In vitro analysis of metabolites from the untreated tissue of Torpedo californica electric organ by mid-infrared laser ablation electrospray ionization mass spectrometry

Abstract

The neuromuscular junction (NMJ), where a motor neuron intercepts and activates a muscle fiber, is a highly versatile and complex subcellular region. Genomic and proteomic approaches using the large ([1 kg) electric organ of Torpedo californica have helped advancing our understanding of this minute (30-50 lm) electric synapse. However, the majority of these studies have focused on mRNA and proteins, therefore neglecting small signaling molecules involved in muscle-nerve 'dialogue'. We developed a novel technique, mid-infrared laser ablation electrospray ionization (LAESI) mass spectrometry (MS), with the potential of detecting a diversity of small signaling molecules in vitro. LAESI uses the native water in the tissue as the matrix to couple the laser pulse energy into the target for the ablation process and enables its direct analysis essentially without sample preparation. Here, we report the detection of metabolites from the untreated frozen tissue of the Torpedo electric organ with LAESI MS at atmospheric pressure. A total of 24 metabolites were identified by accurate mass measurements, natural isotope patterns, and tandem mass spectrometry. Most of the identified metabolites were related to the cholinergic function of the electric synapse (acetylcholine and choline), fatty acid metabolism and acetyl transfer (carnitine and acetylcarnitine), the mitigation of osmotic stress (betaine and trimethylamine N-oxide), and energy production (creatine and creatinine). The biosynthetic precursors of these metabolites and their expected degradation products were also detected indicating that LAESI MS is well suited for tissue metabolomics with the ultimate goal of imaging and in vivo studies.

Ambient mass spectrometry in metabolomics

“…The authors found 24 metabolites associated with this organ, and though quantitation was difficult, they managed to use deuterated standards spiked on the tissue to analyze a few of the most important metabolites. 112 As mentioned above, IMS can be integrated with ambient ionization techniques for imaging experiments. LAESI-travelling wave IMS-quadrupole-time-of-flight-MSI enabled the construction of molecular images of selected ions that were distinguished by drift times in the sagittal sections of mouse brain tissue.…”

Electrochemical profiling using copper nanoparticle-plated electrode for identification of ostrich meat and evaluation of meat grades

“…The MS spectra of these two isobaric amino acids (Fig. 2B, lower panel) showed the protonated molecule [M+H] + at m/z 147, yielding identical fragment ions of m/ z 130 (the loss of NH 3 ) in the MS 2 spectrum, and m/z 84 (the loss of CH 2 O 2 ) in the MS 3 spectrum (Sripadi, Nazarian, Hathout, Hoffman, & Vertes, 2009). Both lysine and glutamine are common amino acid components in meats and account for more than 10% and 15% of total amino acids (Chow & Kuo, 2002) in these four meat species, their appearances as main compositional peaks in the EC chromatogram indicated the specific selectivity of the Cu n SPE to these two isobaric amino acids (see below).…”

Rapid analysis and identification of meat species by laser‐ablation electrospray mass spectrometry (LAESI‐MS)

“…It has been applied to directly analyze the spatial profiling of metabolites on leaf tissue, [14] lipid or metabolite species in mouse brain tissue, [15] and metabolites from the untreated tissue of the Torpedo California electric organ. [16] The direct sampling for in vivo studies without any chemical contamination is the main advantage of LAESI.In this study, five fresh meat samples, chicken, duck, pork, beef and mutton, were analyzed using LAESI-MS. The typical molecular markers of the meat samples were identified using raw mass spectra combining principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA).…”

“…It has been applied to directly analyze the spatial profiling of metabolites on leaf tissue, [14] lipid or metabolite species in mouse brain tissue, [15] and metabolites from the untreated tissue of the Torpedo California electric organ. [16] The direct sampling for in vivo studies without any chemical contamination is the main advantage of LAESI.…”