Validation of the extended Tanaka column characterization protocol by multivariate analysis of chromatographic retention of low-molecular-weight analytes on reversed phase columns using methanol and acetonitrile as organic modifiers

2012 ◽  
Vol 35 (19) ◽  
pp. 2592-2598 ◽  
Author(s):  
Melvin R Euerby ◽  
Matthew James ◽  
Bengt-Olof Axelsson ◽  
Olof Rosén ◽  
Patrik Petersson
Keyword(s):  
Molecular Weight ◽  
Multivariate Analysis ◽  
Reversed Phase ◽  
Chromatographic Retention ◽  
Low Molecular Weight ◽  
Organic Modifiers

scholarly journals Simultaneous Determination of Human Serum Albumin and Low-Molecular-Weight Thiols after Derivatization with Monobromobimane

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3321
Author(s):  
Katarzyna Kurpet ◽  
Rafał Głowacki ◽  
Grażyna Chwatko
Keyword(s):  
Molecular Weight ◽  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Simultaneous Determination ◽  
Reversed Phase ◽  
Fluorescence Labeling ◽  
Detector Response ◽  
Low Molecular Weight

Biothiols are extremely powerful antioxidants that protect cells against the effects of oxidative stress. They are also considered relevant disease biomarkers, specifically risk factors for cardiovascular disease. In this paper, a new procedure for the simultaneous determination of human serum albumin and low-molecular-weight thiols in plasma is described. The method is based on the pre-column derivatization of analytes with a thiol-specific fluorescence labeling reagent, monobromobimane, followed by separation and quantification through reversed-phase high-performance liquid chromatography with fluorescence detection (excitation, 378 nm; emission, 492 nm). Prior to the derivatization step, the oxidized thiols are converted to their reduced forms by reductive cleavage with sodium borohydride. Linearity in the detector response for total thiols was observed in the following ranges: 1.76–30.0 mg mL−1 for human serum albumin, 0.29–5.0 nmol mL−1 for α-lipoic acid, 1.16–35 nmol mL−1 for glutathione, 9.83–450.0 nmol mL−1 for cysteine, 0.55–40.0 nmol mL−1 for homocysteine, 0.34–50.0 nmol mL−1 for N-acetyl-L-cysteine, and 1.45–45.0 nmol mL−1 for cysteinylglycine. Recovery values of 85.16–119.48% were recorded for all the analytes. The developed method is sensitive, repeatable, and linear within the expected ranges of total thiols. The devised procedure can be applied to plasma samples to monitor biochemical processes in various pathophysiological states.

scholarly journals Proteomic Determination of Low-Molecular-Weight Glutenin Subunit Composition in Aroona Near-Isogenic Lines and Standard Wheat Cultivars

2021 ◽  
Vol 22 (14) ◽  
pp. 7709
Author(s):  
Kyoungwon Cho ◽  
You-Ran Jang ◽  
Sun-Hyung Lim ◽  
Susan B. Altenbach ◽  
Yong Q. Gu ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Allelic Variation ◽  
Reversed Phase ◽  
Glutenin Subunit ◽  
Low Molecular Weight ◽  
Near Isogenic Lines ◽  
Wheat Cultivars ◽  
Reliable Determination ◽  
Isogenic Lines

The low-molecular weight glutenin subunit (LMW-GS) composition of wheat (Triticum aestivum) flour has important effects on end-use quality. However, assessing the contributions of each LMW-GS to flour quality remains challenging because of the complex LMW-GS composition and allelic variation among wheat cultivars. Therefore, accurate and reliable determination of LMW-GS alleles in germplasm remains an important challenge for wheat breeding. In this study, we used an optimized reversed-phase HPLC method and proteomics approach comprising 2-D gels coupled with liquid chromatography–tandem mass spectrometry (MS/MS) to discriminate individual LMW-GSs corresponding to alleles encoded by the Glu-A3, Glu-B3, and Glu-D3 loci in the ‘Aroona’ cultivar and 12 ‘Aroona’ near-isogenic lines (ARILs), which contain unique LMW-GS alleles in the same genetic background. The LMW-GS separation patterns for ‘Aroona’ and ARILs on chromatograms and 2-D gels were consistent with those from a set of 10 standard wheat cultivars for Glu-3. Furthermore, 12 previously uncharacterized spots in ‘Aroona’ and ARILs were excised from 2-D gels, digested with chymotrypsin, and subjected to MS/MS. We identified their gene haplotypes and created a 2-D gel map of LMW-GS alleles in the germplasm for breeding and screening for desirable LMW-GS alleles for wheat quality improvement.

scholarly journals Validation and Application of a Derivatization-Free RP-HPLC-DAD Method for the Determination of Low Molecular Weight Salivary Metabolites

2020 ◽  
Vol 17 (17) ◽  
pp. 6158
Author(s):  
Beatrice Campanella ◽  
Tommaso Lomonaco ◽  
Edoardo Benedetti ◽  
Massimo Onor ◽  
Riccardo Nieri ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Low Cost ◽  
Reversed Phase ◽  
Low Molecular Weight ◽  
Single Individual ◽  
Sampling Device ◽  
Non Invasive ◽  
Minimal Sample ◽  
Whole Saliva

Saliva is an interesting, non-conventional, valuable diagnostic fluid. It can be collected using standardized sampling device; thus, its sampling is easy and non-invasive, it contains a variety of organic metabolites that reflect blood composition. The aim of this study was to validate a user-friendly method for the simultaneous determination of low molecular weight metabolites in saliva. We have optimized and validated a high throughput, direct, low-cost reversed phase liquid chromatographic method with diode array detection method without any pre- or post-column derivatization. We indexed salivary biomolecules in 35 whole non-stimulated saliva samples collected in 8 individuals in different days, including organic acids and amino acids and other carbonyl compounds. Among these, 16 whole saliva samples were collected by a single individual over three weeks before, during and after treatment with antibiotic in order to investigate the dynamics of metabolites. The concentrations of the metabolites were compared with the literature data. The multianalyte method here proposed requires a minimal sample handling and it is cost-effectiveness as it makes possible to analyze a high number of samples with basic instrumentation. The identification and quantitation of salivary metabolites may allow the definition of potential biomarkers for non-invasive “personal monitoring” during drug treatments, work out, or life habits over time.

scholarly journals Separation of low molecular weight rapeseed proteins by RP-HPLC-DAD – a short report

2011 ◽  
Vol 24 (No. 1) ◽  
pp. 41-44 ◽  
Author(s):  
A. Kosińska ◽  
ChavanUD ◽  
R. Amarowicz
Keyword(s):  
Molecular Weight ◽  
Sinapic Acid ◽  
Reversed Phase ◽  
Low Molecular Weight ◽  
Short Report ◽  
Uv Spectrum ◽  
Phenolic Constituents ◽  
Separation Of Proteins ◽  
Two Peaks ◽  
Low Molecular Weight Proteins

Low molecular weight proteins were extracted and isolated from rapeseed and analysed using the HPLC-DAD method. The separation of proteins and phenolic compounds was done on the reversed phase C<sub>18</sub> column with a gradient of acetonitrile in water. The chromatogram was characterised by two peaks of low molecular weight proteins with the retention times of 19.92 and 23.24 min. Additional three main peaks of phenolic constituents were recorded on the chromatogram. One of them with maximum of UV spectrum at 328 nm was identified as sinapic acid derivatives.

Reversed-Phase Membrane Inlet Mass Spectrometry Applied to the Real-Time Monitoring of Low Molecular Weight Alcohols in Chloroform

2002 ◽  
Vol 74 (1) ◽  
pp. 300-304 ◽  
Author(s):  
Colin S. Creaser ◽  
David Gómez Lamarca ◽  
Jeffrey Brum ◽  
Christopher Werner ◽  
Anthony P. New ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Molecular Weight ◽  
Real Time ◽  
Reversed Phase ◽  
Low Molecular Weight ◽  
Real Time Monitoring ◽  
The Real ◽  
Membrane Inlet Mass Spectrometry

Optimization of gradient reversed phase chromatographic peak capacity for low molecular weight solutes

2013 ◽  
Vol 1273 ◽  
pp. 57-65 ◽  
Author(s):  
Arianne Soliven ◽  
Imad A. Haidar Ahmad ◽  
Marcelo R. Filgueira ◽  
Peter W. Carr
Keyword(s):  
Molecular Weight ◽  
Reversed Phase ◽  
Peak Capacity ◽  
Low Molecular Weight ◽  
Chromatographic Peak
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