**4.1. QuEChERS procedure**

The need for a simple, rapid, cost-effective and multi-residue method able to provide high quality of analytical results led Anastassiades et al. to develop in the years 2001 and 2002 a new sample treatment method called "QuEChERS". Initially, the methodology was developed for the analysis of veterinary drugs (anthelmintics and thyreostats) in animal tissues, but after realizing its great potential in the extraction of polar and particularly basic compounds, it was also tested with great success on pesticide residue analysis in plant material. The detailed method was first published in 2003 [75].

QuEChERS, acronym of "*Quick, Easy, Cheap, Effective, Rugged and Safe*", is a sample preparation technique entailing solvent extraction with acetonitrile and partitioning with magnesium sulfate alone or in combination with other salts followed by a clean-up step using dispersive solid-phase extraction (d-SPE). This last step is performed by adding small amounts of bulk SPE packing sorbents to the extract. This procedure has attracted the attention of pesticides laboratories worldwide and it is the most commonly employed sample treatment methodology for the multi-residue analysis of pesticides in fruits and vegetables [9,10,76-80]. But today, this methodology is not limited to the analysis of pesticides and its use for the extraction of other families of compounds is tremendously increasing.

The different steps on a typical QuEChERS procedure for the multi-residue analysis of pesticides are shown in **Figure 6**.


**Figure 6.** Schematic view of a typical analytical QuEChERS procedure for the analysis of pesticides as described in [75].

The idea of the QuEChERS procedure was to reduce complicated, laborious and timeconsuming multi-residue sample treatment methods that required high amount of solvents and were therefore expensive. Moreover, some basic, acidic and very polar compounds cannot be satisfactorily extracted with common multi-residue methods. Thus, in order to cover all these analytes, laboratories have to perform specific analysis, and as a consequence, some of these compounds were not being monitored.

144 Chromatography – The Most Versatile Method of Chemical Analysis

**4. QuEChERS** 

**4.1. QuEChERS procedure** 

pesticides are shown in **Figure 6**.

described in [75].

efforts should continue to be made in this promising research area.

material. The detailed method was first published in 2003 [75].

extraction of other families of compounds is tremendously increasing.

(shake 30 s and centrifuge)

1 – Weigh 10 g of Sample (50 mL Teflon tube)

2 – Add 10 mL of Acetonitrile (shake vigorously 1 min)

4 – Add Internal Standard (shake 30 s and centrifuge)

5 – Take aliquote and Add MgSO4 and d-SPE sorbent

3 – Add 4 g of MgSO4 and 1 g NaCl (shake vigorously 1 min)

On-line MIP-SPE pre-concentration methodology has also recently been used by Jing et al. [6], for the determination of trace tetracycline antibiotics (TCs) in egg samples. This approach affords high-throughput analysis (18 min per sample), and also provides high sensitivity and selectivity with recoveries ranging between 91.6 and 107.6%, showing that

The need for a simple, rapid, cost-effective and multi-residue method able to provide high quality of analytical results led Anastassiades et al. to develop in the years 2001 and 2002 a new sample treatment method called "QuEChERS". Initially, the methodology was developed for the analysis of veterinary drugs (anthelmintics and thyreostats) in animal tissues, but after realizing its great potential in the extraction of polar and particularly basic compounds, it was also tested with great success on pesticide residue analysis in plant

QuEChERS, acronym of "*Quick, Easy, Cheap, Effective, Rugged and Safe*", is a sample preparation technique entailing solvent extraction with acetonitrile and partitioning with magnesium sulfate alone or in combination with other salts followed by a clean-up step using dispersive solid-phase extraction (d-SPE). This last step is performed by adding small amounts of bulk SPE packing sorbents to the extract. This procedure has attracted the attention of pesticides laboratories worldwide and it is the most commonly employed sample treatment methodology for the multi-residue analysis of pesticides in fruits and vegetables [9,10,76-80]. But today, this methodology is not limited to the analysis of pesticides and its use for the

The different steps on a typical QuEChERS procedure for the multi-residue analysis of

**Figure 6.** Schematic view of a typical analytical QuEChERS procedure for the analysis of pesticides as

6 – Take aliquote and Analyze (typically GC-MS or LC-MS)

The first two steps of a typical QuEChERS procedure consist in weighing an appropriate amount of sample previously processed and homogenized (for instance 10 g) in a 50 mL Teflon tube (Step 1) and the addition of a solvent for the extraction (Step 2), in general acetonitrile, although the use of other organic solvents such as acetone, THF or ethyl acetate have been described [81].

Then, an extraction-partitioning step takes place by the addition of magnesium sulfate alone or in combination with other salts, generally sodium chloride (Step 3). Acetonitrile is the recommended solvent for QuEChERS because, upon the addition of salts, it is easily separated from water than, for instance, acetone. Ethyl acetate has the advantage of a partial miscibility with water but it can also co-extract lipids and provides lower recoveries during the dispersive SPE. The extraction of lipophilic materials is lower with acetonitrile but this solvent can form two phases with water when samples with high sugar content are manipulated [75]. The addition of salts in Step 3 helps to induce the phase separation. This salting-out effect also influences analyte partition, which of course is also dependent upon the solvent used for extraction. The concentration of salt can also influence the percentage of water in the organic phase and can play an important role in adjusting its polarity. Magnesium sulfate acts as a drying salt to reduce the water phase, thereby helping to improve recoveries by promoting partitioning of the pesticides (or other target compounds) into the organic layer while sodium chloride helps to control the polarity of the extraction solvent. With this, a single extraction-partitioning step is carried out (similarly to an "online" approach) which simplifies the necessity of multiple partitioning steps required in other multi-residue methods. Moreover, this extraction-partitioning step is produced by shaking vigorously for a few minutes, thus preventing more time-consuming steps such as sample blending. At this point, internal standards can be added to the system if necessary (Step 4), followed by shaking again the solution and a centrifugation step that help to separate salts. The use of internal standards can minimize the error generated in the multiple steps of the QuEChERS method. Sometimes the use of more than one internal standard is recommended especially with samples with high fat content because the excessive fat can form an additional layer into which the analytes can also partition [82]. Another advantage of QuEChERS procedure is the fact that, once the extraction-partitioning step is carried out, an aliquot of the extract is used for the next steps, minimizing also the separation or the transfer of the entire extracts frequently employed in other multi-residue methods.

Then, a dispersive solid-phase extraction (d-SPE) clean-up procedure is carried out with an aliquot (for instance 1 mL) of the extract which is placed in a vial containing again magnesium sulfate and small amounts of bulk SPE sorbent materials (Step 5). The vial is

then shaken vigorously or mixed on a vortex mixer to distribute the SPE material and facilitate the clean-up process. This d-SPE step is quite similar to matrix solid-phase dispersion developed by Barker [83, 84], but in d-SPE the sorbent is added to an aliquot of the extract rather than to the original solid sample. Moreover, small amounts of sorbents are used because only a small portion of the extract is subjected to the clean-up procedure, and compared to conventional SPE clean-up methods, d-SPE is less laborious and timeconsuming. Magnesium sulfate is again used in this step as a drying agent to remove water and improve analyte partitioning to provide better clean-up. Primary secondary amine (PSA) is the most common SPE sorbent used in QuEChERS procedure for pesticide analysis. The idea is to use a sorbent able to retain matrix components, but not the analytes of interest. However, depending on the sample matrix, other SPE sorbents can also be used alone or combined with PSA, such as C18, OASIS HLB, and graphitized carbon black sorbents. For instance, for samples with high fat content, PSA mixed with C18 is recommended [85] while for samples with moderate or high levels of chlorophyll and carotenoids (for example carrots), PSA mixed with graphitized carbon black is frequently used [86-88]. After the clean-up, the extract is centrifuged and an aliquot of the supernatant can be concentrated or directly analyzed usually by means of Gas Chromatography-Mass Spectrometry (GC-MS) or Liquid Chromatography-Mass Spectrometry (LC-MS) techniques (Step 6).

Current Trends in Sample Treatment Techniques for Environmental and Food Analysis 147

As commented before, QuEChERS procedure is the sample treatment of choice for the multiresidue analysis of pesticides in fruits and vegetables, and many works can be found in the literature dealing with the simultaneous extraction and clean-up of more than 100 pesticides with acceptable recoveries [75,77,79,80,91]. As an example, Woo Lee et al. developed a new QuEChERS method based on dry ice for the determination of 168 pesticides in paprika [91]. For this purpose, extraction was carried out by using 30 mL of acetonitrile and 10 mL of water, and approximately 10 g of dry ice granules were poured and maintained until layer separation. Clean-up was then carried out by using PSA and GCB sorbents. The separation of the sample extract was induced via the sublimation of dry ice, which occurs at -78.5oC at atmospheric pressure (1 atm). After some minutes of dry ice sublimation with the sample extract, the reduced temperatures of acetonitrile and water ranged from -4.0 to -5.0 oC and from -6.0 to -6.5 oC, respectively, and water was then iced and super cooled. The negative temperatures of the two solvents may reduce their entropies and allowed them to separate. As densities of ice and supercooled water were heavier than that of acetonitrile at 0 oC, water changed to ice and the supercooled water was separated with an acetonitrile layer from the mixed solution. This methodology improved the extraction for flonicamid and its metabolites

which was not satisfactory enough using citrate-buffering QuEChERS method.

Recently, the use of carbon-based nanoparticles has also been described as clean-up sorbent for the analysis of pesticides [10]. In this work, multi-walled carbon nanotubes (MWCNTs) were proposed as reversed-dispersive solid phase extraction (r-DSPE) material for the analysis of 30 pesticides in fruits and vegetables. The amount of MWCNTs influenced the clean-up performance and the recoveries, but the use of only 10 mg MWCNTs was suitable for cleaning up all analyzed matrices and showed to be a good alternative to PSA sorbent. The method was validated for different matrices such as spinach, orange and cabbage with recoveries in the range of 71-100% for all 30 pesticides. QuEChERS has also been proposed for the extraction and clean-up of pesticide residues in other food matrices such as milk [85]. In this last work, recovery of 14 different pesticides residues in milk was investigated to respect the amounts of sodium acetate, PSA and C18 used on the clean-up step. Recoveries for hydrophilic pesticides such as myclobutanil ranged from 82 to 99% while lower values (<80%) were obtained for lipophilic pesticides because they were partially removed by C18

But today, one of the most important features of QuEChERS may be its application to other families of compounds in a variety of matrices different than fruits and vegetables. Some examples of the use of QuEChERS for the extraction of compounds other than pesticides are given in **Table 3**. QuEChERS methodology has already been applied to the analysis of polycyclic aromatic hydrocarbons (PAHs) in fish and shrimp samples [92,93]. Forsbeg et al. developed and validated a modified QuEChERS method for the determination of 33 parent and substituted PAHs in high-fat smoked salmon that greatly enhanced analyte recovery compared to traditional QuEChERS procedure [93]. For this purpose, a mixture of acetone, ethyl acetate and isooctane instead of acetonitrile was employed for the extraction, and different kinds of salts were used for partitioning. The proposed modified QuEChERS

**4.2. Applications of QuEChERS** 

along with other fatty compounds.

Although QuEChERS is quite a simple procedure as can be seen from **Figure 8,** in some cases method development will be necessary depending on the family of compounds to be analyzed, and many modifications over the QuEChERS procedure are being proposed. At the end, compromises will be required to ensure simplicity, speed, broad applicability, high recovery and selectivity. For instance, the control of pH in the extraction step is animportant factor when analyzing pesticides in order to ensure an efficient extraction of pH-dependent compounds such as phenocyalcanoic acids and to minimize degradation of labile pesticides under alkaline or acidic conditions. Buffering with citrate salts has been introduced in the first extraction/partitioning step to adjust the pH at around 5 where most labile pesticides under acidic or alkaline conditions are sufficiently stabilized. The pH control can also be very important in other steps of the QuEChERS procedure to prevent degradation of some compounds. For instance, to improve the stability of alkaline-labile compounds after the PSA clean-up step the final sample extract is slightly acidified by the addition of small amounts of formic acid. Of great importance was the introduction of acetate buffering to achieve a pH value of 6 in order to improve recoveries of pH-dependant analytes [89]. This approach resulted in the official method AOAC 2007.01.

The use of analyte protectants is also often proposed as an optional step previous to GC analysis for those compounds that might tail or breakdown on the capillary GC column interior surfaces, on the inlet liner or on the guard column. A combination of sorbitol, gulonolactone, and ethylglycerol was found to be the most effective analyte protectant to cover the whole range of pesticide compounds [90]. The hydroxyl groups of these protectants can interact with active sites on the chromatographic column and in the flowstream and enhanced the pesticide analyte response. Those protectants are of course not required when LC methods are employed for analysis.

### **4.2. Applications of QuEChERS**

146 Chromatography – The Most Versatile Method of Chemical Analysis

then shaken vigorously or mixed on a vortex mixer to distribute the SPE material and facilitate the clean-up process. This d-SPE step is quite similar to matrix solid-phase dispersion developed by Barker [83, 84], but in d-SPE the sorbent is added to an aliquot of the extract rather than to the original solid sample. Moreover, small amounts of sorbents are used because only a small portion of the extract is subjected to the clean-up procedure, and compared to conventional SPE clean-up methods, d-SPE is less laborious and timeconsuming. Magnesium sulfate is again used in this step as a drying agent to remove water and improve analyte partitioning to provide better clean-up. Primary secondary amine (PSA) is the most common SPE sorbent used in QuEChERS procedure for pesticide analysis. The idea is to use a sorbent able to retain matrix components, but not the analytes of interest. However, depending on the sample matrix, other SPE sorbents can also be used alone or combined with PSA, such as C18, OASIS HLB, and graphitized carbon black sorbents. For instance, for samples with high fat content, PSA mixed with C18 is recommended [85] while for samples with moderate or high levels of chlorophyll and carotenoids (for example carrots), PSA mixed with graphitized carbon black is frequently used [86-88]. After the clean-up, the extract is centrifuged and an aliquot of the supernatant can be concentrated or directly analyzed usually by means of Gas Chromatography-Mass Spectrometry (GC-MS) or

Liquid Chromatography-Mass Spectrometry (LC-MS) techniques (Step 6).

approach resulted in the official method AOAC 2007.01.

required when LC methods are employed for analysis.

Although QuEChERS is quite a simple procedure as can be seen from **Figure 8,** in some cases method development will be necessary depending on the family of compounds to be analyzed, and many modifications over the QuEChERS procedure are being proposed. At the end, compromises will be required to ensure simplicity, speed, broad applicability, high recovery and selectivity. For instance, the control of pH in the extraction step is animportant factor when analyzing pesticides in order to ensure an efficient extraction of pH-dependent compounds such as phenocyalcanoic acids and to minimize degradation of labile pesticides under alkaline or acidic conditions. Buffering with citrate salts has been introduced in the first extraction/partitioning step to adjust the pH at around 5 where most labile pesticides under acidic or alkaline conditions are sufficiently stabilized. The pH control can also be very important in other steps of the QuEChERS procedure to prevent degradation of some compounds. For instance, to improve the stability of alkaline-labile compounds after the PSA clean-up step the final sample extract is slightly acidified by the addition of small amounts of formic acid. Of great importance was the introduction of acetate buffering to achieve a pH value of 6 in order to improve recoveries of pH-dependant analytes [89]. This

The use of analyte protectants is also often proposed as an optional step previous to GC analysis for those compounds that might tail or breakdown on the capillary GC column interior surfaces, on the inlet liner or on the guard column. A combination of sorbitol, gulonolactone, and ethylglycerol was found to be the most effective analyte protectant to cover the whole range of pesticide compounds [90]. The hydroxyl groups of these protectants can interact with active sites on the chromatographic column and in the flowstream and enhanced the pesticide analyte response. Those protectants are of course not As commented before, QuEChERS procedure is the sample treatment of choice for the multiresidue analysis of pesticides in fruits and vegetables, and many works can be found in the literature dealing with the simultaneous extraction and clean-up of more than 100 pesticides with acceptable recoveries [75,77,79,80,91]. As an example, Woo Lee et al. developed a new QuEChERS method based on dry ice for the determination of 168 pesticides in paprika [91]. For this purpose, extraction was carried out by using 30 mL of acetonitrile and 10 mL of water, and approximately 10 g of dry ice granules were poured and maintained until layer separation. Clean-up was then carried out by using PSA and GCB sorbents. The separation of the sample extract was induced via the sublimation of dry ice, which occurs at -78.5oC at atmospheric pressure (1 atm). After some minutes of dry ice sublimation with the sample extract, the reduced temperatures of acetonitrile and water ranged from -4.0 to -5.0 oC and from -6.0 to -6.5 oC, respectively, and water was then iced and super cooled. The negative temperatures of the two solvents may reduce their entropies and allowed them to separate. As densities of ice and supercooled water were heavier than that of acetonitrile at 0 oC, water changed to ice and the supercooled water was separated with an acetonitrile layer from the mixed solution. This methodology improved the extraction for flonicamid and its metabolites which was not satisfactory enough using citrate-buffering QuEChERS method.

Recently, the use of carbon-based nanoparticles has also been described as clean-up sorbent for the analysis of pesticides [10]. In this work, multi-walled carbon nanotubes (MWCNTs) were proposed as reversed-dispersive solid phase extraction (r-DSPE) material for the analysis of 30 pesticides in fruits and vegetables. The amount of MWCNTs influenced the clean-up performance and the recoveries, but the use of only 10 mg MWCNTs was suitable for cleaning up all analyzed matrices and showed to be a good alternative to PSA sorbent. The method was validated for different matrices such as spinach, orange and cabbage with recoveries in the range of 71-100% for all 30 pesticides. QuEChERS has also been proposed for the extraction and clean-up of pesticide residues in other food matrices such as milk [85]. In this last work, recovery of 14 different pesticides residues in milk was investigated to respect the amounts of sodium acetate, PSA and C18 used on the clean-up step. Recoveries for hydrophilic pesticides such as myclobutanil ranged from 82 to 99% while lower values (<80%) were obtained for lipophilic pesticides because they were partially removed by C18 along with other fatty compounds.

But today, one of the most important features of QuEChERS may be its application to other families of compounds in a variety of matrices different than fruits and vegetables. Some examples of the use of QuEChERS for the extraction of compounds other than pesticides are given in **Table 3**. QuEChERS methodology has already been applied to the analysis of polycyclic aromatic hydrocarbons (PAHs) in fish and shrimp samples [92,93]. Forsbeg et al. developed and validated a modified QuEChERS method for the determination of 33 parent and substituted PAHs in high-fat smoked salmon that greatly enhanced analyte recovery compared to traditional QuEChERS procedure [93]. For this purpose, a mixture of acetone, ethyl acetate and isooctane instead of acetonitrile was employed for the extraction, and different kinds of salts were used for partitioning. The proposed modified QuEChERS


Current Trends in Sample Treatment Techniques for Environmental and Food Analysis 149

substantially improved average recovery of 15 PAHs by roughly 38% and led to individual gains of 50-125% for some PAHs such as naphthalene and anthracene among others when compared to traditional Soxhlet extraction with hexane. Acrylamide has also been extracted from various food matrices such as chocolate, peanut butter, and coffee [94]. An accurate determination of acrylamide in foodstuffs was possible using QuEChERS since the use of salt and the PSA sorbent increased the selectivity of the method by reducing the content of more polar matrix coextractives. Hexane was required for high fatty samples such as peanut

The extraction of veterinary drugs residues from animal tissues [98] and from milk [95,99,100] has also been described. For instance in [95] the use of QuEChERS was proposed as a fast sample treatment for a rapid screening method in the identification of 21 veterinary drug residues in milk. In this case, 1% acetic acid in acetonitrile together with a 0.1 M Na2EDTA solution was proposed as extraction solvent, using for partitioning magnesium sulfate and sodium acetate, but no further clean-up step was necessary to attain good results. The analysis of persistent organic pollutants (POPs) such as organochloride pesticides and polychlorinated biphenyls (PCBs) in fish tissues was recently reported using conventional QuEChERS procedure but with the addition of a pre-frozen step for 2 hours at -24oC (by means of a homemade freezing device) before the PSA clean-up step for removal of lipids [96]. After this freezing step, between 60 to 70% of lipids were removed. The reduction of co-extractives increased up to 96% by treatment with calcium chloride and PSA. Extraction of mycotoxins and phytohormones from eggs [12] and vegetables [97], respectively, has also been described using QuEChERS with no further clean-up steps, although in the case of mycotoxins a SPE clean-up step using C18 or Oasis HLB cartridges was sometimes proposed. An interesting application of QuEChERS was recently reported by Gallart-Ayala et al. for the analysis of contaminants migrating into food from food packaging materials [11]. In this case, the extraction of UV Ink photoinitiators such as benzophenone and isopropylthioxanthone (ITX) in several foodstuffs (baby food, fruit juices and wine) packaged in tetra brick containers was performed using a common QuEChERS procedure with PSA sorbent for clean-up. The extraction method proposed showed comparable results in terms of method limits of quantification, run-to-run and day-to-day precisions, and quantification results than a previous SPE method reported for the analysis

of ITX, with the advantage of being 12 times faster (per sample).

publications using QuEChERS will considerably increase in the future.

Summarizing, QuEChERS approach appears to have a bright future not only for the analysis of pesticide residues in foods and other agricultural products but also for the analysis of different families of contaminants either in food or even in other matrices. For instance, QuEChERS has also been proposed in environmental analysis for the extraction of chlorinated compounds from soil samples [101]. The simplicity of its use and the great range of modifications that can be applied, make QuEChERS an ideal extraction procedure to think about when dealing with the extraction of any kind of analytes, and the number of

butter.

**Table 3.** Application of QuEChERS procedure for the extraction of different kind of analytes.

substantially improved average recovery of 15 PAHs by roughly 38% and led to individual gains of 50-125% for some PAHs such as naphthalene and anthracene among others when compared to traditional Soxhlet extraction with hexane. Acrylamide has also been extracted from various food matrices such as chocolate, peanut butter, and coffee [94]. An accurate determination of acrylamide in foodstuffs was possible using QuEChERS since the use of salt and the PSA sorbent increased the selectivity of the method by reducing the content of more polar matrix coextractives. Hexane was required for high fatty samples such as peanut butter.

148 Chromatography – The Most Versatile Method of Chemical Analysis

High-Fat Salmon

Fish tissue

Packaged food (baby food, fruit juices, wine)

Polycyclic Aromatic Hydrocarbons

Veterinary drug

Persistent organic pollutants: 22 organochlorine pesticides + 7 polychlorinated biphenyls (PCBs)

residues Milk

10 mycotoxins Eggs

UV Ink Photoinitiators

Phytohormones Vegetables

Acrylamide Foodstuffs

**Compounds Sample QuEChERS procedure Analysis Reference Extraction-partitioning Clean-up** 

NaC2H3O2 150 mg

MgSO4 + 50 mg PSA 2 mL acetone:ethyl GC-MS [93]

> 150 mg MgSO4 + 50 mg PSA


pre-frozen step (2 hours) 1 g calcium chloride + 900 mg MgSO4 + 150 mg PSA

(C18 or Oasis HLB SPE)

250 mg MgSO4 + 750 mg PSA


LC-

LC-

LC-MS or GC-MS [94]

MS/MS [95]

GC-MS [96]

MS/MS [12]

MS/MS [97]

MS/MS [11]

2 mL acetone:ethyl acetate:isooctane (2:2:1

+ 6 g MgSO4 + 1.5 g

acetate:isooctane (2:2:1

+ 4 g MgSO4 + 1 g NaCl + 1g NaC6H7O7 + 0.5 g

5 ml of hexane (only for high fatty matrices) + 10 mL water + 10 mL acetonitrile

+ 4 g MgSO4 + 0.5 g NaCl

10 mL acetonitrile + 10 ml water + 4 g MgSO4 + 1 g NaCl + 0.5 sodium citrate dibasic + 1 g sodium citrate tribasic

10 mL methanol:water (80:20 *v/v*) with 1% acetic acid+ 4 g MgSO4 + 1 g sodium acetate

10 mL acetonitrile with 1% acetic acid + 4 g MgSO4 + 1 g NaCl + 1g sodium citrate + 0.5 disodium citrate

12 mL acetonitrile + 4 g MgSO4 + 1.5 g NaCl

**Table 3.** Application of QuEChERS procedure for the extraction of different kind of analytes.

10 ml acetonitrile (1% acetic acid) + 10 ml 0.1M Na2EDTA solution + 4g MgSO4 + 1 g sodium

*v/v/v*)

*v/v/v*)

Na2C6H8O8

acetate

The extraction of veterinary drugs residues from animal tissues [98] and from milk [95,99,100] has also been described. For instance in [95] the use of QuEChERS was proposed as a fast sample treatment for a rapid screening method in the identification of 21 veterinary drug residues in milk. In this case, 1% acetic acid in acetonitrile together with a 0.1 M Na2EDTA solution was proposed as extraction solvent, using for partitioning magnesium sulfate and sodium acetate, but no further clean-up step was necessary to attain good results. The analysis of persistent organic pollutants (POPs) such as organochloride pesticides and polychlorinated biphenyls (PCBs) in fish tissues was recently reported using conventional QuEChERS procedure but with the addition of a pre-frozen step for 2 hours at -24oC (by means of a homemade freezing device) before the PSA clean-up step for removal of lipids [96]. After this freezing step, between 60 to 70% of lipids were removed. The reduction of co-extractives increased up to 96% by treatment with calcium chloride and PSA. Extraction of mycotoxins and phytohormones from eggs [12] and vegetables [97], respectively, has also been described using QuEChERS with no further clean-up steps, although in the case of mycotoxins a SPE clean-up step using C18 or Oasis HLB cartridges was sometimes proposed. An interesting application of QuEChERS was recently reported by Gallart-Ayala et al. for the analysis of contaminants migrating into food from food packaging materials [11]. In this case, the extraction of UV Ink photoinitiators such as benzophenone and isopropylthioxanthone (ITX) in several foodstuffs (baby food, fruit juices and wine) packaged in tetra brick containers was performed using a common QuEChERS procedure with PSA sorbent for clean-up. The extraction method proposed showed comparable results in terms of method limits of quantification, run-to-run and day-to-day precisions, and quantification results than a previous SPE method reported for the analysis of ITX, with the advantage of being 12 times faster (per sample).

Summarizing, QuEChERS approach appears to have a bright future not only for the analysis of pesticide residues in foods and other agricultural products but also for the analysis of different families of contaminants either in food or even in other matrices. For instance, QuEChERS has also been proposed in environmental analysis for the extraction of chlorinated compounds from soil samples [101]. The simplicity of its use and the great range of modifications that can be applied, make QuEChERS an ideal extraction procedure to think about when dealing with the extraction of any kind of analytes, and the number of publications using QuEChERS will considerably increase in the future.
