Antibacterial Small Molecules That Potently Inhibit Staphylococcus aureus Lipoteichoic Acid Biosynthesis

The rise of antibiotic resistance, especially in Staphylococcus aureus, and the increasing death rate due to multiresistant bacteria have been well documented. The need for new chemical entities and/or the identification of novel targets for antibacterial drug development is high. Lipoteichoic acid (LTA), a membrane-attached anionic polymer, is important for the growth and virulence of many Gram-positive bacteria, and interest has been high in the discovery of LTA biosynthesis inhibitors. Thus far, only a handful of LTA biosynthesis inhibitors have been described with moderate (MIC=5.34 μg mL⁻¹) to low (MIC=1024 μg mL⁻¹) activities against S. aureus. Herein we describe the identification of novel compounds that potently inhibit LTA biosynthesis in S. aureus, displaying impressive antibacterial activities (MIC as low as 0.25 μg mL⁻¹) against methicillin-resistant S. aureus (MRSA). Under similar in vitro assay conditions, these compounds are 4-fold more potent than vancomycin and 8-fold more potent than linezolid against MRSA.     

Creating new water-soluble Iso- and n-fatty acid arginate hydrochloride with antimicrobial properties

Typically, short- and long-chain lipids from oils exhibit different antimicrobial activities and therefore have been used in agriculture and aquaculture, biomedical therapeutic and antibacterial fields. However, these fatty acids have limitations in terms of bioactive efficacy, thermostability and aqueous solubility. In this study, water-soluble iso-fatty acid arginate hydrochloride derivatives with antimicrobial properties were produced by introducing branched (iso-) chain and other linear- (n-) chain fatty acids to the “arginine” amino acid molecule. The two-step synthetic route was straightforward and provided an efficient 88% and 76% product yields for ethyl n-oleoyl arginate hydrochloride and ethyl iso-oleoyl arginate hydrochloride, respectively. ATR-FT-IR, NMR, and LC-MS-Q-TOF techniques were used to thoroughly characterize and confirm the products. These arginate products had strong antimicrobial activities against Listeria innucua, a Gram-positive bacterium with minimum inhibitory concentrations and minimum bactericidal concentrations ranging from 1.8 µg mL⁻¹ to 29.1 µg mL⁻¹. Therefore, the study demonstrated the development of a novel class of antimicrobial compounds from iso-fatty acids and arginates.     

Free Fatty Acids Reduction in Waste Cooking Oil by Rhodosporidium toruloides and Simultaneous Carotenoids,Lipids, and PAL Enzyme Production in a Two-Phase Culture System

Waste cooking oil (WCO) is a problematic waste product that contains free fatty acids (FFAs), preventing it from being valorized easily as biodiesel and poses an environmental hazard if incorrectly disposed. The use of WCO as a carbon source for Rhodosporidium toruloides (R. toruloides) using a two-phase culture system is developed. The normal growth of R. toruloides when cultured in WCO (OD₆₀₀ 52) reveals its ability to use a hydrophobic substrate as the carbon source compared to glucose (OD₆₀₀ 51.9). Interestingly, the extracellular lipase activity when R. toruloides is grown on WCO is 14.4 U mL⁻¹ compared to when grown on glucose (2.4 U mL⁻¹). Additionally, FFA levels in WCO are reduced from 2% to 0.2% at end of fermentation, suggesting that R. toruloides can consume FFA. Furthermore, higher yield of beneficial products: β-carotene (4.57 µg mL⁻¹), torularhodin (4.2 µg mL⁻¹), fatty acids (1 mg mL⁻¹), and phenylalanine ammonia-lyase (PAL) enzyme (0.12 µmol mg⁻¹) are produced when WCO is the carbon source, compared to glucose (4.1 µg mL⁻¹ β-carotene, 3.0 µg mL⁻¹ torularhodin, 1 mg mL⁻¹ of fatty acids, and 0.096 µmol mg⁻¹ PAL enzyme). This is a first study that shows R. toruloides can grow on hydrophobic carbon source.     

Physicochemical,Antioxidative, and Sensory Properties of Refined Rapeseed Oils

Physicochemical characteristics, antioxidant capacity (AC), and sensory quality of rapeseed oils available on the Polish market were analyzed and compared. The fatty acid composition (saturated fatty acids = 6.91–7.58%, monounsaturated fatty acids = 64.14–66.14%, and polyunsaturated fatty acids = 27.22–30.17%), color (T420 = 54.5–83.8%), amounts of free fatty acids (0.02–0.07%), primary (PV = 0.04–2.04 meq O₂ kg⁻¹) and secondary (AV = 1.02–3.21) oxidation products, phosphorus (0.38–1.62 mg kg⁻¹), chlorophyll (0.002–0.068 mg kg⁻¹), and polycyclic aromatic hydrocarbons (Σ4PAH = 0.00–2.50 μg kg⁻¹) in the commercial rapeseed oils meet the requirements of the European Food Regulation and Codex Alimentarius standards. Moreover, total phenolic content (TPC = 40.3–467.9 mg SA kg⁻¹) in the studied oils significantly differs from each other. However, the AC of rapeseed oils was analyzed using the novel iron oxide nanoparticle-based (IONP = 5552.1 − 18,510.2 μmol TE/100 g) method and the modified ferric reducing antioxidant power (FRAP = 55.7–280.3 μmol TE/100 g), cupric reducing AC (CUPRAC = 79.6–784.0 μmol TE/100 g), 2,2-diphenyl-1-picrylhydrazyl (DPPH = 185.7–516.7 μmol TE/100 g), and 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS = 465.6–2142.6 μmol TE/100 g) assays. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were applied for discrimination of the refined rapeseed oils based on fatty acid composition, physicochemical parameters, AC, and sensory properties.     

Novel 1,4‐Substituted‐1,2,3‐Triazoles as Antitubercular Agents

Tuberculosis (TB) remains a pressing unmet medical need, particularly with the emergence of multidrug‐resistant and extensively drug‐resistant tuberculosis. Here, a series of 1,4‐substituted‐1,2,3‐triazoles have been synthesized and evaluated as potential antitubercular agents. These compounds were assembled via click chemistry in high crude purity and in moderate to high yield. Of the compounds tested, 12 compounds showed promising antitubercular activity with six possessing minimum inhibitory concentration (MIC) values <10 μg mL^?1, and total selectivity for Mycobacterium tuberculosis (Mtb) growth inhibition. A second set of 21 compounds bearing variations on ring C were synthesized and evaluated. This second library gave an additional six compounds displaying MIC values ≤10 μg mL^?1 and total selectivity for Mtb growth inhibition. These compounds serve as an excellent starting point for further development of antitubercular therapies.     

C2 Arylated Benzo[b]thiophene Derivatives as Staphylococcus aureus NorA Efflux Pump Inhibitors

An innovative and straightforward synthesis of second‐generation 2‐arylbenzo[b]thiophenes as structural analogues of INF55 and the first generation of our laboratory‐made molecules was developed. The synthesis of C2‐arylated benzo[b]thiophene derivatives was achieved through a method involving direct arylation, followed by simple structural modifications. Among the 34 compounds tested, two of them were potent NorA pump inhibitors, which led to a 16‐fold decrease in the ciprofloxacin minimum inhibitory concentration (MIC) against the SA‐1199B strain at concentrations of 0.25 and 0.5 μg mL^?1 (1 and 1.5 μm , respectively). This is a promising result relative to that obtained for reserpine (MIC=20 μg mL^?1), a reference compound amongst NorA pump inhibitors. These molecules thus represent promising candidates to be used in combination with ciprofloxacin against fluoroquinolone‐resistant strains.     

Impact of Tetracationic Calix[4]arene Conformation—from Conic Structure to Expanded Bolaform—on Their Antibacterial and Antimycobacterial Activities

The four possible conformers of a new tetrakisguanidino calix[4]arene thought to interact deleteriously with bacterial membranes have been synthesized, characterized, and evaluated for their in vitro cytotoxicity and antibacterial activity against various reference Gram-negative and Gram-positive bacteria, as well as Mycobacterium tuberculosis. It appears that reversal of at least one phenolic unit results in clear increases in their activities. This can be attributed to the evolution towards bolaform structures, which are able to interact more deeply with the bacterial membrane. Indeed, the 1,3-alternate conformer 16 exhibits the best antibacterial activity (MIC<1.0 μg mL⁻¹ on Staphylococcus aureus). Moreover, 16 displays very good antibacterial activities against an isoniazid-resistant strain of M. tuberculosis (MIC=1.2 μg mL⁻¹), associated with the lowest cytotoxicity, thus making it the most potent compound of the series; this could open new ways of research in the field of anti-infective drug development to meet the huge current demand.     

Biological Evaluation of Potent Triclosan‐Derived Inhibitors of the Enoyl–Acyl Carrier Protein Reductase InhA in Drug‐Sensitive and Drug‐Resistant Strains of Mycobacterium tuberculosis

New triclosan (TRC) analogues were evaluated for their activity against the enoyl–acyl carrier protein reductase InhA in Mycobacterium tuberculosis (Mtb). TRC is a well‐known inhibitor of InhA, and specific modifications to its positions 5 and 4′ afforded 27 derivatives; of these compounds, seven derivatives showed improved potency over that of TRC. These analogues were active against both drug‐susceptible and drug‐resistant Mtb strains. The most active compound in this series, 4‐(n‐butyl)‐1,2,3‐triazolyl TRC derivative 3 , had an MIC value of 0.6 μg mL^?1 (1.5 μM ) against wild‐type Mtb. At a concentration equal to its MIC, this compound inhibited purified InhA by 98 %, and showed an IC₅₀ value of 90 nM . Compound 3 and the 5‐methylisoxazole‐modified TRC 14 were able to inhibit the biosynthesis of mycolic acids. Furthermore, mc²4914, an Mtb strain overexpressing inhA, was found to be less susceptible to compounds 3 and 14 , supporting the notion that InhA is the likely molecular target of the TRC derivatives presented herein.     

Influence of different reactor types on Nannochloropsis oculata microalgae culture for lipids and fatty acid production

Greenhouse gases emitted into the atmosphere by burning of fossil fuels cause global warming. One option is obtaining biodiesel. Nannochloropsis oculata was cultured under different light intensities and reactors at 25°C for 21 days with f/2 medium to assess their effects on cell density, lipid, and fatty acids (FAs). N. oculata improved cell density on fed-batch glass tubular reactor (7 L) at 200 μmol E m⁻² s⁻¹, yielding 3.5 × 10⁸ cells ml⁻¹, followed by fed-batch Erlenmeyer flask (1 L) at 650 μmol E m⁻² s⁻¹ with 1.7 × 10⁸ cells ml⁻¹. The highest total lipid contents (% g lipid × g dry biomass⁻¹) were 44.4 ± 0.8% for the reactor (1 L) at 650 μmol E m⁻² s⁻¹ and 35.2 ± 0.2% for the tubular reactor (7 L) at 200 μmol E m⁻² s⁻¹, until twice as high compared with the control culture (Erlenmeyer flask 1 L, 80 μmol E m⁻² s⁻¹) with 21.2 ± 1%. Comparing the total lipid content at 200 μmol E m⁻² s⁻¹, tubular reactor (7 L) and reactor 1 L achieved 35.2 ± 0.2% and 28.3 ± 1%, respectively, indicating the effect of shape reactor. The FAs were affected by high light intensity, decreasing SFAs to 2.5%, and increased monounsaturated fatty acids + polyunsaturated fatty acids to 2.5%. PUFAs (20:5n-3) and (20:4n-3) were affected by reactor shape, decreasing by half in the tubular reactor. In the best culture, fed-batch tubular reactor (7 L) at 200 μmol E m⁻² s⁻¹ contains major FAs (16:0; 38.06 ± 0.16%), (16:1n-7; 30.74 ± 0.58%), and (18:1n-9; 17.15 ± 0.91%).     

Salimyxins and Enhygrolides: Antibiotic,Sponge‐Related Metabolites from the Obligate Marine Myxobacterium Enhygromyxa salina

Unlike their terrestrial counterparts, marine myxobacteria are hardly investigated for their secondary metabolites. This study describes three new compounds ( 1 – 3 ), named salimyxins and enhygrolides, obtained from the obligate marine myxobacterium Enhygromyxa salina. These are the first natural products obtained from Enhygromyxa species. Their structures were elucidated by spectroscopic analysis, including NMR and CD spectroscopy. Enhygrolides are closely related to the nostoclides, which were initially isolated from a cyanobacterium of the genus Nostoc. The salimyxins, representing structurally most unusual degraded sterols, are close to identical to demethylincisterol from the sponge Homaxinella sp. Salimyxin B and enhygrolide A inhibit the growth of the Gram‐positive bacterium Arthrobacter cristallopoietes (MIC salimyxin B, 8 μg mL^?1; enhygrolide A, 4 μg mL^?1).     

Preparation,characterization, and antibacterial activity of cationic nanopolystyrenes

Cationic nanopolystyrenes (CNPSs) were prepared by the polymerization of styrene with a cationic polymerizable emulsifier, N-(2-(methacryloyloxy)ethyl)-N,N-dimethyltetradecane-1-ammonium bromide (MDTB), in the presence of 2,2′-azobis(2-methylpropionamidine) dihydrochloride (APPH). The polymerization kinetics was investigated through monomer conversion versus polymerization time. ¹H NMR spectra and FTIR spectra were used to confirm the structures of MDTB and nanopolymers. The particle size, ζ-potential, molecular weight, and thermal stability were also investigated by Zetasizer, gel permeation chromatography, and thermogravimetric analysis, respectively. The antibacterial activities of the obtained products were evaluated by minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against Staphylococcus aureus ATCC 29913 (S. aureus, a Gram-positive bacterium) and Escherichia coli ATCC 25922 (E. coli, a Gram-negative bacterium). The results indicated that the polymerization has the characteristics of microemulsion polymerization, all nanopolystyrenes are cationic and can be stored in the form of a microemulsion in aqueous phase, CNPSs present stable chemical structure at 190 °C, and their MIC and MBC reach 0.0625 and 0.125 mg mL⁻¹ for E. coli, 0.0325 and 0.0625 mg mL⁻¹ for S. aureus, respectively. Therefore, CNPSs are a promising new type of polymer antimicrobial agent, which can be used in the development of antimicrobial polymer materials and their products. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020 , 137, 48405.     

Antioxidant and Biological Activities of Novel Structured Monoacylglycerol Derivatives with Phenolic Acids

Novel structured monoacylglycerol (MAG)-based phenolic lipids are synthesized from11-bromoundecanoic acid, phenolic acids, and solketal. Selected phenolic acids namely 4-hydroxy benzoic, vanillic, syringic, cinnamic, p-coumaric, sinapic, 4-fluorocinnamic, 4-hydroxyphenyl acetic acid, 3-(4-hydroxyphenyl) propanoic and dihydrocaffeic acids are employed for the synthesis of ten novel MAG-based phenolic lipids. The synthesized phenolic lipids are characterized by FT-IR, NMR, and mass spectra analysis. All the compounds are evaluated for antioxidant, antimicrobial, and cytotoxic activities. MAG derivative 8g of sinapic acid exhibits excellent antioxidant activity in both DPPH assay and inhibition of lipid oxidation assay. MAG derivative 8f bearing p-coumaric acid shows good antimicrobial activity against both Gram-positive and Gram-negative bacterial strains with a minimum inhibitory concentration (MIC) value of 6.25 µm mL⁻¹. All the synthesized compounds are found to exhibit cytotoxicity against B16, DU145, and CHO cell lines, while sinapic and p-coumaric acid derivatives exhibit better activities compared to other derivatives.     

Isoprenoid Biosynthesis Inhibitors Targeting Bacterial Cell Growth

We synthesized potential inhibitors of farnesyl diphosphate synthase (FPPS), undecaprenyl diphosphate synthase (UPPS), or undecaprenyl diphosphate phosphatase (UPPP), and tested them in bacterial cell growth and enzyme inhibition assays. The most active compounds were found to be bisphosphonates with electron‐withdrawing aryl‐alkyl side chains which inhibited the growth of Gram‐negative bacteria (Acinetobacter baumannii, Klebsiella pneumoniae, Escherichia coli, and Pseudomonas aeruginosa) at ～1–4 μg mL^?1 levels. They were found to be potent inhibitors of FPPS; cell growth was partially “rescued” by the addition of farnesol or overexpression of FPPS, and there was synergistic activity with known isoprenoid biosynthesis pathway inhibitors. Lipophilic hydroxyalkyl phosphonic acids inhibited UPPS and UPPP at micromolar levels; they were active (～2–6 μg mL^?1) against Gram‐positive but not Gram‐negative organisms, and again exhibited synergistic activity with cell wall biosynthesis inhibitors, but only indifferent effects with other inhibitors. The results are of interest because they describe novel inhibitors of FPPS, UPPS, and UPPP with cell growth inhibitory activities as low as ～1–2 μg mL^?1.     

Synthesis,Characterization, and Antimicrobial Activity of Silver(I) and Copper(II) Complexes of Phosphate Derivatives of Pyridine And Benzimidazole

Two silver(I) complexes—{[Ag(4‐pmOpe)]NO₃}_n and [Ag(2‐bimOpe)₂]NO₃—and three copper(II) complexes—[Cu₄Cl₆O(2‐bimOpe)₄], [CuCl₂(4‐pmOpe)₂], and [CuCl₂(2‐bis(pm)Ope]—were synthesized by reaction of silver(I) nitrate or copper(II) chloride with phosphate derivatives of pyridine and benzimidazole, namely diethyl (pyridin‐4‐ylmethyl)phosphate (4‐pmOpe), 1H‐benzimidazol‐2‐ylmethyl diethyl phosphate (2‐bimOpe), and ethyl bis(pyridin‐2‐ylmethyl)phosphate (2‐bis(pm)Ope). These compounds were characterized by ¹H, ¹³C, and ³¹P NMR as well as IR spectroscopy, elemental analysis, and ESIMS spectrometry. Additionally, molecular and crystal structures of {[Ag(4‐pmOpe)]NO₃}_n and [Cu₄Cl₆O(2‐bimOpe)₄] were determined by single‐crystal X‐ray diffraction analysis. The antimicrobial profiles of synthesized complexes and free ligands against test organisms from the ATCC and clinical sources were determined. Silver(I) complexes showed good antimicrobial activities against Candida albicans strains (MIC values of ～19 μM ). [Ag(2‐bimOpe)₂]NO₃ was particularly active against Pseudomonas aeruginosa and methicillin‐resistant Staphylococcus epidermidis, with MIC values of ～5 and ～10 μM , respectively. Neither copper(II) complexes nor the free ligands inhibited the growth of test organisms at concentrations below 500 μg mL^?1.     

New Antimicrobial Hexapeptides: Synthesis,Antimicrobial Activities,Cytotoxicity, and Mechanistic Studies

Synthetic antimicrobial peptides have recently emerged as promising candidates against drug‐resistant pathogens. We identified a novel hexapeptide, Orn‐D ‐Trp‐D ‐Phe‐Ile‐D ‐Phe‐His(1‐Bzl)‐NH₂, which exhibits broad‐spectrum antifungal and antibacterial activity. A lead optimization was undertaken by conducting a full amino acid scan with various proteinogenic and non‐proteinogenic amino acids depending on the hydrophobic or positive‐charge character of residues at various positions along the sequence. The hexapeptide was also cyclized to study the correlation between the linear and cyclic structures and their respective antimicrobial activities. The synthesized peptides were found to be active against the fungus Candida albicans and Gram‐positive bacteria such as methicillin‐resistant Staphylococcus aureus and methicillin‐resistant Staphylococcus epidermidis, as well as the Gram‐negative bacterium Escherichia coli; MIC values for the most potent structures were in the range of 1–5 μg mL^?1 (IC₅₀ values in the range of 0.02–2 μg mL^?1). Most of the synthesized peptides showed no cytotoxic effects in an MTT assay up to the highest test concentration of 200 μg mL^?1. A tryptophan fluorescence quenching study was performed in the presence of negatively charged and zwitterionic model membranes, mimicking bacterial and mammalian membranes, respectively. The results of the fluorescence study demonstrate that the tested peptides are selective toward bacterial over mammalian cells; this is associated with a preferential interaction between the peptides and the negatively charged phospholipids of bacterial cells.     

Improving Cell‐Free Protein Synthesis through Genome Engineering of Escherichia coli Lacking Release Factor 1

Site‐specific incorporation of non‐standard amino acids (NSAAs) into proteins opens the way to novel biological insights and applications in biotechnology. Here, we describe the development of a high yielding cell‐free protein synthesis (CFPS) platform for NSAA incorporation from crude extracts of genomically recoded Escherichia coli lacking release factor 1. We used genome engineering to construct synthetic organisms that, upon cell lysis, lead to improved extract performance. We targeted five potential negative effectors to be disabled: the nuclease genes rna, rnb, csdA, mazF, and endA. Using our most productive extract from strain MCJ.559 (csdA^? endA^?), we synthesized 550±40 μg mL^?1 of modified superfolder green fluorescent protein containing p‐acetyl‐L ‐phenylalanine. This yield was increased to ～1300 μg mL^?1 when using a semicontinuous method. Our work has implications for using whole genome editing for CFPS strain development, expanding the chemistry of biological systems, and cell‐free synthetic biology.     

Functionalization of halloysite nanotubes with polyethyleneimine and various ionic liquid forms with antimicrobial activity

Halloysite nanotube (HNT), a natural clay, was modified with branched polyethyleneimine (PEI) to form PEI-HNT using epichlorohydrin (ECH) as coupling agent, then protonated with HCl to obtain H-PEI-HNTs providing [NH₃]⁺[Cl]⁻ functionality for potential antimicrobial properties. Upon PEI modification, zeta potential value of HNTs was increased to +37.3 mV from −34.5 mV and to +41.1 mV for H-PEI-HNTs. Only 1.87 wt % H-element in HNT was increased to 3.03 wt % upon PEI modification along with newly generated elements of N and C at 2.99 and 9.93 wt %, respectively. Moreover, ionic liquid (IL) forms of HNTs with [NH₃]⁺[N(CN)₂]⁻, [NH₃]⁺[PF₆]⁻ and [NH₃]⁺[BF₄]⁻ functionality were generated via anion exchange of H-PEI-HNTs with sodium dicyanamide (SDC), ammonium hexafluorophosphate (AHFP), and sodium tetrafluoroborate (STFB). The antimicrobial properties of the modified, protonated, and IL forms of HNTs were determined via macro dilution, diffusion and agar screening tests against Escherichia coli ATCC 8739, Pseudomonas aeruginosa ATCC 10145, Bacillus subtilis ATCC 6633, Staphylococcus aureus ATCC 6538 strains, and Candida albicans ATCC 10231 strains. It was found that H-PEI-HNTs possesses potent antimicrobial effect compared with the other forms of HNTs with 2–4 mg mL⁻¹ MIC and 8–16 mg mL⁻¹ MBC values via the macro dilution method. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020 , 137, 48352.     

A New Gas Chromatography–Mass Spectrometry Method for Detecting and Quantifying Low Levels of Dimethyl Sulfate in Palm Oil-Based Sulfonated Methyl Esters

Dimethyl sulfate (DMS) is a potential carcinogen that may be formed during the production of α-sulfonated methyl ester (α-SME). To date, there have been no published analytical methods for quantification of DMS in palm oil-based α-SME. The objective of the method development was to establish a simple and reliable method for quantifying and monitoring DMS from palm oil-based α-SME. The sample preparation prior to analysis involved extracting DMS from α-SME using hexane. The gas chromatography–mass spectrometry (GC–MS) separation was performed on an Agilent MS-HP5 column (30 m × 320 μm × 0.25 μm). The MS signal was obtained in the selected ion monitoring (SIM) mode. The method was validated according to the International Conference on Harmonization (ICH) guidelines. The accuracy of the method was measured via percentage recoveries of DMS from spiked α-SME samples at spiking levels of (25, 125, 250, and 500) μg mL⁻¹. The percentage recoveries for all spiking levels were found to be in the range of 88.6–97.4%. The repeatability and interday reproducibility were found to be satisfactory as the relative standard deviations (RSD) were within 11%. The method also has a good linear relationship as indicated by the coefficient of determination of 0.9996 over the range of 1.0–30 μg mL⁻¹. The limits of detection (LOD) and quantification (LOQ) corresponded to 0.8 and 2.3 μg mL⁻¹, respectively. The developed method demonstrated sufficient linearity, intraday and interday precision, sensitivity, and accuracy for determination of DMS in palm oil-based α-SME.