5.2
Impact Factor
Generic selectors
Exact matches only
Search in title
Search in content
Post Type Selectors
Search in posts
Search in pages
Filter by Categories
Corrigendum
Current Issue
Editorial
Erratum
Full Length Article
Full lenth article
Letter to Editor
Original Article
Research article
Retraction
Retraction notice
Review
Review Article
SPECIAL ISSUE: ENVIRONMENTAL CHEMISTRY
5.3
Impact Factor
Generic selectors
Exact matches only
Search in title
Search in content
Post Type Selectors
Search in posts
Search in pages
Filter by Categories
Corrigendum
Current Issue
Editorial
Erratum
Full Length Article
Full lenth article
Letter to Editor
Original Article
Research article
Retraction
Retraction notice
Review
Review Article
SPECIAL ISSUE: ENVIRONMENTAL CHEMISTRY
View/Download PDF

Translate this page into:

Original article
12 (
8
); 5223-5233
doi:
10.1016/j.arabjc.2016.12.020

Ultrasound-assisted emulsification microextraction coupled with salt-induced demulsification based on solidified floating organic drop prior to HPLC determination of Sudan dyes in chili products

, , ,
a Materials Chemistry Research Center, Department of Chemistry and Center of Excellence for Innovation in Chemistry, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand
b Department of Plant Science and Agricultural Resources, Faculty of Agriculture, Khon Kaen University, Khon Kaen 40002, Thailand

⁎Corresponding author at: 123 Mittraphab Road, M. 16, T. Ni-Muang, A. Muang, Khon Kaen University, Department of Chemistry, Faculty of Science, Khon Kaen 40002, Thailand. Fax: +66 43202373. sakcha2@kku.ac.th (Saksit Chanthai)

Received: , Accepted: ,
© The Author(s) 2017
Disclaimer:
This article was originally published by Elsevier and was migrated to Scientific Scholar after the change of Publisher.

Peer review under responsibility of King Saud University.

Abstract

Abstract

  • A novel solidified floating organic drop microextraction was developed for Sudan dyes.

  • Ultrasound was used for green sample preparation and the UAEME-SID-SFOD method.

  • The proposed UAEME-SID-SFOD method does not need a disperser solvent and centrifugation step.

  • The proposed method is simple, rapid, cost-effective, and environmental friendly.

Abstract

This study aimed to analyze the presence of Sudan I–IV in chili products using high performance liquid chromatography and summarized the data performance after ultrasound-assisted emulsification microextraction and salt-induced demulsification with solidified floating organic drop (UAEME-SID-SFOD). After demulsification, the low density solvent was floating instantaneously solidified in an ice-bath; any remaining liquid phase was removed. The solidified part was quickly melted at room temperature and used for analysis. Parameters such as linearity (0.5–2500 ng mL−1, R2 ⩾ 0.999), limits of detection (0.16–0.24 ng mL−1), and limits of quantification (0.53–0.80 ng mL−1) were used to validate this method. The pre-concentration factor for the 5 mL sample solution was 62. Under optimum conditions, the recoveries ranged between 91% and 107%, with relative standard deviations ranging from 1.8% to 3.5% for intra-day (n = 7) and 4.8% to 6.3% for inter-day (n = 5 × 3) analyses. This novel proposed method has great potential applications in most analytical laboratories that use general instrumentation. This method is easy, rapid, and environmentally friendly. In addition, it does not use any disperser solvent and can eliminate the centrifugation step for aqueous and organic phase separation. Therefore, it is expected that the UAEME-SID-SFOD would be a useful method for various analytical sample extraction purposes.

Keywords

Ultrasound
Emulsification
Demulsification
Solidified floating organic drop microextraction
Sudan dyes
Chili products
1

1 Introduction

Sudan dyes (Sudan I–IV; Table 1), are a group of synthetic colorants that belong to the family of lipophilic azo (—N⚌N—) dyes. Traditionally, they are used in products such as waxes, printing inks, polishes, plastics, and oils to improve their appearance (Li et al., 2014). According to the International Agency for Research on Cancer, these dyes are classified as category 3 carcinogens because the dyes and their metabolites possibly cause bladder and liver cancer in humans (Noguerol-Cal et al., 2008). Many countries, including the European Union (EU), have banned the use of Sudan dyes as an additive in the food industry (Di Anibal et al., 2009), and the maximum residue limits (MRLs) for Sudan dyes in any food materials have been set at 0.5–1 mg kg−1; any food material having a higher concentration of Sudan dyes than the MRLs should be withdrawn from the market (Yu et al., 2015). Therefore, to monitor the illegal use of Sudan dyes in food samples, a simple, rapid, and sensitive method should be developed for dye detection in contaminated food products.

Table 1 Chemical structure, formula, MW, and log Kow of Sudan I–IV dyes and the three decanols.
Compound Structure Formula MW (g/mol) log Kow
Sudan I C16H12N2O 248.3 5.51
Sudan II C18H16N2O 276.3 6.60
Sudan III C22H16N4O 352.4 7.63
Sudan IV C24H20N4O 380.4 8.72
1-Dodecanol C12H26O 186.3 5.13
1-Undecanol C11H24O 172.3 4.72
2-Dodecanol C12H26O 186.3 4.70

MW; molecular weight, Kow; partition coefficient.

However, prior to analysis by high performance liquid chromatography (HPLC) to determine trace levels of Sudan I–IV in various samples, the dyes need to be extracted and preconcentrated. Techniques for this purpose have included solid-phase extraction (Qi et al., 2011), on-line molecularly imprinted polymer solid-phase extraction (Zhao et al., 2010), matrix solid-phase dispersion extraction (Enríquez-Gabeiras et al., 2012), supramolecular solvent-based microextraction (López-Jiménez et al., 2010), ionic liquid extraction (Fan et al., 2009), gel permeation chromatography (Zhu et al., 2014) and ultrasound-assisted liquid–liquid extraction (Zacharis et al., 2011).

To remove and/or minimize the environmental impact caused by chemical analysis, green analytical chemistry, a type of green chemistry has been developed as a sustainable concept (Tobiszewski et al., 2010). Over the last decade, ultrasonication has gained popularity as an efficient tool in analytical laboratories. It can be implemented in various ways to enhance the analytical processes, from general uses (e.g., degassing (Eskin, 1995) and cleaning (Tuziuti, 2016) to more specific ones, extraction (Moorthy et al., 2017), derivatization (Bhavani et al., 2019; Prasad et al., 2019), homogenization (Abbas et al., 2014), and emulsification (Alzorqi et al., 2016; Ngan et al., 2019)). In particular, ultrasound-assisted emulsification (UAE) is highly efficient (Saleh et al., 2016; Li et al., 2017). The effect of an ultrasound is based on droplet disruption in the sonicated liquid–liquid system as a result of cavitation. In addition, ultrasound effectively assists in emulsification, without altering the chemical characteristics of the system. Although UAE is now widely used in analytical laboratories to bring solutions into contact, it is occasionally studied or optimized during the development of analytical methods (Ramandi et al., 2017; Prakash Maran et al., 2017).

Recently, a demulsification step following liquid–liquid microextraction (LLME) has been used due to its simplicity and fast extraction. A demulsifier facilitates aqueous and organic phase separation, without the need of centrifugation. In general, demulsifiers are water miscible organic solvents that are mostly used as the dispersive solvent. However, due to decreased partition coefficients of the analytes into the extraction solvent, this approach is not widely used (Chen et al., 2010). Therefore, it is still an interesting challenge to develop demulsified LLME with suitable demulsifiers. Electrolytes or ionic salts, as an alternative choice for demulsifiers, can reduce the zeta potential of the double layer of the partition solution, thereby decreasing the stability of hydrophobic colloids (an emulsion solution) and increasing the ionic strength of the solution. This results in flocculation and coalescence of fine droplets of the extraction solvent (Cañizares et al., 2008). Therefore, the two phases can be separated by adding salts into the emulsion.

A small number of research studies based on solidified floating organic drop microextraction were reported (Mohammadi et al., 2016). A few microliters of the organic solvent (lighter than water) is injected into the aqueous sample and then the mixture is agitated. After centrifugation, the tube is immersed into an ice-bath and then the floated organic solvent is solidified on top of the aqueous surface. The liquid phase is removed and the solidified solvent is quickly melted at room temperature and used for further analysis. The method has advantages including simplicity, good accuracy and precision, short extraction time, low cost, and minimum consumption of organic solvent (Sricharoen et al., 2016).

Here, we designed a novel extraction method for Sudan dyes detection based on UAE microextraction followed by salt-induced demulsification with solidified floating organic drop (UAEME-SID-SFOD) prior to HPLC. In addition, we investigated and optimized the experimental parameters affecting the extraction performance of Sudan I–IV dyes, such as extraction solvent and its volume, pH of sample, ultrasonication time and salt addition for emulsification and demulsification. To address the large volume problem of the conventional solvent demulsified LLME, salts were used as the demulsifier. Subsequently, this method was used to determine Sudan dyes in chili products. Furthermore, the green extraction method, i.e., the ultrasound-assisted extraction, was optimized in detail and used for pre-treatment of food samples prior to the UAEME-SID-SFOD procedure.

2

2 Materials and methods

2.1

2.1 Chemicals and reagents

Sudan dyes I–IV and extraction solvents (1-undecanol, 1-dodecanol, and 2-dodecanol) were obtained from Sigma–Aldrich (USA). In addition, the following chemicals were used: sodium chloride (APS, Australia), sodium sulfate (Carlo Erba, France), magnesium chloride hexahydrate (Carlo Erba, France), magnesium sulfate anhydrous (Panreac, Spain), aluminum chloride hexahydrate (Ajax Finechem Pty Ltd., New Zealand), and aluminum sulfate hexahydrate (Fluka, Switzerland). Dimethyl sulfoxide (99.9%, AR grade) and methanol (HPLC grade) were obtained from RCI Labscan Ltd. (Thailand). Acetone, acetonitrile, hexane, and ethanol were purchased from QRec™ (New Zealand). Deionized water with a resistivity of 18.2 MΩ cm was obtained from a Millipore water purification system (Molsheim, France).

2.2

2.2 Apparatus

Ultrasound-assisted extraction for sample pre-treatment and UAEME was performed using an ultrasonic bath (Sonorex Digitec DT 510 H, Bandelin, Germany) at 35 kHz (640 W). The HPLC system (LC–20A, Shimadzu, Japan) included an LC–20AD HPLC pump (Shimadzu, Japan), a Rheodyne injector with 20-μL sample loop, and a photodiode array detector (SPD-M20A, Shimadzu, Japan). Data were acquired using the Empower software. The HPLC-PDA was recorded at the maximum absorption wavelength of 506 nm. The separation of Sudan I–IV was carried out on A Zorbax-ODS C18 column (4.6 × 250 mm, 5 μm, Shimadzu, Japan) with an isocratic elution using 100% methanol as the mobile phase with a flow rate of 1 mL min−1. The injection volume was 20 μL. Quantitative data were obtained from the peak area using commercial Sudan dyes as the external standard. The four Sudan dyes were separated within 15 min, with the following elution order of their retention time: Sudan I (5.66 min), Sudan II (7.55 min), Sudan III (9.02 min), and Sudan IV (13.82 min).

2.3

2.3 Standard solution preparation

A stock solution of these dyes was prepared in ethanol at a concentration of 100 μg mL−1 and kept at −20 °C. The working solution was then prepared by an appropriate dilution of the stock standard solution with ethanol.

2.4

2.4 Sample preparation

Chili products including chili oil, chili sauce, chili paste, and chili powder, were randomly purchased from local markets at Khon Kaen, northeast Thailand. Prior to Sudan I–IV determination, the ultrasound-assisted extraction method for these samples was optimized based on a previously reported (D. Chen et al., 2013). The sample (0.1 g) was accurately weighed in a 20-mL amber glass bottle, and then, extracted with 5 mL ethanol at 30 °C for 5 min. Subsequently, the extract was centrifuged at 4000 rpm for 5 min to remove any solid material. A vacuum evaporator was used to evaporate the supernatant to dryness at 50 °C. The resulting residue was re-dissolved homogenously with 5 mL deionized water prior to the extraction by UAEME-SID-SFOD.

2.5

2.5 UAEME-SID-SFOD procedure

Fig. 1 illustrates the graphical procedure of UAEME-SID-SFOD. First, a 5-mL aliquot of a standard solution of Sudan I–IV mixture or sample solution was added in a test tube. The ionic strength of the solution was adjusted by adding 6% (w/v) NaCl. Next, 80 μL 1-dodecanol (an extraction solvent) was added to the solution. After extracting in an ultrasound bath at 40 °C for 1 min, the emulsion was formed and the Sudan dyes were extracted into the extraction solvent droplets. Subsequently, addition of 3% (w/v) AlCl3, as a demulsifier, resulted in quick breaking up of the emulsion into two clear separation phases. After the test tube was immersed in an ice-water bath for 5 min, the low density organic solvent that was floating instantaneously solidified on top of the aqueous solution; any remaining liquid phase was removed using a syringe. Next, the solidified ring was quickly melted at room temperature, and the floating extraction phase was then injected into the HPLC.

Graphical procedure of the UAEME-SID-SFOD method.
Figure 1 Graphical procedure of the UAEME-SID-SFOD method.

2.6

2.6 Statistical analysis

Data results were given as the mean ± standard deviation (SD) of three measurements (n = 3). In all graphs, a linear regression analysis was conducted using Microsoft Excel 2013 software.

3

3 Results and discussion

3.1

3.1 Ultrasound-assisted extraction optimization for sample preparation

Ultrasound-assisted extraction enhancement of the extraction yield is primarily attributed to the cavitation bubbles in the solvent produced by the ultrasonic wave passage that causes microjet impacts. In addition, a shockwave-induced damage of sample keeps releasing cell contents into the solvent (Cvjetko Bubalo et al., 2016). We first investigated and optimized the effect of the extraction parameters for sample preparation. Chili paste was used as a model sample. Each experiment was performed in triplicate and the means of the results in terms of percentage recoveries (%R) were used for optimization and were calculated as follows:

(1)
% R = C spiked - C nonspiked C added × 100 where Cspiked, Cnonspiked, and Cadded are the concentration of the analyte after addition of a known amount of standard in the real sample, concentration of the analyte in the real sample, and concentration of a known amount of standard that was spiked in the real sample, respectively (Niazi et al., 2015).

3.1.1

3.1.1 Effects of sample amount and volume of the extraction solvent

Various concentrations of chili paste were obtained by dissolving 0.1, 0.2, 0.3, 0.4, and 0.5 g of sample into 10 mL of methanol. The %R values of all the Sudan dyes were significantly similar ranging from 0.1 to 0.5 g. Therefore, a sample weight of 0.1 g was used for further extraction optimization. The effect of different volumes of methanol (5, 7.5, 10, and 12.5 mL) was studied using 0.1 g chili paste. All the volumes had good extraction %R, but a minimum of 5 mL methanol was needed. Therefore, the extraction solvent volume was set at 5 mL for all subsequent experiments.

3.1.2

3.1.2 Effects of extraction time and temperature

Both extraction time and temperature affect the optimum conditions, and it is desirable to have conditions that are cost-effective. The concentration of dye extracts continuously increased with the increasing extraction time (Fig. 2a). After 5 min of extraction time, no significant differences were observed in the %R of the dyes. Therefore, 5 min was set as the suitable ultrasonic extraction time. In general, the temperature had a positive effect on the extraction yield. The results show no difference in the effect of temperature ranging from 30 °C to 50 °C on the extraction yield of Sudan I–IV. However, when the temperature increases, a larger fraction of soluble organic matter also gets co-extracted, leading to higher matrix effect. In addition, desolvation probably increases due to the elevated temperature. Therefore, an optimum temperature of 30 °C was used as the extraction temperature, although higher temperatures up to 50 °C yielded good results.

Optimization conditions for the ultrasound-assisted extraction method: (a) extraction time and (b) extraction solvent.
Figure 2 Optimization conditions for the ultrasound-assisted extraction method: (a) extraction time and (b) extraction solvent.

3.1.3

3.1.3 Effects of extraction solvent

Despite choosing methanol as a preliminary sample dissolution solvent for extracting these dyes from the sample, six different common organic solvents with different polarities including methanol, ethanol, acetone, acetonitrile, dimethylsulfoxide (DMSO), and hexane were tested. The moderate polarity solvents methanol, ethanol, acetone, acetonitrile, and DMSO extracted high contents of the dyes (Fig. 2b). Therefore, compared to hexane, they were considered good extraction solvents. However, when compared to the other solvent extracts, acetone is easily volatile and highly flammable and its smell likes nail polish remover, when exposed to the air. Acetonitrile is easily ignited by heat, spark or flame and gives off highly toxic hydrogen cyanide fume when heated. The DMSO extract could not easily be evaporated due to its high boiling point (189 °C). In addition, DMSO evaporates slowly at normal atmospheric pressure (Head and McCarty, 1973; Sricharoen et al., 2015). Therefore, methanol and ethanol were the best alternatives. However, because methanol has high toxicity, ethanol was chosen as a green solvent, consistent with a previous report (Rebane et al., 2010).

3.1.4

3.1.4 Analysis of real samples

To confirm whether ultrasound-assisted extraction could be used for sample preparation, all the samples were extracted under optimum ultrasound-assisted extraction conditions (Table 2). All dyes were not detected in the real samples studied. In order to validate the accuracy of the established method, the samples were spiked with Sudan dyes at concentration level of 500 ng mL−1 and subjecting it to the optimized extraction procedure. The %R ranged from 97.34 ± 4.44 to 104.18 ± 4.51% (Sudan I), 92.11 ± 4.01 to 102.35 ± 3.36% (Sudan II), 96.54 ± 4.57 to 108.86 ± 7.88% (Sudan III), and 95.32 ± 7.14 to 110.11 ± 8.76% (Sudan IV), indicating that the ultrasound-assisted extraction method was free of matrix interferences (Table 2). Since the proposed method gives acceptable recovery ranges, it can be used for sample treatment prior to the UAEME-SID-SFOD procedure.

Table 2 The UAE method for determination of Sudan dyes in chili pepper products (n = 3).
Analyte Spiked Recovery (% ± SDa)
(ng mL−1) Chili oil Chili sauce Chili paste Chili powder
Sudan I 0 –
500 104.2 ± 4.51 99.47 ± 4.32 97.34 ± 4.44 100.5 ± 5.33
Sudan II 0 –
500 102.4 ± 3.36 96.54 ± 3.87 99.68 ± 3.21 92.11 ± 4.01
Sudan III 0 –
500 97.68 ± 6.13 108.9 ± 7.88 96.54 ± 4.57 97.33 ± 2.44
Sudan IV 0 –
500 95.32 ± 7.14 110.1 ± 8.76 100.2 ± 3.22 98.11 ± 4.33
a Standard deviation.

3.1.5

3.1.5 Ultrasound-assisted extraction mechanism

The ultrasound-assisted extraction of the target compounds for a given matrix is a complex mechanism involving mass transfer and various possible chemical reactions, which influence the yield and associated biological activities. In general, the effect of sonication on mass transfer directly influences the ultrasonic frequency and sound energy introduced during extraction (Albero et al., 2015). Ultrasound-assisted extraction allows more solvent penetration into the plant body and can break down cell walls; therefore, it is used for extraction of organic and inorganic compounds from food products. Combinations of various physical, mechanical, chemical, and biochemical processes occur when ultrasound is applied in chemical reactions and extraction process (Picó, 2013), which are summarized below.

  1. A cavitation bubble can be generated close to the material surface using ultrasonic waves.

  2. This bubble collapses during a compression cycle and a microjet directing toward the plant matrix is created.

  3. Due to the application of high pressure and temperature during this process, the cell walls are destroyed or the surface at solvent–matrix interfaces is damaged by shock waves and microjets, releasing the analyte into the extraction solvent.

This technique is very effective for extracting natural products from biomass. In addition, the ultrasound-assisted extraction method has numerous advantages, including increased mass transfer, better solvent penetration, less solvent use, low temperature extraction, faster extraction rate, and greater product yield (Tiwari, 2015).

3.2

3.2 UAEME-SID-SFOD optimization

To identify optimum extraction conditions for the UAEME-SID-SFOD method, the effects of various parameters, including type of extraction solvent and its volume, pH of the sample solution, ionic strength, extraction time, and type of the emulsifier and its concentration, were investigated in detail. The optimization was performed with an aqueous solution (5 mL) containing 500 ng mL−1 of each Sudan dye. All the experiments were performed in triplicate and the %R were calculated.

3.2.1

3.2.1 Extraction solvent

It is crucial to select a suitable extraction solvent for all extraction methods. In general, a solvent is selected depending on its insolubility in water and its extraction performance. Moreover, the extraction solvent should have a lesser density than water, good chromatographic behavior, and easy water dispersing ability during the emulsification step. In the proposed method, the extraction solvent should have low density (<1 g mL−1) and low toxicity, with a melting point around room temperature (10–30 °C) and easy solidification property. Therefore, common solvents in the SFODME, such as 1-undecanol, 1-dodecanol, and 2-dodecanol, were selected for study. The following solvent ratios (by volume) were also studied: 1-undecanol:1-dodecanol (1:1), 1-undecanol:2-dodecanol (1:1), and 1-dodecanol:2-dodecanol (1:1). The following experimental conditions were employed: 60 μL extraction solvent, 6% (w/v) NaCl as ionic strength, 2 min extraction time, and 1% (w/v) AlCl3 as an emulsifier, and the results are shown in Fig. 3a. The highest recoveries were obtained when 1-dodecanol alone was used as the extraction solvent, probably because 1-dodecanol has lower polarity than the other solvents. These solvents were water soluble in the following order: 6.9 mg L−1 (1-dodecanol), 16.2 mg L−1 (2-dodecanol), and 43.0 mg L−1 (1-undecanol) (Seebunrueng et al., 2015); this was in agreement with their octanol–water partition coefficients (log Kow), i.e., 5.13, 4.70, and 4.72, respectively. However, there are two possible concerns. The first is the poor polarity and hydrophobicity of the analytes with log Kow values ranging from 5.51 to 8.72 (the log Kow values of Sudan I, II, III, and V are 5.51, 6.60, 7.63, and 8.72, respectively) and its strong ability to form hydrogen bond between its —OH and the phenolic hydroxyl group (—OH) of the analytes. The second concern is the natural matching between the dye and 1-dodecanol because benzene ring in the dye molecular structure results in high affinity with 1-dodecanol. However, the mixed solvent ratios had a synergistic extraction effect for the dyes due to formation of association complexes between the extractant and analyte. Taken together, these results demonstrate that these mixed solvents can also extract the analytes with similar extraction efficiency. Therefore, 1-dodecanol was chosen as the extraction solvent for further experiments.

Optimization conditions for the UAEME-SID-SFOD method: (a) extraction solvent, (b) volume of the extraction solvent, (c) ultrasonication time, (d) ionic strength, (e) type of demulsifier, and (f) demulsifier concentration.
Figure 3 Optimization conditions for the UAEME-SID-SFOD method: (a) extraction solvent, (b) volume of the extraction solvent, (c) ultrasonication time, (d) ionic strength, (e) type of demulsifier, and (f) demulsifier concentration.

3.2.2

3.2.2 Volume of extraction solvent

To determine the minimum volume necessary to guarantee the complete extraction of all the Sudan dyes, different volumes of 1-dodecanol (40, 60, 80, and 100 μL) were used with the same extraction procedure. The %R of all the analytes increased with the increasing elution volume from 40 to 80 μL (Fig. 3b). Subsequently, the analytical signals decreased regularly with increasing extraction solvent volume in the tested range (>80 μL), leading to a dilution effect in high volumes of extraction solvent. Therefore, 80 μL of extraction solvent was used for further experiments.

3.2.3

3.2.3 pH of sample

The pH of the sample solution possibly affects the ultrasound-assisted extraction of the dyes. Therefore, the effect of varying pH value (3–10) was investigated in the presence of 6% (w/v) NaCl and 1% (w/v) AlCl3. pH had no obvious effect on the %R due to the existing neutral molecule that does not noticeably dissociate or protonate in moderately acidic or basic media (Hu et al., 2016; Sun et al., 2011). Accordingly, the pH of the sample solution was not needed to justify for such simple procedure.

3.2.4

3.2.4 Ultrasonication time

The effect of ultrasonication time on the extraction of Sudan I–IV dyes was studied to enhance the extraction yield and simultaneously activate the rapid formation of a fine emulsion solution. The ultrasonication time was varied (0.5, 1, 1.5, 2, and 2.5 min), as shown in Fig. 3c. The %R of the dyes increased with time up to 1 min and then remained constant. Therefore, the ultrasonication time was kept quite short, i.e., within 1 min, which was advantageous for limiting the time needed for the ultrasound-assisted extraction method.

3.2.5

3.2.5 Salt addition for emulsification and demulsification

The solubility of both the target complex and extracting solvent in the aqueous phase usually decreases with an increasing ionic strength. Their back extraction into the organic phase can be promoted due to the salting-out effect (Hassan and Bahrani, 2014; Hassan and Sarkouhi, 2016), aiding the efficient transfer of the analyte into the extraction (organic) phase. The ionic strength was studied by adding salt solution, 0–12% (w/v) NaCl while 1% AlCl3 (w/v) was fixed as a demulsifier in the demulsification step. The %R of the dyes increased up to 6% (w/v) NaCl, and then, remained either constant or slightly decreased (Fig. 3d). Therefore, a minimum of 6% (w/v) NaCl was used to enhance the efficiency of the extraction process. After complete dispersion by an ultrasonic wave in the UAEME-SID-SFOD method, an emulsion was formed and the Sudan dyes were extracted into the extraction solvent by the droplets of 1-dodecanol. Subsequently, the emulsion was broken-up into two clear separation phases by adding a salt solution as a demulsifier (M.-J. Chen et al., 2013). Adding electrolytes into the emulsion solution may reduce the zeta potential of its double layer and its stability, particularly of hydrophobic colloids (Seebunrueng et al., 2015). Therefore, the repulsion of the extraction solvent droplets in the solution decreases, resulting in agglomeration of fine droplets of the extraction solvent that eventually leads to phase separation. Six types of the ionic salts (1% (w/v)), including NaCl, Na2SO4, MgSO4, MgCl2, AlCl3, and Al2(SO4)3, were tested while 6% (w/v) NaCl was fixed in the emulsification step. Both AlCl3 and Al2(SO4)3 showed similar high extraction efficiency (Fig. 3e). A liquid organic droplet floated on top of the surface of the aqueous solution after a short period of time without centrifugation, probably due to the stronger interactions between the larger charge of the salt and emulsion (its ionic strength). The ionic strength (I) of a solution is a function of the concentration of all ions present in that solution, calculated as follows (Sastre de Vicente, 2004):

(2)
I = 1 2 ∑ i = 1 n c i z i 2 where ci is the molar concentration of ion i (M), zi is the charge number of that ion, and the sum is of all the ions in the solution. The ionic strengths of the 1% (w/v) salt solution were 0.1171, 0.2112, 0.3147, 0.3324, 0.4494, and 0.4380 M for NaCl, Na2SO4, MgCl2, MgSO4, AlCl3, and Al2(SO4)3, respectively. However, AlCl3 solution was chosen for further experiments because it is the most effective demulsifier due to its highest ionic strength and fastest phase separation. In addition, while investigating the varying AlCl3 concentration (1–7% (w/v)), 3% (w/v) AlCl3 solution gave the highest %R and the phase separation was clearly completed (Fig. 3f). At lower AlCl3 concentrations, the extraction and aqueous phases were not clearly separated, whereas at higher AlCl3 concentrations (>3%), the %R were constant. Therefore, 3% (w/v) AlCl3 was chosen for further studies.

The most suitable extraction using the proposed method was done with 80 μL of 1-dodecanol and 5.0 mL sample in 6% (w/v) NaCl under ultrasonication for 1 min. Subsequently, to get a clear separation phase, 3% (w/v) AlCl3 was added to demulsify the sample extract. The application of this technique leads to simplification and acceleration of the sample preparation process, which is important in all analytical steps for such accurate results.

3.3

3.3 Analytical performance of the proposed method

The developed UAEME-SID-SFOD method was validated using a low-density solvent for the analysis of all the four Sudan dyes. The analytical features of merit are summarized in Table 3. The linearity of each calibration curve for Sudan I, Sudan II, Sudan III, and Sudan IV was 0.5–2500, 0.8–2500, 0.5–2500, and 0.8–2500 ng mL−1, respectively, and the correlation coefficients were between 0.9990 and 0.9997. The limits of detection (LODs) ranged from 0.16 to 0.24 ng mL−1, with a signal-to-noise ratio (S/N) of 3. The limits of quantification (LOQs) (S/N = 10) ranged from 0.53 to 0.80 ng mL−1. Furthermore, the reproducibility and precision in terms of intra- and inter-day variation were studied by replicate injections of the standard mixture of 500 ng mL−1 each day (n = 7) and on five consecutive days (n = 5 × 3), respectively. The precision, expressed as relative standard deviations (RSDs) of peak area, was <4% and <7% for all the dyes. The enrichment factor (EF) is used to evaluate the extraction efficiency (Es’haghi and Azmoodeh, 2010), which is defined as the ratio of the concentration of the final concentration of analyte in the acceptor solution (Ca,final) and the initial analyte concentration within the sample (Cs,initial):

(3)
EF = C a , final C s , initial
Table 3 Analytical features of merit for the UAEME-SID-SFOD in association with HPLC for determination of Sudan I–IV dyes.
Analyte Linear equation Linear range R2 LOD LOQ Precision (RSD, %) EF
(ng mL−1) (ng mL−1) (ng mL−1) Intra-day (n = 7) Inter-day (n = 5 × 3)
Sudan I y = 4,075,481x − 28,281 0.5–2500 0.9990 0.16 0.53 2.4 5.6 62
Sudan II y = 3,221,028x − 23,954 0.8–2500 0.9994 0.24 0.80 2.1 4.8 62
Sudan III y = 6,219,573x − 28,167 0.5–2500 0.9997 0.16 0.53 1.8 5.2 62
Sudan IV y = 5,411,952x − 45,654 0.8–2500 0.9996 0.24 0.80 3.5 6.3 62

The EF was calculated as 62, which is a remarkable value assuring trace determination of the target dyes. In addition, the HPLC chromatograms of Sudan I–IV dyes (1000 ng mL−1) via direct analysis (without EF) (Fig. 4a) were compared with those via the UAEME-SID-SFOD method (Fig. 4b). The chromatographic signals of these dyes were improved using UAEME-SID-SFOD.

Typical chromatograms of the Sudan dye standards and the studied samples. (a) Direct injection without preconcentration of all standards (1000 ng mL−1), (b) using UAEME-SID-SFOD procedure: concentration of all standards was 500 ng mL−1, (c) chili paste sample and (d) chili paste sample spiked with 500 ng mL−1 of each dyes.
Figure 4 Typical chromatograms of the Sudan dye standards and the studied samples. (a) Direct injection without preconcentration of all standards (1000 ng mL−1), (b) using UAEME-SID-SFOD procedure: concentration of all standards was 500 ng mL−1, (c) chili paste sample and (d) chili paste sample spiked with 500 ng mL−1 of each dyes.

3.4

3.4 Comparison of the proposed method with other related ones

Table 4 summarizes the comparison of the proposed extraction method with literature results for the extraction and determination of Sudan dyes. Cloud point extraction (CPE) based on surfactant-mediated phase separation (Liu et al., 2007) is cheaper, and less toxic surfactants are used. Disadvantages are that low recoveries (80.7–85.4%) may be due to temperature affecting for lose of analytes during the extraction (70 °C in water bath for 30 min). The extraction time was also longer than our proposed method about 30 times and was similar to ionic liquid extraction (Fan et al., 2009). Ionic liquids [C2MIM][PF6] was used as extraction phase in microwave-assisted liquid-liquid microextraction (Hu et al., 2016). Although microwave method provided good extraction time, it requires moderate to high cost for equipment and maintenance. The procedures presented by Chen and Huang (2014) and He et al. (2015) using DLLME based method require dispersive solvent and phase separation by centrifugation. They are not simple due to time consuming. This time-consuming step can be avoided by the proposed UAEME-SID-SFOD method, which is advantageous particularly because an extraction time of only 1 min is enough for complete extraction owing to the large surface area between the fine droplets of the extraction solvent and aqueous sample. The extraction solvent used in the present method is less toxic and a small volume is used. In addition, it does not need organic dispersive solvent and centrifugation step. Therefore, for green extraction, the proposed method is superior to other microextraction techniques for sample preparation.

Table 4 The comparison between UAEME-SID-SFOD and other related methods of determination of Sudan I–IV dyes.
Method Sample Extraction solvent Disperser solvent Extraction Recovery (%) RSD (%) LOD (ng mL−1) LOQ (ng mL−1) Reference
Source Time (min) Intra day Inter day
CPEa Chili powder Triton X-100 (1000 μL) – Water bath (70 °C) 30 80.7–85.4 1.18–3 – 2–4 7–12 Liu et al. (2007)
ILEb Chili products [C2MIM][PF6] (2 mL) – Stirrer (600 rpm) 30 70.7–109.5 2.0–3.5 – 7.0–8.2 – Fan et al. (2009)
IL-MA-LLMEc JUICE [C2MIM][PF6] (0.18 g) – Microwave + hand shaking 2 83.4–115.6 4.8 – 6 3.3–6.2 1.08–1.30 3.60–4.34 Hu et al. (2016)
SFO-DLPMEd Ketchup, chili sauce, chili oil 1-Dodecanol (100 μL) Ethanol (400 μL) Water bath (70 °C) 20 92.6–106.6 3.1–5.2 3.2–4.7 0.1–0.2 – Chen and Huang (2014)
UADLLME-SFOe Tomato sauce, chilli products 1-Dodecanol (30 μL) Acetone (1000 μL) Ultrasonic bath 15 79–92 4.8–7.1 – 1.5 – He et al. (2015)
UAEME-SID-SFODf Chilli products 1-Dodecanol (80 μL) – Ultrasonic bath 1 90.5–107.3 1.8–3.5 4.8–6.3 0.16–0.24 0.53–0.80 This study
a Could point extraction.
b Ionic liquid extraction.
c Ionic liquid microwave-assisted liquid-liquid microextraction.
d Dispersive liquid-phase microextraction with solidification of floating organic droplet.
e Ultrasound-assisted dispersive liquid-liquid microextraction with solidification of floating organic drop.
f Ultrasound-assisted emulsification microextraction and salt-induced demulsification with solidified floating organic drop.

3.5

3.5 Analysis of Sudan dyes in real samples

Sudan I–IV dyes were not detected in any sample. Therefore, the %R was calculated by spiking two levels of Sudan I–IV dyes into the samples (100 and 500 ng mL−1). The results are shown in Table 5, and the chromatograms of the chili paste samples in the absence and presence of Sudan dyes without and with pre-concentration by UAEME-SID-SFOD are illustrated in Fig. 4c and d. Their %R were in the range of 90.54–107.33%, with acceptable precision and <10% RSDs. RSDs show that the sample matrices have no significant effect on the performance of the UAEME-SID-SFOD method. Hence, it is feasible to use the method for the determination of Sudan dyes in chili samples. Because the sample matrices have no significant effect on the performance of the present method, the proposed method can be applied for trace determination of Sudan dyes in these food samples.

Table 5 The determination and recovery study of Sudan I–IV dyes in chili products (n = 3) using UAEME-SID-SFOD and HPLC.
Analyte Spiked Chili oil Chili sauce Chili paste Chili powder
(ng mL−1) Recovery (%) RSDa (%) Recovery (%) RSDa (%) Recovery (%) RSDa (%) Recovery (%) RSDa (%)
Sudan I 0 –
100 94.57 3.11 98.85 5.44 94.35 2.47 101.44 5.02
500 95.33 4.51 92.36 3.82 92.44 1.08 97.32 6.88
Sudan II 0 –
100 92.18 5.24 95.56 2.16 93.11 5.36 105.24 7.24
500 94.24 2.18 97.33 4.64 90.54 4.24 107.33 5.22
Sudan III 0 –
100 100.10 4.11 102.11 6.11 96.68 3.22 98.45 4.56
500 96.58 1.57 101.98 8.56 98.44 2.87 97.71 5.74
Sudan IV 0 –
100 102.34 6.32 97.74 5.14 104.56 3.57 95.34 6.88
500 104.66 5.10 95.11 3.12 100.22 4.11 100.44 4.21
a Relative standard deviation.

4

4 Conclusion

A novel technique of UAEME-SID-SFOD was developed and validated for the analysis of four Sudan dyes in chili products. This method is environmental friendly as a very small amount of organic solvent is used. It shows excellent phase separation of the water/1-dodecanol mixture using AlCl3 as a separation agent (emulsifier) without the use of a centrifugal device and other organic solvents after UAE, and recoveries of 91–107% were obtained. This procedure has several advantages over the conventional DLLME method, including no use of organic liquid as extraction solvent or disperser solvent, fast extraction, simple operation, and cost-effectiveness in terms of both materials and apparatus. Therefore, the UAEME-SID-SFOD method has an immense potential to be used as an alternative green extraction method by ultrasonic application (for both sample pre-treatment and emulsification) for the determination of Sudan dyes in chili products.

Acknowledgments

The authors thank the Higher Education Research Promotion and National Research University Project of Thailand, Office of the Higher Education Commission, through the Food and Functional Food Research Cluster of Khon Kaen University, Materials Chemistry Research Center, Department of Chemistry and Center of Excellence for Innovation in Chemistry (PERCH-CIC), Thailand for financial support of this study.

References

  1. , , , , , . Process optimization of ultrasound-assisted curcumin nanoemulsions stabilized by OSA-modified starch. Ultrason. Sonochem.. 2014;21:1265-1274.
    [Google Scholar]
  2. , , , , , . Ultrasound-assisted extraction of emerging contaminants from environmental samples. TrAC, Trends Anal. Chem.. 2015;71:110-118.
    [Google Scholar]
  3. , , , , . Optimization of ultrasound induced emulsification on the formulation of palm-olein based nanoemulsions for the incorporation of antioxidant β-d-glucan polysaccharides. Ultrason. Sonochem.. 2016;31:71-84.
    [Google Scholar]
  4. , , , , , . Ultrasound assisted Mizoroki–Heck reaction catalyzed by Pd/C: synthesis of 3-vinyl indoles as potential cytotoxic agents. Arabian J. Chem.. 2019;12(8):3836-3846.
    [Google Scholar]
  5. , , , , , . Coagulation and electrocoagulation of oil-in-water emulsions. J. Hazard. Mater.. 2008;151:44-51.
    [Google Scholar]
  6. , , . 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. J. Agric. Food Chem.. 2014;62:5818-5826.
    [Google Scholar]
  7. , , , , , , , , , , , . Development of a liquid chromatography–tandem mass spectrometry with 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. 2013;939:45-50.
    [Google Scholar]
  8. , , , . Low-density extraction solvent-based solvent terminated dispersive liquid-liquid microextraction combined with gas chromatography-tandem mass spectrometry for the determination of carbamate pesticides in water samples. J. Chromatogr. A. 2010;1217:1244-1248.
    [Google Scholar]
  9. , , , , , . Rapid determination of triclosan in personal care products using new in-tube based ultrasound-assisted salt-induced liquid–liquid microextraction coupled with high performance liquid chromatography-ultraviolet detection. Anal. Chim. Acta. 2013;767:81-87.
    [Google Scholar]
  10. , , , , , . Green extraction of grape skin phenolics by using deep eutectic solvents. Food Chem.. 2016;200:159-166.
    [Google Scholar]
  11. , , , , . 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.
    [Google Scholar]
  12. , , , , , . Interference-free determination of illegal dyes in sauces and condiments by matrix solid phase dispersion (MSPD) and liquid chromatography (HPLC–DAD) Food Chem.. 2012;135:193-198.
    [Google Scholar]
  13. , , . Hollow fiber supported liquid membrane microextraction of Cu2+ followed by flame atomic absorption spectroscopy determination. Arabian J. Chem.. 2010;3(1):21-26.
    [Google Scholar]
  14. , . Cavitation mechanism of ultrasonic melt degassing. Ultrason. Sonochem.. 1995;2:S137-S141.
    [Google Scholar]
  15. , , , , , , , . 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.
    [Google Scholar]
  16. , , . Determination of atorvastatin in human serum by salting out assisted solvent extraction and reversed-phase high-performance liquid chromatography–UV detection. Arabian J. Chem.. 2014;7(1):87-90.
    [Google Scholar]
  17. , , . Miniaturized counter current liquid–liquid extraction for organophosphorus pesticides determination. Arabian J. Chem.. 2016;9(1):38-42.
    [Google Scholar]
  18. , , , , , , , . Analysis of Sudan I in food by QuEChERS combined with ultrasound-assisted dispersive liquid-liquid microextraction with solidification of floating organic drop (UADLLME-SFO) prior to HPLC-PAD. Food Sci. Technol. Res.. 2015;21(5):659-664.
    [Google Scholar]
  19. , , . The thermal decomposition of DMSO. Tetrahedron Lett.. 1973;14:1405-1408.
    [Google Scholar]
  20. , , , , , , , . Determination of Sudan dyes in juice samples via solidification of ionic liquid in microwave-assisted liquid-liquid microextraction followed by high-performance liquid chromatography. Food Anal. Methods. 2016;9:2124-2132.
    [Google Scholar]
  21. , , , . Extraction of Sudan dyes from environmental water by hemimicelles-based magnetic titanium dioxide nanoparticles. Environ. Sci. Pollut. Res.. 2014;21:12382-12389.
    [Google Scholar]
  22. , , , . Recovery and purification of cholesterol from cholesterol-β-cyclodextrin inclusion complex using ultrasound-assisted extraction. Ultrason. Sonochem.. 2017;34:281-288.
    [Google Scholar]
  23. , , , , . 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(1):41-45.
    [Google Scholar]
  24. , , , . Supramolecular solvent-based microextraction of Sudan dyes in chilli-containing foodstuffs prior to their liquid chromatography-photodiode array determination. Food Chem.. 2010;121:763-769.
    [Google Scholar]
  25. , , , , . Speciation of Tl(III) and Tl(I) in hair samples by dispersive liquid–liquid microextraction based on solidification of floating organic droplet prior to flame atomic absorption spectrometry determination. Arabian J. Chem.. 2016;9(Suppl. 2):S1510-S1515.
    [Google Scholar]
  26. , , , , , , . Ultrasound assisted extraction of pectin from waste Artocarpus heterophyllus fruit peel. Ultrason. Sonochem.. 2017;34:525-530.
    [Google Scholar]
  27. , , , , . Development of nano-colloidal system for fullerene by ultrasonic-assisted emulsification techniques based on artificial neural network. Arabian J. Chem.. 2019;12(8):4162-4170.
    [Google Scholar]
  28. , , , . Preconcentration and simultaneous spectrophotometric determination of copper and mercury by dispersive liquid–liquid microextraction and orthogonal signal correction–partial least squares. Arabian J. Chem.. 2015;8(5):706-714.
    [Google Scholar]
  29. , , , , , . High-performance liquid chromatography analysis of ten dyes for control of safety of commercial articles. J. Chromatogr. A. 2008;1179:152-160.
    [Google Scholar]
  30. , . Ultrasound-assisted extraction for food and environmental samples. TrAC, Trends Anal. Chem.. 2013;43:84-99.
    [Google Scholar]
  31. , , , , . Ultrasound assisted extraction of bioactive compounds from Nephelium lappaceum L. fruit peel using central composite face centered response surface design. Arabian J. Chem.. 2017;10(Suppl. 1):S1145-S1157.
    [Google Scholar]
  32. , , , , , . Ultrasound assisted Mizoroki–Heck coupling/C—H amination in a single pot: direct synthesis of indole derivatives. Arabian J. Chem.. 2019;12(8):4320-4330.
    [Google Scholar]
  33. , , , , , . Interference-free simultaneous determination of Sudan dyes in chili foods using solid phase extraction coupled with HPLC–DAD. Food Chem.. 2011;125:1462-1467.
    [Google Scholar]
  34. , , , , . Optimization of ultrasonic assisted extraction of fatty acids from Borago officinalis L. flower by central composite design. Arabian J. Chem.. 2017;10(Suppl. 1):S23-S27.
    [Google Scholar]
  35. , , , , . A review of analytical techniques for determination of Sudan I–IV dyes in food matrixes. J. Chromatogr. A. 2010;1217:2747-2757.
    [Google Scholar]
  36. , , , , , , . A possible general mechanism for ultrasound-assisted extraction (UAE) suggested from the results of UAE of chlorogenic acid from Cynara scolymus L. (artichoke) leaves. Ultrason. Sonochem.. 2016;31:330-336.
    [Google Scholar]
  37. , . The concept of ionic strength eighty years after its introduction in chemistry. J. Chem. Educ.. 2004;81:750.
    [Google Scholar]
  38. , , , . Vortex-assisted low density solvent liquid–liquid microextraction and salt-induced demulsification coupled to high performance liquid chromatography for the determination of five organophosphorus pesticide residues in fruits. Talanta. 2015;132:769-774.
    [Google Scholar]
  39. , , , , . A novel extraction method for β-carotene and other carotenoids in fruit juices using air-assisted, low-density solvent-based liquid-liquid microextraction and solidified floating organic droplets. Food Chem.. 2016;203:386-393.
    [Google Scholar]
  40. , , , . A high correlation indicating for an evaluation of antioxidant activity and total phenolics content of various chilli varieties. J. Food Sci. Technol.. 2015;52:8077-8085.
    [Google Scholar]
  41. , , , , , , , , , . 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.
    [Google Scholar]
  42. , . Ultrasound: a clean, green extraction technology. TrAC, Trends Anal. Chem.. 2015;71:100-109.
    [Google Scholar]
  43. , , , . Green analytical chemistry–theory and practice. Chem. Soc. Rev.. 2010;39:2869-2878.
    [Google Scholar]
  44. , . Influence of sonication conditions on the efficiency of ultrasonic cleaning with flowing micrometer-sized air bubbles. Ultrason. Sonochem.. 2016;29:604-611.
    [Google Scholar]
  45. , , , , , . Determination of Sudan I–IV in candy using ionic liquid/anionic surfactant aqueous two-phase extraction coupled with high-performance liquid chromatography. Food Chem.. 2015;173:815-820.
    [Google Scholar]
  46. , , , , , . 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.
    [Google Scholar]
  47. , , , , . 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.
    [Google Scholar]
  48. , , , , , , , . Simultaneous determination of 14 oil-soluble synthetic dyes in chilli products by high performance liquid chromatography with a gel permeation chromatography clean-up procedure. Food Chem.. 2014;145:956-962.
    [Google Scholar]

Fulltext Views
506

PDF downloads
134
View/Download PDF
Download Citations
BibTeX
RIS
Show Sections

Suggested read for related articles: