Skip to main content
PMC home page
Plant Physiology logoLink to Plant Physiology
. 1999 Nov;121(3):1047–1052. doi: 10.1104/pp.121.3.1047

Enhanced Formation of α-Tocopherol and Highly Oxidized Abietane Diterpenes in Water-Stressed Rosemary Plants1

Sergi Munné-Bosch 1, Karin Schwarz 1, Leonor Alegre 1,*
PMCID: PMC59469  PMID: 10557254

Abstract

The lipid-soluble antioxidants α-tocopherol and carnosic acid were studied in field-grown rosemary (Rosmarinus officinalis L.) plants subjected to drought. During summer in the Mediterranean region, the predawn water potential decreased to −3 MPa and the relative water content to 42%, which caused a depletion of the maximum diurnal CO2 assimilation rate by 80%. Meanwhile, the maximum efficiency of photosystem II photochemistry and the chlorophyll content of leaves remained unaltered, indicative of the absence of photooxidative damage. The concentration of α-tocopherol increased by 15-fold and that of carotenoids by approximately 26% in response to water stress. Enhanced formation of the highly oxidized abietane diterpenes isorosmanol (by 25%) and dimethyl isorosmanol (by 40%) was observed during the summer as result of the oxidation of carnosic acid, which decreased by 22%. The large amounts of carnosic acid, α-tocopherol, and carotenoids present in rosemary leaves might contribute to the prevention of oxidative damage in plants exposed to drought.

Plants grown in Mediterranean field conditions cope with the interaction of several stresses, especially during the summer when water deficit, high light, and high temperature limit CO2 fixation (Munné-Bosch and Alegre, 1999). Under such environmental stress conditions, limitations of carbon assimilation within the plant cell result in exposure to excess excitation energy. Although dissipation of excitation energy in the photosystem II (PSII) antennae by non-radiative decay processes and other electron sinks such as nitrogen metabolism and oxygen reduction via photorespiration confer photoprotection to some extent, it has been suggested that electron flux to oxygen via the Mehler reaction increases (Fryer et al., 1998). This results in the formation of activated oxygen species, which can lead to chlorophyll degradation, photodamage, and lipid photooxidation (Smirnoff, 1993; Foyer et al., 1994a; Asada, 1996; Osmond et al., 1997).

To cope with water stress and to avoid photooxidative damage, plants have evolved a series of enzymatic and nonenzymatic antioxidant systems. From the latter, tocopherols and carotenes protect lipid membranes from oxidative stress because they deactivate singlet oxygen, reduce superoxide radicals, and terminate lipid peroxidation by reducing fatty acyl peroxy radicals (Burton and Ingold, 1984; Fryer, 1992; Polle and Rennenberg, 1994). Although α-tocopherol is found throughout the plant kingdom, abietane diterpenes such as carnosic acid, methoxycarnosic acid, carnosol, rosmanol, and isorosmanol have only been found in the genus Salvia and in rosemary (Rosmarinus officinalis L.). Carnosic acid and carnosol are present in large amounts within the plant and possess the highest antioxidant activity of the abietane diterpenes (Nakatani, 1992; Schwarz and Ternes, 1992). It has been demonstrated that these abietane diterpenes are able to inhibit lipid peroxidation and superoxide generation in isolated chloroplasts and microsomes, protecting biological membranes against chemically induced oxidative stresses (Haraguchi et al., 1995; Haraguchi, 1998).

Some authors have proposed an in vivo oxidative pathway for carnosic acid in which enzymatic dehydrogenation and activated oxygen play a key role (Luis et., 1994b; González et al., 1995). In these reactions, formation of highly oxidized abietane diterpenes such as rosmanol, isorosmanol, and dimethyl isorosmanol occurs after the oxidation of carnosic acid (Fig. 1). In contrast to what happens with α-tocopherol (Winston, 1990; Kruk and Strzalka, 1995), carnosic acid is not regenerated once it is oxidized (González et al., 1995).

Figure 1.

Figure 1

Open in a new tab

Formation of abietane diterpenes in rosemary plants. The highly oxidized abietane diterpenes rosmanol, isorosmanol, and dimethyl isorosmanol are formed from carnosic acid by enzymatic dehydrogenation and the participation of activated oxygen (adapted from Luis et al., 1994b).

The aim of this work was to study the effects of drought on photosynthesis and the role of the antioxidants α-tocopherol and abietane diterpenes in drought-induced oxidative stress in rosemary plants growing in Mediterranean field conditions.

MATERIALS AND METHODS

Plant Material and Growth Conditions

Cuttings of rosemary (Rosmarinus officinalis L.) were rooted and grown in 0.5-L pots containing a mixture of soil:peat:perlite (1:1:1, v/v). The plants were maintained in a greenhouse with controlled temperature (24°C/18°C, day/night) and watered twice a week, once with tap water and once with Hoagland's solution. After 1 year of growth, plants of the same height (35 cm) were transplanted to the experimental fields of the University of Barcelona (Spain). Before the plants were transferred, the soil was treated with N:P:K (1:1:1, nutrient ratio) fertilizer at the rate of 100 kg N ha−1. Plants were transplanted on April 10, 1996, and until October 1996 were watered twice a week with 15 mm of water. From November 1996 to August 1997 plants received only natural rainfall. Sixteen plants of approximately the same size were chosen for this study. The experiment was carried out from April 1997 to August 1997, and during this period 5 sunny days were selected for the measurements (April 30, May 28, July 1, July 26, and August 22).

Environmental conditions were monitored by a weather station (Delta-T Devices, Newmarket, UK). Measurements of photosynthetic photon flux density, air temperature, and relative humidity were taken at 1-min intervals, and 5-min means were logged. The photosynthetic photon flux density was measured with a sensor (Quantum Sensor, LI-COR, Lincoln, NE), air temperature and relative humidity were measured with a thermocouple (Vaisala, Helsinki), and the precipitation (in millimeters) was measured with a standard rain gauge. The vapor pressure deficit was determined from relative humidity and air temperature data according to the method of Nobel (1991).

Abietane Diterpenes and α-Tocopherol Determination

Leaves were collected at predawn and midday over the 5-d period, immediately frozen in liquid nitrogen, and stored at −80°C until analysis. Leaves were freeze-dried, and after grinding, leaf samples (1 g) were extracted with 5 mL of methanol containing 5 mg of citric acid and isoascorbic acid per 100 mL and sonicated for 20 s in a sonicator (Sonoplus HD 200, Bandelin, Berlin) equipped with a probe (MS 73, Bandelin). The extract was centrifuged at 1,500g for 3 min at 3°C, and the supernatant was transferred into a volumetric flask. The extraction procedure was repeated four times with 5 mL of methanol each time. The collected supernatants were put through a 110-μm-pore-size cellulose nitrate filter (Schleicher & Schuell, Dassel, Germany). For the measurement of abietane diterpenes, the supernatant was resuspended in an appropriate volume of methanol (1:10, v/v) prior to injection. For α-tocopherol determination the extract was passed through vacuum distillation to eliminate the methanol, de-gassed with nitrogen, and stored at −20°C. Prior to injection, the extract was dissolved in 4 mL of acetonitrile and centrifuged at 1,500g for 3 min at 3°C.

For analysis of α-tocopherol and abietane diterpenes a HPLC method similar to that described by Schwarz and Ternes (1992) was used. α-Tocopherol was separated at room temperature on a 5-μm column (250 × 4 mm; ODS Hypersil, Knauer, Berlin) using acetonitrile:distilled water:2 m citric acid (98:2:0.2, v/v) as an eluant at a flow rate of 1.1 mL min−1. UV detection was carried out at 295 nm (Spectralphotometer, Knauer) and fluorescence detection was carried out at an excitation wavelength of 295 nm and emission at 340 nm (FS 970, Kratos, Ramsey, NJ). One-hundred microliters of sample was injected, and duplicates were run for each extract. α-Tocopherol (98.4% purity, Merck, Rahway, NJ) was used for calibration. Diterpenes were separated on the 5-μm column for 52 min at a flow rate of 0.6 mL min−1. Eluant A consisted of acetonitrile: distilled water:2 m citric acid (51:49:0.83, v/v) and eluant B was acetonitrile:distilled water:2 m citric acid (97:3:0.5, v/v). The UV detection was carried out at 230 nm and the electrochemical detection at +800 mV with a range of 10 nA (model 656/641 VA, Metrohm, Filderstadt, Germany). The injection volume was 20 μL and duplicates were run for each extract. Carnosic acid (98% purity) was used for calibration. All abietane diterpenes were quantified relative to carnosic acid at 230 nm, because the UV spectra of other abietane diterpenes are similar to that of carnosic acid.

Physiological Parameters

The water potential of 10-cm apical shoots was measured using a Scholander-type pressure chamber (ARIMAD-2, ARI Far Charuv-Water Supply Accessories, Ramat Haqolan, Israel). The relative leaf water content was determined as RWC (%) = (fresh weight − dry weight)/(turgid weight − dry weight) × 100. Diurnal cycles of CO2 assimilation rates were performed using a portable measuring system (model 6200, LI-COR) in the field. Net CO2 assimilation rates were calculated from changes in the gas concentration over a 20-s period using the equations developed by von Caemmerer and Farquhar (1981). Six measurements were made at 1.5-h intervals from predawn to sunset. Steady-state modulated chlorophyll fluorescence of leaves was measured using a portable fluorimeter (mini-PAM, Walz, Effeltrich, Germany).

The maximum and relative quantum efficiency of PSII photochemistry (φPSII) was estimated according to the method of Genty et al. (1989) as: φPSII = (Fm′ − Fs)/Fm′ and Fv/Fm = (Fm − Fo)/Fm respectively, where Fm and Fm′ are the maximum fluorescence yields obtained in the dark- and light-adapted state, respectively, Fs is the fluorescence yield at steady-state photosynthesis, Fv is the variable fluorescence yield, and Fo is the basal fluorescence yield obtained in the dark-adapted state. Chlorophyll a + b and the total carotenoid content of leaves were determined spectrophotometrically in 80% (v/v) acetone extracts using the equations described by Lichtenthaler (1987).

RESULTS

Environmental Conditions

The pattern of precipitation during the measurement period is shown in Figure 2. It was characterized by two rainfalls of 59 and 10 mm just before the experiment began, followed by a very dry period from the beginning of May until the end of August, disturbed only by some rainfalls concentrated at the end of June and beginning of July. During the period of the study, the maximum photosynthetic photon flux density moved around 1,800 μmol m−2 s−1, midday air temperature increased from 19°C to 30°C as the season progressed, and the vapor pressure deficit was very high on April 30 and May 28 at midday, with values around 4 kPa (Table I).

Figure 2.

Figure 2

Open in a new tab

Rainfall pattern at the experimental fields of the University of Barcelona during the period of study. Measurement days are indicated by asterisks.

Table I.

Environmental conditions, water relations, and photosynthetic parameters of rosemary plants grown in Mediterranean field conditions

Day PPFD VPD T ψ RWC Amax A φPSII
μmol m−2 s−1 KPa °C MPa % μmol m−2 s−1
30 April 1,783 3.8 19.0 −0.29  ± 0.07 80.18  ± 1.90 10.06 ± 0.61 5.72  ± 0.67 0.26  ± 0.03
28 May 1,885 4.4 24.6 −2.14  ± 0.14 61.45  ± 3.63 3.37 ± 0.23 1.60  ± 0.16 0.27  ± 0.06
1 July 1,816 1.6 21.3 −0.32  ± 0.07 75.35  ± 2.50 9.98 ± 0.72 8.62  ± 1.07 0.34  ± 0.04
26 July 1,884 2.7 26.3 −3.00  ± 0.12 43.48  ± 0.99 2.63 ± 0.28 0.85  ± 0.10 0.33  ± 0.05
22 August 1,654 2.8 30.6 −3.10  ± 0.08 42.67  ± 2.16 1.50 ± 0.15 0.21  ± 0.07 0.32  ± 0.04
Open in a new tab

Values of photosynthetically active photon flux density (PPFD), vapor pressure deficit (VPD), and air temperature (T) at midday are given. Water potential (ψ) and RWC correspond to predawn values. Amax values correspond to measurements taken in the morning, and A and φPSII are for measurements made at midday. Data are the means ± se of six independent replicates.

Plant Water Status and Photosynthesis

Rosemary plants grown in natural field conditions were subjected to different degrees of water stress during the spring and summer of 1997. After the major rainfalls of April, plants had a water potential of −0.3 MPa and a RWC of 80% at predawn (Table I, April 30). As water stress progressed during May, the water potential decreased to −2.14 MPa and RWC to 61% (May 28). Both parameters recovered again to similar pre-drought values after the rainfalls of the end of June (July 1). Nevertheless, there was a second dry period during July and August, which decreased the predawn water potential values from −0.3 to −3 MPa and the predawn RWC values from 75% to 42%, indicating severe stress (Table I, July 26 and August 22).

Maximum diurnal CO2 assimilation rates moved from 10 to 2 μmol m−2 s−1 during this period, depending on plant water status (Table I). Summer drought caused a depletion of 80% in the maximum diurnal CO2 assimilation rate and an almost complete depletion of photosynthesis at midday. Rosemary plants displayed midday depression of photosynthesis during the whole period of the study (except on July 1) and this was not only associated with improved plant water status but also with low vapor pressure deficit during that day (Table I).

Despite the large depletion of photosynthesis at midday in water-stressed plants, the φPSII was maintained unaltered (ANOVA, P < 0.05 probability level) during the whole period of study, with midday φPSII values around 0.30 (Table I). The maximum efficiency of PSII photochemistry and the chlorophyll content of the leaves remained significantly unaltered (ANOVA, P < 0.05 probability level) throughout the experiment at around 0.78 and 30 μg cm−2, respectively (Fig. 3, A and B). The total carotenoid content of the leaves increased by 26% at midday, when RWC values below 50% were reached (Fig. 3C).

Figure 3.

Figure 3

Open in a new tab

Relationship between the RWC at predawn and the ratio of variable to maximum fluorescence (Fv/Fm) (A), chlorophyll a + b (B), carotenoid (C), and α-tocopherol (D) content in rosemary plants. Values at predawn (○) and midday (●) are given. Data are the means of six replicates ± se.

α-Tocopherol and Drought

Figure 3D shows the relationship between the RWC and the α-tocopherol content of leaves. Rosemary plants showed a concentration ranging from 2 to 35 μg of α-tocopherol g−1 dry weight depending on plant water status but irrespective of time of day, and a 15-fold increase in the α-tocopherol content of leaves, when predawn RWC values below 50% were reached on July 26 and August 22. Although the α-tocopherol content of leaves was highly sensitive to RWC decreases imposed by drought, no significant differences were observed between predawn and midday during any of the measurement days.

Abietane Diterpenes and Drought

The relationship between the RWC and the abietane diterpene concentration in plants at predawn and midday is shown in Figure 4. Carnosic acid, the main constituent of abietane diterpenes with a concentration ranging from 2.8 to 4.2 mg g−1 dry weight, followed a completely different pattern from that observed for α-tocopherol in response to water stress. Figure 4A shows that carnosic acid decreased both in response to water stress and during the day. The concentration of carnosic acid at predawn had a strong correlation (r2 =0.97) with RWC decreases in plants, decreasing by 22% when the RWC reached values below 50%. The concentration of carnosic acid also decreased by 12% during the day, although the differences between predawn and midday were smaller when RWC values below 50% were reached.

Figure 4.

Figure 4

Open in a new tab

Relationship between the RWC at predawn and the abietane diterpene content in rosemary plants. A, B, and C show decreases in the concentration of the abietane diterpenes carnosic acid, carnosol, and methoxycarnosic acid in response to water stress and during the day. D and E show enhanced formation of the highly oxidized abietane diterpenes isorosmanol and dimethyl isorosmanol in response to water stress. This behavior is observed very clearly in water-stressed plants at midday. Values at predawn (○) and midday (●) are given. DMIR, Dimethyl isorosmanol. Data are the means of four replicates ± se.

The abietane diterpenes carnosol (Fig. 4B), present in concentrations ranging from 0.5 to 1.3 mg g−1 dry weight, and methoxycarnosic acid (Fig. 4C), present in concentrations ranging from 0.3 to 0.5 mg g−1 dry weight, followed a trend similar to that observed for carnosic acid. In contrast, the highly oxidized abietane diterpenes isorosmanol (Fig. 4D) and dimethyl isorosmanol (Fig. 4E), present in concentrations ranging from 0.28 to 0.36 mg g−1 dry weight and from 0.50 to 0.72 mg g−1 dry weight, respectively, followed a different trend. The differences during the day disappeared in water-stressed plants for these compounds and the concentrations increased at midday by 25% and 40%, respectively, in response to water stress. Rosmanol, another highly oxidized abietane diterpene, was also found in rosemary leaves at low concentrations ranging from 0.10 to 0.12 mg g−1 dry weight throughout the experiment.

DISCUSSION

Rosemary plants maintained constant midday φPSII values throughout the experiment, whereas the net CO2 assimilation rates decreased by 80% when RWC values below 50% were attained. Plants suffered from severe water stress during the summer, but the maintenance of the chlorophyll content of leaves and the constant ratio of variable to maximum fluorescence throughout the experiment show that the photosynthetic machinery was unlikely to be damaged by dehydration (Cornic and Massacci, 1996). Efficient means for protection from photoinhibitory damage may be provided by the thermal dissipation of excess excitation energy, photorespiration that may serve as a sink for the consumption of excess reducing equivalents, and scavenger systems for the removal of reactive oxygen species (Asada, 1994; Foyer et al., 1994b).

A decrease in carnosic acid and carnosol concentration, the most active antioxidants and abundant abietane diterpene compounds, was observed under drought conditions. The concentration of these abietane diterpenes also decreased during the day. By contrast, the concentration of the highly oxidized abietane diterpenes isorosmanol and dimethyl isorosmanol increased in response to water stress, and the differences between the concentrations at predawn and midday completely disappeared in water-stressed plants for these compounds, indicating that the formation of highly oxidized abietane diterpenes increased when plants were subjected to the interaction of water stress and high light during Mediterranean summer. Carnosic acid gives rise to highly oxidized diterpenes such as isorosmanol, dimethyl isorosmanol, and other related compounds by enzymatic dehydrogenation and scavenging of activated oxygen (Luis et al., 1994a; Luis et al., 1994b; González et al., 1995). Therefore, the enhanced formation of highly oxidized abietane diterpenes was most likely to be caused by the antioxidant activity of carnosic acid when plants were subjected to water stress and high light during Mediterranean summer, when photosynthesis was limited and activated oxygen formation may have increased. The decrease observed in the concentration of carnosic acid during the summer was therefore due to its consumption during the summer drought, as well as to the lack of regeneration once oxidized (Luis et al., 1994b; González et al., 1995). Levinsohn et al. (1993) and McGarvey and Croteau (1995) demonstrated that the diterpene cyclase activity decreases in coniferous plants in response to light and water stresses. Thus, the possibility that part of the decrease observed might also be caused by a decrease in the synthesis of carnosic acid should not be ruled out. Although the concentration of carnosic acid decreased during the summer, the large amounts found in rosemary leaves might allow continuous antioxidant activity within the plant cell.

The 15-fold increases in α-tocopherol observed in water-stressed plants might prevent chlorophyll photooxidation, as has been shown previously (Wise and Naylor, 1987; Simontacchi et al., 1993). No differences were observed in the α-tocopherol content of leaves between predawn and midday throughout the experiment, suggesting that although the α-tocopherol concentration of leaves increased in response to water stress, this species was able to efficiently regenerate α-tocopherol during the day. The increase observed in the carotenoid content of leaves could also contribute to prevent chlorophyll degradation and photodamage in rosemary plants.

In summary, our results prove that enhanced formation of highly oxidized abietane diterpenes occurred in rosemary plants as a result of the antioxidant activity of carnosic acid when photosynthesis was limited and activated oxygen formation may have increased. The increases in α-tocopherol and carotenoid concentration, together with the large amounts of carnosic acid found in rosemary leaves, contribute to help the plant withstand water stress and high light during Mediterranean summer.

ACKNOWLEDGMENT

We are very grateful to the staff of the experimental fields of the University of Barcelona for technical assistance.

Footnotes

1

This research was supported by grant no. PB96–1257 from Programa Sectorial de Promoción General del Conocimiento and by a research studentship to S.M. from the University of Barcelona.

LITERATURE CITED

  1. Asada K. Protection and action of active oxygen species in photosynthetic tissues. In: Foyer CH, Mullineaux PM, editors. Causes of Photooxidative Stress and Amelioration of Defence Systems in Plants. Boca Raton, FL: CRC Press; 1994. pp. 77–104. [Google Scholar]
  2. Asada K. Radical production and scavenging in the chloroplasts. In: Baker NR, editor. Photosynthesis and the Environment. Dordrecht, The Netherlands: Kluwer Academic Publishers; 1996. pp. 123–150. [Google Scholar]
  3. Burton GW, Ingold KU. β-Carotene: an unusual type of lipid antioxidant. Science. 1984;224:569–573. doi: 10.1126/science.6710156. [DOI] [PubMed] [Google Scholar]
  4. Cornic G, Massacci A. Leaf photosynthesis under drought stress. In: Baker NR, editor. Photosynthesis and the Environment. Dordrecht, The Netherlands: Kluwer Academic Publishers; 1996. pp. 347–366. [Google Scholar]
  5. Foyer CH, Descourvières P, Kunert KJ. Protection against oxygen radicals: an important defence mechanism studied in transgenic plants. Plant Cell Environ. 1994a;17:507–523. [Google Scholar]
  6. Foyer CH, Lelandais M, Kunert KJ. Photooxidative stress in plants. Physiol Plant. 1994b;92:696–717. [Google Scholar]
  7. Fryer MJ. The antioxidant effects of thylacoid vitamin E (α-tocopherol) Plant Cell Environ. 1992;15:381–392. [Google Scholar]
  8. Fryer MJ, Andrews JR, Oxborough K, Blowers DA, Baker NR. Relationship between CO2 assimilation, photosynthetic electron transport, and active O2 metabolism in leaves of maize in the field during periods of low temperature. Plant Physiol. 1998;116:571–580. doi: 10.1104/pp.116.2.571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Genty B, Briantais JM, Baker NR. The relationship between the quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence. Biochim Biophys Acta. 1989;990:87–92. [Google Scholar]
  10. González AG, Andrés LS, Aguiar ZE, Luis JG. Diterpenes from Salvia mellifera and their biogenetic significance. Phytochemistry. 1995;31:1297–1305. [Google Scholar]
  11. Haraguchi H. Protection against oxidative stresses by plant secondary metabolites (abstract no. 1186) Plant Physiol. 1998;65:S–1676. [Google Scholar]
  12. Haraguchi H, Saito T, Okamura N, Yagi A. Inhibition of lipid peroxidation and superoxide generation by diterpenoids from Rosmarinus officinalis. Planta Med. 1995;61:333–336. doi: 10.1055/s-2006-958094. [DOI] [PubMed] [Google Scholar]
  13. Kruk J, Strzalka K. Occurrence and function of α-tocopherol quinone in plants. J Plant Physiol. 1995;145:405–409. [Google Scholar]
  14. Levinsohn E, Gijzen M, Muzika RM, Barton K, Croteau R. Oleoresinosis in grand fir (Abies grandis) saplings and mature trees: modulation of this wound response by light and water stresses. Plant Physiol. 1993;101:1021–1028. doi: 10.1104/pp.101.3.1021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lichtenthaler HK. Chlorophylls and carotenoids: pigments of photosynthetic biomembranes. Methods Enzymol. 1987;148:350–382. [Google Scholar]
  16. Luis JG, Andrés LS, Fletcher WQ. Chemical evidence for the participation of a perepoxide intermediate in the reaction of singlet oxygen with mono-olefins in relationship with the biogenetic pathway to highly oxidised abietane diterpenes. Tetrahedron Lett. 1994a;35:179–182. [Google Scholar]
  17. Luis JG, Quiñones W, Grillo TA, Kishi MP. Diterpenes from the aerial part of Salvia columbariae. Phytochemistry. 1994b;35:1371–1374. [Google Scholar]
  18. McGarvey DJ, Croteau R. Terpenoid metabolism. Plant Cell. 1995;7:1015–1026. doi: 10.1105/tpc.7.7.1015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Munné-Bosch S, Alegre L. Role of dew on the recovery of water-stressed Melissa officinalis L. plants. J Plant Physiol. 1999;154:759–766. [Google Scholar]
  20. Nakatani N. Natural antioxidants from spices. In: Huang MT, Ho CT, Lee CI, editors. Phenolic Compounds in Food and Their Effects on Health II: Antioxidants and Cancer Prevention. Washington, DC: American Chemical Society; 1992. pp. 72–86. [Google Scholar]
  21. Nobel PS. Physicochemical and Environmental Plant Physiology. San Diego: Academic Press; 1991. [Google Scholar]
  22. Osmond B, Badger M, Maxwell K, Björkman O, Leegod R. Too many photons: photorespiration, photoinhibition and photooxidation. Trends Plant Sci. 1997;2:119–120. [Google Scholar]
  23. Polle A, Rennenberg H. Photooxidative stress in trees. In: Foyer CH, Mullineaux PM, editors. Causes of Photoxidative Stress and Amelioration of Defence Systems in Plants. Boca Raton, FL: CRC Press; 1994. [Google Scholar]
  24. Schwarz K, Ternes W. Antioxidative constituents of Rosmarinus officinalis and Salvia officinalis. Z Lebensm-Unters -Forsch. 1992;195:95–98. doi: 10.1007/BF01201765. [DOI] [PubMed] [Google Scholar]
  25. Simontacchi M, Caro A, Fraga CG, Puntarulo S. Oxidative stress affects α-tocopherol content in soybean embryonic axes upon imbibition and following germination. Plant Physiol. 1993;103:949–953. doi: 10.1104/pp.103.3.949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Smirnoff N. Tansley review no. 52: the role of active oxygen in the response of plants to water deficit and desiccation. New Phytol. 1993;125:27–58. doi: 10.1111/j.1469-8137.1993.tb03863.x. [DOI] [PubMed] [Google Scholar]
  27. von Caemmerer S, Farquhar GD. Some relations between the biochemistry of photosynthesis and the gas exchange of leaves. Planta. 1981;153:376–387. doi: 10.1007/BF00384257. [DOI] [PubMed] [Google Scholar]
  28. Winston GW. Physiochemical basis for free radical formation in cells: production and defences. In: Alscher RG, Cummings JR, editors. Stress Responses in Plants: Adaptation and Acclimation Mechanisms. New York: Wiley-Liss; 1990. pp. 57–86. [Google Scholar]
  29. Wise RR, Naylor AW. Chilling enhanced photooxidation. Plant Physiol. 1987;83:278–282. doi: 10.1104/pp.83.2.278. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES