Degradation of Chlorophyll and Chlorophyllase Activity in Senescing Barley Leaves

Rodríguez, María Teresa; Gonzélez, M.P.; Linares, J.M.

doi:10.1016/s0176-1617(87)80094-9

Cited by 50 publications

(17 citation statements)

References 15 publications

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“…CLH activity increases in leaves that are either senescent [25] or wounded [16], and in response to MeJA treatment [21]. Thus, we examined chlorophyllide formation using naturally-senescent leaves from 8-week-old Arabidopsis plants, those in which senescence was induced by a 4-day dark treatment, and those in which chlorophyll breakdown was induced by MeJA.…”

Section: Resultsmentioning

confidence: 99%

Simple extraction methods that prevent the artifactual conversion of chlorophyll to chlorophyllide during pigment isolation from leaf samples

Tanaka

2013

Plant Methods

101

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BackgroundWhen conducting plant research, the measurement of photosynthetic pigments can provide basic information on the physiological status of a plant. High-pressure liquid chromatography (HPLC) is becoming widely used for this purpose because it provides an accurate determination of a variety of photosynthetic pigments simultaneously. This technique has a drawback compared with conventional spectroscopic techniques, however, in that it is more prone to structural modification of pigments during extraction, thus potentially generating erroneous results. During pigment extraction procedures with acetone or alcohol, the phytol side chain of chlorophyll is sometimes removed, forming chlorophyllide, which affects chlorophyll measurement using HPLC.ResultsWe evaluated the artifactual chlorophyllide production during chlorophyll extraction by comparing different extraction methods with wild-type and mutant Arabidopsis leaves that lack the major isoform of chlorophyllase. Several extraction methods were compared to provide alternatives to researchers who utilize HPLC for the analysis of chlorophyll levels. As a result, the following three methods are recommended. In the first method, leaves are briefly boiled prior to extraction. In the second method, grinding and homogenization of leaves are performed at sub-zero temperatures. In the third method, N, N’-dimethylformamide (DMF) is used for the extraction of pigments. When compared, the first two methods eliminated almost all chlorophyllide-forming activity in Arabidopsis thaliana, Glebionis coronaria, Pisum sativum L. and Prunus sargentii Rehd. However, DMF effectively suppressed the activity of chlorophyllase only in Arabidopsis leaves.ConclusionChlorophyllide production in leaf extracts is predominantly an artifact. All three methods evaluated in this study reduce the artifactual production of chlorophyllide and are thus suitable for pigment extraction for HPLC analysis. The boiling method would be a practical choice when leaves are not too thick. However, it may convert a small fraction of chlorophyll a into pheophytin a. Although extraction at sub-zero temperatures is suitable for all plant species examined in this study, this method might be complicated for a large number of samples and it requires liquid nitrogen and equipment for leaf grinding. Using DMF as an extractant is simple and suitable with Arabidopsis samples. However, this solvent cannot completely block the formation of chlorophyllide in thicker leaves.

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Section: Resultsmentioning

confidence: 99%

Simple extraction methods that prevent the artifactual conversion of chlorophyll to chlorophyllide during pigment isolation from leaf samples

Tanaka

2013

Plant Methods

101

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“…These visual symptoms resulted from the significant decrease of chlorophyll concentration in these organs since Fe is implied in the synthesis of chlorophyll and carotenoid (Morales et al, 1994;Abadia and Abadia, 1993;Pestana et al, 2001). On the other hand, several studies demonstrate that Fe deficiency stimulate the chlorophyllase, enzyme implied in the degradation of chlorophyll, in citrus (Ferrnandez-Lopez et al, 1991) and barley (Rodriguez et al, 1987).…”

Section: Discussionmentioning

confidence: 99%

Genotypic variation of N2-fixing common bean (Phaseolus vulgaris L.) in response to iron deficiency

Krouma¹,

Drevon

Abdelly³

2006

Journal of Plant Physiology

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“…In leaves of oat and barley, modulations of chlorophyllase activities that are weakly correlated with rates of Chl degradation have been reported (Sabater et al 1978, Rodriguez et al 1987. However, the mature green leaves of these species and particularly those of dicotyledons (e.g.…”

Section: Discussionmentioning

confidence: 99%

Development of an assay for Mg-dechelatase of oilseed rape cotyledons, using chlorophyllin as the substrate

Vicentini¹,

Iten²,

Matile³

1995

Physiol Plant

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Development of an assay for Mg-dechelatase of oilseed rape cotvledons. using chlorophvilin as the substrate. -Phvsiol. Plant. 94: 57-63.Chlorophyllin (Chlin). the Mg-chlorin obtained from chlorophyll (Chi) was employed as substrate of Mg-dechelatase. The release of Mg-* was associated with a shift of absorption from 644 to 687 nm. Changes of absorption at 687 nm were taken as a measure of Mg-dechelatase activity present in preparations of oilseed rape thylakoids. Absorption changes were correlated linearly with enzyme dose. The pH optimum of Mg release from Chlin was ca 9 with a broad flank down to pH 7. The reaction showed saturation kinetics with an apparenl K^, value of ca 17 nM. The activity was inhibited in the presence of cysteamine or reduced glutathion. There was no effect of the thiol reagent N-ethyl maleimide. The bulk of dechelatase activity was associated with the chloroplast membranes. The enzyme is partially latent and the appearance of full activity requires the solubilization of thylakoids with detergent. The highest activities were detected in mature green rape cotyledons. During dark-induced senescence the activity declined al roughly the same rate as Chi was lost in the leaf tissue.

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Degradation of Chlorophyll and Chlorophyllase Activity in Senescing Barley Leaves

Cited by 50 publications

References 15 publications

Simple extraction methods that prevent the artifactual conversion of chlorophyll to chlorophyllide during pigment isolation from leaf samples

Simple extraction methods that prevent the artifactual conversion of chlorophyll to chlorophyllide during pigment isolation from leaf samples

Genotypic variation of N2-fixing common bean (Phaseolus vulgaris L.) in response to iron deficiency

Development of an assay for Mg-dechelatase of oilseed rape cotyledons, using chlorophyllin as the substrate

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