The researchers developed a method for the quantitation of four bioactive forms of vitamin B12 (adenosylcobalamin, cyanocobalamin, hydroxocobalamin, methylcobalamin, collectively known as cobalamins) from dietary ingredients and supplements.
The method, outlined in a paper recently published in Food Chemistry, is called High Performance Liquid Chromatography Diode-Array Detection (or HPLC-DAD), developed and validated to provide an easy-to-implement quality control solution for the dietary supplement industry.
According to the Standard Methods Performance Requirements set by the AOAC, an independent nonprofit association that develops standards on a voluntary consensus founded in 1884, all four cobalamins should be determined individually.
Previously used methods compliant to these standards, however, have some pitfalls.
The AOAC official method is efficient in determining total vitamin B12 using common ultraviolet instrumentation, but the method converts all forms of vitamin B12 into cyanocobalamin and therefore does not distinguish the source of them.
Moreover, most of recent published vitamin B12 analytical methods use high-expense instruments such as mass spectrometry, the researchers argued.
Eurofins' director of research and development, Dr Hong You, led the project and was principal investigator.
More vitamin B12 on the market
Growing demand for vitamin B12 products motivated the need to develop an accurate, efficient, and cost-effective method of quantifying it.
“Theoretically, vitamin B12 deficiency is rare in normal healthy population, but vegans or vegetarians, pregnant women, infants, and eating disorder patients can be vulnerable to vitamin B12 deficiency because of their special dietary customs requirements and physiological needs,” the authors wrote.
With the rise of the brain health category in dietary supplements and functional food and beverage, product formulators are increasingly adding vitamin B12 into products, which has been linked to cognitive benefits.
The researchers purchased supplements on iherb.com for testing.
The contents of dietary supplements were combined prior to sample extraction to ensure homogeneity. For tablets, 20 tablets were ground in a coffee grinder and mixed thoroughly. For capsules, the contents of 20 capsules were combined and ground by mortar and pestle thoroughly. For powders, 10 serving size were combined and ground by mortar and pestle thoroughly. Due to the light sensitivity of vitamin B12 cobalamins, all the sample preparation were conducted in the dark room and/or under UV blocking fluorescent light
To demonstrate the method’s selectivity, an excipient blend (considered as the “Placebo” of tablet, capsule, and powder dietary supplements) expected to be free of active vitamin B12 was tested to evaluate potential chromatographic interferences from the typical filler ingredients used in tablet, capsule, and powder dietary supplements.
“This method is relatively simple (no extensive sample prep; straightforward HPLC method with a single column), quite fast, can distinguish four forms of bioactive vitamin B12, and do so with DAD detection,” the researchers reported.
“Being simple and fast also translates into lower cost.”
Source: Food Chemistry
Published online ahead of print, https://doi.org/10.1016/j.foodchem.2019.125010
“Determination of active vitamin B12 (cobalamin) in dietary supplements and ingredients by reversed-phase liquid chromatography: Single-laboratory validation”
Authors: Xiao Qiu, et al.