Fuji, Kaoru

Fujita, Eiichi

Nakamura, Shigetake

Takaishi, Yoshihisa

Kyoto University Research Information Repository

TitleInvestigation of the Non-basic Constituents of Lythrum ancepsMAKINO (Commemoration Issue Dedicated to ProfessorMinoru Ohno On the Occasion of his Retirement)Author(s)Fujita, Eiichi; Fuji, Kaoru; Nakamura, Shigetake; Takaishi,YoshihisaCitationBulletin of the Institute for Chemical Research, KyotoUniversity (1972), 50(3): 206-208Issue Date1972-09-30URL http://hdl.handle.net/2433/76421RightType Departmental Bulletin PaperTextversionpublisherKyoto University  Bull. Inst. Chem. Res., Kyoto Univ. Vol. 50, No. 3, 1972          Investigation of the Non-basic Constituents of                 Lythrum anceps  MAKIN°               Eiichi FUJITA, Kaoru Fun, Shigetake NAKAMURA                        and Yoshihisa TAKAJSHI*                             Received May12,1972       Investigation of the non-basic constituents of Lythrum anceps MAKINO led to isolation and   characterization of ellagic acid and betulic acid.    We have investigated the alkaloidal constituents of Lyihr•urn anceps MAKINO and elucidated the structure and absolute configuration of the novel fourteen alkaloids isolated so far.t' Now, we carried out the investigation for the non-basic constituents of the same plant source.    The whole ground part of the flowering plant was extracted with methanol and the extract was separated into basic and non-basic fractions. The crude non-basic part was treated with methylene chloride to leave an insoluble solid, which was crystallized and purified as yellow needles, m.p.>360°. This compound was found to have the molecular formula C14H6O8i on the basis of analysis and mass spectral data. Its UV and IR spectra suggested the presence of aromatic ring(s), ester group(s), and phenolic hydroxyl group(s) in the molecule. On methylation with diazomethane it gave a crystal-line tetramethyl ether, while it, on acetylation with acetic anhydride and pyridine, afforded a crystalline tetra 0-acetate. Consideration of these facts and site of unsatu-ration led to an assumption that it might be ellagic acid (1).2) Hence, we synthesized ellagic acid and compared it with the natural product. Consequently, their complete identity was firmly established.    The methylene chloride soluble fraction was chromatographed on silica gel column to give betulic acid by elution with methylene chloride. Subsequent elution with acetone(10%)-methylene chloride gave a mixture of oleanolic and ursolic acids. The presence of Q-sitosterol and diisobutyl phthalate in the root was also recognized.                             OH                         HO              0             0                                   0       00 0                                OH                            OH                                1"` li ii • _  0J ,'i (i -r% • Laboratory of Physiological Activity, Kyoto   University, Uji, Kyoto. ( 206)           Investigation of the Non-basic Constituents of  Lythrum anceps MAKINO                           EXPERIMENTAL    Ellagic acid. The dried plant was extracted with methanol under reflux. The extract was concentrated to give a muddy residue, which was suspended with 0.5 hydrochloric acid and allowed to stand under occasional stirring. Filtration effected the separation of alkaloid from non-alkaloidal portion. The residue (non-alkaloid portion) was dried and extracted with methylene chloride. The methylene chloride soluble fraction will be described in the Betulic acid section. The solid which was insoluble in methylene chloride was dissolved in methanol and the solution was refluxed with charcoal. Usual treatment of the filtrate separated from charcoal gave a crystalline compound, which was recrystallized from methanol to yield a pure compound as yellow needles, m.p.>360° (Yield: 1.8% of the dried plant). Anal. Calcd. for C,4H5O8: C, 55.64; H, 2.00; M.W. 302. Found: C, 55.35; H, 2.09; M+ m/e 302. IR v;n'3e. cm-1 : 3550 (OH), 1700, 1200 (ester), 1620, 1580 (aromatic ring). UV %m;;X" nm 255 (e 40800), 366 (e 8800). On addition of alkali, the absorption shifted to the longer wave-length. Its identity with the synthesized ellagic acid was firmly established (m.p., IR, UV). Its methylation with diazomethane gave a tetramethyl ether, m.p. > 300°. Anal. Calcd. for C18H14O8: M.W. 358. Found: M+ m/e 358. A usual acetylation with acetic anhydride and pyridine afforded a tetraacetate, m.p.>300°. Anal. Calcd. for C22H14012: M.W. 470. Found: M+ m/e 470.    Synthesis of ellagic acid. Gallic acid (5 g) was dissolved in acetic acid (50 ml) and the solution was heated under reflux, to which sulfuric acid (2.5 ml) was added. Subsequently, finely powdered potassium persulfate (5 g) was gradually added to the mixture. A somewhat violent reaction occurred and the clear solution rapidly changed to brown. The reaction mixture was allowed to stand for 30 min, after the reaction got gentle. Then, the mixture was poured into water and the precipitate was collected. After its treatment with charcoal in methanol at reflux and filtration, methanol was evaporated off to leave a crystalline residue, which was recrystallized from methanol to yield yellow needles (1.8 g), m.p. > 360°. Anal. Calcd. for C14H608 : C, 55.64; H, 2.00. Found: C, 55.42; H, 2.06.    Betulic acid. The foregoing methylene chloride soluble fraction was chromato-graphed on silica gel column. The methylene chloride eluent gave betulic acid as colorless needles, m.p. 283-285° (yield 1.4 g from 50 g of the methylene chloride soluble substance). Liebermann-Burchard test: positive. Anal. Calcd. for C30H4803: M.W. 456. Found: M+ m/e 456. IR vmBx. cm-1: 3460, 1690, 1645, 890. NMR Oppm (CDC13): 0.7-1.3 (6xCH3), 3.45 (1H, t, J=8 Hz), 4.64, 4.73 (each 1H, s, =CH2). Its methylation with diazomethane gave methyl betulate as colorless needles, m.p. 225-227°. Anal. Calcd. for C31HSO03: C, 79.10; H, 10.71. Found: C, 78.85; H, 10.68. The comparison with an authentic sample of methyl betulate confirmed their identity (m.p., m.m.p., IR and NMR spectra).    Oleanolic acid and Ursolic acid. The acetone (10 %)-methylene chloride eluent of the foregoing chromatography gave colorless needles, m.p. 232-236° (yield 0.5 g from 50 g of the methylene chloride soluble substance). Liebermann-Burchard test: (207) E. FUJITA, K. Fun, S. NAKAMURA AND Y. TAKAISHI         positive. Anal. Calcd. for C30H4803: M.W. 456. Found: M+ m/e 456. IR vmax. cm-1        3400, 1690. NMR Sppm (CDC13): 0.7-1.7 (7xCH3), 3.5 (1H, t, J=8 Hz), 5.5 (1H, s).        The gas chromatography of the methylated product (CH2N2) showed it to be a mixture        of methyl oleanolate and methyl ursolate (1:1) by comparison of the retention time        with the authentic samples. 9-Sitosterol. From the non-basic fraction of the root, a-sitosterol was isolated         as colorless needles, m.p. 137.5-139.5° by a column chromatography on alumina. The         acetate, m.p. 122-124°. Anal. Calcd. for C31145202: C, 81.52; H, 11.48. Found C, 81.54;         H, 11.44. Its comparison with the authentic sample of p-sitosterol acetate confirmed         their identity (m.p., m.m.p., and IR spectrum).            Diisobutyl phthalate. From the benzene eluent of the alkaloid fraction of the         root, an oily neutral substance was isolated. IR vuaaz. cm-' : 1725, 1608, 1580, 1290.        NMR Sppm (CDC13) 0.98 (I2H, d, J=6 Hz), 2.0 (2H, m), 4.1 (4H, d, J=6 Hz), 7.5        (4H, m). Its hydrolysis with 10 % KOH-MeOH under reflux for 5 hrs. gave phthalic        acid. The retention time of its gas chromatogram was completely identical with that        of the authentic sample of diisobutyl phthalate.                                 ACKNOWLEDGMENT            We thank Prof. P. de Mayo, Univ. of Western Ontario, for the gift of betulic acid        and Prof. H. Inouye, Kyoto University, for the samples of oleanolic acid and ursolic          acid.                                    REFERENCES             (1) a) E. Fujita, K. Fuji, K. Bessho, A. Sumi, and S. Nakamura, Tetrahedron Lett., 4595             (1967);                b) E. Fujita, K. Bessho, K. Fuji, and A. Sumi, Chem. Pharm. Bull. (Tokyo), 18, 2216 (1970);                c) E. Fujita, K. Fuji, K. Bessho, and S. Nakamura, ibid., 18, 2393 (1970);               d) E. Fujita, K. Fuji, and K. Tanaka, Tetrahedron Lett., 5905(1968); idem., J. Chem. Soc.             (C), 205 (1971);               e) E. Fujita and K. Fuji, J. Chem. Soc. (C), 1651 (1971);               f) E. Fujita, K. Bessho, Y. Saeki, M. Ochiai, and K. Fuji, Lloydia, 34, 306 (1971);                g) E. Fujita and Y. Saeki, J. Chem. Soc. (D) (Chem. Comm.),368 (1971);                h) idem., Cheni. Pharm. Bull. (Tokyo), 19, 1515 (1971);                i) idem., J. C. S. Perkin I, in press.             (2) P. Nierenstein, J. Chem.Soc., 87, 1412 (1905). (208)

Investigation of the Non-basic Constituents of Lythrum anceps MAKINO (Commemoration Issue Dedicated to Professor Minoru Ohno On the Occasion of his Retirement)

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Investigation of the Non-basic Constituents of Lythrum anceps MAKINO (Commemoration Issue Dedicated to Professor Minoru Ohno On the Occasion of his Retirement)

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