Flow Chart for Cypermethrin from Formulation Sample

Page No. 1/6 FC-PF-248 PESTICIDE FORMULATION & RESIDUE ANALYTICAL CENTRE, PMD, NIPHM, HYDERABAD

Sr. No. in Scope NABL / NON NABL Flow Chart for Analysis of Azadirachtin Content in formulation sample

Date of Analysis

SI. Execution Executed Step No. By 1. Sample No. 2. Name of Sample 2.1 Sample Description

3. Procedure R1 R2 3.1 Preparation of mobile phase: 3.1.1 Mix Acetonitrile and water in the proportion of 35:65 (v/v), pass through membrane filter under vacuum and degas. 3.1.2 Homogenize the mixture using the magnetic stirrer 3.2 Preparation of Make up Phase : 3.2.1 Mix methanol and water in the proportion of 90:10 (v/v), pass through membrane filter under vacuum and degas. 3.2.2 Homogenize the mixture using the magnetic stirrer 3.3 Preparation of Reference Standard Solution: 3.3.1 Note the purity of the standard % % Name of the Laboratory : Pesticide Formulation & Residue Analytical Centre, PMD, NIPHM, Hyderabad Document No. : FC-PF-248 Document Name : Flow Chart for analysis of Azadirachtin content, % by mass Revision No. : 02 Issue Date : 10/09/2013 Revision Date : 26/03/2014 Next Revision Date : 26/03/2016 Prepared By Checked By Approved & Issued By

Mrs. C. Vijaya Lakshmi (Assistant Plant Protection Officer)

Mr. C.V. Rao (Technical Manager)

Dr Abhay Ekbote (Director PM & Quality Manager) Page No. 2/6 FC-PF-248 PESTICIDE FORMULATION & RESIDUE ANALYTICAL CENTRE, PMD, NIPHM, HYDERABAD

3.3.2 Weigh accurately 2 mg a.i. of standard Azadirachtin into a g g 50 ml volumetric flask. 3.3.3 Note the serial number of the balance logbook. 3.3.4 Dissolve and make up to the mark with make up phase (3.2.2). Shake well to homogenize. 3.3.5 Transfer 2 mL of the above solution with the help of pipette into pre wetted (with 3.2.2) RP C18 micro column. 3.3.6 Elute Azadirachtin out of the column by repeatedly washing the column with the make up phase (3.2.2). Collect the elute and washings into 10 ml volumetric flask till the level reaches up to the mark. 3.4 Preparation of sample solution 3.4.1 Note the percent active ingredient content declared on the % % sample. 3.4.2 Weigh accurately sample equivalent to 2 mg a.i. of g g Azadirachtin into 50 ml volumetric flask. 3.4.3 Note the serial number of the balance logbook. 3.4.4 Dissolve and make up to the mark with make up phase (3.2.2). Shake well to homogenize. 3.4.5 Transfer 2 mL of the above solution with the help of pipette into pre wetted (with 3.2.2) RP C18 micro column. 3.4.6 Elute Azadirachtin out of the column by repeatedly washing the column with make up phase (3.2.2.). Collect elute and washings into 10 ml volumetric flask till the Name of the Laboratory : Pesticide Formulation & Residue Analytical Centre, PMD, NIPHM, Hyderabad Document No. : FC-PF-248 Document Name : Flow Chart for analysis of Azadirachtin content, % by mass Revision No. : 02 Issue Date : 10/09/2013 Revision Date : 26/03/2014 Next Revision Date : 26/03/2016 Prepared By Checked By Approved & Issued By

level reaches up to the mark. 3.4.7 Note the serial number of HPLC logbook. 4. HPLC Parameters: 4.1 Detector : UV 4.2 Wave length : 215 nm 4.3 Column Specification: Stainless steel packed with C18 Length x I.D.: 250mm X 4.6mm, 5µ particle size 4.4. Mobile phase: Aetonitrile : Water (35:65)(v/v) Flow rate : 1ml/min 4.5 Injection volume: 20 µl 5. Result: 5.1 Sample chromatogram no 5.2 Standard chromatogram no

6. Calculation:

A1 X M1 Where,

Azadirachtin content, ------X P M1 = mass in ‘g’ of Azadirachtin reference standard

% by mass A2 X M2 solution.

M2 = mass in ‘g’ of sample taken for the test.

A1 = peak area of Azadirachtin in the sample solution.

A2 = peak area of Azadirachtin in the reference Name of the Laboratory : Pesticide Formulation & Residue Analytical Centre, PMD, NIPHM, Hyderabad Document No. : FC-PF-248 Document Name : Flow Chart for analysis of Azadirachtin content, % by mass Revision No. : 02 Issue Date : 10/09/2013 Revision Date : 26/03/2014 Next Revision Date : 26/03/2016 Prepared By Checked By Approved & Issued By

standard solution. P = percent purity Azadirachtin standard. Replicate 1 –

Name of the Laboratory : Pesticide Formulation & Residue Analytical Centre, PMD, NIPHM, Hyderabad Document No. : FC-PF-248 Document Name : Flow Chart for analysis of Azadirachtin content, % by mass Revision No. : 02 Issue Date : 10/09/2013 Revision Date : 26/03/2014 Next Revision Date : 26/03/2016 Prepared By Checked By Approved & Issued By

Replicate 2 –

Result: Sl. No. Name of Test Result Unit Method of Analysis IS 14300 : 1995 1. Active ingredient (Azadirachtin) % (Reaffirmed 2009) Reference /Remarks:

Name Analyzed by Dated signature

Name Checked by Dated signature

Dr Abhay Ekbote (Director PM & Quality Manager)