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William C. Mitchell and Gojko Jelenkovic

The NAD-dependent and NADP-dependent alcohol dehydrogenase activities of strawberries (Fragaria xananassa Duch.) were found to have broad substrate specificities including those alcohols and aldehydes responsible for strawberry aroma and flavor either directly or through their ester products. NAD-dependent activities were greatest against short-chained alcohols, whereas the NADP-dependent activities were most active against aromatic and terpene alcohols. Differences were seen in substrate specificity between receptacle and achene alcohol dehydrogenase activities. Alcohol dehydrogenase activities were found to be developmentally regulated in receptacle tissue and increased during the period of fruit maturation and ripening. Isoelectric focusing of NAD-dependent ADH activities showed that several isozymes of this enzyme exist, that they differ between receptacle and achene tissues, and that they vary among specific genotypes. Our results suggest that NAD- and NADP-dependent ADH activities are integral components of flavor and fragrance volatile production in ripening strawberries.

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Manuela Zude-Sasse, Ulrich Hartmond, Georg Ebert, and Peter Lüdders

Soil flooding reduces partial pressure of oxygen (pO2) in the root zone and often results in a reduction in photosynthesis and growth. In greenhouse studies, rooted stem cuttings of the mango (Mangifera indica L.) rootstock selection 13/1 were exposed to anoxia by saturating the root zone with N2 for up to 52 h. Reduced pO2 in the root zone affected the energy status of the roots and particularly enhanced the phosphorylated and nonphosphorylated pyridine nucleotide charges—the ratio of reduced Nicotinamide-adenine-dinucleotides [NAD(P)H] to total Nicotinamide-adenine-dinucleotide content [oxidized NAD(P)+ plus NAD(P)H]—that drive the redox reaction rates in cell metabolism. Also, the pyridine nucleotide charges in leaves were enhanced, while the photosynthetic rate decreased following reduction in pO2 in the root zone. During up to 4 h of reduced pO2, the ratio of internal CO2 concentration in the mesophyll to ambient CO2 concentration was unchanged. This implies a nonstomatal influence on photosynthesis. In addition, light saturation of photosystem II occurred at lower irradiance (470 μmol·m-2·s-1) resulting in reduced maximum photochemical efficiency below that of the high pO2 controls. After 28 h of reduced pO2, NAD(P) charges in the leaves returned to normal, diminishing its potential effect on net photosynthetic rate.

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A. Talaie and H. Arouie

To determine the effect of NAD on prevention of preharvest fruit drop of `Golden Delicious' apples, a range of studies were conducted for 2 years in the Kamalabad area in Karaj City-Iran. In this experiment NAD at 10 and 15 ppm concentrations were applied in two intervals of 2 and 3 weeks before harvesting (131 and 138 days after full bloom) and at the same time nontreated trees were used as control plants. The results indicate that applying NAD reduces the preharvest fruit drop percentage and then is followed by the yield increase. Fruit drop percentage, fruit weight, length, diameter and volume changes, amount of soluble solid material, total sugar content, total acid content, fruit juice pH, and flesh firmness were measured and analyzed in this experiment. According to the results of this experiment using NAD not only prevents the preharvest fruit drop of `Golden Delicious' apples, but any unsuitable changes on the qualitative property of the fruit are avoided and there even is quality improvements in some cases compared to control trees.

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William C. Mitchell and Gojko Jelenkovic

Assays of enzyme activity, in vivo inhibition studies and the developmental analysis of strawberry (Fragaria × ananassa Duch.) fruit alcohol dehydrogenases (ADH) suggest that both the NAD-(E.C. 1.1.1.1) and the NADP-dependent (E.C. 1.1.1.2) forms of ADH enzymes play integral roles in the development and ripening of fruits. One role of ADH enzymes appears to be the evocation of changes in sugar, soluble solids, acidity and volatile compounds necessary for the normal organoleptic character of strawberry fruits. The data presented includes: 1.) The wide substrate specificity of both ADH enzymes for the “fragrance and flavor alcohols and aldehydes” synthesized by ripe strawberry fruits, 2.) the effect of inhibitors of ADH activity upon strawberry fruit ripening, and 3.) the comparative regulation of NAD- and NADP-ADH enzymes including 4.) the developmental control of ADH enzymes in strawberry fruits.

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Hideaki Yamaguchi, Yoshinori Kanayama, Junichi Soejima, and Shohei Yamaki

Seasonal changes in the amounts of the NAD-dependent sorbitol dehydrogenase (NAD-SDH) (enzyme code, 1.1.1.14) protein in developing apple (Malus pumila Mill var. domestica Schneid) fruit were determined by immunoblotting analysis. The amounts of the enzyme protein were very low in young fruit and rose as fruit matured. The weak correlation between enzyme protein and NAD-SDH activity and also the changes in NAD-SDH specific activity suggested that there could be posttranslational modification to the pre-existing enzyme or isoenzyme(s) of NAD-SDH. The changes in the amounts of NAD-SDH protein did not show the same pattern as those in relative growth rate, which is used to express sink activity, especially in young fruit. The role of NAD-SDH on sink activity in apple fruit, therefore, could not be explained simply by the amount and activity of the enzyme. In young fruit, it seems that enzymes other than NAD-SDH would be more directly related with fruit growth.

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Motoko Iida, Nancy A. Bantog, Kunio Yamada, Katsuhiro Shiratake, and Shohei Yamaki

The regulation of NAD+-dependent sorbitol dehydrogenase (NAD-SDH, EC 1.1.1.14) by sugar was investigated by using sliced tissues of japanese pear (Pyrus serotina Nakai cv. Kousui) fruit in order to determine its role in the mechanism of sugar accumulation in fruit tissue. The results of the activities and steady-state levels of the protein and mRNA indicate that NAD-SDH in japanese pear fruit is among the sugar-inducible genes. By preincubating the sliced tissues for 16 hours in a medium without sugar, NAD-SDH activity declined and reached a stable level that was maintained for up to 40 hours. The washing procedure also reduced the sugar concentration in the apoplast and cytosol of the sliced tissues to low concentrations and enabled them to be manipulated by exogenous applications of carbohydrate solutions. Incubation of tissues in 50 or 100 mm sorbitol for 8 hours led to enhanced expression of the NAD-SDH gene as determined by increased mRNA and protein levels and enhanced enzyme activity. The presence of 100 mm glucose, sucrose, or mannitol also gave significant stimulation on the levels of activity, protein, and mRNA of NAD-SDH compared with those of control tissues bathed in media in which the osmotic potential had been adjusted to that of the sugar solutions by adding polyethylene glycol. However, fructose was ineffective in stimulating NAD-SDH activities and the level of the protein was not enhanced but the level of mRNA was increased. Therefore, it is suggested that NAD-SDH gene transcription is enhanced by each sugar investigated, and fructose appears to be unique as it also influences NAD-SDH at a post-transcriptional level.

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Riccardo Lo Bianco, Mark Rieger, and She-Jean S. Sung

Sorbitol is the major photosynthetic product in peach [Prunus persica (L.) Batsch.]. In sink tissues, sorbitol is converted to fructose via NAD+-dependent SDH. A new procedure is described that allows rapid, simple quantification of SDH activity in growing tissues. The procedure uses only 0.01 to 5 g of fresh tissue per sample, such that a single shoot tip, a single root tip, or ≈5 g of fruit flesh can be assayed for SDH activity. Storage of samples at 4 or -20 °C overnight resulted in significant loss of enzyme activity. Thus, freshly harvested tissues were ground with sand in buffer at 2 °C in a mortar and pestle, and the homogenate was centrifuged at 3000 g n to remove particulate matter and sand. The supernatant was desalted on a Sephadex G-25 column, and the eluent was assayed for SDH activity immediately. Activity was determined by measuring the production of NADH per minute in the assay mixture using a spectrophotometer (340 nm). Tris buffer at pH 9.0 was the best for extraction of peach SDH. Activity of SDH was strongly inhibited by dithiothreitol (DTT) in the extraction mixture and by DTT, L-cysteine, or SDI-158 in the assay mixture, similar to results reported for SDH from mammalian tissues. Peach SDH has a Km of 37.7 mm for sorbitol and a pH optimum of 9.5, similar to those reported for apple (Malus × domestica Borkh.) SDH. Unlike older protocols for SDH activity in plant tissues, the new procedure features reduced sample size (1/10 to 1/100 of that which was previously used), smaller volumes of buffer, fewer buffer ingredients, greatly reduced time for sample preparation, yet comparable or higher values of SDH specific activity. Following the same procedure, SDH activity was also measured in Prunus fremontii Wats., Prunus ilicifolia (Nutt.) Walp., and Marianna 2624 plum (P. cerasifera Ehrh. × P. munsoniana Wight & Hedr.).

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E.W. Stover, P.J. Stoffella, S.A. Garrison, D.I. Leskovar, D.C. Sanders, and C.S. Vavrina

A commercial mixture of 1-naphthaleneacetamide and 1-naphthaleneacetic acid (Amcotone) was applied to tomato (Lycopersicon esculentum Mill.) and pepper (Capsicum annuum L.) at various timings from early bloom through early fruit development to evaluate effects on fruit size and both early and total marketable yield. Amcotone was applied at rates from 10 to 40 mg·L-1, at three sites for each of the species studied. Measured yield response variables in tomato did not differ between the control and Amcotone treatments, regardless of location. Amcotone treatments did not affect yields or fruit size for pepper at the New Jersey or Texas sites. However, at Ft. Pierce, Fla., early marketable yield of pepper was increased in plots receiving three Amcotone applications at 10 mg·L-1, but total marketable yield was significantly reduced in all plots receiving more than two Amcotone sprays, and mean fruit weight was reduced by all Amcotone treatments. Early and total marketable yield of pepper at Ft. Pierce were markedly reduced in plots receiving four applications of 40 mg·L-1, which was a high rate used to assess potential phytotoxicity. While minimal benefit from auxin application was observed in this study, earlier studies suggest that these results may have been influenced by favorable environmental conditions for fruit development or negative effects on unopened flowers during all Amcotone spray applications.

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Dongfeng Liu, Junbei Ni, Ruiyuan Wu, and Yuanwen Teng

., 1999 ; Yamaki, 1986 ). Activity of NAD + -dependent sorbitol dehydrogenase (NAD + -SDH) is positively correlated with sink strength throughout peach ( Prunus persica L.) fruit development, whereas sorbitol oxidase (SOX) activity is not correlated with

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Thomas M. Kon, James R. Schupp, Keith S. Yoder, Leon D. Combs, and Melanie A. Schupp

consistent ( Byers, 1997 ). Because the proposed mode of action of ET is similar to ATS, model-based application timing may be of benefit. Naphthaleneacetamide is a hormonal thinner developed in the 1930s. NAD was developed as a milder analog of 1