Vitamins 58.2. Vitamins soluble in water 1 58.2.1. Thiamine HO S N CH3 5 1 2 1 2 Thiamine, Vitamin B 4 3 5 1 N 4 N + H3C 3-(4-Amino-2-methyl-5-pyrimidinyl)methyl]-5-(2-hydroxyethyl)-4- NH2 methylothiazolium chloride A molecule of thiamine consists of a substituted thiazol ring attached to a substituted pyrimidine ring by a methylene bridge. The substituents in positions 2 and 4 in the pyrimidine ring and 4 and 5 in the thiazol ring are very important for the activity of thiamine. The replacement of the methyl group by an ethyl group in the position 2 of pyrimidine does not have much influence on activity, whereas the C2 butyl derivative acts antagonistically. 2 The acetylation of the amino group in position 4 in the pyrimidine ring HO S N CH3 5 1 2 1 2 weakens4 3 5action, while the replacement of the amino group by a N 4 N + Hhydroxyl3C group creates oxythiamine with antagonistic action NH2 (antivitamin B1). The hydroxyethyl group in the position 5 of the thiazol ring is very important for the activity of thiamine. Its elimination or replacement by another substituent results in the disappearance of activity. The replacement of the hydrogen atom in the position 2 of the thiazol ring by a sulfur atom (Thiothiamine, vitamin S-B1) does not influence activity. In therapy thiamine analogues with an ‘open’ thiazol ring (acetiamine, benfotiamine, fursultiamine, prosultiamine) which act similarly to thiamine, are also used. 3 Some of their properties are even better than those of thiamine, for example lower toxicity, higher stability, prolonged action and better absorption from the GT. In the body, they are a source of vitamin B1 and are used as analgetic agents. HO S N CH S 3 CHO N CH O S 3 N N N N O H3C NH NH 2 CH3 2 PO3H2 Thiothiamine, Vitamin S-B1, SULBONE Benfotiamine, BIOTAMIN O R CHO N CH O H C S 3 S CHO N CH 3 S 3 N N H C O N N 3 HO CH3 NH2 CH3 NH2 Acetiamine, THIANEURON Prosultiamine, R = -CH2-CH2-CH3; DITIAMINA O Fursultiamine, Diavitan R = 4 The daily requirement of witamin B1 depends on a person’s age (low in children, 0.3–1.0 mg/24 h) and sex (slightly higher in men – 1.2–1.5 mg/24 h – than in women – approx. 1.1 mg/24 h). It also depends on nutrition (greater in the case of a diet rich in carbohydrates) and it is higher in pregnant women in (1.5 mg/24 h) and during lactation (1.6 mg/24 h). A deficiency of vitamin B1 increases the concentration of pyruvic acid in the tissues and leads to the beriberi disease, which is characterized by the disturbance of the nervous system (peripheral sensomotor polyneuropathy and encephalopathy) with cardiac insufficiency, edemas and psychic disturbances. In vitamin B1 deficiency medicinal products containing synthetic thiamine are used. They are administered mostly orally in daily doses of 3–9 mg and in serious cases intramuscularly, 10–20 mg once or twice daily. 5 Thiamine is needed for the metabolism of carbohydrates. It is also vital for the supply of energy necessary for the function of the nervous system, the cardiac muscle and skeletal muscles. In the body thiamine is phosphorylated to active thiamine pirophosphate (TPP) and thiamine triphosphate (TTP). TPP is a coenzyme for pyruvate decarboxylase, 2-ketoglutarate dehydrogenase and transketolase. TPP participates in the transfer of the aldehyde group in the pentose phosphate cycle. After oral administration the transport of vitamin B1 depends on the dose. At concentrations < 2 mol active transport by a carrier occurs, while at concentrations >2 mol passive diffusion is observed. The absorption of thiamine is the greatest in the duodenum and the upper and middle sections of the small intestine. Thiamine is not absorbed from the stomach or the distal fragment of the small intestine. Thiamine is absorbed after its phosphorylation in the epithelium. 6 Vitamin B1, after oral administration, is not absorbed completely. Its maximal absorption is 8–15 mg/24 h. The absorption of thiamine is greater when it is administered in divided doses during meals. An excess of administered thiamine is eliminated in urine. The half-time of the -phase elimination of thiamine is 1.0 h. Thiamine is eliminated unchanged and as metabolites – thiaminecarboxylic acid, pyramine and others which have not been identified yet. 7 The stability of thiamine The greatest stability is demonstrated by solutions of thiamine at pH 2, which can be stored for 6 months at 37 0C, without any effect on activity. Rapid degradation is observed at pH > 5, especially in the presence of atmospheric oxygen. The aqueous solutions at pH < 5 can be sterilized for 1 h at 100 0C. Chlorobutanol at a concentration of 0.5% is recommended for the stabilization of thiamine chloride in some pharmacopeal monographs. 8 + HO N CH H /H2O S 3 THIAMINE + N HO N H3C basic and NH2 4-Amino-5-hydroxy- neutral 5-(2-Hydroxyethyl)- environment 4-methylthiazol methyl-2-methyl- pyrimidin H CHO N CH3 N N CH3 SH H C N N 3 N HO N CH3 NH2 NH2 HO H2S thiol form of thiamine Diazepine derivative O2 S The acidic hydrolysis of thiamine splits the bond N-C between the nitrogen atom of the thiazol ring and the carbon atom of the methylene group. In an alkaline environment a thiol form of thiamine is formed, which is very reactive and produces a benzodiazepine derivative after the elimination of hydrogen sulfide. In the presence of oxygen hydrogen sulfide is oxidized to elemental sulfur insoluble in water. 9 58.2.2. Riboflavin CH2OH HO C H HO C H Riboflavin, VITAMINUM B2 HO C H 7,8-Dimethyl-10-(D-1-ribityl)-isoalloxazine CH2 H3C N N O NH H3C N O The term riboflavin is made up of the names of its two components - ribitole and flavin. The word flavin is derived from flavus, the Latin word for yellow. The following relationship between the chemical structure and action of riboflavin is observed: 10 For biological activity the presence of two methyl groups in positions 7 and 8 is necessary; when they are absent or their position is changed action disappears. The replacement of the methyl groups by a chlorine atom produces compounds with antagonistic action. CH2OH A changeHO C ofH ribitol’s configuration from D to L causes action to disappear.HO C H HO C H The imineCH2 group (-NH-) in position 3 is necessary for activity. When H3theC hydrogenN N atomO is replaced by a methyl group in the imino group NH H3actionC disappears.N O Riboflavin is a precursor of riboflavine-5'-phosphate (flavin mononucleotide; FMN) and flavin adenine dinucleotide (FAD). The names nucleotide and dinucleotide are not correct because ribitol is not a pentose and is not fused to flavin with a glycoside bond but these terms are accepted because of their widespread use. 11 . R 2H H3C N N O NH H3C N . O H R FAD = Oxidized form H H3C N N O NH H3C N H R O H H C N N O 3 FADH2 . N H Reduced form H3C N . O H Free radical (FADH) Enzymes containing riboflavin are called flavoproteins. They participate in oxidation-reduction reactions. Flavin coenzymes can exist in any of 3 different red-ox states: oxidised flavin (FAD), semiquinone (free adical) (FADH) or reduced (FADH2) flavin. FAD is converted to semiquinone by one-electron transfer. A second one-electron transfer converts semiquinone to the completely reduced dihydroflavin. 12 The presence of the three red-ox forms makes it possible for flavin coenzymes to participate in one- or two-electron transfers. Because of that flavoproteins calalyse many biochemical reactions together with various acceptors and donors of electrons such as: transmitters of 2 electrons (NAD+, NADP+, DT-diaphorase) one- and two-electron transmitters e.g. quinones one-electron transmitters (cytochrome proteins). 13 Examples of flavoproteins include: dehydrogenases (acylo-CoA dehydrogenase, glutathione reductase, aldehyde dehydrogenase, mitochondrial glycerol-3- phosphate dehydrogenase, succinate dehydrogenase of the citric acid cycle, NADH dehydrogenase of the respiratory chain in mitochondria, dihydrolipoyl dehydrogenase) monooxygenases (lactate oxygenase) dioxygenases oxidases (glucose oxidase, -amino acid oxidase, xanthine oxidase). 14 Vitamin B2 exists in many foods. Sources of this vitamin are liver, white meat, eggs, milk and fresh vegetables. The daily requirement of vitamin B2 is 1–3 mg. A deficiency of vitamin B2 inhibits growth in children. Symptoms of hypovitaminosis are inflammation of oral mucosa and the tongue, angular cheilitis (the cracking of mouth corners), seborrheic dermatitis. Vitamin B2 is used in the treatment of cataract and inflammations of mucous membranes as well as in convalescence after devastating diseases: orally in doses of 3–9 mg daily in severe deficits of vitamin B2, 5–10 mg daily (i.m.). 15 The stability of riboflavin When stored in the solid state and protected from light riboflavin is stable, even at an increased temperature. Under the influence of light and oxygen it is decomposed to lumiflavin. This reaction occurs easily in an alkaline environment under the influence of visible radiation. In neutral and acidic solutions, under the influence of light, lumichrome consisting of various lumiflavins is formed. R CH3 N N O H3C N N O H3C hv/OH- Figure 58.6. NH N H H3C N H3C N O O The decomposition of riboflavin Riboflavin Lumiflavin under the influence of light. hv/H+ H H3C N N O + Lumiflavin NH H3C N O Lumichrom 16 58.2.3.