Báo cáo " Phân lập và khảo sát khả năng loại Hydrogen Sulfite (H2S) của vi khuẩn oxy hóa hợp chất Sulfur (SOB) trong nước kinh Nhiêu Lộc - Thị Nghè" docx

TAP CHl KHOA HOC VACONG NGHE Tap 47, s6 5,2009 Tr. 83-90 PHAN LAP VA KHAO SAT KHA NANG LOAI HYDROGEN SULFITE (H2S) CUA VI KHUAN OXY HOA HOP CHAT SULFUR (SOB) TRONG Nu'ac KINH NHIEU LOC - THj NGHE NGUYEN KHANH HOANG, NGUYEN HOAI PHONG, Hl/A THACH SON, NGUYEN VAN CLFONG I.MODAU Nuac la mpt thanh phan rat quan trpng doi vai su song tren trai dat, day la dieu tat yeu ma mpi nguai deu nhan thu'c dugc. Theo tac gia Timi Ecimovic (2005) be mat trai dat co dien tich 510,072 trieu km' trong do 70,7% dugc bao phii bai nuoc. Trong tong lugng nuac ctia qua dat chi CO 2,5% la nuoc nggt nhung phan lan (68,7%) nuoc nggt lai co mat trong cac niii bang. Vi the, lugng nuac ngpt con nguai co the de dang sii' dung dap trng nhu cau con lai rat it (0,4%) trong tong so nuoc ctia trai dat. Chat lugng ciia nuac co the str dung rat dang dugc mpi nguoi quan tam vi chting phai dap ung dugc nhu cau an,uong, tam, giat. Tuy nhien, cac boat dpng ctia con nguai va thien nhien hang ngay da thai vao nguon nuac rat nhieu hgp chat co the lam cho nguon nuoc tra thanh khong an toan cho nhu cau sir dung ciia con nguoi va he sinh thai trong moi trirong nuoc. Mpt trong nhung hgp chat thai ra do qua trinh boat dpng ctia con nguoi va thien nhien co anh huang rat Ion den chi tieu mau sac va miii ciia nuac chinh la Hydrogen sulfite (H^S). Trong moi truang nuac cac hgp chat sulfur ton tai duoi 3 dang tiiy thuoc vao pH ctia moi truang, khi moi truong co pH 6 thi 90% ton tai dang H2S va 10% a trang thai HS", nhung khi pH moi truong dat gia trj 10 thi sulfur ton tai 100% a trang thai S"". Hydrogen sulfite hoa tan trong nuo'c voi nong dp toi da 0,4 gam/ 100 ml a dieu kien nhiet dp 20°C, khi c6 mat trong nuac chting se gay miii hoi nhu miii trung thoi lam anh huong den gia trj cam quan. Thanh pho Ho Chi Minh vai mang luoi kinh rach va song co tong dien tich be mat bao phii la 141 km" gom 5 he thong kinh rach: Kinh Nhieu Lpc - Thj Nghe (NL-TN) bao phii 33 km"; Kinh Tau Hti - Doi - Te bao phii 37 km"; Kinh Tau Hu - Ben Nghe bao phii 12 km"; Kinh Tan Hoa - Ong Buong- Lo Gom bao phu 16 km"; Kinh Tham Luong - Ben Cat - Vam Thuat bao phu 44 km . He thong kinh rach ctia thanh pho Ho Chi Minh hien nay khong con kha nang tu lam sach khi phai tiep nhan gan nhu toan bp lugng nuoc thai sinh boat va nuac mua ctia thanh pho dong thai voi su lan chiem lam thu hep dien tich bao phii ctia he thong kinh rach dan den tinh trang 6 nhiem ngay cang tram trgng. Phan Ion he thong kinh rach trong pham vi npi thanh va khu vuc dau nguon deu bi 6 nhiem nang va dugc gpi la "Kinh den" hoac "Kinh thtii" bai qua trinh ehnyen hoa cac hgp chat sulfate do nhom vi khuan khir sulfate (SRB) tao thanh hydrogen sulfite igay mill thoi va khi ket hgp voi sat tao thanh FeS gay mau den. Vi the, vao dau nam 2001 voi sir 83 tai trg ctia ngan hang the gioi thanh pho da khai dgng du an ve sinh moi truang vai mue dich cai tao kinh NL-TN va cai tai he thong thoat nuoc ctia thanh pho. Tuy nhien, hien nay du an van con tiln hanh mac dii da dugc ngan hang the giai gia ban den quy I nam 2009. Voi mong muon gop phan vao viec cai thien moi truong thanh pho H6 Chi Minh cu thi gop ph§n cai thien chat lugng nuoc trong h^ thong kinh rach, nhom nghien cuu da tiln hanh phan lap vi khuSn co kha nang oxy boa hgp chat sulfur trong nuac kinh NL-TN sau khi da khao sat kha nang loai hydrogen sulfite co trong nuac voi quy mo phong thi nghiem se tien hanh nghien ciru CO djnh nham tao ra che pham sinh hgc xtr li H2S trong co trong nuac thai sinh boat. 2. NGUYEN LIEU VA PHlTOTNIG PHAP 2.1. Nguyen lieu va hoa chat - Mau nuoc thu nhan tir kinh Nhieu Lgc, Thj Nghe. - Moi truong: PCA, PGA, Peptone, Colistine; Thuoc nhupm Gram (Difco); Bp test djnh danh vi sinh vat: ID32E (bioMerieux). - Til cay (Juoan, France). - Noi hap (Hitachi, Japan). - May do quang UV-VIS (Thermo Electron Cooperation, USA). - Kinh hien vi dien ttr. - Mo hinh thu' nghiem kha nang phan giai sunfite ciia vi khuSn. 2.2. Phu-QTig phap 2.2.1. Phucmg phdp ldy mdu vd chudn bi mdu Hmh 1. Cau Cong Li (cau Nguyin Van Tr6i)- Nai liy miu {Source: www, wikimapia. org) Jl mau dugc lay tai kenh NL-TN a cau Nguyin Van TrSi co vj tri (10''47'29"N; 106 40'55"E), do sau 0,1-0,2 m. Nhiet do cua mau nuac a khoang 26°C. Mau dugc \ky a hai miia mua va miia kho, trong mtia mua (ttr thang 4 den thang 10) va miia kho (ttr thang 11 din thang 3) nam 2007 (bangl). Mau nuac dugc dting dl phan tich ham lugng sunfite va phan lap vi 84 sinh vat. Nuoc dugc lay tai kenh va cho vao erien da triet triing giii' a nhiet do moi truang mang ve phong thi nghiem. Mau dugc kiem tra trong thai gian 24 gid. Bdng I. Chi tiet va thong so ctia mau nuoc So thii' tu 1 2 3 4 5 6 7 8 9 10 11 2 ? ^ Phican Lo Nl N2 N3 N4 N5 N6 N7 N8 N9 NIO Nil SPhdi Thoi gian lay mau Thang 11 -thang 3 Thang 11 -thang 3 Thang 11 -thang 3 Thang 11 -thang 3 Thang 11 -thang 3 Thang 11 -thang 3 Thang 4 - thang 10 Thang 4 -thang 10 Thang 4 -thang 10 Thang 4 -thang 10 Thang 4 -thang 10 ; phdn lap vi sinh vgl.{V Mua Kho Kho Kho Kho Kho Kho Mua Mua Mua Mua Mua Nhiet do CO 26 25,5 26 26 26,5 26 24 24,5 25 25 25,5 larvey- Prescot, 2002): pH 6.5 6,7 6,5 6,5 6,8 6,5 7,1 7,0 7,1 7,0 7,0 Thuy trieu Len Xuong Xuong Xuong Len Xuong Len Xuong Len Xuong Xuong Ba loai moi truong diing de phan lap vi khuan (moi truang Thiosulfate, pH = 8,5; moi truang Beijerick, pH = 7,4; moi truang Thiobacillus thiooxidans, Ph = 4,4). Sir dung pipette da triet triing htit 5 ml mau vao ong nghiem chua san 20 ml dung djch moi truong da triet trting. U a 3TC va theo doi trong thai gian 14 ngay. Hang ngay tien hanh cay khao sat su phat trien ctia vi khuan tii' moi truong long sang moi truong ran. Moi truong ran dugc chuan bj bang each them 1,5% agar (15 g / 1000 ml). 2.2.3. Phu'ong phdp phdn lap vd ldm thudn de chpn vl khudn. Str dung ki thuat c4y phan lap de tao khuan lac rieng le tren moi truang phan lap tuong ling. Tang sinh vi khuan da phan lap tren moi truong tang sinh dang ran va dang long. Tien hanh khao sat tinh chSt sinh hoa, hinh dang, kich thuoc, tinh chat bat mau bang kT thuat thir nghiem sinh hoa, nhupm Gram. 2.2.4. Phicang phdp ldm tdng sinh vi khudn (Harvey- Prescot, 2002) CSy chuyin vi khuSn vao moi truang moi nham tang sinh khoi de phuc vu cong tac khao sat kha nang xu li hydrogen sulfite. Cac moi truong tang sinh gom thach nghieng, dTa Petri va moi truang long. 85 2.2.5. Phuong phdp khdo sdt hdm lucmg HjS (APHA, AWWA and WPCF, TCVN- 4667- 88) Nguyen tic: Phuong phap nay thti' nghiem kha nang lam giam H2S bai cac loai vi khuan oxy hoa hgp chat sunfur kit hgp vai dung dich iode. Tien hanh: - Chuan bj dung djch iode: 20 g KI, 500 ml nuac cit, 5 g iode, HCI (1 : I); dung djch NazSzOj 0,05N. - Chuin dp miu blank: cho 10 ml dung dich iode va 5 ml muac cat va mpt vai gigt HCI vao erlen 250 ml va lie diu. dat cac erlen vao trong toi khoang 15 ml. sau do chuan do vai dung djch Na2S203 0,1 N, sir dung chit chi thj la ho tinh bgt. The ticb Na2S203 0,01 N chuan do dugc gpi la (Vo). Lap lai thi nghiem nhu tren 3 lan. - Chuin do vai mau thtr nghiem bao gom H2S: them 5 ml dung djch H2S vao erlen sau do tiln hanh tuong tu nhu mau blank. Ham lupng H2S dugc xac djnh theo cong thii'c: (F„-F,.).17.V,„^,^^^.1000 ^H,S- y vai: ning do Na2S203 la A^^^ ^^^ = 0,\N ; The tfch H2S: F^ ^ = 5 ml; 17 la khoi lupng (grams) ciia H2S; 1000 la the ticb pha loang; Vx la the ticb cua Na2S203 0.05N dugc str dung de chuan do 5 ml mau H2S; VQ la the tich cua Na2S203 0,05 N dugc str dung de chuan do 5 ml mau blank. 2.3. Khao sat kha nang xir li Hydrogen sulfite ciia vi khuan co trong moi truong - Mau nuac khao sat dugc lay tir kenh NLTN va tien hanh xti' li trong phong thi nghiem duoi dang mo hinh be hieu khi. The tich mau nuac thtr nghiem la 50 lit vai so lugng vi khuan khoang lO^-lO"* CFU/ml.Toc do khi dugc sue vao khoang 10 lit khi/giay nham mue dich co the duy tri dugc mirc DO binh thuong vai dieu kien hieu khi tren 3 mg/l (Frank A.Spellman.2003). Thi nghiem thuc hien 3 lan - Thu nhan mau nuac trong mo hinh sau moi gia va phan ticb bam lugng H2S bang phuong phap lot dong thai xac djnh pH cua mau bang pH ke. -Tien hanh thtr nghiem trong dieu kien khong bo sung vi khuan de doi chtrng. 3. KET QUA VA THAO LUAN 3.1. Phan lap vi khuan co trong mau nuo'c Vi khuan oxi boa cac hgp chat sunfur dugc phan lap ttr mau nuac kenh NL-TN trong ba loai moi truang gom Mi (pH 4,4), M2 (pH 7,2) and M3 (pH 8,5). Cac chting vi khuan nay co kha nang str dung cac hgp chat sunfur (moc tren moi trucmg phan lap chuyen biet) va deu la true khuan gram am. Trong so cac vi khuin phan lap dugc chi 8 chung co kha nang oxi hoa cac hgp chat sunfite (So lieu trinh bay trong bang 2). 8 chting vi khuin oxi boa bgp chat sulfite dugc khao sat hinh dang bang kfnh biln vi dien ttr (hinh anh cua SOB thi hien trong hinh 2). 86 Hinh 2. 8 loai vi khuin dugc chup duai kinh hign vi - N3.1TT (a); N3.3TT (b); N3.4TT (c); N3.6TT (d); N6.1TT(e);N6.2TT(f);N10,l K(g);Nll,l K (h) (Giai thich ki hieu N3.6TT: La vi khuan co so thu ty 6 dugc phan lap Ur miu nuac s6 3 tren moi truong trung tilth; K moi truong kiim) Bdng 2. Tinh chat va so lugng vi khuan phan lap dugc tir mau nuac STT 1 2 3 4 5 6 7 8 9 10 11 Lo Nl N2 N3 N4 N5 N6 N7 N8 N9 NIO Nil Ml 0 0 0 o 0 o 0 0 0 0 0 M2 o 0 X 0 0 X 0 0 0 0 0 M3 0 0 0 0 0 0 0 0 0 X X Miia Kho Kho Kho Kho Kho Kho Mua Mua Mua Mua Mua So vi khuan phan lap 0 0 4 0 0 2 0 0 0 I 1 pH 6,5 6,7 6,5 6,5 6,8 6,5 7,1 7,0 7,1 7,0 7,0 Thuy trieu Xuong Len Xuong Xuong Len Xuong Len Xuong Len Xuong Xuong X. mau CO vi khuin dugc chon; O. mau khong co vi khuan dugc chon. 87 Bang 2 chi ra rang co 4 mau nuoc phan lap dugc vi khuin (2 miu ctia miia mua va 2 mau ctia miia kho). Co 2 mau nuac phan lap dugc vi khuan trong moi trirang trung tinh (M2), trong khi do so con lai phat trien trong moi truong kiem (M3). Qua trinh phan lap vi khuin da chgn dugc 6 chiing vi khuan ttr mau nuac miia kho va 2 chting vi khuin tii' mau nuoc mtia mua. 3.2. Kha nang khir cac hgp chat sunfur cua vi khuan Trong nghien ciru khao sat kha nang xtr li hydrogen sulfite, nong do H2S c6 chi so trung binh khoang 0,59 ppm va gia trj pH ctia mau niroc a dieu kien trung tinh (so lieu phan tich khong dugc the hien trong bai bao cao). So lieu khao sat dugc trinh bay a binh 3 cho thay kha nang oxi boa H2S cua vi khuan phan lap dugc tir mau. Sau 4 gia xu' If chting N3.4 dugc khao sat cho ket qua xu If cao nhat ham lugng H2S giam 99,29%), trong khi do chting N3.3 dugc xu If cho kit qua thap nhat trong 8 chung dugc khao sat chi giam dugc 25%). (%) 99.29 100.00 90.00 80.00 70.00 60.00 50.00 40.00 30.00 20.00 10.00 0.00 Removal H2S after 4 hours treatment period (%) • N3.4 83.OS 81.25 80.00 ONILI 73.44 • N10.1 • N6.1 es.is 54.84 N11.1 N10.1 N3.6 N3.1 • N3.6 25.00 N3.3 0 N3.1 • N6.2 1 • N3.3 Hinh 3. Kha nang xii' li H2S ciia cac chiing vi khuan trong dieu kien mo hinh Trong cac nghien cu'u ctia chting toi cac chting N3.4; Nl I.I; NlO.l; N6.I co kha nang phan giai HTS tren 80%i day la con so rat co y nghTa trong qua trinh lua chgn tmg dung vi sinh vat trong sil' li moi truong vi hieu qua kha cao, trong khi do cac chimg khac kha nang xtr If duoi 80% nhuN3.6;N3.1;N6.2,N3.3. 80 pH 7.8 7.6 7.4. 7.0 J 6.6 -I 6.4 -i- pH Changing Oh 1 h 2h 3h pH Times(h) 4 h Hinh 4: Gia tri pH sau 4 gia khao sat va xii li vai cac chung vi sinh vat phan lap dugc 88 Kha nang phan giai H2S ciia nhu'ng vi khuan phan lap dugc tir nuoc kinh NL-TN sau 4 gia cho thay rang nhung san pham tao thanh khong lam giam pH moi truang. So lieu thi nghiem trong hinh 5 cho thay gia trj pH trung binh trong qua trinh xii' If voi cac chiing vi sinh vat deu tang (so lieu ciia tirng thf nghiem khong the hien trong bao cao). 4. KET LUAN Trong ket qua nghien ciru ciia chiing toi da phan lap dugc 8 chiing vi khuan co kha nang khir H2S til' kenh NLTN. Tat ca chimg deu la vi khuan hieu khi va la true khuan Gram am. Trong 8 chimg nay co 4 chiing N3.4; Nl 1.1; NlO.l; N6.1 co kha nang loai H2S tren 80% trong do chimg N3.4 co kha nang loai H2S cao nhat la 99,29%. San pham tao thanh tii' qua trinh oxi hoa hydrogen sulfite khong lam giam gia trj pH ciia moi truang (khong tao thanh axit) Til' ket qua nghien cti'u co the tiep tuc irng dung nhung chimg vi khuan co kha nang xir li H2S cao bang kT thuat co djnh vi sinh vat nham tao ra sinh pham xti' If moi truong. Tuy nhien, can phai tien hanh khao sat them kha nang xir If va cac dae tfnh sinh hpc ciia cac chimg vi khuan nay de co the thuo'ng mai hoa san pham. TAI LIEU THAM KHAO 1. A. J. H. Janssen, G. Lettinga, and A. de Keizer - Removal of hydrogen sulphide from wastewater and waste gases by biological conversion to elemental sulphur Colloidal and interfacial aspects of biologically produced sulphur particles, Colloids and Surfaces A: Physicochemical and Engineering Aspects 151 (1999)389-397. 2. Alex T. Nielsen, Wen - Tso Liu, Carlos Filipe, Leslie Grady Jr., Soren Molin, and David A. Stahl - Identification of a Novel Group of Bacteria in Sludge from a Deteriorated Biological Phosphorus Removal Reactor, Applied and Environmental Microbiology 65 (3) (1999)1251-1258. 3. Altschul S. F., Thomas L. M., Alejandro A. S., Jinghui Z., Zheng Z., Webb M., and Lipman D. J. - Gapped BLAST and PSI - BLAST: a new generation of protein databases search program. Nucleic Acids Research 25 (17) (1997) 3389-3402. 4. Andrew S. Ball, David B. Nedwell and Rupert G. Perkins - Oxidation of hydrogen sulphide in sour gas by Chlorobium limicola. Enzyme and Microbial Technology 41 (2007)702-705. 5. APHA AWWA and WPCF - Standard Method for Examination of Water and Wastewater 20th ed., Washington, D.C. American Public Health Association, 1998, p. 1325. 6. Aragno M. - Aerobic chemolithoautotrophic bacteria. In J. K. Kristjansson (ed.). Thermophilic bacteria, Boca Raton: CRC Press, Fla, 1991, pp. 7-103. 7. Bitton Gabriel - Wastewater Microbiology, Chichester: John Wiley & Sons, 3rd ed. 2005, p. 768. 8. Brock T. D., and H. G. Schlegel - Autotrophic bacteria. Wis: Springer - Verlag, Madison, 1989, pp. 1 - 15. 89 9. Brune D. C. - Sulfur oxidation by phototrophic bacteria., Biochim. Biophys. Acta 975 (1989)189-221. 10. Brune D. C. - Sulfur compounds as photos\nthetic electron donors. In R. E. Blankenship, M. T. Madigan, and C. E. Bauer, Ano.xygenic photosynthetic bacteria, The Netherlands: Kluwer, Dordrecht, 1995, pp. 847-870. 11. Caroline M. O'Hara - Manual and Automated Instrumentation for Identification of Enterobacteriaceae and Other Aerobic Gram - Negative Bacilli., Clinical Microbiology Reviews 18(1) (2005) 147-162. 72. Chirajyoti Deb, Ranadhir Chakrabort}', Amar N. Ghosh, Nitai C. Mandal, Tusharmouli Mukherjee, Pradesh Roy - A generalized transducing thiophage (TPC - 1) of a facultati\e sulfur chemolithotrophic bacterium, Bosea thiooxidans CT5, of a - Proleobacleria, isolated from Indian soil, FEMS Microbiology Letters 227 (1) (2003) 87-92. 13. Chris Easter, Chris Quigley, Peter Burrowes, Jay Witherspoon and Dirk Apgar - Odor and air emissions control using biotechnology for both collection and wastewater treatment systems. Chemical Engineering Journal 113 (2005)93-104. 14. Christoph Griesbeck et al - Mechanism of Sulfide - Quinone Reductase Investigated Using Site - Directed Mutagenesis and Sulfur Analysis, Biochemistry 41(39) (2002) 11552- 11565. 15. Dam Sao Mai, Nguyen Khanh Hoang Vu Dinh Huy, Abdukarimov Abdusattor, Abdurakhmonov Ibrokhim - The metal corrosion ability of sulfate reducing bacteria isolated from the mine White Tiger, Petrovietnam journal Vietnam (4) (2007) 24-28. 16. Dam Sao Mai, Nguyen Khanh Hoang Vu Dinh Huy, Abdukarimov Abdusattor, Abdurakhmonov Ibrokhim - The biocide effect to sulfate reducing bacteria isolated from the mine White Tiger, Petrovietnam journal Vietnam (5) (2007) 33-37. 17. Daniel C. Brune - Sulfur oxidation by phototrophic bacteria, Biochimica et Biophysica Acta (BBA) - Bioenergetics 97 (5) (1989) 189-221. SUMMARY ISOLA FED AND SURVEYED REDUCE HYDROGEN SULFITE (H2S) ABILLITY OF SULFUR-OXIDIZFNG BACTERIA (SOB) IN THE WATER OF NHIEU LOC - THI NGHE CANAL 8 strains of Sulfur oxidizing bacteria (SOB) were isolated and tested in order to remove hydrogen sulfide (H2S) from Nhieu Loc - Thi Nghe canal in the aerotank pilot scale. The desulphurization ability of the strains N3.4; Nll.l; NlO.l; N6.1;N3.6; N3.1; N6.2; N3.3 were decreased (99.29%; 83.05%; 81.25%; 80.00%; 73.52%; 68.18%; 41.91%; 25.00%) respectively Dia chi: Nhdn bdi ngdy 22 thdng 3 ndm 2009 Nguyen Khanh Hoang, Nguyin Hoai Phong, Htra Thach Son, Truong Dai hpc Cong nghiep Tp. H6 Chi Minh. Nguyen Van Cuang, Khoa KT thuat Y sinh, Truang dai hpc Chung Yuan, Dai Loan. 90 . 8 3-9 0 PHAN LAP VA KHAO SAT KHA NANG LOAI HYDROGEN SULFITE (H2S) CUA VI KHUAN OXY HOA HOP CHAT SULFUR (SOB) TRONG Nu'ac KINH NHIEU LOC - THj. he thong kinh rach: Kinh Nhieu Lpc - Thj Nghe (NL-TN) bao phii 33 km"; Kinh Tau Hti - Doi - Te bao phii 37 km"; Kinh Tau Hu - Ben Nghe