Release of iron from haemoglobin--a possible source of free radicals in diabetes mellitus.

Increased blood glucose in diabetes mellitus stimulates nonenzymatic glycosylation of several proteins, including haemoglobin. Although iron is tightly bound to haemoglobin, it is liberated under specific circumstances yielding free reactive iron. Studies with purified haemoglobin from normal individuals and diabetic patients revealed that concentration of free iron was significantly higher in the latter cases and increased progressively with extent of the disease. In vitro glycosylation of haemoglobin also led to increase in release of iron from protein. This increase in free iron, acting as a Fenton reagent, might produce free radicals, which, in turn might be causing oxidative stress in diabetes.

Comparative studies on the interaction of protoporphyrin with hemoglobin and myoglobin.

The binding parameters of protoporphyrin IX (PPIX) with hemoglobin (Hb) were studied spectrofluorimetrically and the results were compared with those of PPIX interacting with myoglobin (Mb). Two concentration ranges of PPIX (0.3 microM-1.5 microM and 1.5 microM-3.0 microM) were used. For both hemoglobin and myoglobin, the binding affinity constant (K) decreased while the number of binding sites (p) increased as the concentration range of PPIX increased. The interactions occurred in non-cooperative mode. Over a particular PPIX range, the interaction of PPIX with hemoglobin decreased significantly with increasing NaCl molarity indicating a trend in electrostatic interaction, whereas PPIX binding with myoglobin did not change significantly indicating mostly non-electrostatic mode of interaction. Total bound charge (z psi) decreased significantly with increased PPIX concentration range in case of hemoglobin-PPIX interaction, but remained almost same in case of myoglobin-PPIX interactions. Thermodynamic analysis revealed that binding of PPIX to hemoglobin was mostly electrostatic at lower concentration range of PPIX but became less electrostatic at higher concentration range and myoglobin-PPIX interaction, predominantly hydrophobic in nature, became more hydrophobic with increased range of PPIX concentration. The difference in binding modality between PPIX-Hb and PPIX-Mb has been discussed in relation to the state of aggregation of porphyrin as well as the subunit interaction property present and absent in hemoglobin and myoglobin, respectively.

A comparative study of 5'nucleotidase and alkaline phosphatase in human placenta during development.

Activities and a few properties of alkaline phosphatase and 5′-nucleotidase were compared in the developing human placenta. Both the enzymes were mostly membrane-bound and displayed similar developmental patterns with the highest activities at 24/26 weeks of the placenta. L-Phenylalanine, L-tryptophan and L-leucine were inhibitors of alkaline phosphatase, whereas they had no effect on the 5′-nucleotidase. Alkaline phosphatase from a late stage of gestation appeared to be almost heat-stable. An appreciable part of 5′-nucleotidase was also resistant to heat inactivation and this fraction varied with gestational age of the tissue. For both the enzymes, Vmax changed without altering Km values with periods of gestation. Ca2+ , Mg2+ and Mn2+ ions stimulated the alkaline phosphatase activity and Hg2+ , Zn2+ , Cu2+ ,Ni2+ were inhibitory. 5′-Nucleotidase was not activated by any of these cations. EDTA and Concanavalin A inhibited both the enzymes, although the extent of inhibition was different and also varied with gestation.