Journal of Integrative Agriculture 2017, 16(0): Available online at ScienceDirect

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1 Journl of Integrtive Agriulture 2017, 16(0): Aville online t SieneDiret RESEARCH ARTICLE The effet of dehydrogense enzymes tivity in glyolysis on the olour stility of mutton during postmortem XIN Jin-zeng 1, 2, LI Zheng 2, LI Xin 2, LI Meng 2, WANG Ying 2, YANG Fu-min 1*, ZHANG De-qun 2* 1 College of Food Siene nd Engineering, Gnsu Agriulturl University, Lnzhou , P.R.Chin 2 Institute of Food Siene nd Tehnology, Chinese Ademy of Agriulturl Sienes, Beijing , P.R.Chin Astrt This study investigted the influene of tivities of glyerldehyde-3-phosphte dehydrogense (GAPDH) nd ltte dehydrogense-b (LDH-B) on the olour stility of mutton. From 60 sheep, 15 M. longissimus dorsi (LD) musles were seleted on the sis of olour stility (R630/580 nd * vlue) during the storge nd lssified into three groups (5 for one group) s high olour stility (HCS), intermedite olour stility (ICS) nd low olour stility (LCS). The tivities of GAPDH nd LDH-B, instrumentl olour ttriutes, niotinmide denine dinuleuotide (NADH) ontent nd ltte ontent were mesured. The smples in HCS hd higher tivities of GAPDH nd LDH-B thn the smple in the LCS, nd the smples in HCS lso possessed higher NADH ontent nd lower ltte ontent. The higher tivity of dehydrogense enzyme n result in more NADH nd olour stility. The results suggested tht the tivity of GAPDH nd LDH-B my lso ply role in mintining the olour stility. Keywords: mutton, met olour, GAPDH, LDH-B, NADH 1. Introdution Met olour is one of the most importnt qulity ttriutes whih influene onsumer purhsing deisions, euse onsumers often onsider the herry-red olour of red met s n inditor of wholesomeness t the point of sle (Joseph et l. 2012). Disolourtion of fresh met will led to the rejetion y onsumer nd it ws estimted tht the United Sttes met industry inurs more thn one illion dollrs in lost opportunity due to the disolourtion every Reeived 27 Deemer, 2016 Aepted 9 Mrh, 2017 Correspondene YANG Fu-min, E-mil: yfumin@163.om; ZHANG De-qun, E-mil: dequn_zhng0118@126.om 2017, CAAS. All rights reserved. Pulished y Elsevier Ltd. doi: /S (16) yer (Joseph et l. 2012; Sumn nd Joseph 2013). The metmyogloin reduing system ws onsidered s the most importnt ftor tht mintins the stility of met olour during postmortem (Ledwrd et l. 1985; Kim et l. 2009). Niotinmide denine dinuleuotide (NADH) is the ultimte reduing equivlent for the metmyogloin redution of enzyme nd non-enzyme, whih hve een shown to e vitl for delying the disolourtion in the postmortem musle (Yun et l. 2006; Rmnthn et l. 2011). Glyerldehyde-3-phosphte dehydrogense (GAP- DH) nd ltte dehydrogense B (LDH-B) re importnt enzymes in musle glyolysis whih plys key role in forming NADH nd mintining the equilirium of NADH pool in ytoplsm (Brron et l. 1998; Li et l. 2015). Kim et l (2006, 2009) reported tht ddition of ltte ould enhne the olour stility through the replenishment of NADH tlyzed y LDH-B tivity. In ddition, the olour stility tends to e higher in ovine Longissimus lumorum

2 *** et l. Journl of Integrtive Agriulture 2017, 16(0): musles ompred with Semimemrnosus, nd Psos mjor, due to higher LDH-B (Kim et l. 2006, 2009, ). Bsed on the ove report, for olour stility of fresh musle, LDH-B tivity nd its sustrtes re importnt ftors euse of the regenertion of NADH. In ddition, GAPDH is involved in nother formtion pthwy of NADH. In the previously study, glyerldehyde-3-phosphte, the sustrtes of GAPDH, ws proved redue the metmyogloin in eef ground (Sleh nd Wtts 1968). However, the studies out the effets of GAPDH tivity on met olour re limited. And whether the tivity of GAPDH in musles with different olor stility is the sme ws unler. We hypothesized tht the tivity of GAPDH in met with different olour stility is inonsistent, so the ojetive of the urrent study ws to determine the GAPDH tivity nd other iohemil hrteristis of different olour stility of M. Longissimus dorsis (LDs). And lso to evlute the potentil proility of GAPDH regulting the met olour stility. 2. Mterils nd methods 2.1. Rw mterils nd preprtion Sixty mle sheep (Bynnur mutton sheep, 8 months of ge) with n verge rss weight of (24.74±2.05) kg from ommeril plnt (Inner Mongoli Grsslnd Hongo Food Co. Ltd., Chin) were slughtered following the industril prtie. At 30 min postmortem, oth loins ((LDs) were olleted (yielding 120 loins in totl) from 60 lm rsses. Eh musle ws divided into 4 steks nd pled on the styrofom trys, nd wrpped with oxygen-permele polyvinylhloride (PVC, type, Nntong South Asi Plsti Film Co., Ltd, Nntong, Jingsu, Chin; oxygen trnsmission rte=25 ml m 2 24 h Mp 1 ) film. Then the smple trys were stored in n industril refrigertor t (4±2) C nd were tken out for nlysis fter 2 h, 6 h, 1, 2, 4, 6 nd 8 d storge, respetively. One stek ws used for determintion of instrumentl olour nd ph during the storge. Other seven steks were frozen in liquid nitrogen t speifi time, nd then stored t 80 C until used for iohemil nlysis Instrumentl olour The olour of the smples over wrpped with PVC ws mesured through the PVC film fter storge t 4 C for 2 h, 6 h, 1, 2, 4, 6, nd 8 d. Colour ws mesured using Minolt CM-600d spetrophotometer (Koni Minolt SensingIn., Osk, Jpn) with speulr refletne exluded, 8 mm dimeter mesuring perture, illuminnt D65, 10 stndrd oserver nd Commission Interntionle de L Elirge (CIE ) L *, *, * olour sle. The verge vlue of four mesurements on the met surfe ws used. The Minolt instrument reorded refletne vlues in the rnge of 360 to 740 nm t 10 nm intervls. Refletne vlues tht were not diretly mesured y the olour instrument t speifi wve lengths (474, 525 nd 572 nm) were lulted y integrtions. The perentge of myogloins ws determined s desried y Hunt nd King (2012) using the formuls: A 572 A 700 Metmyogloin%=(1.395 ) 100 A 525 A 700 A Deoxymyogloin%=( A 700 ) 100 A 525 A 700 Oxymyogloin%=100% (MM%+DM%) Where, reflex ttenune (A)=log(1/R), A 473, A 525, A 572 nd A 700 is the reflex ttenune t 473, 525 nd 700 nm, respetively. R is refletne. In ddition, the rtio of refletne t 630 nd 580 nm (R630/580) ws lulted s n indiret estimte of surfe olour stility, greter rtio indites lesser mount of metmyogloin rown disolourtion nd thus higher olour stility. Instrumentl olor dt on steks from the sixty rsses were rnked sed on the R630/580 nd * vlue from dy 4 to 8. From this rnking, the five (n=5) high olour-stle (HCS, R630/580, (2.74±0.13)), five (n=5) intermedite olour-stle (ICS, R630/580, (2.21±0.22)) nd five (n=5) low olor-stle (LCS, R630/580, (1.98±0.18)) steks were identified to exmine the moleulr sis of niml-to-niml vrition in olor stility ph vlue The ph vlues were mesured y inserting the glss lomel proe nd temperture sensor of ph meter (Testo205 ph meter, Lenzkirh, Germny) diretly into the rw smple. The devie ws lirted with three uffers (ph 4, 7 nd 10). All mesurements were nlyzed in triplites from whih n verge ws lulted GAPDH tivity The enzyme ws extrted from smples s desried y the method (Bii et l. 2007) with slight modifitions. All steps were rried out on ie during the extrtion. The musle smple (pproximtely 2 g, fresh weight) ws ground nd homogenized using n IKA Ultr-Turrx homogenizer T10 si S25 (IKA, Germny) in 25 mmol Tris-HCl uffer (ph 7.5), ontining 5 mmol EDTA, 10 mmol 2-merptoethnol t rtio of 4 ml g 1 fresh tissue. The superntnt (solule protein frtion) otined fter entrifugtion t g for 45 min ws onsidered s the extrted enzyme. The GAPDH tivity of smples ws mesured with Si-

3 4 *** et l. Journl of Integrtive Agriulture 2017, 16(0): encell olourimetri GAPDH ssy kit (Ctlog # 8148). It ws determined spetrophotometrilly y monitoring NADH genertion t 340 nm LDH-B tivity The tivity of LDH in the smple ws determined sed on the method desried y Kim (2009, ). The musle smple (2 g) ws homogenized in 8 ml of sodium uffer (0.01 mol L 1, ph7.5) for 30 s nd held on ie. The homogente ws filtered through Whtmn # 42 filter pper fter entrifuged t g for 30 min t 4 C. The filtered liquid ws used s enzyme extrt to e mesured. The retion system for the determintion of LDH tivity ontined filtered superntnt (0.1 ml), 172 mmol L 1 NAD (0.1 ml) nd 2.4 ml of Tris (112 mmol L 1 ), KCl (170 mmol L 1 ), L-ltte (56 mmol L 1 ) (ph 9.3). The inresed sorne t 339 nm ws mesured (UV-2401 PC Spetrophotometer, Shimdzu, Kyoto, Jpn) from 30 to 120 s nd used for lulting LDH tivity. The NADH ontent in fresh musle (nmol g 1 ) ws lulted. Units for the determintion of enzyme tivity were expressed s 1 mol min 1 g 1 smple NADH onentrtion NADH ws extrted sed on the methodology desried y Klingenerg (1974) with slight modifitions. Two grms of ovine musle smple ws mixed with 16 ml of 0.5 mol L 1 ooled loholi KOH solution. The mixture ws vortexed for 30 s, then gitted in 90 C wter th for 5 min, nd ooled rpidly to 0 C in n ie th for 5 min. The ph vlue of the musle mixture ws djusted to 7.8 y dding 12 ml triethnolmine-hcl-phosphte mixture (0.5 mol L 1 triethnolmine hydrohloride, 0.4 mol L 1 KH 2 PO 4, 0.1 mol L 1 K 2 HPO 4 ). After holding t room temperture for 10 min to floulte the dentured protein, the musle mixture ws entrifuged t g for 10 min t 4 C (J2-21, Bekmn Instruments, In., Plo Alto, CA) to get the ler superntnt. NADH onentrtion ws mesured sed on the methodology desried y MCormik n Wright (1971). Musle extrt superntnt (0.1 ml) ws mixed with 2.5 ml of 17.5 μg ml 1 dihlorophenolindophenol (DCPIP), 0.5 ml of 0.1 mol L 1 ph 7.4 sodium phosphte uffer, 0.05 ml of 1 mg ml 1 phenzine methosulfte (PMS), 0.1 ml ethnol, nd 0.05 ml lohol dehydrogense (E , Sigm, USA, ontining 0.3 mg protein). The hnge of the sorne t 600 nm ws mesured (UV-2401 PC Spetrophotometer, Shimdzu, Kyoto, Jpn) for 20 min to determine the NADH onentrtion. The NADH ontent in fresh musle tissue (nmol g 1 ) ws lulted with respet to the stndrd urve Ltte nd pyruvte onentrtion The ltte nd pyruvte ssy kit were purhsed from Nnjing Jinheng Bioengineering Institute (Nnjing, Jingsu, Chin). For ltte ssys, the musle smple ws weighed preisely nd dded physiologil sline with the rtio 1 g to 9 ml. The mixture ws homogenized in n ie wter th nd then entrifuged t g for 10 min, nd the superntnt ws olleted for further mesurement. Then the ltte onentrtion ws mesured with ltte ssy kit (A020) t the sorne of 530 nm nd lulted with respet to the ltte stndrd smple. The extrt of pyruvte in the smple ws rried out y ltte method. The pyruvte onentrtion ws mesured with pyruvte ssy kit (A081) t the sorne of 505 nm nd lulted using the pyruvte stndrd smple Sttistil nlysis Instrument olour dt, ph vlues, ltte dt, GAPDH tivity dt, LDH-B tivity dt were nlyzed using the SPSS 22.0 softwre. The results were expressed s mens±stndrd error. Mens were ompred with ANOVA (one-wy nlysis of vrine) to determine the levels of sttistil signifine t 0.05 level. 3. Results 3.1. * vlues nd R630/580 The * vlues (redness) nd R630/580 of LD steks in different olour stility groups were shown in Fig. 1. The * vlue of smple in different groups inresed nd then deresed. Steks in the HCS group hd higher * vlue thn the smples in the other two groups from dy 2 to 8 during storge, nd the differene ws signifint from dy 4 to 8. The vrition of R630/580 vlue ws onsistent with * vlue, whih lso inresed t first dy nd then deresed. The R630/580 vlue of smples in HCS group ws signifintly higher thn the smples in the other two groups from dy 2 to Proportions of myogloin redox forms Myogloin redox forms (%) were presented in Tle 1. The reltive metmyogloin perentge of met smple in three different olour stility groups deresed within 1 d nd then inresed during the remining storge time. The umultion of metmyogloin in the smple of HCS group ws lower thn tht in the LCS group. The reltive deomyogloin perentge of steks in three groups deresed

4 *** et l. Journl of Integrtive Agriulture 2017, 16(0): ontinuously, nd there ws no signifint differene mong the smples in the three groups. Oxymyogloin (%) of steks in different groups inresed during the originl 1 or 2 d fter postmortem nd then deresed. The smple in HCS hd higher perentge of oxymyogloin thn the smple in LCS nd ICS on the lte storge time. The vrition of oxymyogloin ws onsistent with the * -vlue A * vlue B R630/ Storge time Fig. 1 Surfe redness ( * vlue) (A) nd rtio of refletne t 630 to 580 nm (R630/580) (B) of ovine musle during storge for 8 dys t 4 C. HCS, high olour stility group. ICS, intermedite olour stility group. LCS, low olour stility group. Mens etween groups within the sme storge dy with different letters ( ) re different (P<0.05). Error rs represent the stndrd error of the men. Storge time nd R630/ L * vlue, * vlue, hrom nd hue ngle The L * vlue (lightness), * vlue (yellowness), hrom nd hue ngle were presented in Tle 2. L * vlue ws inresed t the first storge time nd then deresed, however, no signifint differene etween the three groups in the sme storge time. The * vlue of smple in the HCS ws higher thn smples in the LCS group during the whole storge time exept dy 1, nd the differene ws signifint from dy 4 to dy 8. The hrom vlue of stek in HCS ws signifint higher thn the smple in the other two groups, exept dy 1. For the hue ngle, the smple in the HCS group ws higher thn the other two groups ph nd ltte There ws no signifint differene of ph 45 mong the three groups s showed in Tle 3. However, the ph 24 of LD steks in HCS ws signifintly higher (P<0.05) thn the other two groups on the first dy fter postmortem. The ltte ontent of steks in different groups ws presented in Fig. 2. The ltte ontent in ll smples inresed within 1 dy nd then were stle during the storge time. The ltte onentrtion in the smple of HCS ws lower thn the smple in ICS nd LCS during storge time exept dy NADH, LDH-B, nd GAPDH The onentrtion of NADH in steks of different groups re shown in Fig. 3. The NADH onentrtion in steks of HCS ws higher thn smple in LCS on the time 2 h, 6 h, 1, 2, nd 6 d. The differene of NADH in steks etween ICS nd LCS ws not ompletely signifint. The tivity of LDH-B in smples of different groups ws Tle 1 M forms (%) of ovine musle t 2 h, 6 h, 1, 2, 4, 6 nd 8 d postmortem (4 C) Time of storge Pigment Ctegory 1) %MetM HCS 25.71±0.35 y 24.36±0.23 y 18.30±0.78 z 23.27±0.58 y 25.59±1.33 y 29.03±1.20 x 32.05±1.17 w ICS 27.09±0.45 y 25.97±1.18 xy 18.93±0.24 z 23.64±0.57 y 27.33±0.32 x 31.40±0.71 w 37.21±1.82 u LCS 28.84±1.11 xy 29.38±1.60 xy 20.19±0.54 z 27.06±0.81 y 27.13±0.12 y 31.30±0.61 x 37.43±1.93 w %DeoM HCS 53.72±4.79 x 51.22±3.89 x 19.14±1.95 y 12.17±0.47 y 11.21±0.59 z 11.76±0.56 z 11.37±0.39 z ICS 53.57±2.54 x 53.91±1.73 x 18.96±1.18 y 13.81±0.81 y 12.97±0.52 z 12.04±0.63 z 11.28±0.90 z LCS 49.19±2.06 x 56.03±1.83 w 19.31±0.99 y 13.11±0.74 z 12.98±0.62 z 12.03±0.43 z 11.28±0.89 z %OxyM HCS 20.57±4.87 z 24.42±4.04 z 62.57±1.26 xy 64.56±0.61 x 63.20±1.04 xy 59.21±0.86 xy 56.58±0.91 y ICS 19.35±2.51 z 20.12±1.36 z 62.11±1.29 w 62.55±0.96 w 59.70±0.82 wx 56.56±1.06 x 51.52±1.65 y LCS 21.97±1.85 y 14.59±2.51 z 60.51±1.19 w 59.84±1.05 w 59.69±0.83 w 57.58±1.08 w 51.30±1.72 x 1) HCS, high olour stility group; ICS, intermedite olour stility group; LCS, low olour stility group. Results re expressed s the men±stndrd error. Mens within the sme group ross storge time with different letters (u z) re different (P<0.05). Mens etween groups within the sme storge time with different letters ( ) re different (P<0.05).

5 6 *** et l. Journl of Integrtive Agriulture 2017, 16(0): illustrted in Fig. 4. The tivity of LDH-B in smple of HCS deresed during the storge time. The smple in the HCS hd higher tivity thn the smple in LCS, nd signifint differene were on the 6 h, 1 d, 4 d nd 6 d. The tivity of LDH-B etween the HCS nd LCS ws not signifint during the most storge time. The tivity of GAPDH in steks of different groups ws shown in Fig. 5. The tivity of GAPDH in the smple of HCS nd ICS ws higher thn the smple in LCS, nd the differene ws found signifint on the 2 h, 6 h, 1 d, 2 d, 4 d fter the postmortem. 4. Disussion 4.1. * vlue nd R630/580 The * vlue is one of the most importnt inditors of the olour stility of fresh musle, nd the redness of fresh musle is minly influened y the proportion of oxymyogloin (Mnini nd Hunt 2005). The * vlue nd proportion of oxymyogloin in steks of HCS were higher ompred with tht in the other two groups from dy 2 to 8 in the present study. In prtil greement with our result, King et l. (2011) reported tht steks with stle olour possessed higher * vlue. During storge, steks in three groups demonstrted derese on * vlue. In greement with our result, previous study Joseph et l. (2012) lso doumented derese in redness of eef musle. The result of R630/580 suggested tht the steks in HCS group hd higher olour stility. Corroorting our result on * vlue, the R630/580 vlue of smple in the three groups deresed during storge. Joseph et l. (2012) lso reported tht 630/580 vlue of two musles (Longissimus lumorum (LL) nd PM) deresed throughout the retil disply L * vlue nd * vlue L * vlue is nother key inditor of the olour stility of fresh musle, in the urrent reserh steks in three groups exhiited no differene on L * vlue during storge exept dy 8. In support, previous study (King et l. 2011; Cnto et l. 2015) demonstrted tht eef musle whih hs different olour stility shows no differene on lightness (L * vlue) during storge. Musles from different soure tht Tle 2 Instrumentl olour of ovine musle t 2 h, 6 h, 1-, 2-, 4-, 6-, nd 8-dy postmortem (4 C). Time of storge Prmeters 1) Ctegory 2) L * HCS 34.08±0.35 z 34.43±0.82 y 40.22±1.38 x 42.27±1.01 x 40.89±0.61 x 39.60±0.33 x 39.30±0.57 x ICS 33.96±0.26 z 36.87±1.60 y 41.29±0.40 x 42.60±0.84 x 42.07±0.80 x 41.45±0.48 x 41.96±0.71 x LCS 33.28±0.86 y 33.21±0.41 y 38.75±0.77 x 40.88±0.52 x 39.73±0.71 x 39.50±0.82 x 39.41±0.77 x * HCS 6.27±0.23 z 6.10±0.26 z 10.56±0.60 y 12.47±0.65 x 13.12±0.60 x 12.29±0.33 x 12.01±0.22 x ICS 5.40±0.36 z 5.31±0.47 z 10.63±0.32 y 11.77±0.49 xy 12.05±0.51 x 11.86±0.44 xy 11.74±0.37 xy LCS 5.23±0.31 y 5.47±0.22 y 10.57±0.49 x 10.46±0.62 x 10.67±0.24 x 9.59±0.39 x 10.36±0.33 x Chrom HCS 10.73±0.43 z 10.32±0.39 z 15.83±0.96 y 18.43±0.66 wx 19.34±0.59 w 18.03±0.43 wx 17.46±0.25 xy ICS 9.53±0.44 y 9.07±0.73 y 15.59±0.53 x 17.33±0.68 x 17.49±0.72 x 16.83±0.62 x 16.11±0.54 x LCS 9.53±0.41 z 9.67±0.24 z 15.38±0.57 x 15.07±0.75 x 15.11±0.37 x 13.11±0.40 y 13.56±0.34 y Hue HCS 35.77±0.60 y 36.28±1.03 y 41.91±0.62 x 42.53±1.31 x 42.68±1.03 x 42.96±0.55 x 43.43±0.44 x ICS 34.44±1.07 z 36.37±2.31 z 43.05±0.69 y 42.84±1.00 y 43.54±0.54 xy 44.83±0.51 xy 46.88±0.98 x LCS 33.31±1.33 z 34.39±1.23 z 43.41±1.09 y 43.93±1.28 xy 44.97±0.71 xy 46.98±0.82 wx 49.81±0.80 w 1) Chrom=( *2 + *2 ) 0.5, Hue ngle=atan ( * / * ) (180/π). 2) HCS, high olour stility group; ICS, intermedite olour stility group; LCS, low olour stility group. Results re expressed s the men±stndrd error. Mens within the sme group ross storge time with different letters (w z) re different (P<0.05). Mens etween groups within the sme storge time with different letters ( ) re different (P<0.05). Tle 3 ph of musle smples in three groups t 45 min nd 24 h postmortem. Ctegory 1) ph 45 ph 24 HCS 6.96±0.02 x 5.94±0.02 y ICS 6.93±0.04 x 5.87±0.02 y LCS 6.97±0.04 x 5.83±0.01 y 1) HCS, high olour stility group; ICS, intermedite olour stility group; LCS, low olour stility group. Results re expressed s the men±stndrd error. Mens within the sme group ross storge time with different letters (x nd y) re different (P<0.05). Mens etween groups within the sme storge time with different letters ( nd ) re different (P<0.05). hve inonsistent olour stility lso hd no signifint differene on L * vlue (Joseph et l. 2012; Cnto et l. 2015). On ontrry, Neethling et l. (2016) reported tht different soure musles from lesok hve inonsistent L * vlue during storge. The steks in the present study signifintly inresed during the storge time. In greement with our result on lightness, Luino et l. (2009) reported tht semimemrnosus musle (SM) from lm possessed n inrese throughout the storge. Likewise, Bo et l. (2016) lso

6 *** et l. Journl of Integrtive Agriulture 2017, 16(0): reported tht L * vlue of ptties mde from mined eef signifintly inresed during the storge time. Go et l. (2013) demonstrted tht LD from lm showed derese on L * vlue during the simulted retil disply, nd this ws not similr with our result. But the L * vlue of eef steks in some previous reported studies (MKenn et l. 2005; King et l. 2010; King et l. 2011; Cnto et l. 2015) were different with our report nd this my due to niml differene. Steks in HCS group hd higher * vlue (yellowness) thn steks in LCS group from dy 4 to 8. In support, King et l. (2011) demonstrted tht longissimus thoris steks with stle olour stility hd higher yellowness throughout storge exept dy 0 nd 1. Cnto et l. (2016) lso reported tht LL from Nellore hd higher * thn PM. Furthermore, MKenn et l. (2005) demonstrted tht musle tht possessed higher olour stility lso mintined greter * vlue thn musle whih hd low or very low olour stility. The result of present reserh exhiited tht storge time inresed the * vlue of steks in three groups. In support Ltte ontent (μmol g 1 ) z z z y y y x x x x x x x x x x x x x x x Storge time Fig. 2 Ltte onentrtions determined in met smple during storge. HCS, high olour stility group. ICS, intermedite olour stility group. LCS, low olour stility group. Mens mong groups t the sme time points with different letters ( ) re different (P<0.05). Error rs represent the stndrd error of the men. Mens within the sme group ross storge dys with different letters (x nd y) re different (P<0.05). NADH ontent (nmol g 1 ) u u u u u u v v v w w w x x x y y y z z Storge time z Fig. 3 The NADH onentrtion in M. longissimus dorsi steks in different olour stility groups during the storge time. HCS, high olour stility group. ICS, intermedite olour stility group. LCS, low olour stility group. Mens etween groups within the sme storge time with different letters (-) re different (P<0.05). Mens within the sme group ross storge dys with different letters (u-z) re different (P<0.05).

7 8 *** et l. Journl of Integrtive Agriulture 2017, 16(0): LDH-B Ativity (μmol min 1 g 1 ) Storge time Fig. 4 LDH-B tivity of M. longissimus dorsi steks in different olour stility groups during the storge time. HCS, high olour stility group. ICS, intermedite olour stility group. LCS, low olour stility group. Mens etween groups within the sme storge time with different letters ( nd ) re different (P<0.05). GAPDH tivity (U g 1 ) Storge time Fig. 5 GAPDH tivity of M. longissimus dorsi steks in different olour stility groups during the storge time. HCS, high olour stility group. ICS, intermedite olour stility group. LCS, low olour stility group. Mens etween groups within the sme storge time with different letters ( nd ) re different (P<0.05). to our results, previous reserh (Luino et l. 2009) hd similr result. In ontrst, Go et l. (2013) reported tht * vlue of LD from lm showed derese throughout the disply ph nd ltte The ph vlue of the musle 24 hours fter slughter ws n importnt ftor whih influened the instrumentl olour (Clnn et l. 2016). The result of present reserh onfirmed tht steks in HCS hd higher olour stility nd ph 24. The result my srie the reson to higher ph, whih n mintin the redution stte of myogloin nd olour stility (Yin nd Fustmn 1993; Hunt nd King 2012; Rihrds 2013). The umultion of ltte in musle ws losely relted to the hnge in ph fter slughter (Mtrneh et l. 2015). The results of our study onfirmed this onlusion. The extent of ph deline in steks of three groups ws linerly ssoited with the inrese of ltte on dy NADH, LDH-B, nd GAPDH The regenertion of NADH ply key role in mintining the

8 *** et l. Journl of Integrtive Agriulture 2017, 16(0): olour stility of met (Rmnthn et l. 2011, ). The steks in HCS group hd higher NADH thn steks in other two groups. Kim et l. (2008) reported tht LL possessed higher olour stility nd NADH ontent thn PM, whih ws prtilly onsistent with our results. GAPDH nd LDH-B re importnt dehydrogense enzymes whih n regenerte the NADH tht is neessry for the redution of metmyogloin. In support, previous reserh (Kim et l. 2009; Rmnthn et l. 2011) demonstrted tht LDH-B ply n importnt role in mintining the olour stility of met. This ws orresponding with our study, whih lso showed tht steks in HCS hd higher LDH-B tivity. Our present result is in prtil greement with the Kim et l. (2008) reports, whih indited tht LL possessed higher olour stility nd tivity of LDH-B thn PM. GAPDH n lso tlyze the formtion of NADH in the glyolyti pthwy. It hd een reported tht the tivity of GAPDH inreses the redution of metmyogloin in eef ground (Sleh nd Wtts 1968), nd steks of LL in olor-stle group hd greter GAPDH ontent thn steks in olor-lile (Cnto et l. 2015). The reserh of Cnto speulted tht the possile reson is tht the musle in olor-stle group whih hve more ontent glyolyti enzymes lso possess higher strong glyolyti metolism nd thus n enhne the prodution of NADH. For the first time, the result of the present study onfirmed tht the musle with high olor stility lso possess higher GAPDH tivity. It n e speulted tht the musle possessed higher tivity of GAPDH whih results in greter ontent of NADH, then the met hd lower rte of umultion of metmyogloin nd onsequently higher olour stility. 5. Conlusion The result of present reserh exhiited tht steks in HCS group hd higher tivity of GAPDH nd LDH-B. These two enzymes possessed higher tivity in the more stle olour steks, nd them my ply role in mintining stle olor in oordintion, s they n produe NADH. Future studies should fous on the reson wht used the inonsistent tivity of these two dehydrogense enzymes, nd the role of them in diverse niml musle, suh s pork nd eef, nd determine the tivity dehydrogense enzymes in glyolysis of norml met like DFD nd PSE with respet to met olor stility. Aknowledgements We grtefully knowledge finnil support from the Ntionl Agriulturl Siene nd Tehnology Innovtion Progrm, the Speil Fund for Agro-sientifi Reserh in the Puli Interest ( ) in Chin nd Chin Agriulture Reserh System (CARS-39). Referenes Bii T, Oukhttr L, Moutoukkil A, Soukri A Purifition nd hrteriztion of glyerldehyde-3- phosphte dehydrogense from europen pilhrd srdin pilhrdus. At Biohimi et Biophysi Sini, 39, Bo Y, Puolnne E, Ertjerg P Effet of oxygen onentrtion in modified tmosphere pkging on olor nd texture of eef ptties ooked to different tempertures. Met Siene, 121, Brron J T, Gu L, Prrillo J E Mlte-sprtte shuttle, ytoplsmi ndh redox potentil, nd energetis in vsulr smooth musle. Journl of Moleulr & Cellulr Crdiology, 30, Clnn H, Jo R H, Pethik D W, Grdner G E Prodution ftors influene fresh lm longissimus olour more thn musle trits suh s myogloin onentrtion nd ph. Met Siene, 119, Cnto A C, Cost-Lim B R, Sumn S P, Monteiro M L, Vin F M, Slim A P, Nir M N, Silv T J, Conte-Junior C A Color ttriutes nd oxidtive stility of longissimus lumorum nd psos mjor musles from Nellore ulls. Met Siene, 121, Cnto A C, Sumn S P, Nir M N, Li S, Rentfrow G, Beh C M, Silv T J, Wheeler T L, Shkelford S D, Gryson A, MKeith R O, King D A Differentil undne of sroplsmi proteome explins niml effet on eef Longissimus lumorum olor stility. Met Siene, 102, Go X G, Xie L, Wng Z Y, Li X M, Luo H L, M C W, Di R T Effet of postmortem time on the metmyogloin redutse tivity, oxygen onsumption, nd olour stility of different lm musles. Europen Food Reserh nd Tehnology, 236, Hunt M C, King A AMSA Met Color Mesurement Guidelines. Amerin Met Siene Assoition. Chmpign, Illinois, USA pp Joseph P, Sumn S P, Rentfrow G, Li S, Beh C M Proteomis of musle-speifi eef olor stility. Journl of Agriulturl & Food Chemistry, 60, Kim Y H Ltte dehydrogense regultion of the metmyogloin reduing system to improve olor stility of ovine musles through ltte enhnement. Disserttion Astrts Interntionl, 69, Kim Y H, Hunt M C, Mnini R A, Seyfert M, Loughin T M, Kropf D H, Smith J S Mehnism for ltteolor stiliztion in injetion-enhned eef. Journl of Agriulturl & Food Chemistry, 54, Kim Y H, Keeton J T, Smith S B, Berghmn L R, Svell J W Role of ltte dehydrogense in metmyogloin redution nd olor stility of different ovine musles. Met Siene, 83, Kim Y H, Keeton J T, Yng H S, Smith S B, Swyer J E, Svell

9 10 *** et l. Journl of Integrtive Agriulture 2017, 16(0): J W Color stility nd iohemil hrteristis of ovine musles when enhned with L- or D-potssium ltte in high-oxygen modified tmospheres. Met Siene, 82, King D A, Shkelford S D, Kuehn L A, Kemp C M, Rodriguez A B, Thllmn R M, Wheeler T L Contriution of geneti influenes to niml-to-niml vrition in myogloin ontent nd eef len olor stility. Journl of Animl Siene, 88, King D A, Shkelford S D, Rodriguez A B, Wheeler T L Effet of time of mesurement on the reltionship etween metmyogloin reduing tivity nd oxygen onsumption to instrumentl mesures of eef longissimus olor stility. Met Siene, 87, Klingenerg M Niotinmide-denine dinuleotides (NAD, NADP, NADH, NADPH) spetrophotometri nd fluorimetri methods. In: Methods of Enzymti Anlysis. 2nd ed. Ademi Press, New York nd London. pp Ledwrd D A Post-slughter influenes on the formtion of metyyogloin in eef musles. Met Siene, 15, Li W, Suve A A NAD(+) ontent nd its role in mitohondri. Methods in Moleulr Biology, 1241, Luino G, Monhn F J, Vst V, Pennisi P, Bell M, Priolo A Lipid nd olour stility of met from lms fed fresh herge or onentrte. Met Siene, 82, Mtrneh S K, Englnd E M, Sheffler T L, Oliver E M, Gerrrd D E Net ltte umultion nd low uffering pity explin low ultimte ph in the longissimus lumorum of AMPKγ3 R200Q mutnt pigs. Met Siene, 110, MCormik D B, Wright L D Niotini id: Anlogs nd oenzymes. In: Colowik S P, Kpln N O (Eds.), Methods in Enzymology, Vol. 18. New York nd London: Ademi Press. pp MKenn D R, Mies P D, Bird B E, Pfeiffer K D, Ellerht J W, Svell J W Biohemil nd physil ftors ffeting disolortion hrteristis of 19 ovine musles. Met Siene, 70, Neethling N E, Sumn S P, Sigge G O, Hoffmn L C Musle-speifi olour stility of lesok (Dmlisus pygrgus phillipsi) met. Met Siene, 119, Rmnthn R, Mnini R A, Ddy G A Effets of pyruvte, suinte, nd ltte enhnement on eef longissimus rw olor. Met Siene, 88, Rmnthn R, Mnini R A, Joseph P, Yin S, Ttiyorwornthm N, Petersson K H, Sun Q, Kond M R Effets of ltte on ground lm olor stility nd mitohondrimedited metmyogloin redution. Food Chemistry, 126, Rihrds M P Redox retions of myogloin. Antioxid Redox Signl, 18, Sleh B, Wtts B M Sustrtes nd intermedites in the enzymti redution of metmyogloin in ground eef. Journl of Food Siene, 33, Sumn S P, Joseph P Myogloin hemistry nd met olor. Annul Review of Food Siene nd Tehnology, 4, Yin M C, Fustmn C Influene of temperture, ph, nd phospholipid omposition upon the stility of myogloin nd phospholipid liposome model. Journl of Agriulturl & Food Chemistry, 41, (Mnging editor WENG Ling-yun)