Comparison of chromatographic ion-exchange resins. I. Strong anion-exchange resins.

A comparative study has been undertaken on various strong anion-exchangers to investigate the pH dependence, titration curves, efficiency, binding strength, and dynamic capacity of the chromatographic resins. The resins tested included: Macro-Prep 25Q, TSK-Gel Q-5PW-HR, Poros QE/M, Q Sepharose FF, Q HyperD 20, Q Zirconia, Source 30Q, Fractogel EMD TMAE 650s, and Express-Ion Q. Testing was performed with five different proteins: Anti-FVII Mab (IgG), aprotinin, BSA, lipolase, and myoglobin. The dependence of pH on retention varies from generally low to very high for proteins with low pI. No direct link between pH dependence on retention and titration curves of the different resins was observed. Efficiency results show the expected trend of lower dependence of the plate height with increasing flow-rate of resins for medium and high pressure operation compared to the soft resins. Binding to the anion-exchange resins as a function of ionic strength may vary depending on the specific protein. Generally, binding and elution at a high salt concentration may be performed with Poros QE/M or Macro-Prep 25Q, while binding and elution at low salt concentration may be done with TSK-Gel Q-5PW. Dynamic capacities are strongly dependent on the specific protein employed and for some resins dependent on the flow-rate. A general good agreement was obtained between this study and data obtained by suppliers for the dynamic capacity. The results of this study may be used for selection of resins for testing in process development, however, the data does not tell anything about specific selectivity differences or resolution between a target protein and a given impurity. None of the resins studied here should be regarded as good or bad, but more or less suitable for a specific purpose, and only testing for the specific application will determine which one is the optimal resin.

Comparison of loading capacities of various proteins and peptides in culture medium and in pure state.

Chromatographic media suppliers most frequently state the capacities of their gels based on either static capacities or frontal analysis experiments of pure proteins, however, these capacity values are often far from the capacities experienced in the production of such proteins. In this work static and dynamic capacities of various pure industrial proteins or peptides are compared to the capacities of the proteins or peptides under similar conditions in their natural culture medium. The results show a significant decrease in the static and dynamic capacities of the proteins or peptides when present in culture medium due to competitive binding of medium proteins. The proteins and peptides included in this study are: lipolase, glucagon-like peptide-1, truncated prothrombin, insulin precursor, and anti-Factor VII monoclonal antibody.

[Dyspepsia in general practice. A study among users of anti-ulcer agents]. / Dyspepsi i almen praksis.

Five doctors in general practice sent identical questionnaires to all those of their listed patients who in the previous year had received one or more prescriptions for an ulcer medication in the groups H2 receptor antagonists, acid pump inhibitors, bismuth and sucralfate preparations. The patients were identified by the pharmacy. Two hundred and fifty-nine questionnaires were returned, and the response rate was 83%. Four point nine percent of an adult population of 7160 received in the year 1993 at least one prescription for ulcer medicine, and 0.87% were in constant treatment. On average the doctors met 49 dyspeptic patients per 1000 adults per year. This is far more than found by gastroenterologists in studies of different ways of handling the dyspeptic patient in general practice.

The Use of RP-HPLC for measuring activation and cleavage of rFVIIa during purification.

A reverse phase HPLC (RP-HPLC) method for analysis of recombinant factor VIIa (rFVIIa) has been developed. The method discriminates between different forms of recombinant FVII (rFVII). To obtain separation of these closely related molecules the method has been optimized with respect to gradient profile and temperature. The method has been used for optimization of purification processes and for kinetic studies. EVidence for autolytic cleavage was obtained. (c) 1995 John Wiley & Sons, Inc.

FVIIa derivatives obtained by autolytic and controlled cathepsin G mediated cleavage.

The heavy chain of coagulation factor VII contains a serine esterase entity. A partial cleavage in the heavy chain occurs during purification and activation of the single-chain zymogen, presumably as a result of autolysis. Neutrophil cathepsin G initially generates a Gla-domainless FVIIa without coagulant activity. However, on extended exposure cleavage also occurs in the heavy chain, resulting in a complete loss of enzyme activity. Four cleavage sites on the heavy chain, two susceptible to trypsin-like autolysis and two susceptible to chymotrypsin-like cathepsin G-mediated catalysis have been identified. The hydrolysis of peptide bonds in the heavy chain might contribute to regulation of the coagulation process in vivo.