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Two-phase flavonoid formation in developing strawberry (Fragaria x ananassa) fruit.

Flavonoids are important secondary metabolites in strawberry as they fulfill a wide variety of physiological functions. In addition, they are beneficial for human health. Previous studies have shown for selected enzymes from the flavonoid pathway that flavonoid biosynthesis shows two peaks during fruit development. We provide optimized protocols for the determination of the activities of the key flavonoid enzymes: phenylalanine ammonia lyase, chalcone synthase/chalcone isomerase, flavanone 3-hydroxylase, dihydroflavonol 4-reductase, flavonol synthase, flavonoid 3-O-glucosyltransferase, and flavonoid 7-O-glucosyltransferase. Using these protocols we were able to demonstrate two distinct activity peaks during fruit ripening at early and late developmental stages for all enzymes with the exception of flavonol synthase. The first activity peak corresponds to the formation of flavanols, while the second peak is clearly related to anthocyanin and flavonol accumulation. The results indicate that flavonoid 3-O-glucosyltransferase activity is not essential for redirection from flavanol to anthocyanin formation in strawberry.

Acyltransferases↗

Isolation of cytochrome P450 inhibitors from strawberry fruit, Fragaria ananassa.

A new glycoside, 2-beta-d-glucopyranosyloxy-4,6-dihydroxyisovalerophenone (3), was isolated from strawberry fruit along with kaempferol-3-beta-D-(6-O-trans-p-coumaroyl)glucopyranoside (1) and kaempferol-3-beta-D-(6-O-cis-p-coumaroyl)glucopyranoside (2). Compounds 1 and 2 inhibited activity of a drug-metabolizing enzyme, CYP3A4.

Cytochrome P-450 CYP3A↗

Effects of cultivar, fruit number and reflected photosynthetically active radiation on Fragaria x ananassa productivity and fruit ellagic acid and ascorbic acid concentrations.

BACKGROUND AND AIMS: A number of strawberry varieties were surveyed for their total ellagic acid concentration, and attempts were made to determine if ellagic acid and ascorbic acid concentrations of two strawberry cultivars could be increased by polythene reflective mulches. METHODS: After adjusting crop yields and cultivation using polythene mulches with two different PAR reflective capacities, field- and polytunnel-grown strawberries were analysed for ellagic acid and ascorbic acid concentrations by HPLC. Comparative measurements of yield and fruit quality were determined along with plant developmental changes. KEY RESULTS: Ellagic acid concentration varied widely with strawberry cultivar (60-341 microg g(-1) frozen weight), as did the ratio of conjugated ellagic acid : free ellagic acid. Also, there was significant year-to-year variation in total ellagic acid concentration with some cultivars. Mulches with different reflective capacities impacted on strawberry production; highly reflective mulches significantly increased growth and yield, the latter due to increases in fruit size and number. CONCLUSIONS: Highly reflective mulches significantly increased total concentrations of ellagic acid and ascorbic acid relative to control in fruit of different cultivars. The potential of agronomic practices to enhance the concentration and amounts of these important dietary bioactive compounds is discussed.

Ascorbic Acid↗

Movement of entomopathogenic nematodes in soils of Fragaria spp.

The mobility of three entomopathogenic nematodes (Steinernema carpocapsae UK and All Strain, and S. glaseri (Nematoda: Steinernematidae) was observed in strawberry fields in Vancouver, WA. Nematodes were sprinkled over the soil surface using a mason jar. Soil and soil-moisture content, soil composition, soil pH and conductivity was reported over the evaluation period. 12 cm deep soil samples were taken after surface application at 9 and 25 days, 7, 26 and 45 and 6, 22, 37 and 50 days at the three different test sites within the surface application site and at 5 cm distance. Soil samples taken were divided into three layers (0-4, 5-8 and 9-12 cm) and exposed once to five Galleria larvae in the laboratory to evaluate nematode presence and movement. Results after surface application demonstrated more vertical movement of S. glaseri into deeper soil layers in comparison to both S. carpocapsae strains.

Animals↗

Isolation and purification of a 316 Da preformed compound from strawberry (Fragaria ananassa) leaves active against plant pathogens.

An antibiotic called fragarin showing activities against bacterial and fungal plant pathogens was isolated and purified by FPLC chromatography from the soluble fraction of strawberry leaves. The molecular weight value determined by mass spectrometry is 316 Da. Fragarin remains fully active after protease treatment or alkaline hydrolysis at 100 degrees C for 20 min. Biological and chemical analyses suggest that fragarin may be a new type of an antimicrobial preformed compound--phytoanticipin--and would constitute a primary non-specific barrier of strawberry defense.

Anti-Bacterial Agents↗

Effects of conditioned medium on activities of PAL, CHS, DAHP synthase (DS-Co and DS-Mn) and anthocyanin production in suspension cultures of Fragaria ananassa.

A conditioned medium (CM) prepared from cell suspension cultures of strawberry stimulated anthocyanin synthesis. The effect was significantly (P<0.05) greater than that of synthetic medium (SM), with macronutrient concentrations, carbohydrate concentrations and pH adjusted to those of CM. The activity of 3-deoxy-D-arabino-heptulosonate 7-phosphate (DAHP) synthase (EC 4.1.2.15) (DS-Mn, DS-Co), phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) and chalcone synthase (CHS, EC 2.3.1.74) were monitored in the CM- and SM-cultured cells. PAL and CHS activities were found to increase significantly (P<0.05) in the CM-cultured cells. CHS transcript levels were higher in the CM-cultured cells compared to transcript abundance in SM-cultured cells. There was no significant difference in the DS-Mn and DS-Co activities of cells grown in conditioned or synthetic media.

Journal Article↗

Comparative study of the products of the peroxidase-catalyzed and the polyphenoloxidase-catalyzed (+)-catechin oxidation. Their possible implications in strawberry (Fragaria x ananassa) browning reactions.

The peroxidase- and polyphenoloxidase-catalyzed oxidations of (+)-catechin yield several products showing different degrees of polymerization, which are apparently responsible for the pigment decay and the associated browning reaction that occurs in processed strawberry fruits and their derived foods. In this work, we have purified both peroxidase and polyphenoloxidase from Oso Grande cv. strawberry fruits, and comparatively analyzed the products of their enzyme-mediated (+)-catechin oxidation. The joint analysis by reversed-phase and size-exclusion HPLC of the (+)-catechin oxidation products obtained with both enzymes indicate that they were qualitatively the same: dehydrodicatechin B4, a (+)-catechin quinone methide, dehydrodicatechin A, a (+)-catechin trimer, and a (+)-catechin oligomer with polymerization degree equal to or greater than 5. The main quantitative differences between the oxidative reactions were the great amount of oligomer formed in the case of the polyphenoloxidase-mediated reaction and the low amount of (+)-catechin reacted in the case of the peroxidase-mediated reaction. One of the possible reasons for such low levels of (+)-catechin consumption in the case of the peroxidase-mediated reaction was the possible inhibition by products of the enzyme-catalyzed oxidation. In fact, the peroxidase-mediated (+)-catechin oxidation was differentially inhibited by dehydrodicatechin A, showing a competitive type inhibition and a k(I) of 6.4 microM. In light of these observations, these results suggest that brown polymer formation, estimated as oligomeric compounds resulting from (+)-catechin oxidation, in strawberries is mainly due to polyphenoloxidase, and although peroxidase also plays an important role, it is apparently auto-regulated by product (dehydrodicatechin A) inhibition.

Catechin↗

Cloning, expression and immunolocalization pattern of a cinnamyl alcohol dehydrogenase gene from strawberry (Fragaria x ananassa cv. Chandler).

Cinnamyl alcohol dehydrogenase (CAD; EC 1.1.1.195) catalyses the conversion of p-hydroxy-cinnamaldehydes to the corresponding alcohols and is considered a key enzyme in lignin biosynthesis. By a differential screening of a strawberry (Fragariax ananassa cv. Chandler) fruit specific subtractive cDNA library, a full-length clone corresponding to a cad gene was isolated (Fxacad1). Northern blot and quantitative real time PCR studies indicated that the strawberry Fxacad1 gene is expressed in fruits, runners, leaves, and flowers but not in roots. In addition, the gene presented a differential expression in fruits along the ripening process. Moreover, by screening of a strawberry genomic library a cad gene was isolated (Fxacad2). Similar to that found in other cad genes from higher plants, this strawberry cad gene is structured in five exons and four introns. Southern blot analyses suggest that, probably, a small cad gene family exists in strawberry. RT-PCR studies indicated that only the Fxacad1 gene was expressed in all the fruit ripening stages and vegetative tissues analysed. The Fxacad1 cDNA was expressed in E. coli cells and the corresponding protein was used to raise antibodies against the strawberry CAD polypeptide. The antibodies obtained were used for immunolocalization studies. The results showed that the CAD polypeptide was localized in lignifying cells of all the tissues examined (achenes, fruit receptacles, runners, leaves, pedicels, and flowers). Additionally, the cDNA was also expressed in yeast (Pichia pastoris) as an extracellular protein. The recombinant protein showed activity with the characteristic substrates of CAD enzymes from angiosperms, indicating that the gene cloned corresponds to a CAD protein.

Alcohol Oxidoreductases↗

Glucanolytic Actinomycetes Antagonistic to Phytophthora fragariae var. rubi, the Causal Agent of Raspberry Root Rot.

A collection of about 200 actinomycete strains was screened for the ability to grow on fragmented Phytophthora mycelium and to produce metabolites that inhibit Phytophthora growth. Thirteen strains were selected, and all produced (beta)-1,3-, (beta)-1,4-, and (beta)-1,6-glucanases. These enzymes could hydrolyze glucans from Phytophthora cell walls and cause lysis of Phytophthora cells. These enzymes also degraded other glucan substrates, such as cellulose, laminarin, pustulan, and yeast cell walls. Eleven strains significantly reduced the root rot index when inoculated on raspberry plantlets.

Journal Article↗