TERPENOIDS

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1. TERPENOIDS SUBMITTED BY: D.SREE RAMA LAKSHMI Dept. Ph. Chemistry GUIDENCE BY: K.VIJAYA KISHORE M.Pharm.(Ph.D) ACHARYA NAGARUNA UNIVERSITY

2. CONTENTS Introduction General proprties Isolation Classification Isoprene rule General methods of structural elucidation Structural elucidation of some drugs

3. INTRODUCTION The term terpene was given to the compounds isolated from terpentine, a volatile liquid isolated from pine trees. Terpenes are the hydrocarbons having the general formula (C H8)n. The term terpenoids represents these hydrocarbons as well as their hydrogenated and dehydrogenated derivatives. Therefore,terpenoids have the general formula (C5)n. Thus, all terpenes are terpenoids but all terpenoids are not terpenes. There are many different classes of naturally occuring compounds. Terpenoids also form a group of naturally occuring compounds majority of which occur in plants, a few of them have also been obtained from other sources. Terpenoids are volatile substances which give plants and flowers their fragrance. They occur widely in the leaves and fruits of higher plants, conifers, citrus and eucalyptus.

4. GENERAL PROPERTIES Physical properties: Terpenoids are colourless liquid. Soluble in organic solvents and insoluble in water. Most of the terpenoids are optically active. Volatile in nature. Boiling point 150° – 180° C. Chemical properties: The are unsaturated compounds. They undergo addition reaction with hydrogen, halogen acids to form addition products like NOCl, NOBr and hydrates. They undergo polmerization and dehdrogenation in the ring. On thermal decomposition, terpenoid gives isoprene as one of the product .

5. ISOLATION OF TERPENOIDS Isolation of essential oils from plant parts method a) Steam distilation b) Solvent extraction c) Maceration d) Adsorption in purified fats/ Enfluerage ii) Separation of terpenoid from essential oils a) Chemical methods b) Physical methods

6. I. ISOLATION OF ESSENTIAL OILS STEAM DISTILLATION

7. b ) SOLVENT EXTRACTION Solvents like hexane and ethanol is used to isolate essential oils. It is used for the plant parts have low amount of essential oil . Plant material are treated with the solvent, it produces a waxy aromatic compound called a "concrete.“ Then it mixed with alcohol, the oil particles are released. Then it passess through a condenser then it separated out. This oil is used in perfume industry or for aromatherapy purposes

8. c) MACERATION Advantage: More plant’s essence is captured. In this method the plant material is converted into moderately coarse powder. Plant material is placed in a closed vessel . To this solvent is added. The mixture is allowed to stand for 1 week, then the liquid is strained. Solid residue is pressed to recover any remaining liquid. Strained and expressed liquids are mixed

9. d) ENFLEURAGE The fat is warmed to 50 C on glass plates. Then the fat is covered with flower petals and it kept for several days until it saturated with essential oils. Then the old petals are replaced by fresh petals ,it repeated. After removing the petals, the fat is treated with ethanol when all the oils present in fat are dissolved in ethanol. The alcoholic distillate is then fractionally distilled under reduced pressure to remove the solvent. Recently the fat is replaced by coconut charcoal, due to greater stability and higher adsorptive capacity.

10. II SEPARATION OF TERPENOID FROM ESSENTIAL OILS a ) CHEMICAL METHOD: Essential oils containing terpenoid hydrocarbon + nitrosyl chloride in chloroform form crystalline adduct of hydrocarbons. Essential oil containing alcohols

11. Terpenoid containing aldehyde and ketone treated with NaHSO 3 , phenyl hydrazine or semicarbazone. After separation it is decomposed to get terpenois. b) PHYSICAL METHOD : Fractional distillation Chromatography

12. Fractional distillation : During fractional distillation of essetial oils, monoterpenoid hydrocarbons get initially distilled followed by their oxgenated derivatives. Distillation of residue under reduced pressure gives sesquiterpenoids.

13. Chromatography In this, essential oils are allowed to flow through alumina/silica which is used as an adsorbent with the principle of separation being adsorption chromatography. Different classes of terpenoids show different chromatograms. These are again subjected to chromatography where the individual terpenoids finally get separated. Vapour phase/gas chromatography, partition chromatography, counter current separation are commonly used for the separation of individual terpenoids.

14. ISOPRENE RULE In 1887, Wallach proposed the isoprene rule . “ It states that the skeleton structures of all terpenoids are built up of isoprene units or 2-methyl 1,3-butadiene”. CH 2 = CH( CH 3 ) -CH=CH 2 The isoprene rule derived from the following facts: The empirical formula of almost all terpenoids is C 5 H 8. The thermal decomposition of all terpenoids gives isoprene as one of the products. Eg: Rubber on destructive distillation yields isoprene as the products.

15. The isoprene rule has been confirmed by the following facts: Isoprene, when heated to 280 C yield a (dipentene). Isoprene may be polymerized to yield a rubber like product Polymerisation ( C 5 H 8 ) n ( Rubber polyterpenoid) n C 5 H 8

16. SPECIAL ISOPRENE RULE This rule proposed by Ingold in 1925. According to this rule “the isoprene units in terpenoids are joined by head to tail linkage or 1,4- linkage ( The branched end of isoprene unit was considered as head and other end as the tail).

17. Violations of isoprene rule Carbon content of certain terpenoids are not a multiple of five. Eg: Cryptone, a naturally occurring ketonic terpenoid contains nine carbon atoms , it cannot be divided into two isoprene units. Cr y pto n e

18. In certain terpenoids isoprene rule is violated. Eg: Lavandulol is composed of two isoprene units are linked through C 3 and C 4.

19. CLASSIFICATION OF TERPENOIDS The terpenoids have general formula (C 5 H 8 ) n . Based on the value of ‘n’ the terpenoids are classified into following:

20. Terpenoids are classified based on the number of rings present in the terpenoids. Acyclic terpenoids Monocyclic terpenoids Bicyclic terpenoids Tricyclic terpenoidsc Tetracyclic terpenoids

21. 1) MONOTERPENOIDS i) Acyclic monoterpenoids: ii) Monocyclic monoterpenoids:

22. iii) Bicyclic monoterpenoids: The size of the first ring (six membered) in terpenoid is same in all these terpenoids but the size of second ring is varies. On the basis of the size of second ring, bicyclic monoterpenoids are further divided into three classes . a) It co n taining 6+ 3 - m e m bered ri n gs ( E g :Caran e ) b) Itcontaining 6+4- membered rings (Pinane)

23. Contining 6+5-membered rings Bornane derivatives – Camphane. Norbornane derivative – Isocamphane, Fenchane,Isobornylane. α - pinene Ca m p h or

24. 2) SESQUI TERPENOIDS Acyclic sequiterpenoid Eg: Farnesol ii) Monocyclic sesquiterpenoid Bisabolene Zinziberene

25. iii) Bicyclic sesquiterpenoids Cadinene iv ) Tricyclic sesquiterpenoids cedrol cedrene

26. 3 ) DITERPENOIDS i) Acyclic diterpenoids Phytol ii) Monocyclic diterpenoids Vitamin A iii) Dicyclic diterpenoid Eg : Agathic acid

27. iv ) Tricyclic diterpenoid Abietic acid

28. 4) TRITERPENOID i) Acyclic triterpenoid Squalene ii ) Tricyclic triterpenoid Eg: Ambrein

29. iv ) Pentacyclic triterpenoid iii) Tetracyclic triterpenoid Lan o sterol A m y rin

30. GENERAL METHODS OF STRCTURAL ELUCIDATION OF TERPENOIDS Analytical method Synthetic method Physical method Molecular rearrangement Synthesis

31. 1) ANALYTICAL METHOD Molecular formula Nature of the oxygen atom Unsaturation Number of rings Oxidative degradation products Dehydrogenation

32. a) Molecular Formula Qualitative method Quantitative method Mass spectroscopy

33. b) NATURE OF OXYGEN ATOM i) Hydroxyl group ROH + ( CH 3 CO) 2 O ROCOCH 3 + CH 3 COOH Acetate

34. Nature of hydroxyl group is revealed by the rate of esterification. Primary alcohols undergo esterification more readily than secondary and tertiary alcohols. ii) Carbonyl group: Carb o n y l gro u p : Alde h y de or Keto n e

35. iii) –CH 2 CO- groups: Terpenoids form oximes with nitrous acid and benzylidene derivative with benzaldehyde

36. iv) C- alkyl group The important C- alkyl group is C- CH 3 group. It is determined by Kuhn-Roth method iii) Carboxyl group If terpenoid soluble in NH 3 and gives effervescence with NaHCO 3 , it indicate the presence of –COOH group. Number of –COOH group is estimated by titration against a standard alkali. Whether the –COOH group is attached to a 1 , 2 or 3 carbon atom is ascertained from the esterification of acids in the following order. Tertiary ˂ secondary ˂ Primary

37. c ) Unsaturation It is determined by the formation of addition products with reagents like hydrogen, halogen, halogen acids, per acids and nitrosyl chloride. Eg: Cadinene undergo hydrogenation to form tetrahydro cadi n en e , it i n dicate t h at cadinene co n tains 2 d o u b le b o n d.

38. d) Number of rings The number of rings is determined from the following table showing the relation between general formula of compound and types of compounds.

39. The molecular formula of citral is C 10 H 16 O , it contain 2 double bonds and one oxygen atom as carbonyl group. Molecular formula of parent hydrocarbon is C 10 H 16 O ≡ C 10 H 16 + 4H (for 2 double bond) + 2H (for carbonyl oxygen) ≡C 10 H 22. The molecular formula C 10 H 22 corresponds to C n H 2n+2 (general formula of acyclic terpenoid), so citral is an acyclic terpenoid

40. e) Oxidative degradation products Ozone: Terpenoid react with ozone to form ozonide it undergo decomposition either hydrolysis or catalytic reduction yields carbonyl compounds . Nitric acid react with nitric acid to form aromatic acid and aliphatic T erpenoids acid.

41. f) Dehydration Terpenoid containing alcoholic or ketonic groups are heated with dehydrating agents (potassium bisulphate, zinc chloride) to form simple aromatic compound with loss of water. g) Dehydrogenation α-terpeneol to dipentene.

42. 2) SYNTHETIC METHOD 1) Catalytic Hydrogenation When aromatic compounds undergo catalytic hydrogenation to form synthetic terpenoids. Eg: Menthol is prepared from thymol an aromatic compound by catalytic hydrogenation

43. 2) Grignard reactions In grignard reagent, methy or isopropyl groups are introduced into compound having carbonyl groups to synthesise large number of terpenoids. 3) Reformatsky reactions In this reaction - halogen substituted ester is treated with a carbonyl compound to form - hydroxyl ester. It is then treated with dil.acid yield - hydroxyl acid which further coverted to an unsaturated acid or a hydrocarbon.

44. 3) PHYSICAL METHOD UV spectroscopy : It is used for the detection of conjugation in terpenoids IR spectroscopy : Used for detecting the presence of a hydroxyl group, an oxo group. Used for distinguish between cis and trans isomer. Used for quantitative measurements (determination of no: of methyl group). NMR spectroscopy : Used for identifying double bonds and determing the nature of endgroups in terpenoid. No: of rings present in terpenoid Orientation of methyl group in terpenoid Presence of –OH group

45. 4) MOLECULAR REARRANGEMENT Molecular rearrangement is used when the degradation reaction gives various products. 5) SYNTHESIS Structure elucidated by the above physical and analytical method is confirmed by its synthesis .

46. STRUCTURAL ELUCIDATION OF CITRAL Constitution of citra l Molecular formula: C 10 H 16 O Presence of two double bond: Citral is treated with bromine or hydrogen, it forms citral tetrabromide. It indicate the presence of two double bond. C 10 H 16 O Br 2 C 10 H 16 O.Br 4

47. Citral o n o z o n ol y s i s y ie l d ac e tone, l aev u lal d e h y de and g y oxal. It indicate that citral is an acyclic compound containing two double bond. c) Presence of an aldehyde group: Formation of an oxime with hydroxylamine indicates the presence of an oxo group in citral. Citral on reduction with Na/Hg it gives an alcohol called geraniol and on oxidation with silver oxide to yield a Geranic acid with same number of carbon atom as citral. Indicate that oxo group in citral is an aldehyde group.

48. d) Citral as an acyclic compound: Formation of above products shows that citral is an acyclic compound containing two double bonds. Corresponding saturated hydrocarbon of citral (molecular Formula C 10 H 22 ) corresponds to the general formula C n H 2n+2 for acyclic compounds, indicating that citral must be an acyclic compound . e) Carbon skeleton of citral Citral is heated with potassium hydrogen sulphate, it gives p- cymene (known compound). F o r m ation o f p -c y m ene a nd product obtained fr o m the oz o n o l y s is reveals that C-skeleton (I) of citral is formed by the joining of two isoprene units in the head to tail fashion. Formation of p-cymene also reveals the position of methyl and isopropyl group in citral.

49. f) Oxidation Citral undergo oxidation with KMnO 4 followed by chromic acid yield acetone, oxalic acid and laevulic acid. These reactions are only explained if the citral has structure (II).

50. Support for the structure (II) Verley found that citral on boiling with aqueous potassium carbonate yielded 6-methyl hept-5-ene-2-one and acetaldehyde. The formation of these can only be explained on the basis of proposed structure of citral (II) if it undergoes cleavage at α,β- double bond. Further methylheptenone undergo oxidation yields acetone and laevulic acid.These can be only explained on the basis of structure (II).

51. Confirmation synthesis of citral by Barbier-Bouveault-Tiemann’s synthesis In this synthesis methyl heptenone is converted to geranic ester by using Reformatsky’s reaction. Geranic ester is then converted to citral by distilling a mixture of calcium salts of geranic and formic acids.

52. Isomerism of citral Two geometrical isomers occur in nature Two isomers are differ in the arrangement of aldehyde group about double bond in 2,3 position. One is cis-citral or Neral and other is trans- citral or geranial.

53. STRUCTURAL ELUCIDATION OF MENTHOL Molecular formula: C 10 H 20 O Menthol forms esters readily with acids it means that it possess an alcoholic group. Menthol then oxidized to yield ketone, menthone (C 10 H 18 O) it indicate that the alcoholic group is secondary in nature. 3) On dehydration followed by dehydrogenation it yields p -cymene. It indicate the presence of p -cymene skeleton (p-menthane skeleton) in two componds.

54. 4) Menthone on oxidation with KMnO 4 yields ketoacid C 10 H 18 O 3. It possess one keto group and one carboxyl group and is called ketomenthylic acid. It readily oxidized to 3-methyladipic acid. These reactions can be explained by considering the following structure of menthol.

55. Menthol was converted to p -Cymene [1-methyl-4- isopropylbenzene], which was also obtained by dehydrogenation of pulegone. Pulegone on reduction yields menthone which on further reduction gives menthol.

56. SYNTHESIS Finally the structure of menthone and menthol have been confirmed by the synthesis given by Kotz and Hese from m-cresol.

57. STRUCTURAL ELUCIDATION OF CAMPHOR CONSTITUTION OF CAMPHOR- Molecular formula - C 10 H 16 O. Presence of keto group It form oxime with hydroxylamine When camphor is distilled with iodine it yields cavacrol. I 2

58. Presence of –CH 2 CO group. When camphor is treated with amyl nitrite and hydrochloric acid, it yields iso nitroso camphor Presence of six membered ring

59. 6. Nature of carbon frame in camphor. when camphor is oxidized with nitric acid, it yields a crystalline dibasic acid , camphoric acid as a camphoric acid possesses the same number of carbon atom as camphor , it means that group must be present in one of the ring of camphor. Further camphoric acid is dicarboxlic acid and its molecular refraction reveals that it is also saturated. Thus during the conversion camphor into camphoric acid , there occur the opening of ring containing the keto group and therefore camphoric acid must be monocyclic compound. When camphoric acid is further oxidized with nitric acid , camphoric acid is obtained.

60. STRUCTURAL ELUCIDATION OF PHYTOL Introduction:- It is a kind of diterpene which comes under the “acyclic diterpene” category. Phytol is an acyclic diterpene alcohol and a constituent of chlorophyll. It is obtained from alkaline hydrolysis of chlorophyll, which is then converted to phytanic acid and stored in fats. It is commonly used as a precursor for the manufacture of synthetic forms of vitamin E and vitamin K1. It is an optically active compound which boils at 145°C at 0.03mm pressure. Molecular Formula: C 20 H 40 O Melting Point: < 25 °C PHY T OL

61. STRUCTURAL ELUCIDA TION Molecular formula: C 20 H 40 O Presence of double bond : When it is catalytically hydrogenated, it adds on one mole of hydrogen to form dihydrophytol indicating that phytol contains one double bond. Presence of primary alcoholic group : Phytol on oxidation with chromic acid yields monocarboxylic acid called phytenic acid which has same no. of C- atom indicating the presence of primary alcoholic group. Ozonolysis of phytol : on ozonolysis it yields glycoaldehyde and a saturated ketone

62. Structure of saturated ketone may be written as follows : Structure of saturated ketone is confirmed by its synthesis from ketone(I):

63. STRUCTURAL ELUCIDATION OF RETINOL It is a kind of diterpene which comes under the “Monocyclic diterpene” category. It is also called Vitamin A Vitamin A is the fat soluble vitamin, is a group of unsaturated nutritional organic compounds that includes retinol, retinal, retinoic acid, and several provitamin A carotenoids (most notably beta- carotene). All forms of vitamin A have a beta-ionone ring to which an isoprenoid chain is attached, called a retinyl group . Molecular Formula: C 20 H 30 O

64. STRUCTURAL ELUCIDATION Molecular formulae : C 20 H 30 O Double bond present: It consumes 5 hydrogen molecules during hydrogenation in presence of Pd catalyst, that means 5 double bonds are present in the structure.

65. Isoprene Units Confirmation: The oxidation of Vit.A with Pot. Permagnate gives 2 molecules of Acetic acid which indicates 2 Isoprene units are present in structure. Methyl group: The oxidation of Vit.A in the presen e of CrO3 which gives 3 molecules of Acetic acid. It means, 3 methyl groups are present.

66. Hydroxy(–OH) group: presence of –OH group can be determined by the formation of acetates with acetic anhydride. Upon Oxidation, Retinol converts to Retinal(aldehyde) and then converts to Retinoic acid. It means, there is primary alchohol present in structure.

67. Beta-Ionone Nucleus: Ozonolysis of retinol gives geronic acid which can directly obtained by ozonolysis of beta-Ionone.It confirms the basic nucleus of beta-Ionone is present in structure.

68. STRCTURAL ELCIDATION OF TAXOL Introduction : It is a type of Complex diterpene. Taxol, a diterpenoid natural product first isolated from Taxus brevifolia, is one of today’s better known anticancer drugs. The paclitaxel molecule consists of a tetracyclic core called baccatin III and an amide tail. The core rings are conveniently called (from left to right) ring A (a cyclohexene), ring B (a cyclooctane), ring C (a cyclohexane) and ring D (an oxetane). MOLECULAR FORMULAE – C 47 H 51 NO 14

69. Nature of O atom : Presence of alcoholic group :

70. Nature of N-atom :

71. SYNTHESIS :

72. STRUCTURAL ELUCIDATION OF SQUALENE Squalene is a natural 30-carbon organic compound originally obtained for commercial purposes primarily from shark liver oil (hence its name, as Squalus is a genus of sharks), although plant sources (primarily vegetable oils) are now used as well, including amaranth seed, rice bran, wheat germ, and olives. Yeast cells have been genetically engineered to produce commercially useful quantities of "synthetic" squalane, which is similar to squalene.

73. STRUCTURAL ELUCIDATION Molecular formulae: C 30 H 50 Presence of double bonds : The molecular formulae of fully saturated squalene was found to be perhydrosqualene with 6H₂ molecule. Absence of conjugated double bonds in squalene: Squalene fails to undergo reduction whwn treated with Na – metal amd amyl alcohol which indicates absence of conjugated double bonds Oxidation of squalene with chromyl chloride: On oxidation with CrO₂Cl₂ in CCl₄ gives Formaldehyde , Acetaldehyde ,Succinic acid Ozonolysis of squalene :

74. STRUCTRAL ELUCIDTION OF CROTENOIDS Carotenoids are the group of non-nitrogenous , yellow , red or orange pigments that are universally distributed in living things. These are also called tetraterpenoids , that are produced by plants and algae as well as several bacteria and fungi. There are over 600 known carotenoids They split into 2 classes xanthophyll and carotenes Tetraterpenoids contain 40 C atoms General structure of carotenoid is a polyene chain consisting of 9-11 double bonds and possibly terminating in rings

75. ALPHA & BETA CAROTENOIDS About 600- 700 different carotenoids are known of which α & β carotene are the most prominent Β carotene is the most known carotenoid and the most often naturally occurring carotene also known as provitamin A