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1 -Original Paper 토양으로부터분리한토착유효미생물을이용한음식물쓰레기의자원화 이상우 함선녀 신택수 김혜경 연익준 * 김광렬 * ( , ) Resource of Food Waste using Indigenous Bacteria Isolated from Soils Sang-Woo Lee Sun Nyeoo Ham Taek-Soo Shin Hye-Kyung Kim Ik-Jun Yeon* Kawng-Yul Kim Department of Environmental Engineering, Chungbuk National University *Department of Environmental Engineering, Chungju National University ABSTRACT : This study was conducted to investigate feasibility of feedstuff for animal using food waste by fermentation mechanism of indigenous microorganism. To achieve this purpose, indigenous bacteria was isolated from soils to use as an inoculant. Enzyme test was performed to verify activity of amylase, protease and lipase using isolated bacteria. Bacteria(H1, D1), which vigorously express the enzyme activity, was selected and used in the fermentation experiments of food waste. From the analysis of 16s rdna sequencing, H1 and D1 were identified as Bacillus subtilis and Paenibacillus polymyxa, respectively. In the fermentation experiment, food waste was mixed with rice bran and popped rice to control moisture and nutrient content. Isolated bacteria(b. subtilis and P. polymyxa) was used as an inoculant. From the measured data such as temperature, ph and ORP, it can be verified that food waste adding the indigenous bacteria was effectively fermented. From the nutritional analysis of manufactured feedstuff, it showed that the contents of crude protein, crude fat and crude fiber were enough to use as feedstuff for animal. In addition, harmful components such as Pb, Hg, Cd, aflatoxin and salmonella concentration were not exceeded permitted standards. Therefore, fermented food waste using indigenous bacteria can be used as feedstuff. Key Words : Food Waste, Fermentation, Indigenous Bacteria, Feedstuff 요약 :. anylase, protease, lipase. 16s rdna sequencing (H1, D1) Bacillus subtilis Paenibacillus polymyxa. 2 (Bacillus subtilis, Paenibacillus polymyxa)., ph ORP.,,. Pb, Hg, Cd, aflatoxin salmonella.. 주제어 :,,, 1. 서론 * ) ,397 ton/day ,372 ton/day. Corresponding author kykim@chungbuk.ac.kr Tel: Fax: , 1997 ( ), ,3),. 2. 4)

2 36 이상우 함선녀 신택수 김혜경 연익준 김광렬,. 5),.,,,. 6),,,, 80%. 7), 재료및방법 2.1. 발효사료화를위한실험재료 ( ) 60 80% -10 mesh., (rice bran) (popped rice) Table 1. Nutrient compositions of food waste and additives Moisture protein fat fiber ash ph NaCl (%) Food waste % Rice bran Popped rice ) Table 토착유효미생물의분리및동정 ( ). 0.9% NaCl 100 ml 5 g 30 1 ml starch agar medium, skim milk agar medium, sierra medium, 30 3 colony,,. 9) 2, universal primer SEL S rrna sense (5'-ATC ATG GCT CAT CAG ATT GAA CGC-3') SEL S rrna antisense(5'-t ACC TTG TTA CGA CTT CTA CCT-3') PCR 16S rdna NCBI GeneBank network service BLAST program. 10) 2.3. 실험장치 5 mm, 250 mm, 450 mm. 450 mm, 500 mm packing. ph, ORP (Oxdiation Reduction Potential) (Mutichannel system, Istek Co.) 음식물사료화를위한반응기운전조건 16 kg, 9 kg, 5 kg 30±5%. J. of KSEE / Vol. 31, No. 1, January, 2009

3 토양으로부터분리한토착유효미생물을이용한음식물쓰레기의자원화 37 Table 2. Experimental conditions of fermentation for food waste Food waste Rice bran Popped rice Moisture Inoculated Material (kg) powder (kg) (kg) (%) microbe F ±5 sterilized F ±5 H1 F ±5 H1, D1 dry oven autoclave (H1, D1) 2, 14. Table 발효사료의영양분및유해물질분석 (crude protein), (crude fat), (crude fiber) Weende Henneberg Stohmann Weende, Fig. 1. 8). Pb, Hg, Cd 3 ICP (Inductively Coupled Plasma Lab8440, Labtam, Co.) B1 ( ). Fig. 1. Feedstuff analysis method. 3. 결과및고찰 3.1. 효소활성검색을통한토착유효미생물분리및동정 (starch agar medium, skim milk agar medium, sierra medium) (Table 3) Amylase 활성검색 Amylase starch agar medium 1% potassium iodide iodine solution colony clear zone, clear zone 0.5 cm + (Table 3) amylase D1 H1 0.7 cm, 0.8 cm clear zone. 11 H1 amylase Protease 활성검색 Protease skim milk agar medium colony clear zone protease. 11) Protease skim milk agar medium clear zone 0.5 cm + Table 3 activity Table 3. Enzyme activity of isolated microorganisms from soils Strain no. Starch agar medium Clear zone Activity (cm) Skim milk agar medium Clear zone Activity (cm) Sierra medium Clear zone Activity (cm) 1 D D D D D Y Y R H H H

4 38 이상우 함선녀 신택수 김혜경 연익준 김광렬 (D1, D3, D5, Y1, R1, H1, H2, H3) clear zone protease, amylase D1 H1 protease. 2 (H1, D1) 2, ph, (ORP) Lipase 활성검색 Lipase sierra medium colony lipase. 11) Lipase sierra medium clear zone 0.5 cm +. Table (D1, D3, D4, Y2, H1) Lipase, D3 H 유효토착미생물동정 Amylase, Protease, Lipase 2 (H1, D1) 16s rdna sequencing Bacillus subtilis(h1), Paenibacillus polymyxa(d1). B. subtilis amylase, cellulase, protease.,,, biotin, vitamin K2,,. 12,13). 14) P. polymyxa, polymyxin B. 15,16) 17) P. polymyxa,, , ph 3 11, 1 Salmonella typhi, Shigella dysentri, EHEC( ), Vibrio, Cholera Salmonella Typhimurium, ETEC( ), Yersinia Enterocolitica, Yersinia Pestis 온도변화 14 (Fig. 2) (F2, F3) B. subtilis P. polymyxa F3 B. subtilis F F1 14 F ), ph 변화,,. amylase, protease, lipase (,,, ),,, (acetate, propionate, butyrate, hydrogen ). 19) 3.3. 분리된토착유효미생물에의한발효사료반응 Fig. 2. Variation of temperature during fermentation. J. of KSEE / Vol. 31, No. 1, January, 2009

5 토양으로부터분리한토착유효미생물을이용한음식물쓰레기의자원화 39 Fig. 3. Variation of ph during fermentation. ph ph. 20) Fig ph, ph. F1 ph ph F2 F3 ph 6.6, 14 F2 F3 ph 산화환원전위 (ORP) 변화 (Oxidation Reduction Potential, ORP). 21) 14 ORP (Fig. 4). F1 74 mv mv. B. subtilis F2 80 mv 4 33 mv 10 (-) mv. F2 B. subtilis 3 Fig. 4. Variation of ORP during fermentation. ORP. B. subtilis P. polymyxa F3 F2 F2 ORP. F3 7 (-) ORP -170 mv. F3 B. subtilis F2 7 P. polymyxa F2 ORP. F2, F3 F1 ORP ORP 제조된발효사료의분석 영양분분석결과,,. 1) F1 2 F2 F3 14 (Table 4) % 10%, 22) 2%. 23),, 4 8%. 22) F1, 19.6%, 16.2% 10.6%

6 40 이상우 함선녀 신택수 김혜경 연익준 김광렬 Table 4. Nutrients compositions of fermented feedstuff Material protein fat fiber F F F Table 5. Analysis of hazardous and toxic components Pb (ppm) Hg (ppm) Cd (ppm) Aflatoxin B1 (ppb) Salmonella D Permitted level < 20 < 0.5 < 2.5 < 50 none Food waste none F none F none. F2 F3 13.7% 15.1%, 11.0% 12.0% 6.9% 8.1% F2 F 유해물질분석결과 Pb, Hg, Cd,, Table 5. Pb Cd 20 ppm 2.5 ppm ppm ppm. Hg 1/ ppb ppb. 4. 결론. 1) 2 16S rdna sequencing Bacillus subtilis, Paenibacillus polymyxa,,. 2) %, %, %. 3) Pb, Hg, Cd.,. 사사 참고문헌 1.,,,, 7(1), (2004). 2.,, (NaCl),, pp (2004). 3.,,,, 7(3), (2001). 4.,,, pp (1998). 5.,,,,,,,, pp (2006). 6. Westendorf, M. L., Zirkel, E. W., and Behnke, K. C., Feeding food or table waste to Livestock, Prof. Anim. Sci., 12, (1996). 7.,,,,,, 19(1), 44 50(2000). 8., (2001). 9. Ronald, M., Handbook of Microbiological Media, CRC Press(2000). 10. Chang, J.-S., Yoon, I.-H., and Kim, K.-W., Isolation and ars Detoxification of Arsenite-Oxidizing Bacteria from Abandoned Arsenic- Contaminated Mines, J. Microbiol. Biotechnol., 17(5), (2007). 11.,, (2001). 12., Penicillin Bacillus subtilis, Korean J. Appl. Microbiol. Bioeng., 1(1), 19 J. of KSEE / Vol. 31, No. 1, January, 2009

7 토양으로부터분리한토착유효미생물을이용한음식물쓰레기의자원화 41 20(1973). 13.,, (2003). 14.,,,,,,, (1993). 15. Paulus, H. and Gray, E., The biosynthesis of polymyxin B by growing cultures of B. polymyxa, J. Biol. Chem., 239, (1964). 16. Kajimura, Y. and Kaneda, M., Fusaricidins B, C and D, new depsipeptide antibiotics produced by Bacillus polymyxa KT-8 isolation, structure elucidation and biological activity, Japan Journal of Antibiotics, 50, (1997). 17., Paenibacillus polymyxa, (2005). 18.,,,,, 28(5), 22 27(2002). 19. Stronach, S. M., Rudd, T., and Lester, J. N., Anaerobic fermentation process in industrial wastewater treatment, Springer-Verlag, Berlin Heidelberg, New York Tokyo(1989) 20.,,,, 16(8), (1994). 21.,, (2002). 22.,, (2002). 23.,,,,,,, 7(2), 65 71(1999).

THE JOURNAL OF KOREAN INSTITUTE OF ELECTROMAGNETIC ENGINEERING AND SCIENCE. vol. 29, no. 10, Oct ,,. 0.5 %.., cm mm FR4 (ε r =4.4)

THE JOURNAL OF KOREAN INSTITUTE OF ELECTROMAGNETIC ENGINEERING AND SCIENCE. 2018 Oct.; 29(10), 799 804. http://dx.doi.org/10.5515/kjkiees.2018.29.10.799 ISSN 1226-3133 (Print) ISSN 2288-226X (Online) Method