Improved sample preparation and rapid extraction of prostacyclin and thromboxane metabolites from biological samples by using reversed-phase octadecylsilyl-silica cartridges

1984 ◽  
Vol 12 (5) ◽  
pp. 834-835 ◽  
Author(s):  
INDRAJIT DAS
Keyword(s):  
Sample Preparation ◽  
Biological Samples ◽  
Reversed Phase ◽  
Rapid Extraction

A Fast and Sensitive Method Combining Reversed-Phase Chromatography with High Resolution Mass Spectrometry to Quantify 2-Fluoro-2-Deoxyglucose and Its Phosphorylated Metabolite for Determining Glucose Uptake

2019 ◽  
Author(s):  
Ashley Williams ◽  
Deborah Muoio ◽  
Guofang Zhang
Keyword(s):  
Glucose Uptake ◽  
Biological Samples ◽  
High Resolution Mass Spectrometry ◽  
Reversed Phase ◽  
Sodium Adduct ◽  
Negative Mode ◽  
Glucose Analogues ◽  
Positive Mode ◽  
Q Exactive ◽  
Glucose Analogue

Quantative measurements of the glucose analogue, 2-deoxyglucose (2DG), and its phosphorylated metabolite (2-deoxyglucose-6-phosphate (2DG-6-P)) are critical for the measurement of glucose uptake. While the field has long identified the need for sensitive and reliable assays that deploy non-radiolabled glucose analogues to assess glucose uptake, no analytical MS-based methods exist to detect trace amounts in complex biological samples. In the present work, we show that 2DG is poorly suited for MS-based methods due to interfering metabolites. We therefore developed and validated an alternative C18-based LC-Q-Exactive-Orbitrap-MS method using 2-fluoro-2-deoxyglucose (2FDG) to quantify both 2FDG and 2FDG-6-P by measuring the sodium adduct of 2FDG in the positive mode and deprotonation of 2FDG-6-P in the negative mode. The low detection limit of this method can reach 81.4 and 48.8 fmol for both 2FDG and 2FDG-6-P, respectively. The newly developed method was fully validated via calibration curves in the presence and absence of biological matrix. The present work is the first successful LC-MS method that can quantify trace amounts of a nonradiolabeled glucose analogue and its phosphorylated metabolite and is a promising analytical method to determine glucose uptake in biological samples.

Investigation of Phenolic Content in Five Different Pine Barks Species Grown in Turkey by HPLC-UV and LC–MS

2021 ◽  
Author(s):  
Mehmet Emin Şeker ◽  
Ali Çelik ◽  
Kenan Dost ◽  
Ayşegül Erdoğan
Keyword(s):  
Sample Preparation ◽  
Orthophosphoric Acid ◽  
Phenolic Content ◽  
Reversed Phase ◽  
Uv Detector ◽  
Epicatechin Gallate ◽  
Rp Hplc ◽  
Phenolic Constituents ◽  
Preparation Step ◽  
Very High

Abstract Investigation of phenolic content from different pine bark species grown in Turkey was performed using a reversed-phase high pressure liquid chromatography with ultraviolet (RP-HPLC-UV) method. All phenolic constituents were separated in <26 min on reversed-phase C18 column with gradient mobile phase that consists of orthophosphoric acid, methanol and acetonitrile. Detections were made on an UV detector at 280 nm and at a flow rate of 1 mL/min. Samples were prepared according to Masqueller’s conventional sample preparation method with slight modifications. To avoid the reduction in extraction efficiency the sample preparation step was carried out under argon atmosphere. The linearity of the method was between 0.9994 and 0.9999. The detection limits for the five phenolic constituents ranged from 0122 to 0.324 mg/L. Catechin and taxifolin were found in all pine barks at a concentration of 0.065 ± 0.002–1.454 ± 0.004 and 0.015 ± 0.001–23.164 ± 0.322 mg/g, respectively. Epicatechin was determined in four pine barks between 0.027 ± 0.001 and 0.076 ± 0.002 mg/g, ferulic acid in two pine barks between 0.010 ± 0.001 and 0.022 ± 0.001 mg/g and epicatechin gallate in only one of the pine barks at 0.025 ± 0.001 mg/g. Finally, the total amount of phenolic compounds and antioxidant capacities of the pine barks were found to be very high.

Recent applications of microextraction sample preparation techniques in biological samples analysis

2021 ◽  
Author(s):  
Seyed Mosayeb Daryanavard ◽  
Hesane Zolfaghari ◽  
Abbi Abdel‐Rehim ◽  
Mohamed Abdel‐Rehim
Keyword(s):  
Sample Preparation ◽  
Biological Samples ◽  
Preparation Techniques

scholarly journals An optimized LC-HRMS untargeted metabolomics workflow for multi-matrices investigations in the three-spined stickleback

PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0260354
Author(s):  
Emmanuelle Lebeau-Roche ◽  
Gaëlle Daniele ◽  
Aurélie Fildier ◽  
Cyril Turies ◽  
Odile Dedourge-Geffard ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Gasterosteus Aculeatus ◽  
High Resolution Mass Spectrometry ◽  
Reversed Phase ◽  
Research Field ◽  
Untargeted Metabolomics ◽  
Whole Body ◽  
Systematic Analysis ◽  
Fish Model ◽  
Ionization Mode

Environmental metabolomics has become a growing research field to understand biological and biochemical perturbations of organisms in response to various abiotic or biotic stresses. It focuses on the comprehensive and systematic analysis of a biologic system’s metabolome. This allows the recognition of biochemical pathways impacted by a stressor, and the identification of some metabolites as biomarkers of potential perturbations occurring in a body. In this work, we describe the development and optimization of a complete reliable methodology based on liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS) for untargeted metabolomics studies within a fish model species, the three-spined stickleback (Gasterosteus aculeatus). We evaluated the differences and also the complementarities between four different matrices (brain, gills, liver and whole fish) to obtain metabolome information. To this end, we optimized and compared sample preparation and the analytical method, since the type and number of metabolites detected in any matrix are closely related to these latter. For the sample preparation, a solid-liquid extraction was performed on a low quantity of whole fish, liver, brain, or gills tissues using combinations of methanol/water/heptane. Based on the numbers of features observed in LC-HRMS and on the responses of analytical standards representative of different metabolites groups (amino acids, sugars…), we discuss the influence of the nature, volume, and ratio of extraction solvents, the sample weight, and the reconstitution solvent. Moreover, the analytical conditions (LC columns, pH and additive of mobile phases and ionization modes) were also optimized so as to ensure the maximum metabolome coverages. Thus, two complementary chromatographic procedures were combined in order to cover a broader range of metabolites: a reversed phase separation (RPLC) on a C18 column followed by detection with positive ionization mode (ESI+) and a hydrophilic interaction chromatography (HILIC) on a zwitterionic column followed by detection with negative ionization mode (ESI-). This work provides information on brain, gills, liver, vs the whole body contribution to the stickleback metabolome. These information would help to guide ecotoxicological and biomonitoring studies.

scholarly journals Improved HPLC Conditions to Determine Eumelanin and Pheomelanin Contents in Biological Samples Using an Ion Pair Reagent

2020 ◽  
Vol 21 (14) ◽  
pp. 5134 ◽  
Author(s):  
Shosuke Ito ◽  
Sandra Del Bino ◽  
Tomohisa Hirobe ◽  
Kazumasa Wakamatsu
Keyword(s):  
Dicarboxylic Acid ◽  
Biological Samples ◽  
Potassium Phosphate ◽  
Structural Diversity ◽  
Tricarboxylic Acid ◽  
Reversed Phase ◽  
Hplc Method ◽  
Original Method ◽  
Ion Pair ◽  
Improved Method

Alkaline hydrogen peroxide oxidation (AHPO) of eumelanin and pheomelanin, two major classes of melanin pigments, affords pyrrole-2,3,5-tricarboxylic acid (PTCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) from eumelanin and thiazole-2,4,5-tricarboxylic acid (TTCA) and thiazole-4,5-dicarboxylic acid (TDCA) from pheomelanin. Quantification of these five markers by HPLC provides useful information on the quantity and structural diversity of melanins in various biological samples. HPLC analysis of these markers using the original method of 0.1 M potassium phosphate buffer (pH 2.1):methanol = 99:1 (85:15 for PTeCA) on a reversed-phase column had some problems, including the short lifetime of the column and, except for the major eumelanin marker PTCA, other markers were occasionally overlapped by interfering peaks in samples containing only trace levels of these markers. These problems can be overcome by the addition of an ion pair reagent for anions, such as tetra-n-butylammonium bromide (1 mM), to retard the elution of di-, tri- and tetra-carboxylic acids. The methanol concentration was increased to 17% (30% for PTeCA) and the linearity, reproducibility, and recovery of the markers with this improved method is good to excellent. This improved HPLC method was compared to the original method using synthetic melanins, mouse hair, human hair, and human epidermal samples. In addition to PTCA, TTCA, a major marker for pheomelanin, showed excellent correlations between both HPLC methods. The other markers showed an attenuation of the interfering peaks with the improved method. We recommend this improved HPLC method for the quantitative analysis of melanin markers following AHPO because of its simplicity, accuracy, and reproducibility.

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