2* * * * * * * * 14C-Tyrosine fed to opium poppy seedlings Two labels areincorporated intopapaverine andnorlaudanosoline,implying that twotyrosines are used.14C-Tyrosine fed toopium poppy seedlings*hydroxylasetyrosineO2If labeled DOPA is fedonly one label is found.*PLPDOPAPLP**Mannich + Arom. Subst.dopamine**thebaineSAM**FADcodeinepapaverinenorlaudanosolinemorphine(in all poppies)
314CO2 FEEDING EXPERIMENT This order was established by growing the poppies in a greenhouse with 14CO2and watching the order of appearanceand buildup of the the radioactivityAt first the rate of incorporation is greaterfor thebaine than for codeine or morphine.After several days, the rate of incorporationinto morphine is greatest. This indicatesthat thebaine is formed first, and that codeineand morphine follow in order.norlaudanosolinethebaineThe same result is obtained with carbon-14labeled tyrosine.codeinemorphineAlso, labeled thebaine is converted to bothcodeine and morphine, but not the other wayaround.Finally, codeine is demethylated to morphine;the reverse is not observed.
4* * * * * * * * * * redrawn (folded) on the next slide ….. SAM FAD norlaudanosolinelaudanosinepapaverineSAMNorlaudanosoline is incorporatedinto papaverine and thebaine at ahigher rate than tyrosine - it’s closer.**redrawn (folded)on the next slide …..**SAMIn a separate set of experiments,it can be shown that the methylgroups originate from methionine(14C labeled CH3) and hence SAM.laudanosoline(-)-reticulene
5. . MULTIPLE LABELS * * * * not an easy synthesis ! thebaine(all labels intact !)(-)-reticulene (5 labels)not an easy synthesis !When multiple labels survive intact it is a good indicationthat the incorporation study is a very valid one!
7. . . * * * * * * ORIGINAL HYPOTHESIS (ALA PUMMERERS KETONE) THEBAINE IT MISLEAD THEM**..Michaeladdition**(+)-salutaridineNADPH*.-H2OTHEBAINE*H+
8. . . * * * * * * REVISION OF ROUTE TO THEBAINE THEBAINE NADPH salutaridinol**(+)-salutaridineallylicdisplacementLabeled salutaridine isincorporated at a highrate into thebaine,codeine and morphine.*.Labeled salutaridinol(both -OH epimers) isalso incorporated at ahigh rate.THEBAINE*
9THE FINAL INTERCONVERSIONS + H2O- H2Oneopinonethebainehemiketalisomerizationof double bonddemethylationNADPHmorphinecodeinecodeinoneLabeled neopinone is converted to codeine in the plant.
10. . . . ANOTHER REVISION * * * * * * * * TRITIUM IS LOST TRITIUM +***(+)-reticulene1,2-dihydroreticulene(-)-reticuleneTRITIUMIS LOST*.(+)-Reticulene appears to beformed first, and than convertedto (-)-reticulene through the1,2-dihydroreticulene intermediate.TRITIUMSURVIVES*(-)-Reticulene is incorporated withthe tritium label intact. (+)-Reticuleneloses the label.(-)-reticulene
11WHAT WAS NOT SHOWN Remember, each of the labeled compounds had to be synthesized with high specificity.Dilution compounds had to isolated or synthesized.Each of the destination compounds had to be specificallydegraded to find the position of the label.At times the percentage incorporation was important toknow - this requires painstaking work.Most times, fortunately, it is sufficient just to know wherethe label ended up.All of these plants had to be grown and tended to.Manipulation of radioactive waste was a major problem(14C has a half-life of 5730 years) it doesn’t go away if spilled.
12Much of this work was done by Sir Alan Rushton Battersby(b. 1925) and his co-workers.Manchester College, EnglandPh.D. Dundee (1959)Reader at Bristol, Liverpool andCambridgeOPTIONAL READINGA.R. Battersby and B.J.T. Harper,“Biogenesis of Morphine”, Chemistry and Industry1958, 364.A.R. Battersby, R. Binks, D.J.Le Count,“Biosynthesis of Morphine”,Proceedings of the Chemical Society, 1960, 287.
13Alkaloids - Nature’s Cure or Curse, Wiley-VCH (2000) ASSIGNMENTSRead:HesseAlkaloids - Nature’s Cure or Curse, Wiley-VCH (2000)“Alkaloid Biogenesis in Papaver Somniferum”,ppTwo of the degradation procedures will be foundhere in Figures 8.16 (C9) and 8.17 (C16). You willalready be familiar with some of this chemistry aswe learned it in the discussion of the structureproof of morphine.