Dispersive Liquid-Phase Microextraction with Solidification of Floating

Article pubs.acs.org/JAFC

Dispersive Liquid-Phase Microextraction with Solidification of Floating Organic Droplet Coupled with High-Performance Liquid Chromatography for the Determination of Sudan Dyes in Foodstuffs and Water Samples Bo Chen†,‡ and Yuming Huang*,† †

State Key Laboratory Breeding Base of Eco-Environments and Bio-Resources of the Three Gorges Reservoir Region, College of Chemistry and Chemical Engineering, Southwest University, Chongqing 400715, People’s Republic of China ‡ Faculty of Environmental Science and Engineering, Kunming University of Science and Technology, Kunming 650500, People’s Republic of China S Supporting Information *

ABSTRACT: Dispersive liquid-phase microextraction with solidification of floating organic drop (SFO−DLPME) is one of the most interesting sample preparation techniques developed in recent years. In this paper, a new, rapid, and efficient SFO− DLPME coupled with high-performance liquid chromatography (HPLC) was established for the extraction and sensitive detection of banned Sudan dyes, namely, Sudan I, Sudan II, Sudan III, and Sudan IV, in foodstuff and water samples. Various factors, such as the type and volume of extractants and dispersants, pH and volume of sample solution, extraction time and temperature, ion strength, and humic acid concentration, were investigated and optimized to achieve optimal extraction of Sudan dyes in one single step. After optimization of extraction conditions using 1-dodecanol as an extractant and ethanol as a dispersant, the developed procedure was applied for extraction of the target Sudan dyes from 2 g of food samples and 10 mL of the spiked water samples. Under the optimized conditions, all Sudan dyes could be easily extracted by the proposed SFO−DLPME method. Limits of detection of the four Sudan dyes obtained were 0.10−0.20 ng g−1 and 0.03 μg L−1 when 2 g of foodstuff samples and 10 mL of water samples were adopted, respectively. The inter- and intraday reproducibilities were below 4.8% for analysis of Sudan dyes in foodstuffs. The method was satisfactorily used for the detection of Sudan dyes, and the recoveries of the target for the spiked foodstuff and water samples ranged from 92.6 to 106.6% and from 91.1 to 108.6%, respectively. These results indicated that the proposed method is simple, rapid, sensitive, and suitable for the pre-concentration and detection of the target dyes in foodstuff samples. KEYWORDS: Sudan dyes, foodstuffs, dispersive liquid-phase microextraction, solidification of floating organic droplet, food analysis

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industry to improving the appearance of the products.2 However, these lipophilic azo dyes are considered to be genotoxic carcinogens, imposing a threat to public health once entering the food chain.3 For instance, Sudan dyes have been widely contacted by painters, hairdressers, etc., resulting in a high occurrence of bladder cancer.4 In particular, Sudan I, a widely used additive in the food industry, has been reported to induce tumors in the liver of some animals, such as mice, and to show a carcinogenic potential for humans.5 Because of this, the International Agency for Research Cancer (IARC) has listed the Sudan dyes as a category 3 carcinogenic to humans.5 Many countries (including the European Union) have banned the use of Sudan dyes as an additive in the food industry.6 However, Sudan dyes are still illegally used in some areas of Europe and Asia, where the contaminations of foodstuffs by Sudan dyes, in particular Sudan I, are reported.7,8 Therefore, to monitor the illegal use of Sudan dyes in foodstuff samples, development of a

INTRODUCTION

Sudan dyes (Sudan I−IV; Scheme 1) are a class of artificially synthesized fat-soluble azo dyes.1 The azo dyes have been widely used as organic colorants in all aspects of the chemical Scheme 1. Chemical Structures of Four Sudan Dyes (Sudan I, Sudan II, Sudan III, and Sudan IV)

Received: February 6, 2014 Revised: June 3, 2014 Accepted: June 3, 2014

© XXXX American Chemical Society

A

dx.doi.org/10.1021/jf5006403 | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry

Article

In present study, on the basis of solidification of the floating organic droplet, a new, rapid, and efficient dispersive liquidphase microextraction method (SFO−DLPME) coupled with HPLC−UV is proposed for the extraction and sensitive detection of banned Sudan dyes (I, II, III, and IV) in foodstuffs and environmental water samples. Hence, the purposes of this work were (1) to confirm the suitability of the SFO−DLPME for the extraction of Sudan dyes, (2) to optimize the predominant factors affecting the enrichment efficiency of SFO−DLPME for Sudan dyes, and (3) to illustrate the feasibility of the developed method by analyzing the target Sudan dyes in foodstuffs and environmental water samples. The results demonstrate that this approach promises fastness, low consumption of organic solvent, and simplicity and is an interesting alternative for the concentration of organic targets in complex matrices.

simple, rapid, and sensitive method for the detection of the target Sudan dyes in the contaminated food stuffs is desirable. In the past few decades, various methods, including highperformance liquid chromatography (HPLC)−ultraviolet (UV),1,7−11 HPLC−mass spectrometry (MS),12 micellar electrokinetic chromatography,13 and enzyme-linked immunoassay,14 have been constructed for the analysis of Sudan dyes. Considering the complicated sample matrix as well as very low concentrations of Sudan dye residues in real samples, it is highly crucial to simultaneously isolate and enrich Sudan dyes using sample preparation methods before instrumental analysis. Various pretreatment procedures,15−19 in particular, solid-phase extraction (SPE),16−19 have been widely used for the separation and pre-concentration of Sudan dyes in foodstuffs. Recently, a different mode of liquid-phase microextraction format using solidification of floating organic droplet (SFO− LPME), which incorporated sampling, extraction, and concentration into a single step, has attracted much attention.20−24 In this technique, a drop of organic solvent is used for extraction. Thus, in comparison to liquid−liquid extraction (LLE), SFO− LPME is a miniaturized LLE technique.23 When the extraction equilibrium is reached, the organic extract droplet is collected facilely via solidifying it in an ice bath because of the lower density than water and suitable melting point of the used organic extraction solvent. Thus, it has been used to the preconcentration of organic and inorganic targets in various matrices.24−30 However, the extraction time was somewhat long because of the long time required for equilibrium. On the other hand, a dispersive liquid-phase microextraction (DLPME)-based pretreatment process has also gained much attention since the first one developed by Assadi and coworkers in 2006.31 In DLPME, target extraction is realized by injection of a suitable mixture of extractant and dispersant into an aqueous sample, resulting in the formation of an emulsion solution. Because of the high dispersion of the extractant in the aqueous phase, the target extraction is completed within a few seconds because of the markedly enhanced contact area between phases. Thus, in comparison to classic liquid-phase microextraction (LPME), DLPME is a time-saving extraction technology and promises the advantages of simplicity and good extraction efficiency with a high pre-concentration factor.32,33 Unfortunately, the commonly used extraction solvents, including CHCl3, CCl4, chlorobenzene, and CS2, are difficult to be separated from the aqueous solution because the density of these solvents are higher than that of water. In addition, all of them are toxic and not environmentally friendly. Furthermore, the extract requires evaporation and reconstitution in an appropriate solvent for HPLC analysis. This leads to the increased analysis time and the risk of analyte losses.23 On the basis of the advantages of both DLPME and SFO− LPME, a new technique coupling dispersive liquid-phase microextraction and solidification of floating organic droplet (SFO−DLPME) has been proposed for extraction and preconcentration of various organic targets, such as polycyclic aromatic hydrocarbons (PAHs),34 phthalate esters,35 chlorinated anilines,36 and antidepressant drugs,37 from aqueous matrices. It combines the best advantages of DLPME and SFO−LPME, eliminating or reducing their disadvantages. However, to the best of our knowledge, no investigation on the use of SFO−DLPME for extraction and enrichment of Sudan dyes from foodstuffs and water samples has been reported.

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EXPERIMENTAL PROCEDURES

Reagents and Chemicals. Sudan I and Sudan II were supplied by Shanghai Chemical Reagents Co. (Shanghai, China). Sudan III and Sudan IV were provided by Kasei Kogyo (Japan). An appropriate amount of four Sudan dyes was dissolved in methanol to obtain the standard stock solution, and the resulting solution was stored at 4 °C in the dark. The methanol was of HPLC grade and provided by Scharlau Chemie S.A. (Scharlau Chemie S.A., Spain). Humic acid (HA) was purchased from Sigma-Aldrich (Shanghai, China). Ultrapure water was supplied by a Milli-Q water purification system in our laboratory. 1-undecanol, 1-dodecanol, acetone, ethanol, HCl, acetonitrile, NaCl, and all other chemicals used were of analytical reagent grade, unless otherwise stated, and were provided by Chongqing Chemical Reagents Co. (Chongqing, China). Instrumental Analysis. The HPLC system (JASCO Corporation, Japan) consisted of PU-2080i binary pumps, CO-2060 column thermostat, and UV-2075 detector. A Hypersil ODS chromatographic column (150 × 4.6 mm, 5 μm) was supplied by Waters Corporation (Milford, MA) and used for separation of four Sudan dyes. Methanol was used as the mobile phase. The flow rate of methanol was set to 1.0 mL min−1. The column temperature was thermostated at 30 °C. The chromatographic peaks of Sudan I−IV were recorded at 506 nm. The injection volume was 20 μL. SFO−DLPME Procedures. In all experiments, to optimize operational parameters of the SFO−DLPME procedure, 20 μg L−1 Sudan I, Sudan II, and Sudan III and 40 μg L−1 Sudan IV in a sample volume of 10 mL were used. The recovery of the target Sudan dyes was used as the criterion for optimizing the parameter for the proposed method. It was acquired on the basis of the ratio of the obtained content of Sudan dyes to the expected content of Sudan dyes, expressed as a percentage. The finally adopted SFO−DLPME procedure was as follows: after 1 g of NaCl (final concentration of 10% as w/v) and 10 mL of the fortified sample are introduced into a 15 mL centrifuge tube, 100 μL of 1-dodecanol and 400 μL of ethanol are rapidly injected by a microsyringe, leading to the formation of a homogeneous turbid solution. Then, the tube is immediately immersed in a water bath at 70 °C for 20 min, followed by centrifugation at 4500 rpm for 5 min, and finally placed in an ice bath for 5 min. The solid organic phase was transferred to a conical vial with the help of a spoon, and then it melts rapidly at room temperature. The melted organic solvent was mixed with 50 μL of methanol, and then 20 μL of the mixture was used for HPLC−UV analysis. Sample Preparation. The foodstuffs, including tomato sauce, chili sauce, and chili oil samples used in this study, were obtained from the local market. For analysis of food samples, about 2 g of the selected food samples was put into a 10 mL glass centrifuge tube and mixed with 5 mL of ethanol. Then, the resulting mixture was shaken for 20 min at 30 °C and then centrifuged at 4000 rpm for 10 min. The transparent solution was transferred to a sample vial, dried under a nitrogen atmosphere, and then diluted to 10 mL with 10% NaCl (w/ B



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v) solution (pH 7.0). Finally, the obtained sample solution was extracted with the optimized extraction protocols. In this work, several aqueous samples, such as river water samples, tap water samples, and wastewater samples were pretreated by the proposed SFO−DLPME method for the analysis of four Sudan dyes. The river water samples were obtained from the Beibei section (Beibei, Chongqing) of the Jialingjiang River. The tap water samples and the sewage water samples were collected from our laboratory and a constructed wetland system for domestic sewage treatment situated in Southwest University (Beibei, Chongqing), respectively. Prior to analysis, all aqueous samples were passed through 0.45 μm micropore membranes and kept in glass bottles at 4 °C in a refrigerator. The extraction of target Sudan dyes from unspiked and spiked water samples was carried out according to the recommended SFO− DLPME procedures.

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RESULTS AND DISCUSSION Optimization of the SFO−DLPME Procedures. Effect of the Kind and Volume of Extraction Solvent. During the

Figure 3. Effect of the concentration of NaCl. Extract solvent, 100 μL of 1-dodecanol; dispersion solvent, 200 μL of ethanol; extraction temperature, 50 °C; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

Figure 1. Effect of the volume of the extract solvent. Concentration of the sample solution, 20 μg L−1 Sudan I, 20 μg L−1 Sudan II, 20 μg L−1 Sudan III, and 40 μg L−1 Sudan IV; pH of the sample solution, 7.0; sample volume, 10 mL; extractant, 1-dodecanol; dispersion solvent, 400 μL of ethanol; extraction time, 20 min; extraction temperature, 70 °C; ion strength, 10% NaCl (w/v). Error bars stand for 1 standard deviation (n = 3).

Figure 4. Effect of the sample solution pH. Extractant, 100 μL of 1dodecanol; dispersion solvent, 200 μL of ethanol; extraction temperature, 50 °C; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

Figure 5. Effect of the extraction time. Extractant, 100 μL of 1dodecanol; dispersion solvent, 400 μL of ethanol; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

Figure 2. Effect of the volume of the dispersion solvent. Extract solvent, 100 μL of 1-dodecanol; dispersion solvent, ethanol; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

the extraction efficiency was mainly determined by the extraction solvent in SFO−DLPME.35 To improve extraction efficiency, three organic solvents, namely, 1-undecanol (melting point of 13−15 °C), 1-dodecanol (melting point of 22−24 °C), and n-hexadecane (melting point of 18 °C), were considered as

extraction process, selecting a suitable extraction solvent is a very important work using the SFO−DLPME method because C



D

a

Nine replicate determinations of 100 ng g−1 Sudan I, 100 ng g−1 Sudan II, 100 ng g−1 Sudan III, and 100 ng g−1 Sudan IV spiked foodstuff samples in 9 consecutive days. bSix replicate determinations of 100 ng g−1 Sudan I, 100 ng g−1 Sudan II, 100 ng g−1 Sudan III, and 100 ng g−1 Sudan IV spiked foodstuff samples in 1 day.

0.15 0.15 0.10 0.20 2.8 1.6 2.4 2.4 4.0 1.6 3.5 3.1 0.9948 0.9923 0.9921 0.9930 1−1250 1−1250 1−1250 1−1250 0.15 0.15 0.10 0.20 2.3 2.6 4.8 3.2 2.0 2.2 3.8 3.7 0.9990 0.9974 0.9934 0.9967 1−1250 1−1250 1−1250 1−1250 0.15 0.15 0.10 0.20 2.8 1.6 2.4 2.4

interdaya intradayb

4.0 1.6 3.5 3.1

intradayb interdaya linear range (ng g−1)

r2

RSD (%)

chili oil

LOD (ng g−1) r2

0.9964 0.9948 0.9921 0.9952 1−1250 1−1250 1−1250 1−1250

where Cp and Ci are the concentrations of Sudan dyes in the floating phase after extraction and in the sample solution before extraction, respectively. As seen, the PFs decreased with an increase in the volume of 1-dodecanol. However, the quantitative recoveries of Sudan dyes were obtained with 100 μL of 1-dodecanol (Figure 1). Therefore, 100 μL of 1-

I II III IV

(1)

targets

PFs = Cp/C i

RSD (%)

the extraction solvents in this work. Because of its high hydrophobicity, n-hexadecane is not miscible with a common dispersive solvent. Here, only 1-undecanol and 1-dodecanol were evaluated for extraction of Sudan dyes in this work. On the basis of the obtained result (see Figure S1 of the Supporting Information), the best extraction efficiency was obtained when 1-dodecanol was used as the extractant. Hence, 1-dodecanol was selected as the extraction solvent in the present work, and its volume was optimized in the range of 50−250 μL. From Figure 1, the recoveries of Sudan dyes rised rapidly when the volume of 1-dodecanol ranged from 50 to 100 μL and then remained stable ranging from 93.8 to 116.9%. Also, the effect of 1-dodecanol (as an extractant) volume on the pre-concentration factor (PF) was supplied in Table S1 of the Supporting Information. Here, PFs were obtained according to the following formula:

linear range (ng g−1)

Figure 7. Effect of the sample volume. Extractant, 100 μL of 1dodecanol; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

tomato sauce

Table 1. Analytical Characteristics of the SFO−DLPME−HPLC Method for Sudan Dyes in Foodstuffs

LOD (ng g−1)


Figure 6. Effect of the solution temperature. Extractant, 100 μL of 1dodecanol; dispersion solvent, 200 μL of ethanol; other conditions were the same as those in Figure 1. Error bars stand for 1 standard deviation (n = 3).

Sudan Sudan Sudan Sudan

intradayb intradaya r2

RSD (%)

chili sauce

LOD (ng g−1)

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and then remained stable, ranging from 93.0 to 105.2%. A total of 400 μL of ethanol was selected to obtain high extraction efficiency in our work. Effect of the Ion Strength and Sample pH. NaCl was used to study the effect of the ionic strength. The tested concentration of NaCl were in the range of 0−20% (w/v). As shown in Figure 3, the addition of NaCl into sample solution could efficiently improve the extraction efficiency of Sudan dyes when the concentration of NaCl was in the range of 0−10%. However, there is no obvious change of extraction efficiency of Sudan dyes if the concentration of sodium chloride exceeds 10%. This could be explained by the salting-out effect. The solubility of the hydrophobic organics in water samples was decreased when the salt was added. This could cause an increase in extraction efficiency.38,39 On the basis of these results, 10% (w/v) NaCl solution was selected in the subsequent experiments. The effect of solution pH, ranging from 4.0 to 11.0, on the extraction of Sudan dyes by SFO−DLPME was studied. From Figure 4, the extraction recoveries of four Sudan dyes remained a minor increase when solution pH ranged from 4.0 to 7.0 and then remained stable in the pH range from 7.0 to 9.0, above which it decreased. This is probably due to their instability in strong acid and alkali medium.40 Hence, below pH 7.0 and above pH 9.0, the recoveries of Sudan dyes decreased. Considering that the best extraction of Sudan dyes was obtained at pH 7.0, the optimal pH 7.0 was adopted as solution pH in this work. Effect of the Extraction Time and Solution Temperature. In SFO−DLPME, analytes are distributed between the sample solution phase and organic phase. Thus, the mass transfer of analyte between the aqueous phase and the organic solvent was greatly affected by the extraction time. To guarantee the equilibrium between phases as well as the maximal extraction of Sudan dyes, the extract time ranging from 0 to 25 min was investigated. According to Figure 5, the influence of the reaction time on recoveries of Sudan dyes was significant. This implies that the transfer of Sudan dyes from sample solution to extraction solvent is slow, which is probably due to the high fatsoluble property of Sudan dyes. A similar result was reported by Regueiro et al.,26 working on extraction of fat-soluble contaminants by ultrasound-assisted emulsification−microextraction. It was found that extraction time presented a key role in the extraction of polycyclic musk fragrances, musk moskene, di-2-ethylhexyl phthalate (DEHP), and di-n-octyl phthalate (DOP). The recoveries of Sudan dyes increased with extraction time up to 20 min, above which no obvious change in extraction recovery was found. Finally, 20 min was selected for the subsequent experiments. For liquid-phase microextraction, the efficient extraction can be achieved by increasing the temperature because an increasing temperature facilitates mass transfer of the targets from the aqueous phase to the organic phase, leading to shortening the analysis time.41,42 Thus, the temperature effect was investigated from 50 to 75 °C by dispersing 1-dodecanol into the water samples containing Sudan dyes. According to Figure 6, the extraction recoveries of Sudan dyes were increased as the solution temperature increased from 50 to 70 °C. However, the higher temperature (>70 °C) resulted in a mild decrease in recovery. Thus, 70 °C was selected as the optimal temperature. Effect of the Sample Volume. Increasing the volume of sample solution could increase the amount of targets migrated

Figure 8. HPLC−UV chromatograms of food sample extraction with SFO−DLPME: (A) chili sauce and (C) chili oil without spiking and (B) chili sauce and (D) chili oil spiked with Sudan dyes standard at the 0.5 μg g−1 level. I, Sudan I; II, Sudan II; III, Sudan III; and IV, Sudan IV.

Table 2. Recoveries of Four Sudan Dyes in Three Foodstuffs after SFO−DLPME spiked levels (μg g−1) Sudan I

Sudan II

Sudan III

Sudan IV

a

0.05 0.1 0.5 0.05 0.1 0.5 0.05 0.1 0.5 0.05 0.1 0.5

tomato sauce recoveriesa (%) 99.3 93.9 92.8 105.9 101.1 93.4 106.6 95.3 98.8 96.3 99.3 97.9

± ± ± ± ± ± ± ± ± ± ± ±

3.6 2.1 0.3 3.1 3.3 4.8 0.9 5.7 4.5 2.9 4.0 5.5

chili oil recoveriesa (%) 95.6 99.5 96.1 95.6 97.3 103.4 101.4 92.6 103.9 95.6 104.4 104.9

± ± ± ± ± ± ± ± ± ± ± ±

4.1 2.3 0.3 2.9 2.5 1.4 5.8 4.5 0.9 6.9 3.3 5.6

chili sauce recoveriesa (%) 95.6 96.4 99.0 97.0 97.1 94.0 97.0 97.1 102.8 98.1 101.9 102.4

± ± ± ± ± ± ± ± ± ± ± ±

4.1 3.9 1.4 1.8 1.5 4.2 2.7 1.5 0.5 2.4 3.2 1.7

Average of three determinations (mean ± SD; n = 3).

dodecanol was chosen as an appropriate extractant to obtain the quantitative results. Effect of the Kind and Volume of Dispersion Solvent. Several organic solvents, namely, ethanol, acetonitrile, acetone, isopropanol, and methanol, were evaluated as the dispersion solvent for extraction of Sudan dyes. The experimental results indicated that acetone, ethanol, acetonitrile, and methanol gave better dispersion ability than that of isopropanol (see Figure S2 of the Supporting Information). In SFO−DLPME, the major consideration for choosing a dispersion solvent is mainly based on its miscibility with the extractant and aqueous phase. Generally, the good disperser solvent has good miscibility with the extraction solvent and sample solution.30 In the selected dispersion solvents, ethanol, methanol, acetone, and acetonitrile are strong polar solvents, whereas isopropanol is a moderate polar solvent. The polarity of the five organic solvents follows the order of ethanol ≈ methanol > acetone > acetonitrile > isopropanol. Therefore, ethanol, methanol, acetone, and acetonitrile could be miscible with the water well, leading to the better dispersion ability than isopropanol. Because of low toxicity and low cost, finally, ethanol was selected as the dispersion solvent for further investigation. Also, the effect of the volume of ethanol ranging from 0 to 1000 μL was evaluated. From Figure 2, the recoveries of Sudan dyes increased when the volume of ethanol increased to 400 μL E



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Table 3. Comparison of Various Analytical Methods for the Detection of Sudan Dye in Foodstuffs analytical method HPLC−DAD

HPLC−UV HPLC−UV

UPLC−MS/MS HPLC−UV HPLC−UV HPLC−UV HPLC−DAD HPLC−UV HPLC−UV HPLC−DAD HPLC−UV LC−MS/MS LC−MS/MS LC−MS/MS

HPLC−UV a

real sample chili- and curry-based sauces and powdered spices tomato sauce and sausage chili powder, dry red pepper, ketchup, and sausage spices and chili foodstuffs red chili powder sausage chili sauce chili products chili powder egg yolk tomato sauce, vinegar sauce, and meat sauce chili powder chili products animal-derived foods and eggs of hen and duck chili powder, pepper chili sauce, sausage, and chili meat tomato sauce, chili oil, and chili sauce

extraction time (min)


RSD (%)

reference

solvent extraction

>60

0−20000

51−100

200−2000

60 30 about 10 >20 >20 about 40 >20 about 10

10−50000 not given 5−2000 not given 50−2500b 20−20000b 0.02−2 0.028−2

87−106 80.7−85.5 86.3−107.5 87.5−103.4 93.2−103 76.8−109.5 87.2−103.5 86−108

0.5−100 2−4 1−5a 3.3−5.0 4.1−5.8 7.0−8.2 2.3−6.1 2.7−7.4

20 12 >10

25−5000 5−5500 0.1−10b

79.9−87.8 72−103 61.9−87.4

1.6−6.2 3−24 0.03−0.12

4.8−9.1 1−15 95%) were obtained when the aqueous-phase volume varied from 5 to 11 mL. Finally, the volume of 10 mL was selected and used as the optimal sample volume in our work. Application of the SFO−DLPME−HPLC Method to Real Foodstuff Samples. For the determination of Sudan dyes in foodstuffs, various solvents, including methanol,10,16 acetonitrile,1,6,17,43 and ethanol,14,44,45 have been used alone for extraction of Sudan dyes in various foodstuffs. Considering that the recovery of Sudan IV could be improved using ethanol as an extraction solvent44 and the ratio of the sample weight to the volume of the extractant has no obvious effect on extraction recovery,46 in our work, about 2 g of the selected food samples was extracted with 5 mL of ethanol. To evaluate the potential application of the SFO−DLPME−HPLC method for the detection of Sudan dyes in real samples, the analytical characteristics were evaluated using the spiked food samples. The results are indicated in Table 1. As seen, the linear ranges, in terms of the peak area and sample concentration for four Sudan dyes, were in the range of 1−1250 ng g−1. The correlation coefficients (r2) were in the range of 0.9921− 0.9990, demonstrating the good linearity of the proposed method. The limit of detection (LOD; S/N = 3) was in the range of 0.10−0.20 ng g−1. Validation of the SFO−DLPME− HPLC method was evaluated by determining the recoveries of the spiked foodstuffs with Sudan I−IV at 0.05, 0.10, and 0.50 μg g−1. The precision of the SFO−DLPME−HPLC method was evaluated by carrying out analysis of the spiked foodstuffs F



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(2) Nohynek, G. J.; Fautz, R.; Benech-Kieffer, F.; Toutain, H. Toxicity and human health risk of hair dyes. Food Chem. Toxicol. 2004, 42, 517−543. (3) Golka, K.; Kopps, S.; Myslak, Z. W. Carcinogenicity of azo colorants: Influence of solubility and bioavailability. Toxicol. Lett. 2004, 151, 203−210. (4) Ahlstrom, L. H.; Eskilsson, C. S.; Bjorklund, E. Determination of banned azo dyes in consumer goods. TrAC, Trends Anal. Chem. 2005, 24, 49−56. (5) Stiborová, M.; Martínek, V.; Rýdlová, H.; Hodek, P.; Frei, E. Sudan I is a potential carcinogen for humans: Evidence for its metabolic activation and detoxication by human recombinant cytochrome P450 1A1 and liver microsomes. Cancer Res. 2002, 62, 5678−5684. (6) Anibal, C. V. Di; Odena, M.; Ruisánchez, I.; Callao, M. P. Determining the adulteration of spices with Sudan I-II-II-IV dyes by UV−visible spectroscopy and multivariate classification techniques. Talanta 2009, 79, 887−892. (7) Zacharis, C. K.; Kika, F. S.; Tzanavaras, P. D.; Rigas, P.; Kyranas, E. R. Development and validation of a rapid HPLC method for the determination of five banned fat-soluble colorants in spices using a narrow-bore monolithic column. Talanta 2011, 84, 480−486. (8) He, L.; Su, Y.; Fang, B.; Shen, X.; Zeng, Z.; Liu, Y. Determination of Sudan dye residues in eggs by liquid chromatography and gas chromatography−mass spectrometry. Anal. Chim. Acta 2007, 594, 139−146. (9) Zhao, C.; Zhao, T.; Liu, X.; Zhang, H. A novel molecularly imprinted polymer for simultaneous extraction and determination of Sudan dyes by on-line solid phase extraction and high performance liquid chromatography. J. Chromatogr. A 2010, 1217, 6995−7002. (10) Yan, H.; Gao, M.; Qiao, J. New ionic liquid modified polymeric microspheres for solid-phase extraction of four Sudan dyes in foodstuff samples. J. Agric. Food Chem. 2012, 60, 6907−6912. (11) Ertaş, E.; Ö zer, H.; Alasalvar, C. A rapid HPLC method for determination of Sudan dyes and Para Red in red chilli pepper. Food Chem. 2007, 105, 756−760. (12) Schummer, C.; Sassel, J.; Bonenberger, P.; Moris, G. Low-level detections of Sudan I, II, III and IV in spices and chili-containing foodstuffs using UPLC-ESI-MS/MS. J. Agric. Food Chem. 2013, 61, 2284−2289. (13) Fukuji, T. S.; Castro-Puyana, M.; Tavares, M. F.; Cifuentes, A. Fast determination of Sudan dyes in chili tomato sauces using partial filling micellar electrokinetic chromatography. J. Agric. Food Chem. 2011, 59, 11903−11909. (14) Wang, Y.; Yang, H.; Wang, B.; Deng, A. A sensitive and selective direct competitive enzyme-linked immunosorbent assay for fast detection of Sudan I in food samples. J. Sci. Food Agric. 2011, 91, 1836−1842. (15) Liu, W.; Zhao, W. J.; Chen, J. B.; Yang, M. M. A cloud point extraction approach using Triton X-100 for the separation and preconcentration of Sudan dyes in chilli powder. Anal. Chim. Acta 2007, 605, 41−45. (16) Long, C.; Mai, Z.; Yang, Y.; Zhu, B.; Xu, X.; Lu, L.; Zou, X. Synthesis and characterization of a novel molecularly imprinted polymer for simultaneous extraction and determination of watersoluble and fat-soluble synthetic colorants in chilli products by solid phase extraction and high performance liquid chromatography. J. Chromatogr. A 2009, 1216, 8379−8385. (17) Yan, H.; Qiao, J.; Wang, H.; Yang, G.; Row, K. H. Molecularly imprinted solid-phase extraction combined with ultrasound-assisted dispersive liquid−liquid microextraction for the determination of four Sudan dyes in sausage samples. Analyst 2011, 136, 2629−2634. (18) Zhang, Z.; Xu, S.; Li, J.; Xiong, H.; Peng, H.; Chen, L. Selective solid-phase extraction of Sudan I in chili sauce by single-hole hollow molecularly imprinted polymers. J. Agric. Food Chem. 2012, 60, 180− 187. (19) Puoci, F.; Garreffa, C.; Lemma, F.; Muzzalupo, R.; Splizzirri, U. G.; Picci, N. Molecularly imprinted solid phase extraction for detection of Sudan I in food matrices. Food Chem. 2005, 93, 349−353.

DLPME method. From Figure S3 of the Supporting Information, the quantitative recovery of Sudan dyes could be reached at less than the 5 mg L−1 level of humic acid. The humic acid contents in the natural waters are less than 4 mg L−1 for river water samples;54 thus, it did not interfere with Sudan dye determination, indicating the good selectivity of the proposed SFO−DLPME method. Second, under the selected extraction conditions indicated above, the analytical performance was tested. The analytical data, such as linear range, correlation coefficients, detection limits, and precision, are shown in Table S3 of the Supporting Information. As seen, good linear relationships could be obtained for four Sudan dyes between the peak area and the sample concentration from 0.2 to 500 μg L−1. The LOD was 0.03 μg L−1. The recoveries of Sudan dyes varied in the range of 91.1−108.6% when 10 mL of aqueous sample at three spiked concentration levels (5, 10, and 20 μg L−1 Sudan dyes) were tested (see Table S4 of the Supporting Information), indicating the utility of the proposed method for the detection of Sudan dyes in environmental water samples. In conclusion, for the first time, we demonstrated that the proposed SFO−DLPME−HPLC method can be used as a useful protocol for pre-concentration and detection of trace Sudan dyes. Our results indicate that SFO−DLPME is an efficient sample preparation technique for the extraction of banned Sudan dyes (I, II, III, and IV) in a complex matrix. The proposed method promises simplicity, less organic solvent consumption, and high sensitivity. Also, it allows for the determination of Sudan dyes in complex matrices with good accuracy and reproducibility.

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ASSOCIATED CONTENT

* Supporting Information S

Pre-concentration factors of Sudan dyes by the SFO−DLPME method (Table S1), recoveries of Sudan dyes in the presence of foreign species (Sudan I−IV concentrations: 20, 20, 20, and 40 μg L−1, respectively) (Table S2), analytical characteristics of the SFO−DLPME−HPLC method for Sudan dyes in aqueous solution (Table S3), determination results of Sudan dyes in environmental water samples (Table S4), effect of different extractants on the extraction efficiency of Sudan dyes (Figure S1), effect of different dispersion solvents on the extraction efficiency of Sudan dyes (Figure S2), and effect of humic acid on the extraction efficiency of Sudan dyes (Figure S3). This material is available free of charge via the Internet at http:// pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*Telephone/Fax: +86-23-68254843. E-mail: [email protected]. Funding

Financial support by the National Natural Science Foundation of China (21277111) is gratefully acknowledged. Notes

The authors declare no competing financial interest.

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REFERENCES

(1) Cornet, V.; Govaert, Y.; Moens, G.; Van Loco, J.; Degroodt, J.-M. Development of a fast analytical method for the determination of Sudan dyes in chili- and curry-containing foodstuffs by highperformance liquid chromatography−photodiode array detection. J. Agric. Food Chem. 2006, 54, 639−644. G



Article

(20) Khalili-Zanjani, M. R.; Yamini, Y.; Shariati, S.; Jönsson, J. A. A new liquid-phase microextraction method based on solidification of floating organic drop. Anal. Chim. Acta 2007, 585, 286−293. (21) Sobhi, H. R.; Yamini, Y.; Esrafili, A.; Abadi, R. H. H. B. Suitable conditions for liquid-phase microextraction using solidification of a floating drop for extraction of fat-soluble vitamins established using an orthogonal array experimental design. J. Chromatogr. A 2008, 1196− 1197, 28−32. (22) Khalili-Zanjani, M. R.; Yamini, Y.; Yazdanfar, N.; Shariati, S. Extraction and determination of organophosphorus pesticides in water samples by a new liquid phase microextraction−gas chromatography− flame photometric detection. Anal. Chim. Acta 2008, 606, 202−208. (23) Vera-Avila, L. E.; Rojo-Portillo, T.; Covarrubias-Herrera, R.; Peña-Alvarez, A. Capabilities and limitations of dispersive liquid− liquid microextraction with solidification of floating organic drop for the extraction of organic pollutants from water samples. Anal. Chim. Acta 2013, 805, 60−69. (24) Farahani, H.; Yamini, Y.; Shariati, S. M.; Khalili-Zanjani, R.; Mansour-Baghahi, S. Development of liquid phase microextraction method based on solidification of floated organic drop for extraction and preconcentration of organochlorine pesticides in water samples. Anal. Chim. Acta 2008, 626, 166−173. (25) Faraji, H.; Tehrani, M. S.; Husain, S. W. Pre-concentration of phenolic compounds in water samples by novel liquid−liquid microextraction and determination by gas chromatography−mass spectrometry. J. Chromatogr. A 2009, 1216, 8569−8574. (26) Regueiro, J.; Llompart, M.; Garcia-Jares, C.; GarciaMonteagudo, J. C.; Cela, R. Ultrasound-assisted emulsification− microextraction of emergent contaminants and pesticides in environmental waters. J. Chromatogr. A 2008, 1190, 27−38. (27) Dadfarnia, S.; Salmanzadeh, A. M.; Shabani, A. M. H. A novel separation/preconcentration system based on solidification of floating organic drop microextraction for determination of lead by graphite furnace atomic absorption spectrometry. Anal. Chim. Acta 2008, 623, 163−167. (28) Bidabadi, M. S.; Dadfarnia, S.; Shabani, A. M. H. Solidified floating organic drop microextraction (SFODME) for simultaneous separation/preconcentration and determination of cobalt and nickel by graphite furnace atomic absorption spectrometry (GFAAS). J. Hazard. Mater. 2009, 166, 291−296. (29) Rivas, R. E.; Lopez-Garcia, I.; Hernandez-Cordoba, M. Microextraction based on solidification of a floating organic drop followed by electrothermal atomic absorption spectrometry for the determination of ultratraces of lead and cadmium in waters. Anal. Methods 2010, 2, 225−230. (30) Yamini, Y.; Rezaee, M.; Khanchi, A.; Faraji, M.; Saleh, A. Dispersive liquid−liquid microextraction based on the solidification of floating organic drop followed by inductively coupled plasma−optical emission spectrometry as a fast technique for the simultaneous determination of heavy metals. J. Chromatogr. A 2010, 1217, 2358− 2364. (31) Rezaee, M.; Assadi, Y.; Milani Hosseini, M. R.; Aghaee, E.; Ahmadi, F.; Berijani, S. Determination of organic compounds in water using dispersive liquid−liquid microextraction. J. Chromatogr. A 2006, 1116, 1−9. (32) Birjandi, A. P.; Bidari, A.; Rezaei, F.; Hosseini, M. R. M.; Assadi, Y. Speciation of butyl and phenyltin compounds using dispersive liquid−liquid microextraction and gas chromatography−flame photometric detection. J. Chromatogr. A 2008, 1193, 19−25. (33) Xiong, J.; Hu, B. Comparison of hollow fiber liquid phase microextraction and dispersive liquid−liquid microextraction for the determination of organosulfur pesticides in environmental and beverage samples by gas chromatography with flame photometric detection. J. Chromatogr. A 2008, 1193, 7−18. (34) Xu, H.; Ding, Z.; Lv, L.; Song, D.; Feng, Y.-Q. A novel dispersive liquid−liquid microextraction based on solidification of floating organic droplet method for determination of polycyclic aromatic hydrocarbons in aqueous samples. Anal. Chim. Acta 2009, 636, 28−33.

(35) Kamarei, F.; Ebrahimzadeh, H.; Yamini, Y. Optimization of ultrasound-assisted emulsification microextraction with solidification of floating organic droplet followed by high performance liquid chromatography for the analysis of phthalate esters in cosmetic and environmental water samples. Microchem. J. 2011, 99, 26−33. (36) Ramkumar, A.; Ponnusamy, V. K.; Jen, J.-F. Rapid analysis of chlorinated anilines in environmental water samples using ultrasound assisted emulsification microextraction with solidification of floating organic droplet followed by HPLC-UV detection. Talanta 2012, 97, 279−284. (37) Ebrahimzadeh, H.; Saharkhiz, Z.; Tavassoli, M.; Kamarei, F.; Asgharinezhad, A. A. Ultrasound-assisted emulsification microextraction based on solidification of floating organic droplet combined with HPLC-UV for the analysis of antidepressant drugs in biological samples. J. Sep. Sci. 2011, 34, 1275−1282. (38) Pan, L.; Adams, M.; Pawliszyn, J. Determination of fatty acids using solid-phase microextraction. Anal. Chem. 1995, 67, 4396−4403. (39) Zhang, Z.; Duan, H.; Zhang, L.; Chen, X.; Liu, W.; Chen, G. Direct determination of anabolic steroids in pig urine by a new SPME−GC−MS method. Talanta 2009, 78, 1083−1089. (40) Sun, S.; Wang, Y.; Yu, W.; Zhao, T.; Gao, S.; Kang, M.; Zhang, Y.; Zhang, H.; Yu, Y. Determination of Sudan dyes in red wine and fruit juice using ionic liquid-based liquid−liquid microextraction and high performance liquid chromatography. J. Sep. Sci. 2011, 34, 1730− 1737. (41) Bagheri, H.; Saber, A.; Mousavi, S. R. Immersed solvent microextraction of phenol and chlorophenols from water samples followed by gas chromatography−mass spectrometry. J. Chromatogr. A 2004, 1046, 27−33. (42) López-Blanco, M. C.; Blanco-Cid, S.; Cancho-Grande, B.; SimalGándara, J. Application of single-drop microextraction and comparison with solid-phase microextraction and solid-phase extraction for the determination of α- and β-endosulfan in water samples by gas chromatography−electron-capture detection. J. Chromatogr. A 2003, 984, 245−252. (43) Qi, P.; Zeng, T.; Wen, Z.; Liang, X.; Zhang, X. Interference-free simultaneous determination of Sudan dyes in chili foods using solid phase extraction coupled with HPLC−DAD. Food Chem. 2011, 125, 1462−1467. (44) Zhang, Y.; Zhang, Z.; Sun, Y. Development and optimization of an analytical method for the determination of Sudan dyes in hot chilli pepper by high-performance liquid chromatography with on-line electrogenerated BrO−-luminol chemiluminescence detection. J. Chromatogr. A 2006, 1129, 34−40. (45) Zhang, J.; Shao, J.; Guo, P.; Huang, Y. A simple and fast Fe3O4 magnetic nanoparticles-based dispersion solid phase extraction of Sudan dyes from food and water samples coupled with highperformance liquid chromatography. Anal. Methods 2013, 5, 2503− 2510. (46) Rebane, R.; Leito, I.; Yurchenko, S.; Herodes, K. A review of analytical techniques for determination of Sudan I−IV dyes in food matrixes. J. Chromatogr. A 2010, 1217, 2747−2757. (47) Fan, Y.; Chen, M.; El-Sepai, F.; Wang, K.; Zhu, Y.; Ye, M. Ionic liquids extraction of Para Red and Sudan dyes from chilli powder, chilli oil and food additive combined with high performance liquid chromatography. Anal. Chim. Acta 2009, 650, 65−69. (48) Yan, H.; Wang, H.; Qiao, J.; Yang, G. Molecularly imprinted matrix solid-phase dispersion combined with dispersive liquid−liquid microextraction for the determination of four Sudan dyes in egg yolk. J. Chromatogr. A 2011, 1218, 2182−2188. (49) López-Jiménez, F. J.; Rubio, S.; Pérez-Bendito, D. Supramolecular solvent-based microextraction of Sudan dyes in chillicontaining foodstuffs prior to their liquid chromatography-photodiode array determination. Food Chem. 2010, 121, 763−769. (50) Piao, C.; Chen, L. Separation of Sudan dyes from chilli powder by magnetic molecularly imprinted polymer. J. Chromatogr. A 2012, 1268, 185−190. (51) Calbiani, F.; Careri, M.; Elviri, L.; Mangia, A.; Pistarà, L.; Zagnoni, I. Development and in-house validation of a liquid H



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chromatography−electrospray−tandem mass spectrometry method for the simultaneous determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli products. J. Chromatogr. A 2004, 1042, 123− 130. (52) Chen, D.; Li, X.; Tao, Y.; Pan, Y.; Wu, Q.; Liu, Z.; Peng, D.; Wang, X.; Huang, L.; Wang, Y.; Yuan, Z. Development of a liquid chromatography−tandem mass spectrometrywith ultrasound-assisted extraction method for the simultaneous determination of Sudan dyes and their metabolites in the edible tissues and eggs of food-producing animals. J. Chromatogr. B: Anal. Technol. Biomed. Life Sci. 2013, 939, 45−50. (53) Sun, H.; Wang, F.; Ai, L. Determination of banned 10 azo-dyes in hot chili products by gel permeation chromatography−liquid chromatography−electrospray ionization-tandem mass spectrometry. J. Chromatogr. A 2007, 1164, 120−128. (54) Li, L.; Huang, Y.; Wang, Y.; Wang, W. Hemimicelle capped functionalized carbon nanotubes-based nanosized solid-phase extraction of arsenic from environmental water samples. Anal. Chim. Acta 2009, 631, 182−188.

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Dispersive Liquid-Phase Microextraction with Solidification of Floating

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