Abstract. 1. Introduction. Journal of Nutrition & Pharmacy Research

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1 Tomto Fruits (Solncee Lycopersicon esculentum Mill.) Feedck Mechnism in The Presence of Exogenous Ethylene under Prolonged Chilling Temperture Storge M. Mkky,, *, K. A. Pschlidis, P. Soni, A. Mngnris, K. Dhim, I. Vssilkoglou, G. Mnggnris 5. School of Agric. Tech., Alexnder Technologicl Eductionl Institution of Thessloniki, 57, Greece.. Agriculturl Systems & Engineering, SERD, Asin Institute of Technology, Pthumthni,, Thilnd.. Dept. of Agriculturl Engineering, Andls University, Pdng, 56, West Sumter, Indonesi.. Technologicl Eductionl Institutions of Thessly, Lriss, Greece. 5. Dept. of Agric. Sciences, Biotech. & Food Science, Cyprus University of Technology, Limssol, Cyprus. *E-mil of the corresponding uthor: Astrct Green mture nd red ripe tomto Rom VF fruits were stored in ir-tight glss jrs (ml), t 5 o C for three weeks in drk. Grouped y similrity in color, three fruits were put into ech jr. Hlf numers of the jrs were injected with ml of.5 ppm Ethylene gs, while the other hlf ws used s control. Four rndom sets of smples, with three replictions, were mesured t 7 dy intervls within three weeks. Upon mesurement, jrs were put into o C room temperture for minutes, nd then the upper tmosphere inside the jr ws nlyzed using gs chromtogrm. The fruits physiology inside ech jr were mesured (skin nd pericrp color nd hrdness, storing nd peeling weight loss, dimension nd trnsversl cross section, solule solid content, nd trittle cidity). The result showed tht red ripe tomto fruits Rom VF stored in 5 o C with exogenous ethylene hve superior ppernce, higher respirtion production nd higher senescent rte s compred to the control. The green mture fruits hd the opposite effect when treted with ethylene, possily due to n uto-induction mechnism in the synthesis process of the enzymes tht re involved in ethylene iosynthesis controlled y the phytohormone ethylene. Thus, green mture fruits, when exposed to CH, reveled negtive feedck mechnism. Keywords: Tomto, CH, Cold Storge, Feedck Mechnism, Auto-Induction Mechnism.. Introduction Most of the fresh products, including tomtoes, were sent y mens of cold-chin (Rodrigue nd Notteoom, ), to prolong products fresh life, nd mintin the fruits in fresh condition t the destintion. Recent studies hve identified tht minimum storge temperture for the tomtoes were round to 5 o C for red ripe fruits (Sltveit, 997) nd to o C for mture-green (Wng, ). The prolems come when tomto were trnsported in mix with the other griculturl products, especilly

2 certin vegetles tht hve lower optimum storge temperture (i.e. 5 o C) (The Engineering ToolBox, ). This condition my led tomto to chilling injury (CI), especilly when exposed to prolonged period of chilling temperture (Wng, ). In this condition, vrious physiologicl nd iochemicl ltertions nd cellulr dysfunctions occur (Rison nd Orr, 99), nd when fruits removed to wrmer tempertures, the chilling injuries symptoms will ecome evident in short time (Wng ), susequently reduced the fruits qulity (Cheng nd Shewfelt, 988), nd mde it susceptile to decy (Mul et l, ). Nevertheless, chilling sensitivity in tomtoes my differentitions ccording to storge temperture nd durtion of exposure (Wng, ), s well s mturity nd degree of ripeness. Over the yers, severl methods in lleviting the chilling injury in tomtoes hve een tried nd hve shown positive results (Wng, ). These methods include short-term hot (Akudk nd Akudk, 7) or cold wter tretments (Ghrezi et l., ), long-term ir het tretments (McDonld et l., 996; Lurie nd Seht, 997) in comintion with ethylene tretment (McDonld et l., 998), intermittent temperture tretment (Artes et l., 998), dipped in chemicl solutions (Ghrezi et l., ), Controlled Atmospheric storge (Tsdelen nd Byindirli, 998), s well s other chemicl pplictions or genetic mnipultion (Meir et l., 996). Nevertheless, these methods require certin preconditions tht increse shipping cost nd wste considerle time of vlule fresh life of the fruits. In ddition, due to the incresing concern for the environment nd helth, options of wxing or chemicl coting to the fruits were limited. On the other hnd, ethylene gsifiction of tomto fruits during storge or shipping show promising future nd could e used s one of possile solutions in reducing the CI risks. The ojectives of this study were to investigte the physicl chnges nd respirtion effects of industril tomto cv. Rom VF upon storing with ethylene tretment in controlled tmospheric condition in 5 ºC temperture, tht is optimum for the trnsport nd storge of mny fruits nd vegetles growth in the Mediterrnen, under drk condition, for weeks. The differences in the prmeters chnge; fruit color, weight, dimensions, hrdness, solule solid content, cidity, nd its climcteric process (vi ripening gses), were oserved in 7 dys intervl until dys.. Mterils nd Methods. Fruits Smples Tomtoes fruits (cv. Rom VF) with uniform size nd mss, hrvested under norml field conditions from ripeness ctegories: red ripe nd green ripe (Fig.) were selected. Complete rndomized lock design ws used to crry out four tretments with three replictions. All smples were clened nd seprted into two groups; red ripe nd green ripe tomtoes. The first two groups of red ripe nd green ripe fruits were used s controls, while the second groups were treted with modified tmosphere condition (MAC). Initil condition of ech fruit ws mesured nd weighed y using n nlyticl lnce (ALS -, KERN & Sohn, Germny), efore pplying the tretment, which served s its initil weight. Fig. Tomto smple with different ripeness, nd 6 were the two selected ripeness ctegories representing the green ripe nd the red ripe, respectively.. Physiology Mesurements 5

3 Fruits color ws recorded in three points long its equtoril with o seprtion distnce using Chrom meter (CIE L) nd expressed s color spce L*, *, * mode. The color mesured in color index s rtio of * nd * (McDonld et l., 996; 998). Air-tight glss jrs continers (ml) were prepred to store the fruits. Every jr s cp ws drilled to crete mm dimeter hole to enle collecting the gs inside the jr for smpling. To void gs inside the jr leked during storge, the hole ws re-seled with trnsprent silicon. This enles the opening to e esily identified nd used to collect gs smple inside the jr y syringe. All jrs were numered; one from ech group will e used to mesure the smples in ech week for three week storge. Jrs of groups nd were used for the red ripe control nd green ripe control smple, while group nd were used for the ethylene treted red ripe nd green ripe fruits. Three tomtoes with similr size, weight, nd color were put inside ech jr. For jrs of group nd, green ripe nd red ripe tomto fruits were put inside, nd then, ech continer ws injected with ml of.5 ppm ethylene gs through the prepred hole on the cps, nd reseled with silicon fterwrd. During storge, ech fruit will e mesured to ssess its climcteric process y men of ssessing its skin nd pericrp; color nd hrdness development, its weight loss; during storge nd fter peeled, the dimension of the fruits nd its trnsversl cross section, solule solid content (SSC), s well s cids level in the fruits using titrtion. The ssessment ws done t 7 dys intervl until dys.. Gs Chromtogrphy Anlyses For the ethylene nd respirtion mesurement of the fruits, ll jrs were kept in inside refrigerted drk nd the temperture ws mintined t 5 o C. On zero, 7,, nd dy, three jrs from ech group ws put into o C room temperture for minutes, nd then one ml gs of the upper tmosphere inside the jr ws drwn with gs-grde syringe (Gold Syringe, Agilent Technology, USA) nd mesured using Gs Chromtogrm mchine (GC system 789B, Agilent Technology, USA). The result of GC mesurement from ech smple ws compred with the stndrd (.5 ppm Ethylene) provided from Agilent. All results were nlyzed y quntittive nd qulittively... Physiology Chnges Assessment After the hedspce gs inside ech continer ws mesured using GC mchine, the continer ws opened, nd the tomto fruits inside ech continer were ssessed. The fruits were re-weighed using Anlyticl lnce (ALS -, KERN & Sohn, Germny), to determine its physiologicl weight loss (Moneruzzmn et l., 9). The fruits colors were mesured y Chrom meter (CR-, Minolt, Jpn) (Luengwili nd Beckles, ). Three points round equtor for ech fruit ws mesured in color using CIE L stndrd chromtogrm, nd verge. The sme mesured points were then peeled to expose its pericrp, nd the color ws recorded nd verge. The color index of the fruits skin nd its pericrp will e used to compre the effects of the tretment to the smples (Roerts et l., ). A stndrd peeler ws used to peel the fruits. The fruit nd its peel were mesured to compre the weight loss during peeling process. The hrdness of the peel nd pericrp from ech fruit were mesured using penetrometer (FT7, Itly) nd the result ws recorded in kg.mm - (Ghrezi et l., ). The peeled fruits were then cut trnsverslly into two prts long its equtor line to ssess the internl ppernce (equtoril section) of fruits pericrp color nd thickness, numer of locules nd its gel formtion, nd its septum width (Kder nd Morris, 976). The peeled fruit ws then individully liquidted using mechnic juicer, nd its SSC ws mesured using refrct meter (ATC-, Atgo, Jpn) (Ghrezi et l., ). Totl trittle cidity ws mesured y tken 5ml of ech fruits liquid, nd then dded with three drops of dye Indictor (phenolphthlein) nd mesured using utomtic titrtion mchine (Tritomtic 5, Crison, Spin) to mesure its cidity (Nir nd Singh, ). The fruits flvor ws mesured y the rtio of SSC nd totl titrtle cid, clculted y dividing SSC percent with the corresponding cid percent nd expressed s rtio (Bldwin et l., 998). 6

4 .5 Sttisticl Anlyses Descriptive sttisticl nlyses y mens of Anlysis of vrince (ANOVA) were used to nlyze the differences etween tretments nd control s well s their vrition mong nd etween groups. The dt pertining to physiologicl nd iochemicl prmeters of the fruit were sujected to sttisticl nlysis using nlysis of vrince (ANOVA) ccording to Fischer s. The level of significnce used in F test nd t test ws set t p.5. Criticl difference vlues were clculted wherever the F test ws found significnt (Ghrezi et l., ). The dt were sujected to SPSS Sttisticl Softwre (IBM Inc., USA).. Results nd Discussion The storing of fruits in lower temperture condition usully suppresses the climcteric nd mny relted ctivities in fruits, ut mkes the fruits prone to chilling injury (Yun et l., ). Adding exogenous ethylene to the storge room might llevite chilling stress to the fruits; however, difference in ripeness stge my ffect the sensitivity of fruits to the ethylene presence (Att-Aly et l., ). In this study, ll of these conditions were set nd ll of the prmeters were mesured. Two groups of tomtoes fruits (c.v. Rom VF) were oserved in 7 dys intervl until dys. Stored t 5 o C in ir seled trnsprent continers t drk for three weeks, ll physicl nd chemicl chnge in ech group ws monitored weekly. The chnge of color in fruits, s well s its pericrp, cn e seen in Fig.. The results indicted tht exogenous ethylene tretment did not significntly mke difference in color development for red ripe tomto under chilled storge, compred to control. It ws oserved oth in fruits skin (Fig. ) nd pericrp (Fig. ), even though in generl, ethylene treted fruits hve slightly less red color s compred to control. Nevertheless, the fruits continue to further develop into fully ripe stge, however, s oserved; color degrdtion ws present in the pericrp fter the second week of storge. On the other hnd, significnt difference ws showed in green-mture fruits. The presence of exogenous ethylene cused the green-mture fruits unle to further develop into reker stge (USDA, 99). The result showed tht different ripeness condition of fruits gve different response to their ripening progress when expose to exogenous ethylene in comintion with chilling temperture storge. Filed to ripen or de-green ws one of indictors of chilling injury (Wng, ).. Greener Color Index ( * / * ) Redder Fruits Color Development Greener Color Index ( * / * ) Redder Pericrp Color Development Fig.. The color ltertion of the smples, mesured y the rtio of * nd * (Color Index) from initil condition until the third week of storge, mesured on () surfce, nd () the pericrp, for ll tretments. 7

5 Weight Loss (%) Fruits Weight loss Weight Loss (%) 5 Peeled Weight loss.5 Fig.. The smples weigh loss when stored for three weeks (), nd fter peeling process (), for ll tretments. Product loss ws nother concern upon storing nd processing crops nd griculturl products for long period. Cold storge hs een known to prevent such losses, nd while comined with ethylene tretments, the results might give vrince ccording to the products types nd conditions. In this study, results showed positive correltion etween storing time nd product weight loss s well s loss upon peeling process (Fig. ). For the red ripe tomtoes, ethylene tretments showed no significnt weight loss upon storge compred to control (Fig. ), ut higher loss oserved fter peeled (Fig. ). On the other hnd, green mture fruits were significntly ffected y the ethylene tretment. The results oserved oth in totl weight loss (Fig. ) nd totl loss fter peeling (Fig. ) were significntly different compred to control. The rte of loss incresed exponentilly fter fruits stored for two weeks. The results once gin showed tht different ripeness stges of fruit significntly ffect the result of exogenous ethylene presence in cold storge condition. The results gree with previous reserch showing tht tomtoes were more sensitive to chilling in less mture stge (Pull, 99). Ethylene (nl.gr -.hr - ) 8 6 Red-Ctrl Green-Ctrl Red-Ethyl Green-Ethyl HS (nl.gr -.hr - ) Red-Ctrl Green-Ctrl Red-Ethyl Green-Ethyl CO (nl.gr -.hr - ) Red-Ctrl Green-Ctrl Red-Ethyl Green-Ethyl CO (nl.gr -.hr - ).5.5 Red-Ctrl Green-Ctrl Red-Ethyl Green-Ethyl c d Fig.. The climcteric, respirtion nd decying process of the fruits, mesured y its () Ethylene, () HS nd (c) CO, nd (d) CO production rte respectively using GC. The climcteric process, respirtion nd decying rte of the fruits were mesured using GC. The climcteric process in fruits mesured s fruits ethylene production rte (Fig. ). Fruits decying rte ws determined y the rte of HS gs produced during storge (Fig. ), while the respirtion rte of the fruits mesured s production rte of CO (Fig. c) nd CO (Fig. d). For the red ripe tomtoes, exogenous ethylene ddition to the fruits shorten the urst ethylene production period, ccelerting the norml process of climcteric, ut with comintion of low temperture storge, cused reduction in their respirtion nd senescence rtes compre to control. For the green mture tomtoes however, the ethylene ddition incresed the respirtion rte, while their ethylene production from fruits s well s its 8

6 senescence rtes ws reduced. This might e corresponding to the fruits condition which filed to enter the reker stges due to chilling injury. The results (Fig. ) lso showed tht the trend of climcteric, respirtion nd senescence did not significntly correspond to ech other, thus suggesting tht ech process might e triggered y different prts of the fruits, nd ffected to nother different prt. Previous results showed tht exposing immture tomto fruit to CH reveled negtive feedck mechnism in reltion to its iosynthesis, where ACC concentrtion nd CH production were suppressed, while CO production incresed significntly (Att-Aly et l., ) s well s CO. However, this condition ws vice-vers to red-ripe tomtoes, where ethylene tretment suppresses the CO (Fig. c) nd CO (Fig. d) production. The urst of CH (Fig. ), HS (Fig. ) nd CO (Fig. c) were oserved in control for the first week, while green mture fruits treted with ethylene showed similr urst only in CO production. In this work, negtive feedck mechnism in reltion to iosynthesis ws oserved when mture green tomto ws exposed to exogenous ethylene, suppress its CH production while multiplying respirtion (CO nd CO) production, possily resulting in lower ACC synthse enzyme concentrtions, nd ctlyzing down-regultion of ripening-ssocited genes. This my hve cused the fruits unle to mture..5. Skin Hrdness Pericrp Hrdness kg/mm... kg/mm.... Fig. 5. The chnge of hrdness of the smples peels (), nd pericrp (), of ll tretments mesured with fruits penetrometer during storge. To understnd the ethylene tretment effect to fruits peels nd pericrp, oth prts firmness were mesured. The results (Fig. 5) showed tht over the storing time, the hrdness of fruits peels nd pericrp were reduced. The ethylene tretment in comintion with storing tomtoes in low temperture slightly reduced the peeling hrdness of the red ripe fruits; while significntly produced hrder skin in the green ripe fruits (Fig. 5). As for the pericrp, only green ripe fruits treted with ethylene showed higher hrdness compred to other smples (Fig. 5). possiility explntion would tht since the de-greening process did not tke plce in the ethylene treted green ripe fruits, less chemicl rection might trnsform the chlorophyll in the pericrp to other compounds. And s such, the cells in the pericrp still constitute much less ripe condition, oserved through its color, nd thus mintin its strength longer compred to the green ripe fruits which lredy hd de-greening process (green ripe control) Brix SSC mg/l Totl Titrtle Acids (citric cid) (SSC/Acid).5 Flvour Index 9

7 c Fig. 6. The chnge of two chemicl components indictor of the smples, mesured y mens of the () SSC, nd () Acidity, in reltion to the (c) flvor index, of ll tretments during storge. The chemicl chnges in the fruits were mesured y the chnge of its solule solid content (SSC), cidity nd the gses produced y the fruits during storge. The SSC nd cidity ply importnt role to the tste of the fruits processing product, the correltion etween this two properties cn e identified s the Brix/Acid index. The results showed tht ethylene tretment for red ripe tomtoes did not significntly ffect their SSC (Fig. 6), nd only slightly incresed the cidity level during tretment (Fig. 6). On the other hnd, the SSC levels of green ripe tomtoes were comprly lower when treted with ethylene (Fig. 6), while their cidity levels tend to rise over the time of storge, in opposite to the green ripe control (Fig. 6). The differences of totl SSC nd titrtle cids etween ethylene treted nd non-ethylene treted of oth red nd green ripe tomtoes will led to differences in tste of the smples fter stored for certin time. Since tste ws one of the importnt fctor tht dictte the products qulity in the tomto processing industry, storing the rw mterils efore process hd to e mnged to prevent the negtive effects to the product tste, while mximizing its shelf life. Decresing the storge temperture, storing the products in the drk s well s mnging ethylene gs from externl sources were wys to mintin the fruits qulity nd end product tste upon stored for longer period. In this study, the tste mesured y compring the SSC to the cidity of the smples. The results (Fig. 6c) indicted tht only the green tomtoes with ethylene tretment hve significnt lower SSC to cid rtio fter the second weeks, while for the red tomtoes, their SSC to cid rtio were considered similr with or without the ethylene tretment. The results indicted tht under low temperture storge, fruits mturity determined the chnge in cidity nd tste while ethylene tretment showed less significnt influence. The results in this work showed some significnt distinction nd enefits in hndling industril tomto fruits (c.v. Rom VF), compred to the previous works. 5. Conclusion In this study, red ripe tomto fruits Rom VF stored in 5 o C with exogenous ethylene hd more superior ppernce fter stored for three weeks, compred to non-ethylene treted fruits. The results lso oserved differences in the respirtion production nd senescent rte. On the other hnd, green mture fruits hd the opposite effect when treted with ethylene. The fruits unle to ripen possily due to utoinduction mechnism in the synthesis process of the enzymes tht involved in ethylene iosynthesis, such s ACC synthse (ACS) enzyme nd ACC oxidse (ACO) enzyme. These enzymes controlled y the phyto-hormone ethylene, when exposed to CH reveled negtive feedck mechnism to immture tomto fruit. This is correlted with the condition where less mture fruits were more sensitive to chilling injury, thus gretly influencing its iosynthesis, where ACC concentrtion nd CH production were suppressed, while CO nd CO production incresed significntly. The results of this work lso showed severl eneficil outputs, especilly upon hndling the processing tomto fruits, s compred to different methods in previous works. Acknowledgments The first uthor would like to thnk the Pnce project for providing the PhD moility scholrship under Ersmus Mundus Action progrm, the Directorte Generl of Higher Eduction of Indonesi for providing the PhD fellowship in Asin Institute of Technology, Thilnd, nd to the LPDP Indonesi, for providing funding for his disserttion work.

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