HPLC Method for Separation of Cannabidiol Hemp Seed Oil with Skin Lipids and Tandem HRMS Technology for Characterization of a Chemical Marker

Cannabidiol (CBD) hemp seed oil is a commercial raw material with antioxidant and anti-inflammatory benefits that has been formulated into body wash and skin care products. The biggest analytical challenge is how to simultaneously quantify CBD and hemp seed oil as they deposited on the skin surface. CBD is easily separated and quantified from skin surface extracts via a HPLC-mass spectrometry methodology. However, the structural skeleton of triacylglycerides (TAGs) in hemp seed oil is same as those from the skin surface sebum. The strong hydrophobicity with subtle structural difference challenges their separation. In this project, a new reverse phase HPLC-high resolution mass spectrometry methodology was developed with a strong mobile phase normal propanol. The separated hemp seed oil TAGs in the chromatogram were identified and characterized using data-dependent acquisition (DDA) technology. Based on the daughter ion characterization, the separated peak with an ammonium adduct at 890.7226 [M + NH4]+ was confirmed as the parent ion of glycerol with three omega-3 fatty acid chains. This is the first time TAG structure with direct HPLC-tandem mass spectrometry technology has been elucidated without a hydrolysis reaction. The confirmed TAG structure with an ammonium adduct at 890.7226 ± 0.0020 can be used as a representative chemical marker for the hemp seed oil quantification.

Feminizing adrenal tumor identified by plasma steroid profiling

Summary Feminizing estrogen-secreting adrenocortical carcinomas (ACCs) are exceedingly rare and carry a poor prognosis. The most common presenting trait is gynecomastia, but enlarged breasts are also a frequent clinical finding in healthy men. Biochemical evaluation may be challenging. As such, there is a high risk of delayed diagnosis and treatment opportunity. Here, we present a case with an estrogen-producing ACC where the abnormal steroid profile obtained at the time of initial workup was essential for the prompt diagnosis. Wider adoption of liquid chromatography mass spectrometry-based steroid assays has potential to improve early diagnosis of feminizing estrogen-secreting ACC. Learning points Feminizing estrogen-secreting adrenocortical carcinomas (ACCs) are a rare, but an important differential diagnosis in men with rapidly developing gynecomastia. Biochemical evaluation is essential for a prompt diagnosis. Steroid hormone profiling using liquid chromatography mass spectrometry technology has the potential to improve early diagnosis of feminizing estrogen-secreting ACC.

Compositional Characteristics of Glucuronide Conjugates in Regional Glechoma hederacea var. longituba Herbal Extracts Using a Set of Polyphenolic Marker Compounds

Glechoma hederacea var. longituba (GHL) is one of many herbal plants widely used in hot herbal teas and in oriental prescriptions to treat various diseases. Although the beneficial effects of GHL may be influenced by differences in the composition of active constituents in the herbal extracts, there are few reports on the compositional characteristics of GHL herbal extracts to date. In this study, liquid chromatography–mass spectrometry technology was used for comparative analysis of constituents in hot-water extracts of GHL samples obtained from various cultivating provinces in South Korea. A set of marker panel consisting of nine polyphenolic compounds, including glucuronide conjugates in particular, was constructed and used to monitor the compositional characteristics in each GHL extract. Our findings show that some of the marker compounds, including rosmarinic acid, were persistently observed as major constituents in the analyses of the 22 GHL sample extracts, whereas, interestingly, other marker compounds such as polyphenol-glucuronic acid conjugates displayed dramatic differences in compositional ratios. This chromatographic approach using the marker compound panel can be applied to qualitatively and quantitatively evaluate compositional characteristics in the GHL extracts, and can also be useful for quality assays of the GHL herbal plant in medicinal and industrial fields.

212 Fatty Acid Analysis and Composition in Meat by Mass Spectrometry

Fatty acids determine the physical and chemical properties of fats. Animal fats, regardless of species, have more saturated and monounsaturated than polyunsaturated fatty acids. The major fatty acids in meat are palmitic (16:0), stearic (18:0), palmitoleic (16:1), oleic (18:1), linoleic (18:2), and linolenic (18:3) acids, among which oleic acid is the most predominant. Arachidonic acid (20:4 cis 5,8,11,14) is an essential fatty acid only found in animal fats and can be used as a quality control indicator in the fatty acid analysis. Fatty acid analysis has been traditionally performed by gas chromatography (GC) of volatile fatty acid derivatives, prominently the methyl esters, and flame ionization detection (FID), in which the carbon chain of fatty acids is degraded to the formylium ion CHO+. The FID is very sensitive and is the most widely used detection method for GC, providing a linear response, i.e., peak area, over a wide range of concentrations. Researchers have been used the FID peak area to calculate the percentages of fatty acids. However, the FID is a “carbon counter” and relies on the “equal per carbon” rule; therefore, at the same molar concentration, fatty acids with a different number of carbons produce different peak areas. The recent development of mass spectrometry technology has improved the specificity of fatty acid detection. Specific target and qualifier ions provide better identification and more accurate quantification of fatty acid concentrations. Although fatty acids can be identified through comparing ion fragmentation with various databases, authentic standards are needed for quantification purposes. Using mass spectrometry, more than 50 fatty acids have been identified in meat samples. Some branched-chain fatty acids may have flavor, safety, and shelf life implications in meat products.

GC/MS and LC/MS Based Serum Metabolomic Analysis of Dairy Cows With Ovarian Inactivity

Metabolic disorders may lead to the inactive ovaries of dairy cows during early lactation. However, the detailed metabolic profile of dairy cows with inactive ovaries around 55 days postpartum has not been clearly elucidated. The objective of this study was to investigate the metabolic difference in cows with inactive ovaries and estrus from the perspective of serum metabolites. According to clinical manifestations, B-ultrasound scan, rectal examination, 15 cows were assigned to the estrus group (E; follicular diameter 15–20 mm) and 15 to the inactive ovary group (IO; follicular diameter <8 mm and increased <2 mm within 5 days over two examinations). The blood was collected from the tail vein of the cow to separate serum 55–60 days postpartum, and then milked and fasted in the morning. Serum samples were analyzed using gas chromatography time-of-flight mass spectrometry technology (GC-TOF-MS) and ultra-high-pressure liquid chromatography-quadrupole-time-of-flight mass spectrometry (UHPLC-QTOF-MS). Differences in serum metabolites were identified using multivariate statistical analysis and univariate analysis. Thirty differentially abundant metabolites were identified between the two groups. In cows with inactive ovaries compared with cows in estrus, 20 serum metabolites were significantly higher (beta-cryptoxanthin (p = 0.0012), 9-cis-retinal (p = 0.0030), oxamic acid (p = 0.0321), etc.) while 10 metabolites were significantly lower (monostearin (p = 0.0001), 3-hydroxypropionic acid (p = 0.0005), D-talose (p = 0.0018), etc.). Pathway analysis indicated that the serum differential metabolites of multiparous cows in estrus obtained by the two metabolomics techniques were mainly involved in β-alanine metabolism and steroid biosynthesis metabolism, while other involved metabolic pathways were related to metabolism of glyoxylate; dicarboxylate metabolism; fructose, mannose, glutathione, glycerolipid, glycine, serine, threonine, propanoate, retinol, and pyrimidine metabolism. This indicates that the abnormalities in glucose metabolism, lipid metabolism, amino acid metabolism, and glutathione metabolism of postpartum dairy cows obstructed follicular development.

MALDI-TOF Mass Spectrometry Technology as a Tool for the Rapid Diagnosis of Antimicrobial Resistance in Bacteria

Species identification by using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a routine diagnostic process for infectious diseases in current clinical settings. The rapid, low-cost, and simple to conduct methodology is expanding its application in clinical microbiology laboratories to diagnose the antimicrobial resistance (AMR) in microorganisms. Primarily, antimicrobial susceptibility testing is able to be carried out either by comparing the area under curve of MALDI spectra of bacteria grown in media with antimicrobial drugs or by identifying the shift peaks of bacteria grown in media including 13C isotope with antimicrobial drugs. Secondly, the antimicrobial resistance is able to be determined through identifying (i) the antimicrobial-resistant clonal groups based on the fingerprints of the clone, (ii) the shift peak of the modified antimicrobial drug, which is inactivated by the resistance determinant, (iii) the shift peak of the modified antimicrobial target, (iv) the peak specific for the antimicrobial determinant, and (v) the biomarkers that are coproduced proteins with AMR determinants. This review aims to present the current usage of the MALDI-TOF MS technique for diagnosing antimicrobial resistance in bacteria, varied approaches for AMR diagnostics using the methodology, and the future applications of the methods for the accurate and rapid identification of AMR in infection-causing bacterial pathogens.

May critical molecular cross-talk between indoleamine 2,3-dioxygenase (IDO) and arginase during human aging be targets for immunosenescence control?

Background This study aimed to identify novel plasma metabolic signatures with possible clinical relevance during the aging process. A biochemical quantitative phenotyping platform, based on targeted electrospray ionization tandem mass spectrometry technology, was used for the identification of any eventual perturbed biochemical pathway by the aging process in prospectively collected peripheral blood plasma from 166 individuals representing the population of São Paulo city, Brazil. Results Indoleamine 2,3-dioxygenase (IDO) activity (Kyn/Trp) was significantly elevated with age, and among metabolites most associated with elevations in IDO, one of the strongest correlations was with arginase (Orn/Arg), which could also facilitate the senescence process of the immune system. Hyperactivity of IDO was also found to correlate with increased blood concentrations of medium-chain acylcarnitines, suggesting that deficiencies in beta-oxidation may also be involved in the immunosenescence process. Finally, our study provided evidence that the systemic methylation status was significantly increased and positively correlated to IDO activity. Conclusions In the present article, besides identifying elevated IDO activity exhibiting striking parallel association with the aging process, we additionally identified increased arginase activity as an underlying biochemical disturbance closely following elevations in IDO. Our findings support interventions to reduce IDO or arginase activities in an attempt to preserve the functionality of the immune system, including modulation of myeloid-derived suppressor cells (MDSCs), T cells, macrophages, and dendritic cells’ function, in old individuals/patients.

Investigating the active substance and mechanism of Jing-Fu-Kang granules via mass spectrometry technology and network pharmacology method

Jing-Fu-Kang granules (JFKG) is a famous Chinese patent medicine for the treatment of cervical spondylosis around the China, whereas the active substance and mechanism are not completely investigated clearly. In the current study, a rapid separation and identification method using UPLC-QE-Orbitrap-MS was established, 97 chemical constituents from JFKG were identified, and 16 prototype components from plasma samples after administration of JFKG were observed within 16 min. The structures of typical compounds were preliminarily speculated by comparing the retention time and fragmentation pattern. Furthermore, multiple databases were used to integrate the compound targets of JFKG, and the disease targets related to cervical spondylosis. After the intersection of the two sets of targets, a protein-protein interaction (PPI) network and a component-target-pathway network were established, then using the DAVID database to perform gene ontology analysis and Kyoto Encyclopedia of Genes and Genomes analysis on the common targets to find related pathways. Finally, a total of 531 common targets and 136 pathways were found to participate in the mechanism. Our findings will help to further confirm the mechanism of JFKG for relieving cervical spondylosis, which will improve the scientific rationality of JFKG in clinical use, and can also assist in guiding doctors.

MStractor: R Workflow Package for Enhancing Metabolomics Data Pre-Processing and Visualization

Untargeted metabolomics experiments for characterizing complex biological samples, conducted with chromatography/mass spectrometry technology, generate large datasets containing very complex and highly variable information. Many data-processing options are available, however, both commercial and open-source solutions for data processing have limitations, such as vendor platform exclusivity and/or requiring familiarity with diverse programming languages. Data processing of untargeted metabolite data is a particular problem for laboratories that specialize in non-routine mass spectrometry analysis of diverse sample types across humans, animals, plants, fungi, and microorganisms. Here, we present MStractor, an R workflow package developed to streamline and enhance pre-processing of metabolomics mass spectrometry data and visualization. MStractor combines functions for molecular feature extraction with user-friendly dedicated GUIs for chromatographic and mass spectromerty (MS) parameter input, graphical quality-control outputs, and descriptive statistics. MStractor performance was evaluated through a detailed comparison with XCMS Online. The MStractor package is freely available on GitHub at the MetabolomicsSA repository.