John Friend

The polysaccharide structure of potato cell walls: Chemical fractionation

Cell walls of potato tubers were fractionated by successive extraction with various reagents. A slightly degraded pectic fraction with 77% galacturonic acid was extracted in hot, oxalate-citrate buffer at pH 4. A further, major pectic fraction with 38% galacturonic acid was extracted in cold 0.1 M Na2CO3 with little apparent degradation. These two pectic fractions together made up 52% of the cell wall. Most of the oxalate-citrate fraction could alternatively be extracted with cold acetate-N,N′,N′-tetracetic acid (CDTA) buffer, a non-degradative extractant which nevertheless removed essentially all the calcium ions. This fraction was therefore probably held only by calcium binding, and the remainder of the pectins by covalent bonds. Electrophoresis showed that both pectic fractions contained a range of molecular types differing in composition, with a high arabinose: galactose ratio as well as much galacturonic acid in the most extractable fractions. From methylation data, the main side-chains were 1,4′-linked galactans and 1,5′-linked arabinans, with smaller quantities of covalently attached xyloglucan. Extraction with NaOH-borate removed a small hemicellulose fraction and some cellulose. The main hemicelluloses were apparently a galactoxyloglucan, a mannan or glucomannan and an arabinogalactan.

Hydrolysis of plant polysaccharides and GLC analysis of their constituent neutral sugars

Abstract The release and degradation of sugars from onion cell walls and potato cell wall polysaccharides were followed during hydrolysis with trifluoroacetic acid so that the optimum hydrolysis conditions could be determined. After 6 hr hydrolysis in 2 M acid at 100°, calculated recovery factors of different monosaccharides from potato pectic fractions ranged from 61 to 96%. Lower yields of monosaccharides were obtained from intact onion cell walls, while the yield from cellulose was less than 0.2%. A new GLC column for the separation of alditol acetates derived from cell wall sugars is described.

Cell wall polysaccharides from onions

Abstract Onion ( Allium cepa ) cell walls were fractionated by successive extraction with oxalate-citrate buffer and with alkali. The substantial oxalate-citrate extracted fraction comprised a range of pectic polysaccharides with varying proportions of neutral side-chains. Methylation analysis of the alkali extract indicated that (1,4′)-linked galactans and a substituted xyloglucan were probably major components. Onions thus resemble dicotyledonous plants more than the Gramineae in their cell wall composition.

Insoluble phenolic compounds and resistance of potato tuber disc to Phytophthora and Phoma

Abstract Tuber discs of two potato cultivars, one (Stormont Enterprise) resistant and the other (King Edward) susceptible to a complex race of Phytophthora infestans were inoculated with sporangia of the fungus or with mycelial discs of Phoma exigua . Penetration by either fungus was impeded faster in Stormont Enterprise than in King Edward; there was less penetration of both cultivars in discs inoculated 24hr after cutting. Browning of discs, measured by a reflectance method, correlated with the browning of cell walls and their ability to stain with Toluidine Blue. The levels of browning correlated with the resistance of the discs to penetration by either fungus. Treatment of the discs with amino-oxyacetic acid (AOA) before inoculation markedly inhibited browning and permitted both fungi to grow further through the tuber tissue. Amino-oxyacetic acid had little effect on the levels of the two sesquiterpenoid phytoalexins, rishitin and lubimin, in Stormont Enterprise tuber discs. These experiments therefore indicate that accumulation of phenolic compounds in cell walls is more important than phytoalexin accumulation in non-race-specific resistance of tuber discs to P. infestans and in resistance to P. exigua . The insoluble phenolic material appears to contain polymerized oxidized chlorogenic acid, esterified p -coumaric and ferulic acids and either one or both lignin and suberin.

The biosynthesis of oxylipins of linoleic and arachidonic acids by the sewage fungus Leptomitus lacteus, including the identification of 8R-Hydroxy-9Z,12Z-octadecadienoic acid.

When the sewage fungus Leptomitus lacteus was grown in liquid culture aerobically and then transferred to medium containing long-chain fatty acids, it produced a number of oxygenated fatty acids. From linoleic acid (18∶2n−6), the major metabolite produced was R-8-hydroxy-9Z,12Z-octadecadienoic acid (8R-HODE), with additional quantities of 8,11-di-HODE, 11,16-di-HODE, and 11,17-di-HODE. Other fatty acid derivatives identified included 7-HODE, 10-HODE, and 13-hydroxy-octadecamonoenoic acid. Arachidonic acid (20∶4n−6) was metabolized primarily to 18- and and 19-hydroxy-eicosatetraenoic acids (18- and 19-HETE) also as R enantiomers, along with smaller quantities of 17-HETE, 9-HETE, 14,15-dihydroxyeicosatrienoic acid and 11,12,19-trihydroxy-eicosatrienoic acid. The oxygenated products of long-chain fatty acids, in particular the biosynthesis of 8R-HODE, a compound classified as a precocious sporulation inducer, were similar to those produced by an unrelated fungal species in the Ascomycota, the take-all fungus Gaeumannomyces graminis. As in G. graminis, the biotransformation of linoleate to 8R-HODE was not significantly inhibited by exposure of the organism to CO. This indicated that the enzyme responsible for 8R-HODE biosynthesis in Leptomitus could be similar to that of G. graminis; yet we did not detect 7,8-di-HODE as a product of 18∶2n−6 metabolism as in G. graminis. CO did inhibit the biosynthesis of 14,15-di-HETE, 18-HETE, and 19-HETE in L. lacteus, which suggested the involvement of a cytochrome P450-type monooxygenase. The biosynthesis of 8R-HODE from 18∶2n−6 was found to occur in certain cell lysates, specifically in low speed (15,000×g) supernatant, following cell disruption.

The role of xanthoxin in the inhibition of pea seedling growth by red light

Abstract Intermittent periods of red light illumination inhibit the growth of dwarf pea seedlings within 1 day and stop length increase completely in 3 days. Cis,trans -xanthoxin is synthesized within the plant during the 1st day of illumination and reaches a maximum level on the 3rd day. The red light treatment also causes an increase in the levels of violaxanthin, linoleic acid and peroxidase, lipoxygenase and carotene-bleaching activities in the plant. The possible control of xanthoxin production is discussed.

Separation of macromolecular components of plant cell walls: electrophoretic methods

Abstract Qualitative quantitative and preparative electrophoretic methods of separating polymeric substances derived from plant cell walls are described. Analyt

Inhibition of phytoalexin accumulation in potato tuber discs by superoxide scavengers

Abstract Non-steroidal anti-inflammatory drugs (NSAIDS) and nordihydroguaiaretic acid (NDGA) inhibited phytoalexin accumulation and browning in potato tuber tis

The oxidation of carotenoids by mitochondria from sugar beet leaves—III : Crocin oxidation by a peroxidase system

Abstract The crocin-destroying system retained in the residue following treatment of mitochondrial preparations from sugar beet leaves with Triton-X-100 does not fall within the lipid-requiring category of catalysts which oxidize carotenoids. The system is similar to indole-acetic acid oxidase of plant tissues with respect to its peroxidase content and reaction with phenolic compounds. The residue contains an indole-acetic acid oxidase which is optimally active at the pH optimum for crocin destruction (pH 3·6); both activities are stimulated by 2,4-dichlorophenol. Catalase present in the preparation appears to have no influence on crocindestroying activity. Generation of hydrogen peroxide was not detected, although there is an absolute requirement for oxygen. The soluble fraction of the leaf contains a thermolabile factor which stimulates crocin destruction by the particulate system. Partial purification and resolution of this factor into several fractions by ion-exchange chromatography is accompanied by purification and fractionation of peroxidase activity. It is concluded that crocin is destroyed by an aerobic oxidation mediated by peroxidase, although the exact requirements for the reaction have not been established. Evidence is presented to support the claim that the particulate system participates in the loss of carotene which occurs when sugar beet leaves are damaged.

Optimisation of 3-hydroxyeicosanoid biosynthesis by the yeast Dipodascopsis uninucleata

The yeast Dipodascopsis uninucleata UOFS Y128 produces 3-hydroxy-5,8,11,14-eicosatetraenoic acid (3-HETE) and 3-hydroxy-5,8,11,14,17-eicosapentaenoic acid (3-HEPE) from arachidonic acid and eicosapentaenoic acid, respectively. The maximal yield of 3-HETE from arachidonic acid was 1.9%: this was achieved by (i) adding arachidonic acid after 46 h growth of a synchronised culture, this being the point at which sexual reproduction and the formation of asci was highest, (ii) using medium buffered to pH 7 in the absence of sucrose and (iii) a fatty acid concentration of 200 mM.