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ALKALOIDS

Alkaloids are a chemically heterogenous group of basic nitrogen containing substances found predominantly in higher plants and also occur in lower plants, animals, microorganisms and marine organisms. Alkaloids usually contain one or two nitrogen atoms although some like ergotamine may contain up to five nitrogen atoms. The term alkaloid was coined in 1819 by the pharmacist Meissner and meant simply alkali-like molecules.' True Alkaloids ' were defined as compounds meeting the additional four qualifications namely: I) Nitrogen is a part of heterocyclic ring Ii) The occurrence of compound is restricted to plant kingdom Iii) The compound has complex molecular structure Iv) The compound manifests significant physiological activity Their manifold pharmacological activities have always excited man's interest, and since early times selected plant products have been used as poisons, euphoriants, psychedelics, stimulants or as medicines. Many of our modern drugs now contain the same compounds or synthetic analogues, and the pharmacological and toxicological properties of these compounds are thus of immense interest and importance.

1.Classification and Structural types


Some of the methods for classification of alkaloids as follows: i) Biosynthetic Origin This method depends on types of precursors or building block compounds used by living organisms to synthesize complex structures. The disadvantage with this method is that the relationship of alkaloids to each other and to their precursors is not always apparent. ii) Pharmacological This classification is based on pharmacological activity. iii) Taxonomic This is done according to the source of compound without reference to the chemical types. iv) Chemical The chemical classification of alkaloids is universally adapted and depends on the type of heterocyclic ring structure present. It is usual to classify alkaloids according to the amino acids (or their derivatives) from which they arise. Thus the most important classes are derived from the amino acids ornithine and lysine; or from the aromatic amino acids phenylalanine and tyrosine; or from tryptophan and a moiety of mavelonoid origin; and a number of compounds are also derived from

anthranilic acid or from nicotinic acid. HETEROCYCLIC RING SYSTEMS

N H pyrrolidine

N H pyrrole

N H piperidine

N pyridine

N quinoline

N isoquinoline indole
H

N H

N H dihydroindole

N quinolizidine

N pyrrolizidine tropane

N N

N N H
C C N

benzylisoquinoline

purine

-phenylethylamine

2. Qualitative tests and general properties


Alkaloids are generally bitter in taste and are optically active.This property is useful for resolution of racemic acids. The alkaloids are mostly crystalline,colourless,non-volatile solids,which are insoluble in water , but soluble in polar organic solvents (methanol,chloroform,ether etc.).Some alkaloids, however,are watersoluble liquids (for example coniine,hygrine,nicotine).Few alkaloids are coloured(for exemple berberine is yellow).Nearly all basic alkaloids form salts with inorganic acids,which are soluble in water and insoluble in organic solvents.The knowledge of solubility of alkaloids and their salts is utilized in their isolation. Alkaloids yield precipitates or characteristic colours when reacted with solutions of molybdic acid,picric acid,chloroplatinic acid,potassium mercuric iodide etc.Therefore the presence of an alkaloids in a plant is detected by using various reagents such as Mayer's reagent (potassium mercuric iodide),Wagner's reagent (iodine dissolved in potassium iodide),Dragendorff's reagent (potassium bismuth iodide),Hager's reagent (saturated solution of picric acid in water),chloroplatinic acid (H2PtCl6), Frohde's reagent (molybdic acid) etc. These precipitates may be amorphous or crystalline and are of various colours:cream(Mayer's),yellow(Hager's),reddish-pinkish-brown(Wagner's and Dragendorff's ) etc.Caffeine and some other alkaloids do not give these precipitates.

3. Isolation and purification


The knowledge of the solubility of alkaloids and their salts is also of considerable pharmaceutical importance.Not only are alkaloid substances often administered in solution but also the differences in solubility between alkaloids and their salts provide methods for the isolation of alkaloids from the plant and their separation from the non-alkaloid substances.For many alkaloids are basic and occur as salts of 2hydroxybutane-1,4--dioic acid(malic acid),or of 1,3,4,5tetrahydroxycyclohexane(quinic acid).They can thus be extracted into acid solution using aqueous HCl,tartaric,or citric acids.Essentially neutral alkaloids like colchicine or piperine,which are in fact amides ,remain in the organic phase ,while most other alkaloids are isolated after basification and extraction into ethyl acetate. The almost invariably subsequent purification of the crude alkaloid mixtures is effected by chromatography using silica or alumina,and then recrystallization of the partially purified compounds from solvent systems like aqueous ethanol ,methanol/chloroform,or methanol/acetone. With few exceptions salts of alkaloids are water-soluble.The following

procedure is used for alkaloids isolation. The powdered plant material is first extracted with petroleum ether for the removal of fats.In a general procedure recommended by Manske the material then is extracted with methanol or ethanol and the dark solution is separated from cellulose and other insoluble materials and evaporated to dryness.Water is added to the residue ,the mixture is acidified to pH 2 and extracted with ethyl acetate or ether for the removal of non-basic molecules. The aqueous solution is basified to pH 10 and extracted with ethyl acetate or ether and organic layer evaporated to dryness to obtain a mixture of water insoluble organic bases.These alkaloids are further purified by fractional crystallization of salts.Alternatively the purification is achieved by using column chromatography or counter-current distribution method. Water-soluble alkaloids are obtained from aqueous solution by the precipitation with ammonium Peinecke's (H[Cr(NH3)2(SCN)4]) solution followed by ion exchange chromatography of the precipitated salt.The freeze-drying of the eluent gives the water-soluble alkaloids. Isolation and purification of an alkaloid from a plant material may not always be a simple procedure as described above because some alkaloid bearing plants contain a complex mixture of several alkaloids.Therefore,the isolation of a pure alkaloid from these plants may become extremely laborious procedure sometimes due to the repeated use of chromatography or crystallizations. Also,in rare instances stream distillation can be employed,for example with low molecular weight or volatile liquid alkaloids such as nicotine ,sparteine and coniine are most conveniently isolated by distillation.An aqueous extract of plant is made alkaline with caustic soda or sodium carbonate and alkaloid is distilled off with steam.The distillate is extracted with solvent to isolate the desired alkaloid.

4.General methods of structure determination


In structure determination of alkaloids,a variety of general chemical methods and more recently physical methods are employed.Many of the following chemical methods have been particularly useful in alkaloids chemistry. In general,elemental composition is obtained from combustion analysis and after determination of molecular weight,molecular formula is calculated.The measurement of optical rotation indicates the presence of optical activity. The alkaloids mostly contains one or more oxygen atoms,which may be present as hydroxyl,methoxy,methylenedioxy,carbonyl ,carbonyl ester,lactone,amide,lactam,epoxide groups or ether linkage.The nature of oxygen atom is ascertained using the following usual chemical reactions.

4.A. Chemical Methods i) Hydroxyl group The acetylation (acetic anhydride, pyridine) or benzoylation (benzoyl chloride,,pyridine) indicates the presence of hydroxyl group together with amino group (-NH or -NH2) in the molecule.If molecule contains hydroxyl group or -NH group then the number of these groups can be estimated by acetylation or Zerewitinoff's method.In the former method, the acetate derivative is hydrolyzed with known volume of 1N NaOH and the amount of base required for hydrolysis (estimated by titration) indicates the number of hydroxyl and any amino groups.In the latter method,hydroxyl and amino groups are estimated by the treatment with methyl magnesium iodide.The volume of methane obtained is measured, which indicates the number of -OH (and any N-H) groups.(One mole of alkaloid containing one -OH or -NH group = 22.4 liters of CH4 at S.T.P). i) Acetylation R-OH + CH3-CO-Cl R-OCO-CH3 R-NH-R1 + CH3-CO-Cl R-N(COCH3)-R1 ii) Zerewitinoff's method R-OH + MeMgI R-OMgI + CH4 R-NH-R' + MeMgI R-N(MgI)-R' + CH4 If hydroxyl group is present it may be alcoholic or Phenolic.Phenolic compounds are soluble in sodium hydroxide and are reprecipitated by carbon dioxide.They give colouration with ferric chloride.If the alkaloid does not respond to thede tests for phenol, the hydroxyl group may be alcoholic. If alcoholic group is present, then the nature of this group (usually secondary,tertiary) is verified by oxidation or by dehydration to unsaturated compound. ii) Carbonyl group The presence of carbonyl group is ascertained by usual reactions with hydroxylamine, semicarbazide or 2,4-dinitrophenyldrazine when the corresponding oxime,semicarbazone or 2,4-dinitrophenyldrazine is formed.The carbonyl group may be present as an aldehyde or a ketone. This distinction can be made from Tollen's reagent.Aldehydes give silver mirror whereas ketones are unreactive. iii) Carboxyl group The solubility of an alkaloid in bicarbonate or ammonia and reprecipitation with carbon dioxide indicates the presence of carboxyl group.The number of carboxyl groups may be determined volumetrically by titration against a standard solution of barium hydroxide using

phenolphthalein as an indicator. The formation of ester on treatment with alcohol in the presence of dehydrating agent also indicates the presence of carboxyl group. iv) Methoxy group The detection and estimation of methoxy group can be achieved using Zeisel's method,which is similar to the Herzig-Meyer method for Nmethyl group determination.In this method aknow weight of alkaloid is boiled (120) with hydroiodic acid,which cleaves methoxy group with the formation of methyl iodide,this is then estimated by absorption in ethanolic silver nitrate and the precipitated silver iodide is filtered,dried and weighted.From the weight of silver iodide ,the number of methoxy group can be calculated.
R-OMe + HI 126 AgNO3 R-OH +MeI AgI

(Estimate ppt gravietrically)

v) Methylenedioxy group (-O-CH2-O-) If an alkaloid,on heating with hydrochloric or sulfuric acid yields formaldehyde,the presence of methylenedioxy group is indicated.Formaldehyde thus obtained can be estimated gravimetrically after conversion to dimedone derivative.
O
O
O

+ HCl

HCHO

Dimedone derivative (estimated gravimetrically) vi) Amide, lactam, ester,lactone groups These groups can be detected and estimated through acid or alkaline hydrolysis. Amide -CONH2 + NaOH -COONa + NH3 Ester -COOR + NaOH -COONa + ROH Lactone
(CH2)n + NaOH
O O OH COONa

(CH2)n

Lactam

(CH2)n + NaOH
HN O NH2 COONa

(CH2)n

vii) Epoxide and ether linkage Epoxide or ether linkages are cleaved by the addition of hydrogen bromide or hydroiodic acid. viii) Nature of nitrogen In the majority of alkaloids nitrogen atom(s) is present in heterocyclic ring therefore it can be secondary or tertiary. The acetylation or benzoylation can distinguish tertiary amine from secondary amine,the former being inert whereas the latter gives acetate or benzoate derivative.This distinction can also be done by treatment with HNO2 or methyl iodide or oxidation with 30% hydrogen peroxide. The presence of N-methyl group is often detected by distillation of amine with sodalime or estimated by the treatment with hydroiodic acid at 150-300 and conversion of methyl iodide produced to silver iodide as mentioned for estimation of methoxy groups. Secondary amine >N-H + HNO2 >N-NO+H2O >NH + CH3I >N-Me + HI Tertiary amine >N + CH3I >N+ -Me I>N + H2O2 >N+-O-+ H2O N-Methyl group >N -Me + CaO CH3NH2 >N -Me + HI N-H + MeI AgI AgNO3 ix) Tertiary methyl group The presence of C-Me group is quantitatively estimated by Kuhn-Roth oxidation (K2Cr2O7/H2SO4) to acetic acid,which is distilled off and titrated against standard base. -C-Me + K2Cr2O7/H2SO4 CH3COOH 4.B. Degradation of alkaloids The following methods are used to find out structural fragments of alkaloid molecules: i. Hofmann exhaustive methylation. ii. Emde's degradation.

iii. von Braun's method. iv. Hydrolysis. v. Alkali fusion. vi. Dehydrogenation. Now we discuss the mainly used methods of Hofmann degradation and Emde's degradation. i) Hofmann degradation The alkaloids is converted into the quaternary ammonium salt by the treatment with excess of alkyl halide and the resulting salt is then converted into the more basic hydroxide by reacting with silver oxide.subsequently the quaternary hydroxide salt is heated at 200 to give an olefin with the elimination of a tertiary amine and water molecule. R-CH2-CH2-N+R3 + OH- R-CH=CH2 + H2O +NR3 The elimination generally proceeds by an E2 elimination in which the hydrogen and quaternary nitrogen groups are present in the transantiparallel configuration. Hofmann's degradation fails in the following cases: i. with tetrahydroquinoline ii. with aromatic heterocyclic rings iii. With compounds having no -hydrogen atom Thus the reaction fails in the case of derivatives of 1,2,3,4,tetrahydrodimethyl quinolium hydroxide,which on pyrolysis merely loses methanol without fission of ring.However,Emde degradation results in cleavage of ring. ii) Emde's degradation The alkaloid is converted to quaternary ammonium salt by refluxing with alkyl halide and the resulting salt is subjected to reductive cleavage by the treatment with sodium amalgam in alkanol or sodium in liquid ammonia or by catalytic hydrogenation.Tetrahydroquinoline and tetrahydroisoquinoline quarternary salts undergo Emde's degradation. 4.C. Physical Methods Recently physical methods are used,in conjunction with chemical reactione to elucidate structure of alkaloids.Such is now the level of sophistication of the associated instruments that it is possible to determine a structure in a matter of days given a few milligrams(or less) of a pure compound. Infrared spectrum gives information about many functional groups.Ultraviolet spectra are used to indicate the nature of unsaturation or aromatic rings. NMR spectroscopy is more versatile for detecting many function groups, the nature of protons,carbons, heterocyclic rings

etc.Mass spectral fragmentation gives the information about molecular weight and degradation of the skeleton. Single crystal X-ray analysis has offered means for determining or confirming stereochemistry as well as distinguishing between alternate structures that appear to fit well for a particular alkaloid.Further support for the stereochemistry can be obtained by using optical rotatory dispersion or circular dichroism studies.