HEPES-buffered Solution Research Articles

Design, Synthesis and Crystal Structure of a Novel Fluorescence Probe for Zn2+ Based on Pyrano[3,2-c] Carbazole.

Zinc is a trace element, which plays an important role in many biological processes. The deficiency of zinc will lead to many diseases. Thus, it is of great significance to develop fast and efficient quantitative detection technology for zinc ions. In this study, a novel fluorescence probe FP2 was designed for Zn2+ quantification based on pyrano[3,2-c] carbazole. The structure of FP2 was characterized by 1HNMR, 13CNMR, HRMS, and X-ray diffraction. In the HEPES buffer solution, FP2 is responsive to Zn2+ and greatly enhanced. The pH value and reaction time were investigated, and the optimum reaction conditions were determined as follows: the pH was 7~9 and the reaction time was longer than 24 min. Under the optimized conditions, the concentration of FP2 and Zn2+ showed a good linear relationship in the range of 0~10 μM, and the LOD was 0.0065 μmol/L. In addition, through the 1H NMR titration experiment, density functional theory calculation, and the job plot of FP2 with Zn2+ in the HEPES buffer solution, the binding mode of FP2 and Zn2+ was explained. Finally, the method of flame atomic absorption spectrometry (FAAS) and FP2 were used to detect the content of Zn2+ in the water extract of tea. The results showed that the FP2 method is more accurate than the FAAS method, which shows that the method described in this work could be used to detect the content of Zn2+ in practical samples and verify the practicability of this method.

Sensitive detection of α-amylase based on host–guest inclusion system of γ-cyclodextrin and dansyl-derived diphenylalanine

A highly sensitive detection system for α-amylase was developed via host–guest complexation between γ-cyclodextrin and dansyl-modified diphenylalanine (FF-Dns). The host–guest inclusion of FF-Dns into the cavity of γ-CD in a HEPES buffer solution (10 mM, pH 7.4) significantly enhanced the fluorescence intensity, and the emission wavelength gradually shifted from 558 to 535 nm. The hydrolysis of γ-CD by the addition of α-amylase released FF-Dns, leading to the recovery of the fluorescence emission characteristics. Therefore, the FF-Dns/γ-CD host–guest complexation system can serve as a platform for the sensitive detection of α-amylase with good selectivity against potential interference. The limit of detection (LOD) of the system was 0.004 U/mL, with a linear working range of 0–6 U/mL. The detection assay was successfully applied in 0.1 % serum, achieving an LOD of 0.017 U/mL and a linear working range of 0–10 U/mL.

Synthesis, structural characterization and antitrypanosomal activity of ferrocenyl-based imines with (2 or 4)-nitrophenyl-2-furan group

Two new metallocene complexes with formulation [Fe(η5-C5H5){(η5-C5H4)-NCH-2-C4H2O-(5-C6H4-2-NO2)}] (3) and [Fe(η5-C5H5){(η5-C5H4)-NCH-2-C4H2O-(5-C6H4-4-NO2)}] (4) have been synthesized and their antitrypanosomal activity has been evaluated. Complexes 3 and 4 were obtained by condensation reaction of ferrocenylamine (1) with respective 5-(2-nitrophenyl)-2-furancarboxaldehyde (2a) or 5-(4-nitrophenyl)-2-furancarboxaldehyde (2b). The complexes were fully characterized by spectroscopic techniques (FT-IR, 1H and 13C{1H} NMR, and HRMS), which confirm their correct obtainment. The crystal structures of 3 and 4 were also determined by single crystal X-ray diffraction. Furthermore, UV–visible studies revealed that compounds 3 and 4 exhibited suitable stability in DMSO:HEPES buffer solution (80:20) throughout 24 h. The in vitro antiparasitic activities of ferrocenyl imine derivatives 3 and 4 were assessed against two species of parasites, Trypanosoma cruzi (T. cruzi) and Trypanosoma brucei (T. brucei). The obtained results revealed that compound 3 (EC50 = 0.44 μM) exhibited nearly a 40-fold greater efficacy as an anti-T. brucei agent compared to derivative 4 (EC50 = 16.0 μM). Moreover, compound 3 demonstrated superior potency when compared with its organometallic analogs (EC50 = 2.42–13.3 μM), which were previously reported by our research group. Interestingly, the EC50 value of 3 was found to be 8 times greater than that of nifurtimox (EC50 = 3.56 μM). The cytotoxicity of the ferrocenyl imines was also evaluated on the L6 rat skeletal myoblast cell line.

Developing new Hydrazine Carbothioamide Molecules for Selective Colorimetric/Fluorometric Detection of Environmentally Essential Hg2+ and Ag+ Metal Ions in Mixed Aqueous Media.

A novel colorimetric and fluorogenic probe L based on hydrazine carbothioamide and 1,8-naphthalimide moieties has been designed and synthesized for the hypersensitive detection of Hg2+ or Ag+ ions. The observed probe L showed colorimetric and fluorometric responses for these studies when Hg2+ or Ag+ was added to the DMSO - HEPES buffer solution (pH = 7). An interference test with other metal ions was determined, and the high selectivity of Hg2+ and Ag+ did not interfere with other metal ions in colorimetric and fluorogenic methods. The possible mechanism of binding of these metal ions and the probe L 1:1 complex was determined by H1 NMR. Additionally, the reversibility of the affinity of probe L with mercury (Hg2+) and silver (Ag+) ions was investigated by adding Na2EDTA. The naked eye detected the "Off-On" type fluorescence sensor in the presence of Hg2+ and EDTA. The tested test strip kits provided a strong probability of probe L with high response and rapid, sensitive detection with Hg2+ ion, which may be suitable for practical use.

Investigating the role of local acidosis in hypothalamic vasopressin cell excitation during hypoxia

The supraoptic nucleus (SON) of the hypothalamus plays a key role in the regulation of fluid homeostasis through the synthesis and release of vasopressin (VP). In this population, VP release can occur at both dendritic and axonal sites, allowing for both localized and distal effects such as renal water retention and vasoconstriction. Recent work from our laboratory demonstrated an inverse neurovascular coupling (iNVC) response in which VP cell activation by acute salt loading resulted in dendritic release of VP, leading to localized vasoconstriction and hypoxia. Additionally, we demonstrated that the hypoxic milieu in turn potentiates VP cell activity and increases firing, suggesting that these mechanism constitutes a positive feedback loop that optimized VP activation in order to cope with the homeostatic challenge (Roy et al., Cell Reports 2021). Still, the biophysical mechanism by which the iNVC-hypoxia elicits increased firing in VP neurons has not yet been explored. We hypothesized that hypoxia-induced acidosis, via activation of acid sensitive ion channels (ASICs) (which are present in SON VP neurons; Ohbuchi et al in 2010) is a key mechanism contributing to VP cell excitation in the iNVC positive feedback loop. We investigated this with whole cell patch clamp electrophysiology using eGFP-VP transgenic rats. We recorded from identified SON VP cells and measured deflections in voltage as bath pH was changed from 7.25 to 6.3 (10 mins). Bath solution was either HCL-buffered artificial cerebrospinal fluid (aCSF) aerated with 95% O2/5% CO2 or HEPES-buffered solution (HBS). We observed an excitatory response to the decreases in pH in most SON VP cells recorded (68.75%, n = 11/16). These responses were reversible and, in many cases, repeatable following a second stimulation. To examine cell-type specificity, ongoing studies are focusing on comparing these responses to those obtained from oxytocin cells. Further, we plan to repeat these experiments in the presence of the ASIC-blocker amiloride and determine whether the hypoxia-induced excitatory effect during iNVC is indeed dependent on ASIC-channels. Together, our results support our hypothesis that tissue acidosis (likely via activation of ASIC-channels) contributes to the excitatory feedback loop evoked during salt-induced iNVC in VP neurons. This abstract was supported by the National Heart Lung and Blood Institute of the National Institutes of Health [R01 HL162575-01 (J.E.S.)]. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Compound 48/80 causes overactive bladder without altering urothelial permeability in mice

The urothelium, a transitional epithelium lining the urinary bladder, is essential for preventing leakage of urine into the underlying tissue. Disruption of this barrier by noxious stimuli or inflammatory mediators increases bladder wall permeability and causes bladder dysfunction. Compound 48/80 is a pro-inflammatory basic secretagogue that causes urothelium-dependent increases in urinary bladder contractility, as well as bladder overactivity in mice. We hypothesized that Compound 48/80 causes bladder dysfunction by increasing urothelial permeability. Intact bladders from male C57BL/6 mice (N=4) were cannulated, filled with HEPES-buffered physiological salt solution containing FITC-labeled-dextran (70 kDa), and pressurized to 25 mmHg. Z-stacks of baseline fluorescence were captured across the bladder wall using 2-photon laser microscopy. Compound 48/80 (50 μg/mL) was then instilled intravesically for 10 minutes before the bladder wall was imaged again. Average FITC intensity plots wall showed no consistent increase in fluorescence with or without Compound 48/80 from the lumen into the bladder wall. There was also no statistical difference in the area under the curve of these line graphs (p=0.5898). Our findings suggest that Compound 48/80 causes overactive bladder without affecting urothelial permeability. Future studies will investigate other secretagogues and lower molecular weight dextrans to clarify the mechanisms by which secretagogues cause a loss of urothelial barrier function. This research is supported by NIH P20-DK127554 (NRT), NIH R01-DK119615 (NRT). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

A novel fluorescent and colorimetric dual-signals peptide-based probe for sequential recognition of Cu(II) and cimetidine and its multi-functional applications

A novel dual-signals peptide-based probe FDKSH was designed and synthesized based on 5-carboxy fluorescein (5-FAM) labelled tetrapeptide backbone (Asp-Lys-Ser-His-NH2). FDKSH exhibited a highly selective fluorescence “on-off” response and obvious colorimetric recognition to Cu2+ in 10 mM HEPES buffer solution at pH 7.4. Meanwhile, the limit of detection (LOD) was found to be 62.1 nM, which was much lower than that of stated by World Health Organization (WHO) in drinking water (30.0 μM). As a new promising cascade probe, the Cu(II)-assisted peptide ensemble probe (FDKSH-Cu2+ ensemble) has very high selectivity for cimetidine by the displacement method with LOD of 63.5 nM. In addition, FDKSH was successfully used to determine Cu2+ and cimetidine in two actual water samples. Additionally, FDKSH was also successfully used for monitoring Cu2+ and cimetidine in living cells and zebrafish, and showed low toxicity and excellent fluorescence imaging performance. It is worth noting that FDKSH-impregnated test strips exhibited significant fluorescence and colorimetric color changes for Cu2+ and cimetidine directly observed by naked eye. What’s more, smartphone with RGB color recognition function provided a new method for semi-quantitative visual detection of Cu2+ and cimetidine by analyzing different concentrations of Cu2+ and cimetidine without expensive instruments.

Evaluation of Effect of Vitamin C on Streptococcus Mutans Induced Secondary Caries: An In vitro Study

This study aimed to assess and understand the protective effect of vitamin C against secondary caries formation around the margins of class V restorations with the help ofan in-vitro bacterial caries model. In this study, a total of 30 mandibular premolars were chosen and assigned to three groups (n=10). In this study, Class V composite restorations were prepared and along with pretreatment of the cavity walls with various solutions like Vitamin C (ascorbic acid solution), HEPES buffer solution and chlorhexidinedigluconate. After incubating specimens with Streptococcus mutans, dentin and enamel were assessed using Fluorescence Microscopy.The study showed Vitamin C (Ascorbic acid) inhibited the formation of secondary caries near the dentin resin interface. No inhibitory effects were observed in enamel margins. From the study, it could be concluded that the incorporation of ascorbic acid in the restorative procedure of resin composite may inhibit secondary caries formation. Neither the application of chlorhexidine nor HEPES buffer had much effect on the inhibitory effect on secondary caries.

Structural studies of Schiff base ligand and its copper complexes: Solvents effect in 1-D polymeric and monomeric copper (II) complexes, computational and sensing studies

The Schiff base ligand (E)-2,4-dibromo-6-(((2-(methylthio)phenyl)imino)methyl)phenol (SL5) and their copper complexes were synthesized. The complex-1 ([Cu (SL5) (CH3COO)]n) and complex-2 ([Cu (SL5) (CH3COO) (CH3OH)]) were obtained by inducing acetic and methanol solvent by post-synthetic modification. Both ligand and metal complexes were characterized by FTIR, SEM, and EDAX analysis. In addition, the photoluminescence behaviour has also been studied. The crystal structures of both ligand and metal complexes were determined by single-crystal X-ray diffraction studies. The structure was solved by direct methods using the SHELXS program and refined by the SHELXL program. The structural changes of ligand in the formation of the metal complexes and the solvent-induced effect of acetic acid and methanol on copper complexes have been studied using crystal structure analysis. The SL5 ligand and complex-2 crystallize in the Monoclinic, P21/n space group and complex-1 crystallize in the Tetragonal, I41/a space group. Complex-1 exhibits six coordinated distorted octahedral geometry. The addition of methanol solvent to complex-1 forms complex-2 and has five coordinated square pyramidal geometry. Complex-1 forms a 1-D coordination polymer chain via Cu1–O3 and it is replaced by the O2–H2‧‧‧O4 interaction with the aid of coordinated methanol solvent in complex-2. The effect of solvents on the coordination sphere directly influences the chelating rings. Complexes-1 exhibits interesting type-II halogen‧‧‧halogen (Br‧‧‧Br) interaction, leading to the formation of tetramer synthons (X4). Halogen-halogen interactions are broken down by adding a methanol solvent, resulting in halogen-hydrogen interactions in complex-2. Crystal structure and quantum computational studies have analyzed the synergetic effect of ligand and solvents on the molecular packing of metal complexes. The Cu(OAc)2SL5 receptor selectively detects sulfide anion in aqueous acetonitrile HEPES buffer solution in the presence of other anions. Further, antibacterial activity has been carried out for MRSA bacteria, and docking studies were carried out for penicillin-binding protein to explore the effect of coordination and solvents.

Clofazimine pKa Determination by Potentiometry and Spectrophotometry: Reverse Cosolvent Dependence as an Indicator of the Presence of Dimers in Aqueous Solutions.

The weakly basic antibiotic and anti-inflammatory drug, clofazimine (CFZ), was first described in 1957. It has been used therapeutically, most notably in the treatment of leprosy. However, the compound is extremely insoluble in aqueous media, and, indeed, there is poor consensus about what its intrinsic solubility is since the reported values range from 0.04 to 11 ng/mL. To understand the speciation and solubilization of CFZ as a function of pH, it is of paramount importance to know the true aqueous pKa. However, there is also poor consensus about the value of the pKa (reported measured values range from 6.08 to 9.11). In the present study, we report the determination of the CFZ ionization constant using two independent techniques. A state-of-the-art potentiometric analysis was performed, drawing on titration data in methanol-water solutions (46-75 wt % MeOH) of CFZ, using the bias-reducing consensus of two different procedures of extrapolating the apparent psKa values to zero cosolvent to approximate the true aqueous pKa as 9.43 ± 0.12 (25 °C, I = 0.15 M reference ionic strength). In parallel, spectrophotometric UV/vis titration data were acquired (250-600 nm at different pH) in 10 mM HEPES buffer solutions containing up to 54 wt % MeOH. The alternating least squares (ALS) method was used in the analysis of the absorbance-pH spectra. Uncharacteristically, the cosolvent UV/vis data in our study showed reverse cosolvent dependence (apparent pKa values increased with increasing cosolvent) which could be explained by a dimerization of the free base. The analysis of UV/vis data obtained from 54 wt % MeOH-water solution containing 20 μM CFZ yielded the apparent pKa 9.51 ± 0.17 (I ≈ 0.005 M). To assess whether self-assembly of CFZ was energetically feasible, density functional theory (DFT) calculations were used to study the putative CFZ dimers in aqueous and methanol media. The DFT-optimized geometries and infrared spectra of CFZ dimers using water and methanol as solvents were calculated and analyzed. Based on the lack of negative frequencies in calculated infrared spectra, it was confirmed that optimized geometries correspond to the true energetic minima. Visual analysis of optimized structures indicates the presence of stacking interactions between two CFZ molecules. The protonation site (the imine nitrogen atom) was determined by 1H NMR spectroscopy.

A simple paper-based nickel nanocluster-europium mixed ratio fluorescent probe for rapid visual sensing of tetracyclines

In this work, a ratiometric fluorometric sensor based on nickel nanoclusters (NiNCs)-europium complex (NiNCs-Eu3+) was constructed for the highly selectivity detection of tetracyclines (TCs) in water samples. In the presence of TCs, the blue fluorescence of the sensor NiNCs-Eu3+ was quenched at 430 nm and the characteristic red fluorescence of Eu3+-TCs appeared at 620 nm because of the combined help of inner filter effect (IFE) and antenna effect. Under the optimized conditions (100 mM Eu3+ (100 µL); temperature (25℃); reaction time (10 min), HEPES buffer solution (pH = 7.0)), the sensor offered a wide detection range of tetracycline (TC) and oxytetracycline (OTC) from 0.1 to 50 μM with the detection limit (LOD) of 25 nM and 21 nM, respectively. Moreover, the sensor was able to detect of TC and OTC in tap and lake water with high recovery rate (89.10%-97.60%). In addition, the portable paper-based sensor was constructed using filter paper embedded with NiNCs-Eu3+. The distinct fluorescent color of the paper-based sensor varied from bright blue to red against different concentrations of TC and OTC. These above findings demonstrated the potential for wide application of as-prepared ratio metric fluorescence sensor for visual detection of TCs in water samples.

Synthesis of Vanillin-Azine as Colorimetric Chemosensor of Sulfide Anion

Vanillin-azine (VA), (4,4'-((1E,1'E)-hydrazine-1,2-diylidenebis(methane-lylidene))bis(2-methoxyphenol) has been synthesized from vanillin and tested as anion colorimetric chemosensors for sulfide anion. The VA was obtained from a condensation reaction between vanillin and hydrazine hydrate with a mol ratio of 2:1 mol for 24 h at room temperature. The structure was elucidated using FTIR, GC-MS, 1H-NMR, and 13C-NMR spectrometers. The VA compound was examined as a colorimetric chemosensor for sulfide anion over several anions of CN−, F−, Cl−, Br−, I−, N3−, CH3COO−, and NO3−. The structure of the product showed agreement with all spectrometric data. The VA chemosensor tests indicated only selective to S2− anion followed by a color change from colorless to light blue in a DMF:HEPES buffer solution (DMF:HBS) medium (9:1, v/v, 10 mM, pH = 7.4). Filter paper strips can detect S2− anion with a color change from white to yellow. The VA chemosensor has a limit of detection (LOD) of 5.4 × 10−4 M, therefore, the VA chemosensor can be applied to detect S2− anion in tap water.

Impact of graft architecture of PEGylated copolymers assembly on hydroxyapatite in the differential regulation of initial cell and bacterial adhesion

Synthetic pure hydroxyapatite (HA), though widely investigated as an artificial orthopedic implant material, has neither anti-infection properties nor innate interfacial bioactivity. Ideally, surface modification of this material needs to be developed with simultaneous anti-microbial capacity and superior cytocompatibility. Herein, graft copolymers of a PAA backbone functionalized with a high density of polyethylene glycol (PEG), and varying grafting densities of cyclic Arg-Gly-Asp-d-Phe-Cys peptides (cRGD) end-grafted onto the PEG chains have been designed and synthesized. Systematic study on the grafted anchoring groups first indicates a synergistic binding mechanism of the copolymer to HA surface, involving electrostatic interaction of the protonated amino (–NH3+) with negatively charged HA, assisted with coordination bonding through the nitrodopamine (ND) group. The PEGylated graft copolymers are demonstrated to firmly adsorb to the HA surface as an ultrathin self-assembly layer from a HEPES buffer solution, showing stability with insignificant desorption and effective inhibition of non-specific protein adsorption evaluated by XPS and QCM-D assays. Direct adhesion assays confirm that HA, modified with the density-optimized cRGD/PEG-functionalized graft polymer, can considerably promote osteoblast attachment, while preventing different strains of bacteria adhesion in the initial phase.

Reversible Self-Assembled Monolayers with Tunable Surface Dynamics for Controlling Cell Adhesion Behavior.

Cells adhering onto surfaces sense and respond to chemical and physical surface features. The control over cell adhesion behavior influences cell migration, proliferation, and differentiation, which are important considerations in biomaterial design for cell culture, tissue engineering, and regenerative medicine. Here, we report on a supramolecular-based approach to prepare reversible self-assembled monolayers (rSAMs) with tunable lateral mobility and dynamic control over surface composition to regulate cell adhesion behavior. These layers were prepared by incubating oxoacid-terminated thiol SAMs on gold in a pH 8 HEPES buffer solution containing different mole fractions of ω-(ethylene glycol)2-4- and ω-(GRGDS)-, α-benzamidino bolaamphiphiles. Cell shape and morphology were influenced by the strength of the interactions between the amidine-functionalized amphiphiles and the oxoacid of the underlying SAMs. Dynamic control over surface composition, achieved by the addition of inert filler amphiphiles to the RGD-functionalized rSAMs, reversed the cell adhesion process. In summary, rSAMs featuring mobile bioactive ligands offer unique capabilities to influence and control cell adhesion behavior, suggesting a broad use in biomaterial design, tissue engineering, and regenerative medicine.

Loss of Chromium(III) From Mixed Cr(III), Fe(III) Serum Transferrins

Abstract Objectives Transferrin, Tf, the protein that transports iron as Fe(III) from the blood to the tissues via endocytosis, is believed to also transport chromium(III), Cr(III). Under physiological conditions, Tf binds and releases Cr(III) rapidly from Cr(III)2-Tf; however, the major form of Tf in the bloodstream is mono-ferric Tf. Given the low concentration of Cr(III) in the bloodstream, the form of Cr(III)-containing Tf that is transported is probably monochromic, monoferric-Tf (Cr(III), Fe(III)-Tf). Given that Tf has two specific metal-binding sites, one in both its C-terminal and the N-terminal lobe, two forms of Cr(III), Fe(III)-Tf can form. The loss of Cr(III) from both forms of Cr(III), Fe(III)-Tf has been examined for the first time. Methods Cr(III), Fe(III)-Tf with Cr(III) in the C-lobe was prepared by adding CrCl3 to N-lobe monoferric Tf in deionized H2O at pH 7.5 containing ∼20 mM bicarbonate. Cr(III), Fe(III)-Tf with Cr(III) in the N-lobe was prepared by adding 1 equivalent of Fe as ferrous ammonium sulfate to Cr(III)2-Tf in 0.1 M HEPES buffer solution with 25 mM bicarbonate at pH 7.4. To examine the release of Cr(III), the pH of solutions of Cr(III), Fe(III)-Tf were acidified from pH 7.4 to pH 5.5. After time intervals, aliquots were removed and frozen for analysis by electron paramagnetic resonance (EPR) spectroscopy, which can distinguish aquated Cr(III) and Cr(III) bound to each of the two metal binding sites of Tf. Results The acidification of solutions of Cr(III), Fe(III)-Tf's in 100 mM HEPES and 25 mM bicarbonate solution, pH 7.4 to pH 5.5 resulted in a rapid loss of Fe(III) followed by a slower loss of Cr(III). The loss of Cr(III) from the Cr(III), Fe(III)-Tf with Cr(III) in the C-lobe is about 4 times faster than that from the same site in Cr(III)2-Tf. For the other site, the Cr loss is about 28 times faster. Thus, having one site vacant from the loss of Fe(III) leads to faster loss of Cr(III) from both lobes, suggesting Cr(III) loss is cooperative. Conclusions Mixed-metal Cr(III), Fe(III) Tf's loose Cr(III) for both metal binding sites in similar fashions to Cr(III) losses from Cr(III)2-Tf. Cr(III) is probably transported as mixed Cr(III), Fe(III)-Tf's. Funding Sources The University of Alabama Bioinorganic Chemistry of Chromium Research Fund.

Structure-performance relationships of four lysosomal markers used for the imaging of HT-29 cancer cells and a cellular model of lysosomal storage disease (Niemann-Pick C)

Four new BODIPY derivatives with a potential tendency to aggregation have been synthesized and characterized by means of NMR techniques, mass spectrometry and UV–Vis/fluorescence spectroscopies. The objective of this study is to determine which structural factors of the new molecules influence most notably the cellular uptake, intracellular location and fluorescence imaging abilities. The behaviour of the compounds in organic solvent and aqueous solution has been studied. In organic solvents (DMSO, ethanol and toluene), the photophysical properties of the new molecules are almost independent of the building blocks used to synthesize the pendant moieties (non-fluorogenic parts). In an aqueous environment (HEPES Buffer Solution, pH 7), at 10 μM, three of the compounds (1, 2 and 4) tend to form weakly emissive nanoparticles (DLS determination) whereas one of them (3) remains soluble and highly fluorescent. In the nanomolar range of concentration, all the compounds are aqueous soluble. The cellular internalisation of the compounds (10 nM) has been studied in human colon adenocarcinoma HT-29 cells by means of flow cytometry and confocal laser scanning microscopy. All the compounds were uptaken by HT-29 cells, but notably molecule 3 (made with lysine as a building block) was the one displaying a higher loading and a more clear lysosomal location (0.88 Pearson's correlation coefficient in colocalization assays using lysosomal fluorescent probe LysoTracker DND-99). Molecule 3 also performed better than the valine derivative 1 as a lysosomal marker in a cellular model (human adrenal carcinoma SW13/cl.2 cells) of lysosomal storage disease (Niemann-Pick type C).

A probe with hydrazinecarbothioamide and 1,8-naphthalimide groups for “turn-on” fluorescence detection of Hg2+ and Ag+ ions and live-cell imaging studies

The probe (L) having hydrazinecarbothioamide and 1,8-naphthalimide moieties was synthesized and evaluated for its metal ion sensing ability. It exhibits a selective and sensitive colorimetric as well as fluorescent recognition of Hg2+ and Ag+ ions in CH3OH - HEPES buffer solution (5 mM, 7:3, v/v, pH = 7.4) in presence of other metal ions. Probe L is weakly fluorescent upon excitation with 410 nm light, but after gradual addition of HgCl2 and AgNO3 enhancement in fluorescent intensity was observed. Detection limits of Hg2+ and Ag+ using L have been found to be 20 nM (Hg2+) and 40 nM (Ag+) over the pH range of 6 – 10 that is suitable for practical application under physiological pH conditions. The reversibility of interaction of L with Hg2+ ion was monitored using Na2EDTA by emission titration. The “OFF-ON’’ fluorescence switching can be observed with naked eye, in which fluorescent “OFF’’ is due to operation of PET process in free L. Whereas upon complexation with these two metal ions, PET is restricted and CHEF process becomes operational. Mass spectral analysis and Job’s plot yielded a binding ratio of 1:1 for both metal ions. NMR titration studies are consistent with binding of Hg2+ or Ag+ to NH group (attached with naphthalimide moiety) and sulfur atom of hydrazinecarbothioamide group. From cytotoxicity assay, 5 μM solution of L was considered in cellular imaging study and the potentiality of the probe L was established by using human breast cancer cell line (MDA-MB-231) and primary human dermal fibroblasts (HDF), through fluorescence cell imaging experiments for tracking both Hg2+ and Ag+ in living cells.

3-(2-Hydroxyphenyl)imidazo[5, 1-a]isoquinoline as Cu(II) sensor, its Cu(II) complex for selective detection of CN− ion and biological compatibility

The heterocyclic probe 3-(2-hydroxyphenyl)imidazo[5, 1-a]isoquinoline (LH) has exhibited specific recognition of Cu2+ ion by forming a complex of formula [Cu(L)2], which in turn showed recognition for CN− ions in CH3CN/aqueous HEPES-buffer solution (5 mM, pH = 7.4, 6:4, v/v). The selective colorimetric and fluorescence response of LH towards Cu2+ was achieved over other competing metal ions. Change of color from colorless to yellowish green upon incremental addition of Cu2+ into LH solution can be easily observed with naked-eye. Emission intensity of LH was quenched in presence of Cu2+ ion due to chelation enhanced quenching (CHEQ) process. From Job’s plot and mass spectral analysis the binding stoichiometry was found to 2:1 between LH and Cu2+. Binding of the probe LH with Cu2+ ion can be reversed by sequestering Cu2+ ion using Na2EDTA solutions. Red shift in UV–visible spectrum upon addition of Cu2+ ion into LH solution was a consequence of decrease in HOMO-LUMO energy gap. From life time decay curve the average fluorescence life time (τ) was found to be 3.8 ns for free LH whereas in presence of Cu2+ τ decreased to 2.4 ns. In addition, so formed [Cu(L)2] complex was able to detect cyanide ion over other anions through removal of copper from the complex, as insoluble [CuCN]x. Limit of detection of LH for Cu2+ and [Cu(L)2] for CN− were found to be 0.45 μM and 0.30 μM respectively. Detection of Cu2+ and CN− were also performed in environmental real water samples. Based on the cytotoxic analysis, 5 μM of LH was selected for determining its fluorescence attributes in cellular imaging in MDA-MB-231 and HDF cells. The cell images showed that intracellular Cu2+ and CN− can be detected using LH and [Cu(L)2] complex respectively.

A 3D Bioprinted Human Meniscus Shape Enriched with Mesenchymal Cells

Background and objectives: Regenerative medicine, with its massive development over the years, has the potential to solve some of the most problematic medical issues, such as functional organ transplantation. The aim of this study was to create a human meniscal shape 3D-printed enriched with human adipose-derived mesenchymal cells. Materials and Methods: Human infrapatellar fat pad was harvested, and mesenchymal cells were isolated. The mesenchymal stem cells were differentiated to the chondrocite lineage and a hydrogel (a nanofibrillar cellulose, sodium alginate, D-mannitol, and Hepes buffer solution combination) cell mixture was bioprinted to create three human-size meniscus structures. The obtained structures were evaluated regarding the cell viability, appropriate size in relation to a native meniscus, and some mechanical characteristics. Results: The human meniscal shape created respected the anatomic characteristic of a native structure. Cell viability of approximately 97% and extracellular matrix formation after the printing process were observed. The mean maximum force for the meniscus with mesenchymal cells was 6.5 N (+/−0.5 N) compared to the mean maximum force for the native meniscus of 10.32 N (+/−0.7 N), which is statistically relevant (p < 0.01). Conclusion: This paper presents the potential of bioprinting viable cell structures that could in the future present enough mechanical strength to replace a human organ, such as a meniscus. There are still limitations regarding the ink and the printing process, but we are confident that these problems will soon be solvable.

Facile synthesis and electrochemical performance of lithium-rich layered oxides with stable hierarchical structure through HEPES-assisted co-precipitation method

Lithium-rich layered oxide (LLO) electrode material is now actively developed as a promising cathode for next-generation lithium-ion batteries due to its high specific capacity. The electrochemical property of the LLO material is mainly regulated by preparation conditions. Here, we report a facile 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (HEPES) assisted co-precipitation method to prepare Li1.2Mn0.54Co0.13Ni0.13O2. The HEPES buffer solution with strong pH buffering capacity not only can keep the pH of the reaction solution stable, but also acts a complexing agent to promote the uniform distribution of elements and in control of the nucleation and growth rate of crystals during the co-precipitation process. The dispersant role of HEPES helps to prepare regular LLO microspheres with unique hierarchical structure, which endows sufficient contact of the particles with electrolyte, shortens Li+ diffusion path and provides high structural stability. The electrode demonstrates superior electrochemical performance and excellent cycling stability at different discharge currents. High discharge capacities of 147.2 and 100.0 mAh g−1 could be achieved after 200 cycles at high current densities of 5 C and 10 C, respectively. The paper provides a novel strategy of the controllable route for high energy storage materials and reveals the correlation between the cation distribution and the electrochemical performance stability of the materials.