The reaction of HOCl and cyanocobalamin: corrin destruction and the liberation of cyanogen chloride.

Overproduction of hypochlorous acid (HOCl) has been associated with the development of a variety of disorders such as inflammation, heart disease, pulmonary fibrosis, and cancer through its ability to modify various biomolecules. HOCl is a potent oxidant generated by the myeloperoxidase-hydrogen peroxide-chloride system. Recently, we have provided evidence to support the important link between higher levels of HOCl and heme destruction and free iron release from hemoglobin and RBCs. Our current findings extend this work and show the ability of HOCl to mediate the destruction of metal-ion derivatives of tetrapyrrole macrocyclic rings, such as cyanocobalamin (Cobl), a common pharmacological form of vitamin B12. Cyanocobalamin is a water-soluble vitamin that plays an essential role as an enzyme cofactor and antioxidant, modulating nucleic acid metabolism and gene regulation. It is widely used as a therapeutic agent and supplement, because of its efficacy and stability. In this report, we demonstrate that although Cobl can be an excellent antioxidant, exposure to high levels of HOCl can overcome the beneficial effects of Cobl and generate proinflammatory reaction products. Our rapid kinetic, HPLC, and mass spectrometric analyses showed that HOCl can mediate corrin ring destruction and liberate cyanogen chloride (CNCl) through a mechanism that initially involves α-axial ligand replacement in Cobl to form a chlorinated derivative, hydrolysis, and cleavage of the phosphonucleotide moiety. Additionally, it can liberate free Co, which can perpetuate metal-ion-induced oxidant stress. Taken together, these results are the first report of the generation of toxic molecular products through the interaction of Cobl with HOCl.

Purification and characterization of corrinoid-compounds from the dried powder of an edible cyanobacterium, Nostoc commune (Ishikurage).

Vitamin B12 content (98.8 +/- 5.6 microg/100 g dry weight) of an edible cyanobacterium, Nostoc commune (Ishikurage) was determined by the Lactobacillus delbrueckii ATCC 7830 microbiological method. Bioautography with vitamin B12-dependent Escherichia coli 215 indicated that N. commune contained two (main and minor) corrinoid-compounds. These corrinoid-compounds were purified to homogeneity from the dried algal cells and characterized. The main and minor purified corrinoid-compounds were identified as pseudovitamin B12 and vitamin B12, respectively, on the basis of silica gel 60 TLC, C18 reversed-phase HPLC, 1H NMR spectroscopy, and UV-visible spectroscopy. These results suggest that the bacterial cells are not suitable for use as a vitamin B12 source, especially in vegetarians.

Purification and characterization of a corrinoid-compound in an edible cyanobacterium Aphanizomenon flos-aquae as a nutritional supplementary food.

The vitamin B12 concentration of the dried cells of Aphanizomenon flos-aquae was determined by both microbiological method with Lactobacillus delbrueckeii ATCC7830 and chemiluminescence method with intrinsic factor. The Aphanizomenon cells contained 616.3 +/- 30.3 micro g (n = 4) of vitamin B12 per 100 g of the dried cells by the microbiological method. The values determined with the chemiluminescence method, however, were only about 5.3% of the values determined by the microbiological method. A corrinoid-compound was purified from the dried cells and characterized. The purified corrinoid-compound was identified as pseudovitamin B12 (an inactive corrinoid-compound for humans) by silica gel 60 TLC, C18 reversed-phase HPLC, ultraviolet-visible spectroscopy, and 1H NMR spectroscopy. The results suggest that the Aphanizomenon cells are not suitable for use as a vitamin B12 source, especially in vegans.

Development of a method for the separation of corrinoids in ovine tissues by HPLC.

A method has been developed using a combination of high-performance liquid chromatography (HPLC) and a radioisotope dilution assay (RIDA) to routinely estimate the distribution of corrinoids (the cobalamins hydroxocobalamin, methylcobalamin and 5'-deoxyadenosylcobalamin, and cobalamin analogues) in liver, plasma, milk, intestinal fluid and faeces. Corrinoids were extracted with a sodium acetate buffer, separated by HPLC and quantified by RIDA. Recoveries of corrinoids were 29% for hydroxocobalamin, 50% for 5'-deoxyadenosylcobalamin and 64% for methylcobalamin. The method allows the routine analysis of many samples and maintains good standards of precision.