Fe in required in various biological and metabolic processes, including DNA synthesis, electron transport, and oxygen transport in eukaryotic cells. So, its deficiency has many health consequences, so it must be supplied by diet. 1-4 Mineral salts are not available for cellular metabolism, so the biological systems of living organisms convert them into an organic and absorbable form. As shown in the food chain in Fig. 1, the chemoautotroph, in the first level of the food chain use inorganic compounds as an energy source and will convert them to organic form during chemical reactions. 5 Animals, will converted Fe into heme. Human diet may be supplied by both of forms. Since iron exists mainly in erythrocytes as the heme compound hemoglobin 2-4, the iron in animal meat is more absorbable in compare to mineral form. The amount of organic iron in diet cannot meet the all need e.g. in children and pregnant women. So, at risk individuals should consume supplements and fortified foods. 6
Based on the latest World Health Organization data in spring 2023, the anemia rate in developed countries has been controlled (Fig. 2) 7, 8. However, the mineral iron, unlike its organic form, has less solubility, so it is not easily absorbed from the digestive tract and has less bioavailability, more toxicity and gastrointestinal side effects. 9 Minerals, when fed in excess, can readily generate free radicals, while organic form has less potential to be toxic. 10,11
In the meantime, biotransformation is a good approach to overcome problems of mineral iron. Human food chain may be enriched with inorganic iron. Microbial biotransformation is more controllable and possible in microorganisms than in plants and animals. 12
Yeasts are known as the best option because they have the potential to act as a small and natural biotransformation factory. They absorb the mineral elements enriched in the growth medium under controlled and specific conditions and convert them into organic form by combining with macromolecules of cells 13,14 Therefore, Iron-fortified yeasts currently being evaluated as a promising source of iron to prevent and reduce iron deficiency in humans and animals 15. Animal studies have shown that organic iron produced in yeast has a greater potential to absorb and improve anemia than the inorganic form. 15,16 The mechanism of iron accumulation in yeast is vague and needs more research, but studies show when iron is abundant in the extracellular environment, the cell accumulates them in cytosol, mitochondria, nuclei and endoplasmic reticula.17-22 So, cell is protected from the toxic effect of Fe, and acts as a resource of iron. 17-23 A tiny fraction of iron accumulates as a solution in the cytosol and vacuoles, while a considerable portion (88 − 68%) is undissolved and chelated to cell constituents. 24 Studies have shown that vacuoles 20 and cell walls 22 are the main iron storage compartment. Iron may be complexed with polyphosphate and organic acids in the vacuole .17
Organisms with the ability to biotransformation and health-promoting properties could be a good carrier for food micronutrient enrichment. Saccharomyces, has an important role in the production of fermented foods and beverages from abcient due to its high nutritional value, bioavailability, safety, and non-toxicity. So, they have been given much attention in the enrichment and production of organic micronutrients, 6,15,25-27 as a good source of Fe, especially for vegans and vegetarians. Also, it has great potential in the bread industry. Nutrients, including iron, are lost in the flour production process, so the bread industry has always tried to restore the lost iron. 28 Previous reports show that the amount of yeasts enrichment and biotransformation depends on various factors, including growth conditions. Martínez-Garay et al., studied a large number of S. cerevisiae with genetic diversity exposed to different concentrations of iron. They uncovered that the amount of iron accumulation is also highly dependent on their genetic pattern. In such a way that when the iron concentration in the medium increases, iron-sensitive strains in terms of molecular mechanism, accumulate iron more and faster than iron-resistant isolates. 29 Kyyaly el al. demonstrated that yeast with the 15 mg/g iron accumulation compared to other concentrations tested and inorganic form, achieved the best baking properties (leavening ability) 15. Also Gaensly et al, concluded that the iron-enriched yeasts maintain its fermentation power. So could be employed as an iron source for supplementing bakery products. 30 But Novsad et al. They found that the fermentation activity of yeast is reduced when enriched with a large amount of iron, which happened here under the influence of a pulsed electric field, so it can only be used as a medicinal supplement. 31
In this research, for the first time, the S. boulardii as an economic, health-giving probiotic was enriched with iron and its biotransformation capacity have been investigated. Also, Box- Behnken design process was used as one of the powerful common statistical tools of response surface method, to study the combined effect among three important variables (concentration of molasses, iron and KH2PO4) and finding their optimal levels on iron-biotransformation by yeast.