TECHNICAL ALPHACYPERMETHRIN

Transcription

1 ALPHACYPERMETHRIN 1. Specification 1.1 Description Full specification: Adopted 10 December 1999 The material shall consist of a homogeneous mixture of technical alphacypermethrin together with related manufacturing impurities and shall be in the form of a white to cream crystalline powder, free from visible extraneous matter and added modifying agents. 1.2 Chemical and physical requirements The material sampled from any part of the consignment (see method WHO/M/1.R1), shall comply with the requirements of section 1.1 and with the following requirements Identity test Where the identity of the active ingredient is in doubt, then it shall comply with at least one additional test. (CIPAC method 454/TC/(M)/2, CIPAC Handbook H, p.15) Alphacypermethrin cis 2 ([IR cis] S and [IS cis] R isomers) content (g/kg basis) The alphacypermethrin cis 2 ([IR cis] S and [IS cis] R isomers) content shall be declared (not less than 910 g/kg), and when determined by the method described in section 2.1 the mean measured content obtained shall not be lower than the declared content Total pyrethroid content. The combined content of the cis and trans isomers of alpha-cyano-3-phenoxybenzyl- 2,2-dimethyl-3-(2,2-dichlorovinyl-)cyclopropanecarboxylate, when determined by the method described in section 2.2, shall not be less than 975 g/kg Volatile hydrocarbon solvent content The maximum volatile hydrocarbon solvent content of the material, when determined by the method described in section 2.3 shall not be higher than 1g/kg. 1

2 1.2.5 Triethylamine content The maximum triethylamine content of the material, when determined by the method described in section 2.4 shall not be higher than 1 g/kg. 1.3 Packing and marking of packages The technical alphacypermethrin shall be packed in suitable clean containers, as specified in the order: All packages shall bear, durably and legibly marked on the container, the following: Manufacturer s name Technical alphacypermethrin Batch or reference number, and date of test Net weight of contents Date of manufacture and the following cautionary notice: Alphacypermethrin is a pyrethroid that acts predominantly on the central nervous system; high dosages have been found to cause tonic seizures in experimental animals. A high concentration in air may be an irritant and contact with the concentrated product may induce a temporary tingling sensation, particularly on the face. It may be hazardous if swallowed. Do not inhale spray mist. Avoid skin contact; wear protective gloves, clean protective clothing, and a face mask (surgical type) when handling the product. Wash hands and exposed skin thoroughly after using. Keep containers out of reach of children and well away from foodstuffs and animal feed and their containers. Alphacypermethrin is toxic to aquatic wildlife. Avoid accidental contamination of water. If poisoning occurs, call a physician. Treatment is symptomatic. 2. Methods of determining chemical and physical properties 2.1 Alphacypermethrin content Outline of method The sample is dissolved in tetrahydrofuran with di-octyl phthalate used as the internal standard. Separation is carried out in the split injection mode using a dimethyl polysiloxane WCOT fused silica capillary column with automatic injection and flame ionisation detector. The alphacypermethrin is determined by comparison with calibration solutions Apparatus Gas chromatograph. Capable of operating over the range 100 to 300 C with a flame ionisation detector, split/splitless injector and autosampler. 2

3 Column. 30 m WCOT fused silica 0.25 mm id and 0.25 µm film thickness coated with dimethyl polysiloxane (chemically bonded), or equivalent. Injector system. Injector split/splitless with fused silica liner containing a 1 cm plug of glass wool. An automatic injector is strongly recommended to ensure the best analytical results. It is important to ensure that the split liner is acid treated, thoroughly deactivated and conditioned before use, to ensure that alphacypermethrin does not epimerise during analysis. Split ratio :1 (split vent at 75 ml/min) Injection Volume 1.0 µl using a autosampler Detector system. Type FID, with no special requirements. Automatic digital integrator or chromatography data system. Compatible with the gas chromatograph. 100 ml volumetric flasks Reagents Tetrahydrofuran, analytical grade. Citric acid. 5% citric acid solution. Dissolve citric acid (25 g) in water (500 ml). Di-octyl phthalate, di-(2-ethylhexyl)phthalate, Internal standard. Select for use a batch which, when chromatographed under the conditions below for the determination of alphacypermethrin gives no peak with a similar retention time to the alphacypermethrin isomers. Cypermethrin analytical standard of known purity (containing a mixture of trans 3, trans 4, cis 1 and cis 2 cypermethrin). Alphacypermethrin working standard, of known alphacypermethrin content Preparation of standard solutions Internal standard solution Dissolve di-octyl phthalate (5.0 g) in tetrahydrofuran (500 ml) (solution I). Ensure sufficient quantity of this solution is prepared for all samples and calibration standards being analysed. The same internal standard solution must be used for standards and samples in the same analytical run. Citric acid. 5% citric acid solution. 3

4 Dissolve citric acid (25 g) in water (500 ml). Alphacypermethrin calibration solution Weigh in duplicate (to the nearest 0.1 mg) about 0.1 g of alphacypermethrin (C A and C B, g) into 100 ml volumetric flasks. Add to each, 70 ml of tetrahydrofuran and agitate the flask gently to dissolve. Add by pipette 10.0 ml of internal standard solution (solution I) and 10 ml of 5% citric acid solution from a measuring cylinder. Make to volume with tetrahydrofuran and mix well. (Solutions C A and C B ). Note: Citric acid is added to stop epimerisation of alphacypermethrin in solution. Prepare a solution without internal standard, by dissolving about 0.1 g of alphacypermethrin standard into a 100 ml volumetric flask. Add approximately 70 ml of tetrahydrofuran and swirl to dissolve. Add by pipette 10 ml of 5% citric acid solution from a measuring cylinder. Make to volume with tetrahydrofuran and mix well. (Solution C O ). Cypermethrin calibration check solution Prepare a solution of cypermethrin, by dissolving about 0.1 g of cypermethrin standard into a 100 ml volumetric flask. Add approximately 70 ml of tetrahydrofuran and swirl to dissolve. Add by pipette 10mL of the internal standard solution to volume with tetrahydrofuran and mix well (Solution C C ) Operating conditions The conditions given below are typical values and may have to be adjusted to obtain optimum results from a given apparatus. Temperatures Column oven 230 C Injector 260 C Detector 300 C Adjust the column oven temperature if required to obtain retention time windows for di-octyl (approximately 13.5 minutes), alphacypermethrin cis 2 isomer (approximately 27.0 minutes) and alphacypermethrin cis 1 isomer (approximately 25.2 minutes). The trans 3 and trans 4 isomers elute at approximately 26.1 and 27.5 minutes respectively. Gas flow rates Helium carrier approximately 0.8 ml min -1 Helium make up gas 60 ml min -1 or optimum for instrument 4

5 Septum purge 2 ml min -1 Hydrogen as recommended for the instrument Air as recommended for the instrument All gases should be purified through molecular sieves. The carrier gas should be further purified through an oxygen trap Sample preparation Sampling Homogenize the bulk material before taking the sample. Take at least 25 g. Rehomogenize the bulk before taking a sub-sample for analysis. Preparation of the sample solutions. Weigh in duplicate (to the nearest 0.1 mg) into 100 ml volumetric flasks sufficient sample (w, g) to contain 0.1 g of alphacypermethrin. Add approximately 70 ml of tetrahydrofuran and agitate the flask gently to dissolve. Add by pipette to each flask 10 ml of di-octyl phthalate solution I and by measuring cylinder, 10 ml of 5% citric acid solution. Make to volume with tetrahydrofuran and mix well (Solutions S A and S B ). Prepare a solution without internal standard, by dissolving about 0.1 g of alphacypermethrin standard into a 100 ml volumetric flask. Add approximately 70 ml of tetrahydrofuran and swirl to dissolve. Add by pipette 10 ml of 5% citric acid solution from a measuring cylinder. Make to volume with tetrahydrofuran and mix well. (Solution S O ) Equilibration of the system. Carry out alternate 1.0 µl injections of solutions I, C o and S o and check whether there are any interfering peaks from impurities. If there are make any necessary corrections but do not use external calibration. Inject calibration solutions C A and C B to equilibrate the system and use the data from these chromatograms to set the integration parameters. Calculate the response factors for these injections to check the stability of the instrument. Response factors should not differ by more than ± 1% of the mean. Inject the solution of cypermethrin in tetrahydrofuran to ensure complete separation of the alphacypermethrin isomers Analysis of sample 5

6 Carry out 1.0 µl injections of calibration solutions C A and C B and sample solutions S A and S B in the following sequence and record the integrated areas of the peaks. Injection sequence C A1, S A1, S A2, C B1, C A2, S B1, S B2, C B Calculation Calculate the relative response factors (f 1, f 2 etc) for the pair of calibration injections which bracket the sample injections eg use C A1 and C B1 for sample injections S A1, S A2 etc and obtain the mean response factor, f. Sample analysis should be repeated if calibration response factors f 1 and f 2 differ by more than ± 2% of the mean f. Relative response factor = H s I r x s x P Where: H s = area of alphacypermethrin cis 2 peak in the calibration solution. I r = area of di-octyl phthalate peak in the calibration solution. s = mass of alphacypermethrin analytical standard in calibration solution (g). P = purity of the alphacypermethrin standard (g/kg). For each sample injection eg S A1, calculate the alphacypermethrin cis 2 content. Alphacypermethrin content = Hw f x Iq x w Where: f = mean relative response factor Hw = mean area of alphacypermethrin cis 2 peak in the sample solution Iq = area of di-octyl phthalate peak in the sample solution w = mass of sample (g) Calculate the alphacypermethrin cis 2 content of the sample as the mean of the four determinations as follows: Sample injection Use relative response factor from Alphacypermethrin cis 2 (g/kg) S A1 C A1 and C B1 Q ) ) X S A2 C A1 and C B1 R ) S B1 C A2 and C B2 S ) ) Y S B2 C A2 and C B2 T ) 6

7 Q and R, S and T should agree to within ± 0.5% of their mean values (X and Y). X and Y should agree within ± 1% of their mean values. Take the mean of the two values x and y as the alphacypermethrin content. 2.2 Total pyrethroid content Outline of method The sample is dissolved in a chloroform/heptane mixture containing 0.2% tetrahydrofuran. Separation is carried out by normal phase high performance liquid chromatography with ultra violet detection. The total pyrethroid content is determined by comparison with calibration solutions Apparatus Liquid chromatograph equipped with a 10 µl loop injector. UV detector capable of operating at 278 nm. Column stainless steel, 250 x 4.6 mm (i.d.) Zorbax SIL normal phase column, or equivalent. Guard column 15 x 3.2 mm (i.d.) 7 µm silica cartridge or equivalent. Injector system capable of reproducible 10 µl injections. Detector system UV, with no special requirements. Automatic digital integrator or chromatography data system compatible with the liquid chromatograph 50 ml, 100 ml volumetric flasks Reagents Cis and trans cypermethrin Analytical standard of known purity Heptane HPLC grade Chloroform HPLC grade (containing 1% ethanol as a preservative) Tetrahydrofuran (THF) ACS reagent (contains butylated hydroxytoluene, BHT, as a preservative) Mobile phase Heptane and chloroform (9 to 1 volume ratio) with 0.2% (v/v) tetrahydrofuran. Measure heptane (2250 ml) and chloroform (250 ml) using a graduated cylinder and add by pipette THF (5 ml) Preparation of standard solutions Cis and trans cypermethrin calibration solution 7

8 Weigh in duplicate (to the nearest 0.1 mg) about 0.05 g each of cis and trans cypermethrin (C C and C D, g) into 100 ml volumetric flasks. Add to each, 5 ml of chloroform to dissolve and dilute to volume with heptane and mix well. (Solutions C C and C D ) Operating Conditions The conditions given below are typical values and may have to be adjusted to obtain optimum results from a given apparatus. Eluting solvent flow rate 2 ml/min Temperature 35 C Injection volume 10 µl Wavelength 278 nm Retention times Cis 1 cypermethrin minutes Cis 2 cypermethrin minutes Trans 3 cypermethrin minutes Trans 4 cypermethrin minutes Sample preparation Sampling Homogenise the bulk material before taking the sample. Take at least 25 g. Rehomogenise the bulk before taking a sub-sample for analysis. Preparation of the sample solutions. Weigh in duplicate (to the nearest 0.1 mg) into 50 ml volumetric flasks sufficient sample (w, g) to contain 0.1 g of technical grade cypermethrin. Add approximately 5 ml of chloroform to dissolve the sample and dilute with heptane. (Solutions S C and S D ) Equilibration of the system Inject calibration solutions C C and C D to equilibrate the system and use the data from these chromatograms to set the integration parameters. Calculate the response factors for these injections to check the stability of the instrument. Response factors should not differ by more than ± 1% of the mean Analysis of sample Carry out 10 µl injections of calibration solutions C C and C D and sample solutions S C and S D in the following sequence and record the integrated areas of the peaks. Injection sequence C C1, S C1, S C2, C D1, C C2, S D1, S D2, C D2 8

9 2.2.9 Calculation Calculate the relative response factors (f 1, f 2 etc.) for the pair of calibration injections which bracket the sample injections e.g. use C C1 and C D1 for sample injections S C1, S C2 etc. and obtain the mean response factor, f. Sample analysis should be repeated if calibration response factors f 3 and f 4 differ by more than ± 2% of the mean f. Relative response factor f = H s s x P Where: H s = area of the four cypermethrin peaks in the calibration solution s = mass of the cypermethrin analytical standard in calibration solution (g) P = purity of the cypermethrin standard (g/kg) For each sample injection eg S C1, calculate the cypermethrin isomer content Cypermethrin isomer content= Hw f x w Where: f = mean relative response factor Hw = mean area of the four cypermethrin isomer peaks in the sample solution w = mass of sample (g) 2.3 Volatile hydrocarbon solvent content Outline of method The volatile hydrocarbon solvent content is determined by measuring the difference in the weight of the sample before and after drying in a vacuum oven Apparatus Vacuum oven capable of maintaining a temperature of C with a vacuum of about 2430" Hg. Desiccator: Note it is recommended to allow the sample to reach room temperature in a desiccator, after removal from the drying oven, to minimise adsorption of atmospheric moisture. Petri Dishes Pyrex, 100 x 15 mm 9

10 2.3.3 Reagents Desiccant e.g. Indicating Drierite Absorbent, or equivalent Procedure In duplicate, accurately weigh Petri dishes (x g). Accurately weigh 5-10 g of the technical material into the Petri dish. (y g). Place the sample in the oven set at a temperature of about 40 C and adjust the vacuum to about 2430" Hg. Dry the sample for about 2.5 hours. Remove the sample from the oven and allow it cool in the desiccator and reweigh (z g). Dry the sample for an additional 30 minutes and redetermine the weight. If the weight is constant, i.e., the relative change between two weighings is less than 0.5%, calculate the percentage volatile hydrocarbon content. Otherwise, continue drying the sample to constant weight. Percentage ( w/w ) volatile hydrocarbon content = 100 (y-z) y-x 2.4 Triethylamine content Outline of method This method is used to determine the content of triethylamine ( TEA ) in alphacypermethrin technical material. The sample is dissolved in a mixture of glacial acetic acid and chlorobenzene and potentiometrically titrated with standard acetous perchloric acid. The amount of TEA is calculated from the equivalence point of the titration curve Apparatus Potentiometric Titrator e.g. Mettler DL25 titrator equipped with a combination glass electrode ( Mettler DG 112), or equivalent. An Epson FX-85 or other printer to record the results. Titrator cups 100 ml to fit the titrator. 5 ml, 100 ml Volumetric pipettes (Class A) 1L and 100 ml Volumetric flasks (Class A) Reagents Acetous perchloric acid. 1 M solution of standard perchloric acid in glacial acetic acid Glacial acetic acid. Reagent grade Potassium hydrogen phthalate A.C.S. Primary standard ( %) Chlorobenzene HPLC Grade 10

11 Acetous perchloric acid 0.01 M. Pipette 100 ml of the 0.1M acetous perchloric acid into a 1L volumetric flask. Dilute to mark with glacial acetic acid. Solution Accurately weigh 204 mg of potassium hydrogen phthalate into a 100 ml volumetric flask. Record the weight to the nearest 0.01 mg. Dissolve in glacial acetic acid Procedure Blank determination (Repeat in triplicate) Add 10mL of chlorobenzene to a titrator and fill to 100 ml with glacial acetic acid. Determine the equivalence point and record the volume at this point. Average the recorded volumes together and use this as the blank value. Standardization of 0.01M Perchloric (Repeat in triplicate) Pipette 5 ml of the potassium hydrogen phthalate solution into a titrator cup. Add 10 ml of chlorobenzene and fill to 100 ml with glacial acetic acid. Attach vessel to titrator, and determine the equivalence point and record the volume at this point. Determine the concentration of the perchloric solution using the calculation found in section Average the results. Analysis of samples All weighings should be accurate to within 0.1 mg. Accurately weigh 3 to 4 g of technical material into the titrator vessel. Record the weight to the nearest 0.1 mg. Add 10 ml of chlorobenzene and fill to 100 ml with glacial acetic acid. Attach the vessel to titrator, and turn on stirrer to dissolve sample. Determine the equivalence point and record the volume at this point Calculation The concentration of the potassium hydrogen phthalate standard solution is calculated by the following equation: where: C 1 = m x V 1 C 1 = concentration of potassium hydrogen phthalate (mmoles/ml) m 1 = mass of potassium hydrogen phthalate (mg) V 1 = volume of potassium hydrogen phthalate solution (ml) 11

12 The concentration of the perchloric solution is determined with the following equation: C2 = C 1 V 1 V 3 - V 4 where: C 2 = concentration of standard perchloric solution (mmoles/ml) C 1 = concentration of potassium hydrogen phthalate solution (mmoles/ml) V 2 = volume of standard potassium hydrogen phthalate solution (ml) V 3 = volume of perchloric to reach equivalence point (ml) V 4 = volume average blank value (ml) The weight percent of triethylamine is determined with the following equation: weight % triethylamine = where: (V 5 -V 4 ) x C 2 x x 100 m 2 V 5 = volume of perchloric used to reach equivalence point (ml) V 4 = volume of average blank value (ml) C 2 = concentration of perchloric solution (mmol/ml) m 2 = mass of alphacypermethrin in sample (mg) 12

13 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 ALPHACYPERMETHRIN AQUEOUS SUSPENSION CONCENTRATE 1. Specification 1.1 Description Full specification: WHO/SIF/61 Adopted 10 December 1999 The material consists of a suspension of fine particles of technical alphacypermethrin, complying with the requirements of the WHO specification, in an aqueous phase together with suitable formulants; After gentle agitation the material shall be homogeneous and suitable for further dilution in water. 1.2 Chemical and physical requirements The material sampled from any part of the consignment (see method WHO/M/1.R1), shall comply with the requirements of section 1.1 and with the following requirements Alphacypermethrin cis 2 ([IR cis] S and [IS cis] R isomers) content (g/kg basis) The content of alphacypermethrin cis 2 ([IR cis] S and [IS cis] R isomers) content (g/kg basis), determined by the method described in section 2.1 shall not differ from the declared content by more than the following amount: Declared content Up to 25 g/kg Above 25 up to 100 g/kg Tolerance permitted ± 15% of the declared content ± 10% of the declared content Higher declared contents are not currently available. The average content of all samples taken shall not be lower than the declared content Alphacypermethrin cis 1 ([IR cis] R and [IS cis] S isomer) content The alphacypermethrin cis 1:cis 2 isomer ratio determined by the method described in section , shall not be higher than 5: ph range The ph of the concentrate, determined by the CIPAC method MT 75 (CIPAC Handbook F, p.205), shall be between 7.0 and

14 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/ Rinsability The maximum non volatile rinsed residue as determined by CIPAC MT (in press) 1 shall not exceed 0.05% Spontaneity of dispersion at 30 C A minimum of 60% of the alphacypermethrin content found under shall be in suspension after 5 min in WHO standard hard water C at 30 ± 2 C as determined by CIPAC method MT 160 (CIPAC Handbook F, p.391) Suspensibility after heat stability treatment In WHO standard hard water. When tested by the method described in section 2.2, a minimum of 70% of the alphacypermethrin content (0.63 g/l) shall be in suspension 30 minutes after agitating a suspension containing (0.9 g/l) of alphacypermethrin prepared in standard hard water at 30 ± 2 C from the suspension concentrate subjected to the heat stability treatment described in section Wet sieve test Not less than 98% of the concentrate shall pass through a 75 µm sieve when tested by the CIPAC method MT 59 (CIPAC Handbook F, p.177) Persistent foam The persistent foam at the top of a 100 ml of suspension prepared in WHO standard soft water, shall not exceed 20 ml after 12 minutes when tested by the CIPAC method described MT 47.2 (CIPAC Handbook F, p.152) Heat stability The suspension after treatment as described in section 2.3 must comply with the requirements of sections 1.2.1, 1.2.2, 1.2.3, 1.2.4, 1.2.5, and of this specification Cold stability After storage testing at 0 ± 1 C for 7 days, according to the CIPAC method MT 39.1 (CIPAC Handbook F, p.128), the product shall continue to comply with 1.2.5, and Packing and marking of packages The alphacypermethrin suspension concentrate shall be packed in suitable clean drums as specified in the order. The shall be made of high density polyethylene or any suitable materials compatible with the product.. 1 Method can be obtained on request from CIPAC secretariat, t Gotink 7, 7261 VE Ruurlo, Netherlands. 14

15 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 All packages shall bear, durably and legibly marked on the container, the following: Manufacturer s name Alphacypermethrin suspension concentrate Alphacypermethrin...g/l Batch or reference number, and date of test Net weight of contents Date of manufacture Instruction for use and the following minimum cautionary notice: Alphacypermethrin is a pyrethroid that acts predominantly on the central nervous system; high dosages have been found to cause tonic seizures in experimental animals. A high concentration in air may be an irritant and contact with the concentrated product may induce a temporary tingling sensation, particularly on the face. It may be hazardous if swallowed. Do not inhale spray mist. Avoid skin contact; wear protective gloves, clean protective clothing, and a face mask (surgical type) when handling the product. Wash hands and exposed skin thoroughly after using. Keep containers out of reach of children and well away from foodstuffs and animal feed and their containers. Alphacypermethrin is toxic to aquatic wildlife. Avoid accidental contamination of water. If poisoning occurs, call a physician. Treatment is symptomatic. 2 Methods of determining chemical and physical properties 2.1 Alphacypermethrin content Outline of method The sample is extracted in tetrahydrofuran with di-octyl phthalate used as the internal standard. Separation is carried out in the split injection mode using a dimethyl polysiloxane WCOT fused silica capillary column with automatic injection and flame ionisation detector. The alphacypermethrin is determined by comparison with calibration solutions Apparatus Gas chromatograph. Capable of operating over the range 100 to 300 C with a flame ionisation detector, split/splitless injector and autosampler. Column. 30 m WCOT fused silica 0.25 mm id and 0.25 µm film thickness coated with dimethyl polysiloxane (chemically bonded), or equivalent. Injector system. Injector Split/splitless with fused silica liner containing a 1 cm plug of glass wool. An automatic injector is strongly recommended to ensure the best analytical results. 15

16 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 It is important to ensure that the split liner is acid treated, thoroughly deactivated and conditioned before use, to ensure that alphacypermethrin does not epimerise during analysis. Split ratio Injection Volume :1 (split vent at 75 ml/min) 1.0 µl using a autosampler. Detector system. Type FID, with no special requirements. Automatic digital integrator or chromatography data system. Compatible with the gas chromatograph 100 ml volumetric flasks Reagents Tetrahydrofuran. Citric acid. 5% citric acid solution. Di-octyl phthalate, di-(2-ethylhexyl)phthalate, Internal standard. Select for use a batch which, when chromatographed under the conditions below for the determination of alphacypermethrin gives no peak with a similar retention time to the alphacypermethrin isomers. Cypermethrin analytical standard (containing a mixture of trans 3, trans 4, cis 1 and cis 2 cypermethrin) of known purity Alphacypermethrin working standard, of known alphacypermethrin content Preparation of standard solutions Internal standard solution. Dissolve di-octyl phthalate (5.0 g) in tetrahydrofuran (500 ml) (solution I). Ensure sufficient quantity of this solution is prepared for all samples and calibration standards being analysed. Citric acid. 5% citric acid solution. Dissolve citric acid (25 g) in water (500 ml). Alphacypermethrin calibration solution. Weigh in duplicate (to the nearest 0.1 mg) about 0.1 g of alphacypermethrin (C A and C B, g) into 100 ml volumetric flasks. Add to each, 70 ml of tetrahydrofuran and agitate the flask gently to dissolve. Add by pipette 10.0 ml of internal standard solution (solution I) and 10mL of 5% citric acid solution from a measuring cylinder. Make to volume with tetrahydrofuran and mix well. (Solutions C A and C B ). Note: Citric acid is added to stop epimerisation of alphacypermethrin in solution. 16

17 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 Prepare a solution without internal standard, by dissolving about 0.1 g of alphacypermethrin standard into a 100 ml volumetric flask. Add approximately 70 ml of tetrahydrofuran and swirl to dissolve. Add by pipette 10 ml of 5% citric acid solution from a measuring cylinder. Make to volume with tetrahydrofuran and mix well. (Solution C O ). Cypermethrin calibration check solution Prepare a solution of cypermethrin, by dissolving about 0.1 g of cypermethrin standard into a 100 ml volumetric flask. Add approximately 70 ml of tetrahydrofuran and swirl to dissolve. Add by pipette 10 ml of the internal standard solution to volume with tetrahydrofuran and mix well (Solution C C ) Operating conditions The conditions given below are typical values and may have to be adjusted to obtain optimum results from a given apparatus. Temperatures Column oven 230 C Injector 260 C Detector 300 C Adjust the column oven temperature if required to obtain retention time windows for di-octyl (approximately 13.5 minutes), alphacypermethrin cis 2 isomer (approximately 27.0 minutes) and alphacypermethrin cis 1 isomer (approximately 25.2 minutes). The trans 3 and trans 4 isomers elute at approximately 26.1 and 27.5 minutes respectively. Gas flow rates Helium carrier approximately 0.8mL min -1 Helium make up gas 60mL min -1 or optimum for instrument Septum purge 2mL min -1 Hydrogen as recommended for the instrument Air as recommended for the instrument All gases should be purified through molecular sieves. The carrier gas should be further purified through an oxygen trap Sample Preparation Sampling Homogenize the bulk material before taking the sample. Take at least 250 g. Rehomogenize the bulk before taking a sub-sample for analysis. Preparation of the sample solutions. 17

18 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 Weigh accurately in duplicate (to the nearest 0.1mg) into 100 ml volumetric flasks sufficient sample (w,g) to contain 0.1 g of alphacypermethrin. Add by measuring cylinder 10 ml of the 5% citric acid solution to each of the flasks and swirl to fully disperse the formulation. Add approximately 70 ml of tetrahydrofuran in approximately 10 to 15 ml portions, swirling in between each addition to fully disperse the sample. Place in an ultrasonic bath for 15 minutes with occasional swirling. Add by pipette 10 ml of the di-octyl phthalate, internal standard solution I and make to volume with tetrahydrofuran and mix well. Filter through a suitable paper eg GF/A or filter unit eg PTFE 0.45 µm to give clear solutions. (Solutions S A and S B ). Prepare a solution without internal standard, by dissolving sufficient sample to contain 0.1 g of alphacypermethrin into a 100 ml volumetric flask. Add by measuring cylinder 10 ml of the 5% citric acid solution to each of the flasks and swirl to fully disperse the formulation. Add approximately 70 ml of tetrahydrofuran in approximately 10 to 15 ml portions, swirling in between each addition to fully disperse the sample. Place in an ultrasonic bath for 15 minutes with occasional swirling. Make to volume with tetrahydrofuran and mix well. Filter through a suitable paper eg GF/A or filter unit eg PTFE 0.45 µm to give a clear solution. (Solution S O ) Equilibration of the system. Carry out alternate 1.0 µl injections of solutions I, C o and S o and check whether there are any interfering peaks from impurities. If there are make any necessary corrections but do not use external calibration. Inject calibration solutions C A and C B to equilibrate the system and use the data from these chromatograms to set the integration parameters. Calculate the response factors for these injections to check the stability of the instrument. Response factors should not differ by more than ± 1% of the mean. Inject the solution of cypermethrin in tetrahydrofuran (C c ) to ensure complete separation of the alphacypermethrin isomers Analysis of sample Carry out 1.0 µl injections of calibration solutions C A and C B and sample solutions S A and S B in the following sequence and record the integrated areas of the peaks. Injection sequence C A1, S A1, S A2, C B1, C A2, S B1, S B2, C B Calculation. Calculate the relative response factors (f 1, f 2 etc) for the pair of calibration injections which bracket the sample injections eg use C A1 and C B1 for sample injections S A1, S A2 etc and obtain the mean response factor, f. Sample analysis should be repeated if calibration response factors f 1 and f 2 differ by more than ± 2% of the mean f. Relative response factor = H s I r x s x P 18

19 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 where: H s = area of alphacypermethrin cis 2 peak in the calibration solution. I r = area of di-octyl phthalate peak in the calibration solution. s = mass of alphacypermethrin analytical standard in calibration solution (g). P = purity of the alphacypermethrin standard (g/kg). For each sample injection eg S A1, calculate the alphacypermethrin cis 2 content. Alphacypermethrin content = Hw f x Iq x w where: f = mean relative response factor Hw = mean area of alphacypermethrin cis 2 peak in the sample solution Iq = area of di-octyl phthalate peak in the sample solution w = mass of sample (g) Calculate the alphacypermethrin cis 2 content of the sample as the mean of the four determinations as follows: Sample injection Use relative response factor from Alphacypermethrin cis 2 (g/kg) S A1 C A1 and C B1 Q ) ) X S A2 C A1 and C B1 R ) S B1 C A2 and C B2 S ) ) Y S B2 C A2 and C B2 T ) Q and R, S and T should agree to within ± 0.5% of their mean values (X and Y). X and Y should agree within ± 1% of their mean values. Take the mean of the two values x and y as the alphacypermethrin content Determination of the Cis 1 to Cis 2 ratio. To determine the cis 1 to cis 2 percentage ratio in a sample Peak area cis 1 x 100 cis 1 % = Total peak area of cis 1 and cis 2 19

20 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 Peak area cis 2 x 100 cis 2 % = Total peak area of cis 1 and cis Suspensibility Outline of method A suspension of known concentration of alphacypermethrin in standard hard water is prepared, poured into a 250 ml graduated cylinder, maintained at constant temperature and allowed to remain undisturbed for 30 minutes. The top 9/10ths are drawn off and the content of alphacypermethrin in the bottom 1/10th is determined, so allowing to evaluate the active ingredient mass still in suspension after 30 minutes Apparatus A 250 m1 graduated cylinder with a ground-glass stopper and a distance of cm between the bottom and the 250 ml calibration mark. A glass tube, about 40 cm long and about 5 mm in internal diameter, pointed at one end to an opening of 2-3 mm, the other end being connected to a suitable source of suction Reagents WHO standard hard water. See method WHO/M/ Procedure Weigh (to the nearest 1 mg) into 100 ml beaker an amount of the sample to form 250 m1 of a suspension containing 0.9 g/l of alphacypermethrin. Add a volume of water at 30 ± 2 C equal to at least twice the mass of the sample taken. Allow to stand for 30 seconds and then stir by hand for 30 seconds with a glass rod 4-6 mm in diameter, at not more than four revolutions per second, making no deliberate attempt to break up any lumps. The immediately transfer the mixture quantitatively to the 250 ml graduated cylinder, using water at 30 ± 2 C for rinsing, and again avoiding mechanical disintegration of lumps. Immediately add sufficient water at 30 ± 2 C to bring the volume up to the 250 ml mark. Insert the stopper and invert the cylinder end over end 30 times at the rate of one complete cycle every 2 seconds. This operation should be carried out as smoothly as possible, keeping the axis of rotation fixed. Allow the graduated cylinder to stand for 30 minutes in a water-bath at 30 ± 2 C taking care that the bath is free from vibrations. Should excessive flocculation occur during the test, the material is unsatisfactory. At the end of the 30 minutes settling period, insert the glass tube into the cylinder and, with a minimum of disturbance, withdraw nine-tenths, of the suspension (i.e. 225 ml) 20

21 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 by means of the suction tube in a period of seconds. This is achieved by maintaining the tip of the glass tube just below the sinking surface of the suspension. Discard the suspension withdrawn Determination of alphacypermethrin in the retained one-tenth of the suspension Quantitatively transfer the bottom one-tenth of suspension (25 ml) from the suspensibility test into a 100 ml volumetric flask with approximately 50 ml of tetrahydrofuran. Add 10 ml internal standard by pipette and 5 ml citric acid solution by measuring cylinder. Dilute to volume with tetrahydrofuran. Place in an ultrasonic bath for 10 minutes. Filter through a suitable disc filter. Determine the alphacypermethrin content of the filtered solution by gas chromatography as described in section injecting duplicate 1.0 µl aliquots of sample and calibration solutions Calculation Mass of alphacypermethrin (g) in the bottom one-tenth of suspension = f 2 x P x w f 1 x 2 x 103 where: f 1 = mean response ratio for the calibration solution f 2 = mean response ratio for the sample solution w = mass (g) of alphacypermethrin standard in the calibration solution P = purity (g/kg) of alphacypermethrin standard Suspensibility % = 10 (b-a) x 100 = 111(b-a) 9 b b where: a = mass (g) of alphacypermethrin in bottom one-tenth of suspension b = mass (g) of alphacypermethrin in the sample used in the suspensibility test (calculated from the mass of sample and its percentage alphacypermethrin content). 2.3 Heat stability treatment 54 C ± 2 C for 14 days (CIPAC method MT 46.1, CIPAC Handbook F, p.149), unless other temperatures and times are requested (FAO Manual on the development and use of FAO specifications for plant protection products, n 149, p.33). 21

22 AQUEOUS SUSPENSION CONCENTRATE WHO/SIF/61 After completion of the heat stability treatment, the samples should not be exposed to heat, bright sunshine, or atmospheric humidity. If required the test should be conducted in commercial type pack. 22