Detection of the Terpenoids of Pinus Halepensis by Spot Tests

Abstract

In pines, oleoresin droplets may be found in all types of living parenchyma cells, but most oleoresin is produced in the thin-walled epithelial cells surrounding the resin ducts or canals. The limpid oleoresin that exudes from incisions cut in the living pine trees, can be separated by steam distillation into a volatile fraction, gum turpentine, and a nonvolatile residue, rosin which when cold sets to a yellow or brown glass.The main components of the volatile fraction of Pin us h a I e pen sis are a-pinene (87%), myrcene (2%) and sesquiterpenes (4%) [1]. Rosin consists mainly of resin acids which have been separated into two types: the abietic and the pimaric.The purpose of this study was to develop specific, selective and sensitive chromogens which could differentiate between the volatile terpenes, resins and lipids, and to apply such in vi t r 0 information for the location of these compounds in the cells of Pin u s h a I e pen sis with the aid of a micros cope.We have found that spotting of a chloroform solution of natural oleoresin on a glass microplate covered with a thin-layer of silica gel G, formed three distinct concentric zones; the central one was that of resin acids, the middle one of terpenes and the outer zone was of lipids. This method constitutes a kind of circle chromatography, and thus enables us to detect and characterize readily the size and the color of each zone. Standards are spotted on the plate for comparison purposes. Rollins and Wood [2] have found that the contrast between the spot of an unsaturated lipid and the background, using various pH indicators, could be increased by bleaching the background with bromine vapor. We found this technique suitable for detection of terpenes and lipids of Pinus since it eliminates any form of heat treatment. ExperimentalPreparation of plates: Microscope slides were spread on glass plates (20X20 em): the suspension was prepared by shaking a mixture of silica gel G and water, 1: 2, for 30 sec and was applied uniformly to a thickness of 0.1 mm with a Desaga applicator.