최종보고서 /

Transcription

1 최종보고서 /

2 최종보고서 /

3 제출문 국립환경과학원장귀하

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5 요약문 1. 사업개요 사업명 주 관 사업기관 2. 연구목적및결과 Ⅰ. 연구필요성및목적 (PM2.5) 23% (USEPA, 2000).., PAH.

6 Ⅱ. 연구목적 (PM2.5). Ⅲ. 연구방법 (AC, HORIBA ) ETC ESC., PM2.5, 16 PAH PAH-TEQ Wang (2011) Ⅳ. 연구결과 PAH BaP bio-teq ETC ESC μg -TEQ/g-PM2.5 ETC ESC. PAH-TEQ 10-6.

7 ... CHO-K1 cell BEAS-2B cell (genetic toxicity) (epigenetic toxicity). 100μg /ml DNA, (apoptosis). 100μg /ml (cm 2 ) ETC ESC μg /cm 2 20 μg /cm 2. BEAS-2B bronchiol( ) ETC ESC (cm 2 ) 44.44μg 20μg. (PM2.5)

8 ., (DPF+DOC),,. 3. 기대효과및활용방안.

9 목차 1. 연구사업명 : 1 2. 연구의배경, 목적및필요성 연구의배경 연구필요성 연구의목표 4 3. 연구내용및방법 연구내용 연구방법 7 4. 연구결과 경유자동차에서미세먼지등오염물질에의한건강위해성영향국 내외자료조사 디젤엔진에서배출되는미세먼지및유해오염물질배출특성평가 경유엔진에서배출되는미세먼지에의한호흡기관련인체위해성평가 결론 기대성과및활용방안 기대성과 활용방안 참고문헌 i -

10 그림목차 그림 1. 폐암발생에대한유전적 (gemetic) 및후생유전적 (epigenetic/non-genotoxic) 독성기전 6 그림 2. Diesel engine system (AC동력계) 9 그림 3. 경유미세먼지 (PM2.5) 시료채취시스템 10 그림 4. ETC모드 10 그림 5. ESC모드 11 그림 6. Overview of ICP-OES 18 그림 7. Genotoxicty test 19 그림 8. Apoptosis signaling pathway 20 그림 9. CHO-K1 cells 21 그림 10. BEAS-2B cells 21 그림 11. DNA 손상측정방법인 Comet assay 22 그림 12. Comet image analysis software에의한 DNA손상정량화방법 23 그림 13. Cytochalasin B 처치후나타나는 binucleated cell 24 그림 14. 염색체손상확인을위한 in vivo Cytochalasin B micronucleus test 24 그림 15. 소핵형성관찰방법 25 그림 16. 경유차량에서배출된입자상물질의구성도 28 그림 17. 입자상물질의호흡을통한유입및침착 31 그림 18. 디젤배기가스중의입자크기 56 그림 19. 운전모드 (ETC모드, ESC모드 ) 에따른 DNA손상평가 69 그림 20. 운전모드 (ETC모드, ESC모드 ) 에따른 chromosome손상 70 그림 21. 운전모드 (ETC모드, ESC모드 ) 에따른 DNA손상평가 71 그림 22. 운전모드 (ETC모드, ESC모드 ) 에따른소핵형성 72 그림 23. 유전독성평가의양성판정 73 - ii -

11 그림 24. p53-null H1299세포및 p53-wild H460세포에서소핵형성확인 75 그림 25. Pifithrin α 처리농도선정 76 그림 26. Pifithrin α에의해 p53이결여된 BEAS-2B세포에서세포사멸 apoptosis 76 그림 27. 위해성평가과정 82 - iii -

12 표목차 표 1. Proposed list mobile souce air toxics 1 표 2. 운전모드특성 11 표 3. 특성시험용연료 ( 경유 ) 분석결과 12 표 4. 16개 PAH류목록 14 표 5. GC-MSD에의한 PAH분석조건 15 표 6. GC-MSD에의한 VOC 분석조건 16 표 7. 경유차량입자상물질에흡착되어있는오염물질 29 표 8. 국외입자상오염물질대기환경기준 31 표 9. 경유차량가스상물질 33 표 10. IARC에의한발암성분류 40 표 11. US EPA에의한발암성분류 (US EPA, 1986*) 41 표 12. Diesel engine exhaust의비발암독성지수 42 표 13. Diesel engine exhaust의발암독성지수 43 표 14. 기간별 PAH의 carcinogenic group (PAH position paper Annexes July 27th 표 15. PAH류의발암성분류 ( 부록Ⅰ참조 ) 45 표 16. 디젤미세먼지에서검출되는 Nitro-PAH 목록 49 표 17. 경유차량배출가스규제대상금속류목록 (listed by ARB) 53 표 18. 디젤배기가스중의입자개수및분진포집량 55 표 19. 디젤배기가스중의 PAH류분석 58 표 20. 디젤배기가스중의휘발성물질농도 59 표 21. 디젤배기가스중의휘발성물질농도 60 표 22. 디젤배기가스중의휘발성물질농도 61 표 23. 디젤배기가스중휘발성물질 (VOC) 에대한비발암독성정보 62 표 24. 디젤배기가스중휘발성물질 (VOC) 에대한발암독성정보 63 표 25. 디젤분진에흡착되어있는금속류측정 64 - iv -

13 표 26. 디젤배기가스중금속류에대한비발암독성정보 65 표 27. 디젤배기가스중금속류에대한발암독성정보 67 표 28. 표면적당노출농도 77 표 29. Lifetime cancer risk assessment에사용되는 parameter 93 - v -

14 1. 연구사업명 : 경유자동차에서발생되는미세먼지 (PM2.5) 의인체위해성영향평가 2. 연구의배경, 목적및필요성 2-1. 연구의배경 경유자동차에서발생되는유해물질 ( 1) (Particulate matter) NO x. 표 1. Proposed list of mobile source air toxics Acetaldehyde Diesel exhaust MTBE Acrolein Ethylbenzene Naphthalene Arsenic compounds Formaldehyde Nickel compounds Benzene n-hexane Polycyclic organic matter 1,3-Butadiene Lead compounds Styrene Chromium compounds Manganese compounds Toluene Dioxin/Furans Mercury compounds Xylene - 1 -

15 (iron), (soot), (tar) (carbonaceous material). 1 μm,,. 2.5μm 미세디젤분진 (DEP PM2.5) polyaromatic hydrocarbon (PAH), heterocyclic compounds, phenols nitroarenes,. IARC Group 2A Probably carcinogen. (reactive oxygen species),,. PAH(polycyclic aromatic hydrocarbons) USEPA 7, benzo(a)pyrene, benz(a)anthracene, chrysene, benzo(b)fluoranthene, benzo(k)fluoranthene, dibenz(a,h) anthracene indeno[1,2,3-cd]pyrene Group B2(probable human carcinogens) 입자상물질저감을위한디젤자동차의후처리장치 후처리기술적용. (PM) NOx - 2 -

16 디젤분진여과장치 (DPF: diesel particulate filter trap). 가열방법및촉매방법 필터를재생. 디젤분진여과장치 (DPF trap) 의재생방법 가열방식 soot 550 ~ 600 촉매방법 soot 250. 디젤분진여과장치 (DPF trap) 의문제점 :. 2 PAH nitration halogenation 연구필요성 PM

17 . 1. ETC ESC p53 (Genetic) (Epigenetic) 연구의목표 경유엔진의 ETC 및 ESC 모드에서발생되는미세먼지 (PM2.5) 의배출특성평 가와미세먼지에의한폐암발생등위해성평가를통해자동차에의한 대기오염개선정책및국민의삶의질을향상시키고자한다

18 3. 연구내용및방법 3-1. 연구내용 경유자동차에서의미세먼지 (PM2.5) 등오염물질에의한건강 위해성영향국 내외자료조사 연구목적 :. 연구내용 : 저감장치가부착된경유엔진에서발생되는미세먼지및유해 오염물질배출특성평가 연구목적 : (ETC ESC). 연구내용 : (ETC ESC) ( PAH )

19 경유자동차에서의미세먼지에의한호흡기관련인체위해성영향평가 연구목적 : (Genetic) (Epigenetic). 연구내용 :. DNA DNA (chromosome),. ( 1). 그림 1. 폐암발생에대한유전적 (genetic) 및후생유전적 (epigenetic/ non-genotoxic) 독성기전 - 6 -

20 3-2. 연구방법 경유자동차에서의미세먼지 (PM2.5) 등오염물질에의한건강 위해성영향국 내외자료조사 연구방법 : (PM2.5) 국내외인체위해성에대한선행연구. 호흡기 (Genetic) (Epigenetic) 경유자동차에서발생되는미세먼지에대한폐암위해성영향평가. 1) 자동차로인한인체위해성관련주요위해물질영향조사 : 2) 미세먼지위해성관련국내외연구자료조사 : 3) 유해오염물질에의한인체위해성영향조사 : - 7 -

21 경유엔진에서발생되는미세먼지및유해오염물질배출특성평가 연구방법 : (ETC ESC) (PM2.5),. (DCM) PAH.. 1) 경유엔진에서주행모드에따라발생되는미세먼지 (PM2.5) 포집및특성분석 경유자동차배출미세먼지특성 : nuclei mode 0.03μm, (thermophoretic force)..,,,. 경유자동차배출미세먼지포집원리 : AC ( HORIBA ) EPA

22 dilution system. Dilution tunnel cyclone PM2.5 ( 2, 3).,,,,. 경유엔진운행조건별미세먼지의입자량분석 : ( ) (ESC ETC) (PM2.5). 20 ~ 30 50% 6 ESC ( 4), ETC ( 5) ( 2). 3. 엔진배기량 7.6L( 시내버스엔진 ) 최대출력 235kw/2200rpm 최대토크 1323NM/1200rpm 압축비 17.1:1 그림 2. Diesel engine system (AC 동력계 ) - 9 -

23 그림 3. 경유미세먼지 (PM2.5) 시료채취시스템 그림 4. ETC 모드

24 그림 5. ESC 모드 표 2. 운전모드특성 ETC 모드 ESC 모드 특징 중속 중부하영역 ( 적재량 50% 정도적재 ) 운전시간 30분 PM2.5 채취시간 :30분 중고속 : 차량속도 70% 영역 중고부하 ( 적재량 70% 정도적재상태 ) 운전시간 : 28분 PM2.5 채취시간 : 28분

25 표 3. 특성시험용연료 ( 경유 ) 분석결과 항목기준분석결과 10% 잔류탄소량 (%) 0.15 이하 0.01 미만 (kg/m 3 ) 815~ 황함량 (ppm) 10 이하 5 다고리방향족 ( 무게 %) 5 이하 3.6 윤활성 ( μm ) 400 이하 215 방향족화합물 ( 무게 %) 30 이하 23 세탄지수 52 이상 52 경유엔진운행조건별미세먼지의입자크기분석 EEPS(Engine Exhaust Particle Sizer, model TSI 3090). EEPS TSI,. 12 ELPI(Electrical Low Pressure Impactor) 100nm SMPS(Scanning Mobility Particle Sizer) (Mobility)

26 SMPS electrometer. EEPS(Engine Exhaust Particle Size; model: 3090EEPS) 5.6 ~ 560nm. 32 (16 /decade), 10L/min 10 3 ~ 10 8 /cm 3 Inlet 1.0μm size cut cyclone 1μm cyclone.. 85V, 470V, 1200V electrometer.. electrometer ( , 2008 Ⅱ). 2) 포집된미세먼지의유기성추출및인체위해성관련유해물질분석 포집된미세먼지의유기성추출 Lewtas (1990). DCM ASE2000 crude. GC-MS PAH 100mg/ml DMSO 0.22μm syringe filter

27 디젤미세입자에흡착된유기성유해물질분석 16 PAH ( 4) GC-MS. Dionex (ASE 200) 11ml Extraction Cell error. ASE, Sample psi 10~15 Dichlormethane 20ml. PAHs Collection Vial. (Rotary evaporator) 1ml Hexane 2~3ml 1ml. Agilent Vial. 5 GC/MSD. 표 4. 16개 PAH류목록 No. Compounds No. Compounds 1 Acenaphthylene 9 Chrysene 2 Acenaphthene 10 Dibenzo(a,h)anthracene 3 Anthracene 11 Fluoranthene 4 Benz(a)anthracene 12 Fluorene 5 Benzo(b)fluoranthene 13 Indeno(1,2,3-cd)pyrene 6 Benzo(k)fluoranthene 14 Naphthalene 7 Benzo(ghi)perylene 15 Phenanthrene 8 Benzo(a)pyrene 16 Pyrene

28 표 5. GC-MSD 에의한 PAH 분석조건 Instrument Analysis conditions Model Aglient 6890N/5975B Column DB-5 ( 60m X 0.25mm X 0.25um ) Temp( ) Rate( /min) Hold(min) Total(min) Column Temp GC/MSD Column Flow 1.0ml/min Split Ratio 5:1 MS ion source temp 230 MS quad temp 150 MS scan range 60~500amu Injection volume 2ul

29 3) 기타인체위해성관련유해물질분석 CO, THC, NOx 및 CO 2 분석 (ESC ETC) CO, THC, NOx CO 2. (Horiba, MEXA-7100) CO, THC, NOx, CO 2, CO CO 2 (NDIR, Nondispersive Infrared), THC (HFID, Heated Flame Ionization Detector), NOx (CLD, Chemiluminescence Detector). VOC분석 (ESC ETC) 1,3-butadiene, benzene, toluene, ethylbenzene, m,p,o-xyelene stylene GC-MSD. 6 Air-Sever Unity Thermal desorber( ).. Purge & Trap Nafion Dryer. TD cold trap -15 VOCs, 300 GC/MSD. 6 GC/MSD

30 표 6. GC-MSD 에의한 VOC 분석조건 Instrument Analysis conditions Purge Prepurge Time(ml/min) 3 Trap Prepurge Time(ml/min) 0.2 Sample Flow Sample Flow Rate(ml/min) 20 TD Trap Low Temp( ) -10 Trap Desorb Trap High Temp( ) 300 Trap Hold Time(min) 5 Split Flow(ml/min) 10 Model Varian CP-3800 / Saturn 2200 Column DB-1 ( 60m X 0.32mm X 5.00um ) Temp( ) Rate( /min) Hold(min) Total(min) GC/MSD Column Temp Column Flow ml/min Split Ratio 5:1 MS scan range 45 ~ 350amu

31 금속류분석 : ICP (Al, Cd, Cr, Cu, Fe, Mn, Ni Zn). Quartz filter Microwave. Microwave,,,. Microwave 60% HNO 3 8ml 98% H2so4 2ml 250W 6, 400w 6, 650w 6, 250w 6. ICP- OES(varian ICP-OES,720-es, 6). ICP-OES 6000 K photomultifiler. 그림 6. Overview of ICP-OES

32 경유자동차에서의미세먼지에의한호흡기관련인체위해성영향 평가 연구방법 : 유전적독성영향 (CHO-K1 cells) (BEAS-2B cells) DNA comet assay micronucleus test ( 6). In vitro comet assay OECD draft In vitro micronucleus test OECD (TG487). 후생유전적독성영향 p53 (pifithrin-α ) apoptosis 유도 p53 ( 7). 그림 7. Genotoxicity test

33 그림 8. Apoptosis signaling pathway 1) 인체위해성관련표적세포등을활용한유전적및후생유전적독성평가 ➀ 미세먼지유기성추출물에의한유전독성 (Genetic) 평가 세포주 (chinese hamster ovary cell) CHO-K1 ( 9) (human normal bronchial epithelial cell) BEAS-2B ( 10)

34 그림 9. CHO-K1 cells 그림 10. BEAS-2B cells DNA손상확인 [Single cell gel electrophoresis(comet assay) 시험 ] DNA Comet assay Singh (1991) alkaline(ph 13) CHO-K1 BEAS-2B ( 11). 12-well plate 80% DMEM well Vehicle 0.1%. 24 well trypsin-edta. PBS 2 PBS 1ml ~ 10 4 cell % low-melting agarose 1.0% normal agarose (1-21 -

35 layer) fully frosted microscope slides (2 layer), agarose low melting agarose (3 layer). lysing solution(1% sodium sarcosinate, 2.5M NaCl, 100mM EDTA, 10mM Tris, ph 10.0 and 1% Triton X-100) 1 unwinding (1mM Na 2 EDTA 300mM NaOH) 25V 30. ethidium bromide(2μg /ml) (Leica DMLB; excitation filter nm, barrier filter 590nm) 400 autoimage analysis system(komet version 5.0; kimetic imaging LtD. UK) 100 ( 12). DNA Olive tail moment(tail distance percentage of DNA in tail) 3 2. 그림 11. DNA 손상측정방법인 Comet assay

36 그림 12. Comet image analysis software 에의한 DNA 손상정량화방법 염색체이상시험 [In vitro 소핵시험 ] clastogen aneugen in vitro cytokinesis-block micronucleus test(cbmn assay). 1 cycle. cytokinesis block cytochalasin B. 2 ( 13). 8-well chamber slide well well 200ml vehicle 0.1%, cytokinesis block cytochalasin B(cyt-B, 3μg /ml). PBS (Phosphate buffered saline) 1% sodium citrate 4 5 1% acetic acid methanol

37 RNAse(0.01mg/ml in 2 SSC) 37 5 slide 2 SSC 5% Giemsa 20 ( 14 15). 2well 1,000. uplight microscope emulsion oil 400 magnification / %. Cytochalasin B (-) Cytochalasin B (+) 그림 13. Cytochalasin B 처치후나타나는 binucleated cell 그림 14. 염색체손상확인을위한 in vitro Cytochalasin B micronucleus test

38 그림 15. 소핵형성관찰방법 ➁ 미세먼지유기성추출물에의한후생유전 (epigenetic) 독성평가 세포주 p53 BEAS-2B p53 pifithrin-α BEAS-2B. p53 p53-wild H460 p53-null H1299 p

39 p53의존성 apoptosis 확인 p53 apoptosis 24 apoptosis apoptotic nucleosome cell death detection ELISA kit(roche Diagnositic). 96-well plate well cytoplasm streptavidin 96 well plate biotinylated mouse antihistone antibody peroxidaseconjugated mouse anti-dna antibody nm 490nm microplate reader apoptosis. 2) 경유자동차미세먼지에의한인체위해성관련연구를통한정책활용

40 4. 연구결과 4-1. 경유자동차에서의미세먼지 (PM2.5) 등오염물질에의한건강 위해성영향국 내외자료조사 : 경유자동차미세먼지에대한 건강위해성연구자료조사 자동차로인한인체위해성관련주요위해물질영향조사,,,. (NOx), (CO), (HC) (CO 2 ), (VOCs)., VOCs, benzene( ), toluene ( ), xylene( ), formaldehyde( ), acetaldehyde( ) 1,3- (1,3-butadiene)

41 .,,. 1) 경유차량에서배출되는입자상물질 ( 52 ) weighing chamber (Soluble Organic Fraction, SOF), (Sulfates), (Solid Fraction, SOL),. 7. 그림 16. 경유차량에서배출된입자상물질의구성도

42 표 7. 경유차량입자상물질에흡착되어있는오염물질 Heterocyclics, hydrocarbons(c14~c35) and PAHs/-derivatives Acids, Cycloalkanes, Alcohols, Esters Alcanoic acids, Halogenated compounds n-alkanes Ketones, Anhydrides Nitrated compounds Aromatic acids Sufonates Quinones Elemental carbon, metals, water, inorganic sulfates and nitrates carbon material(sol) (agglomeration). (thermophoretic force). (dilution) (cooling),, (SOF). SOF. (aliphatic hydrocarbon) 12~20, 18~36 SOF (PAHs; Polycyclic aromatic hydrocarbons). Sulfuric acid sulfur. sulfur sulfur dioxide sulfur trioxide

43 : 0.05 ~ 0.3μm,..,,. 1μm, 미세먼지 (Particulate Matter, PM2.5) 2.5μm 10 TSP PM10 PM PM2.5 15μg /m 3, 24 25μg /m 3, WHO 8μg /m 3, 24 25μg /m 3 PM2.5. PM2.5, 8 25μg /m 3, 24 50μg /m

44 그림 17. 입자상물질의호흡을통한유입및침착 표 8. 국외입자상오염물질대기환경기준 기준시간 PM2.5 ( μg /m 3 ) 한국 1) 미국캐나다호주 유럽연합 WHO 24 시간 년 ) 2015 년시행예정

45 다환방향족탄화수소 (PAHs) PAH. PAH PAH. PAHs,,. PAH.. PAH 2~4 5. PAH PAH PAHs. PAH PAHs (pyrosynthesis) (pyrolysis). PAHs nitration halogenation form. 중금속.,,,.,,,,,,

46 2) 경유차량에서배출되는가스상물질 ( 표 9) 휘발성유기합물 (VOCs) VOCs. VOCs, VOCs. VOCs,,,,,,. 표 9. 경유차량가스상물질 Heterocyclics, hydrocarbons(c1~c10) and PAHs/-derivatives Acids, Cycloalkanes, cycloalkenes, Aldehydes dicarbonyls, alkanoic acids, Ethylene, n-alkanes, Halogenated compounds, n-alkenes, Ketones, Anhydrides, Nitrated compounds, Aromatic acids, Sulfonates, Quinones Acrolein, Ammonia, Carbon dioxide, Carbon monoxide, Benzene, 1,3-Butadiene, Formaldehyde, Formic acid, Hydrogen cyanide, Hydrogen sulfide, Methane, Methanol, Nitric and nitrous acids, Toluene, Nitrogen oxides, nitrous oxide, Sulfur dioxide, water

47 미세먼지위해성관련국내 외연구자료조사 (PM2.5) pubmed google / /, p53 apoptosis.. 바이오디젤및후처리장치에대한독성저감및배출특성조사연구. DOC(diesel oxidation catalyst) DPF(diesel particle filter). DOC DPF NO NO2 PAH nitration (Heeb et al., 2008). nitration nitro-pah. DOC THC (Klein et al., 1998)... Ratcliff et al., Environ Sci Technol , ULSD(ultra-low-sulfur diesel)/ (B20, B100) (DPF) DOC(diesel oxidation catalyst)

48 volatile unburned hydrocarbons, carbonyls PAH/nitro-PAH. ULSD B20, B100 PAH nitro-pah PM. benzene naphthalene B100. soy-biodiesel FAME aromatic ring. DPF PAH total nitro PAH 1-nitropyrene formaldehyde B20, ULSD DPF. Shi et al., J. Toxicol. Environ. Health, 73(19), , (Filter system, diesel DOC, diesel oxidation catalytic converter) Ames test. DOC. SJalava et al., Inhalation Toxicology, 2010; 22(s2): (rapeseed methy ester, RME; hydrotreated fresh vegetable oil, HVO) DOC/POC catalytic converter PM1. PAH PM1 mouse RAW264.7 macrophage (TNFα MIP-2),,. EN590 HVO RME

49 catalyst. RME. Catalyst EN590 HVO RME. catalyst EN590. 유전독성및산화적스트레스. PAH Ames test. pubmed 3 acellular system. Topinka et al., Toxicol Lett ;202(3):186~192 - Topinka (2011) Czech (PM2.5) calf thymus DNA acellular assay (DNA adduct DNA ) PAH. Oh SM et al., Mutat Res. 2011: 16;723(2):142~151 - Oh (2011) Topinka PM2.5. BEAS-2B

50 comet assay in vitro micronucleus test. oxidative stress. André et al., J Appl Toxicol Mar;31(2):131~138 - Dunkerque PM2.5 samonella (TA98, TA102 YG1041), A549 1 alveolar macrophages., DNA adduct DNA DNA-ineteraction mechanism. Gualtieri et al., Toxicol Lett Dec 9. - PM2.5 A549, reactive oxygen species. : xenobiotic stimuli (CYP1A1, CYP1B1, TIPARP, ALDH1A3, AHRR) and to the cell-cell signalling (GREM1) pathways with winter PM2.5 inducing higher fold increases. Moreover the winter fraction modulated also JUN (cell-cell signalling), GDF15, SIPA1L2 (signal transduction), and HMOX1 (oxidative stress). Two genes, epiregulin (EREG) and FOS-like antigen1 (FOSL1). PAH CYP enzyme. Perrone et al., 2010 Mar;78(11): PM1 PM2.5 (, PAH ). A

51 cytokine(il-1, IL-8), comet assay DNA damage. bivariate analysis and multivariate data analysis (Principal Component Analysis, PCA). Al, As, Cr, Cu, Fe, Zn, Ba Mn Ca(++) Mg(++) cell viability DNA damage. 염증반응및 p53관련성 apoptosis작용.. cytokine apoptosis. Apoptosis p53 Bax (Bcl-2-associated X protein). alveolar apoptosis. apoptosis. DEP P53 apoptosis. Li et al., Particle and Fibre Toxicology, 2010, 7(6): 1~12. - aortic endothelial cell (HAEC) pro-inflammation idling (UFP1) urban dynamometer driving schedule (UFP2)

52 (Fe, Zn, Cd, Pb ) (PAH ) UFP1 UFP2 stress response gene(ho-1, OKL38, TF ). UFP2 pro-inflammatory gene IL-8, MCP-1, VCAM NF-kB pro-inflammation. Tal et al., Toxicology and Applied Pharmacology, 2010, - DEP(diesel exhaust particles) MAP kinase NF-kB AP-1. IL-8 airway epithelial cell proinflammatory chemokine. DEP IL-8 IL-8 upstream reporter gene assay. low organic DEP NK-kB IL-8 high organic DEP NK-kB IL-8. DEP pro-inflammatory gene expression. Yun et al., Toxicology in vitro 2009; 23: M774A.1 macrophage p53 p53 Mdm2 (Murine double minute 2). DEP Mdm2 pro=apoptosis protein Bax p53-. p53 pifithrin-α DEP apoptosis p53 J774A.1 apoptosis p

53 유해오염물질에의한인체위해성영향조사. 1) 경유차량에서배출되는입자상물질 Diesel engine exhaust IARC Group 2A( 10), US EPA Integrated Risk Information Sytem(IRIS) likely to be carcinogenic to humans ( 11) National Toxicology Program (NTP) report on Carcinogens(Twelfth edition) (reasonably anticipated to be a human carcinogen). Diesel engine exhaust ( 12) ( 13). 표 10. IARC 에의한발암성분류 Group 1 2A 2B 3 IARC Classification Sufficient evidence in humans or sufficient evidence in animals and strong mechanistic data in humans Limited evidence in humans and sufficient evidence in animals Limited evidence in humans and less than sufficient evidence in animals Inadequate in humans and inadequate or limited in animals 4 Lack of carcinogenicity in humans and in animals

54 표 11. US EPA 에의한발암성분류 (US EPA, 1986*) Group A EPA Classification Human Carcinogen There is enough evidence to conclude that it can cause cancer in humans. B1 B2 C D Probable Human Carcinogen Probable Human Carcinogen Possible Human Carcinogen Not Classifiable as to Human Carcinogenicity There is limited evidence that it can cause cancer in humans, but at present it is not conclusive. There is inadequate evidence that it can cause cancer in humans but at present it is far from conclusive. There is limited evidence that it can cause cancer in animals in the absence of human data, but at present it is not conclusive. There is no evidence at present that it causes cancer in humans. E Evidence of Non-Carcinogenicity for Humans There is strong evidence that it does not cause cancer in humans. * (US EPA) Carcinogen to Human Likely to be carcinogenic to human Suggestive evidence of carcinogenity, but not sufficient to assess human carcinogenic potential Data are inadequate for an assessment of human carcinogenic potential

55 표 12. Diesel engine exhaust 의비발암독성지수 Toxic effects LOAEL a (mg/m 3 ) UF b RfC c ( μg /m 3 ) Reference Pulmonary inflammation and 0.46 histopathology Rat chronic inhalation study 0.144: 30 5 EPA, IRIS Ishinishi et al. (1988) HEC d Rat chronic inhalation study Ishinishi et al. (1988) ~ ~ 5.6 WHO a LOAEL: Low Observable Adverse Effective Level b UF: Uncertainty Factor c RfC: Reference Concentration/Inhalation RfC (Rat chronic inhalation study, Ishinishi et al., 1988) d Conversion Factors and Assumptions Human equivalent concentrations were derived using a mathematical model of diesel particulate matter (DPM) deposition and clearance (Yu et al., 1991), assuming that equal pulmonary surface loadings (in mg DPM/cm²) in rats and humans would be associated with similar effects. First, the model was used to estimate the loading in rats corresponding to the NOAEL, administered 16hr/day, 6 days/week, for 130weeks. Then the model was used to estimate the continuous exposure conditions for humans which would lead to the loading estimated in rats. In performing the modeling, rats were taken to weigh 300g, with a total pulmonary surface area of 4090cm². Human equivalent concentrations were derived using respiratory parameters for a 25 year-old human male having a total pulmonary surface area of 627,000 cm², tidal volume of 0.926L, respiratory frequency of 15 breaths/min, and total daily pulmonary volume of 20m³, with exposure assumed to last 70 years. For more details of the derivation of human equivalent concentrations, see the Support Document (U.S. EPA, 2002)

56 표 13. Diesel engine exhaust 의발암독성지수 Toxic effects Unit Risk ( μg /m 3 ) -1 Reference Lung cancer 6.0x10-4 (1.3x10-4 ~ 2.4x10-3 ) EPA Lung cancer 3.4x10-5 (1.6x10-5 ~ 7.1x10-5 ) WHO 2) 디젤미세먼지에흡착되어있는규제관리대상 PAH 및 nitro-pah 류 PAH carcinogenic group 14. IARC PAH , 2A, 2B, 3 ne(not evaluated) ( I). 16PAHs(EPA 610 PAH) EPA PAH 16. Li (2010) 16 Benzo(ghi)fluoranthene, cyclopenta(cd)pyrene benzo(e)pyrene. Ratcliff (2010) biphenyl, benzo(c)phenan- threne

57 표 14. 기관별 PAH 의 carcinogenic group (PAH position paper Annexes July 27th 2001)

58 표 15. PAH 류의발암성분류 ( 부록 I 참조 ) No. Compounds Groups Structure 1 Acenaphthylene n.e. 2 Acenaphthene 3 a 3 Anthranthrene 3 4 Anthracene 3 5 Benz(a)anthracene 2A b 6 Benzo(a)fluorene 3 7 Benzo(b)fluorene 3 8 Benzo(b)fluoranthene 2B c 9 Benzo(ghi)fluoranthene 3 10 Benzo(j)fluoranthene 2B

59 11 Benzo(k)fluoranthene 2B 12 Benzo(ghi)perylene 3 13 Benzo(c)phenanthrene 3 14 Benzo(a)pyrene 1 d 15 Benzo(e)pyrene 3 16 Chrysene 3 17 Chronene 3-18 Cyclopenta(cd)pyrene 2A 19 Dibenzo(a,h)anthracene 2A

60 20 Dibenzo(a,e)pyrene 2B 21 Dibenzo(a,h)pyrene 2B 22 Dibenzo(a,i)pyrene 2B 23 Dibenzo(a,l)pyrene 2A 24 Fluoranthene 3 25 Fluorene 3 26 Indeno(1,2,3-cd)pyrene 2B 27 5-Methylchrysene 2B

61 28 1-Methylphenanthrene 3 29 Naphthalene 2B 30 Perylene 3 31 Phenanthrene 3 32 Pyrene 3 33 Triphenylene 3 34 retene - ( 출처 : IARC, 1987) 2A ; 인체발암가능성높음, 2B ; 인체발암가능, 3 ; 인체비발암 a b c d

62 3) 디젤미세먼지에흡착되어있는규제관리대상 nitro-pah 류 USEPA (2002) nitroarene LD diesel particle extract ( 16). IARC ( 16, II). DOC PAH nitration (Heeb, 2010) nitro-pah( 16, II)). 표 16. 디젤미세먼지에서검출되는 Nitro-PAH 목록 No. Compounds Groups US EPA a IARC b others c Structure 1 1,3-dinitropyrene 3 o o - 2 1,6-dinitropyrene 2B o o - 3 1,8-dinitropyrene 2B o o - 4 2,7-dinitro-9-fluorenone - o

63 5 3,7-dinitrofluoranthene 2B - o - 6 3,9-dinitrofluoranthene 2B - o - 7 2,7-dinitrofluorene - o methyl-1-nitroanthracene - o Nitroacenaphthene 2B - - o 10 2-nitroanthracene 3 o nitroanthracene 3 o o 12 6-Nitrochrysene 2B o o 13 3-nitrobenzanthrene - o

64 14 7-Nitrobenz[a]anthracene 3 o 15 6-Nitrobenzo[a]pyrene 3 o o 16 4-nitrobiphenyl 3 o 17 1-nitrofluoranthene - o 18 3-nitrofluoranthene 3 o o 19 7-nitrofluoranthene o 20 8-nitrofluoranthene o 21 2-nitrofluorene 2b o o o 22 1-nitronaphthalene 3 o o

65 23 2-Nitronaphthalene 3 o o 24 3-Nitroperylene 3 o 25 3-nitrophenanthrene o o 26 9-nitrophenanthrene o o 27 1-nitropyrene 2B o o o 28 2-Nitropyrene 3 o 29 4-Nitropyrene 2B o o a USEPA, Health Assessment Document for Diesel Engine Exhaust. EPA/600/8-90/057F b 부록 II, IARC C Ratcliff (2010)

66 4) 디젤미세먼지에흡착되어있는규제관리대상금속류 표 17. 경유차량배출가스규제대상금속류목록 (listed by ARB) Metal / Trace element / Ions Cal EPA a US EPA b Other c Aluminum Al o o Antimony Sb o - - Arsenic As o - o Barium Ba o o Beryllium Be o - - Cadmium Cd o - o

67 Copper Cu o o Chromium Cr o - o Cobalt Co o - o Iron Fe o o Lead Pb o o o Manganese Mn o o o Mercury Hg o - - Nickel Ni o - o Selenium Se o - - Silicon Si o - Titanium Ti o - Vanadium V - o Zinc Zn o o Sodium J o o Magnesium Mg o o Sulfur [SO 4 2- ] S o o Calcium Ca o o Phosphorus P o o Potassium K o o Chlorine Cl o - - a Proposed identification of diesel exhaust as a toxic air contaminant, report to the Air Resources Board, as approved by the scientific review panel on April 22, 1998 b US EPA (2002) c Li et al., 2010;

68 4-2. 디젤엔진에서배출되는미세먼지및유해오염물질배출특성평가 배출가스저감장치가장착된소형경유자동차에서주행모드에따라 발생되는미세먼지 (PM2.5) 포집및포집량측정 1) 분진포집량및입자개수측정 EEPS PM2.5 cyclone sampler. 18 ETC ESC mg/hr ETC PM2.5. PM L/min(1.002m3 /hr) 4.63mg/ m mg/ m3. ETC ESC ETC. 표 18. 디젤배기가스중의입자개수및분진포집량 ETC 모드 ESC 모드 분진포집량 4.63 mg /hr 3.45 mg /hr** 흡입풍량 16.7 L/min (1.002 m3 /hr) 분진포집량 4.63 mg / m mg / m3 입자개수 (PN) 5.29 X X vs ETC mode, **p<

69 2) 분진사이즈측정 EEPS 18 ETC ESC 100nm. 92% 1μm (ARB, 1997). 50 ~ 200 m 2 /gram of soot(volkswagen, 1989). 그림 18. 디젤배기가스중의입자크기

70 포집된미세먼지중유기성물질추출 1) 디젤분진에흡착된 16 종의 PAH 류의분석 1μm.. (organic fraction) aldehydes, alkanes alkenes, aliphatic hydrocarbons, PAH PAH (Zielinska, 1990; Johnson, 1994). PAH (ETC, ESC) DCM 16 PAH GC-MASS. 11 PAH naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthrancene, fluoranthene, pyrene, benz(a)anthracene, benzo(a)pyrene benzo(g,h,i)perylene chrysene, benzo(b)fluoranthene, benzo(k)fluoranthene, dibenzo(a,b)anthracene indeno(1,2,3-cd)pyrene ( 19). 2) 디젤분진에흡착된 16 종 PAH 류에대한 PAH-TEQ 산정 PAH IARC ( 14) 12. PAH ETC ESC μg /g ESC. PAH (TEF) PAH PAH-TEQ ETC ESC μg-TEQ/g ETC. (ETC ESC) PAH ESC

71 TEF phenanthrene ETC TEF 1 benzo(a)pyrene PAH-TEQ ETC ( 19). 표 19. 디젤배기가스중의 PAH 류분석 Chemical Materials Carcinogen group a TEF ETC ESC ETC ESC ppm(mg/g) b(a)p-teq(mg/g) Naphthalene 2B Acenaphthylene ne b Acenaphthene Fluorene Phenanthrene Anthrancene Fluoranthene Pyrene Benz(a)anthracene 2A Benzo(a)pyrene Benzo(g,h,i)perylene Chrysene ND ND ND ND Benzo(b)fluoranthene 2B 0.1 ND ND ND ND Benzo(K)fluoranthene 2B 0.1 ND ND ND ND Dibenzo(a,h)anthracene 2A 1 ND ND ND ND Indeno(1,2,3-cd)pyrene 2B 0.1 ND ND ND ND PAHs a PAHs toxic equivalency factor with respect to BaP (Nisbert and LaGoy, 1992) b ne: not evaluated

72 기타인체위해성관련유해물질분석 1) 가스상물질분석 6 ETC CO, THC(total hydrocarbon) CO , g/kwh ESC 0.64, g/kwh. NOx ESC ETC ( 20). 표 20. 디젤배기가스중의휘발성물질농도 단위 (g/kwh) ETC ESC CO 1.63± ±0.11** THC 0.10± ±0.02** NOx 10.80± ±0.18** CO ± ±8.17** vs ETC mode, **p<

73 2) 휘발성물질 (VOCs) 분석 1,3-Butadiene, Benzene, Toluene, Ethylbenzene, styrene m,p-xylene 21 Syrene ETC ESC 1,2-Butadiene Benzene, Toluene, Ethylbenzene m,p-xylene ETC ESC. Toluene Okada (2011) photochemical oxidant ETC ESC 6. 1,3-Butadiene benzene EPA A group( 22) toluene. RfD RfC 23. IARC EPA ( 22) UNIT RISK 24. 표 21. 디젤배기가스중의휘발성물질농도 Chemicals ETC ESC mg/kwh 1,3-Butadiene ±0.38 Benzene 2.06± ±0.04 Toluene 17.62± ±0.42 Ethylbenzene 4.01± ±0.09 m,p-xylene 3.44± ±0.08 Stylene ND ND o-xylene 2.46± ±

74 표 22. 디젤배기가스중의휘발성물질농도 Chemicals Structure Carcinogenic groups IARC a 표 10 USEPA/IRIS b 표 11 1,3-Butadiene 1 Carcinogenic to humans Benzene 1 A Human carinogenic Toluene 3 Inadequate information to assess carcinogenic potential Ethylbenzene 2B D Not classifiable as to human carcinogenicity o,m,p-xylene 3 inadequate for an assessment of human carcinogenic potential Stylene 2B not data a b

75 표 23. 디젤배기가스중휘발성물질 (VOC) 에대한비발암독성정보 Chemicals Critical Effects POINT OF DEPARTURE UF RFC or RFD 1,3-Butadiene RfC (mg/m 3 ) Chronic Inhalation Exposure (2 year) Ovarian atrophy BMCL 10 (HEC) RfD (mg/kg-day) - Benzene RfC (mg/m 3 ) RfD (mg/kg-day) Toluene Decreased lymphocyte count Decreased lymphocyte count BMDL RfC (mg/m 3 ) Neurological effects occupationally exposed workers NOAEL RfD (mg/kg-day) Increased kidney weight BMDL Ethylbenzene RfC (mg/m 3 ) Developmental toxicology NOAEL(HEC) RfD (mg/kg-day) Liver and kidney toxicity NOEL Xylene RfC (mg/m 3 ) Impaired motor NOAEL(HEC) 39 RfD (mg/kg-day) Decreased body NOAEL weight, increased mortality Styrene RfC (mg/m 3 ) CNS effects NOAEL(HEC) RfD (mg/kg-day) Red blood cell and liver effects NOAEL

76 표 24. 디젤배기가스중휘발성물질 (VOC) 에대한발암독성정보 Chemicals Critical Effects Inhalation Unit risk(s) (per μg /m 3 ) Oral Slope Factor(s) (per mg/kg-day) 1,3-Butadiene Leukemia Human carcinogen Benzene Leukemia Human carcinogen ~ ~ Drinking water unit risks ~ (per μg /L) Toluene Inadequate information to assess carcinogenic potential Ethylbenzene Group D: Not classifiable as to himan carcinogenicity Xylene Inadequate for an assessment of human carcinogenic potential Styrene Not assessed under the IRIS program 2) PM2.5 에흡착된중금속분석 ICP 25 Al, Cd, Cr, Cu, Fe, Mn, Ni, Zn Cd ETC Cr ETC ESC (**p<0.01). Li (2010) Na, Mg, Al, P, S, K, Ca, Fe Fe, Zn, Pb, Cd Ba

77 . Jalava (2010) Fe Zn Cd Pb. DOC/POC catalytic converter. RfD/C UNIT RISK SLOPE FACTOR 26. 표 25. 디젤분진에흡착되어있는금속류측정 단위 (mg/l) ETC ESC Al ± ± Cd 0.05 ± 0.10 n.c. Cr 2.30 ± ± 1.81** Cu 7.55 ± ± 0.66 Fe ± ± Mn 0.90 ± ± 0.26 Ni 1.80 ± ± 0.98 Zn ± ±

78 표 26. 디젤배기가스중금속류에대한비발암독성정보 Chemicals Critical Effects POINT OF DEPARTURE UF RFC or RFD Cadmium (Cd) RfC (mg/m 3 ) Not assessed under the IRIS program RfD (mg/kg-day) Significant proteinuria NOAEL Chromium(CrIII) RfC (mg/m 3 ) Not assessed under the IRIS program RfD (mg/kg-day) No effects observed NOAEL(ADJ) Chromium(CrVI) RfC (mg/m 3 ) Nasal septum atrophy (Chromic acid mists NOAEL(ADJ) and dissolved CrIV aerosols) RfC (mg/m 3 ) Lactate dehydrogenase in bronchioalveolar BMC 10 (ADJ) lavage fluid (CrIV particulates) RfD (mg/kg-day) Non reported NOAEL(ADJ)

79 Copper (Cu) RfC (mg/m 3 ) Not assessed under the IRIS program RfD (mg/kg-day) Not assessed under the IRIS program Manganese (Mn) RfC (mg/m 3 ) Impairment of neurobehavioral NOAEL(HEC) function (other 0.05 effect: impairment of neurobehavioral function) RfD (mg/kg-day) CNS effects (other effect: Impairment of NOAEL neurobehavioral 0.14 function) Nickel (Ni) RfC (mg/m 3 ) - RfD (mg/kg-day) Decreased body and NOAEL organ weights 5 Zinc (Zn) RfC (mg/m 3 ) Information reviewed but value not estimated RfD (mg/kg-day) Decreases in erythrocyte Cu, NOAEL Zn-superoxide 0.91 dismutase (ESOD) activity in healthy adult male and female volunteers

80 표 27. 디젤배기가스중금속류에대한발암독성정보 Chemicals Critical Effects Inhalation Unit risk(s) (per μg /m 3 ) Oral Slope Factor(s) (per mg/kg-day) Cadmium (Cd) Lung, trachea, bronchus cancer B1 probable human carcinogen Chromium(CrIII) Group D: Not classifiable as to himan carcinogenicity Chromium(CrVI) Lung cancer A (Human carcinogen inhalation) D (oral route) Copper (Cu) Group D: Not classifiable as to himan carcinogenicity Manganese(Mn) Group D: Not classifiable as to himan carcinogenicity Nickel (Ni) Not assessed under the IRIS program Zinc (Zn) Group D: Not classifiable as to himan carcinogenicity

81 4-3. 경유엔진에서배출되는미세먼지에의한호흡기관련인체 위해성평가 인체위해성관련표적세포등을활용한 유전적 (Genetic) 및후생유전적 (Epigenetic) 평가 1) 미세먼지유기성추출물에의한유전독성평가 ➀ 대표유전독성평가세포인 CHO-K1 세포에서의유전독성평가 DNA 손상 (DNA breakage) ETC ESC DCM chinese hamster ovary cell (CHO-K1 cell) DNA comet assay. Comet assay OECD guideline draft DNA. comet assay 19 H 2 O 2 (**p < 0.01) DNA ETC ESC (**p < 0.01) DNA. ETC ESC DNA ( 19)

82 35 Olive tail moment ** ** ** 5 0 Con H2O2 H 2 O 2 ETC ESC Sample (100mg/ml) 그림 19. 운전모드 (ETC 모드, ESC 모드 ) 에따른 DNA 손상평가

83 CHO-K1 세포에서의염색체이상 ( 소핵형성, micronucleus formation) ETC ESC DCM chinese hamster ovary(cho-k1) cell micronucleus test. 20 MMS(Methylmethanesulfonate) (**p < 0.01) ETC ESC (**p < 0.01). DNA ETC ESC ( 20). 50 MN formation (fold induction) ** ** 0 Con ETC-50 ESC-100 Sample (50 or 100mg/ml) 그림 20. 운전모드 (ETC 모드, ESC 모드 ) 에따른 chromosome 손상

84 ➁ 정상폐세포인 BEAS-2B 세포에서의유전독성평가 DNA 손상 (DNA breakage) ETC ESC BEAS-2B DNA. 21 (H 2 O 2 ), ETC ESC DNA ETC ESC (**p < 0.01) DNA. 80 ** Oilve tail moment ** ## ** 0 Con H2O2 ETC ESC Sample (100mg/ml) 그림 21 운전모드 (ETC 모드, ESC 모드 ) 에따른 DNA 손상평가

85 CHO-K1 세포에서의염색체이상 ( 소핵형성, micronucleus formation) MMS(Methyl methanesulfonate) (**p < 0.01) ETC ESC (**p < 0.01) ( 22) ETC ESC ( 22). Micronucleus formation (#/1,000 cell) Positive mutagens ** ** ** ** ** ** Con ETC-10 ETC-50 ETC-100 ESC-10 ESC-50 ESC-100 그림 22. 운전모드 (ETC 모드, ESC 모드 ) 에따른소핵형성

86 ➂ IN VITRO SYSTEM 을이용한유전독성평가에서의양성판정 23 in vitro in vivo scheme mutagenicity in vitro. in vitro in vitro micronucleus test comet assay. in vitro micronucleus test DNA comet assay. 그림 23. 유전독성평가의양성판정

87 2) 미세먼지유기성추출물에의한후생유전독성평가 DNA p53 DNA. p53 p53 H1299 p53 H460 ETC ESC. 24 ETC ESC. p53. BEAS-2B P53 pifithrin δ apoptosis staurosporine apoptosis 90μM 1 ( 25). 26 ETC apoptosis DNA fragmentation BEAS-2B [p53 wild type, pifithrin α(-)]. p53 pifithrin α BEAS-2B ETC apoptosis. apoptosis p53. apoptosis p

88 40 MN formation (#/1,000 cells) Con ETC CON ETC P53 null- H1299 cell p53 wild- H460 cell 40 MN formation (#/1,000 cells) Con ESC CON ESC P53 null- H1299 cell p53 wild- H460 cell 그림 24. p53-null H1299 세포및 p53-wild H460 세포에서소핵형성확인

89 (A) pifithrin α 농도선정 (90μM) (B) pifithrin α 노출시간선정 (1hr) 그림 25. Pifithrin α 처리농도선정 10 DNA Fragmentation (Fold induction) Con ETC CON ETC P53 (-) pifithrin (+) p53 (+) pifithrin (-) 그림 26. Pifithrin α 에의해 p53 이결여된 BEAS-2B 세포 에서세포사멸 (apoptosis)

90 2) in vitro 실험결과에따른 in vivo 독성예측 in vitro 100μg /ml. 100μg /ml (cm 2 ) 28 ETC ESC 44.44μg /cm 2 20μg /cm 2. BEAS-2B bronchiol( ) ETC ESC (cm 2 ) 44.44μg 20μg. 표 28. 표면적당노출농도 Comet assay Micronucleus test Max. Effect Conc. ( μg /ml) Well volume ( μl ) PM2.5 ( μg /well) Well surface area (cm 2 ) PM2.5 conc ( μg /cm 2 ) In vitro 노출농도에대한인체위해성평가적용예 : Li (2003) ~ 20μm 79μg /m 3 24 nasopharyngeal(npr, ), tracheobroncial (TBR, ) alveolar region(avr, ) 204, μg /cm 2 /24hr

91 . 100μg /ml 20 ~ 44μg /cm 2. in vivo 경유자동차미세먼지에의한인체위해성관련연구를통한정책활용 1) 외국에서의호흡기및급 만성인체위해성평가자료활용을통한정책반영사례 ➀ 디젤미세입자관련국외관련자료 Diesel engine exhaust NTP 12th Report on Carcinogens for Diesel Exhaust Particulates (2011) (PDF) 2. IARC monograph summary, Volume 46 (1989) (PDF) 3. EPA, Health Assessment Document for Diesel Engine Exhaust (2002) 4. Health Canada: It s Your Health Road Traffic and Air Pollution (2004) (PDF) 5. Government of Canada, Sulphur in Diesel Fuel Regulations (SOR/ ): (2006) 6. Government of Canada, On-Road Vehicle and Engine Emission Regulations

92 (SOR/2003-2) 7. Government of Canada, Off-Road Small Spark Ignition Engine Emission Regulations (SOR/ ) 8. Government of Canada, Off-Road Compression Ignition Engine Emission Regulations (SOR/ ) 9. Government of Canada, Marine Spark-Ignition Engine, Vessel and Off-Road Recreational Vehicle Emission Regulations (SOR/ ) 10. INCHEM, Environmental Health Criteria Document 171, Diesel Fuel and Exhaust Emissions (1996) 11. Health Canada, Diesel Research Workshop (2002) 12. Health Canada, Canadian Handbook on Health Impact Assessment, Volume 4: Health Impacts by Industry Sector (2004) (PDF) 13. Health Canada, Air Quality Health Effects (2006) 14. Priority Substances List Assessment Report (CEPA) Respirable Particulate Matter Less Than or Equal to 10 µm (PDF) 15. Health Canada: It s Your Health, Smog and Your Health (2003) (PDF) 16. EPA, Integrated Risk Information System for Diesel Engine Exhaust (2008) 17. NIOSH, Mining and Safety Health Topic: Diesel Engine Exhaust (2009) 18. Health Canada, Evaluation of the Levels of Diesel-related Pollutants on School Buses During the Transportation of Children Executive Summary December (2005) (PDF) ➁ 미세입자국내외규제관리및제도 PM2.5 국내 PM

93 1995 PM2.5 PM10. PM2.5 (PM2.5) PM PM μg, 25μg 3. 미국 ( 미국연방정부규제 ) 1955 [ ] [ ] 1970 [ ] (EPA) PM2.5. EPA CASAC(Clean Air Scientific Advisory Committee) PM10 PM2.5 PM2.5(PM2.5; 24 35μg /m 3 ) PM10 (PM10-PM2.5; 24 70μg /m 3 ). 미국캘리포니아주 (South Coast Air Quality Management SCAGMD)

94 PM10 AQMP(Air Quality Management Program). 유럽연합 (EU) (Clean Air Strategy) PM PM2.5 20μg / m3 PM2.5 25μg / m3. 캐나다 PM PM2.5 CWS(Canada-wide standards) CCNE(Council of Canadian Ministers of the Environment). CWS PM μg / m3, 8μg / m3 100,000. PM2.5. 호주 PM2.5 PM2.5 (, ). 2 (SOA, secondary organic aerosol) VOC NOx PM2.5. ➂ 위해성평가방법에따른위해도평가및규제목표치산정 위해성평가

95 ,. ( ) [ (Risk) = (Hazard) (Exposure)]. (risk assessment) (hazard identification), - (dose-response assessment), (exposure assessment) (risk characterization) 4. (risk management) ( 27). 그림 27. 위해성평가과정

96 위해성평가단계 -. [ ].. 위해성평가계획수립 :,, 기초현황을조사. 평가대상물질을선정 노출량측정계획을수립. 유해성시험 ( 독성시험 ) 에대한계획을수립. 유해성확인 (hazard identification):,,. 발암성물질인지비발암성물질인지를분류. - (No observed adverse effect level; NOAEL). US EPA ( 11) (International Agency for research on Cancer; IARC) ( 12) / /

97 . -,. OECD (Organization for Economic Cooperation and Development), UNEP (United Nations Environment Programme), JECFA (Joint FAD/WHO Expert Comminittee on Food Additives), US EPA IRIS(Integrated Risk Information System).. in vitro.. 용량 - 반응평가 (Dose-response assessment):.. -. (Exposure) : - (, reference Dose, RfD)

98 . 1 (acceptable daily intake, ADI) 1 (tolerable daily intake, TDI). (RfD) (Reference Concentration, RfC). (no observable adverse effect level; NOAEL) (uncertainty factor) ( 1). margin of exposure(moe) NOAEL. MOE - BMD (Benchmark dose). BMD LED10 (lower bound of effective dose, 10%) LED10 - (95% ) 10%. or ( 1) UF: Uncertainty Factor(,,, /,,, ), 100. MF: Modifying Factor( )

99 . 1~ :. 20. N (unit risk) (cancer slope factor). 노출평가 (Exposure assessment):,.,,. - :,, :,,,, :,,,,, (,, ),,,,,

100 - :,, - :,,..,, 1 ( 2). 총노출량 일생동안의일일평균노출량 평균체중 평균수명 ( 2) 총노출량 오염물질농도 접촉율 or 노출기간 흡수분율 (mg/ m3 ), (mg/l), (mg/kg). :,,. ( 15~20 m3 /day), (1.5~2L/day), (kg/day, ). :. 100% 위해도결정 (Risk Characterization):.,. - : (Hazard index) ( 3)

101 (mg/kg/day) (mg/kg/day) 인체노출량 위해지수 기준노출량 ( 3) - 1.,,. ( 4). 총위해지수 ( 4) - OECD (margin of safety margin of exposure). (MOS) POD(point of departure) 1. POD 95% bechmark dose( LED 10 ). - : (individual risk) (population risk). - (q1*) (mg/kg/day) ( 5).,

102 . 개인초과발암위해도 인체노출량 발암력 현오염도수준 단위위해도 ( 5) , 10-6, , ~10-4. ( 6). 인구집단위해도 개인초과발암위해도 노출인구수 ( 6) - (RfD) 1 ( 7) ( 2). 인체노출량 위해지수 기준노출량 ( 7) 총노출량 일생동안의일일평균노출량 평균체중 평균수명

103 총노출량 오염물질농도 접촉율 or 노출기간 흡수분율 - unit risk slope factor 1 ( 8) 개인초과발암위해도 인체노출량 발암력 ( 8) 현오염도수준 개인초과발암위해도 단위위해도 2) 미세먼지인체위해성관련외국의연구동향분석을통한국내적용성제시 Wang (2011) Okada (2001) urban surface dust PAH. PAH. ➀ 디젤미제먼지에흡착된 16 종 PAH 류의 PAH-TEQ 에대한인체발암위해성평가 위해성평가계획수립

104 PAH 16 GS-MASS PAH naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthrancene, fluoranthene, pyrene, benz(a)- anthracene, benzo(a)pyrene benzo(g,h,i)perylene 5 PAH chrysene, benzo(b)fluoranthene, benzo(k)fluoranthene, dibenzo(a,b)anthracene indeno(1,2,3-cd)pyrene.. PAH.. US EPA, IARC, Wang (2011). - Wang (2011) PAH PAH PAH Benzo(a)pyrene(BaP) bio-teq. 19 PAH BaP. PAH (inhalation). 유해성확인 (Hazard Identification) - PAH ( ). 19 PAH IARC. PAH Benzo(a)pyrene group

105 benzo(a)- anthracene naphthalene 2B group. fluorene, phenanthrene, anthracene, fluoranthene, pyrene benzo(g,h,i)perylene group 3. PAH. 용량 - 반응평가 (Dose-Response Assessment) - PAH Risk assessment CSF carcinogenic slope factor [CSF (mg/kg/day) -1 ] Peng (2011). 29 CSF 3.85(mg/kg/day) -1 (Peng, 2011). 노출평가 (Exposure Assessment) - 11 PAH ETC ESC μg /g PAH (TEF) PAH PAH-TEQ. ETC ESC μg-TEQ/g.. 19 PAH-TEQ ( μg-TEQ/g) (4.63mg/ m3 3.45mg/ m3 ) mg/ m mg/ m3.. - ( 9)

106 20m3 /day % 70kg ETC ESC mg/kg/day mg/kg/day. 일일평균노출량오염물질농도 접촉율 노출기간 흡수분율 평균체중 평균수명 ( 9) 표 29. Lifetime cancer risk assessment 에사용되는 parameter 노출변수단위값 오염물질농도 mg/m (ETC) (ESC) 접촉율 (IR inhalation ) m 3 /day 20 노출기간 ( 노출빈도 노출기한 ) day/year year 평균체중 kg 70 평균수명 day 일일평균노출량 mg/kg/day (ETC) (ESC)

107 위해도결정 (Risk Characterization) -. (Cancer slope factor) ( 10). PAH CSF 3.85(mg/kg/day) -1 (Peng, 2011). ETC ESC mg/kg/day mg/kg/day CSF 3.85(mg/kg/day) ETC ESC. 개인초과발암위해도 일일평균노출량 발암력 ( 10) 결과해석에대한제한성 - PAH,

108 4-4. 결론 100~135μg / m3, 150~350μg / m3 15 (Yi, 2010) (PM2.5) 10μg / m3 1.24(95% CI, 1.09~3.52) (Miller et al., 2008), 6% (McCreaner et al.,2007). PAH. p53 apoptosis 경유차량에서배출되는배출가스의배출특성조사 디젤미세분진포집및특성조사 : EEPS PM2.5 cyclone sampler. ETC ESC 4.63mg/ m3 3.45mg/ m ETC. ETC ESC 100nm

109 가스상물질 : CO, THC(total hydrocarbon), CO 2 NOx ETC CO, THC(total hydrocarbon) CO , g/kwh ESC 0.64, g/kwh. NOx ESC ETC. 휘발성물질 (VOC): 1,3-Butadiene, Benzene, Toluene, Ethylbenzene, m,p-xylene 1,2-Butadiene Benzene, Toluene, Ethylbenzene m,p-xylene ETC ESC. Toluene photochemical oxidant (Okada (2011) 1,3-Butadiene benzene A group(usepa). 배출미세먼지에흡착되어있는금속류측정 : ICP Al, Cd, Cr, Cu, Fe, Mn, Ni, Zn Cd ETC Cr ETC ESC (**p<0.01) 경유차량에서배출되는미세먼지유기성추출물의독성평가 p53 apoptosis. 미세먼지유기성추출물의유전독성 (Genotoxicity) 평가 :

110 CHO-K1 cell BEAS-2B cell comet assay micronucleus test 100μg /ml DNA ( ). ETC ESC. 미세먼지유기성추출물의후생유전독성 (epigenetic toxicity) 평가 : p53 H460 H1299 p53 pifithrinα BEAS-2B (apoptosis) ETC ESC. p53 p53- apoptosis. 100μg /ml DNA, (apoptosis). 100μg /ml (cm 2 ) ETC ESC μg /cm 2 20 μg /cm 2. BEAS-2B bronchiol( ) ETC ESC (cm 2 ) μg 20 μg 경유차량에서배출되는 PAH 류의인체발암위해성평가 (AC, HORIBA )

111 ETC ESC (PM2.5) PAH. PAH PAH naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthrancene, fluoranthene, pyrene, benz(a)anthracene, benzo(a)pyrene benzo(g,h,i)perylene. PAH. PAH PAH BaP PAH-TEQ ( μg-TEQ/g) mg/ m3 (ETC ) mg/ m3 (ESC ) mg/kg/day mg/kg/day CSF 3.85 (mg/kg/day) -1 ETS ESC ETC ESC PAH ETC. 일일평균노출량오염물질농도 접촉율 노출기간 흡수분율 평균체중 평균수명 개인초과발암위해도 일일평균노출량 발암력

112 위해성평가결과의제한성 : (PM2.5). (DPF+DOC),, 경유차량에서배출되는배출규제물질목록작성 (PM2.5) 23% (USEPA, 2000).. (PAH, nitro-pah, ). PAH 34, nitro-pah

113 경유차량에서배출되는 PAH 류에대한규제목록 No. Compounds No. Compounds 1 Acenaphthylene 18 Cyclopenta(cd)pyrene 2 Acenaphthene 19 Dibenzo(a,h)anthracene 3 Anthranthrene 20 Dibenzo(a,e)pyrene 4 Anthracene 21 Dibenzo(a,h)pyrene 5 Benz(a)anthracene 22 Dibenzo(a,i)pyrene 6 Benzo(a)fluorene 23 Dibenzo(a,l)pyrene 7 Benzo(b)fluorene 24 Fluoranthene 8 Benzo(b)fluoranthene 25 Fluorene 9 Benzo(ghi)fluoranthene 26 Indeno(1,2,3-cd)pyrene 10 Benzo(j)fluoranthene 27 5-Methylchrysene 11 Benzo(k)fluoranthene 28 1-Methylphenanthrene 12 Benzo(ghi)perylene 29 Naphthalene 13 Benzo(c)phenanthrene 30 Perylene 14 Benzo(a)pyrene 31 Phenanthrene 15 Benzo(e)pyrene 32 Pyrene 16 Chrysene 33 Triphenylene 17 Chronene 34 retene

114 경유차량에서배출되는 Nitro-PAH 류에대한규제목록 No. Compounds No. Compounds 1 1,3-dinitropyrene 16 4-nitrobiphenyl 2 1,6-dinitropyrene 17 1-nitrofluoranthene 3 1,8-dinitropyrene 18 3-nitrofluoranthene 4 2,7-dinitro-9-fluorenone 19 7-nitrofluoranthene 5 3,7-dinitrofluoranthene 20 8-nitrofluoranthene 6 3,9-dinitrofluoranthene 21 2-nitrofluorene 7 2,7-dinitrofluorene 22 1-nitronaphthalene 8 2-methyl-1-nitroanthracene 23 2-Nitronaphthalene 9 5-Nitroacenaphthene 24 3-Nitroperylene 10 2-nitroanthracene 25 3-nitrophenanthrene 11 9-nitroanthracene 26 9-nitrophenanthrene 12 6-Nitrochrysene 27 1-nitropyrene 13 3-nitrobenzanthrene 28 2-Nitropyrene 14 7-Nitrobenz[a]anthracene 29 4-Nitropyrene 15 6-Nitrobenzo[a]pyrene

115 경유차량에서배출되는금속류, 미량원소및이온류에대한규제목록 No. Compounds No. Compounds 1 Aluminum Al 14 Nickel Ni 2 Antimony Sb 15 Selenium Se 3 Arsenic As 16 Silicon Si 4 Barium Ba 17 Titanium Ti 5 Beryllium Be 18 Vanadium V 6 Cadmium Cd 19 Zinc Zn 7 Copper Cu 20 Sodium J 8 Chromium Cr 21 Magnesium Mg 9 Cobalt Co 22 Sulfur [SO 4 2- ] S 10 Iron Fe 23 Calcium Ca 11 Lead Pb 24 Phosphorus P 12 Manganese Mn 25 Potassium K 13 Mercury Hg 26 Chlorine Cl

116 5. 기대성과및활용방안 5-1. 기대성과 < 기술적측면의기대성과 > < 경제 산업적측면의기대성과 > 5-2. 활용방안

117 6. 참고문헌 André V, Billet S, Pottier D, Le Goff J, Pottier I, Garçon G, Shirali P, Sichel F. Mutagenicity and genotoxicity of PM2.5 issued from an urbano-industrialized area of Dunkerque (France). J Appl Toxicol Mar;31(2): ARB (1998) Report to the air resources board on the proposed identification of diesel exhaust as a toxic air contaminant. Part A Exposure assessment, As approached by the scientific review panel on April 22, 1998, ARB (Air Resources Board) 1997 Emission Inventory Technical Support Division, October Diesel Particulate Matter Health Risk Assessment Study for the West Oakland Community: Preliminary Summary of Results, Ferreira-Baptista L, De Miguel E, Geochemistry and risk assessment of street dust in Luanda, Angola: a tropical urban environment. Atmos Environ 2005; 39: Gilman P. et. al., Assessment Document for Diesel Engine Exhaust, EPA, 600,8-90, Greg Michalak, "Diesel Emissions in Fire Stations", Grose M., Sakurai H., Savstrom J., Stolzenburg M.R., Watts W.F., Morgan C.G., Murray I.P., Twigg M.V., Kittelson D.B., McMurry P.H., Chemical and Physical Properties of Ultrafine Diesel Exhaust Particles Sampled Downstream of a Catalytic Trap, Environ. Sci. Technol., 40(17), , Gualtieri M, Longhin E, Mattioli M, Mantecca P, Tinaglia V, Mangano E,

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