Simultaneous determination of tyrosine and dopamine in urine samples using excitation–emission matrix fluorescence coupled with second-order calibration

Simultaneous determination of tyrosine and dopamine in urine samples using excitation–emission matrix fluorescence coupled with second-order calibration

Chinese Chemical Letters 24 (2013) 239–242 Contents lists available at SciVerse ScienceDirect Chinese Chemical Letters journal homepage: www.elsevie...

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Chinese Chemical Letters 24 (2013) 239–242

Contents lists available at SciVerse ScienceDirect

Chinese Chemical Letters journal homepage: www.elsevier.com/locate/cclet

Original article

Simultaneous determination of tyrosine and dopamine in urine samples using excitation–emission matrix fluorescence coupled with second-order calibration Shan-Shan Li, Hai-Long Wu *, Ya-Juan Liu, Hui-Wen Gu, Ru-Qin Yu State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, China

A R T I C L E I N F O

A B S T R A C T

Article history: Received 13 November 2012 Received in revised form 19 December 2012 Accepted 6 January 2013

A ‘‘green’’ and quick analytical method for complex compounds was developed for simultaneous determination of tyrosine (Tyr) and dopamine (DA) in urine samples in this paper. The three-way responsive data recorded by excitation–emission matrix fluorescence (EEM) spectrometer was analyzed using second-order calibration methods based on both parallel factor analysis (PARAFAC) and selfweighted alternating trilinear decomposition (SWATLD) algorithms. The EEM spectra of the analytes were overlapped with the background in urine samples. However the second-order advantage of both PARAFAC and SWATLD methods was exploited, even in the presence of unknown interferences and the satisfactory results can be obtained. Furthermore, the linear ranges of Tyr and DA were determined to be 0.042–6.42 mg/mL and 0.18–4.43 mg/mL, respectively, and the accuracies of both methods were validated by the analytical figures of merit (FOM). ß 2013 Hai-Long Wu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.

Keywords: Second-order calibration PARAFAC SWATLD Tyrosine Dopamine

1. Introduction

2. Experimental

Because second-order calibration methods exploit the ‘‘secondorder advantages’’ [1], they have been widely applied in the field of analytical chemistry in recent years [2,3]. Our laboratory has reported some works about the determination of complex compounds using second-order methods [4,5]. Tyrosine (Tyr) is a semi-essential amino acid in the body and is primarily derived from phenylalanine by phenylalanine hydroxylase [6]. Dopamine (DA) is a biological molecule and it is one of the most important catecholamine neurotransmitters in the mammalian central nervous system [7]. Abnormal concentrations of Tyr and DA have been linked with several neurological disorders such as the debilitating ailment Parkinson’s disease and schizophrenia [8–10]. Several analytical techniques have been used in the past for Tyr and DA detection but each has its disadvantages. For example, chromatographic and electrophoretic methods are very expensive, complex and cumbersome [11]. Another problem is the existence of many interfering compounds when we monitor the concentration of Tyr and DA in biological samples [12]. In this paper, an EEM fluorescence method was proposed to simultaneously determine Tyr and DA in urine samples in which uncalibrated interferences exist with the aid of second-order methods based on PARAFAC [13] and SWATLD [14] algorithms.

Tyr and DA were purchased from the National Institute for Control of Pharmaceutical and Biological Products (Changsha, China). The human urine was obtained from a health volunteer. The standard solutions of Tyr (0.020 mg/mL) and DA (0.036 mg/ mL) were prepared by dissolving with ultra-pure water in a 50 mL volumetric flasks as the stock solution. The solutions were stored in a refrigerator at 4 8C. The water was prepared with a Milli-Q water purification system (Aquapro, China). The working solutions were prepared by appropriate dilution of the stock solution in ultra-pure water. In terms of the therapeutic concentration ranges and the linear analytical ranges of Tyr and DA, 15 samples were prepared for the determination of Tyr and DA in human urine samples. The first nine samples containing Tyr in the concentration range from 0.20 mg/mL to 2.40 mg/mL and DA in the concentration range from 0.36 mg/mL to 3.96 mg/mL. Six urine samples contained not only Tyr and DA, but also urine diluted with water (1:10, v/v). Six urine samples, each in 150 mL, spiked with different amounts of Tyr and DA were diluted to 10 mL with water. The final concentrations of Tyr and DA were within the calibration concentration range necessary to verify the accuracy of secondorder calibration based on PARAFAC and SWATLD algorithms. The spectral of water were recorded in triplicate to subtract the influence of reagent blank solutions. All fluorescence measurements were carried out on a HITACHI F-4500 fluorescence spectrophotometer fitted with a xenon lamp and a 1.0 cm quartz cell. The spectra were obtained by scanning the

* Corresponding author. E-mail address: [email protected] (H.-L. Wu).

1001-8417/$ – see front matter ß 2013 Hai-Long Wu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. http://dx.doi.org/10.1016/j.cclet.2013.01.044

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Fig. 1. Three-way fluorescence spectra for 0.2 mg/mL Tyr and 0.36 mg/mL DA: (a) Tyr and DA; (b) in urine sample.

mixture standard solutions recording at the excitation wavelengths in the range from 290 nm to 380 nm at regular steps of 2.0 nm and emission wavelengths in the range from 218 nm to 290 nm at regular steps of 3.0 nm. The scan rate was 12,000 nm/min. Under the chemical conditions mentioned above, each sample can obtain a two-dimensional data array of size 46  25. The combination of data matrices from 6 samples constituted a three-way data array of size 46  25  6 (excitation wavelengths  emission wavelengths  samples). All computer programs were written in Matlab, and all calculations were carried out on a Windows 7 operating system.

3. Results and discussion The linear ranges of Tyr and DA were investigated before undertaking an experimental design. When Tyr and DA were in the range of 0.042–6.42 mg/mL and 0.18–4.43 mg/mL, respectively, the correlation coefficients were all above 0.9990. The treated excitation–emission matrix fluorescence spectra of Tyr (2.47 mg/mL) and DA (4.39 mg/mL) in the absence of urine and in the presence of urine are plotted in Fig. 1. Fig. 1a shows that the absorbencies of Tyr and DA merge into each other. However after urine addition, the fluorescence intensity of the sample solution sharply increased, resulting in heavy fluorescence overlapping between the analytes and the urine back ground in the chosen region, and sequentially the spectral peaks of Tyr and DA were difficult to identify. Thus simultaneous quantification of Tyr is difficult without previous separation procedures.

Herein, the second-order calibration methods based on PARAFAC and SWATLD algorithms having the well-established ‘‘second-order advantage’’ were adopted to resolve the overlapped spectra and to give satisfactory results. Six prediction samples were prepared with the concentration of Tyr and DA shown in Table 1. The value of core-consistency [15] parameter was analyzed using PARAFAC or SWATLD to estimate the appropriate component number N for each sample. The core-consistency diagnostic test indicated that three factors are the best choice, because there is sharp decrease in core-consistency when more factors are utilized, denoting the three consisting of two target analytes and one interferent from urine. Fig. 2 shows the actual spectral profiles and the profiles from the decomposition of the excitation–emission matrix fluorescence data array obtained for both the calibration and prediction samples using PARAFAC (Fig. 2a1 and b1) and SWATLD (Fig. 2a2 and b2) with the factor number of three. The Fig. 2c1 and c2 showed their relative concentrations. The prediction results with PARAFAC and SWATLD methods are listed in Table 1. For Tyr, the average recoveries gained from PARAFAC and SWATLD are (99.9  3.5)% and (100.0  4.8)%, respectively. For DA, the average recoveries obtained from PARAFAC and SWATLD are (99.6  6.7)% and (98.4  5.6)%, respectively. In the present work, by comparison, both the PARAFAC method and the SWATLD method produced similar results. The FOM, including SEN, SEL [16,17], LOD, LOQ and the rootmean-square error of prediction (RMSEP) were also given in Table 1. An inspection of the values quoted in Table 1 will allow us to observe that both methods could gain good results and the results were very similar with each other. In order to investigate

Table 1 Predicted results and figures of merit for urine samples using PARAFAC and SWATLD. Tyrosine Actual (mg/mL)

Dopamine Predicted (mg/mL) PARAFAC

0.412 0.618 0.824 1.030 1.442 1.648 Average recovery (%) RMSEP (mg/mL) SEL SEN (mL/ng) LOD (mg/mL) LOQ (mg/mL)

0.434 0.618 0.798 0.988 1.418 1.620

(105.2) (105.3) (96.9) (95.8) (98.4) (98.3)

99.9  3.5 0.037 0.21 0.29 1.51 4.57

Actual (mg/mL) SWATLD 0.444 0.660 0.791 0.974 1.413 1.598

(107.7) (106.8) (96.0) (94.6) (98.0) (97.0)

100.0  4.8 0.058 0.21 0.29 1.47 4.46

3.294 2.928 2.562 2.192 1.464 1.098

Predicted (mg/mL) PARAFAC

SWATLD

3.042 2.760 2.395 2.150 1.503 1.280

3.048 2.768 2.370 2.134 1.483 1.231

(92.4) (94.3) (93.5) (98.1) (102.6) (116.5)

99.6  6.7 0.20 0.16 0.53 2.28 6.92

(92.5) (94.5) (92.5) (97.4) (101.3) (112.1)

98.4  5.6 0.19 0.16 0.52 2.34 7.10

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Fig. 2. The PARAFAC-resolved spectral profiles (a1, b1) and the SWATLD-resolved spectral profiles (a2, b2) and the actual target analytes spectral profiles: (a) excitation and (b) emission. And the corresponding resolved relative concentrations (c1, c2).

the precision and repeatability of the methods, the analysis of urine samples containing Tyr and DA were repeated for 3 days. All of the results are shown in Table 2. From Table 2, we could see all the RSDs of predicted concentrations using our methods in interday were found to be less than 3%. These results of the inter-day Table 2 Precision and repeatability of the methods: RSDs of inter-day experiments. PARAFAC

First day average recovery (%) Second day average recovery (%) Third day average recovery (%) Inter-day RSD (%)

SWATLD

Tyr

DA

Tyr

DA

99.97 99.82 97.55 1.37

99.57 95.56 100.46 2.65

99.99 99.23 98.41 0.80

98.38 98.82 100.14 0.92

RSD further prove that both of the methods can give accurate results, but the performance of SWATLD is slightly better than that of PARAFAC. 4. Conclusion In this study, we have successfully developed a ‘‘green’’ and fast method for quantitative analysis of tyrosine and dopamine in human urine samples by using second-order calibration methods, for the excitation–emission matrix fluorescence (EEM) data, based on the parallel factor analysis (PARAFAC) and the self-weighted alternating trilinear decomposition (SWATLD) algorithms, respectively. The results revealed that both parallel factor analysis and self-weighted alternating trilinear decomposition methods show good results despite the fluorescence spectra of tyrosine and

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