Heat production partition in sheep fed above maintenece from indirect calorimetry data

Open Jou nal o Animal Sciences, 2015, 5, 86-98 Published Online Ap il 2015 in SciRes. h p://www.sci p.o g/jou nal/ojas h p://dx.doi.o g/10.4236/ojas.2015.52011 How o ci e his pape : C iscioni, P., del Ca men López, M., Zena, V. and Fe nández, C. (2015) Hea P oduc ion Pa i ion in Sheep Fed abo e Main enance om Indi ec Calo ime y Da a. Open Jou nal o Animal Sciences, 5, 86-98. h p://dx.doi.o g/10.4236/ojas.2015.52011 Hea P oduc ion Pa i ion in Sheep Fed abo e Main enance om Indi ec Calo ime y Da a Pa icia C iscioni1, Ma ía del Ca men López1, Vic o Zena2, Ca los Fe nández1* 1Resea ch Cen e ACUMA, Animal Science Depa men , Poly echnic Uni e si y o Valencia, Valencia, Spain 2In e uni e si y Ins i u e o Bioenginee ing Resea ch and Technology O ien ed o he Human Being, Uni e sidad Poli écnica de Valencia, Valencia, Spain Email: *cj e [email protected] .es Recei ed 11 Feb ua y 2015; accep ed 23 Ma ch 2015; published 27 Ma ch 2015 Copy igh © 2015 by au ho s and Scien i ic Resea ch Publishing Inc. This wo k is licensed unde he C ea i e Commons A ibu ion In e na ional License (CC BY). h p://c ea i ecommons.o g/licenses/by/4.0/ Abs ac The objec i e o his s udy is o compa e he pa i ion o hea ene gy (HE) in wo sheep b eeds by indi ec calo ime y and in eg al calculus. An expe imen was conduc ed wi h wo Spanish na i e sheep b eeds (d y and non-p egnan ) which we e ed wi h pelle ed mixed die s abo e main en- ance. Six Gui as and six Manchegas b eed sheep we e selec ed (58.8 ± 3.1 and 60.2 ± 3.2 kg body weigh , espec i ely). All sheep we e ed wi h he same concen a e mixed a ion (0.300 kg ce eal s aw as o age and 0.700 kg concen a e) in wo meals. Hal he daily a ion was o e ed a 800 h and ano he hal a 1600 h. The sheep had ee access o wa e . Sheep we e alloca ed in me abolic cages; ene gy balance and gas exchange we e assessed in each sheep. The s a is ical analyses in- cluded he ixed e ec o b eed and andom e ec o sheep. The me abolic ene gy (ME) o main- enance ep esen ed 69% o he o al ME in ake and he a e age was 354 kJ pe kg o me abolic body weigh (kg0.75 BW) on a e age. The basal me abolism (HeE) was g ea e (P < 0.05) in Gui a han Manchega b eed (270 ± 18 s. 247 ± 15 kJ/kg0.75 BW and day). As sheep we e ed wi h abo e main enance, he e ained ene gy in he body accoun ed o 22% o he ME in ake and 77% o he ME in ake was los as hea . 51% o he MEI was con e ed o HeE; 5% was los as physical ac i i y o s anding and lying down (HjE), and 13% was associa ed wi h he p ocess o eeding and wo k o diges ion and me abolism (HdE). Wi hin HdE, 47% ep esen ed he cos o in ake and eeding and 54% he cos o diges ion and me abolism. No di e ences in HE pa i ion be ween b eeds we e ound, al hough Gui a b eed showed less e iciency o ene gy e en ion han Manchega b eed. The e o e, his s udy demons a ed a en a i e app oach o pa i ioning HE, combining indi ec calo ime y and in eg al calculus. *Co esponding au ho . P. C iscioni e al. 87 Keywo ds Sheep, Hea P oduc ion Pa i ion 1. In oduc ion In he las ew decades a g ea numbe o e o s ha e been de o ed o measu ing ene gy expendi u e in animals. Animals p oduce hea om a a ie y o me abolic p ocesses such as main enance, he mo egula ion, physical ac i i y and p oduc ion (e.g. deposi ion o body issue, and milk p oduc ion). Indi ec calo ime y has played an impo an ole in measu ing his ene gy eleased as hea o hea p oduc ion (HE). In open ci cui espi a ion cham- be s, hea p oduc ion can be calcula ed based on he measu emen s o O2 consump ion and CO2 p oduc ion. The di ec measu emen o HE by indi ec calo ime y o e s he oppo uni y o e alua e a ia ions among animals in line wi h hei geno ype, pheno ype o en i onmen al condi ions. Hea ene gy can be u he pa i ioned in o ha associa ed wi h basal me abolism, olun a y ac i i y, p oduc o ma ion, diges ion and abso p ion, he mal egu- la ion, hea o e men a ion, and was e o ma ion and exc e ion. Pa i ion o HE in o meaning ul physiological o me abolic componen s is he mos di icul and con o e sial aspec o all eeding sys ems [1]. The calcula ion o hea inc emen in p oducing animals needs he pa i ioning o o al HE be ween a componen due o main en- ance and a componen due o p oduc ion. The as ing hea p oduc ion (FHP) pos eeding depends on he p e- ious eeding le el, and leng h o as ing, and o en includes a con ibu ion o physical ac i i y. The e o e, ac- i i y ela ed o al HE is he mos a iable componen o o al ene gy expendi u e [2]. Calo ime ic echniques can p o ide di ec access o o al HE, and expe imen al in e en ions and compu a- ional echniques a e equi ed o disen angle i s componen s [3]. The objec i e o his s udy is o p esen he me hodology de eloped in ou labo a o y o moni o o al HE and he hea inc emen associa ed wi h eeding in wo sheep b eeds by employing indi ec calo ime y and ma he- ma ical calculus. 2. Ma e ial and Me hods 2.1. Animals and Feeding. The expe imen al p ocedu e was app o ed by he Animal Use and Ca e Commi ee o he Poly echnic Uni e si y o Valencia (Spain) and ollowed he codes o p ac ice o animals used in expe imen al wo ks p oposed by he Eu opean Union [4]. Twel e mul ipa ous, d y and non-p egnan sheep o wo Spanish na i e b eeds we e used in his expe imen . Gui a b eed belong o Valencia P o ince and Manchega b eed o Cas illa-La Mancha P o ince (Spain), bo h a e au och honous b eeds and almos no in o ma ion abou ene gy pa i ioning is a ailable om hese b eeds. Ene gy me abolism in o ma ion would be use ul o ene gy equi emen s pu poses. Six Gui as and six Manchegas b eed sheep we e selec ed, which had simila body weigh (58.8 ± 3.1 and 60.2 ± 3.2 kg o BW, espec i ely). I is a ma u e weigh o Gui a [5] and almos o Manchega [6]; ma u e weigh in emales Gui a and Manchega anged be ween 50 - 60 and 65 - 75 kg, espec i ely. The expe imen was conduc ed as a andom design wi h he wo b eeds as ixed e ec . All sheep we e ed he same concen a e mixed a ion (0.300 kg ce eal s aw as o age and 0.700 kg concen a e) in wo meals. Hal he daily a ion was o e ed a 800 h and hal a 1600 h, espec i ely. Sheep had ee access o wa e . The concen a e was mixed and pelle ed along wi h he p emix. I s chemical composi ion alues on d y ma e (DM) basis was 92.62% o ganic ma e (OM), 17.19% c ude p o ein (CP), 47.33% neu al de e gen ibe (NDF), 25.37% acid de e gen ibe (ADF), 8.45% s a ch and 18.94 MJ o g oss ene gy (GE) pe kg DM, ollowing he ecommenda ion o [7] o sheep. Chemical composi ion o ce eal s aw was: 91.30% OM, 4.21% CP, 77.54% NDF, 50.33% ADF and 0.91% o s a ch. 2.2. Expe imen al Schedule and Measu emen s Sheep we e ed wi h expe imen al die s in pens o 10 days. Once adap ed o he die , he sheep we e alloca ed in me abolic c a es o 10 mo e days, in he moneu al condi ions (20˚C o 23˚C as de e mined by a Hobo p obe, ONSET da a logge s, Cape Cod, MA, USA). Feed in ake, e usals, u ine and aeces we e collec ed, weighed and P. C iscioni e al. 88 eco ded daily o each sheep o e a collec ion pe iod o 5 days. Faeces we e collec ed in wi e-sc een baske s placed unde he loo o he me abolic c a es and u ine was collec ed h ough a unnel in o plas ic bucke s con- aining 100 mlsulphu ic acid o main ain a pH below 3 o la e analysis. Samples o o age and concen a e, e- usals, aeces and u ine we e s o ed a −20˚C, and hen pooled o chemical analysis. Ruminal luid samples we e collec ed by s omach ube be o e he mo ning eeding on he las day o he sample collec ion pe iod. Ru- minal luid pH was immedia ely de e mined using a Model 265A po able pH me e (O ion Resea ch Inc., Be - e ly, MA, USA). A uminal luid sample was acidi ied wi h H2SO4 and ozen un il la e de e mina ion o am- monia ni ogen (ammonia-N). Samples o analysis o ola ile a y acids (VFA) we e mixed wi h H3PO4 and kep ozen un il analysis. The body weigh o each animal was aken a he beginning o he die adap a ion pe iod and a he end o he ecollec ion pe iod. Gaseous exchange was measu ed o each ewe du ing 24 h (6 sheep pe b eed, one animal pe day) using a mobile open-ci cui espi ome y sys em (head-hood) designed o small uminan s. As hal he daily a ion was o e ed a 800 h and hal a 1600 h, wice a day he head hood d awe was opened, lea ing he ood and a bucke o wa e inside. In o de o measu e as ing, h ee sheep pe b eed we e andomly selec ed, and dep i ed o ood o 3 days and he gaseous exchange measu emen was done on day 4. The espi ome y sys em has a head hood, a low me e (The mal Mass Flowme e Sensy low VT-S, ABB, Alzenau, Ge many) and ai suc ion p o ided by a cen i ugal an (CST60 Sole Palau Inc., Pa e s del Vallès, Ba celona, Spain). The me hane (CH4) and ca bon dioxide (CO2) concen a ion we e measu ed using he in a ed p inciple and oxygen (O2) was measu ed by he pa amagne ic p inciple (Easy low Gas Analyze , model 3020, ABB, Alzenau, Ge many). Al hough he uni was an au ocalib a ed model, he analyse s we e calib a ed wi h e e ence gases be o e each es . Fe nández e al. [8] desc ibed he mobile open-ci cui espi ome y sys em used and he di e ences now a e ha we use a head hood ins ead o a acemask and, he gas exchange da a acquisi- ion is con inuous. We desc ibed i b ie ly: The sys em was capable o eco d da a a in e als o 1 second. Gas analyze uni and low me e was connec ed o he compu e (Fuji su Siemens Li ebook Se ies, Pen ium 4 lap op, Munich, Ge many) by an uni e sal se ial bus (USB) connec o . A se ial communica ion p o ocol was used wi h i s p og ammable logic con olle s (MODBUS O ganiza ion, Inc., Hopkin on, MA, USA; www.modbus.o g) o communica ion be ween he analy ical de ices (analyse uni and low me e ) and he compu e . The elec onics p o o yping pla o m A duino (www.a duino.cc) was used o send da a om he gas analyse uni o LabVIEW7.1 (Na ional Ins umen s, Aus in, Texas, USA) by RS-232 p o ocol in eal ime. The low me e was moni o ed by a 10 bi analog o digi al con e e (model DS2438; Maxim In eg a ed P oduc s, Inc. Sunny ale, CA, USA). The whole sys em was calib a ed injec ing pu e N2 in o he head box [9], de e mined g a ime ically using a p ecision scale. Calib a ion ac o s we e calcula ed acco ding o [10]. The CH4 and CO2 p oduc ion and O2 con- sump ion we e calcula ed as desc ibed by [11]. An ini ial a mosphe ic ai sample was collec ed and he gas con- cen a ions we e used as e e ence o calcula ions. 2.3. Chemical Analysis Feed, eed e usal and eces samples we e i s d ied in a o ced ai o en a 55˚C o 48 h hen g ound o pass a 1 mm sc een be o e analysis. U ine was d ied by lyophiliza ion. Chemical analyses o he die , e usals and eces we e conduc ed acco ding o me hods o [12] o DM and ash. DM o die s and eces was de e mined by o en- d ying a 102˚C ± 2˚C o 24 h. Ash concen a ion was measu ed by incine a ion in an elec ic mu le u nace a 550˚C o 6 h o de e mine OM. The NDF and ADF we e measu ed in an ANKOM Fibe Analyze (A220, ANKOM Technologies, Fai po , NY, USA) acco ding o [13] and [12], espec i ely. NDF was de e mined using sodium sul i e and alpha amylase. S a ch con en was de e mined by enzyma ic me hod (α-amylase ob ained om Sigma-Ald ich, S einheim, Ge many) acco ding o [14]. The ni ogen (N) om eed, eces and u ine we e analysed by he Dumas p inciple (T uSpec CN; LECO Co po a ion, S . Joseph, MI, USA). Mul iplying N by a ac o o 6.25 con e ed he esul s o CP. The GE con en o he d ied samples ( eed, eces and u ine) was ana- lyzed by combus ion in an adiaba ic bomb calo ime e (Gallenkamp Au obomb; Loughbo ough, UK). NH3-N con en o uminal luid samples was analyzed by he Kjeldahl p ocedu e (2300 Kjel ec Analyze Uni Foss Teca o , Hille ød, Denma k). De e mina ion o uminal VFA was based on he me hod desc ibed by [15] using a gas ch oma og aph (Fisons 8000 se ies; Fisons Ins umen s SpA, Milan, I aly) equipped wi h a spli / spli less injec o and lame ioniza ion de ec o . P. C iscioni e al. 89 2.4. Calcula ions Theme abolizable ene gy in ake (MEI) was calcula ed as he di e ence be ween g oss ene gy in ake (GEI) and ene gy losses in eces, u ine and CH4 (wi h an ene gy equi alen alue o 39.5 kJ/L CH4 [16]. The HE was de e mined om measu emen s o O2 consump ion, CO2 and CH4 p oduc ion, and u ine N (Nu ), using he equa ion [16]: HE (kJ) = 16.18 × O2 + 5.02 × CO2 − 2.17 × CH4 − 5.99 × Nu whe e gases we e exp essed in li e s pe hou s and Nu in g ams pe day. The body issue ene gy (REbody) was calcula ed as MEI - HE. The ene gy associa ed wi h he oxida ion o p o ein (OXP), ca bohyd a e (OXCHO) and a (OXF) was cal- cula ed by he me hod o [17] and [18] o uminan s. The p oduc ion o ola ile a y acid om ca bohyd a e e men a ion is ollowed by CO2 and CH4 p oduc ion. A a io CO2:CH4 o 3:1 and 1.7:1 o high g ain and high o age die s, espec i ely [19]. The CO2 p oduc ion om oxida ion (CO2x) was calcula ed as CO2 − (CO2:CH4 × CH4). The calcula ions we e ca ied as ollowing: OXP = 6.25 × Nu × 18.42 (kJ/g), OXCHO = (−2.968 × O2 + 4.174 × CO2x − 2.446 × Nu ) × 17.58 (kJ/g), OXF = (1.719 × O2 − 1.719 × CO2x − 1.963 × Nu ) × 39.76 (kJ/g). Then, he HE om oxida ion (HxE) was: HxE (kJ) = 16.18 × O2 + 5.02 × CO2x − 5.99 × Nu . Gases we e exp essed in li e s pe hou s and Nu in g ams pe day. The non p o ein espi a o y quo ien om oxida ion o nu ien s (RQnpx) was de e mined as: RQnpx = (CO2x – (Nu × 6.25 × 0.774))/(O2 – (Nu × 6.25 × 0.957). As we men ioned abo e, e ained o eco e ed ene gy was de e mined as he di e ence be ween MEI and he HE. Hea ene gy associa ed wi h REbody (H E) and ME used o issue gain (ME ) we e based on an assumed e - iciency o die a y ME use o issue gain o 0.75 [20]. The e o e, ME o main enance (MEm) was es ima ed by di e ence be ween MEI and ME , and he e iciency o use o ME o main enance (km) as basal me abolic a e o e MEm. The N balance (N e ained) was de e mined as well, by di e ence among N in ake and eces plus u ine. 2.5. Hea P oduc ion Pa i ion The o al HE consis o many componen s: basal me abolism (HeE), hea associa ed wi h olun a y ac i i y (HjE), hea o p oduc o ma ion (H E), hea o he mal egula ion (HcE), hea o syn hesis and exc e ion o was e p oduc s (HwE), hea o diges ion (HdE) and hea o e men a ion (H E). This ene gy e ms was de ined acco d- ing o [1]. HE = HeE + HjE + H E + HcE + HwE + HdE + H E. Due o he animals being in a he moneu al and non-s ess ul en i onmen , HcE was assumed o be ze o. The NRC [1] de ines he hea inc emen o eeding (HiE) as: HiE = H E + HdE + H E + HwE. In ou ial we conside ed HwE negligible and, H E = HE – HxE. The e o e, he HE was pa i ioned be ween componen s due o eed in ake, physical ac i i y, and basal me a- bolic a e. This pa i ioning is desc ibed as ollows. HE = HeE + HjE + HiE. The main enance equi emen s consis o basal me abolic a e plus he ac i i y inc emen , and a e usually de- ined as FHP; he e o e we de ine FHP as a sum o wo componen s; HeE and HjE. The HeE co esponded o he minimum ene gy expendi u e o es ing, heal hy, non- ep oduc i e, as ing and adul animal ha a e in a he - moneu al en i onmen du ing he inac i e ci cadian phase [21]. The ac i i y inc emen was assumed in ou s udy as he di e ence be ween FHP and HeE. HjE = FHP – HeE. The e m de ined by NRC [1] as hea o diges ion (HdE) includes he ene gy cos o ea ing, umina ion, wo k o diges ion and nu ien me abolism. The HdE was de e mined as ollows: HdE = HE – HeE – H E – HjE. Then, HdE was di ided in HE o ea ing (HdEe) and HdE o diges ion and me abolism (HdEdm). The cos o P. C iscioni e al. 90 ea ing is de ined in he li e a u e as he ac o p ehending, bi ing, chewing, sali a ing and swallowing h oughou a ime pe iod (use o be 15 minu es, acco ding o [22] and [2]). In ou s udy he cos o ea ing (HdEe) is guided by he inc ease in HE om he momen ha eed is o e ed on he eede o 3 hou s la e , and we measu e he peak a ea unde he cu e. The HdEdm was ob ained by di e ence and we assumed included he wo k o diges- ion and me abolism. The wo HdE componen s include umina ion and mus include he ene gy cos (oxygen consump ion) o gas oin es inal and hepa ic o gans, al hough each o hese ac i i ies was no quan i ies. HdE = HdEe + HdEdm. Figu e 1 shows some nomencla u e desc ibed abo e and we can ealize ha he pa i ion o HdE in HdEe and HdEdm is heo ical, based on he cu e pa e n o he me abolic a e (HE). 2.6. Da a Analyses The quan i ica ion o pa i ioning HE was de eloped by in eg al calculus in R ( e sion 2.12.2, R Founda ion o S a is ical Compu ing, Viena, Aus ia). Du a ion o gas exchange was ixed a 1 day, and he ime uni was mi- nu es. The in eg a ed a eas we e: HE = The To al Hea P oduc ion in ed and as ed animal was de e mined using in eg al o cu es o med by da a om indi ec calo ime y. HiE = Di e ence o in eg als o HE and FHP. FHP = In eg al o as ing hea p oduc ion, di ided in HeE and HjE. HjE = Di e ence o in eg als FHP and HeE. HeE alue was de e mined as he a ea unde he cu e de ined by he alue epea ed mo e equen ly (mode) o he mean alues o da a ob ained om he espi ome y o 3 as ing animals, once he highes alues we e d opped, which could be caused by mo emen s. HdEe = Di e ence o in eg als o o al HE and FHP in an app oxima e 3-hou pe iod a e eeding. H E = Is he di e ence be ween in eg als o HE and HxE. De ini e in eg al o a signal can be in e p e ed as he a ea unde he cu e. This nume ic in eg a ion allows us o e alua e he de ined in eg al o a con inuous unc ion in a closed in e al wi h he desi ed accu acy, and i consis s in he adjus men o a polynomial o a se o poin s and hen in eg a ing hem. The in eg al o each cu e was based on he apezium me hod. The sum o hese a eas o apeziums can be app oxima ed o he in eg al ha ep esen s he a ea unde he cu e, acco ding o he ollowing equa ion: Figu e 1. Example o hea p oduc ion (HE) pa i ioning and hei componen s: as ing hea p oduc ion (FHP), basal me abolism (HeE), hea associa ed wi h olun a y ac i i y (HjE), hea o diges ion (HdE), hea o eeding du ing 3 hou s (HdEe), hea a e 3 hou s o eeding o nex in ake (HdEdm). FHP = HeE + HjE; HdE = HdEe + HdEdm. P. C iscioni e al. 91 1 Δ n h ii ih S x α αα = = ⋅ ∑ whe e S α ep esen he a ea o empo al se ies i x α o in e als Δi α o one minu e o inc ease du ing 24 hou s co esponding o he animal α. In his s udy he S α and i x α alues can be eplaced by he desc ip ions in Table 1. The e ec s o b eed on hea pa i ion we e analyzed using he PROC MIXED o [23]. The expe imen was conduc ed in a andomized design and he model o he dependen a iables included he ixed e ec o b eed and andom e ec o sheep. The ollowing s a is ical model was used: Y = μ + α (B) + sheep + ε whe e Y is he de- penden a iable, μ is he o e all mean, B is he b eed (Manchega and Gui a), sheep is andom e ec and ε is he andom e o . E ec s we e decla ed signi ican a P < 0.05 and P- alues be ween 0.05 and 0.10 we e con- side ed as a end. S uden ’s es was used o compa ison be ween b eeds. 3. Resul s The da a on ene gy in ake and ou pu s o he sheep, eco ding du ing he calo ime ic measu emen , a e p e- sen ed in Table 2. No signi ican di e ence was obse ed o he ene gy balance be ween b eeds. HE pa i ioning a e shown in Table 3. The as ing me abolism in sheep was measu ed a day 4 a e h ee days o s a a ion, when espi a ion quo ien has usually allen o abou 0.70 [24]. No di e ences we e obse ed o FHP be ween b eeds (258 kJ/kg0.75 BW and day, on a e age). The basal me abolism (HeE) was signi ican ly (P < 0.05) highe o Gui a han Manchega b eed (270 s. 247 kJ/kg0.75 BW and day, espec i ely) and no di - e ences o ac i i y we e ound (27 kJ/kg0.75 BW and day). Table 1. De ini ion o he a eas and empo al se ies used in he nume ical in eg a ion. S α i x α hn-h1 HE ( o al hea p oduc ion) Heαi (hea p oduc ion pe hou ) 24 hou s FHP ( as ing hea p oduc ion) hpαi (hea p oduc ion pe hou ) 24 hou s HiE (hea inc emen ) hiEαi (hea inc emen pe hou ) 24 hou s HdEe (Hea inc emen pos inges ion) hdEcgαi (hea inc emen pe hou ) App ox. 3 hou s H E (hea o e men a ion) h Eαi (hea inc emen pe hou ) 24 hou s Table 2. D y ma e in ake (g/d) and ene gy balance (kJ/kg0.75 BW) in wo sheep b eeds. B eeds SEM1 P- alue Gui as Manchegas LW2, kg 58.0 60.2 1.16 0.981 DMI3, g/d 883.3 984.8 45.23 0.287 Ene gy balance, kJ/kg0.75 BW GEI4 803 854 25.3 0.34 E 5 eces 226 260 28.8 0.56 E u ine 41 29 5.5 0.31 E me hane 50 46 3.5 0.66 ME6 487 518 13.4 0.260 RE7body 99 124 14.1 0.42 REp o ein 48 70 7.4 0.11 RE a 52 54 12.6 0.98 ME gain 132 165 18.8 0.42 ME main enance 354 353 8.3 0.95 km8 0.77 0.70 0.021 0.099 1SEM = s anda d e o o he mean; 2BW = body weigh ; 3DMI = d y ma e in ake; 4GEI = g oss ene gy in ake; 5E = ene gy; 6ME = me abolizable ene gy in ake; 7RE = e ained ene gy; 8km = e iciency o ME. P. C iscioni e al. 92 Table 3. HE pa i ioning (kJ/kg0.75 BW) and HE pa i ioning pe MEI (%) in wo sheep b eeds. B eeds SEM1 P- alue Gui as Manchegas HE pa i ioning, kJ/kg0.75 BW HE2 387 394 5.5 0.56 H E3 33 41 4.7 0.42 HeE4 270 247 4.0 0.02 HjE 5 23 30 1.2 0.453 FHP6 293 277 2.8 0.19 HdE7 54 71 8.8 0.39 HdEe8 24 31 5.3 0.57 HdEdm9 30 40 5.4 0.16 HiE10 61 77 8.7 0.40 HE pa i ioning pe MEI11, % RE12body/MEI 20 23 2.4 0.58 REp o ein/MEI 10 14 1.3 0.12 RE a /MEI 11 10 2.4 0.81 HE/MEI 80 77 2.4 0.58 H E/MEI 7 8 0.8 0.57 HeE/MEI 56 48 1.8 0.02 HjE/MEI 5 6 0.2 0.01 FHP/MEI 60 54 1.8 0.06 HdE/MEI 11 14 2.0 0.49 HdEe/MEI 5 6 1.2 0.61 HdEdm/MEI 5 8 1.2 0.21 HiE/MEI 13 15 2.0 0.52 1SEM = s anda d e o o he mean; 2HE = hea p oduc ion; 3H E = hea o p oduc o ma ion; 4HeE = basal me abolism; 5HjE = hea associa ed wi h olun a y ac i i y; 6FHP = as ing hea p oduc ion; 7HdE = hea o diges ion; 8HdEe = hea o ea ing; 9HdEdm = hea o diges ion and me abolism; 10HiE = hea inc emen o eeding; 11MEI = me abolizable ene gy in ake; 12RE = e ained ene gy. The a ia ion in HE associa ed wi h eeding (HdE) was no signi ican ly di e en be ween b eeds (62 kJ/ kg0.75 BW and day, on a e age). No signi ican di e ences we e obse ed o he wo componen s ela ed o hen; ea ing-chewing- umina ion (HdEe) and umina ion-diges ion-me abolism (HdEdm). The p opo ional con ibu ion o HxE due o oxida ion o nu ien s is shown in Table 4. No di e ences we e obse ed o HxE and OXF and, di e ences we e ound in OXCHO and OXP; lowe OXCHO (P < 0.05; 75 s. 129 kJ/kg0.75 BW) and highe OXP (P < 0.01; 52 s. 30 kJ/kg0.75 BW) in Gui a han in Manchega b eed o sheep. No signi ican di e ences we e obse ed in N balance, only Gui a b eed shown g ea e (P < 0.05) alues in u ine N compa ed wi h Manchega b eed: 0.65 s. 0.35 g/kg0.75 BW, espec i ely (Table 5). Table 6 shows he uminal pa ame e s like, pH, ammonia-N and VFA. Highe ammonia-N alues (P < 0.05) we e ob ained in- Gui a b eed compa ed wi h Manchega b eed (22.8 s. 17.81 mg/dL, espec i ely) and nume ical lowe VFA (P = 0.056) in Gui a compa ed wi h Manchega (33.49 s. 37.46 mmol/L). 4. Discussion 4.1. HE Pa i ion: Ac i i y The HjE es ima ed in ou expe imen included only he ac o s anding and lying down. The a e age alue o HjE, in sheep alloca ed in me abolic cages was 27 kJ/kg0.75 BW and day. This alue ep esen s he 8.5% o he HeE (exp essed on FHP he alues is 10% on a e age). NRC [25] sugges ed ha ene gy equi emen s o ac i - i y we e 10% o FHP o s all ed sheep. P. C iscioni e al. 93 Table 4. Ene gy (kJ/kg0.75 BW) associa ed wi h he oxida ion o nu ien s and hei pe cen age o e HxE in wo sheep b eeds. B eeds SEM1 P- alue Gui as Manchegas Oxida ion o nu ien s, (kJ/kg0.75 BW) HxE2 381 389 5.8 0.52 H E3 7 6 0.7 0.65 OXP4 52 30 4.6 0.01 OXCHO5 72 129 19.1 0.04 OXF6 256 229 16.2 0.43 OXP/HxE, % 14 8 1.2 0.02 OXCHO/HxE, % 19 33 5.0 0.05 OXF/HxE, % 68 59 4.5 0.36 RQnpx7 0.81 0.87 0.033 0.521 HxE/MEI8, % 78 76 2.3 0.60 H E/MEI, % 1.3 1.2 0.14 0.521 1SEM = s anda d e o o he mean; 2HxE = hea p oduc ion o oxida ion; 3H E = hea o e men a ion; 4OXP = ene gy associa ed wi h he oxida ion o p o ein; 5OXCHO = ene gy associa ed wi h he oxida ion o ca bohyd a e; 6OXF = ene gy associa ed wi h he oxida ion o a ; 7RQnpx = non p o- ein espi a o y quo ien om oxida ion o nu ien s; 8MEI = me abolizable ene gy in ake. Table 5. Ni ogen balance (g/kg0.75 BW) in wo sheep b eeds. B eeds SEM1 P- alue Gui as Manchegas N2 in ake 1.28 1.19 0.033 0.211 N eces 0.30 0.35 0.031 0.429 N u ine 0.65 0.35 0.060 0.003 N e ained 0.33 0.49 0.050 0.109 RP o ein3, g/d 43 64 6.2 0.09 RFa 4, g/d 27 29 8.1 0.58 Gain, g/d 199 285 23.3 0.12 1SEM = s anda d e o o he mean; 2N = ni ogen; 3RP o ein = e ained p o ein; 4RFa = e ained a . Table 6. pH, ammonia N and ola ile a y acids (VFA) in wo sheep b eeds. B eeds SEM1 P- alue Gui as Manchegas pH 7.30 7.22 0.172 0.652 Ammonia N2, mg/dL 22.80 17.81 3.256 0.048 To al VFA3, mmol/L 33.49 37.46 3.276 0.056 VFA, mmol/L Ace ic 23.47 26.43 1.552 0.081 P opionic 6.70 5.80 0.982 0.18 Isobu y ic 0.88 0.68 0.111 0.07 Bu y ic 3.55 3.15 0.573 0.55 Iso ale ic 1.28 0.93 0.142 0.24 N- ale ic 0.51 0.32 0.024 0.48 N-cap oic 0.07 0.09 0.003 0.35 1SEM = s anda d e o o he mean; 2N = ni ogen; 3VFA = ola ile a y acids. P. C iscioni e al. 94 Calo ime ic s udies ha e es ablished he ollowing ene gy cos o a ious physical ac i i ies by uminan s animals [26], and s anding compa ed wi h lying esul in a cos o 10 kJ/kg BW and day. In ou s udy we as- sumed ha Manchega made mo e posi ion changes han Gui a b eed (63 compa ed wi h 48 posi ion changes, espec i ely). Goa s no mally p e e s anding while eeding and, in ou s udy mo e ac i i y (s anding up) and mo e DMI was ound in Manchega sheep han Gui a b eed (985 s. 883 g DMI/d, espec i ely). Howe e , al- hough nume ically di e en , DMI was no signi ican and conside ing DMI as pe cen age o BW, bo h b eeds showed simila DMI (1.6% BW). I we exp ess HjE pe g am o DMI we ob ain 0.55 and 0.66 kJ HjE/g DMI o Gui a and Manchega b eed, espec i ely. The e o e, he g ea e alue o physical ac i i y in Manchega b eed was no due o nume ically highe DMI. 4.2. HE Pa i ioning: Feeding The HE has been shown o inc ease du ing eeding in sheep [22] like in o he mammals. Con inuous measu e- men s o espi a o y exchange consis en ly show ha HE in sheep inc eases apidly by 40% - 80% du ing a cou se o a meal. This inc ease pe sis ed e en h ough meals las ing up o 2 hou s bu declines he ea e apidly o a es no mo e han 15% - 20% g ea e han hose eco ded be o e a meal (Figu e 1). Mos o he s udies calcula ed he cos o ea ing o a sho pe iod o ime (15 minu es) and he a e o in ake was eco ded (g DM pe minu e o bi es pe minu e). This cos is calcula ed om he inc emen in HE abo e he a e age HE o he p e- eeding pe iod. And i was ela ed o he ype and amoun o eed consumed and also o he ime spen on ea ing. Ou me hodology was comple ely di e en , wi h wo eeding pe iods and 24 hou s o con inuous HE measu emen , he measu es we e based on he g ea e peak o HE a e eeding combined wi h nume ical in eg a ion. The e o e ou esul s a e no compa able due ha we did no eco d he a e o in ake. We ound ha he cos o in ake du ing 6 hou s ( wo meals) we e 28 kJ HdEe/kg0.75 BW and day on a e age, and he cos o diges ion and me abolism succeeding 18 hou s was 35 kJ HdEdm/kg0.75 BW and day. The cos o in ake wi h wo meals ep esen he 47% o he o al HdE, while 54% would be he cos o diges ion and me abolism o he sheep on he me abolic cages wi h almos minimal physical ac i i y. The HdEe o wo meals accoun o 6% o he MEI, while 7% o he MEI was expended in HdEdm, and no compa able esul was ound om he li e a- u e (Table 3). The H E was mo e ela ed o he cos o me hane p oduced han o al cos o e men a ion, and due o his eason H E was de e mined by di e ence be ween HE and HxE, wi h alues o 7 kJ/kg0.75 BW, on a - e age (Table 4). In ou s udy, whe e sheep we e ed abo e main enance wi h mixed die and concen a e pelle ed and, non- locomo ion and he mal s ess was conside ed, he HiE ep esen ed 14% o he MEI. The o al ene gy cos o inges ion and diges ion a e consis en ly 40% o 50% o HiE o o ages bu a e less o ba ley pelle s; 24% - 37% [27]. 4.3. ME o Main enance Feeding le el in ou ial was es ima ed as 1.5 imes main enance, hence he sheep was in posi i e issue ene gy balance, and some o he MEI is being di ec ed owa ds issue ene gy gain. The MEm was es ima ed by di e - ence be ween MEI and ME and he alue ob ained was 354 kJ/kg0.75 BW and day on a e age. The e iciency o use o ME o main enance was 0.74 on a e age (sligh ly highe han NRC [25]), because we ha e a concomi- an ene gy use o main enance and gain. The e o e, nex a e age alues we e obse ed (Table 3); he MEm ep esen ed he 69% o he o al MEI in his s udy. As sheep we e ed abo e main enance, he REbody accoun o 23% o he MEI and, 77% o he MEI was los as hea (HE). Wi hin HE, 51% o he MEI d o e o basal me abolism (HeE), 5.3% was los as physical ac i i y o s anding and lying down (HjE), and 13% was associa ed wi h he p ocess o eeding and wo k o di- ges ion and me abolism. Re iewing he li e a u e, we ound a iabili y in de e mina ion o es ima ion o MEm. Manchega sheep had a alue o 383 kJ/kg0.75 BW in he s udy [28] and day and [29] [30] in Gui a sheep ound a alue o 352 kJ/kg0.75 BW and day and 391 kJ/kg0.75 BW, espec i elyand an e iciency o use o ME o main enance o 0.64 (km). The es ima ed MEm equi emen in Segu eña sheep by linea eg ession was 374 kJ/kg0.75 BW and he e i- ciency o u ilisa ion o ME o main enance (km) was 0.72 [31]. Nine epo s [32] sugges MEm anging om 305 o 460 kJ/kg0.75 BW, and [33] es ima ed a MEm o Menz sheep o he E hiopian highlands 422 kJ/kg0.75 BW and day. The sys ems [25] and [34] adop ed an a e age alue o 321 and 397 kJ/kg0.75 BW, espec i ely due o he ac ha FHP s. eeding ial o , calo ime y s udies s. o he me hods, gi e di e en alues. O he s [35]