247 Riboflavin Deficiency

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Proceedings of The 6th Annuual International Conference Syiah Kuala University (AIC Unsyiah) in conjunction with The 12th International Conference on Mathematics, Statistics and Its Application (ICMSA) 2016

October 4-6, 2016, Banda Aceh, Indonesia

Riboflavin Deficiency: What Do We Really Know?

1*Juwita, 2Marisa

1Department of Biochemistry, Faculty of Medicine, University of Syiah Kuala, Darussalam, Banda Aceh 23111, Indonesia; 2Department of Nutrition, Faculty of Medicine, University of Syiah Kuala, Darussalam, Banda Aceh 23111, Indonesia;

*Corresponding Author: [email protected]

Abstract

Riboflavin is a vitamin B namely vitamin B2 is a watersoluble vitamin and stable to the high temperatures condition. These vitamin has a chemical structure of an isoalloxazine ring, bound to ribitol side chains. Source of riboflavin can be derived from vegetable or animal such as milk, eggs, and dark green vegetables. The active form of riboflavin is FAD (Flavin Adenine Dinucleotide) and FMN (Flavin Mononucleotide), serves as a prosthetic group in a variety of enzymes, in particular as a catalyst for oxidation-reduction reactions. Riboflavin through its active form which is FAD and FMN, have an important role in energy production in the form of ATP and the metabolism of carbohydrates, fats, protein, amino acid synthesis, as well as the activation of other vitamins. Deficiency of this vitamin can cause a variety disorders. This review aims to discuss the riboflavin deficiency and its effects, in particular against tampering hyperhomocysteinemia (a risk factor for cardiovascular disease), migraine, anemia, and cataracts.

Key Words: Riboflavin, FAD, FMN, deficiency, hyperhomocysteinemia

Introduction Riboflavin (aka vitamin B2) is a watersoluble vitamin and has the chemical name 7,8-dimethyl-10-ribityl- isoalloxazine (Powers, 2003). The chemical structure is an isoalloxazine ring bound to a ribitol side chain. The form of riboflavin which most common is FAD (flavin adenine dinucleotide), and then FMN (flavin mononucleotide) (Belinda, 2014). These forms of flavin (FAD and FMN) can bind covalently or non-covalently with the enzyme. Riboflavin is relatively stable to heat, but can be quickly degraded through exposure of light (Powers, 2003). The chemical structure of riboflavin and its flavin (FMN and FAD) can be seen in figure 1 below.

Figure 1. Chemical Structure of Riboflavin, FMN, and FAD (Mazzotta et al., 2014).

Source of Riboflavin Riboflavin can be obtained from various sources of food, such as milk and its dairy products, cereals, meat, fish, fruits, and vegetables (especially dark green vegetables that contain high concentrations of riboflavin). Breakfast with cereal and milk is very good to maintain adequate intake of riboflavin (Powers, 2003). Mediterranean Diets, which are characterized by high consumption of fruits, vegetables, and

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cereals, but low consumption of simplex carbohydrates, olive oil, and red wine, are associated with increased levels of vitamins (including vitamin B2) and minerals in the body (Kennedy, 2015).

Role and Mechanism of Riboflavin Riboflavin plays an important role in the body's metabolism. It is a precursor to the formation of FMN and FAD molecules, the biologically active forms of riboflavin. These flavins act as coenzymes which are important for the activity of enzymes involved in energy metabolism (Agostoni et al., 2013; Dai and Koh, 2015). Metabolic reactions that produce energy include oxidation-reduction reaction, electron transfer chain, amino acid oxidation, fatty acid oxidation, and the citric acid cycle (Agostoni et al., 2013; Anshoori and Saedisomeolia, 2014; Kennedy, 2015).

Riboflavin can work together with other B vitamins such as B6, B9, and B12 (Belinda, 2014). Coenzyme FMN and FAD have a role in synthesis and conversion of other vitamins such as niacin, folate, and vitamin B6. These compounds also have a role in protein synthesis of hemoglobin, synthesis of nitric oxide, oxidation of xanthine, P450 enzymes, and fatty acid metabolism (Ma et al., 2008; Belinda, 2014). Some enzymes which are requiring flavin can be seen in Table 1 below.

Table 1. Enzymes with Flavin as Coenzyme Enzyme Coenzyme Function Dihydrolipoyl dehydrogenase FAD Energy metabolism Fatty acyl-CoA dehydrogenase FAD Fatty acid oxidation Succinate dehydrogenase FAD Krebs cycle NADH dehydrogenase FMN Respiratory chain Xanthine dehydrogenase FAD Purine catabolism Glutathione reductase FAD Reduction GSSG to GSH Methylene-tetrahydrofolate reductase FAD 5-Methyl-ethyl- tetrahydrofolate Pyridoxine phosphate oxidase FMN Vitamin B6 metabolism Monoamine oxidase FAD Neurotransmitters metabolism Source: Thorne Research (2008)

Riboflavin also have antioxidant properties. Riboflavin may increase endogenous antioxidant through its role as a coenzyme in glutathione redox cycle. Glutathione reductase enzymes needs FAD as coenzyme and NADPH to convert the oxidized glutathione (GSSG) to the reduced glutathione (GSH). Hydrogen ion from NADPH, transport by FAD to GSSG, so that GSSG could be converted to GSH. GSH is an endogenous antioxidant which can inactivate free radicals such as hydrogen peroxide (H2O2). GSH role as an antioxidant is mediated by the glutathione peroxidase enzyme (GPx). GPx transport hydrogen ion from GSH to H2O2, in order to convert H2O2 to H2O (Anshoori and Saedisomeolia, 2014). Therefore collaboration of glutathione reductase, FAD, NADPH, GSH, and glutathione peroxidase can reduce free radicals level in the body, as seen in Figure 2 below.

Figure 2. Collaboration of Glutathione Reductase, FAD, NADPH, GSH, and Glutahione Peroxidase to Inactivate Free Radicals (Anshoori and Saedisomeolia, 2014).

Coenzyme flavin has a role in the cycle of folate and methionine. In the folate cycle, flavin acts as a coenzyme for the methyltetrahydrofolate reductase (MTHFR), while in the methionine cycle flavin works together with methionine synthase reductase (MTRR) (Kennedy, 2015). Both MTHFR and MTRR enzymes play an important role in homocysteine metabolism (García-Minguillán et al., 2014). In the folate cycle,

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along with FAD, MTHFR enzyme catalyzes the conversion of 5,10-dimethyltetrahydrofolate (CH2-THF) to 5-methyltetrahydrofolate (CH3-THF) (Shane, 2008; Kennedy, 2015). This reaction provides a methyl group, which is important in remethylation process of homocysteine to form methionine (García- Minguillán et al., 2014; Kennedy, 2015), as can be seen in Figure 3 below.

Figure 3. FAD Role in Folate and Methionine Cycle (Shane, 2008).

Riboflavin Intake Riboflavin intake is very important for all ages, especially infants and children, to help the body produces energy (Agostoni et al., 2013). Recommended Daily Intake (RDI) for adult men and women is 1,3 and 1,1 mg/day, respectively (Institute of Medicine, 1998).

Table 2. Riboflavin recommended daily intake (RDI) Criteria Age (Year) Intake Recomendation Children 1-3 0,5 mg/day 4-8 0,6 mg/ day Boys 9-13 0,9 mg/ day 14-18 1,3 mg/ day Girls 9-13 0,9 mg/ day 14-18 1,0 mg/ day Men 19-70 1,3 mg/ day Women 19-70 1,1 mg/ day Elderly Men >70 1,3 mg/ day Elderly Women >70 1,1 mg/ day Pregnancy 14-50 1,4 mg/ day Lactation 14-50 1,6 mg/ day Source: Institute of Medicine (1998)

Riboflavin Deficiency Deficiency can result from inadequate intake or due to malabsorption (Kennedy, 2015). In Cambodia, Whitfield et al (2015) found the riboflavin and thiamin deficiencies in women of childbearing age in both rural and urban areas (Whitfield et al., 2015). Although riboflavin plays an important role in metabolism, and riboflavin deficiency may occur in many countries, but this deficiency is not lethal because its efficient used in the body (Murray et al., 2009). Clinical signs and symptoms appear after inadequate intake of riboflavin about 3-8 months (Thorne Research, 2008). Riboflavin deficiency will affect vision, and cause symptoms such as weaknesses, pain,

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itching or burning eyes, or cheilosis. Moreover, it can cause cancer, nerve degeneration, peripheral neuropathy, dermatitis, increased risk of cardiovascular disease, and anemia (Belinda, 2014; Kennedy, 2015). If deficiency affects the brain, there will be symptoms of personality changes, fatigue, and brain dysfunction (Kennedy, 2015). Yazdanpanah et al. (2008) found that low intake of riboflavin is associated with high risk of fractures in postmenopausal women. Some disorders caused by riboflavin deficiency include hyperhomocysteinemia, cataracts, anemia, and migraine.

Hyperhomocysteinemia Amino acid homocysteine may be toxic if the levels are excessive in the body (Kennedy, 2015). Level of homocysteine is influenced by genetic and nutrition factors. High level of homocysteine in circulation is associated with an increased risk of cardiovascular disease (Ganji and Kafai, 2004; McNulty et al., 2006). High level of homocysteine is also associated with lower level of riboflavin in individuals with polymorphisms of MTHFR enzyme has reduced enzyme activity (García-Minguillán et al., 2014). Riboflavin supplementation to individuals with MTHFR polymorphisms results in lower homocysteine levels significantly. One of efforts to lower homocysteine level is riboflavin and folic acid fortification in food intake, so the risk of heart disease and stroke will decline (McNulty et al., 2006). Ganji and Kafai (2004) found that intake of milk, yoghurt, cereals, and vegetables (all of which rich in riboflavin and folic acid) can significantly reduce the level of homocysteine in circulation.

Cataracts Cataract is one of the eye-lens disorders that mainly caused by oxidative stress process (Chiu and Taylor, 2007). Riboflavin has antioxidant effect. FAD as an active form of riboflavin, works as a coenzyme for glutathione reductase. This enzyme is FAD-dependent, which catalyzes the conversion of GSSG into GSH (Anshoori and Saedisomeolia, 2014). GSH is an important endogenous antioxidant, and works to prevent damage to lens of the eye due to ROS. To perform its function as an antioxidant, GSH must be converted from GSSG continuously, and this process requires FAD (Mazzotta et al., 2014).

Riboflavin deficiency is associated with the incidence of cataracts due to inadequate coenzyme FAD required for the activity of the glutathione reductase enzyme (Mazzotta et al., 2014). The low level of this vitamin is a risk factor for the vulnerability of the eyes from oxidative stress that may lead to the formation of cataracts (Bhat et al., 1993). High intake of riboflavin or riboflavin supplementation can reduce the risk of cataracts and cope with or recover the early stages of a cataract (Bhat et al., 1993; Chiu and Taylor, 2007).

Anemia Anemia can be defined as a reduction of hemoglobin (Hb) concentration in red blood cells (Northrop- Clewes and Thurnham, 2013), resulting in the inadequate oxygen supply to the cells of the body. Riboflavin has an important role in the formation of red blood cells (erythropoiesis). Given high-dose riboflavin to the anemic patients has proved to overcome anemia quickly (Belinda, 2014). A study has shown improvements or positive effects of riboflavin and retinol supplementation to pregnant women have anemia and iron deficiency (Ma et al., 2008). Young women in the United Kingdom who are deficient in riboflavin showed the improvement or increased both hemoglobin status and significant riboflavin levels due to riboflavin supplementation. Increased riboflavin level may lead to an increase in the number of red blood cells in circulation and thus the concentration of hemoglobin (Powers et al., 2011).

Migraine Migraine is a chronic disorder that is often found, could be due to dysfunction of energy metabolism in the mitochondria (Schoenen et al., 1997). Metabolism in mitochondria involves various important molecules, such as FAD and FMN in terms of electron transfer in oxidation-reduction reactions. Riboflavin as a precursor of FAD and FMN are needed for these types of reactions. Administration of riboflavin as a prophylactic dose of 400 mg/day can reduce the severity and duration of migraine attacks (Schoenen et al., 1997). Riboflavin may increase the activity of complex I and II electron transfer in mitochondria in terms of energy production. Research by Nambiar et al. (2011) also found that the administration of riboflavin 100 mg/day as prophylaxis can reduce the frequency, duration, and severity of migraine attacks. Thus, riboflavin can be used as a prophylactic for patients suffered from migraine.

Conclusion Riboflavin (aka vitamin B2) has an important role in the metabolism, through FMN and FAD as it active form. Both of this flavin are essential for the enzymes activity in metabolism such as energy produces metabolism, glutation cycle, and folate-methionine cycle. The daily intake of riboflavin is low, but it is efficiently used for the metabolism reactions. Riboflavin deficiency may occur due to inadequate intake

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or malabsorption. There will be variety of clinical manifestation and disorders associated with the deficiency. Therefore, high intake or supplementation of riboflavin is suggested to avoid deleterious impacts of riboflavin deficiency.

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