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Biofortification of vegetables with micronutrients

Biofortification of vegetables with micronutrients

An extra portion of zinc

Powerful forearms, a pipe in the mouth, a sailor's hat: It takes him only seconds to open and empty the can of spinach. He faces his next brawl with superhuman strength. This is how we all know Popeye, the sailor. The secret of his strength is in the high iron content of spinach. This idea got innumerable parents to trying to get their children to “like” the ­rather unpopular vegetable. Unfortunately, some things are quite wrong here: While spinach is tasty and healthy, it is not a wonder drug. As in most plant tissues, its iron content is around 30 µg/g – and not 10 times as much, as it was often assumed because the dry and fresh weights were once swapped. However, it is perfectly accurate that iron gives strength. It is an essential micro nutrient.

However, the World Health Organisation WHO estimates that about half the global population is suffering from an iron undersupply [1]. A deficit like this is ­referred to as “hidden hunger” today, because the consequences are often not evident at once. Insufficient intake of essential minerals and vitamins can lead to severe health impairment and millions of years of life lost by sickness and premature death (DALYs=disability-adjusted life years in the language of epidemiology). Many people are suffering not just from iron deficits. The “Big Four” also include zinc, iodine and vitamin A. Estimates suggest that zinc deficiency threatens up to 2 Bn people. Figures for iodine are similar. Approx. 20% of the children in the world suffer from Vitamin A deficiency. Globally, iron and zinc deficits are on place 9 and 11 of the most frequent causes of death respectively [1].

Why are iron and zinc essential?

Nothing can live without these metals. Iron can be present in two oxidation states under physiological conditions and therefore has been recruited for innumerable redox reactions during evolution. Cellular respiration, for example, would be impossible without iron-sulphur proteins. Metals are indispensable for the interaction of proteins with small molecules as well. Most of the iron in our body is localised in the haemo­globin of the red blood cells, where it binds oxygen. An easy to diagnose consequence of iron deficit therefore is anaemia. The consequences if iron deficit include reduced mental and physical performance.
Zinc is part of approx. 9% of the proteins in eukaryotes [2]. Zinc is found in enzymes of all six classes as a catalytic co-factor, particularly frequently in hydrolases. Zinc is an important structure element particularly in DNA-binding regulatory proteins (zinc-­finger motif). In spite of the diversity of biological functions of zinc, a deficit has been difficult to determine, in contrast to the situation with iron. The evidence for a deficit is mainly based on the positive effects of zinc addition to the food (= zinc supplementation). The results here are clear. Meta analyses document a significant reduction of diarrhoea (about 20%), pneumonia (about 15%) and mortality (about 18%) in children supplied with additional zinc [3]. A case study in the latest World Health Report of the WHO cites comparable figures for children under two in ­Bangladesh to whom 70mg zinc per week were administrated [4]. The positive effect on the immune defence that can be derived from these findings has also been evidenced for older people. Both zinc and iron deficits in early childhood also result in stunted growth that cannot be balanced out even by sufficient supply in later development stages.


Fig. 1 Typical natural variation of the zinc contents of different plant foods. The averages and maximums from the HarvestPlus database according to Pfeiffer & McClafferty [2007] Crop Sci. are shown. 47, p. 88–p. 105.


Fig. 2 The hyper-metal-accumulating plant Arabidopsis halleri. It grows in metal-polluted habitats as shown in this figure (strong zinc and lead contamination in the area of a zinc foundry) as well as uncontaminated soils. A. halleri is able to accumulate more than 10,000 and up to 50,000 µg of zinc/g dry weight anywhere. (I would like to thank Ricardo Stein, Romário Melo; University Bochum and Stephan Höreth; University Bayreuth, for the photograph).

How can supply with iron and zinc be improved?

In the light of the severe consequences of “hidden hunger”, improved supply with essential minerals has become an important goal around the world. A body of laureates of the Nobel Price for economy, evaluating possible solutions for the most urgent global welfare problems in the “Copenhagen Consensus”, created a ranking with three measures for fighting “hidden hunger” among the first five positions in 2008. In 2012, this body assigned bundled measures to improve micro nutrient supply the highest priority (www.copenhagenconsensus.com). The food industry is also paying increasing attention to the “Big Four”.

Supplementation, e.g., with zinc tablets or fortification of foods by addition of iodine and iron, can counteract deficiencies but requires an infrastructure such as working distribution paths. Additionally, specifically the locally produced agricultural products are the main food source in many regions of the world. Therefore, diversification of food and increase of micro nutrient content of the plant staple foods (= biofortification) would promise much more success in implementation and range. Increasing the diversity of their diet, e.g. by adding more meat and vegetables, however, often fails due to poverty.

Strategies of biofortification

For about ten years, the basics and implementation of biofortification have been subject to increasingly intense research. Some important activities are bundled in the HarvestPlus programme, which is essentially funded by the Gates Foundation (www.­harvestplus.org). The goal is to increase the micro nutrient contents in the most important food plants to about two to three times the current values. “Hidden hunger” is expected to reduce significantly by this increase.

Generally, there are three strategies: Changing the growing practices, classic plant breeding and bioengineering The zinc content of grain seeds specifically can be increased by corresponding fertilisation. Like supplementation and fortification, this method requires the corresponding infrastructure and capital. Particularly in African countries, however, even fertilisation with the nutrients of nitrogen and phosphate, which are most important for quantity, is not common. Therefore, the most sustainable progress is expected by permitting development of micro-nutrient-rich varieties of the most important food plants and establishing them.

Natural Variation of the Micro Nutrient Content

Breeding of agricultural crops with a higher micro nutrient density requires the corresponding genetic potential, i.e. a natural variation of these properties. In fact, the zinc and iron contents in the consumed organs of some agricultural crops can differ by a factor of 2–4 (fig.1). The first success is based on this range. A few weeks ago, for example, a rice type developed in the HarvestPlus programme with its zinc content raised by about 50% on average was introduced in Bangladesh (www.harvestplus.org/content/media).

Bioengineering approaches

Introduction of additional genes could achieve higher increases, is applicable even with species where classic breeding is virtually impossible (e.g. banana) or could even add new properties. The likely most prominent example for biofortification is the “Golden Rice” that was developed by scientists in Zürich and Freiburg. This rice synthesises provitamin A not only in its leaves but also in the starch-rich endosperm of the grain. Eating about 100g of “Golden Rice” per day would cover 60% of the Vitamin A-demand of an 8-year old child. This could help millions of children. Nevertheless, the “Golden Rice” is not cultivated yet due to strong political resistance.

Zinc and iron biofortification is an even more complex biological problem than vitamin biosynthesis. Metal ions cross a long distance from the ground to the seeds. Their reactivity - the reason why they were recruited during evolution - requires a precisely regulated network of transport and storage processes. Potentially harmful interactions with proteins and other cellular components must be suppressed by complex formation with designated ligands. At the same time, the metal ions in and between cells must be mobile. We do not have comprehensive understanding of how this metal homoeostasis works in detail in any organism. Some of the processes involved have been explained molecularly in the meantime. The examination of metal-hyper-accumulating plants that may acquire zinc concentrations in their leaves up to 1000 times as high as those of virtually any other organism is one successful research approach (fig.2). The molecular analysis of this extreme property has, e.g., disclosed the key role of zinc-pumping P-type-ATPases and the metal-ligand nicotianamine for the long-distance transport of zinc from the roots to the leaves (fig.3). Genetic engineering of the synthesis of nicotianamine in rice permitted the development of plants containing three times as much iron and zinc in their grains [7,8]. The changed expression of three genes for metal homeostasis in a rice species grown in Myanmar led to an increase of the iron content in the grain by 3.4 times. In Myanmar, 75% of the children and 71% of the pregnant women suffer from anaemia caused by iron deficiency [9].


Fig.3 Mechanisms of the long-distance transport of zinc from the roots to leaves and seeds This simplified chart shows cell types of the roots and the xylem as part of the transporting tissue. The absorption of zinc ions from the soil solution through the plant membrane is conveyed by specialist transporters. The formation of complexes of the metal ligand nicotianamine with Zn(II) ensures the mobility of Zn(II) in the cell-cell transport. Zn(II) is finally pumped to the xylem by P-type-ATPases such as HMA4 and can thus reach the above-ground organs.

Bioavailability of iron and zinc

In the end, the micro nutrient content in the plant products is not solely decisive for biofortification. Their bioavailability, i.e. the share that can actually be absorbed by the human digestive tract, is essential as well. The reactivity of iron and zinc ions mentioned permits highly stable complexes to be formed with food components such as phytate (Myo-Inositol-hexakisphosphate), which prevent absorption in intestinal epithelial cells. Only about 15–35% of the zinc and 5–15% of the iron taken up in the food are absorbed by the body. The bandwidth is due to the influence of other food components and one reason for the benefits of larger variety in one's diet. For example, ascorbic acid supports iron intake, because it reduces the less-available Fe(III) to better-available Fe(II). Again, breeding and bioengineering can help. Low-phytate-species of some cultural crops have been developed. However, reduction of the phosphate storer phytate has often coincided with yield reductions. The above genetically engineered rice with increased nicotianamine synthesis can highly effectively balance out the iron deficit of mice suffering from anaemia and thus evidently provides bioavailable iron [7].

It can be expected that our molecular understanding of metal homoeostasis in plants will increase quickly over the next years, leading to growing potentials for biofortification by breeding and bioengineering. It remains to be hoped that we as a society will learn to assess crops according to their properties instead of according to how they are developed as quickly as possible. Then we should take the first step and at least admit those genetically engineered species that only use variations available in their natural gene pool. These are types where only genes from the same species or a closely related, sexually compatible species, have been transferred (=“Cisgenesis”).

Literature

[1] WHO (2002) http://www.who.int/whr/2002/en/whr02_en.pdf
[2] Andreini, C. et al. (2006), J. Proteome Res. 5, 3173–3178
[3] Gibson RS (2012), Zinc deficiency and human health: etiology, health consequences, and future solutions. Plant Soil 361, 291–299
[4] WHO (2013) http://www.who.int/whr/2013/report/en/index.html.
[5] Hanikenne, M. et al. (2008), Nature 453, 391–395
[6] Deinlein, U. et al. (2012), Plant Cell 24, 708?–723
[7] Lee, S. et al. (2009), Proc Natl Acad Sci USA 106, 22014–22019
[8] Lee, S. et al. (2011), Plant Biotechnol. J. 9, 865–873
[9] Aung, M.S. et al. (2013), Front. Plant Physiol. 4, 158

Photo: © istockphoto.com| Andrew Rich, mstay

L&M int. 1 / 2014

The articles are publishes in issue L&M int. 1 / 2014.
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