Current Bioactive Compounds 2017, 13, 3-17

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REVIEW ARTICLE

ISSN: 1573-4072 eISSN: 1875-6646 Realizing the Potential of Health-Promoting Rosehips from Dogroses (Rosa sect. Caninae)

Hilde Nybom* and Gun Werlemark

Balsgård, Department of Plant Breeding, Swedish University of Agricultural Sciences, Fjälkestadsvägen 459, SE29194 Kristianstad, Sweden

Abstract: Systematics and Genetics: Medicinal effects of rosehips are attributed mainly to dogroses, i.e. species in Rosa sect. Caninae. These are mostly pentaploid and have a unique meiosis with asymmetrical chromosome distribution. Offspring in reciprocal crosses differ widely since seedlings resemble the seed parent more than the pollen parent.

Chemical Contents: Flesh (shells) of rosehips from dogroses contain high levels of antioxidants, mainly polyphenols and ascorbic acid, as well as carotenoids and vitamins B and E. The seeds are rich in polyunsaturated fatty acids and are used for skin treatment and cosmetics. Both shells and seeds, separately or A R T I C L E H I S T O R Y together, are used for health-promoting food and food additives.

Received: March 9, 2016 Revised: May 25, 2016 Medicinal Effects: The most well-documented studies concern the positive effects on inflammatory proc- Accepted: May 31, 2016 esses, especially in patients suffering from osteoarthritis. The exact medicinal role of different rosehip DOI: compounds have not been investigated in detail, and the relative importance of fruit flesh versus seed is 10.2174/15734072126661606070906 35 also not yet determined.

Plant Production and Breeding: Rosehips are mainly harvested by hand from wild or naturalized dogrose bushes. Once the key compounds have been identified, commercial output of rosehips with pre- dictable quality would be facilitated by careful optimization of plant material, cultivation conditions, and post-harvest and processing procedures. Some cultivars have been bred in Europe but breeding goals have not been clearly defined and commercial production is limited. Modern genomic tools have improved breeding programs in many crops but polyploidy and the canina meiosis present considerable challenges for research and breeding in dogroses. Keywords: Antioxidant, functional food, medicinal plant, phytotherapy, plant breeding, plant taxonomy.

1. INTRODUCTION The well-known genus Rosa contains numerous species of commercial interest. Several wild species have contributed to the highly appreciated and economically important ornamental rose cultivars, and, in the case of R. damascena, also to the rose oil industry. Other rose species have remained more anonymous but some have considerable potential due to their fruits: the rosehips. Commercially marketed rosehips are obtained from several different species. One important group is the dogroses, all of which belong to the section Caninae (Fig. 1). These are long-lived woody peren- nials, found mainly in woodland margins and in disturbed habitats such as roadsides and open pastures.

*Address correspondence to this author at the Balsgård, Department of Plant Breeding, Swedish University of Agricultural Sciences, Fjälkestadsvägen 459, SE29194 Kristianstad, Sweden; Tel: +46 703 444 369; E-mail: Fig. (1). Rosehips of the dogrose species Rosa dumalis. Photo Staf- [email protected] fan Andersson.

Dogroses were sometimes cultivated in the Middle Ages difficult group to classify. Numerous dogrose taxa have been for use as medicinal plants, using almost all parts of the defined according to morphological characters such as leaflet plants: rosehips, seeds, petals, leaves and roots. Presently, shape, hip shape, length of pedicel, presence or absence of dogroses are sometimes planted as ornamentals, especially in glandular hair, shape of prickles, persistence of sepals, shape public areas where they are appreciated for their hardiness of style head orifice, leaf pubescence, and plant shape. At the and soundness. In addition, dogroses are used as rootstocks end of the 19th century, the total number of taxa described, for the budding of ornamental roses, but they are gradually including varieties, reached 4266 [2]. being replaced by rootstocks derived from other species with Section Caninae is divided into a number of subsections; superior characteristics. In the last two decades, modern sci- Trachyphyllae with R. marginata (syn. R. jundzillii) as the ence has, however, shown that rosehips from dogroses con- only species, Rubrifoliae with R. glauca (syn. R. rubrifolia) tain compounds of interest for functional food and nutraceu- ticals as well as for medicinal drugs. Efforts are now being as the only species, Vestitae with, e.g., R. mollis, R. sherardii, R. tomentosa and R. villosa (syn. R. pomifera), Ru- made to identify the active ingredients and their mode of bigineae with, e.g., R. agrestis, R. inodora, R. micrantha and action. R. rubiginosa, Tomentellae with R. balsamica and R. abiet- Objectives of this review include: ina, and Caninae with, e.g., R. canina, R. corymbifera, R. dumalis, R. montana and R. stylosa. Occasionally, R. stylosa 1. An overview of the complex taxonomy of dogroses is instead treated within its own subsection, Stylosae. since proper determination of the analysed plant material is essential for future optimization of commercially 2.2. DNA-Based Dogrose Taxonomy manufactured and marketed products, Morphological characters are generally insufficient for 2. An insight into the unique transmittal of chromosomes, unambiguous classification of dogroses, even when applied and therefore also traits, from parents to offspring since to plants in comparative garden trials [3, 4]. In dogroses, this has profound effects on the origination of dogrose many taxa have, however, turned out to be even more diffi- plants and their use, cult to distinguish when analysed with DNA markers, and 3. An account of chemical compounds with a potential samples from closely related species often overlap to a high role in the health-promoting effects of rosehips, degree. In one AFLP-based study conducted on >900 dogrose specimens sampled in a transect across Europe, the 4. Short descriptions of some of the studies that have re- three major subsections Caninae, Rubigineae and Vestitae ported medicinal effects of dogroses, and were clearly discriminated [5]. Some support was also indi- 5. Efforts to link the above-mentioned research areas, and cated for the smaller subsections Trachyphyllae, Rubrifoliae suggest how improvements towards well-informed and Stylosae, all of which have been hypothesized to derive utilization of rosehips can be achieved. from crosses between dogroses and taxa in other sections. Within the subsections, many species overlapped consid- 2. ROSE TAXONOMY erably. In dogroses, geographic distances often appear to be Proper inventorizing and evaluation of genetic resources at least as important as morphology-based classification in are a cornerstone in plant domestication. For the develop- explaining genotypic similarities between analyzed speci- ment of rosehip-based health-promoting products, potentially mens [4, 5]. Possibly, such species have arisen multiple valuable species should be collected and analysed. The genus times in multiple locations and are, therefore, polyphyletic. Rosa is however quite complex, and delimitation of sections In another study, AFLP and DNA microsatellite (SSR) and species is controversial. Numerous taxa have been de- markers were applied to discriminate among three species scribed at different ploidy levels, and inter-taxon hybridiza- within the subsection Vestitae [6]. Again, considerable over- tion is common. Although a modern classification of roses is lap was reported but clustering of the samples reflected dif- needed, the well-known Rehder system updated by Wisse- ferences in ploidy level (all species contained both tetra- and mann [1] is still much used. Three small subgenera, Hulthe- pentaploid specimens) more than in species affiliation or mia, Hesperhodos and Plathyrhodon, and one large, Rosa, geographical origin. are recognized. 3. CYTOLOGY AND REPRODUCTION Subgenus Rosa is further subdivided into 10 different sections. One of the largest is section Caninae, dogroses, The basic chromosome number in the genus Rosa is x = with approximately 50 species. The other sections are 7. Diploid rose species have two sets of chromosomes, 2n = Pimpinellifoliae (15 species), Gallicanae (one species and 2x = 14. In meiosis, these 14 chromosomes pair up to form several hybrid taxa), Carolinae (about five species), Rosa seven bivalents as expected, and inheritance is asymmetri- (about 80 species, formerly known as section Cinna- cally biparental. Many rose species as well as almost all or- momeae), Synstylae (about 25 species), Indicae (three spe- namental cultivars are instead tetraploid, with 2n = 4x = 28. cies), Banksianae (possibly two species), Laevigatae (one These can form bivalents or multivalents, but inheritance is species) and Bracteatae (one or two species). still symmetrically biparental.

2.1. Classical Dogrose Taxonomy 3.1. Canina Meiosis Medicinal effects of rosehips are mostly ascribed to the Dogroses are usually pentaploid (2n = 5x = 35) although dogroses but these have unfortunately proved to be the most some are tetraploid or hexaploid. All dogroses have peculiar Health-Promoting Rosehips Current Bioactive Compounds 2017, Vol. 13, No. 1 5 and asymmetric canina meiosis, first described in a cytologi- cal study [7] and later verified (e.g. [8]). Regardless of Seed parent ploidy level, only seven bivalents are formed in the first meiotic division (Fig. 2). The remaining chromosomes occur exclusively as univalents. These univalents are not included in viable pollen grains, which contain only the seven divided bivalent chromosomes. In contrast, all univalents are transmitted to one of the daughter cells in the female meiosis, and finally included in the viable egg cells, which therefore con- New pentaploid offspring tain 21, 28 or 35 chromosomes depending on the ploidy level. The resulting seedlings obtain the full chromosome number but inheritance is asymmetric: only 15–25% (depending on ploidy level) of the chromosomes are inherited from the pollen parent, whereas the remainder are obtained from the seed parent (Fig. 3). Consequently, inheritance is matroclinal, i.e. offspring is more similar to the maternal Pollen parent parent than to the paternal parent [9, 10]. Fig. (3). Origination of a new offspring with two pentaploid dogrose plants as parents. Each parent has 5 x 7 chromosomes, with M1 the two bivalent-forming chromosomes shown to the left in each quintuplet. Chromosomes transmitted by the seed parent and by the A1 pollen parent, respectively, are shown within the rectangle. Draw- ing Hilde Nybom.

M2 Application of MAC-PR has revealed the transmittal of different chromosomes in dogroses, by pollen cells and egg cells, respectively [13, 14]. Bivalent formation in meiosis A2 takes place between seven pairs of chromosomes. The two chromosomes in a pair consistently share the same SSR alleles, and a very homogenous set is therefore transmitted by X both pollen cells and egg cells to all offspring. By contrast, XX X X the maternally transmitted univalent chromosomes carry more variable SSR alleles. When only bivalent-residing alleles are taken into consideration, similarity among geno- Fig. (2). Meiosis in a tetraploid dogrose; female on the left side and types is relatively constant regardless of whether these geno- male on the right side. M1 metaphase 1, A1 anaphase 1, M2 meta- types belong to the same species or not. By contrast, univa- phase 2, A2 anaphase 2. On the female side, the two alternative lent-allele variation is more closely associated with pre- developments of daughter cells resulting from the first meiotic divi- sumed levels of relatedness among genotypes. sion are shown, leading to viable and non-viable egg cells, respectively. Chromosomes from the bivalent-forming genomes in grey 3.3. Implications of the Canina Meiosis and chromosomes from the univalent-forming genomes in red. Drawing Hilde Nybom. SSR-based analyses of dogroses suggest that the genomic configuration can be interpreted as each species having one homogenous, bivalent-forming diploid genome and two, three or four univalent-forming haploid genomes depending 3.2. Chromosome Transmittal on ploidy level [13, 14]. A maximum of four different alleles DNA fingerprint bands obtained with single-locus meth- in pentaploid dogroses, with one allele occurring in two cop- ods like SSR analysis are easily scored in diploids, and can ies (presumably residing on the bivalent-forming chromo- be traced through several generations of parents and off- somes), has been reported for the single-copy genes LEAFY spring [11]. In polyploid plants, one primer pair often pro- and cGAPDH and for one ribosomal locus (nrITS), suggest- duces multiple bands, and scoring is less straight-forward. ing that the SSR-based model is valid also for other parts of To fully utilize the information content of these DNA pro- the genome [15]. files, segregation patterns must be determined. This requires Sequencing of the nrDNA internal transcribed spacer the ability to score allele dosage in contrast to just the pres- (nrITS-1) region has revealed five major groups of se- ence or absence of an allele. The MAC-PR approach (micro- quences in the genus Rosa [16]. One of these groups, C-type, satellite DNA allele counting - peak ratios) determines allele occurs only in the dogroses, while three other groups are copy number based on quantitative differences between mi- found in both dogroses and species belonging to other sec- crosatellite allele peak ratios, and therefore allows the pre- tions. In another sequencing study, five different rDNA gene cise determination of allelic configuration in each studied families were identified from extensive cloning analysis of sample [12]. the nrITS-1 region [17]. The -family, shown to be identical 6 Current Bioactive Compounds 2017, Vol. 13, No. 1 Nybom and Werlemark with the C-type group, occurred in all three investigated 4. CHEMICAL CONTENTS dogrose species whereas the other four gene families oc- The rosehip is an accessory fruit (pseudofruit) originating curred in one or two species each. Thus a putative ‘proto- from a single flower with numerous pistils. After fertiliza- canina’ rDNA type is hypothesized to correspond to the - tion, each pistil develops into a separate achene (a monocar- (C-type) family found in modern species. pellate, indehiscent fruit), which contains a single seed inside Implications of the canina meiosis have been investigated a hard coat. The achenes are often termed ‘seeds’ even in in studies on possible genome-dependent differences in gene scientific literature. Usually 15–40 achenes occur together transcription. Analysis of three dogrose species using RT- inside an urn-shaped structure with fleshy, red-coloured CAPS (reverse transcription - cleaved amplified polymor- walls. phic sequences) suggests that the -family is considerably Comparison across studies is complicated due to the fact upregulated compared to other rDNA families [18]. By con- that chemical analyses are sometimes conducted on the en- trast, no differences in transcription levels were found be- tire rosehip, and sometimes on just the fleshy parts without tween bivalent- and univalent-residing copies of three genes seeds. Assessments are also affected by the use of fresh, fro- investigated in another study [15]. zen or dried rosehips as well as by the drying method (e.g., An epigenetic influence on the transcription of dogrose at room temperature, in heated air or by freeze-drying) and genes has been reported [19]. Using cDNA-AFLP (amplified by the use of different extraction methods and laboratory fragment length polymorphism of coding DNA) and MSAP protocols. In addition, few authors are aware of the taxo- (methylation-sensitive amplified polymorphism), maternally nomic complexity within section Caninae. The analyzed biased epigenetic patterns could be identified in spontane- material is often reported as belonging to well-known spe- ously occurring, reciprocal dogrose hybrids. Moreover, these cies, like R. canina, although entirely different species patterns correlated more closely with morphological varia- names might have been accorded by a taxonomist. In the tion than with microsatellite DNA variation. The underrepre- review provided here, species names must therefore be sented paternal genome was also transcribed but the percent- treated with discretion. age of paternally expressed fragments differed between plant 4.1. Environmental Influence organs, with a higher amount found in sepals compared to leaves. Both global and local environmental conditions have profound effects on yield and quality of rosehips. Already 3.4. Mating System, Apomixis and Hybridization Melville and Pyke [23] found a correlation between latitude Reproductive system and breeding behavior affect the and level of vitamin C in English rosehips; a colder climate amount and partitioning of genetic variation in wild popula- presumably leads to a higher level of vitamin C. Similarly, an increase in vitamin E content with altitude has been re- tions, and in the offspring of plants used in breeding pro- ported [24]. Variation in chemical contents, e.g. vitamin C, grammes. Dogrose species do not propagate vegetatively, can also stem from year-to-year variation in precipitation, except over very short distances, and thus most plants seen temperature and insolation [25-29] as well as from variation in nature derive from a separate seed. Most polyploid rose in the ripening stage of the analysed rosehips [26, 29, 30]. species, including the dogroses, are self-fertile. The ability to set seed after selfing together with the canina meiosis, leads 4.2. Rosehip Compounds to severely restricted genetic recombination in dogroses. Reactive oxygen and nitrogen species (ROS and RNS, In addition, dogroses can produce seeds without prior respectively) are formed in human bodies as part of normal fertilization of the egg cell, i.e. by apomixis. Since pollina- cellular metabolism and can lead to oxidative stress causing tion is required for triggering the embryo development, serious damage and contributing to, e.g., proliferation of apomixis is pseudogamous. Originally, apomixis in dogroses cancer cells and cardiovascular diseases. Plant cells can, in was hypothesized from the morphological homogeneity of contrast to animal cells, produce antioxidant compounds that dogrose offspring and their similarity with the seed parent. may neutralize the harmful ROS and RNS. Recently, apomictically derived seedlings have been une- quivocally demonstrated with DNA markers in the offspring In a comparative study of many different plant species, from experimental pollinations between different dogrose rosehips from dogroses had four times higher total antioxi- species [13, 14, 20, 21]. The number of apomictically de- dant concentration compared to the second highest, the rived seedlings varied from 0 to 10% in these crosses. The crowberry (Empetrum hermaphroditum) [31]. The antioxi- extent to which apomixis occurs also after intraspecific pol- dant activity of rosehips derives mainly from polyphenols, lination has never been determined, nor its role in the repro- carotenoids and the vitamins B, C and E. Many of the anti- duction of wild populations. oxidants also protect one another from oxidation, leading to synergistic effects. This antioxidative effect prompted an All dogrose taxa appear to be cross-compatible, as is also investigation of the potential of replacing sodium ascorbate true for the whole genus Rosa, and interspecific hybridiza- and sodium nitrite with a rosehip extract as a stabilizer for tion may be responsible for many speciation events in dogro- color and texture in German sausages [32]. The rosehip ex- ses. Thus, SSR-based analysis of the hexaploid dogrose spe- tract protected against protein oxidation but could not fully cies R. micrantha suggests that it has arisen from hybridiza- replace sodium nitrite. tion between R. rubiginosa on one hand, and either R. canina Vitamin C is required for the synthesis of collagen, ten- or R. corymbifera, on the other hand [22]. dons and bones. High levels of vitamin C can prevent growth Health-Promoting Rosehips Current Bioactive Compounds 2017, Vol. 13, No. 1 7 of some cancer cells in vitro and in vivo, while other cells tive role against atherosclerosis and heart diseases [44]. Total remain unaffected [33]. Apparently, vitamin C contributes to polyphenol content in rosehips of Polish dogrose species a cytotoxic effect in cancer cells by acting as a pro-oxidant reached 9.3–15.2% of DW, with flavonols making up the together with an elevated copper level [34]. Amount of vita- major part [45]. Hvattum [46] investigated the composition min C in rosehips ranges from 0.5 up to 6 g/100 g DW de- of flavonoids in R. canina; one anthocyanin, several gly- pending on genotype, species and environmental factors [25, cosides (a molecule with sugar bound to a non-sugargroup) 35-39]. of quercetin and taxifolin, and eriodictyol were identified. Salminen et al. [47] reported that the major phenolic com- Vitamin E comprises several lipid-soluble antioxidant pounds were proanthocyanidin glycosides, while the flavon- compounds. Deficiency of this vitamin can be observed in oid glycosides only occurred in small amounts. Some of the patients with a defect in the -tocopherol transfer protein proanthocyanidin glycosides were larger than trimers, which produced in the liver; the sensory neurons deteriorate caus- is unusual for plants. Olsson et al. [48] measured the amount ing peripheral neuropathy and ultimately death [40]. of flavonoids in different berries, and found only a moderate Whether a supplement of vitamin E can also lower the risk amount of quercetin, 87 g/g DW, in rosehips together with of cardiovascular disease and cancer has been debated [41, 88 g/g DW of quercetin-glycosides. By comparison, lin- 42]. In this context, the tocopherols are of particular interest gonberries, Vaccinium vitis-idaea, had 10 times more quer- since they contribute substantially to vitamin E activity. In cetin and twice as much quercetin-glycosides. But the rose- rosehips of R. rubiginosa and R. dumalis, concentration of - hips also had 222 g/g DW of “other flavonols” whereas tocopherol is approximately 100–150 g/g DW but the con- lingonberries only had 25 g/g DW of such compounds. tent is even higher in the black-fruited R. spinosissima (189.5 g/g DW) in section Pimpinellifoliae [43] (Fig. 4). In One polyphenol of particular interest is the tannin ellagic another study with R. canina from Turkey, content of - acid, which together with quercetin is the most abundant tocopherol was, however, reported to reach only 34 g/g polyphenol in rosehip tea [49], and has been claimed to have DW in the fruit and 8 g/g DW in the seeds [36]. beneficial effects against cancer and cardiovascular diseases. Apparently, ellagic acid acts synergistically with quercetin to induce apoptosis in certain cancer cells [49]. In a study on 14 Polish rose species, rosehips were shown to contain up to 1065 g/g DW of ellagic acid but with considerable interspecific variation [50]). These amounts are lower than the levels found in e.g. raspberries, but comparable to the levels in strawberries. In yet another study, ellagic acid in dogrose leaves reached 19.42 mg/g DW, i.e. almost 20 times higher than in the fruits [45]. The antioxidative effect of rosehips stems mainly from the polyphenols, as demonstrated by testing the ROS activity in R. canina preparations devoid of vitamin C but containing several different phenolics [51]. Gao et al. [52] obtained the same result where comparing the antioxidant effect in phenolic, ascorbic and lipophilic extracts in two different assays. More than 90% of the total activity was due to the phenolic compounds, whereas only 8.6% activity could be ascribed to Fig. (4). Rosehips of Rosa spinosissima. Photo Staffan Andersson. the ascorbic acid and even less to the lipids. They also found that a pure phenolics extract had a higher antioxidative capacity than extracts with both phenolic and ascorbic fractions. Consequently, a strong correlation has been demon- Vitamin B or folate is an essential vitamin required for 9 strated between total phenolics content and the antioxidative DNA and RNA formation, and deficiency can lead to accu- capacity in berries, including rosehips [53]. Although the mulation of e.g. homocysteine which is a risk factor for cardiovascular disease. Impaired cell replication is first noticed differences in total phenolics contents were rather restricted, ranging from 55 GAE (gallic acid equivalent)/g DW (R. vil- in rapidly replicating tissue such as blood and can therefore losa) to 84 GAE/g DW (R. dumalis), a 6-fold difference was cause anaemia. Pregnant females are recommended to eat up found in total antioxidant contents among different dogrose to 400 g/day to ensure proper cellular replication and species [54]. growth of the fetus. Content of folate in rosehips is very high: 400–600 g/100 g DW depending on species [25] but Carotenoids are organic, fat-soluble pigments that pro- sensitive to oxidative degradation, making it vulnerable to duce red to yellow colours in plants; xanthophylls have oxy- losses during post-harvest handling and processing. gen and carotenes (-carotene, -carotene which is a pre- cursor of vitamin A, and lycopene) lack oxygen. The caro- Polyphenols, like flavonoids, tannins and lignin, consti- tenoids also act as antioxidants and apparently have an- tute the most abundant antioxidants in our diet. The plants timutagenic and tumour-preventing properties. The reported produce polyphenols as a defence against parasites, insects content of total carotenoids in dogroses range widey, from and herbivores, and for pigments and flavor of flowers and fruits. The flavonoids inhibit LDL (low density lipoproteins) 25 to 2400 mg/kg [29, 52, 55-57]. Lycopene is one of the more important carotenoids with 162 to 390 mg/kg DW [29, oxidation and platelet aggregation, and may have a protec- 8 Current Bioactive Compounds 2017, Vol. 13, No. 1 Nybom and Werlemark

56] which is more than in many fresh tomatoes. Rosehips possibly due to enzymatic degradation during the long expo- therefore have potential as an important alternative source sure to high moisture conditions [65]. for lycopene in the food industry. Some of the carotenoids Commercial companies use hot-air dryers but most of the increase substantially during fruit maturation but consider- desirable compounds are thermolabile and oxygen sensitive, able differences have been reported between species and between years [29, 58]. and therefore drying results in loss of colour and vitamins, as well as degradation of nutrients [66, 67]. Losses can be re- For consumer acceptance, sugars and volatile aroma duced by careful optimization of the temperature profile re- compounds may also play an important role. Total sugar sulting in about 50–65% retention for ascorbic acid [65, 68]. content in rosehips typically vary from 12 to 18 g/100 g FW For preservation of carotenoids, low temperatures have been [39] or from 20 to 36 g/100 g DW [59]. In one study, glu- reported as more beneficial, presumably because these have cose was the major sugar followed by fructose, and both many double bonds and are susceptible to oxidation en- increased significantly during fruit ripening [60], while con- hanced by, e.g., light and increased temperature [37]. In contents of glucose and fructose were quite even in another trast, Mabellini et al. [65] found little effect of drying tem- study [59]. A large number of volatiles have been docu- perature on the retention of carotenids. mented in the fruits of five rose taxa, four of which belonged The hips are either dried whole or after being cut into to sect. Caninae [59]; a large concentration of alcohols and smaller pieces [68, 69]. Whole-dried rosehips usually swell aldehydes was found in all samples while monoterpenes and sesquiterpenes were more variable. when the drying is initiated since moisture is trapped inside the epidermis. Strålsjö et al. [25] noted a reduction in drying 4.3. Seed Compounds time from 11 h for whole-dried fruit to 1 h and 45 min for cut fruits. In the same study, retention of folate and vitamin Presently, many cosmetic and pharmaceutical industries C benefitted from a high temperature (90°C) with hips being seek to replace synthetic products with natural ingredients, cut into slices. Improved vitamin C retention can also be and a number of traditionally used plants have therefore re- achieved by drying with a CO2 mixture [68] or by freeze- ceived a renewed interest. Rosehip seed oil has long been drying [37]. Vacuum drying at low temperature (60°C) has used in South America for treating all sorts of skin ailments yielded superior retention of antioxidant capacity and con- as well as being used as a cosmetic. Recently, some cold- tent of phenolics when applied to rosehip pulp for production pressed rosehip seed oils have also been introduced to the of fruit leather [70]. food market in Europe. A commonly marketed commodity is rosehip powder. In The oil content of the seeds is 5–18%, depending on spe- Turkey, the fruit is first dried, then crushed and thereafter put cies and extraction method. Among fatty acids in this oil, into hot water for mashing at 85°C under anaerobic condi- 97% is linoleic (omega-6), -linolenic (omega-3), oleic, tions. The mashing continues for one hour and the mash is palmitic, and stearic acid, while the final 3% consists of 12 then pulped with screens down to 0.5 mm pore size to obtain additional acids [61]. In several reports, linoleic acid makes the rosehip pulp which, after drying, is marketed as a pow- up approximately 40–56%, -linoleic acid and its derivatives der. During this process, more than 85% of the ascorbic acid 20–30%, and oleic acid 14–20% [36, 61, 62] of the total fatty is commonly lost [66]. acid content. Szentmihályi et al. [63] investigated several species and attributed the detected variation, 36–55% lino- Processing often proceeds beyond the production of dried leic acid and 20–27% -linolenic acid, to genetic differ- material. In a recent study, a pasteurized beverage was ences. Nowak [61] also found that relative ratios between manufactured from commercial rosehip powder and sea some acids differed between, e.g., R. dumalis samples and R. buckthorn (Hippophae rhamnoides) puree, and subsequently canina samples, two species that are very difficult to sepa- stored up to 35 days at 4°C or 22°C [71]. Samples were ana- rate with molecular markers. lyzed at regular intervals during the storage period, for content of ascorbic acid, carotenoids, major phenolics, toc- In addition to the high level of polyunsaturated fatty ac- ochromanols, total phenols and total antioxidant capacity. ids, seed oils from rosehips also contain comparatively high The largest changes occurred during the first 5 days of stora- levels of phytosterols, tocopherols, carotenoids and phenolic ge, after which a decrease was noted mainly for ascorbic compounds [62], suggesting that rosehip oil could become acid. It was concluded that packaging of the beverages sho- competitive in the food market. Moreover, these compounds uld be carried out at higher temperatures while storage sho- may, together with the fatty acids, provide some of the me- uld take place at low temperatures (4°C), thereby decreasing dicinal effects of rosehip seeds reported below. oxidation of all bioactive compounds except ascorbic acid.

4.4. Processing Traditionally, rosehip seed oil is extracted by cold- pressure (not over 40°C) for dermo-drugs and food, or by Rosehips are seldom used in a raw state, and postharvest organic solvents for the manufacturing of cosmetics [65]. treatments must therefore be optimized for retention of the Yield is increased by applying a solvent to the cracked seeds valuable compounds [64]. For commercial purposes, rose- and using, e.g., Soxhlet, ultrasound or microwave-assisted hips are usually dehydrated, sometimes after first making a extraction [72, 73]. Other solvents with low boiling tempera- pulp. Sometimes, harvested fruits are just spread out on the tures such as hexane and ethylic ether, have also been intro- ground for drying in the sun but this can lead to fungal con- duced. As a non-toxic alternative, supercritical CO2 extrac- tamination. Moreover a prolonged low-temperature dehydra- tion where the CO2 is compressed into liquid, is useful [72] tion can result in 90% reduction of vitamin C content [37], and superior for recovery of carotenoids [63]. To further Health-Promoting Rosehips Current Bioactive Compounds 2017, Vol. 13, No. 1 9 increase the oil yield, seeds can be pre-treated with specific been carried out in order to study the pharmacological effects enzymes, and then pressed at specific temperature and under of extracts made from rosehip powder. In an early example, moisture conditions [74]. Kharazmi and Winther [77] treated polymorphonuclear leukocytes (PMNs; important and abundant inflammatory cells 5. UTILIZATION AS FOOD AND MEDICINE involved in the pathogenesis of, e.g., arthritis) with extracts of rosehip powder (Hyben-Vital®). A chemotaxis assay Dogroses have long been used for the manufacturing of demonstrated significant inhibition of PMN movement. various food products. The fleshy parts of the rosehips are Similarly, a chemiluminescence assay with activated PMNs used fresh or dried, for the production of juice, jam, jelly, demonstrated significant inhibition of an oxidative burst re- marmalades, fruit leather, cream soups, bakery products and sponse. tea, and also as ingredients in ice-cream, yoghurt, candy and various liquors. Cold-pressed oils have recently been intro- An important role of the antioxidants in protection duced to the food market where their high levels of polyun- against inflammatory processes was suggested by Daels- saturated fatty acids and phytosterols, tocopherols and caro- Rakotoarison et al. [51], who showed that the polyphenols of tenoids should make them highly competetive [62]. Occa- R. canina rosehips are effective ROS scavengers and act sionally, rose leaves and petals are also used, especially in against the tested ROS species in human blood. Some years teas and other infusions. later, Widén et al. [78] reported that sequentially eluted Since interest in healthy food is increasing, fruit and ber- rosehip exctracts contribute to the protection of human eryth- ries are now appreciated not only as a tasty snack but also for rocytes. their health-promoting substances. Therapeutic activities of In addition to antioxidants, other substances are appar- rosehips have been described against the common cold, dia- ently also involved. One substance that has received consid- betes, gastro-intestinal disorders and gastric ulcers, and dif- erable attention is the galactolipid GOPO®, (2S)-1,2-di-O- ferent kinds of inflammatory disorders, e.g., osteoarthritis. [(9Z,12Z,15Z)-octadeca-9,12,15-trienoyl]-3-O--d- Uses as a mild laxative and a diuretic have also been re- galactopyranosyl glycerol, which has been isolated from ported. The mechanisms behind these effects are, however, rosehip powder (Hyben-Vital®) and has been shown to be largely unknown. Rosehips contain very high levels of poly- efficient in reducing chemotaxis of PMNs [79]. In 2011, phenols, once thought to act mainly as antioxidants in the Schwager et al. [80] showed that both rosehip powder and body but now regarded as an important factor in detoxifica- isolated GLGPG (syn. GOPO®) modulate the onset and de- tion and induction of genes involved in energy metabolism, velopment of ostheoarthritis by a decrease in nitric oxide and chronic anti-inflammatory state and endogenous-antioxidant by interfering with the activity of an inflammatory cytokine network at the cellular level [75]. called interleukin-1 (IL-1). The rosehip powder also de- According to the scientific literature, the most commonly creased the migration of neutrophils thereby delaying or pre- employed dogrose species for commercial food and food venting the initial steps of the disease. The reduced chemo- additives are R. canina and R. rubiginosa (syn. R. eglante- taxis and catabolic processes induced by the rosehip powder ria), but there has been much confusion among names, and thus provide a basis for cartilage protection. number of species involved may be considerably higher. Effects involving fatty acids have also been suggested. A Studies on medicinal effects are usually reported as based on lipophilic extract of rosehip powder (Hyben-Vital®) was thus R. canina even when employing commercially available pulp shown to inhibit the activity of COX-1 and -2 (cycloox- or powder of wild-collected rosehips imported from Chile ogenase, potent inflammatory mediators in the arachidonic where the naturalized species R. rubiginosa is much more acid cascade) in vitro [81]. A subsequent in-depth study common [76]. In addition, there is sometimes no mentioning demonstrated that this inhibition is closely associated with of whether the material has been manufactured from entire the contents of linoleic and -linolenic acid [82]. Wenzig et fruits (i.e. flesh and seeds), or from the flesh only (often al. [83] concluded that in addition to identified fatty acids termed ‘shells’). Many have used a rosehip powder from (triterpenoids and oleic, linoleic and -linolenic acids), other ® Hyben Vital in Denmark, containing mainly Danish-grown lipophilic constituents are likely to play a role in the ob- rosehips from the R. canina cultivar ‘Lito’, with approxi- served inhibition of COX-1, COX-2 and LTB4. mately equal amounts (DW) of fruit flesh and seeds, and standardized to a defined GOPO® content (see below). This Lately, there has been some debate about whether to use ® ® whole rosehips or only the fruit flesh. Two different rosehip product is now marketed as Movo (humans) and LitoVet ® (animals), but was previously marketed as LitoZin®, which is powders, with seeds (Litozin ) and without seeds (supplier sometimes synonymized with LitoMove® in commercial Martin Bauer GmbH & Co Kg, Germany), were compared information. The latter is, however, instead manufactured by Wenzig et al. [83] for phytochemical composition as well from wild-collected rosehips in Chile, and is standardized to as in vitro anti-inflammatory and radical-scavenging effects, an added vitamin C content. with the powder without seeds proving more effective in all assays. Powders with and without seeds (both provided by 5.1. Anti-Inflammatory and Antiulcerogenic Effects Axellus, mostly trading in wild-collected Chilean rosehips), were also used for in vitro analyses with human peripheral 5.1.1. In vitro Studies blood leukocytes and primary chondrocytes [84]. Although The most well-documented medicinal effects of rosehips both extracts modified a large number of biomarkers (pro- involve their anti-inflammatory action combined with, e.g. teins as well as expressed genes) associated with inflamma- cartilage-protective properties. Various in vitro studies have tory damage and cartilage erosion, the extract without seeds 10 Current Bioactive Compounds 2017, Vol. 13, No. 1 Nybom and Werlemark again appeared to be more efficient, and also contained more collected Chilean rosehips) produced overall less pain in chondroprotective and anti-inflammatory constituents. patients with osteoarthritis of the knee [96]. Results were, however, comparable between powders and doses (powder While almost all in vitro studies until now have been without seed was administered at regular dose and at half conducted with material of only one dogrose species, at least dose, respectively) but with a small tendency for half-dose one study compared wild-collected Serbian rosehips of a flesh-only powder to yield a better result in the self- dogrose (R. canina) with a non-dogrose (R. arvensis, section evaluation test. Synstylae). Significant activity against COX-1 and 12-LOX pathway metabolites was thus assessed with both a methanol Pain relief and a decrease in C-reactive protein and extract of dried hips and a purée of R. canina while only a creatinine levels in the blood of patients treated with rose- minor effect was noted for a purée of R. arvensis and no ef- hips have been well documented, but an animal-based study fects were obtained from neither water nor methanol extracts indicated beneficial effects also on bone structure. A mixture of fresh or dried fruits of the latter species [85]. Interestingly, of rosehip powder (provided by Atlas, trading in wild- these two species also differ substantially in chemical con- collected Chilean rosehips) and green tea seeds was thus tents, with R. canina having 5–10 times higher vitamin C shown to have a significant protective effect on monosodium content, and 10 times higher values in a FRAP assay measur- iodoacetate-induced osteoarthritis in rats, assessed from ing antioxidant activity. changes in behavior as well as changes in the actual bone structure [97]. 5.1.2. Animal Models Finally, it should be mentioned that there are also some Several animal model-based studies have been conducted studies that report a lack of effects. In one double-blind, pla- for investigating anti-inflammatory and antiulcerogenic ef- cebo-controlled study, osteoarthritis patients were given a fects of rosehip. Rats fed a R. canina extract apparently de- powder containing 90% fruit flesh and 10% seeds [98]. veloped protection againt ethanol-induced gastrointestinal Symptoms decreased only to the level reported by the pla- lesions [86, 87]. In another rat-based study, extract from R. cebo group, while C-reactive protein in the blood remained canina was used in combination with a Lactobacillus strain unchanged in both groups. Similarly, a short-term (one and resulted in a decreased ROS activity in injured rat colon ® month) trial with administration of LitoZin to patients diag- [88]. The authors speculate that this combination could be nosed with rheumatoid arthritis as well as to healthy con- useful as a pre-treatment to tissue injuries in the colon sys- trols, failed to identify any changes in plasma antioxidative tem, and in connection with organ transplants. When testing capacity and C-reactive protein in either group [99]. Much Setarud (IMOD®), a patented drug prepared from extracts of more research is obviously needed on the effects of using Rosa canina, Tanacetum vulgare and Urtica dioica, on rats rosehip powders of different origins (species, geographic with induced colitis, a significant reduction of both macro- location) and formulation (amount of flesh versus seeds), as scopic and histological damage was observed [89]. Possibly, well as appropriate dosage and minimum duration of treat- the drug inhibited the exacerbation of the inflammatory con- ment. dition caused by, e.g., the IL-1-gene. 5.1.3. Clinical Trials 5.2. Diabetes and Obesity Winther et al. [90] reported that daily intake of rose-hip Rosehips have been used traditionally as a remedy powder for four weeks by healthy volunteers and patients against diabetes in Anatolia. Modern research has now suffering from osteoarthritis, resulted in reduced serum C- shown that components extracted from rosehip seeds inhibit reactive protein (CRP) levels and reduced chemotaxis of body weight gain in mice, and produce a dose-dependent peripheral blood neutrophils. Since then, several clinical reduction of visceral fat [100]. Since this extract also im- trials have been performed on patients diagnosed with os- proves glucose tolerance, it might be useful for the treatment teoarthritis of the hip, knee or hand; see, e.g.,[91-94]. An of diabetes, obesity and metabolic syndrome. Interestingly, improved functional capacity of the affected joints, reduced although extracts from fruit and seed of R. canina achieved a pain and an increased activity in the daily life of these pa- substantial inhibitory effect on body weight gain in mice, tients has been demonstrated, especially after a longer time especially visceral fat, food intake was not affected [101]. (preferably several months) of administration. Very few side The principal active constituent in the latter study was trans- effects such as constipation, diarrhea or allergy have been tiliroside, a substance known to reduce blood glucose levels recorded. after glucose loading. Orhan et al. [102] administered an ethanol extract of rosehips (locally collected and dried R. In a follow-up of a previous in vitro study reporting supe- canina) to rats during 7 days, and found a remarkable hypo- rior results for flesh-only powders [83], Chrubasik- glycemic effect in animals with induced diabetes. Andersson Hausmann et al. [95] compared the effects of administering ® et al. [103] showed that a rosehip dietary supplementation two different rosehip powders, with seeds (LitoZin ) and (commercially available purée made of ground R. canina) without seeds (supplier Martin Bauer GmbH & Co Kg, Ger- reduced lipid accumulation in the liver of mice and lowered many), to patients suffering from long-term low back pain the total plasma cholesterol. Therefore, they proposed that syndrome or knee pain. Significant pain relief was obtained, rosehips exert an antidiabetic effect by downregulation of the but neither generic nor disease-specific outcome variables hepatic lipogenic program. were able to detect any differences between the two powders. In another study, ingestion of powders with and with- In contrast, no effects were noted on body weight or glu- out seeds (provided by Axellus, mostly trading in wild- cose tolerance when obese humans ingested a rosehip drink (made from dried Chilean rosehips without seeds) in a ran- Health-Promoting Rosehips Current Bioactive Compounds 2017, Vol. 13, No. 1 11 domized, double-blind, cross-over investigation [104]. Most form of vitamin A, tretionin. The oil can be used for treating likely, the daily intake of active rosehip ingredient was lower burns, eczema and acne, and made into a salve that is much in this investigation compared to in the studies by Ninomiya appreciated in the cosmetic industry because of its skin reju- et al. [101] and Andersson et al. [103], and the metabolic venating capacity. Although the extraction process often status of the study group was therefore different. Still, the requires high temperatures, the oil retains its high antioxi- study showed that a daily consumption of rosehip drink dur- dant capacity to a higher extent than other skin salves of ing six weeks significantly lowered the systolic blood pres- plant origin [73]. Today, there are hundreds of rosehip oil sure and plasma cholesterol levels, thereby also decreasing products in the market, originating mainly from Chile, but the risk of cardiovascular diseases. also from United States and China.

5.3. Mutagenesis and Cancer 6. GENETICS AND PLANT BREEDING Another area of rosehip research concerns anti-mutagenic Although most plant domestication processes start with and anti-carcinogenic effects. In a study, using with the the collection of desirable plant organs (e.g. leaves, fruits or mutagenic substance sodium azide on the bacterium Salmo- roots) from wild-growing populations, further progress usu- nella typhimurium, an extract from R. canina thus reduced ally involves optimized field or orchard cultivation of care- the mutagenicity by 44% [105]. Inhibition of cancer cell pro- fully selected plant genotypes. For production of rosehips for liferation in vitro has been demonstrated for extracts from medicinal purposes, there is obviously an urgent need for various fruits and berries, with time- and dose-dependent determination of the most desirable traits, and then selecting effects of many substances. The inhibitory effect is often – and also possibly improving the plant material accordingly but not always – correlated with the vitamin C level although [109]. vitamin C itself does not have this inhibitory effect. Olsson For successful field production and breeding of rosehips, et al. [48] demonstrated an inhibition of colon and breast cancer cells by rosehip extracts whereas the control group information is needed not only on the traits involved in fruit quality, but also on traits that govern, e.g., yield and sound- treated with only vitamin C did not show the same inhibi- ness. Some information is available for ornamental roses but tion. However, the vitamin C level was correlated with the the traits may be judged differently. Dogroses flower during inhibition of cancer cell proliferation and with some caro- only 1–2 weeks in contrast to most ornamental roses that tenoids, and a synergistic effect including several of the bio- flower much longer. The recessively inherited trait known as chemical compounds was therefore suggested. recurrent blooming would, however, be a serious draw-back Tumbas et al. [49] tested different fractions of rosehip tea in rosehip production, especially in machine-harvested or- phytochemicals (vitamin C, flavonoids and phenolic acids) chards where repeated harvestings would be required. Stems on the cell growth of different cancer cell lines, but antipro- without prickles would facilitate manual orchard work but liferative activity was obtained only with the polyphenol would also render the plants as more vulnerable to grazing fraction while vitamin C and flavonoids contributed to the animals. The arching growth type (e.g. R. canina) has been antioxidant activity. Cagle et al. [106] showed that a rosehip reported to be dominant over the more desirable erect (and extract reduced proliferation also of human brain glioblas- non-climbing) growth type (e.g. R. rubiginosa) in dogroses toma cells, and that this was achieved more effectively than [110]. Other growth type- and vigour-related traits studied in by the frequently used chemo-therapeutic drug Temo- ornamental roses like length of the flowering stems and plant zolomide. The rosehip extract thus inhibited different cancer height, as well as branch number, plant spread and prickle cell proliferation pathways and enhanced the effect of Temo- number, are under polygenic control [111, 112]. zolomide when used concomitantly. However, this extract did not promote apoptosis of the cancer cells. Polygenic resistance towards most of the common fungal diseases like blackspot (Marssonina rosae), powdery mildew 5.4. Skin Remedies (Podosphaera pannosa), rust (Phragmidium spp.) and Sphaceloma-leafspot is apparently common in dogroses Healthy subjects in a double-blind clinical study were [113, 114]. In one study, R. rubiginosa had no symptoms measured for facial skin wrinkles, moisture and elasticity although other species in the same field were heavily in- after ingesting a rosehip powder with 50% fruit flesh and fested, suggesting that this particular genotype may have 50% seeds (Hyben Vital®) [107]. In addition, blood samples monogenic resistance [114]. were evaluated for leakage of hemoglobin through cell membranes to evaluate cell longevity. Results showed posi- 6.1. DNA Markers tive effects with time (up to 8 weeks of ingestion) of the Correct identification of plant material is needed for the same magnitude as those obtained with the skin rejuvenant astaxanthin in the control group. Rosehip extracts have also evaluation of germplasm and for plant breeding, but is often difficult to achieve with morphological traits. In contrast, been shown to decrease melanogenesis in guinea pigs by molecular markers (especially SSR analyses) have proven to reducing the melanin content through the action of mainly be very efficient for discrimination among different rose proanthocyanidins [108] and may become useful for the pre- genotypes. To improve allele identification and avoid stutter- vention of hyperpigmentation and for the removal of age- ing, SSR repeats with three or more basepairs are often used related pigmentation spots. in both ornamental roses and dogroses [13, 115]. A variation Commercial skin remedies are, however, usually made on this theme is the EST (expressed sequence tags) -SSR from rosehip oil instead of entire rosehips. The most well- markers developed for ornamental roses [116] and for R. known substance in this oil is trans-retinoic acid, the acid roxburghii [117]. 12 Current Bioactive Compounds 2017, Vol. 13, No. 1 Nybom and Werlemark

So far, large-scale marker-based screenings of dogroses velopmental stages and under different stress conditions have mostly been undertaken for the purpose of taxonomic [123]. The web interface ROSAseq was created together with classification. When applied to several genotypes within the the ROSAcyc resource pathway database, thus providing ac- same population, between-genotype variation is restricted cess to a valuable transcriptome database and an overview of [118]. Interestingly, putatively invasive populations of R. gene expression patterns. rubiginosa in the Nordic countries [14] as well as in Argen- Another RNA-Seq library was recently generated from tina [119] show considerably less variation than native popu- multiple sources, including tetraploid ornamental roses (cut lations in Central Europe. roses and garden roses) as well as diploid R. multiflora, re- 6.2. Genetic Linkage Maps sulting in 13,390 full-length coding regions corresponding to orthologous genes in the Fragaria vesca database [124]. A variety of DNA markers have been applied for the de- Since RNA-Seq data also have the potential to reveal multi- velopment of genetic linkage maps for both diploid [120] ple SNPs that can be used for genotyping and marker-trait and autotetraploid ornamental roses [121]. Map-based identi- associations, a total of 68,893 SNPs were subsequently fication of genes is based on associations between DNA placed on the WagRhSNP 68K axiom array. This microarray markers and phenotyping data, scored in a designated map- will be suitable for genotyping as well as for identification of ping population. The use of such maps has facilitated the genes governing commercially important traits and for fine- resolution of genes and QTL (quantitative trait loci) affecting tuning the localization of these genes on already existing traits of relevance for ornamental roses, like flower mor- genetic linkage maps. A large part of these 68.000 SNPs phology, colour and scent, but also some more universally would, in all likelihood, be informative also in dogroses. important traits like plant morphology (prickles and growth vigour) and disease resistance (black-spot and powdery mil- A transcriptomics study of particular relevance also for dew) [121]. Collection of accurate phenotyping data is, how- dogroses, was recently conducted on the vitamin C-rich Rosa ever, very challenging for quantitative traits like chemical roxburghiensis belonging to subgenus Platyrhodon [117]. A contents in plant tissue, since scorings should be undertaken total of 498 unique putative transcripts were identified, some during several seasons and preferably also in clonally dupli- of which are known to regulate fruit development including cated material maintained in separate cultivation trials at major enzymes in the ascorbate biosynthesis pathway. different geographic locations. 6.4. Dogrose Breeding Until now, no genetic linkage maps have been developed for dogroses. Possibly, one could use a strategy similar to A few dogrose cultivars have been developed from selec- what has been applied for diploids, i.e. a double pseudo- tions in material obtained from wild populations, e.g. ‘Mech- testcross; two highly heterozygous parents are crossed, and thilde von Neuerburg’ (R. rubiginosa) ‘Sylwia’, ‘Sylwana’ markers segregating in one or both parents are evaluated in and ‘Plovdiv 1’ (R. canina) and ‘Karpatia’ (R. villosa) [125]. the offspring. Distances between these markers are deter- With the aim to produce high-yielding plants for fruit pro- mined by, e.g., Chi-square analyses of the segregation ratios duction, inter-sectional crosses have also been conducted, obtained in the offspring. In the case of dogroses, some of resulting in e.g. the Pillnitzer Vitamin-Rose ‘PiRo 3’ devel- the parental markers are transmitted to all offspring because oped in the 1960s at Pillnitz in Germany from a cross be- they are either homozygous on the bivalents, or – for the tween R. dumalis (sect. Caninae) and R. pendulina var. seed parent only – occur on the univalents [13, 14, 20, 21]. salaevensis (sect. Rosa ) (Fig. 5). This cultivar is reported to Remaining markers are transmitted to approximately 50% of have a good yielding capacity together with high soluble the offspring and therefore reside on the bivalents, and solids content [126] and a vitamin C content of ca. 1150 would thus be amenable for linkage analysis. mg/100 g fruit flesh [127]. Other hybridogenous cultivars are ‘Nektar’ (R. rugosa), ‘Besshipnyjj’ (R. webbiana R. 6.3. Transcriptomics rugosa), ‘Vitaminnyjj’ (R. majalis R. webbiana), and ‘Pozdnespelyij’, ‘Uralskij Champion’ and ‘Voroncovskijj’ A different approach towards the identification of genes governing traits of interest, is to start from genes that are which have a similar background [125, 127, 128]. All of actively expressed in the genome. Based on the sequencing these have early-ripening fruits, rather soft fruit flesh and a of EST (expressed sequence tags) of ornamental rose, and relatively mild taste with faint aroma. development of microarrays containing 350 and 5000 More recent selection programs aimed at identifying va- polynucleotides, respectively, candidate genes have been rieties for health food and medicinal purposes, have resulted identified for several traits of importance for flower initia- in the R. canina cultivar ‘Lito’ grown exclusively by its tion, flower organ morphogenesis and senescence, and scent owner, Hyben Vital® in Denmark, and several as yet un- biosyntehsis (review in [122]). named selections of different dogrose species, like R. du- Subsequently, RNA-Seq (also known as whole transcrip- malis and R. rubiginosa [26], and R. dumalis, R. canina, R. montana and R. pulverulenta [129]. While R. canina may tome shotgun sequencing) was applied by using next genera- ® tion sequencing to cover a major part of the transcriptome of have very high levels of GOPO , neither plant habit or yield the diploid ornamental rose cultivar ‘Old Blush’. Samples is desirable for commercial plant production. Most of the were taken of different types of tissue and under different other species are characterized by a compact and easily biotic and abiotic stress conditions. Information was ob- pruned plant habit, comparatively good resistance against tained for approximately 21,000 putative genes, and how fungal diseases, many and large hips, and high levels of these are expressed in different plant organs at different de- ascorbic acid. Health-Promoting Rosehips Current Bioactive Compounds 2017, Vol. 13, No. 1 13

to the matroclinal inheritance, direction of the cross should be chosen so that the plant with the most valuable traits is used as seed parent. Still, crosses do not always produce as expected. Some aberrations during dogrose reproduction have been demonstrated with SSR analysis [13, 14]. Thus, an unreduced eggcell is sometimes fertilized, giving rise to a hexaploid offspring from a pentaploid seed parent, as determined by the occurrence of six alleles and also verified by flow cytometry. In addition, fertilization is occasionally achieved by diploid or aneuploid pollen [14]. Finally, some crosses may give rise to apomictically derived offspring.

CONCLUSION AND FUTURE PROSPECTS Although many promising studies have been published lately, more research is clearly needed to ascertain the health-promoting effects of rosehips. Both laboratory and Fig. (5). Rosehips of the cultivar ‘PiRo 3’. Photo Li-Hua Zhu. clinical experiments should be carried out using well-defined and thoroughly documented plant material, e.g. species determination incl. properly deposited herbarium specimens, Matroclinal inheritance is expected in dogroses since the geographic location, and harvesting and post-harvest proce- seed parent transmits all of its unchanged and often diverse dures. Since considerable inter-specific variation in chemical univalents to the next generation. By contrast, the biparen- contents has been demonstrated, a wide range of genetically tally transmitted bivalent chromosomes have a similar allelic different plant materials should be screened to identify the configuration if plants of the same species are cross- optimal choice for different applications of rosehip-derived pollinated, thus precluding a major influence of different products. A possible interaction between genotype and geo- paternal alleles. In interspecific hybrid plants, the situation graphic location must be investigated. An optimization of the may be somewhat different. Although observations of some growing conditions of the plants as well as harvesting and traits have demonstrated the expected matroclinal inheritance processing of the rosehips should also help to improve the for cuticular wax characters [130], intermediate or even pa- efficiency of the final products. ternal inheritance has instead been reported for other traits [131]. In species with a regular meiosis, hybrid plants often In the second step, the plant material could also be fur- display a mixture of parental traits due to the occurrence of ther improved by plant breeding. Crosses between different dominance effects, and this could also be the reason why genotypes have the potential to optimize character combina- some characters in dogrose crosses appear to be paternally tions but they must be carefully planned according to the inherited. Incomplete dominance or codominance could also special situation deriving from polyploidy, apomixis and – lead to intermediate inheritance, as is often seen in the col- above all – the unique canina meiosis and resulting matrocli- oration of flowers. In addition, effects of epigenetic, tissue- nal inheritance. Further complications due to possible silenc- specific processes have been indicated in a recent study of ing of univalent-residing genes, and epigenetic influence dogroses [19]. Dogrose hybrids sometimes display charac- must also be taken into consideration. ters that are extreme by related to either parent, e.g., an in- Plant breeding efficiency will be much improved if key creased amount of glandular hair and prickles [21]. The genes can be identified, e.g. genes affecting yield and fruit cause for this transgressive inheritance could be a comple- quality including various health-promoting compounds. Fur- mentary action of alleles, and/or recessive genes becoming ther development of transcriptomics-based studies is likely homozygous in the hybrid while being heterozygous and to become very valuable for identification of genes involved undetected in the parental plants. in relevant metabolic pathways. In addition, an international Directed crosses between dogroses on one hand, and spe- consortium, “Rose Genome Sequencing Initiative”, is pres- cies or cultivars in other sections on the other hand, are usu- ently undertaking the first sequencing of a rose genome, ally conducted with the dogrose as the seed parent. Unless namely a homozygous genotype (HzRc-RDP12) derived the pollinator is diploid, the offspring becomes highly poly- from the diploid ‘Old Blush’ [133]. The rose genome se- ploid (6x–8x). Pollinating pentaploid R. dumalis and R. ru- quence will be useful for discovering molecular and genetic biginosa genotypes with pollen from tetraploid ornamental mechanisms that control many horticultural traits in orna- cultivars produced mainly hexaploid offspring, with four mentals, and possibly also for discovering yield- and healthy genomes from the seed parent and two from the pollen par- compounds-related traits in dogroses. ent according to SSR analysis [132]. Instead, using the dogrose species as pollen parent has the same impact on the CONFLICT OF INTEREST ploidy of the offspring as when using a diploid species, but The authors confirm that this article content has no con- seed set is usually poor since dogroses have inferior pollen flict of interest. viability. Crosses between two dogrose species produce offspring ACKNOWLEDGEMENTS with the same chromosome number as the seed parent. Due Declared none. 14 Current Bioactive Compounds 2017, Vol. 13, No. 1 Nybom and Werlemark

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