1. Haems are unstable under aerobic conditions in the presence of thiols, which are used to activate the ferrochelatase enzyme; catalase inhibits this degradation of haem. In addition, thiols ...interfere with the determination of protohaem as its pyridine haemochromogen derivative. 2. Three ferrochelatase assays are described that minimize interference by these two reactions. Two of these assays involve measurement of porphyrin utilization, one spectrophotometrically and the second spectrofluorimetrically. The third assay measures haem formation by a pyridine haemochromogen technique. Results obtained with these three methods were in close agreement at a GSH concentration of 4mm. 3. The stimulatory effect of GSH on ferrochelatase has been confirmed. The spectrum of the haem formed is dependent on GSH concentration; at high GSH concentrations (20mm) the haem is in the reduced state, but at low concentration (4mm) the spectrum of the product resembles that of an oxidized haemoprotein such as ferrihaemoglobin. 4. The inhibitory effect of oxygen on ferrochelatase activity has been confirmed by spectrophotometric assay of porphyrin disappearance.
Ferritin, an iron-containing protein widely diffused in nature, has the important biochemical function of being the principal reserve and regulator for Fe3+ levels in blood and tissue. Due to its ...natural, effectively non-toxic iron content, ferritin has been the object of strong interest in the development of pharmaceutical products for use in iron deficiency treatment. Therefore, the need has arisen for the analytical characterization of this industrial product, be it in the hydroglyceric solution or dry powder form. The main considerations for the characterization of industrial ferritin are the following: a) iron content in both the unmodified product and the precipitated protein; b) protein content and protein/iron ratio; c) protein identification by means of polyacrylamide gel electrophoresis (PAGE); d) confirmation of protein identity and evaluation of molecular weight by means of size exclusion chromatography; e) identification and evaluation of other components of bulk ferritin preparations such as preservatives, excipients for lyophilization, water and concomitant proteins. Eight different samples were examined using the above considerations. Among the qualitative and quantitative results reported, of particular interest are those obtained by means of size-exclusion high performance liquid chromatography (SEC-HPLC). With a UV "diode array" detector it is possible to discern the peaks for ferritin and its molecular aggregates from those of concomitant proteins and preservatives; furthermore, it is possible to evaluate their molecular dimensions. Using this method, the ferritin monomer and other protein fractions can be quantitatively analyzed either by calculating area percent distribution of the chromatographic peaks or by comparing the sample with a highly pure external standard of ferritin.
We consider mean first-passage times (MFPTs) for systems driven by non-Markov gamma and McFadden dichotomous noises. A simplified derivation is given of the underlying integral equations and the ...theory for ordinary renewal processes is extended to modified and equilibrium renewal processes. The exact results are compared with the MFPT for Markov dichotomous noise and with the results of Monte Carlo simulations.
PDF
