Influence of Citrus Rootstocks in Bioactive Compounds of Clementines

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1 Journl of Food nd Nutrition Reserch, 2017, Vol. 5, No. 8, Avilble online t Science nd Eduction Publishing DOI: /jfnr Influence of Citrus s in Bioctive Compounds of Clementines Pilr Legu 1,*, Frncisc Hernández 1, Mrí Ángeles Forner-Giner 2 1 Miguel Hernández University, Deprtment of Plnt Sciences nd Microbiology, Reserch Group in Plnt Production nd Technology, Ctr. de Beniel, Orihuel, Alicnte, Spin 2 Centro de Citricultur y Producción Vegetl. Instituto Vlencino de Investigciones Agrris (IVIA), Aprtdo Oficil, Moncd, Vlenci, Spin *Corresponding uthor: p.legu@umh.es Abstrct Citrus fruit is one of the most importnt commercil fruit crops in the world. It is essentil to use citrus rootstocks in this crop, since ll vrieties, whether, ornges, tngerines, lemons nd grpefruits re highly sensitive to Phytophthor spp., present in lmost ll soils. Selection of optiml rootstock is crucil for commercil cultivtion of citrus fruit, s the effect fruit production, tree size, diseses, nd dpttion to certin soil conditions. Besides, rootstock my ffect internl qulity prmeters of citrus fruits, including juice totl, soluble solids, cidity, ntioxidnt cpcity, sugrs nd orgnic cids content, flvor. In this study, we evluted the effect of three rootstocks (Crrizo citrnge, C-35 citrnge nd Forner-Alcide 5) on physicochemicl prmeters, sugrs nd orgnic cids, totl ntioxidnt ctivity, totl phenolic, crude fibre content nd flvonoids, of Clemenrubí nd Orogrós clementines. The results indicte tht the effect of rootstock on the physicochemicl prmeters, ntioxidnt ctivity, totl phenolic, sugrs nd orgnic cids nd flvonoids of clementines fruits is rther complex phenomenon tht gretly depends on specific interctions between the rootstock nd ech prticulr scion vriety. C-35 citrnge rootstock produced the heviest nd bigger size fruits, while the rootstock Forner-Alcide 5 produced the fruit with the mjor flvonoid content. Keywords: clementines, orgnic cids, polyphenolic profile, rootstock, sugrs Cite This Article: Pilr Legu, Frncisc Hernández, nd Mrí Ángeles Forner-Giner, Influence of Citrus s in Bioctive Compounds of Clementines. Journl of Food nd Nutrition Reserch, vol. 5, no. 8 (2017): doi: /jfnr Introduction Citrus fruit is one of the most importnt commercil fruit crops in the world. Spin produces 2.4 million tons of mndrins, being the second lrgest producer in the world, nd being the lrgest exporter [1]. It is essentil to use citrus rootstocks in this crop, since ll vrieties, whether; ornges, tngerines, lemons nd grpefruits re highly sensitive to Phytophthor spp., present in lmost ll soils. s hve lrge impct on scion growth hbit nd fruit yield nd qulity [2]. Besides, rootstock my ffect internl qulity prmeters of citrus fruits, including juice totl, soluble solids, titrtble cidity, ntioxidnt cpcity, sugrs nd orgnic cids content, flvor [3,4,5]. Crrizo citrnge [Citrus sinensis (L.) Osb. Poncirus trifolit (L.) Rf.] is currently the most importnt rootstock used in Spin, employed in over 80% of nursery production [6]. Recently some Spnish Orchrds hve been plnted with C-35 citrnge (C. sinensis P. trifolit) rootstock; it is resistnt to the citrus nemtode (Tylenchuslus semipenetrns Cob.) nd trees on this rootstock lso rech smller size thn Crrizo citrnge [7]. Forner-Alcide 5 is new citrus rootstock relesed from the citrus rootstock breeding progrm of Instituto Vlencino de Investigciones Agrris (IVIA), this rootstock is resistnt to Citrus Tristez Virus (CTV), tolernt to slinity [8], iron chlorosis [9], nd citrus nemtode [10]. In ddition, it produces high yiled nd excellent qulity fruit [11]. Citrus fruits nd derived products hve beneficil effect on humn helth becuse of their nutritionl nd ntioxidnt properties, nd their consumption is ssocited with decrese in the incidence of crdiovsculr disese nd reduced risk of certin cncers [12]. In the present study, we evluted, for the first time, the effect of three rootstocks (Crrizo citrnge, C-35 citrnge nd Forner-Alcide 5 (FA-5)) on physicochemicl prmeters, sugrs nd orgnic cids, totl ntioxidnt ctivity, totl phenolic, crude fibre content nd flvonoids, of Clemenrubí nd Orogrós clementines. Both clementines re erly muttions from Oronules nd re being registered in EU nd commercilized in Spin. The informtion generted will help breeders to select the best rootstock for their plnttions of clementines.

2 546 Journl of Food nd Nutrition Reserch 2. Mterils nd Methods 2.1. Plnt Mteril nd Experimentl Design Seeds of Crrizo citrnge nd C-35 were obtined from the germplsm collection of rootstocks t IVIA nd the seeds of the FA-5 from the plnts obtined in the citrus rootstock breeding progrm. All nursery processes were performed in n phid-proof greenhouse, with cooling system temperture rnging between 18 nd 27 C nd reltive humidity of bout 80%. The trees were grown in the nursery using stndrd prctices nd, were plnted in Algemesi (Vlenci, Spin). The plot ws plnted in My, 2005 under screenhouse in order to void sun-burning. The experimentl design ws rndomized complete block, with twelve trees per plot. The soil ws sndy-lom, (ph 8.2, 38.2% snd, 40.5% silt, 21.3% cly, 1.4% orgnic mtter nd 26.8% CCO3). The grove ws cultivted, pruned nd spryed, ccording to locl prctices. The soil ws kept free of weeds using post-emergence herbicides Preprtion of Juice Smple The clementines (Citrus clementin Hort. ex Tn.) cv. Clemenrubí nd Orogrós, fruits were hrvested t optimum mturity nd immeditely trnsferred to the lbortory of the Deprtment of Plnt Science nd Microbiology, University Miguel Hernández Elche, Spin. Reserch ws crried out using 30 kg of clementines from ech vriety/rootstock combintion. Forty-five homogeneous fruits (bsed on color, size, nd bsence of defects) were selected per scion/stock for physicl nd chemicl determintions. Three subsmples (ech one composed by 10 fruits) were mde t rndom, nd then the fruits from ech subsmple were cutting in hlf nd creful hndsqueezing in commercil kitchen juicer. The freshly squeezed juices were centrifuged t 15,000 rpm nd 4ºC for 20 min (Sigm 3-18K centrifuge, Sigm Lborzentrifugen GmbH, Osterode nd Hrz, Germny) nd were kept in freezer t temperture of -18ºC until nlysis, while the remining 15 fruits per scion/stock were freeze-dried for polyphenolic compounds determintion Physicochemicl Anlyses For ech fruit, the following prmeters were mesured: mximum width or equtoril dimeter, ED (mm), nd fruit length, FL (mm), using digitl clliper (model CD-15 DC; Mitutoyo (UK) Ltd, Telford, UK) with 0.01 mm ccurcy; fruit weight, FW (g) using digitl scle Srtorius (model BL-600, with n ccurcy of 0.01 g). Totl soluble solids (TSS) were determined ccording to [11] with hnd-held refrctometer (Atgo Co., model N-1, Tokyo, Jpn, 0.2 Brix ccurcy) nd expressed s degrees Brix t 20 C. Titrtble cidity (TA) nd ph were determined using n utomtic titrtion device (877 Titrino plus, Metrohm ion nlyses CH9101, Herisu, Switzerlnd) with 0.1 N NOH up to ph 8.1 using 1 ml of diluted juice in 25 ml of distilled H 2 O, nd results were expressed s g citric cid equivlent per 100/g fw, since this is the dominnt orgnic cid in ornge. The rtio, lso known s the ripening index (RI), ws clculted s the reltion between TSS nd TA. All nlysis ws performed in triplicte. Colour determintions were mde for juice (n=3). The CIELAB L* (brightness or lightness; 0 = blck, 100 = white), * ( * = greenness, +* = redness) nd b* ( b* = blueness, +b* = yellowness) color vribles were mesured using Chrom Meter (C-300 Minolt Corp., Osk, Jpn) coupled to Minolt DP-301 dt processor. These vlues were then used to clculte Hue ngle degree [Hº* = rctng (b*/*)], where 0º = red-purple; 90º = yellow, 180º = bluish-green nd 270º = blue nd Chrom [C* = ( *2 + b *2 )] 1/2, indictive of the intensity or color sturtion. The crude fibre (CF) contents were determined by digester, n Ankon fiber nlyzer (model A220, Mcedon, NY, USA), following the officil methodology estblished by the Spnish Ministry of Agriculture, Fisheries nd Food [13]. Results were expressed s g 100 g -1 dw (dry weight). Three repetitions per vriety/rootstock combintion were crried out Anlysis of Sugrs nd Orgnic Acids Contents Individul orgnic cids nd sugrs were lso determined using three juice smples for ech scion/stock combintion s described by [14]. Briefly, one milliliter of the extrct ws filtered through 0.45 μm Millipore filter nd then injected into HPLC (Hewlett-Pckrd, series 1100, Wilmington Del., USA). The elution system consisted of 0.1% phosphoric cid with flow rte of 0.5 ml min -1. The orgnic cids were eluted through Supelco column [SupelcogelTM C-610H column (30 cm 7.8 mm)] nd Supelgurd column (5 cm 4.6 mm, Supelco, Inc., Bellefonte, PA, USA) nd detected by bsorbnce t 210 nm. For sugr determintions, the sme HPLC, elution system, flow rte nd columns were used. Sugrs were detected with refrctive index detector (Hewlett-Pckrd, series 1100, G1362A, Wilmington Del., USA). A stndrd curve of pure orgnic cids (oxlic, citric, trtric, mlic, cetic, fumric, succinic nd L-scorbic cids) nd stndrd curve of pure sugrs (glucose, mltose, fructose, sucrose nd sorbitol) purchsed from Sigm (Poole, Dorset, UK) were used for quntifiction. Clibrtion curves, obtined by triplicte injection of stndrd solutions, were used for quntifiction purposes nd showed good linerity (R2>0.999). Results for both individul orgnic cids nd sugrs were expressed s g 100 ml -1. Three repetitions per vriety/rootstock combintion were crried out Totl ntioxidnt ctivity (TAA) TAA ws quntified by spectrophotometry s described by [15]. This procedure llowed determining both the hydrophilic nd lipophilic TAA in the sme extrction. Briefly, for ech subsmple, 5 ml of citrus juice were homogenized in 5 ml of 50 mm phosphte buffer ph=7.8 nd 3 ml of ethyl cette, then centrifuged t g for 15 min t 4 ºC. The upper frction ws used for TAA due to lipophilic compounds (L-TAA) nd the lower for TAA due to hydrophilic compounds (H-TAA). In both

3 Journl of Food nd Nutrition Reserch 547 cses, TAA ws determined in triplicte in ech extrct using the enzymtic system composed of the chromophore 2,2 -zino-bis-(3-ethylbenzothizoline-6-sulfonic cid) dimmonium slt (ABTS), the horse rdish peroxidse enzyme nd its oxidnt substrte (hydrogen peroxide), in which ABTS + rdicls re generted nd monitored t 730 nm. The decrese in bsorbnce fter dding the extrct ws proportionl to TAA of the smple. A clibrtion curve ws performed with Trolox ((R)-(+)-6-hydroxy- 2,5,7,8-tetrmethyl-cromn-2-crboxylic cid) (0 to 20 nmol) from Sigm (Mdrid, Spin), nd results re expressed s mg of Trolox equivlent (TE) 100 g -1 fw. Three repetitions per vriety/rootstock combintion were crried out Totl Phenolic Content by Folin Cioclteu s Regent Totl phenolic content (TPC) ws quntified using Folin-Cioclteu regent s described by [16]. Absorption ws mesured for the smple t 760 nm using spectrophotometer (ThermoSpectronic Heγios γ, Englnd). Three repetitions per vriety/rootstock combintion were crried out nd results were expressed s mg of gllic cid equivlent (GAE) 100 g -1 fw HPLC-DAD-ESI-MS n Anlysis, Identifiction nd Quntifiction of Phenolic Compounds 50 mg of lyophilized smple ws mixture with 1 ml of MeOH nd then mixing ws crried out by vortex, nd the extrction ws performed in n ultrsonic bth for 1 min t room temperture. The extrct ws centrifuge t 4, 000 rpm for 4 min nd pssed through 0.45 μm PTFE filter (Wters, Milford, USA) prior to injection into the chromtogrphic system. Chromtogrphic nlysis ws performed on n Agilent 1100 series HPLC-ESI-DAD-MS n Ion Trp (Wldbronn, Germny). A reversed-phse Agilent Pursuit XRs 5 C18 column ( mm i.d. nd prticle size 5 μm, Wldbronn, Germny). Wter/formic cid (95:5, v/v) nd cetonitrile were used s mobile phses A nd B, respectively, with flow rte of 0.8 ml min -1. The grdient strted with 5% of solvent B, reching 60% solvent B t 37 min, nd 98% t 40 min, which ws mintined up to 2 min. The injection volume ws 10 μl. Chromtogrphic comprison with nlyticl stndrds, bsorbnce spectr, nd mss spectr, using MS n (dt not shown) experiments, were used to identify compounds. Flvnones nd C-glucoside were monitored nd quntified t 360 nm. Three repetitions per vriety/rootstock combintion were crried out nd results were expressed s mm of hesperiden for flvnones nd s quercetin for C-glucoside Sttisticl Anlyses All dt were subjected to two-wy nlysis of vrince, ANOVA [fctors: (i) clementines cultivr nd (ii) rootstocks]. Lest significnt difference (LSD) multi-comprison test ws used to determine significnt differences mong tretments. Significnce of differences ws represented s p Sttisticl nlyses were performed using SPSS 20.0 for Windows (SPSS Science, Chicgo, IL, USA). 3. Results nd Discussion 3.1. Physicochemicl Prmeters The results of the physicochemicl prmeters re presented in Tble 1. The fctor rootstock significntly ffected fruit weight nd equtoril dimeter. C-35 citrnge produced the heviest nd bigger size fruits (Tble 1). A study reported bout Nvelin ornge cultivr tht C-35 citrnge lso induced the gretest weight nd fruit size [11]. The interction rootstock vriety ws significntly for chemicl prmeters (ph, TSS, TA nd RI). In generl, Clemenrubí vriety grfted on Crrizo citrnge nd FA-5 hs higher content in TSS (Figure 1). Regrding, the interction rootstock vriety for TA Clemenrubí grfted on the three rootstocks ssyed showed higher vlues of TA compred with Orogrós (Figure 1c), while with respect to ph ws Orogrós which showed higher vlues (Figure 1d). [17] reported bout Lne Lte nvel ornge on four citrus rootstocks tht the rootstock ffect on fruit juice cidity, nd RI; lso, [5] in study bout effects of rootstock/scion combintions on the flvor of citrus fruit found tht TSS nd cidity levels in juice fruits depend of rootstock/scion combintion; however, other uthors found tht the effects of rootstock on fruit juice cidity nd RI were non-significnt [18,19,20]. The ripening index (TSS/TA) is n importnt prmeter relted to qulity chrcteristics of citrus fruits [21] nd it is lso the best criterion in correlting fruit qulity with consumer cceptnce [22]. Orogrós grfted on C-35 citrnge showed the higher ripening index (Figure 1b). In generl, RI vlue for cceptnce of consumers since plesnt tste is chieved when it is greter thn 10 [23]. Besides, Orogrós fruits reched higher vlues of juice TSS nd lower TA fruits grfted on C-35 citrnge nd they could hve improved the flvor of the juice nd therefor the commercil qulity nd consumer cceptnce. Dt obtined in this study from citrus rootstock showed the influence of rootstock nd vriety in the physicl nd chemicl fruit prmeters. Besides, it cler tht, the effect of rootstocks on some physicl nd chemicl prmeters of citrus fruits is rther complex phenomenon tht gretly depends on specific interctions between the rootstock nd ech prticulr scion vriety. Color is considered one of the most importnt externl fctors of fruit qulity, s fruit ppernce gretly influences consumers. In temperte countries, this non-destructive method cn be pplied in the field s well s in the industry to ccurtely show the pprent mturtion degree of the fruit [24]. The fctor rootstock significntly ffected ll color prmeters; however, the interction rootstock vriety only ffected * nd Hº* prmeters (Tble 2). Fruits of trees grfted on Crrizo citrnge ws more luminous in color juice (higher L* prmeter). On the other hnd, trees grfted onto Crrizo citrnge nd FA-5 produced fruit with the most intense ornge colored juice (higher * prmeter) (Tble 2). In Figure 2e, it shows tht Clemenrubí grfted on C-35 citrnge presented the lowest vlues of *, while Orogrós showed the higher vlues. The fruits with best juice color were produced on FA-5.

4 548 Journl of Food nd Nutrition Reserch Tble 1. Effect of citrus rootstocks on fruit weight, size, ph, totl soluble solids, titrtble cidity nd ripening index of clementines cv. Clemenrubí nd Orogrós Prmeters FW (g) ED (mm) FL (mm) ph TSS TA RI Crrizo citrnge 83.8 b b 54.2 b b 11.1 C-35 citrnge b FA b 55.7 b b b Clemenrubí Orogrós *** *** NS ** NS ** *** NS NS NS *** NS *** *** x NS NS NS ** ** *** *** (LSD). FW, fruit weight; ED, equtoril dimeter; FL, fruit length; TA, titrtble cidity (g citric cid L -1 ); TSS, totl soluble solids (ºBrix); RI, ripening index (TSS/TA) Tble 2. Effect of citrus rootstocks on juice colour of clementines cv. Clemenrubí nd Orogrós Prmeters L* * b* C* Hº* Crrizo citrnge 40.5 b C-35 citrnge 39.5 b 1.8 b 7.7 b 7.9 b 76.8 FA c b 7.7 b 75.4 b Clemenrubí Orogrós *** ** *** *** *** NS *** NS NS *** x NS *** NS NS *** (LSD). L*= lightness; *= green/red coordinte; b*= blue/yellow coordinte; C*= chrom; Hº*= hue ngle. Figure 1.

5 Journl of Food nd Nutrition Reserch 549 Figure 1. Interction grphics rootstock x vriety for TSS (), RI (b), TA (c), ph (d), * (e), Hº* (f), sucrose (g), citric cid (h), mlic cid (i), TPC (j), H-TAA (k), L-TAA (l), CF (m), Api-6,8-di-C-glc (n), Nr-7-O-rut (o) nd Hesp-7-O-rut (p)

6 550 Journl of Food nd Nutrition Reserch Tble 3. Effect of citrus rootstocks on orgnic cids nd sugrs (g 100 ml -1 ) of clementines juice cv. Clemenrubí nd Orogrós Prmeters Glucose Fructose Sucrose Citric Mlic Ascorbic Crrizo citrnge b b C-35 citrnge b FA b Clemenrubí Orogrós NS NS NS *** ** NS *** *** * *** NS * x NS NS ** *** ** NS (LSD). Tble 4. Effect of citrus rootstocks on totl phenolic content (TPC), totl ntioxidnt ctivity (TAA) nd crude fibre (CF) of clementines cv. Clemenrubí nd Orogrós Prmeters TPC H-TAA L-TAA CF Crrizo citrnge b b C-35 citrnge c FA Clemenrubí Orogrós NS *** NS NS NS ** NS ** x ** *** ** ** (LSD). TPC, mg GAE 100 g -1 fw; TAA, mg TE 100 g -1 fw; CF, g 100 g -1 dw Sugrs nd Orgnic Acids The fctor vriety significntly ffected sugrs nd orgnic cids content, while the fctor rootstock vriety only ffected sucrose, citric nd mlic cids (Tble 3). Clemenrubí fruits on C-35 citrnge, FA-5 nd Crrizo citrnge hd significntly more glucose nd fructose tht Orogrós fruits. However, significnt interction rootstock vriety ws found for sucrose of the fruit juice, so the Orogrós fruits hd more sucrose tht Clemenrubí fruits grfted on C-35 citrnge nd FA-5 (Figure 1g). The lowest citric cid vlues were obtined on Crrizo citrnge, while the highest citric cid vlues were obtined on C-35 citrnge nd FA-5 (Tble 3). The Clemenrubí fruits grfted on C-35 citrnge showed the higher citric cid (Figure 1h). The highest mlic cid vlues were recorder in Clemenrubí nd Orogrós trees grfted on Crrizo citrnge (Figure 1i). Our results show how interction rootstock vriety influence sugrs nd orgnic cids of Clementine Totl Antioxidnt Activity, Totl Phenolic Content nd Crude Fiber The effects of rootstock/scion combintions on ntioxidnt ctivity, totl phenolic nd crude fibre of clementines hve not been studied in systemtic nd comprehensive mnner previously. The results obtined indicte tht the interction rootstock vriety significntly ffected to ntioxidnt ctivity, totl phenolic nd crude fiber (Tble 4 nd Figure 1 j,k,l,m). The highest TPC vlues were recorded in Orogrós trees grfted on C-35 citrnge nd FA-5, however the Clemenrubí trees recorded the higher vlues of TPC grfted on Crrizo citrnge (Figure 1 j). Results from TAA (both H-TAA nd L-TAA) showed tht the rootstock FA-5 induced the higher H-TAA for Clemenrubí fruits while tht it induced the lower H-TAA for Orogrós fruits (Figure 1 k); however the rootstock Crrizo citrnge induced the higher L-TAA for Clemenrubí fruits nd the lower for Orogrós fruits (Figure 1 l). The crude fiber content lso ws ffected by

7 Journl of Food nd Nutrition Reserch 551 interction rootstock vriety (Tble 4, Figure 1 m). In this cse, lso the rootstock FA-5 induced the higher crude fiber content for Clemenrubí fruits while tht it induced the lower vlues of crude fiber for Orogrós fruits (Figure 1 m). These results show the high degree of interction between rootstock/scion Identifiction nd Quntifiction of Phenolic Compounds Flvonoids re widely distributed in fruits nd ech species is chrcterized by prticulr flvnone glycoside pttern [12]. Severl uthors hve evluted the effect of the rootstock in reltion to flvonoid content in Citrus sp. nd observed tht it could be n importnt gronomic fctor in totl flvonoid content [25]. In this study six flvonoids re detected in clementines cv. Clemenrubí nd Orogrós (Figure 2). The interction rootstock vriety significntly ffected to flvnones nd C-glucosides. As shown in Tble 5, the Orogrós cv, grfted onto FA-5 rootstock presented the highest mounts of Api-6,8-di-C-glc, Nr-7-O-rut, nd Hes-7-O-r flvonoids (Figure 1n, o, nd p). In this study, we found significntly differences between rootstock in reltion to flvonoid content. So, the rootstock FA-5 produced the mjor flvonoid content; while Crrizo citrnge nd C-35 citrnge rootstocks showed similr tendencies with not significnt differences. Depending on the fruit development stge, the presence nd /or concentrtions of flvonoids cn be ffected. So, most Citrus species ccumulte substntil quntities of flvonoids during their orgn development [26]. Tble 5. Effect of citrus rootstocks on polyphenols profile (Flvnones nd C-glucosides) of clementines cv. Clemenrubí nd Orogrós Prmeters Api-6,8-di-C-glc Dios-6-C-glc Api-7-O-rut Nr-7-O-rut Hes-7-O-rut Didymin Crrizo citrnge b b b b 7.8 b b C-35 citrnge b b b 9.6 b b FA Clemenrubí Orogrós ** * * * ** NS ** * NS * NS NS x *** NS NS * ** NS (LSD) Figure 2. Chromtogrm of polyphenolic profile (Flvnones nd C-glucosides) of clementines (1 Api-6,8-di-C-glc (11.6 min); 2 Dios-6-C-glc (12.9 min); 3 Didymin (16.0 min); 4 Nr-7-O-rut (17.8 min); 5 Api-7-O- rut (18.2 min); 6 Hes-7-O-rut (19.5 min))

8 552 Journl of Food nd Nutrition Reserch 4. Conclusions To our knowledge this is the first study tht exmined the influence of citrus rootstocks (Crrizo citrnge, C-35 citrnge nd F-5) in bioctive compounds of clementines cv. Clemenrubí nd Orogrós. The present results indicte tht the effect of rootstock on the physicochemicl prmeters, ntioxidnt ctivity, totl phenolic, sugrs nd orgnic cids nd flvonoids of clementines fruits is rther complex phenomenon tht gretly depends on specific interctions between the rootstock nd ech prticulr scion vriety. So, C-35 citrnge rootstock produced the heviest nd bigger size fruits, while the rootstock FA-5 produced the fruit with the mjor flvonoid content. Acknowledgements This work ws funded by the Instituto Ncionl de Investigción y Tecnologí Agrri y Alimentri (RTA ), Generlitt Vlencin, nd FEDER funds. References [1] FAO (Food nd Agriculture Orgniztion of the United Ntions). FAOSTAT [on line] Avilble: [Accessed My 17, 2017]. [2] Cstle WS. Creer perspective on citrus rootstocks, their development nd commerciliztion. HortScience 45: 11-15, [3] Cstle WS, Bldwin JC, Murro RP, Littel R. Performnce of Vlenci sweet ornge trees on 12 rootstocks t two loctions nd n economic interprettion s bsis for rootstock selection. HortScience 45: , 2010b. [4] Cnturis-Aviles, T, Muro FAA, Stuchi ES, d Silv SR, Espinoz-Nuñez E. Tree performnce nd fruit yield nd qulity of Okitsu Stsum mndrin grfted on 12 rootstocks. Sci Hortic 113: , [5] Benjmin G, Tietel Z, Port R. Effects of rootstock/scion combintions on the flvor of citrus fruit. J Agric Food Chem. 61: , [6] MAPA (2013). Estdístics Agrris [on line] Avilble: s/gricultur/defult.spx [Accessed My 2, 2017]. [7] Cmeron JW, Soost RK. C35 nd C32: citrnge rootstocks for citrus. HortScience 21: , [8] Forner-Giner MA, Primo-Millo E, Forner JB. Performnce of Forner-Alcide 5 nd Forner-Alcide 13, Hybrids of Cleoptr Mndrin Poncirus Trifolit, As Slinity-Tolernt Citrus s. 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