Detection of Phenylpropanoids in Citrus Leaves Produced in Response to Xanthomonas citri subsp. citri.

Citrus canker (CC), caused by the bacterial pathogen Xanthomonas citri subsp. citri, impacts citrus production in many areas of the world by reducing yields, degrading tree health, and severely blemishing the outer peels of fresh fruit. The relative susceptibility to CC among different species of Citrus varies from the highly susceptible lime (Citrus × aurantifolia), sweet orange (C. × sinensis), and grapefruit (C. × paradisi) to the much less susceptible calamondin (C. × microcarpa) and kumquat (C. japonica). This investigation compares the responses to infection with X. citri subsp. citri of these five genotypes with respect to phenylpropanoid compound profiles and relative increases or decreases of specific compounds postinoculation. In response to X. citri subsp. citri infection, all hosts possessed increased concentrations of phenylpropanoids in leaf tissue, whereas the similarly treated nonhost orange jessamine (Murraya paniculata) did not. Several of the tested genotypes exhibited notably increased production of fluorescent phenylpropanoids, including umbelliferone, herniarin, auraptene, scoparone, and others. The profiles of these compounds and their levels of production varied among the tested species yet all investigated Citrus genotypes exhibited increased concentrations of phenylpropanoids regardless of their degree of susceptibility to X. citri subsp. citri. Kumquat and calamondin, the tested genotypes least susceptible to X. citri subsp. citri, also exhibited the highest levels of the dihydrochalcone 3',5'-di-C-glucosyl phloretin, the aglycone portion of which, phloretin, is a known antibiotic, although levels of this compound were not affected by inoculation with X. citri subsp. citri.

Controlled-release of Chlorine Dioxide in a Perforated Packaging System to Extend the Storage Life and Improve the Safety of Grape Tomatoes.

A controlled-release chlorine dioxide (ClO2) pouch was developed by sealing a slurry form of ClO2 into semipermeable polymer film; the release properties of the pouch were monitored in containers with or without fruit. The pouch was affixed to the inside of a perforated clamshell containing grape tomatoes, and the effect on microbial population, firmness, and weight loss was evaluated during a 14 day storage period at 20 °C. Within 3 days, the ClO2 concentration in the clamshells reached 3.5 ppm and remained constant until day 10. Thereafter, it decreased to 2 ppm by day 14. The ClO2 pouch exhibited strong antimicrobial activity, reducing Escherichia coli populations by 3.08 log CFU/g and Alternaria alternata populations by 2.85 log CFU/g after 14 days of storage. The ClO2 treatment also reduced softening and weight loss and extended the overall shelf life of the tomatoes. Our results suggest that ClO2 treatment is useful for extending the shelf life and improving the microbial safety of tomatoes during storage without impairing their quality.

Development of delayed bitterness and effect of harvest date in stored juice from two complex citrus hybrids.

BACKGROUND: Mandarins and mandarin hybrids have excellent flavor and color attributes, making them good candidates for consumption as fresh fruit. When processed into juice, however, they are less palatable, as they develop delayed bitterness when stored for a period of time. In this study the kinetics of delayed bitterness in two citrus mandarin hybrid siblings, 'Ambersweet' and USDA 1-105-106, was explored by sensory and instrumental analyses. In addition to the bitter limonoids, other quality factors (i.e. sugars, acids, pH, soluble solids content (SSC), titratable acidity (TA) and the ratio SSC/TA) were also measured. RESULTS: The two citrus hybrid siblings had different chemical profiles, which were perceived by taste panels. USDA 1-105-106 developed delayed bitterness when the juice was stored for more than 4 h, similar to juice from 'Navel' oranges, but 'Ambersweet' did not. Bitterness in 'Ambersweet' was more affected by harvest maturity, as juice from earlier harvest had lower SSC but higher TA and bitter limonoids. CONCLUSION: Since juice of USDA 1-105-106 shows delayed bitterness when stored for more than 4 h, this cultivar is not suitable for juice processing. Our finding that siblings can differ in chemical and sensory properties emphasize the importance of post-processing storage studies before releasing cultivars for juice.

Antimicrobial activity of controlled-release chlorine dioxide gas on fresh blueberries.

The effect of chlorine dioxide (ClO2) gas on the safety and quality of blueberries was studied. In vitro studies revealed that both ClO2 gas fumigation and ClO2 direct contact in water killed food pathogen bacterium Escherichia coli and fruit decay pathogen fungus Colletotrichum acutatum. In vivo studies were conducted using noninoculated berries and berries inoculated with postharvest decay and foodborne pathogens. Berries were inoculated with either E. coli (5.2 log CFU/g) or C. acutatum (3.9 log CFU/g). Inoculated fruit were dried for 2 h at room temperature in a climate-controlled laboratory and packed in perforated commercial clamshells, with or without ClO2 pads, and stored at 10°C for up to 9 days. The effects of ClO2 on microbial populations and fruit firmness were monitored during storage. In the inoculation experiment, treatment with ClO2 reduced populations of E. coli and C. acutatum by 2.2 to 3.3 and 1.3 to 2.0 log CFU/g, respectively. For the noninoculated blueberries, the initial total aerobic bacteria count and the yeast and mold count were 4.2 and 4.1 log CFU/g, respectively. ClO2 treatment reduced total aerobic bacteria count and yeast and mold count by 1.5 to 1.8 and 1.3 to 1.7 log CFU/g, respectively. The firmness of both inoculated and noninoculated blueberries was maintained by ClO2 treatment. Thus, controlled-release ClO2 gas fumigation technology shows promise as an effective and practical antimicrobial agent in commercial clamshell packaging of blueberry and other fruits.

Effects of chitosan-essential oil coatings on safety and quality of fresh blueberries.

UNLABELLED: Chitosan coating plus different essential oils was developed and applied to fresh blueberries in order to find more natural treatments to preserve fresh fruit quality and safety during postharvest storage. Studies were 1st performed in vitro where wild-type Escherichia coli and Penicillium digitatum were grown in suitable media, and then subjected to 6 essential oils. Three compounds, carvacrol (CAR), cinnamaldehyde (CIN), and trans-cinnamaldehyde (ECIN) had high antimicrobial capacity and were selected for an in vivo study for postharvest storage of blueberries. The selected essential oils, 0.5% each, were added into a chitosan solution and coated on fresh blueberries. After storage at 5, 10, and 20 °C for various days, fruit firmness and microbial populations were evaluated. The chitosan coating substantially decreased bacteria and yeasts/molds on the fruit, and all 3 essential oils added to the antimicrobial activities. Further dosage experiments showed that the antimicrobial activity remained even when lowering CAR concentration to 0.1% and ECIN to 0.2%. Chitosan, CAR, and ECIN also maintained fruit firmness. Our results suggest that chitosan coatings containing essential oils are effective in extending the shelf life of fresh blueberries. PRACTICAL APPLICATION: Blueberries are high-value fruit with strong antioxidant capacity and other health-promoting benefits. However, microbial food safety is an increasing concern, and decay and softening limits their storability. A combination of ≥ 0.1% CAR or ≥ 0.2% ECIN with a chitosan coating effectively reduced softening of fresh berries and decay by inhibiting microbial growth.

Effect of extraction, pasteurization and cold storage on flavonoids and other secondary metabolites in fresh orange juice.

BACKGROUND: Fresh orange juice is perceived to be more wholesome than processed juice. Fresh juice may have nutrients and phytonutrients that differ from pasteurized or processed juice. RESULTS: 'Hamlin' and 'Valencia' oranges were extracted using a commercial food service juicer, pasteurized or not, resulting in fresh-commercial juice (FCJ) or pasteurized FCJ (FCPJ) for comparison with pasteurized processed juice (PPJ) in 2009, and gently hand-squeezed 'Valencia' juice (HSJ) in 2010 for nutrient and phytonutrient content. Regardless of pasteurization, FCJ/FCPJ contained 25-49% lower insoluble solids than the PPJ, while in HSJ the insoluble solids content was between that of FCJ and PPJ. The major orange juice flavonoid glycosides were twofold higher in PPJ than in FCJ/FCPJ and HSJ, indicating that the extraction and finishing process led to more peel tissue in the juice than fresh juice extraction methods. The total phenolic content (TPC) in the juices followed a similar pattern to the flavonoid glycoside content. The polymethoxylated flavones (PMFs), associated with peel oil, occurred at the highest levels in the FCJ/FCPJ and lowest in HSJ. Limonoids and alkaloids occurred at higher levels in PPJ and HSJ than in FCJ/FCPJ. CONCLUSION: The high peel oil content of FCJ/FCPJ resulted in higher PMF levels compared to PPJ and HSJ, while flavonoid glycosides, limonoids and alkaloids, which occur at high concentrations in the inner peel albedo, occurred at higher concentrations in PPJ.

Protein sequestration of lipophilic furanocoumarins in grapefruit juice.

The sequestration of grapefruit furanocoumarins by foods was investigated by characterizing the binding between these compounds and foods with contrasting protein, fat, and carbohydrate compositions. Individual grapefruit furanocoumarins exhibited contrasting affinities to foods, where the lipophilic bergamottin and several structurally related dimers bound to foods more tightly than the more polar 6',7'-dihydroxybergamottin. From the investigation of different classes of macromolecules in foods, water-soluble proteins were found to be the major constituents responsible for furanocoumarin sequestration. Studies using bovine serum albumin as a model protein demonstrated the dissociation of grapefruit furanocoumarins from the insoluble juice cloud particles and the subsequent formation of water-soluble bovine serum albumin-furanocoumarin complexes. Fluorescence binding assays further demonstrated the binding of bergamottin and 6',7'-dihydroxybergamottin to bovine serum albumin. These results demonstrate that proteins can be sequestration agents of these important dietary furanocoumarins.

Effect of extraction method on quality of orange juice: hand-squeezed, commercial-fresh squeezed and processed.

BACKGROUND: Fresh orange juice is perceived to be more wholesome than processed juice. Fresh juice may have flavor and nutrients that differ from pasteurized or processed juice. RESULTS: 'Hamlin' and 'Valencia' oranges were extracted using a commercial food service juicer, pasteurized or not, resulting in fresh-commercial juice (FCJ) or pasteurized juice (FCPJ) for comparison with pasteurized processed juice (PPJ) in 2009, and gently hand-squeezed 'Valencia' juice (HSJ) in 2010 for quality attributes. There was higher peel oil, lower pectin content, and less cloud loss in FCJ/FCPJ compared to PPJ and HSJ regardless of pasteurization. Titratable acidity was generally higher and the ratio of solids to acids lower in FCJ/FCPJ or HSJ compared to PPJ. FCJ/FCPJ had generally higher levels of most aroma volatiles than did PPJ and, overall, the highest esters and terpenes, while methanol and ethanol levels were highest in HSJ. For sensory evaluation, FCJ/FCPJ had more peel oil and PPJ more cooked flavor than other samples, while 'Valencia' HSJ was preferred over the other juices. CONCLUSION: High peel oil content and thermo-pasteurization process decreased cloud loss of orange juice. Extraction and finishing processes rather than pasteurization or oil content were major factors in influencing orange juice flavor quality.

Biotransformations of 6',7'-dihydroxybergamottin and 6',7'-epoxybergamottin by the citrus-pathogenic fungi diminish cytochrome P450 3A4 inhibitory activity.

Penicillium digitatum, as well as five other citrus pathogenic species, (Penicillium ulaiense Link, Geotrichum citri Link, Botrytis cinerea P. Micheli ex Pers., Lasiodiplodia theobromae (Pat.) Griffon & Maubl., and Phomopsis citri (teleomorph Diaporthe citri)) were observed to convert 6',7'-epoxybergamottin (1) into 6',7'-dihydroxybergamottin (2), bergaptol (3), and an opened lactone ring metabolite 6,7-furano-5-(6',7'-dihydroxy geranyloxy)-2-hydroxy-hydrocoumaric acid (4). Metabolism of 2 by these fungi also proceeded to 4. The structure of 4 was established by high resolution mass spectrometry and (1)H and (13)C NMR techniques. The inhibitory activity of 4 towards human intestinal cytochrome P450 3A4 (CYP3A4) was greatly decreased (IC(50) >172.0 µM) compared to 2 (IC(50)=0.81 µM).

Effect of 1-methylcyclopropene on tomato flavour components, shelf life and decay as influenced by harvest maturity and storage temperature.

BACKGROUND: In Florida, tomatoes are harvested green (GR), which includes mature green (MG) and immature green (IG) fruits, and stored at low temperature (13 °C), resulting in poor flavour. Flavour improvement might be achieved if fruits were harvested with some colour (to eliminate IG fruits) and/or stored at higher temperature with the ripening inhibitor 1-methylcyclopropene (1-MCP). RESULTS: 'Florida 47' tomatoes were harvested at GR (MG + IG), breaker (BR), turning (TR) and pink (PK) stages, treated (+) or not (-) with 1-MCP and stored at 13 and/or 18 °C. 1-MCP treatment resulted in a gain of up to 6 days of shelf life depending on harvest maturity and storage temperature. Storage at 18 °C rather than 13 °C resulted in an increase in internal red colour, soluble solids (SS)/titratable acidity (TA) ratio and levels of many volatiles studied. The shelf life of BR (+) MCP fruits stored at 13 °C was similar to that of MG fruits. IG fruits exhibited lowest levels of SS, TA and ten volatiles compared with BR fruits. CONCLUSION: Harvesting tomatoes with colour (BR), thereby eliminating IG fruits, and treating with 1-MCP resulted in better quality with adequate shelf life.

Surface treatments and coatings to maintain fresh-cut mango quality in storage.

BACKGROUND: Edible coatings may extend fresh-cut fruit storage by preventing moisture loss and decreasing gas exchange. This study evaluated the effect of an antibrowning dip (calcium ascorbate, citric acid and N-acetyl-L-cysteine), followed or not with carboxymethylcellulose (CMC) or carrageenan coatings on quality of fresh-cut mangoes stored at 5 °C for up to 20 days. A fourth treatment, only used in one of four experiments, consisted of chitosan. Treatments were applied on 'Tommy Atkins', 'Kent' and 'Keitt' mangoes harvested from Homestead (FL), and on imported store-bought mangoes. RESULTS: The antibrowning dips maintained the best visual quality during storage for all cultivars, as indicated by higher b*, hue and L*. The CMC coating maintained similar visual quality, but carrageenan or chitosan decreased L* and b*. The antibrowning dip containing calcium ascorbate reduced firmness loss on cut pieces of 'Keitt', 'Kent' and store-bought mangoes. The antibrowning treatment maintained higher titratable acidity for 'Kent' and 'Keitt', resulting in lower sensory sweetness. CONCLUSION: This study with repeated experiments showed that calcium ascorbate with citric acid and N-acetyl-L-cysteine maintained cut mango slices attractiveness in storage by keeping light color in both varieties. The addition of a polysaccharide coating did not consistently improve quality.

Effect of Liberibacter infection (huanglongbing or "greening" disease) of citrus on orange juice flavor quality by sensory evaluation.

UNLABELLED: Some anecdotal reports suggest that infection of citrus trees with Candidatus Liberibacter asiaticus (Las), the suspected causal agent of huanglongbing (HLB) disease, imparts off flavor to orange juice. It is of interest to the industry to know how Las infection affects juice quality with respect to cultivar, maturity, or processing method. Hamlin, Midsweet, and Valencia oranges were harvested over 2 y from trees that tested negative (Las-) or positive (Las+) for Las from different groves and included normal looking (nonsymptomatic) and symptomatic fruit (small, green, and lopsided) from Las+ trees. In the 1st year, fruit were manually juiced, while in the 2nd year, a commercial process was used. Juice from Las+ trees was compared to juice from Las- trees in difference-from-control tests, and by descriptive analysis. Results showed large variability due to tree, harvest date, and cultivar. Juice from Hamlin Las+ trees tended to be more bitter and sour than its Las- counterpart. In contrast, hand processed Valencia juice from Las+ trees was perceived to have some off-flavor and bitterness compared to control, but the following year, commercially processed Valencia juice from Las+ trees was perceived to be only slightly more sour than juice from Las- trees for the April harvest, and to be sweeter for the June harvest. When juice from individual replicates was pooled to be more representative of a commercial situation, there was no difference between Las+ and Las- juice in Valencia. Trained panel differences were noted for juice from Hamlin Las+ fruit, especially for symptomatic fruit. PRACTICAL APPLICATION: Assumptions that juice made from oranges harvested from Huanglongbing (from infection with Liberibacter sp.) affected trees is off-flavored appeared to be generally more true for Hamlin juice than for Midsweet or Valencia, especially for Hamlin juice made from symptomatic fruit. For Midsweet and Valencia, flavor differences between juice made from fruit harvested from diseased or healthy trees varied greatly between trees, season, and even processing method. Under a commercial processing situation, where juice is blended from several varieties, seasons, and multiple locations, it is expected that off-flavor will not be a major problem.

Removal of furanocoumarins in grapefruit juice by edible fungi.

Furanocoumarins (FCs) in the human diet irreversibly inhibit human cytochrome P450 3A4 (CYP 3A4) and are responsible for the "grapefruit/drug""interaction phenomenon. Previously, we reported that FCs in grapefruit juice (GFJ) bind to autoclaved Aspergillus niger, and this binding reduced the GFJ inhibition of CYP 3A4. However, A. niger is not an edible fungus, and thus, potentially similar binding by edible fungi was also characterized. In this study, autoclaved Morchella esculenta, an edible ascomycete, removed much of the FC content in GFJ, resulting in decreased inhibition of CYP 3A4 activity by the GFJ. Three other edible fungi, Monascus purpureus, Pleurotus sapidus, and Agaricus bisporus, were evaluated for their binding with two of the major FCs in GFJ, 6',7'-dihydroxybergamottin (DHB) and bergamottin (BM). These autoclaved edible fungi removed these FCs from GFJ, similar to M. esculenta, indicating that binding is a general, passive interaction between FCs and fungal hyphae. The removal of FCs was independent of pH in GFJ. Dried fungal material of M. esculenta was also effective in removing FCs from GFJ and occurred with GFJ samples prepared from both fresh grapefruit and GFJ concentrate.

Binding of furanocoumarins in grapefruit juice to Aspergillus niger hyphae.

Furanocoumarins (FCs) in grapefruit are involved in the "grapefruit/drug interactions" in humans, in which the FCs inhibit the intestinal cytochrome P450 3A4 (CYP 3A4) activity responsible for metabolizing certain prescribed medications. These interactions have adversely affected the grapefruit industry and have led a need to develop a process to remove the FCs from grapefruit juice (GFJ) in a manner that retains much of the original juice sensory attributes. In our experiments, grapefruit juice was incubated with Aspergillus niger, and the compositional changes in hydroxycinnamates, flavonoid glycosides, and the FCs were monitored. Many of the FCs and 7-geranyloxycoumarin were efficiently taken up by the fungal tissue, whereas no uptake occurred with the polar hydroxycinnamates, flavonoid glycosides, and a few of the polar FCs. This biosorption was also observed with autoclaved A. niger, indicating that the uptake of non-polar FCs by the fungal hyphae was due to adsorption rather than metabolism. The binding of the FCs to autoclaved fungus was complete within 4 h, and the level of binding was proportional to the amount of autoclaved fungal hyphae used. This removal of the FCs from GFJ led to a reduced inhibition of CYP 3A4 activity in in vitro assays by both GFJ and GFJ extracts.

Aspergillus niger metabolism of citrus furanocoumarin inhibitors of human cytochrome P450 3A4.

Fungi metabolize polycyclic aromatic hydrocarbons by a number of detoxification processes, including the formation of sulfated and glycosidated conjugates. A class of aromatic compounds in grapefruit is the furanocoumarins (FCs), and their metabolism in humans is centrally involved in the "grapefruit/drug interactions." Thus far, the metabolism by fungi of the major FCs in grapefruit, including 6', 7'-epoxybergamottin (EB), 6', 7'-dihydroxybergamottin (DHB), and bergamottin (BM), has received little attention. In this study, Aspergillus niger was observed to convert EB into DHB and a novel water-soluble metabolite (WSM). Bergaptol (BT) and BM were also metabolized by A. niger to the WSM, which was identified as BT-5-sulfate using mass spectrometry, UV spectroscopy, chemical hydrolysis, and (1)H and (13)C nuclear magnetic resonance spectroscopy. Similarly, the fungus had a capability of metabolizing xanthotoxol (XT), a structural isomer of BT, to a sulfated analog of BT-5-sulfate, presumably XT-8-sulfate. A possible enzyme-catalyzed pathway for the grapefruit FC metabolism involving the cleavage of the geranyl group and the addition of a sulfate group is proposed.