Exploring the Impact of Subtle Differences in the Chemical Structure of 1-Alkylsulfates and 1-Alkylsulfonates on Their Interactions with Human Serum Albumin.

The objective of this study was to examine the interactions between anionic surfactants, specifically 1-alkylsulfonates (KXS) and 1-alkylsulfates (SXS) ions, with human serum albumin (HSA). A combination of experimental techniques, including isothermal titration calorimetry (ITC), steady-state fluorescence spectroscopy (SF), and molecular dynamics-based approaches was employed to gain a comprehensive understanding of these processes. It has been demonstrated that the subtle variations in the charge distribution on the anionic surfactant headgroups have a significant impact on the number of binding sites, the stoichiometry of the resulting complexes, and the strength of the interactions between the surfactants and the protein. Additionally, we established that the affinity of the investigated ligands to specific regions on the protein surface is governed by both the charge of the surfactant headgroup and the length of the aliphatic hydrocarbon chain. In summary, the findings highlight the crucial role of charge distribution on surfactant functional groups in the binding mode and the thermodynamic stability of surfactant-protein complexes.

Shorter Alkanesulfonate Carbon Chains Destabilize the Active Site Architecture of SsuD for Desulfonation.

Bacteria have evolved to utilize alternative organosulfur sources when sulfur is limiting. The SsuE/SsuD and MsuE/MsuD enzymes expressed when sulfur sources are restricted, are responsible for providing specific bacteria with sulfur in the form of alkanesulfonates. In this study, we evaluated why two structurally and functionally similar FMNH2-dependent monooxygenase enzymes (MsuD and SsuD) are needed for the acquisition of alkanesulfonates in some bacteria. In desulfonation assays, MsuD was able to utilize the entire range of alkanesulfonates (C1-C10). However, SsuD was not able to utilize smaller alkanesulfonate substrates. Interestingly, SsuD had a similar binding affinity for methanesulfonate (MES) (15 ± 1 µM) as MsuD (12 ± 1 µM) even though SsuD was not able to catalyze the desulfonation of the MES substrate. SsuD and MsuD showed decreased proteolytic susceptibility in the presence of FMNH2 with MES and octanesulfonate (OCS). Tighter loop closure was observed for the MsuD/FMNH2 complex with MES and OCS compared to SsuD under comparable conditions. Analysis of the SsuD/FMNH2/MES structure using accelerated molecular dynamics simulations found three different conformations for MES, demonstrating the instability of the bound structure. Even when MES was bound in a similar fashion to OCS within the active site, the smaller alkane chain resulted in a shift of FMNH2 so that it was no longer in a position to catalyze the desulfonation of MES. The active site of SsuD requires a longer alkane chain to maintain the appropriate architecture for desulfonation.

Sulfonate-isosteric replacement examined within heroin-hapten vaccine design.

Heroin overdose and addiction remain significant health and economic burdens in the world today costing billions of dollars annually. Moreover, only limited pharmacotherapeutic options are available for treatment of heroin addiction. In our efforts to combat the public health threat posed by heroin addiction, we have developed vaccines against heroin. To expand upon our existing heroin-vaccine arsenal, we synthesized new aryl and alkyl sulfonate ester haptens; namely aryl-mono-sulfonate (HMsAc) and Aryl/alkyl-di-sulfonate (H(Ds)2) as carboxyl-isosteres of heroin then compared them to our model heroin-hapten (HAc) through vaccination studies. Heroin haptens were conjugated to the carrier protein CRM197 and the resulting CRM-immunoconjugates were used to vaccinate Swiss Webster mice following an established immunization protocol. Binding studies revealed that the highest affinity anti-heroin antibodies were generated by the HMsAc vaccine followed by the HAc and H(Ds)2 vaccines, respectively (HMsAc > HAc≫HDs2). However, neither the HMsAc nor H(Ds)2 vaccines were able to generate high affinity antibodies to the psychoactive metabolite 6-acetyl morphine (6-AM), in comparison to the HAc vaccine. Blood brain bio-distribution studies supported these binding results with vaccine efficiency following the trend HAc > HMsAc â‰« H(Ds)2 The work described herein provides insight into the use of hapten-isosteric replacement in vaccine drug design.

Clinical Effect of the Acrylonitrile-Co-Methallyl Sulfonate Surface-Treated Membrane as a Cytokine Adsorption Therapy for Sepsis due to Acute Panperitonitis: A Retrospective Cohort Study.

INTRODUCTION: Sepsis is a systemic inflammatory response syndrome caused by infectious diseases, with cytokines possibly having an important role in the disease mechanism. Acrylonitrile-co-methallyl sulfonate surface-treated (AN69ST) membrane is expected to improve the outcomes of patients with sepsis through cytokine adsorption. OBJECTIVE: This study aimed to investigate the clinical effect of the AN69ST membrane in comparison to standard continuous renal replacement therapy (CRRT) membranes for panperitonitis due to lower gastrointestinal perforation. METHODS: Using the Diagnosis Procedure Combination database, we identified adult patients with sepsis due to panperitonitis receiving any CRRT. Propensity score matching was used to compare patients who received CRRT with the AN69ST membrane (AN69ST group) and those who received CRRT with other membranes (non-AN69ST group). The primary outcome measure was in-hospital mortality. RESULTS: A total of 528 and 1,445 patients were included in the AN69ST group and in the non-AN69ST group, respectively. Propensity score matching resulted in 521 pairs. There was no significant difference in in-hospital mortality (32.1 vs. 35.5%; p = 0.265) and 30-day mortality (41.3 vs. 42.8%, p = 0.074) between the AN69ST group and the non-AN69ST group. CONCLUSION: There is no significant difference in-hospital mortality between CRRT with the AN69ST membrane and CRRT with standard CRRT membranes for panperitonitis due to lower gastrointestinal perforation. These results indicate that the AN69ST membrane is not superior to the standard CRRT membrane.

Intercalated Poly (2-Acrylamido-2-methyl-1-propanesulfonic Acid) into Sulfonated Poly (1,4-Phenylene ether-ether-sulfone) Based Proton Exchange Membrane: Improved Ionic Conductivity.

A series of hybrid proton exchange membranes were synthesized via in situ polymerization of poly (2-acrylamido-2-methyl-1-propanesulfonic acid) PMPS with sulfonated poly (1,4-phenylene ether-ether-sulfone) (SPEES). The insertion of poly (2-acrylamido-2-methyl-1-propanesulfonic acid) PMPS, between the rigid skeleton of SPEES plays a reinforcing role to enhance the ionic conductivity. The synthesized polymer was chemically characterized by fourier-transform infrared spectroscopy (FT-IR) and nuclear magnetic resonance 1H NMR spectroscopy to demonstrate the successful grafting of PMPS with the pendent polymer chain of SPEES. A variety of physicochemical properties were also investigated such as ion exchange capacity (IEC), proton conductivity, water uptake and swelling ratio to characterize the suitability of the formed polymer for various electrochemical applications. SP-PMPS-03, having the highest concentration of all PMPS, shows excellent proton conductivity of 0.089 S cm-1 at 80 °C which is much higher than SPEES which is ~0.049 S cm-1. Optimum water uptake and swelling ratio with high conductivity is mainly attributed to a less ordered arrangement polymer chain with high density of the functional group to facilitate ionic transport. The residual weight was 93.35, 92.44 and 89.56%, for SP-PMPS-01, 02 and 03, respectively, in tests with Fenton's reagent after 24 h. In support of all above properties a good chemical and thermal stability was also achieved by SP-PMPS-03, owing to the durability for electrochemical application.

Preparation, Characterization, Morphological and Particle Properties of Crystallized Palm-Based Methyl Ester Sulphonates (MES) Powder.

Methyl ester sulphonates (MES) have been considered as an alternative green surfactant for the detergent market. Investigation on the purification of methyl ester sulphonates (MES) with various carbon chains of C12, C14, C16 and C16-18 derived from palm methyl ester is of great interest. These MES powders have been repeatedly crystallized with ethanol and the purity of MES has increased to a maximum of 99% active content and 96% crystallinity index without changing the structure. These crystallized MES with high active content have 1.0% to 2.3% moisture content and retained its di-salt content in the range of 5%. The crystallized MES C16 and C16-18 attained excellent flow characteristics. Morphology, structural and its crystallinity analyses showed that the crystals MES had good solubility properties, stable crystal structure (ß polymorphic) and triclinic lateral structure when it is in high active content. The brittleness of MES crystals increased from a ß' to a ß subcell. Crystal with high brittleness has the potential to ease production of powder, which leads to a reduction in the cost of production and improves efficiency.

Rational Design of Near-Infrared Aggregation-Induced-Emission-Active Probes: In Situ Mapping of Amyloid-ß Plaques with Ultrasensitivity and High-Fidelity.

High-fidelity mapping of amyloid-ß (Aß) plaques is critical for the early detection of Alzheimer's disease. However, in vivo probing of Aß plaques by commercially available thioflavin derivatives (ThT or ThS) has proven to be extremely limited, as evident by the restriction of enrichment quenching effect, low signal-to-noise ( S/ N) ratio, and poor blood-brain barrier (BBB) penetrability. Herein, we demonstrate a rational design strategy of near-infrared (NIR) aggregation-induced emission (AIE)-active probes for Aß plaques, through introducing a lipophilic π-conjugated thiophene-bridge for extension to NIR wavelength range with enhancement of BBB penetrability, and tuning the substituted position of the sulfonate group for guaranteeing specific hydrophilicity to maintain the fluorescence- off state before binding to Aß deposition. Probe QM-FN-SO3 has settled well the AIE dilemma between the lipophilic requirement for longer emission and aggregation behavior from water to protein fibrillogenesis, thus making a breakthrough in high-fidelity feedback on in vivo detection of Aß plaques with remarkable binding affinity, and serving as an efficient alternative to the commercial probe ThT or ThS.

Oxidation-Mediated, Zwitterionic Polydopamine Coatings for Marine Antifouling Applications.

We synthesized a zwitterionic dopamine derivative ( ZW-DOPA) containing both catechol and amine groups, and we demonstrated an excellent marine antifouling surface by controlling the oxidation of ZW-DOPA. The oxidation was mediated by the deprotonation of catechol or the addition of an oxidant (ammonium persulfate (AP) or sodium periodate (NaIO4)). The oxidation and subsequent molecular transformation of ZW-DOPA was investigated over time by UV-vis spectroscopy. Among the different oxidation conditions tested, NaIO4-induced ZW-DOPA coating was the most efficient and successfully formed on various substrates, such as titanium dioxide, stainless steel, and nylon. Compared with uncoated substrates, ZW-DOPA-coated substrates showed high resistance to marine diatom adhesion. Considering the ease of use and substrate independence of the ZW-DOPA coating, this method shows promise as a basis for inhibiting marine fouling on a variety of substrates used in the marine industry and aquatic environments.

Sulfonated Copolymers as Heparin-Mimicking Stabilizer of Fibroblast Growth Factor: Size, Architecture, and Monomer Distribution Effects.

Fibroblast growth factors (FGF) are involved in a wide range of biological processes such as cell proliferation and differentiation. In living organisms, the binding of FGF to its receptors are mediated through electrostatic interactions between FGF and naturally occurring heparin. Despite its prevalent use in medicine, heparin carries notable limitations; namely, its extraction from natural sources (expensive, low yield and extensive purification), viral contamination, and batch-to-batch heterogeneity. In this work a range of synthetic homopolymers and copolymers of sodium 2-acrylamido-2-methylpropanesulfonate were evaluated as potential FGF stabilizers. This was studied by measuring the proliferation of BaF3-FR1c cells, as a model assay, and the results will be compared with the natural stabilization and activation of FGF by heparin. This study explores the structure-activity relationship of these polysulfonated polymers with a focus on the effect of molecular weight, comonomer type, charge dispersion, and polymer architecture on protein stabilization.

The construction of DNAzyme-based logic gates for amplified microRNA detection and cancer recognition.

Benefiting from the simple DNAzyme and a duplex-specific nuclease, a series of microRNA-stimulated DNAzyme logic gates were rationally assembled for the highly accurate detection of multiple low-abundant microRNA biomarkers that correspond to the specific aberrant microRNA expression patterns of cancer tissues, thus showing a great potential in early diagnosis.

Triple-helix molecular-switch-actuated exponential rolling circular amplification for ultrasensitive fluorescence detection of miRNAs.

We have formulated a rapid and high-efficiency fluorescent biosensing platform based on a target-activated triple-helix molecular switch (THMS)-conformation-change-induced exponential rolling circular amplification (RCA) strategy for the ultrasensitive detection of miR-21. In this strategy, there are several aspects that are worthwhile. First, the functionalized THMS, comprising a typical triplex structure (T-A·T), specific recognition sequence for nicking endonuclease, complementary sequence for miR-21, and RCA product-annealing sequence, was concurrently used to perform signal transduction with one fluorophore and one quencher. As compared to the traditional double-helix molecular switches or molecular beacons, one of the biggest differentiating factors is that the properties of THMSs are independent of any specific binding sequence that they may contain. As far as we know, this is the first time that an ingeniously designed THMS not only contains the primer for exponential RCA, but also functions as the tracer for fluorescence assay. In the presence of miR-21, targets can induce conformation changes in THMS with the release of the trapped DNA segment (P), which, in turn, can activate the first run of the RCA process. Meanwhile, the RCA reaction is also initiated by the formation of a similar primer (SP) as the trapped DNA through a continuous "extension-nicking" reaction. Secondly, the resultant first-generation RCA product consists of numerous tandem repeated regions that can attach to countless THMSs, resulting in the release of the trapped DNA segment (P) for initiating the second run of the RCA reaction. Significantly, a large amount of THMSs were continuously consumed to yield a remarkably strong fluorescent signal. In addition, this biosensor was demonstrated to exhibit improved sensitivity owing to the high efficiency and rapid amplification kinetics of the exponential RCA and high selectivity toward miR-21 with a limit of detection as low as 1.1 aM. Hence, the target-mediated THMS-conformation-change-initiated exponential RCA strategy presents an optimal detection performance toward analytes for potential applications in related fundamental research and clinical diagnosis.

3-18F-fluoropropane-1-thiol and 18F-PEG4-1-thiol: Versatile prosthetic groups for radiolabeling maleimide functionalized peptides.

The efficient radiosynthesis of biomolecules utilizing minute quantities of maleimide substrate is important for availability of novel peptide molecular imaging agents. We evaluated both 3-18F-fluoropropane-1-thiol and 2-(2-(2-(2-18F-fluoroethoxy)ethoxy)ethoxy)ethane-1-thiol (18F-fluoro-PEG4 thiol) as prosthetic groups for radiolabeling under physiological conditions. The precursor employed a benzoate for protection of the thiol and an arylsulfonate leaving group. The radiofluorination was fully automated on an Eckert & Ziegler synthesis system using standard Kryptofix222/K2CO3 conditions. In order to minimize the amount of biological molecule required for subsequent conjugation, the intermediates, S-(3-18F-fluoropropyl) benzothioate and 18F-fluoro-PEG4 benzothioate, were purified by HPLC. The intermediates were isolated from the HPLC in yields of 37-47% and 28-35%, respectively, and retrieved from eluate using solid phase extraction. Treatment of the benzothioates with sodium methoxide followed by acetic acid provided the free thiols. The desired maleimide substrate in acetonitrile or phosphate buffer was then added and incubated at room temperature for 15 min. The final radiolabeled bioconjugate was purified on a separate HPLC or NAP-5 column. Maleimides utilized for the coupling reaction included phenyl maleimide, an Evans Blue maleimide derivative, a dimeric RGDfK maleimide (E[c(RGDfK)]2), two aptamer maleimides, and PSMA maleimide derivative. Isolated radiochemical yields (non-decay corrected) of maleimide addition products based on starting 18F-fluoride ranged from 6 to 22% in a synthesis time of about 90 min. 18F-thiol prosthetic groups were further tested in vivo by conjugation to E[c(RGDfK)]2 maleimide in a U87MG xenograft model. PET studies demonstrated similar tumor accumulation of both prosthetic groups. 18F-fluoro-PEG4-S-E[c(RGDfK)]2 displayed a somewhat favorable pharmacokinetics compared to 18F-fluoropropyl-S-E[c(RGDfK)]2. Bone uptake was low for both indicating in vivo stability.

A self-cleaning, mechanically robust membrane for minimizing the foreign body reaction: towards extending the lifetime of sub-Q glucose biosensors.

Long-term, subcutaneously implanted continuous glucose biosensors have the potential to improve diabetes management and reduce associated complications. However, the innate foreign body reaction (FBR) both alters the local glucose concentrations in the surrounding tissues and compromises glucose diffusion to the biosensor due to the recruitment of high-metabolizing inflammatory cells and the formation of a dense, collagenous fibrous capsule. Minimizing the FBR has mainly focused on "passively antifouling" materials that reduce initial cellular attachment, including poly(ethylene glycol) (PEG). Instead, the membrane reported herein utilizes an "actively antifouling" or "self-cleaning" mechanism to inhibit cellular attachment through continuous, cyclic deswelling/reswelling in response to normal temperature fluctuations of the subcutaneous tissue. This thermoresponsive double network (DN) membrane is based on N-isopropylacrylamide (NIPAAm) and 2-acrylamido-2-methylpropane sulfonic acid (AMPS) (75:25 and 100:0 NIPAAm:AMPS in the 1st and 2nd networks, respectively; "DN-25%"). The extent of the FBR reaction of a subcutaneously implanted DN-25% cylindrical membrane was evaluated in rodents in parallel with a PEG-diacrylate (PEG-DA) hydrogel as an established benchmark biocompatible control. Notably, the DN-25% implants were more than 25× stronger and tougher than the PEG-DA implants while maintaining a modulus near that of subcutaneous tissue. From examining the FBR at 7, 30 and 90 days after implantation, the thermoresponsive DN-25% implants demonstrated a rapid healing response and a minimal fibrous capsule (~20-25 µm), similar to the PEG-DA implants. Thus, the dynamic self-cleaning mechanism of the DN-25% membranes represents a new approach to limit the FBR while achieving the durability necessary for long-term implantable glucose biosensors.

Automated production of [68 Ga]Ga-DOTANOC and [68 Ga]Ga-PSMA-11 using a TRACERlab FXFN synthesis module.

The interest in gallium-68 labelled positron-emission tomography probes continues to increase around the world. However, one of the barriers for routine clinical use is the cost of the automated synthesis units for relatively simple labelling procedures. Herein, we describe the adaptation of a TRACERlab FXFN synthesis module for the automated production of gallium-68 radiopharmaceuticals using a cation-exchange cartridge for postprocessing of the 68 Ge/68 Ga generator eluate. The recovery of activity from the cartridge was 95.6% to 98.9% using solutions of acidified sodium chloride (5 M with pH = 1-3). The radiosyntheses of [68 Ga]Ga-DOTANOC and [68 Ga]Ga-PSMA-11 were performed using acetate sodium buffer or 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid, with a total duration of 21 and 23 minutes, respectively, including generator elution and radiopharmaceutical dispensing. Activity yields were 77% ± 2% for [68 Ga]Ga-PSMA-11 and 68% ± 3% for [68 Ga]Ga-DOTANOC (n > 100). The labelled peptides had a radiochemical purity exceeding 97%, and all quality control parameters were in conformity with the limits prescribed by the European Pharmacopoeia.

Molecular Rotors for Universal Quantitation of Nanoscale Hydrophobic Interfaces in Microplate Format.

Hydrophobic self-assembly pairs diverse chemical precursors and simple formulation processes to access a vast array of functional colloids. Exploration of this design space, however, is stymied by lack of broadly general, high-throughput colloid characterization tools. Here, we show that a narrow structural subset of fluorescent, zwitterionic molecular rotors, dialkylaminostilbazolium sulfonates [DASS] with intermediate-length alkyl tails, fills this major analytical void by quantitatively sensing hydrophobic interfaces in microplate format. DASS dyes supersede existing interfacial probes by avoiding off-target fluorogenic interactions and dye aggregation while preserving hydrophobic partitioning strength. To illustrate the generality of this approach, we demonstrate (i) a microplate-based technique for measuring mass concentration of small (20-200 nm), dilute (submicrogram sensitivity) drug delivery nanoparticles; (ii) elimination of particle size, surfactant chemistry, and throughput constraints on quantifying the complex surfactant/metal oxide adsorption isotherms critical for environmental remediation and enhanced oil recovery; and (iii) more reliable self-assembly onset quantitation for chemically and structurally distinct amphiphiles. These methods could streamline the development of nanotechnologies for a broad range of applications.

Micellar Aggregation Behavior of Alkylaryl Sulfonate Surfactants for Enhanced Oil Recovery.

Alkylaryl sulfonate is a typical family of surfactants used for chemically enhanced oil recovery (EOR). While it has been widely used in surfactant-polymer flooding at Karamay Oilfield (40 °C, salinity 14,000 mg/L), its aggregation behavior in aqueous solutions and the contribution of aggregation to EOR have not been investigated so far. In this study, raw naphthenic arylsulfonate (NAS) and its purified derivatives, alkylaryl monosulfonate (AMS) and alkylaryl disulfonate (ADS), were examined under simulated temperature and salinity environment of Karamay reservoirs for their micellar aggregation behavior through measuring surface tension, micellar size, and micellar aggregation number. It was found that all three alkylaryl sulfonate surfactants could significantly lower the surface tension of their aqueous solutions. Also, it has been noted that an elevation both in temperature and salinity reduced the surface tension and critical micellar concentration. The results promote understanding of the performance of NAS and screening surfactants in EOR.

Synthesis of AMPSA Polymeric Derivatives Monitored by Electrical Conductivity and Evaluation of Thermosensitive Properties of Resulting Microspheres.

Four stimuli-responsive polymers of N-isopropylacrylamide (NIPA) and 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPSA) and crosslinked derivatives by N,N'-methylene bisacrylamide (MBA) were synthesized: PNA, PAMPSA, PNAM, PAMPSAM. The effect of the cross-linker and methyl sulphonic acid (-CH3-SO3H) group on particle size, aggregation and volume phase transition temperature (VPTT) was investigated. Polymeric particles were synthesized via the surfactant free precipitation polymerization (SFPP) at 70 °C in the presence of cationic initiator 2,2'-azobis[2-methylpropionamidine] dihydrochloride (AMP) system. Chemical composition and morphology of investigated samples were evaluated using ATR-FTIR spectroscopy, ¹H-NMR spectrometry and SEM-EDS techniques. The hydrodynamic diameters (HD), zeta potential (ZP), and polydispersity index (PDI) in aqueous dispersions were assessed by dynamic light scattering (DLS) between 18⁻42 °C. HD values at 18 °C for PNA, PAMPSA, PNAM, PAMPSAM polymers were approx. 32, 730, 715, 665 nm, and ZP values were -1.36, -0.01, 8.90, -0.09 mV, respectively. The VPTT range was observed between 29 and 41 °C. PDI's for PNA and PNAM were low and varied between 0.276 and 0.460, and between 0.119 and 0.056, respectively. PAMPSA and PAMPSAM were characterized by higher PDI in the range 0.728⁻0.959 and 0.658⁻0.836, respectively. The results confirmed the thermal sensitivity of the synthesized polymers and indicated a significant polydispersity and aggregation tendency of the resulting molecules. The conductivity results were applied for the interpretation of the polymerization process.

Optical Control of Metal Ion Probes in Cells and Zebrafish Using Highly Selective DNAzymes Conjugated to Upconversion Nanoparticles.

Spatial and temporal distributions of metal ions in vitro and in vivo are crucial in our understanding of the roles of metal ions in biological systems, and yet there is a very limited number of methods to probe metal ions with high space and time resolution, especially in vivo. To overcome this limitation, we report a Zn2+-specific near-infrared (NIR) DNAzyme nanoprobe for real-time metal ion tracking with spatiotemporal control in early embryos and larvae of zebrafish. By conjugating photocaged DNAzymes onto lanthanide-doped upconversion nanoparticles (UCNPs), we have achieved upconversion of a deep tissue penetrating NIR 980 nm light into 365 nm emission. The UV photon then efficiently photodecages a substrate strand containing a nitrobenzyl group at the 2'-OH of adenosine ribonucleotide, allowing enzymatic cleavage by a complementary DNA strand containing a Zn2+-selective DNAzyme. The product containing a visible FAM fluorophore that is initially quenched by BHQ1 and Dabcyl quenchers is released after cleavage, resulting in higher fluorescent signals. The DNAzyme-UCNP probe enables Zn2+ sensing by exciting in the NIR biological imaging window in both living cells and zebrafish embryos and detecting in the visible region. In this study, we introduce a platform that can be used to understand the Zn2+ distribution with spatiotemporal control, thereby giving insights into the dynamical Zn2+ ion distribution in intracellular and in vivo models.

High-Throughput Measurement of Small-Molecule Enantiopurity by Using Flow Cytometry.

Fluorescence-activated cell sorting (FACS) offers a powerful approach to high-throughput library screening in directed evolution experiments. However, FACS is rarely used in the evolution of stereoselective enzymes, due to the difficulty of designing fluorescence-based assays for measuring enantiopurity. Here, we describe a new FACS-based enantiopurity analysis approach that overcomes these limitations by using enantiomeric DNA biosensors labeled with orthogonal fluorophores. By co-encapsulating the biosensors with a mixture of target enantiomers in microfluidic droplets, we could demonstrate the use of FACS to differentiate between droplets having various levels of target enantiopurity. We envision the utility of this method for high-throughput screening of enantiopurity in the directed evolution of stereoselective enzymes, thereby facilitating the discovery of new asymmetric biocatalysts for the synthesis of pharmaceuticals and other high-value chemicals.

Aminoethane Sulfonic Acid Magnesium Salt Inhibits Ca2+ Entry Through NMDA Receptor Ion Channel In Vitro.

The effect of a cerebroprotective agent magnesium bis-aminoethanesulfonate (laboratory code FS-LKhT-317) on intracellular calcium concentration was studied by the fluorescent imaging technique on neuroglial cell culture from Spraque-Dawley rat hippocampus. The substance produced a pronounced inhibitory effect and suppressed NMDA receptor activity in concentrations of ≥50 µM. The observed effects were reversible or partially reversible and were detected by a decrease in Ca2+ signal amplitude in neurons in response to NMDA applications in a Mg2+-free medium and by inhibition of Ca2+ pulses in magnesium-free medium (elimination of magnesium block).