Shoots, roots, and seeds of corn (Zea mays L., cv. Michigan 500), oats (Avena sativa L., cv. Au Sable), and peas (Pisum sativum L., cv. Wando) were analyzed for their superoxide dismutase content using a photochemical assay system consisting of methionine, riboflavin, andpnitro blue tetrazolium. The enzyme is present in the shoots, roots, and seeds of the three species. On a dry weight basis, shoots contain more enzyme than roots. In seeds, the enzyme is present in both the embryo and the storage tissue. Electrophoresis indicated a total of 10 distinct forms of the enzyme. Corn contaned seven of these forms and oats three. Peas contained one of the corn and two of the oat enzymes. Nine of the enzyme activities were eliminated with cyanide treatment suggesting that they may be cupro-zinc enzymes, whereas one was cyanideresistant and may be a manganese enzyme. Some of the leaf superoxide dismotases were found prmarily in mitochondrin or chloroplasts. Peroxidases at high concentrations interfere with the assay. In test tube assays of crude extracts from seedlings, the interference was neglgible. On gels, however, peroxidases may account for two of the 10 superoxide dismutase forms.Superoxide dismutases are metalloproteins catalyzing the dismutation of the superoxide free radical (02--) to molecular oxygen and H202. This enzymic activity was first described by McCord and Fridovich (17) with a cupro-zinc protein (erythrocuprein) from bovine erythrocytes. Similar cupro-zinc proteins with SOD3 activity were subsequently isolated from various eukaryotic sources (8). Manganese-containing proteins with SOD activity were later found in prokaryotes and in mitochondria of eukaryotes (8). Iron proteins from Escherichia coli (10) and algae (15) were recently shown to possess SOD activity.Cupro-zinc SOD has already been isolated from tissues of higher plants: pea seeds (19), spinach leaves (2, 15), and wheat germ (4). Isozymes have also been reported for wheat and spinach cupro-zinc enzyme (4,15). A cyanide-resistant enzyme from wheat germ has been described as manganese-SOD (4). Occurrence of manganese enzyme in spinach chloroplasts has been reported (15), but is not certain (7). There is no report of iron enzyme in higher plants.A comparative study of the SOD distribution in plants has not been conducted thus far. Distribution of the enzyme within the plant and intraspecific differences may be of interest and were the objective of the present study. Embryos and scutella were excised from seeds which had been allowed to imbibe at 25 C for 10 hr and were rinsed three times with distilled H20. Endosperm was obtained from dry corn and oat seeds from which the embryo-bearing end had been removed. Endosperm was drilled from the cut surface by using a size 60 wire bit fitted on a battery operated drill. The hulls of oat seeds were removed, unless the seeds were to be germinated.Preparation of Extracts. The tissues were thoroughly ground with a cold mortar and pestle in an ice bath, until no fibrous residue could be seen. The...

Superoxide dismutase was purified from pea (Pisum sativum L., cv. Wando) seeds and corn (Zea mays L., cv. Michigan 500) seedlings. The purified pea enzyme eluting as a single peak from gel exclusion chromatography columns contained the three electrophoreticaUy distinct bands of superoxide dismutase characterizing the crude extract. The purified corn enzyme eluted as the same peak as the pea enzyme, and contained five of the seven active bands found in the crude extract. The similar molecular weights and the cyanide sensitivities of these bands indicated that they are probably isozymes of a cupro-zinc superoxide dismutase. One of the remaining corn bands was shown to be a peroxidase.Superoxide dismutase accounted for 1.6 to 2.4% of the water-soluble protein in seedlings of corn, peas, and oats (Avena sativa L., cv. Au Sable). The superoxide dismutase activity per plant and per milligram water-soluble protein considerably increased during germination of oats and during greening and hook opening of peas.In a previous study, considerable amounts of SOD3 were found to be present in roots, shoots, seeds, and seed parts of oats, corn, and peas (4). Electrophoresis indicated multiple forms of the enzyme. Significant differences in quantity and forms of the enzyme were observed between species and between organs within a species. The objective of this study was to substantiate further the occurrence of the enzyme in higher plants and to examine the observed differences between species. For this purpose the enzyme was purified. Changes of SOD activity during seedling growth were also studied. MATERIALS AND METHODSEnzyme Purification. Unless otherwise stated, all operations were performed at 0 to 4 C. Dry pea seeds (Pisum sativum L., cv. Wando) were soaked in distilled H20 for about 15 hr. The resulting 1650 g wet weight was crushed with an electric mortar and pestle and homogenized with 1 liter of 0.1 M K2HPO4 in a Waring Blendor. The resulting pH was 7.5. After stirring, the slurry was filtered/squeezed through six layers of cheesecloth. The filtrate was centrifuged twice at 13000g for 30 min in a Sorvall RC2-B refrigerated centrifuge.The supernatant was subjected to the Tsuchihashi (chloroform-ethanol) treatment essentially as described by Weisiger and Fridovich (10). None of the pea SOD enzymes is inactivated by this treatment. The supernatant was mixed with 0.25 volume of ethanol and 0.15 volume of chloroform and stirred for 15 min. It was then clarified by centrifugation at 13000g for 15 min. Chloroform that was separated out during centrifugation was removed by suction. The supernatant was decanted, solid K2HPO4 (20 g/l) was added, and the two phases were separated after 30 min. The less dense, ethanol-rich phase was collected, chilled, and centrifuged at -15 C. Additional chloroform separating out during centrifugation was removed by suction, and the ethanolic phase was decanted.Chilled acetone (0.5 volume) was added to the ethanolic phase while stirring. The precipitate was removed by centrifugation. Addit...

Alfalfa meal and chloroform extracts of the meal have increased the growth and yield of several plant species. A crystalline substance isolated from the active fraction of alfalfa meal increased the dry weight and water uptake of rice seedlings when sprayed on the foliage or applied in nutrient culture. The substance was identified as triacontanol by mass spectrometry. Sprays containing this compound also increased the growth of corn, and barley grown in soil. Authentic triacontanol produced a similar response over a wide range of concentrations on rice grown in nutrient cultures and tomatoes grown in soil.

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Triacontanol (TRIA), a common constituent of plant waxes, was first shown in 1977 to be an active growth substance which at nanomolar concentrations increased the growth and yield of crops. TRIA is used to increase crop yields on millions of hectares, particularly in Asia. Many investigators have shown that it affects several basic metabolic processes including photosynthesis, nutrient uptake, and enzyme activity. However, the initial site of action has not been elucidated. TRIA rapidly elicits a second messenger (TRIM) in rice (Oryza sativa L.), which at nanomolar concentrations causes plants to respond in a manner similar to TRIA. TRIM has been identified as 9-0-L(+)-adenosine (9H-purin-6-amine, 9-j0-L-ribofuranosyl). During the process of isolating and identifying 9-jf-L(+)-adenosine, it was shown that this enantiomer, which previously has not been reported as occurring in nature, made up about 1% of the total adenosine pool in roots from untreated rice seedlings.TRIA' and OCTA (Fig. 1) are primary alcohols which are ubiquitous in the environment. OCTA inhibits the activity of TRIA at equimolar concentrations (16), and both compounds elicit the second messengers, OCTAM and TRIM, respectively, in rice (Oryza sativa L.) seedlings (17). TRIM has been identified as L(+)-adenosine ( Fig. 1) 'Abbreviations: TRIA, triacontanol; OCTA, octacosanol; TRIM, triacontanol second messenger; OCTAM, octacosanol second messenger; D(-)-adenosine, 9-f3-D(-)-adenosine; L(+)-adenosine, 9-3-L(+)-adenosine or 9H-purin-6-amine, 9-f-L-ribofuranosyl.

No abstract

The source of wheat (Triticum aestivum L.) seed may affect crop growth and yield. Within a genotype, seed that has a higher protein content and/or is larger will produce more vigorous seedlings and sometimes higher yields. Spring and winter wheat seed collected from five cultivar trials grown at several different locations were used to determine the relative effect of genotype and environment on seed protein and seed size, and these factors were related to the dry weight of seedling shoots (vigor). Both environment and genotype affected the protein content of the seed. Regardless of genotype or environment, seedling vigor was consistently related to seed protein. Seedling vigor was also related to seed size, but when seed size was eliminated as a factor by using uniformly sized seed, the seed protein content and vigor relationships were significant.

Triacontanol ICH.(CH2)2sCH2OHI increased growth in vitro of cell cultures of haploid tobacco (Nicotiana tabacum). The fresh weight of cell cultures of tomato (Lycopersicon esculentum), potato (Solanum tuberosum), bean (Phaseolus vulgaris), and barley (Hordeum vulgare x H. jubatum) was also increased. The increase in growth of tobacco callus seems to have been due to an increase in cell number. Another long chain alcohol, octocosanol ICH(CH2)2fiCH2OHI, did not increase the growth of tobacco cell cultures.