The impact and future of artificial intelligence in medical genetics and molecular medicine: an ongoing revolution.

Artificial intelligence (AI) platforms have emerged as pivotal tools in genetics and molecular medicine, as in many other fields. The growth in patient data, identification of new diseases and phenotypes, discovery of new intracellular pathways, availability of greater sets of omics data, and the need to continuously analyse them have led to the development of new AI platforms. AI continues to weave its way into the fabric of genetics with the potential to unlock new discoveries and enhance patient care. This technology is setting the stage for breakthroughs across various domains, including dysmorphology, rare hereditary diseases, cancers, clinical microbiomics, the investigation of zoonotic diseases, omics studies in all medical disciplines. AI's role in facilitating a deeper understanding of these areas heralds a new era of personalised medicine, where treatments and diagnoses are tailored to the individual's molecular features, offering a more precise approach to combating genetic or acquired disorders. The significance of these AI platforms is growing as they assist healthcare professionals in the diagnostic and treatment processes, marking a pivotal shift towards more informed, efficient, and effective medical practice. In this review, we will explore the range of AI tools available and show how they have become vital in various sectors of genomic research supporting clinical decisions.

Molecular dynamics simulations reliably identify vibrational modes in far-IR spectra of phospholipids.

The properties of self-assembled phospholipid membranes are of essential importance in biochemistry and physical chemistry, providing a platform for many cellular life functions. Far-infrared (far-IR) vibrational spectroscopy, on the other hand, is a highly information-rich method to characterize intermolecular interactions and collective behaviour of lipids that can help explain, e.g., chain packing, thermodynamic phase behaviour, and sequestration. However, reliable interpretation of the far-IR spectra is still lacking. Here we present a molecular dynamics (MD) based approach to simulate vibrational modes of individual lipids and in an ensemble. The results are a good match to synchrotron far-IR measurements and enable identification of the molecular motions corresponding to each vibrational mode, thus allowing the correct interpretation of membrane spectra with high accuracy and resolving the longstanding ambiguities in the literature in this regard. Our results demonstrate the feasibility of using MD simulations for interpreting far-IR spectra broadly, opening new avenues for practical use of this powerful method.

A Comprehensive Study of the Radical Scavenging Activity of Rosmarinic Acid.

Rosmarinic acid (RA) is reported in separate studies to be either an inducer or reliever of oxidative stress, and this contradiction has not been resolved. In this study, we present a comprehensive examination of the radical scavenging activity of RA using density functional theory calculations in comparison with experimental data. In model physiological media, RA exhibited strong HO• radical scavenging activity with overall rate constant values of 2.89 × 1010 and 3.86 × 109 M-1 s-1. RA is anticipated to exhibit excellent scavenging properties for HOO• in an aqueous environment (koverall = 3.18 × 108 M-1 s-1, ≈2446 times of Trolox) following the hydrogen transfer and single electron transfer pathways of the dianion state. The neutral form of the activity is equally noteworthy in a lipid environment (koverall = 3.16 × 104 M-1 s-1) by the formal hydrogen transfer mechanism of the O6(7,15,16)-H bonds. Chelation with RA may prevent Cu(II) from reduction by the ascorbic acid anion (AA-), hence blocking the OIL-1 pathway, suggesting that RA in an aqueous environment also serves as an OIL-1 antioxidant. The computational findings exhibit strong concurrence with the experimental observations, indicating that RA possesses a significant efficacy as a radical scavenger in physiological environments.

Study of the Mechanism and Kinetics of the Radical Scavenging Activity of 2-Mercaptoimidazole.

2-Mercaptoimidazole (2MI) is related to natural ovothiols that are recognized as powerful radical scavengers. Yet, despite early reports of its potent antioxidant properties, 2MI received little attention. Specifically, its radical scavenging activity against typical free radicals like HO• and HOO• has not yet been studied in terms of its mechanism and kinetics. In this project, density functional theory (DFT) simulations were used to assess the antiradical activity of 2MI. Calculations indicate that 2MI can demonstrate anti-HO• activity in both lipid and aqueous environments (koverall of 1.05 × 1010 and 2.07 × 1010 M-1 s-1, respectively). The calculated kinetics is extremely close to the experimental data in water (pH = 7.0), resulting in a kcalculated/kexperimental ratio of 1.73, validating the accuracy of the computational method and its usefulness for assessing radical scavenging activity in silico. In lipid media, the HOO• radical scavenging activity of 2MI is faster than that of common typical natural scavengers such as ascorbic acid, Trolox, and trans-resveratrol; hence, 2MI is a powerful radical scavenger in nonpolar media.

C-amidation of substitutedß3oligoamides yields novel supramolecular assembly motif.

N-acylated substitutedß3oligoamides are known to form unique supramolecular nanorods based on a 3-point hydrogen bond self-assembly motif. This motif is an intermolecular extension of the hydrogen bonding network that stabilizes the 14-helix secondary structure unique toß3oligoamides. Acetylation of the N-terminus of the molecule provides the necessary third hydrogen bond pair of the motif. Here, the possibility of introducing the third hydrogen bond pair via amidation of the C terminus is investigated. While similar in purpose, this modification introduces a chemically distinct new self-assembly motif, also removing the bulky carboxyl group that does not fold into the 14 helix positioning instead as a side chain. Three substitutedß3oligoamide variants with the base sequence LIA (where the letters denoteß3residues with side chains analogous to α amino acids) were compared: N-acylated Ac-ß3[LIA] as a reference, C-amidatedß3[LIA]-CONH2, andß3[LIA] with free unmodified N and C termini as a negative control. The three variants were dissolved in water to promote self-assembly. The self-assembly was characterised using mid- and far-infrared spectroscopy, small angle x-ray scattering (SAXS) and atomic force microscopy (AFM). IR measurements confirmed that all three samples were in a similar conformation, consistent with pseudo 14-helical secondary structures. Far-infrared spectroscopy measurements ofß3[LIA]-CONH2showed distinct peaks consistent with highly organised skeletal modes, i.e. regular supramolecular assembly, that was largely absent from the other two oligoamides. Modelling of SAXS data is consistent with elliptical cylinder structures resulting from nanorod bundling for bothß3[LIA]-CONH2and Ac-ß3[LIA], but not in the unmodified sample. Consistently, AFM imaging showed large nanorod bundling structures inß3[LIA]-CONH2, varied bundling structures in Ac-ß3[LIA], and only aggregation inß3[LIA]. Amidation showed much more organised and robust assembly compared to acetylation, providing a new, easy to synthesize self-assembly motif for helical nanorod assembly that is similar but distinct to N-acylation.

Theoretical and Experimental Studies of the Antioxidant and Antinitrosant Activity of Syringic Acid.

Syringic acid (SA) is a natural phenolic acid found in vegetables, fruits, and other plant-based foods. A range of biological activities were proposed for this compound including anticancer, antimicrobial, anti-inflammation, and anti-diabetic activities, as well as antioxidant and antinitrosant properties. In this study, the focus is on the latter two. The HO•, HOO•, NO, and NO2 scavenging activities of SA were evaluated in physiological environments by kinetic and thermodynamic calculations. The computed rate constants of the HO• radical scavenging of SA were 4.63 × 109 and 9.77 × 107 M-1 s-1 in polar and nonpolar solvents, respectively. A comparison with the experimentally determined rate constant in aqueous solution yields a kcalculated/kexperimental ratio of 0.3, thus the computed kinetic data are reasonably accurate. SA exhibited excellent HOO• and NO2 scavenging activity in water (koverall(HOO•) = 1.53 × 108 M-1 s-1 and koverall(NO2) = 1.98 × 108 M-1 s-1), whereas it did not show NO scavenging activity in any of the studied environments. In lipid medium, SA exhibited weak activity. Thus, in polar environments, the HOO• radical scavenging of SA is 1.53 times higher than that of ascorbic acid. Consistently, SA is a promising antioxidant and antinitrosant agent in polar environments.

In Silico Study of the Radical Scavenging Activities of Natural Indole-3-Carbinols.

Enzymatic hydrolysis of indole glucosinolates in the human body yields the Indole-3-carbinol family of compounds (I3Cs). I3Cs are implicated in the self-healing processes of the human body and thus used as over the counter dietary supplements characterized as potential antioxidants. In this study, the antioxidant activity of natural indole-3-carbinol derivatives were investigated against the HOO• radical by using thermodynamic and kinetic calculations. It was found that natural I3Cs containing only C8-H and/or N1-H bonds such as indole-3-carbinol, 4-methoxy-indole-3-carbinol, and 1-methoxy-indole-3-carbinol exhibit weak antioxidant properties. However, the presence of OH groups in the indole ring significantly increases the radical scavenging in aqueous as well as lipid media. In particular the HOO• scavenging activity of 1-hydroxyl-indole-3-carbinol far exceeds that of Trolox, 2.0 times in aqueous medium and 352.9 times in lipid (pentyl ethanoate) medium. This suggests that 1-hydroxyl-indole-3-carbinol is one of the most powerful known antioxidants in lipid environments, pointing at potential dietary application in preventive and regenerative medicine.

Nanoviscosity Measurements Revealing Domain Formation in Biomimetic Membranes.

Partitioning of lipid molecules in biomimetic membranes is a model system for the study of naturally occurring domains, such as rafts, in biological membranes. The existence of nanometer scale membrane domains in binary lipid mixtures has been shown with microscopy methods; however, the nature of these domains has not been established unequivocally. A common notion is to ascribe domain separation to thermodynamic phase equilibria. However, characterizing thermodynamic phases of single bilayer membranes has not been possible due to their extreme dimensions: the size of the domains falls to the order of tens to hundreds of nanometers whereas the membrane thickness is only a few nanometers. Here, we present direct measurements of phase transitions in single bilayers of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) phospholipid mixtures using quartz crystal microbalance-based nanoviscosity measurements. Coexisting thermodynamic phases have been successfully identified, and a phase diagram was constructed for the single bilayer binary lipid system. It was demonstrated that domain separation only takes place in planar membranes, and thus, it is absent in liposomes and not detectable in calorimetric measurements on liposome suspensions. On the basis of energetic analysis, the main transition was identified as the breaking of van der Waals interactions between the acyl chains.

A QCM-D and SAXS Study of the Interaction of Functionalised Lyotropic Liquid Crystalline Lipid Nanoparticles with siRNA.

Biophysical studies were undertaken to investigate the binding and release of short interfering ribonucleic acid (siRNA) from lyotropic liquid crystalline lipid nanoparticles (LNPs) by using a quartz crystal microbalance (QCM). These carriers are based on phytantriol (Phy) and the cationic lipid DOTAP (1,2-dioleoyloxy-3-(trimethylammonium)propane). The nonlamellar phase LNPs were tethered to the surface of the QCM chip for analysis based on biotin-neutravidin binding, which enabled the controlled deposition of siRNA-LNP complexes with different lipid/siRNA charge ratios on a QCM-D crystal sensor. The binding and release of biomolecules such as siRNA from LNPs was demonstrated to be reliably characterised by this technique. Essential physicochemical parameters of the cationic LNP/siRNA lipoplexes-such as particle size, lyotropic phase behaviour, cytotoxicity, gene silencing and uptake efficiency-were also assessed. The SAXS data show that when the pH was lowered to 5.5 the structure of the lipoplexes did not change, thus indicating that the acidic conditions of the endosome were not a significant factor in the release of siRNA from the cationic lipidic carriers.

Analytical approaches to study domain formation in biomimetic membranes.

Domains in biological membranes are linked to a range of biochemical life functions and thus understanding the fundamental physico-chemical drivers of domain formation is one of the key problems of biophysics and chemical biology. The phospholipid bilayer that is the structural basis of the biomembrane is a complex multicomponent mixture, and hence domain formation may be the result of thermodynamic phase equilibria, or specific sequestration of certain lipids; possibly both. There are several obstacles in the way of studying domains and thermodynamic phases in biomembranes: the complexity of the lipid mixture, the two dimensional nature of the membrane and the variety of superstructures the lipid membrane can fold into. Complexity is addressed by the introduction of biomimetic membranes, simplified mixtures of synthetic lipids. Most studies of lipid phase equilibria have been conducted using a biomimetic membrane. This review is intended to address the challenges posed to analytical methodology by the membrane dimensions, while also discussing the question of the reference state. Four key methods are assessed for their strengths and weaknesses in identifying domains and thermodynamic phases in membranes: differential scanning calorimetry, fluorescence microscopy, atomic force microscopy and quartz crystal microbalance. It is demonstrated that, while these methods provide complementary information and thus should be used in tandem, quartz crystal microbalance based nano-viscosimetry measurements offer a breakthrough in measuring phase transition temperatures, and allow the compilation of phase diagrams, of single bilayers of lipid mixtures. By comparing the structural phases of the lipids used for the different methods, it is also shown that the membrane curvature in vesicular lipid samples inhibits the formation of domains which are only observed in flat lamellar membranes, or giant unilamellar liposomes where the role of curvature is negligible.

Design Principles of Peptide Based Self-Assembled Nanomaterials.

The ability to design functionalized peptide nanostructures for specific applications is tied to the ability of controlling the morphologies of the self-assembled superstructures. That, in turn, is based on a thorough understanding of the structural and environmental factors affecting self-assembly. The aim of designing self-assembling nanostructures of controlled geometries is achieved via a combination of directional and non-directional second order interactions. If the interactions are distributed in a geometrically defined way, a specific and selective supramolecular self-assembly motif is the result. In this chapter we detail the role of non-covalent interactions on the self-assembly of peptides; we will also discuss different types of peptide building blocks and design rules for engineering unnatural supramolecular structures.

Cholesterol Rich Domains Identified in Unilamellar Supported Biomimetic Membranes via Nano-Viscosity Measurements.

Understanding the distribution of cholesterol in phospholipid membranes is of key importance in membrane biophysics, primarily since cholesterol enriched regions, rafts, are known to play a special role in protein function. In this work, quartz crystal microbalance with dissipation (QCM)-based viscosity measurements were used to study cholesterol-induced domain formation in partially suspended single bilayer membranes. 1,2-Dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and its mixtures with different amounts of cholesterol were studied. QCM temperature ramping experiments identified domains of different phase transition temperatures in the mixed membranes. The phase transition of DMPC shifted from 23.4 °C toward lower temperatures with increasing cholesterol content. A second, continuous but much broader, transition peak has been observed for the DMPC: cholesterol mixtures suggest that a separate cholesterol rich domain coexists with the DMPC rich domain. Importantly, the sharp DMC phase transition peak gradually diminished and eventually disappeared over 15% cholesterol content, suggesting that the cholesterol rich domain has a definite stoichiometry and once this cholesterol concentration is reached the DMPC-rich domain disappears. DSC control experiments do not show the second domain, suggesting that the phase separation only happens in nontensioned (flat) membranes.

Self-assembled nanomaterials based on beta (ß(3)) tetrapeptides.

ß(3)-amino acid based polypeptides offer a unique starting material for the design of self-assembled nanostructures such as fibres and hierarchical dendritic assemblies, due to their well-defined helical geometry in which the peptide side chains align at 120° due to the 3.0-3.1 residue pitch of the helix. In a previous work we have described the head-to-tail self-assembly of N-terminal acetylated ß(3)-peptides into infinite helical nanorods that was achieved by designing a bioinspired supramolecular self-assembly motif. Here we describe the effect of consecutively more polar side chains on the self-assembly characteristics of ß(3)-tetrapeptides Ac-ß (3)Ala-ß(3)Leu-ß(3)Ile-ß(3)Ala (Ac-ß(3)[ALIA]), Ac-ß(3)Ser-ß(3)Leu-ß(3)Ile-ß(3)Ala (Ac-ß(3)[SLIA]) and Ac-ß (3)Lys-ß (3)Leu-ß(3)Ile-ß (3)Glu (Ac-ß(3)[KLIE]). ß(3)-tetrapeptides complete 1 1/3 turns of the helix: thus in the oligomeric form the side chain positions shift 120° with each added monomer, forming a regular periodic pattern along the nanorod. Dynamic light scattering (DLS) measurements confirmed that these peptides self-assemble even in highly polar solvents such as water and DMSO, while diffusion-ordered NMR spectroscopy revealed the presence of a substantial monomeric population. Temperature dependence of the size distribution in DLS measurements suggests a dynamic equilibrium between monomers and oligomers. Solution casting produced distinct fibrillar deposits after evaporating the solvent. In the case of the apolar Ac-ß(3)[ALIA] the longitudinal helix morphology gives rise to geometrically defined (∼70°) junctions between fibres, forming a mesh that opens up possibilities for applications e.g. in tissue scaffolding. The deposits of polar Ac-ß(3)[SLIA] and Ac-ß(3)[KLIE] exhibit fibres in regular parallel alignment over surface areas in the order of 10 µm.

Structural analysis of bioinspired nano materials with synchrotron far IR spectroscopy.

Bioinspired fibres and hierarchical nano-materials are based on the self-assembly of organic building blocks such as polypeptides. Confirming the core structure of such materials is often challenging as they lack the long-range order required by crystallographic methods. Far-IR spectroscopy characterizes the vibrational modes of large molecular units. These vibrational modes are very sensitive to angle strain and second order interactions such as hydrogen bonding. As such, far-IR spectra hold information about the secondary structure and interactions of large biomolecules. Here we analyze the far-IR vibrational spectra of fibrous nano-materials based on three isomeric unnatural tripeptides, Ac-ß(3)Leu-ß(3)Ile-ß(3)Ala, Ac-ß(3)Ile-ß(3)Ala-ß(3)Leu, and Ac-ß(3)Ala-ß(3)Leu-ß(3)Ile. These peptides have well described self-assembly characteristics, forming one-dimensional nanorods that impose tight conformational constraints on the constituent molecules. The synchrotron far-IR spectroscopic results were interpreted by using density functional theory (DFT) modelling based vibrational analysis. The sensitivity of the spectra to peptide conformation was assessed by comparing the experimental spectra with DFT predictions. In high dielectric implicit solvent, intramolecular hydrogen-bonding is inhibited and thus the energy minimized peptide structure remains close to the 14-helix folding characteristic of substituted ß(3)-peptides, giving good agreement between the experimental and predicted vibration spectra. In contrast, energy minimization in vacuum alters the peptide conformation leading to intramolecular hydrogen bonds, and hence the predicted vibration spectra do not agree with the experimental data. Therefore, our results demonstrate the ability of far-IR spectroscopy to identify correct structural predictions and thus open the way for using far-IR spectroscopy for the characterization and structural analysis of bioinspired nano-materials and potentially their interactions with surfaces, ionic environments and other biomolecules. Far-IR structural analysis is particularly powerful in case of one- and two-dimensional materials such as fibres, hydrogels and thin layers where standard crystallographic analysis is not available.

Labeling phospholipid membranes with lipid mimetic luminescent metal complexes.

Lipid-mimetic metallosurfactant based luminophores are promising candidates for labeling phospholipid membranes without altering their biophysical characteristics. The metallosurfactants studied exhibit high structural and physicochemical similarity to phospholipid molecules, designed to incorporate into the membrane structure without the need for covalent attachment to a lipid molecule. In this work, two lipid-mimetic phosphorescent metal complexes are described: [Ru(bpy)2(dn-bpy)](2+) and [Ir(ppy)2(dn-bpy)](+) where bpy is 2,2'-bipyridine, dn-bpy is 4,4'-dinonyl-2,2'-bipyridine and ppy is 2-phenylpyridine. Apart from being lipid-mimetic in size, shape and physical properties, both complexes exhibit intense photoluminescence and enhanced photostability compared with conventional organic fluorophores, allowing for prolonged observation. Moreover, the large Stokes shift and long luminescence lifetime associated with these complexes make them more suitable for spectroscopic studies. The complexes are easily incorporated into dimyristoil-phosphatidyl-choline (DMPC) liposomes by mixing in the organic solvent phase. DLS reveals the labeled membranes form liposomes of similar size to that of neat DMPC membrane. Synchrotron Small-Angle X-ray Scattering (SAXS) measurements confirmed that up to 5% of either complex could be incorporated into DMPC membranes without producing any structural changes in the membrane. Fluorescence microscopy reveals that 0.5% label content is sufficient for imaging. Atomic Force Microscopic imaging confirms that liposomes of the labeled bilayers on a mica surface can fuse into a flat lamellar membrane that is morphologically identical to neat lipid membranes. These results demonstrate the potential of such lipid-mimetic luminescent metal complexes as a new class of labels for imaging lipid membranes.

Viscoelastic changes measured in partially suspended single bilayer membranes.

For studies involving biomimetic phospholipid membrane systems, such as membrane-protein interactions, it is crucial that the supported membrane is biomimetic in its physical properties as well as in its composition. Two often overlooked aspects of biomimicry are the need for unrestrained lipid mobility, reflected in the viscoelastic properties of the membrane, and sufficient space between the membrane and the support for the insertion of transmembrane proteins. Here we show for a series of DMPC-based membranes that a partially suspended single bilayer membrane can be formed on functionalized gold surface without tethering. These membranes exhibit sufficient freedom of motion to represent the viscoelastic properties of a free lamellar bilayer membrane as demonstrated by determining the phase transition temperatures of these single bilayer membranes from the viscosity change upon chain melting using the dissipation signal of a quartz crystal microbalance (QCM-D). Atomic force microscopy imaging confirmed confluent, smooth membrane coverage of the QCM-D sensor that completely obscured the roughness of the sputtered gold surface. High-force AFM imaging was able to push membrane patches into the valleys of the gold morphology, confirming the inherently suspended nature of the MPA supported membrane. We show that the correlation between frequency and dissipation changes in the QCM-D sensograms is a sensitive indicator of the morphology of the membrane.

Computational assessment of the radical scavenging activity of cleomiscosin.

Coumarinolignans such as cleomiscosin A (CMA), cleomiscosin B (CMB), and cleomiscosin C (CMC) are secondary metabolites that were isolated from diverse plant species. Cleomiscosins (CMs) have numerous interesting biological activities, including noteworthy cytotoxicity of cancer cell lines along with hepatoprotective and assumed antioxidant activities. In this present study, the antioxidant properties of three cleomiscosins were investigated with a focus on the structure-activity relationship using thermodynamic and kinetic calculations with the M06-2X/6-311++G(d,p) method. The results show that CMs, including CMA, CMB, and CMC, are weak antioxidants in apolar environments, with k overall of 7.52 × 102 to 6.28 × 104 M-1 s-1 for the HOO˙ radical scavenging reaction in the gas phase and 3.47 × 102 to 6.44 × 104 M-1 s-1 in pentyl ethanoate. Remarkably, the difference in the fusion of phenylpropanoid structure with coumarin via two ortho-hydroxyl groups (CMA and CMB) does not cause any noticeable effect on their antioxidant activity, while the presence of a methoxy substitute on the aromatic ring of phenylpropanoid units (CMC) increases the reaction rate to about 61 to 84 times faster than that of CMA. In contrast, the studied CMs exhibit a good antioxidant capacity in polar environments, with a k overall range from 4.03 × 107 to 8.66 × 107 M-1 s-1, 102-103 times faster than that of Trolox, equal to that of ascorbic acid and resveratrol. The angular fusion of the phenylpropanoid and coumarin structures, as well as the methoxy substitution on the aromatic ring of the phenylpropanoid unit of the studied CMs, do not have any considerable effect on their antioxidant activity under the studied conditions.

Applying Machine Learning for Antibiotic Development and Prediction of Microbial Resistance.

Antimicrobial resistance (AMR) poses a serious threat to human health worldwide. It is now more challenging than ever to introduce a potent antibiotic to the market considering rapid emergence of antimicrobial resistance, surpassing the rate of antibiotic drug discovery. Hence, new approaches need to be developed to accelerate the rate of drug discovery process and meet the demands for new antibiotics, while reducing the cost of their development. Machine learning holds immense promise of becoming a useful tool, especially since in the last two decades, exponential growth has occurred in computational power and biological big data analytics. Recent advancements in machine learning algorithms for drug discovery have provided significant clues for potential antibiotic classes. Apart from discovery of new scaffolds, the machine learning protocols will significantly impact prediction of AMR patterns and drug metabolism. In this review, we outline power of machine learning in antibiotic drug discovery, metabolic fate, and AMR prediction to support researchers engaged and interested in this field.

The radical scavenging activity of monocaffeoylquinic acids: the role of neighboring hydroxyl groups and pH levels.

Caffeoylquinic acids (CQAs) are well-known antioxidants. However, a key aspect of their radical scavenging activity - the mechanism of action - has not been addressed in detail thus far. Here we report on a computational study of the mechanism of activity of CQAs in scavenging hydroperoxyl radicals. In water at physiological pH, the CQAs demonstrated ≈ 104 times higher HOO˙ antiradical activity than in lipid medium (k(lipid) ≈ 104 M-1 s-1). The activity in the aqueous solution was determined by the hydrogen transfer mechanism of the adjacent hydroxyl group (O6'-H) of the dianion states (Γ = 93.2-95.2%), while the single electron transfer reaction of these species contributed 4.8-6.8% to the total rate constants. The kinetics estimated by the calculations are consistent with experimental findings in water (pH = 7.5), yielding a kcalculated/kexperimental = 2.4, reinforcing the reliability and precision of the computational method and demonstrating its utility for evaluating radical reactions in silico. The results also revealed the pH dependence of the HOO˙ scavenging activity of the CQAs; activity was comparable for all compounds below pH 3, however at higher pH values 5CQA reacted with the HOO˙ with lower activity than 3CQA or 4CQA. It was also found that CQAs are less active than Trolox below pH 4.7, however over pH 5.0 they showed higher activity than the reference. The CQAs had the best HOO˙ antiradical activity at pH values between 5.0 and 8.6. Therefore, in the physiological environment, the hydroperoxyl antiradical capacity of CQAs exhibits similarity to renowned natural antioxidants including resveratrol, ascorbic acid, and Trolox.

Synthesis and computational evaluation of the antioxidant activity of pyrrolo[2,3-b]quinoxaline derivatives.

Pyrrolo[2,3-b]quinoxaline derivatives are known to possess antioxidant, anticancer, and antibacterial properties. Here we report the successful synthesis of five derivatives of 3-hydroxy-3-pyrroline-2-one through substitution. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay was employed to evaluate the antioxidant activity of the compounds. Out of these, ethyl 1,2-diphenyl-1H-pyrrolo[2,3-b]quinoxaline-3-carboxylate (3a) demonstrated the greatest potential as a radical scavenger. Thermodynamic and kinetic calculations of the radical scavenging activity indicated that 3a exhibited HO˙ radical scavenging activity with the overall rate constant of 8.56 × 108 M-1 s-1 in pentyl ethanoate; however, it was incapable of scavenging hydroperoxyl radicals in nonpolar media. In non-polar environments, the hydroxyl radical scavenging capability of 3a is fairly similar to that of reference antioxidants such as Trolox, melatonin, indole-3-carbinol, and gallic acid. Hence, in the physiological lipid environment, 3a holds promise as a scavenger of HO˙ radicals.