Antibacterial and Antioxidant Activity of Synthetic Polyoxygenated Flavonoids

Flavonoids are an abundant class of naturally occurring compounds with broad biological activities, but their limited abundance in nature restricts their use in medicines and food additives. Here we present the synthesis and determination of the antibacterial and antioxidant activities of twenty-two structurally related flavonoids (five of which are new) by scientifically validated methods. Flavanones (FV1–FV11) had low inhibitory activity against the bacterial growth of MRSA 97-7. However, FV2 (C5,7,3′,4′ = OH) and FV6 (C5,7 = OH; C4′ = SCH3) had excellent bacterial growth inhibitory activity against Gram-negative E. coli (MIC = 25 µg/mL for both), while Chloramphenicol (MIC = 25 µg/mL) and FV1 (C5,7,3′ = OCH3; 4′ = OH) showed inhibitory activity against Gram-positive L. monocytogenes (MIC = 25 µg/mL). From the flavone series (FO1–FO11), FO2 (C5,7,3′,4′ = OH), FO3 (C5,7,4′ = OH; 3′ = OCH3), and FO5 (C5,7,4′ = OH) showed good inhibitory activity against Gram-positive MRSA 97-7 (MIC = 50, 12, and 50 µg/mL, respectively), with FO3 being more active than the positive control Vancomycin (MIC = 25 µg/mL). FO10 (C5,7= OH; 4′ = OCH3) showed high inhibitory activity against E. coli and L. monocytogenes (MIC = 25 and 15 µg/mL, respectively). These data add significantly to our knowledge of the structural requirements to combat these human pathogens. The positions and number of hydroxyl groups were key to the antibacterial and antioxidant activities

Prenylated Flavonoids with Potential Antimicrobial Activity: Synthesis, Biological Activity, and In Silico Study

Prenylated flavonoids are an important class of naturally occurring flavonoids with important biological activity, but their low abundance in nature limits their application in medicines. Here, we showed the hemisynthesis and the determination of various biological activities of seven prenylated flavonoids, named 7–13, with an emphasis on antimicrobial ones. Compounds 9, 11, and 12 showed inhibitory activity against human pathogenic fungi. Compounds 11, 12 (flavanones) and 13 (isoflavone) were the most active against clinical isolated Staphylococcus aureus MRSA, showing that structural requirements as prenylation at position C-6 or C-8 and OH at positions C-5, 7, and 40 are key to the antibacterial activity. The combination of 11 or 12 with commercial antibiotics synergistically enhanced the antibacterial activity of vancomycin, ciprofloxacin, and methicillin in a factor of 10 to 100 times against drug-resistant bacteria. Compound 11 combined with ciprofloxacin was able to decrease the levels of ROS generated by ciprofloxacin. According to docking results of S enantiomer of 11 with ATP-binding cassette transporter showed the most favorable binding energy; however, more studies are needed to support this result.Fil: Osorio, Mauricio. Universidad Técnica Federico Santa María, Departamento de Química. Chile.Fil: Caravajal, Marcela.Universidad Técnica Federico Santa María. Centro de Biotecnología CB-DAL. Chile.Fil: Vergara, Alejandra. Universidad Técnica Federico Santa María. Centro de Biotecnología CB-DAL. Chile.Fil: Butassi, Estefanía. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Farmacognosia. Argentina.Fil: Butassi, Estefanía. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Farmacognosia. Argentina.Fil: Zaccchiano, Susana. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Farmacognosia. Argentina.Fil: Mascayano, Carolina. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Ciencias del Ambiente. Chile.Fil: Montoya, Margarita. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología. Chile.Fil: Mejías, Sophia. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología. Chile.Fil: Cortez-San Martín, Marcelo. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología. Chile.Fil: Vásquez-Martínez, Yesseny. Universidad de Santiago de Chile. Facultad de Ciencias Médicas. Escuela de Medicina. Programa Centro de Investigaciones Biomédicas Aplicadas. Chil

Biotransformation of Natural and Synthetic Isoflavonoids by Two Recombinant Microbial Enzymes

Isolation and synthesis of isoflavonoids has become a frequent endeavor, due to their interesting biological activities. The introduction of hydroxyl groups into isoflavonoids by the use of enzymes represents an attractive alternative to conventional chemical synthesis. In this study, the capabilities of biphenyl-2,3-dioxygenase (BphA) and biphenyl-2,3-dihydrodiol 2,3-dehydrogenase (BphB) of Burkholderia sp. strain LB400 to biotransform 14 isoflavonoids synthesized in the laboratory were investigated by using recombinant Escherichia coli strains containing plasmid vectors expressing the bphA1A2A3A4 or bphA1A2A3A4B genes of strain LB400. The use of BphA and BphB allowed us to biotransform 7-hydroxy-8-methylisoflavone and 7-hydroxyisoflavone into 7,2′,3′-trihydroxy-8-methylisoflavone and 7,3′,4′-trihydroxyisoflavone, respectively. The compound 2′-fluoro-7-hydroxy-8-methylisoflavone was dihydroxylated by BphA at ortho-fluorinated and meta positions of ring B, with concomitant dehalogenation leading to 7,2′,3′,-trihydroxy-8-methylisoflavone. Daidzein (7,4′-dihydroxyisoflavone) was biotransformed by BphA, generating 7,2′,4′-trihydroxyisoflavone after dehydration. Biotransformation products were analyzed by gas chromatography-mass spectrometry and nuclear magnetic resonance techniques

Antimicrobial, anti-inflammatory and antioxidant activities of polyoxygenated chalcones

It was synthesized nine polyoxygenated chalcones with a potential and safe use as antioxidant, antimicrobial and anti-inflammatory therapies. Chalcones obtained by Claisen-Schmidt condensation were studied as antioxidant, inhibitors of human 5-lipoxygenase, antifungal, antibacterial and antibiotic resistance modifiers. Two chalcones with catecholic moieties were able to strongly decrease the minimum inhibitory concentration (MIC) of methicillin against methicillin-resistant Staphylococcus aureus, increase the antiradical activity and significantly inhibit the human 5-lipoxygenase. Only one of these chalcones was active synergistically with methicillin. Chalcones with methoxyl substituents at different positions displayed the best activities against Cryptococcus neoformans. Only one chalcone showed good activity against the plant pathogenic bacteria Pseudomonas syringae whose half maximal inhibitory concentration (IC50) value (2.5 µg mL-1) was similar to that observed with the antibiotic streptomycin (2.9 µg mL-1). These simple chalcones have safe potential uses in antioxidant, antimicrobial and anti-inflammatory therapies.Fil: Vásquez-Martínez, Yesseny A. Universidad de Santiago de Chile. Facultad de Ciencias Médicas. Programa Centro de Investigaciones Biomédicas Aplicadas; Chile.Fil: Osorio, Mauricio E. Universidad Técnica Federico Santa María. Departamento de Química. Laboratorio de Productos Naturales; Chile.Fil: San Martín, Diego A. Universidad Técnica Federico Santa María. Departamento de Química. Laboratorio de Productos Naturales; Chile.Fil: Carvajal, Marcela A. Universidad Técnica Federico Santa María. Centro de Biotecnología (CB-DAL); Chile.Fil: Vergara, Alejandra P. Universidad Técnica Federico Santa María. Centro de Biotecnología (CB-DAL); Chile.Fil: Sanchez, Elizabeth. Universidad Técnica Federico Santa María. Centro de Biotecnología (CB-DAL); Chile.Fil: Raimondi, Marcela. Universidad Nacional de Rosario. Facultad de Farmacia y Bioquímica. Área Farmacognosia; Argentina.Fil: Zacchino, Susana. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Área Farmacognosia; Argentina.Fil: Mascayano, Carolina. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Ciencias del Ambiente; Chile.Fil: Torrent, Claudia. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Ciencias del Ambiente; Chile.Fil: Cabezas, Francisco. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Ciencias del Ambiente; Chile.Fil: Mejias, Sophia. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología; Chile.Fil: Montoya, Margarita. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología; Chile.Fil: Cortez-San Martín, Marcelo. Universidad de Santiago de Chile. Facultad de Química y Biología. Departamento de Biología; Chile

Prenylated flavonoids with potential antimicrobial activity: synthesis, biological activity, and in silico study

Prenylated flavonoids are an important class of naturally occurring flavonoids with important biological activity, but their low abundance in nature limits their application in medicines. Here, we showed the hemisynthesis and the determination of various biological activities of seven prenylated flavonoids, named 7–13, with an emphasis on antimicrobial ones. Compounds 9, 11, and 12 showed inhibitory activity against human pathogenic fungi. Compounds 11, 12 (flavanones) and 13 (isoflavone) were the most active against clinical isolated Staphylococcus aureus MRSA, showing that structural requirements as prenylation at position C-6 or C-8 and OH at positions C-5, 7, and 4′ are key to the antibacterial activity. The combination of 11 or 12 with commercial antibiotics synergistically enhanced the antibacterial activity of vancomycin, ciprofloxacin, and methicillin in a factor of 10 to 100 times against drug-resistant bacteria. Compound 11 combined with ciprofloxacin was able to decrease the levels of ROS generated by ciprofloxacin. According to docking results of S enantiomer of 11 with ATP-binding cassette transporter showed the most favorable binding energy; however, more studies are needed to support this result.Fil: Osorio, Mauricio. Universidad Técnica Federico Santa María; ChileFil: Carvajal, Marcela. Universidad Técnica Federico Santa María; ChileFil: Vergara, Alejandra. Universidad Técnica Federico Santa María; ChileFil: Butassi, Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; Argentina. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Departamento de Química Orgánica. Área Farmacognosia; ArgentinaFil: Zacchino, Susana Alicia Stella. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Departamento de Química Orgánica. Área Farmacognosia; ArgentinaFil: Mascayano, Carolina. Universidad de Santiago de Chile; ChileFil: Montoya, Margarita. Universidad de Santiago de Chile; ChileFil: Mejías, Sophia. Universidad de Santiago de Chile; ChileFil: Cortés San Martín, Marcelo. Universidad de Santiago de Chile; ChileFil: Vásquez Martínez, Yesseny. Universidad de Santiago de Chile; Chil

Molecular and Phylogenomic Analysis of a Vancomycin Intermediate Resistance USA300LV Strain in Chile

Antimicrobial resistance is a major global health problem, and, among Gram-positive bacteria, methicillin-resistant Staphylococcus aureus (MRSA) represents a serious threat. MRSA causes a wide range of infections, including bacteremia, which, due to the limited use of β-lactams, is difficult to treat. This study aimed to analyze 51 MRSA isolates collected in 2018 from samples of patients with bacteremia from two hospitals of the Metropolitan Health Service of Santiago, Chile, both in their resistance profile and in the identification of virulence factors. In addition, genomic characterization was carried out by the WGS of an isolate that was shown to be the one of greatest concern (N°. 42) due to its intermediate resistance to vancomycin, multiple virulence factors and being classified as ST8 PVL-positive. In our study, most of the isolates turned out to be multidrug-resistant, but there are still therapeutic options, such as tetracycline, rifampicin, chloramphenicol and vancomycin, which are currently used for MRSA infections; however, 18% were PVL positive, which suggests greater virulence of these isolates. It was determined that isolate N°42 is grouped within the USA300-LV strains (ST8, PVL+, COMER+); however, it has been suggested that, in Chile, a complete displacement of the PVL-negative ST5 clone has not occurred

A Open One-Step RT-qPCR for SARS-CoV-2 detection

<p>Sequences of the plasmids needed to purify the required enzymes for a standardized One-Step open RT-qPCR protocol to detect SARS-CoV-2 RNA in clinical samples. Supplementary Information</p><p>Filetype GB</p&gt

One-Step RT-qPCR cycling conditions.

The COVID-19 pandemic has resulted in millions of deaths globally, and while several diagnostic systems were proposed, real-time reverse transcription polymerase chain reaction (RT-PCR) remains the gold standard. However, diagnostic reagents, including enzymes used in RT-PCR, are subject to centralized production models and intellectual property restrictions, which present a challenge for less developed countries. With the aim of generating a standardized One-Step open RT-qPCR protocol to detect SARS-CoV-2 RNA in clinical samples, we purified and tested recombinant enzymes and a non-proprietary buffer. The protocol utilized M-MLV RT and Taq DNA pol enzymes to perform a Taqman probe-based assay. Synthetic RNA samples were used to validate the One-Step RT-qPCR components, demonstrating sensitivity comparable to a commercial kit routinely employed in clinical settings for patient diagnosis. Further evaluation on 40 clinical samples (20 positive and 20 negative) confirmed its comparable diagnostic accuracy. This study represents a proof of concept for an open approach to developing diagnostic kits for viral infections and diseases, which could provide a cost-effective and accessible solution for less developed countries.</div

Purification of M-MLV RT, Taq DNA pol and Pfu-Sso7d enzymes.

SDS-PAGE of M-MLV RT (A) Taq DNA pol (B) and Pfu-Sso7d (C) purification. Samples were prepared by volume; 3 μL for pellet (P), clarified lysate (CL), and flowthrough (F); 12 μL for wash (W) and elution samples from the Ni-NTA and heparin purifications. The observed molecular weight of the purified proteins matches the one predicted based on their amino acid sequences: 80 kDa for M-MLV RT, 96 kDa for Taq DNA pol, and 100 kDa for Pfu-Sso7d. The His lane in the heparin purification panel in (B) and (C) corresponds to a pooled Ni-NTA purified protein sample before heparin purification. SDS-PAGE gels in (A) and (C) were made at 10% polyacrylamide (PA), whereas 12% PA was used for (B). (TIF)</p

CDC SARS-CoV-2 N1 and N2 probe-based One-Step RT-qPCR assays performed with synthetic RNA using homebrew M-MLV RT and Taq DNA pol.

(A-B) Representative amplification curves using N1 and N2 CDC-approved double-quenched probes. Each curve represents a specific dilution of SARS-CoV-2 synthetic N RNA used as template: 1.4 x 107 copies approximately (red line), 1.4 x 106 (yellow line), 1.4 x 105 (green line), 1.4 x 104 (light blue line), 1.4 x 103 (blue line), 1.4 x 102 (purple line), 1.4 x 101 (light brown line) and no template control (NTC, gray line). Amplification curves for synthetic N RNA from MERS-CoV (black dashed line) and SARS-CoV-1 (orange dashed line) are also indicated. Characteristic Cq values are indicated on the upper left side of each panel. N.d.: non-detected (no Cq reported).</p