Glycine Ethyl Ester Research Articles

Functional profiling of the rhizospheric Exiguobacterium sp. for dimethoate degradation, PGPR activity, biofilm development, and ecotoxicological risk

This study introduces an indigenous bacterial strain, Exiguobacterium sp. (L.O), isolated from sugarcane fields in Sevur, Tamil Nadu, which has adapted to prolonged exposure to dimethoate. The strain demonstrated the capability to utilize 150 ppm of dimethoate as its sole carbon source, achieving a remarkable degradation rate of 95.87% within 5 days in mineral salt media. Gas chromatography–mass spectrometry (GC–MS) analyses identified the presence of intermediate by-products formed during degradation, like methyl diethanol amine and aspartyl glycine ethyl ester. Notably, phosphorothioic O, O, S-acid, an expected end product in the degradation of dimethoate, was also identified, further confirming the strain’s effective metabolic breakdown of the pesticide. Further degradation study and analysis of changes in functional group was performed by FTIR, and a hypothetical degradation pathway was elucidated showing the course of dimethoate metabolism by the strain. Exiguobacterium sp. (L.O) also displayed significant plant growth-promoting traits, including the production of HCN, IAA, and ammonia and the formation of biofilms, which enhance its utility in agricultural applications. The ecotoxicity study revealed the degradation by-products exhibited reduced toxicity compared to the parent compound dimethoate, highlighting the strain’s potential not only for bioremediation but also for supporting sustainable agricultural practices. This research presents a novel application of Exiguobacterium sp. (L.O), integrating the bioremediation of the organophosphate pesticide dimethoate with agricultural enhancement. This approach is critical for addressing the challenges associated with pesticide pollution in agricultural practices. This study is likely the first to demonstrate the application of this strain in the degradation of dimethoate, as suggested by an extensive review of the literature.

1,3-dipolar cycloaddition as a method for the synthesis of dipyrrolidinyl- and dipyrrolylketones

In the reactions of 1,3-dipolar cycloaddition of a twofold excess of aryl aldimines of glycine ethyl ester with diarylideneacetones and diarylidenecyclohexanones in the presence of silver acetate, the corresponding dipyrrolidinylketones were obtained. Diethyl 4,4′-carbonylbis(3,5-diarylpyrrolidine-2-carboxylates) obtained from diarylideneacetones undergo aromatization under the action of N-bromosuccinimide to form diethyl 4,4′-carbonylbis(3,5-diaryl-1H-pyrrole-2-carboxylates). The selectivity of the reactions and the structure of the products were determined using correlation NMR spectroscopy and X-ray diffraction analysis.

Directional Management Self‐Additive Spacer Cations for Stable 2D Ruddlesden–Popper Perovskite Solar Cells with Efficiency over 21%

Abstract2D Ruddlesden–Popper perovskites are highly regarded materials for improving the stability of perovskite solar cells (PSCs). Wherein, self‐additive 2D perovskites have recently been proposed to provide substantial strategies for managing the crystallization kinetics and bulk defects. For a profound understanding of the formation mechanisms, herein, with selecting three self‐additive 2D perovskites as demonstrations, a comprehensive analysis of the self‐additive behavior combing experimental and theoretical calculations is conducted. Self‐additive 2D perovskites exhibit more suitable formation energies and strong interaction, which is conducive to realize the self‐additive effect to form more stable structure. As demonstrated, glycine (Gly)‐based spacer cations played a pivotal role in the nucleation and growth of 2D perovskites by adjusting the aggregation state of colloids precursor, resulting in excellent‐quality films with large average grain size (≈3 µm). Meanwhile, theoretical analysis of electronic distribution and binding energies (Eb) revealed that glycine ethyl ester (Gly‐E) perovskite possesses highly robust internal interactions, which will effectively mitigate defect formation and enhance device stability. Endowing with the above outstanding feature, Gly‐E devices exhibited an optimized PCE of 21.60%, one of the highest PCEs among all 2D RP PSCs (n ≤ 6). The findings provide a basis for the rational design of self‐additive 2D perovskites and achieving highly‐performance 2D RP PSCs.

Asymmetric Phase-Transfer Alkylation of Readily Available Aryl Aldehyde Schiff Bases of Amino Acid Ethyl Esters.

Asymmetric phase-transfer alkylation of the N-(arylmethylene)-α-alkylamino acid ethyl esters and N-(arylmethylene)glycine ethyl esters was found to be catalyzed by the (R)- or (S)-Simplified Maruoka Catalyst with high efficiency and excellent enantioselectivity. This approach was successfully applied to the enantioselective formal synthesis of the angiotensin II type 2 receptor (AT2R) antagonists Olodanrigan and LX9211, and the practical aspect is demonstrated by the kilogram-scale synthesis of a key intermediate for the synthesis of LX9211.

Amides of moronic acid and morolic acid with the tripeptides MAG and GAM targeting antimicrobial, antiviral and cytotoxic effects.

A series of amides of selected plant triterpenoids, moronic acid and morolic acid, with the tripeptides MAG and GAM, was designed and synthesized. Two required tripeptides 5 and 10 were synthesized by a step-wise chain elongation of the ethyl esters of either glycine or l-methionine at their N-terminus using Boc-protected amino acids in each step. The tripeptides 5 and 10 were used for the synthesis of 13-23, the derivatives of moronic acid (11) and morolic acid (12), to get a series of amide derivatives of the less frequently studied triterpenoids 11 and 12. The target compounds, and their intermediates, were subjected to an investigation of their antimicrobial, antiviral and cytotoxic activity. Selectivity of the pharmacological effects was found. Generally, the target compounds inhibited only the G+ microorganisms. Compound 16 inhibited Staphylococcus aureus (I = 99.6%; c = 62.5 μM) and Enterococcus faecalis (I = 85%; c = 250 μM). Several compounds showed moderate antiviral effects, both anti-HIV-1, 19 (EC50 = 57.0 ± 4.1 μM, CC50 > 100 μM), 20 (EC50 = 17.8 ± 2.1 μM, CC50 = 41.0 ± 5.2 μM) and 23 (EC50 = 12.6 ± 0.82 μM, CC50 = 38.0 ± 4.2 μM), and anti-HSV-1, 22 (EC50 = 27.7 ± 3.5 μM, CC50 > 100 μM) and 23 (EC50 = 30.9 ± 3.3 μM, CC50 > 100 μM). The target compounds showed no cytotoxicity in cancer cells, however, several of their intermediates were cytotoxic. Compound 21 showed cytotoxicity in HeLa (IC50 = 7.9 ± 2.1 μM), G-361 (IC50 = 8.0 ± 0.6 μM) and MCF7 (IC50 = 8.6 ± 0.2 μM) cancer cell lines, while being non-toxic in normal fibroblasts (BJ; IC50 > 50 μM).

Palladium‐Catalyzed Asymmetric Trifluoromethylated Allylic Alkylation of N‐Substituted Glycine Ethyl Esters with α‐(Trifluoromethyl)alkenyl Acetates

AbstractAn efficient strategy for asymmetric trifluoromethylated allylic alkylation of easily available N‐substituted glycine ethyl esters with α‐(trifluoromethyl)alkenyl acetates has been developed. Catalyzed by a [Pd(C3H5)Cl]2/(R)‐BINAP, various trifluoromethyl‐containing N‐substituted glycine ethyl ester derivatives are afforded with good yields and excellent enantioselectivities. The product can be readily converted into diverse fluoro‐substituted species, which shows the practicability of this method.

Fbg αC 389-402 Enhances Factor XIII Cross-Linking in the Fibrinogen αC Region Via Electrostatic and Hydrophobic Interactions.

Coagulation Factor XIII (FXIII) stabilizes blood clots by cross-linking glutamines and lysines in fibrin and other proteins. FXIII activity in the fibrinogen αC region (Fbg αC 221-610) is critical for clot stability and growth. Fbg αC 389-402 is a binding site for thrombin-activated FXIII, (FXIII-A*), with αC E396 promoting FXIII-A* binding and activity in αC. The current study aimed to discover additional residues within Fbg αC 389-402 that accelerate transglutaminase activity toward αC. Electrostatic αC residues (E395, E396, and D390), hydrophobic αC residues (W391 and F394), and residues αC 328-425 were studied by mutations to recombinant Fbg αC 233-425. FXIII activity was monitored through MS-based glycine ethyl ester (GEE) cross-linking and gel-based fluorescence monodansylcadaverine (MDC) cross-linking assays. Truncation mutations 403 Stop (Fbg αC 233-402), 389 Stop (Fbg αC 233-388), and 328 Stop (Fbg αC 233-327) reduced Q237-GEE and MDC cross-linking compared to wild-type (WT). Comparable cross-linking between 389 Stop and 328 Stop showed that FXIII is mainly affected by the loss of Fbg αC 389-402. Substitution mutations E396A, D390A, W391A, and F394A decreased cross-linking relative to WT, whereas E395A, E395S, E395K, and E396D had no effect. Similar FXIII-A* activities were observed for double mutants (D390A, E396A) and (W391A, E396A), relative to D390A and W391A, respectively. In contrast, cross-linking was reduced in (F394A, E396A), relative to F394A. In conclusion, Fbg αC 389-402 boosts FXIII activity in Fbg αC, with D390, W391, and F394 identified as key contributors in enhancing αC cross-linking.

Reactions of arylaldimines of glycine ethyl ester with halo-substituted arylideneacetones

The reaction of 1,3-dipolar cycloaddition of para -halosubstituted mono- and diarylideneacetones and azomethine ylides generated in situ from N -arylimines of glycine ethyl ester in the presence of silver acetate and a half time excess of triethylamine afforded tetrasubstituted ethyl pyrrolidine carboxylates. For para -halogensubstituted dibenzyledeneacetone, a minor product with a different regioselectivity was observed. The selectivity of the reaction and the structure of the products were determined using NMR correlation spectroscopy. The regioselectivity of the cycloaddition was interpreted using DFT/PBE calculations of the transition state energies of the reactions.

Fbg αC 389-402 enhances factor XIII crosslinking in the fibrinogen αC region via electrostatic and hydrophobic interactions.

Coagulation Factor XIII (FXIII) hinders fibrinolysis via transglutaminase activity in fibrin and other proteins. FXIII activity in the fibrinogen αC region (Fbg αC 221-610) is critical for clot stability and growth. Fbg αC 389-402 acts as a binding site for thrombin-activated FXIII, (FXIII-A∗), with αC E396 promoting FXIII-A∗ binding and activity. The current study aimed to discover additional residues within Fbg αC 389-402 that accelerate transglutaminase activity toward αC (233-425). αC E395 and D390 were proposed to form ionic interactions with FXIII that improve binding. αC W391 and F394 were hypothesized to augment FXIII-fibrinogen αC binding through favorable hydrophobic contacts with FXIII residues. Truncated Fbg αC 328-425 was also examined for any potential impact on FXIII activity. Fbg αC D390, W391, F394, E395, E396, and residues 328-425 were studied by select mutations to recombinant Fbg αC 233-425. FXIII activity was quantified by monitoring glycine ethyl ester (GEE) crosslinking to αC Q237 over time by mass spectrometry. Truncation mutations “403 Stop” (Fbg αC 233-402), “389 Stop” (Fbg αC 233-388), and “328 Stop” (Fbg αC 233-327) experienced reduced Q237-GEE crosslinking compared to wild-type (WT). Comparable crosslinking between “389 Stop” and “328 Stop” shows that FXIII is mainly affected by the loss of Fbg αC 389-402. Substitution mutations E396A, D390A, W391A, and F394A decreased crosslinking relative to WT, whereas E395A, E395S, E395K, and E396D had no effect. Similar FXIII-A∗ activities were observed for double mutants (D390A,E396A) and (W391A,E396A), relative to D390A and W391A, respectively. In contrast, crosslinking was further reduced in (F394A,E396A), relative to F394A. In conclusion, Fbg αC 389-402 boosts FXIII activity in Fbg αC, with D390, W391, and F394 identified as additional residues that enhance αC crosslinking.

Transglutaminase Activities of Blood Coagulant Factor XIII Are Dependent on the Activation Pathways and on the Substrates.

Factor XIII (FXIII) catalyzes formation of γ-glutamyl-ε-lysyl crosslinks between reactive glutamines (Q) and lysines (K). In plasma, FXIII is activated proteolytically (FXIII-A*) by the concerted action of thrombin and Ca2+. Cellular FXIII is activated nonproteolytically (FXIII-A°) by elevation of physiological Ca2+ concentrations. FXIII-A targets plasmatic and cellular substrates, but questions remain on correlating FXIII activation, resultant conformational changes, and crosslinking function to different physiological substrates. To address these issues, the characteristics of FXIII-A* versus FXIII-A° that contribute to transglutaminase activity and substrate specificities were investigated. Crosslinking of lysine mimics into a series of Q-containing substrates were measured using in-gel fluorescence, mass spectrometry, and UV-Vis spectroscopy. Covalent incorporation of fluorescent monodansylcadaverine revealed that FXIII-A* exhibits greater activity than FXIII-A° toward Q residues within Fbg αC (233-425 WT, Q328P Seoul II, and Q328PQ366N) and actin. FXIII-A* and FXIII-A° displayed similar activities toward α2-antiplasmin (α2AP), fibronectin, and Fbg αC (233-388, missing FXIII-binding site αC 389-402). Furthermore, the N-terminal α2AP peptide (1-15) exhibited similar kinetic properties for FXIII-A* and FXIII-A°. MALDI-TOF mass spectrometry assays with glycine ethyl ester and Fbg αC (233-425 WT, αC E396A, and truncated αC (233-388) further documented that FXIII-A* exerts greater benefit from the αC 389-402 binding site than FXIII-A°. Conformational properties of FXIII-A* versus A° are proposed to help promote transglutaminase function toward different substrates. A combination of protein substrate disorder and secondary FXIII-binding site exposure are utilized to control activity and specificity. From these studies, greater understandings of how FXIII-A targets different substrates are achieved.

Weed Control, Rice Safety, and Mechanism of the Novel Paddy Field Herbicide Glyamifop

Glyamifop (R&D code: FG001), (R)-(2-(4-(6-chlorobenzoxazol-2-oxy) phenoxy) propionyl) glycine ethyl ester is a newly developed aryloxyphenoxypropionate (HRAC Group 1) herbicide for weed control in paddy fields. This work determined the effect of Glyamifop on weeds and its safety for rice in the glasshouse. Glyamifop controlled the common gramineous weeds in paddy fields at 100 g a.i. ha−1: the fresh weight inhibition rates of Echinochloa crus-galli, Leptochloa chinensis, Setaria viridis, Eragrostis japonica, Digitaria sanguinalis and Panicum bisulcatum were all above 90%. It has almost no inhibitory effect on broad-leaved and cyperaceae weeds, such as Eclipta prostrata and Cyperus iria. Glyamifop inhibited cyhalofop-butyl-resistant L. chinensis, penoxsulam-resistant E. crus-galli and quinclorac-resistant E. crusgalli var. zelayensis by 100%, 99.98% and 96.37%, respectively, at 100 g a.i. ha−1, based on the fresh weight. The selectivity index of Glyamifop foliage treatment in the rice varieties japonica ‘Huaidao 5’, indica ‘Xiangliangyou 900’ and glutinous ‘Zhennuo 29’ was 5.93, 6.81 and 4.91, respectively; therefore, Glyamifop is safe for the 3 different rice varieties. Fresh weight rice inhibition rates were 7.18%, 2.99% and 7.93% at the 2.5-, 3.5- and 5.5-leaf stage, respectively, and the selectivity index was 5.18, 6.04 and 7.93, respectively, indicating that Glyamifop was safe for rice at these leaf stages. L. chinensis ACCase activity decreased with increasing Glyamifop concentration, and the inhibitory effect was similar to that of cyhalofop acid; this confirmed that Glyamifop is an ACCase inhibitor. In conclusion, Glyamifop has potential for the management of gramineous weeds as it has good activity against weeds that are resistant to common herbicides in paddy fields.

Crystallization and defects regulation of efficient inverted perovskite solar cells via glycine ethyl ester hydrochloride

Crystal quality and defects at grain boundaries (GBs) or grain surfaces are two dominant factors in perovskite solar cells (PSCs) that determine device performance. Here, a novel amino acid salt named glycine ethyl ester hydrochloride (GEECl) is first introduced into perovskite to improve crystal quality and reduce GBs. Owing to fewer GBs and better vertically extended crystals, PSCs with GEECl exhibit improved carrier transport ability. The defects at GBs or grain surfaces can be well controlled by filling the MA+ vacancy with –NH3+ and the interaction between the carbonyl group (C═O) and uncoordinated Pb2+. As a result, the GEECl-modified devices based on MAPbI3 achieved a higher efficiency of 19.86 % with an FF close to 80 %. Furthermore, the GEE+ cations locate at GBs or grain surfaces protect the perovskite films against moisture, which enhanced the stability of PSCs. This work provides a promising route to further accelerate the development of PSCs.

Combination of Glycinamide and Ascorbic Acid Synergistically Promotes Collagen Production and Wound Healing in Human Dermal Fibroblasts.

The purpose of this study is to present a novel strategy to enhance collagen production in cells. To identify amino acid analogs with excellent collagen production-enhancing effects, human dermal fibroblasts (HDFs) were treated with 20 kinds of amidated amino acids and 20 kinds of free amino acids, individually at 1 mM. The results showed that glycinamide enhanced collagen production (secreted collagen level) most effectively. Glycine also enhanced collagen production to a lesser degree. However, other glycine derivatives, such as N-acetyl glycine, N-acetyl glycinamide, glycine methyl ester, glycine ethyl ester, and glycyl glycine, did not show such effects. Glycinamide increased type I and III collagen protein levels without affecting COL1A1 and COL3A1 mRNA levels, whereas transforming growth factor-β1 (TGF-β1, 10 ng mL−1) increased both mRNA and protein levels of collagens. Ascorbic acid (AA, 1 mM) increased COL1A1 and COL3A1 mRNA and collagen I protein levels. Unlike TGF-β1, AA and glycinamide did not increase the protein level of α-smooth muscle actin, a marker of differentiation of fibroblasts into myofibroblasts. The combination of AA and glycinamide synergistically enhanced collagen production and wound closure in HDFs to a level similar to that in cells treated with TGF-β1. AA derivatives, such as magnesium ascorbyl 3-phosphate (MAP), 3-O-ethyl ascorbic acid, ascorbyl 2-O-glucoside, and ascorbyl tetraisopalmitate, enhanced collagen production, and the mRNA and protein levels of collagens at 1 mM, and their effects were further enhanced when co-treated with glycinamide. Among AA derivatives, MAP had a similar effect to AA in enhancing wound closure, and its effect was further enhanced by glycinamide. Other AA derivatives had different effects on wound closure. This study provides a new strategy to enhance cell collagen production and wound healing using glycinamide in combination with AA.

A One-Pot Synthesis of Disubstituted Thiazoles from Chalcone C–H Bonds, Elemental Sulfur, and Glycine Ethyl Ester

AbstractA method for assembling chalcones, glycine ethyl ester hydrochloride, and elemental sulfur toward a synthesis of 4,5-disubstituted thiazoles is reported. The transformation presumably proceeds via a sequence of β-C–H amination, annulation, and dealkoxycarbonylation. This tactic represents a rare example of a method for obtaining such disubstituted thiazoles directly from chalcone C–H bonds.

Functionalized 5‐Amino‐4‐cyanoxazoles, their Hetero‐ and Macrocyclic Derivatives: Preparation and Synthetic Applications

AbstractAn approach to a series of new 5‐amino‐4‐cyanoxazoles is described. Synthesis of the title compounds relied on a two‐step sequence including heterocyclization of 2‐amido‐3,3‐dichloroacrylonitriles with aliphatic secondary amines (dimethylamine, morpholine), primary aliphatic amines with active functional groups (2‐aminoethanol and glycine ethyl ester), and aniline. An efficient and straightforward protocol introduces a carboxylate group at the C‐2 position of 5‐amino‐4‐cyanoxazoles, connected to the heterocycle directly or through an aliphatic linker. This carboxylic group is an attractive motif that can be found in a variety of drug‐relevant compounds and also used for further modifications. Furthermore, efficient transformations of selected trisubstituted compounds were used to demonstrate their rich synthetic potential – e. g., as precursors to 2‐(4‐cyano‐5‐(dimethylamino)oxazol‐2‐yl)acetamides, oxazole‐containing macrocyclic structures, 2‐(oxazol‐2‐yl)acetamides, amino pyrazoles, 3‐(4‐cyano‐5‐aminoxazol‐2‐yl)coumarins, and oxazole amino acids.

An efficient approach to diarylethene-amino acid photochromic fluorescent hybrids

• An effective approach to creating photo-controlled diarylethene-amino acid hybrids. • X-ray diffraction study, spectral luminescent and photochromic properties. • Thermally and photochemically reversible intramolecular photocyclization. • Dependence of the photoinduced form stability on the structure. • Modulation of emission under successive irradiation with light of different wavelengths. An effective approach to the synthesis of diarylethene-amino acid hybrids DE-Gly, DE-AABA and DE-DAA ( 5a-d - 7-a-d ) was developed via condensation of furan-2,5-dione-based diarylethenes (DE) and ethyl esters of glycine (Gly), α-aminobutyric acid (AABA) and D-aspartic acid (DAA) with moderate to good yields. According to X-ray diffraction data, DE-Gly hybrid 5a exists in an antiparallel conformation with a distance of 4.549 Å between the reactive carbon atoms C(1)-C(11), potentially capable of forming a single bond, which is suitable for the conrotatory photocyclization reaction allowed by the Woodward-Hoffman rules. The structures of the obtained compounds were proved by 1 H, COZY, HSQC, HMBC and 13 C NMR spectroscopy. The hybrids DE-Gly, DE-AABA and DE-DAA absorb at 443–456 nm, which corresponds to their existence in a ring-open form O, and display fluorescence at 531–612 nm. Irradiation with light of 436 nm results in their rearrangement into ring-closed colored nonfluorescent isomers C. Backwards re-opening occurs under the action of visible light (λ > 500 nm). Spectral kinetic investigation of 5a,c revealed that the efficiency of photocyclization, as well as the emission quantum yield, decreases with the growth of solvent polarity. The ring-closed isomer C of sterically hindered 5a (R 2 = R 3 = Me) is stable at room temperature, whereas for 5c (R 3 = H), ring opening readily occurs with the speed increasing with the polarity of the solvent. The internal emission of sterically hindered hybrids 5a, 6a and 7a (R 2 = R 3 = Me) is reversibly modulated in a binary response with good fatigue resistance under successive irradiation with light of 436 and 540 nm.

Composites made of polyorganophosphazene and carbon nanotube up-regulating osteogenic activity of BMSCs under electrical stimulation

Bone is an electrically responsive tissue, so electroactive materials that can deliver electrical cues to bone are helpful for enhancing regeneration under electrical stimulation (ES), and conductive materials are crucial in ES transmission to determine osteogenesis. Compared with polyesters, biodegradable polyorganophosphazenes (POPPs) show superiority in the field of bone tissue engineering thanks to their rich phosphorus/nitrogen contents, suggesting that the combination of POPPs-based conductive substrates with ES may achieve synergistic enhancements on osteogenesis. Herein, conductive composite films were fabricated by blending poly[(alanine ethyl ester)-(glycine ethyl ester)]phosphazene (PAGP) with carbon nanotubes (CNTs). After surface modification with polydopamine (PDA), bone marrow mesenchymal stromal cells (BMSCs) were cultured on the films under ES, using the cells cultured on conductive films composed of poly(L-lactide) (PLLA) and CNTs as controls. The BMSCs on PAGP/CNT films demonstrated significantly faster proliferation rates and stronger osteogenic differentiation potentials than those on PLLA/CNT films, while cell attachments on the two PDA-coated substrates were similar. Under appropriate ES, further increases in the expressions of osteogenic markers as alkaline phosphatase, collagen I and calcium deposition were identified in comparison with the cases without ES. The contributions of the osteocompatible POPPs, the substrate conductivity and the ES treatment to enhanced osteogenesis suggested new strategies for the design of bone repair materials.

Separation of the host-guest system for ferrocene derivatives in octahedral nanocages by electrochemical ionization

An octahedral nanosized M6L4 cage complex is assembled from six 90° Pd(II) compounds [Pd(phen)(NO3)2, phen = 1,10-phenanthroline)] and four 120° trident ligands [2,4,6-tri(4-pyridyl)-1,3,5-triazine]. The size of the cage complex is verified by TEM image and electronic structure calculation. The cage complex can act as a host container that allows the encapsulation of suitable sized guest molecules, such as electroactive molecules (one is water-soluble, ferrocene methanol) and (the other is water-insoluble, ferrocene glycine ethyl ester). The host–guest complexes are characterized by NMR, IR, UV and electrochemical studies. Interestingly, under a 1.0 V electro-stimuli oxidation voltage, guest molecules encapsulated in the cage cavity are expelled upon oxidation (from neutral to monocations) respectively, which can evade from the cage cavity through controlling the reduction/oxidation state of small guest molecules.

Synthesis of Potential Haptens with Morphine Skeleton and Determination of Protonation Constants.

Vaccination could be a promising alternative warfare against drug addiction and abuse. For this purpose, so-called haptens can be used. These molecules alone do not induce the activation of the immune system, this occurs only when they are attached to an immunogenic carrier protein. Hence obtaining a free amino or carboxylic group during the structural transformation is an important part of the synthesis. Namely, these groups can be used to form the requisite peptide bond between the hapten and the carrier protein. Focusing on this basic principle, six nor-morphine compounds were treated with ethyl acrylate and ethyl bromoacetate, while the prepared esters were hydrolyzed to obtain the N-carboxymethyl- and N-carboxyethyl-normorphine derivatives which are considered as potential haptens. The next step was the coupling phase with glycine ethyl ester, but the reactions did not work or the work-up process was not accomplishable. As an alternative route, the normorphine-compounds were N-alkylated with N-(chloroacetyl)glycine ethyl ester. These products were hydrolyzed in alkaline media and after the work-up process all of the derivatives contained the free carboxylic group of the glycine side chain. The acid-base properties of these molecules are characterized in detail. In the N-carboxyalkyl derivatives, the basicity of the amino and phenolate site is within an order of magnitude. In the glycine derivatives the basicity of the amino group is significantly decreased compared to the parent compounds (i.e., morphine, oxymorphone) because of the electron withdrawing amide group. The protonation state of the carboxylate group significantly influences the basicity of the amino group. All of the glycine ester and the glycine carboxylic acid derivatives are currently under biological tests.

Integrated Approach for Characterizing Bispecific Antibody/Antigens Complexes and Mapping Binding Epitopes with SEC/MALS, Native Mass Spectrometry, and Protein Footprinting.

Bispecific antibodies (BsAbs), with a unique mechanism of recognizing two different epitopes or antigens, have shown potential in various therapeutic areas. Molecular characterization of BsAbs' epitopes not only allows for detailed understanding of their mechanism of actions but also guides the design and selection of drug candidate molecules. In this study, we illustrate the practical utility of an integrated approach, including size exclusion chromatography with multiangle light scattering and native mass spectrometry (MS) for the biophysical characterization of complex formation of a BsAb with two target antigens, cluster of differentiation 3 (CD3) and B-cell maturation antigen (BCMA). MS-based protein footprinting strategies, including hydrogen/deuterium exchange MS, fast photochemical oxidation of proteins, and carboxyl group footprinting with glycine ethyl ester, were further applied to determine BsAb's binding epitopes. This combination approach provides molecular details on the binding mechanisms of BsAb to the two distinct antigens with rapid output and high resolution.