Lecture №26

1. Lecture №26 Vitamins as medicines: general characteristic, methods of receiving, classification, analysis, storage, precondition of the usage in medical practice. Vitamins of the aliphatic and alicyclic rows. prepared ass.MedvidI.I.

2. Vitamins - are organic compounds of different chemical structures, which in small amounts required for the normal organism living. They are a part of enzyme systems, which are biological catalysts of the chemical reactions in living cells and are involved in metabolism. The human and animals can not synthesize vitamins or synthesize them in insufficient quantities (nicotinic acid) and therefore must obtain them from food. In some cases vitamins are formed in the tissues as a result of chemical transformation of substances that are their precursors (provitamins). In 1912 y. polish scientist K. Funkproposed term “vitamins” that means “amines necessary for life”.

3. Classification of vitamins By the solubility: fat soluble (A, D, E, F, K ) Water-soluble (group В, С, synthetic analogs of К, РР, Р) By letters: by the disease that occurs at the insufficient amount of this vitamin(С – scurvy (antiscorbutic), А –antixerophthalmic, В – beri-beri, Е – one that ensuresprocreation. additional figures at the expanding of the group of vitamins(В1 – В15, Вс) Chemical classification.

4. Chemical classification of vitamins Vitamins of the aliphatic row: а) Derivatives of the unsaturated polyoxy-- lactones (ascorbic acid (vitamin C)); b) Derivatives of gluconic acidesters (pangamic acid (vitamin В15)); c) Derivatives of- aminoacids (pantothenic acid (vitaminВ3)). Vitamins of the alicyclic row: а) cyclohexyl isoprenoids(retinols (vitamins of A group)); b) Cyclohexanolethylenehydrindanoic (calciferols (vitamins of groupD)). Vitamins of the aromatic row (naphthoquinone derivatives): c) natural vitamins of К group(phylloquinone– К1, menaquinone – К2); d) synthetic analogues of vitamins ofК group (menadione, vikasol).

5. Vitamins of the heterocyclic row: 1) Chromane derivatives а) tocopherols (vitamins of Е group); b) bioflavonoids (vitamins of Р group). 2) Pyridine derivatives а) nicotinic acid and its amide (vitamine of РР group); b) oxymethylpyridine vitamines (vitamins of В6 group). 3) pyrimidine derivatives (thiamin (vitamin В1). 4) Pterine derivatives (folic acid (vitamin Вс). 5) Isoalloxazine derivatives or flavin vitamins (riboflavin(vitamin В2)) 6) Corrine derivatives – cobalamins (vitamins of В12 group).

6. Obtaining of vitamins Synthetic and semi-synthetic methods (С, А, Е, D, В1, В2, РР and others). From plant and animal raw materials , microorganisms (С - fruit-bloom, Р –waste of the tea industry, D –natural sterines, А –fish oil, Е –vegetable fats, В12 –product of microbiological synthesis of antibiotics, effluents). Properties White crystals or crystalline powder (rutin - a green- yellow, folic acid - a yellow or yellow-orange, riboflavin - yellow-orange, cyanocobalamin - dark red). Tocopherol acetate-light yellow, transparent, dense, oily liquid with slight odor. Vitamins are unstable under the action of light, air and heat (some of them).

7. Identification of vitamins There are not general reactions, except nitrogen determination by the Kjeldahl method. Often the group reaction(on the pyridine cycle etc.) More often - reactions that depend on the individual characteristics of the chemical structure. According to theSPhU requirements – 10preparations (D, С, РР, nicotinamide В6, В2, В1 (hydrobromide), В1 (hydrochloride), Вс, В12: ІR-, UV- spectroscopy, chromatography.

8. Assay of vitamins Biological methods - determining of the biological activity of vitamins. Studies conducted on rats, pigeons, guinea pigs, which are moved on a diet (exclude the study vitamin from the food). Then determines which of the study vitamin can cure or save the animal from avitaminosis. Parallel conduct similar studies with the standard drug. Activity in IU (international unit) - the notional amount of the standard drug in mg or mcg (). For one unit is considered to be the minimum amount of vitamin that cures or prevents the animal from avitaminosis. 1 IU is different in different vitamins (1 IU of vitamin A corresponds to 0,344 g of tocopherol acetate, vitamin D - 0,25 g of ergocalciferol). Currently, the most frequently for quantification use physical, physico-chemical and chemical methods.

9. USAGE Parts of biocatalysts (enzymes) that cause different functions in the body metabolism. Hypo-and avitaminosis of the some vitamin. In addition, vitamins of В group –CNS diseases, heart diseases; В12 – anemia; РР – a violation of the peripheral circulation; Е – for the pregnancy keeping; D – rachitis; А – skin and eye disease. Multivitamins–support during the cold season, reduce the risk of heart diseases and others. Side effects - allergic reactions (Multi-tabs, Kinder-biovital), dyspepticdisorders. Supervitaminosis – vitamines А, D, К; essentiale Н (vitamins of Вgroup) – at the expense of long reception.

10. Vitamins of the aliphaticrow Ascorbic acid belongs to the derivatives of polyoxy-γ-lactones ofthe unsaturated carboxylic acids, On the other hand it can be attributed to the vitamins of heterocyclic series as furan derivative. It is widely distributed in nature. Especially rich in it is flora: fresh vegetables, fruits, etc. In industry, ascorbic acid synthesized from D-glucose. To the vitamins ofaliphatic row, derivatives of gluconic acid esters, pangamic acid belongs (vitamin В15). In medical practice is used its calcium salt. Pangamic acid is a part of the rice bran, yeast, blood, liver. To the vitamins of aliphatic row, dertivatives of β-aminoacids, pantothenic acid belongs. Yeast, eggs, liver and egg yolk are rich on pentothenic acid. In medical practice is used its calcium salt.

11. Ascorbic acid (Acidum ascorbicum) Vitamin С (SPhU) (R)-5-[(S)-1,2- Dihydroxyethyl]-3,4-dihydroxy-5H-furan-2-on  -lactone-2,3-dehydro-L-gulonic acid

12. Extraction of ascorbic acid Available in fresh vegetables - cabbage, beets, lettuce, tomatoes, potatoes, berries - strawberries, currents, fruits - lemon, orange. t is also found in milk, eggs, rose hips, fennel, etc.. For the extraction of ascorbic acid from the rose hips produced water extract, which is thickened in vacuum to the thicken of syrup. From the remainder precipitated ballast substances with alcohol and ether, and the filtrate evaporated to the dry state. Residue is clearing by crystallization or chromatographic method. The basic amount of ascorbic acid is now extracted synthetically, based on D-glucose, that can be transformed by the reduction in the D-sorbitol and D-sorbitol by enzymatic oxidation - in the L-sorbose To protect the alcohol group of sorbose, at first it is condensed with two molecules of acetone, and then oxidize diacetone sorbose by КMnO4. Obtained diacetone ketoglutaric acid is saponificated to ketogulonic acid.

14. Properties of ascorbic acid White or almost white crystalline powder of or colorless crystals that change color under the action of air and moisture. Easily soluble in water, soluble in 96% alcohol, practically indissoluble in ether. Melts at about 190 ° C with decomposition. By endiol group ascorbic acid shows both acidic and restorative properties. Acidic nature is cased by the mobility of hydrogen atom in hydroxyl group in position 3; at the alkali titration ascorbic acid behaves like a monobasic acid.

15. Ascorbic acid oxidizes in two stages : • reversible oxidation to dehydroascorbic acid (keto-form); • irreversible process of oxidation, which leads to the formation of furfural:

16. Identification of ascorbic acid • By the physico-chemical constants : UV- andIR- spectroscopy, determination of рН and the specific optical rotation. • Add to the solution of ascorbic acid diluted nitrate acid and silver nitrate solution – silver gray metallic precipitate falls:

17. Unpharmacopoeialreactions: • To 1 ml of obtained solution add 2 drops of iron (III) chlorideR2 solutionand after1 min. 1 drop of of potassium ferricyanideR; blue color appeares: • To 1 ml ofobtained solution add 5-7 drops of0,05 М iodine solution; solution becomes transparent: 3FeCl2 + 2K3[Fe(CN)6] → Fe3[Fe(CN)6]2 + 6KCl.

18. To 1 ml of obtained solution add 1 drop of CuSO4solution and 2-3 drops of 1 % NH4CNS solution; white precipitate forms: • To the aqueous solution of the preparation add NaHCO3andFeSO4, shake and leave to stand; a dark purple color appears which disappears at the addingof H2SO4dil.:

19. at the adding by drops 2,6- dichlorophenolinedophenolate solution to the solution of ascorbic acid blue color disappears:

20. Ascorbic acid is easily oxidized at the interaction with : • Phosphoric-molybdenum acid (the products of blue color form); • methylene blue (discoloration occurs due to formation of leucobase); • Potassium permanganate(discoloration; • Fehling reagent (red color)

21. Assay of ascorbic acid Iodometry, direct titration(SPhU). Indicator – starch. Em .= М. m./2 Alkalimetry, direct titration . Indicator – phenolphthalein). E.m. = М. m.

22. Iodatometry, direct titration in acidic medium in the presence of potassium iodide, indicator – starch. At the point of equivalence excess of potassium iodate causes the blue color of solution. E.m. = М. m./2. • Titration bysodium 2,6-dichlorophenolindophenolate solution (for the determination of ascorbic acid in row material). E.m. = М. m./2 • Other redox methods(Iodochlorometry, cerimetry and others).

23. Ascorbic acid storage In nonmetallicairtight containers, in the dark place. Usage of ascorbic acid Takes part in oxidative-reduction processes, in carbohydrates metabolism and tissue regeneration, the formation of steroid hormones. Daily demand of a healthy person - 80-100 mg. In preventive and therapeutic purposes at the scorbutus (scurvy), bleeding of various etiologies, infectious diseases and poisonings, liver and kidneys diseases. Issue: tablets, dragee by 0,025; 0,05; 0,1; 0,5 g; 5% and 10% solutions for injections. Soluble tablets by 1,0 g.

24. Calcium pangamate(Calcii pangamas), vit. В15, Kalham By the chemical structure pangamic acid is an ester ofD-gluconic and dimethylaminoaceticacids (dimethylglycine). Substance, except for calcium pangamate contains calcium gluconate (25%) and calcium chloride (6%).

25. Extraction of pangamic acid Isolated from rice bran, yeast, blood, liver. Synthesized from D-glucose: Properties. White, sometimes yellowish crystalline powder with characteristic odor. Hygroscopic. Easily soluble in water and hardly soluble in organic solvents.

26. Identification of calcium pangamate By the physico-chemical constants: IR spectroscopy. Substance gives reactions on calcium (4 pharmacopoeial reaction). Residue of gluconic is confirmed by the reaction with iron (III) salts by the formation of light green color. Alkaline hydrolysis. After the heating of substance with solution of sodium hydroxide smell of amines appears. The reaction of formation of colored iron hydroxamate (esteric group):

28. Assay of calcium pengamate Given that the substance except calcium pangamate, also includes calcium gluconate and calcium chloride, quantitatively determine the content of: Nitrogen (3,6 - 4,2%) -acidimetry in non-aqueous medium; Calcium (5,8 - 7,4%) – complexonometry method; Chlorides (not more than 2,2%) - a method of reverse argentometry by Folgard; The amount of carboxyl groups(11 - 15%) - by ion exchange chromatography.

29. STORAGE Inglassairtight containers, in a dry place. Usage of calcium pangamate Activates the transfer of oxygen to the cells of tissues and takes part in the methylation of biosubstrates, improves lipid metabolism. Similar to vitamins preparation. Calcium pangamate is used for the treatment of various forms of atherosclerosis, cirrhosis, alcoholism and other diseases. Issue : tablets by 0,05 g.

30. Calcium pantothenate(Calcii pantothenas), vit. В3 Calcium salt ofD-(+)-α,γ-dioxy-β,β-dimethylbutyryl-N-amide-β'-aminopropionic acid (β-alanine). Properties. White fine crystalline powder without smell. Easily soluble in water, very little soluble in organic solvents.

31. Extraction of calcium pantothenate In large quantities contained in caviar, yeast, liver, egg yolk, rice. Synthetically is extracted by the following scheme:

32. Identification of calcium pantothenate By the physico-chemical constants: the specific rotation from +25 to +28° (5 % aqueous solution) Substance gives reactions on calcium. With the solution of copper (II) sulfate in an alkaline medium substance forms a blue complex (β-alanine):

33. Residue of α,γ -dioxy- β,β –dimethylbutyric aciddetermine after alkaline hydrolysis. . Substance is boiled with sodium hydroxide solution, after cooling acidified by acid chloride solution and added iron (III) chloride - yellow color appears:

34. The reaction of colored iron hydroxamate formation :γ-butyrolactone

35. Assay of calcium pantothenate Determine the content of: Nitrogen (5,7 – 6,0%) – Kjeldahl method; Calcium (8,2 – 8,6%) - complexonometry method. Test on purity Specific impurity. Pantoyl lactone –is determined spectrophotometricaly in the visible part of the spectrum by the reaction of iron hydroxammate formation.

36. Storage In glass airtight containers, in a dry place at the room temperature. Usage of calcium pantothenate Participates in hydrocarbon and fat metabolism, stimulates the formation of corticosteroids and is part of coenzyme A. The daily demand - 10-12 mg. Since it is formed during the vital functions of Escherichia coli, its avitaminosis is not observed. To treat neuralgia, eczema, allergies, polyneuritis, and other diseases related with violation of metabolism and at the inflammatory processes. Issue: tablets by 0,1 g.

37. Vitamins of the alicyclic row Vitamins of the alyciclic row include retinols (vitamins of А group) and calciferols (vitamins ofD group). In the core of the retinol molecule is trimethylcyclohexanoic cycle associated with tetraenol conjugate chain, in the end of which is hydroxyl or aldehyde group. Retinol was obtained from the liver of fish in 1909 year. In 1928 y., Euler found that some plants are substances that have provitamin activity, scilicet are the precursors of vitamins. Provitamins of vitamin A are α-, β-and γ-carotene. . To vitamins of D group, which is called calciferols or antirachitis include derivatives of cyclohexanol ethylene-hydrindane. Is opened few vitamins of D group: D1-D7 similar by the chemical structure, physico-chemical properties and pharmacological action. The practical usage hame the following vitamins: vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). Vitamins D2 and D3 are in egg yolks, caviar, butter, milk. A significant number of them accompanied by retinol in liver and adipose tissue of fish and marine animals. In medicine is used retinol acetate and ergocalciferol.

38. Retinol acetate(Retinoli acetas) vit. А Trans-9,13-Dimethyl-7-( 1,1,5-trimethylcyclohexene5-yl-6)-nonatatraene-7,9,11,13-ol-15 acetate

39. Extraction of retinol Hydrolysis of fish liver by 15 % КОН solution in the atmosphere of inert gas is used for the vitamin A obtaining. Main quantity of vitamin A is obtained synthetically from citral by the following scheme:β-ionone

40. Properties of the retinol acetate White or pale yellow crystals with a weak odor. Extremely unstable under the action of air oxygen and light (easily oxidizes by the atmospheric oxygen, especially at elevated temperatures and light, with the formation of heronic acid). Practically insoluble in water, soluble in 95% alcohol, chloroform, ether and oils. Identification of retinol acetate The reaction with antimony (III) chloride in chloroform environment - blue color appears. The reaction with aluminum chloride - blue color appears. Reactions on the unsaturated bonds (discoloration of iodine, bromine water, reduction of phosphorwolframic acid, AuCl3).

41. Assay of retinol acetate UV-spectrophotometry Photocolorimetry by the reaction withSbCl3 Storige of retinol acetate Due to the fact that the substance is easily oxidizes, store it in sealed ampoules in a stream of nitrogen, which keep from the action of light, at the temperatures no more than +5°. Oil solutions of retinol acetate are stored in chock-filled, well corked cups of dark glasses at the temperatures no more than +10 °С.

42. Usage of retinol acetate In the treatment of avitaminosis, diseases and lesions of the skin, eye diseases. Prescribed as pills, granules, oil solutions orally, intramuscularly and topically. During the treatment is necessary to consider the possibility of hypervitaminosis. Daily dose for a healthy person - 1 mg treatment dose - up to 10 mg (33000 UA), but not more than 30 mg (100 000 UA). 1 IU = 0,344 microgram or 100000 IU = 0,0344 g of retinol acetate. There are two forms of retinol: acetate and palmytate (more stable then acetate). Issue. VitaminА gel. caps. By 2500, 8000, 12000, 50000, 100000 IU. Caps. Aevit – vit. А + Е – caps. №24 (retinol 30000 IU, tocopherol 70 IU).

43. Vit. A are carotenes, which contained in fruits, carrots, red peppers and others.-Carotene in the human body decomposes under the influence of liver enzymes into two molecules of vitamin А, - and -carotenes form only one molecule of vitamin А.βαγ

44. Ergocalciferol (Ergocalciferolum), vit. D2 (SPhU) (5Z,7E,22E)-9,10-Secoergosta-5,7,10(19),22-tetraene-3β-ol Cyclopentanophenantrene General formulaof calciferols

45. Extraction of ergocalciferol VitaminD2by its structure is similar to steroids. This vitamin is obtained by the UV-radiation of ergosterine, which contains inyeast, the uterine horn etc.. Process of ergosterine transformation in ergocalciferol conducts through the formation of lumisterine and tachysterine:

46. Properties of ergocalciferol White or slightly yellowish crystalline powder or white or almost white crystals. Practically insoluble in water, easily soluble in 96%alcohol, soluble in fatty oils. Sensitive to air, heat and light. Solutions in volatile solvents are volatile and must been used immediately after preparation. In the solutions is possible the depending on temperature and time reversible isomerization in pre-ergocalciferol. Activity of the substance is caused by both components. Solution of ergocalciferol in oil - is a transparent oily liquid from light yellow to dark yellow.

47. Identification of ergocalciferol By the physico-chemical constants : IR-spectroscopy. Steroid part of molecule cases Liberman reaction (chloroform solution of the drug at the shakingwith acetic anhydride and sulfate acid becomes red, color transforms to purple, then to blue and finally to green). At the interaction with antimony (III) chloride solution in the presence of acetylchloride formed orange-pink color. This reaction is also used for determination of impurities by TLC and assay by the photocolorimetry method. Sobel-Meyer’s reaction (with glycerin-1 ,3-dichlohydrine) - green coloration. Shaltegger’s reaction (drug is boiled with benzene solutions of aldehydes (vanillin, furfural, anisic, etc.),then add perchlorate acid) - red color appears.

48. Assay fo ergocalciferol Liquid chromatography(SPhU). Photocolorimetry Storage of ergocalciferol Ergocalciferol is stored in an airtight container under nitrogen in a dark place at a temperature from 2 °С to 8 °С. The contents of opened containers should be used immediately. Medical forms of ergocalciferol are stored in chock-filled, well corked cups of dark glasses, because it easily oxidizes by the air oxygen, under the action of light gradually decomposes and forms toxic products.

49. Cholecalciferol (Cholecalciferolum), vit. D3 (5Z,7E)-9,10-Secocholesta-5,7,10(19)-triene-3β-ol At first was obtained from fish oil. Provitamin of vitamin D3is 7-dihydrocholesterol Because of the presence of cholesterol and 7-dehydrocholesterol in the humane skin lipids content it is possible to synthesize vitamin D3under the action of sun radiation or UV-radiation on the surface of the humane body.

50. Usage of calciferols In the medical practice use alcoholic (0,5 %)and oil (0,125 %)solutions of vitamin D2 to prevent and treat rachitis, but also at the bone diseases associated with violations of calcium metabolism. Vitamins ofDgroup areeffective for treatment of all forms of erythematosus and other skin diseases. Daily demand for a healthy man– 1000 IU. VitaminD3 – cholecalciferol. TabletsVideine (2000 іand5000 IU). Vit. А+D3 – aqueoussolution bottles by 10 ml. Fish Oil.