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M.L. Fishman, B. Levaj, D. Gillespie, and R. Scorza

Abbreviations. ASP, alkaline-soluble pectin; CSP, chelate-soluble pectin; HPSEC, high performance size exclusion chromatography; IV, intrinsic viscosity; M, molecular weight; MF, melting flesh; NMF, nonmelting flesh, R g , radius of gyration; WAF

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Yong-Soo Hwang, D.J. Huber, and L.G. Albrigo

Cell wall composition and structure were examined in visually normal (N), granulated (G), and collapsed (VC) juice vesicles of `Marsh Seedless' grapefruit (Citrus paradisi) Macf.). According to gel-filtration data, VC appeared to be associated with a modification of water-soluble (WSP) and chelate-soluble (CSP) pectin molecular weight (Mr); small-Mr pectins increased, whereas large-J4. pectins decreased. The difference in M = of pectins did not appear to be mediated by polygalacturonases. Molecular weight of hemicelluloses did not differ. Granulated vesicles contained about two times more structural polysaccharides (pectins, hemicelhdose, and cellulose) than N vesicles, although hemicellulose and pectin M = modification were absent. Ion-exchange profiles of WSP, CSP, and hemicelhrlose fractions of VC and G vesicles were not different from those of N vesicles. Individual cells in vesicles with G and these vesicles themselves were much larger than those of N vesicles, whereas cells in VC were partially or completely collapsed.

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Eunice Melotto, L. Carl Greve, and John M. Labavitch

Acid hydrolysis-generated pectic oligomers have been shown to affect ripening of tomato fruit by inducing both acceleration of reddening and increased ethylene biosynthesis (Campbell & Labavitch, 1991 Plant Physiol 97:706-713). In the present work, homogeneous size classes of these oligomers were demonstrated to have different impacts on ethylene production of tomato fruit pericarp discs. Endogenous oligomeric material of the same size classes was isolated from ripening tomato tissues and also tested for biological activity. They promoted some aspects of ripening as shown by increased ACC and ethylene production, which suggests that pectic oligomers are potential regulators of the ripening process in tomatoes. A metabolic origin for these oligomers is suggested by the fact that they are produced by in vitro polygalacturonase I treatment of polygalacturonic acid or tomato pectin.

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Jong-Pil Chun, Jae-Chang Lee, and Yong-Soo Hwang

Pectins isolated from three cultivars with different maturity were compared to find a potential role of pectin modification on the fruit softening during fruit development and ripening. There was an increase of total pectins in developing fruit and no significant decrease of pectins was confirmed even after storage in `Tusgaru' (30 days) and `Fuji' (120 days), whereas soluble pectins, except NaOH-soluble ones, gradually increased in all cultivars. Gel-filtration profile and ion exchange chromatographic evidence of soluble pectins revealed that pectin degradation in apple fruit may not be associated with softening. However, a degree of esterification probably has an important role on softening of fruits. Further results will be discussed in the presentation.

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Michael Wisniewski, Glen Davis, and Katherine Bowers

Our previous research has indicated that the pit membrane regulates deep supercooling of xylem parenchyma in woody plants. This area of the cell wall is composed of three layers that may be rich in pectins. Since pectins may define the porosity of the cell wall they may also regulate deep supercooling. The present study examined pectin distribution in ray cells using monoclonal antibodies, that recognize un-esterified (JIM5) and methyl-esterified (JIM7) epitopes of pectin, in conjuction with immunogold electron microscopy. Antibodies were obtained courtesy of J. Paul Knox, John Innes Inst., U.K. Dormant and non-dormant tissues of Prunus persica, Cornus florida and Salix babylonica were utilized. Labelling with JIM7 revealed that methyl-esterified pectins were abundant and evenly distributed within the primary cell wall and amorphous layer. Labelling with JIM5 revealed that un-esterified pectins were located specifically within the pit membrane, in the outer region of the primary cell wall. No differences were observed between species, however, preliminary data indicated that JIM5 labelling was greater in dormant than in non-dormant tissues.

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Supreetha Hegde and Niels O. Maness

Softening to a normal melting flesh texture in peaches involves a combined participation between polymers located in the middle lamella and primary cell wall. Pectins located in the primary cell wall polysaccharide matrix which cosolubilize when hemicellulose is extracted with KOH have received less attention than the chelator or sodium carbonate soluble pectin likely to be associated with the middle lamella. We conducted a series of extractions for cell walls prepared from softening peach fruit (47, 30, and 15 N firmness) using 0.5 m imidazole, sodium carbonate and a graded series of KOH. Hemicellulose-associated pectin was a substantial proportion of most KOH extracts (30 to 50 mole percent) and fractionated on size exclusion chromatography as a high apparent molecular weight peak which became more prominent as fruit softened and could be separated from two lower apparent molecular weight peaks by anion exchange chromatography. The nature of a hemicellulose-pectin interaction in peach was apparently by physical entrapment, versus covalent cross-linking. Softening related changes in hemicellulose-associated pectin will be addressed.

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Denise M. Tieman, Kurt D. Kausch, Delila M. Serra, and Avtar K. Handa

Transgenic `Rutgers' 37-81^ tomato (Lycopersicon esculentum Mill.) homozygous for a pectin methylesterase (PME) antisense gene, which lowers PME activity and increases levels of soluble solids, was compared to azygous (a segregating line of 37-81^ with 0 copies of the introduced gene) and wild-type `Rutgers' in the field during Summer 1992 and 1993 to determine the effects of the introduced PME antisense gene on tomato plant growth, fruit set, fruit yield, and fruit processing attributes. Fresh and dry weight accumulation in transgenic plants was similar to wild-type `Rutgers' and azygous 37-81^ lines during 1992 and 1993, indicating that the introduced PME antisense gene did not affect biomass accumulation. Transgenic plants showed an increase in fruit number and yield in 1992 compared to wild-type `Rutgers' and azygous 37-81^, but no differences were observed among the three genotypes in 1993. Average fruit weight did not show significant differences among the three genotypes in 1992, but was lower in azygous and transgenic plants than wild-type plants in 1993. Transgenic fruit had higher soluble and total solids and higher pH than control fruit, but shelf life was somewhat shorter in transgenic fruit. Overall, these data indicate that introduction of the PME antisense gene, which improves the processing quality of tomatoes, does not adversely affect fruit yield or vegetative growth of plants.

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Supreetha Hegde and Niels Maness

The mechanism of softening was studied in a rapidly softening peach cultivar `Belle of Georgia' by assessing changes in pectins and hemicellulose from enzymically inactive cell walls. Cell wall preparations were sequentially extracted with imidazole and sodium carbonate (pectin extracts), and potassium hydroxide (hemicellulose extracts). The pectin extracts were particularly enriched in galacturonic acid, arabiiose and rhamnose, and contained only small amounts of hemicellulose associated sugars. Hemicellulose extracts were enriched in xylose, glucose, mannose, and fucose. More tightly bound hemicellulose fractions contained considerable amounts of pectin associated sugars. The proportion of pectin associated sugars in hemicellulose extracts was greater for cell wall extracts of softened fruit. Some possible relationships between pectin/hemicellulose solubility and fruit softening will be presented. Work was supported by USDA grant 90-34150-5022 and the Oklahoma Agricultural Experiment Station.

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Y. Liu, B.S. Patil, H. Ahmad, and D.T. Gardiner

Pectin is a class of complex polysaccharides that function as hydrating agents and cementing materials for the cellulose network. Pectin has various health benefits, such as decreasing serum cholesterol levels, alleviating diabetes mellitus, and preventing cancer. It has been reported that the cancer prevention effect is closely related to the structure of pectin (galactose-rich, molecular weight <10,000, and methylation degree 50% to 70%). This study was conducted to investigate the variation of grapefruit pectin content due to harvest time. `Rio Red' grapefruit on sour orange rootstock grown at Texas A&M Univ.-Kingsville Citrus Center were harvested every 2 months and analyzed for pectin content, galacturonic acid concentration, methylation degree, and neutral sugar composition. Results showed that lamella contains more pectin than flavedo and albedo. In the lamella, the edible section, the uronic acid content ranged from 85% to 90% from August to April the following year. Methylation degree increased from August (31.89%) to April (46.99%). Total neutral sugar content of lamella pectin decreased from 110.54 to 61.77% mg·g -1. Galactose, arabinose, and rhamnose are the major sugar contents of pectin (85%), and glucose content increased with the season from 3.14 to 13.34 mg·g-1. Molecular weight of pectin was also determined.

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Domingos P. F. Almeida and Donald J. Huber

Pectin solubility in ripening tomato fruit is typically studied in vitro, employing isolated cell walls; however, it is unknown whether in vitro studies address the actual changes in the status of pectins in the fruit in situ. In vivo pectin solubilization was examined in a pressure-extracted apoplastic fluid obtained from ripening and chill-injured tomato fruit with down-regulated polygalacturonase (PG) activity and untransformed wild-type. Pectin levels in apoplastic fluid increased 3-fold during ripening and were not affected by PG levels. In contrast, PG strongly affected pectin levels in bulk, enzymically active pericarp fluid. There was a 14-fold increase in bulk pectin levels during ripening of PG-antisense fruit and a 36-fold increase in wild-type fruit. Pectin levels in the apoplastic fluid of fruit stored at 5 °C for 14 days were 40% lower than that of freshly harvested mature-green fruit, but increased significantly upon transfer of fruit to 15 °C. Monomeric galactose in the apoplastic fluid increased from 41 mg·mL–1 at the mature-green stage to 67 mg·mL–1 in ripe fruit. Bulk levels of galactose were 3- to 4-fold higher than apoplastic levels. After low-temperature storage galactose levels were 50% and 20% lower than in freshly harvested fruit for the bulk and apoplastic fluids, respectively. These results indicate that in vivo pectin solubilization is restricted and largely independent of PG. Low-temperature storage reduces in vivo pectin solubilization, an effect that is reversed upon transfer of fruit to higher temperature following cold storage.