Advances in Mass Spectrometry-Metabolomics Based Approaches.

Highly selective and sensitive analytical techniques are necessary for microbial metabolomics due to the complexity of the microbial sample matrix. Hence, mass spectrometry (MS) has been successfully applied in microbial metabolomics due to its high precision, versatility, sensitivity, and wide dynamic range. The different analytical tools using MS have been employed in microbial metabolomics investigations and can contribute to the discovery or accelerate the search for bioactive substances. The coupling with chromatographic and electrophoretic separation techniques has resulted in more efficient technologies for the analysis of microbial compounds occurring in trace levels. This book chapter describes the current advances in the application of mass spectrometry-based metabolomics in the search for new biologically active agents from microbial sources; the development of new approaches for in silico annotation of natural products; the different technologies employing mass spectrometry imaging to deliver more comprehensive analysis and elucidate the metabolome involved in ecological interactions as they enable visualization of the spatial dispersion of small molecules. We also describe other ambient ionization techniques applied to the fingerprint of microbial natural products and modern techniques such as ion mobility mass spectrometry used to microbial metabolomic analyses and the dereplication of natural microbial products through MS.

Electromembrane Extraction of Naphthenic Acids in Produced Water Followed by Ultra-High-Resolution Mass Spectrometry Analysis.

Naphthenic acids comprise one of the most toxic compounds of the produced water released from offshore oil platforms. Therefore, developing and applying faster, simpler, and more efficient analytical methods for analyzing naphthenic acids are urgently needed. Electromembrane extraction (EME) uses the electrokinetic migration of target ions through a porous membrane. Herein, the EME method was applied to extract naphthenic acids from produced water. The EME method was optimized, and the optimal conditions encompassed decanol as the organic solvent, the sample with pH 10.0, 5 min of extraction at 200 V, and the ratio 4:1 (borate buffer/matrix, v/v). Electrochemical impedance spectroscopy confirmed charged species' migration from produced water through the EME. Subsequently, all extracts were analyzed by ultra-high-resolution mass spectrometry. The EME efficiency was assessed by comparing the extraction results to the liquid-liquid extraction (LLE) method results. Analytical results showed good linearity for both solvent and matrix curves (R2 > 0.98). Low detection limits ranged from 0.10 to 0.13 µg mL-1 and quantification limits from 0.36 to 0.45 µg mL-1. Precision and accuracy values ranged from -13.3% to 16.5%. These values fit the proposed method, demonstrating that the EME was more efficient than LLE in naphthenic acid extraction. The EME method preferably extracted aromatic compounds with double-bond equivalence from 6 to 8. The EME coupled with ultra-high-resolution mass spectrometry was demonstrated as a promising analytical approach to naphthenic acid extraction as an efficient and more environmentally friendly alternative to conventional extraction methods.

Laser ablation electrospray ionization mass spectrometry imaging as a new tool for accessing patulin diffusion in mold-infected fruits.

This work describes the use of laser ablation electrospray ionization mass spectrometry imaging (LAESI imaging) to investigate the diffusion of the mycotoxin patulin from rotten to healthy areas of fruits. Slices of mold-infected and uninfected (control) apples and strawberries were prepared, and this was the only sample preparation step used. An infrared laser beam (2.94 µm) was used to irradiate the slices, resulting in the ablation of sample compounds directly ionized by electrospray and analyzed by mass spectrometry. Multivariate curve resolution - alternating least squares was applied in unfolded LAESI images to obtain relative quantity information. Patulin was not detected in the control samples but was seen in all mold-infected fruits. LAESI images showed the diffusion of patulin from the rotten area to unaffected parts of the fruits. This study points out the advantage of LAESI imaging over traditional analytical methods used to study the diffusion of mycotoxins in fruits.

Structural and biosynthetic studies on eremophilenols related to the phytoalexin capsidiol, produced by Botrytis cinerea.

A thorough study of the fermentation broth of three strains of Botrytis cinerea which were grown on a modified Czapek-Dox medium supplemented with 5 ppm copper sulphate, yielded five undescribed metabolites. These metabolites possessed a sesquiterpenoid (+)-4-epi-eremophil-9-ene carbon skeleton which was enantiomeric to that of the phytoalexin, capsidiol. The isolation of these metabolites when the fungus was stressed, suggests that they may be potential effectors used by B. cinerea to circumvent plant chemical defences against phytopathogenic fungi. The biosynthesis of these compounds has been studied using 2H and 13C labelled acetate.

New AChE inhibitors from microbial transformation of trachyloban-19-oic acid by Syncephalastrum racemosum.

Trachyloban-19-oic acid (1) is a diterpene very abundant in nature and its structural modification can furnish new bioactive compounds. Biotransformation of 1 by fungus Syncephalastrum racemosum provided three derivatives, two hydroxylated products (2-3) and one product of rearrangement (4). Products 3 and 4 have never been reported so far, to the best of our knowledge. Structure of 3 was formed after oxidation and rearrangement of compound 2. Compounds 1-4 were evaluated for inhibition of acetylcholinesterase, enzyme linked to the symptomatic control of Alzheimer's disease. All the compounds presented inhibitory activity higher than starting material 1, and product 3 presented IC50 = 0.06 µM, which is about six times higher than activity found for galanthamine (IC50 = 0.38 µM), the positive control used in this assay.