Fatty Acids of Roots of Selected Species

In order to interpret the structural and functional roles of the lipids in roots, the composition of the lipids of these organs must be determined. A survey was made of the fatty-acid composition of roots of selected crop species. Seeds were germinated under sterile conditions; the lipids were extracted from the roots, methyl ester were prepared of the fatty acids, and the fatty acids were analyzed by gas chromatography. Two groups of plants were found—one which contained larger percentages of linoleic acid in the roots and another which contained larger percentages of linolenic acid. In both groups the most abundant saturated fatty acid was palmitic. The fatty acids of tissue samples taken at different distances from the tip of the root were determined. It was found that the amount and relative percentages of the individual fatty acids varied developmentally. The fatty acids of major lipid classes of corn roots (Zea mays L.) were determined. Most of the fatty acids were esterified in the phospholipids. Apparently the glycolipids are only minor constituents of the lipids of roots. This is in contrast to the lipid composition of photosynthetic tissues in which the glycolipids are major constituents. INTRODUCTION A considerable amount of effort has been expended to identify the lipids and fatty-acid composition of commercially valuable seeds and of photosynthetic tissues, especially leaves and chloroplasts (Hitchcock and Nichols, 1971). The most abundant fatty acid of green leaves is a-linolenic acid, followed by palmitic acid, the most abundant saturated fatty acid. In photosynthetic tissues (leaves), the palmitic acid is associated with a variety of parental lipids, whereas the a-linolenic acid is most commonly esterified in monogalactosyl diglyceride (Nichols and James, 1968). The fatty acids of the roots of a few species have been identified (Kaimal and Lakshminarayana, 1970; Holman and Nichols, 1972) where the most 'Manuscript received June 14, 1973. THE OHIO JOURNAL OF SCIENCE 74(3): 150, May, 1974 No. 3 ROOT FATTY ACIDS 151 abundant saturated fatty acid is palmitic and the most abundant unsaturated fatty acid is generally linoleic (C18:2-18 carbon atoms and 2 double bonds). This paper reports the results of efforts to identify the fatty-acid composition of young roots of some crop species, as well as to identify how these fatty acids of roots vary developmentally. The fatty acid composition of major classes of lipids was also identified. Because of the high concentration of lipids in biological membranes and the possible role of the fatty acids in the function of these membranes, identification of the fatty acid composition of these lipids is important. MATERIALS AND METHODS Growth of Roots Eight different cultivars were used in this study. These were: squash (Cucurbita maxima Duchesne, Burpee's Blue Hubbard), bush bean {Phaseolus vulgaris L., Burpee's Tenderpod), pumpkin (Cucurbita pepo L., Burpee's Small Sugar), watermelon (Citrullus vulgaris Schrad., Burpee variety unknown), sweet corn (Zea mays L., Burpee's Honeycross-Yellow Hybrid), cucumber (two forms, both Cucumis sativus L., Burpee's A and C and Burpee's London), and oats (Avena sativa L., variety unknown; obtained from a local farm store). All operations until the time of harvest were performed in a sterile transfer room. The seeds were first surface sterilized for 45 sec in a 10% Chlorox solution and were then rinsed with sterile distilled water. The seeds germinated at approximately 25°C in the dark in sterile Petri dishes containing sterile distilled water for a period sufficient to obtain enough root material (1-2 gm). In all cases a specific length of root measured from the apex was used for fatty-acid analysis. Segments of corn roots cut at different distances from the apex were used in the studies of development. A known weight (ca. 1-2 g) of tissue was harvested in each case. Lipid Extraction and Fatty-Acid Analysis The harvested root tissue was boiled for 3 min in chloroform-methanol (2:1, v/v). The tissue was then homogenized in chloroform-methanol, filtered, and the filter washed clean (three washings) with the solvent. The extract was then washed through an acidified aqueous layer and dried with anhydrous sodium sulfate. The fatty acids were transesterified with methanolic-HCl (2.5% HC1, w/w) under reflux for 1.5 hr in an atmosphere of nitrogen (Kates, 1964). The methyl esters were purified by passage through silicic acid columns. The fatty acid methyl esters were resolved by gas chromatography on a 10-ft column of 10% DEGS on HMDS-treated acid-washed Chromosorb W. Separations were made isothermally at 185°C. The detector was calibrated with known weights of fattyacid methyl ester standards obtained from Applied Science Company, State College, Pa. Column Chromatography of Lipids Extracts from corn roots were chromatographed into general classes of lipids and then the fatty-acid composition of each of these classes was determined. The aliquot used for lipid analysis was washed with water, dried with anhydrous sodium sulfate, and was then placed on a 10-g column of Unisil silicic acid (Clarkson Chemical Company, Williamsport, Pa.). Neutral lipids were eluted with 500 ml of chloroform, monogalactosyl diglyceride was eluted with 100 ml of chloroformacetone (1:1, v/v), digalactosyl diglyceride was eluted with 100 ml of acetone, and phospholipids were eluted with 100 ml each of chloroform-methanol (9:1, v/v), chloroform-methanol (1:1, v/v), and methanol. The data given in the tables are all averages of three determinations except those in table 1, where the data are averages of only two determinations. 152 R. MITCHELL, ET AL. Vol. 74 RESULTS AND DISCUSSION The fatty-acid compositions of roots of seven species or varieties are given in table 1. The saturated fatty acid found in highest percentage was palmitic (C16:0), and the unsaturated fatty acid found in highest percentage was linolenic (C18:3). These two fatty acids are also most abundant in photosynthetic tissues, although photosynthetic tissues generally contain a higher percentage of C18:3. The relative amounts of the different fatty acids found in the roots of five of these species or varieties resemble the situation found in etiolated barley leaves (Newman et at., 1973).