Alkaloids Lecture Notes PDF

Extraction and Isolation of Alkaloids - In addition to the alkaloids, the plant contains several other constituents such as proteins, fats, waxes, Pigments and other constituents. - Therefore, isolation of alkaloids from alkaloid-bearing plants necessitates the following; General procedure i...

Extraction and Isolation of Alkaloids - In addition to the alkaloids, the plant contains several other constituents such as proteins, fats, waxes, Pigments and other constituents. - Therefore, isolation of alkaloids from alkaloid-bearing plants necessitates the following; General procedure involved in separation and isolation of alkaloids. I- Preparation of the sample II- Liberation of the free alkaloid bases with a suitable alkali. III- Extraction of the alkaloidal bases with an organic solvent. IV- Purification of the crude alkaloidal extract V- Fractionation of the total alkaloids. VI- Separation of the individual alkaloids as single pure compounds I- Preparation of the sample 1- The plant material should be first reduced to a moderately coarse or fine powder to facilitate extraction of the alkaloids. 2- Organs which are rich in fats and oils (such as seed) must be defatted by extraction with petroleum ether or hexane to avoid emulsion formation during extraction. II- Liberation of the free base - Alkaloids occur in the plant mostly as salts of acids, e.g. (Tannates, oxalates or meconates …etc) - The free base of alkaloids can be liberated before extraction with a suitable alkali. * Choice of the alkali depend on; 1- The chemical properties of the alkaloidal base: The use of strong alkali is not permissible in the following cases; a- Ester alkaloids  e.g. Atropine and Cocaine. b- Phenolic alkaloids  e.g. Morphine, Cephaline c- Alkaloids containing lactone ring Pilocarpine 2- The presence of fatty materials: Strong alkali cannot be used on fatty drugs as the soap formed on saponification causes the formation of troublesome emulsions. *Ammonium hydroxide is the most widely used alkali. This due to the following; a- It is sufficiently basic to liberate most common alkaloidal bases. b- It also has the advantage of being volatile and easily removed on evaporation of the solvent. N.B. *In case of the alkaloidal salt which is difficult to decomposed by the alkali as alkaloidal tannate in Cinchona bark.  They firstly ttt with dil. HCl  then  The hydrochlorides salt formed are ttt with suitable alkali  Alkaloidal base. III- Extraction of the alkaloidal bases. - Extraction of the alkaloidal bases from their plant material can be achieved by solvents of three different types. 1-Extraction with water-immiscble solvents - Chloroform, ether, methylene chloride and benzene may be used. - CHCl3 extracts all alkaloids with a variable degree. except the quaternary bases e.g. tubocurarine - The quaternary bases are more soluble in water than in organic solvent. 2-Extraction with water-miscible solvents. - Ethanol or methanol is used for extraction. - No pretreatment with alkali is required, due to the alcohols equally dissolve the alkaloidal salts and their free bases which present in plant. 3- Extraction with acidulated water - The drug is extracted with acidulated water  then made alkaline  alkaloids base extracted with immiscible solvent. - It is not selective because it dissolve large quantity of other materials present in plant as sugars, mucilage, coloring matter, proteins and tannins. IV-Purification of the crude alkaloidal extract. By: 1- Re-extracting by using suitable organic solvent after shaking with acid soln.  The acidic soln. is rendered alkaline with NH4OH. 2- Formation of alkaloidal complex with a suitable precipitating agent  Filteration, crystallization  Decomposition of complex to obtain the free alkaloids (eg. Picric acid complex decompose by NH4OH soln.) 3- Distillation can be used to separate volatile alkaloids from non-volatile ones. V- Separation of single pure alkaloids by different methods depending on their nature: as 1- Fractional crystallization 2- Gradient pH technique: - Provides a suitable methods for separation of alkaloids which varying in their basicity. - In this technique, the crude alkaloid mix. is dissolved in 2 % tartaric acid soln. Ext. with organic solvent  Ext. neutral and very weakly basic alkaloids). - The pH of the aqueous solution is then raised by a 0.5 pH increments to pH 9  where extracting at each pH with organic solvent. - This technique enable the gradual separation of alkaloids from weakly basic to strongly basic alkaloids.  Scheme for isolation of alkaloids Plant material Petroleum ether Marc Extract 1- MeOH or 95% EtOH 2- Concentrate 3- Partition between CHCl3 (EtOAc) and 2 % Tartaric acid. 2 % Tartaric acid CHCl3 (EtOAc) Neutral or weakly Basic Basify with ammonia Alkaloids Or Na carbonate CHCl3 (EtOAc) Basic aqueous soln. Strong Basic Alkaloids May contain Quaternary Alkaloids. Quantitative determination methods of alkaloids 1- Volumetric methods. 2- Gravimetric methods 3- Colourmetric and spectrophotometric method 4- UV and IR absorption methods 5- Fluorimetric methods 6- Polarimetric methods 7- Chromatographic methods Classification of alkaloids -There are many systems for the classification; 1- According to their biosynthetic precursor. Phenyl alanine - Alkaloids classified into 6 groups according to Tryptophane 6 amino acids precursors. Histidine 2- According to their occurrence in different families. 3- According to the nature of basic chemical structure Anthranilic acid which are derived from them. Lysine As, Some of the common nuclei of alkaloids are; Ornithine  Basic nucleus of Alkaloids N N N N H H N N N H N H Pyrrole Pyrrolidine Pyridine Pyrimidine Imidazole Piperidine N N CH3 N N Pyrrolizidine H Nor-hipinane Tropane Indole N NH N N N N N N H Quinoline Isoquinoline Purine Carboline  Common nucleus of Alkaloids H3C N CH3 N N N H H Hydroindole + Pyrrolidine (in Physostigma alkaloids) Isoquinoline + Phenanthrene N H (Aporphine) Indole + Hydroquinoline (in Ergot alkaloids) Phenanthrine CH2 H2C N Steroidal N Quinuclidine Tropolone (in Cinchona alkaloids) Phenanthridine - In our study Alkaloids will be classified according to their basic nuclei. I- Alkaloids of Phenylalkylamine group. - They are known as alkaloidal amines. - They don’t contain heterocyclic nitrogen atoms 1- Ephedrine - It occurs alone in Ephedra sinica and E. equisetina F. Gnetaceae (1-3 %) Or associated with nor-pseudoephedrine in E. vulgaris.(2.5-3 % Totai alkaloids) OH CH-CH-CH3 CH-CH-CH3 OH NH-CH3 D-ψ-Ephedrine NH-CH3 L-Ephedrine Structural similarity between Ephedrine and the - HO OH animal hormone adrenaline (Epinephrine) (Therefore they have similar pharmacological HO CH-CH2 action But Ephedrine has many advantages as NH-CH3 (effective orally and its action is more prolonged.) *Extraction of Ephedrine From plant material 1- By steam distillation. 2- By Ext. the plant material with benzene  shaking with HCl acid  L-ephedrine & pseudoephedrine are extracted and separated by; 1- Solubility in H2O (Ephedrine is much more soluble than Pseudoephedrine) 2- By their oxalates, Ephedrine oxalate is much less soluble in cold H2O than the other 3- By their hydrochlorides, Ephedrine HCl is less soluble in CHCl3 and alcohol. Therapeutic uses: 1- In treatment of bronchial asthma as bronchodilator. 2- In treatment of certain allergic disorders and as a nasal decongestant. ) CuSO4 test (Chen’s test:): Ephedrine solution in H2O + 1 drop of 5% CuSO4 + NaOH Purple colour Shake with ether Purple colour in the ethereal layer and the aqueous layer turns blue in colour  Cathe (Khat) Alkaloids Cathine [(+)-norpseudoephedrine] and Cathinone. b) Sources: Khat (Catha edulis, F: Celastraceae) and Ephedra vulgaris (F: Gnetaceae).  They act by inducing the release of catecholamines. They are CNS stimulants, decrease fatigue and produce a feeling of well-being (euphoria). Khat may cause addiction.  Ch. Test: Soln. + CuSO4 + NaOH Purple colour OH O H C C CH3 H H C C CH3 NH2 NH2 d-nor-Ψ-ephedrine (Cathine) Cathinone  Cathinone is an amphetamine - like stimulant causes excitement, a pleasuring effect, loss of appetite, drowsy hallucinations and euphoria. Negatibe effects such as Dilated pupils (mydriasis), increased heart rate and blood pressure Withdrawal symptoms that may follow prolonged khat use include lethargy, mild depression, nightmares, and slight tremor. In 1980, the World Health Organization classified khat as a drug of abuse that can produce mild to moderate psychological dependence. It is a controlled/illegal substance in many countries, but is legal for sale and production in many others. Cathinone is not very stable and breaks down to produce cathine and norephedrine. 3- Mescaline - It is present in Cactus lophophora, F. Cactaceae. Pharmacologically: it causes disturbance of normal mental functions causing euphoria and hallucination. 4- Capsaicin - Capsaicin is a phenolic compound present as extremely pungent principal in the fruit of Capsicum spp. F. Solanaceae - Capsicum imparts a pungent taste to water. - The pungency is destroyed by oxidation. - Capsaicin does not possess basic characters owing to the amide group  thus can be Capsaicin extracted with ether from acidic medium. *Therapeutic uses - Capsicum are used as counter-irritant to relief Rheumatism and Lumbago. Tests for Identity: 1- Alcohol solution + FeCl3 bluish green color. 2- Capsacine + H2SO4 + small amount of sugar violet colour.  II- Alkaloids of Tropolone group Colchicum autumnale, Liliaceae Colchicum alkaloids Colchicum seeds and corms are obtained from Colchicum autumnale (Fam. Liliaceae). The structures of colchicum alkaloids are characterized by the presence of a seven- membered pseudoaromatic unsaturated ring, referred to as "Tropolone nucleus". Ferric chloride test: Used to differentiate colchicine from colchiceine. FeCl3 garnet-red colour FeCl3 olive green colour Quantitative estimation of colchicine in plant material The green colour developed with colchiceine and FeCl3 used quantitatively for estimation of colchicine (hydrolyzed colchiceine).  Uses In treatment of gout (relief of pain and inflammation around joints). It appears to act primarily as an anti-inflammatory agent, and does not affect uric acid metabolism, which needs to be treated with other agents, e.g. a xanthine oxidase inhibitor such as allopurinol. Anticancer agent: (Colchicine binds to tubulin in the mitotic spindle, preventing polymerization and assembly into microtubules as do Podophyllotoxin and vincristine). Used as plant hormone to produce polyploidy in seedlings. High blood uric acid level Acute arthritis Breakdown of product monosodium urate stone in kidney of the body’s purine (nucleic acid) metabolism. ♂>♀ Familial metabolic disease  Idiopathic decrease in uric acid excretion (75%) Increase uric acid Aetiology production due Impaired uric acid excretion secondary of raised to increased to thiazide diuretics, uric acid cell turn over chronic Renal failure level (tumors), increase uric acid synthesis High dietary purine intake  II- Pseudoalkaloid A- A modified diterpene (Taxol) Source: Bark of Taxus species  (Yew tree) e.g. Taxus brevifolia F. Taxaceae. It is isolated from the stem bark of these trees in very small amounts: -About three trees are needed to provide enough drug to treat one cancer patient. -The bark from 4000 trees is needed for the production of 1 kg of taxol. It is used for treatment of ovarian cancer. Clinical trials have proved its effectiveness in lung cancer, breast cancer, malignant melanoma and other tumors. It induces the polymerization of the protein Tubulin leading to formation of stable non-functional microtubules (that are unable to support mitosis).  B- Steroidal alkaloids 1- Solanum alkaloids Several alkaloids occur in different Solanum species as Solanum nigrum, S. tubersum; Family Solanaceae. 1- Solanine  2- Solasonine Hydrolysed by mineral acids Test for identity 1. Solanum alkaloids + hot amyl or ethyl alcohol cooling jelly (characteristic for solanum alkaloids). 2. Antimony trichloride solution red colour. 3. Colourimetric assay; alkaloidal sol. + H2SO4 red colour (aldehyde formation). Uses Solanum alkaloids used as precursor to produce steroidal hormones