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COLLABORATIVE INTERNATIONAL PESTICIDES ANALYTICAL COUNCIL LIMITED Commission Internationale des Méthodes d'Analyse des Pesticides (CIMAP) Long lasting insecticidal nets (LN) are becoming more and more important in the control and prevention of diseases like malaria. In order to meet an urgent need for methods to characterise LN, CIPAC provides selected methods as download. By downloading these methods you accept the following conditions of use: • these methods are provided subject to changes without prior notice • the copyright of these methods is with CIPAC • CIPAC declines any responsibility when using these methods PERMETHRIN
ISO common name Permethrin Chemical name 3-Phenoxybenzyl (1RS)-cis,trans-3-(2,2-dichlorovinyl) -2,2-dimethylcyclopropanecarboxylate (IUPAC); (3-Phenoxyphenyl)methyl 3-(2,2-dichloroethenyl)-2,2 -dimethylcyclopropanecarboxylate (CA) RMM 391.29 v.p. 2.06 x 10-7 Pa (20°C), In water: 11.1µg/l (20±0.5°C/pH7.0-9.3); soluble in organic solvents Yellow to yellowish brown oil and solidifies on lowering temperature PERMETHRIN TECHNICAL
331/TC/m/-

1 Sampling. Take at least 100 g.
2 Identity tests
2.1 GLC. Use the GLC method below. The retention time of permethrin for the
sample solution should not deviate by more than 1% from that for the
permethrin working standard solution and intensities of the permethrin isomers
should give the same pattern as in the working standard solution.
2.2 Infrared. Prepare a film between NaCl plates and scan from 4000 to 400
cm-1. The spectrum produced from the sample should not differ significantly
from that of the standard.
3 Permethrin
OUTLINE OF METHOD The content of permethrin in the test samples are
determined by capillary GC using flame ionisation detection and triphenyl
phosphate as internal standard, and the trans-isomer ratio is calculated from
the chromatogram obtained.
The content of permethrin is the total content of cis- and trans-isomers.
REAGENTS
Acetone analytical grade
Permethrin working standard technical product of certified purity. Store
Triphenyl phosphate internal standard. Must not show peaks with the same retention times as cis-permethrin and trans-permethrin. Internal standard solution. Dissolve triphenyl phosphate (1.0 g) in acetone (100 ml). Ensure that a sufficient quantity of this solution is prepared for all samples and calibration standards to be analysed. Calibration solution. Homogenise the permethrin working standard. When the permethrin working standard is waxy solid or partly waxy solid homogenise it by warming it to melting and by stirring. Prepare calibration solutions in duplicate. Weigh (to the nearest 0.1 mg) 90 to 110 mg (s mg) of permethrin working standard into a vial or stoppered flask (100 ml). Add by pipette internal standard solution (5 ml) and dissolve completely. Add by measuring cylinder acetone (45 ml) and mix well (Solutions CA and CB). APPARATUS Gas chromatograph equipped with a split/splitless injection and a flame Capillary column fused silica, 30 m x 0.25 (i.d.) mm, film thickness: 0.25 µm, coated with crosslinked dimethyl polysiloxane (DB-1 or equivalent) Electric integrator or data system PROCEDURE (a) Gas chromatographic conditions (typical): fused silica, 30 m x 0.25 (i.d.) mm, film thickness: 0.25 µm, coated with crosslinked dimethyl polysiloxane (DB-1 or equivalent) (b) Linearity check. Check the linearity of the detector response by injecting 1 µl of solutions with permethrin concentrations 0.5, 1 and 2 times that of the calibration solution before conducting analysis. (c) System equilibration. Prepare two calibration solutions. Inject 1 µl portions of the first one until the response factors obtained for two consecutive injections differ by less than 1.0%. Then inject a 1 µl portion of the second solution. The response factor for this solution should not deviate by more than 1.0% from that for the first calibration solution, otherwise prepare new calibration solutions. (d) Preparation of sample solution. Homogenise the sample. When the sample is waxy solid or partly waxy solid homogenise it by warming it to melting and by stirring. Prepare sample solutions in duplicate for each sample. Weigh (to the nearest 0.1 mg) 90 to 110 mg (w mg) of permethrin into a vial or stoppered flask (100 ml). Add by pipette internal standard solution (5 ml) and dissolve completely. Add by measuring cylinder acetone (45 ml) and mix well (Solutions SA and SB). (e) Determination. Inject in duplicate 1 µl portions of each sample solution bracketing them by injections of the calibration solutions as follows; calibration solution CA, sample solution SA, sample solution SA, calibration solution CB, sample solution SB, sample solution SB, calibration solution CA, and so on. Measure the relevant peak areas. (f) Calculation of permethrin content. Calculate the mean value of each pair of response factors bracketing the two injections of a sample and use this value for calculating the permethrin contents of the bracketed sample injections. fi = individual response factor f Hs = total peak area of permethrin (cis-permethrin + trans-permethrin) in Hw = total peak area of permethrin (cis-permethrin + trans-permethrin) in Ir = peak area of the internal standard in the calibration solution Iq = peak area of the internal standard in the sample solution s = mass of permethrin working standard in the calibration solution (mg) w = mass of sample taken (mg) P = purity of permethrin working standard (g/kg) Repeatability r
= 9 g/kg at 953 g/kg active ingredient content = 9 g/kg at 951 g/kg active ingredient content Reproducibility R = 23 g/kg at 953 g/kg active ingredient content
= 18 g/kg at 951 g/kg active ingredient content (g) Calculation of trans-isomer ratio. Hwt = peak area of trans-permethrin in the sample solution Hwc = peak area of cis-permethrin in the sample solution PERMETHRIN LONG LASTING INSECTICIDAL NET
331/LN/m/-

1 Sampling. Take at least 100 g.

2 Identity tests
2.1 GLC. As for 331/TC/m/2.1
2.2 Infrared. Extract the sample with suitable solvent. Filter and evaporate
the solvent. Proceed as for 331/TC/m/2.2
3 Permethrin As for 331/TC/m/3 except:
REAGENTS
Heptane analytical grade
Internal standard solution. Dissolve triphenyl phosphate (1.0 g) in heptane (150 ml). Ensure that a sufficient quantity of this solution is prepared for all samples and calibration standards to be analysed. Calibration solution. Homogenise the permethrin working standard. When the permethrin working standard is waxy solid or partly waxy solid homogenise it by warming it to melting and by stirring. Prepare calibration solutions in duplicate. Weigh (to the nearest 0.1 mg) 72 to 88 mg (s mg) of permethrin working standard into a vial or stoppered flask (200 ml). Add by pipette internal standard solution (10 ml) and dissolve completely. Add by measuring cylinder heptane (90 ml) and mix well (Solutions CA and CB). PROCEDURE (d) Preparation of sample solution. Clean scissors with acetone before use. Cut the sample with the scissors into 5 – 10 mm square. Prepare sample solutions in duplicate for each sample. Weigh (to the nearest 0.1 mg) sufficient sample to contain 36 to 44 mg (w mg) of permethrin into a vial or stoppered flask (100 ml). Add by pipette internal standard solution (5 ml) and by measuring cylinder heptane (45 ml). Place the vial or stoppered flask in a water bath (85 – 90°C) for 45 minutes. Shake the vial or stoppered flask once or twice during extraction. Filter a portion of each sample solution through a filter paper prior to analysis (Solutions SA and SB). (f) Calculation of permethrin content. Calculate the mean value of each pair of response factors bracketing the two injections of a sample and use this value for calculating the permethrin contents of the bracketed sample injections. fi = individual response factor f Hs = total peak area of permethrin (cis-permethrin + trans-permethrin) in Hw = total peak area of permethrin (cis-permethrin + trans-permethrin) in Ir = peak area of the internal standard in the calibration solution Iq = peak area of the internal standard in the sample solution s = mass of permethrin working standard in the calibration solution (mg) w = mass of sample taken (mg) P = purity of permethrin working standard (g/kg)
Repeatability r
= 1.6 g/kg at 20.3 g/kg active ingredient content = 1.3 g/kg at 20.0 g/kg active ingredient content
= 0.9 g/kg at 18.7 g/kg active ingredient content
Reproducibility R = 1.9 g/kg at 20.3 g/kg active ingredient content
= 1.5 g/kg at 20.0 g/kg active ingredient content = 1.5 g/kg at 18.7 g/kg active ingredient content
4 Surface concentration and release index
REAGENTS As for 331/TC/m/3 except:
Internal standard solution for calibration solution. Dissolve triphenyl phosphate
(0.1 g) in acetone (200 ml) to prepare a stock solution. Transfer by pipette the stock solution (5 ml) to a volumetric flask (50 ml). Make up to volume with acetone and mix well. Ensure that a sufficient quantity of this solution is prepared for all calibration standards to be analysed. Internal standard solution for sample solution. Transfer by pipette the stock solution (5 ml) to a volumetric flask (500 ml). Make up to volume with acetone and mix well. Ensure that a sufficient quantity of this solution is prepared for all samples to be analysed. Calibration solution. Homogenise the permethrin working standard. When the permethrin working standard is waxy solid or partly waxy solid homogenise it by warming it to melting and by stirring. Prepare calibration solutions in duplicate. Weigh (to the nearest 0.1 mg) 90 to 110 mg (s mg) of permethrin working standard into a volumetric flask (100 ml) and make up to volume with acetone and mix well. Transfer by pipette this solution (1 ml) to a volumetric flask (20 ml), make up to volume with acetone and mix well. Transfer by pipette this solution (5 ml) to a vial (20 ml), add by pipette internal standard solution for calibration solution (5 ml) and mix well (Solutions CA and CB).
APPARATUS As for 331/TC/m/3 except:
Constant temperature oven capable of controlling temperature within the range
Rotary evaporator

PROCEDURE As for 331/TC/m/3 except:
(a) Gas chromatographic conditions (typical):
(b) Linearity check. Check the linearity of the detector response by injecting 1 µl of solutions with permethrin concentrations 0.1, 1 and 2.5 times that of the calibration solution before conducting analysis. (c) System equilibration. Prepare two calibration solutions. Inject 1 µl portions of the first one until the response factors obtained for two consecutive injections differ by less than 2.0%. Then inject a 1 µl portion of the second solution. The response factor for this solution should not deviate by more than 2.0% from that for the first calibration solution, otherwise prepare new calibration solutions. (d) Preparation of sample solution. Clean scissors and tweezers with acetone before use. Prepare sample solutions in triplicate for each sample Note 1). Cut ca. 5 cm x 5 cm net samples with the scissors. Weigh accurately, to the nearest 0.1 mg, of each sample (w mg). Transfer it with the tweezers into a vial (20 ml). Add by pipette internal standard solution for sample solution (10 ml). Cap the vial and shake the solution by hand for 1 minute Note 2). Take out the netting with the tweezers and discard the solution. Let the netting dry at room temperature for ca. 10 minutes. Transfer it into a vial (20 ml) with the tweezers. Cap the vial and place it in a temperature-controlled oven set at 70°C. Heat the sample for 2 hours Note 3). After heating, remove the vial from the oven and let it equilibrate to room temperature. Add by pipette internal standard solution for sample solution (10 ml) into the vial. Cap the vial and shake the solution by hand for 1 minute. Take the netting out with the tweezers. Let the netting dry at room temperature for ca. 10 minutes. Transfer the sample solution from the vial into a round-bottom flask. Rinse the vial with acetone (about 1 ml) and transfer it into the same round-bottom flask. Evaporate the solution in vacuo to dryness. Add by pipette acetone (2 ml) into the flask to dissolve the residue (Solutions for "surface concentration at post-wash 1", SA1, SB1 and SC1). Transfer the dried netting into a vial (20 ml) with the tweezers. Repeat heating, internal standard solution adding, evaporating and dissolving procedures as above. (Solutions for "surface concentration at post-wash 2", SA2, SB2 and SC2). Again, let the netting dry at room temperature for ca. 10 minutes. Transfer the dried netting into a vial (20 ml) with the tweezers. Repeat heating, internal standard solution adding, evaporating and dissolving procedures as above. (Solutions for "surface concentration at post-wash 3", SA3, SB3 and SC3). Note 1: Analytical error is larger than average content determinations. Triplicate determinations, therefore, are recommended. Note 2: The shaking speed is about 30 times per 10 seconds. Note 3: Put vials in a covered cardboard box while heating so that the vials are not exposed to direct stream of warm air. (e) Determination. Inject in duplicate 1 µl portions of each sample solution bracketing them by injections of the calibration solutions as follows; calibration solution CA, sample solution SA1, sample solution SA1, calibration solution CB, sample solution SB1, sample solution SB1, calibration solution CA, and so on. Measure the relevant peak areas. (f) Calculation of surface concentration. Calculate the mean value of each pair of response factors bracketing the two injections of a sample and use this value for calculating the surface concentrations of the bracketed sample injections. fi = individual response factor f Hs = total peak area of permethrin (cis-permethrin + trans-permethrin) in Hw = total peak area of permethrin (cis-permethrin + trans-permethrin) in Ir = peak area of the internal standard in the calibration solution Iq = peak area of the internal standard in the sample solution s = mass of permethrin working standard taken (mg) w = mass of sample taken (mg) P = purity of permethrin working standard (g/kg) (g) Calculation of release index. Calculate the mean value of the two injections of sample solutions SA3, SB3 and SC3 by the equations described in (f), and the release index for each piece of the netting. B = mean value of surface concentration at post-wash 2 (µg/g) C = mean value of surface concentration at post-wash 3 (µg/g) Fig. 2 Example of gas chromatogram of Permethrin TC Fig. 3 Example of gas chromatogram of Permethrin LN

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