THE CHEMISTRY OF THE RED AND BLUE PIGMENTS OF FLOWERS AND FRUITS. PART I11 E ~ S H.T H U N T ~ SMASSACAUSETTS S, INSTITUTE
OP
TECHNOLOGY, CAMBRIDGE,
MASSACHU~ETT~
Methylated Anthocyanins and Anthocyanidines Among the anthocyanins which were isolated by Willstatter were some from the flowers of the peony, petunia, wild mallow, hollyhock, and from the skins of the grape, huckleberry, and berries of the wild vine (Virginia creeper). The analyses of these substances convinced him that the corresponding anthocyanidines were mono or dimethyl ethers of cyanidine or delphinidine. The homogeneity of the glucosides was not beyond question in every case, however. Furthermore, upon attempting to locate the position of the methoxyl group by recognition of the fragments into which the anthocyanidine was split up by alkali fusion, it developed that the treatment was much too harsh and yielded inhomogeneous non-crystallime mixtnres in which the methoxyl groups had evidently been partially removed by hydrolysis. For this reason Willstatter was unable to fix the position of the methoxyl groups in the anthocyanidine molecule, and his published reports on these cases indicated that he was well aware of their shortcomings. Willstatter in this work on methylated anthocyanidmes had universally employed potash fusions a t 140-250'. Early attempts to effect the complete degradation of cyanidine and delphinidine with strong aqueous solutions of alkali a t 100-110" had apparently convinced him that while the phloroglucinol part of the molecule was readily obtainable the degradation of the other half of the molecule to a phenol carboxylic acid was incomplete. Upon attempting to find milder means of effecting this cleavage for the anthocyanidine methyl ethers, Karrer (Ref. 18) was surprised to iind that mere boiling for several hours with l(t1.570 sodium hydroxide, or even 10% barium hydroxide, in an atmosphere of hydrogen not only sutficed to yield the phloroglucinol component but gave the unsaponified methyl ethers of the corresponding phenol carboxylic acids. Since this procedure gave homogeneous crystalline acids in which the position of the methoxyl group was known, Karrer was able to assign definite constitutions to the cases which Willstatter had been unable definitely to characterize. As a result of this advance in technic Karrer offered convincing evidence that instead of a group of methylated anthocyanidines there have so far been isolated only two, a monomethyl ether of cyanidine and a dimethyl ether of delphinidine. The first is called peonidine from its occurrence as a diglucoside in the petals of the dark red peony: since alkaline hydrolysis by Karrer's method gives phloroglucinol and vanillic acid (XI) its structure
is definitely characterized as that represented in XII. The second on similar treatment gives phloroglucinol and syringic acid (XIII) : C1
OCH,
OCHa
OCHS H O O C ~ O H OCHz
HOOC-H
Syringic add (XIII)
Vanillic add OrI)
Syringidine chloride (=V)
the corresponding anthocyanidine is designated as syringidine (XIV).' During the progress of Karrer's work the synthesis of peonidine was reported from Robinson's laboratory (Ref. 5) : the method employed was an extension of the general synthetic procedure which had already proved its worth by the synthesis of the three fundamental anthocyanidmes. [See end of Part I, THISJOURNAL, 5, 1396 (Nov., 1928).] In the course of effecting this simplification in our knowledge of the methylated anthocyanidines Karrer has shown that the pigments supposed by Willstatter to have been pure individuals, were really mixtures of varying proportions of two anthocyanins, one the well-known delph'min, the other a diglucoside of syringidine mentioned above. Indeed the pigment obtained from the huckleberry, and supposed by Willstatter to be a galactoside of a distinct monomethyl ether of delphinidine (myrtillidine), was found to consist of a mixture of delphinidine and syringidine galactosides and glucosides, the anthocyanidines being in about equal proportion. Althaein from the hollyhock contained a similar mixture of glucose compounds of the two anthocyanidiies, but no galactose. In a group of three other pigments (oenin, cyclamin, and ampelopsin), syringidine forms 80-90% of the fundamental base. The pigment malvin, from the wild mallow, on the other hand, is notable as the most homogeneous of all the natural anthocyanins yet studied and consists exclusively of syringidine diglucoside. With these qualifications the occurrence of the methylated anthocyanins is indicated in Table IV. The parenthetical numbers refer to the bibliography at the end of this article. Karrer and Wididmer (Ref. 25) have recently reported the rrnurence in the primrose of an anthocyanin whose sugar-freepigment is a trimethyl ether of delphinidine in which two methoxyls are located as in syringidine: the third is in the phloroglucinol nucleus, hut whether in pmition 5 or 7 has not yet been determined.
TABLE IV ANTHOCYANIN DERIVATIVES oP METHYLATED ANTHOCY~IDINES Peonidinc (Cyanidine 3'-monomethyl ether1
I I
Svingidine (Dclphinidine 3',br-dimethyl ether1
Mo~oc~ucos~ms Ampelopsin-berries of wild vine (Virginia creeper) (23, 18) Oenin-grape
(20, 23, 18)
Cydamin--eyclamen (18) Althaein-hollyhock (21, 18) Mvrtillin-hncMeberry (20, 23, 18)
* WiUstitter (5) has also reported petunin as a diglucoside of a monomethyl delphinidine: in the light of Karrer's work, however, it is possible that it may be found on reexamination t o be derived from syringidine. Oenin, the syringidine monoglucoside which imparts the beautiful blue color of the grape, has a special interest. Willstatter (20, 23) had concluded that it was a monoglucoside of a dimethyl ether of delphinidine given by him the designation oenidine. In a paper submitted on November 23, 1926, for publication in the Helvetica Chimica Acta, Karrer (Ref. 18) has shown that while Willstatter's general characterization is correct, the anthocyanidine possesses neither of the alternative formulas suggested by Willstatter, but is simply syringidine. Eighteen days before the deposit of Karrer's paper for publication, Anderson and Nabenhauer of the New York State Agricultural Experiment Station a t Geneva, where experiments on the pigments of New York grapes have for some time been in progress, reported in the J. Am. Chem. Soc. (Ref. 29) that permanganate oxidation of the acetyl derivative of oenidine gave syringic acid and that the corresponding anthocyanidine consequently must possess the structure now designated by Karrer's term syringidine. This appears to add another to the already long list of instances in which substantially identical conclusions have been simultaneously reached by independent workers in widely separated laboratories.
The Position of the Carbohydrate Residue in the Anthocyanins It has thus been shown that although some twenty-eight substances responsible for the colors of red and blue flowers and fruits have now been isolated, they are glucosides of only five individual anthocyanidines, and two of these are very simple derivatives of certain of the remaining three fundamental parent substances. Examination of Tables I and IV discloses,
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CHEMISTRY OB REDAND BLUEPIGMENTS. I11
55
however, that within any given group there frequently occur several isomeric combinations of the same anthocyanidine with the same number of molecules of the same sugar. In some of these instances i t is probable that the anthocyanins will be found to be identical; in other cases, since the molecules are built up of the same units, the difference must involve the way in which these are put together. The answer to this new structural problem involves either the particular position a t which the carbohydrate is attached to the anthocyanidine or the position of the carbohydrate structure by which it is attached, or both. Karrer has recently reported (Ref. 19) great progress in the study of the first of these questions: i. e., the determination of the position on the anthocyanidine structure to which the carbohydrate residue is joined. By means of oxidation with hydrogen peroxide he endeavored so gently to break up the molecule of malvin that smaller fragments containing the carbohydrate residue in a recognizable position might be obtained. While the effects produced were not exactly in this sense, their consideration led to the preliminary hypothesis that the carbohydrate was attached t o position 3 of the anthocyanidine nucleus. This hypothesis was then brilliantly confirmed by another ingenious mode of attack involving methylation of the anthocyanins themselves, followed by subsequent removal of the sugar group and identification of the unmethylated position which it must have originally occupied. As a result it is now certain that in monardin, peonin, cyanin, and malvin a t least, and probably in the case of all the other anthocyanins as well, the carbohydrate residue is attached through the position 3-hydroxyl of the anthocyanidine nucleus (XV). OCH,
C1
Ho/Ad
I I
\-
Ol(=?Z
\
