Desorption Electrospray Ionization Cyclic Ion Mobility-Mass Spectrometry Imaging for Traumatic Brain Injury Spatial Metabolomics.

Lipidomics focuses on investigating alterations in a wide variety of lipids that harness important information on metabolic processes and disease pathology. However, the vast structural diversity of lipids and the presence of isobaric and isomeric species creates serious challenges in feature identification, particularly in mass spectrometry imaging experiments that lack front-end separations. Ion mobility has emerged as a potential solution to address some of these challenges and is increasingly being utilized as part of mass spectrometry imaging platforms. Here, we present the results of a pilot mass spectrometry imaging study on rat brains subjected to traumatic brain injury (TBI) to evaluate the depth and quality of the information yielded by desorption electrospray ionization cyclic ion mobility mass spectrometry (DESI cIM MSI). Imaging data were collected with one and six passes through the cIM cell. Increasing the number of passes increased the ion mobility resolving power and the resolution of isobaric lipids, enabling the creation of more specific maps. Interestingly, drift time data enabled the recognition of multiply charged phosphoinositide species in the complex data set generated. These species have not been previously reported in TBI MSI studies and were found to decrease in the hippocampus region following injury. These changes were attributed to increased enzymatic activity after TBI, releasing arachidonic acid that is converted to eicosanoids to control inflammation. A substantial reduction in NAD and alterations in other adenine metabolites were also observed, supporting the hypothesis that energy metabolism in the brain is severely disrupted in TBI.

Spatial lipidomics maps brain alterations associated with mild traumatic brain injury.

Traumatic brain injury (TBI) is a global public health problem with 50-60 million incidents per year, most of which are considered mild (mTBI) and many of these repetitive (rmTBI). Despite their massive implications, the pathologies of mTBI and rmTBI are not fully understood, with a paucity of information on brain lipid dysregulation following mild injury event(s). To gain more insight on mTBI and rmTBI pathology, a non-targeted spatial lipidomics workflow utilizing high resolution mass spectrometry imaging was developed to map brain region-specific lipid alterations in rats following injury. Discriminant multivariate models were created for regions of interest including the hippocampus, cortex, and corpus callosum to pinpoint lipid species that differentiated between injured and sham animals. A multivariate model focused on the hippocampus region differentiated injured brain tissues with an area under the curve of 0.99 using only four lipid species. Lipid classes that were consistently discriminant included polyunsaturated fatty acid-containing phosphatidylcholines (PC), lysophosphatidylcholines (LPC), LPC-plasmalogens (LPC-P) and PC potassium adducts. Many of the polyunsaturated fatty acid-containing PC and LPC-P selected have never been previously reported as altered in mTBI. The observed lipid alterations indicate that neuroinflammation and oxidative stress are important pathologies that could serve to explain cognitive deficits associated with rmTBI. Therapeutics which target or attenuate these pathologies may be beneficial to limit persistent damage following a mild brain injury event.

Spatial Lipidomics Maps Brain Alterations Associated with Mild Traumatic Brain Injury.

Traumatic brain injury (TBI) is a global public health problem with 50-60 million incidents per year, most of which are considered mild (mTBI) and many of these repetitive (rmTBI). Despite their massive implications, the pathologies of mTBI and rmTBI are not fully understood, with a paucity of information on brain lipid dysregulation following mild injury event(s). To gain more insight on mTBI and rmTBI pathology, a non-targeted spatial lipidomics workflow utilizing ultrahigh resolution mass spectrometry imaging was developed to map brain region-specific lipid alterations in rats following injury. Discriminant multivariate models were created for regions of interest including the hippocampus, cortex, and corpus callosum to pinpoint lipid species that differentiated between injured and sham animals. A multivariate model focused on the hippocampus region differentiated injured brain tissues with an area under the curve of 0.994 using only four lipid species. Lipid classes that were consistently discriminant included polyunsaturated fatty acid-containing phosphatidylcholines (PC), lysophosphatidylcholines (LPC), LPC-plasmalogens (LPC-P) and PC potassium adducts. Many of the polyunsaturated fatty acid-containing PC and LPC-P selected have never been previously reported as altered in mTBI. The observed lipid alterations indicate that neuroinflammation, oxidative stress and disrupted sodium-potassium pumps are important pathologies that could serve to explain cognitive deficits associated with rmTBI. Therapeutics which target or attenuate these pathologies may be beneficial to limit persistent damage following a mild brain injury event.

Determination of thyroid hormones in dietary supplements using liquid chromatography-tandem mass spectrometry.

More than 27 million Americans have some kind of thyroid disease. Numerous dietary supplements claiming to support healthy thyroid function and healthy metabolism and balance or promote weight loss are available for purchase in retail stores and on the internet. In the literature, there have been reports of adverse events associated with the consumption of thyroid hormone-containing products. In this study, an LC-MS/MS method was developed and validated for the analysis of thyroxine (T4), 3,3',5-triiodo-l-thyronine (T3), 3,3',5'-triiodothyronine (rT3), 3,5-diiodothyronine (3,5-T2) and 3,3'-diiodothyronine (3,3'-T2) in dietary supplements. Sonication with methanol was used for the extraction of free hormones from nonglandular products. The tissue-bound hormones from glandular thyroid products were extracted using a modified enzymatic digestion, in which the matrix was first extracted by sonication with methanol and then by enzymatic digestion with proteases. Both extraction methods provided acceptable recovery values between 78% and 116%. Fifty-eight products making claims related to thyroid management were purchased over the internet from 2017-2018 and quantitatively analyzed for five hormones using the validated methods. Eleven out of 19 glandular products were found to contain quantifiable amounts of hormones. Maximum daily servings were also calculated for each product based on label information. The maximum amount of T4, T3, and rT3 per daily serving in the glandular products were up to 210, 32, and 7.6 µg/day, respectively. In the case of nonglandular products, which were labeled to contain plant extracts, vitamins, minerals, diiodo compounds, and so forth, the amounts of 3,5-T2 and 3,3'-T2 were up to 740 and 2700 µg/day, respectively.

Analysis of bitter orange dietary supplements for natural and synthetic phenethylamines by LC-MS/MS.

Citrus aurantium, commonly known as bitter orange, is a popular dietary supplement ingredient sold worldwide. Bitter orange supplements are sold primarily as weight management and sports performance products and have gained popularity after Ephedra products were banned from the US market. Supplements containing synephrine are reported to exhibit adverse cardiovascular effects especially in the presence of caffeine. In this study, an LC-MS/MS method was established to quantify five natural amines (synephrine, octopamine, tyramine, N-methyltyramine, and hordenine) and four synthetic phenethylamines (phenylephrine, methylsynephrine, etilefrine, and isopropyloctopamine) in dietary supplements sold in the US. The method was validated and found to have acceptable performance to accurately measure analytes in complex botanical products. The average recoveries from a blank matrix were 88-125% with an RSD of 0.5-7.0%. Fifty-nine products labeled to contain bitter orange peel, extract, or its amines were purchased and their amine content was measured. Several products were found to contain higher amounts of amines than that expected from a typical bitter orange extract. Of the 23 products that made label claims for synephrine, only 5 products (22%) were within 80-120% of labeled synephrine content. The presence of synthetic amines, methylsynephrine (up to 240 mg/daily serving), and isopropyloctopamine (up to 76 mg/daily serving), whose effects in humans are not known, were detected in six products and one product, respectively. While the use of methylsynephrine and isopropyloctopamine are not permitted in dietary supplements, hordenine, N-methyltyramine, and octopamine are currently listed on the FDA's Dietary Supplement Ingredient Advisory List.

Systematic revision of the Tubifera casparyi-T. dictyoderma complex: Resurrection of the genus Siphoptychium and introduction of the new genus Thecotubifera.

The genus Siphoptychium is resurrected on the basis of comparative morphology and phylogeny of partial nuc 18S rDNA (18S) and translation elongation factor 1-alpha (EF1A) nucleotide sequences. The genus is characterized by the firm upper surface of the pseudoaethalium, accreted but easily separable sporothecae, a tubular or fibrous columella, and spores with a reticulate ornamentation consisting of 7-9 meshes across the diameter. In addition to the currently known single species S. casparyi (= Tubifera casparyi), two new members of Siphoptychium are described: S. violaceum from coniferous forests of Europe, east Asia, and southeast Asia, and S. reticulatum from temperate and subarctic regions of North America and alpine regions of Europe. A second genus, Thecotubifera, is described to accommodate Tubifera dictyoderma. The fruiting body of this species is transitional between a pseudoaethalium and a true aethalium. It is covered by a contiguous membranous cortex formed by the fused tips of the sporothecae, a feature typical for aethalia. However, the inner portions of sporothecae remain discernible, a feature more typical for pseudoaethalia. Columellae of Th. dictyoderma are formed by perforated plates, and the spores have a reticulate ornamentation consisting of 2-5 meshes across the diameter. For Th. dictyoderma, we could confirm records only for tropical regions and Japan, whereas all studied European specimens, including those mentioned in current monographs, represent species of Siphoptychium.

A critical revision of the Tubifera ferruginosa complex.

Based on a combination of morphological and molecular investigations, a critical revision of the widely distributed myxomycete Tubifera ferruginosa is presented. A phylogeny of the morphospecies, based on partial 18S nuc rDNA sequences, displays several clearly distinct clades, all differing by a genetic distance (p distance) of at least 0.15, with the distance within the clades below 0.11. These molecular differences correlate with morphological characters, such as the shape of sporothecal tips, the color of immature fructifications and the ultrastructure of the inner surface of the peridium. The combination of morphological and molecular data provides evidence that T. ferruginosa is actually a species complex, representing at least seven species. These are T. ferruginosa sensu stricto, T. applanata, T. corymbosa, T. dudkae, T. magna, T. montana and T. pseudomicrosperma. Among these T. applanata and T. dudkae (as Reticularia dudkae) were described recently based on morphological characters and the 18S nuc rDNA phylogeny confirmed their separation. Another four species, T. corymbosa, T. magna, T. montana and T. pseudomicrosperma, are described here. We propose an epitype for T. ferruginosa sensu stricto and recognize subsp. ferruginosa and subsp. acutissima within this species. All studied taxa of the T. ferruginosa complex are shown to lack a capillitium. Structures formerly described as capillitium represent the hyphae of fungi occurring within the fructifications.

Perichaena longipes, a new myxomycete from the Neotropics.

A new species of myxomycete, Perichaena longipes, is described from 56 sporocarp specimens that appeared in moist chamber cultures prepared with samples of decaying plant materials collected in Panama, Costa Rica and Brazil. This new species is distinguished from the morphologically similar species P. pedata on the basis of the much longer stipe, lighter peridium and the unique ornamentation of the capillitium. The nuc 18S ribosomal DNA sequences obtained from four specimens of P. longipes support the distinction of this new taxon and its separation from P. pedata. Furthermore, maximum likelihood phylogeny supports earlier evidence that species currently within the genus Perichaena do not form a monophyletic clade. Instead they appear to form three separate branches within the bright-spored clade. The first clade includes P. longipes together with several species of Trichia and Metatrichia, the second includes P. pedata and P. chrysosperma, and the third clade is composed of P. corticalis, P. depressa and P. luteola.

The genus Alwisia (Myxomycetes) revalidated, with two species new to science.

Based on morphological investigations and a phylogeny constructed with partial sequences of the SSU rDNA gene, we revalidate the genus Alwisia and propose the combination Alwisia bombarda Berk. & Broome to be used against Tubifera bombarda (Berk. & Broome) G.W. Martin. Two new species, Alwisia morula and A. repens, are described based on material collected respectively in Costa Rica and Australia. Both new species lack a capillitium and possess individually stalked subspherical sporothecae. Alwisia repens differs from A. morula by its procumbent stalks and iridescent peridium. A comparison of 83 sequences of species in the genera Lycogala, Reticularia and Tubifera with a recent two-gene phylogeny of the bright-spored myxomycetes resulted in a similar topology of both Bayesian and maximum likelihood trees and placed A. bombarda, A. morula and A. repens in one well delimited clade within Reticulariaceae.

A new species of Alwisia (Myxomycetes) from New South Wales and Tasmania.

Based on a morphological investigation of a series of specimens collected in New South Wales and Tasmania and a phylogeny constructed with partial 18S ribosomal RNA gene sequences, we describe a new species Alwisia lloydiae; the fourth species within the recently revalidated genus Alwisia. This new species is characterized by short ovate sporothecae with mostly free stalks, morphologically resembling the recently described A. morula. However, the new species possesses a tubular capillitium that suggests an affinity with A. bombarda. The capillitium of the new species is ornamented with globular warts, and this feature separates it from all other members of the genus.