Post on 29-Dec-2015
ANALYSIS OF GAMMA-LINOLENIC AND OTHER FATTY
ACIDS IN THE EDIBLE PART OF BORAGE (Borago officinalis L.).
M. del Ríoa, A. de Harob
a C.I.D.A. Apdo. 4240. 14.080 CORDOBA
b Inst. Agricultura Sostenible C.S.I.C. Apdo. 4084. 14080 CORDOBA.
ABSTRACT
Borage (Borago officinalis L.) is currently used as a vegetable in the North of Spain.
The edible parts of the plant are the basal leaf petioles, leaves and stem. The objective
of this study has been to determine the oil, gamma-linolenic acid and other fatty acids
content in the tissues of borage. The leaves were the most important source of lipid
with 9 % dry weight, while basal leaf petioles and stem were 1.8 % and 0.92 %
respectively. Alfa-linolenic acid and GLA are formed by desaturation of linoleic acid
by delta-6 and delta-15-desaturases, respectively, but the activity of the delta-15-
desaturase predominates. Alfa-linolenic was the major constituent with about 37% in
tissues photosyntetics and 24% in stem and basal leaf petioles. Basal leaves and main
stem leaves showed the highest content in GLA 15.8 % and 15.5% respectively. In
contrast to the seeds, all the tissues analyzed had stearidonic acid (18:4 n-3). The major
fatty acids were: palmitic, alfa-linolenic and stearidonic acids in leaves, and palmitic,
linoleic and alfa-linolenic acids in stem and basal leaf petioles. Inverse relation was
found between oleic acid and GLA and between linoleic vs. alfa-linolenic and
stearidonic acids in basal leaves. In main stem leaves and basal leaf petioles was also
found inverse relation between alfa-linolenic vs. oleic and linoleic acids.
.
INTRODUCTION
Gamma-linolenic acid (GLA, C:18, n-6,9,12, cis) is one essential fatty acid
precursor of indispensable compounds in the body such as prostaglandins (PGE1) and
leukotrienes (LT), often known as eicosanoids. Many factors have been shown to
interfere the bioconvertion of linoleic to GLA like ageing, stress, diabetes, high alcohol
intake, high cholesterol levels, viral infections and nutritional deficiencies. Since there
is some evidence that 6-desaturase activity is impaired in each of these situations, with
an exogenous supply of GLA these may be ameliorated [1]. Seeds of several plants
such as evening primrose (Oenothera biennis) and the common borage (Borago
officinalis) and the fruits of Ribes spp., have been found to contain substantial
proportions of GLA as a constituent of their lipids [2, 3, 4 ].
White flowered borage is currently used as a vegetable in the North of Spain, but no
attention has been paid till now to study as source of GLA. The parts of the plant that
are preferentially eaten are the basal leaf petioles (before the plant is completely
developed and before flowering) leaves and stem.
Lipids containing GLA are unusual constituents of tissues in higher plants, which are
generally rich in alfa-linolenic acid (all-cis-9,12,15-octadecatrienoic acid). Apparently a
plant that synthesizes substantial amounts of GLA acid in the seeds, should form this
acid in the leaves. However the leaves of the evening primrose plants contain alfa-
linolenic as major constituent of photosynthetic tissues but not GLA [5].
The objective of the present study was to determine the oil and fatty acids content in
the edible portion of the borage with emphasis on GLA acid.
MATERIAL
Seeds of 12 accessions of white flowered borage (B. officinalis L.) were provided by
The Center of Agricultural Research of Rioja . Borage plants were grown in a 16 h.
photoperiod at 15ºC and an 8 h. dark at 10ºC. Collections of the basal leaf petioles and
basal leaves were made before the plants were completely developed and before
flowering. The stem and main stem leaves were collected after flowering.
METHOD
Lipid Extraction and Analysis. Oil content was determined according to the
Association of Official Analytical Chemists. (AOAC, 1984, Method 7062) [6].
The fatty acid composition of oil was determined by GLC (Gas Liquid
Chromatography) of the corresponding methyl esters, using as quantifier
methylheptadecanoic acid as an internal standard . Digestion, transmethylation of lipids
from leaves, petioles and stem and extraction of fatty acid methyl esters in one step was
realized according to Garcés and Mancha [7]. The samples were heated at 80ºC with a
reagent containing methanol: heptane: toluene: 2-2-dimetoxypropane: H2SO4
(37:36:20:5:2). After 120 min. two phases were formed, the upper one containing the
fatty acid methyl esters ready for GLC analysis. The analysis were performed using a
Perkin Elmer Autosystem Gas Chromatograph equipped with flame ionization detector
and split injector. The chromatograph is equipped with a capillary column (25m x 0.25
mm, i.d. 0.25 mm film) with acidified polietilenglycol as the stationary phase. Oven
temperature was programmed from 190ºC to 210ºC at a rate of 2ºC/min. The
temperature of the detector and injector were 275ºC and 200ºC respectively. Nitrogen
was used as carrier gas.
RESULTS
The data given in Table I show the oil and fatty acids content in the portion edible of
borage in comparison with the content of seeds [8]. Leaves showed the highest content
of oil with a mean value of 9% dry weight. Basal leaf petioles and stem had a mean
value of 1.8% and 0.92% respectively. The major fatty acid in all the tissues. was alfa-
linolenic acid . Fatty acids composition was very similar in basal leaves and main stem
leaves of borage plant. The leaf lipids contained as the major constituents, about 15%
palmitic acid, 37% alfa-linolenic and 17% steridonic acids. Nevertheless, basal leaf
petioles and stem showed as the major fatty acids, about 20% palmitic acid, 19%
linoleic acid and 24% alfa-linoleic acid.
The GLA was present in all the tissues analyzed. The basal leaf petioles and stem
showed the highest content of GLA with a mean value of 15.8 and 15.5 respectively.
In contrast to the borage seeds, stearidonic acid was synthesized in substantial
amounts in all the tissues evaluated, with the highest levels, 17.04 and 18.72 in basal
and main stem leaves. Short-chain fatty acids such as lauric and miristic acids and
traces of C20-C24 fatty acids were found in the tissues evaluated.
Inverse relation was found between oleic acid and GLA and between linoleic vs. alfa-
linolenic and stearidonic acids in basal leaves. In main stem leaves and basal leaf
petioles was also found inverse relation between alfa-linolenic vs. oleic and linoleic
acids.
DISCUSSION
In contrast to the borage seeds, the lipids of basal leaf petioles, leaves and stem of
plant contained two n-3 acids alfa-linolenic and stearidonic acids. Alfa-linolenic acid
and GLA were detected in all the tissues of borage, which indicates that both delta-15
and delta-6 desaturases are active, but the effect of the delta-15 desaturases
predominates. Therefore alfa-linolenic is the major octadecatrienoic acid in the tissues
of borage. These results support that alfa-linolenic acid is channeled almost exclusively
to the phospholipids and glycolipids, which are the major constituents of the cellular
membranes. Whereas GLA is esterified mainly in the storage lipids, i.e triacylglicerols
of borage seeds, such as occur with evening primrose plant [9]. New studies are being
realized in our group of investigation for determining the proportions of these lipids in
the tissues of borage.
The inverse relation found between GLA and oleic acid in basal leaves suggests that
GLA is formed to desaturation of oleic acid., which corroborates the hypothetical
biosynthetic pathways to fatty acids in borage seeds that proposes Gunstone,[10] and
corroborates results obtained by Cherry et al. [11] and Whipkey et al. [12], in which the
sequence of alternating desaturations from oleic acid to GLA is significant. However,
others patterns of desaturations must be considered for explain the n-3 acid synthesis.
Considering the inverse relation found in basal leaves, between linoleic acid vs. alfa-
linoleic and stearidonic acids, as well as in main stem leaves and basal leaf petioles
between alfa linoleic acid vs. oleic acid and linoleic acid the results are compatible with
the following sequential desaturations:
oleic acid ® linoleic acid ® gamma-linolenic acid ¯ alfa-linolenic acid (18:3, n-3). ¯ stearidonic acid (18:4, n-4).
Additional works are being realized in order to explain the probable biosynthetic
pathways to fatty acids in borage.
ACKNOWLEDGMENTS
The authors wish to extend their appreciation to Dr. Vicente Dominguez (Centre of
Agricultural Research of Rioja) for providing the accessions of borage.
REFERENCES
1 D. F. Horrobin. Plant Lipid Res. Vol. 31, No. 2, (1992) 163.
2 B. J. F. Hudson. J. Am. Oil Chem. Soc. 61 (1984) 540.
3 H. Traitler, H. Winter, U. Richli and Y. Ingenbleek. Lipids. 19 (1984) 923.
4 R. B. Wolf, R. Kleiman and R. E. England. J. Am. Oil Chem. Soc. 60 (1983)
1858.
5 K. D. Mukherjee and I. Kiewitt. J. Agric. Food Chem., Vol. 35, (1987) 1009.
6 AOAC . Official Methods of Analysis, Williams, Arlington, 1984.
7 R. Garcés and M. Mancha. Analytical Biochemistry, Vol. 211 , (1993) 139.
8 M. del Río, J.M.Fernández and A. de Haro. Grasas y aceites. 44 (1993) 125.
8 G. , Lotti and M. F., Quartacci. Agrochimica. 34 . (1990) 243.
9 F. D. Gunstone. Plant Lipid Res. Vol. 31, No. 2, (1992) 145.
10 J. H. Cherry, L. Bishop, P. M., Hasegawa and H. R., Leffler. Ibid. 24 .(1985). 237
11 A., Whipkey, J. E. Simon and J. Janick. JAOCS. 65. (1988) 979.
TABLE I
Fatty Acid Composition and oil content of the edible portion of borage (data are given as peak area percentages).
oil content
a
12:0 b 14:0 16:0 16:1 18:0 18:1 18:2
main stem leaf 93.740.92-
8.24
0.180-0.79
14.3012.42-18.08
1.60.31-4.52
2.951.54-5.63
3.061.13-5.67
9.667.67-11.74
basal leaf 93.31
0-8.011
0-8.715.73
12.46-18.191.13
0-1.823.07
2.06-4.493.24
1.01-6.159.34
7.04-11.14
basal leaf petioles 1.8 ----------0.89
0-2.3420.9
19.09-21.911.63
0-3.843.59
0-7.153.78
0-8.4417.86
13.14-20.98
stem 0.91 ----------1.37
0-2.1321.11
17.39-26.581.41
0.5-2.041.87
0-4.282.23
0-5.2621.05
16-97-28.17
seed 34.3 ---------- ---------10.88
8.5-18.7 -----------3.86
2.4-9.218.43
9.9-30.238.12
26.7-44.5
a Oil content is given as percentages of dry weight.
b Fatty acids are designated by number of carbon atoms:number of double bonds. c The n-6 (or omega-6) and the n-3 (or omega-3) define the position of the first
double bond in the molecule starting from the carbon atom at the methyl of the
chain.dIncluding 20:0, 20:1, 22:0 and 22:1.eIncluding 20:1 and 22:1.