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    Sample chapter alkaloids - Pharmaceutical Chemistry of Natural Products by V. Alagarsamy--order +91 8527622422 Sample chapter alkaloids - Pharmaceutical Chemistry of Natural Products by V. Alagarsamy--order +91 8527622422 Document Transcript

    • CHAPTER 5 Alkaloids INTRODUCTI0N Alkaloids are pharmacologically active, complex organic compounds containing one or more nitrogen atoms, characteristically as primary, secondary or tertiary amines, which provide basicity to the alkaloid. The term alkaloid (alkaloid means alkali-like) cannot be defined exactly, as there is no clear-cut boundary between alkaloids and naturally occurring complex amines. In practice, those substances obtained from plant sources and answer for the standard qualitative tests are called alkaloids. The name protoalkaloid is applied to compounds such as ephedrine and colchicine which are not having certain properties of typical alkaloids. They are found mainly in plants, but to lesser extent in animals and microorganisms. By keeping the knowledge of alkaloids in view, alkaloids may be defined as basic nitrogen containing (usually in a heterocyclic system) plant products, possessing marked pharmacological action at very low dose level. Generally, from an amino acid, the nitrogen atoms in alkaloids are instigated and the carbon skeleton of the particular amino acid precursor is also mostly retained in the structure of alkaloid. However, through the transamination reactions a major group of alkaloids are found to obtain their nitrogen atoms, incorporating only the nitrogen from an amino acid, while the remaining part of the molecule may be derived from acetate or shikimate, or may be terpenoid or steroid in origin. Alkaloids are generally bitter in taste and optically active (except papaverine), usually levorotatory in nature (exception is coniine, which is dextrorotatory), colourless (except berberine which is yellow; harmaline and betanidine which are reddish), crystalline solids (except nicotine and coniine which are liquids) and soluble in organic solvents like chloroform and ethanol but insoluble in water. As they are basic in nature and form salts with the acids, some of the alkaloids exist as salts called quaternary amines (e.g. cinchona alkaloids with quininic acid) while some of them exist as free bases (e.g. nicotine). Some of the alkaloids also occur as glycosides (e.g. solanum alkaloids) and esters (e.g. atropine). Biological activity of the alkaloids frequently depends on the amine function being transferred into a quaternary amine at physiological pH by protonation.Chapter-05.indd 149 6/29/2012 10:19:07 AM
    • 150 Pharmaceutical Chemistry of Natural Products Alkaloids were first isolated successfully in the nineteenth century and the alkaloid-containing drugs were marketed. The structure of the first alkaloid, namely coniine was established in the year 1870 by Schiff. In search of plant drugs with anticancer activity, catharanthus alkaloids and paclitaxel came into the market. NOMENCLATURE OF ALKALOIDS There is no systematic nomenclature of alkaloids because of their complex molecular structure and some historical reasons. They are named by adopting different methods as described below: 1. According to their physiological action: Examples include emetine (Greek word: emetikos means to vomit), morphine (German word: morphine means God of Dreams) and narcotine (Greek word: narkoo means benumb). 2. According to the plants from which they are obtained: Examples include papaverine from Papaver somniferum and berberine from Berberis vulgaris. 3. Prefixes like epi, iso, neo, pseudo or Greek letters are used to name isomeric or slightly modified structures: The prefix nor indicates the structure which does not have a methyl group attached to the nitrogen atom (e.g. norephedrine). 4. According to the name of the discoverer: For example, pelletierine (discovered by P.J. Pelletier). 5. The minor alkaloids are named by adding one prefix or suffix to the name of principal alkaloids: For example, cinchonidine derived from cinchonine. CLASSIFICATION OF ALKALOIDS Alkaloids are classified in different methods. Some of these methods are described briefly and the chemical classification is described in detail: 1. Pharmacological classification: It is based on the clinical use or pharmacological activity. Examples include analgesic alkaloids and cardioactive alkaloids. 2. Taxonomic classification: It is based on the family or genus, without reference to the chemical type of alkaloid present (e.g. solanaceous alkaloids). However, the most common classification is according to the genus in which they occur (e.g. rauwolfia alkaloids and cinchona alkaloids). 3. Biosynthetic classification: It is based on the type of precursors or building block compounds used by plants to synthesize alkaloids. This is a more fundamental method than chemical classification. For example, morphine, papaverine, narcotine and colchicine may be listed as phenylalanine- and tyrosine-derived bases. 4. Chemical classification: It is based on the chemical structure of the alkaloid. The chemical classification of alkaloids is universally adopted and depends on the basic ring structure present. For example, atropine is a tropane alkaloid; quinine is considered as a quinoline- type alkaloid; papaverine is an isoquinoline and reserpine, strychnine and ergometrine are indole alkaloids.Chapter-05.indd 150 6/29/2012 10:19:07 AM
    • Alkaloids 151 Alkaloids True alkaloids Proto/amino alkaloids Pseudo alkaloids Example: Steroidal, terpenoidal alkaloids Contain heterocyclic nitrogen Simple amines Not derived from amino acids · but from acyl CoA units Derived from amino acids Example: Conestine, caffeine Based on the chemical nature, alkaloids are further classified into two major groups as mentioned below: 1. Heterocyclic or typical alkaloids 2. Nonheterocyclic or atypical alkaloids [protoalkaloids (or) biological amines] They are further subdivided as follows: Heterocyclic Alkaloids 1. Pyridines and piperidines CH3 H H O N N O C C C C C N CH3 COOCH3 H H N O Piperine Nicotine Arecoline OCH3 OH CN O O N N O N Me N H CH3 H CH3 Conine Lobeline Pelletierine RicinineChapter-05.indd 151 6/29/2012 10:19:07 AM
    • 152 Pharmaceutical Chemistry of Natural Products 2. Quinolines H H CHϭCH2 CHϭCH2 H2C H2C H H HO CH2 HO CH2 N N H H H3CO H3CO N N Quinine Quinidine H H CHϭCH2 CHϭCH2 H2C H2C H H HO CH2 HO CH2 N N H H N N Cinchonine Cinchonidine 3. Isoquinolines H3CO O H2C N O N CH3 H3CO O H O H ClϪ CH2 H3CO O Nϩ O C OCH3 H3CO OCH3 OCH3 OCH3 OCH3 Noscapine Berberine chloride PapaverineChapter-05.indd 152 6/29/2012 10:19:08 AM
    • Alkaloids 153 H3CO H3CO N N H3CO H3CO H H CH3 CH3 H H H H H H OCH3 OCH3 HN HN OCH3 OH Emetine Cephaline H3C OCH3 CH3 O ϩ N OH OH O H H H3CO O Nϩ H3C N H3CO OH H CH3 (Ϯ)-Tubocurarine Galanthamine 4. Phenanthrenes CH3 CH3 CH3 N N N H H H O O O OH H3CO OH H3CO OCH3 HO Morphine Codeine ThebaineChapter-05.indd 153 6/29/2012 10:19:08 AM
    • 154 Pharmaceutical Chemistry of Natural Products 5. Indole alkaloids H H3C C HNOC HOOC NH NH CH2OH N N CH3 CH3 Ergometrine Lysergic acid CH3 CH3 OH R1 H CONH O R1= CH3NHCOO — N R2= CH3 R3= CH3 H N N N N O O H CH3 H CH2 R2 R3 H Physostigmine HN Ergotamine N H3CO N H H O H OCH3 O H C C O OCH3 H3CO OCH3 OCH3 ReserpineChapter-05.indd 154 6/29/2012 10:19:08 AM
    • Alkaloids 155 OH N N N CH2CH3 H H H O H H C N Vinblastine R Ϫ CH3 COOCH3 H3CO Vincristine R Ϫ CHO OH H N Yohimbine CH2CH3 H H H3CO N OCOCH3 HO C R1 N R OCH3 O R2 N O O Strychnine R1 = R2 = ϪH Brucine R1 = R2 = ϪOCH3 6. Pyrrole and pyrrolidines O N CH2 C CH3 N+ COOϪ CH3 Me Me Hygrine Stachydrine 7. Tropane alkaloids C6H5 C6H5 O NиCH3 OCOϪCH NиCH3 OCOCH CH2OH CH2OH Atropine HyoscineChapter-05.indd 155 6/29/2012 10:19:08 AM
    • 156 Pharmaceutical Chemistry of Natural Products COOR CH2OH OR’ NиCH3 NиCH3 OOCиCH H C6H5 R= CH3 Hyoscyamine R’= C6H5CO (Benzoyl) Cocaine 8. Imidazole or glyoxalines H H H3C NиCH3 O O N Pilocarpine 9. Purines O O O H3CиN NиCH3 H3C.N NH HN NиCH3 C C C N N N N N N O O O CH3 CH3 CH3 Caffeine Theophylline Theobromine 10. Terpenoid alkaloids OH OCH3 OCH3 OCOC6H5 OH H3C N OCOCH3 HO H2C OCH3 OCH3 AconitineChapter-05.indd 156 6/29/2012 10:19:08 AM
    • Alkaloids 157 11. Steroidal alkaloids CH3 H3C N CH3 CH3 N CH3 CH3 CH3 H3C N HO H3C Solanidine Conessine Nonheterocyclic Alkaloids HO OH H H OH C N C CH3 H H3C CH3 HC CH2 N H OH Ephedrine (Ϯ)-Adrenaline O H3COCO O OH O H3C CH3 C NH C CH3 O CH3 O OH AcO HO O C O TaxolChapter-05.indd 157 6/29/2012 10:19:09 AM
    • 158 Pharmaceutical Chemistry of Natural Products MeO NHCOCH3 CH3 HO CH2 CH2 N MeO CH3 OMe O Colchicine Hordenine OMe OCH3 H3CO OCH3 N(ϩ) N (Ϫ) CH2 CH2 O H3COOC NH2 Mescaline Serpentine QUALITATIVE CHEMICAL TESTS FOR ALKALOIDS General tests answered by all alkaloids are as follows: 1. Dragendorff’s test: To 2–3 mL of the alkaloid solution add few drops of Dragendorff’s reagent (potassium bismuth iodide solution). An orange brown precipitate is formed. 2. Mayer’s test: To 2–3 mL of the alkaloid solution add few drops of Mayer’s reagent (potassium mercuric iodide solution). White brown precipitate is formed. 3. Hager’s test: To 2–3 mL of the alkaloid solution add few drops of Hager’s reagent (saturated solution of picric acid). Yellow precipitate is formed. 4. Wagner’s test: To 2–3 mL of the alkaloid solution add few drops of Wagner’s reagent (iodine– potassium iodide solution). Reddish brown precipitate is formed. 5. For opium alkaloids: These alkaloids are present as salts of meconic acid. Opium is dissolved in water, filtered and to the filtrate, ferric chloride solution is added by which deep reddish purple colour is obtained. The colour persists even upon adding hydrochloric acid. 6. For tropane alkaloids (Vitalis–Morin reaction): Tropane alkaloid is treated with fuming nitric acid, followed by evaporation to dryness and addition of methanolic potassium hydroxide solution to an acetone of nitrated residue. Violet colouration takes place because of the presence of tropane derivative. 7. For purine alkaloids (murexide colour reaction): Caffeine is taken in a Petri dish to which hydrochloric acid and potassium chlorate are added and heated to dryness. A purple colour is obtained by exposing the residue to vapour of dilute ammonia. The purple colour is lost upon addition of alkali. Caffeine (and other purine alkaloids) gives murexide colour reaction.Chapter-05.indd 158 6/29/2012 10:19:09 AM
    • Alkaloids 159 ISOLATION OR PRODUCTION OF ALKALOIDS Alkaloid bearing plant usually contains a complex mixture of alkaloids, hence isolation and purification of an alkaloid from a plant is always not a simple process. Further, the presence of organic acids, glycosides, etc., present in the plants may complicate the isolation process. Thus, isolation of pure alkaloid may often become a laborious procedure. The steps involved in the isolation of an alkaloid may be summarized as follows: 1. The presence of an alkaloid in a plant is ascertained by using the various alkaloidal reagents. (Refer the qualitative tests mentioned above.) 2. The next step is the separation of relatively small amount of alkaloids from large amount of extraneous plant materials. 3. The final step is the separation and purification of individual alkaloids from the crude mixture. The isolation of an alkaloid from the crude powdered drug is schematically represented below. Powdered drug containing alkaloid salts (tannates, oxalates, etc.) 1. Defat if necessary with petroleum ether 2. Moisten and render alkaline with Na2CO3 or K2CO3 or NH3 or Ca(OH)2 Free alkaloids Extract the alkaloid with organic solvent like CHCl3, ether Total extract Concentrate and shake the extract with succes- sive quantities of acid like dil. H2SO4 Aqueous acid solution of alkaloid Residual organic fraction (Pigments, fats and occasionally very weak Make alkaline with Na2CO3 bases or chloroform soluble alkaloid sulphates) and extract alkaloids with an immiscible solvent like CHCl3 Residual aqueous fraction Organic solution of the alkaloid bases Distil off the solvent Crude alkaloid mixture Fractional crystallization, fractional precipitation, column chromatography, partition chromatography, gas chromatography or by counter current extraction Individual alkaloidsChapter-05.indd 159 6/29/2012 10:19:09 AM
    • 160 Pharmaceutical Chemistry of Natural Products DETERMINATION OF MOLECULAR STRUCTURE OF ALKALOIDS: GENERAL METHODS As the molecular structure of alkaloids is complex, only recently few of the complex alkaloids’ structures were elucidated. The various chemical methods performed to determine the structure of alkaloids is as follows: Molecular Formula Determination The first step in structural elucidation is the determination of molecular formula and optical rotatory power. Elemental composition and hence the empirical formula is found by combustion analysis. Hydrolysis Simple fragmentation by hydrolysis with water, acid or alkali yields simple fragments which are then analysed separately. For example, atropine on hydrolysis yields tropine and tropic acid. Determination of Unsaturation The unsaturation can be determined by adding bromine, halogen acids or by hydroxylation with KMnO4 or by reduction (using either LiAlH4 or NaBH4). Functional Group Determination By using the usual standard chemical tests or by infrared (IR) spectroscopy, functional nature of the alkaloids is determined. Functional nature of oxygen: The oxygen atom may be present in the form of alcoholic hydroxyl (–OH), phenolic hydroxyl (–OH), methoxyl (OCH3), acetoxy (–OCOH3), benzoxyl (–OCOC6H5), carboxyl (–COOH), aldehyde (–CHO), ketone (C=O) and methylene dioxide group (–O–CH2–O–). These groups are characterized by the chemical tests as follows: Phenolic hydroxyl group (=C–OH) It is identified by the following tests:  Soluble in alkali and reprecipitation by CO2.  Violet colouration with neutral ferric chloride.  Yields ester on acetylation. This reaction can be used to determine the number of phenolic –OH.  Yields ether on reaction with alkyl halide.Chapter-05.indd 160 6/29/2012 10:19:09 AM
    • Alkaloids 161 Alcoholic hydroxyl group (–C–OH) It yields ester on acetylation and benzoylation (but negative answer for phenolic –OH)—refer above tests. This is confirmed by oxidation, dehydration, dehydrogenation and by spectroscopy (IR and NMR). Alcohols are of three different types: 1°, 2° and 3°, and they are usually distinguished by their oxidation products. Primary alcohol (O) (O) R—CH2—OH R—CHO R—COOH Aldehyde with Carboxylic acid same no. of with same no. of ‘C’ as in alcohol ‘C’ as in alcohol and aldehyde Secondary alcohol (O) (O) R—CH2—CH—CH3 R—CH2—C—CH3 R—CH2—C—OH OH O O Ketone with Carboxylic acid same no. of with fewer no. of C as in alcohol C with respect to alcohol and ketone But in cyclic structure, 2° alcohol yields different oxidation products as shown below: OH O COOH (O) (O) COOH Ketone with Acid with same no. of same no. of C as in alcohol C as in alcohol Tertiary alcohol CH3 (O) (O) R—CH2—C—CH3 R—CH2—C—CH3 R—CH2—C—OH OH O O Ketone with Carboxylic acid lesser no. of with fewer no. of C with respect to C with respect to alcohol alcohol and ketoneChapter-05.indd 161 6/29/2012 10:19:09 AM
    • 162 Pharmaceutical Chemistry of Natural Products The number of hydroxyl (OH) groups present in the compound is determined by the following methods: Acetylation method: H+ R—OH + (CH3CO)2O R—O—CO—CH3 By determining the amount of acetic anhydride that reacted with alcohol to form an ester, the number of hydroxyl groups is determined. Zerewitinoff active hydrogen determination method: When alcohol is heated with CH3MgI, methane is obtained. By measuring the methane so formed, the amount of alcohol can be determined. I R—OH ϩ CH3MgI CH4 ϩ Mg OR –OH = CH4 = 22.4 L of alcohol at normal temperature and pressure. Carbonyl group The presence of aldehydes and ketones is detected by their reaction with hydroxylamine, semicarbazide and phenylhydrazine to form the corresponding oxime, semicarbazone and phenylhydrazone, respectively. NH2OH. HCl C O C N OH ϩ HCl ϪH2O Oxime By determining the HCl formed, the ketones are estimated quantitatively. Ph—NHNH2 C O C N NH — Ph ϩ HCl ϪH2O Phenylhydrazone NO2 O2 N NHNH2 NO2 C O O2 N NH NϭC Dinitrophenylhydrazone O O H2N — NH — C — NH2 C O C N NH C NH2 ϪH2O Semicarbazone The aldehydes and ketones are distinguished by their oxidation or reduction products. The carbonyl groups of aldehydes, ketones and carboxyl are further confirmed by their spectral data such as IR, ultraviolet (UV) and NMR.Chapter-05.indd 162 6/29/2012 10:19:09 AM
    • Alkaloids 163 Carboxyl group (–COOH) The presence of carboxyl group is determined by the following:  Its solubility in weak bases such as NH3, NaHCO3 and Na2CO3.  Esterification with alcohols.  Specific IR and NMR signals.  Quantitatively by acid–alkali titration: Performed by titrating the carboxylic acid with NaOH using phenolphthalein as an indicator. By knowing the volume of NaOH consumed the number of –COOH groups are determined. Ester group (RCOOR) Esters and related groups like amides and lactones are detected by their reaction with water, dilute acids or alkali to the hydroxyl and acidic compounds. By elucidating the acid and alcohol, the nature of alkaloids is determined. R—COOR’ R—COOH ϩ R’OH R—CONH2 R—COOH ϩ NH3 NaOH R — CH — CH2 — CH2 R — CH — CH2 — CH2 O CO OH COONa Alkoxy group (–OR) Determined by Zeisel’s method—alkoxy group such as methoxy on reacting with hydroiodic acid followed by silver nitrate yields equal amount of silver iodide. From the amount of silver iodide formed, the number of alkoxy groups is calculated. 126°C AgNO3 OCH3 ϩ HI –OH ϩ CH3I AgI + CH3NO2 Boil Estimation of C-methyl group (Kuhn Roth method): By estimating the acetic acid formed upon oxidation, the C-methyl groups are quantified. K2Cr2O7/ H2SO4 C CH3 CH3COOH Functional nature of nitrogen: Most alkaloids contain ‘N’ in their ring structure, which may exist as 2° or 3°.Chapter-05.indd 163 6/29/2012 10:19:10 AM
    • 164 Pharmaceutical Chemistry of Natural Products The 2°and 3° amines are distinguished as follows:  2° Amines (acetylated or benzoylated) undergo Libermann’s nitroso reaction.  2° Amines take up 2 moles of alkyl halide to form 4° ammonium salt.  3° Amines take up 1 mole of alkyl halide to form 4° ammonium salt. (O) 3° N N N O 30% H2O2 Amine oxide Distillation Alkaloid Methylamine, Dimethylamine, Trimethylamine –(indicates the presence of 1 or 2 alkyl groups attached to amine ‘N’) –Ammonia (indicates the presence of primary amine) Further, the nature of ‘N’ is confirmed by degradation methods such as Hoffmann Exhaustive Methylation (HEM). The N-alkyl groups are estimated by Herzig–Meyer method: HI AgNO3 N CH3 N H ϩ CH3I AgI 150–300°C EtOH Under pressure* HI AgNO3 N C2H5 N H ϩ C2H5I AgI 150–300°C EtOH Under pressure* *Differs form –OR (alkoxy group) estimation From the amount of silver iodide formed, the number of N-alkyl groups is calculated. Degradation of Alkaloids Degradation of alkaloids gives rise to some identifiable products of known structure and hence by knowing structure of the degraded products and the changes occurred during the degradation it is convenient to know the structure of the original molecule. Different degradation reactions carried out in elucidating the structure of alkaloids are as follows: 1. HEM method 2. Emde method 3. Von Braun’s (VB) method for 3° cyclic amines 4. Reductive degradation 5. Oxidation 6. Zinc distillation 7. Alkali fusion 8. Dehydrogenation 1. HEM method: Originally this method was applied by Willstater in 1870 for naturally occurring alkaloids. It was further developed by Hoffmann and hence it is known as HEM. Principle of this method is that the quaternary ammonium hydroxides yield olefin with the cleavage of carbon– nitrogen linkage upon heating with the loss of water molecule (H from β-carbon atom with respect to N and OH from the 4° ammonium hydroxide).Chapter-05.indd 164 6/29/2012 10:19:10 AM
    • Alkaloids 165 Quaternization is done by complete methylation of the amine followed by hydrolysis with moist Ag2O or KOH. 2 CH3I Moist Ag2O R—CH2—CH2—NCH3 R—CH2—CH2—Nϩ(CH3)3I R—CH2—CH2—Nϩ(CH3)3OHϪ (AgOH) –H2O ⌬ –N(CH3)3 RCHϭCH2 Olefin This method can be applied to the reduced ring system but fails with unsaturated analogues and hence, the unsaturated rings are first saturated and then HEM is performed. H2–Ni 2CH3I ⌬ AgOH ϪH2O OHϪ N N Nϩ N H (CH3)2 (CH3)2 (i) CH3I (ii) AgOH Isomerism ⌬ -H2O -N(CH3)3 Nϩ OHϪ (CH3)3 As β-hydrogen atom is needed to cleave C–N bond and eliminate water molecule, the HEM fails on the ring system that does not have β-hydrogen atom. For example, in the degradation of isoquinoline, the cleavage of N does not occur at the final step as there is no β-hydrogen with respect to ‘N’. Na—EtOH (i) 2CH3I N NH (ii) AgOH Nϩ (CH3)2 OHϪ ⌬ ϪH2O (i) 2CH3I Nϩ (CH3)3 OHϪ (ii) AgOH N (CH3)2Chapter-05.indd 165 6/29/2012 10:19:10 AM
    • 166 Pharmaceutical Chemistry of Natural Products However, there are some cases in which HEM fails even if the β-hydrogen atom is present (the following reaction explains this). (i) 2CH3I ⌬ (ii) AgOH ϪCH3OH N NϩOHϪ N H (CH3)2 CH3 (95%) Ϫ H2O N (CH3)2 (5%) 2. Emde method: Emde modification may be used in the above two cases, where HEM failed. In this method, 4° ammonium halide is reduced with sodium amalgam in aqueous ethanol or Na–liquid NH3 or catalytically. Na—EtOH (i) 2CH3I N NH (ii) AgOH Nϩ (CH3)2 OHϪ ⌬ ϪH2O (i) 2CH3I (ii) AgOH Nϩ (CH3)2 OHϪ N (CH3)2 (Beta hydrogen absent) Na–liq. NH3 (or) Na–Hg H2O–EtOH CH3 (Alpha methyl styrene)Chapter-05.indd 166 6/29/2012 10:19:11 AM
    • Alkaloids 167 Tetrahydroquinoline is degraded as follows: Na–liq. NH3 Nϩ IϪ N N H3C CH3 H3C CH3 (CH3)2 3-Phenyl-N,N-dimethyl O-Propyldimethyl aniline propylamine Emde degradation on tetrahydroisoquinoline also proceeds as follows: Nϩ (CH3)2 IϪ Nϩ (CH3)2 H2–Pt (or) (I) CH3I Na–Hg (ii)Na–Hg HEM N (CH3)2 CH3 CH3 3. VB method: (a) For 3° cyclic amines: The 3° N atom in the ring upon reaction with CNBr followed by hydrolysis yields brominated 2° amine. BrCN Hydrolysis Ϫ Br CH2Br HBr CH2Br N Nϩ N NH R CN R CN R R 2° Amine This method is applied on compounds which do not respond to HEM. Ring opening takes place differently in VB and HEM method which is shown in the following degradation. HEM VB CH3 N N (CH3)2 N (CH3)2 CN CH2BrChapter-05.indd 167 6/29/2012 10:19:11 AM
    • 168 Pharmaceutical Chemistry of Natural Products In general, CNBr cleaves the unsymmetrical amines to yield the bromides or shorter bromides. However, in the VB method only dealkylation may occur without ring cleavage in some cases. COOCH3 COOCH3 COOCH3 (i) HCl CNBr (ii) ϪCO2 N — CH3 OCOC6H5 N — CN OCOC6H5 N—H OCOC6H5 (b) For 2° cyclic amines: C6H5COCl PBr5–Br2 Distil under Br—(CH2)5Br + C6H5CN reduced pressure N N N 1,5-Dibromo pentane H COC6H5 Br—C—C6H5 Br 4. Reductive degradation: Ring system is opened by treating with HI in many cases. HI (or) CH3—(CH2)3—CH3 + NH3 300°C n-Pentane N N H 5. Oxidation: Oxidation gives valuable information about the fundamental structure of alkaloids and the position and nature of functional groups, side chains, etc. For example, picolinic acid obtained upon oxidation of coniine indicates that the coniine is an α-substituted pyridine. (O) Coniine N COOH Picolinic acid By varying the strength of oxidizing agents, a variety of products may be obtained. Different types of oxidizing agents used are as follows: 1. For mild oxidation: H2O2, O3, I2. 2. For moderate oxidation: acid or alkali KMnO4, CrO3 in CH3COOH. 3. For vigorous oxidation: K2Cr2O7–H2SO4, concentrated HNO3 or MnO2–H2SO4. 6. Zinc distillation: Distillation of alkaloid over zinc dust degrades it into a stable aromatic derivative. Zinc dust Morphine Phenanthrene distillationChapter-05.indd 168 6/29/2012 10:19:11 AM
    • Alkaloids 169 The reaction indicates that morphine is possessing phenanthrene nucleus. 7. Alkali fusion: Fusion of alkaloids with solid KOH gives simple fragments from which the nature of alkaloid can be derived. Fusion with Papaverine Substituted isoquinoline KOH The reaction indicates papaverine is containing isoquinoline nucleus. HO COOH KOH Adrenaline fusion HO Protocatechuic acid The reaction indicates adrenaline is a monosubstituted catechol derivative. 8. Dehydrogenation: Distillation of alkaloid with catalysts such as S, Se and Pd yields simple and recognizable products from which the gross skeleton of the alkaloid may be derived. Thus with the help of degradation, nature of various fragments obtained, nature of nucleus and type of linkages are established. The fragments obtained are arranged in the possible ways with the possible linkages and the one that will explain all the properties is selected and confirmed by synthesis. Optical activity of an alkaloid helps greatly in establishing the structure of alkaloid. Physical Methods in Conjunction with Chemical Methods The developments in spectroscopic methods not only curtailed time consumption as compared to degradation studies, but also helped in determining the molecular structure of complex alkaloids. Morphine structure was established after the developments of the below-mentioned physical methods. The complete structure of vindoline (including configuration) has been established by spectroscopic methods. Thelepogine, another alkaloid structure, has been established using X-ray analysis technique without performing any of the chemical analysis. The important physical methods used in structural elucidation of alkaloids are as follows: 1. IR spectroscopy 2. UV spectroscopy 3. NMR spectroscopy 4. Mass spectroscopy 5. X-ray analysis 6. Optical rotatory dispersion (ORD) and circular dichroism 7. Conformational analysis IR spectroscopy: This method is used to identify the presence of functional groups such as –OH, –NH2, –NH and –C=O. The groups such as –OCH3, –NCH3, –OH, –NH2 and –NH can be detected by IR spectroscopy but quantified by NMR spectroscopy. NMR spectroscopy: This method helps to detect protons of alkyl, alkenyl, N-methyl, O-methyl, C-methyl, aryl and heteroaryl groups, etc. It also helps in quantitative estimation of these groups. Aromatic and heteroaromatic protons are exactly quantified by using NMR spectroscopy.Chapter-05.indd 169 6/29/2012 10:19:11 AM
    • 170 Pharmaceutical Chemistry of Natural Products UV spectroscopy: UV spectrum of a compound is characteristic of chromophoric system and not the whole compound. Hence, it helps to establish the likely structural type or class of the alkaloid under investigation. Mass spectroscopy: This method is used to confirm the proposed molecular structure of the alkaloid by determining the molecular weight of compounds and the fragments of the degradation products. It also helps to confirm the side chain or attached groups by analysing the fragmentation pattern. X-ray analysis: This method is used to distinguish the various possible structures of alkaloids. ORD and circular dichroism: This method is used to confirm the structure of optically active stereoisomers. Conformational analysis: It is an experimental technique used to establish the stereochemistry as well as physical properties and chemical reactivity of alkaloids. Synthesis The above-mentioned chemical and analytical work helps to propose a tentative structure (or structures) of the alkaloid under investigation. Synthesis always gives additional evidence for the assigned structure even though the physical methods (mentioned above) provide final proof of the proposed structure. ALKALOIDS OF PHARMACEUTICAL IMPORTANCE Source of medicinally important alkaloids with their pharmacological properties and uses are depicted in Table 5.1. Table 5.1 Source pharmacological properties and uses of alkaloids S. no. Alkaloids Source (Family) Pharmacological properties/uses 1. Piperine Piper nigrum and other Piper spp Aromatic, stimulant, stomachic, carminative, (Piperaceae) condiment stimulates taste buds and gastric juice. 2. Nicotine Nicotiana tobaccum (Solanaceae) Stimulant effects on heart and nervous system. 3. Arecoline Areca catechu Parasympathomimetic, anthelmentic drug. (Palmae) 4. Lobeline Lobelia nicotianaefolia (Campanulaceae Used in asthma and as respiratory stimulant. or Lobeliaceae) 5. Pelletierine Punica granatum (Euphorbiaceae) Anthelmintic against tapeworm. Astringent in the treatment in diarrhoea. 6. Quinine, quinidine, Cinchona officinalis and other Cinchona Antimalarial, bitter stomachics, antipyretic cinchonine, spp (Rubiaceae) cinchonidineChapter-05.indd 170 6/29/2012 10:19:12 AM
    • Alkaloids 171 S. no. Alkaloids Source (Family) Pharmacological properties/uses 7. Quinidine C. officinalis and other Cinchona spp Antiarrhythmic (Rubiaceae) 8. Morphine Papaver somniferum (Papaveraceae) Hypnotic, sedative and analgesic. It sedates respiratory centre, emetic centre and the cough reflux. 9. Codeine P. somniferum (Papaveraceae) Used in the treatment of cough. 10. Thebaine, papaverine, P. somniferum (Papaveraceae) Hypnotic sedative and analgesic. noscapine, narceine 11. Berberine Various genera of Berberidaceae In the treatment of cutaneous Leishmaniasis. (Ranunculaceae, Papaveraceae) 12. Emetine, cephaline Cephaelis ipecacuanha, Cephalis Expectorant in small doses and emetic in accuminata (Rubiaceae) higher doses. Antiprotozoal against Entamoeba histolytica. 13. Tubocurarine Chondrodendeon tomentoscum Neuromuscular blocking agent, skeletal (Menispermaceae) muscle relaxant. 14. Galanthamine Leucojum aestivum (Amaryllidaceae) Used in the treatment of Alzheimer’s disease 15. Ergometrine Claviceps purpurea (Clavicipitaceae) Oxytocic in obstetrics. 16. Ergotamine C. purpurea (Hypocreaceae) Specific analgesic in the treatment of migraine. 17. Lysergic acid amide Rivea corymbosa As a psychotomimetic. (Convolvulaceae) 18. Physostigmine Physostigma venenosum (Leguminosae) Parasympathomimetic (ophthalmic) activity. 19. Reserpine Rauwolfia serpentine As an antihypertensive and in the treatment of (Apocynaceae) neuropsychiatric disorder. 20. Serpentine R. serpentine As an antihypertensive and in the treatment of (Apocynaceae) neuropsychiatric disorder. 21. Yohimbine Aspidosperma spp In the treatment of erectile dysfunction. (Apocynaceae) 22. Vincristine Catharanthus roseus As an antineoplastic, for treating leukaemia in (Apocynaceae) children, as hypotensive and in the treatment of diabetes. 23. Vinblastine C. roseus In the treatment of Hodgkin’s disease. (Apocynaceae) 24. Strychnine Strychnos nux-vomica Stomachic, tonic, stimulant to CNS, CVS and (Loganiaceae) respiratory. Increases blood pressure and hence used in the treatment of heart failure. 25. Brucine S. nux-vomica Stomachic, tonic, stimulant to CNS, CVS and (Loganiaceae) respiratory. Increases blood pressure and hence used in the treatment of heart failure. 26. Hygrines Erythroxylum coca Local anaesthetic and stimulant. (Erythroxylaceae) 27. Atropine, Hyoscine, Atropa belladonna, Datura stramonium Parasympatholytic agent, decreases saliva, Hyoscyamine and Hyoscyamus spp sweat, gastric juice. (Solanaceae) 28. Cocaine Cocca spp (Erythroxylaceae) Local anaesthetic, CNS stimulant. 29. Pilocarpine Pilocarpus jaborandi (Rutaceae) Physiological antagonist of atropine, increases sweating, used in the treatment of glaucoma.Chapter-05.indd 171 6/29/2012 10:19:12 AM
    • 172 Pharmaceutical Chemistry of Natural Products S. no. Alkaloids Source (Family) Pharmacological properties/uses 30. Caffeine Coffea arabica (Rubiaceae) CNS stimulant and weak diuretic. 31. Theophylline Thea sinensis CNS stimulant and weak diuretic. (Theaceae) 32. Theobromine Thea sinensis CNS stimulant and weak diuretic. (Theaceae) 33. Aconitine Aconitum napellus and other Aconitum In the treatment of neuralgia, sciatica, spp (Ranunculaceae) rheumatism, inflammation, analgesic and cardiac depression. 34. Conessine Holarrhena antidysenterica Antiprotozoal, used in the treatment of (Apocynaceae) dysentery. 35. Ephedrine Ephedra gerardiana Sympathomimetic and used in the treatment and other Aconitum spp of asthma. (Ephedraceae) Used in the treatment of allergic condition such as hay fever. 36. Adrenaline Adrenal glands Sympathomimetic 37. Taxol Taxus brevifolia Anticancer (paclitaxel) (Taxaceae) 38. Colchicine Colchicum autumnale In the treatment of gout and rheumatism. and other Aconitum spp Antitumour activity. (Liliaceae) Abbreviations: CNS, central nervous system; CVS, cardiovascular system. Some of the pharmaceutically important alkaloids are described with their structural elucidation herein. Tropane Alkaloids Atropine 5 4 O H 6 2 8 N(CH3) O C C 3 1 7 3 2 CH2OH 1 8-Methyl-8-aza-bicyclo[3.2.1]octan-3-yl-3-hydroxy-2-phenylpropanoate Atropine is a naturally occurring belladonna alkaloid that is extracted from the belladonna plant. It is the racemic mixture of L-hyoscyamine, and hence it can also be called (±) hyoscyamine. It is the tropine ester of racemic tropic acid and occurs mainly in the roots of deadly nightshade (Atropa belladonna), thorn apple (Datura stramonium) of the Solanaceae family along with L-hyoscyamine (an optically active form). Mode of action: It is a competitive antagonist for the muscarinic acetylcholine receptor. Atropine reduces the parasympathetic activity of all muscles and glands regulated by the parasympathetic nervous system through muscarinic (M) receptors. M1 and M3 receptors function through GqChapter-05.indd 172 6/29/2012 10:19:12 AM
    • Alkaloids 173 protein and activate membrane-bound phospholipase C, generating inositol triphosphate and diacylglycerol which releases calcium ions to produce depolarization in glands and smooth muscles. M2 receptors open through Gi proteins to activate potassium channels resulting in hyperpolarization (decreases the cardiac function). Atropine equally blocks the M1, M2 and M3 subtypes of receptors and antagonizes the actions. Properties and uses: It is a white, crystalline powder or colourless crystals. Freely soluble in alcohol and well soluble in water. The greater molar potency of atropine helps it to block several moles of acetylcholine. The umbrella-like atropine molecule may mechanically or electrostatically inactivate adjacent receptors on the cell surface so that these receptors are also unavailable for acetylcholine or other parasympathomimetic stimulants. It is well absorbed from the gastrointestinal (GI) tract and distributed throughout the body. It crosses the blood–brain barrier to enter the central nervous system (CNS), where large doses produce stimulant effects and toxic doses produce depressant effects. Atropine is also absorbed systemically when applied locally to mucous membranes. The drug is rapidly excreted in the urine. Atropine is used in ophthalmology for its mydriatic action on eye, and it is also used to relieve night sweats as it diminishes salivary and gastric secretions. Structural elucidation The structure of atropine is established as follows: 1. Molecular formula: The molecular formula of atropine is found to be C17H23NO3. 2. Atropine is an ester: Atropine on hydrolysis yields tropine and (±) tropic acid. Therefore, atropine is a tropine ester of tropic acid (tropine tropate). Ba(OH)2 C17H23NO3 ϩ H2O C8H15NO ϩ C9H10O3 Atropine Tropine Tropic acid Structure of tropic acid (C9H10O3)  Results of the usual standard tests reveal that tropic acid is found to possess one 1° alcohol (–OH) and –COOH group.  Upon strong heating tropic acid is converted to atropic acid, which on further oxidation yields benzoic acid. This indicates atropic acid and tropic acid have benzene ring with a side chain. COOH ⌬ (O) C9H10O3 ϪH2O C9H8O2 Therefore, atropic acid is having one –COOH, one double bond and one benzene nucleus. CH2 C6H5CиCOOH C6H5CHϭCH.COOH I II Structure II is known to be cinnamic acid, hence I is atropic acid.  As atropic acid is obtained by dehydration of tropic acid, addition of water molecule to atropic acid yields tropic acid—compound III or IV.Chapter-05.indd 173 6/29/2012 10:19:12 AM
    • 174 Pharmaceutical Chemistry of Natural Products CH3 CH2OH C6H5иCиCOOH C6H5иCиCOOH OH H III IV The structure (IV) is found to be correct, as it has one 1° alcohol [first point under the section ‘Structure of Tropic Acid (C9H10O3)’] and it is confirmed by synthesis. Synthesis of tropic acid from acetophenone CH3 CH3 CH3 CH2 CH2Cl HCN OH HCl OH Heat ether C6H5иCϭO C6H5иC C6H5иC C6H5иCиCOOH C6H5иCHиCOOH CN COOH ϪH2O HCl Acetophenone Atrolactic acid Na2CO3 H2 O CH2OH C6H5иCHиCOOH (ϩ)-Tropic acid Structure of tropine (C8H15NO)  Results of the usual standard tests reveal that the ‘N’ atom is found to be present as 3° N.  By Herzig–Meyer method, it is found to possess one N—CH3 group.  Results of the usual standard tests reveal that it is found to possess one 2° alcohol (–OH) group (benzoylation and oxidation reaction).  Ladenburg performed the following reactions on tropine. HI (H) Distil Zn dust Tropine Tropine iodide Tropane CH3Cl ϩ nor-Tropane hydrochloride C8H15NO C8H14NI C8H14N C7H13N N C2H5 2-Ethylpyridine On this basis, Ladenburg proposed that tropine is a reduced pyridine derivative. or N CH2CH2OH N CHOHиCH3 CH3 CH3Chapter-05.indd 174 6/29/2012 10:19:12 AM
    • Alkaloids 175 Oxidation of tropine: Tropinic acid is a dicarboxylic acid obtained upon oxidation (possessing same number of carbon atom as in alcohol) and hence the alcoholic group in tropine must be present in ring structure and hence, Ladenburg structure is discarded. CrO3 CrO3 C8H15NO C8H13NO C8H13NO4 Tropine Tropinone (Ϯ)-Tropinic acid Furthermore, it found that tropinone yields dibenzylidene derivative which is a characteristic of –CH2–CO–CH2. This confirms that the ketone and alcohol in tropine is linked to a ring structure. In HEM, tropinic acid yields pimelic acid. Formation of pimelic acid confirms the presence of seven-membered carbon chain in tropinic acid and tropine. HEM 4H C8H13NO4 C7H8NO4 HOOCи(CH2)5иCOOH Tropinic acid Piperylene Pimelic acid carboxylic acid Presence of five-membered ring is confirmed on the basis of the formation of N-methylsuc- cinimide from tropinic acid on oxidation. H2C CH CH2COOH H2C CO CrO3 NиCH3 NиCH3 H2SO4 H2C CH COOH H2C CO Tropinic acid N-Methylsuccinimide Thus, the structure of tropinone and tropine can be proposed as follows: 7 1 2 H2C CH CH2 H2C CH CH2 1 2 3 7 3 NиCH3 CO NиCH3 CHOH NиCH3 OH 6 6 4 H2C CH CH2 H2C CH CH2 5 5 4 Tropinone Tropine All the foregoing reactions of tropine are explained with the above structure as mentioned below:  Formation of 2-ethyl pyridine from tropine HI [HI] HCl Zn NиMe OH NиMe I NиMe Distil MeCl ϩ NH N CH2CH3 Tropine Dihydrotropidine Nordihydrotropidine 2-Ethylpyridine (tropane)Chapter-05.indd 175 6/29/2012 10:19:13 AM
    • 176 Pharmaceutical Chemistry of Natural Products  Formation of tropinone and tropinic acid from tropine PhCHO CHPh CH2CO2H NиMe OH NиMe O NиMe NиMe O CO2H CHPh Tropine Tropinone Tropinic acid Dibenzylidenetropionone  Formation of tropilidene (cycloheptatriene) from tropine (i) MeI H2SO4 (ii) AgOH (i) MeI NиMe OH ϪH2O NиMe NиMe2 ϩ OH – Vacuum (ii) AgOH distil (iii)Vacuum distil Tropine Tropilidene NMe2  Formation of pimelic acid from tropinic acid CH2CO2H CH2CO2H CHCO2H CHCO2H (i) MeI (i) MeI NиMe Heat (ii) AgOH (ii) AgOH ϩ NиMe2OH Ϫ Na–Hg CO2H (iii)Heat CO2H CO2H CO2H CO2H CO2H Tropinic acid Pimelic acid NMe2 The proposed structure of tropine is confirmed by the synthesis. Willstatter’s synthesis: (i) HI KOH Me2NH O(ii) [HI] Br2 I C2H5OH Br Br NMe2 Suberone Cycloheptene Exhaustive methylation NMe2 Br Br Quinoline Br2 Me2NH HBr 150°C (i) Na/EtOH Cycloheptatriene Cycloheptadiene (ii) Br2/HBr BrChapter-05.indd 176 6/29/2012 10:19:13 AM