Liquid Chromatography At Critical Conditions Research Articles

Quantitative Characterization of Solid Epoxy Resins Using Comprehensive Two Dimensional Liquid Chromatography Coupled with Electrospray Ionization-Time of Flight Mass Spectrometry

A comprehensive multidimensional liquid chromatography system coupled to Electrospray Ionization-Mass Spectrometry (LCxLC-ESI-MS) was developed for detailed characterization and quantitation of solid epoxy resin components. The two orthogonal modes of separation selected were size exclusion chromatography (SEC) in the first dimension and liquid chromatography at critical conditions (LCCC) in the second dimension. Different components present in the solid epoxy resins were separated and quantitated for the first time based on the functional groups and molecular weight heterogeneity. Coupling LCxLC separations with mass spectrometry enabled the identification of components resolved in the two-dimensional space. Several different functional group families of compounds were separated and identified, including epoxy-epoxy and epoxy-alpha-glycol functional oligomers, and their individual molecular weight ranges were determined. Repeatability obtained ranged from 0.5% for the main product to 21% for oligomers at the 0.4% concentration level.

On‐line coupling of liquid chromatography at critical conditions with electrospray ionization tandem mass spectrometry for the characterization of a nitroxide‐mediated poly(ethylene oxide)/polystyrene block copolymer

Coupling of liquid chromatography at critical conditions (LCCC) with on-line mass spectrometry (MS) detection was implemented via an electrospray ionization (ESI) interface, using a mobile phase containing the cationizing agent. Critical conditions established for poly(ethylene oxide) were used to characterize a poly(ethylene oxide)/polystyrene block copolymer (PEO-b-PS) in both MS and MS/MS modes. As co-oligomer molecules were successfully separated according to the PS block size, structural information could be reached from simplified MS spectra. The microstructure of this copolymer, synthesized by nitroxide-mediated polymerization, could further be unambiguously characterized in LCCC/ESI-MS/MS experiments since the PS block size could be reached by both the co-oligomer chromatographic behavior and its MS/MS pattern.

Microwave assisted synthesis and characterization of end functionalized poly(propylene oxide) as model compounds

End functionalized poly(propylene oxide)s with n-alkyl endgroups are synthesized by anionic ring opening polymerization of propylene oxide (PO) in bulk employing controlled microwave heating in sealed vessels. Different alcohols ranging from n-butanol to n-octadecanol are used as initiator and 2–10 propylene oxide units are added on average. A continuous decrease of internal pressure during the sealed vessel experiment reflects the consumption of PO monomer and the completion of the reaction is confirmed by a drop of the internal pressure to zero. The products are characterized by size exclusion chromatography (SEC), liquid chromatography at critical conditions (LCCC) and liquid adsorption chromatography (LAC). SEC with coupled density and refractive index (RI) detection provides information on the chemical composition along the molar mass distribution. LCCC allows a separation of the individual polymer homologous series according to their end groups. In LAC, the alkyl terminated chains elute later than the non-functional chains and can be separated to the baseline according to the number of repeat units.

Liquid chromatography at the critical conditions in pure eluent: An efficient tool for the characterization of functional polystyrenes

Functional polymers are usually separated by liquid chromatography at critical conditions (LC-CC) using a desorli/adsorli mixture as eluent. In this paper, we describe LC-CC using dimethylformamide (DMF) as pure eluent at 72 °C and its application to the analysis of functional polystyrenes. The critical adsorption point is reached by tuning the temperature so that the lack of repeatability is no longer a problem. We analysed several batches of polystyrenes, exhibiting different molecular weights and functionalities. We were able to differentiate di-, mono- and non-functional polystyrenes, bearing carboxylic acid, hydroxyl or nitroxide moieties. The behaviour of these polystyrenes expressed by the experimental results was compared with their behaviour described by the LC-CC theory through the calculation of the effective interaction parameter of each end group. The technique also makes it possible to quantify the fractions of different functional polymers in a blend.

Liquid Chromatography at Critical Conditions: Comprehensive Approach to Sequence-Dependent Retention Time Prediction

An approach to sequence-dependent retention time prediction of peptides based on the concept of liquid chromatography at critical conditions (LCCC) is presented. Within the LCCC approach applied to biopolymers (BioLCCC), the specific retention time corresponds to a particular sequence. In combination with mass spectrometry, this approach provides an efficient tool to solve problems wherein the protein sequencing is essential. In this work, we present a theoretical background of the BioLCCC concept and demonstrate experimentally its feasibility for sequence-dependent LC retention time prediction for peptides. BioLCCC model is based on three notions: (a) a random walk model for a macromolecule chain; (b) an entropy and energy compensation for the macromolecules within the adsorbent pore; and (c) a set of phenomenological parameters for the effective interaction energies of interactions between the amino acid residues and the adsorbent surface. In this work, the phenomenological parameters have been obtained for C18 reversed-phase HPLC. Note, that contrary to alternative additive models for retention time prediction based on summation of the so-called "retention coefficients", the BioLCCC approach takes into account the location of amino acids within the primary structure of a peptide and, thus, allows the identification of the peptides having the same composition of amino acids but differing by their arrangement. As a result, this new approach allows prediction of retention time for any possible amino acid sequence in particular HPLC experiments. In addition, the BioLCCC model lacks of main drawbacks of additive approaches that predict retention time for sequences of limited chain lengths and provide information about amino acid composition only. The proposed BioLCCC approach was characterized experimentally using LTQ FT LC-MS and LC-MS/MS data obtained earlier for Escherichia coli. The HPLC system calibration was performed using peptide retention standards. The results received show a linear correlation between predicted and experimental retention times, with a correlation coefficient, R2, of 0.97 for a peptide standard mixture and 0.9 for E. coli data, respectively, with the standard error below 1 min. The work presents the first description of a BioLCCC approach for high-throughput peptide characterization and preliminary results of its feasibility tests.

High-Temperature Liquid Chromatography at Critical Conditions: Separation of Polystyrene from Blends with Polyethylene and Ethylene-Styrene Block Copolymers

ABSTRACT Liquid chromatography at critical conditions (LCCC) is an important tool for the separation of complex polymers according to chemical composition. For ambient temperatures more than 150 different LCCC separation systems are known, while at temperatures that are suitable for chromatography of polyolefins (i.e. > 130°C) not a single system is known from literature. In this article we present LCCC conditions for polystyrene at a temperature of 140°C and their application to the analysis of polymer blends composed of polystyrene and polyethylene or styrene-ethylene block copolymers.

Development of high-temperature separation techniques for the chemical composition analysis of ethylene - methyl methacrylate block copolymers

AbstractEthylene - methyl methacrylate block copolymers are semicrystalline polymers that dissolve in organic solvents only at high temperatures. Accordingly, microstructure analysis by solution methods must be conducted at temperatures above 130°C. For the analysis of block copolymers of different compositions several analytical techniques were used, including high-temperature size-exclusion chromatography (SEC), hyphenated SEC-FTIR, and CRYSTAF (crystallisation analysis fractionation). While SEC with refractive index detection indicated a certain multimodality of the samples, SEC coupled with FTIR revealed that the samples were chemically inhomogeneous and may contain homo- and copolymer fractions. The presence of polyethylene and poly(methyl methacrylate) homopolymers in the copolymer samples was confirmed by CRYSTAF analysis, when the total concentration as well as the carbonyl group distribution were monitored separately. Chromatographic separation of the different sample components was achieved when liquid chromatography at critical conditions (LC-CC) was used. For the first time, true high-temperature LC-CC methods were developed operating at a column temperature of 140°C. As the stationary phase, silica gel was used. Suitable mobile phases were binary mixtures of 1,2,4-trichlorobenzene or 1,2- dichlorobenzene with cyclohexanone. Using LC-CC, the samples were separated into the copolymer and the homopolymer fractions.

Polystyrene- block-poly(ethylene oxide) from nitroxide mediated polymerization: detection of minor species by coupled chromatographic techniques

A polystyrene- block-poly(ethylene oxide) (PS- b-PEO) copolymer is synthesised in an original manner by nitroxide mediated polymerization of styrene from a PEO macroalkoxyamine. Minor amounts of side products are unavoidably present in the final polymer, including: traces of macroalkoxyamine, PEO- b-PS- b-PEO triblock copolymer—a product of the recombination of PEO- b-PS macro-radical during the initial stage of the polymerization—and hPS—a product of thermal initiation. These species do not represent more than a little percentage of the total composition. They are hardly detectable with classical SEC. We use liquid chromatography at critical conditions (LC-CC), 2D chromatography and LC-CC-NMR to get an exhaustive description of the chemical composition distributions (CCD) and of the molar mass distributions (MMD) of all the minor species quoted above. The results are consistent with the synthesis route and prove the accuracy of the LC-CC based techniques in analysing even very small amounts of species.

Star‐shaped poly(L‐lactide)s with variable numbers of hydroxyl groups at polyester arms chain‐ends and directly attached to the star‐shaped core—Controlled synthesis and characterization

AbstractStar‐shaped poly(L‐lactide)s (PLAs) bearing variable numbers of secondary hydroxyl groups at linear arms chain‐ends and primary hydroxyl groups directly attached to dipentaerithritol core (DPE) ((HO)6−xDPE(PLA‐OH)x, where x = 1–6) were prepared and then analyzed by means of size exclusion chromatography (SEC), 1H NMR spectroscopy, MALDI‐TOF mass spectrometry, and eventually by Liquid Chromatography at Critical Conditions (LC‐CC). First, starting from DPE(OH)6 a series of polyols with various number of hydroxyl groups has been obtained ((BnO)6−xDPE(OH)x, where Bn denotes benzyl moiety and x = 1–6). The living ring‐opening polymerization of L‐lactide (LA) with (BnO)6−xDPE(OH)x/tin(II) octoate mixtures as initiating and catalytic system led to star‐shaped (BnO)6−xDPE(PLA‐OH)x polymers with molar masses (Mn) controlled by LA and DPE concentrations ratio in the feed. Finally, deprotection (via hydrogenation) gave a series of (HO)6−xDPE(PLA‐OH)x PLA's. SEC (with Multiangle Laser Light Scattering Detector (MALLS)), NMR, and MALDI‐TOF analyses confirmed the assumed structures and Mn's of the prepared (BnO)6−xDPE(PLA‐OH)x and (HO)6−xDPE(PLA‐OH)x PLA's. LC‐CC measurements revealed that for (BnO)6−xDPE (PLA‐OH)x series the elution volumes increase monotonically with the increasing number of –PLA‐OH arms in one macromolecule and are independent on the given PLA molar mass because of the critical conditions. Contrary to the polymers having the protected core hydroxyl groups, the elution volume for (HO)6−xDPE(PLA‐OH)x series decreases with the increasing number of ‐PLA‐OH arms reaching a minimum value for 4‐arm PLA and then slightly increases for 5‐ and 6‐arm PLA. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 43: 6116–6133, 2005

Comprehensive two-dimensional liquid chromatography and hyphenated liquid chromatography to study the degradation of poly(bisphenol A)carbonate

Size exclusion chromatography (SEC), gradient polymer elution chromatography (GPEC) and liquid chromatography at critical conditions (LC-CC) have been developed and applied to observe chemical changes in poly(bisphenol A)carbonate (PC) due to hydrolytic degradation. Especially LC-CC appeared to be very successful to observe differences in functionality of PC as result of hydrolytic degradation. Observed differences due to degradation could be identified by (semi) on-line coupling to matrix assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The differences in functionality could be attributed to the formation of different end-groups, i.e. OH end-groups. In addition, comprehensive two-dimensional liquid chromatography (2D-LC) has been applied successfully to study the hydrolytic degradation of PC. LC-CC × SEC showed that the formation of PC with different end-groups occurred over the whole molecular mass range. This information could not be obtained with the separate liquid chromatographic techniques, thereby illustrating the added value of 2D-LC.

Semi-Quantitative Determination of Free Cyclic and Linear Polytetrahydrofurane in Coatings Using Critical Chromatography

Liquid chromatography at critical conditions (LCCC) is being used more frequently, especially for the determination of cyclic structures in a matrix of the parent linear molecules, analyses that cannot be performed easily by other techniques. In practice, analyses are often more complex than separating the linear from the cyclic structures, e.g. in the determination of low concentrations of free (linear and cyclic) polyols like polytetrahydrofurane (pTHF) in functional polymers and coatings. To this end different chromatographic phase systems at the critical conditions for the polyols were investigated to obtain the maximum amount of information. The identification of different oligomeric series was performed by means of an on-line coupling of LCCC with atmospheric pressure chemical ionization – mass spectrometry (APCI-MS). Mass summated chromatograms were introduced to improve the signal to noise ratio of specific oligomeric series. MS cannot be used for quantitative purposes here; therefore the evaporative light scattering detector (ELSD) and the refractive index (RI) detectors were evaluated for routine analyses. Specific tailing problems observed with a drift tube in the ELSD are discussed. Two LCCC systems (reversed phase and normal phase) were compared for best selectivity

Use of gradient, critical, and two‐dimensional chromatography in the analysis of styrene‐ and methyl methacrylate‐grafted epoxidized natural rubber

AbstractThe growing number of heterogeneous polymeric species that are being synthesized places increasing demands on existing analytical techniques. Although size‐exclusion chromatography (SEC) has established itself as a powerful analytical tool, it has its limits when complex polymers, e.g., graft copolymers, must be analyzed. In this case, complementary techniques such as gradient HPLC and liquid chromatography at critical conditions (LCCC) are more favorable. The present study describes the synthesis and analysis of methyl methacrylate‐ and styrene‐grafted epoxidized natural rubber by different chromatographic techniques. The grafting efficiency was evaluated by gradient HPLC under normal and reversed phase conditions. Methyl methacrylate‐grafted ENR50 was further analyzed by LCCC, where separation of the rubber and grafted rubber occurred according to chemical composition but was independent of the molar mass of the methyl methacrylate homopolymers. This was followed by the combination of LCCC and SEC, where separation was achieved in two dimensions. Relevant deductions were made of both the chemical composition distribution and the molar mass distribution of the functional groups of methyl methacrylate‐grafted ENR50. © 2003 Wiley Periodicals, Inc. J Appl Polym Sci 88: 2530–2538, 2003

Characterization of star-shaped poly( l-lactide)s by liquid chromatography at critical conditions

A series of linear and star-shaped poly( l-lactide)s (PLA's) have been prepared by living polymerization of l,l-dilactide (LA) and analyzed by liquid chromatography at critical conditions (LC-CC). For the analysis of the PLA's LC-CC conditions have been used corresponding to silica gel as the stationary phase and a mixture of 1,4-dioxane/ n-hexane (56.25/43.75 by vol%) at 50 °C as the mobile phase. At the critical point of adsorption, a series of linear C 4H 9-PLA-OH's having molar masses ( M n) in the range from 2.3×10 3 to 7.4×10 4, prepared by ring-opening polymerization of LA initiated with Sn(OC 4H 9) 2 (THF, 80 °C), showed no dependence of the elution volumes on molar mass. In subsequent experiments, star-shaped PLA's bearing various numbers of PLA-OH arms (R-(PLA-OH) x ) have been prepared in a controlled synthesis starting from various polyols (R-(OH) x ) containing exclusively primary hydroxyl groups: diethyleneglycol ( x=2), trimethylolpropane ( x=3), di(trimethylolpropane) ( x=4), dipentaerithritol ( x=6), and poly(3-ethyl-3-hydroxymethyloxetane) (〈x〉=13.4) and LA monomer. As coinitiator/catalyst tin(II) octoate (Sn(Oct) 2) has been used (bulk polymerization, 120 °C). 1H NMR analysis of the resulting star-shaped polymers revealed that all OH-groups in the polyols started growth of the PLA chains. The series of star-shaped PLA's have been analyzed by LC-CC as well as by two-dimensional (2D) chromatography (i.e. LC-CC versus size exclusion chromatography (SEC)) with regard to possible structural imperfections. It has been shown, that the LC-CC elution volumes of the resulting R-(PLA-OH) x increase with the number of PLA-OH arms, allowing discrimination of the individual R-(PLA-OH) x 's in their mixture. An exponential increase of the retention volume as a function of the number of arms has been found. Eventually, LC-CC measurements of the elution volumes carried out for acetylated star-shaped PLA's (R(PLA-OOCCH 3) x ) have shown that for the interactions of the R-(PLA-OH) x macromolecules with the column packing the hydroxyl end-groups are mostly responsible.

Separation and quantification of the linear and cyclic structures of polyamide-6 at the critical point of adsorption

The linear and cyclic structures of polyamide-6 were separated by liquid chromatography at critical conditions (LCCC) and identified with different mass spectrometric (MS) techniques and quantitated by LCCC with evaporative light-scattering detection (ELSD). Electrospray ionization MS was not suitable to identify the higher cyclic structures. For this purpose, matrix-assisted laser desorption ionization time-of-flight MS performed better and cyclic and linear structures were oligomerically resolved and separately identified in the mass spectrometer. The highest cyclic structure present and detected was the cyclic pentacontamer. It could be demonstrated that cyclic and linear oligomers follow different ionization and fragmentation routes/patterns. Quantification with ELSD of the components separated by LCCC using a universal calibration curve or an iterative procedure was developed. An area correction to account for different peak widths of coeluting components improves precision and accuracy of the calibration curve and improves quantitation accuracy for the samples analyzed. With these corrected values, no molecular mass dependency was observed for the cyclic and linear structures. Under critical conditions, the linear and cyclic structures of polyamide-6 were separated, identified and quantified.