Biol. Chem. Research. 42 Isolation of Saccharomyces Spp from Manure of Beef Bali Cattle as a Probiotics Properties and has CMC-ase Activity to Improve Nurient Quality of Rice Bran.

I solation of……………………………….Rice Bran Candraw ati, et al., 20 14 Bile resistance : The resist ance t o bile w as exam ined according to m et hod of Hyronim us et al . 2000. Each st rain w as inoculat ed in M RS brot h w ith 0.5 or 1 w v bile salt Difco. Cultures w ere incubat ed at 37 C for 24 h, and t he numbers of viable cell w ere det erm ined and compared t o a cont rol w it hout bile salt on M RS agar plat es. CM C-a se activity: W eighed as m uch as 11 g of OM EA and 3 g of CM C-ase, t hen dissolved in distilled w at er. Heat in a w at erbat h and aft er it did st relilisasi on autoklav. Refrigerat e at a t em perat ure of 45-50 C, t hen poured on Pet ri dishes and left t o solidify. Isolat es t hat have been cultured in a nut rient brot h for 24 hours. Take a paper disk wit h t w eezers and t hen dipped in a solution of nut rient broth t hen put on a pet ri dish cont aining Om ea media and CM C-ase. Leave for 24 hours. Aft er 24 hours of clear zone w idth measurem ent s are gen erat ed by using a vernier caliper. The size of t he clear zone and the apparent absence of clear zone, an indicator of the abilit y of the microbes t o break down cellulose, as w ell as fast and slow arise t he clear zone VanDevoorde and Verst raet e, 1987; Kanti, 2007. Fermented of Rice Bran The isolat e of Saccharomyces spp.S-6 and S-7 w hich has been approved from bile salt and poult ry digest ive t ract in vit ro t est could assim ilat e cholest erol for probiotics agency and t w o isolat es have CM C-ase act ivit y. The st udy w as carried out at t he Bioscience Laboratory of Udayana Universit y, Bali, Indonesia. Ferm ent at ion of com mercial rice bran w as prepared as follow s. Comercial rice bran w as used. Approxim at ely 0.20 2 x 10 6 spores Saccharomyces spp. S-6 and S-7 isolat e cult ure w as added to 100 g of st eam ed rice bran. Then, w ater w as added t o bring t he product t o 50 cont ent and left up t o 2 days for ferm ent at ion. Aft er t hat , ferment ed rice bran was dried at 45 C for six hours and t hen it was ground for analysis. Unferm ent ed rice bran w as also ground for it s chem ical analysis. Retention and excretion of nutrients In order t o det ermine t he nut rient digest ibilit y and met abolizable energy M E value of t he rice bran. The amount of rice bran used w as 50 g. This amount as based on preliminary assays w it h Lohman brow n laying hens using rice bran. All the birds w ere deprived of feed for 24 h t o ensure t hat their aliment ary canals w ere empt y from feed residues. They w er e t hen force-fed w it h the specific amount of rice bran ferment ed and unferment ed. St ainless st eel funnel w it h 40 cm st em was used in force f eeding t echnique M ust afa et al., 2004. Wat er w as available ad libit um during t he experiment al period. The t ot al excr et a w ere collect ed in plast ic t rays. The excret a samples w ere frozen, allow ed coming to equilibrium w it h the at mospheric moisture, w eighed, and groun through a 1 mm sieve. Samples of excret a and rice bran w ere subject ed t o appropriat e analysis t o det ermine DM , OM , CP, CF, and energy, respect ively. Laboratory analyses Dry mat t er DM , organic matt er OM , CP and ash det erm inat ions w ere done according t o the Assocciat ion of Official Analyt ical Chem ist s l994. The CP cont ent of t he diet s w as det erm ined using t he Kjeldahl procedure AOAC, 1994. Crude fibre in t he feeds w as det erm ined using t he procedure of Van Soest et al. l991 on oven-dried samples. Gross energy GE w as m easured wit h an adiabat ic oxygen bomb calorim et er Parr, USA,

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Vol. 3 1, No. 1: 3 9-52 20 14 I solation of……………………………….Rice Bran Candraw ati, et al., 20 14 Calculations The data w ere used t o calculat e AM E value according to t he follow ing form ulat e M ust afa et al., 2004: AM E apparent m et abolizable energy = IE – FE. Where IE = ingest ed energy; FE = fecal energy voided by the fed birds. St atistical Analysis All dat a w ere subject ed t o a one-w ay analysis of variance t est St eel and Torrie, 1989. St at ist ical significances among t reat m ent means w ere det erm ined by m et hod of New M ult iple Range Test of Duncan when t he F value w as significant at 5 level RESULTS Resilience isolat e Saccharomyces spp. t he low pH is one of t he charact erist ics required t o be met by a candidat e or a probiot ic that could be developed into a pot ential probiotic. On t his t est , t he m edium pH w as adjust ed t o 1.5, 3.0, 4.5, and 6.0 by using HCl hydrochloric acid, because HCl has charact erist ics similar t o stom ach acid. Resist ant to highly acidic nature needs t o be ow ned by t he candidat e probiotic, because t he applicat ion lat er, t his probiot ic candidat e must pass very acidic st omach conditions, before reaching t he colon. Observat ions as present ed in Table 1. In this st udy, as m any as 10 isolat es Saccharomyces spp . isolat ed from catt le feces w as t est ed resist ance t o various pH, at pH 1.5, 3.0, 4.5, and 6. Two isolat es w ere unable t o grow at pH 1.5, w hich isolates t he S-1 and S-2. Saccharomyces spp .S-1 isolat es even sim ply could not grow at all pH t reat m ents given. Only 8 isolat es w ere able t o grow w ell, nam ely Saccharomyces spp.S-4 t o S-10 isolat es. There is a t endency of t he higher pH, especially at pH 6, most of t he isolat es has decreased t he number of colony life. Saccharomyces spp colonies grow w ell at pH 3, even isolat e S-3 and S-5 Saccharomyces spp colony number reached 5.531 and 5.093 log colony per gram . Table 1. The number of colony isolates Sa ccharomyces spp Log colony g at various pH. Code Isolat e pH 1.5 3 4.5 6 S 1 - - - - S 2 - 3.98 3.88 3.32 S 3 3.45 5.53 4.41 4.13 S 4 4.33 3.65 3.58 4.56 S 5 4.12 5.09 4.99 4.12 S 6 4.19 4.35 4.36 4.40 S 7 4.29 4.42 3.99 3.58 S 8 4.06 3.617 4.318 3.86 S 9 4.31 4.41 3.95 3.92 S 10 4.16 4.33 3.97 3.87 Descript ions: S-1 s d S-10 is a Saccharomyces spp isolat es w ere isolat ed from manure of beef Bali cat tle

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