Journal of Environmental Health Sciences (한국환경보건학회지)

Korean Society of Environmental Health (한국환경보건학회)
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Synthesis and Isolation of Monoacetyl-DCB and Diacetyl-DCB from 3,3대-dichlorobenzidine(DCB)

디클로로벤지딘으로부터 대사물질의 합성과 분리방법에 대한 연구

  • Lee, Jin-Heon (Department of Environmental Education, Kongju National University) ;
  • Lee, Beom-Gyu (Division of Physics and Chemistry, Chosun University)
  • Published : 2003.06.01
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Abstract

3,3-dichlorobenzidine is suspected to be cancinogenic in experimental animal and human. Several studies have investigated excretion of metabolites in urine, hemoglobin adduction and cancer incidence among workers occupationally exposed to 3,3'-dichlorobenzidine. In these researches, metabolites of 3,3'-dichlorobenzidine had a very important role, and were required as highly purity. The purpose of this study was synthesis and isolation of its metabolites from 3,3'-dichlorobenzidine. 3,3'-dichlorobenzidine was partially dissolved in benzene, ether, ethanol and methanol, and completely dissolved in 70% acetic acid on mixtures of citric acid containing less than 1% DCB, pyridine, a mixture of 0.5N NaOH and toluene(1:2), and phenol saturated with 20 mM TRIZA base. DCB, monoacetyl-DCB and diacetyl-DCB were measured by using gas chromatography/mass spectrometry(GC/MS). Detection for checking them was nitrogen phosphorous detection mode(NPD), and for identifying them was selected ion monitoring mode(SIM). The base peaks were 252 m/z in DCB, 252, and 294 m/z in monoacetyl-DCB, and 252, 294 and 336 m/z in diacetyl-DCB, respectively. Diacetyl-DCB was synthesized by titrating DCB solution of pyridine with sufficient acetyl chloride. Precipitation was diacetyl-DCB, which was purity of 98.7%. And its supernatant was composed of DCB, monoacetyl-DCB and diacetyl-DCB. By using acetic acid as controller of acetylation, monoacetyl-DCB was isolated from diacetyl-DCB . And residual pyridine was removed by using acetone. The purity of monoacetyl-DCB was 98.8%.

3,3'-디클로로벤지딘(DCB)는 실험동물에 발암물질로 밝혀졌고, 사람에게 암을 유발시킬 수 있는 발암물질로 의심되고있다. 많은 연구자들이 사업장에서 DCB에 폭로된 근로자들을 대상으로 뇨중에 배설된 대사물질, 헤모글로빈 부가체, 그리고 암 발생율 등에 대하여 연구를 하고 있다. 이러한 연구를 하기 위해서는 표준물질로 되어 있는 DCB의 대사물질이 꼭 필요하다. 따라서 본 연구의 목적은 DCB를 이용하여 이들의 대사물질을 합성하여 표준물질로 사용코자 합니다. DCB는 벤젠. 에테르, 에탄올, 메탄올 등에 부분적으로 용해되지만, 구연산이 1 % 이하로 함유된 70% 아세트산, 피리딘, 0.1N NaOH와 톨로엔이 1:2로 섞인 혼합물, 20 mM TRIZA염으로 포화된 페놀 등에는 완전히 용해되기 때문에 본 연구에서는 DCB를 피리딘에 녹여서 사용하였다. DCB와 대사물질인 mono-acetyl-DCB 및 diacetyl-DCB는 가스크로마노그래피(GC/MS)로 분석하였고, 검출기는 NPD와 SIM를 사용하였다. DCB의 기본피크는 252 m/z이었고, mono-acetyl-DCB의 기본피크는 252와 294 m/z로 구성되어 있었으며. diacetyl-DCB의 기본피크는 252, 294, 336 m/z로 구성되어 있었다. Discetyl-DCB는 피리딘에 용해된 DCB에 염소아세틸를 충분히 적정하여 합성하였다. 이렇게 얻은 diacetyl-DCB의 순도는 98.7%이었다. 침전물위에 있는 용해물질 속에는 DCB. mono-acetyl-DCB, diacetyl-DCB가 함유되어 있었는데, 아세트산을 아세틸화를 조절하는 물질로 사용하여 DCB를 모두 아세틸화시키었고, diacetyl-DCB로부터 mono-acetyl-DCB를 분리하여 추출하였다. 추출된 mono-acetyl-DCB는 아세톤으로 세척하여 98.8%의 순도를 얻었다.

Keywords

References

  1. Sagelsdoff, P.. Haengg. R., Heuberger, B., JoppichKuhn, R., Jung, R., Weideli, H. J. and Joppich, M. : Lack of bioavailability of dichlorobenzidine from diarylide azo pigments : molecular dosimetry for hemoglobin and DNA adducts, Carcinogenesis, 17(3). 507-514, 1996 https://doi.org/10.1093/carcin/17.3.507
  2. IARC. : Monographs of the carcinogenic risk of chemicals to humans: some industrial chemicals and dyestuffs. 3,3'-dichlorobenzidine and its hydrochloride. Int. Agency Res. Cancer, 29, 239-256, 1982b https://doi.org/10.1002/ijc.2910290303
  3. Rinde. E. and Troll, W. : Metabolic reduction of benzidine azo dyes to benzidine in rhesus monkey, J. Natl. Cancer Inst., 45, 181-182, 1975
  4. IARC. : 3,3'-Dichlorobenzidine and its hydrochloride, Monographs on the carcinogenic risk of chemicals to humans. Internal Agency for Research on Cancer. Lyon, France, 29, 239-256, 1982a
  5. Kadlubar, F. F., Miller, J. A. and Miller, E. C. : Hepatic microsomal N-glucuronidation and nucleic acid binding of N-hydroxy arylamines in relation to urinary bladder carcinogenesis, Cancer Res., 37, 805-814, 1977
  6. Joppich-Kuhn, R.. Hanggi, R., Sagelsdorff, P., Smith, A. E., Weideli, H. J. and Joppich, M. : Determination of dichlorobenzidine-hemoglobin adducts by GC/MS-NCI Int. Arch. Occup. Environ. Health, 69, 240-246, 1977 https://doi.org/10.1007/s004200050142
  7. Zwirner-Baier, I. and Neumann, H. G. : Biomonitoring of aromatic V:acetylation and deacetylation in the metabolic activation of aromatic amines as determined by hemoglobin binding, Arch. Toxicol., 72, 499-504, 1998 https://doi.org/10.1007/s002040050534
  8. Decad, G. M., Snyder, C. D. and Mitoma, C. : Fate of water insoluble and water soluble dichlorobenzidine-based pigments in Fscher 344 rats, J. Toxic. Environ. Health, 11, 455-465, 1983 https://doi.org/10.1080/15287398309530358
  9. Kellner, H. M., Christ, O. E. and Lotzsch, K. : Animal studies on the kinetics of benzidine and 3,3'-dichlorobenzidine, Arch. Toxikol., 34, 61-79, 1973
  10. Iba, M. M. : Effect of acute 3,3'-dichlorobenzidine administration on rat hepatic enzymeic and nonenzymic microsomal lipid peroxidation and antioxidant status, Res. Commun. Chem. Pathol. Pharmacol. 56(2), 243-252, 1987
  11. Iba, M. M. and Thomas, P. E. : Activation of 3,3'-dichlorobenzidine in rat liver microsomes to mutagens : involvement of cytochrome P-450d. Carcinogenesis 9(5), 717-723, 1988 https://doi.org/10.1093/carcin/9.5.717
  12. Morales, R., Rappaport, S. M. and Hermes, R. E. : Air sampling and analytical procedures for benzidine, 3,3'-dichlorobenzidine, and its salts, Am. Ind. Hyg. Assoc. J., 40, 970-978, 1978
  13. Roberts, E. and Rossano, A. J. : A sensitive colorimetric determination of 3,3-dichlorobenzidine in urine, Am. Ind. Hyg. Assoc. J., 43, 80-83, 1983