(21,22), and ions (6, 10). Accordingly, the loss of K+ has been shown to be one of the first indications of membrane permeability changes under a variety of stresses (5,17,18,20).The net loss of K+ from ozone-treated Chlorella cells suspended in a Tris buffer has been measured using a cation-specific electrode (6). The electrode method measured only the net loss of K+ at a low external K+ concentration. Thus, the effect of ozone upon the K+ influx and efflux could not be studied separately under physiological conditions. The use of Tris as a buffer induces K+ loss itself, thus making the interpretation of K+ loss difficult (7). For influx measurements, I09 cells were added to a total volume of 5 ml of medium in a stirred and temperature-regulated cuvette (38 C). Label ("6RbCl, New England Nuclear) was added prior to cell addition and subsequent gassing. Either oxygen or ozone in oxygen, produced and measured as previously described (6, 11), was introduced into the agitated solution through a 50-,ul micropipette. At specific times, a 200-jul aliquot of the cell suspension was withdrawn, filtered on a Millipore filter (0.45 ,um), and washed with 5 ml of unlabeled medium. The cell sample on the filter was removed to a drying stand, bleached with I drop of Purex, and placed in a toluene-Triton cocktail for scintillation counting (14).Uptake rates were calculated from the amount of 8'Rb present in the cells at various time intervals, based upon the external specific radioactivity of Rb+ and K+.In experiments during which the initial uptake kinetics were examined, l-ml aliquots from a batch of cells (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)

Ozone-treated cells of the photosynthetic green alga ChloreUa sorokiniana var. pacificensis exhibit an exponential decline in viability, as measured by their ability to form colonies on agar plates. Postexposure conditions appear to have little, if any, effect on this rate of decline. Except in young (early exponential phase) cells, culture age did not affect this rate. The decline in cell viability was correlated with the production of malondialdehyde, arising from the oxidative breakdown of an ozonide of unsaturated fatty acid material. The loss of fatty acids is substantiated by gas-liquid chromatography. A loss of 5 X 10`5 moles of fatty acid per cell corresponds to 75% nonviable cells after 50 minutes of ozone exposure.

Exposure of Chlorela sorokiniwa (07-11-05) to ozone inhibits photosynthesis. In this study, the effects of ozone on 02 evolution and fluorescence yields are used to characterize this inhibition. At an ozone dose of about 3 micromnoles delivered to 2 x 10' cells, the photosynthetic rate of the cells is inhibited 50%, as indicated by a decrease in bicarbonate-stimulated 02 evolution (control rate, 1.4 ± 0.3 x 10"1 moles per ceil per minute). Plant productivity is decreased by ozone exposure in three major ways: (a) induction of stomatal closure in sensitive plants, which protects them from further injury but reduces the plant's photosynthetic capacity; (b) reduction in the chloroplast's ability to carry out efficient photosynthesis (8, 10); and (c) induction of necrosis and chlorosis which generally appears from 1 to 2 days after ozone exposure.Chlorella has been used previously to study ozone effects on ion transport pathways (1, 11). In the present study, we describe ozone alterations of photosynthetic competence in Chlorella as they are measured by 02 evolution and fluorescence yield kinetics. In addition, we assess the effects of an externally applied osmotic pressure used to vary the internal water potential. MATERIALS AND METHODSChlorella sorokiniana (07-11-05) were grown autotrophically as described previously (2). Either 02 or ozone was bubbled into 10 ml of a stirred buffered phosphate solution containing 2 x 108 cells/ml for varying times (11). A 1.0-ml cell sample was removed and, after addition of 20 mm NaHCO3, assayed for 02 evolution. 02 evolution was measured by a YSI Clark 02 electrode in a water-jacketed cuvette (total volume about 2 ml) maintained at 38°C. The cuvette was illuminated by red light with wavelengths greater than 600 nm and at an intensity of 50 mw/cm2 (about 80-90%o saturating). The cuvette was constructed so that the electrode just fit into the top opening, minimizing diffusion of 02 from the air but allowing additions by microsyringe. Light intensity was varied with Kodak neutral density filters and measured with a Licor radiometer (corrected to PAR).Ozone was generated in an 02 gas stream by UV irradiation and quantitated colorimetrically by reaction with a neutral solution of KI (3).Fluorometer measurements, using both actinic and measuring beams of light, utilized a noncommercial fluorometer (partially described in ref. A broad-band blue actinic beam of light (450 ± 50 nm; 11 kergs cm-2 s-1) was produced by tungsten bulb projector and blue cutoff filter (Coming CS-4-96), a Wratten blue filter (Type 48A), and a 500-nm cut-offfilter (Optics Technology, Palo Alto, CA). Actinic light passed into the sample along the same optical axis as the measuring beam, but at a 1800 angle to it.Fluorescence was measured at a 900 angle from the optical axis of the measuring beam by an S-20 photomultiplier (Type 9558B, E.M.I.). The photomultiplier was screened by a red cut-off filter (Corning CS-2-60), a red Wratten cellulose filter (Type 26) and a 680-nm interference filter (half-width, ...

No abstract