Extraction and quantitation of dolichol and dolichyl phosphate in soybean embryo tissue |
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Authors: | Jack W. Rip Kenneth K. Carroll |
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Affiliation: | 1. Department of Geological Sciences, The University of Alabama, Tuscaloosa, AL 35487, United States;2. Department of Plant and Environmental Sciences, New Mexico State University, Las Cruces, NM 88003, United States |
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Abstract: | A procedure for the quantitative extraction of both dolichol and dolichyl phosphate (Dol-P) in plant tissue (soybean embryos) into diethyl ether from an alkaline saponification mixture is described. A complete and quantitative separation of total dolichol and total Dol-P is then obtained based on their respective solubilities in diethyl ether and water. After separation dolichol and Dol-P can both be analyzed and quantitated directly by reverse-phase HPLC on C18 columns without additional purification. The two major homologs of dolichol and Dol-P are those with 17 and 18 isoprene units. The total dolichol and total Dol-P contents of dry embryos were 96.3 +/- 0.8 and 5.3 +/- 0.1 micrograms/g, respectively. The post-HPLC recoveries for dolichol and Dol-P were 101 +/- 2 and 84 +/- 3% respectively, using [1-14C]dolichol and Dol-P containing 20 isoprene units as recovery standards. Dol-P estimations could be carried out on material equivalent to as little as 65 mg embryo tissue. |
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Keywords: | dolichyl phosphate dolichol germination lipids HPLC |
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