Journal of Applied Botany and Food Quality 86, 37 - 46 (2013), DOI:10.5073/JABFQ.2013.086.006 1Julius Kühn-Institute (JKI), Federal Research Centre for Cultivated Plants, Institute for Ecological Chemistry, Plant Analysis and Stored Product Protection, Quedlinburg, Germany 2Hansabred GmbH & Co. KG, Dresden, Germany Diversity of volatile patterns in sixteen Fragaria vesca L. accessions in comparison to cultivars of Fragaria ×ananassa D. Ulrich1*, K. Olbricht 2 (Received April 4, 2013) Summary of the latter was described as much more sweetish-aromatic than those of the F. ×ananassa cultivars but with some astringent and Fragaria vesca is the most distributed wild species in the genus bitter impressions (ULRICH et al., 2007). F. vesca is characterized by Fragaria. Due to this biogeography, a high diversity is to expect. outstanding flowery notes like violet and acacia. But especially in During two harvest seasons, sixteen accessions from different lo- the white mutant F. vesca f. alba (Ehrh.) Staudt, these impressions cations from the most eastern habitat at Lake Baikal in Siberia, from sometimes were described by the testers with negative statements Middle and Southern Europe and Northern Europe with Scandinavia like over-aromatic and perfume-like. By gas chromatography- and Iceland were investigated as well as two of the three described olfactometry (GCO) experiments, the flowery impressions were North American subspecies and three F. vesca cultivars. Five very assigned to the content of the aromatic ester methyl anthranilate distinct European F. ×ananassa cultivars were chosen to serve as a whereas the herbaceous impressions are caused by a high content comparison. Beside brix value and acid contents, the aroma patterns of terpenoids. including 67 volatile compounds were quantified by stir bar sorptive Fragaria vesca always was involved in breeding experiments. extraction followed by gas chromatography with mass spectrometric Since the typical high aromatic flavor was missed in the F. detection. The diversity of important volatiles of Fragaria vesca and ×ananassa cultivars, cross-breeding started already in 1918 by F. ×ananassa is discussed regarding biogeography, domestication, Elisabeth Schiemann (KUCKUCK , 1980). Successful ways of species breeding and the so-called funnel effect. introgression with the cultivated strawberry using cross-breeding and chromosome doubling were first described by SCOTT (1951). Introduction Hereby, decaploid fertile clones could be obtained. The first deca- ploid cultivars were introduced under the denomination Fragaria The wild strawberry Fragaria vesca L. (wood strawberry) is an old × vescana by BAUER and BAUER (1979), combining characteristics food, ornamental, and medicinal plant. Already the Roman poet Ovid of F. vesca and F. ×ananassa. Synthetic octoploids were obtained mentioned Fragaria twice and very likely meant Fragaria vesca from the inclusion of Fragaria vesca in the breeding pedigree by (DARROW , 1966). In the Middle Ages, the wood strawberry became EvAN S (1977) and BOR S and SULLIVAN (2005). Nowadays, breeding a Christian symbol and is to find in many paintings from that time. programmes with F. vesca focus on the inheritance of methyl Fragaria vesca is the most widely distributed species of the genus anthranilate and several resistances (PINKER et al., 2012). Fragaria and appears circumpolar throughout Europe, Northern The aroma of the wood strawberry was investigated in detail for the America, and Northern Asia. Since the Middle Ages, the so-called first time by DRAWERT et al. (1973). The content of 40 identified ‘semperflorens’ or ‘alpine’ forms of F. vesca ssp. vesca have been main peaks in F. vesca exceeds those of the F. ×ananassa cultivar cultivated in European gardens (SHULAEV , 2010) together with cv. 'Revata' by the factor of 4.7, which is in agreement with later F. moschata L. and F. viridis Duch., two other wild species occur- findings of ULRICH et al. (1997) (factor 3.2 in comparison to ring in European forests. After the 1750’s the cultivated wild-types cv. 'Elsanta'). Additionally, AHARONI et al. (2004) discussed the were replaced by the garden strawberry (F. ×ananassa Duch.) with differences in the patterns of volatile organic compounds (VOCs) its bigger fruits and a unique pleasant flavor. between F. vesca and F. ×ananassa. In this paper the differences F. vesca (2n = 2x = 14) has a very small genome of 240 Mb. As are interpreted as a gain and loss of metabolic diversity caused by a diploid herbaceous plant with remontant behavior in case of the domestication and breeding. In plant breeding this process also is forma semperflorens, it became the model plant for molecular studies known as funnel effect (ULRICH et al., 1997; ULRICH and OL B RICHT , in the Rose family (Rosaceae) (HOLLEN D ER et al., 2012). In 2010, 2011) or bottlenecking (DOE B LEY et al., 2006). A general shortcome genome sequencing was completed for a Hawaiian accession of the of several published investigations of VOC profiles is the neglect wood strawberry (SHULAEV et al., 2010). of the environmentally caused dynamic of the volatile patterns. It The octoploid F. ×ananassa Duch. (2n = 8x = 56), which derives is unquestioned that beside genetic prerequisites outer factors like from a spontaneous hybridization of the two octoploids F. chiloensis maturity at harvest, location, climate, irrigation, harvest time as well (L.) Miller and F. virginiana Miller, is not a direct descendant of as the attack by pathogens have an eminent influence on biosynthetic F. vesca. However, genome research revealed that the two parental network and hence on the development of the whole volatile patterns species of the garden strawberry derived from different diploid (quantitative and qualitative!) (FORNEY et al., 2000; AZO D ANLOU ancestors including F. vesca. (ROU ss EAU -GUEUTIN et al., 2009; et al., 2004; OL B RICHT et al., 2011). To quote an example, JOUQUAN D SHULAEV et al., 2010). et al. (2008) observed changes in ester contents of fruit from Since the Middle Ages, wild-type strawberries were praised for their cultivated strawberries between two harvest dates of the same season intense flavor (DARROW , 2005). The aroma of wild strawberry is up to 350 %. This volatile dynamic was in the same order like those perceived as very aromatic and more herbaceous than those of F. caused by the cultivars. Consequently, the use of samples from only ×ananassa cultivars (HIRVI and HONKANEN , 1982). Therefore, few or even one harvest date gives results which represent only a F. vesca serves as a comparison with regard to flavor. In direct ‘snapshot’ of the possible dynamic of metabolite patterns. comparison of cultivars and wild-types, the sensory impression Therefore, the topic of this investigation is to study the diversity of volatile patterns in a collection of 16 F. vesca accessions in * Corresponding author comparison to selected F. ×ananassa cultivars in two harvest years 38 D. Ulrich, K. Olbricht by using a sampling strategy which prevents the influence of maturity each F. ×ananassa cultivar were harvested (Fig. 2). The weight per and harvest date during one season. F. vesca accession varied depending on the fruit size (for example between 115 g with 198 fruits for 'Multiplex' and 1054 g with 902 fruits for 'Red Wonder' in 2012). All fruits were immediately Materials and methods frozen at minus 20 °C after the sepals were removed. Plant material and cultivation Fragaria vesca is the most distributed wild species of the genus and exists with four subspecies, six formae and several cultivars Extraction of fruit volatiles by immersion stir bar sorptive (STAU D T , 1962; STAU D T , 1989). Due to this biogeography, a high extraction (imm-SBSE) diversity is to expect. Therefore, 16 accessions from different To prepare an enzyme inhibited strawberry juice, all frozen fruits of locations from the most eastern habitat at Lake Baikal in Siberia, one accession or cultivar of the whole season were pooled. One mass from Middle and Southern Europe and Northern Europe with part of fruits without sepals were homogenized in one volume part Scandinavia and Iceland were investigated as well as two of the three of a solution of 18.6 % (m/v) NaCl by a household mixer for 2 min. described North American subspecies and three cultivars (Fig. 1). The homogenate was centrifuged 4000 rpm for 30 min. One hundred The used accessions are listed in Tab. 1. Five very distinct European milliliter of the supernatant were mixed with 10 μl internal standard F. ×ananassa cultivars were chosen to serve as a comparison. They (0.1 % (v/v) 2,6-dimethyl-5-hepten-2-ol dissolved in ethanol). For were selected with regard to their breeding history and demonstrate each sample, three head-space vials containing 3 g NaCl each for with different aroma patterns the process of domestication over a saturation were filled with 10 ml of the supernatant, sealed with period of nearly 80 years. magnetic crimp caps including septum, and stored at 4 °C for up to All Fragaria vesca accessions were cultivated in the field at three weeks. Dresden-Weixdorf, Germany, longitude 13.8°, latitude 51.145°, An aliquot of 8 ml of the saturated homogenate but without the solid 200 m above sea level, on sandy loam on gravel ground (German NaCl deposit was transferred in an empty glass vial for volatile classification: 'Ackerzahl' 37). Each accession was growing in a isolation by SBSE. A stir bar with 0.5 mm film thickness and 10 mm quadrate of 1 sqm containing plants of different ages according to length coated with polydimethysiloxan (PDMS) was placed in the their natural habitats. Neither chemical treatment nor extra fertilizer liquid (Gerstel, Mülheim an der Ruhr, Germany). The stir bar was were applied, except for two applications of insecticide during the moved at 350 rpm at room temperature for 45 min. After removal time of flowering in order to prevent damage by the strawberry from the strawberry juice, the stir bar was rinsed with purified water, weevil (Anthonomus signatus Say) in both years.