Chlorophyllase Activity Research Articles

Changes in Chlorophylls and Carotene Contents of Green and Bleached Lima Bean Seeds During Development and Maturation1

Abstract Green seeds from lima bean (Phaseolus lunatus L.) cvs. G 2 and Green Fordhook 861, contain chlorophylls a and b in a ratio of about 2:1. Chlorophyll content per seed increases during development, reaches a max about 5 weeks after pod formation, then declines sharply during maturation. Bleached seeds have less chlorophyll, especially chlorophyll a, and carotene than non-bleached seeds. In developing and mature lima bean seeds max chlorophyllase activity appears during the stage which corresponds with max chlorophyll content rather than the stage of most rapid loss of chlorophyll. The enzyme is particulate, located in the chloroplast membranes, and has an optimum pH of 8.5 ± 0.2. The low carotene content of seeds of green-seeded commercial cultivars might contribute to their sensitivity to light. Several cultivars that were obtained from other countries. (Plant Introductions) contain 10 to 20 times as much carotene as the commercial cultivars tested. These “Introductions” provide genetic material for increasing the carotene content of lima bean seeds. High carotenes would improve nutritional value of beans and might reduce or prevent bleaching.

The role of chlorophyllase and light in the decomposition of chlorophyll from marine phytoplankton

In a study of the biochemical aspects of the decomposition of chlorophyll, measurement of the activity of the enzyme chlorophyllase in marine phytoplankton was attempted in vitro by observing the conversion of chlorophyll to chlorophyllide. Our experiments indicate that in darkness and at room temperatures any enzymatic activity, if present, is immeasurable by the techniques employed. As a consequence, we have considered the photo-bleaching of chlorophyll as a possible means of decomposition. This has been identified as a photo-oxidation and the decomposition of the pigment is extremely rapid. Our experiments show that a similar photo-oxidation exists in fractured cells and degraded pigments found in the feces of marine herbivores. Although the rate of photo-bleaching is slower in vivo than in vitro, we postulate that this is a principle mechanism for pigment decomposition.

Chlorophyllase of sugar-beet leaves

The chlorophyllase of sugar-beet leaves was characterized and partly purified by gel-filtration and ion-exchange chromatography on various types of Sephadex. A rapid method was developed for determining the chlorophyllase activity of fractions from columns. Indications of more than one form of the enzyme may be explained by its tendency to associate or aggregate with itself or with other substances. Association is greater in 0·02 M citrate than in citrate containing 0·5 M NaCl. The enzyme is strongly adsorbed to negatively charged materials, which may explain its improved extractibility when the sodium chloride concentration of the medium is increased. Measurements of some properties of the enzyme are affected by insolubility of the substrate.

Ribonuclease and Chlorophyllase Activities in Senescing Leaves

AbstractThe activities of two enzymes, ribonuclease and chlorophyllase were investigated during the senescence of leaves. Ribonuclease activities were measured in primary leaves of Phaseolus vulgaris, and related to the levels of nucleic acid, protein and chlorophyll. Similarly, changes in chlorophyllase activity during senescence of leaves of Raphanus sativus were measured and related to chlorophyll. During senescence the levels of each enzyme as well as its respective substrate declined. Retardation of senescence, by excision of young tissue from intact plants or by treatment of detached leaves with cytokinins resulted in a maintainace of both the substrate and enzyme levels. It was concluded that high levels of ribonuclease and chlorophyllase activity are not linked directly with the degradation of RNA and chlorophyll during leaf senescence.

Physiology of virus-infected plants

Little information is available on the sequence of physiological changes from virus inoculation to full development of disease symptoms. In this paper, we discuss (1) activity of chlorophyllase, (2) ferrous and ferric iron changes, (3) inorganic and organic phosphorus, and (4) respiration in pigeon pea sterility mosaic infected pigeon pea plants and cassava mosaic infected cassava plants. In both healthy and diseased plants, chlorophylla andb increased with age. However, these were significantly lower in diseased plants than in healthy plants, from early stages. Chlorophyllase activity increased with age, such increase being much steeper in diseased plants. There appeared to be a progressive conversion of ferrous Fe to ferric Fe in diseased leaves. Diseased leaves at all ages had higher levels of total P. There was a greater conversion of inorganic P to the organic form as the disease progressed. Respiration increased with progress of disease up to the production of full symptoms, and dropped thereafter reaching levels lower than those in healthy leaves of corresponding age. In PSMV-affected pigeon pea leaves activity of catalase, peroxidase, ascorbic acid oxidase and cytochrome oxidase increased while polyphenol oxidase activity decreased. Mitochondrial nitrogen was much higher in diseased leaves than in healthy leaves.

Chlorophyllase Activity in Apples and Bananas during the Climacteric Phase

A COMMON denominator of the ripening process in many fruits is the loss of chlorophyll and subsequent yellowing of the fruit skin. Biale and Young1 have stated that loss of chlorophyll is largely responsible for the ground colour changes observed in ripening fruits. Chlorophyllase functions in vitro both in the attachment and removal of the phybol side chain to the porphyrin nucleus of chlorophyll and prochlorophyll moieties2–4, and is probably important to the in vivo catabolism of chlorophyll in fruits4,5.

The respiration climacteric in apple fruits.: The action of hydrolytic enzymes in peel tissue during the climacteric period in fruit detached from the tree

The respiration rate and the activity of four hydrolytic enzymes, chlorophyllase, lipase, ribonuclease, and acid phosphatase, were determined at intervals during the development of the respiration climacteric in detached apple fruits stored at 12°. The activities of lipase, ribonuclease and acid phosphatase rose to a peak at about the time the peak in respiration was reached and then decreased as the respiration fell. Acid phosphatase and ribonuclease are found in the soluble protein fraction in our preparations. This suggests that they are not bound within particles similar to the lysosomes which play an important role in the senescence of animal cells. Chlorophyllase activity rose before the commencement of the rise in respiration and continued beyond the peak. As the activity of chlorophyllase increased, the chlorophyll content of the tissue decreased. The possible relationship between chlorophyllase, lipase, the degradation of chlorophyll and the disorganization of the chloroplast lamellae which occurs during the climacteric is discussed.

Studies on sterility mosaic disease of pigeon pea

The reduction in the contents of chlorophyll in virus diseased pigeon pea leaves was observed to be as high as 60·9 per cent. Carotene and xanthophyll contents of diseased leaves also showed a decrease. The activity of chlorophyllase was increased due to virus infection. The total carbohydrate contents of diseased leaves were reduced. The synthesis of sucrose in diseased leaves was at a lower rate than in the healthy. This reduction in the ability to synthesize sucrose indicated a derangement of photosynthetic activity in diseased plants. Evidences were obtained to show that the translocation of sugars was reduced and that the nature of sugars translocated was altered in the diseased plants.

Relationship between Chlorophyll and Carotenoid Biosynthesis in Dirty-Red (Green-Flesh) Mutant in Tomato

In a tomato mutant, dirty red, both chlorophylls and carotenoids are present in the ripe fruit resulting in a dirty-red flesh color. Since it had been proposed that the phytol chain of the chlorophyll molecule may be utilized as a precursor for carotenoid during maturation, the dirty-red mutant was studied to determine the effect of chlorophyll retention on carotenoid synthesis. An inheritance study revealed that dirty red is controlled by a single recessive gene and is synonymous or allelic to the green-flesh mutant (gf). Analyses of chlorophylls a and b indicated that the action of the dirty-red gene becomes evident as the ripening process commences. The chlorophyll contents of the parental strains (Rutgers and dirty red) and the F2 red and dirty-red recombinants were almost equal at the immature green stage. Upon ripening, the dirtyred fruits contained some chlorophyll, while none was detected in the normal red fruits. The chlorophyllase activities of the red and dirty-red F2 progenies were determined. At the green stage, both segregates showed almost equal enzyme activities, indicating that, at this early stage, there was no loss or impairment of enzyme activity in the dirty-red plants. At incipient color, the dirty-red segregates showed higher chlorophyllase activity than the red F2 progeny. No enzyme activity was detected in the ripe fruits of red segregates, while the dirty-red recombinants exhibited slight chlorophyllase activity. Tests indicated that chlorophyllase from either normal or dirty-red plants could degrade chlorophyll from either source. These results suggest that impaired degradation by this enzyme is not responsible for the chlorophyll content of the ripe dirty-red fruits. The dirty-red gene also influences carotenoid synthesis and accumulation, and a relationship between the two pigment systems was demonstrated. The p3-carotene, y-carotene, lycopene, phytofluene, and phytoene contents were determined. Where the dirty-red factor was homozygous, chlorophyll was present in the ripe fruits, and the carotenoid content was lower than in normal red sibs. The differences in carotenoid contents between the two types were highly significant in the F2 recombinant classes.

Činitelé ovlivňující přesnost stanovení ehroroíylů a a b pomocí jejich rozdělení papírovou chromatografií a kolorimetrického měření eluátů.

Quantitative determination of chlorophyll a and β can be made by paper chromatography of acetone extracts of plant material with colorimetric measurement of the eluates from the separated zones.From the suitable solvent systems which give adequate separation of the pigments at a distance of 20 cm. from the start,Hager's mixture (1955) separates the chlorophylls better than the toluene-isopropanol (400: 1 v/v.) mixture, which, however, is better for the separation of carotenoids. Twice the amount of chlorophyll is separated on Whatman 31 ET paper, equally well and with the same time of development, as on Whatman No. 3 paper, on which it is possible to separate a maximum of about 15 μg of chlorophyll pigments per 1 em. start length. Losses on elution are, however, higher on using Whatman 31 ET paper. In plants with a high chlorophyllase activity, the error of determining chlorophyll a andb is greatly reduced if the leaves are placed for 1 min. in boiling water before extraction. For elution of chlorophylla andb from paper it is better to use anhydrous acetone, for chlorophyllides 80% acetone.A comparison of the procedure investigated with the method of two-wave length spectrophotometric measurement of crude acetone extracts showed that in view of the average 10% loss, the chromatographic method is hardly suitable for determining the absolute amounts of chlorophylla andb, although the relation (a/b) can be determined with similar precision by both methods. Moreover, in view of the greater amount of work involved the chromatographic method can only be recommended for confirming the results of spectrophotometrie determination. Quantitative determination of chlorophylls from the area of the spot or from the "RF" value can only be of an informative character.

Activity and Partial Purification of Chlorophyllase in Aqueous Systems

SUMMARY A procedure is described for the 500-fold purification of chloro- phyllase from etiolated rye seedlings. Evidence is presented that the final preparation is a lipoprotein-sodium deoxycholate complex. An assay procedure with an aqueous medium is presented. p-Chloromercuribenzoate and ferricyanide inhibit the enzyme, whereas iodoacetamide and diisopropyl fluorophos- phate do not, at the concentrations tested. The possible role of the enzyme in the biosynthesis and breakdown of chlorophyll and in the formation of lamellas of the chloroplast is discussed. REFERENCES 1. 2. 3. k: 6. 7. 8. 9. 10. 11. 12. 13. 14. WILLST~TTER, R., AND STOLL, A., Untersuchungen Ober Chloro- phyll, Springer-Verlag, Berlin, 1913, p. 172. KROSSING, G., Biochem. Z., 306, 359 (1940). ARDAO, C., AND VENNESLAND, B., Plant Physiol., 36, 368 (1960). MAYER, H., Planta, 11, 294 (1930). HOLDEN, M., Biochem. J., 78, 359 (1961). WEAST, C.

Chlorophyllase Activity of Spinach Chloroplastin

When spinach chloroplasts are treated with digitonin, a chlorophyll-lipoprotein complex is solubilized. Careful addition of ethanol to the clear green solution gives a green precipitate which is one of the components (factor 1) of cytochrome c photooxidase (1, 9, 10). Since factor 1 is a chlorophyll-lipoprotein complex, it may properly be called chloroplastin. The present paper is concerned with the determination of the chlorophyllase activity of this chloroplastin.