Some important classes of alkaloids Class Precursors Examples _____________ Piperidine alkaloids L-lysine (C 5 N)

1. Some important classes of alkaloids Class PrecursorsExamples_____________ Piperidine alkaloids L-lysine (C5N) Piperine Pyrrolidine/tropane alkaloids L-ornithine(C4N) Cocaine, scopolamine Pyridine alkaloids L-Trp or L-Asp Niacin (Vit. B3), nicotine Catecholamines L-Tyr (C6C2N) Dopamine, adrenaline, mescaline and tetrahydroisoquinolinesanhalamine Opiates 2 L-Tyr units Morphine, tubocurarine Phenylalanine-derived L-Phe(C6C3N) Capsaicin, ephedrine Indole alkaloids L-TrpSerotonin, ergotamine, LSD, vinblastine Quinoline alkaloids “ quinine, camptothecin Purine alkaloids L-Gly, L-Gln, L-Asp Caffeine, theobromine Building blocks from the acetate, shikimate, or deoxyxylulose phosphate pathways are also frequently incorporated into the alkaloid structures. Many alkaloids acquire their N via transamination reactions (catalyzed by Vitamin B6). Most alkaloids are quite toxic and produced by the plant as a defense against herbivores.

2. Opiates: morphine and related compounds • Focus: • pieces/precursors • key assembly steps • structure-activity • physiological interactions First methylation Precursor is formed by combining two C6C2 units, both originating from tyrosine

3. Two stereoisomers lead to different classes of alkaloids From (S)-N-methyl coclaurine Further methylation is common Berberineis found in many members of the Berberidaceae, the Ranunculaceae, and other families used in traditional medicine. Berberine has antiamoebic, antibacterial, and anti-inflammatory properties.

4. Curare (Chondrodendrontomentosum) Arrow poison prepared in the rain forests by native South American tribes from bark and stems may contain >30 different plants, C. tomentosumis the main one. Radical dimerization of the two coclaurine isomers produces a dimer. Tubocurarine is an ACh agonist, once it gets into the bloodstream, it relaxes voluntary muscles to paralysis by blocking nerve impulses at neuromuscular junction. Synthetic derivatives used during surgery as muscle relaxant.

5. Oh oh…I think I just stuck myself… better get out my cholinesterase inhibitor pen!

6. Making the morphine skeleton Key steps: oxidative coupling to make the 4th ring 2) reduction and acetylation promotes formation of the furan ring 3) Demethylation then reduction of the ketone

7. “antitussive”

8. Opium • 80% of the world’s illegal opium is grown in Afghanistan, supported by the Taliban. • $ 4 billion/year industry • Northern alliance initiative to destroy poppy fields pits farmers and corrupt local officials against military • U.S. troops are supporting efforts to provide farmers with crop alternatives like pomegranates or grapes • However, many farmers told in 2007 that they will receive assistance in establishing legal crops are still waiting for help R. Draper, “Opium Wars”, National Geographic, Feb. 2011.

9. Properties of opiates Brain has opioid receptors (d, k and m) that mediate adenylcyclase, lower cAMP levels, increase K+ conductance, close Ca2+ channels, decrease transmission of pain signals Morphine’s use as analgesic involves overall CNS depression so side effects include drowsiness, lethargy, constipation, unconsciousness

10. Structural basis for bioactivity of opiates Enkephalins and endorphins are endogenous opioid peptide ligands produced during labor, other times of stress or shock “Runner’s high” ACh inhibitors, rapidly degraded “Morphine rule”: structure must contain an aromatic ring attached to a quarternary C (Cq) with a 2-C linker to a tertiary (3o) amine

11. Drugs derived from morphine semi-synthetically Similar activity, can be produced from morphine by methylation Heroin was introduced into cough syrup in 1898 Acetylation decreases polarity, so it crosses blood-brain barrier quickly. More addictive though. Synthetic analogues Effective analgesic and less addictive Used in cough medicine  Antitussive but not analgesic “Demerol” 50-100X more active than morphine due to lipophilicity Orally active, produces less euphoria, less withdrawal Fast, but short-acting analgesia

12. From L-Tryptophan: Indole alkaloids Produced from L-tryptophan plus an isoprene unit, the indole alkaloids have polycyclic ring structures Fungi from Claviceps genus are best-known producers because they infect some grain crops, but the indoles are produced by other fungi including AspergillusandPenicillium Lysergic acid is probably the most well-known, as it is the precursor for hallucinogen LSD (lysergic acid diethylamide) and the ergot alkaloids – mixed agonist/antagonist effects on 5-HT (serotonin) receptors leads to hallucinations serotonin skeleton

13. Ergot – not a fun guy Ergot is the dried sclerotium of the fungus Clavicepspurpureathat develops on the ovary of rye and other grasses consumed by humans or animals. The poisonous properties of ergots are caused by a group of indole alkaloids, the ergot alkaloids or ergolines. Consumption of ergot-infected rye produces a disease called ergotism. Ergot poisoning affects two types of receptors: 1) The a-adrenergic receptors for norepinephrinewhich causes • GI upsets, e.g. diarrhea, abdominal pains, and vomiting. • Circulatory changes, e.g. coldness of hands and feet due to vasoconstriction of the blood vessels to the extremities

14. Ergot – not a fun guy 2) Ergot also acts on the serotonin (5-HT) receptors and the a-dopaminergic receptors causing neurological symptoms: Headache, vertigo, convulsions, psychotic disturbances, hallucinations Vasoconstriction can cause restricted blood flow in small terminal arteries, death of the tissue, gangrene, and even the loss of hands, feet, or limbs. Gangrenous ergotism was known as St. Anthony’s Fire because the Order of St. Anthony cared for the sufferers during the Middle Ages in Europe when outbreaks of the disease in humans and animals were relatively frequent.

15. Indole-isoprenoid is modified by attachment of a small peptide (Phe & Pro usually) Ergotamine’s vasoconstrictive activity has led to its use in treatment of migraines

16. For more information:A Trip Through LSD and the Ergot Alkaloids Melissa Medeiros Natural Products May 2006

17. Amaryllidaceae alkaloids synthetic derivative trans-dihydronarciclasine narciclasine (abundant) pancratistatin Compounds 1 – 6 are found in Narcissus pseudonarcissus (daffodils) and other plants from the Amaryllidaceae family. Pancratistatin (1) selectively induces apoptosis in cancer cells through a mitochondrial pathway, with little effect on normal cells. However, it’s not very abundant. Narciclasine (3) disrupts organization of the actin cytoskeleton of cancer cells at 30-90 nM. Compound (5) also has potent anticancer activity while its cis-isomer has none. Problems: low water-solubility, low natural abundance of (1) and (5), and toxicity of (3) ... Narciclasine inhibits CYP3A4 (major liver detoxifying cytochrome), which can damage the liver.

18. SAR shows this part of structure is key... Goal of study: prepare trans-dihydro derivative of narciclasine that has the anti-cancer activity without the liver toxicity.

19. Protection of the OH groups followed by hydrogenation usually produces a mixture of cis- and trans-isomers. Since the cis-isomer (7) has no anticancer activity, reaction conditions were modified until maximum yield of trans isomer was achieved. Compound 5 did not inhibit any of the cytochromes tested. Result: an effective way to prepare a less toxic drug!

20. More indole alkaloids...from Uncariatomentosa Complex polycyclic indole alkaloids are prevalent in species such as Uncariatomentosa (Cat’s claw) see: Heitzman, M. E.; Neto, C. C.; Winiarz, E.; Vaisberg, A. J., Hammond, G. B.* Review: Ethnobotany, phytochemistry and pharmacology of Uncaria (Rubiaceae), Phytochemistry, 66: 5-29 (2005).

21. Traditional uses of U. tomentosa Aqueous extracts of bark or roots used by Ashaninka in Peruvia rainforest: • Allergies, asthma • Wound healing, viral infections, fevers • Arthritis, gastric ulcers, rheumatism, inflammation • Contraception, menstrual irregularities, childbirth • Some of these activities are associated with alkaloids • Uncaria also contains flavonoids, catechins, cinchonains, caffeic acid and ursolic acid – these may play a role in some of the bioactivities.

23. Weaker dopamine receptor antagonist – some locomotor depression observed Pentacyclicoxindole alkaloids have immune-boosting effect, upregulate B & T cell production Tetracyclicoxindole alkaloids block immune factor, may also inhibit central dopamine release, depressing movement

24. More indole alkaloids...terpeneindole alkaloids with multicyclic structures Coupling of tryptamine (decarboxylatedTrp) with the secoiridoid secologanin, followed by a variety of cyclizations forms terpeneindole alkaloid skeletons with lots of structural diversity

25. Some interesting terpeneindole alkaloids Yohimbine • From Pausinystaliayohimbe • Folk use: Aphrodisiac, weight loss • Pharmacology: dilates blood vessels, binds a-adrenergic receptors Reserpine, deserpidine • From Rauwolfiaserpentina • Folk use: treat snake bite, insanity • Pharmacology: lowers blood pressure, tranquilizer, depletes stored catecholamines

26. Skeletal rearrangements and dimerization lead to vincristine/vinblastine structure • Intermediates form through rearrangement of isoprenoid moieties Anti-cancer agent, used to treat leukemia and lymphoma.

27. Strychnine • From dried seeds of Strychnosnux-vomica • Biosynthesis: Rearrangements in strictosidine • Highly toxic convulsant • Binds to glycine binding sites in spinal cord, causes asphyxiation • Has been used as a rat poison

28. Quinine – Antimalarial agent from bark of Cinchona trees Discovered in 1600’s , cured a Peruvian countess Depolymerizes toxic byproducts of Plasmodium Quinine was originally added to tonic water as a prophylactic against malaria – gin was later added to mask the bitter taste 

29. Some important classes of alkaloids Class PrecursorsExamples_____________ Piperidine alkaloids L-lysine (C5N) Piperine Pyrrolidine/tropane alkaloids L-ornithine(C4N) Cocaine, scopolamine Pyridine alkaloids L-Trp or L-Asp Niacin (Vit. B3), nicotine Catecholamines L-Tyr (C6C2N) Dopamine, adrenaline, mescaline and tetrahydroisoquinolinesanhalamine Opiates 2 L-Tyr units Morphine, tubocurarine Phenylalanine-derived L-Phe(C6C3N) Capsaicin, ephedrine Indole alkaloids L-TrpSerotonin, ergotamine, LSD, vinblastine Quinoline alkaloids “ quinine, camptothecin Purine alkaloids L-Gly, L-Gln, L-Asp Caffeine, theobromine Building blocks from the acetate, shikimate, or deoxyxylulose phosphate pathways are also frequently incorporated into the alkaloid structures. Many alkaloids acquire their N via transamination reactions (catalyzed by Vitamin B6). Most alkaloids are quite toxic and produced by the plant as a defense against herbivores.

30. From amino acids to purines to caffeine and xanthine alkaloids Purine heterocyclic ring system is derived from amino acids and various one C donors muscle relaxant

31. The xanthines Caffeine, Theobromine, and Theophylline • The purine alkaloids caffeine, theobromine, and theophylline are all methyl xanthines that commonly co-occur in plants. Major sources are stimulant beverages and foods such as tea, coffee, cocoa, and cola. • Xanthines competitively inhibit phosphodiesterase, causing an increase in cyclic AMP and adrenaline release. This leads to CNS stimulation, relaxation of bronchial smooth muscle, and induction of diuresis. Inhibition of TNF-a and leukotriene synthesis is thought to occur, reducing inflammation and innate immunity. • The effects vary among the three compounds. Caffeine is the best CNS stimulant. As a vasoconstrictor it can be combined with a therapeutic agent to increase effectiveness (e.g. compound analgesics). It has weaker diuretic action. • Theobromine has little stimulant action, but has more diuretic activity and also muscle relaxant properties. Theophylline also has low stimulant action and is an effective diuretic, but it relaxes smooth muscle better than caffeine or theobromine and is frequently used in slow-release formulations.

33. Caffeine and adenosine • Caffeine readily crosses the blood-brain barrier, and once in the brain, the principal mode of action is as a nonselective antagonist of adenosine receptors (competitive inhibition) • Adenosine is found in every part of the body, but it has special functions in the brain. Concentrations of brain adenosine are thought to be increased by metabolic stresses such as anoxia or ischemia. It also may have a specific role in control of the sleep-wake cycle. • Brain adenosine may also protect the brain by suppressing neural activity and increasing blood flow through A2A and A2B receptors located on vascular smooth muscle. By counteracting adenosine, caffeine reduces resting cerebral blood flow – it’s a vasoconstrictor. • Adenosine is released in the brain through a complex mechanism. It is not likely that adenosine is the primary neurotransmitter for any group of neurons, but rather is released together with other transmitters by a number of neuron types.

34. So you can’t start the day without Joe?You’re not alone! • Is caffeine addictive? Several classes of adenosine receptors are known, and there is evidence that A 2A receptors interact with the dopamine system, which is involved in reward and arousal. • Tolerance: because caffeine is primarily an antagonist of adenosine receptors, regular caffeine consumers may adapt to its continuous presence by increasing the number of adenosine receptors. • This reduces the stimulatory effects (tolerance adaptation) and makes one much more sensitive to adenosine, so that reducing caffeine intake results in withdrawal symptoms.

35. From Drugs and the Human Body, 6th edition, K. Liska

36. Chocolate and pet poisoning • Dogs are most often affected, due to their ability to find chocolate and the common 'sweet tooth' they develop, but cats and other mammals are susceptible to the toxic effects of chocolate, too. • Theobromine is the major stimulant in chocolate. Its effects: • CNS and cardiovascular stimulant • Increases blood pressure (mild) • Nausea and vomiting • Are some chocolates more toxic than others?Yes. Unsweetened chocolate contains 8-10 times the amount of Theobromine as milk chocolate. Semi-sweet or dark chocolate falls roughly in between the two. • The toxic dose of Theobromine (or caffeine) for pets is 100-200mg/kg body weight. However, reports by the ASPCA have noted problems at doses much lower than this - i.e. 20mg/kg. • Using the 20mg/kg as a measure of "problems can be seen” a 50 lb (23 kg) dog would have to consume 9 oz of milk chocolate. Some dogs won't see problems at this rate but others may. http://vetmedicine.about.com/cs/nutritiondogs/a/chocolatetoxici.htm

38. Background • A Dutch cohort study in 2002 and others found that higher coffee consumption correlates with lower risk of Type-2 diabetes. • Coffee constituents that have been previously shown to affect glucose metabolism include caffeine, chlorogenic acid, lignans, trigonelline. • Caffeine also reported to induce energy expenditure by boosting thermogenesis. • However some cohort studies suggest that decaf also reduces risk. • Is caffeine the active constituent against diabetes?

39. Mouse model • KK-A3’ mice spontaneously develop type-2 diabetes by 6 weeks old • Obesity, hyperglycemia, high triglycerides and fatty liver also develop • Hyperinsulinemia is observed, suggesting that insulin resistance occurs in peripheral tissues. • Altered signaling in adipose tissues: decreased adiponectin (a adipocytokine) and increased MCP-1 (monocytechemoattractant protein) and TNF-a which induce insulin resistance. • Study looked at mechanism of coffee’s action against type-2 diabetes and the effect of caffeine alone on development of hyperglycemia.

40. Experiments • Expt 1: Diabetic mice were given diluted coffee vs. plain drinking water for 5 weeks ad libitum • Blood drawn and blood glucose measured weekly • Insulin tolerance test given at 5 weeks • At study end, mice were sacrificed and tested for serum insulin, triglycerides, total cholesterol, and pro-inflammatory markers MCP-1 and IL-6. • Liver and adipose tissues were removed and examined for cholesterol, TG and phospholipid content. • Expt 2: Same study was repeated with caffeine solution (250ug/mL) vs. plain drinking water

41. Coffee treatment lowered blood glucose and insulin tolerance in the mouse model

42. Coffee treatment significantly reduced liver weight, most types of body fat, liver lipid content, insulin and TAG levels, and inflammatory markers

43. Conclusions • Coffee improved insulin sensitivity and decreased hyperglycemia (high blood sugar) • Expression of inflammatory cytokines such as MCP-1, IL-6 and TNFa in adipose tissue was lower in the coffee treatment group. • Coffee decreased adipose tissue inflammation, thus improving insulin resistance. • Caffeine alone also reduced hyperglycemia, inflammation in some but not all fat tissues. • Both coffee and caffeine improved fatty liver.