Capillary electrokinetic chromatography of insulin and related synthetic analogues.

With the implementation of recombinant DNA technology in the pharmaceutical industry, some synthetic insulins have been developed in order to improve the therapy of diabetes. These analogues differ only slightly in the amino acid sequence, therefore displaying a great challenge for analytical chemistry. Within the work presented in this paper, capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC) with sodium dodecylsulphate (SDS) as micelle-forming agent, and microemulsion electrokinetic chromatography (MEEKC) with microemulsions consisting of SDS, n-octane and 1-butanol were investigated for the separation of human insulin and five synthetic analogues. Best results were achieved with a solvent-modified MEKC system consisting of 100mM sodium dodecyl sulphate and 15% acetonitrile in 10mM borate buffer (pH 9.2). A similar system based on perfluorooctanoic acid as micelle-forming agent in ammonium acetate (pH 9.2) was successfully employed for the hyphenation with a quadrupole/time-of-flight mass spectrometer via a sheath-flow interface. In this case, detection limits at 10mg/L could be achieved.

Studies on the metabolic fate of 111In-labeled antibodies.

Indium-111-labeled antibodies, though providing superior photon flux to iodine-labeled antibodies, can exhibit high levels of accumulation in some non-target organs. In an effort to understand the nature of this non-target uptake we have evaluated the molecular weight of 111In species retained in several tissues by radio-FPLC (sizing chromatography) following injection of [111In]DTPA 5G6.4, a murine monoclonal antibody, into normal mice. Blood, liver and kidneys were removed, and liver and kidneys were homogenized at several time points after antibody injection. The proportion of 111In-containing species was found to vary with the tissue and with time. Analysis of blood showed only radiolabeled antibody. In the liver, several 111In species were identified with molecular weights compatible with intact antibody, [111In]transferrin, and low molecular weight complexes, with an increase in the proportion of [111In]transferrin and low molecular weight species occurring over time. While the same molecular weight species were also identified in the kidneys, the kidneys contained the largest percentage of low molecular weight species which increased over time. When 125I-labeled 5G6.4 was injected and the tissues similarly analyzed, only radioactive material with the molecular weight of intact antibody was detected. Comparison of two methods of purification of [111In]labeled antibody after labeling revealed a significant difference in the organ uptake of radiolabeled products for 111In. Although dialysis was sufficient for the removal of labile 111In, as determined by TLC, subsequent sizing chromatography on Bio-Gel P-60 dramatically dropped the hepatic and renal uptake of 111In relative to blood and diminished the proportion of the low molecular weight species present on sizing FPLC of extracts from tissues.(ABSTRACT TRUNCATED AT 250 WORDS)