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Int J Mol Sci ; 22(12)2021 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-34207373


A nucleic acid aptamer that specifically recognizes methicillin-resistant Staphylococcus aureus (MRSA) has been immobilized on magnetic nanoparticles to capture the target bacteria prior to mass spectrometry analysis. After the MRSA species were captured, they were further eluted from the nanoparticles and identified using matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS). The combination of aptamer-based capture/enrichment and MS analysis of microorganisms took advantage of the selectivity of both techniques and should enhance the accuracy of MRSA identification. The capture and elution efficiencies for MRSA were optimized by examining factors such as incubation time, temperature, and elution solvents. The aptamer-modified magnetic nanoparticles showed a capture rate of more than 90% under the optimized condition, whereas the capture rates were less than 11% for non-target bacteria. The as-prepared nanoparticles exhibited only a 5% decrease in the capture rate and a 9% decrease in the elution rate after 10 successive cycles of utilization. Most importantly, the aptamer-modified nanoparticles revealed an excellent selectivity towards MRSA in bacterial mixtures. The capture of MRSA at a concentration of 102 CFU/mL remained at a good percentage of 82% even when the other two species were at 104 times higher concentration (106 CFU/mL). Further, the eluted MRSA bacteria were successfully identified using MALDI mass spectrometry.

Aptâmeros de Nucleotídeos/química , Nanopartículas de Magnetita/química , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Staphylococcus aureus Resistente à Meticilina/citologia , Técnica de Seleção de Aptâmeros/métodos
Molecules ; 24(12)2019 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-31216770


Considering the importance of ultra-performance liquid chromatography-electrospray ionization-quadrupole time of flight-tandem mass spectrometry (UPLC-ESI-QTOF-MS/MS) hyphenated techniques for analysis of secondary metabolites from crude extracts, the present study was aimed at identification of secondary metabolites in acetone extract of the lichen Usnea longissima. From our study, 19 compounds were tentatively identified through comparison of exact molecular masses from their MS/MS spectra, mass fragmentation studies and comparison with literature data. In addition, potent cytotoxic activity of U. longissima extract prompted us to isolate four compounds, 18R-hydroxy-dihydroalloprotolichesterinic acid (19), neuropogolic acid (20), barbatic acid (21), and usnic acid (22) from this extract which were adequately identified through mass spectrometry and NMR spectroscopy. All four compounds displayed cytotoxic activity. Barbatic acid (21) manifested doxorubicin equivalent activity against A549 lung cancer cell line with IC50 of 1.78 µM and strong G0/G1 accumulation of cells. Poly ADP-ribose polymerase (PARP) cleavage confirmed that it induced cytotoxic activity via apoptosis. Finally, our work has discerned the depside, barbatic acid (21) from crude extract as a candidate anti-cancer molecule, which induces cell death by stepping up apoptosis.

Ascomicetos/efeitos dos fármacos , Ascomicetos/metabolismo , Cromatografia Líquida de Alta Pressão , Metabolômica , Ácidos Ftálicos/farmacologia , Metabolismo Secundário , Espectrometria de Massas por Ionização por Electrospray , Acetona , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Cromatografia Líquida de Alta Pressão/métodos , Relação Dose-Resposta a Droga , Humanos , Metabolômica/métodos , Conformação Molecular , Estrutura Molecular , Ácidos Ftálicos/química , Espectrometria de Massas por Ionização por Electrospray/métodos
J Mass Spectrom ; 53(10): 963-975, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29987857


Sumatriptan succinate, a selective 5-HT1B receptor agonist, was subjected to forced degradation studies as per to International Conference on Harmonization-specified conditions. The drug exclusively showed its degradation under basic, photolytic, and oxidative stress conditions, whereas it was found to be stable under acidic, thermal, and neutral conditions. Eight (DP-1 to DP-8) degradation products were identified and characterized by UPLC-ESI/MS/MS experiments combined with accurate mass measurements. The effective chromatographic separation was achieved on Hibar Purospher STAR, C18 (250 × 4.6 mm, 5 µm) column using mobile phase consisting of 0.1% formic acid and methanol at a flow rate of 0.6 mL/minute in gradient elution method. It is noteworthy that 2 major degradation products DP-3 and DP-7 were isolated using preparative HPLC and characterized by advanced NMR experiments. The degradation pathway of the sumatriptan was established, which was duly justified by mechanistic explanation. In vitro cytotoxicity of isolated DPs was tested on normal human cells such as HEK 293 (embryonic kidney cells) and RWPE-1 (normal prostate epithelial cells). This study revealed that they were nontoxic up to 100 µm concentration. Further, in silico toxicity of the drug and its degradation products was determined using ProTox-II prediction tool. This study revealed that DP-4 and DP-8 are predicted for immune toxicity. Amine oxidase A and prostaglandin G/H synthase 1 are predicted as toxicity targets for DP-3, DP-4, and DP-6 whereas DP-1 and DP-2 are predicted for amine oxidase A target.

Cromatografia Líquida/métodos , Espectroscopia de Ressonância Magnética/métodos , Sumatriptana/análise , Sumatriptana/química , Espectrometria de Massas em Tandem/métodos , Sobrevivência Celular/efeitos dos fármacos , Estabilidade de Medicamentos , Células HEK293 , Humanos , Espectrometria de Massas por Ionização por Electrospray/métodos , Sumatriptana/toxicidade