1

Validation Data of 127

Pesticides Using a Multiresidue
Method
by LC-MS/MS and GC-MS/MS
in Olive Oil

PESTICIDE RESIDUE LABORATORY

UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 3

CONTENTS

1. Aim and Scope 1

2. Short Description 1

3. Apparatus and Consumables 1

4. Chemicals 1

5. Procedure 2

5.1 Sample Preparation 2

5.2 Recovery Experiments for Method Validation 2

5.3 Extraction 2

5.4 Measurement 3

5.5 Instrumentation and analytical conditions for the LC-MS/MS system 3

5.6 Instrumentation and analytical conditions for the GC-MS/MS system 4

6. Validation of the method 5

6.1 Recoveries and within-laboratory reproducibility

6.2 Limits of Quantification 5

6.3 Linearity 5

7. Conclusions 8

8. References 8

Appendix I - Mass Transitions 9

Appendix II - Validation Results 13

PESTICIDE RESIDUE LABORATORY

UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
1

1. Aim and Scope

This report describes a validation data of 127 pesticides using a multiresidue method
by LC-MS/MS and GC-MS/MS in olive oil.

2. Short Description

The homogeneous sample is diluted with n-hexane and extracted with acetonitrile.
The purification is carried out using a SPE cartridge with C18 (500 mg) as sorbent. The
extract obtained is analysed by GC-MS/MS and LC-MS/MS.

3. Apparatus and Consumables

 Automatic pipettes, suitable for handling volumes of 10 L to 500 L and 1 mL to

3 mL.
 50 ml centrifuge tubes with screw caps (e.g. Sharlab S. L, Spain, article-no 027 –
409926).
 Centrifuge, suitable for the centrifuge tubes employed in the procedure and
capable of achieving at least 1000 rpm.
 C18 SPE cartridges (e.g. Scharlau Extrabond C18 500 mg 6mL).
 Syringes, e.g. 2 mL disposable syringes.
 Syringe filters, 0.45 m pore size.
 Injection vials, 2 ml, suitable for LC and GC auto-sampler.
 Inserts of 300 L suitable for injection vials (e.g. Waters insert 300 L 6X29 mm with
preinstalled plastic spring).
 Test tubes.
 Volumetric flasks.
 Concentration Workstation, e.g. TurboVap LV, Zymark.
4. Chemicals
 n-Hexane 96% HPLC grade
 Acetonitrile HPLC grade
 Methanol HPLC grade
 Cyclohexane GC grade
 Ultrapure water
 Pesticides Standards, e.g. Dr. Ehrenstorfer, Sigma Aldrich.

Page 1
PESTICIDE RESIDUE LABORATORY
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04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 2

5. Procedure
5.1 Sample preparation

Samples were prepared according to the procedure described in the

document “Method Validation and Quality Control Procedures for Pesticide Residues
Analysis in Food and Feed” (Document N° SANCO/12495/2011). The samples were
homogenised by shaking and were stored at room temperature before analysis. As
olive oil is a processed sample it should be analysed within the stated shelf life.

5.2 Recovery Experiments for Method Validation

The olive oil employed did not contain any of the pesticides analysed. Olive oil
obtained from organically grown olives is recommended for the analysis.

The validation method was performed at two fortification levels (0.50 mg/Kg
and 0.05 mg/Kg). Five spiked samples were analyzed at each level.

Working standard solutions containing all the pesticides used for the validation
method were prepared in two different solvents, acetonitrile for LC-MS/MS and
cyclohexane for GC-MS/MS.

5.3 Extraction

1. Weigh 1.5 g ± 0.1 g sample in 50 mL centrifuge tube.

2. Add 1.5 mL of n-hexane. Shake the tube for 20 seconds.
3. Add 6 mL of acetonitrile.
4. Shake vigorously for 10 minutes.
5. Centrifuge for 2 min at 1000 rpm .
6. Condition a C18 SPE cartridge with 5 mL acetonitrile (ACN)
7. Introduce 3 mL of the extract from the upper phase (ACN) into the C18 SPE
cartridge.
8. Elute with 3 mL of ACN.
9. Collect the eluate and repeat the elution with 3 mL of ACN.
10. Take 1 mL of the eluate and evaporate to dryness.

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3

11. Reconstitute with 1 mL of ACN: H2O 1:9 (v:v). Sake the sample in the vortex and
filter with a 0.45 m syringe filter into an injection vial suitable for HPLC-MS/MS.
This extract contains 0.083 g of sample per mL.
12. Transfer 1.5 mL eluate into a test tube. Evaporate to dryness
13. Add 0.5 mL cyclohexane
14. Sake the sample in the vortex.
15. Filter into an injection vial suitable for GC-MS/MS. This extract contains 0.249 g of
sample per mL.

5.4 Measurement

Both LC and GC systems were operated in multiple reaction monitoring mode

(SRM). SRM experiments were carried out to obtain the maximum sensitivity for the
detection of the target molecules. For confirmation of the studied compounds, two
SRM transitions and a correct ratio between the abundances of the two optimised SRM
transitions (SRM2/SRM1) were used, along with retention time matching.
The mass transitions used are presented in Appendix I.

5.5 Instrumentation and analytical conditions for the LC- MS/MS system

LC-MS/MS System 3200 Q TRAP, Applied Biosystems:

- Column: Atlantis T3 3 μm 2.1x200 mm

- Column temperature: 40 ºC
- Mobile phase A: 2 mM ammonium formate in water, 0.1% formic acid.
- Mobile phase B: methanol
- Injection volume: 5 μL
- Autosampler temperature: 10 ºC

Page 3
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
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Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 4

FLOW RATE AND ELUTION GRADIENT

Time (min) A (%) B (%) Flow (μL/min)
0,0 95 5 300
1,0 95 5 300
1,1 70 30 300
10,0 0 100 300
13,0 0 100 300
13,1 95 5 300

5.6 Instrumentation and analytical conditions for the GC-MS/MS system

- GC: Agilent 7890 Series

- Autosampler: Agilent 7683 Injector and sample tray
- Inlet: Splitless
- Carrier gas: Helium
- Inlet pressure: 22.73 psi
- Inlet temperature: 250ºC
- Injection volume: 1 μL
- Oven temperature program:

Hold Time
Rate (ºC/min) Value Run Time (min)
(min)
Initial 70 2 2
Ramp 1 25 150 0 5.2
Ramp 2 3 200 0 21.867
Ramp 3 8 280 10 41.867

- Analytical column: Agilent JW HP-5ms 30 m x 250 μm x 0.25 μm

- Retention time locking: Chlorpyrifos methyl locked to 16.596 min
- Spectrometer: Agilent 7000B Series
- Source temperature: 280 ºC
- Quadrupole temperature: Q1 and Q” = 150 ºC
- Gas flows: Nitrogen at 1.5 mL/min, Helium at 25 ml/min

Page 4
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
5

6 Validation of the method

6.1 Recoveries and within-laboratory reproducibility
The results corresponding to the mean recovery and within-laboratory
reproducibility in terms of relative standard deviation (RSD) at both fortification levels
are summarized in Appendix II.
Document N° SANCO/12495/2011 recommends mean recovery values within the
range 70-120% and RSD<20%. The majority of the validation results fulfil the criteria for
the acceptance mentioned above.

6.2 Limits of Quantification

The limits of quantification (LOQ) of the pesticides analysed in olive oil are 0.050
mg/kg as maximum, taking into account the dilution factor applied during the
extraction process.

6.3 Linearity

Linearity was evaluated in matrix, using matrix-matched calibration curves.

A) Pesticides analysed by LC-QqQ-MS/MS

A matrix-matched calibration curve was constructed for each pesticide at five

calibration levels; 4, 10, 20, 50 and 100 g/L.
The criterion for the acceptance of the linearity (R2 ≥ 0.95) was fulfilled for all
analytes in the method. Figures 1 and 2 show some examples of calibration curves in
olive oil.

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 6

Figure 1: Calibration curve for dimethoate

DIMETHOATE y = 1771,4x - 1059,1

R2 = 0,9974
200000

150000
Area

100000

50000

0
0 10 20 30 40 50 60 70 80 90 100
Standard Concentration (ppb)

Figure 2: Calibration curve for omethoate

OMETHOATE y = 3145,7x - 6269,4

R2 = 0,9955
350000
300000
250000
200000
Area

150000
100000
50000
0
0 10 20 30 40 50 60 70 80 90 100
Standard Concentration (ppb)

B) Pesticides analysed by GC-MS/MS

A calibration curve was constructed for each pesticide at five calibration levels, 10,
20, 50, 100 and 200 g/L. The criterion for the acceptance of the linearity (R2 ≥ 0.95) was
fulfilled for all analytes in the method. Figures 3 and 4 show some examples of
calibration curves in olive oil.

Page 6
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UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
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7

Figure 3: Calibration curve for benfluralin

y = 233,11x 1,087
BENFLURALIN
R2 = 0,9998

80000
70000
60000
50000
Area

40000
30000
20000
10000
0
0 20 40 60 80 100 120 140 160 180 200
Standard Concentration (ppb)

Figure 4: Calibration curve for cypermethrin

CYPERMETHRIN y = 240,66x 1,0831

R2 = 0,9962
90000
80000
70000
60000
50000
Area

40000
30000
20000
10000
0
0 20 40 60 80 100 120 140 160 180 200
Standard Concentration (ppb)

Page 7
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
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 8

7 Conclusions

In spite of the fact that there are some recoveries and RSD values slightly lower or
higher than the values recommended by the SANCO document of method validation,
the results obtained are considered acceptable within the studied range, as in certain
justified cases, typically with multi-residue methods, recoveries outside this range may
be accepted, especially if they are consistent. Due to the high complexity of the matrix
and considering the good repeatability of the method, we accepted those recoveries
in the range 60–140%, which is the range proposed by the guideline for routine analysis
working with multiresidue methods.

This report aims to provide information to laboratories that analyse pesticide

residues in olive oil or are interested in it.

8 References

 Method Validation and Quality Control Procedures for Pesticide Residues Analysis in
Food and Feed (Document N° SANCO/12495/2011)
 http://www.eurl-pesticides.eu

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 9

APPENDIX I: MASS TRANSITIONS

MASS TRANSITIONS FOR LC-MS/MS (3200 Q TRAP, Applied Biosystems)

R.T. R.T.
PESTICIDE TRANSITION PESTICIDE TRANSITION
(min) (min)
223.1 > 126.2 276.2 > 244.2
ACETAMIPRID 11.50 DIMETHENAMID 16.10
223.1 > 72.8 276.2 > 168.2
208.2 > 116.2 230.1 > 199.1
ALDICARB 12.70 DIMETHOATE 11.50
208.2 > 89.1 230.1 > 125.1
240.1 > 223.3 388.2 > 301.1
ALDICARB SULFONE 8.80 DIMETHOMORPH 16.00
240.1 > 86.1 388.2 > 165.2
207.2 > 89.1 327.1 > 205.1
ALDICARB SULFOXIDE 8.40 DIMOXYSTROBIN 17.10
207.2 > 132.1 327.1 > 116.1
404.1 > 372.3 326.1 > 70.0
AZOXYSTROBIN 15.70 DINICONAZOLE 17.90
404.1 > 344.2 326.1 > 43.1
192.1 > 160.2 215.2 > 106.3
CARBENDAZIM 9.10 DMST 13.90
192.1 > 105.2 215.2 > 79.1
222.0 > 165.1 330.1 > 121.0
CARBOFURAN 13.80 EPOXICONAZOLE 16.90
222.0 > 123.0 330.1 > 74.9
238.0 > 163.2 328.1 > 159.0
CARBOFURAN 3-OH 11.30 ETACONAZOL 16.80
238.0 > 181.0 328.1 > 55.2
412.0 > 365.9 304.1 > 217.1
CARFENTRAZONE-ETHYL 17.20 FENAMIPHOS 17.00
412.0 > 345.9 304.1 > 202.0
222.1 > 104.0 FENAMIPHOOS- 336.1 > 249.1
CHLORIDAZON 11.60 13.90
222.1 > 77.1 SULPHONE 336.1 > 202.0
291.0 > 72.0 FENAMIPHOS- 320.1 > 233.1
CHLOROXURON 14.70 13.70
291.0 > 46.0 SULPHOXIDE 320.1 > 202.0
250.0 > 169.1 208.1 > 95.1
CLOTHIANIDIN 11.00 FENOBUCARB 15.80
250.0 > 132.0 204.1 > 152.2
213.1 > 72.1 422.2 > 366.3
CLORTOLURON 14.70 FENPYROXIMATE 19.30
213.1 > 46.0 422.2 > 138.1
199.1 > 128.2 165.0 > 72.1
CYMOXANIL 12.10 FENURON 11.20
199.1 > 111.2 165.0 > 120.0
225.1 > 72.0 408.1 > 182.0
DESMETHYL PIRIMICARB 10.00 FLAZASULFURON 15.40
225.1 > 168.0 408.1 > 227.0
238.1 > 112.2 364.0 > 152.2
DICROTOPHOS 1.30 FLUFENACET 16.70
238.1 > 126.7 364.0 > 124.1
406.1 > 251.1 383.1 > 173.0
DIFENOCONAZOLE 17.80 FLUOPICOLIDE 16.90
406.1 > 75.0 383.1 > 109.0
339.0 > 72.0 459.1 > 427.2
DIMEFURON 15.50 FLUOXASTROBIN 16.50
339.0 > 167.1 459.1 > 188.1
256.1 > 224.2 376.0 > 307.2
DIMETHACHLOR 15.40 FLUQUINCONAZOLE 16.70
256.1 > 148.2 376.0 > 108.1

Page 9
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04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
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 10

R.T. R.T.
PESTICIDE TRANSITION PESTICIDE TRANSITION
(min) (min)
334.0 > 247.3 294.1 > 70.0
FLURTAMONE 15.80 PACLOBUTRAZOLE 16.20
334.0 >178.3 294.1 > 125.0
222.3 > 165.5 248.0 > 202.1
FLUTRIAFOL 14.80 PARAOXON - METHYL 12.90
222.3 > 120.3 248.0 > 90.1
284.1 > 104.0 329.0 > 125.0
FORCHLORFENURON 15.30 PENCYCURON 17.80
284.1 > 228.2 329.0 > 89.0
353.0 > 168.0 296.2 > 91.0
HEXYTHIAZOX 18.90 PETHOXAMID 16.80
353.0 > 228.0 296.2 > 131.2
255.9 > 208.9 318.0 > 160.2
IMIDACLOPRID 10.80 PHOSMET 15.60
255.9 > 175.0 318.0 > 77.0
321.0 > 119.0 367.9 > 145.0
IPROVALICARB 16.60 PICOXISTROBIN 16.90
321.0 > 90.9 367.9 > 145.0
248.9 > 159.9 218.1 > 105.2
LINURON 16.00 PYMETROZINE 7.80
248.9 > 132.9 218.1 > 78.0
315.0 > 99.0 387.9 > 194.0
MALAOXON 13.80 PYRACLOSTROBIN 17.50
315.0 > 127.1 387.9 > 163.0
203.1 > 175.0 242.1 > 186.1
METAMITRON 11.30 TERBUTRYN 16.10
203.1 > 104.1 242.1 > 68.1
320.1 > 70.1 201.9 > 175.0
METCONAZOLE 17.60 THIABENDAZOLE 9.90
320.1 > 125.2 201.9 > 131.0
225.9 > 169.0 252.9 > 126.0
METHIOCARB 16.00 THIACLOPRID 12.10
225.9 > 121.1 252.9 > 73.0
258.0 > 122.0 291.8 > 211.0
METHIOCARB SULPHONE 11.70 THIAMETHOXAM 9.80
258.0 > 201.0 291.8 > 181.0
METHIOCARB 241.9 > 185.0 293.9 > 69.0
11.00 TRIADIMEFON 16.40
SULPHOXIDE 241.9 > 122.0 293.9 > 197.1
163.0 > 88.0 295.9 > 70.0
METHOMYL 9.40 TRIADIMENOL 16.50
163.0 > 106.0 295.9 > 227.2
417.9 > 175.0 408.9 > 186.0
METOSULAM 13.80 TRIFLOXYSTROBIN 17.90
417.9 > 140.1 408.9 > 145.0
224 .0 > 127.0 346.0 > 278.2
MONOCROTOPHOS 9.80 TRIFLUMIZOLE 18.00
224.0 > 193.0 346.0 > 43.1
275.1 > 88.1 318.1 > 70.0
NEBURON 17.20 TRITICONAZOLE 16.80
275.1 > 114.1 318.1 > 125.1
213.9 > 125.0 336.0 > 187.0
OMETHOATE 8.10 ZOXAMIDE 17.60
213.9 > 183.0 336.0 > 159.0
237.0 > 72.1
OXAMYL 9.00
237.0 > 90.0

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 11

MASS TRANSITIONS FOR GC-MS/MS (Agilent 7000B Series)

R.T. R.T.
PESTICIDE TRANSITION PESTICIDE TRANSITION
(min) (min)
206 > 176
266 > 148
BENALAXYL 26.74 DICLORAN 12.56 206 > 124
148 > 105
176 > 148
292 > 264 394 > 266
BENFLURALIN 11.72 DIFLUFENICAM 27.78
292 > 160 266 > 218

311 > 279 ENDOSULFAN 241 > 206

BIFENOX 29.18 22.64
311 > 216 (ALPHA ISOMER) 239 > 204

341 > 185 ENDOSULFAN 241 > 206

BROMOPROPYLATE 28.62 26.16
183 > 155 (BETA ISOMER) 239 > 204

273 > 193 ENDOSULFAN 387 > 253

BUPIRIMATE 24.81 26.76
273 > 108 SULPHATE 387 > 217

323 > 267 125 > 89

CHLORFENVINPHOS 20.98 - 21.57 ESFENVALERATE 34.69
267 > 159 125 > 99

247 > 227 231 > 175

CHLORFENAPYR 25.22 ETHION 26.00
247 > 197 231 > 129

197 > 169 158 > 97

CHLORPYRIFOS 19.23 ETHOPROPHOS 10.74
197 > 107 200 > 158

251 > 139

CHLORPYRIFOS - 286 > 271 FENARIMOL 30.40
16.59 139 > 111
METHYL 286 > 93
277 > 125
299 > 221 FENITROTHION 18.07
CHLORTAL-DIMETHYL 19.43 277 > 109
291 > 109
265 > 210
268 > 205
FENPROPATHRIN 28.99 181 > 152
CHLORTHIOPHOS 26.12 325 > 269
181 > 127
269 > 205
233 > 165
188 > 147 FLUSILAZOLE 24.59
CHLOZOLINATE 21.38 233 > 152
331 > 259
250 > 200
206 > 151 FLUVALINATE-TAU 34.70 – 34.85
250 > 55
CYFLUTHRIN 32.2 - 32.5 163 > 127
163 > 91 242 > 95
FURALAXYL 21.92
181 > 152 301 > 225
CYPERMETHRIN 32.69 - 33.02 163 >127
163 > 91 314 > 245
IPRODIONE 28.39
314 > 56
253 > 93
DELTAMETHRIN 36.00
181 > 152 136 > 108
ISOCARBOPHOS 19.60
230 > 212
304 > 179
DIAZINON 14.47
179 > 137 199 > 121
ISOFENPHOS-METHYL 20.72
241 > 199

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 12

R.T. R.T.
PESTICIDE TRANSITION PESTICIDE TRANSITION
(min) (min)
181 > 152 283 > 96
LAMBDA - PROCYMIDONE 21.96
30.37 163 > 127 283 > 67
CYHALOTHRIN
163 > 91
181 > 145 337 > 267
PROFENOFOS 23.90
LINDANE 13.46 219 > 183 208 > 63
219 > 145
259 > 191
173 > 117 PROPICONAZOLE 26.94 - 27.15 259 > 173
MALATHION 18.80
173 > 99 259 > 69

222 > 220 173 > 145

MEPANIPYRIM 23.06 PROPYZAMIDE 13.95
222 > 193 173 > 109

206 > 162 221 > 193

METALAXYL 17.34 PYRAZOPHOS 30.71
206 > 132 232 > 204

145 > 85 147 > 132

METHIDATHION 22.30 PYRIDABEN 31.53
145 > 58 147 > 117

179 > 152 340 > 199

MYCLOBUTANIL 24.44 PYRIDAFENTHION 28.51
179 > 125 340 > 97

235 > 139 136 > 96

NUARIMOL 27.44 PYRIPROXYFEN 29.86
314 > 139 136 > 78

170 > 141 146 > 118

o-PHENYLPHENOL 8.78 QUINALPHOS 21.64
170 > 115 146 > 91

252.1 > 252 252 > 127

OXYFLUORFEN 24.73 TEBUCONAZOLE 27.43
252 > 196 250 > 125

291 > 109 197 > 141

PARATHION 19.27 TEFLUTHRIN, CIS- 15.08
291 > 79 177 > 127

PARATHION – 263 > 109 231 > 175

16.59 TERBUFOS 13.79
METHYL 263 > 81 231 >129

252 > 162 336 > 218

PENDIMETHALIN 20.99 TETRACONAZOLE 18.86
252 > 161 336 > 204

274 > 125 TOLCLOFOS - 265 > 250

PHENTHOATE 21.71 16.81
274 > 121 METHYL 265 > 93

182 > 111 161 > 134

PHOSALONE 29.67 TRIAZOPHOS 26.46
182 > 138 161 > 106
306 > 264
238 > 145 TRIFLURALIN 11.64 264 > 206
PICOLINAFEN 28.79
238 > 190 264 > 160
238 > 166 212 > 172
PIRIMICARB 15.68 VINCLOZOLIN 16.63 212 > 145
166 > 96
212 > 109
305 > 290
PIRIMIPHOS - METHYL 18.31
290 > 125

Page 12
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 13

APPENDIX II: VALIDATION RESULTS

MEAN MEAN
RECOVERY% RSD% RECOVERY% RSD%
PESTICIDE TECHNIQUE R2
(Spiking Level (n = 5) (Spiking Level (n = 5)
0,05 mg/Kg) 0,5 mg/Kg)
ACETAMIPRID 64% 3% 84% 4% LC-MS/MS 0.9922
ALDICARB (RD) 89% 24% 76% 7% LC-MS/MS 0.9988
ALDICARB SULFOXIDE 89% 19% 82% 5% LC-MS/MS 0.9900
ALDICARB SULFONE 95% 16% 85% 11% LC-MS/MS 0.9958
AZOXYSTROBIN 99% 6% 91% 2% LC-MS/MS 0.9999
BENALAXYL 73% 8% 85% 3% GC-MS/MS 0.9988
BENFLURALIN 71% 8% 76% 2% GC-MS/MS 0.9996
BIFENOX 73% 8% 84% 6% GC-MS/MS 0.9952
BROMOPROPYLATE 66% 7% 72% 6% GC-MS/MS 0.9990
BUPIRIMATE 80% 7% 84% 5% GC-MS/MS 0.9990
CARBENDAZIM 101% 9% 74% 4% LC-MS/MS 0.9940
CARBOFURAN (RD) 62% 2% 72% 5% LC-MS/MS 0.9999
CARBOFURAN 3-OH 70% 16% 79% 5% LC-MS/MS 0.9952
CARFENTRAZONE ETHYL 74% 18% 70% 5% LC-MS/MS 0.9931
CHLOFENVINPHOS 82% 11% 96% 10% GC-MS/MS 0.9970
CHLORFENAPYR 84% 14% 91% 8% GC-MS/MS 0.9933
CHLORIDAZON 73% 10% 81% 4% LC-MS/MS 0.9996
CHLOROXURON 68% 13% 86% 2% LC-MS/MS 0.9980
CHLORPYRIPHOS 72% 5% 74% 3% GC-MS/MS 0.9969
CHLORPYRIPHOS METHYL 74% 9% 88% 7% GC-MS/MS 0.9970
CHLORTAL-DIMETHYL 73% 4% 79% 3% GC-MS/MS 0.9985
CHLORTHIOPHOS 68% 7% 75% 7% GC-MS/MS 0.9987
CHLOZOLINATE 73% 12% 87% 7% GC-MS/MS 0.9997
CLORTOLURON 67% 7% 67% 1% LC-MS/MS 0.9998
CYFLUTHRIN 108% 8% 93% 6% GC-MS/MS 0.9930
CYHALOTHRIN LAMBDA 88% 6% 88% 4% GC-MS/MS 0.9957
CYMOXANIL 82% 5% 78% 7% LC-MS/MS 0.9966
CYPERMETHRIN 98% 6% 88% 4% GC-MS/MS 0.9930
DELTAMETHRIN 96% 7% 90% 4% GC-MS/MS 0.9990
DIAZINON 76% 6% 82% 6% GC-MS/MS 0.9999
DICLORAN 79% 7% 79% 8% GC-MS/MS 0.9981
DICROTOPHOS 87% 6% 81% 7% LC-MS/MS 0.9996
DIFENOCONAZOLE 88% 5% 102% 6% LC-MS/MS 0.9980
DIFLUFENICAN 73% 9% 87% 2% GC-MS/MS 0.9995
DIMEFURON 85% 10% 96% 1% LC-MS/MS 0.9998
DIMETHACHLOR 79% 15% 69% 5% LC-MS/MS 0.9974
DIMETHENAMID 78% 12% 82% 5% LC-MS/MS 0.9978

Page 13
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 14

MEAN MEAN
RECOVERY% RSD% RECOVERY% RSD%
PESTICIDE TECHNIQUE R2
(Spiking Level (n = 5) (Spiking Level (n = 5)
0,05 mg/Kg) 0,5 mg/Kg)
DIMETHOATE (RD) 90% 3% 75% 7% LC-MS/MS 0.9972
OMETHOATE 105% 4% 95% 7% LC-MS/MS 0.9934
DIMETHOMORPH 81% 8% 89% 4% LC-MS/MS 0.9992
DIMOXYSTROBIN 62% 8% 71% 6% LC-MS/MS 0.9999
DINICONAZOLE 69% 14% 86% 7% LC-MS/MS 0.9954
DMST 69% 14% 79% 5% LC-MS/MS 0.9992
ENDOSULFAN SULPHATE (RD) 84% 8% 90% 12% GC-MS/MS 0.9985
ENDOSULFAN (ALPHA ISOMER) 53% 9% 56% 4% GC-MS/MS 0.9985
ENDOSULFAN (BETA ISOMER) 62% 15% 71% 6% GC-MS/MS 0.9995
EPOXICONAZOLE 85% 21% 92% 6% LC-MS/MS 0.9992
ESFENVALERATE/FENVALERATE 91% 7% 89% 5% GC-MS/MS 0.9921
ETACONAZOL 71% 7% 83% 5% LC-MS/MS 0.9992
ETHION 83% 9% 95% 7% GC-MS/MS 0.9983
ETHOPROPHOS 69% 7% 83% 2% GC-MS/MS 0.9997
FENAMIPHOS (RD) 75% 12% 82% 3% LC-MS/MS 0.9990
FENAMIPHOS SULFOXIDE 77% 10% 85% 3% LC-MS/MS 0.9980
FENAMIPHOS SULFONE 75% 13% 89% 2% LC-MS/MS 0.9998
FENARIMOL 67% 6% 74% 6% GC-MS/MS 0.9983
FENITROTHION 105% 10% 104% 12% GC-MS/MS 0.9963
FENOBUCARB 76% 7% 80% 4% LC-MS/MS 0.9936
FENPROPATHRIN 72% 7% 77% 5% GC-MS/MS 0.9969
FENPYROXIMATE 83% 4% 70% 2% LC-MS/MS 0.9936
FENURON 82% 4% 80% 2% LC-MS/MS 0.9996
FLAZASULFURON 110% 11% 102% 7% LC-MS/MS 0.9996
FLUFENACET 75% 20% 86% 4% LC-MS/MS 0.9942
FLUOPICOLIDE 81% 16% 78% 4% LC-MS/MS 0.9998
FLUOXASTROBIN 92% 9% 86% 3% LC-MS/MS 0.9964
FLUQUINCONAZOLE 94% 18% 87% 7% LC-MS/MS 0.9984
FLURTAMONE 91% 9% 92% 3% LC-MS/MS 0.9996
FLUSILAZOLE 80% 8% 81% 11% GC-MS/MS 0.9992
FLUTRIAFOL 78% 16% 72% 5% LC-MS/MS 0.9999
FLUVALINATE 107% 6% 102% 6% GC-MS/MS 0.9980
FORCHLORFENURON 105% 10% 82% 2% LC-MS/MS 0.9960
FURALAXYL 68% 10% 82% 10% GC-MS/MS 0.9986
HEXYTHIAZOX 89% 11% 68% 4% LC-MS/MS 0.9976
IMIDACLOPRID 65% 8% 74% 3% LC-MS/MS 0.9998
IPRODIONE 94% 14% 97% 20% GC-MS/MS 0.9947
IPROVALICARB 98% 19% 99% 6% LC-MS/MS 0.9980
ISOCARBOPHOS 95% 11% 110% 18% GC-MS/MS 0.9967
ISOFENPHOS-METHYL 78% 7% 93% 6% GC-MS/MS 0.9993
LINDANE 74% 7% 79% 10% GC-MS/MS 0.9991
LINURON 87% 15% 88% 8% LC-MS/MS 0.9970
MALATHION (RD) 91% 11% 111% 8% GC-MS/MS 0.9909
MALAOXON 76% 11% 74% 4% LC-MS/MS 0.9973
MEPANIPYRIM 74% 9% 84% 5% GC-MS/MS 0.9987

Page 14
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 15

MEAN MEAN
RECOVERY% RSD% RECOVERY% RSD%
PESTICIDE TECHNIQUE R2
(Spiking Level (n = 5) (Spiking Level (n = 5)
0,05 mg/Kg) 0,5 mg/Kg)
METALAXYL 74% 13% 84% 6% GC-MS/MS 0.9996
METAMITRON 68% 7% 83% 3% LC-MS/MS 0.9978
METCONAZOLE 90% 5% 93% 6% LC-MS/MS 0.9996
METHIDATHION 100% 11% 114% 17% GC-MS/MS 0.9953
METHIOCARB (RD) 73% 12% 92% 6% LC-MS/MS 0.9986
METHIOCARB SULFOXIDE 88% 9% 90% 4% LC-MS/MS 0.9996
METHIOCARB SULFONE 78% 7% 78% 6% LC-MS/MS 0.9996
METHOMYL 86% 11% 81% 10% LC-MS/MS 0.9966
METOSULAM 85% 11% 106% 2% LC-MS/MS 0.9978
MONOCROTOPHOS 72% 4% 93% 5% LC-MS/MS 0.9990
MYCLOBUTANIL 77% 9% 75% 11% GC-MS/MS 0.9991
NEBURON 66% 13% 74% 5% LC-MS/MS 0.9990
NUARIMOL 67% 8% 76% 10% GC-MS/MS 0.9979
o-PHENYLPHENOL 66% 7% 82% 4% GC-MS/MS 0.9997
OXAMYL 61% 11% 95% 10% LC-MS/MS 0.9938
OXYFLUORFEN 101% 10% 96% 4% GC-MS/MS 0.9906
PACLOBUTRAZOLE 84% 10% 73% 2% LC-MS/MS 0.9966
PARATHION-METHYL (RD) 95% 11% 108% 12% GC-MS/MS 0.9904
PARAOXON METHYL 78% 8% 83% 2% LC-MS/MS 0.9990
PENCYCURON 92% 5% 96% 2% LC-MS/MS 0.9994
PENDIMETHALIN 77% 6% 78% 3% GC-MS/MS 0.9988
PETHOXAMID 68% 8% 79% 4% LC-MS/MS 0.9988
PHENTHOATE 83% 9% 99% 9% GC-MS/MS 0.9957
PHOSALONE 102% 6% 107% 15% GC-MS/MS 0.9981
PHOSMET 88% 13% 79% 7% LC-MS/MS 0.9994
PICOLINAFEN 74% 7% 86% 2% GC-MS/MS 0.9992
PICOXISTROBIN 116% 11% 103% 6% LC-MS/MS 0.9994
PIRIDAFENTHION 95% 7% 108% 15% GC-MS/MS 0.9984
PIRIMICARB (RD) 65% 13% 76% 9% GC-MS/MS 0.9997
DESMETHYL PIRIMICARB 82% 3% 84% 4% LC-MS/MS 0.9988
PROCYMIDONE 77% 9% 84% 6% GC-MS/MS 0.9983
PROFENOFOS 81% 10% 94% 15% GC-MS/MS 0.9899
PROPICONAZOLE 71% 8% 78% 5% GC-MS/MS 0.9993
PROPYZAMIDE 72% 8% 84% 3% GC-MS/MS 0.9998
PYMETROZINE 78% 9% 81% 3% LC-MS/MS 0.9992
PYRACLOSTROBIN 84% 15% 96% 6% LC-MS/MS 0.9942
PYRAZOPHOS 92% 7% 102% 12% GC-MS/MS 0.9927
PYRIDABEN 68% 7% 71% 6% GC-MS/MS 0.9984
PYRIPROXYFEN 65% 7% 72% 2% GC-MS/MS 0.9987
QUINALPHOS 74% 8% 84% 7% GC-MS/MS 0.9977
TEBUCONAZOLE 84% 9% 77% 11% GC-MS/MS 0.9934
TEFLUTHRIN 66% 3% 73% 1% GC-MS/MS 0.9996
TERBUFOS 65% 5% 75% 9% GC-MS/MS 0.9999
TERBUTRYN 53% 12% 83% 5% LC-MS/MS 0.9974
TETRACONAZOLE 75% 11% 81% 9% GC-MS/MS 0.9991
THIABENDAZOLE 74% 10% 76% 6% LC-MS/MS 0.9986

Page 15
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551
 16

MEAN MEAN
RECOVERY% RSD% RECOVERY% RSD%
PESTICIDE TECHNIQUE R2
(Spiking Level (n = 5) (Spiking Level (n = 5)
0,05 mg/Kg) 0,5 mg/Kg)
THIACLOPRID 59% 4% 68% 1% LC-MS/MS 0.9994
THIAMETHOXAM (RD) 80% 10% 93% 3% LC-MS/MS 0.9982
CLOTHIANIDIN 59% 44% 80% 12% LC-MS/MS 0.9972
TOLCLOFOS-METHYL 73% 6% 83% 2% GC-MS/MS 0.992
TRIADIMEFON 100% 23% 96% 3% LC-MS/MS 0.9978
TRIADIMENOL 44% 36% 101% 36% LC-MS/MS 0.9955
TRIAZOPHOS 103% 9% 105% 15% GC-MS/MS 0.9981
TRIFLOXYSTROBIN 104% 21% 116% 9% LC-MS/MS 0.9580
TRIFLUMIZOLE 76% 9% 86% 4% LC-MS/MS 0.9998
TRIFLURALIN 71% 8% 78% 5% GC-MS/MS 0.9998
TRITICONAZOLE 71% 17% 80% 6% LC-MS/MS 0.9984
VINCLOZOLIN 72% 13% 84% 6% GC-MS/MS 0.9986
ZOXAMIDE 98% 14% 105% 9% LC-MS/MS 0.9958

Page 16
PESTICIDE RESIDUE LABORATORY
UNIVERSITY OF ALMERIA
AGRO-FOOD ANALYSIS SERVICE
CITE I
Dirección General de Innovación Agraria y Ganadería
Dpto. Hidrogeología y Química Analítica
Consellería de Agricultura, Pesca y Alimentación
Ctra. Sacramento s/n. La Cañada de San Urbano
GENERALITAT VALENCIANA
04120 ALMERÍA, SPAIN
Pintor Goya, 8, 46100 Burjassot-VALENCIA, SPAIN
Tel (+34) 950 01 50 34/55 31
Tel (+34) 963 631 551