Sj. van der Wal

Use of an evaporative light scattering detector in reversed-phase high-performance liquid chromatography of oligomeric surfactants

Abstract The sensitivity of the evaporative light scattering detector was found to be dependent on the analyte concentration and the organic modifier concentration: under reversed-phase gradient conditions calibration is necessary for each analyte. Detection limits for alcohol and carboxylic acid homologues were determined. The advantages of helium over carbon dioxide as a nebulization gas for both volatile and non-volatile or thermolabile analytes are a >30°C lower operating temperature and a five-fold better signal-to-noise ratio. High-resolution separations of polyether alkyl alcohols were effected by reversed-phase gradient high-performance liquid chromatography with ultraviolet transparent mobile phases. Evaporative light scattering detection was compared with refractive index and ultraviolet absorbance detection at 190 nm and found to be superior in signal-to-noise ratio as well as baseline drift. A column-switching system was designed to allow complete compositional analysis of technical samples of ethoxylated alkyl alcohols and their carboxylic acid derivatives.

Boxcar chromatography : A new approach to increased analysis rate and very large column plate numbers

Abstract A new form of column-switching is described which provides significantly faster separations in routine applications of either gas or liquid chromatography. The principle of the method is the partial separation of one or a few compounds of interest on a first column, with diversion of the resulting fraction to a second column. The second column will then be filled with several samples at any given time. High-efficiency separations (10 5 N 7 ) are possible within reasonable per-sample times.

Comparative study of several phase systems for the separation of estrogen conjugates by high-pressure liquid chromatography

Summary A number of liquid-solid and liquid-liquid phase systems with aqueous mobile phases were evaluated for the chromatographic separation of estrogen conjugates. The dependence of retention, selectivity and efficiency on the type of stationary phase, the composition, pH and viscosity of the mobile phase and the temperature was investigated. Polar chemically bonded stationary phases on silica, except LiChrosorb AN, were found to be inferior for the chromatography of steroid conjugates, and so were pellicular and superficially porous anion exchangers and anion exchangers with a polystyrene matrix. Microparticulate octadecyl- and methyl-silica and octadecyl-silica coated with liquid anion exchanger were found to be suitable stationary phases for chromatographic profiling of complex estrogen conjugate mixtures, and were compared with anion-exchange cellulose.

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.

Low viscosity organic modifiers in reversed-phase HPLC

SummaryThree new organic modifier solvent systems for reversed-phase HPLC are recommended which can increase the speed of analysis up to twofold over conventional modifier systems, without a significant sacrifice in general selectivity.

Photometric detection at 185 nm for high-performance liquid chromatography with either isocratic or gradient elution : Assay of mixtures of polyethylene glycol oligomers

Abstract The use of acetonitrile—water mobile phases for reversed-phase high-performance liquid chromatography allows photometric detection at wavelengths as low as 185 nm in both the isocratic and gradient elution modes. At 185 nm almost all compound types exhibit usable detection sensitivity, making the UV spectrophotometer a “universal” detector. Application of detection at 185 nm for the gradient elution separation of various polyethylene glycols allows the routine analysis of mixtures of these compounds, either in unesterified samples or in samples containing both the unesterified and esterified derivatives.

Separation of estrogen glucuronides, sulphates and phosphates on ion-exchange cellulose by high-pressure liquid chromatography

Abstract In continuation of previous work, the selectivity of anion-exchange cllulose systems with respect to thirty estrogen conjugates was studied. The influence of the type and particle size of cellulose anion exchangers, the type and concentration of the counter anion, pH and temperature was investigated. On the basis of the results the optimal choice of the ion-exchange material, the eluent and the operating conditions is demonstrated. The results of the optimization are exemplified by a number of chromatograms showing rapid separations, including the determination of the estrogen conjugate profile of human pregnancy urine.

Automated preseparation derivatization on a capillary electrophoresis instrument

Abstract Fully automated derivatization prior to separation and the separation conditions were established for the determination of d -valine in an excess of l -valine by preseparation derivatization with o-phthalaldehyde and N-acetylcysteine, separation of the resulting diastereomers by micellar electrokinetic capillary chromatography and absorbance detection. The precision and detection limit with absorbance detection are a factor of three worse than those achieved with HPLC and fluorimetric detection. The application of the method at high enantiomeric excess or with chemically impure samples is limited by the detector of the instrument.

Monitoring the in vitro enzyme-mediated degradation of degradable poly(ester amide) for controlled drug delivery by LC-ToF-MS.

To scrutinize materials for specific biomedical applications, we need sensitive and selective analytical methods that can give more insight into the process of their biodegradation. In the present study, the enzymatic degradation of multiblock poly(ester amide) based on natural amino acids, such as lysine and leucine, was performed with serine proteases (α-chymotrypsin (α-CT) and proteinase K (PK)) in phosphate-buffered saline solution at 37 °C for 4 weeks. Fully and partially degraded water-soluble products were analyzed by liquid chromatography hyphenated with time-of-flight mass spectrometry using an electrospray interface (LC-ESI-ToF-MS). Tracking the release of monomeric and oligomeric products into the enzyme media during the course of enzymatic degradation revealed the preferences of α-CT and PK toward ester and amide bonds: both α-CT and PK showed esterase and amidase activity. Although within the experimental time frame up to 30 and 15% weight loss was observed in case of α-CT and PK, respectively, analysis by size exclusion chromatography showed no change in the characteristic molecular-weight averages of the remaining polymer. This suggests that the enzymatic degradation occurs at the surface of this biomaterial. A sustained and linear degradation over a period of 4 weeks supports the potential of this class of poly(ester amide)s for drug delivery applications.

Endgroup-based separation and quantitation of polyamide-6,6 by means of critical chromatography

Polyamide-6,6 is a polycondensation product from the two monomers adipic acid and 1,6-hexamethylenediamine. Depending on the reacted amount of these monomers, different ratios of amine and carboxylic acid endgroups can be formed. Besides linear chains, cyclic polyamides will also be formed. Using critical chromatography, polyamide-6,6 can be separated independently of molar mass. Retention is based solely on endgroup functionality. It is demonstrated that high-molecular-mass polyamide-6,6 (Mw approximately 20,000-30,000) can be separated using this approach. The separation was optimized by using different parameters, such as percentage modifier, temperature and pressure. The concentration of phosphoric acid was used for selective retention of the different endgroup functionalities. Using this property, a new method called critical gradient chromatography was performed where the mobile phase changes from a weak to a strong solvent with respect to the endgroup functionality, while retaining the critical conditions of the backbone unit. Quantification using UV detection is discussed.