1550 J.C.S. Perkin IAlkaloids of the Genus Erythroxylum. Part 1. E. monogynum Roxb.RootsBy Joseph T. H. Agar and William C. Evans," Department of Pharmacy, The University, Nottingham NG7 2RDThree new bases from the root-bark of Erythroxylum monogynum have been identified on the basis of chemicaland spectroscopic evidence ; they are 1 aH,5aH-tropan-3a-yI 3,4,5-trimethoxybenzoate, 1 aH.5aH-tropane-3a.GP-diol3- (3.4.5-trimethoxycinnarnate) 6- benzoate, and 1 aH,5aH-tropane-3cc,6/3,7/3-triol3- (3,4, Ei-trimethoxy-benzoate). 1 aH,5aH-Tropan-3a-y1 3.4.5-trimethoxycinnarnate was also isolated. Alkaloids were not detectedin the root-wood.FROM the leaves of the Indian tree Erythroxylwn mono- cinnamylcocaine have been isolated; the stem-wood is agynum Roxb. (Erythroxylaceae), (-)-ecgonine and source of terpenoids2 We have noted the presence, inR.N. Chopra and N. N. Ghosh, Arch. Pharm., 1938,276, 340. the root-bark, of a number of new alkaloids includingJ. C. Fairlie. R. McCrindle, and R. D. H. Murray, J . Chem. J. T. H. Agar, W. C. Evans, and P. G. Treagust, J . Pharm.Pharmacol., 1974, 26, Suppl., 111P. Soc. (C), 1969, 2115, and references cited therein1976 15511 aH, 5aH-t ropan-3a-yl 3,4,5-trimethoxybenzoate ; theknown bases laH,5aH-tropan3a-yl 3,4,5-trimethoxy-cinnamate, tropine, and $-tropine were also recorded.We present here the evidence for the structures of thenew alkaloids.Partition, and preparative t.1.c. of the root-barkextract afforded seven principal basic fractions repre-senting 0.85% of the dried material.A fraction havingRP 0.48 (system C; see Experimental section) gave acrystalline hydrobromide, C,,H,,NO,,HBr, and a picrate.Alkaline hydrolysis of the base gave tropine (1) and3,4,5-trimethoxybenzoic acid ; this observation, sup-plemented by n.m.r. and mass spectroscopic measure-ments, and by partial synthesis, established the newbase as laH,5aH-tropnn-3a-y1 3,4,5-trimethoxybenzo-ate. The 3P-isomer possessed properties (RB values,m.p., and n.m.r. data) dissimilar to those of the naturalproduct and its presence in the root-bark was not detec-ted.A base of R P 0.35 (system C), isolated as the picrate,was identified as laH,5aH-tropan-3a-y1 3,4,5-tri-methoxycinnamate (3), which has been reported as aconstituent of Erytlzroxylunz ellipticum leaves.* It wascompared with the analogous 3P-compound which ap-peared to be absent from the bark.The non-crystalline base, C,,H3,N07, of highest RF(0.90 ; system C) possessed a substituted hydroxytropanenucleus (mass spectrum), two dissimilar carbonyl groups,and substituted aromatic systems (i.r.).Hydrolysisgave benzoic acid, 3,4,5-trimethoxycinnamic acid andtropane-3a,6P-diol (4). These units were clearly accom-modated in the mass spectrum and an ion of m/e 333,consistent with structure (11) (cj. ref. 5) established thepresence of the trimethoxycinnamoyl group at C-3;no peak at m/e 217 was observed thus eliminating benzoyl-substitution at this position. On this evidence, thealkaloid can be identified as laH,5crH-tropane-3a,6p-diol3-(3,4,5-trimethoxycinnamate) 6-benzoate (5).Thedetermination of its optical rotation was precluded bythe small amount of sample available. The structurewas confirmed by partial synthesis of the (*)-isomerfrom ( &)-6P-hydroxytropan-3-one ( 7 ) . Conversion ofthis ketone into the benzoyl derivative (8) followed bystereospecific reduction with Raney nickel (W-7) andhydrogen yielded tropane-3a,6p-diol 6-benzoate (6).The 3~-orientation of the hydroxy-group was confirmedby n.m.r. spectroscopy (triplet, Wi 13 Hz, at 6 4.16;cf. quintet, Wi ca. 28 Hz for 3pderivatives). Esterific-ation with trimethoxycinnamoyl chloride yielded thediester, (&)-(5), which possessed the same i.r. and massspectroscopic properties and R F values as the naturalbase ; the product gave a picrate, C,,H3,N07,C,H,N,07,with m.p.5 "C greater than that of the picrate of thenatural compound.A fourth crystalline base of RF 0.17 (system C) had4 S. R. Johns, J. A. Lamberton, and A. A. Sioumis, Austral. J .5 E. C. Blossey, H. Budzikiewicz, M. Ohashi, G. Fodor, and C.Chem., 1970, 23, 421.Djerassi, Tetrahedron, 1964, 20, 585.ester (vmX 1700 cm-l) and free hydroxy-group (vmx.3400 cm-l) i.r. absorption. Mass spectrometry of thepicrate gave, for the base, the molecular formulaC,,H,,NO,; other ions indicated the presence of tri-methoxybenzoic acid and tropane-3~,6P,7P-triol(9) units.Hydrolysis afforded the relevant acid and the triol. AnN /MeQ3 RON /Me R20a R'O( 1 ) R = H ( 4 ) R' = R2= H121 R = (Me013C6H2.CO ( T r n b ) (5) R1 = Tmc, R2 =Bz( 3 ) R = (Me013C6H2*CH:CH-C0 [Tmc) (61 R' = H, R2= BZR O k 0( 7 ) R = H (9) R = H(8) R =Bz (10) R = TmbM eOR( 1 1 ) m / e 333, R = Tmc(12) m / e 307, R = Tmbion at m/e 307 in the mass spectrum was consistent withthe fragment (12), which would arise from an esterifyingsubstituent of teloidine at C-3.The a-orientation of thetrimethoxybenzoyl moiety was indicated by the band-width (W4 14 Hz) of the H-3 n.m.r. signal a t 6 5.23;integration showed the presence of two hydroxy-groups.From the above it follows that this new alkaloid islaH,5aH-tropane-3a,6p,7p-triol 3-(3,4,5-trimethoxy-benzoate) (10).A number of other bases were detected in other frac-tions of the chromatographed plant extract, but theirsmall quantity and their isolation as mixtures, have sofar precluded their precise identification.Extracts ofthe root-wood, after fractionation by t.l.c., did not givepositive results in tests for alkaloids.The considerable range of tropane ester alkaloidspresent in the root-bark of E. monogynum is noteworthy;to date, only in Datura and Anthocercis,6 of the Solanaceae,has the co-occurrence of tropine, dihydroxytropane,and teloidine esters also been recorded. Erythroxylum,in common with these genera, possesses free $-tropineand tropine in the roots but only the latter also occursas a variety of esters. Outside Datura, Erythroxylum6 W. C. Evans, A. Ghani, and V. A. Woolley, Phytochemistry,1972, 11, 2527; W.C. Evans and P. G. Treagust, ibid., 1973, 12,2605J.C.S. Perkin I 1552appears to be the first genus from which a bisacyloxy-tropane involving two different acids has been isolated;a full survey of alkaloids within the genus (ca. 200 sj$.)may well offer considerable chemotaxonomic interest.EXPERIMENTAL1.r. spectra (KBr discs) were recorded with a Unicam SP200 instrument; n.m.r. spectra were obtained for solutionsin CDCl, with a Perkin-Elmer R 10 spectrometer (tetra-methylsilane as internal standard) ; mass spectra weredetermined with A.E.I. MS902 and MS9 instruments.G.1.c. was performed with a Pye 104 series chromatograph(OV-101 on a solid, silanised support). For t.1.c. and p.1.c.alumina (Merck G) plates with (A) ether; (B) ether-ethanol(1 : 1); or (C) chloroform-ethanol (98: 2), and (D) silicaplates with acetone-ammonia (s.g.0.88) (4 : 1) were used.Separated alkaloids were detected with U.V. light and iodo-platinate reagent. The conditions for the hydrolysis ofesters were as recorded previously.7Plant Matevial.-Dried root-bark and herbarium speci-mens were obtained from the region of Madras and Coimba-tore, India.Extraction of A ZkaZoids.-In typical experiments powderedroot-bark (100 g) was mixed with calcium hydroxide (20 g)and water (40 ml), set aside overnight, and exhaustivelyextracted with ether (4 1). The concentrated extract (25 ml)was passed through kieselguhr (1 5 g) supporting IN-sulphuricacid (7.5 ml). After extraneous material had been washedfrom the column with ether, the bases were collected inammoniacal chloroform.Evaporation afforded a crudealkaloid mixture which, in ether (2 ml), was submitted topartition chromatography a t pH 8.0 and partially fraction-ated by successive elution with light petroleum (b.p. 40-60°), ether, and chloroform. Pure alkaloids were obtainedby the use of a second partition column and by p.1.c. (systemlaH,5aH-Tropan-3a-yZ 3,4,5-Trimetlzoxybenzoate (2) .-Abase isolated by p.1.c. and having Rip 0.48 (system C)afforded a hydrobromide, fine needles from ethanol, m.p.212" (decomp.) (Found: C, 51.7; H, 6.3; N, 3.4. C18H2,-NO,,HBr requires C, 51.9; H, 6.3; N, 3.4%), and a picrate,prisms from aqueous ethanol, m.p. 201" (Found: C, 50.7;H, 5.1; N, 9.8.C18H,,N0,,C,H3N,07 requires C , 51.1;H, 5.0; N, 9.9yo), vmx. 1 700 cm-1 (ester C=O), m/e 42, 82,83, 94, 96, and 124 (100%) (characteristic of tropine ester^),^212 (lay0, trimethoxybenzoic acid), 195 (15%, acyl unit),and 335 (39%, M+). The base recovered from the picrateshowed 6 1.64br, 1.79br, and 2.00 (8 H, s, s, and m, respec-tively, 2-, 4-, 6-, and 7-H,), 2.21 (3 H, s, NMe), 3.06br(2 H, m, 1- and 5-H), 3.99 (9 H, s, 3 x OMe), 5.12 (1 H, t,W+ 11 Hz, 3P-H), and 7.17 (2 H, s, ArH) . Alkaline hydroly-sis of the base (0.02 g) gave 3,4,5-trimethoxybenzoic acid(0.01 g), m.p. and mixed m.p. 170°, identical (i.r., mass, andn.m.r. spectra) with authentic material (Found: C, 56.5;H, 6.0. Calc. for ClOH12O5: C, 56.6; H, 5.7%), and tropine,identical (Rip in system D, and RF and i.r.spectrum oftigloyl ester) with authentic material.Tropine (0.1 g) and 3,4,5-trimethoxybenzoyl chloride(0.27 g) were intimately mixed and maintained at 100 "C for6 h. The resulting solid mixture was dispersed in ammoniasolution (30 ml; 5 ~ ) and liberated basic material was col-lected in chloroform (3 x 30 ml). Evaporation and p.1.c.gave a product which, on treatment of the sulphate withsodium picrate solution, furnished 1 aH,5aH-tro@a-3a-yl3,4,5-trirnethoxybenzoate picrate (0.082 g), m.p. 201" (Found:C).C, 50.9; H, 5.3; N, 9.3%), identical with the naturalderivative (m.p., mixed m.p., and i.r., n.m.r., and massspectra).The 3P-Iso?ner of the Base (2).-Esterification of 4-tropine with 3,4,5-trimethoxybenzoyl chloride as above,followed by picrate formation, gave laH,5aH-tropan-3p-yZ3,4,5-trimethoxybenzoate picrate, prisms from aqueous ethanol,m.p. 226" (Found: C, 51.4; H, 5.2; N, 9.8%).The base,needles, m.p. 98", had Rip 0.75 (system C), 6 5.16 (1 H, m,W, 21 Hz, 3a-H).laH,5aH-Tropan-3a-yZ 3,4,5-Trirnethoxycinnanzute (3) .-This base, RF 0.35 (system C), was isolated by p.1.c. ofcomponents of ethereal eluates from partition columns, andwas identified by i.r., n.m.r., and mass spectroscopy (Found:M', 361.1899. Calc. for C,,H,,NO,: M , 361.1889). Thealkaloid formed a picrate, prisms from aqueous ethanol,m.p. 210" (Found: C, 52.5; H, 5.0; N, 9.1. C20H27N0,,C,H,N,O,requires C, 52.9; H, 5.1; N, 9.5%). The productresulting from esterification of the tropiiie and 3,4, 5-tri-methoxycinnamoyl chloride [as for the trimethoxybenzoate(Z)] gave a picrate (0.090 g), m.p.210", identical with thatderived from the natural product (Found: C, 53.0; H, 5.0;The 3P-Isorner of the Base (3).-Esterification of $-tropine with 3,4,5-trimethoxycinnarnoyl chloride, as above,gave a base, Rip 0.73 (system C), which furnished with sodiumpicrate solution laH,5aH-tropan-3P-y2 3,4,5-trimethoxy-cinnanzate picrate, small prisms from aqueous ethanol, m.p.253" (decomp.) (Found: C, 51.8; H, 5.8; N, 8.9%).la-,5aH-TroPane-3a, 6P-diol 3-( 3,4,5-Trimethoxycinnanzate)6-Benzoate (5) .-P.1.c. of the combined light petroleumeluates from partition columns gave basic material (5) (RF0.9 in system C) which on salt formation yielded a hydro-bromide, prisms from hot water, m.p.195-200" (Found:M+, for basic unit, 481.2091. C2,H3,N07 requires M , 481.2101), vmx 1701 (doublet, 2 x ester GO) and 1624 cm-l(CHzCH), and an auvichloride, plates from dilute hydro-chloric acid, m.p. 148" (Found: &I+, for basic unit, 481.2105),m/e (characteristic signals from spectra of aurichloride andhydrobromide) 105 (C,H,*CO) 122.0365 (Calc. for PhC0,H :122.0368), 221.081 1 [Calc. for (MeO),*C,H,*CH:CHCO22 1.08 131, 238 [ (MeO) ,*C,H2*CH:CH*C02H], and 333.1564[C1,H,,NO,, formula (1 l), requires 333.15761. The picrate,prisms from aqueous ethanol, had m.p. 224". The productsof alkaline hydrolysis of the new base were examined byg.1.c. The components of the mixed acid fraction, in ether,after conversion into their methyl esters with diazomethane,had the same retention times as methyl benzoate andmethyl 3,4,5-trimethoxycinnamate ; the basic unit, re-covered from aqueous solution, gave peak enhancementwhen mixed with, and had the same retention time as( &)- laH,5aH-tropane-3a,6p-diol.(&)-laH,5031-tropane-3a,6P-diol 6-benzoate (6) (0.25 g),mixed with an excess of 3,4,5-trimethoxycinnamoyl chloride(0.7 g) and maintained a t 100 "C for 2 h under a dry airstream, gave, after p.1.c.(system A), a base (5) characterisedas the picrate, prisms from aqueous ethanol, m.p. 228"(Found: C , 55.6; H, 5.1; N, 7.3. C,,H,,NO,,C,H,N,O,requires C, 55.8; H, 4.8; N, 7.9%). The i.r. and massspectroscopic data of this compound were the same as for thenatural product derivatives; mixed m.p.225".laH,5aH-TroPane-3a,6P-dioZ 6-Benzoate (6) .-( &)-6P-Hydroxytropan-3-one (7) (2.0 g) was treated (140 "C; 1 h)with benzoyl chloride (1.6 g), the mixture in chloroform was7 W. C. Evans and V. A. Major, J . CAem. SOC. ( C ) , 1966, 1621.N, 9.5%)1976 1553washed with sodium carbonate solution ( 2 ~ ; 3 X 30 ml),and basic material was isolated by standard procedures.P.1.c. (system C) and recovery of the alkaloid, R p 0.90 (systemB), gave 3-oxotropan-6p-yl benzoate (S), isolated as thepicrate (1.2 g), prisms from aqueous ethanol, m.p. 172"(Found: C, 51.6; H, 4.3; N, 11.2. C15H17N0,,C,H,N,0,requires C, 51.6; H, 4.1; N, 11.5%), vmX. 1710 cm-l(doublet, 2 x GO), mle 84, 112, 138, 154, and 259 (M+).The ketone (8) (0.5 g) on reduction with hydrogen andRaney nickel (W-7) at room temperature and atmosphericpressure for 18 h gave a base (6) isolated as (f)-laH,5aH-tropane-3a,6p-dioZ 6-benzoate picrate (0.27 g), prisms fromaqueous ethanol, m.p.203" (Found: C, 51.1; H, 4.5; N,11.7. Cl,HIBNO,,C,H,N,O, requires C, 51.4; H, 4.5; N,11.15%), vmaX 1 695 (GO) and 3 410 cm-l (OH), 6 1.7-2.5(6 H, m, 2-, 4-, and 7-H,), 2.58 (lH, s, 3-OH, lost on deuteri-ation), 2.71 (3 H, s, NCH,), 3.49 (2 H, m, 1- and 5-H), 4.16(1 H, t, W4 14 Hz, 3-H), 5.94 (1 H, q, 6-H), 7.51 (3 H, m,aromatic m- and p-H), and 8.05 (2 H, m, aromatic o-H),wz/e 96, 113, 156, ahd 261 ( M f ) .laH , 5aH-Tropane-Sec, 6PY7P-triol 3- (3,4,5-Trimethoxybcnzo-ate) (lo).-The first fractions of the chloroform eluate of apartition column, pH 6.8, used for the resolution of a crudealkaloid fraction afforded a crystalline solid, m.p.112' ; thesolid (10) was characterised as the picrate, prisms fromaqueous ethanol, m.p. 188" (Found: l'+, 367.1621.C18H,,N07 requires M , 367.1631), vmx. 1 700 (ester G O ) and3 380 cm-l (OH). The base (lo), recovered from the picrate,showed 6 2.56 (3 H, s, NCH,), 3.12br (2 H, s, 1- and 5-H),3.92 (9 H, s, OCH,), 4.56 (2 H, s, 6- and 7-OH), 5.23 (1 H, m,W* 14 Hz, 3-H), and 6.73 (2 H, s, aromatic o-H), m/e 172(teloidine unit, C8HlPNO3), 195 [(MeO),C,H,CO], 212.0684(calc. for C10H1205: 212.0685), and 307 [(MeO),.C,H,-CO*O*C,HlON]. Hydrolysis of the base (24 mg) withaqueous barium hydroxide gave 3,4,5-trimethoxybenzoicacid (9 nig), m.p. and mixed m.p. 171", M f 212, identical(i.r. spectrum) with the authentic compound, together withtropane-3a,6(3,7p-triol (9), identical (RF values of free baseand its triacetate) with an authentic specimen.Boot-wood-The powdered root-wood (1 g ) , treated withcalcium hydroxide (0.2 g) and water (0.4 ml), was exhaustedwith ether and submitted t o standard procedures for theisolation of alkaloids. Thin-layer chromatograms (systemC) of the extract were sprayed with iodine and iodoplatinatereagents.We are indebted to Dr. C. L. Madan, Jamniu Tawi, andthe Tropical Products Institute, London, for the supply ofplant material.[6/121 Received, 19th Janzzary, 1976
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