Towards hemerythrin-based blood substitutes: comparative performance to hemoglobin on human leukocytes and umbilical vein endothelial cells.

Hemerythrin is a dioxygen-carrying protein whose oxidative/nitrosative stress-related reactivity is lower than that of hemoglobin, which may warrant investigation of hemerythrin as raw material for artificial oxygen carriers ('blood substitutes'). We report here the first biological tests for hemerythrin and its chemical derivatives, comparing their performance with that of a representative competitor, glutaraldehyde-polymerized bovine hemoglobin. Hemerythrin (native or derivatized) exhibits a proliferative effect on human umbilical vein endothelial cell (HUVEC) cultures, as opposed to a slight inhibitory effect of hemoglobin.

Hemoglobin-albumin crosslinked copolymers: reduced prooxidant reactivity.

We have previously reported that derivatization of hemoglobin with periodate-modified sugar derivatives such as oxidized adenosine triphosphate (oATP) leads to an increase in prooxidant reactivity at the heme. Here, we report that copolymerization of hemoglobin with serum albumin alleviates this problem completely, to the extent where the copolymer even has a slightly lower autooxidation rate compared to native hemoglobin. A similar, although not as potent, effect is obtained when hemoglobin is derivatized with oATP in the presence of small-molecule antioxidants instead of albumin.

A new polyethyleneglycol-derivatized hemoglobin derivative with decreased oxygen affinity and limited toxicity.

A new protocol is described for derivatization of hemoglobin with polyethyleneglycol (PEG) via reaction of the unmodified native hemoglobin with an activated amine-reacting polyethylene glycol derivative which, unlike protocols previously described, leads to formation of a peptide bond between hemoglobin and PEG. Dioxygen binding and peroxide reactivities of the derivatized hemoglobin are examined, and found to be within reasonable limits, with the particular observation that, unlike with a few other derivatization protocols, the dioxygen affinity is slightly lower than that of native Hb. In cell culture tests (human umbilical vein epithelial cells, HUVEC), the derivatization protocol induces no toxic effect.

Ascorbate peroxidase activity of cytochrome c.

The peroxidase-type reactivity of cytochrome c is proposed to play a role in free radical production and/or apoptosis. This study describes cytochrome c catalysis of peroxide consumption by ascorbate. Under conditions where the sixth coordination position at the cytochrome c heme iron becomes more accessible for exogenous ligands (by carboxymethylation, cardiolipin addition or by partial denaturation with guanidinium hydrochloride) this peroxidase activity is enhanced.

Derivatization of haemoglobin with periodate-generated reticulation agents: evaluation of oxidative reactivity for potential blood substitutes.

Periodate modification of the sugar moiety in sugars, including adenosine triphosphate (ATP), has previously been employed in order to prepare dialdehyde-type reagents, which were then utilized in crosslinking reactions on haemoglobin, yielding polymerized material with useful dioxygen-binding properties and hence proposed as possible artificial oxygen carriers ('blood substitutes'). Here, the periodate protocol is shown to be applicable to a wider range of oxygen-containing compounds, illustrated by starch and polyethylene glycol. Derivatization protocols are described for haemoglobin with such periodate-treated crosslinking agents, and the dioxygen-binding properties and redox reactivities are investigated for the derivatized haemoglobins, with emphasis on pro-oxidative properties.