Jennifer R. DeEll

Practical applications of chlorophyll fluorescence in plant biology

Preface. List of Figures. List of Tables. Chlorophyll Fluorescence Nomenclature. 1. An Introduction to Chlorophyll Fluorescence J. Harbinson, E. Rosenqvist. 2. Chlorophyll Fluorescence: A General Description and Nomenclature E. Rosenqvist, O. van Kooten. 3. Applications of Chlorophyll Fluorescence in Forestry and Ecophysiology G.H. Mohammed, P. Zarco-Tejada, J.R. Miller. 4. The Application of Chlorophyll Fluorescence to Study Light, Temperature, and Drought Stress Y. Fracheboud, J. Leipner. 5. Applications of Chlorophyll Fluorescence in Ecotoxicology: Heavy Metals, Herbicides, and Air Pollutants R. Popovic, D. Dewez, P. Juneau. 6. The Application of Chlorophyll Fluorescence in the Aquatic Environment C. Wilhelm. 7. Use of Chlorophyll Fluorescence in Postharvest Quality Assessments of Fruits and Vegetables J.R. DeEll, P.M.A. Toivonen. 8. The Potential of Chlorophyll Fluorescence in Fruit Breeding S. Khanizadeh, J.R. DeEll. Index.

A modified total oxyradical scavenging capacity assay for antioxidants in plant tissues

The total oxyradical scavenging capacity (TOSC) assay of Winston et al. [Free Radical Biol. Med. 24 (1998) 480] was modified through the development and incorporation of a standard curve, yielding a relatively simple but reliable method to quantify the total water-soluble antioxidant capacity (TAC) of plant tissues. The basis of the TOSC assay is the oxidation of α-keto-γ-methiolbutyric acid (KMBA) by 2,2′-azobis-amidinopropane (ABAP) with the evolution of ethylene as the quantifiable end product. A standard curve was established by using a serial dilution (0–100 μM) of the antioxidant Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) which scavenges peroxyl radicals formed from the thermal homolysis of ABAP, thereby reducing the rate of ethylene evolution. Regression analysis of the standard curve yielded a second-degree polynomial (R2=0.9975) that was used to predict the TAC (μMoles Trolox equivalents) of plant tissues. The precision and utility of the method was tested by determining TAC of skin tissue from ‘Delicious’ and ‘Empire’ apples (Malus domestica Borkh.) at various maturities, following cold storage, and after treatment with 1-methylcyclopropene (1-MCP). The TAC of ‘Delicious’ apples was twofold greater than that of ‘Empire’ apples, and remained relatively constant for up to 60 days in cold storage, then significantly decreased thereafter. This decrease during the latter stages of cold storage may be related to the development of superficial scald in this cultivar. ‘Delicious’ and ‘Empire’ fruit treated with 1-MCP had also a significantly higher TAC at various maturities after 120 days of cold storage. This is the first study to show that a treatment with 1-MCP can potentially maintain some intrinsic functional food quality parameters (i.e. antioxidant levels) in stored fruit. The establishment of a standard curve for use, in conjunction with the TOSC assay, resulted in a precise and highly reproducible predictor of TAC in apple fruit, while also permitting a rapid turnover of tissue samples for analysis.

Use of Chlorophyll Fluorescence in Postharvest Quality Assessments of Fruits and Vegetables

Chlorophyll fluorescence was initially developed as a tool for studying photosynthesis. The basis for this analysis is that changes in membranes or membrane-bound constituents of the chloroplast lead to changes in the fluorescence emission characteristics (Rosenqvist and van Kooten, 2002). In postharvest physiological studies, it has been determined that chloroplasts are one of the most sensitive membrane systems, similar in sensitivity to mitochondrial membranes (Toivonen, 1992). Thus, chlorophyll fluorescence changes can be potentially the most sensitive measure of membrane changes or perturbations in the plant cell. This fact permits postharvest researchers to gain useful information on early responses to postharvest stress in chloroplast-containing fruits and vegetables (DeEll et ah, 1999). A wide range of fruits and vegetables have shown chlorophyll fluorescence changes that were useful for prediction of stress response, including those that may not have been thought to contain significant amounts of chlorophyll, such as mature apples.

Timing of postharvest 1-methylcyclopropene treatment affects Bartlett pear quality after storage

DeEll, J. R. and Ehsani-Moghaddam, B. 2011. Timing of postharvest 1-methylcyclopropene treatment affects Bartlett pear quality after storage. Can. J. Plant Sci. 91: 853-858. This study investigated the effects of postharvest 1-methylcyclopropene (1-MCP) treatment timing on the ripening and physiological disorders of Bartlett pears during cold storage and subsequent shelf-life. Pears were held for 1, 3 or 7 d at 3°C after harvest and then treated with 0.3 µL L-1 1-MCP for 24 h at 3°C. Fruit quality attributes were evaluated after 4 mo of cold storage at 0.5°C, plus 1 to 11 d at 22°C. All 1-MCP treatments reduced ethylene production, as well as delayed fruit softening and yellow color development. However, the most substantial benefit of 1-MCP observed was the marked reduction in disorders, especially senescent scald and internal breakdown. The results suggest that 1-MCP treatment 3 d after harvest provided the best balance of reduced disorder development during storage and the ability of Bartlett pears to soften adequately thereafter. Fruit treated with 1-MCP at 1 d after harvest did not soften as much as those treated 3 or 7 d after harvest, while treatment after 7 d provided less control of disorders than treatment after 1 or 3 d.

Oxidative metabolism is associated with physiological disorders in fruits stored under multiple environmental stresses

In combination with low temperature, controlled atmosphere storage and 1-methylcyclopropene (ethylene antagonist) application are used to delay senescence of many fruits and vegetables. Controlled atmosphere consists of low O2 and elevated CO2. When sub-optimal partial pressures are used, these practices represent multiple abiotic stresses that can promote the development of physiological disorders in pome fruit, including flesh browning and cavities, although there is some evidence for genetic differences in susceptibility. In the absence of surface disorders, fruit with flesh injuries are not easily distinguished from asymptomatic fruit until these are consumed. Oxidative stress metabolites tend to accumulate (e.g., γ-aminobutyrate) or rapidly decline (e.g., ascorbate and glutathione) in vegetative tissues exposed to hypoxic and/or elevated CO2 environments. Moreover, these phenomena can be associated with altered energy and redox status. Biochemical investigations of Arabidopsis and tomato plants with genetically-altered levels of enzymes associated with the γ-aminobutyrate shunt and the ascorbate-glutathione pathway indicate that these metabolic processes are functionally related and critical for dampening the oxidative burst in vegetative and fruit tissues, respectively. Here, we hypothesize that γ-aminobutyrate accumulation, as well energy and antioxidant depletion are associated with the development of physiological injury in pome fruit under multiple environmental stresses. An improved understanding of this relationship could assist in maintaining the quality of stored fruit.

Effects of elevated CO2 and 1-methylcyclopropene on storage-related disorders of Ontario-grown Empire apples

Deyman, K. L., Chiu, G., Liu, J., Brikis, C. J., Trobacher, C. P., DeEll, J. R., Shelp, B. J. and Bozzo, G. G. 2014. Effects of elevated CO2 and 1-methylcyclopropene on storage-related disorders of Ontario-grown Empire apples. Can. J. Plant Sci. 94: 857-865. The impact of 1-methylcyclopropene (1-MCP) application on CO2-induced physiological injury in Empire apple fruit during controlled atmosphere storage was assessed over a 3-yr period using an experimental design involving multiple treatment replicates. Fruit harvested at optimal maturity from one or two orchards were treated with or without 1 µL L-1 1-MCP, then chilled at 0 or 3°C under various CO2 partial pressures (5, 2.5 or 0.03 kPa CO2) in the presence of 2.5 kPa O2 for up to 46 wk using a split-plot design. Fruit were sampled periodically for assessment of flesh browning and external peel injury. The maximal incidence of external CO2 injury varied from 15 to 100% over the 3 yr, and the most rapid development of this disorder was evident at 5 kPa CO2. The incidence of external CO2 injury as a function of storage time was influenced by orchard location and storage temperature. Moreover, the incidence of flesh browning at 0°C and 5 kPa CO2 was influenced slightly by orchard; this disorder was never higher than 30%, and the impact of elevated CO2 was inconsistent across years. Notably, there was no evidence for negative effects of 1-MCP on the incidence of storage-related disorders.

1-Methylcyclopropene treatment modifies postharvest behavior of Fantasia nectarines

The effects of 1-methylcyclopropene (1-MCP), an inhibitor of ethylene action, on the ripening and quality of Fantasia nectarines were examined. Fruit were harvested from two commercial orchards and subsequently exposed to 1 μL L-1 of 1-MCP for 24 h at 0°C. Following treatment, fruit were held at 0°C for 0, 2, or 4 wk, and then assessed for quality during a ripening period at 23°C. 1-MCP treatment improved postharvest firmness retention in nectarines after 0 and 2 wk at 0°C plus 4 days at 23°C. Soluble solids concentration (SSC) was lower in nectarines treated with 1 MCP and held for 0 or 4 wk at 0°C, compared with similar non-treated fruit. The peel ground color change from green to yellow was also delayed by 1-MCP. Nectarines treated with 1-MCP exhibited less CO2 and hydrophobic volatile production during 14 days at 23°C, compared with non-treated fruit. The overall inhibition of fruit ripening by 1-MCP appears transitory in Fantasia nectarines. Chilling injury was observed after 4 wk of storage at 0°C a...

Chlorophyll Fluorescence: A New Technique to Screen for Tolerance of Strawberry Flowers to Spring Frost

Abstract Most strawberry cultivars have flowers that are sensitive to temperatures below 0°C. The development of early or very early cultivars with frost resistant flowers is essential in climates with a danger of spring frosts. Traditionally, breeding programs have used visual screening methods to evaluate the damage to pistils and anthers caused by frost. These methods rely on natural seasonal conditions, are time consuming, and do not provide accurate information on the exact temperature that caused the damage. The objective of this study was to evaluate the use of chlorophyll fluorescence (CF) to estimate the low temperature susceptibility of 64 strawberry cultivars. Strawberry flowers were exposed to continuous low temperatures (0°C for 24 h, 1°C for 24 h, -2°C for 24 h, and finally -3°C for 24 h) and CF was measured following the treatments. Variable fluorescence (Fv) decreased somewhat with time in all genotypes when the flowers were held at -3°C, however, the reduction varied with cultivar. The slight reduction of Fv in the more chilling-tolerant cultivars was not significant, while significant linear or quadratic declines were observed in the more chilling susceptible cultivars. Overall, chlorophyll fluorescence appears to be an effective, simple method for evaluating the low temperature susceptibility of strawberry genotypes.

Plant Glyoxylate/Succinic Semialdehyde Reductases: Comparative Biochemical Properties, Function during Chilling Stress, and Subcellular Localization

Plant NADPH-dependent glyoxylate/succinic semialdehyde reductases 1 and 2 (cytosolic GLYR1 and plastidial/mitochondrial GLYR2) are considered to be of particular importance under abiotic stress conditions. Here, the apple (Malus × domestica Borkh.) and rice (Oryza sativa L.) GLYR1s and GLYR2s were characterized and their kinetic properties were compared to those of previously characterized GLYRs from Arabidopsis thaliana [L.] Heynh. The purified recombinant GLYRs had an affinity for glyoxylate and succinic semialdehyde, respectively, in the low micromolar and millimolar ranges, and were inhibited by NADP+. Comparison of the GLYR activity in cell-free extracts from wild-type Arabidopsis and a glyr1 knockout mutant revealed that approximately 85 and 15% of the cellular GLYR activity is cytosolic and plastidial/mitochondrial, respectively. Recovery of GLYR activity in purified mitochondria from the Arabidopsis glyr1 mutant, free from cytosolic GLYR1 or plastidial GLYR2 contamination, provided additional support for the targeting of GLYR2 to mitochondria, as well as plastids. The growth of plantlets or roots of various Arabidopsis lines with altered GLYR activity responded differentially to succinic semialdehyde or glyoxylate under chilling conditions. Taken together, these findings highlight the potential regulation of highly conserved plant GLYRs by NADPH/NADP+ ratios in planta, and their roles in the reduction of toxic aldehydes in plants subjected to chilling stress.

Ancient Plant Glyoxylate/Succinic Semialdehyde Reductases: GLYR1s Are Cytosolic, Whereas GLYR2s Are Localized to Both Mitochondria and Plastids

Plant NADPH-dependent glyoxylate/succinic semialdehyde reductases 1 and 2 (GLYR1 and GLYR2) are considered to be involved in detoxifying harmful aldehydes, thereby preserving plant health during exposure to various abiotic stresses. Phylogenetic analysis revealed that the two GLYR isoforms appeared in the plant lineage prior to the divergence of the Chlorophyta and Streptophyta, which occurred approximately 750 million years ago. Green fluorescent protein fusions of apple (Malus x domestica Borkh.), rice (Oryza sativa L.) and Arabidopsis thaliana [L.] Heynh GLYRs were transiently expressed in tobacco (Nicotiana tabaccum L.) suspension cells or Arabidopsis protoplasts, as well in methoxyfenozide-induced, stably transformed Arabidopsis seedlings. The localization of apple GLYR1 confirmed that this isoform is cytosolic, whereas apple, rice and Arabidopsis GLYR2s were localized to both mitochondria and plastids. These findings highlight the potential involvement of GLYRs within distinct compartments of the plant cell.