Generic method for systematic phase selection and method development of biochromatographic processes. Part I. Selection of a suitable cation-exchanger for the purification of a pharmaceutical protein

Even if the first protein therapeutics are now for more than 20 years on the market the selection of suitable adsorbents for the preparative downstream processing (DSP) of these biomolecules as well as the method development towards process conditions are still based mainly on 'trial and error'. Therefore, theses processes are not perfectly efficient, but indeed very time consuming and laborious. In this study a novel systematic method is introduced to find a suitable adsorbent (not necessarily the best one) with appropriate separation parameters for a specific separation with reduced effort. Following this strategy, the adsorbents must first be packed into columns under preparative conditions and then characterized completely with regard to, e.g. pressure drop, k'-values, plate heights (HETP curves), selectivity and capacity by using test substances, which are similar in their characteristics (molecular mass, size, charge distribution, hydrophobicity) to the target proteins. With the database once determined, a preselection of most suitable adsorbents including separation parameters is made regarding chromatographic and also economical properties. After this, preparative experiments must be conducted with a reduced number of adsorbents to figure out the individual influence of side components. This approach is demonstrated for the separation of an exemplary industrial protein mixture using cation-exchange chromatography (CEX). Characterization of different weak CEX-adsorbents is illustrated. After comparing these phases with each other, a first preselection and a prediction of suitable adsorbents is made. In the following preparative separation conditions (load, velocity, gradient) are determined for the preparative separations using the database and results of some additional experiments. The final comparison of separation performance in preparative scale confirms this selection and so the applicability of the new method.     

New antimalarial drugs

Approximately 40% of the world population live in areas with the risk of malaria. Each year, 300-500 million people suffer from acute malaria, and 0.5-2.5 million die from the disease. Although malaria has been widely eradicated in many parts of the world, the global number of cases continues to rise. The most important reason for this alarming situation is the rapid spread of malaria parasites that are resistant to antimalarial drugs, especially chloroquine, which is by far the most frequently used. The development of new antimalarial drugs has been neglected since the 1970s owing to the end colonialism, changes in the areas of military engagement, and the restricted market potential. Only in recent years, in part supported by public funding programs, has interest in the development of antimalarial drugs been renewed. New data available from the recently sequenced genome of the malaria parasite Plasmodium falciparum and the application of methods of modern drug design promise to bring significant development in the fight against this disease.     

Determination of heavy metal complexes with humic substances by HPLC/ICP-MS coupling using on-line isotope dilution technique

An isotope dilution mass spectrometric (IDMS) method has been developed for the simultaneous determination of the complexes of 11 heavy metals (Ag, Cd, Cu, Mo, Ni, Pb, Tl, U, W, Zn and Zr) with humic substances (HS) by coupling HPLC with ICP-MS and applying the on-line isotope dilution technique. The HPLC separation was carried out with size exclusion chromatography. This HPLC/ICP-IDMS method was applied to samples from a brown water, ground water, sewage and seepage water as well as for a sample containing isolated fulvic acids. The total contents of heavy metals and of their complexes were analyzed in these samples with detection limits in the range of 5–110 ng/L. The analysis of heavy metal/HS complexes from the different waters resulted in characteristic fingerprints of the distribution pattern of heavy metals in the separated HS fractions. A comparison between the total heavy metal concentrations and their portions bound to humic substances showed distinct differences for the various metals. Simultaneous ¹²C detection was used for the characterization of HS complexes not identified by UV detection and for the determination of relative DOC concentrations of chromatographic peaks. Received: 21 February 1997 / Revised: 27 May 1997 / Accepted: 28 May 1997     

NMR spectroscopic characterization of millisecond protein folding by transverse relaxation dispersion measurements

The cold shock protein CspB adopts its native and functional tertiary structure on the millisecond time scale. We employed transverse relaxation NMR methods, which allow a quantitative measurement of the cooperativity of this fast folding reaction on a residue basis. Thereby, chemical exchange contributions to the transverse relaxation rate (R(2)) were observed for every residue of CspB verifying the potential of this method to identify not only local dynamics but also to characterize global events. Toward this end, the homogeneity of the transition state of folding was probed by comparing Chevron plots (i.e., dependence of the apparent folding rate on the denaturant concentration) determined by stopped-flow fluorescence with Chevron plots of six residues acquired by R(2) dispersion experiments. The coinciding results obtained for probes at different locations in the three-dimensional structure of CspB indicate the ability and significance of transverse relaxation NMR to determine Chevron plots on a residue-by-residue basis providing detailed insights on the nature of the transition state of folding.     

Chemie polyfunktioneller liganden LXIII. Adamantankäfigverbindungen mit den heteroelementen arsen,stickstoff und silizium

The reaction of 1,1,1-tris(diiodarsinomethyl)ethane, CH₃C(CH₂AsI₂)₃ (I), with i-C₃H₇NH₂, n-C₄H₉NH₂, C₆H₅NH₂, p-CH₃C₆H₄NH₂ and [(CH₃)₃Si]₂NH in the presence of (C₂H₅)₃N as auxiliary base in THF gives the adamantane cage compounds CH₃C(CH₂AsNC₃H₇)₃ (III), CH₃C(CH₂AsNC₄H₉)₃ (IV), CH₃C(CH₂AsNC₆H₅)₃ (V), CH₃C(CH₂AsNC₆H₄CH₃)₃ (VI) and CH₃C[CH₂AsNSi(CH₃)₃]₃ (VII). VII is also obtained in the reaction of I with NaN[Si(CH₃)₃]₂. The by-product (CH₃)₃SiO(CH₂)₄I (VIII) could be isolated in both syntheses of VII. All compounds have been characterized by mass spectrometry and infrared, Raman and ¹H NMR spectroscopy.     

Neue Oxozincate der Alkalimetalle: Rb2[ZnO2] und Cs2[ZnO2]

New Oxozineates of Alkali Metals: Rb₂[ZnO₂] and Cs₂[ZnO₂] Colorless single crystals of the hitherto unknown Rb₂[ZnO₂] (a = 9.55₈, b = 6.33₅, c = 15.91 Å, β = 118.6°, z = 8, d_pyk = 4.11, d_rö = 4.21 g · cm^?3) and Cs₂[ZnO₂] (a = 9.85₁, b = 6.61₉, c = 16.26 Å, β = 116.8°) have been prepared, which crystallize monoclinic, P2₁/c – D. (For parameters see text.) Unexpected there are “isolated” groups [Zn₄O₈]. Half of the Zn atoms exhibit the unusual coordination number 3 towards O^2?. The Madelung Part of Lattice Energy, MAPLE, and the Effective Coordination Number, ECoN, the latter by means of Mean Fictive Ionic Radii, MEFIR, are calculated and discussed.     

Boride-based nano-laminates with MAX-phase-like behaviour

MAX-phases being usually composed of transition metals, group A elements and carbon/nitrogen are considered interesting materials for many applications because of their tremendous bulk modulus, “reversible” plasticity, and machinability. This is mainly due to their unique kind of bonding comprising covalent, ionic as well as metallic bonds providing “easy” planes of rupture and deformability due to the layered crystal structures.In transition metal boride systems, similar types of bonding are available. In particular the W₂B₅-structure type and its stacking variations allow the synthesis of strongly layered crystal structures exhibiting unique delamination phenomena.The paper presents ab initio calculations showing the similarities of bonding between the ternary carbides and the corresponding ternary or quaternary borides. Formation of boride-based nano-laminates from auxiliary liquid phases, from the melt as well as during sintering and precipitation from supersaturated solid solutions will be discussed by means of SEM and TEM studies. The role of impurities weakening the interlayer bonding will be addressed in particular. The pronounced cleavage parallel to the basal plane gives rise for crack deflection and pull-out mechanisms if the laminates are dispersed in brittle matrices such as boron carbide, silicon carbide or other transition metal borides.     

Excess Molar Volumes and Liquid–Liquid Equilibria of the Ionic Liquid 1-Methyl-3Octyl-Imidazolium Tetrafluoroborate Mixed with Butan-1-ol and Pentan-1-ol

Experimental densities were measured for the system 1-methyl-3-octyl-imidazolium tetrafluoroborate [OMIM][BF₄] + butan-1-ol, + pentan-1-ol at 298.15 K and ambient pressure using a vibrating tube densimeter, taking into account the influence of the viscosity correction. Excess molar volumes V^E have been determined. V^E is quite small and negative in the alcohol-rich range of the mixture composition and positive in the alcohol-poor range. LLE data of [OMIM][BF₄] + pentan-1-ol have been measured using a laser light scattering cell for detecting cloud points at different compositions in the temperature range of 282–292 K. A miscibility gap with an upper critical solution temperature (UCST) of 292 K has been found.     

Green chemistry and sustainability metrics in the pharmaceutical manufacturing sector

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