發展微透析取樣配合流動注入火焰式原子吸收光譜儀之連線分析系統進行離體與活體動物血液中可透析鎂之研究

On-line microdialysis sampling coupled with FAAS for in-vitro and in-vivo monitoring of diffusible magnesium in blood of living animals

李正發(Cheng-Fa Lee)

指導教授 ： 黃友利 副教授 ； 陳百薰 教授

高雄醫學大學/醫學院/醫學研究所/碩士(2003年)

摘要

微透析配合原子吸收光譜法進行金屬分析是相當特殊與新穎的連線分析方法。在本研究中我們建立微透析取樣（Microdialysis, MD）配合流動注入系統（Flow injection system, FI）和火焰式原子吸收光譜儀（Flame atomic absorption spectrometry, FAAS）的連線分析系統，利用所建立的連線分析系統可以進行離體或活體動物血液樣品中可透析鎂濃度的偵測。首先將灌流液注入微透析透針（Microdialysis probe）中進行透析，再利用六相線上注入閥的收集環來收集透析液，最後藉由攜帶溶液與霧化器的攝入吸引力，將收集到的透析液導入FAAS中進行鎂的連線偵測。本研究所建立MD-FI-FAAS連線分析方法使用0.9％（w/v）的超純氯化鈉溶液作為灌流液，灌流速度設定為40 μl/min，以0.2％（v/v）的超純硝酸溶液作為攜帶溶液，將霧化器攝入流速設定為6 ml/min。將所發展的分析方法應用於透析液中鎂的測定，完成一個樣品分析的流程共需70秒（樣品收集60秒，鎂訊號的讀取10秒）。本研究分析方法的驗證如下：線性範圍從0 mg/l到300 mg/l；偵測極限為0.53 mg/l（3SD/m, n = 7）；在全血、血清與血漿中做鎂的標準添加，其添加回收率均在100％上下。對鎂標準溶液、全血、血漿以及血清樣品進行再現性分析之intra-assay （n = 6）與inter-assay （n = 5）的精密度 （RSD%）均小於20％。為了評估MD-FI-FAAS連線分析方法的準確性，本研究利用no net flux （NNF）法進行鎂標準溶液、血漿樣品以及大白兔血液中可透析鎂濃度的比較。此外，本研究發現如果產生溶血現象時，將會增加全血中可透析鎂的濃度。最後將所建立的MD-FI-FAAS連線分析方法應用在靜脈注射硫酸鎂（40 mg/kg）之活體大白兔上，來進行血液中可透析鎂的原位、動態與連續分析。

關鍵字

火焰式原子吸收光譜儀 ； 鎂 ； 微透析 ； 可透析鎂 ； 連線分析系統

並列摘要

Microdialysis coupled with atomic absorption spectrometry （AAS） is a unique and novel hyphenated method for the determination of metals. In the present study, we have established an on-line microdialysis sampling coupled with flow injection （FI） flame atomic absorption spectrometry （FAAS） system for in-vitro and in-vivo monitoring of the diffusible magnesium （mg） in blood of living animals. Microdialysates perfused through implanted microdialysis probes were collected with a sample loop of an on-line injection valve and directly introduced into the atomizer of FAAS by a carrier solution and a nebulizer uptake flow. Ultrapure saline solution （0.9% NaCl） was used as the perfusion solution at a flow rate of 40 ?l/min through the microdialysis probe. The 0.2% HNO3 solution was used as the carrier solution at a nebulizer uptake rate of 6 ml/min. The interval for each determination was 70 s （60 s sampling time and 10 s read time）. The optimized conditions of the MD-FI-FAAS were performed for measurement of Mg in microdialysate. The performance characteristics of this on-line microdialysis-FI-FAAS system for Mg were as follows: linearity range, 0-300 mg/l; detection limit （n = 7）, 0.53 mg/l; recovery for spiked whole blood, plasma and serum samples, ≒ 100 %; intra-assay （n = 6） and inter-assay （n = 5） precisions （RSD%） for aqueous Mg standard solutions and three whole blood, plasma and serum samples, < 20%. The accuracy of the proposed on-line method was checked using no net flux （NNF） method. Interference resulted from hemolysis would markedly increase the concentration of diffusible Mg in whole blood by the proposed method. The use of an on-line MD-FI-FAAS system permitted the in situ, dynamic and continuous in vivo monitoring of diffusible Mg in blood of living rabbits after administrating with MgSO4 (40 mg/kg).

並列關鍵字

FAAS ； Magnesium ； Mg ； Microdialysis ； diffusible ； On-line analysis

參考文獻

1. A. Elamin and T. Tuvemo, Diabetes Res. Clin. Pract., 1990,10, 203.

2. R. J. Elin, Overview of problems in the assessment of the magnesium in cellar process and medicine, 4th Int. Symp. Magn., 1985, pp.67-76, Blanksurgh, Va (Kargr, Basel).

3. F. Guerrero-Romero and M. Rodríguez-Moraź, Acta Diabetol., 2002, 39, 209.

4. I. M. Papzachariou, A. Martinex-Isla, E. Efthimiou, R. C. N. Willamson and S. I. Girgis, Clin. Chim. Acta, 2000, 302, 145.

5. B. T. Altura, T. L. Shirey, C. C. Young, J. Hiti, K. Dell’Orfano, S. M. Handwerker and B. M. Altura, Methods Find. Exp. Clin. Pharmacol., 1992, 14, 297.