Chemistry of Natural Compounds, Vol. 49, No. 5, November, 2013 [Russian original No. 5, September–October, 2013]
STRUCTURE AND BIOLOGICAL ACTIVITY OF SEVERAL CLASSES OF COMPOUNDS FROM THE BROWN ALGA Sargassum pallidum
N. I. Gerasimenko,* S. V. Logvinov, N. G. Busarova, and E. A. Martyyas
UDC 582.272.7:577.115.083:577.115.3
Sargassum pallidum (Phaeophyceae) is broadly distributed in the Yellow Sea and the Sea of Japan, on the southeastern shores of Sakhalin Island, and on the southern islands of the Kuril archipelago. Algae of the genus Sargassum, including S. pallidum, produce chemical compounds of various structures such as polysaccharides (in particular, sulfated fucan), plastoquinones, chromanols, chromenes, sterols, and lipids [1–4] and are used in Eastern medicine because they exhibit antitumor, antioxidant, antiviral, and antimicrobial activities [4–8]. Lipids, e.g., monogalactosyldiacylglycerols from S. thunbergii exhibited antifungal activity against Candida albicans [9]; from S. fulvellum, fibrinolytic activity [10]. A sulfoglycerolipid from S. wightii showed antibacterial activity [11]. During a study of the lipid composition of S. pallidum at various stages of the life cycle and at various times of the year, we detected substances that were not previously found in this alga, the contents of which were rather noticeable and had a distinct seasonal nature, in addition to the lipids typical for algae of the genus Sargassum [12]. These substances were isolated in order to study their structural properties. Lipid extracts from freshly collected specimens were obtained and separated into pure components according to the method described earlier by us [13]. The substances were purified over additional columns. The elutions were monitored by TLC on silica gel plates (Silica gel 60F254, Merck, Germany) using the previously described solvent systems [13, 14]. The neutral lipids (NL) contained according to TLC and lipid standards the usual lipids triacylglycerols (TAG) and free sterols in addition to significant quantities of free fatty acids (FFAs) and two compounds were detected. These were compound 1 that was located significantly below the FFAs and compound 2 that had an Rf value slightly greater than that of the TAG. Work up of the chromatogram with H2SO4 (10%) and ashing of it at 180°C gave a lilac color. We detected compound 3 in the glyceroglycolipids. It gave a positive reaction with anthrone reagent and was located on the chromatogram between monogalactosyl- and digalactosyldiacylglycerols. Compound 4 gave a positive reaction with Dragendorff’s reagent and had an Rf value on the chromatograms using the solvent systems for a lipid betaine [14] that was analogous to the Rf value of a standard isolated by us by preparative TLC from the alga S. miyabe, in which this compound and diacylglycerohydroxymethyltrimethylalanine (DGTA) were identified [15]. In our opinion, compounds 1 and 3 and FFAs are normal metabolic products of the alga and not artifacts formed during the extraction process. The structures of the compounds were identified by PMR, 13C NMR, and 2D spectra and comparison of them with the literature. The compounds were identified as 1,2-diacyl-3-hydroxy-sn-glycerol (1, 1,2-DAG), di-(2-ethylhexyl)-phthalate (2, DEHP), 1-O-acyl-3-O-(E -D-galactopyranosyl)-sn-glycerol (3, monogalactosylmonoacylglycerol, MGMG), and 1,2-diacylglycero-3-O-2c-(hydroxymethyl)-(N,N,N-trimethyl)-E-alanine (4, DGTA). The fatty-acid (FA) compositions of the lipids were studied using GC and GC/MS [13]. 1,2-Diacyl-3-hydroxy-sn-glycerol (1). 1H NMR spectrum (500 MHz, CD3OD, G, ppm, J/Hz): 4.38 (1Í, dd, J = 12.0, 3.3, Í-1), 4.15 (1Í, dd, J = 12.0, 6.6, Í-1), 5.23 (1Í, m, Í-2), 3.65 (2Í, d, J = 5.4, Í-3). According to our results, the resonance of the C-2 H atom was situated at weaker field. This disagreed with data reported earlier for DAG from olives and vegetable oils [16, 17].
G. B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-East Branch, Russian Academy of Sciences, 690022, Vladivostok, Prosp. 100 Let Vladivostoku, 159, Russia, e-mail:
[email protected]. Translated from Khimiya Prirodnykh Soedinenii, No. 5, September–October, 2013, pp. 795–796. Original article submitted July 24, 2013. 0009-3130/13/4905-0927
©2013
Springer Science+Business Media New York
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The principal FAs in 1,2-DAG from S. pallidum were 14:0 (18.9%) and 16:0 (66.4% of total FA). The fraction of FA 16:1n-7, 18:0, 18:1n-9, 18:3n-3, 18:2n-6, and 20:1n-9 was 14.7% (of total FA). Di-(2-ethylhexyl)-phthalate (2, CAS 117-81-7). PMR and 13C NMR spectral data agreed with those described for DEHP from leaves of Nauclea officinalis [18]. 1-O-Acyl-3-O-(E-D-galactopyranosyl)-sn-glycerol (3). 1H MNR spectrum (500 MHz, CD3OD, G, ppm, J/Hz): 4.16 (1Í, dd, J = 4.6, 11.3, Í-1), 4.13 (1Í, dd, J = 6.1, 11.3, Í-1), 3.99 (1Í, m, Í-2), 3.90 (1Í, dd, J = 5.2, 10.5, Í-3), 3.65 (1Í, dd, J = 4.6, 10.5, Í-3), 4.22 (1Í, d, J = 7.6, Í-1c), 3.53 (1Í, dd, J = 7.6, 9.7, H-2c), 3.46 (1Í, dd, J = 9.6, 3.3, H-3c), 3.82 (1Í, dd, J = 3.3, 1.1, H-4c), 3.50 (1Í, m, J = 1.2, 5.4, 6.8, H-5c), 3.71 (1Í, dd, J = 5.4, 11.4, H-6c), 3.75 (1Í, dd, J = 6.8, 11.4, H-6c). 13Ñ NMR spectrum (125 MHz, CD3OD, G, ppm): 67.18 (Ñ-1), 70.26 (Ñ-2), 72.50 (Ñ-3), 105.94 (Ñ-1c), 73.19 (Ñ-2c), 75.48 (Ñ-3c), 70.90 (Ñ-4c), 77.40 (Ñ-5c), 63.11 (Ñ-6c). The structure was determined using COSY, HSQC, and HMBC spectra. The MGMG contained FAs 14:0 (10.4%), 16:0 (57.2%), 18:0 (16.0%), 18:1n-9 and 18:3n-3 (16.4% of total FA). 1,2-Diacylglycero-3-O-2c-(hydroxymethyl)-(N,N,N-trimethyl)-E-alanine. 1H MNR spectrum (700 MHz, CD3OD, G, ppm, J/Hz): 4.38 (1Í, dd, J = 12.0, 3.0, Í-1), 4.15 (1Í, dd, J = 12.0, 7.0, 3.8, Í-1), 5.22 (1Í, m, Í-2), 3.62 (1Í, dd, J = 5.9, 2.0, Í-3), 3.59 (1Í, dd, J = 5.4, 1.1, Í-3), 3.79 (1Í, dd, J = 5.6, 9.4, Í-1c), 2.99 (1Í, m, Í-2c), 3.58 (1Í, m, J = 5.6, 1.5, Í-3c), 3.99 (1Í, dd, J = 13.2, 7.9, Í-4c), 3.45 (1Í, dd, J = 2.0, 13.1, 2.0, Í-4c), 3.16 (9Í, s, Í-5c–7c (Ìå)3). 13Ñ NMR spectrum (175 MHz, CD3OD, G, ppm): 64.50 (Ñ-1), 72.13 (Ñ-2), 71.05 (Ñ-3), 73.11 (C-1c), 45.78 (C-2c), 179.04 (C-3c), 68.47 (C-4c), 54.72 (Ñ-5c–7c, (Ìå)3). 13C NMR spectra were similar to those described previously for cryptophyte alga Cryptomonas CR-1 [14]. Resonances were assigned and the mutual locations of groups were determined using COSY, HSQC, and HMBC spectra. DGTA from S. pallidum comprised FAs decreasing in content in the order 16:0 (32.5%), 20:4 n-6 (21.9%), 18:1 n-9/18:3 n-3 (8.4%), 14:0 (8.1%), 18:2 n-6 (6.5%), 16:1 n-7 (3.3%), 20:3 n-3 (2.7%), 20:5 n-3 (2.4%), 20:0 (3.8%). Among the other FAs, 18:0, 15:0, 16:1n-5, 20:1n-9, 20:2n-9, and trace quantities of 17:0, 18:3n-6, 18:4n-3, 18:1n-7, 22:1n-9, and 22:0 were detected. Antimicrobial activity of the compounds was determined using the method described by us earlier [13]. DAG were slightly active (1.5 mm from the well edge) against Fusarium oxysporum. MGMG were active against Gram-positive bacterium Staphylococcus aureus and fungus F. oxysporum (each 4.0 mm from the well edge). DGTA inhibited the growth of yeast-like fungus Candida albicans (2.5 mm from the well edge), Aspergillus niger, and F. oxysporum (5.0 and 4.0 mm from the well edge, respectively). DEHP did not exhibit antimicrobial activity whereas this same phthalate from flowers of Calotropis gigantea showed a broad spectrum of antimicrobial activity [19], like the structurally similar phthalate (di-n-octylphthalate) that was observed earlier in S. wightii [20]. According to the researchers, its presence in the alga was not the result of environmental contamination. We also did not observe DEHP in seawater taken at the alga collection site and in the organic solvents used in the work.
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