Biotechnology of natural products: from allergens to the production of aroma glucosides

Biotechnology of natural products: from allergens to the production of aroma glucosides
Fig. 2.

The professorship Biotechnology of Natural Products (BINA) at the Technical University Munich studies the biosynthetic pathways of natural products in fruits to improve food safety and quality and enable their biotechnological production.

Natural products are low-molecular weight metabolites produced naturally by any organism. They include very small molecules such as ethanol, but also more complex structures, such as the natural sweetener stevioside. Although they occur in small quantities, they have interesting biological activities and chemical structures. In particular, plants are a rich source of bioactive natural products.

The professorship Biotechnology of Natural Products (BINA) has been set up at the Technical University Munich with a contribution made by Degussa in 2003. The research activities of this group comprise studies on the isolation and identification as well as the biosynthesis and the metabolism of plant metabolites with the aim to understand the biological roles of natural products that influence the quality of foods due to their technological and physiological properties.

Instrumental-analytical, biochemical and molecular biological methods are employed for natural product analysis and production. Of special interest is the formation and transformation of secondary plant metabolites such as aroma chemicals and pigments during fruit ripening and the characterisation of the genes and enzymes that participate in this process. With the advent of whole genome sequences of important crop plants such as grapevine, apple, and strawberry, genetic and biochemical analyses of entire gene families putatively involved in the production of plant metabolites became feasible and are exploited by the group.

The scientific results enable quality driven breeding of crop plants and the bio-catalytic production of natural active ingredients for food and cosmetic industry.

The allergy potential of strawberries and tomatoes is affected by the genotype

Studies performed in Sweden, Spain and Germany have demonstrated a correlation of the formation of the natural anthocyanin pigments and the content of Bet v 1 homologous allergens in strawberry fruit, providing a first hint of the biological function of allergens in plants. Thus, besides their generally accepted health-beneficial effects e.g. lowering the risk of cardiovascular diseases and cancer, fruits contain proteins which can elicit food allergies.

During the last decades, the prevalence of allergies increased drastically. In Northern Europe, sensitization against Bet v 1, the major allergen from birch pollen (Betula verrucosa), dominates. Antibodies produced in response to a primary sensitisation to the birch pollen allergen can cross-react with similar allergen epitopes from e.g. apple, strawberry and tomato leading to birch-pollen related food allergy.

Strawberry fruits and tomatoes are among the most widely consumed fruits and vegetables worldwide, however, many people are allergic to them, especially if they have been diagnosed with birch pollen allergy. Symptoms of an immunological reaction to strawberries and tomatoes can affect the skin (urticaria or dermatitis), irritate mucous membranes and trigger a runny nose, and can also lead to abdominal pain. Food allergy sufferers develop symptoms after eating fresh fruit or vegetables, while processed products are often tolerated.

BINA has developed a new method to quantify the level of Bet v 1 homologous allergens in strawberry fruits and tomatoes and determined the effect of the genotype, cultivation, climate, and food processing on the allergen amount. In order to analyse a broad spectrum, genotypes were selected in both cases, which differed in size, shape, and colour. Furthermore, the influence of organic and conventional cultivation conditions as well as various processing methods ranging from sun-drying and oven-drying to freeze-drying of the fruits, were investigated. It was assumed that the concentration of the allergenic protein varies with the colour of the ripe fruit, the state of growth, and the processing method.

Twenty-three different-coloured tomato varieties and 20 strawberry genotypes of different sizes and shapes were examined to determine the genetic effect for the expression of the allergenic protein in the fruits. The concentration of the allergen in both types of fruit varied greatly between genotypes. Thus, the major driver for the contrasting allergen contents was the genetic background of the fruits. In addition, the heat sensitivity of the proteins could be confirmed. If the fruits were exposed to heat during the drying process, their allergy potential was reduced. However, the influence of cultivation conditions (conventional and organic) on the allergy content was minor.

Consequently, the proteins investigated in the studies (Sola l 4.02 in tomatoes and Fra a 1 protein in strawberries) may in future serve as markers for the breeding of hypoallergenic tomato and strawberry varieties and thus, improve the fruit quality for people suffering from birch pollen allergy.

Production of aroma glucosides

The consumption of fruits and vegetables is regarded as essential in a modern balanced diet. One of the main quality attributes of fruits is flavour, which can comprise several hundreds of volatile compounds but only a handful of them exceed their odour thresholds and contribute to the overall aroma. Aroma chemicals are volatile compounds, which are emitted by plants and serve their producers both as appreciated attractants to lure beneficial animals and repellent toxins to deter pests in a species-specific and concentration-dependent manner. Plants have evolved solutions to cope with the challenge to provide sufficient volatiles without poisoning themselves. Uridine-diphosphate sugar dependent glycosyltransferases (UGTs) acting on volatiles is one important part of this sophisticated system, which balances the levels of bioactive metabolites and prepares them for cellular and long-distance transport as well as storage but still enable the remobilisation of disarmed toxins for the benefit of plant protection. UGTs transfer a sugar from an activated carbohydrate donor to an acceptor molecule, mostly an alcohol and thus can form aroma glucosides.

The cosmetics industry

Recently aroma glucosides have been attracted interest by food and cosmetic industry. The focus in use for industrial flavour or fragrance solutions has generally been on the active aroma compound (the aglycone) and the advantage of the functionality of the glycoside for flavour or fragrance formulations e.g. in food or feed products, cleaning liquids and washing powder has been ignored. These are all areas where there is a high use and demand for flavour or fragrance formulations, especially formulations with improved functionality compared to existing formulations. Aroma chemicals are volatile and predominantly poorly water-soluble, which limits their number of industrial applications. In contrast, aroma glycosides show increased water solubility and are more stable than their corresponding aglycones. Although aroma glycosides are odourless, they offer the possibility of controlled release of the bound aroma compound. Thus, aroma glycosides have been proposed for inhibiting unpleasant odour of sanitary and pet-care products, as oral deodorant to reduce faeces odour, to impregnate garments, as ingredients of long-lasting deodorants, to mask perspiration-related odour in beddings, as ingredients of massage oil and bath preparations for sustained release of perfumes, use in air freshener, and so on.

As technical efforts and costs for DNA sequencing dramatically dropped during the last decade, the number of plant genome sequences increased significantly, thus providing opportunities to functionally characterize the UGT gene families in plants. These studies yielded the first UGT genes that encode efficient biocatalysts for the production of aroma glucosides, which have applications in the food, feed, chemical, cosmetic, and pharmaceutical industries as slow release aroma chemicals.

Industry and commercialisation

Aroma glucosides such as those naturally occurring in fruits and leaves can now be produced on an industrial scale at reasonable costs by a process developed by BINA. The biotechnological process applies the same plant enzymes (glycosyltransferases) that are used by nature. 4GENE, a spin-off of BINA, is commercialising aroma glucosides as FLAVOR-ON-DEMAND, worldwide (see Fig. 2). The start-up company develops and produces flavour and fragrance precursors (aroma glucosides) by a whole-cell bio-catalytic process. The project was supported by the EXIST research transfer programme of the German government and is the first winner of the ACHEMA prize in the category Industrial Biotechnology in 2015. The customers can currently choose from a growing portfolio of 22 glycosides.

By means of appropriate triggers such as heat, changing pH value, microbial or enzymatic degradation the aroma glycosides release the bound flavour and fragrance molecules. In personal care products such as deodorants, liners or pads the enzymatic activation can be realised by wetting with body fluids or by microbial degradation (related to the natural skin flora). In perfumes with their direct head note and longer lasting base note the scent effects can be significantly extended by slow hydrolysis of corresponding aroma glucosides. Many aroma glucosides suitable for activation are already available. The broad and continuously growing portfolio in combination with the possible options for activation and release enables for an almost infinite number of potential applications. The directed release of flavours and fragrances can improve the sensory profile of new products, even food products. This creates unique opportunities.

 

Professor Dr Wilfried Schwab

Biotechnology of Natural Products

Technische Universität München

+49 8161 71 2912

wilfried.schwab@tum.de

4GENE

+49 157 86415814

heimo.adamski@4gene.de

This is a commercial article that will appear in SciTech Europa Quarterly issue 28, which will be published in September, 2018.

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