Effects of Ohmic Heating on Lipase Activity, Bioactive Compounds and Antioxidant Activity of Rice Bran

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1 Austrlin Journl of Bsic nd Applied Sciences, 3(4): , 2009 ISSN Effects of Ohmic Heting on Lipse Activity, Bioctive Compounds nd Antioxidnt Activity of Rice Brn P. Loypimi, A. Moonggrm, P. Chottnom 1 Deprtment of Food Technology nd Nutrition, Fculty of Technology, Mhsrkhm University, Mhsrkhm Thilnd Abstrct: The study imed to investigte the effect of stbilistion using ohmic heting (OMH) on lipse ctivity, bioctive compounds nd ntioxidnt ctivity of rice brn. The moisture content of rice brn smples ws djusted to 20, 30 nd 40 %. Alterntive current electricity (50Hz), with three levels of electricl field strengths (EFS) of 75,150 nd 225 Vcm were pplied, in order to stbilise the rice brn. Lipse ctivity, free ftty cid (FFA), nd thiobrbituric cid rective substnces (TBARS) of stbilised rice brn during storge t 4C were investigted. The influence of OMH on the concentrtion of phenolic compound, átocopherol, ãoryznol nd the ntioxidnt ctivity of smples were lso evluted. The results indicted tht OMH ws n effective method for stbilistion of rice brn with moisture ddition. According to this study, the EFS t Vcm nd MC of rice brn 3040 % were the optimum conditions to retrd the incresing of FFA content, lipse ctivity nd lipid oxidtion during storge. Moreover, the stbilised rice brn using OMH yielded the highest levels of phenolic compound, átocopherol, ãoryznol, nd ntioxidnt ctivity. Key words: Ohmic heting, rice brn, lipse ctivity, ntioxidnt ctivity INTRODUCTION Rice brn is the outer lyer of the rice kernel nd by product of the rice milling process. Severl studies hve reported tht rice brn is n excellent source of nutrients nd bioctive compounds including proteins, vitmins, dietry fibers, tocopherols, tocotrienols nd ãoryznol (Qureshi et l., 2002; Ryynnen et l., 2004; Ausmn et l., 2005; Minhjuddin et l., 2005; Imsngun et l., 2008). Rice brn is lso stisfctory source of ft with rnge between 1220% (Mrshll nd Wdsworth, 1994). Since rice brn contins high ft content, rpid deteriortion of crude ft by lipse immeditely occurs, following the milling process nd yields free ftty cids (FFA) nd glycerol. The ft hydrolysis cuses the rice brn unsuitble for humn consumption nd lowers the oil yield (Li et l., 2005). Therefore, immeditely fter the milling process, inctivtion of lipse nd inhibition of the formtion of FFA re necessry. The process tht produces stble rice brn through the dectivting of enzymes which inhibit the deteriortion of rice brn is clled stbilistion. Vrious stbilistion methods, pplied to protect rice bn oil degrdtion, hve been reported such s steming (Julino, 1985; Azrin et l., 2008), extrusion (Kim et l., 1987), microwve heting (Lkkkul et l., 2004); (Rmeznzdeh et l., 2000) nd ohmic heting (Lkkkul et l., 2004). Ohmic heting or Joule heting is food processing method in which n lternting electricl current is pssed through food smple. Ohmic heting is distinguished from other electricl heting method by the presence of electrodes contined in the food; the frequency nd the wveform of the electric field impose between the electrodes. Most foods contin ionic species such s slts nd cids, therefore, electric current cn be mde to pss through food nd generte het inside it (FDA, 2000). The production of n insideout heting pttern mkes ohmic heting much fster thn conventionl outsidein heting. With proper formultion, electricl energy cn be converted to het uniformly in food mtrices. The efficiency of OMH is dependent on the conductive nture of the food to be processed (Zolti nd Sweringen, 1996). As result, OMH provides rpid nd uniform heting nd high qulity product with miniml chnges of structure, nutrition, or orgnoleptic. Moreover, the use of OMH for food processing is clener nd more environmentlly friendly. Potentil pplictions for OMH include blnching, evportion, dehydrtion, fermenttion, nd extrction. Corresponding Author: Anuchit Moongngrm, Deprtment of Food Technology nd Nutrition, Fculty of Technology, Mhsrkhm University, Mhsrkhm Thilnd Tel: Fx: Emil: nuchitc@yhoo.co.th 3642

2 Severl studies hve investigted number of spects for the ppliction of OMH within the food industry, for exmple, its potentil to increse dye diffusion in beet (Hlden et l., 1990), extrction of fruit juices (Lim nd Sstry, 1999; Wng nd Sstry, 2000) nd the enhncement of the drying process of sweet potto (Zong nd Lim, 2003). Lkkkul et l., (2004) reported tht ohmic heting ws n lterntive method for stbilistion of rice brn becuse it incresed the oil yield of rice brn to mximum of 92%, whilst only 53% of oil ws extrcted from the control smples. There hve not been ny investigtions undertken on the effect of ohmic heting on the stbility of lipse, the level of phytochemicls nd ntioxidnt ctivity of rice brn. Therefore this study ws crried out to investigte the effect of ohmic heting on lipse ctivity, to evlute the effect of ohmic heting on phytochemicls nd ntioxidnt ctivity rice brn obtin from different ohmic heting conditions. MATERIALS AND METHODS Chemicls: Stndrd átocopherol ws purchsed from SigmAldrich Chemicl Co., (St. Louis, Mo, USA). HPLC grde methnol, cetonitrile, hexne, ethyl cette nd ethnol were purchsed from BHD (Poole, UK). Gmm oryznol stndrd ws purchsed from Tsuno Food Industril Co., Ltd. (Wkym, Jpn) Gllic cid, 1,1 diphenyl2 picrylhydrzyl (DPPH), bythyltedhydroxynisole (BHA) nd buthyltedhydroxytoluene (BHT) were obtined from Fluk Chemicl (Buchi, Switzerlnd). All chemicls nd regents were nlyticl grde. Rice Brn Preprtion: Rice brn (Oryz stiv L. CV. RD6 smples were purchsed from rice milling fctory in Mhsrkhm province. Freshly milled rice brn ws immeditely pssed through 20 mesh sieve to remove broken rice nd husks. The initil moisture content of rice brn ws determined (AOAC, 2002) before subjecting to stbilistion. Rice Brn Stbilistion Using Ohmic Heting: The moisture content of the smples ws djusted to three different levels: 20, 30 nd 40 % by dding deionized wter (Rmeznzdeh et l., 2000) nd then they were heted by ohmic heting. This stbilistion process of rice brn ws finished within 12 h. The ohmic heting unit ws lbscle ohmic heter equipped with titnium electrodes nd enclosed on Teflon tee. The 180g of rice brn smple ws plced between the electrodes. An lternting current of 50 Hz with three different levels of electricl field strengths (EFS) (75, 150, nd 225 Vcm ) ws pplied. A 1.5 KW cpcity dielectric heter operting t MHz ws used. Two electrodes, size 40x30 cm, were connected to the dielectric heter. The bottom electrode ws mde from 20 guge luminum sheet, whilst the top electrode ws mde from 20 guge luminum wire mesh, which fcilitted the escpe of moisture from the smple through the top electrode during heting. A smple holder, size 14.4x14.4xl.5 cm, ws fbricted out of 3 mm thick Teflon sheet. Teflon ws used since it hs very low dielectric loss fctor of 0.08 (Sreenrynn nd Chttopdhyy, 1986).) nd hence it bsorbs prcticlly no energy from rdio frequency electric field. The top electrode of the dielectric heter ws plced bove the smple holder on Teflon spcers, leving n ir gp of 0.5 cm bove the surfce of the smple. The verge electric field intensity pplied ws 0.5 K Vcm. The voltge, current nd temperture were continuously mesured by using dt logger. The smple ws removed from the heter fter it reched the mximum temperture nd it ws cooled to room temperture before plcing in the smple bg (polyethylene). The rice brn which ws stbilised by steming method t 105 C for 15 min (Julino, 1985) nd rw rice brn served s control. The stbilised smples were stored t 20 C upon nlysis. Determintion of Effect of OMH on Rice Brn Chrcteristics: The stbilised rice brn nd rw rice brn were stored t 4 C. FFA, TBARS, nd lipse ctivity mesurement ws mde fter storge for 0, 7, 14 nd 21 dys. Determintion of Lipse Activity: Smple Extrction: The smple extrction ws undertken by following the method reported by Prbhu et l., (1999) with some modifictions: rice brn (5g) ws deftted by stirring for 30 min with 30 ml of nhexne. The deftted brn ws llowed to irdry for 1 h, so tht prcticlly ll hexne ws removed nd then 30 ml of 0.5mM CCl in 50mM phosphte buffer ph 7.0 ws dded. The deftted brn ws stirred t 10ºC for 30 min. The

3 suspension ws then centrifuged t 3000 rpm for 15 min t 4ºC. The superntnts were collected s the crude lipse extrct nd pplied for lipse ctivity mesurement. Enzyme Assy: Lipse ctivity ws mesured ginst pnitrophenylbutyrte (pnpb) nd pnitrophenyllulte (pnpl) substrtes. The ssy ws ccomplished by following the method described by Htzinikolou et l., (1999). Solution A: 40 mg of pnpb or pnpl were dissolved in 12 ml isopropnol solution. Solution B: 0.4g Triton X00 nd 0.1g gum Arbic were dissolved in 90 ml of 0.1M potssium phosphte buffer (ph 7.0). The substrte solutions were prepred by dropwise ddition of 0.3 ml solution A (pnpb) or 0.2 ml solution A (pnpl) into 3.0 ml solution B under intense mixing (vortex). These emulsions were stble for 1 h t room temperture. For the pnpb substrte system, 0.1 ml of crude lipse extrct ws dded to 3.3 ml of substrte solution nd the rection rte fter incubting t 35 C for 5 min ws mesured by spectrophotometer t 410 nm. For the pnpl substrte system, 0.1 ml of crude lipse extrct ws dded to 3.2 ml of the corresponding substrte solution nd the mixture ws incubted in shking wterbth t 35 C for 20 min. The rection ws terminted by boiling for 5 min following centrifugtion 6000 rpm for 10 min the bsorbnce of the cler superntnt ws mesured t 410 nm. In both cses, smple without enzyme tken the sme tretment ws used s blnk. One unit (U) of enzyme ctivity ws defined s the mount of enzyme required for the libertion of 1 μmol pnp per minute under the ssy conditions. Determintion of Free Ftty Acid (FFA): The FFA vlue ws determined by AOCS officil method (C 540 procedure) (AOCS, 2004). FFA ws clculted s oleic cid nd expressed s percentge of the totl lipids Determintion of Thiobrbituric Acid Rective Substnces (TBARS): The extent of lipid oxidtion ws monitored by the formtion of TBARS. The TBARS vlue ws determined every seven dy of storge by following the method reported by Mielnik et l., (2003) with some modifictions 5g of rice brn smples with 30 ml of 7.5% queous solution thichlorocetic cid (TCA) ws sonicted for 20 min using sonictor (Vibrcell VC 130, Sonics, USA). The slurry ws centrifuged t 3600 rpm for 15 min. The superntnt (5.0 ml) of upper lyer ws collected nd mixed with 5.0 ml of 20 mm queous thiobrbituric cid (TBA) in test tube. The smples were incubted t 100 ºC for 35 min in wterbth nd cooled for 10 min in cold wter. Absorbnce ws mesured t 532 nm by spectrophotometer (Spectronic Genesys 5, USA) ginst blnk contining 5.0 ml distilled wter nd 5.0 ml TBA regent. The results expressed s milligrm mlondildehyde per kilogrms of rice brn were clculted from the stndrd curve of 1, 1, 3, 3tetrethyoxypropne (TEP). Determintion of Bioctive Compounds: Determintion of Totl Phenolic Content (Iqbl et l., 2005): The rection ws initited by mixing 0.2 ml of pproprite diluted rice brn extrct, 0.8 ml of freshly prepred diluted FolinCiocltue regent nd 2 ml of 7.5% sodium crbonte. The volume of the resulting mixture ws djusted to 7 ml by deionized wter nd plced in the drk for 2 h to ensure completion of the rection. The bsorbnce of the resulting bluecolored mixture ws mesured t 765 nm by spectrophotometer (Shimzu, Jpn). Gllic cid ws used s the clibrtion stndrd nd the results were clculted s gllic cid equivlent (mg) per grm (g) of brn. Determintion of átocopherol nd ã Oryznol (Chen nd Bergmn, 2005; Azrin et l., 2008): Rice brn (1g) ws extrcted by following the method reported by (Azrin et l., 2008). Prior to HPLC nlysis, the extrcts were filtered through 0.45 mm syringe filter. An nlysis of ã oryznol nd á tocopherol ws performed, using the reversed phse high performnce liquid chromtogrphy (RPHPLC), ccording to the method reported by Chen nd Bergmn (2005), with some modifictions. The Shimdzu HPLC system (model L6200A), equipped with Photo diode rry detector (Shimdzu, Jpn) nd computer system, ws pplied. Detection ws operted t 292 nd 325 nm, simultneously. The spectr, from 250 to 600 nm, were recorded for ll peks. The extrcted smples were injected through gurdcolumn nd seprted on C 18 column (4.60 x 150mm, 4 μm) (Phenomenex, USA). Grdient elution ws then pplied. Mobile phses A, B, nd C were methnol, wter nd buthnol, respectively. The grdient ws s follows: 02 min 92% A, 4% B nd 4% C: 1225 min liner grdient, from 4% B to 3 % B nd 4% C to 5 % C, with flow rte of 1.5 ml /min nd injection volume of 20 ml. The átocopherol ws detected t 292 nm nd ãoryznol 3644

4 ws detected t 325 nm. Chromtogrms were recorded nd pek res were used to clculte the content of ãoryznol nd tocopherol, ginst the stndrd curve of stndrds. Determintion of Antioxidnt Activity: 6.1 Extrction of Rice Brn: Finely ground brn (5.0g) ws extrcted in 80% methnol (25 ml) by plcing the mixture in sonictor for 10 minutes. The mixture ws filtered nd residue ws then subjected to the sme procedure twice. The residue ws extrcted with 0.15 mol/l hydrochloric cid nd the extrcts were combined nd filtered through filter pper, evported to dryness under reduced pressure t 45ºC by rotry evportor (Buchi, Switzerlnd) nd used for ntioxidnt ctivity nlysis. Antioxidnt Activity: Antioxidnt ctivity ws evluted using three different methods: DPPH Rdicl Scvenging Activity: To evlute the free rdicl scvenging ctivity, the extrct ws llowed to rect with stble free rdicl 1,1diphenyl2picrylhydrzyl (DPPH) by following the method described by Dsgupt nd De (2004). The methnolic extrct (smple extrct, 0.1 ml) ws dded to 3 ml of the 0.004% methnol solution of DPPH. Absorbnce t 517 nm ws mesured fter 30 min nd the percent inhibition ctivity ws clculted s (ACA S) / A C) 100 (A C = Absorbnce without extrct; A S = Absorbnce with extrct). The results were expressed s the concentrtion of test compound to inhibit 50% of free rdicl (IC ). Lipid Peroxidtion Assy: Lipid peroxidtion ws used to mesure the lipid peroxide formed in egg yolk homogente s lipidrich medi ((Dsgupt nd De, 2004). Egg yolk homogente (0.5 ml of 10% v/v) nd 0.1 ml of extrct were dded to test tube nd mde up to 1.0 ml with distilled wter. Subsequently 0.05 ml of 0.07M FeSO4 ws dded to induce lipid peroxidtion nd the mixture ws incubted t room temperture for 30 min. The 1.5 ml of 20% cetic cid (ph 3.5) nd 1.5 ml of 0.8% (w/w) thiobrbituric cid in 1.1% sodium dodecyl sulphte were combined. The resulting mixture ws vortexed nd heted t 95 C for 60 min. After cooling, 5.0 ml of butnol were dded to ech tube nd centrifuged t 3000 rpm for 10 min. The bsorbnce of superntnt ws mesured t 532 nm. Inhibition of lipid peroxidtion ws expressed s IC vlues. Totl Antioxidnt Cpcity Assy: Antioxidnt ctivity ws determined by totl ntioxidnt cpcity (Dsgupt nd De, 2004). 0.3 ml of the extrct ws mixed with 3.0 ml of regent solution (0.6 M sulfuric cid, 28 mm sodium phosphte nd 4 mm mmonium molybdte). The tubes were incubted t 95 C for 90 min. After the mixture hd cooled t room temperture, the bsorbnce ws mesured t 695 nm ginst blnk. The ntioxidnt ctivity is expressed s the number of equivlents of scorbic cid, gllic cid, tocopherols nd butyltes hydroxytoluene. Sttisticl Anlysis: The mens nd stndrd devitions of FFA content, TBARS vlues, lipse ctivity nd ntioxidnt ctivity were reported in triplicte determintions for ech smple. Twowy nlysis of vrince (ANOVA) in completely rndomize design (CRD) using 3 3 fctoril ws used to test significnt differences of moisture content nd the levels of electricl field strengths. Multiple comprison tests were performed by using Duncn multiple rnge tests to determine the significnt difference between tretments. Sttisticl significnce ws declred t P < RESULTS AND DISCUSSION Electricl Resistnce nd Conductivity of Rice Brn: Rice brn smples with different levels of moisture content were subjected to mesure the electricl resistnce nd conductivity. The results showed tht rice brn smples with higher moisture content hd less electricl resistnce wheres the electricl conductivity ws high in higher moisture content smples s indicted in Tble 1. This demonstrtes tht electricl conductivity is function of food components. Ionic components nd moisture mobility increse electricl conductivity, whilst lipids nd fts decrese it (Bengston et l., 2006)

5 Tble 1: The electricl resistnce nd conductivity of rice brn with different moisture contents. Moisture content(%) of rice brn Electricl resistnce(ohm) Electricl conductivity(s m ) 8.10* ± ± ± ± ± ± ± ± *Moisture content of rice brn before dding with wter Vlues re mens of triplicte smples Effect of Applying OMH on Temperture Chnges of Rice Brn: The fresh brn ws plced in the OMH unit nd the temperture chnges were trcked by dt locker plced in the core of the unit. The rice brn smple with the higher moisture content indicted higher of temperture (Fig.1). The rice brn smple, with 40% moisture content nd electric field strength of 225 volt/cm, showed the most rpid chnge in temperture nd the highest temperture of 124.3C. The results were similr to those in study by Yongswtdigul et l,. (1995); Lkkkul, et l., (2004). However, the increse of temperture ws dependent upon severl fctors such s the physicl property of the food, chmber re nd distnce between electrodes (Cstro et l., 2004). The increse in temperture ws relted to tht of the electric field strength, linerly, s shown in Fig. 1 nd Tble 1. The electricl conductivity ws linerly correlted with temperture when the electricl field ws sufficiently high (t lest 60 VCm ) (Bengston et l., 2006) Fig. 1: The temperture chnges of rice brn ffected by different conditions of OMH (MC20/E75 MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) The Effect of Different Stbilistion Conditions on Lipse Activity: Lipse is n endogenous of rice brn nd it is highly effect on rice brn shelf life, becuse there is decomposition of lipids in the brn into free ftty cid by this enzyme. In order to mesure the rice brn lipse ctivity, the substrte specificity, pnitrophenylbutyrte (pnpb) nd pnitrophenyllulte (pnpl) were used. The results re shown in Fig 2 nd 3, respectively. It is cler tht the rw rice brn indicts the highest lipse ctivity. The results were supported by Icier et l., (2006) who used ohmic heting (50 Vcm, 54 s) to obtin the effective peroxidse inctivtion of pe puree. (Cstro et l., 2004) showed tht the electric field hs significntly effect on the lipoxygense (LOX) inctivtion with pproximtely 5 times lower D vlues thn tht of conventionl heting (wter bth). However, they lso found tht some enzyme such s pectinse nd âglctosidse (âgal) were not ffected by the presence of the electric field. The presence of n electric field cn influence biochemicl rections by chnging moleculr spcing nd incresing interchin rections. Moreover, the presence of the electric field my remove the metllic prosthetic groups present in the enzymes LOX nd PPO thus cusing the enhncement of ctivity loss (Cstro et l., 2004). 3646

6 Fig. 2: º Lipse ctivity of stbilised rice brn stored for 21 dys t 4 C using pnpb s substrte(mc20/e75 MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) Fig. 3: º Lipse ctivity of stbilised rice brn stored for 21 dys t 4 C using pnpl s substrte (MC20/E75MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) Effect of OMH on the Chnge of Free Ftty Acid: The influences of ohmic heting on brn stbility s determined by the chnge in FFA re indicted in Fig. 4. The FFA concentrtion of rw rice brn ws significntly higher thn those of stbilised rice brn when storge time incresed. The FFA content of rw rice brn ws more thn 10% fter storge for 21 dys wheres the FFA content of rice brn, stbilised t different conditions, slightly incresed to pproximtely 3%. Therefore, the stbilised rice brn ws still suitble for humn consumption, ccording to (To et l., 1993), but the brn with over 5% FFA is considered unsuitble for humn consumption. The rte of FFA formtion 3647

7 Fig. 4: Effect of ohmic heting on free ftty cid (FFA) content (%) of rice brn during storge t 4 C (MC20/E75MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) ws over 5 fold, in unstbilised rice brn, whilst tht of stbilised rice brn ws pproximtely 2 fold within 21 dys under study condition (4ºC). The content of FFA ws positively relted to the ctivity of lipse, s indicted in Fig. 2 nd 3. OHM could retrd the forming of FFA compred with rw rice brn nd stemed rice brn. However, there ws no significnt difference between the rice brn smples treted with different electric field strengths nd moisture contents. These results re supported by (Lkkkul et l., 2004), who showed tht the FFA content in untreted brn stored t 4 C for 6 weeks incresed from 3.96% to 18.03% whilst the FFA of ohmic heted brn smples (21% moisture content), which were stored identiclly s the untreted brn incresed from 3.25% to 5.54%. Sreenrynn nd Chttopdhyy (1986) obtined n increse in FFA from 4.2% to 6.2% during six week cold storge period by using dielectric heting (0.5 kv/cm, MHz) to tret rice brn with moisture content of 21%. Fig. 5: The effect of ohmic heting on TBARS vlues of rice brn stbilised (mlondildehyde (mg)kg of rice brn) (MC20/E75MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) 3648

8 Effect of OMH on Thiobrbituric Acid Rective Substnces (TBARS): The TBARS vlues for ohmiclly heted nd steming stbilised rice brn smples s compred to the rw rice brn re displyed in Fig.5. In the rw rice brn, the TBARS vlues showed the most rpid incresed nd reched their mximum t 14 dys of storge. This could be due to (in the rw rice brn) lipse still being ctive nd hydrolyses lipids in rice brn freeing ftty cids nd thus being redy for oxidtion. The vlues for TBARS, of rw rice brn, were incresed up to 2 weeks of storge nd then decresed. This my due to some TBARS is developed by the lipid oxidtion to other oxidtion products (Ryu nd Cheigh, 1980). The lowest TBARS vlues were observed in the rice brn with moisture content of 30% nd being ohmiclly heted by n electric field strength t 150 Vcm with the level of 1.98 mg mlondildehyde (MDA)kg dry solid fter 21dys. This my be due to the fct tht, in this condition, the het could penetrte nd effectively destroyed lipse nd it combined with the prevention effect of rice brn to retrd the development of oxidtion product in rice brn. The stbilised rice brn cn reduce lipid oxidtion becuse it contins number of ntioxidnts nd this result could lso indicte by the ntioxidnt results nd bioctive compounds of rice brn s shown in next section of the pper. The ntioxidnt ctivity of compounds is often described by its bility to dely the onset of utoxidtion by scvenging rective oxygen species, or the bility to ct s chin breking ntioxidnts to inhibit the propgtion phse of lipid utoxidtion (Yun et l., 2005). Effect of OMH on Totl Phenolic Compounds, átocopherol nd ãoryznol Content: Results showed the totl phenolic content rnging from 1.79 to 3.28 mg gllic cid equivlent g of rice brn. The tocopherol nd goryznol content rnged from μg/g nd μgg, respectively (Tble 2). Stbilised rice brn with moisture content of 3040 % nd EFS of Vcm yielded significntly higher mount of phenolic compounds, tocopherol nd goryznol thn those of rice brn t 20% moisture content nd rice brn stbilised by heting. The totl phenolic, tocopherol, nd goryznol content of the rice brn stbilised by OMH were comprble to those of rice brn reported by(chotimrkorn et l., 2008). Tble 2: The effect of ohmic heting on bioctive compounds of rice brn (mgg ) Stbilistion condition Bioctive compound Totl phenolic átocopherol ãoryznol e MC20/E ± 0.02 c c ± ± 0.19 d MC20/E ± 0.05 b b ± ± 0.20 c b b MC20/E ± ± ± 0.22 b MC30/E ± 0.03 b ± ± 0.12 MC30/E ± ± ± 0.07 MC30/E ± ± ± 0.06 MC40/E ± ± ± 0.05 MC40/E ± ± ± 0.04 MC40/E ± ± ± 0.03 c Steming 2.46 ± 0.13 c b ± ± 0.16 d d d Rw brn 1.79 ± ± ± 0.27 Vlues re mens of triplicte smples (on dry weight bsis) d Mens within column with different numbers re different (P 0.05) MC stnds for moisture content nd E stnd for electricl field strength (Vcm ) Effect of OMH on Antioxidnt Activity: DPPH Rdicl Scvenging Activity: The ntioxidnt ctivity by DPPH rdicl scvenging method, s expressed by the concentrtion of the mount extrct to inhibit 50% of stble free rdicl (IC 50), showed tht the reference stndrds, scorbic cid nd BHT, were the strongest ntioxidnt (Fig. 6). The rice brn with moisture content of 20% (225 EFS), 30% nd 40%, in combintion with the electricl field strengths of 75,150, 225 Vcm, indicted the strongest ntioxidnt ctivity This suggests there exists n optiml moisture content for ohmic heting nd the provision of higher moisture content does not significntly impct on the ctivity of ntioxidnt. 2. Lipid Peroxidtion Assy: Lipid peroxidtion of egg yolk lipids undergo rpid nonenzymtic peroxidtion when incubted in the presence of ferrous sulphte (Dsgupt nd De. 2004).The effect of ohmic heting on lipid peroxidtion system re shown in Fig. 7. The results indicted tht the rice brn with higher moisture content (3040%) with higher EFS levels ppered to increse the oxidtive stbility expressed by the higher ctivity thn rice brn stbilised by steming nd rw rice brn. 3649

9 Fig. 6: DPPH rdicl scvenging ctivity (IC 50) of rice brn (mg/ml)ic 50 = concentrtion of the extrct to inhibit 50% of DPPH rdicl (MC20/E75MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm 1; Vlues re mens of triplicte smples) Fig. 7: Lipid peroxil rdicl scvenging ctivity (IC50) of rice brn (mg/ml) IC50 = concentrtion of the extrct to inhibit 50% of free rdicl (MC20/E75MC40/E225 = Rice brn with moisture content of 20, 30 nd 40% combined with electricl field strength of 75, 150 nd 225 Vcm; Vlues re mens of triplicte smples) 3. The Totl Antioxidnt Cpcity Assy: The totl ntioxidnt cpcity of brn extrct is expressed s the number of equivlent ntioxidnt stndrds (scorbic cid, gllic cid, átocopherol nd BHT). The totl ntioxidnt cpcity of stbilised rice brn smples s compred with tht of rw rice brn is displyed in Tble 3. The rice brn with moisture content of 3040 % nd EFS t Vcm indicted greter ntioxidnt cpcity in ll ntioxidnt stndrd equivlents. These results were relted to the concentrtion of phenolic compounds, tocopherol nd goryznol, which re ntioxidnt. 3650

10 Tble 3: Totl ntioxidnt cpcity of stbilised rice brn (ohmic heting, steming) nd rw rice brn (mg ntioxidnt stndrd equivlent /g rice brn). Stbilistion condition Totl ntioxidnt ctivity Ascorbic cid Gllic cid Tocopherol BHT f f f MC20/E ± 0.05 f 1.53 ± ± ± 0.06 e e e MC20/E ± 0.15 e 1.94 ± ± ± 0.18 d d d MC20/E ± 0.11 c 2.18 ± ± ± 0.13 c c c MC30/E ± 0.04 b 2.40 ± ± ± 0.05 MC30/E ± ± ± ± 0.09 b b b MC30/E ± ± ± ± 0.06 bc c c MC40/E ± 0.11 cd 2.11 ± ± ± 0.13 bc c c MC40/E ± 0.05 cd 2.09 ± ± ± 0.05 bc c c MC40/E ± 0.04 d 2.06 ± ± ± 0.04 e e e Steming 5.65 ± 0.07 d 2.04 ± ± ± 0.08 g f g g Rw brn 4.63 ± ± ± ± 0.17 Vlues re mens of triplicte smples (on dry weight bsis) g Mens within column with different numbers re different (P 0.05) MC stnds for moisture content nd E stnd for electricl field strength (Vcm ) Conclusions: The results of this study suggested tht ohmiclly heting rice brn with the ddition of moisture is n effective stbilistion method. According to the studied conditions nd the results, the optimum conditions to obtin stbilised rice brn re through n djustment of the moisture content of the rice brn from 3040% nd lso the ppliction of n electricl field strength between Vcm. The benefits were seen by n increse in the moisture content of the rice brn nd EFS of lternting current to improve the stbility nd preserve the bioctive compounds. However, in this study, the ohmic heting unit ws the lb scle. In order to develop this to lrger scle, n intensive study of mny spects is needed. ACKNOWLEDGMENTS Finncil support provided by Mhsrkhm University, Thilnd nd the Thilnd Reserch FundMster Reserch Grnts (TRFMAG, 2008) is grtefully cknowledged. We cknowledge dvice nd consulttion on the configurtion of the ohmic heting unit by Tnongsk Moontree, Electricl Engineer nd Lecturer in the Fculty of Technology, Mhsrkhm University. REFERENCES AOAC, Officil Methods of Anlysis. Assocition of Officil Anlyticl Chemist Interntionl.17 th Ed., AOAC Interntionl, Githersburg, MD, USA. AOCS, Officil methods nd recommended prctices of the Americn Oil Chemists Society, Americn Oil Chemists Society, Chmpign. Ausmn, L.M., N. Rong nd R.J. Nicolosi, Hypocholesterolemic effect of physiclly refined rice brn oil: studies of cholesterol metbolism nd erly therosclerosis in hypercholesterolemic hmsters. Journl of Nutritionl Biochemistry, 16(9): Azrin, A., I. Mznh nd A.H. Azizh, Extrction nd determintion of oryznol in. ASEAN Food Journl., 15(1): Bengston, R., E. Birdsll, S. Feilden, S. Bhttiprolu, S. Bhle, M. lim, Ohmic heting nd induction heting. In Hui, Y. H. (Ed.). Hndbook of Food Science, Technolgy, nd Engineering (Vol. 3). FL: CRC Press, pp: 120. Cstro, I., B. Mcedo, J.A.Teixeir nd A.A. Vicente, The effect of electric field on importnt foodprocessing enzymes: Comprison of inctivtion kinetics under conventionl nd ohmic heting. Journl of Food Science, 69(9): C696C701. Chen, M.H. nd C.J. Bergmn, A rpid procedure for nlysing rice brn tocopherol, tocotrienol nd gmmoryznol contents. Journl of Food Composition nd Anlysis., 18(23): Chotimrkorn, C., S. Benjkul nd N. Silli, Antioxidnt components nd properties of five longgrined rice brn extrcts from commercil vilble cultivrs in Thilnd. Food Chemistry., 111(3): Dsgupt, N. nd B. De, Antioxidnt ctivity of Piper betle L. lef extrct in vitro. J. Food chemistry, 88(2): FDA, Kinetics of microbil inctivtion for lterntive food processing technologies: ohmic nd inductive heting. Avilble from < Retrieved 22 June,

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