Characterization of lycopene hydrocolloidal structure induced by tomato processing.

Tomato juice and paste are special type of dispersions, composed of suspended particles (pulp) dispersed in a colloidal liquid medium (serum). The bright red appearance of soluble solid separated by high speed centrifugation denoted the presence of lycopene in this fraction. Since lycopene is a hydrophobic compound it is not expected to appear in the water soluble fraction. HPLC analysis indicated presence of substantial amount of lycopene in soluble fraction which was confirmed by the appearance of lycopene crystals when observed under Transmission Electron Microscope (TEM). Considerable amount of pectin in the soluble fraction led to hypothesis that pectin facilitated the formation of hydrocolloidal system of suspended lycopene during processing. Enzyme treatment confirmed this hypothesis when pectinase effectively disrupted colloidal system and precipitated lycopene. Necessity of the divalent ions to retain the suspension signified the electrostatic interactions in the matrix surrounding lycopene crystals.

Molecular Mechanisms and Pathways as Targets for Cancer Prevention and Progression with Dietary Compounds.

A unique feature of bioactive food ingredients is their broad antioxidant function. Antioxidants having a wide spectrum of chemical structure and activity beyond basic nutrition; display different health benefits by the prevention and progression of chronic diseases. Functional food components are capable of enhancing the natural antioxidant defense system by scavenging reactive oxygen and nitrogen species, protecting and repairing DNA damage, as well as modulating the signal transduction pathways and gene expression. Major pathways affected by bioactive food ingredients include the pro-inflammatory pathways regulated by nuclear factor kappa B (NF-κB), as well as those associated with cytokines and chemokines. The present review summarizes the importance of plant bioactives and their roles in the regulation of inflammatory pathways. Bioactives influence several physiological processes such as gene expression, cell cycle regulation, cell proliferation, cell migration, etc., resulting in cancer prevention. Cancer initiation is associated with changes in metabolic pathways such as glucose metabolism, and the effect of bioactives in normalizing this process has been provided. Initiation and progression of inflammatory bowel diseases (IBD) which increase the chances of developing of colorectal cancers can be downregulated by plant bioactives. Several aspects of the potential roles of microRNAs and epigenetic modifications in the development of cancers have also been presented.

Complex formation of blueberry (Vaccinium angustifolium) anthocyanins during freeze-drying and its influence on their biological activity.

Biological activity of polyphenols is influenced by their uptake and is highly influenced by their interactions with the food matrix. This study evaluated the complex formation of blueberry polyphenols with fruit matrixes such as pectin and cellulose and their effect on the biological and antiproliferative properties of human colon cell lines HT-29 and CRL 1790. Free or complexed polyphenols were isolated by dialyzing aqueous or methanolic blueberry homogenates. Seven phenolic compounds and thirteen anthocyanins were identified in blueberry extracts. Blueberry extracts showed varying degrees of antioxidant and antiproliferative activities, as well as α-glucosidase activity. Fruit matrix containing cellulose and pectin, or purified polygalacturonic acid and cellulose, did not retain polyphenols and showed very low antioxidant or antiproliferative activities. These findings suggest that interactions between polyphenols and the food matrix may be more complex than a simple association and may play an important role in the bioefficacy of blueberry polyphenols.

Biochemical basis for functional ingredient design from fruits.

Functional food ingredients (nutraceuticals) in fruits range from small molecular components, such as the secondary plant products, to macromolecular entities, e.g., pectin and cellulose, that provide several health benefits. In fruits, the most visible functional ingredients are the color components anthocyanins and carotenoids. In addition, several other secondary plant products, including terpenes, show health beneficial activities. A common feature of several functional ingredients is their antioxidant function. For example, reactive oxygen species (ROS) can be oxidized and stabilized by flavonoid components, and the flavonoid radical can undergo electron rearrangement stabilizing the flavonoid radical. Compounds that possess an orthodihydroxy or quinone structure can interact with cellular proteins in the Keap1/Nrf2/ARE pathway to activate the gene transcription of antioxidant enzymes. Carotenoids and flavonoids can also exert their action by modulating the signal transduction and gene expression within the cell. Recent results suggest that these activities are primarily responsible for the health benefits associated with the consumption of fruits and vegetables.

Microarray analysis of ripening-regulated gene expression and its modulation by 1-MCP and hexanal.

Hexanal, an inhibitor of phospholipase D, has been successfully applied for the pre- and post-harvest treatment of fruits, vegetables and flowers. Changes in gene expression induced by hexanal and the ethylene antagonist 1-MCP, were analyzed by microarray using TOM2 tomato oligo-array containing approximately 12 000 unigenes. Mature green tomato fruits were treated with 1-MCP and hexanal, RNA isolated after 10 days of storage, and labeled cDNA synthesized for microarray analysis. A large variation in gene expression profile was observed in 1-MCP-treated fruits. Genes for ethylene biosynthetic pathway enzymes such as ACC- synthase/oxidase, ethylene receptor and ethylene response factors were heavily down-regulated in 1-MCP-treated fruits. In addition, genes for key enzymes involved in cell wall degradation and carotenoid development pathways were down-regulated. Hexanal treatment significantly down-regulated ACC-synthase, and to a lesser extent, other components of ethylene signal transduction. By contrast to MCP-treated fruits, hexanal-treated fruits gradually ripened and showed higher levels of lycopene and ß-carotene. GC-MS analysis of volatiles showed a higher level of major volatile components in hexanal-treated fruits. Similarities in the modulation of gene expression by hexanal and 1-MCP suggest that hexanal, in addition to being a PLD inhibitor, may also act as a weak ethylene inhibitor.

Cloning, expression and functional characterization of the C2 domain from tomato phospholipase Dα.

C2 domains exist as highly conserved N-terminal or C-terminal calcium- and lipid-binding motifs comprising nearly 130 amino acids, responsible for recruiting proteins to the membrane during signal transduction. In this study, the sequence corresponding to the N-terminal 164 amino acids of a full length cDNA of phospholipase Dα from tomato fruit was cloned in pET28(b) vector and expressed in E. coli as a His-tagged protein. Recombinant C2 domain showed micromolar affinity towards Ca(++) with a maximum of 2 high affinity binding sites. Interaction of C2 domain with synthetic unilamellar vesicles, evaluated by protein- lipid fluorescence resonance energy transfer, showed maximum affinity towards phosphatidic acid, and virtually no binding with phosphatidylcholine. The binding towards phosphoinositides was reduced with increasing degree of phosphorylation. Acid- and chaotropic salt- titrations indicated an electrostatic, rather than a hydrophobic mode of interaction between C2 domain and the phospholipid vesicles. Conformational analyses of the recombinant C2 domain showed a much longer calcium binding loop region, a far less electropositive phosphoinositide-binding region, unique calcium binding pockets with high electro-negativity, and other features that are distinct from the typical C2 domains of phospholipase A2 and Protein kinase C α, signifying the uniqueness of Phospholipase Dα in fruit developmental events.

Grape and wine polyphenols down-regulate the expression of signal transduction genes and inhibit the growth of estrogen receptor-negative MDA-MB231 tumors in nu/nu mouse xenografts.

The antitumor properties of the Merlot grape (and Merlot wine) polyphenols were evaluated in relation to their ability to modulate gene expression in developing tumors using an athymic nude mouse model transplanted with the estrogen receptor-negative MDA-MB231 cells. Groups of mice were fed a modified AIN 93G diet (Research Diets Inc, New Brunswick, NJ) with the experimental groups receiving 100 mg/kg body weight equivalent of polyphenols by gavage 3 times per week. After 1 week of acclimation and another week of polyphenol supplementation, MDA-MB231 cells were transplanted and the growth patterns of the tumors monitored. After 33 days of tumor growth, the animals were euthanized, the tumors isolated, and gene expression profiles analyzed using signal transduction and cell cycle arrays. The development of tumors was almost totally arrested in grape polyphenol-treated mice. Total polyphenols isolated from the wine were more effective in reducing tumor growth as compared with a hydrophobic polyphenol fraction isolated from the wine, showing a 50% and 60% reduction in tumor growth on day 33, respectively. Analysis of gene expression showed that genes such as CDK2, FAS, LEF1, PRKCE, and PTGS2, belonging to the NFkappaB, phospholipase C, and calcium signaling pathways, were down-regulated in tumors that developed in grape polyphenol-treated mice. Several genes related to cell cycle regulation, such as CDK5RAP1, RBBP8, and SERTAD1, were up-regulated in these tumors. Changes in the expression of these genes were less pronounced in tumors of wine polyphenol-treated mice. The study highlights the potential influences of dietary polyphenolic components on gene expression in estrogen receptor-negative tumors and its relation to inhibition of tumor growth.

Physico-chemical characteristics of nanovesicle-carbohydrate complexes in grape juice concentrate.

It is generally assumed that polyphenols, such as anthocyanins in fruit juice, exist in a free soluble state and are readily available for absorption in the gastro-intestinal tract. In the present study, we have investigated the interaction of polyphenols with soluble carbohydrate polymers, such as pectin and lipid nanovesicles, that are generated during homogenization of the fruit tissue during juice extraction. A commercially available grape juice concentrate contained nearly 25% of polyphenol fraction bound to macromolecules that were nondialyzable. Treatment of dialyzed juice with cellulase, pectinase, and beta-galactosidase did not cause the release of bound polyphenols; however, treatment with triton X-100 caused an increased release of bound polyphenols. The dialyzate contained relatively more -3-O glucosides and -3-O-acetoyl glucosides in comparison to the bound fraction which was enriched in -3-O-coumaroyl glucosides, suggesting qualitative differences in the bound and the free anthocyanin composition. Electron microscopic analysis of the juice fractions revealed the presence of electron-dense nanovesicle-fiber complexes ranging from 10 to 200 nm in diameter. Such complexes were absent in the dialyzate fraction. Cellulase treatment did not change the morphology of the complexes; however, treatment with pectinase and beta-galactosidase disrupted the complexes, releasing vesicular structures, suggestive of the pectin nature of the fibrous matrix. The dialyzed and the dialyzate fractions also showed differences in their 1H NMR and fluorescence spectral characteristics. The dialyzed fraction containing polyphenol-pectin complexes showed no superoxide scavenging capacity, reduced hydroxyl radical scavenging activity, and high 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity, indicating potential changes in functionality because of the complex formation.

Effects of phosphorus fertilizer supplementation on processing quality and functional food ingredients in tomato.

Even though several types of phosphorus fertilizers are used in crop production, the influence of phosphorus on produce quality is not well understood. Several quality attributes of tomato juice were analyzed in relation to phosphorus supplementation during a three-year field study (2000-2002). In addition to the recommended phosphorus fertilization, phosphorus supplementations, either through soil (low and high) or through foliar spray (hydrophos, seniphos), were tested. In general, soil and foliar phosphorus supplementation did not provide a statistically significant increase in yield. Tomato juice was evaluated for various quality characteristics including pH, titratable acidity, precipitate weight ratio, total solids, serum viscosity, Brookfield viscosity, color, lycopene levels, vitamin C, and flavor volatiles. Changes observed in several quality parameters were marginal, statistically insignificant and influenced by the season. Therefore, it appears that phosphorus supplementation may not significantly affect the processing quality parameters in tomato fruits.

Effects of phosphorus fertilizer supplementation on antioxidant enzyme activities in tomato fruits.

The effects of soil and foliar phosphorus supplementation on the activities and levels of superoxide dismutase (SOD), guaiacol peroxidase (POX), and ascorbate peroxidase (APX) in tomato fruits were evaluated by determining enzyme activities and isoenzyme analysis. Both protein levels and enzyme activities varied depending on the variety and season. In general, phosphorus supplementation did not alter SOD, POX, and APX activities significantly;however, some treatments showed season- and stage-specific enhancement in activities as noticed with hydrophos and seniphos supplementation. Three different SOD isozymes were observed, and these isozymes showed very similar staining intensities in response to P application and during the three developmental stages studied. Two major isozymes of POX and two different APX isozymes were observed at all the developmental stages. The results suggest that antioxidant enzyme activities may be influenced by the availability of phosphorus, but are subject to considerable variation depending on the developmental stage and the season.

Selective cytotoxicity of a red grape wine flavonoid fraction against MCF-7 cells.

Red wine is a rich source of polyphenolic components such as anthocyanins and flavonoids. The inhibitory effects of red wine polyphenolics on human breast cancer cells have been demonstrated earlier, but their effects on normal cells have not been fully established. Red wine (Merlot) was fractionated by hydrophobic interaction chromatography and different flavonoid fractions with increasing hydrophobicity were obtained. These fractions were tested for their inhibitory effect on human breast cancer cells (MCF-7), normal human mammary epithelial cells (HMEC), and a non-tumorigenic MCF-10A cell line. By contrast to the authentic flavonoids such as quercetin, naringenin and catechin which inhibited the growth of HMEC much more than that of MCF-7 cancer cells, a red wine fraction, that was comprised mainly of the flavonoid aglycones, showed maximal inhibition of the growth of breast cancer cells, with relatively low cytotoxicity towards HMEC and MCF-10A cells. In the presence of this flavonoid fraction, the normal cells grew normally, whereas the breast cancer cells underwent a change in morphology into spherical forms. Cytotoxicity analyses suggested that these cells had become apoptotic. The efficiency of inhibition of cell proliferation by various flavonoid fractions appeared to be related to their inhibition of calcium and calmodulin-promoted phosphodiesterase activity, suggesting that flavonoids may interfere with calcium second messenger function. The results suggest that certain grape wine ingredients have anticancer properties and these ingredients may be helpful for developing designer functional foods with cancer-preventive properties.