Mass spectrometry imaging of levofloxacin distribution in TB-infected pulmonary lesions by MALDI-MSI and continuous liquid microjunction surface sampling.

A multi-modal mass spectrometry imaging (MSI) and profiling approach has been applied to assess the partitioning of the anti-TB fluoroquinolone levofloxacin into pulmonary lesions. Matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) and a commercial liquid microjunction surface sampling technology (LMJ-SSP), or flowprobe, have been used to both spatially profile and image drug distributions in lung tissue sections from TB-infected rabbits following oral administration of a single human-equivalent dose. Levofloxacin levels were highest at 6 h post-dose in normal lung, cellular granuloma, and necrotic caseum compartments. The drug accumulated in the cellular granuloma regions with lower amounts partitioning into central caseous compartments. Flowprobe imaging at 630 µm (limited by the probe tip diameter) enabled visualization of drug distribution into lesion compartments, including limited differentiation of relative drug abundance in cellular versus caseous regions of the lesions. MALDI-MSI analysis at 75 µm provided more detailed drug distribution, which clearly accumulated in the cellular region immediately surrounding the central caseum core. Imaging and profiling data acquired by flowprobe and MALDI-MSI were validated by quantitative LC/MS/MS analysis of lung and granuloma homogenates taken from the same animals. The results of the investigation show flowprobe imaging and sampling as a rapid and sensitive alternative to MALDI-MSI for profiling drug distributions into tissues when spatial resolution of data below the threshold of the probe diameter is not required.

Real-time metabolomics on living microorganisms using ambient electrospray ionization flow-probe.

Microorganisms such as bacteria and fungi produce a variety of specialized metabolites that are invaluable for agriculture, biological research, and drug discovery. However, the screening of microbial metabolic output is usually a time-intensive task. Here, we utilize a liquid microjunction surface sampling probe for electrospray ionization-mass spectrometry to extract and ionize metabolite mixtures directly from living microbial colonies grown on soft nutrient agar in Petri-dishes without any sample pretreatment. To demonstrate the robustness of the method, this technique was applied to observe the metabolic output of more than 30 microorganisms, including yeast, filamentous fungi, pathogens, and marine-derived bacteria, that were collected worldwide. Diverse natural products produced from different microbes, including Streptomyces coelicolor , Bacillus subtilis , and Pseudomonas aeruginosa are further characterized.

Liquid microjunction surface sampling probe fluid dynamics: characterization and application of an analyte plug formation operational mode.

The recently discovered sample plug formation and injection operational mode of a continuous flow, coaxial tube geometry, liquid microjunction surface sampling probe (LMJ-SSP) was further characterized and applied for concentration and mixing of analyte extracted from multiple areas on a surface and for nanoliter-scale chemical reactions of sampled material. A transparent LMJ-SSP was constructed and colored analytes were used so that the surface sampling process, plug formation, and the chemical reactions could be visually monitored at the sampling end of the probe before being analyzed by mass spectrometry of the injected sample plug. Injection plug peak widths were consistent for plug hold times as long as the 8 min maximum attempted (RSD below 1.5%). Furthermore, integrated injection peak signals were not significantly different for the range of hold times investigated. The ability to extract and completely mix individual samples within a fixed volume at the sampling end of the probe was demonstrated and a linear mass spectral response to the number of equivalent analyte spots sampled was observed. Using the color and mass changing chemical reduction of the redox dye 2,6-dichlorophenol-indophenol with ascorbic acid, the ability to sample, concentrate, and efficiently run reactions within the same plug volume within the probe was demonstrated.

Liquid microjunction surface sampling probe fluid dynamics: computational and experimental analysis of coaxial intercapillary positioning effects on sample manipulation.

A coaxial geometry liquid microjunction surface sampling probe (LMJ-SSP) enables direct extraction of analytes from surfaces for subsequent analysis by techniques like mass spectrometry. Solution dynamics at the probe-to-sample surface interface in the LMJ-SSP has been suspected to influence sampling efficiency and dispersion but has not been rigorously investigated. The effect on flow dynamics and analyte transport to the mass spectrometer caused by coaxial retraction of the inner and outer capillaries from each other and the surface during sampling with a LMJ-SSP was investigated using computational fluid dynamics and experimentation. A transparent LMJ-SSP was constructed to provide the means for visual observation of the dynamics of the surface sampling process. Visual observation, computational fluid dynamics (CFD) analysis, and experimental results revealed that inner capillary axial retraction from the flush position relative to the outer capillary transitioned the probe from a continuous sampling and injection mode through an intermediate regime to sample plug formation mode caused by eddy currents at the sampling end of the probe. The potential for analytical implementation of these newly discovered probe operational modes is discussed.

Direct sampling and analysis from solid-phase extraction cards using an automated liquid extraction surface analysis nanoelectrospray mass spectrometry system.

Direct liquid extraction based surface sampling, a technique previously demonstrated with continuous flow and autonomous pipette liquid microjunction surface sampling probes, has recently been implemented as a liquid extraction surface analysis (LESA) mode on a commercially available chip-based infusion nanoelectrospray ionization (nanoESI) system. In the present paper, the LESA mode was applied to the analysis of 96-well format custom-made solid-phase extraction (SPE) cards, with each well consisting of either a 1 or a 2 mm diameter monolithic hydrophobic stationary phase. These substrate wells were conditioned, loaded with either single or multi-component aqueous mixtures, and read out using the commercial nanoESI system coupled to a hybrid triple quadrupole/linear ion trap mass spectrometer or a linear ion trap mass spectrometer. The extraction conditions, including extraction/nanoESI solvent composition, volume, and dwell times, were optimized in the analysis of targeted compounds. Limit of detection and quantitation as well as analysis reproducibility figures of merit were measured. Calibration data was obtained for propranolol using a deuterated internal standard which demonstrated linearity and reproducibility. A 10× increase in signal and cleanup of micromolar angiotensin II from a concentrated salt solution was demonstrated. In addition, a multicomponent herbicide mixture at ppb concentration levels was analyzed using MS(3) spectra for compound identification in the presence of isobaric interferences.