‹#› 1 Selected reactions of organic compounds © Department of Biochemistry (Jiří Dostál) 2010 ‹#› 2 This lecture is an extract of main ideas from chapters 1-14 + 29, Medical Chemistry II •Hemiacetals, acetals, aldimines •Derivatives of acids (esters, anhydrides, amides) •Dehydrogenations of various substrates •Oxygenation, hydroxylation, deoxygenation •Conjugate vs. Redox pair •Transamination of amino acids •Reactions of citric acid cycle ‹#› 3 Mutual reactions of selected compounds Acid Aldehyde Thiol Alcohol Alcohol ester hemiacetal - ether Thiol thioester thiohemiacetal sulfide Amine salta / amideb aldiminec Aldehyde - aldold Acid anhydride aAcid-base reaction. bCondensation (water eliminated). cCalled also Schiff base. dOnly in strongly alkaline environment. See also Med. Chem. II Appendix 1 ‹#› 4 Hemiacetals and acetals Hemiacetals are made by addition of alcohol to carbonyl group. Acetals are made by substitution of the hydroxyl group of hemiacetal by the alkoxyl group (-OR) of alcohol. substitution addition hemiacetal (unstable) cyclic hemiacetal is stable acetal ‹#› 5 Aldimines (Schiff bases) are formed by addition-elimination reaction Schiff base unstable intermediate (aminohemiacetal) C=NH or C=NR is imino group ‹#› 6 Schiff bases in the organism •Non-enzymatic glycation of proteins (Med. Chem. II, p. 40) •Transamination of AA – intermediate with pyridoxal phosphate (see later, lecture Amino acids) •Crosslinks in collagen (Lys ... alLys) (Med. Chem. II, p. 42) •Linkage of retinal to opsin (biochemistry of vision) ‹#› 7 Esters of carboxylic acids are made by condensation reaction with alcohols and water is liberated Esterification in vitro requires acidic catalysis. Enzyme esterifications have different mechanism using acyl-CoA. The reverse reaction is the hydrolysis of ester. acetic acid ethyl acetate ‹#› 8 Distinguish: hydrolysis × hydration substrate + H2O ® product 2 OH substrate + H2O ® + product 1 H product OH H Hydrolysis = decomposition of substrate by the action of water (typical in esters, amides, peptides, glycosides, anhydrides) Hydration = addition of water (to unsaturated substrates) ‹#› 9 Acetylsalicylic acid has acidic group and ester group salicylic acid acetylsalicylic acid ‹#› 10 Selected inorganic acids H2SO4 H3PO4 HNO2 HNO3 ‹#› 11 Esters of sulfuric acid sodium dodecyl sulfate (SDS) is an anionic surfactant sulfuric acid alkyl sulfate (alkyl sulfuric acid) dialkyl sulfate ‹#› 12 Compare: alkyl sulfate ´ alkanesulfonate alkyl sulfate alkanesulfonate bond C-O 4 O atoms around S made by esterification bond C-S 3 O atoms around S made by sulfonation R-OH + HO-SO2-OH ® R-O-SO2-OH + H2O R-H + SO3 ® R-SO3H ‹#› 13 Esters of nitric acid glycerol trinitrate (glyceroli trinitras) isosorbide dinitrate (isosorbidi dinitras) R-OH + HO-NO2 ® R-O-NO2 + H2O both exhibit vasodilatation effect ‹#› 14 Monoesters of phosphoric acid glucose glucose 6-phosphate kinase ‹#› 15 Diesters of phosphoric acid are linkage elements in nucleic acids 3’,5’-phosphodiester bond ‹#› 16 Diesters of phosphoric acid are linkage moieties in phospholipids ‹#› 17 Organophosphates thiophosphoric acid fluorophosphoric acid cyanophosphoric acid methylphosphonic acid methylfluorophosphonic acid sarin Compare: sulfonic ac. (C-S) ´ phosphonic ac. (C-P) ‹#› 18 Compare the structures alkyl sulfate alkanesulfonate alkyl phosphate alkanephosphonate ‹#› 19 Carboxylic acid anhydride + - H2O condensation ‹#› 20 Phosphoric acid anhydride is diphosphoric acid (diphosphate)* + - H2O * Historical term is „pyrophosphate“ (e.g. Harper’s Biochemistry) condensation occurs in ATP, ADP, NAD+, FAD ‹#› 21 Compare: diphosphate ´ bisphosphate diphosphate (anhydride) fructose 1,6-bisphosphate (double ester) ‹#› 22 Hexakisphosphate of inositol is phytic acid ‹#› 23 Mixed anhydride of carboxylic acid and phosphoric acid is acyl phosphate + - H2O acyl phosphate Acyl phosphates are macroergic compounds • 3-phosphoglyceroyl phosphate (1,3-bisphosphoglycerate) • carbamoyl phosphate condensation ‹#› 24 1,3-Bisphosphoglycerate glycerol glyceric acid glycerate 1,3-bisphosphoglycerate anhydride ester ‹#› 25 aminoacyl-AMP (mixed anhydride) + ATP + 2 Pi + H2O Activation of amino acid by ATP in proteosynthesis ‹#› 26 Carbamoyl phosphate carbonic acid H2CO3 carbamoyl (acyl of carbamic ac.) carbamoyl phosphate carbamic acid phosphate ‹#› 27 Amines and acids can react in two ways + + - H2O alkylammonium salt N-alkylamide Low temperature: acid-base reaction High temperature: condensation acid base (alkylamine) ‹#› 28 Amides are polar non-electrolytes Free el. pair on nitrogen is in conjugation with double bond ‹#› 29 Compare properties Feature Amines Amides General formula El. pair on nitrogen Basic properties Salt formation In water behaves as Polar compound pH of aqueous solution R-NH2 free and available yes yes weak electrolyte yes basic R-CO-NH2 conjugated with C=O no no non-electrolyte yes neutral ‹#› 30 Urea is diamide of carbonic acid carbonic acid urea ‹#› 31 Compare Feature Urea Uric acid Chemical name Latin name In water Solubility in water Aqueous solution is Reducing property Salt formation Catabolite of carbonic ac. diamide urea non-electrolyte excelent neutral no no amino acids 2,6,8-trihydroxypurine acidum uricum weak diprotic acid poor* weakly acidic yes Þ antioxidant yes (two types) adenine and guanine * Depends on pH, in acidic pH precipitates from solution !!!! H ‹#› 32 Lactone versus Lactam lactone is cyclic ester lactam is cyclic amide - H2O - H2O ‹#› 33 Compare properties Carboxylic acid Ester Amide weak electrolyte polar soluble in H2O non-electrolyte non-polar insoluble in H2O non-electrolyte polar soluble in H2O ‹#› 34 Polarity of organic compounds (see Medical Chemistry II, chapter 4) hydrocarbons halogen derivatives ethers esters ketones amines amides alcohols carboxylic acids polarity non-polar compounds ‹#› 35 Dehydrogenations of various substrates Substrate Product alkane primary alcohol secondary alcohol endiol aldehyde hydrate hemiacetal / cyclic hemiacetal hydroxy acid p-diphenol thiol amino acid alkene aldehyde ketone diketone carboxylic acid ester / lactone oxo acid p-quinone disulfide imino acid see Med. Chem. II Appendix 3 ‹#› 36 Dehydrogenations in enzyme reactions •Substrate loses 2 H atoms from typical groups: • primary alcohol group -CH2-OH (e.g. ethanol, cholin) • secondary alcohol group >CH-OH (lactate, malate) • endiol group HO-(R)C=C(R)-OH (vitamin C) • secondary amine group >CH-NH2 (amino acid) • saturated hydrocarbon group -CH2-CH2- (fumarate, acyl-CoA) •Product acquires a double bond (C=O, C=NH, CH=CH) •2 H atoms are transferred to cofactor ‹#› 37 Enzyme dehydrogenations require the cooperation of three components enzyme + substrate + cofactor (lactate dehydrogenase) (lactate) (NAD+) suffix -ase vitamin B derivative + reduced substrate oxidized cofactor + 2nd redox pair 1st redox pair dehydrogenase oxidized substrate reduced cofactor ‹#› 38 Dehydrogenation of ethanol (simplified scheme = redox pair) acetaldehyde ‹#› 39 Dehydrogenation of ethanol (complete reaction with cofactor, two redox pairs) NAD+ = nicotinamide adenine dinucleotide alcohol dehydrogenase acetaldehyde ‹#› 40 Oxidation of methanol (simplified scheme) formaldehyde formic acid dehydrogenation oxygenation ‹#› 41 Two ways of glycerol oxidation dehydrogenation on C2 dehydrogenation on C1 oxygenation on C1 glyceraldehyde glyceric acid dihydroxyacetone ‹#› 42 Oxidation of ethylene glycol proceeds stepwise with a number of intermediates ethylene glycol glycolic acid glycolaldehyde glyoxalic acid oxalic acid ‹#› 43 Dehydrogenation of aldehyde hydrate unstable intermediate carboxylic acid aldehyde aldehyde hydrate ‹#› 44 Dehydrogenation of hemiacetal hemioacetal ester cyclic hemiacetal gluconolactone (glucopyranose) - 2H - 2H ‹#› 45 Hydroxy acids as dehydrogenation substrates lactic acid (lactate) acidum lacticum malic acid (malate) acidum malicum b-hydroxybutyric acid b-hydroxybutyrate ‹#› 46 Dehydrogenation of lactic acid (lactate) lactate pyruvate ‹#› 47 Other examples Ketone bodies CAC reaction acetoacetate Beta-hydroxybutyrate malate oxaloacetate ‹#› 48 α,b-Dehydrogenation of acyl (catabolism of fatty acids) saturated acyl-CoA α,β-unsaturated acyl-CoA ‹#› 49 Dehydrogenation of amino compounds R-CH2-NH2 + FMN ® R-CH=NH + FMNH2 amine imine 2-amino acid 2-imino acid imino group >C=NH ‹#› 50 The hydrolysis of imino acid gives oxo acid and free (toxic) ammonia ‹#› 51 Dehydrogenation of diphenol benzene-1,4-diol (hydroquinone) p-benzoquinone (aromatic ring) (not aromatic) ‹#› 52 Dehydrogenation of -SH substrates proceeds with two molecules (mild oxidation)* thiol dialkyl disulfide disulfide bridges in proteins * Strong oxidation occurs with one molecule to give sulfonic acid. ‹#› 53 Oxygenation of aldehyde aldehyde carboxylic acid ½ O2 Direct: Indirect: aldehyde aldehyde hydrate carboxylic acid ‹#› 54 Hydroxylation of phenylalanine tetrahydrobiopterine (BH4) is a donor of 2H to make water from the second oxygen atom phenylalanine tyrosine ‹#› 55 Biochemical hydrogenations •Substrate gains 2 H •Mainly from NADPH + H+ •Reduction syntheses (FA, cholesterol) -CH=CH- + NADPH+H+ ® -CH2-CH2- + NADP+ ‹#› 56 Hydrogenation of ubiquinone ubiquinone ubiquinol ‹#› 57 Deoxygenation is quite rare reaction D-ribose 2-deoxy-D-ribose NADPH+H+ + H2O + NADP+ ‹#› 58 Distinguish: A donor of H+ = acid A donor of H = reductant An acceptor of H+ = base An acceptor of H = oxidant ! Redox versus Conjugate pair ‹#› 59 Conjugate pair lactic acid lactate (conjugate base) Redox pair lactate pyruvate Example 1 ‹#› 60 L-Ascorbic acid is a weak diprotic acid Two conjugate pairs: Ascorbic acid / hydrogen ascorbate Hydrogen ascorbate / ascorbate two enol hydroxyls pKA1 = 4.2 pKA2 = 11.6 Example 2 ‹#› 61 L-Ascorbic acid has reducing properties (antioxidant) ascorbic acid dehydroascorbic acid (reduced form) (oxidized form) ‹#› 62 Uric acid (lactim) is a weak diprotic acid uric acid hydrogen urate urate pKA1 = 5.4 pKA2 = 10.3 2,6,8-trihydroxypurine Example 3 ‹#› 63 Uric acid has reducing properties (physiological antioxidant) various products hydrogen urate radical relatively stable (oxidized form) Hydrogen urate anion affords one electron R· is oxygen radical like ·OH, superoxide. hydrogen urate anion (reduced form) ‹#› 64 Transaminations of amino acids ‹#› 65 Catabolic pathway of nitrogen (in blue colour) • dietary proteins ® AA (stomach, intestine) •transamination of AA in cells ® glutamate •dehydrogenation + deamination of glutamate ® NH3 •detoxication of ammonia (liver) ® urea nitrogen output nitrogen input ‹#› 66 Transamination •amino group is transferred from AA to 2-oxoglutarate •enzyme: aminotransferase •cofactor: pyridoxal phosphate •amino acid gives the corresponding oxo acid •the second product is glutamate ‹#› 67 General scheme of transamination glutamate 2-oxo acid amino acid 2-oxoglutarate aminotransferase reversible reaction ‹#› 68 Dehydrogenation deamination of glutamate NAD(P)+ main source of ammonia in human body glutamate 2-iminoglutarate 2-oxoglutarate glutamate dehydrogenase ‹#› 69 Transamination of alanine ALT = alanine aminotransferase 2-oxoglutarate alanine pyruvate ‹#› 70 Transamination of aspartate aspartát oxalacetát 2-oxoglutarate glutamate aspartate oxaloacetate ‹#› 71 Remember •pyruvate / lactate = redox pair •pyruvate / alanine = transamination pair • •oxaloacetate / malate = redox pair •oxaloacetate / aspartate = transamination pair ! ‹#› 72 Reactions of citric acid cycle How is CO2 made from acetyl-CoA? ‹#› 73 Citric acid cycle •Initial substrate: acetyl-CoA •Three types of products: • 2´ CO2 ® eliminated from body by lungs • 4´ reduced cofactors ® reoxidized in respiratory chain • 1´ GTP ® substrate-level phosphorylation ‹#› 74 Condensation of oxaloacetate with acetyl-CoA oxaloacetate acetyl-CoA citrate ‹#› 75 Isomeration of citrate to isocitrate citrate isocitrate secondary hydroxyl group tertiary hydroxyl group ‹#› 76 Decarboxylation and dehydrogenation of isocitrate isocitrate 2-oxoglutarate ‹#› 77 Decarboxylation and dehydrogenation of 2-oxoglutarate 2-oxoglutarate succinyl-CoA thioester macroergic intermediate ‹#› 78 Substrate phosphorylation gives GTP + CoA-SH succinate succinyl-CoA guanosine triphosphate ‹#› 79 Dehydrogenation of succinate succinate fumarate ‹#› 80 Hydration of fumarate Hydration is not a redox reaction fumarate L-malate ‹#› 81 Dehydrogenation of L-malate re-enters CAC L-malate oxaloacetate