April 2014, Volume 77, Issue 7, pp 581–589A Stability-Indicating HPLC Method for the Determination of Nitrosylcobalamin (NO-Cbl), a Novel Vitamin B12 AnalogFirst Online: 14 March 2014Received: 21 November 2013Revised: 09 February 2014Accepted: 20 February 2014DOI: 10.1007/s10337-014-2645-5Cite this article as: Dunphy, M.J., Sysel, A.M., Lupica, J.A. et al. Chromatographia (2014) 77: 581. AbstractNitrosylcobalamin (NO-Cbl), a novel vitamin B12 analog and anti-tumor agent, functions as a biologic ‘Trojan horse’, utilizing the vitamin B12 transcobalamin II transport protein and cell surface receptor to specifically target cancer cells. A stability-indicating HPLC method was developed for the detection of NO-Cbl during forced degradation studies. This method utilized an Ascentis® RP-Amide (150 mm × 4.6 mm, 5 μm) column at 35 °C with a mobile phase (1.0 mL min−1) combining a gradient of methanol and an acetate buffer at pH 6.0. Detection wavelengths of 450 and 254 nm were used to detect corrin and non-corrin-based products, respectively.
NO-Cbl, synthesized from hydroxocobalamin and pure nitric oxide gas, was subjected to degradative stress conditions including oxidation, hydrolysis and thermal and radiant energy challenge. The method was validated by assessing linearity, accuracy, precision, detection and quantitation limits and robustness. The method was applied successfully for purity assessment of synthesized NO-Cbl and for the determination of NO-Cbl during kinetic studies in aqueous solution and in solid-state degradation assessments. This HPLC method is suitable for the separation of cobalamins in aqueous and methanolic solutions, for routine detection of NO-Cbl and for purity assessment of synthesized NO-Cbl. Additionally, this method has potential application in identification and monitoring of diseases involving altered nitric oxide homeostasis where vitamin B12 therapy is utilized to scavenge excess nitric oxide, subsequently resulting in the in vivo production of NO-Cbl.Keywords12Copyright informationWhat B12 Supplement Should I Take?
I get this question all the time, so I finally decided to write up a blog post to point people to. Cyanocobalamin is the most common form of B12 supplement and is the form found in fortified foods. It doesn’t occur much in nature, but it is the most stable form of vitamin B12, and the cheapest. It has been the most studied form and has consistently been shown to be effective. Unless there are extenuating circumstances, I recommend that people take cyanocobalamin as their B12 supplement. Cyanocobalamin contains a molecule of cyanide, but the amount of cyanide in cyanocobalamin supplements is so small as to be physiologically insignificant (more info). Some people have cyanide metabolism defects, and they should not take cyanocobalamin. If you are asking yourself right now if you might be one of those people, you can assume the chances are close to zero. People with cyanide poisoning should not take cyanocobalamin. If you have not previously suspected that you might have cyanide poisoning, then you can assume you don’t have it.
People with chronic kidney problems should probably take a non-cyanocobalamin form of B12 (more info). Finally, cigarette smokers might want to take a non-cyanocobalamin form of vitamin B12 as they can build up cyanide in their system. This is theoretical – I have never come across a vegan cigarette smoker who reported vitamin B12 deficiency based on taking cyanocobalamin and the Institute of Medicine has concluded that “The effect of smoking on the B12 requirement thus appears to be negligible (1).” Methylcobalamin is one of the two co-enzyme forms of vitamin B12, the other being adenosylcobalamin (known by many other names, including “dibencozide”). When I first got into the B12 issue, dibencozide was all the rage, now it’s methylcobalamin for some reason. The two forms have different functions in the body and both are necessary (more info). Methylcobalamin is promoted by some alternative health practitioners and the supplement industry as superior to cyanocobalamin, primarily because it is a co-enzyme form of vitamin B12.
Based on many studies and case reports in the scientific literature in which cyanocobalamin has successfully cured vitamin B12 deficiency, it appears that the body can convert cyanocobalamin into methylcobalamin without any problem. The body also has to convert methylcobalamin or cyanocobalamin into adenosylcobalamin for B12 to carry out all of its functions. (A fourth form of vitamin B12, hydroxocobalamin, is the form typically found in animal products and B12 injections; it must also be converted into the co-enzyme forms.) But is there any harm in taking methylcobalamin over cyanocobalamin? Probably not, but methylcobalamin is thought not to be as stable as cyanocobalamin and therefore higher doses are recommended, a minimum of 1,000 µg per day. Recommendations for cyanocobalamin are much lower (more info). Some people with chronic fatigue report getting more relief from adenosylcobalamin than either methylcobalamin or cyanocobalamin (more info), while other people report feeling better only when taking both co-enzyme forms (adenosyl- and methyl-).
Cyanocobalamin supplements are ubiquitous, cheap, and well-studied, while methyl and adenosyl are much more of an unknown entity. Unless you have a good reason to be using them, I recommend cyanocobalamin. Some companies claim to have a natural, living, plant, or raw source of vitamin B12. The B12 can come from seaweed or other unstated sources. Unless a label lists the source of vitamin B12 as cyanocobalamin, methylcobalamin, adenosylcobalamin, or hydroxocobalamin, I would not rely on it. Specific Brands of Vitamin B12 I do not have recommendations regarding any specific brands of vitamin B12 supplements. As far as cyanocobalamin goes, I assume all sublingual or chewable tablets to be effective. I do not have an opinion on B12 skin patches or sprays. As far as methylcobalamin supplements, as I stated above, I’m not as confident about them, and I have no opinion on any specific brand name. 1. Food and Nutrition Board, Institute of Medicine. Dietary Reference Intakes for Thiamin, Riboflavin, Niacin, Vitamin B6, Folate, Vitamin B12, Pantothenic Acid, Biotin, and Choline.