Review 18-11-2011 final.pmd

Madras Agric. J., 98 (7-9): 282-285, September 2011 QuEChERS Method for Determination of Some Chlorinated
Hydrocarbon and Synthetic Pyrethroid Residues in Sheep Meat
by Gas Chromatography-Electron Capture Detector
M. Paramasivam*, R. Harischandra Naik and S. Chandrasekaran
Pesticide Toxicology Laboratory, Department of Agricultural Entomology Tamil Nadu Agricultural University, Coimbatore - 641 003 A rapid, simple and efficient multiresidue method was developed and optimized for the
identification and quantification of organochlorine pesticides (OCP) and synthetic pyrethroids
(SPs) in sheep meat samples. The method consists of a modified Quick, Easy, Cheap, Effective,
Rugged, and Safe (QuEChERS) sample preparation method. Samples were extracted with
acetonitrile, and the extracts were cleaned up by dispersive solid phase extraction with
primary secondary amine (PSA) sorbent and anhydrous magnesium sulphate. Determination
and quantification of OCP residues was carried out using a GC-ECD. Mean recoveries were
found in the ranges 70-110 % and 84-99 % for the investigated OCPs and SPs, respectively, with
the RSD was less than 20%. This method was found more efficient and reliable enabling more
number of samples to be analysed in less time. Moreover lipid removal was achieved to a
large extent to get desired result.
Keywords: QuEChERS, Multiresidue analysis, Sheep meat, Chlorinated hydrocarbons, Synthetic
Sheep meat is contaminated with pesticides in tissues are based on an extraction step with ethyl feed, water and during exposure of animals to pest acetate (Muhammad et al., 2010), acetone (Barbini control activity directly or indirectly. Biological activity et al., 2007) and hexane-acetonitrile (Darko and of pesticides is not only restricted to the target Acquaah, 2007) followed by partition and clean-up organisms but also extends to non targets animals in order to remove the fatty interference, followed by (Singh and Dhaliwal, 1993). In India for crop a gas chromatographic determination with electron protection, the usage of organochlorines, particularly, capture or mass spectrometry detection (GC-ECD, cyclodienes are banned but restricted use of these chemicals is allowed in the case of public health Unlike many earlier pesticide residue analysis programme, especially malaria control programme.
methods developed for traditional chromatographic Pyrethroids have been used widely throughout the detection systems (e.g. Ultra Violet/vis absorbance, world for insect control in agriculture and to combat fluorescence, element-selective detectors), the human and animal parasites (Mastovska et al., QuEChERS (Quick, Easy, Cheap, Effective, Rugged 2006). Pesticides used for cattle disease control and Safe) approach takes advantage of the wide can affect public health and international trading of analytical scope and high degree of selectivity and food products, if maximum residue levels (MRL) are above those stipulated by FAO and WHO.
chromatography (GC and LC) coupled. GC, GC-MS Chlorinated hydrocarbons have low acute toxicity but possess greater potentials for causing chronic Spectrometry (LC-MS/MS) have become the main toxicity due to liphophilic nature. The OCP residues analytical tools in most pesticide monitoring in poultry egg samples varied from BDL to 0.031 µg/g laboratories to meet world standards. Thus the (Suganthy et al., 2009). Pesticide residues streamlined features, practical benefits and excellent accumulating in the tissues of animals lead to bio- accumulation in different levels and ultimately to preparation approach lead to the great popularity of biomagnification. Government of India and QuEChERS concept (Anastassiades et al., 2003, Directorate of Plant Protection and Quarantine in Lehotay et al., 2010). The QuEChERS has been the year 2008, banned diclofenac, a veterinary drug applied with success on several food matrices such due to hazards biocidal nature to folken birds.
as fruits and vegetables (Anastassiades et al., The most commonly reported methods for the 2003,) and low-fatty (2-20%) food matrices, such as analysis of pesticide residues in meat and animal milk, egg, and avocado (Lehotay et al., 2005).
*Corresponding author email: [email protected] In fat samples, conventional methods for the market. The meat sample (1 kg) was blended with analysis of pesticide residues usually involve high volume blade homogeniser and used for the laborious and time-consuming clean-up steps.
Moreover, analytical problems associated with lipids Recovery studies
extraction when using gas chromatographic system,especially electron capture detector, due to matrix interferences, may cause signal suppression or sample was mixed with a known amount of OCPs enhancement. In multi-residue analysis, the sample and SPs mixture in a 50ml polypropylene centre preparation process cannot be selective to remove fuge tube. In controlled sample, same amount of chemical compounds of the matrix if they have hexane was added. Both tubes were kept for 30 similar properties as the analytes. Modifications of minutes to allow the pesticide standard reference materials to get imbibed in to the ground meat.
solvents, salt volumes, and clean-up sorbents. The Extraction and cleanup
aim of the study was to develop a simple and efficientmultiresidue analysis in sheep meat. In order to The extraction of pesticide residue from meat shorten the analytical procedure during extraction sample was followed by QuEChERS method with modification (Anastassiades et.al. 2003). An aliquotof 10 g. of homogenised sample was weighed in to Materials and Methods
a 50 ml of polypropylene centrifuge tube and 20 ml Chemicals
of acetonitrile was added and hand shakenimmediately for 2 minutes. Four grams of anhydrous Acetone, acetonitrile, and hexane of HPLC grade, magnesium sulphate and 1 g. of sodium chloride anhydrous magnesium sulphate, sodium chloride were further added and shaken immediately for 30 of analytical grade all from M/s. Merck (Mumbai, seconds with the closed screw cap. The mixture India), primary secondary amine (PSA) from M/s.
was then centrifuged at about 10000 rpm for 10 minutes using a centrifuge (Plasto Craft®) to Preparation of standards
separate the sediments and water from acetonitrile.
Next the 4 ml (equivalent to the 2 g.of sample) of supernatant of acetonitrile layer obtained after Accustandard; Inc, (New Haven, CT06513, USA).
salting out was loaded in to a 15 ml centrifuge tube Each insecticide was dissolved in hexane to make containing 100 mg of primary secondary amine and a 1000 µg/g stock standard solution. Intermediate and working standard solutions were prepared from immediately followed by the centrifugation at about stock solution by serial dilution technique. Five 5000 rpm at 5 minutes . After centrifugation, 2 ml of working standard solutions ranging from 0.01, 0.05, supernatant was taken out in to the turbovap tube 0.1, 0.5, and 1 µg/g were prepared from standard and concentrated to dryness by using the Turbovap mixture for a linearity check. Spiking solutions were evaporator (Caliper Life Sciences, Russelsheim, used for fortifying the sample and also for the Germany) at 40°C with a gentle stream of nitrogen calculation after appropriate dilution. All the solutions and the residues were redissolved in 1 ml of hexane were protected against light with aluminium foil and and subsequently filtered through membrane filter paper to remove the excess of colouring materials Instrumental parameters
and transferred in to GC vials. The concentration ofsample represented by the test solution was 1g/ml.
A GC-2010 (Shimadzu, Japan), equipped with The detection and quantification was carried out by ECD with 63Ni radio isotope 370 MBQ (10mci) as source of detector in gas chromatography. Capillaryfussed silica column (J&W, scientific, USA), DB-5, Results and Discussion
30m X 0.25 mm id X 0.25 µm film thickness. Carrier In the present work, the QuEChERS method was gas: nitrogen 2.0 ml/min (constant flow mode), make followed to extract the pesticide residues from the up gas-nitrogen; Injector temperature 250°C and sheep meat. This approach can significantly reduce the analysis time and solvent consumption.
The GC column oven was initially set at a Anastassiades et al., (2003) reported a QuEChERS temperature of 160° C for 1 min, increased @ 3°C / method of pesticide extraction by using acetonitrile min to 200°C, held for 2 min and increased @ 4°C as a vortex mixture, the cleanup procedure was / min to 220°C held for 4 min finally increased @ performed by dispersive solid phase extraction 5°C / min to 250°C held for 15 min. Split injection at using the primary secondary amine and finally the a volume of 1 µl by Shimadzu AOC 20i auto injector extract was injected in to GC-ECD system. Electron capture detector was found to be fairly sensitive toall selected pesticides under standardised GC- Sample preparation
ECD parameters. Blank as well as control sample Meat samples were collected from the local did not show any peak that could be attributed to the Table 1. Average recovery and relative standard
Table 1 and 2 show recoveries and relative deviation of chlorinated hydrocarbon from sheep
standard deviation (RSD) values obtained for spiked meat samples by modified QuEChERS method
samples at spiking level. These values evidencedthat the method achieved acceptable recoveries for all pesticides (70-120%), with RSD lower than 15%.
Most pesticides gave 70-120% acceptablerecoveries with associated RSD<15%, therefore developed method may be used as a quantitative More lipophilic pesticides (alpha HCH, delta HCH, aldrin, and dieldrin) gave recoveries of 70-80 % (Table 1) and reasonably consistent (RSD<10%).
Lower recoveries were unavoidable in some pesticides for the satisfactory chromatographic performance, as well to remove co-extractive fats.
Consistent recoveries and accurate quantitation of most lipophilic residues will require either the use of an internal standard matched closely to the lipophilic analytes, the use of an extracted matrix- matched calibration or to correct the results for the Table 2. Average recovery and relative standard
deviation of synthetic pyrethroids from sheep
meat samples by modified QuEChERS method
* Means of three replication, SD- standard deviation, RSD-Relative standard deviation studies of pesticides. All the pesticides gave well resolved peaks. However, permethrin, beta- cyfluthrin, fenvalerate and deltamethrin gave twopeaks due to separation of isomers and the mean of their two peakes was used to calculation. The linearity range of multiresidue pesticides was worked out by plotting the response against a standard and ranged from 0.01 to 1 µg / g.
Selection and optimization of extraction and cleanup procedure are very important in multiresidue analysis. The extraction of pesticide residue depends upon the polarity of the pesticides as well as the type of pesticide matrix. Owing to the wide range of polarity and solubility exhibited by the * Means of three replication, SD- standard deviation, RSD-Relative standard deviation compounds investigated, acetonitrile was selectedas one of the solvents for extraction of pesticides Among synthetic pyrethroids, alpha cypermethrin because of its effectiveness of polar and non polar (74.40%) recorded the lowest recovery value of 74.40 pesticides from the diverse range of matrices. The % and deltamthrin I recorded the highest value of clean up step was modified by the addition of 100mg 110.23 %.Fenpropathrin (76.28), lambda cyhalothrin PSA to the remaining fats. The added PSA effectively (78.93%),permethrin I (91.34%), permethrin II removed many polar matrix components, such as (95.32%), beta cyfluthrin I (81.65%), beta cyfluthrin organic acids, certain polar pigments, and sugars, II (82.50%), fenvalerate I (85.53%), fenvalerate II from the food extracts. The co-extractives in the (100.77%), and deltamethrin II (77.73%) showed extract, because of insufficient cleanup of sample acceptable range of recovery values. Analytical cause rapid deterioration of gas chromatographic problems associated with the presence of lipids system especially electron capture detector, thereby were not observed in spiked samples after the precluding reliable results. In the QuEChERS analysis of routine sequences: Retention time drifted method originally used for pesticide extraction, 10 about 0.1% and response drifted lower than 30% at ml of acetonitrile was added to 10 g. of sample followed by addition of magnesium sulphate and Detector was found quite sensitive not only for salting out step to remove water and PSA to bind organochlorine but also synthetic pyrethriods. The with organic acids polar colouring material or limit of detection (LOD) of the method was worked glucosides. The sample matrix was less disturbed out based on the signal to noise ratio of 1:3 and when using primary secondary amines which limit of quantification (LOQ) based on 1:10 ratio.
ECD was found to be fairly sensitive for all the 24 pesticides under standardised GC parameters all determination of the residues of four pyrethroids in the pesticides gave well resolved peaks. The meat by GC-ECD and confirmation by GC-MS. Anal. QuEChERS method takes the advantage of several Bioanal. Chem., 389: 1791-1798.
features of acetonitrile to provide a rather selective Darko, G. and Acquaah, S.O. 2007. Levels of organo isolation of pesticide residue over a wide polarity chlorine pesticides residues in meat. Int. J. Environ.
Sci. Tech., 4: 521-524.
Lehotay, S.J., Kyung, A. S., Hyeyoung. K., Urairat, K.C., The proposed method is sensitive, quick, easy, Wusheng. F., Katerina, M., Eunha, H. and Natchanun, and allows the multiresidue determination of 24 representative pesticides including OCPs and SPs preparation methods for the analysis of pesticide pesticides in fat samples, with recoveries ranging residues in fruits and vegetables. J. Chromo. A.,
1217: 2548-2560.
from 70% to 120% for most of them with theoptimized conditions. The proposed method shows Lehotay, S.J., Mastovska, K. and Yun, S.J. 2005. Evaluation of two fast and easy methods for pesticide residue good sensitivity and recovery and allows for rapid analysis in fatty food matrices. J. AOAC Int., 88:
analysis. This method requires only small volumes of solvents per sample and needs no special Mastovska, K. and Lehotay, S.J. 2006. Rapid sample equipments. Thereby, this method could be used preparation method for LC/MS/MS or GC/MS analysis for routine analysis in laboratories for national and of acrylamide in various food matrices. J. Agric. community monitoring programs of different Food Chem., 54: 7001-7008.
families of pesticides in animal fat with equipment Muhammad, F., Akhtar, M., Rahman, Z.U., Farooq, H.U., that allows quantitation and confirmation in one Khaliq, T. and Anwar, M.I. 2010. Multi-residue determination of pesticides in the meat of cattle inFaisalabad-Pakistan. Egyptian Acad. J. Biol. Sci., References
2: 19-28.
Anastassiades, M., Lehotay, S.J., Stajnbaher, D. and Singh, B. and Dhaliwal, G.S. 1993. Pesticide contamination Schenck, F.J. 2003. Fast and easy multiresidue of fatty foods in developing countries. In: Dhaliwal,
method employing acetonitrile extraction / partitioning G.S. and Singh (Eds), Pesticides: Their Ecological and "Dispersive Solid-Phase Extraction" for the Impact in Developing Countries. Commonwealth determination of pesticide residues in produce, J. AOAC Int., 86:412-431.
Suganthy, M., Kuttalam, S. and Chandrasekaran, S. 2009.
Barbini, D.A., Vanni, F., Girolimetti, S. and Dommarco, R.
Monitoring of chlorinated hydrocarbon insecticides 2007. Development of an analytical method for the and chlorpyriphos residues in market samples of egg.
Madras Agric. J., 96: 396-397.
Received: August 12, 2011; Accepted: September 15, 2011

Source: http://www.masutnau.in/pdf/98/98-7-9/98-7-9-282-285.pdf