Ceria nanocubic-ultrasonication assisted dispersive liquid-liquid microextraction coupled with matrix assisted laser desorption/ ionization mass spectrometry for pathogenic bacteria analysis

摘要

A new ceria (CeO_2) nanocubic modified surfactant is used as the basis of a novel nano-based microextraction technique for highly sensitive detection of pathogenic bacteria (Pseudomonas aeruginosa and Staphylococcus aureus). The technique uses ultrasound enhanced surfactant-assisted dispersive liquid-liquid microextraction (UESA-DLLME) with and without ceria (CeO_2) followed by matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS). In order to achieve high separation efficiency, we investigated the influential parameters, including extraction time of ultrasonication, type and volume of the extraction solvent and surfactant. Among various surfactants, the cationic surfactants can selectively offer better extraction efficiency on bacteria analysis than that of the anionic surfactants due to the negative charges of bacteria cell membranes. Extractions of the bacteria lysate from aqueous samples via UESA-DLLME-MALDI-MS were successfully achieved by using cetyltrimethyl ammonium bromide (CTAB, 10.0 μL, 1.0 × 10~(-3) M) as surfactants in chlorobenzene (10.0 μL) and chloroform (10.0 μL) as the optimal extracting solvent for P. aeruginosa and S. aureus, respectively. Ceria nanocubic was synthesized, and functionalized with CTAB (CeO_2@CTAB) and then characterized using transmission electron microscopy (TEM) and optical spectroscopy (UV and FTIR). CeO_2@CTAB demonstrates high extraction efficiency, improve peaks ionization, and enhance resolution. The prime reasons for these improvements are due to the large surface area of nanoparticles, and its absorption that coincides with the wavelength of MALDI laser (337 nm, N_2 laser). CeO_2@CTAB-based microextraction offers lowest detectable concentrations tenfold lower than that of without nanoceria. The present approach has been successfully applied to detect pathogenic bacteria at low concentrations of 10~4-10~5 cfu/mL (without ceria) and at 10~3-10~4 cfu/mL (with ceria) from bacteria suspensions. Finally, the current approach was applied for analyzing the pathogenic bacteria in biological samples (blood and serum). Ceria assist surfactant (CeO_2@CTAB) liquid-liquid microextraction (LLME) offers better extraction efficiency than that of using the surfactant in LLME alone.