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Efficiency of apples, strawberries, and tomatoes for reduction of oxidative stress in pigs as a model for humans.

OBJECTIVE: The present study investigated the potency of apples, strawberries, and tomatoes in decreasing oxidative stress in pigs as a model for humans. METHODS: Forty-eight growing pigs were individually penned in metabolic cages and divided into six groups. All groups received isocaloric daily rations composed of a basal diet supplemented with starch, linseed oil, linseed oil and apples, linseed oil and strawberries, linseed oil and tomatoes, or linseed oil, apples, strawberries and tomatoes. Oxidative stress was evaluated at the beginning and at the end of the 22-d experimental period by measuring the degree of DNA damage in mononuclear blood cells, the concentration of malondialdehyde (MDA) in plasma, the MDA excretion rate, the concentration of tocopherols in plasma, plasma total antioxidant status, and erythrocyte glutathione peroxidase concentration. RESULTS: The results confirmed that the linseed oil diet significantly increased MDA formation in the body and DNA damage to mononuclear blood cells in comparison with the starch diet. Although the MDA concentration in plasma was significantly decreased in all fruit groups, urine MDA excretion rate was decreased only in the linseed oil/apple and linseed oil/apple/strawberry/tomato groups. Supplementation with fruit significantly decreased DNA damage, was more pronounced in the linseed oil/apple and linseed oil/apple/strawberry/tomato groups. Significantly lower glutathione peroxidase concentrations were observed only in the linseed oil/strawberry group. CONCLUSIONS: Our findings support the hypothesis that supplementation with apples, strawberries, or tomatoes effectively decreases oxidative stress by decreasing MDA formation in the body and by protecting mononuclear blood cells against increased DNA damage. This effect was particularly pronounced in the group supplemented with a fruit mixture; among the single fruit supplements, the most beneficial effect was obtained with apples.

Analysis of Variance↗

Determination of diflubenzuron in apples by gas chromatography.

A method for the determination of residues of the insecticide diflubenzuron, 1-(4-chlorophenyl)-3-(2,6-difluorobenzoyl)urea, in apples using gas chromatography with electron-capture detection has been developed and validated. The three solvents ethyl acetate, acetone and dichloromethane were tested for extraction of diflubenzuron residues from apples. Dichloromethane gave the highest recovery and the lowest background and was chosen as the extraction solvent. After extraction the residue of diflubenzuron was derivatized with heptafluorobutyric anhydride. The derivative was purified by silica solid-phase extraction using toluene as the eluent. The external standard calibration was linear over the range 0.05-1.0 microgram/ml and the limit of quantification was 0.03 mg/kg apple using 25 g samples. Recovery of diflubenzuron from spiked apples (0.1-0.8 mg/kg) was 80-88% with a relative standard deviation of less than 10% (n = 5). The method was applied to the determination of diflubenzuron residues in apples from a treated field.

Acetates↗

High-performance liquid chromatography with diode-array detection for the determination of phenolic compounds in peel and pulp from different apple varieties.

Quantitative analysis of phenolic compounds from four apple varieties (Golden and Red Delicious, Granny Smith and Green Reineta) using high-performance liquid chromatography with diode-array detection was carried out. For each variety, both peel and pulp were analysed. The identification of phenolic compounds was made by comparing their retention times and UV spectra with those of standards. The results revealed differences between pulp and peel in all cases studied. The highest levels of phenolic compounds were found in the peel. High levels of catechins and flavonol glycosides, especially rutin, were found in apple peels. Chlorogenic acid was the major peak in the pulp for all apple varieties studied except for Granny Smith. Significant quantitative differences between the apple varieties were also found, the Golden Delicious variety showing the lowest content of phenolic compounds and Green Reineta variety the highest. The recovery of phenolic compounds from both peel and pulp was measured in all apple varieties. The values ranged between 95 and 105%, indicating close to quantitative recovery for the method used.

Caffeic Acids↗

Plant-based heterologous expression of Mal d 2, a thaumatin-like protein and allergen of apple (Malus domestica), and its characterization as an antifungal protein.

Mal d 2 is a thaumatin-like protein and important allergen of apple fruits that is associated with IgE-mediated symptoms in apple allergic individuals. We obtained a full-length cDNA clone of Mal d 2 from RNA isolated from ripe apple (Malus domestica cv. Golden Delicious). The cDNA's open reading frame encodes a protein of 246 amino acid residues including a signal peptide of 24 residues and two putative glycosylation sites. The deduced amino acid sequence of the mature Mal d 2 protein results in a predicted molecular mass of 23,210.9Da and a calculated pI of 4.55. Sequence comparisons and molecular modeling place Mal d 2 among those pathogenesis-related thaumatin-like proteins that contain a conserved acidic cleft. In order to ensure the correct formation of the protein's eight conserved disulfide bridges we expressed Mal d 2 in Nicotiana benthamiana plants by the use of a tobacco mosaic viral vector. Transfected N.benthamiana plants accumulated Mal d 2 to levels of at least 2% of total soluble protein. MALDI-TOF mass spectrometric analyses of the recombinant Mal d 2 and its proteolytic fragments showed that the apple-specific leader peptide was correctly cleaved off by the host plant and that the mature recombinant protein was intact and not glycosylated. Purified recombinant Mal d 2 displayed the ability to bind IgE from apple-allergic individuals equivalent to natural Mal d 2. In addition, the recombinant thaumatin-like Mal d 2 exhibited antifungal activity against Fusarium oxysporum and Penicillium expansum, implying a function in plant defense against fungal pathogens.

Allergens↗

Multiplex RT-PCR-ELISA compared with bioassay for the detection of four apple viruses.

A sensitive and reliable multiplex RT-PCR-ELISA technique for the detection of Apple chlorotic leaf spot virus, Apple stem pitting virus, Apple mosaic virus and Apple stem grooving virus was developed. This technique is compared with the method used commonly for indexing by woody indicators, which is time consuming and expensive. For the RT-PCR-ELISA technique, the amplified products were labeled with digoxigenin during the RT-PCR by incorporation of a digoxigenin labeled primer. After hybridization of the PCR products to specific capture oligonucleotides, which were bound covalently to the surface of NucleoLink strips, anti-digoxigenin antibodies were used for detection. More than 100 samples were tested in parallel by indexing and multiplex-RT-PCR-ELISA. All infections detected by woody indicators were also detected by multiplex RT-PCR-ELISA. Furthermore, additional infections were only found by multiplex RT-PCR-ELISA. The colourimetric detection of multiplex-RT-PCR products was at least as sensitive and sometimes slightly more sensitive than detection by gel electrophoresis. The results show that this molecular technique is more reliable for the detection of the above mentioned apple viruses than indexing by woody indicators, thereby helping to reduce cost and time during the certification of plant material.

Biological Assay↗

Time-resolved fluorescence immunoassay (TRFIA) for the detection of Escherichia coli O157:H7 in apple cider.

An immunoassay based on immunomagnetic separation and time-resolved fluorometry was developed for the detection of E. coli O157:H7 in apple cider. The time-resolved fluorescent immunoassay (TRFIA) uses a polyclonal antibody bound to immunomagnetic beads as the capture antibody and the same antibody labeled with europium as the detection antibody. Cell suspensions of 10(1) to 10(8) E. coli O157:H7 and K-12 organisms per ml were used to test the sensitivity and specificity of the assay. The sensitivity of the assay was 10(3) E. coli O157:H7 cells with no cross-reaction with K-12. Pure cultures of E. coli O157:H7 (10(1) to 10(5) CFU/ml) in apple cider could be detected within 6 h, including 4 h for incubation in modified EC broth with novobiocin and 2 h for the immunoassay. When apple cider was spiked with 1 to 10(3) CFU/ml of E. coli O157:H7 and 10(6) CFU/ml of K-12, our data show that the high level of K-12 in apple cider did not impede the detection of low levels of O157:H7. The minimum detectable numbers of cells present in the initial inoculum were 10(2) and 10(1) CFU/ml after 4- and 6-h enrichment. The TRFIA provides a rapid and sensitive means of detecting E. coli O157:H7 in apple cider.

Antibodies, Bacterial↗

Survival of acid-adapted or nonadapted Escherichia coli O157:H7 in apple wounds and surrounding tissue following chemical treatments and storage.

This study evaluated survival/growth of acid-adapted or nonadapted Escherichia coli O157:H7 inoculated (4 log CFU/wound) in wounds (10 mm deepx6 mm diameter) of apples. Wounds were inoculated with a green fluorescent protein (GFP)-expressing derivative of a rifampicin-resistant strain of E. coli O157:H7 ATCC 43895 and allowed to attach (1 h). Apples were dipped (2 min) in solutions (approximately 25 degrees C) of water (W), 5% acetic acid (AA), 5% hydrogen peroxide (HP), 0.02% sodium hypochlorite (SH), or not treated (NT), and stored at 25 degrees C. Survivors were determined in cores (10-mm deep) of the apple wounds (12 mm from center of wound; inner core) and surrounding tissue (18 mm from center of wound; outer core) after homogenizing the samples in Dey-Engley (D/E) neutralizing broth and plating on tryptic soy agar (TSA) and TSA supplemented with 100 microg/ml rifampicin (35 degrees C, 48 h) after 0, 2 and 5 days. Average bacterial populations at day-0 were 4.0 and 2.0 logs in the inner and outer core, respectively. In the inner core of the untreated apples populations increased to 7.0 logs at day-2, while counts did not exceed 3.0 logs in the outer core during storage of all treatments. Previous acid-adaptation of the cultures did not affect survival of the pathogen. Dipping in W, AA and SH did not reduce initial bacterial populations, while at day-2 of storage inner core counts from W, AA and SH reached 7.1, 5.5 and 6.9 logs, respectively. In contrast, HP reduced initial counts in the inner core by approximately 1.5 logs, but they increased to 7.0 logs by day-2. Populations of all treatments reached 6.6-7.2 logs in the inner core by day-5. Thus, sanitizer treatment did not effectively reduce nor inhibit growth of E. coli O157:H7 contamination in apple wounds and surrounding tissue.

Acetic Acid↗

The antifungal properties of chitosan in laboratory media and apple juice.

The antimicrobial properties of chitosan glutamate, a derivative of chitin, were investigated in laboratory media and apple juice against 15 yeasts and moulds associated with food spoilage in order to assess the potential for using chitosan as a natural food preservative. Of the seven strains of filamentous fungi studied, chitosan reduced the growth rate of Mucor racemosus at 1 g/l whilst concentrations of 5 g/l were required to completely prevent growth of three strains of Byssochlamys spp. on agar plates incubated at 25 degrees C for 3 weeks. Three strains of filamentous fungi were resistant to the antifungal effects of chitosan at 10 g/l. The presence of chitosan in apple juice (pH 3.4) at levels ranging from 0.1 to 5 g/l inhibited growth at 25 degrees C of all eight spoilage yeasts examined in this study. The initial effect of chitosan in apple juice was biocidal with viable numbers reduced by up to 3 log cycles. Following an extended lag phase, some strains recovered and resumed growth to levels similar to those observed in unsupplemented apple juice. The most sensitive strain was an isolate of Zygosaccharomyces bailii obtained from a spoiled carbonated beverage; this yeast was completely inactivated by chitosan at 0.1 and 0.4 g/l for 32 days of storage at 25 degrees C. The most resistant strain was Saccharomycodes ludwigii, an isolate from spoiled cider: a level of addition of 5 g/l of chitosan was required to inactivate this strain and to maintain yeast-free conditions in apple juice for 14 days at 25 degrees C. Growth inhibition and inactivation of filamentous moulds and yeasts, respectively, was concentration-, pH- and temperature-dependent. It was concluded that chitosan was worthy of further study as a natural preservative for foods prone to fungal spoilage.

Ascomycota↗

Genetic clues to the origin of the apple.

Molecular genetic markers complement archaeological, breeding and geographical investigations of the origins, history and domestication of plants. With increasing access to wild apples from Central Asia, along with the use of molecular genetic markers capable of distinguishing between species, and explicit methods of phylogeny reconstruction, it is now possible to test hypotheses about the origin of the domesticated apple. Analyses of nuclear rDNA and chloroplast DNA (cpDNA) sequences indicate that the domesticated apple is most closely related to series Malus species. Moreover, the occurrence of a shared 18-bp duplication in the cpDNAs of wild and cultivated apple supports the close relationship between them. Hypotheses about the hybridization and the origin of the domesticated apple cannot be rejected completely until more variable, phylogenetically informative markers are found.

DNA, Ribosomal Spacer↗

Binding of antibodies against birch pollen antigens/allergens to various parts of apples as studied by immuno-gold electron microscopy.

The clinically and biochemically observed correlation between birch pollen allergy and hypersensitivity to apples was investigated by immunocytochemical techniques. For this purpose, apple tissue was fixed in p-formaldehyde and embedded in Lowicryl K4M resin at -35 degrees C. Ultrathin sections were cut and successively incubated with rabbit antibodies against birch pollen antigens/allergens and protein A/gold. Specific antibody binding sites were detected throughout the apple fruit (peel, fruit flesh, seed). Control sections incubated with normal rabbit IgG antibodies and protein A/gold showed minimal background staining. It was concluded from the results of immunocytochemical labelling that apple tissue and birch pollen contain similar molecular structures which lead to the observed cross-reactions. The present immunocytochemical results confirm biochemical investigations reporting partial structural identity of antigens/allergens in birch and apple.

Allergens↗

[Exercise-induced anaphylaxis after apple intake].

Exercise-induced anaphylaxis frequently is related to food ingestion, so that it receives the name of "food-dependent exercise-induced anaphylaxis". The food identification is difficult in some patients. We report two patients with apple-dependent exercise-induced anaphylaxis. STUDY: hematimetries, biochemistries, VMA and 5-HIAA in urine. Complement levels, antinuclear antibodies and viruses serology. ALLERGOLOGIC STUDY: skin prick test with a common inhalant. Skin prick test and prick by prick with different foods. Total Ig E level and specific Ig E determination by the CAP-Pharmacia System. Oral challenge test with apple, exercise test and exercise challenge test after eating an apple. Prick by prick test with fresh apple was positive. Specific Ig E determination by the CAP-Pharmacia System revealed a positive result. We get to reproduce the episode with an exercise challenge test after eating an apple in the same conditions of usual exercise in both patients.

Adolescent↗

Seasonal variation in food allergy to apple.

The aim of the study was to investigate the possibility of a seasonal variation in reactivity to apples in 27 birch pollen allergic patients. Before and during the birch pollen season 1998, the patients were subjected to double-blind, placebo-controlled food challenges (DBPCFCs) with grated fresh Golden Delicious apple followed by an open food challenge with whole fresh apple. The clinical reactions elicited during the challenges were evaluated both by the patients and the investigators. Moreover, the skin reactivity and the in vitro reactivity to apple were evaluated by skin prick test (SPT), leukocyte histamine release (HR), measurement of specific IgE, and immunoblotting experiments. The sensitivity of the DBPCFC, when compared with the result of the open challenge, was 0.74 (14/19) before the season and 0.80 (16/20) during the season. None of the patients reacted to the blinded challenge without a subsequent reaction to the open challenge. One placebo reaction was registered both before and in season, but not in the same patient. The patient scores of the first positive challenges, and the maximal scores of each combined blinded and open challenge session, were significantly increased during the pollen season (P<0.05). The scores of the open challenge were significantly higher than the scores of the DBPCFC both before the season and during the in-season challenges (P<0.05). Specific IgE against Golden Delicious increased during season (P<0.05), while neither SPT, HR, nor immunoblotting experiments could confirm an increase in reactivity. In conclusion, the results of the oral challenge tests indicated an increase in clinical reactivity to apples during the birch pollen season in birch pollen allergic individuals.

Blotting, Western↗

Comparative content of some bioactive compounds in apples, peaches and pears and their influence on lipids and antioxidant capacity in rats.

The aim of this study was to compare some bioactive compounds in apples, peaches and pears and their influence on lipids and antioxidant capacity in rats. The content of total polyphenols (g/100g) was 0.23 +/- 0.03; 0.22 +/- 0.03 and 0.68 +/- 0.1 in peeled fruits and 0.48 +/- 0.04, 0.47 +/- 0.04 and 1.2 +/- 0.12 in peels of peaches, pears and apples, respectively. Caffeic, p-coumaric and ferulic acids and the total radical-trapping antioxidative potential (TRAP) values in peeled apples and their peels were significantly higher than in peaches and pears, respectively. Contrarary, no significant differences in the content of dietary fiber among the studied fruits were found. The content of all studied indices in peels was significantly higher than peeled fruits (p < 0.05 ). A good correlation between the total polyphenols and the TRAP values was found in all fruits. Diets supplemented with apples and to a less extent with peaches and pears have improved lipid metabolism and increased the plasma antioxidant potential especially in rats fed with added cholesterol. The highest content of biologically active compounds and the best results in the experiment on rats makes apple preferable for dietary prevention of atherosclerosis and other diseases.

Journal Article↗

Grape and apple wines volatile fermentation products and possible relation to spoilage.

The main volatile by-products of the alcoholic fermentation of grape wine, cider and apple pulp wine were investigated to determine if any correlated with spoilage resistance in the latter two. Spoilage was visually detected after seven days in low-alcohol grape wine in comparison to 11 and 16 days in cider and apple pulp wine, respectively. Acetaldehyde, ethyl acetate, methanol, propanol, isobutanol and amyl alcohols were the main fermentation by-products detected in all three wines. There were highest concentrations of acetaldehyde, ethyl acetate, methanol and propanol in grape wine and, therefore, these by-products could not be implicated in spoilage resistance in apple wines. Increased concentrations of isobutanol and amyl alcohols, however, in cider and apple pulp wine in comparison to grape wine might have been the reason for spoilage resistance in the apple wines.

1-Propanol↗

Activity and concentration of polyphenolic antioxidants in apple: effect of cultivar, harvest year, and storage conditions.

Consumers' increasing interest in the relationship between diet and health is a sign for food producers to pay more attention to potential health-protecting compounds in new product development and food processing. From a production chain perspective the choice of the raw material that is used is important for the health-protecting potential of the end product. Four apple cultivars (Jonagold, Golden Delicious, Cox's Orange, and Elstar), which can be used as fresh apples or in processed apple products, were compared with regard to flavonol, catechins, phloridzin, and chlorogenic acid concentrations and antioxidant activity. Jonagold apples possessed the highest flavonoid concentration and the highest antioxidant activity. To study seasonal differences, apples from three different harvest years were analyzed, but in three cultivars no effect on flavonoid concentration and antioxidant activity was observed. Long-term storage, both at refrigerator temperature and under controlled atmosphere conditions, was found not to influence flavonoid concentration or antioxidant activity.

Antioxidants↗

Effect of 1-methylcyclopropene on volatile emission and aroma in cv. Anna apples.

The rapidly ripening summer apple cultivar Anna was treated with 0.1 micro L(-1) and 1 microL L(-1) 1-methylcyclopropene (MCP) at harvest and kept at 20 degrees C, or stored for 5 weeks at 0 degrees C and then transferred to 20 degrees C. Total volatiles were not reduced by treatment with 0.1 microL L(-1) MCP, but were 70% lower in fruits treated with 1 microL L(-1) MCP than in untreated fruits. Ethylene production was 50% and 95% inhibited by 0.1 microL L(-1) and 1 microL L(-1) MCP, respectively. The volatiles produced by fruit at harvest were predominantly aldehydes and alcohols, with some acetate esters as well as 2-methyl butyl acetate and beta-damascenone. During ripening, the acetate and butyrate esters increased greatly and alcohols and aldehydes decreased. MCP-treated apples retained more alcohols, aldehydes, and beta-damascenone volatiles than did untreated apples. Sensory evaluation found that control and 0.1 microL L(-1) treated apples developed more fruity, ripe, and overall aromas, but the preference was for the 1 microL L(-1) treated apples with a less ripe aroma.

Alcohols↗

Gas chromatographic-olfactometric characterization of aroma compounds in two types of cashew apple nectar.

Cashew apple nectar is a secondary product from the production of cashew nuts and possesses an exotic tropical aroma. Aroma volatiles in pasteurized and reconstituted (from concentrate) Brazilian cashew apple nectars were determined using GC-MS and split, time-intensity GC-olfactometry (GC-O)/GC-FID. Methional, (Z)-1,5-octadien-3-one, (Z)-2-nonenal, (E,Z)-2,4-decadienal, (E,E)-2,4-decadienal, beta-damascenone, and delta-decalactone were identified for the first time in cashew apple products. These compounds plus butyric acid, ethyl 3-methylbutyrate, 2-methylbutyric acid, acetic acid, benzaldehyde, homofuraneol, (E)-2-nonenal, gamma-dodecalactone, and an unknown were the most intense aroma volatiles. Thirty-six aroma volatiles were detected in the reconstituted sample and 41 in the pasteurized sample. Thirty-four aroma active components were common to both samples. Ethyl 3-methylbutyrate and 2-methylbutyric acid were character impact compounds of cashew apple (warm, fruity, tropical, sweaty). Using GC-pFPD, 2-methyl-3-furanthiol and bis(2-methyl-3-furyl) disulfide were identified for the first time in cashew apple. Both were aroma active (meaty).

Anacardium↗

Effects of processing treatment and sorbate addition on the flavor characteristics of apple cider.

Processing treatments used to produce a microbiologically "safe" apple cider were evaluated to determine the impact of these treatments on the overall flavor characteristics. Apple cider with (0.1%) and without (0%) potassium sorbate was subjected to four processing treatments: untreated, irradiated at 2 kGy, irradiated at 4 kGy, and pasteurized. Volatile flavor compounds were isolated from the cider using solid-phase microextraction methods with gas chromatographic analysis. A trained descriptive analysis panel evaluated sensory attributes. The effects of the processing treatment were dependent on the presence of sorbate in the apple cider. Irradiation treatments resulted in a decrease in the content of esters characteristic of apple flavor and an increase in the content of alcohols and aldehydes formed through lipid oxidation reactions. The presence of sorbate reduced the effects of the irradiation treatments on these volatile flavor compounds. Sensory panelists, however, detected higher intensities of undesirable flavor attributes, including "cardboard flavor", and lower intensities of the desirable "apple flavor" in irradiated cider with added sorbate.

Alcohols↗