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Glycolipids and fatty acids of some seaweeds and marine grasses from the black sea
Phytochemistry,Vol. 30, No. 7, pp. 2279 2283, Printed III Great Britain.
0031-9422/91 $3.00+0.00 Q 1991PergamonPressplc
1991
GLYCOLIPIDS AND FATTY ACIDS OF SOME SEAWEEDS MARINE GRASSES FROM THE BLACK SEA VALERY
M. DEMBITSKY,
ELENA
E. PECHENKINA-SHUBINA
AND
and OLGA A. ROZENTSVET
Laboratory of Chemistry of Natural Compounds, Institute of Ecology of the Volga River Basin, USSR Academy of Sciences, Togliatti 445003, U.S.S.R. (Receioed in
Key Word Index-Marine
reoised form 17 October 1990)
red algae; marine green algae; marine grasses; glycolipids; fatty acids.
Abstract-Fourteen species of marine red algae belonging to the Rhodophyta, 13 species of marine green algae belonging to the Chlorophyta and two species of the marine grass from the Embryophyta were examined for their glycolipid and fatty acid compositions. Characteristic of red algae was their high content of 20:4 and 20: 5. The green algal species were unusual in containing 16 :4 varying from 4.9 to 23.1% of the total fatty acids; 16 :0, 18 : 1 and 18 : 3 acids were also found in high amounts. Glycolipid contents of total lipid extracts of red algae ranged from 16.4 to 31.8 pmol g-’ dry wt, while those in green algae varied from 10.5 to 31.8. The red algae contained almost equal quantities of monoglycosyldiacylglycerol (MGDG), diglycosyldiacylglycerol (DGDG) and sulphaquinovosyldiacylglycerol with some exceptions. MGDG and DGDG were the major glycolipids in green algae and marine grasses, respectively.
siphonia subulzjkra to 46.5%. As minor components,
INTRODUCTION
Fatty acids found in marine algae have aroused considerable interest among researchers [l-5]. Although general trends of fatty acid distribution in various algal types have been elucidated C&12], further experimental work has lost none of its importance; differences in fatty acid composition can be notable even among closely related species [l, 3, 4, 7, S] as well as within the same species depending on their natural milieu [ 131. Studies of algal fatty acids can also be useful because of the remarkable role of algae as nutrients for many marine organisms and to the high concentration of polyunsaturated acids (20:4 and 20: 5) which, when included the diet, can reduce the probability of heart and vascular deseases [14-161. Recently, Gerwick et al. [17-201 reported on the isolation of several new bioactive fatty acids from the red seaweeds Ptilota jiiicina, Murrayella periclados, Platysiphonia miniata and Farlowia mollis. These fatty acids (icosanoids)
are natural products with potential biomedical applications. Our earlier work addressed the phospholipids contained in some red [21] and green algae [22] from the Black Sea, as well as glycolipids, phospholipids and fatty acids of brown algae from the same habitat [23]. The present paper reports further results from our examination of marine algal lipids.
RESULTS AND DISCUSSION
Fatty acids of red algae are of great interest, certain red algal species contain as much as 61.2% of 20:4 and 20: 5 acids (Table 1). High concentrations of 16:0 were also found in several species, for example, in Ceramium strictum, this acid amounted to 52.1% and in Poly-
15 : 0, 16 : 2,16 :4 and 18 :4 were found in many red algal species. It would appear that other red algae may contain somewhat higher levels of 20:5 than those found in the red algae from the Black Sea. Thus, the amount of this acid in Palmaria palmata is 70% [24] whilst in P. stenogona [9] it is reportedly 72.7%. Another red algae, Halosaccion ramentaceum, contains 63.7% of this acid [7]. Stefanov et al. [25], investigating the composition of two red algal species from the Black Sea, Corallina granifera and Phyllophora nervosa, found 30.3% of 20: 5 in the former (our result is 56.1%) and 5.6% in the latter (our result is 5.1%). Arachidonic acid in Phyllophora nervosa amounted to 44.3% according to the above investigators, while in our study it was 32.5%. From the reported data, it can be easily seen that one and the same species inhabiting the coastal waters of Bulgaria and those of the U.S.S.R., feature certain differences which may depend on the conditions existing in their respective environments. According to our data, the total amounts of 20:4 and 20: 5 change from 16.1%, in Apoglossum ruscifolium, to 61.2% in Corallina granifera. Examination of the green algae for their fatty acid composition revealed the presence of 16:0 varying in the examined species from 16.9 to 34.5% (Table 2). Characteristic of green algae was the presence of 16:4 (n-3) in amounts varying from 4.9 to 23.4%; 16:3 has not been estimated by us, but according to certain authors [2, 51, its concentration can be as high as lO--12% in Codium fragile. Unlike red and brown algae, green algae contain large amounts of 16:3 and 16:4 polyunsaturated acids [2, 6-8, 24, 26, 27-J. Johns et al. [8] have proposed that one of these acids, namely 16:4, can be taxonomically important among green macrophytic algae, while in plants of other divisions it is found in not more than trace
2279
Table 1. Fatty acid composition of red marine algae (wt % of total fatty acids) Species
14:o
15:o
16:O
16: 1*
16:2
1.4kO.2
-
39.4 f 0.4
10.3 kO.4
0.7kO.l
-
25.9kO.2
5.6+0.2
-
52.1 & 1.0
9.7kO.3
--
0.5+0.1
16:4
18:O
18: l*
18:2*
1s:3*
18:4
20:4*
20:s
2.1 +0.2
2.8kO.3
19.9kO.4
2.6kO.l
2.1 kO.3
3.3 +0.2
10.3 +0.5
5.8 +0.3
3.4kO.l
0.9kO.l
2.2kO.l
6.0 + 0.2
2.6kO.l
3.5kO.2
12.3 kO.3
43.4kO.8
5.6 k 0.4
0.9kO.2
1.6+0.1
0.8_+0.2
-
1.4_+0.1
5.9kO.2
26.1 kO.5
27.4kO.4
14.3 +0.6
0.9kO.3
0.6&0.1
1.4kO.2
11.8kO.3
3.2 f 0.2
2.7 +0.4
0.8 + 0.2
5.8 +O.l
21.4kO.6
0.6 & 0.2
18.4 * 0.7
2.3 +0.2
0.5kO.l
0.6kO.2
0.9+0.1
6.6kO.4
3.4 f 0.3
3.7 LO.2
0.8+0.1
5.lkO.3
56.1+ 1.1
6X&-0.4
1.3&0.1
37.51tro.9
6.3 F0.2
__
1.4*0.1
3.OkO.2
15.6+0.5
4.4kO.3
1.8kO.2
1.3kO.3
9.9kO.6
10.7_+0.4
2.1+0.1
-
24.3 & 0.7
4.4kO.3
._
3.2kO.l
10.1,0.2
2.3kO.l
2.OkO.l
0.6kO.l
21.5 +0.4
29.5 f 0.7
3.2kO.l
--
28.5 +0.8
12.1 kO.4
1.4kO.3
0.6kO.l
5.4kO.3
9.9kO.l
4.8kO.l
2.3kO.l
4.2kO.2
0.9+0.1
31.0+0.3
5.1 kO.2
0.5+0.1
0.7kO.l
2.210.2
9.3kO.3
1.7kO.l
0.9 _+0.2
0.9 & 0.0
0.5+0.1
22.7kO.3
6.3 $- 0.2
1.2kO.l
0.5 +o.o
8.7+0.2
10.6+0.4
3.0 f 0.2
1.1+0.1
3.9 + 0.2
1.1+0.1
41.2+ 1.0
7.1 kO.3
1.0+0.1
3.2kO.l
2.1kO.2
1.2*0.0
0.9+0.1
3.3kO.l 6.9kO.3
--
29.4 +0.6 46.5 +0.9
10.9*0.4 4.3 +0.3
0.5_+0.0
1.6kO.2 4.5LO.l
11.3_fo.3 3.3 +0.2
3.OkO.2 1.1+0.1
5.1+0.2
2.2kO.l
41.8kO.4
4.720.3
5.OkO.3
13.8kO.3
1.2kO.2
Apoglossum ruscifofium Callitkamnion corymbosum Ceramium strictum Chondoria desyphylla Corallina gran@ra Dasyopsis apkulata Gelidium latijolium Kyfinia virgatuia Laurencia coronopus Peyssonnelia dubyi Phy Nophora nerwsa Polysiphonia elongata P. subulijera Spermathamnion stricturn
Values are mean + s.d. (n = 3-6). *More than one isomer may be present. -Not detected
0.7kO.2
-
_ 0.7_+0.1
6.3 + 0.2
25.5 +0.5
11.6+0.4
31.2kO.4
20.1 + 0.4
24.4kO.5
32.5 L-O.8
5.1 +0.3
1.4kO.l 0.7+0.1
3.4 + 0.3 1.6kO.2
34.5 +0.6 31.1+0.7
0.8kO.l
8.1 +0.3
17.3kO.4
0.7+0.1
Table 2. Fatty acid composition of green marine algae (wt % of total fatty acids) SpeCiCS
14:o
Liryopsis hypmidm B. phmma Chaetomorphn cm.s.sa 2.350.2 ClmiopbO~~ .%?ricea 4.6 f 0.3 c. tvlgabda Entocladia lJ&i& 1.1kO.l Entemwwrpha lima ll.lkO.3 E. pdifera 0.9kO.l Rhizockmium implextm Ulothrix paw 0.6 + 0.0 ma rigida 1.0+0.1 ma sp. 0.7+0.0 Vrospora penrcillr~ormis
16:4
18:O
18:1*
4.9 + 0.2 lO.lkO.4
-
0.9 * 0.2
3.1+0.1
38.9 + 0.7 14.5*0.4
0.9*0.1
12.9k0.S 12.4 f 0.3
0.4 + 0.2 a.2+0.3
l.lkO.3 29kO.l
2.7kO.2 0.8kO.2
2.9kO.l -
2.1 fO.l
7.4*0_2
3.3 +0.2
18.3 f0.2
2.0*0.1
15.1 f0.3
8.4 + 0.2
3.9kO.l
29+0.1
2.1 f0.2
s.s+O.3 3.7kO.l
1.9kO.l
8.5 f 0.2 43.7kO.9
3.Sf0.2 8.8kO.3
26kO.l 1.2+0.0
2.2+0.1 0.3 * 0.0
7.1 f0.3 0.6+0.1
8.3 f 0.2 1.9kO.2
9.8 + 0.4
4.2 + 0.3
13.7kO.4
5.1 f0.3
12.1 f0.4
10.7kO.4
2.OkO.l
4.6 f 0.5
2.5 f0.2
1.0+0.1
23.4k 1.0 17.8 +0.4
2.OkO.l
7.8k0.S 7.9kO.2
4.7 f 0.3 1.3kO.l
7.6 f 0.3 26.7 k 0.8
S.l+O.Z 13.1kO.5
5.5+0.1 1.0 LO.1
3.6 k 0.2 1.2+0.1
6.7kO.3 1.3rtO.2
o.s*o.1
124k0.S
8.2 & 0.3
3.2kO.l
0.9rtO.l
1.0+0.2
IS:0
16:0
16: l*
16:2
-
28.6 f 0.7 20.0+0.4
6.7 f 0.3 26.2 f 0.9
0.9*0.1
24.0 + 0.6
7.3 * 0.3
1.4+0.1
30.8 If 0.5 34.5 * 0.4
23.3 f 0.3 5.3kO.2
-
19.6 + 0.4
11.4+(x5
0.6+0.1
24.8 + 0.6 24.3 f 0.7
1.1 kO.2
30.1+0.5
1.2 f 0.0
23.5 rf:0.8 16.9 + 0.4 29.2kO.3
8.4+0.3 3.8kO.2 6.lkO.4
18.4kO.6
19.6~0.5
0.7kO.l 0.5+0.1 -
2.2kO.l
-
-
7.9 f 0.3 1.8 f0.2
7.7 f 0.4 8.6 f 0.5 5.750.2
3.9kO.l 1.9kO.l 0.4kO.l
6.6kO.4
2.3kO.l
-
18:2’
18:3*
18:4
20:4*
22:s
22:s
22~6
14.8kO.4
12.0+0.3
8.3kO.4
8.5kO.2 147+03 44.5* 1.0
9.3kO.3 0.5*0.1 9.8kO.4
14.4kO.6 14.9rf-0.4 1.1 fO.0
9.3 * 0.4 10.1+0.6 1.6 f 0.2
3.8 kO.3 2.1 fO.l 0.9+0.1
5.0*0.1 18.3 +0.4
3.1 kO.2 3.4 f 0.3
1.3f0.2
18.1kO.5
2.4kO.l
5.6kO.3
7.4 + 0.4
5.6 +0.2
ll.SkO.3
1.7kO.l
0.8kO.l
Footnotes as Table 1
Table 3. Fatty acid composition of green seagrasses (wt % of total fatty acids) species
14:o
ls:o
16:0
16:l’
16:2
16:4
18:0
18:l.
18:2*
18:3’
18:4
20:4*
20:s
22:s
22~6
Zostera marina
0.7 * 0.2 a9ko.i
0.5+0.1
19.4kO.7 17.2ka3
2.2kO.2 1.3kO.3
0.5*0.1
0.6 + 0.1 1.1*0.1
2.lkO.2 1.s*o.1
11.4+0.4 10.3 f0.5
14.1 f0.6 12.8kO.3
37.4 +0.8 35.9kO.9
0.9 f 0.2 2.550.3
o.srto.1 0.6 f 0.0
8.4kO.3 13.7kO.S
0.7*0.1 0.5*0.1
1.1 &O.l 0.9 f 0.2
z. luma
Footnotes as Table 1.
2282
V. M.
DEMBITSKY et al.
amounts. Distribution of 16:4 in green algae deserves special attention and investigation. In the examined species, the total amounts of polyunsaturated 20:4 and 20: 5, ranged from 0.9 to 20.4%. Data available on four species of green algae from the Black Sea [25] has already shown that 16: 3 (n-3) in Ulva rigida and Bryopsis hypnoidea amounts to 11.1 and 14.7%, respectively. Fatty acid compositions of the two seagrasses Zostera marina and Z. nana, were similar (Table 3), although the proportions of 20: 5 were different, amounting to 8.4 and 13.7%, respectively. Seagrasses differed from the macrophytic algae in containing higher proportions of 18:2 (n-6) and 18 : 3 (n-3) (Table 3): many plant species have concentrations of 18: 3 that exceed those of 18:2 [9, 28-311 with only a few exceptions when their proportions are reversed [28]. Seagrasses also differ from macrophytes in having low amounts of acids with more than three double bonds, and containing saturated fatty acids greater than Cz,, [30, 313. Fatty acid composition of seagrasses makes them similar to higher terrestrial plants [32], but differing from the latter in the presence of acids with chain lengths of 20 or more, although in low amounts.
Glycolipids, being the major class of polar lipids of algae and plants, have attracted considerable attention from investigators [22,33,34-J. Examination of glycolipid composition of red algae in the present study showed variation of MGDG content from 19.6 to 46.1% of total glycolipids, of DGDG, from 22.7 to 51.3%, and of SQDG, from 23.7 to 39.9. These were identified by comparison with authentic compounds as well as specific spray reagents, such as r-naphthol [35] and periodateSchiff’s reagent [36]. The total glycolipids was estimated as 16.4-31.5 pmol g- ’ dry wt (Table 4). The distribution of fatty acids in various classes of glycolipids reported by Araki et al. [33] for the red alga Porphyra yezoensis is of interest. Thus, 20: 5 contained in MGDG amounted to 73.8%, and in an MGDGl fraction, to 93%. According to Pettitt et al. [34], 20:5 may exhibit strong seasonal changes; MGDGZ contained 80.3% of this acid in winter, but only 60.5% in summer. There was no such strong seasonal difference found for MGDGl in Polysiphonia lanosa, the content of which was 46.5% in winter and 43.5% in summer. In green algae, MGDG was the major glycolipid varying from 40.7 to 51.3%. DGDG and SQDG occurred
Table 4. Glycolipid composition of red marine algae (% total glycoliplds) Species
MGDG
DGDG
SQDG
GL*
Apoglossum ruscijolium Callithamion corymbosum Ceramium strictum Chondiria dasyphylla Corallina granafera Dasyopsis apiculata Gelidium latijolium Kylinia uirgatula Laurencia coronopus Peyssonnelia dubyi Phyllophora nervosa Polysiphonia elongata P. subulijera Spermathamnion strictum
29.9 35.8 30.3 33.7 36.6 46.1 40.0 42.3 19.6 31.6 28.5 37.4 34.2 34.1
42.4 29.1 34.9 26.4 28.2 30.2 32.5 28.9 51.3 38.0 36.1 22.1 28.4 40.0
21.1 34.5 34.8 39.9 35.2 23.1 21.5 28.8 29.1 30.4 35.4 39.9 31.4 25.3
27.1 20.4 16.8 19.7 31.8 24.5 26.4 18.2 30.6 16.4 21.5 23.4 26.9 31.5
Values are means & s.d. (n = 3-6). *GL---the sum of MGDG, DGDG. SQDG pmol FA g-’ dry wt
Table 5. Glycolipid composition of green marine algae (W total glycolipids) Species
MGDG
DGDG
SQDG
GL*
Bryopsis hynoidea B. plumosa Chaetomorpha crassa Ctadophora sericea C. vagabunda Enteromorpha lmza E. prolifera Entocladia viridis Rhizoclonium implexum Ulotrix jlacca Ulva rigida Ulva sp.
41.8 43.6 41.3 46.9 44.6 48.4 41.6 51.3 40.7 46.1 49.9 54.1
27.3 29.9 24.9 16.1 18.4 21.2 19.4 18.8 22.8 20.8 15.1 10.5
30.9 26.5 33.8 37.0 37.0 30.4 33.0 29.9 36.5 32.5 35.0 35.4
25.1 23.5 20.9 16.8 18.1 19.3 21.5 31.8 20.4 21.5 26.4 19.0
Footnotes as Table 4.
Lipids from marine algae and grasses Table 6. Glycolipid
composition of seagrasses glycolipids)
(%
total
Species
MGDG
DGDG
SQDG
GL*
Zostera marina 2. nana
33.6 34.8
51.8 50.3
14.6 14.9
10.5 14.8
Footnotes as Table 4.
2283
8. Johns, R. B., Nichols, P. D. and Perry, G. J. (1979) Phytochemistry 18, 799. 9. Khotimchenko, S. V. and Svetashev, V. I. (1983) Biol. Norya (Vladivostok) 5, 45. 10. Pohl, P. and Zurheide, F. (1979) in Advances in Biochemistry ana’ Physiology of Plant Lipids Vol. 3, pp. 427-453.
Elsevier/North-Holland, Amsterdam 11. Pohl, P. and Zurheide, F. (1982) in Marine Algae in Pkarmaceutical Science Vol. 2, pp. 65-80. Walter de Gruyter, Berlin. 12. Kayama, M., Araki, S. and Sato, S. (1989) in Marine Biogenic Lipids? Fats and Oils Vol. 2, pp. 3348. 13. Khotimchenko, S. V. and Svetashev, V. 1. (1987) Biol. iworya (Vladivostok) 6, 3.
in the range 10.5-29.9% and 26.5-37.0%, respectively (Table 5). In the seagrasses, Zostera nana and Z. marina, glycolipids were found in different proportions, MGDG
ranging from 33.4 to 34.8%, DGDG from 50.3 to 51.8% and SQDG from 14.6 to 14.9% (Table 6). Glycolipids from the green algae studied showed variations from 10.5 to 31.8 pmol g-l dry wt, and in seagrasses, from 10.5 to 14.8 pmol g- ’ dry wt. Our earlier data on phospholipids and fatty acids found in some seagrasses [37] and the data presented by other authors [38] show substantial correlation. EXPERIMENTAL Algalmaterial. Algae were harvested in July 1987 in the northeast of the Black Sea (the Bays of Feodosia and Karadagh) at O-10 m depth Freshly collected algae were carefully cleaned of extraneous matter and only clean tissue was used for homogenization in a high-speed unit. Lipid extraction. Ground algal tissue was treated with CHCl,-MeOH (1: 1). Lipids were extracted in three stages and the extracts combined. CHCls-MeOH-H,O (8: 1:4) was then added. Following evapn in a N, stream, the resulting extract was dissolved in dry CHCI, and stored at -20”. Lipid iso~t~on and ident~cation. Glycolipids were identified according to procedures described in ref [23]. They were isolated by TLC on silica gel [39] using CHCl,-Me&O-MeOH-HCO,H-H,O (25: 10:5:5:2) and Me,CO-C,H,-HCO,H-H,O (200: 30:3:8). Glycolipids were visualized with non-destructive fuchsin reagent [40]. Me esters of fatty acids were prepd according to ref. [41] and determined by FIDGC as described previously [42].
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41. Carreau, J. P. and Dubacq, J. P. (1978) J. Chramatogr. 151, 384. 42. Dembitsky, V. M. and Nebylitayn, B. D. (1980) Bioorgan. Khim. 6,1542.
0031-9422/91 $3.00+0.00 Q 1991PergamonPressplc
1991
GLYCOLIPIDS AND FATTY ACIDS OF SOME SEAWEEDS MARINE GRASSES FROM THE BLACK SEA VALERY
M. DEMBITSKY,
ELENA
E. PECHENKINA-SHUBINA
AND
and OLGA A. ROZENTSVET
Laboratory of Chemistry of Natural Compounds, Institute of Ecology of the Volga River Basin, USSR Academy of Sciences, Togliatti 445003, U.S.S.R. (Receioed in
Key Word Index-Marine
reoised form 17 October 1990)
red algae; marine green algae; marine grasses; glycolipids; fatty acids.
Abstract-Fourteen species of marine red algae belonging to the Rhodophyta, 13 species of marine green algae belonging to the Chlorophyta and two species of the marine grass from the Embryophyta were examined for their glycolipid and fatty acid compositions. Characteristic of red algae was their high content of 20:4 and 20: 5. The green algal species were unusual in containing 16 :4 varying from 4.9 to 23.1% of the total fatty acids; 16 :0, 18 : 1 and 18 : 3 acids were also found in high amounts. Glycolipid contents of total lipid extracts of red algae ranged from 16.4 to 31.8 pmol g-’ dry wt, while those in green algae varied from 10.5 to 31.8. The red algae contained almost equal quantities of monoglycosyldiacylglycerol (MGDG), diglycosyldiacylglycerol (DGDG) and sulphaquinovosyldiacylglycerol with some exceptions. MGDG and DGDG were the major glycolipids in green algae and marine grasses, respectively.
siphonia subulzjkra to 46.5%. As minor components,
INTRODUCTION
Fatty acids found in marine algae have aroused considerable interest among researchers [l-5]. Although general trends of fatty acid distribution in various algal types have been elucidated C&12], further experimental work has lost none of its importance; differences in fatty acid composition can be notable even among closely related species [l, 3, 4, 7, S] as well as within the same species depending on their natural milieu [ 131. Studies of algal fatty acids can also be useful because of the remarkable role of algae as nutrients for many marine organisms and to the high concentration of polyunsaturated acids (20:4 and 20: 5) which, when included the diet, can reduce the probability of heart and vascular deseases [14-161. Recently, Gerwick et al. [17-201 reported on the isolation of several new bioactive fatty acids from the red seaweeds Ptilota jiiicina, Murrayella periclados, Platysiphonia miniata and Farlowia mollis. These fatty acids (icosanoids)
are natural products with potential biomedical applications. Our earlier work addressed the phospholipids contained in some red [21] and green algae [22] from the Black Sea, as well as glycolipids, phospholipids and fatty acids of brown algae from the same habitat [23]. The present paper reports further results from our examination of marine algal lipids.
RESULTS AND DISCUSSION
Fatty acids of red algae are of great interest, certain red algal species contain as much as 61.2% of 20:4 and 20: 5 acids (Table 1). High concentrations of 16:0 were also found in several species, for example, in Ceramium strictum, this acid amounted to 52.1% and in Poly-
15 : 0, 16 : 2,16 :4 and 18 :4 were found in many red algal species. It would appear that other red algae may contain somewhat higher levels of 20:5 than those found in the red algae from the Black Sea. Thus, the amount of this acid in Palmaria palmata is 70% [24] whilst in P. stenogona [9] it is reportedly 72.7%. Another red algae, Halosaccion ramentaceum, contains 63.7% of this acid [7]. Stefanov et al. [25], investigating the composition of two red algal species from the Black Sea, Corallina granifera and Phyllophora nervosa, found 30.3% of 20: 5 in the former (our result is 56.1%) and 5.6% in the latter (our result is 5.1%). Arachidonic acid in Phyllophora nervosa amounted to 44.3% according to the above investigators, while in our study it was 32.5%. From the reported data, it can be easily seen that one and the same species inhabiting the coastal waters of Bulgaria and those of the U.S.S.R., feature certain differences which may depend on the conditions existing in their respective environments. According to our data, the total amounts of 20:4 and 20: 5 change from 16.1%, in Apoglossum ruscifolium, to 61.2% in Corallina granifera. Examination of the green algae for their fatty acid composition revealed the presence of 16:0 varying in the examined species from 16.9 to 34.5% (Table 2). Characteristic of green algae was the presence of 16:4 (n-3) in amounts varying from 4.9 to 23.4%; 16:3 has not been estimated by us, but according to certain authors [2, 51, its concentration can be as high as lO--12% in Codium fragile. Unlike red and brown algae, green algae contain large amounts of 16:3 and 16:4 polyunsaturated acids [2, 6-8, 24, 26, 27-J. Johns et al. [8] have proposed that one of these acids, namely 16:4, can be taxonomically important among green macrophytic algae, while in plants of other divisions it is found in not more than trace
2279
Table 1. Fatty acid composition of red marine algae (wt % of total fatty acids) Species
14:o
15:o
16:O
16: 1*
16:2
1.4kO.2
-
39.4 f 0.4
10.3 kO.4
0.7kO.l
-
25.9kO.2
5.6+0.2
-
52.1 & 1.0
9.7kO.3
--
0.5+0.1
16:4
18:O
18: l*
18:2*
1s:3*
18:4
20:4*
20:s
2.1 +0.2
2.8kO.3
19.9kO.4
2.6kO.l
2.1 kO.3
3.3 +0.2
10.3 +0.5
5.8 +0.3
3.4kO.l
0.9kO.l
2.2kO.l
6.0 + 0.2
2.6kO.l
3.5kO.2
12.3 kO.3
43.4kO.8
5.6 k 0.4
0.9kO.2
1.6+0.1
0.8_+0.2
-
1.4_+0.1
5.9kO.2
26.1 kO.5
27.4kO.4
14.3 +0.6
0.9kO.3
0.6&0.1
1.4kO.2
11.8kO.3
3.2 f 0.2
2.7 +0.4
0.8 + 0.2
5.8 +O.l
21.4kO.6
0.6 & 0.2
18.4 * 0.7
2.3 +0.2
0.5kO.l
0.6kO.2
0.9+0.1
6.6kO.4
3.4 f 0.3
3.7 LO.2
0.8+0.1
5.lkO.3
56.1+ 1.1
6X&-0.4
1.3&0.1
37.51tro.9
6.3 F0.2
__
1.4*0.1
3.OkO.2
15.6+0.5
4.4kO.3
1.8kO.2
1.3kO.3
9.9kO.6
10.7_+0.4
2.1+0.1
-
24.3 & 0.7
4.4kO.3
._
3.2kO.l
10.1,0.2
2.3kO.l
2.OkO.l
0.6kO.l
21.5 +0.4
29.5 f 0.7
3.2kO.l
--
28.5 +0.8
12.1 kO.4
1.4kO.3
0.6kO.l
5.4kO.3
9.9kO.l
4.8kO.l
2.3kO.l
4.2kO.2
0.9+0.1
31.0+0.3
5.1 kO.2
0.5+0.1
0.7kO.l
2.210.2
9.3kO.3
1.7kO.l
0.9 _+0.2
0.9 & 0.0
0.5+0.1
22.7kO.3
6.3 $- 0.2
1.2kO.l
0.5 +o.o
8.7+0.2
10.6+0.4
3.0 f 0.2
1.1+0.1
3.9 + 0.2
1.1+0.1
41.2+ 1.0
7.1 kO.3
1.0+0.1
3.2kO.l
2.1kO.2
1.2*0.0
0.9+0.1
3.3kO.l 6.9kO.3
--
29.4 +0.6 46.5 +0.9
10.9*0.4 4.3 +0.3
0.5_+0.0
1.6kO.2 4.5LO.l
11.3_fo.3 3.3 +0.2
3.OkO.2 1.1+0.1
5.1+0.2
2.2kO.l
41.8kO.4
4.720.3
5.OkO.3
13.8kO.3
1.2kO.2
Apoglossum ruscifofium Callitkamnion corymbosum Ceramium strictum Chondoria desyphylla Corallina gran@ra Dasyopsis apkulata Gelidium latijolium Kyfinia virgatuia Laurencia coronopus Peyssonnelia dubyi Phy Nophora nerwsa Polysiphonia elongata P. subulijera Spermathamnion stricturn
Values are mean + s.d. (n = 3-6). *More than one isomer may be present. -Not detected
0.7kO.2
-
_ 0.7_+0.1
6.3 + 0.2
25.5 +0.5
11.6+0.4
31.2kO.4
20.1 + 0.4
24.4kO.5
32.5 L-O.8
5.1 +0.3
1.4kO.l 0.7+0.1
3.4 + 0.3 1.6kO.2
34.5 +0.6 31.1+0.7
0.8kO.l
8.1 +0.3
17.3kO.4
0.7+0.1
Table 2. Fatty acid composition of green marine algae (wt % of total fatty acids) SpeCiCS
14:o
Liryopsis hypmidm B. phmma Chaetomorphn cm.s.sa 2.350.2 ClmiopbO~~ .%?ricea 4.6 f 0.3 c. tvlgabda Entocladia lJ&i& 1.1kO.l Entemwwrpha lima ll.lkO.3 E. pdifera 0.9kO.l Rhizockmium implextm Ulothrix paw 0.6 + 0.0 ma rigida 1.0+0.1 ma sp. 0.7+0.0 Vrospora penrcillr~ormis
16:4
18:O
18:1*
4.9 + 0.2 lO.lkO.4
-
0.9 * 0.2
3.1+0.1
38.9 + 0.7 14.5*0.4
0.9*0.1
12.9k0.S 12.4 f 0.3
0.4 + 0.2 a.2+0.3
l.lkO.3 29kO.l
2.7kO.2 0.8kO.2
2.9kO.l -
2.1 fO.l
7.4*0_2
3.3 +0.2
18.3 f0.2
2.0*0.1
15.1 f0.3
8.4 + 0.2
3.9kO.l
29+0.1
2.1 f0.2
s.s+O.3 3.7kO.l
1.9kO.l
8.5 f 0.2 43.7kO.9
3.Sf0.2 8.8kO.3
26kO.l 1.2+0.0
2.2+0.1 0.3 * 0.0
7.1 f0.3 0.6+0.1
8.3 f 0.2 1.9kO.2
9.8 + 0.4
4.2 + 0.3
13.7kO.4
5.1 f0.3
12.1 f0.4
10.7kO.4
2.OkO.l
4.6 f 0.5
2.5 f0.2
1.0+0.1
23.4k 1.0 17.8 +0.4
2.OkO.l
7.8k0.S 7.9kO.2
4.7 f 0.3 1.3kO.l
7.6 f 0.3 26.7 k 0.8
S.l+O.Z 13.1kO.5
5.5+0.1 1.0 LO.1
3.6 k 0.2 1.2+0.1
6.7kO.3 1.3rtO.2
o.s*o.1
124k0.S
8.2 & 0.3
3.2kO.l
0.9rtO.l
1.0+0.2
IS:0
16:0
16: l*
16:2
-
28.6 f 0.7 20.0+0.4
6.7 f 0.3 26.2 f 0.9
0.9*0.1
24.0 + 0.6
7.3 * 0.3
1.4+0.1
30.8 If 0.5 34.5 * 0.4
23.3 f 0.3 5.3kO.2
-
19.6 + 0.4
11.4+(x5
0.6+0.1
24.8 + 0.6 24.3 f 0.7
1.1 kO.2
30.1+0.5
1.2 f 0.0
23.5 rf:0.8 16.9 + 0.4 29.2kO.3
8.4+0.3 3.8kO.2 6.lkO.4
18.4kO.6
19.6~0.5
0.7kO.l 0.5+0.1 -
2.2kO.l
-
-
7.9 f 0.3 1.8 f0.2
7.7 f 0.4 8.6 f 0.5 5.750.2
3.9kO.l 1.9kO.l 0.4kO.l
6.6kO.4
2.3kO.l
-
18:2’
18:3*
18:4
20:4*
22:s
22:s
22~6
14.8kO.4
12.0+0.3
8.3kO.4
8.5kO.2 147+03 44.5* 1.0
9.3kO.3 0.5*0.1 9.8kO.4
14.4kO.6 14.9rf-0.4 1.1 fO.0
9.3 * 0.4 10.1+0.6 1.6 f 0.2
3.8 kO.3 2.1 fO.l 0.9+0.1
5.0*0.1 18.3 +0.4
3.1 kO.2 3.4 f 0.3
1.3f0.2
18.1kO.5
2.4kO.l
5.6kO.3
7.4 + 0.4
5.6 +0.2
ll.SkO.3
1.7kO.l
0.8kO.l
Footnotes as Table 1
Table 3. Fatty acid composition of green seagrasses (wt % of total fatty acids) species
14:o
ls:o
16:0
16:l’
16:2
16:4
18:0
18:l.
18:2*
18:3’
18:4
20:4*
20:s
22:s
22~6
Zostera marina
0.7 * 0.2 a9ko.i
0.5+0.1
19.4kO.7 17.2ka3
2.2kO.2 1.3kO.3
0.5*0.1
0.6 + 0.1 1.1*0.1
2.lkO.2 1.s*o.1
11.4+0.4 10.3 f0.5
14.1 f0.6 12.8kO.3
37.4 +0.8 35.9kO.9
0.9 f 0.2 2.550.3
o.srto.1 0.6 f 0.0
8.4kO.3 13.7kO.S
0.7*0.1 0.5*0.1
1.1 &O.l 0.9 f 0.2
z. luma
Footnotes as Table 1.
2282
V. M.
DEMBITSKY et al.
amounts. Distribution of 16:4 in green algae deserves special attention and investigation. In the examined species, the total amounts of polyunsaturated 20:4 and 20: 5, ranged from 0.9 to 20.4%. Data available on four species of green algae from the Black Sea [25] has already shown that 16: 3 (n-3) in Ulva rigida and Bryopsis hypnoidea amounts to 11.1 and 14.7%, respectively. Fatty acid compositions of the two seagrasses Zostera marina and Z. nana, were similar (Table 3), although the proportions of 20: 5 were different, amounting to 8.4 and 13.7%, respectively. Seagrasses differed from the macrophytic algae in containing higher proportions of 18:2 (n-6) and 18 : 3 (n-3) (Table 3): many plant species have concentrations of 18: 3 that exceed those of 18:2 [9, 28-311 with only a few exceptions when their proportions are reversed [28]. Seagrasses also differ from macrophytes in having low amounts of acids with more than three double bonds, and containing saturated fatty acids greater than Cz,, [30, 313. Fatty acid composition of seagrasses makes them similar to higher terrestrial plants [32], but differing from the latter in the presence of acids with chain lengths of 20 or more, although in low amounts.
Glycolipids, being the major class of polar lipids of algae and plants, have attracted considerable attention from investigators [22,33,34-J. Examination of glycolipid composition of red algae in the present study showed variation of MGDG content from 19.6 to 46.1% of total glycolipids, of DGDG, from 22.7 to 51.3%, and of SQDG, from 23.7 to 39.9. These were identified by comparison with authentic compounds as well as specific spray reagents, such as r-naphthol [35] and periodateSchiff’s reagent [36]. The total glycolipids was estimated as 16.4-31.5 pmol g- ’ dry wt (Table 4). The distribution of fatty acids in various classes of glycolipids reported by Araki et al. [33] for the red alga Porphyra yezoensis is of interest. Thus, 20: 5 contained in MGDG amounted to 73.8%, and in an MGDGl fraction, to 93%. According to Pettitt et al. [34], 20:5 may exhibit strong seasonal changes; MGDGZ contained 80.3% of this acid in winter, but only 60.5% in summer. There was no such strong seasonal difference found for MGDGl in Polysiphonia lanosa, the content of which was 46.5% in winter and 43.5% in summer. In green algae, MGDG was the major glycolipid varying from 40.7 to 51.3%. DGDG and SQDG occurred
Table 4. Glycolipid composition of red marine algae (% total glycoliplds) Species
MGDG
DGDG
SQDG
GL*
Apoglossum ruscijolium Callithamion corymbosum Ceramium strictum Chondiria dasyphylla Corallina granafera Dasyopsis apiculata Gelidium latijolium Kylinia uirgatula Laurencia coronopus Peyssonnelia dubyi Phyllophora nervosa Polysiphonia elongata P. subulijera Spermathamnion strictum
29.9 35.8 30.3 33.7 36.6 46.1 40.0 42.3 19.6 31.6 28.5 37.4 34.2 34.1
42.4 29.1 34.9 26.4 28.2 30.2 32.5 28.9 51.3 38.0 36.1 22.1 28.4 40.0
21.1 34.5 34.8 39.9 35.2 23.1 21.5 28.8 29.1 30.4 35.4 39.9 31.4 25.3
27.1 20.4 16.8 19.7 31.8 24.5 26.4 18.2 30.6 16.4 21.5 23.4 26.9 31.5
Values are means & s.d. (n = 3-6). *GL---the sum of MGDG, DGDG. SQDG pmol FA g-’ dry wt
Table 5. Glycolipid composition of green marine algae (W total glycolipids) Species
MGDG
DGDG
SQDG
GL*
Bryopsis hynoidea B. plumosa Chaetomorpha crassa Ctadophora sericea C. vagabunda Enteromorpha lmza E. prolifera Entocladia viridis Rhizoclonium implexum Ulotrix jlacca Ulva rigida Ulva sp.
41.8 43.6 41.3 46.9 44.6 48.4 41.6 51.3 40.7 46.1 49.9 54.1
27.3 29.9 24.9 16.1 18.4 21.2 19.4 18.8 22.8 20.8 15.1 10.5
30.9 26.5 33.8 37.0 37.0 30.4 33.0 29.9 36.5 32.5 35.0 35.4
25.1 23.5 20.9 16.8 18.1 19.3 21.5 31.8 20.4 21.5 26.4 19.0
Footnotes as Table 4.
Lipids from marine algae and grasses Table 6. Glycolipid
composition of seagrasses glycolipids)
(%
total
Species
MGDG
DGDG
SQDG
GL*
Zostera marina 2. nana
33.6 34.8
51.8 50.3
14.6 14.9
10.5 14.8
Footnotes as Table 4.
2283
8. Johns, R. B., Nichols, P. D. and Perry, G. J. (1979) Phytochemistry 18, 799. 9. Khotimchenko, S. V. and Svetashev, V. I. (1983) Biol. Norya (Vladivostok) 5, 45. 10. Pohl, P. and Zurheide, F. (1979) in Advances in Biochemistry ana’ Physiology of Plant Lipids Vol. 3, pp. 427-453.
Elsevier/North-Holland, Amsterdam 11. Pohl, P. and Zurheide, F. (1982) in Marine Algae in Pkarmaceutical Science Vol. 2, pp. 65-80. Walter de Gruyter, Berlin. 12. Kayama, M., Araki, S. and Sato, S. (1989) in Marine Biogenic Lipids? Fats and Oils Vol. 2, pp. 3348. 13. Khotimchenko, S. V. and Svetashev, V. 1. (1987) Biol. iworya (Vladivostok) 6, 3.
in the range 10.5-29.9% and 26.5-37.0%, respectively (Table 5). In the seagrasses, Zostera nana and Z. marina, glycolipids were found in different proportions, MGDG
ranging from 33.4 to 34.8%, DGDG from 50.3 to 51.8% and SQDG from 14.6 to 14.9% (Table 6). Glycolipids from the green algae studied showed variations from 10.5 to 31.8 pmol g-l dry wt, and in seagrasses, from 10.5 to 14.8 pmol g- ’ dry wt. Our earlier data on phospholipids and fatty acids found in some seagrasses [37] and the data presented by other authors [38] show substantial correlation. EXPERIMENTAL Algalmaterial. Algae were harvested in July 1987 in the northeast of the Black Sea (the Bays of Feodosia and Karadagh) at O-10 m depth Freshly collected algae were carefully cleaned of extraneous matter and only clean tissue was used for homogenization in a high-speed unit. Lipid extraction. Ground algal tissue was treated with CHCl,-MeOH (1: 1). Lipids were extracted in three stages and the extracts combined. CHCls-MeOH-H,O (8: 1:4) was then added. Following evapn in a N, stream, the resulting extract was dissolved in dry CHCI, and stored at -20”. Lipid iso~t~on and ident~cation. Glycolipids were identified according to procedures described in ref [23]. They were isolated by TLC on silica gel [39] using CHCl,-Me&O-MeOH-HCO,H-H,O (25: 10:5:5:2) and Me,CO-C,H,-HCO,H-H,O (200: 30:3:8). Glycolipids were visualized with non-destructive fuchsin reagent [40]. Me esters of fatty acids were prepd according to ref. [41] and determined by FIDGC as described previously [42].
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