Prodelphinidin polymers: definition of structural units

摘要

1186 J.C.S. Perkin I Prodelphinidin Polymers : Definition of Structural Units By Lai Yeap Foo and Lawrence J. Porter,* Chemistry Division, Department of Scientific and Industrial Research, Petone, New Zealand The two basic structural units of condensed prodelphinidin polymers have been defined by generation of the C-4 carbocations, corresponding in stereochemistry to (+)-gallocatechin (1 ) and (-) -epigallocatechin (2). and trap- ping them as their phloroglucinol adducts (4) and (7). Two naturally occurring dimeric prodelphinidins with stereochemistry based on that of gallocatechin have also been isolated. THEchemistry of the procyanidins, a widely distributed class of phenolic secondary plant products, is now well understood. The structure and distribution of the singly linked B-type procyanidin dimers has been studied le3 and the anomalous lH n.m.r.spectra of their acetate and methyl ether derivatives have been explained on the basis of rotational isomeri~m.~?~ Additionally, the structure of the doubly linked A-type dimers has been successfully assigned by 13C n.m.r. spectroscopy 6-8 and K. Weinges, W. Kaltenhauser, H.-D. Mafx, E. Nader, I?. Nader; J. Perner, and D. Seiler, Annalen, 1968, 711, 184. R. S. Thompson, D. Jacques, E. Haslam, and R. J. N. Tanner, J.C.S. Perkin I, 1972, 1387. F. Delle Monache, F. Ferrari, and G. B. Marini Bettolo, Gazzetta, 1971, 101, 387. A. C. Fletcher, L. J. Porter, and E. Haslam, J.C.S. Chem. Comm., 1976, 627. the distribution of this class of procyanidins in the plant kingdom has also been ~tudied.~ In contrast, little is known of the chemistry of the prodelphinidins apart from the fact that a wide range of woody plants yield delphinidin when methanolic extracts of their fruit or leaves are heated with butanol and hydro- chloric acid.9 The flavan-3-01s corresponding to this group of phenolic polymers, (+)-gallocatechin (1) and (-)-epigallocatechin (2)are well known natural products A.C. Fletcher, L. J. Porter, E. Haslam, and R. K. Gupta, J.C.S. Pevkin I, 1977, 1628. D. Jacques, E. Haslam, G. R. Bedford, and D. Greatbanks, J.C.S. Chem. Comm., 1973, 518. D. Jacques, E. Haslam, G. R. Bedford, and D. Greatbanks, J.C.S. Perkin I, 1974, 2663. G. Schilling, K.Weinges, 0.Miiller, and W. Mayer, Annalen, 1973, 1471. E. C. Bate-Smith, Phytochemistry, 1975, 14, 1107. and were isolated and characterised long ago.lo*l1 How- ever, although at least four, lsquo; leucodelphinidins rsquo; have been reported with flavan-3,4-diol structures assigned,12 their proposed formulations have not been supported by compelling evidence. In view of the failure so far either to isolate l3or to demonstrate the existence l4of flavan- 3,4-diols in plant material containing proanthocyanidins possessing a 5,7-dihydroxylated ring A, it seems probable that the above lsquo; leucodelphinidins rsquo; possess structures analogous to the B-type procyanidin series1 The isolation of proanthocyanidin polymers from OH OH (1) (2 1 vegetative tissue may now be achieved under very mild conditions which apparently cause little degradation.15 The structure of the constituent monomer units of these polymers may readily be determined by capture by OH.ordquo;rdquo;OH OHOH + (3) phloroglucinol of the carbocation (3)generated from the polymer under mild acid condition^.^ RESULTS AND DISCUSSION Two proanthocyanidin polymers were isolated from the leaves of crown vetch (Coronilla varia L.) and flowering currant (Ribes sanguineurn Pursh.) .Each polymer is a rich source of prodelphinidin units, as shown by the delphinidin : cyanidin ratios of 2.5 : 1 and 6.8 : 1 respectively, determined by degradation with hot butanol and hydrochloric acid. These ratios are close to previously determined value^.^^ l5 Treatment of each polymer with phloroglucinol under acid conditions and separation of the products by chromatography yielded the prodelphinidin-phloro-glucinol adducts.The Ribes sanguineurn polymer yielded the phloroglucinol adduct (4) with a flavan unit of stereochemistry corresponding to that of (+)-gallo- catechin. The structure of the adduct was readily determined by the spectral properties of the phenol (4), nona-acetate (5), and octamethyl ether (6). The lH n.m.r. spectra of (4)-(6) were all identical with those of the corresponding derivatives of the phloroglucinol adduct of the carbocation possessing the (+)-catechin lo D. G. Roux, Nature, 1957, 179, 158. l1 M. Tsujimura, Sci. Pafiers Inst. Phys. Chem.Res., Tokyo,1929, 10, 253. K. Weinge?, W. Bahr, W. Ebert, K. Goritz, and H.-D. Marx. Fovtsclzr. Chem. org. Nuturstoffe, 1969, 27, 158. 1187 stereochemistry5 except for the H-2 and H-6 absorp-tions. The spectra of (5)and (6) showed large couplings in the heterocyclic ring, which defines the 2,3-trans-3,4- trans-stereochemis t ry ; the sharp two-pro t on singlet, 6 7.33 (acetate derivative) arises from the 2rsquo;-and 6rsquo;-protons, and thus defines the ring B oxidation pattern. ORrsquo; -Rrsquo;oooRrsquo;ORrsquo; Rrsquo; R* (4) H H (5) Ac Ac (6) Me H The CoroniZZa varia polymer yielded the ( -)-epigalloca-techin-phloroglucinol adduct (7) which was accompanied by the (-)-epicatechin-phloroglucinol adduct (9), pre-viously synthesised from Butea frondosa The structure of (7) was established by the close correspon- dence of the lH n.m.r.spectrum of its nona-acetate (8) with that of the octa-acetate derived from (9).5 ORrsquo; OH Rrsquo; RZ (9) (7) H H (0) Ac Ac The a,values for (5),(6),and (8) corresponded both in sign and in relative magnitude to those for the cor-responding procyanidin structure^,^ thereby establishing that the prodelphinidin adducts possess the same absolute configuration. The lower molecular weight flavonoids of the Ribes sanguineurn leaves extract were separated from the polymer by ethyl acetate extraction. Two-dimen-sional paper chromatography revealed that this extract contained at least eight flavonoids and flavonoid glyco- sides, in addition to eight components reactive to vanillin l3 E.Haslam in lsquo; The Flavonoids.rsquo; eds. T. J. Mabry, H. Mabry,and J. B. Harborne, Chapman and Hall, London, 1975, p. 505. l4 D. Jacques, C. T. Opie, L. J. Porter, and E. Haslam, J.C.S. Perkin I, 1977, 1637. 16 W. T. Jones, R. B. Broadhurst, and J. W. Lyttleton,Pitytocitemislry, 1976, 15, 1407. 1188 (flavans; see Table 1). The flavans included a major component, number 5, with a mobility characteristic of a proanthocyanidin dimer.5 TABLE1 Paper-chromatographic RF values of the major flavans of Ribes sanguineum leaves RF Component no. a b Structure 1C 0.59 0.52 Epicatechin2 0.45 0.43 Gallocatechin 3c 0.37 0.45 Epigallocatechin4 0.29 0.58 5c 0.27 0.34 Prodelphinidin B-4 6 0.19 0.48 7 0.10 0.31 8c 0.09 0.45 a t-Butyl alcohol-acetic acid-water (3 : 1 : 1 v/v).15 Acetic acid. Major components. On the basis of RP dataJ alone. The ethyl acetate soluble fraction was separated into a number of fractions containing monomeric flavanoids, and finally an homogeneous sample of the major pro- delphinidin (Table 1, number 5). The earlier fractions containing mixtures of lower molecular weight flavonoids were acetylated and separated by preparative t.1.c. This yielded the acetates of the major flavan-3-01, (-)-epigallocatechin (2), and a small amount of (+)-gallocatechin (1). Other flavonoids isolated were the HoQ'OH 0 (10) R = H (11) R = OH acetates of naringenin (lo), eriodictyol (ll),and two unidentified flavonoid acetates.The major prodelphinidin was treated with phenyl- methanethiol and acetic acid in ethanoL2 Paper chromatography revealed that it was rapidly cleaved to yield epigallocatechin plus a more mobile flavan. Treat-ment of the prodelphinidin with diazomethane yielded a decamethyl ether, M+' 750, confirming its molecular constitution. The major prodelphinidin also yielded a dodeca-acetate (12). The lH n.m.r. spectrum of this derivative established the 2,3-trans-3,4-trans-s t ereo- chemistry of the upper flavan unit (J2,39.6, J3,4 10.0 Hz). The 2,3-cis-stereochemistry of the lower flavan unit was established by the existence of distinct signals for H-2 and H-3 and the rather narrow envelope of signals for the C-4 methylene protons.This is in con- trast to the presence of a lower flavan unit with 2,3- trans-stereochemistry, where the H-2 and H-3 signals are coincident and those of H-4 are very broad. The presence of trihydroxylated B-rings was confirmed by the presence of two sharp, approximately two-proton singlets at 6 6.87 and 7.00, corresponding to the major rotational isomer, and a minor pair at 6 7.31 and 7.33. J.C.S. Perkin I The structure of the major prodelphinidin acetate therefore corresponds to (12) and by analogy with the procyanidins this prodelphinidin may be referred to as B-4 (13). The lH n.m.r. spectrum of (12) is in fact very similar to the published1 spectrum of procyanidin B-4 deca-acetate apart from differences in the aromatic region.OR (12) R = Ac (13) R = H The occurrence of a dominant proanthocyanidin with opposite stereochemistry for the upper and lower flavan units parallels procyanidin synthesis in Rubus species, where procyanidin B-4 dominates and is accompanied by (-)-epicatechin (2).295 Similarly Ribes sanguineurn contains an almost homogeneous prodelphinidin polymer containing gallocatechin units, and prodelphinidin B-4 (13) is accompanied by (-)-epigallocatechin (2) as the dominant monomeric flavan-3-01. This implies that prodelphinidin biosynthesis is probably under the same control as procyanidin biosynthesis, where a flav-3-en-3- 01 has been implicated as the key biosynthetic inter- mediat e.14J6 A further prodelphinidin containing a mixed ring B oxidation pattern was isolated from Salix ca$rea. The isolation of procyanidin B-6 from this source resulted in a fraction being obtained where B-6 was contaminated (14) with two other proanthocyanidins. Acetylation yielded the acetate of B-6 plus an homogeneous sample of the acetate of the major proanthocyanidin contaminant.The mass spectrum of the acetate revealed a molecular ion at m/e 1056, and this and subsequent fragmentations were consistent with a proanthocyanidin containing eleven hydroxy-groups. This was confirmed by the analytical figures and lH l6 E. Haslam, C. T. Opie, and L. J. Porter,Phytochemistry, 1977,16,99. n.m.r. spectrum, which was identical with that of procyanidin B-3 deca-acetate apart from a higher proton integration in the acetate region and a different appear- ance of the aromatic region, 6 6.9-7.2.The outstanding feature was a sharp, approximately two-proton singlet at 6 6.94, confirming the presence of a trihydroxylated ring B. However, two structural alternatives remained, viz. that the extra hydroxy-group could reside in either the upper or the lower flavan unit. This was resolved by the observation (see Experimental section) that the acetate functions are rapidly hydrolysed under the conditions of pigment generation from proanthocyanidins BunOH-5yo HC1 (4 : 1); 100 "C; 2 h and the acetate derivative yields the phenolic anthocyanidin. In fact the anthocyanidin may be generated from the phenol or the acetate with equal facility.This confirmed that the EXPERIMENTAL Mass spectra were measured on an A.E.I. MS30 instru-ment and 1H n.m,r. spectra were obtained at 60 MHz with a Varian EM 360 spectrometer. Sephadex LH-20 chro- matography with ethanol was carried out as described by Haslam and his co-workers .2 Paper chromatography was carried out in 6 acetic acid (solvent A) and butan-2-01- acetic acid-water (14 : 1 : 5 v/v) (solvent B). Anthocyani-dins were generated from the proanthocyanidins as des- cribed by Bate-Smith.ls Under the conditions of this reaction (butan-l-ol-5yo HC1; 100 "C; 2 h) the antho- cyanidin pigment was generated with equal facility from both proanthocyanidins and their acetate derivatives. The reaction was tested for several procyanidins and pro- delphinidins, and their acetates.Extraction of Prodelphinidin Polymers.-The leaves of Ribes sanguineurn (2200 g) were extracted in four batches TABLE2 lH N.m.r. data for flavan derivatives (6 J/Hz ; SiMe, internal standard; CDC1, solvent, unless otherwise specified) Compound Gallocatechin hexa-acetate Ring A 6.66, 6.70 H-6, H-8 Ring B 7.16(s) H-2', H-6' Phloroglucinolring-protons 2-H 3-H 4-H 2'-H 5.17(br s) 3'-H 5.1 7(m) 4'-H, 2.8-3.0(m) Acetates 2.16, 2.26 Epigallocatechin hexa-acetate 6.56, 6.66 P.01 7.20(s) 5.06(s) 5.40(m) 2.8-3.0(m) 2.16, 2.26 (4) a (5) 6.62, 6.79 5.95(s) P.21 ~2.31 6.65(s)7.33(s) 6.04(br s)7.02(s) 4.54b 5.00(d) 10.0 4.54) 5.78(t) 9.6 4.54b 4.70(d) 9.4 1.76, 2.00, 2.04, 2.33, 2.40 (9), acetate Prodelphinidin B-4 acetate Salin prodelphinidin acetate (12)p (14)f 6.02, 6.20 6.64, 6.78 6.60, 6.77 6.59, 6.77 AB quartet6.48(br s) 12.21 POI 12.41 6.69(s) 7.20(s) 7.22, 7.33 6.87(s), 7.00(s) 7.31(s), 7.33(s) 6.94(s)6.9-7.2 6.18(s) 6.84, 7.00 6.84, 6.93 6.67(s) *e 6.62(s) e ~2.41 P.51 4.62(d) 9.0 5.48(s) 5.47(s) 4.90(d) 4.49(d) 9.4 9.6 4.28(t) 9.0 5.14(dd) 5.16(dd) 5.79(t) 5.60(t) 9.7 9.6 4.68(d) 9.0 4.48(d) 4.48(d) ~2.51 2.4 4.63(d) 4.76 10.0 10.0 5.10(s) 5.06(br s) 5.31(m) 5.06(m) 2.8-3.9 2.4-3.0 3.40br, 3.42 3.75, 3.80 3.87, 3.92 c 1.86, 1.93 2.04.2.17 2.27; 2.34 1.87, 1.92, 2.04, 2.18, 2.30, 2.34 1.7-2.3 1.68, 1.93, 1.96, 2.24, 2.25, 2.32 a Solvent 'H- acetone. b Heterocyclic ring-proton chemical shifts are virtually coincident for both the procyanidin and prodelphinidin adducts; signal appears as a broad singlet.c A methyl ether; peaksinacetatecolumn refers tomethoxyresonances. d Apart from thering B resonances, the shifts for the major rotamer only could be assigned. e For lower flavan unit. f Major rotamer only. Salix caprea proanthocyanidin was a prodelphinidin and the acetate must possess structure (14). The occurrence of proanthocyanidins with flavan units of different ring B oxidation patterns is not novel. Apart from the considerable number of compounds isolated from Acacia species containing 5-deoxyflavan units, two other more pertinent examples exist. A proantho- cyanidin with (-)-epiafzelechin linked through C-4 to (-)-4'-O-methylepigallocatechin has been isolated from Ouratea, Prionostemma, and Maytenus species l7 and a procyanidin with (+)-catechin linked through C-4 to (-)-epiafzelechin was isolated from the fruit of Wistaria sinensis.I8 The above results almost certainly imply that in addition to the two ' normal' series of B-type pro- cyanidins and prodelphinidins possessing homogeneous oxidation patterns in the rings B of both the upper and lower units, two further series must exist with ' crossed ' oxidation patterns.These series, added to the rarer propelargonidins and the corresponding flavan-3-01s (+)-afzelechin and (-)-epiafzelechin, imply that at least 72 B-series proanthocyanidins possessing 5,7-dihydroxylated rings A exist in nature.l7 F. Delle Monache, M. Pomponi. G. B. Marini-Bettolo, I. L. d'Albuquerque, and 0.G. de Lima, Phytochernistvy, 1976, 15, 573. with acetone-water (7 : 3 v/v) according to the method of Jones et al.,15 with an additional six-fold ethyl acetate extraction after the light petroleum extraction. This procedure yielded 15 g of ethyl acetate-soluble flavonoids plus 40 g of crude freeze-dried prodelphinidin polymer. The Coronilla varia prodelphinidin polymer was isolated in a similar way.15 (2R,3S,4S)-4-(2,4,6-Trihydroxyphenyl)jIavan-3,3',4',5,5',-7-hexaol.-The Ribes sanguineurn polymer (20.0 g) and phloroglucinol hydrate (40.0 g) were stirred in ethyl acetate- O.~M-HCI(1: 4; 1000 ml) at 20 "C for 16 h. The aqueous phase was extracted with ethyl acetate (6 x 150 ml) to yield 25 g of crude product.The bulk of the phloro- glucinol was removed by crystallisation from ethanol, and the remaining ethanol solution was chromatographed on Sephadex LH-20 (5 x 40 cm) in ethanoL2 The eluate was collected as 15 ml fractions and these were combined to yield the crude adduct. Rechromatography in the same system on a smaller column yielded a chromatographically homogeneous sample of the product as a light tan powder (1.1g), Rp(A) 0.40, RRp(B)0.39. Reaction with butan-l-ol- HCl yielded delphinidin chloride and phloroglucinol. Acetylation of the phenol (acetic anhydride-pyridine) and purification by preparative t.1.c. silica; benzene-I* K. Weinges, K. Goritz, and F.Nader, Annalen, 1968, 715,164. 19 E. C. Bate-Smith, Phytochemistry, 1973, 12, 907. acetone (4: 1 v/v) gave a single product, RF 0.40, and precipitation from methanol-water yielded an amorphous nona-acetate (Found: C, 57.9; H, 4.4. C,gH,amp;, requires C, 57.9; H, 4.5); M+' 808; a58925-38.3"(c 0.2, CHCl,). Methylation (diazomethane in ether-methanol) yielded a major methyl ether from preparative t.1.c. silica; ethyl acetate-light petroleum (b.p. 40-60 "C) (6 :4 v/v), RF 0.58, and crystallisation from methanol yielded the octa- methyl ether, plates, m.p. 149-151 "C (Found: C, 64.4; H, 6.15. C2gH34010 requires C, 64.2; H, 6.3); m/e 542 (M)+*,333, 301 (loo),181, and 167; a58925-141' (c 0.25, CHCl,) . (2R,3R,4R)-4-(2,4,6-TrihydroxyphenyZ)jZavan-3,3',4',5,5',-7-hexaol.-The polymer from Coronilla varia (7.4 g) was treated with phloroglucinol hydrate (15 g) and the products were worked up and separated as described above.Chro-matography effected satisfactory separation from the accom- panying procyanidin adduct, but the product (0.60 g) RF(A) 0.39, RF(B) 0.381 could not be separated from procyanidin oligomers. The crude phenol (0.47 g) was therefore acetylated (acetic anhydride-pyridine) . Purific-ation by preparative t.1.c. silica; benzene-acetone (4 : 1 v/v) gave a major product with RF 0.49 and precipitation from methanol-water yielded the amorphous nona-acetate (Found: C, 57.6; H, 4.6. C,gHamp;g requires C, 57.9; H, 4.5); amp;I+' 808; a58925f83.3" (c.0.2, CHC1,). (2R,3R,4R)-4-(2,4,6-Trihydroxyphenyl)jZavan-3,3',4',5,7-pentaoZ.-The prodelphinidin adduct was accompanied by the corresponding procyanidin adduct, which was obtained as a tan solid (0.17 g),RF(A) 0.57, RF(B) 0.52. The phenol was acetylated to yield the octa-acetate, isolated and purified as above, RF 0.59 silica; benzene-acetone (4: 1 v/v, and crystallised from methanol; m.p. 154-155 "C, (lit.,5 148-150 "C) (Found: C, 59.15; H, 4.7. Calc. for Camp;,,O18: C, 59.2; H, 4.6); M+' 750. Separation of Ribes F1avanoids.-The ethyl acetate fraction (15 g) from the prodelphinidin polymer preparation was chromatographed on Sephadex LH-20 (5 x 40 cm) and the eluate was collected as 15 ml fractions. Prodelphinidin B-4. Fractions 275-400 yielded an almost homogeneous sample of the major prodelphinidin (0.53 g), RF(A) 0.38, RF(B) 0.28.Reaction with phenyl- methanethiol in acetic acid and ethanol yielded (-)-epigallocatechin, RF(A) 0.29, RF(B) 0.37, co-chromato-graphed with authentic material isolated from earlier fractions, plus two more mobile products, RF(A) 0.19, RF(B) 0.80, and RF(A) 0.22, RF(B) 0.86, presumably cor- responding to the benzylthio-derivatives of the gallocatechin carbocation.2 A homogeneous sample of the dodeca-acetate was obtained by reaction of the prodelphinidin with acetic anhydride- pyridine to yield a major product, RF 0.25 silica; benzene- acetone (4: 1 v/v), m.p. 154-156 "C (methanol) (Found: C, 57.9; H, 4.4. C,4H5002, requires C, 58.2; H, 4.5); a58925-77.0" (c 0.2, CHCl,).The prodelphinidin also yielded a deca-methyl ether (diazomethane in ether-methanol), m/e 750 (M+'),541, 509, 331, 299, 210, and 181 (100). Other FZavonoids.-The remaining fractions from the 2O J. W. Clark-Lewis, L. W. Jackman, and T. M. Spotswood,Austral. J. Chem., 1964, 17, 632. J.C.S. Perkin I chromatography of the ethyl acetate-soluble fraction all contained complex mixtures of flavonoids. The compo- nents were separated by acetylation (acetic anhydride- pyridine) of the mixtures and separation by preparative t.1.c. silica; benzene-acetone (4 : 1 v/v). Major flavo- noids found in each fraction were as follows. (1) Fractions 150-275 yielded eriodictyol, a dark u.v.- absorbent spot on paper, RF(A) 0.24, RF(B) 0.85.The tetra-acetate had RF 0.62, m/e 456 (M+'),414, 372, 330, 288, 153, and 136. The 'H n.m.r. spectrum revealed the characteristic ABX system for the heterocyclic ring pro- tons,20 G(CDC1,) H-2 5.50, H-3ax 3.02, and H-3eq 2.79 (J2.3(sx.) 13.2, J2,yeq.) 3-11 J3(eq.).3(ax.) 20.6 Hz). Eriodict~ol was accompanied by a small amount of naringenin, RF0.70, recognised from the mass spectrum of its triacetate, m/e 398 (Mf), 356, 314, 272, 153, and 120, and its 'H n.m.r. spectrum. (2) Fractions 75-125 contained a number of flavonoid glycosides and the major flavan-3-01 component, (-)-epigallocatechin, RF(A) 0.29, RF(B) 0.37, isolated as its hexa-acetate, RF 0.50; m/e 558 (M+'),516, 498, 456, 432, 414, 390, 372, 348, 330, and 288; a5se25-27.0' (c 0.24, CHCl,); the 'H n.m.r.spectrum (CDCl,) was identical to that of authentic (-)-epicatechin penta-acetate apart from the appearance of H-2' and H-6' signals as a two-proton singlet, 6 7.22. The fractions also contained a small amount of gallocatechin, RF(A) 0.38, RF(B) 0.46, which yielded a hexa-acetate, RF 0.60, with a mass spectrum identical with that of epigallocatechin hexa-acetate, and 'H n.m.r. spectrum (CDC1,) the same as that of authentic (+)-catechin penta-acetate apart from the H-2' and H-6' signals appearing as a two-proton singlet at 6 7.25. Prodelphinidin from Salix caprea Catkins.-During studies on the procyanidin polymer from Salix caprea a collection was obtained where the polymer contained significant amounts of prodelphinidin units.The ethyl acetate extract contained procyanidins B-3 and B-6 and other lower-mobility proanthocyanidins. Sephadex chro- matography yielded a sample of procyanidin B-6 mixed with three other proanthocyanidins, the major component of which was a prodelphinidin, RF(A) 0.43, RF(B) 0.34. The mixture could not be separated by further chromatography, and was acetylated; the products were separated pre- parative t.1.c. on silica in benzene-acetone (4: 1 vlv) to yield procyanidin B-6 hexa-acetate,5 RF0.46, plus the major prodelphinidin undeca-acetate, Rp 0.32, as an amorphous solid purified by precipitation from methanol-water (Found: C, 58.7; H, 4.8. C5,H4,024 requires C, 59.1; H, 4.6); M+' 1056, a58g20-68.8" (c 0.61, CHCl,). Reaction with butan-1-01-5yo HC1 (4: 1 v/v) at 100 "C produced delphinidin chloride. We thank Dr. W, T. Jones, Applied Biochemistry Divi- sion, Department of Scientific and Industrial Research, New Zealand, for a gift of CoroniZZa varia L. polymer; Dr. E. Haslam, Department of Chemistry, Sheffield University, for permission to publish the data on the Salix caprea pro- delphinidin ; Professor A. D. Campbell, University of Otago, for microanalyses; and Mr. S. A. Gwyn for mass spectra. 7/2014 Received, 15th November. 19773