Development of an enzymatic assay for ethanolamine in plasma (original) (raw)
Related papers
Analytical Biochemistry, 1968
In recent years, a number of methods for the quantitative determination of the phospholipids by thin-layer chromatography have been reported (l-4). These methods depend on the separation of phospholipids on thin layers of silica gel G or similar adsorbents developed with mixtures of chloroform, methanol, and water, some of which also contain ammonium hydroxide or acetic acid. The phospholipid-containing areas of the adsorbent material were transferred to tubes or columns and eluted with various solvent mixtures and the quantity of phosphorus in the eluates was determined.
Clinical Chemistry and Laboratory Medicine (CCLM)
Objectives Fast and reliable ethanol assays analysis are used in a clinical context for patients suspected of ethanol intoxication. Mostly, automated systems using an enzymatic reaction based on ethanol dehydrogenase are used. The manuscript focusses on the evaluation of the performance of these assays. Methods Data included 30 serum samples used in the Belgian EQA scheme from 2019 to 2021 and concentrations ranged from 0.13 to 3.70 g/L. A regression line between target concentrations and reported values was calculated to evaluate outliers, bias, variability and measurement uncertainty. Results A total of 1,611 results were taken into account. Bias was the highest for Alinity c over the whole concentration range and the lowest for Vitros for low concentrations and Cobas 8000 using the c702 module for high concentrations. The Architect and Cobas c501/c502 systems showed the lowest variability over the whole concentration range. Highest variability was observed for Cobas 8000 using th...
Clinical enzymology and its applications
2008
Clinical Enzymology Enzymes nomenclature, classification, structure and specificity Enzymes, isoenzymes and their relevance in diagnosis Regulation of enzyme levels in serum and plasma Selection of enzyme tests Measurement of isoenzymes and isoforms Some important enzymes in clinical practice Aspartate transaminase (AST) and Alanine transaminase (ALT) Creatine Kinase (CK) Lactate dehydrogenase (LDH) Alkaline phosphatase Acid phosphatase γ-Glutamyltransferase (GGT) Amylase Lipase Cholinesterases 5’ nucleotidase (5'NT) Functional tests of renal, liver and gastric fluid Gastric function tests Renal function tests Liver function tests (LFT)
Biochemical Pharmacology, 1974
The effect of acute ethanol ingestion on hepatic phospholipid metabolism has been investigated in the rat. No significant increase in the content of either lecithin or phosphatidylethanolamine occurred in the liver after 12 hr from treatment. The triglyceride content of the liver increased threefold. An increase of lecithin and phosphatidylethanolamine synthesis by the Kennedy pathway was found in vitro in the homogenates and microsomal fractions prepared from the ethanol-treated rats. A higher rate of conversion of phosphorylcholine and phosphorylethanolamine to CDP-choline and CDP-ethanolamine was also found in the experimental animals. The breakdown of CDP-choline to phosphorylcholine was noticeably decreased in the liver fractions of the ethanol-treated rats. No changes in the sequential methylation pathway for lecithin synthesis in the liver were observed after acute ethanol ingestion.
Analysis of the alcohol biomarker phosphatidylethanol by NACE with on-line ESI-MS
ELECTROPHORESIS, 2010
Phosphatidylethanol (Peth), a group of aberrant phospholipids formed in cell membranes in the presence of ethanol, has been recently proposed as biomarker of chronic alcohol abuse. The aim of this study was to develop a new analytical method, based on NACE online coupled with a mass spectrometer for the analysis of Peth in blood. For this purpose an iontrap mass spectrometer equipped with an orthogonal ESI source operating in negative ion mode was used. An alkaline solution of ammonium acetate 5 mM (pH 9) in water/ methanol (MeOH) (80:20 v/v) was delivered as coaxial sheath liquid. All experiments were performed using an uncoated fused-silica capillary (90 cm  75 mm id). The effects of variable percentages of ACN, MeOH, 2-propanol, dichloromethane, along with variable concentrations of ammonium acetate were investigated for the separation of Peth. Collectively, a separation medium composed of ACN (45% v/v), 2-propanol (20% v/v), dichloromethane (20% v/v), MeOH (10% v/v), water (5% v/v), and ammonium acetate (25 mM) was chosen. The estimated LOD was 0.1 mM, while LOQ was 0.4 mM. Within-run (intra-day) and between-run (inter-day) precision was always lower than 15%. The method proved to be robust and reliable. The MS detector allowed the simultaneous identification of several Peth homologues, and the use of an internal standard (phosphatidylbutanol) with similar electrophoretic properties of that of Peth increased quantitation effectiveness.
Biosynthesis of rat brain phosphatidylethanolamines from intracerebrally injected ethanolamine
Brain Research, 1977
2-aH]Ethanolamine was injected intracerebrally into male rats and the brains of the animals immediately removed by particular procedures at regular intervals over the first 1200 sec. The incorporation of radioactivity into brain phosphorylethanolamine, cytidine-5'-diphosphate (CDP) ethanolamine and phosphatidylethanolamines was examined and quantitated. The nature of phosphatidylethanolamine molecular subspecies, which became labelled, was also investigated after isotope administration. Phosphorylethanolamine, CDP-ethanolamine and phosphatidylethanolamines were all labelled already 5 sec after the administration of labelled ethanolamine. The specific radioactivities of different phosphatidylethanolamine molecular subspecies varied according to the time elapsed from the injection to the sacrifice of the animals. This last result, together with the data on time course of labelling of ethanolamine phosphoglycerides and their precursors, provides indications that this base may be incorporated into lipids not only by net synthesis pathway, but also by base-exchange reaction.
Journal of inherited metabolic disease, 2018
Since organic acid analysis in urine with gaschromatography-mass spectrometry (GC-MS) is a time-consuming technique, we developed a new liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF/MS) method to replace the classical analysis for diagnosis of inborn errors of metabolism (IEM). Sample preparation is simple and experimental time short. Targeted mass extraction and automatic calculation of z-scores generated profiles characteristic for the IEMs in our panel consisting of 71 biomarkers for defects in amino acids, neurotransmitters, fatty acids, purine, and pyrimidine metabolism as well as other disorders. In addition, four medication-related metabolites were included in the panel. The method was validated to meet Dutch NEN-EN-ISO 15189 standards. Cross validation of 24 organic acids from 28 urine samples of the ERNDIM scheme showed superiority of the UPLC-QTOF/MS method over the GC-MS method. We applied our method to 99 patient urine samples with 32 differe...
Metabolomics
Introduction Current markers of Parkinson's disease (PD) fail to detect the early progression of disease state. Conversely, current omics techniques allow the investigation of hundreds of molecules potentially altered by disease conditions. Based on evidence previously collected by our group in a mouse model of PD, we speculated that a particular set of circulating lipids might be significantly altered by the pathology. Objectives The aim of current study was to evaluate the potential of a particular set of N-acyl-phosphatidylethanolamines (NAPEs) as potential non-invasive plasma markers of ongoing neurodegeneration from Parkinson's disease in human subjects. Methods A panel of seven NAPEs were quantified by LC-MS/MS in the plasma of 587 individuals (healthy controls, n = 319; Parkinson's disease, n = 268); Random Forest classification and statistical modeling was applied to compare Parkinson's disease versus controls. All p-values obtained in different tests were corrected for multiplicity by controlling the false discovery rate (FDR). Results The results indicate that this panel of NAPEs is able to distinguish female PD patients from the corresponding healthy controls. Further to this, the observed downregulation of these NAPEs is in line with the results in plasma of a mouse model of Parkinson's (6-OHDA). Conclusions In the current study we have shown the downregulation of NAPEs in plasma of PD patients and we thus speculate that these lipids might serve as candidate biomarkers for PD. We also suggest a molecular mechanism, explaining our findings, which involves gut microbiota.
Journal of Lipid Research, 1977
The formation of product by ethanolaminephosphotransferases (EC 2.7.8.1) and cholinephosphotransferases (EC 2.7.8.2) in microsomal fractions from brains and livers of mature rats is increased several fold by 1,2diacyl-,s11-glycerols. With the addition of l-alkyl-2-acyl-mglycerols, we have found an ll-fold increase with brain microsomes and a 20-fold increase with liver microsomes in the synthesis of choline ether lipids (l-alkyl-2-acyl-and 1-alk-1 '-enyl-2-acyl-,~~z-glycero-3-phosphorylcholines). For the synthesis of ethanolamine ether lipids (1-alkyl-2-acyland 1-al k-1 '-enyl-2-acyl-.cn-glycero-3-phosphorylethanolamines), the stimulation of alkylacylglycerols was 7-fold for brain microsomes and 18-fold for liver microsomes. The alkylacyl glycerols (8 mM) also inhibited the synthesis of diacyl phosphoglycerides by 44 to 6596, indicating that the same ethanolaminephosphotransferases and cholinephosphotransferases are utilized for the synthesis of alkylacyl phosphoglycerides and diacyl phosphoglycerides. A desatur-ation of the alkyl groups may take place in the same reaction mixture. The rate of incorporation of phosphorylcholine into alkenylacyl glycerophosphorylcholines (choline plasmalogens) with alkylacylglycerols, cytidine diphosphate choline, and liver microsomes was 15 nmoles per mg protein per hour. The in vitro synthesis of choline plasmalogens with alkylacylglycerols had not been observed previously. The corresponding rate of incorporation of phosphorylethanolamine into ethanolamine plasmalogens was 10 nmoles per mg protein per hour, a value greater than any of the previously reported values for ethanolamine plasmalogen formation from alkylacyl gl ycerophosphorylethanolamines. Supplementary key words cholinephosphotransferase (EC 2.7.8.2). ethanolaminephosphotransferase (EC 2.7.8.1). plasmalogen. phosphatidylethanolamine. phosphatidylcholine. cytidine-5'-diphosphate choline. cytidine-5'-diphosphate ethanolamine. l-alkyl-2-acyl-sn-glycero-3-phosphorylcholines. I-alkyl-2-acyl-,m glycero-3-phosphorylethanolamines