Riboflavin (vitamin B2) is a hydrophilic (water-soluble) vitamin of the B group. It is visually distinguished from most hydrophilic vitamins by its intense yellow fluorescent color, which is reflected in its name (flavus: yellow). Historical names of riboflavin include ovoflavin, lactoflavin and uroflavin, which refer to the first isolation of this substance. In 1932, Warburg and Christian obtained the “yellow ferment” from yeast and identified it as a coenzymatically active flavin mononucleotide (FMN). The structure of riboflavin was elucidated in 1933-34 by Kuhn and Wagner-Jauregg and synthesized in 1935 by Kuhn, Weygand, and Karrer. In 1938, the discovery of flavin adenine dinucleotide (FAD) as a coenzyme of D-amino acid oxidase was made by Wagner. The basic structure of vitamin B2 is the tricyclic isoalloxazine ring system, which has pronounced redox properties (reduction/oxidation properties). Attached to the N10 atom of the isoalloxazine molecule is ribitol, a pentavalent alcohol sugar that is critical for vitamin efficacy. The biologically active compound of vitamin B2 is 7,8-dimethyl-10-(1-D-ribityl)isoalloxazine. The IUPAC (International Union of Pure and Applied Chemistry) proposed the term riboflavin as a short name.Like thiamine (vitamin B1), riboflavin possesses a high degree of structural specificity, so that even slight changes in the molecular structure can be accompanied by a reduction or loss of vitamin efficacy or – in certain cases – by an antagonistic (opposite) mode of action. Replacement of the ribityl residue by galactose (→ galactoflavin) results in the strongest antagonistic effect and quickly leads to clinical vitamin B2 deficiency. When replacing the ribitol side chain with other carbohydrate analogs, such as arabinose and lyxose, the antagonism is weaker and, in some cases, pronounced only in some animal species, such as the rat. To unfold biological activity, riboflavin must be phosphorylated at the C5 atom of the ribitol side chain under the action of riboflavin kinase (enzyme that transfers a phosphate residue by cleaving adenosine triphosphate (ATP)) (→ flavin mononucleotide, FMN) and subsequently adenylated (→ flavin adenine dinucleotide, FAD) by a pyrophosphorylase (enzyme that transfers an adenosine monophosphate (AMP) residue while consuming ATP). FMN and FAD are the major derivatives (derivatives) of riboflavin and act as coenzymes of oxidases and dehydrogenases. In animal and plant organisms, more than 100 enzymes, and in mammals more than 60 enzymes, are known to be FMN- or FAD-dependent – so-called flavoproteins or flavin enzymes, respectively. Vitamin B2 is very heat stable, oxygen sensitive and highly sensitive to UV light compared to other vitamins. Riboflavin and non-protein-bound flavin derivatives are readily photolytically degraded (cleavage of a molecule under the influence of UV light) to vitamin-inactive lumichrome (dimethylisoalloxazine) or lumiflavine (trimethylisoalloxazine), in which the aliphatic side chain is partially or completely cleaved. For this reason, products containing vitamin B2 should be stored in an airtight container and protected from light.
Synthesis
Riboflavin is synthesized by plants and microorganisms and enters the animal organism through the food chain. Consequently, vitamin B2 is widely distributed in plants and animals and is present in numerous foods.
Absorption
In food, riboflavin occurs in free form, but primarily as protein-bound FMN and FAD – flavoprotein. Riboflavin is released by gastric acid and nonspecific phosphatases and pyrophosphatases (enzymes that hydrolytically (with water retention) cleave phosphate residues) of the upper small intestine. The absorption (uptake via the intestine) of free riboflavin in the upper small intestine, especially in the proximal jejunum (empty intestine), is subject to a dose-dependent dual transport mechanism. In the physiological (normal for metabolism) range up to about 25 mg, riboflavin is actively absorbed in response to a sodium gradient by means of a carrier following saturation kinetics. Above physiological doses, absorption of vitamin B2 additionally occurs by passive diffusion [1, 2, 4-6, 8]. The absorption rate of riboflavin after intake of physiological doses is on average between 50-60%.Uptake of the B vitamin in the dietary composite and the presence of bile acids promote absorption. Presumably, the delayed gastric emptying rate and prolonged gastrointestinal transit time play a role in promoting contact with the absorbing surface. In the intestinal mucosa cells (mucosal cells), part of the absorbed (ingested) free riboflavin is converted to FMN by riboflavin kinase and subsequently to FAD by a pyrophosphorylase to keep the concentration of free vitamin B2 as low as possible and to ensure further absorption. However, the majority of absorbed free vitamin B2 is converted to its coenzymatically active forms FMN and FAD in the liver after portal vein transport.
Transport and distribution in the body
Free riboflavin, FMN, and FAD are released from the liver into the bloodstream. There, most of vitamin B2 is present as FAD (70-80%) and FMN and only 0.5-2% in free form. Riboflavin and its derivatives are transported in the blood plasma in protein-bound form. Main binding partners are plasma albumins (80 %), followed by specific riboflavin-binding proteins (RFBPs) and globulins, especially immunoglobulins. For transport into target cells, vitamin B2 is dephosphorylated under the action of plasmatic phospatases (enzymes that hydrolytically (under water retention) cleave phosphate residues), since only free, unphosphorylated riboflavin can pass cell membranes by diffusion. Intracellularly (inside the cell), conversion and fixation into the coenzyme forms occurs again – metabolic trapping. Almost all tissues are capable of forming FMN and FAD. Particularly high conversion rates are found in the liver, kidney, and heart, which therefore have the highest concentrations of riboflavin-70-90% as FAD, <5% as free riboflavin. As with all hydrophilic (water-soluble) vitamins, with the exception of cobalamin (vitamin B12), the storage capacity of vitamin B2 is low. Tissue stores exist in the form of protein- or enzyme-bound riboflavin. In the case of a deficiency of apoprotein or apoenzyme, excess riboflavin cannot be stored, resulting in a reduced riboflavin stock.In adult humans, about 123 mg of vitamin B2 is retinated (retained by the kidney). This amount is sufficient to prevent clinical deficiency symptoms for about 2-6 weeks – with a biological half-life of circa 16 days. Riboflavin-binding proteins (RFBPs) are important for both transport processes and metabolism (metabolism) of vitamin B2. In the liver and kidney, specific actively working transport systems have been demonstrated that contribute to the enterohepatic circulation (liver-gut circulation) and tubular reabsorption (reabsorption in the renal tubules) of riboflavin to some extent according to individual requirements. According to animal studies, riboflavin transport to the central nervous system (CNS) is also subject to an active mechanism and homeostatic regulation (self-regulation) that protects the CNS from both under- and over-supply. In women in gravidity (pregnancy), specific RFBPs have been discovered to maintain a gradient in blood serum from maternal (maternal) to fetal (fetal) circulation. Thus, even if the mother’s vitamin B2 supply is inadequate, the riboflavin supply necessary for fetal growth and development is largely ensured.While estrogens stimulate the synthesis of RFBPs, poor nutritional status leads to RFBP deficiency.
Metabolism
The metabolism of riboflavin is controlled by hormones and RFBPs depending on individual vitamin B2 status. Riboflavin-binding proteins and hormones, such as triiodothyronine (T3, thyroid hormone) and aldosterone (adrenocortical hormone), regulate the formation of FMN by stimulating riboflavin kinase activity. Subsequent synthesis of FAD by pyrophosphorylase is controlled by end product inhibition to prevent FAD excess. The coenzymes FMN and FAD are provided by modulating (modifying) the activity of the respective enzymes only to the extent required by the organism according to its needs.Under conditions of decreased serum T3 levels and/or decreased concentration of RFBPs, as in malnutrition (undernutrition/malnutrition) and anorexia (loss of appetite; anorexia nervosa: anorexia), a decrease in plasma FAD concentration and a substantial increase in free riboflavin, normally present only in trace amounts, in erythrocytes (red blood cells) are observed.
Excretion
Excretion of vitamin B2 occurs predominantly through the kidney as free riboflavin. As much as 30-40% of 7-hydroxymethyl-, 8-hydroxymethyl-, or 8-alpha-sulfonylriboflavin and trace amounts of other metabolites (intermediates) are renally eliminated (excreted by the kidneys). After high-dose supplementation of vitamin B2, 10-hydroxyethylflavin may appear in the urine as a result of bacterial degradation. The coenzyme forms FMN and FAD cannot be detected in urine. Clearance (excretion) data indicate that approximately half of the plasmatic riboflavin is eliminated in the urine. Renal clearance is higher than glomerular filtration. A healthy adult excretes 120 µg of riboflavin or more in the urine in 24 hours. Riboflavin excretion < 40 mg/g creatinine is an indicator of vitamin B2 deficiency. Patients requiring dialysis due to renal failure (chronic renal failure/acute renal failure) are at increased risk for vitamin B2 deficiency because riboflavin is lost during dialysis (blood purification).Less than 1% of vitamin B2 is eliminated in the bile with feces (via stool). The elimination or plasma half-life (the time that elapses between the maximum concentration of a substance in blood plasma to the fall to half that value) depends on the riboflavin status and the dose supplied. While a rapid elimination half-life is 0.5-0.7 hours, a slow plasma half-life varies from 3.4-13.3 hours. There is no linear relationship between dietary vitamin B2 intake and renal riboflavin excretion. While below tissue saturation (≤ 1.1 mg vitamin B2/day) the rate of elimination changes only insignificantly, there is a marked increase in riboflavin excretion – break point (> 1.1 mg vitamin B2/day) when saturation is reached. In gravidity (pregnancy), due to the induction (introduction, in the sense of increased formation) of riboflavin-binding proteins, the excretion of vitamin B2 via the kidney is reduced. A decreased excretion rate is also found in tumor disease (cancer) because patients have increased serum concentrations of immunoglobulins that bind vitamin B2.
Lipid-soluble derivatives of riboflavin
Lipid-soluble (fat-soluble) compounds, such as tetrabutyric acid or tetranicotinyl derivatives of riboflavin, can be prepared by esterification of the hydroxyl (OH) groups of the ribitol side chain. Compared to the native (original), hydrophilic (water-soluble) vitamin, the lipophilic (fat-soluble) riboflavin derivatives exhibit better membrane permeability (membrane traversability), improved retention (retention), and slower turnover (turnover). Preliminary studies show beneficial effects of these derivatives in blood coagulation disorders and the treatment of dyslipidemia. In addition, the use of the lipid-soluble riboflavin compounds-alone or in combination with vitamin E-may prevent the accumulation (buildup) of lipid peroxides as a result of exposure to carbon tetrachloride or to carcinostatic agents, such as adriamycin.
