CHEM 425a/552 Natural Products D. Pavia

1. CHEM 425a/552 Natural Products D. Pavia ANNOUNCEMENTS - Buff Problem due today (F Oct 5) • Quiz on Monday Oct 8 • heterocyclic rings and alkaloid classification by rings • New assignment: Olive Problem due Friday, Oct 12

2. b-PHENYLETHYLAMINES PROTOALKALOIDS

3. b-PHENYLETHYLAMINES Structural Element The simplest Class of Alkaloids Not considered alkaloids by some - structures are too simple and only one or two biochemical steps away from mainstream metabolic compounds. All are derived from the amino acids phenylalanine or tyrosine: FACT: most of the alkaloids come from amino acids tyrosine phenylalanine

4. MISTLETOE b-Phenylethylamine Tyramine

5. b-Phenylethylamines THE SIMPLEST ALKALOIDS found in tyramine mistletoe viscum album acacia spp. tyrosine b-phenylethylamine old cheese (Emmental) bacterial action PLP - CO2 Just one biochemical step away from phenylalanine or tyrosine! Decarboxylation by the coenzyme pyridoxyl-5’-phosphate (vitamin B6) phenylalanine

6. MSTLETOE I

7. Viscum album

8. EPHEDRA Ephedrine

9. Ephedrine Recently banned from organic weight-loss supplements! found in ephedra spp Extract of ephedra was used in the early american west as a cure for asthma (Mormon Tea). ephedrine Chinese medicine “Ma Huang” for hay fever. Sympathomimetic Amine “Organic” appetite suppressant. compare R=H amphetamine R= H Norepinephrine R = CH3 methamphetamine R = CH3 Epinephrine (adrenalin) neurotransmitter / hormone “speed” or crystal “meth”

10. UTAH Ephedra

13. BARLEY Hordenine

14. Barley Hordenum spp

15. Hordenine found in Mexican cactus (Anhalonium fissuratum) Barley (sprouting) ( Hordenum spp ) hordenine Mistletoe ( Phoradenron spp ) Where does this come from? See next section first.

16. SAdM Nature’s Methylating Reagent S-ADENOSYLMETHIONINE SAM

17. S-Adenosylmethionine folic acid N-formylfolic acid PLAY IT, SAM ! N-methylfolic acid methionine .. .. homocysteine SN2 cysteine ONE-CARBON METABOLISM ATP + .. S-adenosylmethionine

18. METHYLATION BY SAM S-adenosylmethionine + .. Good nucleophiles will displace the methyl group (SN2 reaction). :Nu The trivalent positively-charged sulfur makes a good leaving group. Sulfur becomes divalent (normal) and loses the charge. .. .. Nu The nucleophile has become “methylated”. CH3

19. S-Adenosylmethionine is the principal methylating agent in the secondary metabolism of plants. These methylations can be carried out : .. : O .. .. O CH3 .. SAM .. : O H .. .. .. CH3 CH3 NH2 NH N SAM SAM CH3 .. .. : : : : SAM ….. in some cases The enzymes that carry out these processes are fairly non-discriminating. Almost any substrate with an -OH of -NH group is fair game.

20. BACK TO HORDENINE BIOSYNTHESIS

21. Hordenine BIOSYNTHESIS * tyrosine hordenine 1 4 - CO2 PLP SAM SAM N-methyltyramine tyramine 2 3 1 2 3 4 Feeding Experiment AMOUNT OF RADIOACTIVITY 14C-labelled tyrosine fed to barley sprouts TIME

22. SAM (14C-Me ) * SAM (14C-Me ) * Hordenine BIOSYNTHESIS * * tyrosine hordenine - CO2 PLP * N-methyltyramine tyramine Feeding experiment - SAM with 14C-labelled methyl group

23. HYDROXYLASES

24. HYDROXYLASES (MONOOXYGENASES) It is fairly easy for a plant to place a hydroxyl group in an aromatic ring by use of a hydroxylase enzyme. If one hydroxyl is present, it is even easier to add the second. The coenzyme XH2 in plants is a tetrahydropteridine O2 / XH2 hydroxylase Cu2+ cofactor + H2O + X In mammals XH2 is ascorbic acid. O2 / XH2 hydroxylase Note that while tyrosine is given as an example, most of the tyrosine in plants is not made this way, it originates from a pathway called the shikimic acid pathway. + H2O + X

25. - 2H+ The change the pteridine undergoes. - 2e- X XH2 ONE PROPOSED MECHANISM FOR THE HYDROXYLATION * E·Cu+ * + O2 + XH2 + H-O-H + X * E·Cu+ 2 + XH2 E·Cu+ + XH· + H+ E·Cu+ + O2 [ E·CuO2 ] XH· + SUBST-H2 XH2 + SUBST-H· [ E·CuO2 ] + SUBST-H· [ E·CuO2 · SUBST-H· ] [ E·CuO2 · SUBST-H· ] + XH2 SUBST-OH + H2O + X + E·Cu+

26. PEYOTE ALKALOIDS Mescaline

27. PEYOTE see assigned readings (last slide) for the details Lophophora williamsii Grows sparsely in the American Southwest, abundantly in arid high plains in Mexico. Indians have used it for centuries in religious ceremonies. Peyote was a serious item of trade amongst the native americans. It was used by tribes many hundreds of miles from its source. Hallucinogen vivid color hallucinations (vast film of fluctuating colors) nausea thirst mescaline The main alkaloid is mescaline Peyote “buttons” (cactus segments) were chewed by the indians

28. P e y o t e

30. MANY PLANTS (LIKE PEYOTE) CONTAIN FAMILIES OF RELATED ALKALOIDS The hydroxylating and methylating enzymes are fairly non-specific - often you will find many alkaloids in a plant that are similar in structure, but differ in the number of hydroxyl, methoxyl and N-methylamino groups found in their structures. ….. see the next slide In many cases, one will be a precursor of one of the others and they may be related by a definite sequence of biosynthesis. M A B C D E X Y Z

31. SOME OF THE PEYOTE ALKALOIDS mescaline anhalonidine anhalamine pellotine anhalinine anhalidine methylenedioxy group lophophorine

32. The order of hydroxylation or methylation may not be specific in some cases, in others the enzymes will control the order of introduction forcing a specific sequence or order of appearance of the compounds. Methylenedioxy groups are thought to be introduced in the following manner: .. O2 .. : : .. + NADPH NADP + H2O Enz-B-H

33. we will see in the next lecture how the one and two carbon pieces are added to form the rings tyrosine mescaline POSSIBLE BIOSYNTHETIC SEQUENCES FOR PEYOTE ALKALOIDS Can you fill in the coenzymes and other reagents?

34. PHARMACOLOGY

35. TRANSMISSION OF THE SIGNAL ALONG A NERVE PATH NERVE AXON transmission of the signal is electrical synaptic vesicles (NE inside) MAO transmission of the signal is chemical SYNAPSE Na+ K+ NEUROTRANSMITTER (norepinephrine = NE) K+ binding regenerates the electrical signal Na+ neurotransmitter is released and removed by an enzyme after the binding; it is transported back to the synapse by the binding opens a channel through which K+ and Na+ ions flow generating an electrical signal catechol O-methyl transferase COMT

36. TOPOLOGY OF THE NERVOUS SYSTEM NEUROTRANSMITTERS AND HORMONES central nervous system CNS SYMPATHETIC NERVOUS SYSTEM (Adrenergic) nerve cell adrenal gland Adrenalin Noradrenalin blood stream ORGAN receptor sites binding causes a response

37. norepinephrine (excitory) acetylcholine (inhibitory) serotonin (inhibitory) INTERCONNECTIONS IN THE BRAIN

38. MECHANISMS OF ACTION b-phenylethylamines minic NE Amphetamines cause immediate release of NE stimulate postsynaptic receptors Ephedrine (agonist) eventually deplete stores of NE Cocaine blocks readsorption of NE by the presynaptic terminus continues stimulation Mescaline stimulation similar to amphetamines not known why it stimulates color hallucinations and amphetamines do not

39. ASSIGNMENTS Olive Problem ……..due Friday, October 12 READ: Schultes, Hofmann, and Ratsch, Plants of the Gods, Healing Arts Press (2001) “What are Plant Hallucinogens?” etc., pp 10-30, “Who Uses Hallucinogenic Plants?”, pp 62-64, “The Tracks of the Little Deer”, pp 144-155 (Peyote) “Cactus of the Four Winds”, pp 166-169 (San Pedro). ….. a major reading assignment, but quite enjoyable.