Ensiling characteristics and aerobic stability of temperate grasses containing different concentrations of water soluble carbohydrates

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1 REPORT TO THE STAPLEDON MEMORIAL TRUST Ensiling chrcteristics nd eroic stility of temperte grsses contining different concentrtions of wter solule crohydrtes Aner A. Rodríguez-Crís Deprtment of Animl Science, University of Puerto Rico, Myguez Cmpus Host Institution: Institute of Biologicl, Environmentl nd Rurl Sciences (IBERS) Aerystwyth University Period of Fellowship My 28 to July 4, Introduction An increse in wter solule crohydrte (WSC) concentrtion hs een ssocited with higher energy vlues in grss forge nd utiliztion efficiency of grss nutrients for milk production (Miller et l., 2001). Likewise, effective forge preservtion s silge requires n optimum WSC content in the vegettive mteril to generte fermentcion with low ph, high lctic cid content, nd low mmoni production. In temperte grss forges, glucose, fructose, nd sucrose, re the primry WSC in the vegettive mteril cell content (Smith, 1973). New vrieties of perennil ryegrss contining high (>15%) WSC content hve een develop in the UK. Other temperte grsses such s timothy nd cockfoots grsses contin moderte or low levels of solule sugrs (Sveinsson nd Bjrndotti, 2004, Smd et l., 2004). Differences in WSC concentrtions mong temperte grsses my ffect the qulity of the fermenttion process nd eroic stility of the resulting silge. This experiment ws conducted to determinte the ensiling chrcteristics nd eroic stility of the temperte grsses; Perennil ryegrss (Lolium perenme vr. AerMgic), Timothy grss (Phleum pretense vr. S48), nd Cocksfoot (Dctylis glomerte vr. Aertop) contining different concentrtions of WSC on dry mtter sis. Mteril nd Methods Crop mteril Four replicte plots (36*10 mt) of Perennil ryegrss, Timothy grss, nd Cocksfoot were used s vegettive mteril to determine the ensiling chrcteristics nd eroic stility of the three grss species. Forges were hrvested under frm conditions. Forge from the four plots of ech species were mixed nd wilted during 24 hours efore ensiling. After wilting, vegettive mteril ws chopped in forge hrvester to pieces of 3 5 cm length nd trnsported to the lortory. A 500g representtive smple from ech species ws tken for nutritionl vlue nlyses using stndrd procedures (AOAC, 1990, Vn Soest, et l., 1991). Fermenttion Process Mini-silos with net volume of 1.8 litre were used to determine the ensiling chrcteristics of the three grsses. Thirty-six mini-silos were tightly filled with out 650 g of the fresh crop, 1

2 immeditely seled, nd kept t constnt temperture of C. For ech forge species three smples were collected fter 2, 7, 14, nd 21 dys of fermenttion. Smples were stored for lter determintion of ph, chemicl nlysis, concentrtion of voltile ftty cids (e.g. cetic, utyric nd propionic cids), lctic cid, ethnol nd mmoni N (g/kg of totl nitrogen). In vitro dry mtter (IVDVD) nd NDF (IVNDFD) degrdility from ech forge species were estimted fter 21 dys of fermenttion using the ANKOM procedure. Aeroic Stility After 14 nd 21 dys of fermenttion, g replictes of silge from ech forge specie were sujected to n eroic stility test wherey the silge ws exposed to ir for 7 dys. Temperture s the criterion to determine eroic stility ws mesured using dt loggers tht recorded redings once per six hours from thermocouple inserted into the mid point of representtive silge smples erted in open polystyrene oxes. The oxes were keep t constnt monitored room temperture (20-22 C). Aeroic deteriortion ws denoted y dys (or hours) until the strt of sustined increse in temperture of more thn 3 C ove room temperture. Sttisticl Anlysis Chemicl composition, ph, nd fermenttion products dt were nlyzed s completely rndomized design with 3 (forge specie) y 4 (dy of fermenttion) fctoril rrngement of tretments using the Generl Linel Model Suroutine of SAS (1990). The ANOVA model ws s follows: Y ijk = µ + A i + B j + (A*B) ij + E ijk Where: Y ijk = Individul response vrile mesured (i.e. chemicl components, ph) µ = Overll men A i = Effect of forge specie B j = Effect of fermenttion dy A*B ij = Interction of forge specie * fermenttion dy E ijk = rndom residul error For in vitro DM nd NDF degrdility completely rndomized design ws utilized with 3 replictes per forge species. For ech model, significnt min effect nd interctions mong mens were seprted using Tukey t-test (SAS, 1990). Aeroic stility dt were nlyzed ccording to split plot design with 3 (forge specie) * 2 (lenght of fermenttion) * 7 (dys of eroic exposure) *29 (hour points during the eroic exposure period ) fctoril rrngement, usign the silo s repetitive mesurement during the 7 dys of eroic xposure. Y ijk = µ + A i + B j + (A*B) ij + C k + (A*C) ik + (B*C) jk + (A*B*C) ijk + D l + (A*D) il + (B*D) jl +(A*B*D) ijl + (C*D) kl + (A*C*D) ikl + (B*C*D) jkl + (A*B*C*D) ijkl + E m + F ijklmn 2

3 Where: Y ijk = Individul response vrile mesured (temperture) µ = Overll men A i = Effect of forge specie B j = Effect of length of fermenttion (14 or 21 dys) A*B ij = Interction of forge specie* length of eroic exposure C k = Effect of dy of eroic exposure A*C i k = Interction of forge specie* dy of eroic exposure B*C jk = Interction length of fermenttion * dy of eroic exposure A*B*C ijk = Interction forge species * length of fermenttion * dy of eroic exposure D l = Effect of hour of eroic exposure A*D il = Interction forge specie * hour of eroic exposure B*D ij = Interction length of fermenttion * hour of eroic exposure A*B*D ijl = Interction forge species * length of fermenttion * hour of eroic exposure C*D kl = Effect of dy of eroic exposure * hour of eroic exposure A*C*D ikl = Interction forge species * dy of eroic exposure * hour of eroic exposure B*C*D jkl = Interction length of fermenttion * dy of eroic exposure * hour of eroic exposure. A*B*C*D ijkl = Interction forge specie *length of fermenttion * dy of eroic exposure * hour of eroic exposure Em = rndom effect of silo = rndom residul error E ijklmn Results nd Discussion Initil chemicl composition of the vegettive mteril utilized in this experiment is typicl of vlues reported for grsses hrvested in temperte climtes (Tle 1; Sveinsson nd Bjrndotti, 2004, Smd, et l., 2004). All three grsses fter wilting showed etween 28 to 33% DM, to 92.75% OM, nd 7.24 to 7.62% IM. As expected, perennil ryegrss hd the highest (P<0.05) WSC content s compred to timothy grss nd cocksfoot. There were lso significnt differences mong grss species in initil CP, NDF, ADF nd hemicelulose contents. Perennil ryegrss hd lower (P<0.05) cell-wll percentge nd ligno-cellulolitic frction mong the grsses evluted, however, CP content ws 2.44 nd 2.58 percentge units higher in timothy grss thn perennil ryegrss nd cocksfoot, respectively. The chemicl chrcteristics of the three grsses during the course of 21 dys of ensiling re shown in Tle 2. During the whole fermenttion process DM content ws higher (P<0.05) in cocksfoot thn timothy grss nd perennil ryegrss grss silge, ut orgnic nd inorgnic mtter contents were similr. Crude protein content ws higher (P<0.05) in fermented timothy grss thn in the other two grss species. During the entire ensiling process WSC ws higher (P<0.05) nd totl cellwll, ligno-celulose frctions, nd hemicelulose content were lower (P<0.05) in perennil ryegrss silge thn timothy nd cocksfoot silge. In this experiment, the initil WSC content in the vegettive mteril rnged from to %. 3

4 Tle 1. Chemicl composition of the temperte grsses Perennil ryegrss, Timothy grss nd Cocksfoot efore ensiling Component (%) 1 Timothy grss Perennil grss Corkfoots Dry Mtter Orgnic Mtter Inorgnic Mtter Crude Protein WSC c NDF ADF Hemicelulose 1, Dry Mtter Bsis 2 Wter Solule Crohydrtes 3 Hemicelulose = NDF ADF, Mens with unlike superscripts in the sme row differ (P<0.05) A decresed of 4.18, 5.07, nd 2.80 percentges in the WSC content in timothy grss, perennil ryegrss, nd cocksfoot ws oserved during the initil 2 dys of fermenttion, respectively. This decrese in solule sugrs content is consistent with n ctive fermenttion during the ensiling process. However, for ll three grsses very slow fermenttion ws oserved s evidenced y reltively high ph nd low lctic cid concentrtions (Figures 1 nd 2). In the three fermented grsses nerly identicl ph ws oserved during the first two dys of fermenttion; therefter, the ph in perennil ryegrss silge ws lower (P<0.05) from dy 4 to 21 of fermenttion thn in timothy nd cocksfoot. Lctic cid content fter 2 dys of fermenttion ws lso higher (P<0.05) in perennil ryegrss silge thn in the other two grsses. Acetic cid (Figure 3) nd ethnol (Figure 4) content were lso higher (P<0.05) in perennil ryegrss silge thn timothy nd cocksfoot from dy 4 to 14, nd fter 14 nd 21 dys of fermenttion, respectively. For ll silges the reltion NH 3 - N/Totl-N ws similr nd lower thn 8%, which indictes low level of proteolysis during the fermenttion process (Figure 5). The quntittive indictors of fermenttion of the three temperte grsses ensiled were less thn expected, ut the present trends re s those reported in other relted experiments (Downing et l., 2008). Perennil ryegrss conserved s silge showed etter fermenttion chrcteristics thn the other two grsses evluted, s evidenced y lower ph nd higher lctic cid content. Vrition in chemicl composition nd epiphytic micro-flor hs een reported in forges hrvested in temperte climtes t the sme stge of growth. In this experiment, the wilting period efore ensiling might lso hve influence the initil microil popultions or the pprent low ctivity level of micro-flor ssocited with the fermenttion. A chnging micro-flor during the ensiling process using different sustrtes (i.e. lctic cid), implies very heterogenous microil popultion, which might lso help to explin the slow fermenttion process. The possily greter numers of undesirle microorgnisms present in the wilted-chopped forge prior to ensiling nd during the fermenttion period, or n inility for the desirle orgnisms to compete, might lso ffect their growth rte nd metolic ctivity. 4

5 Tle 2. Effects of ensiling dy on chemicl composition of Perennil ryegrss, Timothy grss nd Cocksfoot silge Item Fermenttion Dy Timothy grss Perennil ryegrss Cocksfoot Chemicl Composition (%) Dry Mtter Orgnic Mtter Inorgnic Mtter Crude Protein WSC 1, c c NDF ADF Hemicelulose 1, Dry Mtter Bsis, 2 Wter Solule Crohydrtes, 3 Hemicelulose = NDF ADF,,,c Mens with unlike superscripts in the sme row differ (P<0.05) 5

6 7 6.5 ph Fermenttion dy Timothy grss Perennil ryegrss Cocksfoot Figure 1. ph of timothy grss, perennil ryegrss, nd cocksfoot silge Mens with unlike superscripts differ (P<0.05) g/kg DM c c c 10 c Fermenttion dy Timothy grss Perennil ryegrss Cocksfoot Figure 2. Lctic cid content of timothy grss, perennil ryegrss, nd cocksfoot silge Mens with unlike superscripts differ (P<0.05) 6

7 g/kg DM c Timothy grss Perennil ryegrss Cocksfoot Fermenttion dy Figure 3. Acetic cid content of tinothy grss, perennil ryegrss, nd cocksfoot silge Mens with unlike superscripts within the sme fermenttion dy differ (P<0.05) g/kg DM Fermenttion dy Timothy grss Perennil ryegrss Cocksfoot Figure 4. Ethnol content of timothy grss, perennil ryegrss, nd cocksfoot silge Mens with unlike superscripts within the sme fermenttion dy differ (P<0.05) 7

8 % Fermenttion dy Timothy grss Perennil ryegrss Cocksfoot Figure 5. Reltion NH 3 -N/totl-N of timothy grss, perennil ryegrss, nd cocksfoot silge In this experiment the IVDMD nd IVNDFD vlues of the resulting silges were similr, nd 61.61, nd 71.39, nd 74.26, nd 60.98% for timothy grss, perennil ryegrss nd cocksfoot silge, respectively (Figure 6) % IVDMD IVNDFD Timothy grss Perennil ryegrss Cooksfoot Figure 6. IVDMD nd IVNDFD of timothy grss, perennil ryegrss, nd cocksfoot silge The eroic instility of silges is evidenced y n increse in temperture s result of the utiliztion of residul WSC nd lctic cid y eroic cteri, yest nd fungi. In this experiment the three temperte grss silges fermented during 14 nd 21 dys, were exposed to ir during 168 h nd temperture monitored every 6 hours. For oth lengths of fermenttion, temperture ws higher (P<0.05) in cocksfoot silge thn in those of timothy grss nd perennil ryegrss (Tle 3). During the 7 dys of eroic exposure, temperture in perennil ryegrss silge ws lower (P<0.05) thn 8

9 tht oserved in timothy grss (dys 2 thru 5) nd cocksfoot (dys 1 thru 7; Tle 4). Temperture in cocksfoot silge ws lso higher (P<0.05) thn for timothy grss the lst three of eroic exposure. For ll silges, the gretest chnges in temperture were oserved during the first 6 to 36 h of eroic exposure (Figure 7). However, cocksfoot silge hd higher (P<0.05) temperture thn timothy grss nd perennil ryegrss silge during the initil 12 to 36 h of eroic exposure, respectively. Tle 3. Effect of length of fermenttion on temperture of Perennil ryegrss, Timothy grss nd Cocksfoot, silge fter exposure to ir during 7 dys Temperture C Length of Fermenttion (d) Perennil ryegrss Timothy Grss Cocksfoot , Mens with unlike superscripts in the sme row differ (P<0.05) Tle 4. Effect of dy of eroic exposure on temperture of Perennil ryegrss, Timothy grss nd Cocksfoot silge Temperture C Aeroic Exposure (d) Perennil ryegrss Timothy Grss Cocksfoot ,,c Mens with unlike superscripts in the sme row differ (P<0.05) An increse of 3 C ove mient temperture ws oserved fter 12 hour of eroic exposure in cocksfoot silge, nd fter 18 hour in fermented timothy grss nd perennil ryegrss. In summry, in this experiment slow fermenttion ws oserved in the three ensiled temperte grsses, ut perennil ryegrss silge showed lower ph nd higher lctic cid content thn silges of the other two grsses. A short eroic stility ws oserved in silges of the three temperte grsses, cocksfoot ieng the most unstle to eroic conditions. 9

10 C Timothy grss Perennil ryegrss Corkfoots RT RT+3 Figure 7. Effect of hour of eroic exposure on temperture of timothy grss, pereenil ryegrss nd corkfoots silge Literture Cited AOAC Officil methods of nlysis of the Assocition of Officil Anlyticl Chemists. 15th edition. Arlington, VA. Downing, T.W., A. Buyserie, M. Gmroth, nd P. French Effect of wter solule crohydrtes on fermenttion chrcteristcis of ensiled perennil ryegrss. The Professionl Animl Science 24: Miller, L.A., J.M. Moory, D.R. Dvies, M.O. Humphreys, S.D. Scolln, J.C. McRe, nd M.K. Theodorou Incresed concentrtion of wter-solue crohydrte in perennil ryegrss (Lolium perenene L.): milk production from lte- lcttion diry cows. Grss nd Forge Science 56: Smd, Y, T. Tki, nd T. Ymd Genetic vrition in wter solule cohydrte in diverse cultivrs of Dctylis glomert L. during vegettive growth. Austrlin Journl of Agriculturl Reserch 55(11): SAS Inst., SAS/STAT User s Guide (Relese 6.12). SAS Inst., Inc., Cry, NC. Smith, D The nonstructurl crohydrtes. In: Chemistry nd Biochemistry of Herge. Vol. 1 G.W. Butler nd R.W. Biley, ed. Acdemic Press, NY. Sveinsson, T nd L. Bjrndottir Timothy productivity nd forge qulity; possiilities nd limittions. NJF Seminr 384. Akureyri, Icelnd. Vn Soest, P. J., J. B. Roertson nd B. A. Lewis Methods for dietry fier, neutrl detergent fier, nd non-strch polysccrides in reltion to niml nutrition. J. Diry Sci. 74: