Clinal Variation in the Production of Usnic Acid in Cladonia subtenuis along Light Gradients

Abstract

Clinal variation in the usnic acid content of thalli of Cladonia subtenuis along determined light gradients was demonstrated spectrophotometrically. Integrated measurements of light incidence were made using ozalid paper light sensors. The percentage of usnic acid in the dry podetia is directly related to the percentage of full sunlight in the microhabitat of the plants. Extreme values ranged from 0.13% in plants from a site of low light intensity (3% full sunlight) to 2.82% in plants from a relatively open site (51% full sunlight). The yellowish color in the podetia of sun populations was not markedly altered by extraction of the usnic acid. Cladonia subtenuis (Abb.) Evans is a common lichen in the southeastern United 1 This research was supported in part by U.S. Public Health Research Grant GM-08345 from the Division of General Medical Sciences, National Institutes of Health, to Dr. Chicita F. Culberson. I thank Dr. Culberson for her help in this study. 2 Department of Botany, Duke University, Durham, North Carolina 27706. This content downloaded from 207.46.13.172 on Fri, 07 Oct 2016 06:14:56 UTC All use subject to http://about.jstor.org/terms 1969] RUNDEL: CLINAL VARIATION IN USNIC ACID PRODUCTION 41 States, occurring on soil in openings or at the edge of pine woods and on banks along old roadcuts. This species has long been known for great variation between sun and shade forms. This variation was noted by Evans (1944) when he raised the taxon to specific rank. Evans recognized four forms including f. cinerascens, a grayish-green shade form with little yellow color and little or no usnic acid, which he distinguished from the typical f. subtenuis or sun form, yellowish in color and containing usnic acid. These two forms have been separated solely on the basis of the degree of yellow coloring of the podetia which has been assumed to be related to the content of usnic acid, a yellow lichen substance. A very yellow color of the podetia has been considered to indicate a high content of usnic acid, a gray-green color to indicate little or no usnic acid. In the present study an attempt has been made to relate a quantitative assessment of the usnic acid content in C. subtenuis to the light incidence of the habitat. Usnic acid is one of the most widespread of lichen substances and a considerable body of literature concerning it exists. Few studies, however, have concerned themselves with quantitative determinations of the usnic acid content of lichens. Laaskso and Gustafsson (1952) used a ferric chloride reagent to make quantitative measurements of usnic acid sepctrophotometrically for Cladonia alpestris. Another spectrophotometric method was used by Ramaut et al. (1966) in a study of the chemotaxonomic significance of usnic acid in Cladonia tenuis, C. leucophaea, and C. impexa. In their study the height of the absorption curve at 380 my for a benzene extract of usnic acid from the lichens studied was used to measure usnic acid concentration. A more refined spectrophotometric analysis for usnic acid has been used in the present research. Usnic acid in benzene solution has a very strong ultraviolet absorption peak at 285 m/, and this peak provides a more satisfactory measurement of the usnic acid content than the absorption at 380 my where no peak occurs. MATERIALS AND METHODS Mats of Cladonia subtenuis were collected in Duke Forest (Durham County, North Carolina) from a range of ecological sites in which the percentage of full sunlight reaching the microhabitat of each mat had been measured. Light measurements at these sites were made with deciduous trees in full leaf. From each small mat of C. subtenuis studied, approximately 0.5 g of the terminal 5 mm of the podetia were collected and dried to constant weight in a desiccator over concentrated sulfuric acid. Material in obvious fruiting condition was not used. The dried sample was weighed and immediately extracted. The lichen was extracted in 15 ml of benzene for 10 minutes on a steam table. The filtered solutions from four successive extractions were pooled and diluted to 500 ml with benzene. Three ml of this solution were run on a Bausch & Lomb Spectronic 505 spectrophotometer against a benzene reference to give an absorption curve for the 280-330 my range (Fig. 1). The usnic acid content of the samples was determined from the absorption peak at 285 me. A calibration curve relating concentration of usnic acid in milligrams per 500 ml of benzene was obtained using solutions of known concentration of usnic acid. The relationship between peak height and usnic acid concentration is linear over the range measured. From the calibration chart the usnic acid concentration of each sample was determined; these data and the original dry weight of each sample enabled the calculation of the percentage of usnic acid by weight in each sample. Integrated measurements of light incidence on thallus mats in the field were made with ozalid paper light sensors (Friend, 1961). Packets of light-sensitive ozalid paper were exposed for 24 hours on the surface of the mats to be studied. The number of papers bleached by the sun at a given sample point was compared to that bleached in a totally This content downloaded from 207.46.13.172 on Fri, 07 Oct 2016 06:14:56 UTC All use subject to http://about.jstor.org/terms 42 THE BRYOLOGIST [Volume 72