CC BY
79
DOI: 10.14258/jcprm.2018033717
UDC 582.632+547.913.5+547.914.4
6-HYDROXYISOCARYOPHYLLENE AND ISOCARYOPHYLLENIC ACID FROM BIRCH VEGETATIVE BUDS
© D.N. Vedernikov1, S. V. Teplyakova1, O. V. Khoroshilova2
1St. Petersburg State Forest Technical University named after S.M.Kirov, Institutsky per., 5, St. Petersburg, 194021 (Russia), e-mail: [email protected] 2Saint-Petersburg State University, Institute of Chemistry of St. Petersburg State University, Universitetsky Prospekt, 26, Peterhof, Petrodvortsovy borough, St. Petersburg, North-West Federal District, 199034. (Russia), e-mail: [email protected]
The new isocaryophyllene derivatives: 6-hydroxyisocaryophyllene [(lR,4Z,6R,9S)-8-methylene-ll,ll-dimethyl-bicyclo[7.2.0]undec-4-ene-6-ol], epoxide of 6-hydroxyisocaryophyllene [(6R)-hydroxy-(4R,5S)-epoxyisocaryophyllene [(lR,4R,5S,6R,9S)-4.5-epoxy-8-methylene-ll,ll-dimethylbicyclo[7.2.0]undeco-4-ene-6-ol], (6R)-acetoxyisocaryophyllene [(6R)-acetoxy-(lR,4Z,9S)-8-methylene-ll,ll-dimethylbicyclo[7.2.0]undec-4-ene], isocaryphyllenic acid [(1R,4E,9S)4-carboxy-8-methylene-ll,ll-dimethylbicyclo[7.2.0]undeco-4-ene] were first detected in the birch vegetative buds. 6-Hydroxyisocaryophyllene and epoxide of 6-hydroxyisocaryophyllene are isolated by from the Betula pendula Roth, birch buds etheric extract by chromatography on silica gel. (6R)-Acetoxyisocaryophyllene was synthesized. The structure of 6-hydroxyisocaryophyllene and epoxide of 6-hydroxyisocaryophyllene isolated from the buds were determined by NMR spectroscopy. Caryophyllenic acids are isolated from the ether extract with an aqueous solution of alkali. Caryophyllenic acids are separated by chromatography on silica gel. The structures of caryophyllenic acid and isocaryphyllenic acid isolated from the Betula grandifolia Litv., B. albo-sinensis Burk., B.fusca Pall, ex Georg, B. obscura A. Kotula, B. litwinowii Doluch., B. hallii Howell, B. grandifolia Litv. birch buds were determined by X-ray diffraction analysis. The physico-chemical characteristics and NMR data of 6-hydroxyisocaryophyllene, epoxide of 6-hydroxyisocaryophyllene and all the isolated acids are given. The obtained mixtures of compounds were analyzed by gas chromatography - mass spectrometry (GC-MS). The gas chromatographic retention indices of all identified compounds were determined.
Keywords: 6-hydroxyisocaryophyllene, epoxide of 6-hydroxyisocaryophyllene, isocaryophyllenic, caryophyllenic acid, vegetative buds of birch, NMR spectroscopy, X-ray diffraction analysis, gas chromatography - mass spectrometry.
We have earlier reported the hydrocarbon extract sesquiterpene alcohols composition of pendent white birch (.Betula pendula Roth.) vegetative buds. The fraction contains 6-hydroxycaryophyllene, 6-hydroxyhumulene, 14-hydroxycarophyllene [1]. The main components of the investigated birch 14-hydroxy-caryophyllene, 6-hydroxycarophyllene and their esters. But on the GC-MS chromatogram, there was an intense peak of an unknown compound whose mass spectrum was similar to the mass spectrum of 6-hydroxycaryophyllene. The same pattern was observed in the analysis the birch buds Betula grandifolia Litv., B. albo-sinensis Burk., B. fusca I'all.ex
Khoroshilova Olesya Valerievna - graduate student, specialist structure of compounds that give similar mass spectra, of RSA, e-mail: [email protected]
Corresponding author.
Introduction
Vedernikov Dmitriy Nikolaevich - Department of Technology of Wood Chemical Products, Wood Chemistry and Biotechnology, Professor, D.Sc., Associate Professor, e-mail: [email protected]
Teplyakova Svetlana Vitalievna - Department of Technology of Wood Chemical Products, Wood Chemistry and Biotechnology, graduate student, e-mail: [email protected]
Georg, B. obscura A. Kotula, B. litwinowii Doluch. , birch of golden, bog birch B. hallii Howell, B. grandifolia Litv. extract obtained by extraction with ether. Two peaks of other compounds were present on chromatograms. Whose mass spectra were little distinguishable. The aim of this work was to establish the
Experimental
Vegetative buds of pendent white birch Betula pendula (100 g (65 g absolutely dry buds) of buds) were collected in April 2016 in the Kirishi district of the Leningrad Region. Buds of bog birch B. hallii (28 g (17.4 g a.d.b.)) were harvested in March 2017 in the arboretum of St. Petersburg Forestry University. The wet, crushed buds were extracted with methyl tert-butyl ether (MTBE) in a Soxhlet apparatus. The yield of extract from a.d.b.of pendent white birch was 41%, of the bog birch buds - 42%. The extracts were treated with a 1% aqueous solution of NaOH to remove acids. The yield of acids from the pendent white birch extract was 33%, the acid yield from the bog birch extract - 87%. The obtained neutral substances of the extract (17.8 g) of pendent white birch buds and the acids of bog birch extract (6.36 g) were separated by preparative liquid chromatography on silica gel with gradient elution using petroleum benzine (PE) (boiling range 40-70 °C) as an eluent with the addition of up to 30% MTBE. The fraction of sesquiterpene alcohols (8.35% of the MTBE extract) eluted from the PE column supplemented with 5% MTBE. The fraction was re-chromatographed into individual compounds. Unknown sesquiterpene alcohol (0.120 g) (1) (eluted earlier than other sesquiterpene alcohols: 6-hydroxycaryophyllene and 6-hydroxy-gumulene). The fraction of oxides of sesquiterpene alcohols eluted with the addition of 7% MTBE.
Sesquiterpene acids eluted PE with an addition of 25 to 30% MTBE. The acid (3) (2.21 g) was eluted before the acid (4) (2.42 g). The acid fractions were methylated with diazomethane before analyzing by gas chromatog-raphy-mass spectrometry (GLC-MS) since the acids did not appear as peaks in the chromatogram.
The chromatographic analysis was carried out using a 6850A Agilent chromato-mass spectrometer (Agilent Technologies, Inc.) with a model G2629A gas chromatograph equipped with a model G2577A HP5973 Network selective mass spectrometry detector. The flow rate of carrier gas (He) was lmL per min. Samples of 2 |iL were injected in the split mode at a ratio of 20 : 1. The injector temperature was 270 °C. The transfer line were kept at 270 °C. The quadrupole temperature was 270 C. The ion source was kept at 230 °C. The ionizing energy was 70 eV. Mass range was from 40 to 550 m/z.
To fractionate samples, a HP-5MS quartz column (30 m x 0.25 mm; film thickness 0.25 |im) with a 5% phenylmethyl-siloxane stationery phase was used. The thermostat temperature was programmed to increase from 100 to 270 °C at a rate of 5 °C min"1. The oven was held at this temperature for 30 min.
The gas chromatographic retention indices (RI) of the analyzed substances were determined using the retention indices of «-alkancs (Ci6- Ci9) as standard compounds (Sigma-Aldrich)). The standard compounds were chosen so that the retention times of the studied substances fell between those of the reference alkanes. Retention indices were calculated following the determination of the coefficients of the following equation: RI = ax2 + bx + c, where RI and x represent the retention index and retention time, respectively. All calculations were performed using an Advanced Grapher program (version 2.08).
High resolution mass spectrum (HR-ESI) was recorded on a Bruker-micrOTOF instrument using an elec-trospray method. The scanning interval is 50-1200 m/z. The ion polarity is positive, the voltage of the ion source capillary is 4500 V, the gas pressure at spraying is 0.4 bar, and the dry gas flow rate is 4.01 / min. The solvent was methanol.
NMR spectra were recorded using a NMR spectrometer Jeol ECX-400A (400 and 100 MHz for 1H and 13C spectra, respectively) and CDC13 as a solvent (5-scale). As internal standards residual signals CHC13 (5 H 7.25 ppm) were used.
IR spectra were recorded on a FTIR-8400S Shimadzu instrument using the FT-IR reflection technique.
Optical rotation angle was determined on the device: Automatic Polarimeter AA65. Solvent-chloroform. The length of the cuvette is 3 cm.
Thin layer chromatography was performed on Merck Silica gel 60 F254 plates. The spots on the TLC plates were sprayed with a 10% solution of sulfuric acid in ethanol with the addition of vanillin.
For single crystal X-ray diffraction experiment crystals were fixed on a micro mount and placed on an Agilent Technologies SuperNova (4), (5) using CuKa and MoKa monochromated radiation diffractometers, respectively. All of crystals were measured at a temperature of 100K. The structures were solved by the Direct methods and refined by means of the SHELXL program [2] incorporated in the OLEX2 program package [3]. The crystal-lographic data and some parameters of refinement are placed in Table 1. Empirical absorption correction was applied in CrysAlisProprogram complex [4] using spherical harmonics, implemented in SCALE3 ABSPACK scaling algorithm. Supplementary crystallographic data for this paper have been deposited at Cambridge Crystallographic
Data Centre (CCDC 1583801. 1583802 for isocaryophyllenic acid (4) and caryophyllenic acid (5), respectively) and can be obtained free of charge via www.ccdc.cam.ac.uk/data_request/cif. Physic-chemical and Spectral Characteristics of the Extracted Compounds
(6R)-Hydroxyisocaryophyllene [(lR,4Z,6R,9S)-8-methylene-ll.ll-dimethylbicyclo[7.2.0]undec-4-ene-6-ol] (1). Oil, RI= 1642. IR-spectrum (v, sm1): 3385(OH), 3057(=CH), 2953(CH2), 2923(CHZ), 2851(CH2), 1713, 1708, 1633(C=CH2), 1462(-С(СНз)=СН-), 1378, 1367, 1265, Ю25(СН-ОН), 894(C=CH2), 740, 704, 581.
Mass-spectrum (m/z): 220 (1), 205 (7), 187 (11), 177(11), 149 (21), 131 (43), 121 (41), 109(74), 107(45), 105 (46), 95(58), 93(54), 91(69), 79(81), 69(79), 41(100).
High resolution mass spectrometry (HR-ESI): m/z 243.1720(100) [M+Na]+, 244.1752(16.3) [M+Na+1]+, 245.1782(1.3) [M+Na+2], Mass calculation C15H240: m/z 243.1725(100), 244.1758(16.2), 245.1792(1.2).
(6R)-Acetoxyisocaryophyllene [(6R)-acetoxy-(lR,4Z,9S)-8-methylene-ll.ll-dimethylbicyclo[7.2.0]-undec-4-ene] (2). Oil, RI= 1742. [a]D20+32.8° (c= 0.945. CHC13). IR-spectrum (v, см1): 2950, 2929, 2859, 1737 (C=0), 1630.1451. 1368.1242 (0-C=0), 1020. 964. 888(C=CH2), 850.
Mass-spectrum (m/z): 262 (<1), 247(<1), 220(3), 202(17), 189 (21), 159(31), 145(23), 133(76), 131(61), 119(32), 109(46), 105(51), 91(63), 79(51), 67(21), 65(14), 55(23), 43(100).
(6R)-hydroxy-(4R,5S)-epoxyisocaryophyllene [(lR,4R,5S,6R,9S)-4.5-epoxy-8-methylene-ll.ll-dimethyl-bicyclo[7.2.0]undeco-4-ene-6-ol] (3). Oil, RI=1808.
Mass-spectrum (m/z): 236 (<1), 221(2), 205(3), 203(6), 193(4), 189(5), 175(8), 161(12), 147(13), 135 (14), 121(45), 109(61), 95(76), 82(50), 79(78), 69(100), 55(47), 41(94), 29(18).
High resolution mass spectrometry (HR-ESI): m/z 259.1674(100) [M+Na]+; 260.1708(16.5) [M+Na+1]+, 261.1741(2.0) [M+Na+2]+. Mass calculation C15H2402 +Na: m/z 259.1674(100), 260.1708(16.2), 261.1741(1.2).
lU NMR in CDC13 (5): 0.99 (3H, s, H-12), 1.00 (3H, s, H-13), 1.52 (3H, s, H-14), 1.55 (1H, dd, J9-ioa=9.2, H-lOa), 1.56 (2H, m, H-2), 1.67 (lH,m, H-3a), 1.76 (1H, dd, J10a-iop=10.5, 9-iop=9.2. H-10(3), 1.77 (1H, ddd, Ji_9=9.2, i.2a=11.7, i_2|3=3.5, H-l) 1.91 (1H, dd, J6-7a=9.4,7a_7p=12.0, H-7a), 1.96 (1H, dd, J6.7p=3.4. H-7(3), 2.04 (1H, m, H-3P), 2.50 (1H, ddd, J9-ioa=8.9; J9_10p=10.0, H-9), 2.86 (1H, d, J5-6=4.1, H-5), 4.30 (1H, ddd, J6.7p=6.5, H-6), 5.03 (1H, br.s, H-15(3), 5.15 (1H, br.s H-15a).
13C NMR in CDCI3 (5): 16.9 (q, C-14), 22.4 (q, C-13), 27.6 (t, C-2), 29.9 (q, C-12), 32.9 (s, C-ll), 40.00 (t, C-3), 40.2 (t, C-10), 41.4 (t, C-7), 46.9 (d, C-9), 49.3 (s, C-4), 55.4 (d, C-l), 64.2(d, C-6), 67.6 (d, C-5), 147.7 (s, C-8), 114.0(t, C-15).
Isocaryophyllenic acid [1 R,4E,9S)4-carl)oxy-8-methvlene-l 1.11-dimcthylbicvclo [7.2.0]undeco-4-ene] (4).
Tm = 103-104 °C (from hexane). RI of methyl ester - 1726. [a]D26 -46.1° (c=1.447. CHC13). IR-spectrum (v, см1): 2947, 2920, 2854, 2650, 2565, 1674(C=C-COOH), 1635(C=C), 1450, 1423(HRC=CH2), 1307, 1288, 1257, 1226,
1199, 1188, 941, 922, 887, 771, 740, 532.
Mass-spectrum of methyl ester (m/z): 248(6), 233(18), 216(6), 201(11), 189(35), 179(63), 173(25), 165(13), 156(2), 147(88.5), 133(76), 119(57), 105(60), 91(100), 79(61), 69(79), 59(14), 53(28).
High resolution mass spectrometry (HR-ESI): m/z 235.1697(100) [M+H]+, 236.1740(16.6) [M+H+l]+, 237.1774(1.3) [M+H+2J+. Mass calculation C15H2202 +H: m/z 235.1698(100), 236.1732(16.2), 237.1765 (1.2). TLC: system: hexane-MTBE=10:3. RpO.55 (pink).
Caryophyllenic acid [lR,4Z,9S)4-carboxy-8-methylene-ll.ll-dimethylbicyclo [7.2.0]undeco-4-ene] (5).
Tm = 90-91 °C (from hexane). RI of methyl ester - 1688. [a]D26 -31.5 (c=1.642. CHC13). IR-spectrum (v, см"1): 3070, 3016, 2924, 2854, 2781, 2604, 1670(C=C-COOH), 1628(C=C), 1439, 1419(HRC=CH2), 1369, 1303, 1269,
1200, 1169, 956, 933, 887, 783, 624, 574.
Mass spectrum of methyl ester (m/z): 248(2), 233(16), 216(6), 201(9), 189(31), 179(60), 173(25), 165(14), 156(3), 147(100), 145(30), 133(62), 119(72), 105(57), 91(96), 79( 80), 69(75), 59(14), 53(28).
High resolution mass spectrometry (HR-ESI): m/z 235.1697(100) [M+H]+, 236.1740(16.6) [M+H+l]+, 237.1774(1.3) [M+H+2J+. Mass calculation C15H2202 +H: m/z 235.1698(100), 236.1732(16.2), 237.1765(1.2). TLC: system: hexane-MTBE=10:3. RpO.63 (lilac).
Caryophyllenic acid pyrazoline methyl ester кислоты (6).Тпл=113-115 °C (from hexane). RI=1731. IR-spectrum (v, см"1): 2950, 2861, 1737, 1639, 1463, 1438, 1373, 1248, 1208, 1171, 1047, 891.
High resolution mass spectrometry (HR-ESI): m/z 291.2067 (100) [M+H]+; 292.2100(19.6) [M+H+l]+; 293.2133(2) [M+H+2J+. Mass calculation: C17H26N202+H: m/z 291.2103 (100); 292.2134(18.4); 293.2094(1.6);
main peaks: m/z 313.1886 (100) [M+Na]+; 314.1919(20.0) [M+Na+1]+; 315.1952(2.0) [M+Na+2]+. Mass calculation: C17H26N202+Na: m/z 313.1894 (100); 314.1928(18.4); 315.1961(1.6).
Mass spectrum (m/z): 262 (10), 247(10), 234(5), 219(13), 215(13), 205(10), 203(34), 193(8), 187(20), 173(11), 161(39), 147(47), 133(66), 119(33), 109(20),107(100), 93(57), 91(87), 79(85), 69(39), 41(59).
13 C NMR in CDC13 (8): 21.9 q (C-13) 29.8, q (C-12), 23.9 t (C-6), 25.9 t (C-2), 34.5 s (C-ll), 36.3 t(C-3), 36.4 t (C-7), 36.91 (C-10),36.9 d (C-9), 42.7d (C-l) 52.0 q (OCH3), 58.8 d (C-5), 83.6 t (CH2-N=N), 98.4 s (C-4), 111.2 t (C-15), 151.2 s (C-8), 168.0 s (C-14).
Results and discussion
The structure of compound (1) was established on the basis of two-dimensional NMR spectra and analysis of the spin-spin interaction constants. We were compared the NMR spectra of the two compounds, since the mass spectrum (1) practically coincides with the mass spectrum of (6R) -hydroxycaryophyllene [(lR,6R,4E,9S)-8-methylene-11,1 l-dimethylbicyclo[7.2.0]undeco-4-ene-6-ol, RI = 1670]. In the 13C NMR and 13C NMR spectra of DEPT 135. a similar set of carbon signals associated with different numbers of hydrogen atoms was present, as in the spectrum of 6-hydroxycaryloryllene [5]. In the 1H NMR spectrum, proton signals of three methyl groups with centers at 0.98; 0.97; 1.62 ppm were observed (Table 1). The last signal belongs to the protons of the CH3-group, which is bonded to the carbon atom (C-4) of the double bond. The structure also contained an exomethylene group, as evidenced by proton signals at 4.92 and 4.85 ppm, trisubstituted double bond (proton signal at 1H at 5.20 ppm (H-C5)) and adjacent secondary alcohol group (proton signal at 1H at 4.55 ppm (H-C6)). The proton signal at the carbon atom (C-6) bound to the OH group was noted as a triplet of doublets, and the proton signal at the double bond carbon atom in the cyclic structure gave the signal as a doublet. Thus, the structure of the compound should have the following fragment:
—C=CH-CH-CH2— I I 2
CH3 OH
Two doublets of doublets with the intensity of one proton with centers at 2.55 and 2.42 ppm on the NMR spectrum refer to the protons of the methylene group next to the carbon atom with the secondary hydroxyl. The second side of the methylene group should contain a completely substituted carbon atom (C-8) (the absence of the protons interaction of the methylene group with other protons). The displacement of the signals of the protons of the methylene group into a weaker field suggests a close proximity of the exomethylene group. That is, in the structure of the molecule of the compound there is a fragment:
—C=CH-CH-CH2—C=CH2 I I I
CH3 OH 1
The isolated compound is a derivative of caryophyllene, since on the 1H NMR spectrum, the signals of methyl groups in the form of singlets with centers of 0.97 and 0.98 ppm (3H-C13, 3H-C12) proton signals at carbon of two double bonds are visible. One combination of atoms refers to the exomethylene group, and another combination of atoms belongs to the group with trisubstituted carbons, which is characteristic of caryophyllene.
A secondary hydroxyl group is present in the structure of the compound. The hydroxyl group should be located next to the double bond. Two-dimensional COSY NMR spectrum was obtained to confirm the existence of the above assumed fragments, which showed interactions between neighboring hydrogen atoms. Two-dimensional spectrum of C-H was obtained to assign hydrogen signals to specific carbon atoms. The fragments found were similar to the (6R)-hydroxycaryophyllene fragments, but chemical shifts and spin-spin interaction constants differed. Thus, J5_6 for hydroxy-caryophyllene is 10.4 Hz, and for the isolated compound it is 7.7. which makes the signal narrower. It was assumed that the isolated compound is (6R)-hydroxyisocaryophyllene.
Two-dimensional NMR spectrum of NOESY was analyzed to confirm the structure. The cross peaks that appeared on the spectrum corresponded to the structure of (6R)-hydroxy-isocaryophyllene (Fig.l). Earlier it was reported that 14-hydroxyisocaryophyllene and isocaryophyllene-14-al were found in essential oils of various birches, including pendent white birch [6].
Fig. 1. Selected NOE correlations of the compound (1)
Table 1. Data on the 'H NMR and 13C NMR spectra of the isolated compounds (1) and (2) (5H, 5C)
The isolated alcohol was proacetylated with a mixture of acetic anhydride and pyridine and the obtained product (2) was found on the chromatogram of the hydrocarbon extract of the buds, since acetates of sesquiterpene alcohols are present in the buds of birch [7].
The epoxide (3) corresponding to the alcohol (1) is also present in the birch bud extract, and was isolated by preparative chromatography. The epoxy of 6-hydroxycaryophyllene is also present in the extract of the birch buds [7].
J/Hz: for (1): 1-9=9.1; l-2a=11.9; 1-2(3=3.7; 2a-3(3=8.9; 2(3-3(3=4.0; 2a-2(3=13.8; 2(3-3a=4.5; 3a-2a=4.6; 3a-3(3=13.6; 3(3-14=1.1; 15a-15(3=1.5;
5-14=2.0; 5-6=7.7; 6-7(3=3.6; 6-7a=8.4; 7a-7(3=13.0; 9-10a =9.1; 9-10(3= 9.6; 10a-10(3 =10.6;
for (2): 1-9=9.1; l-2a=11.0; 1-2(3=4.0; 2a-3(3 =8.9; 2(3-3(3=4.0; 2a-3(3=8.9; 2a-2(3=13.6; 2(3-3a=4.5; 2a-3a=4.9; 3a-3(3=13.2; 3(3-14=1.1; 6-7a =8.4;
6-7(3=3.6; 6-5=7.7; 7a-7(3 =4.0; 9-10a =9.1; 9-10(3 = 10.2; 10a-10p =10.6; 15(3-7a=1.2
The isolated compounds of the bog birch buds are acids, since the compounds were shown on GC-MS chromatograms only after preliminary methyla-tion with diazomethane, dissolved in an aqueous-alkaline solution when extracting the etheric extract of the buds. Further, the structure of the isolated acids was established by X-ray structural analysis. The analysis showed that both compounds crystallize in PE at + 4 °C (Table 2) and form crystals consisting of two conformers in a 1 : 1 ratio (Fig.2. 3).
Two-dimensional spectra of ^^H ROESY, COSY, HMQC, HMBC were taken to establish the correspondence of carbon and hydrogen atoms to the signals on the NMR spectra, The results are shown in Table 3 below and Figures 4 and 5.
Acids (4) and (5) are geometric isomers that crystallize, forming conformers
The :H NMR spectrum of compound (5) was similar to the spectrum given for caryophyllenic acid isolated from extracts of a plant of the genus Lichnofora [8].
C atom 1 2
SC,ppm SH, ppm SC,ppm 5H, ppm
1 49.4 d 1.69 ddd 50.1 d 1.68 ddd
2 25.41 a 1.62 m P 1.52 m 25.5 t oil.58 m (31.54 m
3 29.21 a 1.83 m P 2.30 m 29.2 t al.94 m P 2.44 m
4 136.0 s - 138.0 s -
5 130.0 d 5.20 d 125.5 d 5.14d
6 70.9 d 4.55 ddd 72.3 d 5.60ddd
7 45.41 a 2.55 dd P 2.42 dd 42.5 t a 2.39 dd P 2.56 dd
8 151.9s - 151.0 s -
9 40.3 d 2.44 ddd 39.8 d 2.47 ddd
10 39.71 a 1.53 dd P 1.72 dd 39.7 t al.56 dd (31.72 dd
11 33.0 s - 33.0 s -
12 29.9 q 0.98 s 30.lq 0.98 s
13 23.0 q 0.97 s 23.2 q 0.99s
14 22.8 q 1.62 s 22.9 q 1.63 s
15 112.3 t a 4.92 br.s P 4.85 br.s 113.2 t a 4.91 br.s P 4.97 br.s
CH3COO - - 21.6 q 2.04 s
CHjCOO - - 170.5 s -
Table 2. Summary crystallographic data for (4) and (5)
Compound (4) (5)
Empirical formula C15H22O2 C15H22O2
Formula weight Orthorhombic Monoclinic
Temperature/K 6.1029(3) 9.2537(2)
Crystal system 13.0180(5) 11.4795(3)
Space group 34.244(2) 13.4148(4)
a/A 90 90
b/A 90 104.500(3)
c/A 90 90
aJ° 2720.6(2) 1379.64(6)
p/° 234.32 234.32
y/° P2i2i2i P2i
Volume/A3 0.580 0.073
Z 100(2) 100(2)
Pcalcg/Cm3 8 4
¡i/mm"1 1.144 1.128
F(000) 0.15 x 0.05 x 0.05 0.8x0.3x0.15
Crystal size/mm3 CuKa MoKa
Radiation 12799 30724
Angle range 26(°) 5365 6308
Total reflections 7.264-144.970 5.998-55.000
Unique reflections 4386 6133
Reflections with I>2a(I) 0.0721 0.0401
Ri„t 0.0736 0.0265
R-sigma 0.0747 0.0324
S 0.1916 0.0856
Ri [I>2a (I)] 0.0926 0.0335
wR2 [I>2a (I)] 0.2156 0.0867
R, [all data] 1.081 1.058
wR2 [all data] 0.46; -0.30 0.20; -0.18
Pmœc, Pmde/A3)/ & A"3
CCDC Number
Ri = SFo - |FC||/S|F0|; mR2 = {SHF02 -Fc2)2]/SHF02)2]}1/2;
w =l/[c72(F02)+(aP)2 + bP], where где P = (F02 + 2Fc2)/3; 5 = {E[w(F02 - Fc2)]/(n -p)}1 where n is the number of reflections and p is the number of refinement parameters.
14
COOH
11 10
12
12
Fig. 2. Confonners (4) according to X-ray diffraction
Fig. 3. Confonners (5)
Fig. 4. Selected NOE and HMBC correlations of the compound (4)
HOOC
HOOC
Fig. 5. Selected NOE and HMBC correlations of the compound (5) Table 3. Data on the 'H NMR and 13C NMR spectra of the compounds (4) and (5)* (5H, 5C, ppm)
C atom (4) (5)
8C,ppm 8H, ppm 8C,ppm 8H, ppm
1 52.Id 1.81 ddd 51.6 d 1.63 ddd
2 27.4 t a 1.48m P 1.66m 30.0 t a 1.66m P 1.85m
3 23.9 t a 2.45m P 2.30 m 34.8 t a 2.68m P 2.63 m
4 132.2 s - 130.2 s -
5 144.9 d 7.00 dd 147.0 d 6.23 br. t
6 28.71 a 2.49 m P 2.41 m 31.2 a 2.23 m P 3.22 m
7 8 34.0 t 154.6s a 2.41m P 2.29 m 33.5 t 151.8s 2.19m
9 40.2 d 2.50 ddd 48.9 d 2.46 ddd
10 40.3 t a 1.74 dd P 1.56 dd 40.3 t 1.59 dd 1.78 dd
11 33.4 s - 33.9s -
12 30.1 q 0.99 s 30.2 q 1.00 s
13 23.0 q 0.96 s 22.5 q 0.98 s
14 173.7s - 174.3 s -
15 111.6 t a 4.87 d P 4.80 d 114.3 t a 5.01 d P 4.86 d
* 'H and 13C NMR spectra were obtained at 50 ° C.
J, Hz: for (4): l-2a=9.4; 1-2(3=4.0; 2a-3(3=11.5; 2(3-3(3=4.2; 2(3-3a=12.6; 2a-2(3=13.8; 3a-2a=5.3; 3a-3(3=13.7; 15-15 =1.5; 5-6(3=9.3; 5-6a=6.1; 6-7(3=4.0; 6-7a=8.9; 7a-7(3=13.0; 1-9=9.7; 9-10a =2.0; 9-10(3= 9.1; 10a-10(3=10.4.
J, Hz: for (5): 1-9=8.9; l-2a=11.0; 1(3-2(3=1.4; 2(3-3a=2.5; 3a-2a=7.0; 2(3-2a=9; 2a-3(3 =13.0; 2(3-3(3 =2.0; 3a-3(3 =13.5; 5-6(3=7.7; 5-6a=7.6; 7a-6a=4.0; 7(3-6(3=7.0; 6(3-7a=10.0; 7(3-7a=11.0; 6(3-6a=13.0; 9-10a =8.5; 9-10(3=9.6; 10a-10(3=10.8.
H3COO1
Analysis of caryophyllenic acids fractions after methylation with diaz-omethane often gave not two peaks, but 4 by the GLC-MS method. At the same time, the corresponding peak to the methyl ester of caryophyllenic acid (5) decreased. The compound to which the peak corresponded on a chromato-gram with a retention index of 1731 was separated from the methyl ester of caryophyllenic acid by column chromatography on silica gel, using PE as the eluent with up to 3% diethyl ether.
(6)
The spectral data of the by-product of methylation coincided with the spectral data of pyrazoline methyl ester of caryophyllenic acid (5) [8]. On the
chromatogram obtained by the GLC-MS method, as well as the peak of the compound with the retention index of 1747. the corresponding mass spectrum, which completely corresponds to the mass spectrum (6), but the intensity of this peak is 10 times smaller. Presumably, this is the peak of the methyl ester of isocaryophyllenic acid.
It is assumed that the presence of compound (5) in the extracts of plants of the genus Lychnophorinae leads to the development of antitumor, antinociceptive and anti-inflammatory activities. The alcohol extract of this plant, containing (5) inhibits the activity of xanthine oxidase [9].
6-Hydroxyisocaryophyllene was isolated from Betula pendula birch buds. Structure was determined by NMR spectroscopy. Caryophyllenic acid and isocaryphyllenic acid were discovered in the Betula grandifolia, B. albo-sinensis, B. fusca, B. obscura, B. litwinowii, B. hallii, B. grandifolia vegetative buds. The structure of the acids were determined by X-ray diffraction analysis. Thus, in the birch buds, together with the (E) isomers, there are also (Z) isomers.
Acknowledgements
XRD study have been prepared at the X-ray Diffraction Centre of St.Petersburg State University.
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For citing: Vedernikov D.N., Teplyakova S.V., Khoroshilova O.V. Khimiya Rastitel'nogo Syr'ya, 2018, no. 3,pp. 115-122. DOI: 10.14258/jcprm.2018033717.
Conclusions
References
Received January 30. 2018
Revised March 23. 2018
