Mono and sesquiterpenic compounds reported in
Plant secondary metabolites are synthesized for their protection and regulation purposes. Quite often, due to their properties, these metabolites have relevant organoleptic and biological properties and can play important roles in human health and general well-being. A relevant case study in this context is berries and flowers from Sambucus nigra L., which have been used for generations in folk medicine. Although those effects are mainly linked to phenolic compounds, mono and sesquiterpenic secondary metabolites may also play a key role. Despite their potential, S. nigra mono and sesquiterpenic compounds are yet largely unexplored. Complex and dynamic external and internal plant-related phenomena deeply affect terpenes profile, as metabolism, abiotic and biotic stresses, and understanding these phenomena is the first step for S. nigra berries and flowers’ valuation. This chapter will cover aspects linked to elder plant uses, mono and sesquiterpenic composition, and the influence of preharvest and postharvest effects over these metabolites. This knowledge is crucial for scientists and industries to understand and improve the quality of S. nigra-based products.
- Sambucus nigra L.
- secondary metabolites
- mono and sesquiterpenic compounds
A growing interest in the exploitation of natural products as sources of bioactive compounds with potential health benefits has been observed, particularly in the consumption of plant-based products that are able to prevent, ameliorate, or even treat chronic diseases with increasing incidence in the advent of the twenty-first century . The elderberry plant (
Understanding those effects could be of extreme importance, given that mono and sesquiterpenic-based extracts are commercially important, namely for pharmaceutical/nutraceutical, agronomic, food, sanitary, and cosmetic industries. For instance, limonene and linalool, two of the most used monoterpenic compounds, are often employed in perfumes, creams, soaps, as flavor additives for food, as fragrances for household cleaning products and as industrial solvents . In the particular case of potential health benefits-related applications, these compounds have been reported as exhibiting hepatoprotection , anti-inflammatory , analgesic , and antioxidant  activities, among others . These effects are strongly dose-dependent, which reinforce the need to study in detail the terpenic composition of
The challenge to understand the impact of pre- and postharvest processes over the mono and sesquiterpenic compounds is the first step to further establish approaches that can control the variables that have a significant effect over these processes. Hence, the present chapter is devoted to a detailed discussion of
2. Elderberry and elderflower applications
Flowers and berries from
This species has gained attention due to its diverse uses, assimilating in markets from food and herbal industries. In 2010,
One of the main uses of elderberries is the production of natural food colorants, juices, and concentrates, due to their high content in phenolic compounds . Those are also exploited for the formulations of decoctions [2, 19], infusions , juice and syrup/concentrate , extracts, supplements, pies, ice creams, jellies, juices, beverages, beers, wines, liqueurs, and fruit bars [27, 28, 29]. In addition to color, flavor (taste and aroma) is also an important parameter in the consumer perception and product acceptance . Due to the pleasant and characteristic floral aroma, elderflowers are often used as flavoring agents  for the preparation of infusions, decoctions [19, 20, 21, 22], pastry products , nonalcoholic cordials, and fermented beverages [18, 23]. Elderflowers are characterized by an intense, pleasant, and characteristic aroma, currently named as elderflower aroma [32, 33]. Despite the role of esters, alcohols, and aldehydes, monoterpenes, as limonene, terpinolene, and terpinene, present a relevant contribution for the elderflowers fruitiness aroma , and more exotic notes, such as woody and spicy, have been attributed to some mono and sesquiterpenic compounds [32, 34].
S. nigramono and sesquiterpenic metabolites composition
Terpenic compounds form a large and structurally diverse family of secondary metabolites derived from C5 isoprene units, with over 35,000 known structures . The volatile and semi-volatile ones, that is, mono and sesquiterpenic compounds, result from two main biosynthetic routes, starting from the mevalonate and the methylerythritol phosphate pathways (Figure 1). These are produced through the activity of a large family of enzymes, the mono and sesquiterpene synthases and cyclases, but others are formed through transformation of the initial products by acylation, dehydrogenation, oxidation, and other reaction types, such as acetylation [35, 36]. For instance, in Figure 1, illustrates the biosynthesis of linalool, caryophyllene, and humulene, three compounds present in
Information about mono and sesquiterpenic compounds from elderflowers and elderberries is still scarce and disperse. Thus, to systematize this data, the information related with ripe berries and fresh flowers and minimally processed products, such as infusions, syrups, and juices, is presented in Table 1. The reported studies are mainly focused on analytes’ identification rather than on their quantification, however, when available, quantitative data is also provided.
|3-Carene||✓||✓||[31, 38, 39]|
|✓||—||[31, 32, 37]|
|d-Limoneneb||✓||2.24–9.92||[31, 32, 37, 38, 41, 42, 43]|
|Myrcene||✓||✓||[31, 37, 38, 42]|
|Ocimeneb||✓||1.55–9.32||[31, 32, 34, 41]|
|✓||✓||[32, 34, 37, 41]|
|✓||✓||[31, 37, 38]|
|✓||✓||[32, 34, 41]|
|✓||✓||[31, 32, 34, 41]|
|Terpinolene||✓||✓||[31, 32, 37, 38]|
|Camphor||✓||✓||[37, 41, 42]|
|1,8-Cineole||✓||✓||[31, 34, 41, 42]|
|Citral||✓||✓||[31, 37, 41]|
|Citronellol||✓||✓||[34, 37, 42, 44]|
|Geraniolb||✓||1.05–7.21||[31, 32, 37, 41]|
|Hotrienolb||✓||2.56–8.08||[31, 34, 37, 40, 41, 42, 43, 44]|
|Linaloolb||✓||1.18–128.89||[31, 37, 40, 41, 42, 43, 45]|
|✓||✓||[31, 37, 40, 42, 44]|
|✓||✓||[31, 37, 40, 42, 44]|
|✓||—||[31, 32, 34, 40, 42, 44]|
|✓||—||[31, 32, 34, 40, 42, 44]|
|Linalool methyl ether||✓||—|||
|Menthol||✓||✓||[37, 41, 42]|
|Nerol||✓||✓||[31, 32, 37, 41]|
|Nerol oxideb||✓||1.02–7.80||[31, 34, 41, 42, 45]|
|✓||✓||[31, 34, 37, 40, 41, 42, 45]|
|✓||1.68–8.34||[31, 34, 37, 40, 41, 42, 45]|
|✓||70.85–2699.56||[31, 37, 41, 42, 45]|
|Terpinen-4-ol||✓||✓||[31, 32, 37, 41]|
|✓||✓||[31, 32, 34, 37]|
|✓||✓||[31, 37, 42]|
So far, 89 mono and sesquiterpenic compounds are reported in elderflowers (64) and elderberries (61). Recent studies using an advanced gas chromatographic methodology (comprehensive two-dimensional gas chromatography coupled with time-of-flight mass spectrometry detection—GC × GC-ToFMS) have contributed to substantially increase the knowledge about
As evidenced in Table 1, of the 64 volatile terpenic compounds reported from elderflowers, 40 are oxygen-containing structures. As shown in Figure 2, the peak intensities of the monoterpenic metabolites predominate, representing up to 99 and 77% of the overall elderflowers and elderberries terpenic content, respectively [31, 37]. Linalool oxide (in the pyranoid form) is a major component from fresh elderflowers, accounting for up to 87% (relative to the overall GC peak area) . Other authors reported that hotrienol (14%, w/w), rose oxide (5%, w/w), linalool (4%, w/w), and linalool oxide (furanic forms, 3%, w/w) were the major monoterpenic metabolites from dried elderflowers  (chemical structures illustrated in Figure 3).
Regarding ripe elderberries, limonene and
4. Factors that modulate mono and sesquiterpenic profile
4.1. Preharvest impact
Crop quality could be defined as a set of agronomic/commercial, organoleptic, and nutritional qualities that are variable among (1) distinct species but also among different cultivars within the same species (genetic factors); (2) different climatic conditions, such as water availability and light exposition; and (3) different agronomic conditions, such as cultivation systems, fertilization, and harvesting date . Altogether, these preharvest factors may have an impact on the final quality of the elderberry fruits and flowers; however, the information about these effects is scarce. The impact of preharvest factors is often focused on parameters with direct agronomic and commercial relevance, as plant yield, fruit size, sugar content and acidity (e.g., reviews on
The production of terpenic metabolites depends on the physiological and developmental stage of the plant [10, 37]. Fruit ripening, in particular, is a crucial phenomenon that affects different physiological and biochemical processes, which are determinant to the development of nutritional and organoleptic characteristics . The fruit organoleptic characteristics such as taste, color, and aroma are important quality and consumer acceptance-determining features [30, 49].
During ripening, major events occur, including cell expansion and softening, dismantling of the photosynthetic apparatus, and degradation of chlorophyll . Elderberry ripening takes place from the 1 to 2-month period, starting with a green appearance and they ripen over a period of 6–8 weeks from July to September (depending on the geographic location). When elderberries become ripe, they have a characteristic deep purple color . The accumulation of sugars (expressed as total soluble solids [TSS]) and decrease in acidity (pH and titratable acidity [TA]) have been routinely used by growers as a decision-making parameter to establish the harvesting moment and even the commercial price of the berries [18, 23, 37]. The ripe elderberries’ pH ranges from 3.8 to 4.8; TA ranges from 0.48 to 1.43 g citric acid/100 g FW berries, while TSS ranges from 10.1 to 17.5°Brix [37, 38, 50, 51]. Figure 4 illustrates the impact of ripening in those tree parameters on elderberries harvested in a Portuguese location (Tarouca, Távora and Varosa Valley), in the harvest season of 2013.
During the ripening process, several other phenomena occur, namely biosynthesis and degradation of a wide range of secondary metabolites that may have direct relevance in elderberry sensorial characteristics. A recent metabolomics-based study that exploited the effects of the developmental stages of different cultivars on the volatile terpenic components  demonstrated that the variability of monoterpenic compounds (
Plant cultivars generally differ in yield, organoleptic, and nutritional characteristics [23, 46], and their genetic background is a factor that influences quality traits . In the particular case of elderberries, cultivars are classified based on their morphological characteristics and yield , as no definitive taxonomic DNA-based studies have been conducted in this species. Although, efforts have been made for their classification with molecular data. For instance, Portuguese
It is reported that elderberry yield ranges anywhere from 1 to over 30 kg per bush, depending on cultivar [54, 55]. This aspect, together with the fact that several cultivars are nowadays explored for the formulation of various products, where formula standardization is required, implying the comparison of cultivars’ composition, can play a significant role in their application (e.g., ) and then become important decision tool for producers. The fact that mono and sesquiterpenic synthesis is encoded by a variety or cultivar-related genes implies that their levels can be cultivar-dependent, which, on the one hand, might be used to trace its varietal origin  and, on the other hand, can be used to better manage their final product and to maximize the commercial value of the crop. An exploratory study, suggested a possible cultivar effect over the mono and sesquiterpenic compounds profile from fresh elderflowers ; however, more consolidated data is still required to sustain the stated remarks, namely in what concerns the number of analyzed samples and different harvesting years.
The specific cultivar metabolite profile may imply differences at the sensorial level in
Despite the studies reported earlier, a more comprehensive understanding of the influence of preharvest parameters will require their analysis in an integrated approach, including, among others, climate, agricultural practices, soil, and harvesting year to fully understand how these affect the biochemical mechanisms involved in the formation of mono and sesquiterpenic metabolites from elderflowers and berries and also to improve its valorization potential, particularly when related with health benefits and relevant sensorial characteristics. Also, the influence of climate change on the
4.2. Postharvest impact
Postharvest management includes a set of postproduction practices comprising, among others, cleaning to eliminate undesirable elements and improve product appearance, sorting, cooling, control of variables such as temperature and relative humidity, and packing, ensuring that the product complies with the established quality standards for fresh and processed products [58, 59]. Postharvest practices may deeply affect the quality of a product in many aspects such as chemical and sensorial characteristics but also their potential health benefits, and ultimately, it may affect product’s acceptability and marketability . Therefore, reliable and objective quality-control tools to measure the impact of postharvest practices (ideally integrated with preharvesting conditions) over product quality and in the present appraisal on sensory quality are essential.
Elderflowers and elderberries go through different postharvest handling and storage conditions that precede processing, to prepare stable formulations for commercialization. Figure 5 illustrates the main steps from harvesting for the storage of elderberries and elderflowers and the main chemical changes that may occur throughout these processes [31, 33, 42, 60, 61, 62, 63].
The knowledge of the impact of handling and storage conditions on the terpenic metabolites of
After 1 year of storage, a decrease of the total terpenic content up to 47% for frozen elderflowers; up to 67 and 71% when vacuum packed and kept under light exposure and without light exposure, respectively; up to 82% for air-dryed elderflowers; and up to 85% for freeze-dried elderflowers (Figure 6) . Under vacuum packing, there was no significant impact from light exposure. Linalool oxides were suggested as markers of the impact of the studied postharvest conditions over the volatile terpenic metabolites of elderflowers .
Drying methodologies, as air-drying or freeze-drying, often fail to completely preserve volatile aroma compounds , as reported in dried elderflowers, mainly due to diffusion and evaporation losses [31, 33, 69, 70]. Drying of elderberries or their products promotes a water activity reduction, contributing to the preservation of the samples against microbial contamination and also decreases the degradation of anthocyanins , by increasing their stability . Other strategies have been used to preserve the elderberries’ bioactive components or to enhance their nutritional value, as for instance, their processing with pulsed ultraviolet light to enhance the phenolic content . However, no studies were performed so far on mono and sesquiterpenic fractions of elderberries.
Storage time also plays an important role in the mono and sesquiterpenic composition illustrated by the fact that 15 compounds, including rose oxides, hotrienol, linalool,
Some components, such as hotrienol, were observed to increase during storage of elderflowers, which could be associated with the action of enzymes, such as glucosidases, that unbound the volatile components from glycosides present in the matrix . Non-oxygen-containing structures, that is, monoterpenes and sesquiterpenes, also increased under certain postharvest conditions (Figure 6), again assuming that
The modifications in
5. Concluding remarks
Plant secondary metabolites play key role in the plants’ protection and communication processes. Beyond that, these components, and particularly, mono and sesquiterpenic compounds, are nowadays explored in industrial sectors due to their pleasant aroma characteristics and potential on the prevention and management of human diseases. The exploitation of
Thanks to FCT/MEC for the financial support to the QOPNA Research Unit (FCT UID/QUI/00062/2013), through national funds and where applicable co-financed by the FEDER, within the PT2020 Partnership Agreement. This work was developed within the scope of the project CICECO - Aveiro Institute of Materials, POCI-01-0145-FEDER-007679 (FCT Ref. UID/CTM/50011/2013), financed by national funds through the FCT/MEC and when appropriate co-financed by FEDER under the PT2020 Partnership Agreement. Â. Salvador thanks the grant AgroForWealth: Biorefining of agricultural and forest by-products and wastes: integrated strategic for valorization of resources toward society wealth and sustainability (CENTRO-01-0145-FEDER-000001), funded by Centro2020, through FEDER and PT2020, and Operational Group Sambucus Valor: integrated valorization of elderberry plant according to the patterns of healthy consumption: from the plant to the formulation of new value-added food products, funded by PDR 2020, Measure 1, “Innovation,” Formation of Operational Groups (PDR2020-101-031117, Partnership no146/Initiative no341).