以往因受限於分析技術的不足，文獻中鮮少有報導能直接測得活體動物體內砷物種動態的變化情形，而長久以來，砷物種在活體動物體內的轉換代謝機轉，又一直是生物醫學界亟欲窺探的重要資訊。為達到現場（in-situ）進行活體動物體內（in-vivo）連續（continuous）監測砷物種動態（dynamic）變化的目的，本研究已成功地建立二套微型化（Miniaturized）microdialysis-microbore HPLC-ICP-MS與microdialysis-microbore HPLC-HG-ICP-MS之線上（on-line）分析技術，並實際針對活體動物體內三氧化二砷代謝過程中，砷物種的動態（dynamic）變化趨勢提供了直接的證據。
為開發適合上述連續監測活體動物體內砷物種動態變化的分析系統，本研究依層析、微透析及ICP-MS儀器測定等三大部分分別進行最佳化操作條件的探討。在完成各關鍵操作參數的最佳化後，本研究即針對所建立之連線系統的分析效能進行探討，實驗結果顯示，對本研究欲測定四個砷物種As(III)、DMA、MMA及As(V)而言，最低可達到的偵測極限分別為0.37, 0.56, 1.42 及 1.88 ng/mL，測定時間的解析度亦可縮短至30分鐘，在長時間的監測過程中，所得到之分析結果的穩定度（RSD值）則均可維持在10 %以內。

在確認利用所建立之連線分析系統，進行活體動物體內砷物種連續監測可行性的過程中，本研究係將微透析探針佈植到大白鼠的肝臟及膀胱內，並以腹腔注射的方式將As2O3注入大白鼠的體內，再進行大白鼠的肝臟及膀胱尿液中砷物種濃度變化的連續動態監測。結果顯示，當大白鼠接受到高劑量無機砷的注射後，其肝臟中及膀胱尿液中隨即出現大量的As(III)與DMA，且DMA出現的時間較As(III) 約慢了60 分鐘，此一結果與文獻中推測之無機砷代謝機制頗為吻合，顯示本研究所建立之連線分析技術確實已具備了現場(in-situ)，體內(in-vivo)及連續(continuous)監測活體動物體內砷物種動態變化的可行性。

The toxicological and biological roles of trace elements are reported highly dependent on their chemical forms and/or oxidation states. In the past decade, the understanding of the impact of various species of trace elements on biological and environmental systems has been remarkably advanced, due mainly to the significant improvement in the speciation techniques. In view of the need of pharmacokinetics studies, the metabolic and biotransportation information of arsenicals in target organs has been considered indispensable in adequately characterizing the effectiveness and impact of arsenic trioxide. Based on aforementioned requirements, an in vivo, in-situ and continuous analytical system was developed to monitor the dynamic variation of different arsenic metabolites in anesthetized rat.
In our study, two miniaturized hyphenation techniques, named microdialysis-micro HPLC-ICP-MS and microdialysis-micro HPLC-HG-ICP-MS, were developed. To extract analyte species from target organs, Ringer’s solution was used to perfuse arsenicals from target organs at flow rate of 0.5 mL/min. An on-line injector (CMA 160) equipped with a 10-mL sample loop and a hydride generator was used to interface microdialysis probe, microbore-LC column and ICP-MS. Under the optimized conditions, the detection limits obtained with the proposed method can be as low as 0.37, 0.56, 1.42 and 1.88 ng/mL for As(III), DMA, MMA and As(V), respectively. The stability of this method can be controlled down to approximately 10 % after continuous operation up to 360 min, permitting to continuously monitor the variation of analyte arsenic species in anesthetized rat after the administration of arsenic trioxide. Based on the achieved analytical results, an in vivo, in-situ and continuous monitoring technique for the investigation of dynamic variation of trace arsenicals in anesthetized rat has been performed by this study. On behalf of the novelty of our proposed method, further characterization of the detoxification and metabolic mechanism of arsenic in various organisms has become possible.

1. D. M. Templeton, Anal. Bioanal. Chem., 2003, 375(1062-1066).
2. T. E. Robinson, J. B. Justice Jr., Microdialysis in the Neurosciences,
1st ed. Elsevier, 1991, p3.
3. F. C. Cheng, Y. Shih, Y. J. Liang, L. L. Yang, C. S. Yang, Journal of Chromatography B, 1996, 682(195-200).
4. Y. Huang, Z. Zhang, J. Lv, H. Cheng, Analytica Chimica Acta, 2000,
419(175-184).
5. S. Asai, T. Kohno, Y. Ishii, and K. Ishikawa, Analytical Biochemistry,
1996, 237(182-187).
6.吳玉琛, 吳祖方, 陳甫州,化學, 1998, 56(327-334).
7. M. I. Davies, Analytica Chimica Acta, 1999, 379(227-249).
8. J. A. Stenken, Analytica Chimica Acta, 1999, 379(337-358).
9. A. L. Quellec, S. Dupin, P. Genissel, S. Saivin, B. Marchand, and G.
Houin, Journal of Pharmacological and Toxicological Methods, 1995,
33(11-16).
10. R. Clement, J. Malinovsky, G. Dollo, P. L. Corre, F. Chevanne, R. L.
Verge, Journal of Pharmaceutical and Biomedical Analysis, 1998,
17(665-670).
11. S. Menacherry, W. Hubert, J. B. Justice, Anal. Chem., 1992, 64(577-
583).
12. K. V. Belle, S. Sarre, G. Ebinger, Y. Michotte, European Journal of
Pharmaceutical sciences, 1995, 3(273-280).
13. M. I. Davies, J. D. Cooper, S. S. Desmond, C. E. Lunte, S. M. Lunte,
Advanced Drug Delivery Reviews, 2000, 45(169-188).
13. Y. Zhao, X. Liang, C.E. Lunte, Analytica Chimica Acta, 1995,
316(403-410).
14. T. Buttler, C. Nilsson, L. Gorton, G. Marko-Varga, T. Layrell, Journal
of Chromatography A, 1996, 725(41-56).
15. K. L. Snyder, C. E. Nathan, A. Yee, J. A. Stenken, Analyst, 2001, 126
(1261-1268).
16. M. S. M. Martinez, B. G. Hurtado, C. C. Gandarillas, J. M. Lanao, A. S. Navarro, Analytica Chimica Acta, 2002, 459(143-150).
17. N. Torto, L. Gorton, T. Laurell, G. Marko-Varga, Trends in analytical chemistry, 1999, 18(252-260).
18. P. J. Tsai, J. P. Wu, N. N. Lin, J. S. Kuo, C. S. Yang, Journal of
chromatography B: Biomedical applications, 1996, 686(151-156).
19. H. Wang, H. Zou, A. Feng, Y. Zhang, Analytica Chimica Acta, 1997, 342(159-165).
20. T. H. Tsai, Y. F. Chen, I. F. Chen, C. F. Chen, Journal of
chromatography B, 1999, 729(119-125).
21. C. S. Yang, P. J. Tsai, W. Y. Chen, W. J. Tsai, J. S. Kuo, Journal of
chromatography B, 1999, 734(1-6).
22. N. Torto, J. Mwatseteza, G. Sawula, Analytica Chemica Acta, 2002, 456(253-261).
23. A. Weston, P. R. Brown, HPLC and CE Principles and Practice,
Academic Press, 1997, p242.
24. H. Garraud, A. Woller, P. Fodor, O. Donard, Analusis, 1997, 25(25-
31).
25. J. P. Chervet, M. Ursem, J. P. Salzmann, Anal. Chem., 1996, 68(1507-
1512).
26. K. E. Lawerence, G. W. Rice, and V. A. Fassel, Anal. Chem., 1984, 56(289-292).
27. K. E. LaFreniere, V. A. Fassel and D. E. Eckels, Anal. Chem., 1987, 59, 879
28. D. R. Wiederin, F. G. Smith and R. S. Houk, Anal. Chem., 1991, 63, 219
29. C. K. S. Sam, H. m. Pang, R. S. Houk, Anal. Chem., 1992, 64(2444-2450).
30. C. K. S. Sam , N. Robert, R. S. Houk, Analyst. 1992. 117(577-582).
31. J. M. Carey, F. A. Byrdy, J.A. Caruso, J. Chromatogr. Sci, 1993, 31, 330
32. M. J. Powell, D. W. Boomer, Anal. Chem., 1995, 67(2474-2478).
33. J. S. Carin, J. T. Kiely, J. Anal. At. Spectrom. 1996, 11(525-527).
34. G. E. M. Hall, J. C. Pelchat, G. Gauthier, J. Anal. At. Spectrom.
1998, 14(205-213).
35. S. A. Pergantis, E. W. Heithmar and T. A. Hinners, Anal. Chem., 1995,
67, 4530
36. F. Vanhaecke, M. V. Holderbeke, L. Moens, R. Dams, J. Anal. At. Spectrom. 1996, 11(543-548).
37. S. Augagneur, B. Medena, J. Szpunar, R. Lobinski, Anal. Chem., 1996, 11, 713
38. C. Dubuisson, E. Poussel, J. M. Mermnet, J. Anal. At. Spectrom.
1997, 12, 281
39. S. D. Lofthouse, G. M. Greenway, S. C. Stephen, J. Anal. At.
Spectrom. 1997, 12( 1373-1376).
40. A. Tangen, R. Tones, T. Greibokk, W. Lund, Anal. Chem., 1997,
12(667-670).
41. A. Woller, H. Garraud, J. Boisson, A. M. Dorthe, P. Fodor, O. F. X. Donard, J. Anal. At. Spectrom. 1998, 13(141-149).
42. M. A. Tarr, G. Zhu, R. F. Browner, Anal. Chem., 1993, 65, 1689
43. H. Liu, A. Montasser, Anal. Chem., 1994, 66(3233-3242).
44. S. H. Nam, J. S. Lim, A. Montaser, J. Anal. At. Spectrom. 1994, 9(1357-1362).
45. S. A. Pergantis, E. M. Heithmar, T. A. Hinners, Anal. Chem., 1995, 67(4530-4535).
46. C. B¢Hymer, K. L. Sutton, J. A. Caruso, J. Anal. At. Spectrom. 1998, 13(855-858).
47. J. W. Olesik, J. A. Kinzer, B. Harkleroad, Anal. Chem., 1994, 66, 2022
48. H. Liu, A. Montaser, Anal. Chem., 1994, 66, 3233
49. H. Vanhoe, L. Moens, R. Dams, J. Anal. At. Spectrom. 1994, 9, 815
50. S. Saverwyns, X. Zhang, F. Vanhaecke, R. Cornelis, L. Moens, Richard Dams, J. Anal. At. Spectrom. 1997, 12(1047-1052).
51. Y. C. Sun, Y. S. Lee, T. L. Shiah, P. L. Lee, W. C. Tseng, M. H. Yang,
Journal of Chromatography A, 2003, 1005(207-213).
52. K. Sutton, R. M. C. Sutton, J. A. Caruso, Journal of Chromatography A, 1997, 789(85-126).
53.G. Zoorob, M. Tomlinson, J. Wang, J. Caruso, J. Anal. At. Spectrom.
1995, 10, 853.
54. M. J. Powell, D. W. Boomer, D. R. Wiedrin, Anal. Chem., 1995, 67,
2474.
55. S. C. K. Shum and R. S. Houk, Anal. Chem., 1993, 65, 2972.
56. S. A. Pergaantis, W. Winnik, D. Betowski, J. Anal. At. Spectrom.
1997, 12(531-536).
57. O. F. X. Donard, J. A. Caruso, Spectrochimica Acta Part B, 1998,
53(157-163).
58. E. D. Goldberg, J. Geol., 1954, 62, 249
59. B. L. Carson, H. V. Ellis III, J. L. McCann, Toxicology and Biological Monitoring of Metals in Humans, Lewis Publishers. Inc., 1986.
60. H. Pendias, A. Kabata-Pendias, Trace Element in Soils and Plants, CRC Press, Inc., 1985.
61. 黃雅莉等, 台灣烏腳病高盛行地區居民尿中砷物種分析研究(初步結果), 1995年生物偵測技術研討會, 行政院勞委會勞工安全衛生研究所, 民國八十四年三月.
62. 鍾禹德, “發展超臨界流體萃取與原子吸收光譜連線測定技術進行固態樣品中甲機汞與無機汞離子的物種分析方法研究”,國立清華大學碩士論文, 民國八十八年七月
63. A. S. Medel, Spectrochimica Acta Part B, 1998, 53(197-211).
64. B. Godlewska, A. Hulanicki, Chem. Anal, 1993, 38(267-278).
65. J. C. Van Loon, Analytical Chemistry, 1979, 51(1139A-1150A).
66. K. T. Suzuki, Analytical Biochemistry, 1980, 102(31-34).
67. T. Hirschfeld, Anal. Chem., 1980, 52, 297A
68.W. C. Tseng, M. H. Yang, T. P. Chen, Y. L. Huang, Analyst, 2002, 127(560-546).
69. B. K. Mandal, K. T. Suzuki, Talanta, 2002, 58(201-235).
70. U.S. Department of health and human services agency for toxic substances and disease registry division of health education and promotion, 2000, Case Studies in Environmental Medicine, ”Arsenic Toxicity”, http://www.atsdr.cdc.gov/HEC/CSEM/arsenic/arsenic.pdf
71. X. C. Le, X. Lu, M. Ma, W. R. Cullen, H. V. Aposhian, B. Zheng, Anal. Chem, 2000, 72(5172-5177).
72. M. Vahter, Applied organometallic chemistry, 1994, 8(175-182).
73. K. T. Suzuki, B. K. Mandal, Y. Ogra, Talanta, 2002, 58(111-119).
74. A. H. Hall, Toxicology Letters, 2002, 128(69-72).
75. M. Vahter, Toxicology Letters, 2000, 112-113(209-217).
76. M. Styblo, L.D. Razo, L. Vega, D. R. Germolee, E. L. LeCluyse, G. A. Hamilton, W. Reed, C. Wang, W. R. Cullen, D. J. Thomas, Arch Toxicol, 2000, 74(289-299).
77. D. J. Thomas, M. Styblo and S. Lin, Toxicol. Appl. Pharmacol., 2001, 176(127-144).
78. J. P. Buchet, R. Lauwerys, and S. Facchetti. Evaluation of exposure to inorganic arsenic in man. Analytic Techniques for Heavy Metals in Biological Fluids, 1st ed. Elsevier, Amsterdam, The Netherlands, 1983, p75-90.
79. M. S. H. Lam, R. J. Ignoffo, Cancer practice, 2001, 9(155-157).
80. K. E Jarvis, A. L. Gray and R. S. Houk, “Handbook of Inductively
Coupled Plasma Spectrometry”, Chapman and Hall, New York(1989).
81. S. Mazan, N. Glon, G. Cretier, J.L. Rocca and J.M. Mermet, J. Anal. At. Spectrom., 1998,17(366-370).
82. S.F. Boulyga, H.J. Dietze, and J.S. Becker, Mikrochim. Acta., 2001, 137(93-103).
83. M.A. Dexter, P.K. Appelblad, C.P. Ingle, J.H. Batey, H.J. Reid, and B.L. Sharp, J. Anal. At. Spectrom., 2002,17(183-188).
84. Q. Xie, and R. Kerrich, J. Anal. At. Spectrom., 2002,17(69-74).
85. C.P. Ingle, P.K. Appelblad, M.A. Dexter, H.J. Reid, and B.L. Sharp, J. Anal. At. Spectrom., 2001,16(1076-1084).
86. I. Feldmann, N. Jakulowski, and D. Stuewer, Fresenius. J. Anal. Chem., 1999, 365(415-421).
87. I. Feldmann, N. Jakulowski, and D. Stuewer, Fresenius. J. Anal. Chem., 1999,365(422-428).
88. A. Montaser, “Inductively Coupled Plasma Mass Spectrometry”,
Wiley-VCH, New York(1998).
89. C. P. Dubuisson, E.Mermnnnet, J. Anal. At. Spectrom., 1997,12,281.
90. M. I. Davies, J. D. Cooper, S. S. Desmond, C. E. Lunte, Advanced Drug Delivery Reviews, 2000, 45(169-188).
91. 黃傳捷,”利用感應耦合電漿質譜儀進行生物樣品中微量元素及其物種分析研究”,國立清華大學博士論文,民國八十六年七月
92. I. Matin, M. A. Lopez-Gonzalvez, M. G, C. Camara, M. A. Palacios, Journal of Chromatography B, 1995, 666(101-109).
93. M. Styblo, L. D. Del Razo, E. L. LeCluyse, G. A. Hamilton, C. Wang, W. R. Cullen, D. J. Tomas, Chem. Res. Toxicol, 1999, 12(560-565).