Background

This story began at the National Research Council of Canada (NRC) in the 1990s with a scientific study undertaken by Avivagen’s founding scientists, stimulated by the intense interest in the possibility that β-carotene has non-vitamin A, preventive anticancer activity. Work continued at Avivagen in the ensuing decades, exploring oxidation as the mode of action rather than an antioxidant action of β-carotene itself. Ultimately, a previously unrecognized, readily-formed, biologically active substance was discovered that is a very plausible source of β-carotene’s non-vitamin A activity. The substance, formed as the main product whenever and wherever β-carotene undergoes non-enzymatic, spontaneous oxidation, is a copolymer of β-carotene with oxygen, containing ~30% oxygen by weight. Recognition of the widespread existence of carotenoid copolymer compounds in Nature opens the door to fully realizing the health potential of carotenoids and, in particular, of β-carotene. The existence of these compounds now also provides an attractive explanation, at least in part, for the human health benefits associated with dietary fruit and vegetable consumption.

β-Carotene and the more than 600 other members of the carotenoid family are characterized by an extended system of conjugated carbon double bonds that not only is the source of their intense coloration but also makes the compounds highly susceptible to reaction with the oxygen in air.

There’s More to β-Carotene than Vitamin A

Although β-carotene is best known as an excellent source of vitamin A there is another facet of β-carotene chemistry that Avivagen discovered has important health implications. Oxygen plays a critical role in unlocking the full potential of β-carotene. Whereas vitamin A is produced when oxygen, guided by a highly specific enzyme resident in the gut, splits the β-carotene molecule in half, in the absence of the enzyme, oxygen in air spontaneously and indiscriminately attacks the β-carotene molecule. This occurs regardless of environment, whether in a plant or in a laboratory.

Spontaneous uptake of oxygen by solid β-carotene standing in air. β-Carotene adds oxygen with an up to 30% increase in mass.

In the spontaneous, non-enzymatic reaction oxygen strongly prefers to add to β-carotene to form a polymer, rather than cut the β-carotene molecule into smaller pieces. The main product turns out to be a β-carotene-oxygen copolymer containing up to 8 molecules of O2 per β-carotene on average. This compound had not been reported before [1].

Non-Vitamin A Effects?

β-Carotene is reported to have beneficial non-vitamin A health effects, which appear to be associated with immune function benefits. A summary of literature results for supplementation with β-carotene has been provided in a European Food Safety Authority (EFSA) panel report on the safety and efficacy of β-carotene as a feed additive [2]:

“The positive effects in cows or pigs reported in the literature include decreased service per conception, increased number of viable embryos/reduced embryonic mortality, improved embryo quality, improved immunity with reduced incidence of retained placenta and metritis, increased pregnancy rate, increased percentage of milk fat (with unaffected milk yield), improved protection of the mammary gland against infection as a result of increased intracellular killing of microbes by phagocytes, and higher plasma progesterone and oestradiol levels in the cat…”. “Chew and co-workers reported that beta-carotene improves the immune status and decreases the incidence of reproductive disorders in peripartum cows by increasing lymphocyte and phagocyte function…”.

However, the EFSA panel further states:

“The data on the effects of supplemented beta-carotene on immunity and reproduction remain inconsistent and no conclusions can be drawn”. This comment underscores the lack of reproducibility of data when β-carotene itself is used as the supplement. Also, the possible inadvertent involvement of vitamin A is cited as a confounding factor.

Non-Vitamin A Effects are Real: The β-Carotene Copolymer is the Active Agent

By allowing pure β-carotene dissolved in a solvent to react fully with oxygen, Avivagen discovered a substance called OxBC (Oxidized β-Carotene), which is composed entirely of oxidation products that are principally β-carotene-oxygen copolymers. Importantly, OxBC contains no β-carotene or vitamin A and has no vitamin A activity. With the OxBC tool it became possible to show that the immunological activity of OxBC is a direct result of the activity of the β-carotene-oxygen copolymers [3].

The discovery of immunological activity in OxBC and the clear and consistent health benefits subsequently observed when OxBC is provided orally to livestock and companion animals confirm that the non-vitamin A effects of β-carotene are real and, furthermore, explain why β-carotene itself is unable to fulfil this function unless it first has undergone some amount of spontaneous oxidation to form OxBC. Benefits in poultry and swine are obtained at just low parts-per-million levels of OxBC in finished feed, well within the limits of levels of β-carotene were it available from plant sources, and quite apart from and complementary to the effects of supplemental vitamin A.

Real Products. OxC-beta™ Commercial Formulations for Non-Vitamin A Function

For commercial purposes the β-carotene copolymer compound is most conveniently and economically prepared in the form of OxBC produced synthetically by a global β-carotene manufacturer. The presence of very minor amounts of numerous co-generated, low molecular weight, norisoprenoid compounds is reflective of the process occurring naturally in plant materials, and that their presence does not interfere with the activity of the predominant β-carotene copolymer product. OxBC is formulated into premixes, known commercially as OxC-beta™ Livestock for livestock feeds and incorporated into Vivamune™ and Oximunol™ supplements for companion animals.

Natural Occurrence of OxBC in Plant Materials

The β-carotene copolymer, as well as other carotenoid copolymer compounds, are ubiquitous, forming naturally in variable and oftentimes significant quantities in carotenoid-containing plant materials, especially during processing, such as dehydration, and during storage. This is simply the result of exposure to air. Examples relevant to livestock feeds include alfalfa, grass and seaweed products, and in human nutrition, carrot and tomato powders [4].

Illustration of the effect of dehydration and storage on β-carotene oxidation in carrots. The formation of geronic acid, a marker of β-carotene oxidation and the formation of copolymer product, in dehydrated, powdered carrot can be seen to be closely linked to the oxidative loss of β-carotene. Note: carrots contain crystalline β-carotene.

Corroboration of the natural occurrence of the β-carotene copolymer compound has been provided independently by Schaub, Beyer and co-workers. In accounting for the loss of β-carotene in an extensive variety of provitamin A plant crop foods during storage, this group has reported that spontaneous, non-enzymatic oxidation of β-carotene leads to the concurrent formation of β-carotene copolymer compounds as the major product [5].

Something’s Missing in Livestock Feeds

In animal nutrition, β-carotene is primarily regarded as a source of vitamin A. Originally vitamin A was provided by adding certain β-carotene-rich plant materials or forages to livestock feeds. Sometimes β-carotene is referred to as provitamin A, reflecting the importance of this precursor function. However, significant losses of β-carotene occur in plant materials during processing and storage, with associated losses in vitamin A activity. Consequently, plant material sources of β-carotene included for vitamin A function have been replaced in modern livestock feeds with supplements that include synthetic vitamin A instead, along with other vitamins, micronutrients and minerals. Consequently, most modern livestock feeds lack or are deficient in forages containing β-carotene and β-carotene-oxygen copolymer compounds, and possibly other phytonutrients. Although oxidative loss of β-carotene means there is some inevitable loss of vitamin A activity from a plant source, the associated appearance of β-carotene copolymer compound potentially provides complementary non-vitamin A activity.

Meeting an Unfilled Food Animal Need for Non-Vitamin A Function with OxC-beta™ Livestock

Feeding trials in multiple animal species have demonstrated the clear and reproducible health benefits of providing supplemental carotenoid-oxygen copolymers in the form of OxC-beta™ Livestock, over and above supplemental vitamin A. As there is no vitamin A or β-carotene present in OxC-beta™ Livestock and it is without vitamin A activity, any benefits observed with OxC-beta supplementation therefore must be independent of both vitamin A and intact β-carotene itself. The trial results, combined with supporting in vitro studies clearly point to β-carotene copolymer being the actual source of the observed health benefits.

OxC-beta™ Livestock’s demonstrated beneficial effects in trials with thousands of animals in different countries and under a variety of conditions are consistent with this naturally-occurring, active form of β-carotene contributing to a better-balanced, healthier feed ration by compensating for the lack or deficiency of what is a previously unrecognized forage component.

Companion Animals Also Benefit from OxC-beta™ Products

With over 1 million administrations of Oximunol™ tablets and Vivamune™ chews provided to canines over 8 years, clearly observed benefits include supporting or promoting healthy skin and coat, healthy joint function, normal mobility and normal intestinal function.

Oxidation of β-carotene is a unifying concept that explains both the micronutrient’s vitamin A and non-vitamin A activities. The extended system of double bonds common to all carotenoids means these other compounds also are capable of related non-vitamin A activities.


References

  1. Burton, G. W.; Daroszewski, J.; Nickerson, J. G.; Johnston, J. B.; Mogg, T. J.; Nikiforov, G. B. ß-Carotene autoxidation: oxygen copolymerization, non-vitamin A products and immunological activity. J. Chem. 2014, 92, 305-316. DOI: 10.1139/cjc-2013-0494 (Open Access).
  2. EFSA Scientific Opinion on the safety and efficacy of beta‐carotene as a feed additive for all animal species and categories. EFSA Journal 2012, 10, 2737 (Open Access).
  3. Johnston, J. B.; Nickerson, J. G.; Daroszewski, J.; Mogg, T. J.; Burton, G. W. Biologically active polymers from spontaneous carotenoid oxidation. A new frontier in carotenoid activity. PLoS ONE 2014, 9, e111346 (Open Access).
  4. Burton, G. W.; Daroszewski, J.; Mogg, T. J.; Nikiforov, G. B.; Nickerson, J. G. Discovery and Characterization of Carotenoid-Oxygen Copolymers in Fruits and Vegetables with Potential Health Benefits. J Agric Food Chem 2016, 64, 3767-3777 (Open Access).
  5. Schaub, P.; Wust, F.; Koschmieder, J.; Yu, Q.; Virk, P.; Tohme, J.; Beyer, P. Nonenzymatic beta-Carotene Degradation in Provitamin A-Biofortified Crop Plants. J Agric Food Chem 2017, 65, 6588-6598 (an abstract of the article is available at: https://pubs.acs.org/doi/abs/10.1021/acs.jafc.7b01693).