REACCIONES DE CARBOHIDRATOS Reactions of monosaccharides • Carbonyl reactions: • • • • • • • Osazone formation Cyanohydrin reaction Reduction Oxidation Action of base Action of acid Ring chain tautomerism • Alcohol reactions: • Glycoside formation • Ether formation • Ester formation Mutarrotación Glucosa (Dextrosa); Dextrosa); Dextro--rotatoria Dextro rotatoria.. 3 Mutorrotación D-glucosa glucosa:: Cicla de 2 formas → Estructuras Furanosas o Piranosas. Piranosas. D-Ribose (C (C--5): Estructuras Furanosa o Piranosa Conformaciones Silla o Bote de una estructura Piranosa. Piranosa. 2 possible chair conformations of β-D-glucose Epimerización H de C2 Removido → Ión Enolate. Enolate. Reprotonación:: Δ Estereoquímica de C2. Reprotonación 8 Rearreglo Enodiol • En medio Básico: Básico: La posición del C=O puede Cambiar. Cambiar. • Los CHOs deben permanecer en slns ácidas o neutras para preservar su identidad identidad.. 9 Reducción • Puede ser: – Cataliticamente: NaBH4 (H2/Ni,Pt,Pd) – Enzimaticamente. o (H2 + Catalizador) • Productos : Polioles o Alco- Azucares (Alditoles) • En general; se nombran adicionando la terminación itol a la raíz del nombre del azúcar. – Glucosa → Sorbitol (Glucitol) – Manosa → Manitol – Fructosa → Manitol + Sorbitol – Gliceraldehido → Glicerol ALDITOLES Rxs Oxidación Aldosas oxidadas → 3 9pos de ácidos – Ácidos Aldónicos Aldónicos:: Aldehído → Carbonilo (Glucosa → ácido glucónico glucónico)) – Ácidos Uronicos Uronicos:: Aldehído permanece intacto ⇒ Se oxida el primer alcohol del otro extremo de la molécula.. molécula • Glucosa → Ácido Glucuronico • Galactosa → Ácido Galacturonico – Ácidos Sacáricos (ácidos glicáricos glicáricos)) – Oxidación en ambos extremos de un monosacárido monosacárido)) • Glucosa → Ácido Sacárico • Galactosa → Ácido Múcico • Manosa → Ácido Manárico Oxidación con Bromo • El agua de bromo oxida el aldehído formando un Ácido Aldónico. Chapter 23 14 Oxidación con Ácido Nítrico • Oxida el aldehído y el alcohol terminal. • Forma ácido aldárico. 15 Oxidación con Reactivo de Tollens. Tollens. • Reactivo de Tollens Rx con el aldehído, pero el álcali del medio promueve el rearreglo enodiol, así Rxnan también las cetosas. plata. • Azucares reductores forman un espejo de plata Chapter 23 16 Formas Oxidadas DD-Glucosa Azucares no Reductores • Glicósidos son acetales, estables a álcalis, no dan fácilmente reacciones de oxidación. • Disacáridos y Polisacáridos son también acetales y azucares no reductores. 18 Formation of Glycosides • React the sugar with alcohol in acid. • Since the open chain sugar is in equilibrium with its α- and β-hemiacetal, both anomers of the acetal are formed. • Aglycone is the term used for the group bonded to the anomeric carbon. => Chapter 23 19 Ether Formation • Sugars are difficult to recrystallize from water because of their high solubility. • Convert all -OH groups to -OR, using a modified Williamson synthesis synthesis, after converting sugar to acetal, stable in base. 20 Ester Formation Acetic anhydride with pyridine catalyst converts all the oxygens to acetate esters. 21 Formation of osazones • Identification of sugars. sugars. Phenylhydrazine.. • Rx Monosaccharide + Phenylhydrazine • A crystalline compound with a sharp melting point will be obtained obtained.. • D-fructose and D-mannose Osazone as D-glucose glucose.. • Seldom used for identification identification;; we now use HPLC or mass spectrometry. spectrometry. Osazone Formation Both C1 and C2 react with phenylhydrazine. 23 Ruff Degradation Aldose chain is shortened by oxidizing the aldehyde to -COOH COOH,, then decarboxylation decarboxylation.. 25 Ruff degradation Series of reactions that removes the reducing carbon ( C=O ) from a sugar and decreases the number of chiral centers by one; one; used to relate configuration. configuration. CHO H OH H OH CH 2 O H Br 2 H 2O CO 2 H H OH H OH CH 2 O H Ca 2+ CHO H OH CH 2 O H D-(+)-glyceraldehyde H 2O 2 Fe 3 + CO 2 H OH H OH CH 2 O H Cyanohydrin formation • Rxn Aldose + HCN • Used to increase the chain length of monosaccharides • Results in a Cyanohydrin hydrolyzed Acid Reduced Aldehyde.. Aldehyde • Known as the Fischer Fischer--Kiliani synthesis • Can prepare all monosaccharides from Dglyceraldehyde Kiliani--Fischer synthesis. Kiliani Series of reactions that extends the carbon chain in a CHO by one carbon and one chiral center. CHO HCN H OH CH2OH C N C N COOH COOH H H OH HO +,H O H 2 H OH HO H + H OH H OH H OH H OH CH2OH CH2OH CH2OH CH2OH CHO H OH H OH CH2OH diastereomers separable Na(Hg) O C H OH H OH H2C O lactone -H2O Kiliani--Fischer Synthesis Kiliani • This process lengthens the aldose chain. • A mixture of C2 epimers is formed. 29 Periodic Acid Cleavage • Periodic acid cleaves vicinal diols to give two carbonyl compounds. • Separation and identification of the products determine the size of the ring. 30 Base-catalyzed isomerisation of monosaccharides H O H HO H H OH H OH OH CH2OH Glucose Enolization H OH OH HO H H OH H OH CH2OH Ca(OH)2 H2O H O H O H HO H H OH H OH OH CH2OH HO HO H H H H OH OH CH2OH + Recovered glucose, 67% CH2OH + Mannose, 2% O HO H H OH H OH CH2OH Fructose, 30% Glycosides Glycoside formation A glycoside hydroxyl CH2OH OH H H OH H HO OH H OH α-D-Glucopyranose (a hemiacetal) CH2OH OO H H OH H HO H H OH HCl, CH3OH CH3 + CH2OH OH H H OH H HO O CH3 H OH Methyl β-D-glucopyranoside (a glycoside (an acetal) + H CH2OH OH H H OH H + HO O H H OH H CH3 CH2OH O H + H OH H C H HO H OH O HO CH3 CH2OH CH2OH + OH OO H H H H H+ OH H OH H + HO HO H O H OH H OH O OH CH2OH Another example of a glycoside: OH H3CO OH H O O H H Doxorubicin (an anticancer drug) H CH3 HO O H NH2 1. Alkylation at the glycoside position. H CH3 2. Complete alkylation OH OH HO HH H H OH OH OH OCH3 H3CO OH (CH3O)2SO2 HH H H OCH3 NaOH OCH3 H3CO + OCH3 H3CO OOCH3 HH H H H OCH3 OCH3 3. Complete acylation OH OAc AcO O OH HO H Acetic anhydride HH H HH H H OH H OAc Py OH OH OAc AcO + OAc AcO OOAc HH H H H OAc OAc N y C Glycoside Formation OH OH O OH H H CH3OH O CH 3 H H OH H OH HO H H H+ H HO H H OH •Stable in water •Does not undergo mutorotation OH Acetal Hemi-acetal O OH O O OH H NH H OH H HO OH NH NH O OH O H OH H H N O H H HO H H OH N-glycoside Glucose oxidase • Glucose oxidase converts glucose to gluconic acid and hydrogen peroxide • When the reaction is performed in the presence of peroxidase and O-Dianisidine a yellow color is formed • This forms the basis for the measurement of urinary and blood glucose • Testape Testape,, Clinistix, Clinistix, Diastix (urinary glucose) • Dextrostix (venous glucose) Glycoconjugates.. Glycoconjugates Formation of spans and tweens OH CH2 O SORBITOL CH OH OH THF compound OH 1,4-SORBITAN O O CH2 O O C CH2 (O-C2H4) O C R R O CH (O-C2H4)nOH CH OH HO (C2H4-O)x OH (O-C2H4)x OH OH SPANS (form W/O emulsions) TWEENS (form O/W emulsions) Action of strong acids on monosaccharides • Monosaccharides are normally stable to dilute acids, but are dehydrated by strong acids • D-ribose when heated with concentrated HCl yields furfural (commercial route for the production of THF (tetrahydrofuran tetrahydrofuran)) • D-glucose under the same conditions yields 5hydroxymethyl furfural Action of base on sugars • Sugars are weak acids and can form salts at high pH. pH. • A 1,2-enediol salt is formed as the result. result. • This allows the interconversion of D-mannose, D-fructose and D-glucose glucose.. • The reaction is known as the Lobry de Bruyn Bruyn-Alberta von Eckenstein Reaction. Reaction. Action of base on sugars • Enediols obtained by the action of base are quite susceptible to oxidation when heated in the presence of an oxidizing agent • Copper sulfate is frequently used as the oxidizing agent and a red preciptate of Cu2O is obtained • Sugars which give this reaction are known as reducing sugars • Fehling’s solution : KOH or NaOH and CuSO4 • Benedict’s solution: Na2CO3 and CuSO4 • Clinitest tablets are used to detect urinary glucose in diabetics Glucose measurement methods • Most methods are enzymatic methods – 3 enzyme systems are currently used to measure glucose: • Glucose oxidase • Glucose dehydrogenase • Hexokinase • These reactions produce either a product that can be measured photometrically or an electrical current that is proportional to the initial glucose concentration Glucose dehydrogenase methods α -D-glucose mutarotase β -D-glucose + NAD ΜΤΤ + NADH β -D-glucose glucose dehydrogenase diaphorase D-gluconolactone + NADH MTTH (blue color) +NAD glucose dehydrogenase Gluconolactone + PQQH glucose + pyrroloquinoline quinone (PQQ) PQQH 2 + 2[Fe(CN) 6 ] -3 2[Fe(CN) 6 ] -4 PQQ + 2[Fe(CN) 2[Fe(CN) 6 ] -3 + 2e - 6] -4 + 2H + Glucose oxidase methods: colorimetric method glucose oxidase β -D-glucose + O 2 D-gluconolactone + H 2O 2 D-gluconolactone + H 2O gluconic acid H 2O 2 + chromogenic oxygen acceptor (ortho-dianisidine, 4 aminophenazone, ortho-tolidine) peroxidase colored chromogen + H 2O Glucose oxidase methods: electronic sensing method β -D-glucose + 2[Fe(CN) 6 ]-3 glucose oxidase + H 2O 2[Fe(CN) 6]-4 D-gluconic acid + 2[Fe(CN) 6]-4 + 2H + 2[Fe(CN) 6]-3 + 2e - glucose oxidase β -D-glucose + O 2 H 2O 2 D-gluconolactone + H 2O 2 2H + + O 2 + 2e - A blood test for glucose levels Dye H C H HO O2 Colored Dye OH H2O2 O C OH H H Glucose Oxidase HO O OH H H OH H OH H OH H OH CH2OH D-Glucose CH2OH D-Gluconic Acid (An Aldonic Acid) An example of an enzymatic oligosaccharide synthesis used for the production of a clinical candidate OH OH O HO OH HO NHAc HO O CMP-Sialic acid OR O OH α2,3-Sialyltransferase HO AcHN CO2– OH O OH OH O O NHAc HO O OH HO N-Acetyllactosamine (from chemical synthesis) GDP-Fucose α 1,3-Fucosyltransferase OH OH HO HO AcHN CO2– OH O HO H3C OH OH O NHAc O O O O OH OH O OH OR Sialyl Lewis x: An anti-inflammatory agent developed by Cytel O OH OR Present limitations of enzymatic oligosaccharide synthesis: 1. The nucleotide sugars are expensive 2. Not all enzymes are available in recombinant form 3. The enzymes do not accept many unnatural sugars, thus modified oligosaccharides cannot be readily made 4. Most glycosyltransferases are inhibited by high concentrations of the nucleotide product; thus, large scale synthesis is difficult Many groups are seeking solutions to these problems so stay tuned!…. The present challenge: Integrating oligosaccharide synthesis with protein chemistry to make glycoproteins OH OH H3C OH OH O OH HO OH NHAc CO2– O O HO O O O O O AcHN OH OH HO OH OH HO HO OH CO2– O O O HO O O AcHN AcHN O HO HO N H CH3(H) O Chemical synthesis of carbohydrates is important for… • Studying their biological activities • Development of new therapeutic compounds Examples of approved carbohydrate drugs: Substance Acarbose AMVISC Hyalgan Lovenox Miglitol ORTHOVISC Relenza Tamiflu SOLARASE Topamax Voglibose Indication Diabetes Opthalamic surgery Osteoarthritis Cardiovascular disease Diabetes Osteoarthritis Influenza Influenza Actinic keratosis Epilepsy Diabetes Company Bayer AG Anika Therapeutics FIDIA/Sanofi Aventis Bayer AG Anika Therapeutics Glaxo Smithkline Roche Hyal Pharmaceuticals J&J Takedo/Abbott Modificación del Almidón Almidón Modificación Física: Física: Modificación Química: Pregelatinización Enlaces cruzados ((cross cross linked linked)) Dextrinización Esterificados Hidroxialkilados Fosfato monoesterificados Xantatos Dextrinización Jarabe de glucosa Modificación Biotecnológica: Dextrinización Jarabe de glucosa Jarabe de glucosaglucosafructosa