Enzymatic activity of three sugarcane varieties under salt stress

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1 ISSN Revist rsileir de Engenhri gríol e mientl v., n.9, p.86-8, 6 Cmpin Grnde, P, UE/UFCG DOI: Enzymti tivity of three sugrne vrieties under slt stress Meri F. Crvlho, Mrus M. Corre, Gin C. Crvlho, Fernndo C. Rolim Neto, Gessi P.. Mrinho & Sunmit. de ndrde Universidde Federl Rurl de Pernmuo/Deprtmento de Tenologi Rurl/Progrm de Pós-grdução em Engenhri mientl. Reife, PE. E-mil: meri.fonse@hotmil.om (Corresponding uthor); metri@dtr.ufrpe.r; fernndortxo@yhoo.om.r Instituto Federl de Edução Ciêni e Tenologi do Piuí/Deprtmento de Tenologi gríol. Vlenç, PI. E-mil: ginrvlho@gmil.om Universidde Federl Rurl de Pernmuo/Unidde dêmi de Engenhri gríol mientl. Reife, PE. E-mil: gessi.mrinho99@gmil.om; mriosntos9@hotmil.om.r Key words: slinity ntioxidnt enzymes resistne S T R C T Sugrne is speies of gret eonomi importne to rzil. The study of enzymti tivity hs helped to identify vrieties of sugrne tolernt to slt stress, for plnting in rzilin semirid regions, where there is the ourrene of sline soils. The ojetive of this study ws to evlute the enzymti tivity of three vrieties of sugrne sujeted to stress used y slinity. The experiment ws onduted in greenhouse t the Federl Rurl University of Pernmuo, in ompletely rndomized design with four replites. The lef lde smples were olleted for nlysis of solule proteins nd determintion of the tivity of tlse nd sorte peroxidse. The protein ontent ws higher for R9696 nd lower for the vrieties R9579 nd R However, in R86755, there ws n inrese in the tivity of sorte peroxidse nd tlse, while the vrieties R9579 nd R9696 showed redution in the tivity of these enzymes. These results indite tht the vriety R86755 hs more effiient mehnism in plnt protetion ginst the tion of retive oxygen speies. Plvrs-hve: slinidde enzims ntioxidntes resistêni tividde enzimáti de três vrieddes de n-de-çúr so estresse slino R E S U M O n-de-çúr é um espéie de grnde importâni eonômi pr o rsil. O estudo d tividde enzimáti tem juddo identifir vrieddes de n-de-çúr tolerntes o estresse slino pr plntio ns regiões semiárids rsileirs ns quis oorre inidêni de solos slinizdos. Ojetivou-se, neste estudo, vlir tividde enzimáti de três vrieddes de n-de-çúr sumetids o estresse usdo pel slinidde. O experimento foi onduzido em s de vegetção n Universidde Federl Rurl de Pernmuo. Foi feito um delinemento inteirmente sulizdo, om qutro repetições. Form oletds mostrs do limo folir pr nálise ds proteíns solúveis e determinção d tividde ds enzims tálse e sorto peroxidse. O teor de proteíns foi mior pr vriedde R9696 nd menor pr s vrieddes R9579 e R86755; entretnto se pereeu, n vriedde R86755, umento d tividde d sorto peroxidse e d tálse enqunto que ns vrieddes R9579 e R9696 oorreu um redução d tividde desss enzims ujos resultdos indim que vriedde R86755 present um menismo mis efiiente n proteção d plnt ontr ção de espéies retivs de oxigênio. Ref. 4-5 Reeived 7 Sep, 5 epted 4 Jul, 6 Pulished ug, 6

2 Enzymti tivity of three sugrne vrieties under slt stress 87 Introdution Sugrne is the min rw mteril for the prodution of sugr nd lohol (Mrin & Nssif, ). It hs environmentl importne for produing ethnol nd showing high tmospheri C fixtion rte, for prolonged period of time, nturlly ontriuting to the improvement of the environment nd reduing the greenhouse effet used y the urning of fossil fuels (Gllrdo & ond, ; Sundfeld & Mhdo, ). In northest of rzil, where sugrne prodution is very signifint (CON, 4), the re of soils degrded y slinity nd sodiity hs inresed due to the expnsion of irrigted res in mrginl lnds, indequte mngement of wter nd soil, defiient dringe nd the use of sline wters in irrigtion. It is known tht the effet of slinity on plnts uses loss of qulity nd yield, ffeting plnt growth through osmoti stress nd toxiity used y the exessive umultion of slts in the leves (Siriporndulsil et l., ). s resistne to the stress, plnts rete ntioxidnt defense mehnisms ple of neutrlizing the ytotoxiity of retive oxygen speies (ROS) (grwl & Pndey, 4). mong the vrious enzymes tht t in this system, tlses (CT), sorte peroxidse (PX) nd superoxide dismutses (SODs) stnd out (Miller et l., ). Through the tivity of the enzymes, plnts protet their ells from the toxi effets of ROS (Foyer & Shigeok, ; ros et l., 4). This study imed to evlute the responses of the ntioxidnt system, sugrne vrieties nd the defense mehnisms of these plnts to the slt stress, nd identify vrieties more tolernt to this stress. Mteril nd Methods The experiment ws rried out in greenhouse t the Federl Rurl University of Pernmuo (UFRPE), in Reife- PE, rzil, using the illets of three sugrne vrieties tolernt to slinity: R9579, R9696 nd R86755; provided y the Sugrne Experimentl Sttion of Crpin (EECC). The ompletely rndomized experimentl design ws used in x 5 ftoril sheme, whih orresponded to three sugrne vrieties whose seedlings were irrigted with five levels of slinity (, 5,, 5 nd mm of NCl); eh tretment omprised four replites. Plnts were limted for dys nd potssium nd nitrogen fertiliztions were performed ording to rop needs, sed on the Mnul of Fertiliztion for the stte of Pernmuo (IP, 8). In order to determine the replenishments with sline solutions, the pot pity weight ws determined nd, fter fixed irrigtion intervl of four dys, the pots were weighed gin. The differene etween the redings (kg) determined the volume of solution (L) to e pplied in eh irrigtion event. For the enzymti nlyses, the leves were olleted fter thirty dys, when smples of the lef lde of the third fully expnded lef were olleted, ounting from the pex to the se of the plnt, immeditely stored nd luminum pper envelopes nd immersed in liquid nitrogen (LN ) for fst freezing of the mteril. Then, ll the mteril ws stored in freezer t - ºC until the nlyses. One extrtion of proteins ws performed using smples tht were frozen, merted nd homogenized with uffer of mm potssium phosphte (ph 7.5), mm of EDT (ethylenediminetetreti id), mm DL-dithiothreitol nd % PVPP (polyvinyl polypyrrolidone) t temperture of -4 ºC in the reltionship of. g Ml - (w:v) (zevedo et l., 998). The homogenized mteril ws entrifuged t, rpm for 5 min. The superntnt solution ws stored t - ºC nd used s soure for the determintion of totl proteins (TP) nd tivity of the enzymes tlse (CT) nd sorte peroxidse (PX). Enzymti tivities were expressed in enzymti unit. One unit of the enzyme is defined s the mount of enzyme tht uses n inrese of. sorne unit per minute, expressed s U min - mg - of solule protein. The onentrtion of solule proteins ws determined using S (ovine serum lumin) s the stndrd. The retion ws performed with the ddition of µl of the smple (previously diluted) t ml of the regent rdford nd inuted t mient temperture, for 5 min. Then, the reding ws performed in spetrophotometer t 595 nm. CT tivity ws determined through the method desried y zevedo et l. (998) with modifitions, in solution ontining ml of the uffer mm potssium phosphte (ph 7.5) nd 5 µl of hydrogen peroxide (H ) t mm. The retion ws initited y the ddition of 5 µl of the protein extrt nd the tivity ws determined through the deomposition of H for 6 s, sed on the ltertions t 4 nm t the temperture of 5 ºC, in spetrophotometer. PX tivity ws determined through retion omposed of 65 μl of the uffer 8 mm potssium phosphte (ph 7.5), μl of 5 mm sorte, μl of M EDT, μl of mm H nd 5 μl of the protein extrt. PX tivity ws determined through the monitoring of sorte oxidtion rte t 9 nm, t ºC, for 6 s, in spetrophotometer. The dt were sujeted to nlysis of vrine (NOV), using the progrm SISVR. For ll dt, the quntittive ftor reltive to the slinity levels ws sttistilly nlyzed through regression, iming the fit of equtions in order to orrelte the slinity levels with the studied vriles. The mens of the tretments were ompred y Tukey test t.5 proility level. Results nd Disussion The results of the evlutions of solule proteins (SP) nd enzymti tivity (tlse (CT), sorte peroxidse (PX)) re respetively presented in Figures to nd Tle. Considering the evlution of solule proteins, the vriety R9696 sttistilly differed from R9579 nd R86755 (Tle nd Figure ). The inrese in slinity my use different responses mong the protein ontents. These differenes my e relted to intrinsi hrteristis of the genotypes, s reported y Siriporndulsil et l. (). Figure nd Tle show men redution of 58% in the protein ontent for the tretment of highest sline onentrtion R. rs. Eng. grí. mientl, v., n.9, p.86-8, 6.

3 88 Meri F. Crvlho et l. Tle. Evlution of solule proteins nd the enzymti tivity of tlse nd sorte peroxidse in sugrne vrieties sujeted to different slinity levels Vriety NCl levels (mm) 5 5 Regression eqution R² Solule proteins (mg g - of DM) R y =.x -.77x R d y = 4E-5x -.4x R y = 5E-5x -.57x Ctlse (µmol H mg - of protein min - ) R y = -.6x +.65x R y =.55x +.75x R y = -.484x +.65x +..6 sorte peroxidse (µmol H mg - of protein min - ) R y = -.649x + 7.9x R y =.86x +.98x R y = -.6x x Sme lowerse letters etween vrieties do not differ y Tukey test (p <.5) 7.. SP (mg g - de DM) CT (µmol H mg - of proteinmin - ) C SP (mg g - de MF) R9579 R9696 R86755 R9579 R9696 R CT (µmol H mg - of proteinmin - ) R9579 R9696 R86755 R9579 R9696 R Sline onentrtion of NCl (mm) Mens of vriety followed y sme letters do not differ y Tukey test t.5 proility level Figure. Evlution of solule proteins (SP) (mg g - of DM) of sugrne leves. () Comprison etween SP vlues otined for the vrieties sujeted to different levels of slt stress; () Comprison etween SP vlues otined for the vrieties t eh studied sline level ( mm), ompred with the ontrol. ris et l. (996), for sugrne, oserved redution of 54.% in the umultion of solule proteins, for the highest level of wter defiit. Silv (), in studies with six sugrne genotypes sujeted to wter stress, oserved men redution of 6% in solule proteins for the severe stress, in reltion to the ontrol. Prid & Ds (5) reported tht, under stress onditions, the protein ontent in the plnts my derese due to the inrese of proteolysis. In ddition, ording to Gill & Tutej 5 5 Sline onentrtion of NCl (mm) Mens of vriety followed y sme letters do not differ y Tukey test t.5 proility level Figure. Evlution of the tlse (CT) tivity (µmol H mg - of protein min - ) of sugrne leves. () Comprison etween CT vlues otined for the vrieties sujeted to different levels of slt stress; () Comprison etween CT vlues otined for the vrieties t eh studied sline level (), ROS umultion my severely ffet ell funtionlity nd integrity with ltertions in redox homeostsis, dmges to mromoleules nd denturtion of proteins. Cstrillo et l. () highlighted tht, t ellulr level, proteins n e dmged y the lk of hydrtion, whih promotes n inrese of retive oxygen speies. In the evlution of tlse nd sorte peroxidse enzymti tivities (Tle nd Figures nd ( nd )), R. rs. Eng. grí. mientl, v., n.9, p.86-8, 6.

4 Enzymti tivity of three sugrne vrieties under slt stress 89. PX (µmol H mg - of prote i nmin - ). PX (µmol H mg - of proteinmin - ) R9579 R9696 R86755 R9579 R9696 R86755 the vriety R86755 promoted higher vlues ompred with R9579 nd R9696, inditing its greter pity of dpttion to slt stress. The vriety R9696 presented the lowest vlue nd, therefore, lower pity of dpttion to stress. These dt orroorte those oserved y Willdino et l. (), who evluted the slt stress on the vriety R8655 nd oserved more effiient mehnism in the protetion of the plnt ginst the tion of retive oxygen speies, prtiulrly H, due to the inrese in the tivities of the enzymes CT nd PX. The inrese in PX nd CT tivity indites the importne of these enzymes in the detoxifition of H nd protetion ginst oxidtive dmges (Ci et l., ). Moris (), studying the omined tion of ioti stress ftors, reported tht the enzymti ehvior of the vriety R86755 ws effiient in the regultion nd detoxifition of the ROS produed in the plnt ell, inditing greter tolerne of this vriety to the different types of stress. Figures nd nd Tle present the results of the enzymes CT nd PX, respetively, ompring the vrieties nd the pplied sline levels. In generl, there ws n inrese in PX nd CT tivity in ll vrieties when sujeted to mm NCl. Deuner et l. () oserved tht seeds of owpe genotypes mintin high germintion nd PX nd CT tivity until the NCl onentrtion of mm. The C 5 5 Sline onentrtion of NCl (mm) Mens of vriety followed y sme letters do not differ y Tukey test t.5 proility level Figure. Evlution of the sorte peroxidse (PX) tivity (µmol H mg - of protein min - ) of sugrne leves. () Comprison etween PX vlues otined for the vrieties sujeted to different levels of slt stress; () Comprison etween PX vlues otined for the vrieties t eh sline level vriety R86755 showed higher tion of these enzymes in the repir of dmges used y slinity, due to the signifint inrese in PX nd CT tivity, in omprison to the results for R9579 nd R9696. sed on the results found for the ontrol nd the NCl onentrtion of 5 mm, the lower the stress, the lower the tivity of these enzymes, s reported y zevedo (). Mny uthors hve oserved tht the inrement in slinity uses inrese in the tivity of the enzymes CT nd PX, leding to redution in the deleterious effets of H (Mi et l., ; ogdllh et l., ). However, Crvlho et l. (), studying the limtion of rie plnts to slt stress, oserved redution of CT when plnts were exposed to NCl. ording to zevedo et l. (), the pity of tivtion of the ntioxidnt system is one of the min mehnisms tht llow the plnt to tolerte the oxidtive stress. Silv (), nlyzing six sugrne genotypes sujeted to severe wter stress, oserved response of up to.% in PX tivity. Quintel et l. (5), evluting wter stress intensity nd durtion on the pthosystem sugrne x Meloidogyne inognit, oserved n inrese in PX tivity without ffeting CT tivity in the tested wter tretments. In mize nd soyen rops, the enzymes CT nd PX ted in similr wy under extreme situtions, inresing s result of the severity of the pplied stress (Vsonelos et l., 9). zevedo () oserved inrese in CT nd PX tivity in sugrne for inresing levels of slinity. Conlusions. In the initil growth stge, the sugrne vriety R86755 showed higher tivity of the enzymes tlse nd sorte peroxidse, in omprison to R9696 nd R9579, whih is n indition of its higher tolerne to slinity.. t the sline onentrtion of mm NCl, there ws greter prodution of the enzymes tlse nd sorte peroxidse for the vriety R The vriety R9696 showed higher vlue of solule proteins ompred with the other studied vrieties. Literture Cited ogdllh, G. M.; Serg, M. M.; Quik, P. W. Fine nd orse regultion of retive oxygen speies in the slt tolernt mutnts of rnyrd grss nd their wild-type prents under slt stress. Physiologi Plntrum, v.8, p.6-7,. org/./j x grwl, S.; Pndey, V. ntioxidnt enzyme responses to NCl stress in Cssi ngustifoli. iologi Plntrum, v.48, p , 4. ris, M. I..; Delgdo, E. O.; Crmente, R. V. Cmios fisiológios de l ñ de zúr nte el défiit hídrio. Méxio: Universidd utónom Chpingo..ed p. zevedo, R..; ls, R. M.; Smith, R. J.; Le, P. J. Response of ntioxidnt enzymes to trnsfer from elevted ron dioxide to ir nd ozone fumigtion, in leves nd roots of wild type nd tlse-defiient mutnt of rley. Plnt Physiology, v.4, p.8-9, R. rs. Eng. grí. mientl, v., n.9, p.86-8, 6.

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