Rukiin folaattien määritys ja pitoisuuksien lisääminen elintarvikeprosesseilla

The average daily intake of folate, one of the B vitamins, falls below recommendations among the Finnish population. Bread and cereals are the main sources of folate, rye being the most significant single source. Processing is a prerequisite for the consumption of whole grain rye; however, little is known about the effect of processing on folates. Moreover, data on the bioavailability of endogenous cereal folates are scarce. The aim of this study was to examine the variation in as well as the effect of fermentation, germination, and thermal processes on folate contents in rye. Bioavailability of endogenous rye folates was investigated in a four-week human intervention study. One of the objectives throughout the work was to optimise and evaluate analytical methods for determining folate contents in cereals. Affinity chromatographic purification followed by high-performance liquid chromatography (HPLC) was a suitable method for analysing cereal products for folate vitamers, and microbiological assay with Lactobacillus rhamnosus reliably quantified the total folate. However, HPLC gave approximately 30% lower results than the microbiological assay. The folate content of rye was high and could be further increased by targeted processing. The vitamer distribution of whole grain rye was characterised by a large proportion of formylated vitamers followed by 5-methyltetrahydrofolate. In sourdough fermentation of rye, the studied yeasts synthesized and lactic acid bacteria mainly depleted folate. Two endogenous bacteria isolated from rye flour were found to produce folate during fermentation. Inclusion of baker s yeast in sourdough fermentation raised the folate level so that the bread could contain more folate than the flour it was made of. Germination markedly increased the folate content of rye, with particularly high folate concentrations in hypocotylar roots. Thermal treatments caused significant folate losses but the preceding germination compensated well for the losses. In the bioavailability study, moderate amounts of endogenous folates in the form of different rye products and orange juice incorporated in the diet improved the folate status among healthy adults. Endogenous folates from rye and orange juice showed similar bioavailability to folic acid from fortified white bread. In brief, it was shown that the folate content of rye can be enhanced manifold by optimising and combining food processing techniques. This offers some practical means to increase the daily intake of folate in a bioavailable form.B-vitamiinien ryhmään kuuluvat folaatit esiintyvät useina muotoina, vitameereina. Folaatteja tarvitaan yhden hiiliyksikön siirtoreaktioissa mm. DNA:n ja RNA:n synteesissä sekä aminohappoaineenvaihdunnassa. Folaattien puutoksen tiedetään aiheuttavan megaloblastista anemiaa ja sikiöillä hermostoputken sulkeutumishäiriöitä. Näiden vitamiinivaikutusten lisäksi folaateilla on todettu terveyttä edistäviä vaikutuksia, kuten pienentynyt riski sairastua tiettyihin syöpiin sekä sydän- ja verisuonitauteihin. Folaattianalytiikka on haastavaa, sillä folaatit esiintyvät elintarvikkeissa usein pieninä pitoisuuksina ja tuhoutuvat herkästi lämmön, valon ja hapettumisen vaikutuksesta. Suomalaisten folaattisaannista noin 40% on peräisin viljatuotteista. Lähtökohtana väitöskirjatyölle oli se, että vaikka ruis on merkittävin yksittäinen folaattilähde suomalaisessa ruokavaliossa, rukiin folaateista, niiden hyväksikäytettävyydestä ja elintarvikeprosessien vaikutuksista ei ole paljonkaan tietoa. Tutkimuksen tavoitteena oli tutkia folaattipitoisuuksien vaihtelua lajikkeiden välillä sekä keinoja lisätä pitoisuuksia elintarvikeprosesseilla. Lisäksi tutkittiin rukiin luontaisten folaattien hyväksikäytettävyyttä. Mikrobiologinen menetelmä oli luotettava ja tarkka kokonaisfolaattipitoisuuden määrittämisessä, ja suuren erotuskyvyn nestekromatografia (HPLC) soveltui hyvin vitameerien pitoisuuksien määrittämiseen. HPLC:llä määritetyt vitameerien yhteenlasketut pitoisuudet olivat kuitenkin noin 30% pienemmät kuin mikrobiologisesti määritetyt kokonaisfolaattipitoisuudet. Rukiin folaateista suurin osa oli formyloituja muotoja sekä 5-metyylitetrahydrofolaattia. Leivinhiivan käyttäminen ruisleivonnassa nosti folaattitasoa niin, että leivässä saattoi olla enemmän folaatteja kuin ruisjauhossa. Koeputkifermentaatioissa hiivojen todettiin syntetisoivan folaatteja, kun taas maitohappobakteerit pääosin kuluttivat niitä. Idätys paitsi nosti folaattipitoisuuksia, myös aiheutti selviä muutoksia vitameerijakaumaan: idätetyn rukiin tärkeimmät vitameerit olivat 5-metyylitetrahydrofolaatti sekä tetrahydrofolaatti. Juuri-iduissa folaattipitoisuudet olivat erityisen suuria. Folaattien synteesi idätyksen aikana pystyi kompensoimaan folaattien tuhoutumista idätystä seuraavissa lämpökäsittelyissä. Rukiista ja appelsiinimehusta saadut luontaiset folaatit olivat yhtä hyvin hyväksikäytettävissä kuin valkoiseen leipään lisätty foolihappo. Väitöskirjatyössä osoitettiin, että rukiin folaattipitoisuuksia pystytään merkittävästi kasvattamaan prosessoinnin keinoin ja että verrattain pienikin lisä rukiin luontaisia folaatteja vaikuttaa positiivisesti folaattistatukseen

Bioaccessibility of Folate in Faba Bean, Oat, Rye and Wheat Matrices

Cereals and legumes are rich in folate. However, due to the instability of folate, processing and digestion can induce significant folate loss. In this paper, folate bioaccessibility of faba bean, oat, rye and wheat flours and pastes was studied using a static in vitro digestion model. Folate bioaccessibility depended on food matrices, varying from 42% to 67% in flours and from 40% to 123% in pastes. Digestion was associated with the inter conversion of formyl folates, as well as the increase of oxidised vitamers and decrease of reduced vitamers. Especially in faba bean, 5-methyltetrahydrofolate showed surprisingly good stability both in digestion and heat treatment, resulting in high bioaccessibility. The physiological concentration of ascorbic acid did not stabilise folate in digestion; however, a higher level helped to maintain reduced vitamers. Heat treatment (10-min paste making) could improve folate bioaccessibility by liberating folate from the food matrices and by altering folate vitamer distribution.Peer reviewe

The bioaccessibility of folate in breads and the stability of folate vitamers during in vitro digestion

Both the liberation and stability of endogenous folate are relevant to the bioaccessibility of folate. Since folates are unstable, in addition to studying the natural folate content in foods, bioaccessibility should be considered. To understand folate changes during digestion, a mixture of standard folate compounds was subjected to a static in vitro gastrointestinal digestion assay. Next, different types of bread were analysed to study how food matrices influence folate bioaccessibility. Folates were identified and quantitated by a UHPLC-PDA/FL method. Folic acid and 10-formylfolic acid were stable throughout the digestion, and the conversions among formyl folates and 5,10-methenyltetrahydrofolate were triggered at the gastric phase. Tetrahydrofolate began to degrade during the oral phase and was lost completely during the gastric phase. During the intestinal phase, 5-methyltetrahydrofolate began to degrade and suffered a 60% loss. With bread matrices, folate conversions and the decrease of reduced folates were also common, but the extent of changes varied. Generally, rye breads had the highest (80–120%) bioaccessibility of folate, while oat breads had the lowest (31–102%). The high proportion of 5-methyltetrahydrofolate could result in low bioaccessibility because of its relatively low stability during digestion in bread matrices. An increase in 10-formylfolic acid content was observed for all the breads, but 10-formyldihydrofolate seemed to be more stable in rye breads than in oat and wheat breads. The results showed that folates undergo significant changes during digestion and that food matrices could be modified to affect these changes towards better folate bioaccessibility.Peer reviewe

Quantification of folate in the main steps of traditional processing of tef injera, a cereal based fermented staple food

Injera is an Ethiopian fermented flatbread preferably made from whole grain cereal (tef). Tef it is increasingly used to produce gluten-free pasta and bread, but the folate content of teff and products made from it remains unknown. Given that folate deficiencies lead to several health disorders, the aim of this study was to quantify folate in each of the three main steps of traditional processing of tef injera. Total folate contents of tef flour, fermented batter and injera were determined through microbiological assays using Lactobacillus rhamnosus (ATCC 7469). Folate content of tef flour was 8.7 mu g/100 g of dry matter content, which is in the same range as the richest cereals like oats. The increase in folate content due to fermentation was highly variable (60-148%). Cooking always led to folate losses, with a maximum of 52.8%. Altogether, injera processing increased folate retention between 38.0 and 121.8%. Folate content of injera was 14.3 mu g/100 g on fresh weight-basis. Tef injera can contribute up to 10% of the recommended nutrient intake of folate for children aged 1-3 and women of reproductive age. Although the folate content of teff is already high, future studies should focus on optimizing the folate content of injera.Peer reviewe

5-Methyltetrahydrofolate Is a Crucial Factor in Determining the Bioaccessibility of Folate in Bread

This study investigated the bioaccessibility of folate in wheat bread baked with different ingredients and processing methods. Next, different matrices were spiked with 5-methyltetrahydrofolate, gallic acid (GA), or both to investigate the stability of 5-methyltetrahydrofolate during in vitro digestion. The folate bioaccessibility in bread varied from 44 to 96%. The inclusion of whole-grain or faba bean flour significantly improved both folate content and bioaccessibility. Baking with yeast increased the folate content by 145% in bread but decreased folate bioaccessibility compared to the bread without added yeast because of the instability of 5-methyltetrahydrofolate. Spiking experiments confirmed oxidation as a critical reason for 5-methyltetrahydrofolate loss during digestion. However, GA protected this vitamer from degradation. Additionally, 5-methyltetrahydrofolate was less stable in whole-grain wheat matrices than other matrices. This study demonstrated that the stability of 5-methyltetrahydrofolate is crucial for folate bioaccessibility in bread, and methods for stabilizing this vitamer should be further studied.Peer reviewe

Lactobacillus plantarum P2R3FA Isolated from Traditional Cereal-Based Fermented Food Increase Folate Status in Deficient Rats

Folate deficiencies are widespread around the world. Promoting consumption of folate-rich foods could be a sustainable option to alleviate this problem. However, these foods are not always available. Cereals, being a staple food, could contribute to folate intake. They are fermented prior to consumption in many African countries, and fermentation can modify the folate content. In Ethiopia, injera is a widely consumed fermented flat bread. The main drivers of its fermentation are lactic acid bacteria (LAB). The aim of this work was to isolate and identify folate-producing LAB from injera fermented dough and to evaluate their ability to increase folate status after depletion in a rat model. Among the 162 strains isolated from 60 different fermentations, 19 were able to grow on a folate-free culture medium and produced 1 to 43 µg/L (24 h, 30 °C incubation). The four highest folate producers belonged to the Lactobacillus plantarum species. The most productive strain was able to enhance folate status after depletion in a rat model, despite the relatively low folate content of the feed supplemented with the strain. Folate-producing L. plantarum strain has potential use as a commercial starter in injera production

Lactobacillus plantarum P2R3FA Isolated from Traditional Cereal-Based Fermented Food Increase Folate Status in Deficient Rats

Folate deficiencies are widespread around the world. Promoting consumption of folate-rich foods could be a sustainable option to alleviate this problem. However, these foods are not always available. Cereals, being a staple food, could contribute to folate intake. They are fermented prior to consumption in many African countries, and fermentation can modify the folate content. In Ethiopia, injera is a widely consumed fermented flat bread. The main drivers of its fermentation are lactic acid bacteria (LAB). The aim of this work was to isolate and identify folate-producing LAB from injera fermented dough and to evaluate their ability to increase folate status after depletion in a rat model. Among the 162 strains isolated from 60 different fermentations, 19 were able to grow on a folate-free culture medium and produced 1 to 43 µg/L (24 h, 30 °C incubation). The four highest folate producers belonged to the Lactobacillus plantarum species. The most productive strain was able to enhance folate status after depletion in a rat model, despite the relatively low folate content of the feed supplemented with the strain. Folate-producing L. plantarum strain has potential use as a commercial starter in injera production

Comparative Analysis Reveals Changes in Some Seed Properties in Amaranth Mutant Variety ‘Zobor’ (A. hypochondriacus × A. hybridus)

The aim of our long-term research program is to improve the quality and quantity of amaranth production through mutation breeding using γ-radiation. In this paper, we present the characterization of the new variety ‘Zobor’ of A. hypochondriacus × A. hybridus developed by radiation-induced mutagenesis of hybrid K-433. Multiyear phenotypic characterization of an important yield parameter (1000-seed weight) showed that the studied mutant variety ‘Zobor’ has an advantage in seed weight over the nonirradiated control seeds of K-433 with predictable performance of this yield trait. ‘Zobor’ exhibited changes in seed morphometric parameters, starch particle size, and pasting properties with no change in amylose content and swelling power. Moreover, the seeds of ‘Zobor’ showed the significantly highest folate content among selected amaranth varieties. The mutant variety could, therefore, be interesting for the development of functional foods and as a low-management crop, attractive for cultivation in Europe

Ultra-high performance liquid chromatographic and mass spectrometric analysis of active vitamin B12 in cells of Propionibacterium and fermented cereal matrices

A sensitive and selective method is needed to analyse in situ produced vitamin B12 in plant-based materials, potential new dietary sources of vitamin B12. A UHPLC/UV method was developed and validated for the determination of human active vitamin B12 in cell extracts of Propionibacterium freudenreichii subsp. shermanii and after immunoaffinity purification in extracts of cereal matrices fermented by P. freudenreichii. An Acquity HSS T3 C18 column resulted in a baseline separation, a calibration curve of excellent linearity and a low limit of detection (0.075 ng/5 μL injection). As confirmed by UHPLC–MS, the active vitamin B12 could be separated from pseudovitamin B12. The recovery of vitamin B12 from purified spiked cereal matrices was good (>90%; RSD < 5%). A nutritionally relevant amount of active vitamin B12 was produced by P. freudenreichii in cereal malt matrices (up to 1.9 μg/100 g) in 24 h at 28 °C.Peer reviewe

Riboflavin, niacin, folate and vitamin B12 in commercial microalgae powders

This study aimed to investigate the riboflavin, niacin, folate and B12 content in microalgae powders. Riboflavin was determined with an ultra-high-performance liquid chromatographic (UHPLC) method after extraction and a two-enzyme treatment. Niacin analysis involved mild acidic hydrolysis, with niacin quantitated as the sum of nicotinic acid and nicotinamide using a UHPLC method. Both a microbiological (MBA) and a UHPLC method was used for vitamin B12 quantification as a cyanocobalamin (CNCb1) and UHPLC-mass spectrometry was used to confirm the vitamin B12 form. Total folate was determined both with MBA and as a sum of the vitamers with a UHPLC method after extraction and tri-enzyme treatment. The riboflavin and niacin content varied from 21 to 41 mu g/g and 0.13-0.28 mg/g, respectively, in Chlorella sp., Spirulina (Arthrospira sp.) and Nannochloropsis gaditana powders. Chlorella powders were, on average, richer in total folate (19.7 mu g/g) than Spirulina powders were (3.5 mu g/g). The sum of the folate vitamers determined with UHPLC matched better with the microbiological total folate content in Chlorella than in Spirulina powders. Pseudovitamin B12 was the predominant form over active vitamin B12 in Spirulina powders, whereas Chlorella sp. and N. gaditana powders solely contained active vitamin B12 up to 2.1 mu g/g.Peer reviewe