中文摘要
　　麻黃的使用已有久遠的歷史，自古即用以發汗、利尿、治療氣喘、支氣管炎、腎水腫等。主要活性成分為三對麻黃生物鹼分別為：左旋麻黃鹼與右旋偽麻黃鹼﹔左旋去甲基麻黃鹼與右旋去甲基偽麻黃鹼﹔左旋甲基麻黃鹼與右旋甲基偽麻黃鹼。由於麻黃種類繁多，市售麻黃中所含的麻黃生物鹼含量有很大不同，因此麻黃生物鹼的定量工作十分重要。在麻黃生物鹼的分析方法中包含薄層層析、氣相層析、高效能液相層析與毛細管電泳等方法，然而這些方法需要經過前處理去除干擾物或藉由衍生化來增加靈敏度，且結果並沒有皆令人滿意。本研究室提出一利用核磁共振光譜定性定量市售麻黃植物中麻黃生物鹼含量的方法，可改善傳統分析方法的困擾，且對麻黃生物鹼更加具有選擇性。此方法能同時定性定量五種以上的麻黃生物鹼，且不需要任何標準品製備檢量線。
　　本篇論文以核磁共振光譜儀完成麻黃植物中五個麻黃生物鹼的分析與定量，分別為左旋麻黃鹼、右旋偽麻黃鹼、左旋去甲基麻黃鹼、右旋去甲基偽麻黃鹼與左旋甲基麻黃鹼。在麻黃植物的基原判斷分別完成草麻黃、木賊麻黃和中麻黃，其中草麻黃中左旋麻黃鹼含量約為右旋偽麻黃鹼的2~3倍；在中麻黃的分析中左旋麻黃鹼與右旋偽麻黃鹼的含量與草麻黃則為相反，右旋偽麻黃鹼含量約為左旋麻黃鹼的2~3倍；在木賊麻黃中麻黃生物鹼則以左旋麻黃鹼為主要量。
　　利用此方法可快速鑑定市售含麻黃藥材之科學中藥中所使用的麻黃品種，且可同時定量分析所含的麻黃生物鹼含量，具有極佳的濃度-積分值線性相關。並針對國內市售的四家藥廠含有麻黃的29種中藥製劑，完成各類麻黃生物鹼的含量分析，由分析結果可知四家藥廠所製造的麻黃製劑中麻黃生物鹼的含量有很大的差別，並對各家藥廠的麻黃生物鹼相對含量提出討論。另外在不含麻黃的中藥製劑中分析出麻黃生物鹼，並由核磁共振光譜儀圖譜與麻黃生物鹼的含量分析，對麻黃生物鹼的來源提出推論。
　　以核磁共振光譜儀作為分析工具，提供一快速、準確、方便的分析方法同時定性定量麻黃生物鹼類似物。並且本方法可以應用於市售藥物、麻黃製品及食品的檢驗。

Abstract
　The use of Ephedra, known as Ma-huang, can be traced back to ancient times, since it has been used as a diaphoretic, diuretic and antiasthmatic, as well as agents for the treatment of bronchitis and acute nephritic edema. The main active components of Ephedra are three pairs of optically active diastereomeric ephedrine analogues: l-ephedrine, d-pseudoephedrine, l-norephedrine, d-norpseudoephedrine, l-methylephedrine and d-methylpseudoephedrine. Ephedra used in Chinese medicine showed quite variable quality because a number of species comprised its source in the market, which makes the identification of ephedrine essential. The methods for analysis of Ephedra alkaloids include TLC, GC, HPLC and CE; however, these methods required elaborate-clean-up and derivatization procedures in order to enhance sensitivity and to remove the compounds that interfere with the target compounds, which were not always satisfactory. The present study provides a qualitative and quantitative analysis of the ephedrine in Ephedra herbs in the market. The developed method not only eases the trouble of conventional analysis methods, but also increases the selectivity of ephedrine; in addition, it allows simultaneous determination of more than five ephedrine analogues without any calibration curves.
　A qualitative and quantitative analysis method for the determination of five ephedrine analogues from Ephedra herbs using 1H-NMR spectroscopy was described. It allows rapid and simultaneous determination of ephedrine analogues l-ephedrine, d-pesudoephedrine, l-norephedrine, d-norpesudoephedrine, and l-methylephedrine without any pre-cleaning steps. This method was applied for the analysis of Ephedra materials including E. sinica Stapf, E. equisetina Bge, and E. intermedia Schrenk for ephedrine analogues. In E. sinica Stapf, the quantity of ephedrine is two to three times more than that of pesudoephedrine. On the contrary, in E. intermedia Schrenk, the quantity of pesudoephedrine is two to three times more than that of ephedrine. In E. equisetina Bge, ephedrine is the major content.
　The method allows rapid detection of the exact kind of Ephedra used in scientific Chinese medicine formula containing Ephedra materials in the market and provides simultaneous quantitative analysis of the ephedrine in the formula; it has excellent concentration integral liner relationship. The study also focused on the quantitative analysis of ephedrine in 29 Chinese medicine preparations containing Ephedra provided by four domestic pharmaceutical companies. The quantity of ephedrine in these Ephedra preparations resulted in great variation, and the relative quantity of ephedrine in the preparation of each company was discussed. Moreover, some Chinese medicine preparations without ephedrine have been detected the existence of ephedrine, and the source of the ephedrine was identified by the 1H-NMR method.
　1H-NMR can serve as an analysis tool, providing a rapid, accurate, and convenient method for qualitative and quantitative analysis of ephedrine analogues from Ephedra species. This method can be applied to the analysis of commercial pharmaceutics or products of Ephedra and dietary supplements.

參 考 文 獻
1. Anonymous. The Ephedras. The Lawrence Review of Natural Products, St. Louis, MO, November 1995, pp1–2.
2. S. Mahdihassan, F. S. Mehdi. Soma of the Rigveda and an attempt to identify it. Am. J. Chin. Med. 1989, 17, 1–8.
3. M. Blumenthal, A. Goldberg, J. Brinckmann. Ephedra. Herbal medicine. In: Expanded Commission E Monographs. American Botanical Council, Austin, Texas, 2002, pp110–117.
4. WHO. Herba Ephedra. WHO Monographs on Selected Medicinal Plants, World Health Organization, Geneva. 1999, 1, pp145–153.
5. http://www.gao.gov/archive/1999/h299090.pdf.
6. P. Shekelle, M. Hardy, S. C. Morton, M. Maglione, M. Suttorp, E. Roth, L. Jungvig, W. A. Mojica, J. Gagné, S. Rhodes. Ephedra and ephedrine for weight loss and athletic performance enhancement: Clinical efficacy and side effects. Evidence Report/Technology Assessment. USA, 2003, 76.
7. http://www.fda.gov/
8. R. A. Price. Systematics of the gentales : A review of morphological and molecular evidence. Int. J. Plant Sci. 1996, 157, 40–49.
9. S. Caveney. New observations of the secondary chemistry of world Ephedra (Ephedraceae). Am. J. Bot. 2001, 88 , 1199–1208.
10. http://www.csc.pku.edu.cn/
11. http://www.tcmhealth.gov.tw/
12. 國家中醫藥管理局《中華本草》編委會。中華本草，第二冊，上海科學技術出版社，上海，1999，pp349-357。
13. 孟景春，周仲英，中醫學概論。知音出版社，台北，1991，p164。
14. 顏焜熒，圖解常用中藥處方。南天，台北，1984。
15. 莊松榮製藥廠有限公司研發部，莊松榮複方選輯。莊松榮製藥廠有限公司，高雄，2000。
16. 王綿之，許濟群，方劑學。知音出版社，台北，2001。
17. Y. M. Liu, S. J. Sheu, S. H. Chiou, S. C. Chang, Y. P. Chen. A comparative study on commercial samples of Ephedrae herba. Planta Med. 1993, 59, 376–378.
18. S. J. Sheu. Identification by chemical analysis of the botanical sources of commercial samples of Chinese herbal drugs. J. Food Drug Anal. 1997, 5, 285–294.
19. J. G. Hardman, L. E. Limbird, A. Gilman. Goodman and Gilman’s The Pharmacologic Basis of Disease. McGraw-Hill, New York, 2001.
20. T. H. Burnham, K. K. Novak, W. L. Bell, S. L. Schweain. Ephedrine. In: Drug Facts and Comparisons. St Louis, MO: Facts and Comparisons. 2002. p. 692.
21. A. Astrup, S. Toubro, N. J. Christensen, F. Quaade. Pharmacology of thermogenic drugs. Am. J. Clin. Nutr. 1992, 55, 246–248.
22. L. Bukowiecki, L. Jahjah, N. Follea. Ephedrine, a potential slimming drug, directly stimulates thermogenesis in brown adipocytes via beta-adrenoreceptors. Int. J. Obes. 1982, 6, 343–350.
23. A. G. Dulloo, J. Seydoux, L. Girardier, P. Chantre, J. Vandermander. Green tea and thermogenesis: Interactions between catechin-polyphenols, caffeine and sympathetic activity. Int. J. Obes. Relat. Metab. Disord. 2000, 24, 252–258.
24. W. R. Martin, J. W. Sloan, J. D. Sapira, D. R. Jasinski. Physiologic, subjective, and behavioral effects of amphetamine, methamphetamine, ephedrine, phenmetrazine, and methylphenidate in man. Clin. Pharmacol. Ther. 1971, 12, 245–258.
25. 許長安，常用藥物治療手冊。2001，pp942–943。
26. F. T. Noggle, J. DeRuiter, C. R. Clark. Liquid chromatographic determination of the enantiomeric composition of methamphetamine prepared from ephedrine and pseudoephedrine. Anal. Chem. 1986, 58, 1643–1648.
27. J. S. Zhang, Z. Tina, Z. C. Lou. Simultaneous determination of six alkaloids ephedrae herba by high performance liquid chromatography. Planta Med. 1988, 54, 69–70.
28. J. S. Zhang, Z. Tian, Z. C. Lou. Quality evaluation of twelve species of Chinese Ephedra (Mu Huang). Acta Pharm. Sin. 1989, 24, 865–871.
29. H. X. Liang, R. G.. Yu, Q. H. Yang, K. Y. Ni. Determination of ephedrine, pesudoephedrine and strychnine in jiu-fen san by HPLC. Acta Pharm. Sin. 1990, 25, 849–853.
30. H. X. Liang, R. G. Yu, Q. H. Yang, K. Y. Ni. Use of the powell method to optimize the composition of HPLC mobile phase –The separation of ephedrine and pesudoephedrine. Acta Pharm. Sin. 1990, 26, 49–52.
31. C. Imaz, D. Carreras, R. Navajas, C. Rodriguez, A. F. Rodriguez, J. Maynar, R. Cortes. Determination of ephedrines in urine by high performance liquid chromatography. J. Chromatogr. 1993, 631, 201–205.
32. X. Jin, K. R. Cui. Resolution of the enantiomers using (+)-FLEC ephedrines chiral derivatization and reversed-phase liquid chromatography. Acta Pharm. Sin. 1994, 29, 122–127.
33. X. Jin, S. Wang, C. J. Zhang. Quantitative analysis of ephedrines in urine by HPLC. Acta Pharm. Sin.. 1994, 29, 375–379.
34. G. Shao, F. Wu, D. S. Wang, R. Zhu, X. Luo. Quantitative analysis of l-ephedrine and d-pesudoephedrine in plasma by High Performance Liquid Chromatography with fluorescence detection . Acta Pharm. Sin. 1995, 30, 384–389.
35. B. J. Gurley, P. Wang, S. F. Gardner. Ephedrine-type alkaloid content of nutritional supplements containing Ephedra sinica (Ma-huang) as determined by high performance liquid chromatography. J. Pharm. Sci. 1998, 87, 1547–1553.
36. N. Okamura , H. Miki, T. Harada, S. Yamashita,Y. Masaoka, Y. Nakamoto, M. Tsuguma, H. Yoshitomi, A. Yagi. Simultaneous determination of ephedrine, pseudoephedrine,norephedrine and methylephedrine in Kampo medicines by high-performance liquid chromatography. J. Pharm. Biomed. Anal. 1999, 20, 363–372.
37. C. Imaz , R. Navajas, D. Carreras, C. Rodrýguez, A. F. Rodrýguez. Comparison of various reversed-phase columns for the simultaneous determination of ephedrines in urine by high-performance liquid chromatography. J. Chromatogr. A. 2000, 870, 23–28.
38. M. K. Lee, B. H. Cheng, C. T. Che, D. P. H. Hsieh. Cytotoxicity assessment of Ma-huang (Ephedra) under different conditions of preparation. Toxicol. Sci. 2000, 56, 424–430.
39. S. J. Sheu, M. H. Huang. Determination of Ephedra alkaloids by high-performance liquid chromatography. Chromatographia. 2001, 54, 117–119.
40. H. X. Li, M. Yu. Ding, K. Lv, A. Y. Yu. Simultaneous separation and determination of ephedrine alkaloids and tetramethylpyrazine in ephedra sinica Stapf by HPLC. J. Liq. Chromatogr. Relat. Technol. 2002, 25, 313–320.
41. N. Wu, W. Feng , E. Lin , G. Chen, J. Patel , T. M. Chan, B. Pramanik . Quantitative and structural determination of pseudoephedrine sulfate and its related compounds in pharmaceutical preparations using high-performance liquid chromatography. J. Pharm. Biomed. Anal. 2002, 30, 1143–1155.
42. K. Aljazaf, T. W. Hale, K. F. Ilett, P. E. Hartmann, L. R. Mitoulas, J. H. Kristensen, L. P. Hackett. Pseudoephedrine: effects on milk production in women and estimation of infant exposure via breast milk. Br. J. Clin. Pharmacol. 2003, 56, 18–24.
43. B. T. Schaneberg, S. Crockett, E. Bedir, I. A. Khan. The role of chemical fingerprinting: Application to Ephedra. Phytochemistry. 2003, 62, 911–918.
44. M. Ichikawa, M. Udayama, K. Imamura, S. Shiraishi, H. Matsuura. HPLC determination of (1)-pseudoephedrine and (2)-ephedrine in Japanese herbal medicines containing Ephedra herb using solid-phase extraction. Chem. Pharm. Bull. 2003, 51, 635–639.
45. R. A. Niemann, M. L. Gay. Determination of ephedrine alkaloids and synephrine in dietary supplements by column-switching cation exchange high-performance liquid chromatography with scanning-wavelength ultraviolet and fluorescence detection. J. Agric. Food Chem. 2003, 51, 5630–5638.
46. M. C. Roman. Determination of ephedra alkaloids in urine and plasma by HPLC-UV: Collaborative study. J. AOAC Int. 2004, 87, 15–24.
47. L. Li, J. F. Cui, K. R. Cui, M. Z. Wang. The identification of ephedrine and its analogues in urine by gas chromatography. Acta Pharm. Sin. 1990, 26, 66–71.
48. J. F. Cui, C. Q. Niu, J. S. Zhang. Determination of six Ephedra alkaloids in Chinese Ephedra (Ma Huang) by gas chromatography. Acta Pharm. Sin. 1991, 26, 852–857
49. M. J. LeBelle, C. Savard, B. A. Dawson, D. B. Black, L. K. Katyal, F. Zrcek. Chiral identification and determination of ephedrine, pseudoephedrine, methamphetamine and methcathinone by gas chromatography and nuclear magnetic resonance. Forensic Sci. Int. 1995, 71, 215–223.
50. L. B. Hansen. A stable gas chromatography-mass spectrometry analysis system to characterize Ma Huang products found in health foods and supplements. J. Pharm. Sci. 2001, 90, 943–948.
51. M. H. Spyridaki, C. J. Tsitsimpikou, P. A. Siskos, C. G. Georgakopoulos. Determination of ephedrines in urine by gas chromatography-mass spectrometry. J. chromatogr., Biomed. appl. 2001, 758, 311–314.
52. P. V. Eenoo, F. T. Delbeke, K. Roels, P. D. Backer. Simultaneous quantitation of ephedrines in urine by gas chromatography-nitrogen-phosphorus detection for doping control purposes. J. chromatogr., Biomed. appl. 2001, 760, 255–261.
53. H. X. Li, M. Y. Ding, K. Lv, J. Y. Yu. Separation and determination of ephedrine alkaloids and tetramethylpyrazine in Ephedra sinica Stapf by gas chromatography-mass spectrometry. J. Chromatogr. Sci. 2001, 39, 370–374.
54. B. M. El-Haj, A. M. Al-Amri, M. H. Hassan, H. S. Ali, R. K. B. Khadem. The use of cyclohexanone as a "derivatizing" reagent for the GC-MS detection of amphetamines and ephedrines in seizures and the urine. Forensic sci. int. 2003, 135, 16–26.
55. P. R. Stout, K. L. Klette, C. K. Horn. Evaluation of ephedrine, pseudoephedrine and phenylpropanolamine concentrations in human urine samples and a comparison of the specificity of DRI amphetamines and Abuscreen online (KIMS) amphetamines screening immunoassays. J. Forensic. Sci. 2004, 49, 160–164.
56. Y. M. Liu, S. J. Sheu. Determination of ephedrine and pseudoephedrine in Chinese herbal preparations by capillary electrophoresis. J. Chromatogr. 1993, 637, 219–223.
57. M. Chicharro, A. Zapardiel, E. Bermejo, J.A. Perez, L. Hernandez. Direct determination of ephedrine and pseudoephedrine in human urine by capillary zone electrophoresis. J. Chromatogr. 1993, 622, 103–108.
58. Z. Aturki, S. Fanali. Use of β-cyclodextrin polymer as a chiral selector in capillary electrophoresis. J. Chromatogr. A, 1994, 680, 137–146.
59. M. Chicharro, A. Zapardiel, E. Bermejo, J.A. Perez, L. Hernandez. Direct determination of ephedrine alkaloids and epinephrine in human urine by capillary zone electrophoresis. J. Liq. Chromatogr. 1995, 18, 1363–1381.
60. S. Chinaka, S. Tanaka, N. Takayama, K. Komai. Simultaneous chiral analysis of methamphetamine and related compounds by capillary electrophoresis. J. Chromatogr. B. 2000, 749, 111–118.
61. Y. T. Iwata, A. Garcia, T. Kanamori, H. Inoue, T. Kishi, I. S. Lurie. The use of a highly sulfated cyclodextrin for the simultaneous chiral separation of amphetamine type stimulants by capillary electrophoresis. Electrophoresis. 2002, 23, 1328–1334.
62. Y. Kazuo, F. Kimiko. Qualitative and quantitative analysis of Ephedra alkaloids in ephedrae herba carbon-13 nuclear magnetic resonance. Chem. Pharm. Bull. 1979, 27, 43–47.
63. M. H. George. Determination of ephedrine, pseudoephedrine, and norephedrine in mixtures (bulk and dosage forms) by proton nuclear magnetic resonance spectroscopy. J. AOAC Int. 1995, 78, 946–954.
64. H. K. Kim, Y. H. Choi, W. T. Chang, R. Verpoorte. Quantitative analysis of ephedrine analogues from Ephedra species using 1H-NMR. Chem. Pharm. Bull. 2003, 51, 1382–1385.
65. R. Suedee, C. Songkram, A. Petmoreekul, S. Sangkunakup, S. Sankasa, N. Kongyarit. Direct enantioseparation of adrenergic drugs via thin-layer chromatography using molecularly imprinted polymers. J. Pharm. Biomed. Anal. 1999, 19, 519–527.
66. M. McCooeye, L. Ding, G. J. Gardner, C. A. Fraser, J. Lam, R. E. Sturgeon, Z. Mester. Separation and quantitation of the stereoisomers of Ephedra alkaloids in natural health products using flow injection-electrospray ionization-high field asymmetric waveform ion mobility spectrometry-mass spectrometry. Anal. Chem. 2003, 75, 2538–2542.
67. T. A.van Beek, G. P. Lelyveld. Thin layer chromatography of bilobalide and ginkgolides A, B, C and J on sodium acetate impregnated silica gel. Phytochem. Anal. 1993, 4, 109–114.
68. I. Duarte, A. N. Barros, P. S. Belton, R. Righelato, M. Spraul, E. Humpfer, A. M. Gil. High-resolution nuclear magnetic resonance spectroscopy and multivariate analysis for the characterization of Beer. J. Agric. Food Chem. 2002, 50, 2475–2481.
69. Y. H. Choi, H. K. Choi, A. Hazekamp, P. Bermejo, Y. Schilder, C. Erkelens, R. Verpoorte. Quantitative analysis of bilobalide and ginkgolides from Ginkgo biloba leaves and ginkgo products using 1H-NMR. Chem. Pharm. Bull. 2003, 51, 158–161.
70. L. Bala, G. A. Nagana Gowda, U. C. Ghoshal, A. Misra, M. Bhandari, C. L. Khetrapal. 1H NMR spectroscopic method for diagnosis of malabsorption syndrome: a pilot study. NMR Biomed. 2004, 17, 69–75.
71. H. K. Kim, Y. H. Choi, Teus J. C. Luijendijk, R. A. Vera Rocha, R. Verpoorte. Comparison of extraction methods for secologanin and the quantitative analysis of secologanin from symphoricarpos albus using 1H-NMR. Phytochem. Anal. 2004, 15, 257–261.
72. K. M. Holtman, H. M. Chang, J. F. Kadla. Solution-state nuclear magnetic resonance study of the similarities between milled wood lignin and cellulolytic enzyme lignin. J. Agric. Food Chem. 2004, 52, 720–726.
73. G. L. Gall, I. J. Colquhoun, M. Defernez. Metabolite profiling using 1H NMR spectroscopy for quality assessment of green tea, Camellia sinensis (L.). J. Agric. Food Chem. 2004, 52, 692–700.
74. I. F. Duarte, A. N. Barros, C. U. Almeida, M. Spraul, A. M. Gil. Multivariate analysis of NMR and FTIR data as a potential tool for the quality control of beer. J. Agric. Food Chem. 2004, 52, 1031–1038.
75. Y. H. Choi, H. K. Kim, A. Hazekamp, C. Erkelens, A. W. M. Lefeber, R. Verpoorte. Metabolomic Differentiation of Cannabis sativa Cultivars Using 1H NMR Spectroscopy and Principal Component Analysis. J. Nat. Prod. 2004, 67, 953–957.
76. G. M. Hanna. NMR regulator analysis: determination and characterization of Chinese-herb aristolochic acids. Pharmazie. 2004, 59, 170–174.
77. C. Y. Li, C. H. Lin, C. C. Wu, K.H. Lee, T. S. Wu. Efficient 1H nuclear magnetic resonance method for improved quality control analyses of ginkgo constituents . J. Agric. Food Chem. 2004, 52, 3721–3725.
78. C. J. Pouchert, J. Behnke. The Aldrich library of 13C and 1H FT NMR spectra. Aldrich chemical company, inc. Edit. 1, 1993, 2, USA.
79. Karl G. Gunderson, Michael J. Shapiro, Robert A. Doti and Jerry W. Skiles. Simultaneous determination of enantiomeric purity and erythro/threo relationship in chiral 1,2-aminoalcohols by NMR using (R)-(+)-t-butylphenylphosphinothioic acid. Tetrahedron: asymmetry. 1999, 10, 3263–3266.
80. M. Fujita, T. Hiyama. Fluoride ion catalyzed reduction of aldehydes and ketones with hydrosilanes. Synthetic and mechanistic aspects and an application to the threo-directed reduction of a-substituted alkanones. J . Org. Chem. 1988, 53, 5405–5415.
81. M. Nishimura, S. Minakata, T. Takahashi, Y. Oderaotoshi, M. Komatsu. Asymmetric N1 unit transfer to olefins with a chiral nitridomanganese complex: Novel stereoselective pathways to aziridines or oxazolines. J. Org. Chem. 2002, 67, 2101–2110