Konfitüren, Säfte und Trocknung biologisch angebauter Lebensmittel: Was geschieht mit Allergenen während der Verarbeitung?

Food allergy is an increasing health problem, whereby plant allergens are one of the major source eliciting allergenic reactions. Ninety percent of birch pollen allergic patients develop intolerances to fruits and vegetables. Antibodies against Bet v 1, the major birch pollen allergen, cross-react with Bet v 1-homolog proteins present in a variety of fruits and vegetables. Bet v 1 and Bet v 1-homolog proteins belong to the family 10 of pathogenesis-related proteins (PR-10). In plants PR-10 proteins are induced in various stress conditions, e.g. pathogen attack, wounding or environmental factors. The amount of allergens is influenced by pre- and post-harvesting factors, such as cultivar, organic or conventional cultivation and storage conditions as well as different processing methods. The aim of the project is to develop an Enzyme Linked Immunosorbent Assay (ELISA) to determine the allergen content in fruits (apple, strawberry, plum, tomato) and processed food products (apple juice, strawberry jam). The influence of organic versus conventional farming on the allergen content will be investigated. Furthermore, the effects of different processing techniques will be analyzed

Effect of tomato variety, cultivation, climate and processing on Sola l 4, an allergen from Solanum lycopersicum

Tomatoes (Solanum lycopersicum) are one of the most consumed vegetables worldwide. However, tomato allergies in patients suffering from birch pollen allergy occur frequently. Due to highly similar protein structures of the tomato allergen Sola l 4 and the major birch pollen allergen Bet v 1, patients cross-react with allergenic proteins from tomato as well as other fruits or vegetables. The aim of this study was to quantify Sola l 4 in various tomatoes differing in color, size and shape for identification of varieties with a reduced allergen level. Therefore, an indirect competitive ELISA using a specific polyclonal Sola l 4 antibody was developed. In addition, two varieties, both cultivated either conventionally or organically and furthermore dried with different methods, were analyzed to investigate the influence of the cultivation method and processing techniques on Sola l 4 level. Within 23 varieties, Sola l 4 content varied significantly between 0.24 and 1.71 μg Sola l 4/g FW. The tomato cultivars Rugantino and Rhianna showed the significantly lowest level, whereas in cultivars Farbini and Bambello the significantly highest concentration was determined. Drying of tomatoes in the oven and by sun resulted in a significant decrease. The thermal instability was verified for the recombinant Sola l 4 emphasizing the results for the native protein in dried tomato samples. Overall, the Sola l 4 content is cultivar-dependent and no correlation between color and Sola l 4 amount was found. During the drying process of tomatoes Sola l 4 level was significantly reduced due to thermal instability. Growing conditions have a minor effect whereas seasonal effects show a more pronounced impact. These findings could extend the knowledge about the allergen level of different tomato varieties and may help to improve food safety to potentially increase the life quality of patients suffering from birch pollen allergy

Effect of the Strawberry Genotype, Cultivation and Processing on the Fra a 1 Allergen Content

Birch pollen allergic patients show cross-reactivity to vegetables and fruits, including strawberries (Fragaria × ananassa). The objective of this study was to quantify the level of the Fra a 1 protein, a Bet v 1-homologous protein in strawberry fruits by a newly developed ELISA, and determine the effect of genotype, cultivation and food processing on the allergen amount. An indirect competitive ELISA using a specific polyclonal anti-Fra a 1.02 antibody was established and revealed high variability in Fra a 1 levels within 20 different genotypes ranging from 0.67 to 3.97 μg/g fresh weight. Mature fruits of red-, white- and yellow-fruited strawberry cultivars showed similar Fra a 1 concentrations. Compared to fresh strawberries, oven and solar-dried fruits contained slightly lower levels due to thermal treatment during processing. SDS-PAGE and Western blot analysis demonstrated degradation of recombinant Fra a 1.02 after prolonged (>10 min) thermal treatment at 99 ◦ C. In conclusion, the genotype strongly determined the Fra a 1 quantity in strawberries and the color of the mature fruits does not relate to the amount of the PR10-protein. Cultivation conditions (organic and conventional farming) do not affect the Fra a 1 level, and seasonal effects were minor

Higher expression of the strawberry xyloglucan endotransglucosylase/hydrolase genes FvXTH9 and FvXTH6 accelerates fruit ripening

Fruit softening in Fragaria (strawberry) is proposed to be associated with the modification of cell wall components such as xyloglucan by the action of cell wall‐modifying enzymes. This study focuses on the in vitro and in vivo characterization of two recombinant xyloglucan endotransglucosylase/hydrolases (XTHs) from Fragaria vesca, FvXTH9 and FvXTH6. Mining of the publicly available F. vesca genome sequence yielded 28 putative XTH genes. FvXTH9 showed the highest expression level of all FvXTHs in a fruit transcriptome data set and was selected with the closely related FvXTH6 for further analysis. To investigate their role in fruit ripening in more detail, the coding sequences of FvXTH9 and FvXTH6 were cloned into the vector pYES2 and expressed in Saccharomyces cerevisiae. FvXTH9 and FvXTH6 displayed xyloglucan endotransglucosylase (XET) activity towards various acceptor substrates using xyloglucan as the donor substrate. Interestingly, FvXTH9 showed activity of mixed‐linkage glucan:xyloglucan endotransglucosylase (MXE) and cellulose:xyloglucan endotransglucosylase (CXE). The optimum pH of both FvXTH9 and FvXTH6 was 6.5. The prediction of subcellular localization suggested localization to the secretory pathway, which was confirmed by localization studies in Nicotiana tabacum. Overexpression showed that Fragaria × ananassa fruits infiltrated with FvXTH9 and FvXTH6 ripened faster and showed decreased firmness compared with the empty vector control pBI121. Thus FvXTH9 and also FvXTH6 might promote strawberry fruit ripening by the modification of cell wall components

Lebensmittelchemie 2012

Neben der Identifizierung von Aromastoffen durch die molekulare Sensorik sind die Aufklärung ihrer ‧Biosynthese sowie ihre biotechnische Produktion ins Zentrum der Aromastoffforschung gerückt. Geruch und Geschmack vermitteln die Qualität eines Lebensmittels. Den Geruch bestimmen flüchtige Aromastoffe, die heute vielen industriell hergestellten Lebensmitteln zugesetzt werden, um herstellungsbedingte Verluste auszugleichen oder das Aroma abzurunden. Der Verbraucher akzeptiert dabei eher natürliche Aromen. Der Begriff „natürlich“ darf nach EU-Recht allerdings nur dann zur Bezeichnung eines Aromas verwendet werden, wenn es ausschließlich Aromaextrakte oder natürliche Aromastoffe enthält.1) Dazu zählen biotechnisch erzeugte Aromastoffe, die in puncto Nachhaltigkeit für Aromenhersteller eine Alternative sind zu den klassischen natürlichen Aromen aus pflanzlichen Rohstoffen wie ätherischen Ölen. Biotechnische Verfahren dienen vermehrt dazu, strukturell komplexe Aromastoffe billiger und in größeren Mengen herzustellen, als dies mit chemischen Synthesen oder durch Pflanzenextraktion möglich ist.2) Fortschritte in Enzymtechnik, Gentechnik, Bioprozess-Monitoring und Produktisolierung bieten viel versprechende Möglichkeiten für biotechnische Aromastoffproduktion.3) Spezifische Strategien sind hierfür aber nur dann weiterzuentwickeln, wenn Enzymologie und Regulation der Aromastoffbiosynthese auf molekularer Ebene verstanden sind. Daher ist die biochemische Forschung zur Aufklärung von Biosynthesewegen in Pflanzen ein wichtiger Bestandteil der Aromabiotechnik

Metabolic engineering of apple by overexpression of the MdMyb10 gene

Flavonoids are low-molecular-weight phenolic compounds that are widely distributed in the plant kingdom. They have different roles in plant resistance to biotic and abiotic stresses. The transcription factor gene MdMyb10 (Gene Bank: DQ267896) was introduced into two apple (Malus domestica Borkh.) cultivars i.e. ‘Holsteiner Cox (HC)’ and ‘Gala’ via Agrobacterium-mediated transformation. The regenerated shoots were selected on kanamycin containing media. The presence of additional MdMyb10 gene in putative shoots was confirmed by PCR, RT-PCR and Southern blotting. Expression level of introduced MdMyb10 gene was analyzed by quantitative real time PCR. The results confirmed a dramatic increase in overexpression of MdMyb10 in the transgenic plants, up to 1261 and 847-folds for cultivars Holsteiner Cox and Gala, respectively compared to non-transformed negative control plants. HPLC-MS was used to determine the levels of different flavonoid compounds in both non-transgenic and transgenic plants. In MdMyb10 ‘HC’ transgenic plants, some of the polyphenols analyzed were enhanced while others were reduced in comparison to their levels in the non-transgenic plants. On the other hand, all of the analyzed polyphenol classes were induced in MdMyb10 ‘Gala’ transgenic plants in comparison to their levels in the non-transgenic plants. In the present study, the flavonoid pathway was successfully modified in apple by overexpressing the MdMyb10 transcription factor to validate the hypothesis of increased effect on plant disease resistance.Islamic Development Ban

Die Testung von Äpfeln auf ihre Allergenität

Apples have several allergens which, when eaten, lead to symptoms in the mouth within 5-10 min-and therefore cannot be eaten by those suffering from apple allergies. In Germany around 7.5 million people have developed specific antibodies against the main allergen (Mal d 1) in apples and are thus sensitized. At least 3.5 million of them develop the sometimes-considerable allergic symptoms as an expression of an Oral Allergy Syndrome. So far there is no drug therapy for this allergy. Apple allergy sufferers can therefore only do without apples entirely, or eat apples that have been heated beforehand or look for varieties that contain few allergens and can therefore be described as "allergy-friendly" apple varieties. Solely determinations of allergens in the laboratory cannot predict whether an apple can be eaten by apple allergy sufferers without allergic symptoms; clinical trials are required for this. We describe a standardized clinical, oral provocation test that can be used to characterize a low-allergen, allergy-friendly apple or apple variety. The results of such at least three-year tests can be used to award the ECARF seal for allergy-friendly products

Final report for the CORE Organic Plus funded project “Drying, Juices and Jams of Organic Fruit and Vegetables: what happens to Desired and Non-Desired compounds? FaVOR-DeNonDe” Period covered: 30 March 2015 - 29 March 2018

The main activity was related to the description of qualitative data on commonly consumed fruit (strawberry, plum, apple) and vegetables (tomato and sweet bell pepper) along some processing chains (jam, juices and dried products), pointing out the role of different sources of variability, such as the type of cultivation (Conventional, CONV or Organic, ORG), the different cultivars, the sampling year, and the type of processing. A particular attention has been paid on the type of processing, where the use of innovative plant was performed, in comparison with traditional or home-made processing typologies. This for the need, especially for local and small farmers, of small and simple plants to process agricultural products, in order to add value for their productions, for achieving high quality and safe products. For this scope three types of approaches for processing were used: - a miniaturized multifunctional processing line (MT), already validated in previous Projects, in comparison with home-made technique (HM) for jam strawberry production; - a pneumatic press, a rack-press and a belt-press compared for the production of apple juices; - an innovative drier, completely supplied by solar energy (SUN) compared with a traditional forced air oven-drier (OVEN) was used for the production of prunes, dried tomatoes and sweet bell peppers. The list of analyzed samples is shown on the Final Report Document: SCHEME OF ANALYZED SAMPLES (ORG is Organic, CONV is Conventional) The processed samples were produced in open field on two consecutive sampling years (2015 and 2016), on experimental fields. A third year of sampling was added, considering the evaluation made on raw products. Moreover, for tomato and sweet bell pepper, the production made from open fields of private organic growers, was considered, so obtaining samples in two consecutive years (2016 and 2017). Hence, a further comparison of the quality indexes was made on raw products in three consecutive years (2015, 2016 and 2017). The control samples were represented by the lyophilized material, generally referred as “raw” or “not treated” sample. The considered qualitative data regarded "Desired" and "Non Desired" compounds, being the desired ones those potentially healthy and tasty for humans, such as phytochemicals, antioxidants and volatiles, and the non desired some anti-nutritional traits, such as the mycotoxin patulin and the presence of allergens. The samples were also evaluated by their sensory properties, by using a trained panel. As for the allergen analysis, they were analyzed by indirect competitive ELISA for determination of Bet v 1-related protein, or pathogenesis-related proteins (PR-10), causing birch pollen related fruit allergies. A big amount of data was collected, with a consequent variability among them. The general consideration about desired compounds that can be made is that, if a well detectable difference in composition has been found in raw product, this difference was also retained in the processed one, with the processing technique strongly influencing the qualitative parameters of the final products. In this context, better results in terms of antioxidant presence were found for: - apple juices obtained from belt-press; - strawberry jams obtained with miniaturized small-scale plant; - prunes, tomatoes and sweet bell pepper obtained with oven drying. On the other hand, it has to be pointed out that processing methods causing a lower retention on the phytochemical contents often show a better response in term of sensory properties, as found in apple juices and in solar-dried samples of strawberries, plums and tomatoes. Some differences were found among the assayed cultivars, and, finally, among the comparison of the system of cultivation (ORG vs CONV). Specifically, the system of cultivation mainly influenced the antioxidant content in apples of traditional cultivars of Estonia obtained from aged plants; as for strawberry, it was found an higher amount of antioxidants in organic samples in one year of three, and a significant increase in ascorbic acid was found in the assayed local variety of organic bell pepper, in all three years, but not in the corresponding commercial hybrid. On the other hand, the plum cultivar Jubileum, assayed from Denmark and Norway in two years, resulted not adapted for organic cultivation for the content in ascorbic acid, constantly giving lower indexes in all samples in comparison with fruits from conventional cultivation. As regards the influence of the sampling years, in samples of apples, plums, tomatoes and sweet bell pepper, a tendency to an increase in phytochemicals and antioxidant indexes was detected in 2016 with respect to 2015 samples, due to the very different climate conditions of these two years. Interestingly, for the non desired traits, no clear relationship was found between the presence of patulin and allergens for the systems of cultivations, but significant variations were found with the difference in processing technique, with the sampling years and with the assayed cultivars. Interestingly, for some apple cultivars from Estonia low levels of the allergen Mald1 were detected. Concluding, the sources of variability among the analyzed samples gave, in order of influence, the following ranking: - 1. Processing techniques; - 2. Years of sampling; - 3. Cultivars; - 4. Growing methods

Down-regulation of Fra a 1.02 in strawberry fruits causes transcriptomic and metabolic changes compatible with an altered defense response

The strawberry Fra a 1 proteins belong to the class 10 Pathogenesis-Related (PR-10) superfamily. In strawberry, a large number of members have been identified, but only a limited number is expressed in the fruits. In this organ, Fra a 1.01 and Fra a 1.02 are the most abundant Fra proteins in the green and red fruits, respectively, however, their function remains unknown. To know the function of Fra a 1.02 we have generated transgenic lines that silence this gene, and performed metabolomics, RNA-Seq, and hormonal assays. Previous studies associated Fra a 1.02 to strawberry fruit color, but the analysis of anthocyanins in the ripe fruits showed no diminution in their content in the silenced lines. Gene ontology (GO) analysis of the genes differentially expressed indicated that oxidation/reduction was the most represented biological process. Redox state was not apparently altered since no changes were found in ascorbic acid and glutathione (GSH) reduced/oxidized ratio, but GSH content was reduced in the silenced fruits. In addition, a number of glutathione-S-transferases (GST) were down-regulated as result of Fra a 1.02-silencing. Another highly represented GO category was transport which included a number of ABC and MATE transporters. Among the regulatory genes differentially expressed WRKY33.1 and WRKY33.2 were down-regulated, which had previously been assigned a role in strawberry plant defense. A reduced expression of the VQ23 gene and a diminished content of the hormones JA, SA, and IAA were also found. These data might indicate that Fra a 1.02 participates in the defense against pathogens in the ripe strawberry fruits