Comprehensive Two-Dimensional Liquid Chromatography Analysis of a Block Copolymer

The properties of a polymeric material are influenced by its underlying molecular distributions, including the molecular-weight (MWD), chemical-composition (CCD), and/or block-length (BLD) distributions. Gradient-elution liquid chromatography (LC) is commonly used to determine the CCD. Due to the limited solubility of polymers, samples are often dissolved in strong solvents. Upon injection of the sample, such solvents may lead to broadened or poorly shaped peaks and, in unfavourable cases, to “breakthrough” phenomena, where a part of the sample travels through the column unretained. To remedy this, a technique called size-exclusion-chromatography gradients or gradient size-exclusion chromatography (gSEC) was developed in 2011. In this work, we aim to further explore the potential of gSEC for the analysis of the CCD, also in comparison with conventional gradient-elution reversed-phase LC, which in this work corresponded to gradient-elution reversed-phase liquid chromatography (RPLC). The influence of the mobile-phase composition, the pore size of the stationary-phase particles, and the column temperature were investigated. The separation of five styrene/ethyl acrylate copolymers was studied with one-dimensional RPLC and gSEC. RPLC was shown to lead to a more-accurate CCD in shorter analysis time. The separation of five styrene/methyl methacrylate copolymers was also explored using comprehensive two-dimensional (2D) LC involving gSEC, i.e. SEC × gSEC and SEC × RPLC. In 2D-LC, the use of gSEC was especially advantageous as no breakthrough could occur.

A sequential liquid chromatographic (LC) procedure for the determination of polycyclic aromatic hydrocarbons (PAH) in a petroleum crude oil and other complex mixtures is described. The procedure includes normal-phase LC on an aminosilane column to isolate fractions containing isomeric PAH and reversed-phase LC on a polymeric C18 column to separate the individual PAH isomers. Appropriate perdeuterated PAH are added to the sample so that each isomeric fraction will contain one internal standard. The perdeuterated PAH are excellent internal standards for this sequential LC procedure. Perdeuterated PAH have normal-phase and reversed-phase LC retention characteristics similar to those of the parent PAH. In the normal-phase LC separation, the perdeuterated PAH elute in the same fraction as the parent PAH. In the reversed-phase LC separation, the perdeuterated PAH elute first and are generally resolved from the parent PAH. The optimized spectrofluorometric detection of each PAH analyte is accomplished by programming appropriate sets of excitation and emission wavelengths to correspond with the elation time of each analyte on the polymeric C18 column. The analytical results obtained from this procedure for the analysis of a shale oil sample [Standard Reference Material (SRM) 1580] and a petroleum crude oil (SRM 1582) are compared to values obtained by gas chromatography - mass spectrometry.

A comprehensive study on solvent effect and establishment of n-hexane solvent system based normal-phase liquid chromatography × reversed-phase liquid chromatography for isolation of natural products

The first oligonucleotide therapeutic was approved by the Food and Drug Administration in 1998, and since then, 12 nucleic acids have been commercialised as medicines. To be approved, the oligonucleotides need to be identified and characterised as well as its related impurities. Different methods exist, but the most commonly used is ion-pairing reversed-phase liquid chromatography with tandem mass spectrometry. The separation obtained depends on the mobile phase and column used. Other methods have been developed, notably by using hydrophilic interaction chromatography and two-dimensional high performance liquid chromatography. Furthermore, ion-pairing reversed-phase high performance liquid chromatography ultra-violet spectroscopy detection and mass spectrometry has been optimised for the analysis of methylated nucleobases due to the utilisation of this modification in the drugs. This review covers the recent advancements in the analysis and characterisation of oligonucleotides in 2021 by high performance liquid chromatography mass spectrometry, notably by hydrophilic interaction chromatography and two-dimensional liquid chromatography but also the different parameters that influence the analysis by ion-pairing reversed-phase high performance liquid chromatography, the characterisation of methylated nucleobases, and the recent software developed for oligonucleotides.

A not-stop-flow online normal-/reversed-phase two-dimensional liquid chromatography–quadrupole time-of-flight mass spectrometry method for comprehensive lipid profiling of human plasma from atherosclerosis patients

Phospholipids (PL) are components of increased value in nutritional applications, especially those rich in polyunsaturated fatty acids (PUFA) such as α‐linolenic acid (ALA, 18:3 n‐3) and linoleic acid (LA, 18:2 n‐6). Currently, the oil processing industry generates by‐products in the form of oilseed and plant cakes that can be valorized for food/feed applications. With the aim of diversifying PL sources and improving their identification in terms of fatty acid (FA) composition, plant cakes obtained from five different sources (camelina, flaxseed, hemp, sesame, walnut) were extracted and analyzed by lipidomics approach using three analytical methods of mass spectrometry (MS): (i) shotgun; (ii) reverse phase liquid chromatography (RPLC) in multiple reaction monitoring (MRM) mode; and (iii) normal phase liquid chromatography (NPLC) with data dependant scan MS fragmentation. We obtained complementary information: while shotgun allowed identifying PL species, RPLC associated to MRM enabled to access high sensitivity and to focus on minority species. Then, NPLC in data dependant scan led to the identification of lysophospholipids and cardiolipin species besides phosphatidylcholine (PC). Among, the five plant cakes analyzed, camelina cake showed the highest percentage of ALA. If ALA was also found in flaxseed and hemp cakes, it was in a lesser extent. Moreover, ALA was located on the sn‐2 position of the glycerol backbone, a valuable position for nutritional applications.Practical applications: The food industry generates large amounts of by‐products, and most of them remain unused. Among them, degumming pastes, i.e., by‐products from the oil refining process, could be valuable sources of phospholipids (PL). These phospholipids present interesting properties, as emulsifiers, thickening agents, or antioxidants, for food or cosmetic industries. Moreover, depending on the plant source, the fatty acid composition in phospholipids could be different from that of the oil (mainly composed of triglycerides instead of phospholipids), especially for α‐linolenic acid (ALA, 18:3 n‐3). Among, the five plant cakes analyzed (camelina, flaxseed, hemp, sesame, walnut), camelina cake showed the highest percentage of ALA. If ALA was also found in flaxseed and hemp cakes, it was in a lesser extent. Moreover, ALA was located in the sn‐2 position of the glycerol, a valuable position for nutritional applications. On the whole, using powerful and complementary analytical procedures for PL characterization allowed widening the potential sources of this type of lipids.Workflow of the plant cake lipid analysis based on three analytical methods of mass spectrometry.

Predicting the behaviour of polydisperse polymers in liquid chromatography under isocratic and gradient conditions

Separation and analysis of phenolic acids from Salvia miltiorrhiza and its related preparations by off-line two-dimensional hydrophilic interaction chromatography × reversed-phase liquid chromatography coupled with ion trap time-of-flight mass spectrometry

Adsorption mechanism of triterpenoid saponins in reversed-phase liquid chromatography and hydrophilic interaction liquid chromatography: Mogroside V as test substance

Lipid oxidation is one of the major causes of food spoilage for lipid-rich foods. In particular, oil-in-water emulsions, like mayonnaises and spreads, are prone to oxidation due to the increased interfacial area that facilitates contact between the lipids and hydrophilic pro-oxidants present in the water phase. Polar, amphiphilic lipid species present at the oil/water interface, like the mono- (MAGs) and di-acylglycerols (DAGs), act as oxidation starters that initiate subsequent oxidation reactions of the non-polar lipids in the oil droplets. A comprehensive two-dimensional liquid chromatography (LC×LC) method with evaporative light-scattering detection (ELSD) was set up to study the composition of the complex mixture of oxidized polar and non-polar lipids. The LC×LC-ELSD method employs size exclusion chromatography (SEC) in the 1D (1st dimension) to separate the various lipid species according to size. In the 2D (2nd dimension), normal-phase liquid chromatography (NPLC) is used to separate the fractions according to their degree of oxidation. The coupling of SEC with NPLC yields a good separation of the oxidized triacylglycerols (TAGs) from the large excess of non-oxidized TAGs. In addition, it allows the isolation of non-oxidized DAGs and MAGs that usually interfere with the detection of a variety of oxidized products that have similar polarities. This method facilitates elucidating how lipid composition affects oxidation kinetics in emulsified foods and will aid in the development of more oxidation-stable products.

In this study, we evaluate the use of various two-dimensional liquid chromatographic methods to characterize water-soluble, synthetically grafted bio-polymers, consisting of long poly(acrylic acid) chains and short maltodextrin grafts. The confirmation of the presence of grafting and the estimation of its extent is challenging. It is complicated by the limited solubility of polymers, their structural dispersity and chemical heterogeneity. Moreover, the starting materials (and other reagents, reaction products and additives) may be present in the product. Reversed-phase liquid chromatography (RPLC), hydrophilic-interaction liquid chromatography (HILIC) and size-exclusion chromatography (SEC) were used to characterize the product, as well as the starting materials. Additionally, fractions were collected for off-line characterization by infrared spectroscopy and mass spectrometry. The one-dimensional separation methods were found to be inconclusive regarding the grafting question. Breakthrough (the early elution of polymer fractions due to strong injection solvents) is shown to be a perpetual problem. This issue is not solved by comprehensive two-dimensional liquid chromatography (LC × LC), but information demonstrating the success of the grafting reaction could be obtained. SEC × RPLC and HILIC × RPLC separations are presented and discussed.

Automated sample clean-up procedure for organophosphorus pesticides in several aquatic organisms using normal phase liquid chromatography

Glycerolipid and cholesterol ester analyses in biological samples by mass spectrometry

Antiparasitic efficacy of flavonoids identified from Psoralea corylifolia against Tetrahymena piriformis in guppy (Poecilia reticulate)

Trace analysis of organic compounds by high-performance liquid chromatography with photoionization detection