A new chemiluminescence system for determination of Copper(II)

Zhang Xiaoyan, Fan Xiaodong^#, Liu Xiaoli, Du Jianqiang
(Department of Chemical Engineering, Northwest University, Xi'an 710069, China; ^#Department of Chemical Engineering, Northwestern Polytechnical University, Xi'an 710072, China)

Received May.31, 2002.

Abstract This paper reports a new complex chemiluminescence system, i.e., Luminol-H₂O₂ catalyzed by CuR₂^2- complex which is called New Copper Reagent. The detection limit is 6.6×10^-8 g /L Cu(II). The linear dynamic range is 1×10^-7~1×10^-4g/L. The relative standard deviation(n=5) for 4×10^-5 g/L Cu(II) is 2.7%. Foreign ions, such as Fe(II), Fe(III) and Co(II), interfere when exist in more than 10-fold ratio(w/w) to Cu(II), and several ions can be tolerated when exist in higher ratio to Cu(II). The tolerable capacity of interference ions is relatively large. The precision, accuracy and selectivity of the method are excellent. The procedure has been satisfactorily applied to determine trace copper in immunogloblulin, foods and Tabellae Moroxydini Hydrochloride.
Keywords Chemiluminescence, Luminol, Salicylic acid, Copper.

1. INTRODUCTION
In comparison with other kinetic determination methods, chemiluminescence (CL) analysis is rapid, sensitive and does not require expensive instruments^[1]. Some CL methods for copper determination have been reported, but many metal ions interfere the results.
    Yan et al. have reported a determination of copper in the luminol CL system after ion-chromatography separation of copper, but the CL system itself has poor selectivity. In recent years, therefore, a search has been made for a better CL system^[2-4].CuR^2-₂(CuR^2-₂= Copper Amino Acid Chelate)has been observed to give chemiluminescence when treated with hydrogen peroxide in alkaline medium. Several metal ions were found to speed the reaction^[1,5]. Cu(II) gives the largest effect. The reaction has now been adopted for the determination of trace copper.

2. EXPERIMENTAL
2.1 Instrument and reagents   
CL meter, model YHF-1(made in China), including photomultiplier tube, fused-silica reaction cell and a flat-bed recorder. A pH meter, model 25 (made in Shanghai , China) was used for pH measurements.
    Standard copper solution, 1.0mg/mL, prepared by dissolving the required weight of analytical grade CuSO₄·5H₂O in the requisite volume of redistilled water, and diluted further as required; Luminol (MERCK, Germany) solution, 1×10^-2 mol/L, and 2.5×10^-4mol/ L luminol working solution was prepared from it; Salicylic acid solution, 1×10^-3 mol/L; Hydrogen peroxide solution, 0.25 mol/L; Buffer solution, 0.1M KOH-0.1M H₃BO₄, was adjusted to pH 10.5-11.5.
2.2 Procedure         
Clean and dry the reaction cell, then inject 2.0 ml of 2.5×10^-4 mol/L luminol solution, 1.0ml of 2.5 mol/L hydrogen peroxide through one side-tube, and inject 1.00～2.00ml of CuR₂^2- test solution through the other side-tube, measure the peak height of the recorded kinetic curve of the CL reaction. Drain the cell through a side-tube.

3. RESULTS AND DISCUSSION
3.1 The CL reaction  
A mixture of luminol solution and hydrogen peroxide in alkaline solution, with CuR₂^2- as catalyst, emits light, which reaches maximum intensity 3 seconds after injection of the CuR₂^2- test solution, in a further 1 second the light intensity falls to 50% of the maximum. Various buffer solutions such as 0.1M KOH-0.1M H₃BO₃ (pH 10.5-11.5), 0.1M NaHCO₃-0.1M Na₂CO₃ (pH 11.0), 0.1M NH₄Cl-NH₃(pH 10.5) were tested, and the first one gave the highest emission intensity ( Fig.1 ).

Fig.1 Chemiluminescence dynamic curve
1:Luminol-H₂O₂ system
2:Luminol-H₂O₂-Cu (II) system
3:Luminol-H₂O₂-CuR₂^2- system
[luminol]=2.5×10^-4mol/L
[H₂O₂]=0.25 mol/L
[Cu (II)]=4×10^-5 g/L
0.1M KOH-0.1M H₃BO₃, pH 11.0

3.2 Optimization conditions
Fig.2 illustrates the results of the study of optimization of the luminol and peroxide concentrations. The luminol and peroxide concentrations were 2.5×10^-4 mol/L and 0.20 mol/L respectively. Fig.3 illustrates the effect of the acidity of CuR₂^2- solution; pH 3 was obtained as optimal.
3.3 Calibration and detection limit
Fig.4 shows that light intensity is directly proportional to Cu (II) concentration from 1×10^-6-1×10^-4 g/L. The detection limit is 6.6×10^-8 g/L, defined as the concentration corresponding to the mean background signal plus twice is a standard deviation.

Fig.3 Chemiluminescence intensity as function of pH of the Cu (Ⅱ) Solution
Intial condition: [luminol]=2.5×10^-4 mol/L, [H₂O₂]=0.25 mol/L, [Cu (Ⅱ)]=4×10^-5g/L, 0.1M KOH-0.1M H₃BO₃, pH 11

Fig. 4 Calibration curve for Cu (II)

3.4 Interferences
An extensive interference study gave the results shown in Table 1. Although Fe (II), Fe (III) and Cr (III) interfere the results when exist at greater than 10-fold ratio (w/w) to Cu (II), they can be screened with 1.0×10^-5 mol/L sodium citrate, which does not affect the CL intensity.

Table 1 Influence of coexistent ions

3.5 Determination of copper in immunogloblulin, foods and tabelle moroxydini hydrochloride 
The sample is dissolved by heating 0.500g Tabellae Moroxydini Hydrochloride with 5ml of concentrated nitric acid in a beaker on a controlled hot-plate until nearly dry, then add 1ml of concentrated perchloric acid. It is heated until all white fumes disappeared, and then make up to a volume in a 50ml volumetric flask. A known volume of this solution is transferred into a 25ml volumetric flask, 1.5ml salicylic acid is added, pH is adjusted to 3 with nitric acid, and the solution is diluted to a volume with redistilled water, and analysed as already described. Typical results are showed in Table 2.

Table 2 Determination of Copper in immunogloblulin, foods and Tabellae Moroxydini Hydrochloride (mean of five determination )

REFERENCES             
[1] Zhang Xiaoyan, Li Shaoqing. Chemiluminescence analytical technology. Xi'an Map Publishing House. 2000,189-192.
[2] Yan B, Worsfold P. J. Anal. Chim. Acta. 1990, 236-237.
[3] Zhuang Huisheng, Chen Guonan, Wang Qing'e. Anal. Chem., 2000, 28 (5): 573-576.
[4] Zhang Wuming, Zhong Shiming, Hu Hejin. Analyst, 1989, 17 (10): 922-924.
[5] Li Shaoqing. Chinese Chem. Letter. 1991, 2 (9): 701.
[6] PRC Certified Reference Materials Catalog, State Bureau of Technical Supervision, 1995.

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