isolation of saccharomyces spp from manure of beef bali ......isolation of saccharomyces spp from...

Official Reprint PDF Isolation of Saccharomyces Spp

from Manure of Beef Bali Cattle as a Probiotics Properties and has CMC-

Quality of Rice Bran By

Desak Putu Mas Ari Candrawati, D.A. Warmadewi, and Igng Bidura

ISSN 0970-4973 (Print) ISSN 2319-3077 (Online/Electronic) J. Biol. Chem. Research Volume 31 (1) 2014 Pages No. 39-52

Journal of Biological and Chemical Research (An International Journal of Life Sciences and Chemistry) Indexed and Abstracted: Index Copernicus International (Europe), Chemical Abstract Services (U.S.A.), Research Bible (Japan), Directory of Research Journals Indexing (DRJI), Indian Science.in, Database Electronic Journals Library (Germany), Open J-Gate, Google Scholar, J Gate e-Journal Portal, Info Base Index, International Impact Factor Services (Singapore) and Eye Source Published by Society for Advancement of Sciences®

ase Activity to Improve Nutrient

J. Biol. Chem. Research. Vol. 31, No. 1: 39-52 (2014) (An International Journal of Life Sciences and Chemistry) Ms 30/2/160/2013, All rights reserved ISSN 0970-4973 (Print) ISSN 2319-3077 (Online/Electronic) Desak Putu Mas Ari Candrawati Igng Bidura

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RESEARCH PAPER Received: 09/11/2013 Revised: 20/12/2013 Accepted: 03/01/2014

Isolation of Saccharomyces Spp from Manure of Beef Bali Cattle as a Probiotics Properties and

of Rice Bran Desak Putu Mas Ari Candrawati, D.A. Warmadewi, and Igng Bidura

Faculty of Animal husbandry, Udayana University, Denpasar-Bali, Jl. PB. Soedirman, Denpasar, Indonesia

ABSTRACT This experiment was carried out to study both on the CMC-ase activity and probiotics agent of Saccharomyces spp isolation from manure of beef Bali cattle samples can be used in order to alleviate the negative effect of rice bran as feed. Eighteen of Lohman Brown laying hens was assigned to three treatments in a completely randomized design. Each treatment has six replications with one bird per replication (individual cage). The treatments were (i) unfermented rice bran as control; (ii) fermented rice bran by Saccharomyces spp.S-6 isolate; (iii) fermented rice bran by Saccharomyces spp.S-7 isolatse, respectively. The report on the first experiment showed that five isolates of Saccharomyces spp (Sc.S-5; S-6; S-7; S-8; and S-9) were isolated from manure of beef Bali cattle samples in the first experiment. The whole isolates of Saccharomyces spp showed resistant grew on both in different temperature (100-450C), acid conditions (2,5-4,5), and bile salt. But only two isolates (Sc.S-6 and S-7) were potensial as probiotics sources and has CMC-ase activity. The study showed that fermentation of rice bran using both of Saccharomyces spp.S-6 and S-7 culture could improve significant differences (P<0.05) on digestibility of its dry matter (DM), organic matter (OM), crude protein (CP), crude fibre (CF), and increased its metabolizable energy of rice bran. The content of dry matter, organic matter, and gross energy of rice bran were no effected significantly different (P>0.05) by fermentation. On the other hand, fermentation caused increasing crude protein (CP) and crude fibre (CF) of the rice bran. It was concluded that two isolates (Sc.S-6 and S-7) were isolated from manure of beef Bali cattle samples, both were the potensial as a probiotics sources and has CMC-ase activity and its utilization in the rice bran fermentation could increase nutrient composition and digestibility of rice bran. Key words: Saccharomyces Spp., Probiotics, Crude Fiber, Digestibility, and Rice Bran.

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has CMC-ase Activity to Improve Nutrient Quality

Isolation of……………………………….Rice Bran Candrawati, et al., 2014

INTRODUCTION During recent years, numerous studies have been undertaken to obtain scientific evidences for beneficial effect of Saccharomyces spp as promising probiotic. Such beneficial effects are considered to include the protection from pathogen, enhancement of the immune system, antimutagenic and anticarinogenic effects and the reduction of serum cholesterol. Chen et al. (2005) reported that dietary supplementation of complex probiotic slightly improved digestibility of nutrients. The use of Saccharomyces cerivisiae (Sc.) culture as a probiotics sourches in poultry production as become an area of great interest, because continued use of probiotics in animal feeds may result in the presence of antibiotics residues in animal products (Han et al., 2001). Gut microfloral enzymes are beneficial to the nutrition of the host because they increase the digestion of nutrients, especially in the lower intestine. Previous experiments showed that the inclusion of microorganisms in the diets improved feed conversion efficiency and digestibility (Chen et al., 2005). Piao et al. (l999) showed that 0, 10% yeast added to a diet could reduce animal wastes. Yeast culture improved posphorus utilization in growing chickens. Inconsistent reports about the effect of probiotics may be due to several aspects such as strains of bacteria, dose level, diet composition, feeding strategy, feed form, and interaction with other dietary feed additives (Chesson, 1994). Rice bran is a by-product of rice milling industry, which is abundantly available (approximately 10% of paddy weight) during the harvesting season. Unfortunately this by-product contains toxic factors such as trypsin inhibitor, phytic acid as phytate, and high content of crude fiber. These anti-nutritive factors have been reported by Kahlique et al. (2003) causing reduction of feed intake and depress performance of poultry. These toxic factors are phytic acid as phytate and crude fiber (CF). These anti-nutritive factors have been reported to reduce feed intake and depress performance of poultry. Bach Knudsen (2001) reported that CF has been defined as the complex macromoleculer substances in food plants that are not degraded by mammalian digestive enzymes. With the exception of lignin, all of the materials called CF are carbohydrates in nature. CF is thought to mediate protective effects on the colonic epithelium through their fermentation products and fecal bulking capacity (Wang et al., 2004). Feeding high fiber resulted in a lowered rate of lipogenesis and tendency of an increased capacity to utilize acetyl-CoA in pigs (Zhu et al., 2003). Non starch polysaccharides (NSP) are the carbohydrate components of CF and are the predominant substrates for anaerobic fermentation. The potency of rice bran as energy source for poultry depends on its cell wall content, degree of microbial fermentation in poultry large intestine, absorption and production of the volatile fatty acid (Wang et al., 2004). The potensial of rice bran as energy sources for poultry depends considerably on such factors as cell wall content, degree of microbial fermentation in the large intestine, and extent of absorption and utilization of the volatile acids produced (Kahlique et al., 2003). The use of rice bran in poultry diets has been highly limited by the presence of phytic acid and other minor anti-nutritional factors like tannin and non-starch polysaccharides (NSP). The high levels of NSP in rice bran are limiting its unrestricted use in poultry feeding. J. Biol. Chem. Research. 40 Vol. 31, No. 1: 39-52 (2014)


These NSPs are known to increase the gut viscosity, reduce nutrient absorption in the intestine and affected indirectly the growth and performance of birds (Rhames et al. 2006; Cao et al., 2003). Many studies have clearly demonstrated that, supplemented of probiotics to diets rich in NSP results in a significant reduction in the intestinal viscosity, enhances energy, and protein utilization (Bidura et al., 2012; Bidura et al., 2009). Probiotics are defined as the viable microorganisms that exhibit a beneficial effect on health of the host by improving its intestinal microbial balance. Yeast culture is one of the most extensively studied probiotic. Traditionally, S.cerevesiae have been used for food products such as baking industries, “tape”, and are considered as organisms that can be used also for a pharmaceutical (Ahmad, 2005). The use of antibiotics as routine feed additives has been baned in same countries because of public concern over possible antibiotics residual effects and the development of drug-resistent bacteria. Probiotics have been introduced as an alternative to antibiotics; however, their effects on poultry production are not consistent, resulting in uncertainties and skepticism for development of the products. There are many types of probiotic preparations in the market. Many studies have been conducted to test the afficacy of such preparations on animal growth and performance. Several studies with broiler have indicated that probiotics preparations improve live weight gain and feed conversion rate (Bidura et al., 2012; 2009; 2008; Sutarpa et al., 2011). Hong et al. (2004) reported that fermentation of feed using Aspergilus oryzae increased digestibility of its DM and CP. Therefore, it is susgested that fermented of rice bran feeding by Saccharomyces spp isolates as a CMC-ase akctivity (isolation from manure of beef Bali cattle) can be used in order to alleviate the negative effect of rice bran as feed. METERIAL AND METHODS Animals and experimental design Eighteen of Lohman Brown laying hens was assigned to three treatments in a completely randomized design. Each treatment has six replications with one bird per replication (individual cage). All of the birds were fed experimental diets for two days. The treatments were (i) unfermented rice bran as control; (ii) fermented rice bran by Saccharomyces spp.S-6 isolate; (iii) fermented rice bran by Saccharomyces spp.S-7 isolate; respectively. The objectives of this study are to determine the nutrient digestibility and the ME value of rice bran using Lohman Brownn laying hens at 42 weeks of age. Probiotics properties Acid tolerance: to determine the trancsit tolerance to low pH, the method of Corzo and Gilliland (1999) was used with slight modifications. Strain was grown in MRS broth at 370C for 24 h. A 0.5 ml aliquot of the bacterial culture was inoculated in 10 ml of phosphate buffered saline adjusted to pH 2 with 4 N HCl. Phosphate buffer was prepared by dissolving NaCl (9 g/l), Na2HPO42H2O (9 g/l), and KH2PO4 (1.5 g/l) in destiled water. The initial bacterial concentration was 106-8 cfu/ml. Culture was incubated at 370C. After 0; 0.5; 1; 2; and 4 h incubation, cell were serially diluted tenfold in 0.1 M sodium phosphate buffer (pH 7.2), and the viable cells were enumerated on MRS agar plate at 370C for 2 days. J. Biol. Chem. Research. 41 Vol. 31, No. 1: 39-52 (2014)


Bile resistance: The resistance to bile was examined according to method of Hyronimus et al. (2000). Each strain was inoculated in MRS broth with 0.5 or 1% (w/v) bile salt (Difco). Cultures were incubated at 370C for 24 h, and the numbers of viable cell were determined and compared to a control (without bile salt) on MRS agar plates. CMC-ase activity: Weighed as much as 11 g of OMEA and 3 g of CMC-ase, then dissolved in distilled water. Heat in a waterbath and after it did strelilisasi on autoklav. Refrigerate at a temperature of 45-50 0C, then poured on Petri dishes and left to solidify. Isolates that have been cultured in a nutrient broth for 24 hours. Take a paper disk with tweezers and then dipped in a solution of nutrient broth then put on a petri dish containing Omea media and CMC-ase. Leave for 24 hours. After 24 hours of clear zone width measurements are generated by using a vernier caliper. The size of the clear zone and the apparent absence of clear zone, an indicator of the ability of the microbes to break down cellulose, as well as fast and slow arise the clear zone (VanDevoorde and Verstraete, 1987; Kanti, 2007). Fermented of Rice Bran The isolate of Saccharomyces spp.S-6 and S-7 which has been approved from bile salt and poultry digestive tract in vitro test could assimilate cholesterol for probiotics agency and two isolates have CMC-ase activity. The study was carried out at the Bioscience Laboratory of Udayana University, Bali, Indonesia. Fermentation of commercial rice bran was prepared as follows. Comercial rice bran was used. Approximately 0.20% (2 x 106 spores) Saccharomyces spp. S-6 and S-7 isolate culture was added to 100 g of steamed rice bran. Then, water was added to bring the product to 50% content and left up to 2 days for fermentation. After that, fermented rice bran was dried at 450C for six hours and then it was ground for analysis. Unfermented rice bran was also ground for its chemical analysis. Retention and excretion of nutrients In order to determine the nutrient digestibility and metabolizable energy (ME) value of the rice bran. The amount of rice bran used was 50 g. This amount as based on preliminary assays with Lohman brown laying hens using rice bran. All the birds were deprived of feed for 24 h to ensure that their alimentary canals were empty from feed residues. They were then force-fed with the specific amount of rice bran (fermented and unfermented). Stainless steel funnel with 40 cm stem was used in force feeding technique (Mustafa et al., 2004). Water was available ad libitum during the experimental period. The total excreta were collected in plastic trays. The excreta samples were frozen, allowed coming to equilibrium with the atmospheric moisture, weighed, and groun through a 1 mm sieve. Samples of excreta and rice bran were subjected to appropriate analysis to determine DM, OM, CP, CF, and energy, respectively. Laboratory analyses Dry matter (DM), organic matter (OM), CP and ash determinations were done according to the Assocciation of Official Analytical Chemists (l994). The CP content of the diets was determined using the Kjeldahl procedure (AOAC, 1994). Crude fibre in the feeds was determined using the procedure of Van Soest et al. (l991) on oven-dried samples. Gross energy (GE) was measured with an adiabatic oxygen bomb calorimeter (Parr, USA), J. Biol. Chem. Research. 42 Vol. 31, No. 1: 39-52 (2014)


Calculations The data were used to calculate AME value according to the following formulate (Mustafa et al., 2004): AME (apparent metabolizable energy) = IE – FE. Where IE = ingested energy; FE = fecal energy voided by the fed birds. Statistical Analysis All data were subjected to a one-way analysis of variance test (Steel and Torrie, 1989). Statistical significances among treatment means were determined by method of New Multiple Range Test of Duncan when the F value was significant at 5% level RESULTS Resilience isolate Saccharomyces spp. the low pH is one of the characteristics required to be met by a candidate or a probiotic that could be developed into a potential probiotic. On this test, the medium pH was adjusted to 1.5, 3.0, 4.5, and 6.0 by using HCl (hydrochloric acid), because HCl has characteristics similar to stomach acid. Resistant to highly acidic nature needs to be owned by the candidate probiotic, because the application later, this probiotic candidate must pass very acidic stomach conditions, before reaching the colon. Observations as presented in Table 1. In this study, as many as 10 isolates Saccharomyces spp. isolated from cattle feces was tested resistance to various pH, at pH 1.5, 3.0, 4.5, and 6. Two isolates were unable to grow at pH 1.5, which isolates the S-1 and S-2. Saccharomyces spp.S-1 isolates even simply could not grow at all pH treatments given. Only 8 isolates were able to grow well, namely Saccharomyces spp.S-4 to S-10 isolates. There is a tendency of the higher pH, especially at pH 6, most of the isolates has decreased the number of colony life. Saccharomyces spp colonies grow well at pH 3, even isolate 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 Saccharomyces spp (Log colony/g) at various pH. Code Isolate pH

1.5 3 4.5 6 S1 - - - - S2 - 3.98 3.88 3.32 S3 3.45 5.53 4.41 4.13 S4 4.33 3.65 3.58 4.56 S5 4.12 5.09 4.99 4.12 S6 4.19 4.35 4.36 4.40 S7 4.29 4.42 3.99 3.58 S8 4.06 3.617 4.318 3.86 S9 4.31 4.41 3.95 3.92 S10 4.16 4.33 3.97 3.87 Descriptions: S-1 s / d S-10 is a Saccharomyces spp isolates were isolated from manure of beef Bali cattle J. Biol. Chem. Research. 43 Vol. 31, No. 1: 39-52 (2014)


In the test of resistance to bile salts as seen in Table 2 shows, that the ten isolates were able to survive and grow on a medium containing bile salts at concentrations of 0.2 mM and 0.4 mM, whereas at a concentration of 0.6 mM no isolates can survive. Microbial isolates resistance to bile salts is used to assess the ability to survive the digestive tract isolates contained bile salts on the surface of the intestine. Probiotics will be dealing with the environment in the small intestine, in which there are bile or bile salts are removed by the liver through the gall bladder, after successfully passing the acidic conditions in the stomach. Therefore, in the process of developing new probiotic, or a new probiotic candidate must be able to pass the test of resistance to bile or bile salts were performed in vitro. Based on the nature of the resistance shown by some isolates, strai-indicates that the strain has the potential to be developed into a potential probiotic.

Table 2. Ability of yeast Saccharomyces spp isolates were resistant living in bile salts. Isolate Bile salt concentration (absorbance)

0.2 mM 0.4 mM 0.6 mM S1 + (0.244) + (0.169) - (0.067) S2 + (0.248) + (0.168) - (0.059) S3 + (0.253) + (0.163) - (0.081) S4 + (0.252) + (0.158) - (0.064) S5 + (0.243) + (0.166) - (0.073) S6 + (0.224) + (0.144) - (0.036) S7 + (0.192) + (0.133) - (0.031) S8 + (0.145) + (0.151) - (0.051) S9 + (0.126) + (0.132) - (0.044) S10 + (0.138) + (0.149) - (0.035)

Description:

- : A <0.1 (not bear bile salt) + : A 0.1 - 0.5 (a bile salt-resistant) + + : A 0.5 - 1.0 (bile salt-resistant) + + + : A> 1.0 (highly resistant of bile salts) S-1 s / d S-10 is a Saccharomyces spp isolates were isolated from manure of beef Bali cattle

CMC-ase test is to test the ability of the yeast Saccharomyces spp isolates in degrading crude fiber. It can be measured from the resulting clear zone diameter (Table 3). From the results of this study were only yeast Saccharomyces spp isolates S-5, S-6, S-7, S-8, and the yeast Saccharomyces spp.S-9 who has the ability to digest fiber. Looks yeast Saccharomyces spp isolates S-6 has the most wide clear zone, while the S-8 has a clear zone at least. J. Biol. Chem. Research. 44 Vol. 31, No. 1: 39-52 (2014)


This means that isolates S-6 has kemampuam in digesting crude fiber than the highest S-8. More detailed, clear zone diameter and difference generated by the two isolates are presented in Figure 1. Table 4 shows the nutrient the content of crude protein and gross energy were slightly increased by fermentation. On the other hand fermentation caused decreasing dry matter (DM), organic Matter (OM), and crude fibre (CF) of the rice bran. These results indicated that carbohydrate was used for microbial growth and the reduction of dry matter. The result indicated that all of nutrient digestibility of fermented rice bran by Saccharomyces spp culture were increased significantly (P<0,05) different rather than unfermented rice bran.

Figure 1. Saccharomyces spp.S-8 isolates zone is smaller than the isolate Saccharomyces spp.S-6 (right). Table 3. CMC-ase activity test yeast Saccharomyces spp isolates based on the diameter

of the clear zone caused. Isolat Clear zone diameter (cm) S-1 - S-2 - S-3 - S-4 - S-5 3.60 S-6 4.51 S-7 4.00 S-8 1.85 S-9 3.80 S-10 -

Description: S-1 s / d S-10 is a Saccharomyces spp isolates were isolated from manure of beef Bali cattle. J. Biol. Chem. Research. 45 Vol. 31, No. 1: 39-52 (2014)

Isolate S8 Isolate S6

Clear zone


Table 4 shows the nutrient digestibility and metabolizable energy of unfermented rice bran (UFR) and fermented rice bran (FR) ingredient. The digestibility of crude protein, organic matter, and crude fiber, were slightly increased significantly different (P<0,05) by the fermentation, both by Saccharomyces spp.S-6 and S-7, respectively. The metabolizable energy of FR was increased significantly different (P<005) than metabolizable energy of UFR. Fermented of feeding ingredient were caused increased of crude protein, organic matter, and crude fibre digestibilities, resvectively than UFR ingredients. Chemical composition and nutrient digestibility of rice bran (fermented compared to the unfermented) were shown in Table 4 as below: Table 4. Chemical composition and nutrient digestibility of unfermented and fermented rice bran by Saccharomyces Spp.S-6 and S-7 isolates culture (in % Dry Matter). Rice Bran Parameters Unfer-

mented (UFR)

Fermented by S-6 Isolate (FR)

Fermented by S-7 Isolate

SEM

Chemical composition: Dry Matter (%) 89.75a 89.64a 89.62a 0.453 Organic Matter (%) 90.93a 90.87a 90.22a 0.318 Crude Protein (%) 11.88b 14.69a 14.78a 0.149 Crude Fibre (%) 15.96b 17.65a 18.19a 0.285 Gross Energy (Kcal/kg) 3258.31a 3272.22a 3274.97a 44.528 Nutrient digestibility (%):

Dry matter digestibility (%) 25.48c 28.24b 29.81a 0.185

Organic Matter digestibility (%) 26.16b 30.07a 30.25a 0.314

Crude Protein digestibility (%) 37.38b 51.77a 53.23a 1.106

Crude fibre digestibility (%) 15.92c 24.10b 26.29a 0.252

Metabolizable energy (Kcal/kg) 1696.38b 1898.44a 1940.69a 19.512

Note: UFR = Unfermented rice bran; FR = Fermented rice brang by 0.20% yeast culture The different superscript at the same row is significantly different (P<0.05)

DISCUSSION The results showed that only 5 isolates of Saccharomyces spp potential as probiotic agents and have the CMC-ase activity. To five isolates showed resistance to acid and bile salts that is characteristic of probiotic bacteria. Saccharomyces is a unicellular microorganisms lacking chlorophyll, grew well at 30oC and pH 4.8. pH value in the gastrointestinal tract of poultry in each section, are: crop (4.5), proventriculus (4.4), J. Biol. Chem. Research. 46 Vol. 31, No. 1: 39-52 (2014)


gizzard (2.6), duodenum (5.7-6.0), jejunum (5.8), ileum (6.3), colon (6.3), and ceca (5.7) (Hidayat, 2010). Some advantages of Saccharomyces fermentation process, these microorganisms are rapidly proliferating, resistant to high alcohol content, resistant to high temperatures, has held steady and rapid nature of adaptation. According to Ahmad (2005), temperature optimum environment for yeast growth is 25-300C and a maximum temperature of 35-470C. Some advantages Saccharomyces microorganisms in the fermentation process that is rapidly proliferating, resistant to high alcohol content, resistant to high temperatures, has held steady and rapid nature of the adaptation. CMC-ase enzyme activity (endo-1,4-beta-glukonase) is a clear zone around the colony indicated that the microbes have a strong extracellular cellulase enzyme activity. The size of the clear zone and the apparent absence of clear zone, an indicator of the ability of the microbes to break down cellulose, as well as fast and slow arise the clear zone (Van Devoorde and Verstraete, 1987). Utama et al. (2006) reported that the yeast Saccharomyces cerevisiae is yeast capable of producing the enzymes amylase and selulolase, so as to increase the digestibility of protein and crude fiber such as cellulose and hemicellulose, as has been overhauled in the form of a simple monosaccharide. Cellulolytic yeasts capable of producing the enzyme 1,4 beta-endo-glukonase, 1,4 beta-exo glukonase, and beta-glucosidase that can degrade components of crude fiber into soluble carbohydrates (Howard et al., 2003) According Utama et al. (2006), fermentation feed purpose is to improve the usability of feed and elimination of anti- nutritional substances, as well as utilize biomass formed. Rice bran fermented with yeast will be able to increase the microbial biomass, so that the crude protein content of rice bran increased. Also reported that the fermentation process and the product is affected by the type and number of microbes, substrat types, pH, and temperature during the fermentation process. Biomass is a form of mass from the biological processes of microorganisms. Microorganisms capable of converting material into proteins. The fermentation process has the objective to produce a product (material feed) that have a nutrient content, texture, better biological availability, and reduce substance antinutrisi. Suparjo et al. (2003) stated that the fermentation of rice bran with 0.2% Aspergillus niger cultured for 72 hours can markedly increase protein and phosphorus content of rice bran, on the contrary lower crude fiber content and acid phytat rice bran. According Sumarsih et al. (2007), the longer the storage time of feed ingredients when fermented by Trichoderma viride, the more the loss of dry matter and organic matter material, instead increasing the crude protein content and materials decreased crude fiber content material. Also reported that the longer the fermentation process or materials storage time, the increased dry matter digestibility and organic matter materials. According to Jaelani et al. (2008), the increase in ME content of palm kernel cake (palm kernel cake/meal) as a result of fermentation by the fungus T. reesei of 1824.13 kcal/kg to 1930.44 kcal/kg suspected because of the degradation of mannan polysaccharides exist in palm kernel by fungus T. reesei into simpler forms (monosaccharides) that produces enough energy value better than in the form of polysaccharides mannan. J. Biol. Chem. Research. 47 Vol. 31, No. 1: 39-52 (2014)


The same thing was reported also by Sabini et al. (2000) which states that the fungus T. reesei is able to degrade a polysaccharide mannan mannotriosa, mannobiosa, and monnosa. According to Jaelani et al. (2008), fermented palm kernel cake can markedly increase the crude protein content compared to palm kernel cake without fermentation. Table 3 shows the chemical composition of unfermented rice bran (UFR) and fermented rice bran (FR) ingredient. The digestibility both of crude protein, organic matter, crude fibre, and metabolizable energy were slightly increased by S.cerevisiae fermentation. S. cerevisiae can not effect on crude fiber digestibility of rice bran. Becouse, among the cell wall polysaccharides of rice bran known as nonstarch polysaccharides (NSP) are celluloses, pectins, and oligosaccharides. NSPs can not be degraded enzymitically in the digestive systems of the birds due to the lacking of enzymes degrading the NSPs in their digestive systems (Choct, 2002). The digestibility of crude protein, crude fiber, and ME were slightly increased by the fermentation. These results indicated that carbohydrates other than fibres were used for microbial growth (Saccharomyces cerevisiae) and the reduction of nitrogen free extract resulted in increased concentration of the other components. Yi et al. (l996) reported that supplementation of microbial in diets improved N retention in broiler chickens and in vitro digestibility of vegetable protein. Also, Chen et al. (2005) reported that addition of 0.20% complex probiotic (L.acidophilus and S.cerivisae) in basal diets were inceased digestiblities of dry matter. Fermented of rice bran ingredient by ragi (Saccharomyces cereviseae) had better digestibilities, because Saccharomyces cereviseae in the gastrointestinal tract can part of an probiotics sourches. Also, Piao et al. (l999), suggested that probiotics in the gastro intestinal tract can improve protein and energy retention on the body of birds. These fungal are effective in degrading of the complex compounds such as beta-glucans and arabinoxylans (Bedford and Classen, 1992). Biofermentation of rice bran by Saccharomuces spp.S-6 and S-7 isolates culture had better digestibility, because Saccharomuces spp culture in the gastro intestinal tract can to be part of probiotic sources. Saccharomyces cerevisiae as part of probiotic could improve protein and energy retention in the gastro intestinal tract of the birds (Piao et al., l999). Cho et al. (2007) reported that supplementation of microbe in diet could improve the bioavailability of dietary. Wang et al. (2004) reported that the inclusion of fiber sources such as wheat bran or potato starch reduced the maintenance of energy requirement. These fungi are effective in degradating the complex compounds such as beta-glucans and arabinoxylans (Dubey, 2006). Chen et al. (2005) reported that dietary supplementation of complex probiotic slightly improved digestibility of nutrients. Inconsistent reports about the effect of probiotics may be due to several aspects such as strains of bacteria, dose level, diet composition, feeding strategy, feed form, and interaction with other dietary feed additives (Chesson, 1994). The high level of non-starch polysaccharides (NSP) in rice bran is limiting its unrestricted use in poultry feeding. The NSP is known increasing the gut viscosity, reduce nutrient absorption in the intestine and affect indirectly the growth and performance of bird (Rhames et al., 2006; Cho et al., 2007). J. Biol. Chem. Research. 48 Vol. 31, No. 1: 39-52 (2014)


Many studies have clearly demonstrated that the addition of probiotics culture or enzymes to diets rich in NSP resulted in a significant reduction of the intestinal viscosity, enhances energy and protein utilization. Wang et al. (2004) reported that degree of microbial fermentation in the large intestine improves the bioavailability of dietary. Yi et al. (l996) reported that supplementation of microbial in diets improved N retention in broiler chickens and in vitro digestibility of vegetable protein. Also, Chen et al. (2005) reported that addition of 0.20% complex probiotic (L. acidophilus and S. cerivisae) in basal diets increased digestiblity of DM and CP. Hong et al. (2004) reported that fermentation of feed using Aspergilus oryzae increased digestibility of its DM and CP. The inclusion of soluble dietary fiber (wheat bran) increased daily NSP excreted from feces (Wang et al., 2004; Suprapti et al., 2008), increased both of metabolizable energy and crude protein contents of palm kornel meal (Jaelani et al., 2008). Chen et al. (2005) reported that dietary supplementation of complex probiotic slightly improved digestibility of nutrients.

CONCLUSION It was concluded that there are two isolate (Sc.S-6 and S-7) were isolated from manure of beef Bali cattle samples, both were the potensial as a probiotics sources and has CMC-ase akctivity and its utilization in the rice bran fermentation could increase nutrient composition and digestibility of rice bran. ACKNOWLEDGEMENTS Authors are entirely grateful for the chemical analyses of sample by staff of Nutrition Laboratory, Fakulty of Animal Husbandry, Udayana University, Denpasar-Bali. The authors also want to thank the Ditbinlitabmas, Dirjen Dikti, Depdiknas in Jakarta for funding this research. A part of the present study was financially supported by Ditbinlitabmas, Dirjen Dikti, Depdiknas in Jakarta. REFERENCES Ahmad, R. Z. 2005. Used of Khamir Saccharomyces cerevisiae for feeding. Wartazoa Vol.

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Corresponding author: Dr. Igng Bidura, Faculty of Animal Husbandry, Udayana University, Denpasar-Bali, Jl. PB. Soedirman, Denpasar, Indonesia Email: [email protected] [email protected] Phone / Fax. 0361-702771 J. Biol. Chem. Research. 52 Vol. 31, No. 1: 39-52 (2014)

isolation of saccharomyces spp from manure of beef bali ......isolation of saccharomyces spp from...

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