 M -1 s -1 ). Furthermore, the assay system has been successfully used for detecting endogenous β-Gal in living ovarian cancer cells and can passively targeted to identify β-Gal in organelle level and determine its subcellular location with satisfactory accuracy. We anticipate that the new fluorophore cyanovinylene dye herein may inaugurate new opportunities for the development of excimer emission sensors.

Single-cell phenotypic profiling of circulating tumor cells (CTCs) in the blood of cancer patients can reveal vital tumor biology information. Even though various approaches have been provided to enrich and detect CTCs, it remains challenging for consecutive CTC sorting, enumeration, and single-cell characterizations. Here, we report an integrated microfluidic device (IMD) for single-cell phenotypic profiling of CTCs that enables automated CTCs sorting from whole blood following continuous single-cell phenotypic analysis while satisfying the requirements of both high purity (92 ± 3%) of cell sorting and high-throughput processing capacity (5 mL whole blood/3 h). Using this new technique we test the phenotypes of individual CTCs collected from xenograft tumor-bearing mice and colorectal (CRC) patients at different tumor stages. We obtained a correlation between CTC characterization and clinical tumor stage and treatment response. The developed IMD offers a high-throughput, convenient, and rapid strategy to study individual CTCs toward minimally invasive cancer therapy prediction and disease monitoring and has the potential to be translated to clinic for liquid biopsy.

A smartphone-powered medical dongle as a miniaturized electrochemical analyzer associated with an enzymatic β-hydroxybutyrate test strip for accurate characterization of blood ketone in peripheral whole blood at the point-of-care, which is capable of providing critical guidance for the evaluation and treatment of diabetic ketoacidosis (DK) and diabetic ketosis acid (DKA), is reported. The measured results of blood ketone by the medical dongle were compared with the clinical results from a bulky biochemical analyzer, and the analysis showed good agreement. The proposed medical smartphone-powered dongle was demonstrated to be a very promising platform as a miniaturized electrochemical analyzer for point-of-care monitoring of the critical biochemical parameters such as blood ketone and a good solution for mobile health management.

(13)C-Metabolic Flux Analysis ((13)C-MFA) is rapidly being recognized as the authoritative method for determining fluxes through metabolic networks. Site-specific (13)C enrichment information obtained using NMR spectroscopy is a valuable input for (13)C-MFA experiments. Chemical shift overlaps in the 1D or 2D NMR experiments typically used for (13)C-MFA frequently hinder assignment and quantitation of site-specific (13)C enrichment. Here we propose the use of a 3D TOCSY-HSQC experiment for (13)C-MFA. We employ Non-Uniform Sampling (NUS) to reduce the acquisition time of the experiment to a few hours, making it practical for use in (13)C-MFA experiments. Our data show that the NUS experiment is linear and quantitative. Identification of metabolites in complex mixtures, such as a biomass hydrolysate, is simplified by virtue of the (13)C chemical shift obtained in the experiment. In addition, the experiment reports (13)C-labeling information that reveals the position specific labeling of subsets of isotopomers. The information provided by this technique will enable more accurate estimation of metabolic fluxes in large metabolic networks.

This paper reports on a novel method for the rapid determination of the shale porosity by double headspace extraction gas chromatography (DHE-GC). Ground core samples of shale were placed into headspace vials and DHE-GC measurements of released methane gas were performed at a given time interval. A linear correlation between shale porosity and the ratio of consecutive GC signals was established both theoretically and experimentally by comparing with the results from the standard helium pycnometry method. The results showed that (a) the porosity of ground core samples of shale can be measured within 30 min; (b) the new method is not significantly affected by particle size of the sample; (c) the uncertainties of measured porosities of nine shale samples by the present method range from 0.31 to 0.46 p.u.; and (d) the results obtained by the DHE-GC method are in a good agreement with those from the standard helium pycnometry method. In short, the new DHE-GC method is simple, rapid, and accurate, making it a valuable tool for shale gas-related research and applications.

We developed and validated a new paper-based assay for the detection of human blood type. Our method involves spotting a 3 μL blood sample on a paper surface where grouping antibodies have already been introduced. A thin film chromatograph tank was used to chromatographically elute the blood spot with 0.9% NaCl buffer for 10 min by capillary absorption. Agglutinated red blood cells (RBCs) were fixed on the paper substrate, resulting in a high optical density of the spot, with no visual trace in the buffer wicking path. Conversely, nonagglutinated RBCs could easily be eluted by the buffer and had low optical density of the spot and clearly visible trace of RBCs in the buffer wicking path. Different paper substrates had comparable ability to fix agglutinated blood, while a more porous substrate like Kleenex paper had enhanced ability to elute nonagglutinated blood. Using optimized conditions, a rapid assay for detection of blood groups was developed by spotting blood to antibodies absorbed to paper and eluted with 200 μL of 0.9% NaCl buffer directly by pipetting. RBCs fixation on paper accurately detected blood groups (ABO and RhD) using ascending buffer for 10 min or using a rapid elution step in 100/100 blood samples including 4 weak AB and 4 weak RhD samples. The assay has excellent reproducibility where the same blood group was obtained in 26 samples assessed in 2 different days. Agglutinated blood fixation on porous paper substrate provides a new, simple, and sensitive assay for rapid detection of blood group for point-of-care applications., (© 2011 American Chemical Society)

Agglutinated blood transports differently onto paper than stable blood with well dispersed red cells. This difference was investigated to develop instantaneous blood typing tests using specific antibody-antigen interactions to trigger blood agglutination. Two series of experiments were performed. The first related the level of agglutination and the fluidic properties of blood on its transport in paper. Blood samples were mixed at different ratios with specific and nonspecific antibodies; a droplet of each mixture was deposited onto a filter paper strip, and the kinetics of wicking and red cell separation were measured. Agglutinated blood phase separated, with the red blood cells (RBC) forming a distinct spot upon contact with paper while the plasma wicked; in contrast, stable blood suspensions wicked uniformly. The second study analyzed the wicking and the chromatographic separation of droplets of blood deposited onto paper strips pretreated with specific and nonspecific antibodies. Drastic differences in transport occurred. Blood agglutinated by interaction with one of its specific antibodies phase separated, causing a chromatographic separation. The red cells wicked very little while the plasma wicked at a faster rate than the original blood sample. Blood agglutination and wicking in paper followed the concepts of colloids chemistry. The immunoglobin M antibodies agglutinated the red blood cells by polymer bridging, upon selective adsorption on the specific antigen at their surface. The transport kinetics was viscosity controlled, with the viscosity of red cells drastically increasing upon blood agglutination. Three arm prototypes were investigated for single-step blood typing.

Paper-based microfluidic patterns have been demonstrated in recent literature to have a significant potential in developing low-cost analytical devices for telemedicine and general health monitoring. This study reports a new method for making microfluidic patterns on a paper surface using plasma treatment. Paper was first hydrophobized and then treated using plasma in conjunction with a mask. This formed well defined hydrophilic channels on the paper. Paper-based microfluidic systems produced in this way retained the flexibility of paper and a variety of patterns could be formed. A major advantage of this system is that simple functional elements such as switches and filters can be built into the patterns. Examples of these elements are given in this study.

This paper demonstrated an in situ headspace gas chromatography (GC) technique for monitoring bacterial growth using a commercial GC system with a multiple headspace extraction (sampling) mode. The technique was based on measuring the carbon dioxide mass in the headspace of a closed sample vial during the bacteria growth. A mathematic equation was derived in order to calculate the integrated amount of carbon dioxide produced by bacterial growth during the incubation. The method can be used to monitor the bacterial growth rate in a given cultural medium. The present method is very simple, sensitive, and safe.

Photoacoustic rapid-scan FT-IR spectroscopy was used to collect spectra of paper samples printed with mineral- and vegetable-oil-based inks at different concentrations. Partial least-squares (PLS) analysis and principal component analysis (PCA) were combined to form a model that, with data collected in the 3600-3200, 3000-2800, and 1800-1000 cm-1 spectral regions, enables one to predict the concentration of ink in printed samples. Prediction is statistically robust provided the selected optical path difference (OPD) velocities are within the range 0.05-1.00 cm/s.

Pinus radiata kraft pulps with varying carboxyl content were studied using Fourier transform infrared photoacoustic spectroscopy (FT-IR-PAS). The examined pulp samples, with Kappa number ranging from 20.8 to 128, originated from pulping experiments conducted in flow-through reactors utilizing varying effective alkali, temperature, and cooking time. A partial least-squares (PLS) analysis was used to formulate a model that correlates the spectral data with the carboxyl content of pulp. Using three principal components, the resultant PLS model could explain approximately 98.5% of the variance in the X-matrix (spectral features) and 96.8% of the variance in the Y-matrix (measured carboxyl content). The FT-IR-PAS technique in combination with PLS analysis predicts the carboxyl content of the pulps with a high degree of accuracy. This method is much faster than the conventional titration methods and also not destructive to the pulp sample.

In the present study, hardwood and softwood pulps were characterized by employing Fourier transform infrared photoacoustic spectroscopy (FT-IR-PAS). The pulp samples examined originated from Swedish sulfite and kraft pulp mills, which utilize different cooking processes and modern bleaching technologies. Partial least-squares (PLS) analysis was used to correlate the spectral data obtained with the kappa (K) numbers and carbohydrate compositions of the pulp samples determined by enzymatic hydrolysis and subsequent capillary zone electrophoresis. Using four principal components, the present PLS model based on photoacoustic FT-IR spectra could explain 85% of the variance in the X matrix and 81% of the variance in the Y matrix. The FT-IR-PAS technique in combination with PLS was found to accurately predict the contents of carbohydrates, i.e., xylose, glucose, mannose, arabinose, galactose, and hexenuronic acid residues, as well as the content of lignin measured in terms of K numbers and corrected K numbers of the pulps. From these predictions, the contents of xylan, glucomannan, and cellulose can also be predicted. The content of 4-O-methylglucuronic acid residues is, however, more difficult to predict accurately, using this approach.

Detailed quantitative analysis of lignin monomer composition comprising p-coumaryl, coniferyl, and sinapyl alcohol and p-coumaraldehyde, coniferaldehyde, and sinapaldehyde in plant has not been studied from every point mainly because of artifact formation during the lignin isolation procedure, partial loss of the lignin components inherent in the chemical degradative methods, and difficulty in the explanation of the complex spectra generally observed for the lignin components. Here we propose a new method to quantify lignin monomer composition in detail by pyrolysis-gas chromatography (Py-GC) using acetylated lignin samples. The lignin acetylation procedure would contribute to prevent secondary formation of cinnamaldehydes from the corresponding alcohol forms during pyrolysis, which are otherwise unavoidable in conventional Py-GC process to some extent. On the basis of the characteristic peaks on the pyrograms of the acetylated sample, lignin monomer compositions in various dehydrogenative polymers (DHP) as lignin model compounds were determined, taking even minor components such as cinnamaldehydes into consideration. The observed compositions by Py-GC were in good agreement with the supplied lignin monomer contents on DHP synthesis. The new Py-GC method combined with sample preacetylation allowed us an accurate quantitative analysis of detailed lignin monomer composition using a microgram order of extractive-free plant samples.

Spectra of highly acidic oligosaccharides obtained by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) can be greatly enhanced in quality by coating the MALDI probe surface with a film consisting of a perfluorosulfonated ionomer (Nafion) prior to the addition of the sample-matrix mixture. For comparison, three mixtures containing highly acidic oligo- and polysaccharides derived from partial acidic hydrolysis of alginate, pectin, or carboxymethyl cellulose (CMC) were analyzed by employing probes with an uncoated gold surface or a surface coated with a Nafion or nitrocellulose film. The negative ion-mode MALDI spectra of the oligouronates (oligomers containing mannuronic/guluronic and galacturonic acid residues) obtained using uncoated or nitrocellulose-coated probes consisted of a series of broad, multiple peaks. These multiple peaks were assigned to the molecular ions of the nondissociated [M - H]- and partially sodiated [MnNa - H]-, where n = 1, 2, or 3, oligomers. In contrast, the corresponding spectra obtained with Nafion-coated probes contained only a single series of sharp peaks originating from the molecular ions ([M - H]-) of nondissociated oligomers exhibiting chain lengths of as many as approximately 15 uronic acid residues. The Nafion coating was apparently capable of removing the sodium counterions remaining in the deposit of the sample-matrix mixture on the probe, thereby greatly enhancing the signal-to-noise ratios of the peaks in the spectra. In a similar manner, higher quality spectra could also be obtained by using Nafion-coated probes for analysis of the oligouronates and CMC oligomers by positive ion-mode MALDI-MS.

A new chromatographic method for the determination of oxidants, such as peroxyborates and peroxides, present in a pulp matrix has been developed. The new method is characterized by its high reproducibility, its low limit of detection, and its high selectivity. Thioanisole (methylphenyl sulfide) is shown to selectively and quantitatively react with oxidants that are present within a pulp matrix. The experimental protocol proposed requires an HPLC system with a normal-phase column. The method allows for the quantitative monitoring of the thioanisole starting material, as well as the products of oxidation, methylphenyl sulfoxide and methylphenyl sulfone. After a 2-day reaction period, the analysis time for a sample is less than 20 min. The detection limit is 2.7 × 10(-6) M for the sulfoxide and sulfone and less than 5.0 × 10(-7) M for the thioanisole. This novel approach for monitoring oxidants present on solid lignocellulosic matrixes may provide pulp and paper manufacturers with a new tool for the study of the long-term bleaching effectiveness of peroxy-containing chemical additives.