Gas chromatography/mass spectrometry analysis of reaction products of Chemical Weapons Convention-related 2-(N,N-dialkylamino)ethylchlorides with 2-(N,N-dialkylamino)ethanols.

RATIONALE: The Chemical Weapons Convention (CWC) mandates rigorous screening of chemical weapons and their potential degradation/reaction products, which is essential to identify such products in suspected samples. The reaction between 2-(N,N-dialkylamino)ethylchlorides and 2-(N,N-dialkylamino)ethanols (precursors/degradation products of VX agents) produces a new class of reaction products that are not explored. METHODS: The reaction products, bis(2-N,N-dialkylaminoethyl)ethers (1-10), were synthesized using established synthetic procedures, and gas chromatography/mass spectrometry (GC/MS) (electron ionization [EI] and chemical ionization [CI]) methods were developed for their identification. The GC/MS experiments were performed on an Agilent GC/MSD system using an HP-5MS capillary column. Methane gas was used as the CI reagent gas for GC/CIMS experiments. GC/retention index (RI) values of 1-10 were calculated using the retention times of the hydrocarbon mixture and the analytes. RESULTS: The GC/EI spectra of 1-10 exhibited [M-H]+ ions and distinctive fragments that provided valuable structural information. The selective fragmentation of the alkyl groups on nitrogen facilitated the discrimination of possible isomeric compounds. Interpretation of EI fragments in the high mass region is important for unambiguous identification of 1-10, because the major ions significantly match other CWC-related compounds containing the 2-(N,N-dialkylaminoethyl) group. GC/CI (methane) spectra included M+., [M + H]+, [M-H]+, reagent-specific adduct ions, and a few structure-indicative fragments. The spiking experiments in soil and water samples revealed that the target analytes were stable, easily extractable, and detectable using GC/MS. CONCLUSIONS: The reaction products, 1-10, could be successfully synthesized and characterized using GC/MS (EI and CI). The GC/MS and GC/RI data provide important insights into the unambiguous identification of the target molecules in challenging CWC verification and are helpful in participation in the Organization for the Prohibition of Chemical Weapons proficiency tests.

Plasma Metabolite Profiling Identifies Nondiabetic Chronic Pancreatitis Patients With Metabolic Alterations Progressing to Prediabetes Before HbA1c.

INTRODUCTION: Diabetes (T3cDM) secondary to chronic pancreatitis (CP) arises due to endocrine dysfunction and metabolic dysregulations. Currently, diagnostic tests are not available to identify patients who may progress from normoglycemia to hyperglycemia in CP. We conducted plasma metabolomic profiling to diagnose glycemic alterations early in the course of disease. METHODS: Liquid chromatography-tandem mass spectrometry was used to generate untargeted, targeted plasma metabolomic profiles in patients with CP, controls (n = 445) following TRIPOD guidelines. Patients were stratified based on glucose tolerance tests following ADA guidelines. Multivariate analysis was performed using partial least squares discriminant analysis to assess discriminatory ability of metabolites among stratified groups. COMBIROC and logistic regression were used to derive biomarker signatures. AI-ML tool (Rapidminer) was used to verify these preliminary results. RESULTS: Ceramide, lysophosphatidylethanolamine, phosphatidylcholine, lysophosphatidic acid (LPA), phosphatidylethanolamine, carnitine, and lysophosphatidylcholine discriminated T3cDM CP patients from healthy controls with AUC 93% (95% CI 0.81-0.98, P < 0.0001), and integration with pancreatic morphology improved AUC to 100% (95% CI 0.93-1.00, P < 0.0001). LPA, phosphatidylinositol, and ceramide discriminated nondiabetic CP with glycemic alterations (pre-diabetic CP); AUC 66% (95% CI 0.55-0.76, P = 0.1), and integration enhanced AUC to 74% (95% CI 0.55-0.88, P = 0.86). T3cDM was distinguished from prediabetic by LPA, phosphatidylinositol, and sphinganine (AUC 70%; 95% CI 0.54-0.83, P = 0.08), and integration improved AUC to 83% (95% CI 0.68-0.93, P = 0.05). CombiROC cutoff identified 75% and 78% prediabetes in validation 1 and 2 cohorts. Random forest algorithm assessed performance of integrated panel demonstrating AUC of 72% in predicting glycemic alterations. DISCUSSION: We report for the first time that a panel of metabolites integrated with pancreatic morphology detects glycemia progression before HbA1c in patients with CP.

LC-MS/MS Analysis of Reaction Products of Arginine/Methylarginines with Methylglyoxal/Glyoxal.

Methylglyoxal (MGO) and glyoxal (GO) are toxic α-dicarbonyl compounds that undergo reactions with amine containing molecules such as proteins and amino acids and result in the formation of advanced glycation end products (AGEs). This study aimed at investigating the reactivity of arginine (Arg) or dimethylarginine (SDMA or ADMA) with MGO or GO. The solutions of arginine and MGO or GO were prepared in PBS buffer (pH 7.4) and incubated at 37 °C. Direct electrospray ionization-high-resolution mass spectrometry (ESI-HRMS) analysis of the reaction mixture of Arg and MGO revealed the formation of Arg-MGO (1:1) and Arg-2MGO (1:2) products and their corresponding dehydrated products. Further liquid chromatography (LC)-MS analyses revealed the presence of isomeric products in each 1:1 and 1:2 product. The [M + H]+ of each isomeric product was subjected to MS/MS experiments for structural elucidation. The MS/MS spectra of some of the products showed a distinct structure indicative fragment ions, while others showed similar data. The types of products formed by the arginines with GO were also found to be similar to that of MGO. The importance of the guanidine group in the formation of the AGEs was reflected in similar incubation experiments with ADMA and SDMA. The structures of the products were proposed based on the comparison of the retention times and HRMS and MS/MS data interpretation, and some of them were confirmed by drawing analogy to the data reported in the literature.

Evaluation of amino acids and other related metabolites levels in end-stage renal disease (ESRD) patients on hemodialysis by LC/MS/MS and GC/MS.

End-stage renal disease (ESRD) is a rapidly increasing health problem, and every year, about 2 million ESRD cases are reported worldwide. Hemodialysis (HD) is the vital renal reinstatement therapy for ESRD, and HD patterns play a crucial role in patients' health. Plasma metabolomics is the potential approach to understanding the HD process, effectiveness, and patterns. The lack of protein vitality is a primary problem for HD patients, and the quantities of amino acids intracellularly and in the blood are considered to be a symbolic index of protein metabolism and nutrition conditions. In the current study, LC/MS/MS and GC/MS methods were developed for 29 targeted plasma metabolites and validated as per ICH bioanalytical method validation M10 guidelines. The 29 metabolites included 20 proteinogenic amino acids and nine other related metabolites. The methods were employed to measure the absolute quantities (µM) of the targeted metabolites in HD patients (n=60) before and after dialysis (PRE-HD and POST-HD), and compared with the healthy control (HC) group (n=60). Phenylacetylglutamine was found to be higher in both PRE-HD (72.88±14.5 µM) and POST-HD (26.62±7.9 µM), when compared to HC (1.61±0.6 µM). On the other hand, glutamic acid was lower in PRE-HD (14.90±6.5 µM), and POST-HD (13.6±6.1 µM) than that of HC (245.4±37.8 µM). The dialytic loss was found to be 52-45% for arginine, lysine, and histidine, while it was 38-26% for glycine, cysteine, proline, alanine, threonine, glutamine, valine, and methionine. The dialytic loss was low (≤12%) for aspartic acid, glutamic acid, asparagine, leucine, tyrosine, tryptophan, and isoleucine. Graphical abstract adapted from mass spectrometry templates by Biorender.com retrieved from https://app.biorender.com/biorender-templates .

Assessment of uremic toxins in advanced chronic kidney disease patients on maintenance hemodialysis by LC-ESI-MS/MS.

INTRODUCTION: In the advanced stage of chronic kidney disease (CKD), electrolytes, fluids, and metabolic wastes including various uremic toxins, accumulate at high concentrations in the patients' blood. Hemodialysis (HD) is the conventional procedure used worldwide to remove metabolic wastes. The creatinine and urea levels have been routinely monitored to estimate kidney function and effectiveness of the HD process. This study, first from in Indian perspective, aimed at the identification and quantification of major uremic toxins in CKD patients on maintenance HD (PRE-HD), and compared with the healthy controls (HC) as well as after HD (POST-HD). OBJECTIVES: The study mainly focused on the identification of major uremic toxins in Indian perspective and the quantitative analysis of indoxyl sulfate and p-cresol sulfate (routinely targeted uremic toxins), and phenyl sulfate, catechol sulfate, and guaiacol sulfate (targeted for the first time), apart from creatinine and urea in PRE-HD, POST-HD, and HC groups. METHODS: Blood samples were collected from 90 HD patients (both PRE-HD and POST-HD), and 74 HCs. The plasma samples were subjected to direct ESI-HRMS and LC/HRMS for untargeted metabolomics and LC-MS/MS for quantitative analysis. RESULTS: Various known uremic toxins, and a few new and unknown peaks were detected in PRE-HD patients. The p-cresol sulfate and indoxyl sulfate were dominant in PRE-HD, the concentrations of phenyl sulfate, catechol sulfate, and guaiacol sulfate were about 50% of that of indoxyl sulfate. Statistical evaluation on the levels of targeted uremic toxins in PRE-HD, POST-HD, and HC groups showed a significant difference among the three groups. The dialytic clearance of indoxyl sulfate and p-cresol sulfate was found to be < 35%, while that of the other three sulfates was 50-58%. CONCLUSION: LC-MS/MS method was developed and validated to evaluate five major uremic toxins in CKD patients on HD. The levels of the targeted uremic toxins could be used to assess kidney function and the effectiveness of HD.

Lutein and ß-carotene biosynthesis in Scenedesmus sp. SVMIICT1 through differential light intensities.

The study evaluated the biosynthesis of lutein and ß-carotene by Scenedesmus sp. SVMIICT1 under five different light intensities (50, 250, 500, 750 and 1000 µE/m2/s). Liquid chromatography/mass spectrometry (LC/MS) was used to determine relative quantities of lutein and ß-carotene. Relatively, high lutein content of 1.43 ± 0.04 and 0.70 ± 0.02 mg/g was found with 50 and 500 µE/m2/s conditions respectively. ß-Carotene content was quantified as 0.15 ± 0.01, 0.1 ± 0.01 and 0.12 ± 0.02 mg/g with 50, 250 and 500 µE/m2/s conditions respectively. The light intensities altered photosystem II and photosystem I. At 50 µE intensity, high chlorophyll content and high photosystem II quantum efficiency (FV/FM) was observed. Low FV/FM ratio of around 0.3 was detected in high light intensities (750 µE and 1000 µE) due to photoinhibition. Lipid fraction increased with increasing light intensity and the fatty acid profiles were similar in all five conditions.