Elution Profiles (elution + profile)

Distribution by Scientific Domains

Selected Abstracts

Lyophilization to improve drug delivery for chitosan-calcium phosphate bone scaffold construct: A preliminary investigation

Benjamin T. Reves
Abstract Lyophilization was evaluated in chitosan-calcium phosphate microspheres and scaffolds to improve drug delivery of growth factors and antibiotics for orthopedic applications. The dual delivery of an antibiotic and a growth factor from a composite scaffold would be beneficial for treatment of complex fracture sites, such as comminuted fractures and segmental bone defects. The aim of this investigation was to increase the loading capacity of the composite by taking advantage of the increased porosity, due to lyophilization, and to produce an extended elution profile using a secondary chitosan-bead coating. The physiochemical properties of the composite were investigated, and loading and elution studies were performed with alkaline phosphatase (ALP), bone morphogenetic protein-2 (BMP-2), and amikacin. Lyophilization was found to increase the surface area of scaffolds by over 400% and the porosity of scaffolds by 50%. Using ALP as a model protein, the loading capacity was increased by lyophilization from 4.3 ± 2.5 to 24.6 ± 3.6 ,g ALP/mg microspheres, and the elution profile was extended by a supplemental chitosan coating. The loading capacity of BMP-2 for composite microspheres was increased from 74.4 ± 3.7 to 102.1 ± 8.0 ,g BMP-2/g microspheres with lyophilization compared with nonlyophilized microspheres. The elution profiles of BMP-2 and the antibiotic amikacin were not extended with the supplemental coating. Additional investigations are planned to improve these elution characteristics for growth factors and antibiotics. © 2009 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009 [source]

Poster Sessions AP13: Novel Techniques and Technologies

J. K. Yao
Studies of the antioxidant defense system and the monoamine metabolic pathways are often complicated by cumbersome analytical methods, which require separate and multistep extraction and chemical reaction procedures. Thus, measurements of multiple parameters are limited in relatively small biological samples. High performance liquid chromatography (HPLC) coupled with a Coulometric Multi-Electrode Array System (CMEAS) provides us a convenient and most sensitive tool to measure low molecular weight, redox-active compounds in biological sample. The deproteinized sample was analyzed on a HPLC coupled with a 16-channel CMEAS, which incremented from 60 to 960 mV in 60 mV steps. Each sample was run on a single column (Meta-250, 4.6 × 250 mm) under a 150-minute complex gradient that ranged from 0% B (A: 1.1% pentane sulfonic acid) to 20% B (B: 0.1 m lithium acetate in mixture of methanol, acetonenitrile and isopropanol), with a flow rate of 0.5 mL/min. We have developed an automated procedure to simultaneously measure various antioxidant, oxidative stress marker, and monoamine metabolites in a single column with binary gradient. No other chemical reactions are necessary. In order to reduce the running time and yet achieve a reproducible retention time by the autosampler injection, our gradient elution profile was modified to produce a shorter equilibration time and to compensate for the initial contamination of mobile phase B following the first injection. Without the use of two columns in series and peak suppresser/gradient mixer, we have simplified the previously published method to measure over 20 different antioxidants, oxidative stress markers and monoamine metabolites simultaneously in biological samples. [source]

Application of hydrophilic interaction chromatography retention coefficients for predicting peptide elution with TFA and methanesulfonic acid ion-pairing reagents

Chad E. Wujcik
Abstract Hydrophilic retention coefficients for 17 peptides were calculated based on retention coefficients previously published for TSKgel silica-60 and were compared with the experimental elution profile on a Waters Atlantis HILIC silica column using TFA and methanesulfonic acid (MSA) as ion-pairing reagents. Relative peptide retention could be accurately determined with both counter-ions. Peptide retention and chromatographic behavior were influenced by the percent acid modifier used with increases in both retention and peak symmetry observed at increasing modifier concentrations. The enhancement of net peptide polarity through MSA pairing shifted retention out by nearly five-fold for the earliest eluting peptide, compared with TFA. Despite improvements in retention and efficiency (Neff) for MSA over TFA, a consistent reduction in calculated selectivity (,) was observed. This result is believed to be attributed to the stronger polar contribution of MSA masking and diminishing the underlying influence of the amino acid residues of each associated peptide. Finally, post-column infusion of propionic acid and acetic acid was evaluated for their potential to recover signal intensity for TFA and MSA counter-ions for LC-ESI-MS applications. Acetic acid generally yielded more substantial signal improvements over propionic acid on the TFA system while minimal benefits and some further reductions were noted with MSA. [source]

Design of Affinity Tags for One-Step Protein Purification from Immobilized Zinc Columns

Richard S. Pasquinelli
Affinity tags are often used to accomplish recombinant protein purification using immobilized metal affinity chromatography. Success of the tag depends on the chelated metal used and the elution profile of the host cell proteins. Zn(II)-iminodiacetic acid (Zn(II)-IDA) may prove to be superior to either immobilized copper or nickel as a result of its relatively low binding affinity for cellular proteins. For example, almost all Escherichiacoli proteins elute from Zn(II)-IDA columns between pH 7.5 and 7.0 with very little cellular protein emerging at pH values lower than 7.0. Thus, a large portion of the Zn(II)-IDA elution profile may be free of contaminant proteins, which can be exploited for one-step purification of a target protein from raw cell extract. In this paper we have identified several fusion tags that can direct the elution of the target protein to the low background region of the Zn(II)-IDA elution profile. These tags allow targeting of proteins to different regions of the elution profile, facilitating purification under mild conditions. [source]

Determination of the cis-trans isomerization barrier of enalaprilat by dynamic capillary electrophoresis and computer simulation

Oliver Trapp
Abstract Dynamic capillary electrophoresis (DCE) and computer simulation of the elution profiles with the stochastic model has been applied to determine the isomerization barriers of the angiotensin converting enzyme inhibitor enalaprilat. The separation of the rotational cis-trans isomeric drug has been performed in an aqueous 20 mM borate buffer at pH 9.3. Interconversion profiles featuring plateau formation and peak broadening were observed. To evaluate the rate constants kcis,trans and ktrans,cis of the cis-trans isomerization from the experimental electropherograms obtained by dynamic capillary electrophoresis, elution profiles were analyzed by a simulation with iterative convergence to the experimental data using the ChromWin software which requires the total migration times of the individual isomers tR, the electroosmotic break-through time t0, the plateau height hplateau, the peak widths at half height of the individual isomers wh, as well as the peak ratio of the isomers as experimental data input. From temperature-dependent measurements between 0° and 15°C the thermodynamic parameters ,G, ,H and ,S, the rate constants kcis,trans and ktrans,cis and the kinetic activation parameters ,G,, ,H,, and ,S, of the cis-trans isomerization of enalaprilat were obtained. From the activation parameters the isomerization barriers at 37°C were calculated to be ,G,trans,cis = 87.2 kJ·mol,1 and ,G,cis,trans = 91.9 kJ·mol,1. [source]

Dispersion in non-ideal packed beds

AICHE JOURNAL, Issue 2 2010
U. M. Scheven
Abstract This work reanalyzes published time series dispersion traces (Han et al., AIChE J. 1985;31:277,288) from step input tracer passages recorded at different locations along the length of a packed bed filled with monodisperse solid spheres. The intrinsic dispersivity and sample dependent dispersion are separated by imposing a heuristic model where coarse grained axial advection velocities vary in a plane perpendicular to the flow but not along the direction of flow. The derived intrinsic dispersivity agrees with the predicted value (Scheven et al., Phys Rev Lett. 2007;99:054502-1,054502-4). It serves as a bench mark for different implementations of simulations coupling Stokes flow and diffusion in random geometries, and for experimental tests of injection and packing methods. Conceptually, a well defined and fittable effective dispersivity is introduced in an analytical framework describing dispersion data obtained in non-ideal packed beds, where elution profiles cannot be fitted to the solution of the one dimensional advection diffusion equation. © 2009 American Institute of Chemical Engineers AIChE J, 2010 [source]

Coumarins and phenolic fingerprints of oak and Brazilian woods extracted by sugarcane spirit

Alexandre Ataide da Silva
Abstract A total of 25 sugarcane spirit extracts of six different Brazilian woods and oak, commonly used by cooperage industries for aging cachaça, were analyzed for the presence of 14 phenolic compounds (ellagic acid, gallic acid, vanillin, syringaldehyde, synapaldehyde, coniferaldehyde, vanillic acid, syringic acid, quercetin, trans -resveratrol, catechin, epicatechin, eugenol, and myricetin) and two coumarins (scopoletin and coumarin) by HPLC-DAD-fluorescence and HPLC-ESI-MSn. Furthermore, an HPLC-DAD chromatographic fingerprint was build-up using chemometric analysis based on the chromatographic elution profiles of the extracts monitored at 280,nm. Major components identified and quantified in Brazilian wood extracts were coumarin, ellagic acid, and catechin, whereas oak extracts shown a major contribution of catechin, vanillic acid, and syringaldehyde. The main difference observed among oak and Brazilian woods remains in the concentration of coumarin, catechin, syringaldehyde, and coniferaldehyde. The chemometric analysis of the quantitative profile of the 14 phenolic compounds and two coumarins in the wood extracts provides a differentiation between the Brazilian wood and oak extracts. The chromatographic fingerprint treated by multivariate analysis revealed significant differences among Brazilian woods themselves and oak, clearly defining six groups of wood extracts: (i) oak extracts, (ii) jatobá extracts, (iii) cabreúva-parda extracts, (iv) amendoim extracts, (v) canela-sassafrás extracts and (vi) pequi extracts. [source]

Ultra scale-down approach to correct dispersive and retentive effects in small-scale columns when predicting larger scale elution profiles

N. Hutchinson
Abstract Ultra scale-down approaches represent valuable methods for chromatography development work in the biopharmaceutical sector, but for them to be of value, scale-down mimics must predict large-scale process performance accurately. For example, one application of a scale-down model involves using it to predict large-scale elution profiles correctly with respect to the size of a product peak and its position in a chromatogram relative to contaminants. Predicting large-scale profiles from data generated by small laboratory columns is complicated, however, by differences in dispersion and retention volumes between the two scales of operation. Correcting for these effects would improve the accuracy of the scale-down models when predicting outputs such as eluate volumes at larger scale and thus enable the efficient design and operation of subsequent steps. This paper describes a novel ultra scale-down approach which uses empirical correlations derived from conductivity changes during operation of laboratory and pilot columns to correct chromatographic profiles for the differences in dispersion and retention. The methodology was tested by using 1 mL column data to predict elution profiles of a chimeric monoclonal antibody obtained from Protein A chromatography columns at 3 mL laboratory- and 18.3 L pilot-scale. The predictions were then verified experimentally. Results showed that the empirical corrections enabled accurate estimations of the characteristics of larger-scale elution profiles. These data then provide the justification to adjust small-scale conditions to achieve an eluate volume and product concentration which is consistent with that obtained at large-scale and which can then be used for subsequent ultra scale-down operations. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source]

The stereodynamics of 1,2-dipropyldiaziridines

CHIRALITY, Issue 2 2010
Oliver Trapp
Abstract N-alkylated trans -diaziridines are an intriguing class of compounds with two stereogenic nitrogen atoms which easily interconvert. In the course of our investigations of the nature of the interconversion process via nitrogen inversion or electrocyclic ring opening ring closure, we synthesized and characterized the three constitutionally isomeric diaziridines 1,2-di- n -propyldiaziridine 1, 1-isopropyl-2- n -propyldiaziridine 2, and 1,2-diisopropyldiaziridine 3 to study the influence of the substituents on the interconversion barriers. Enantiomer separation was achieved by enantioselective gas chromatography on the chiral stationary phase Chirasil-,-Dex with high separation factors , (1-isopropyl-2- n -propyldiaziridine: 1.18; 1, 2-diisopropyldiaziridine: 1.24; 100°C 50 kPa He) for the isopropyl substituted diaziridines. These compounds showed pronounced plateau formation between 100 and 150°C, and peak coalescence at elevated temperatures. The enantiomerization barriers ,G, and activation parameters ,H, and ,S, were determined by enantioselective dynamic gas chromatography (DGC) and direct evaluation of the elution profiles using the unified equation implemented in the software DCXplorer. Interestingly, 1-isopropyl-2- n -propyldiaziridine and 1,2-diisopropyldiaziridine exhibit similar high interconversion barriers ,G, (100°C) of 128.3 ± 0.4 kJ mol,1 and 129.8 ± 0.4 kJ mol,1, respectively, which indicates that two sterically demanding substituents do not substantially increase the barrier as expected for a distinct nitrogen inversion process. Chirality, 2010. © 2009 Wiley-Liss, Inc. [source]