A Nanoparticle-Based Solid-Phase Extraction Procedure Followed by Spectrofluorimetry to Determine Carbaryl in Different Water Samples

Nesse estudo, um novo método baseado em nanopartículas de magnetita Fe3O4 (MNPs) foi desenvolvido para a extração, preconcentração e determinação de traços de carbaril de amostras de água do meio ambiente. Fe3O4 MNPs foram sintetizadas e modificadas pelo surfactante dodecil sulfato de sódio (SDS) e aplicadas com sucesso na extração de carbaril e na sua determinação através de espectrofluorimetria. Fatores que afetam a adsolubilização do carbaril, como quantidade de SDS, pH, tempo de espera, solvente de desorção e volume máximo de extração foram otimizados. Sob as condições selecionadas, foi possível extrair carbaril quantitativamente. Recuperações (84,5-91,9%) e desvio padrão relativo (6,2%) aceitáveis foram alcançados ao analisar amostras de água adulterada. Um fator de concentração de 20 foi obtido na extração de 100 mL de amostras de água do meio ambiente. Os limites de detecção e quantificação encontrados foram 2,1 e 6,9 μg L-1, respectivamente. O método proposto foi aplicado com sucesso na extração e determinação de carbaril em amostras de água do meio ambiente.


Introduction
Recently, nanoparticles (NPs) have attracted substantial interest in the scientific community for the sample extraction.Compared with micrometer-sized particles used in the solid phase extraction (SPE), NPs offer a significantly higher surface area-to-volume ratio and a short diffusion route, resulting in a higher extraction capacity; rapid dynamics of extraction and higher extraction efficiencies. 1,2lso, NPs' surface functionality can be easily modified to achieve the selective sample extraction or cell collection. 1,2n these NP-based extraction procedures, the NPs were collected after extraction either by centrifugation or by filtration, which would be difficult in some cases such as with large volume samples. 1 More recently, magnetite nanoparticles (MNPs) have received increasing attention.MNPs as adsorbents can be easily collected by using an external magnetic field placed outside of the extraction container. 1Thus separation process can be performed directly in crude samples containing suspended solid material without additional centrifugation or filtration, which makes separation easier and faster. 1,3ecause of their large surface area and unique physical and chemical properties, they have been widely applied in the analytical chemistry field.These MNPs (as bare or modified Fe 3 O 4 adsorbents) can be used for the extraction and determination of trace amounts of organic 1,[4][5][6][7][8][9][10][11][12][13][14][15][16] and inorganic 2,[17][18][19][20] compounds from different samples.
The adsorption of ionic surfactants on MNPs can form hemimicelles and admicelles.8][13][14][15][16][17] In these methods, the sorbents were produced by the adsorption of anionic 6,8,13,17 or cationic 5,15,16 surfactants on the surface of MNPs.By using hemimicelles in SPE, a new kind of adsorbents is obtained which has many advantages, such as high extraction yields, high breakthrough volumes, rapid separation and rapid elution of analytes.Furthermore, no clean-up steps are required additionally. 5,7,15etermination of trace level pesticides in aquatic environment is very important due to their intense use in agriculture and to their persistence as well. 1 These compounds cause a variety of neurotoxic and endocrine disruptor effects.Thus, World Health Organization (WHO) and various national governmental institutions have established residue limits and published guidelines for quantification of pesticide residues in waters. 21,22arbaryl is a broad spectrum pesticide belonging to the N-methylcarbamate group and functioning as a reversible inhibitor of cholinesterase (ChE) activity.Carbaryl moderately binds to soil and has potential to leach to groundwater.It is not persistent in soil since it can be hydrolyzed, photodegraded, oxidized and degraded by microbes.In alkaline or neutral water, hydrolysis is the major degradation route for carbaryl, with half-lives ranging from a few hours to a few days.It is also subject to microbial degradation in natural water.Photolysis plays a role in the degradation process, significantly reducing degradation half-life of carbaryl.The major degradation product is 1-naphthol.In general, carbaryl is slightly toxic to mammals, moderately to highly toxic to aquatic organisms and honeybees. 23n Australia, the guideline for carbaryl in drinking water has recommended the health value (HV) to be 0.03 mg L -1 .The environmental quality standards in Japan are established the level for carbaryl as 0.05 mg L -1 in public water.Canadian drinking-water quality maximum acceptable concentration (MAC) value is 0.09 mg L -1 . 1 Analyses of carbamates by high performance liquid chromatography (HPLC) and by mass spectrometry (MS) coupled with HPLC and gas chromatography (GC) in environmental samples, either with direct or with derivatization methods, have been widely used. 24wever, GC based analysis of many carbamates is hindered by the thermal degradation of the analytes and precolumn derivation is required. 25,26On the other hand, the costs of HPLC and HPLC-MS methods are high.They usually use high volumes of toxic solvents and their separation procedure is time-consuming.Sometimes you have to develop an extremely complex gradient for the separation, specific preprocessing procedures for samples or sophisticated cleanup strategies. 27herefore, there is an increasing demand for developing sensitive, simple and reliable methods for determining pesticides in aquatic environments.Luminescence spectroscopy is considered as a sensitive and selective analytical technique.It offers remarkable analytical features for the determination of organic pollutants.However, its use (especially fluorescence) for analysis of organic pesticide residues has been limited by the fact that relatively few of these compounds are strongly luminescent. 28n this work, Fe 3 O 4 MNPs were synthesized and modified by the sodium dodecyl sulfate (SDS) surfactant.Then, a SPE procedure based on these modified NPs was developed for the extraction of carbaryl from environmental water samples and its determination by a simple, rapid and inexpensive spectrofluorimetric method.Predominant factors influencing the synthesis of SDS-coated Fe 3 O 4 NPs as well as carbaryl extraction and determination were investigated and optimized.

Apparatus
All fluorescence measurements were made using a Shimadzu RF-5301 PC spectrofluorophotometer (Kyoto, Japan) equipped with a 150 W Xenon lamp and 1.00 cm quartz cells.Instrument excitation and emission slits both were adjusted to 5 nm.A centrifuge from Hettich (EBA 20 model/Andreas Hettich GmbH & Co. KG, Tuttlingen, Germany) with 15 mL calibrated centrifuge tubes (Hirschmann, EM techcolor, Germany) was used to accelerate the phase separation process.The pH-meter model M120 (Halstead, Essex, England CO9 2DX) supplied with a glass combined electrode was used for the pH measurements.The mixtures were shaked using a Unimax 1010 Shaker-Inkubator (Heidolph, Germany).
A stock standard solution at a concentration of 500 µg mL -1 was prepared by dissolving appropriate amount of carbaryl in methanol and stored under dark conditions in refrigerator when not in use.This solution was stable for 1 month.Working standard solutions were obtained daily by appropriately diluting this stock solution with ultrapure water.The water used for sample preparation was deionized and purified using a Milli-Q system (Advantage A 10, Millipore, France).

Preparation and characterization of MNPs
Fe 3 O 4 NPs were prepared by the co-precipitation method with some modifications. 17So, the mole ratio of Fe 3+ /Fe 2+ was approximately 1.7:1 instead of approximately 2:1 in the cited reference.Firstly, 4.6 g of FeCl 3 •6H 2 O, 2.0 g of FeCl 2 •4H 2 O, and 0.85 mL of HCl (12 mol L -1 ) were dissolved in 25 mL of deionized water which was degassed with N 2 before use.The resulted clear yellowish green solution was added drop-wise into 250 mL of 1.5 mol L -1 NaOH solution (heated to 80 °C), under vigorous stirring with N 2 passing continuously through the solution during the reaction.Upon addition, the solution turned black and was then stirred magnetically at 1000 rpm for 30 min.After the reaction, the obtained precipitate was separated from the reaction medium by magnetic field, washed with 200 mL of deionized water four times, and then re-suspended in 250 mL of deionized water.The average particle size of the obtained MNPs was observed by using a scanning electron microscope (SEM), model vega2 (Czech Republic).

Recommended procedure
For the batch adsorption experiments, a 100 mL sample solution containing 2-100 µg mL -1 of carbaryl (or real water sample), 5 mL of NPs solution and 400 µg mL -1 of SDS solution were put into a 250 mL beaker and the pH of the mixture was adjusted to the desired value with 0.1 mol L -1 HCl.Secondly, the mixture was shaked at 200 rpm and allowed to complete the extraction process for 5 min.Subsequently, a strong magnet was placed at the bottom of the beaker, and the SDS-coated Fe 3 O 4 NPs was isolated from the solution.After about 5 min, the solution became limpid and supernatant solution was decanted.Finally, the preconcentrated target analyte was eluted from the isolated particles with methanol (2 × 2.5 mL) to desorb the target analyte with the aid of stirring.The fluorescence intensity of final solution (i.e., 5 mL) was measured at 334 ± 3 nm with the excitation wavelength set at 279 ± 3 nm.

Sample collection
All water samples were obtained from different districts of Azerbaijan-e-sharghi. Tap water samples were taken from our lab in Tabriz and a well water sample came from the Jolfa.River water samples were collected from Aras river (Jolfa).Bottled mineral water samples were obtained from local markets in Tabriz.All samples were collected randomly and filtered through 0.45 µm filter paper before use in order to remove suspended solids.The filtered water samples were stored at 4 °C until analysis.Then, a 100 mL portion of clear samples was subjected to the extraction and spectrofluorimetric determination.The spiked water samples were made by adding certain amounts of carbaryl standard solution to the real water samples of fixed volume.

Synthesis of Fe 3 O 4 MNPs
Co-precipitation is a facile and convenient way to synthesize Fe 3 O 4 NPs from aqueous Fe 2+ /Fe 3+ salt solutions by the addition of a base under inert atmosphere at room temperature or at elevated temperature.The size, shape and composition of MNPs depends on the type of salts used (e.g., chlorides, sulfates, nitrates), the Fe 2+ /Fe 3+ ratio, the reaction temperature, the pH value and ionic strength the media. 13n this work, different conditions were investigated for the NPs synthesis consisting of: using chloride or sulfate salts, changing the Fe 2+ /Fe 3+ ratio from 1:1 to 1:2.5, adjusting the sample pH with NH 4 OH or NaOH and finally the addition of base to Fe 2+ /Fe 3+ solution or reverse order.The results showed that using this procedure, the quality of MNPs is fully reproducible once the synthesis is performed using chloride salts, the Fe 2+ /Fe 3+ ratio fixed at 1:1.7 and addition of Fe 2+ /Fe 3+ mixture to NaOH solution (heated at 80 °C).The SEM image (Figure 1) of the dispersed Fe 3 O 4 NPs in water shows that the synthesized MNPs have rather high surface area and also the substructures with dimensions less than 42 nm could be observed.

Effect of pH
In the mixed hemimicelle based SPE, value of pH plays a critical role in the hemimicelles formation and the target compound extraction.The surface charge density of Fe 3 O 4 NPs is a main factor affecting the extraction of analyte and its amount varies strongly with the pH values. 13The effect of pH on the adsorption of carbaryl was investigated in the range between 1.5-9.5 by using 0.1 mol L -1 HCl or NaOH solution for pH adjustment (see Figure 2).The percent adsorption and thus analytical signal was increased by increasing pH and reached maximum at pH 3.0, then remained nearly constant up to pH 3.5 and decreased at higher pH values.Thus, a pH of 3.0 was chosen for all subsequent experiments and HCl solution used for the pH adjustment.
The surface charge of bared Fe 3 O 4 NPs is neutral at the pH ca.6.5 (pH zpc = pH zero point charge). 8,13,17Therefore, it is concluded that the surface of these particles has a positive charge at acidic pHs.The positively charged surface of Fe 3 O 4 NPs in acidic solutions was favorable for the adsorption of anionic surfactants and thus targeted analyte.
When pH was above the isoelectric point of the Fe 3 O 4 NPs, the positive charge density on the surface of the Fe 3 O 4 NPs is decreased, thus the adsorption of SDS molecules on NPs surfaces becomes less favorable 8,11,13,17 and this could lead to a remarkable depression in the analytical signal.However, in quite acidic medium, the analytical signal was decreased probably due to dissolution of Fe 3 O 4 NPs at pHs below 2 or protonation of SDS molecules, which could reduce the hemimicelles formation efficiency. 13e amounts of Fe 3 O 4 NPs and sample volume Compared to ordinary sorbents (micron-size particle sorbents), NP sorbents have higher surface areas.Therefore, satisfactory results can be achieved with fewer amounts of NP sorbents.The influence of MNPs content was studied by adding different amounts of Fe 3 O 4 suspension, ranging from 1-6 mL.The results showed that an amount of 5 mL of suspension was the optimum value.
Effect of sample volume on the adsorption of carbaryl was studied so, the 50, 100, 150 and 200 mL solutions containing 100 µg L -1 carbaryl were selected.Then adsorption and desorption processes were performed under the optimum conditions as described in experimental section.The results showed that the carbaryl present in the volumes up to 100 mL was completely and quantitatively adsorbed with NPs.The adsorption then remained constant at higher volumes.Therefore, for determination of trace quantities of carbaryl in samples, a sample volume of 100 mL was selected.

Effect of SDS amount
It was observed that MNPs can't adsorb carbaryl from aqueous solution at all, while their SDS modified form adsorbed carbaryl efficiently.The adsorption of surfactants on the surface of mineral oxides is a favorable process and based on the added surfactants, they can form various aggregates on the surface (i.e., hemimicelles, mixed hemimicelles and admicelles), below and above the critical micelles concentration (CMC). 6,8,17In this study, SDS was added to the solution at concentrations lower than its CMC (2.3 g L -1 ) 29 to modify the surface of Fe 3 O 4 NPs.The effect of SDS amounts on the adsorption of carbaryl was considered in the ranges of 15-180 mg for the surfactant when 100 mL of water containing 100 µg L -1 of carbaryl was used (see Figure 3).According to the results, with increase in the SDS amount, the adsorption amount of the carbaryl increased remarkably.The increase in adsorption can be explained by the gradual formation of SDS aggregates on the Fe 3 O 4 NP surface and the carbaryl is adsorbed gradually.Maximum adsorption was  obtained when SDS amounts were between 50-90 mg.At higher amounts of SDS, the adsorption of carbaryl decreased gradually due to formation of SDS aggregates in the solution, which can compete with formation of SDS aggregates on the surface of Fe 3 O 4 .Given these findings, 80 mg of SDS (final concentration of 400 µg mL -1 ) was selected for the next studies.

Extraction time
In the SPE process, the shaking time is one of the prime factors influencing the target analytes extraction.The extraction time profiles were studied by varying the mixing time of MNPs-sample suspension in the range of 1-10 min.Results reveals that a rapid extraction could occurr in about 5 min.The high surface area of MNPs along with homogeneous distribution of the nano-sorbent throughout the sample could be the possible reasons for achieving such a fast extraction process.Thus, a short shaking time of 5 min was selected for adsorption in subsequent experiments.

Desorption condition
Organic solvents can rapidly and completely disrupt the mixed hemimicelles and therefore the analyte is removed from the surface of NPs.In this work, methanol, ethanol, acetone and acetonitrile were studied for desorption of analyte from the SDS-coated Fe 3 O 4 NPs by gently shaking the solution.The maximum signal was observed when methanol was used.Therefore, desorption of analyte was carried out by using 2 × 2.5 mL of methanol.For achieving the more efficient desorption condition, desorption time was investigated in the range of 1-10 min.A duration time of 5 min appeared to be sufficient for complete desorption.

Analytical parameters
Under the optimum experimental conditions, calibration graphs were obtained by extraction of 100 mL of standard solutions and under the experimental conditions specified in the procedure.The calibration curve for the detection of carbaryl was linear over the concentration range of 2 to 100 µg L -1 .The corresponding fitted equation was FI = 7.23(± 0.51) C + 14.18(± 0.92) (n = 9) and the coefficient of correlation (r 2 ) was 0.9989.In this equation, the FI is the fluorescence intensity and C is the carbaryl concentration in µg L -1 .The relative standard deviation (RSD) obtained for the repetitive determinations of 50 µg L -1 of carbaryl was found to be 6.2% (n = 6).The limit of detection (LOD) and quantification (LOQ) by using the criterion 3S b /m and 10S b /m, were found to be 2.1 and 6.9 µg L -1 , respectively, where S b is the standard deviation of the blank measurements and m is the calibration slope.
In order to highlight the advantageous features of the proposed method, its performance in the extraction of carbaryl was compared with other methods reported in the literature.The distinct features of these methods are summarized in Table 1.Compared with references 21,[30][31][32][33][34] which use GC-MS or HPLC for the determination of targeted analyte, the proposed method does not require high investment and maintenance costs of the instruments.][35][36][37] Mukdasai et al. 37 has used a two-step micro-extraction method for the extraction of carbaryl.This method is based on the derivatization reactions for both carbaryl and its SPE sorbent, which is a laborious and time-consuming method.This method uses a high concentration factor (i.e., 2730); however, our LOD is approximately ten times better than that reported in this work.The second method uses graphene (G) grafted silica-coated Fe 3 O 4 nanocomposite for the extraction of carbaryl and its determination by HPLC. 38Some reported features in this work are better than our method.But a simple, rapid and low-cost spectrofluorimetric method in combination with a different sorbent has been adopted in this work for the extraction and determination of carbaryl.

The validation and application of the method
The present method was validated by different water samples spiked with known amount of carbaryl.As shown in Table 2, the overall recoveries of carbaryl added to water samples were in the range of 75.2 to 87.0% which were acceptable except for the river water samples.The addition of a neutral salt such as NaCl, can increase these values (up to 84.5-91.9%)due to known salting out effect.
Figure 4 shows typical excitation and emission spectra for carbaryl in different samples.The coincidence of excitation and emission spectra in this figure along with acceptable recoveries indicated that no significant matrix effect occurred in the proposed method.In order to test the applicability of the proposed method, four different water samples were collected and analyzed under the optimum conditions.The results have been summarized in Table 2.

Figure 1 .
Figure 1.SEM image of the dispersed Fe 3 O 4 NPs.

Figure 2 .
Figure 2. The effect of pH on the analytical signals.

Figure 3 .
Figure 3.The effect of SDS amount on the analytical signals.

Figure 4 .
Figure 4. Excitation and emission spectra after NP based-SPE: (a) reagent's blank, (b) a blank spiked with the concentration in the range of LOQ, (c) river water sample, (d) river water spiked with carbaryl at 60 µg L -1 and (e) standard solution of carbaryl (75 µg L -1 ).

Table 1 .
Analytical characteristics of different methods used for extraction and determination of carbaryl

Table 2 .
Results of recoveries of spiked water samples a Recoveries in the presence of NaCl (5% m/v); nd = not detected.