Pesticide Analytical Manual Vol. I APPENDⅨX‖/ PROTOCOL E PROTOCOL B: PROCEDURE FOR TESTING CHEMICALS THROUGH SECTION 402 BACKGROUND Method: Section 402 Chemical Type: Applicable to chemicals with acid or phenol structure. Chemicals are methylated before determination by glC, because esters and ethers are more easily chromatographed than acids and phenols Commodity Type: Applicable to wide variety of commodities, both fatty and nonfatty Different extraction steps are used depending on commodity PAM I Tables: Table 402-a DATA DEVELOPMENT Gas Chromatography Because the method being tested includes methylation of the analyte, glC chan acteristics of both the acid/ phenol and its methylation product are collected Perform the following operations Dissolve methyl ester/ether reference standard, if available, in 10% acetone/isooctane(v/v) to prepare stock standard solution. Dilute wit in Protocol C for chromatography of methyl ester/ ether reference standard. Report results on Reporting Form C Stop work if methyl ester/ether does not cause response on any detector when chromatographed on any appropriate column Dissolve acid/phenol reference standard in acetone to prepare stock standard solution. Dilute as needed with acetone. Also dilute with 50% methylene chloride/hexane to prepare solution suitable for testing re- covery through gel permeation chromatography(GPC).(Note: 2, 4,5-T will not dissolve in 50% methylene chloride/hexane, so solutions in that solvent mixture must be prepared by diluting acetone stock solution. Follow directions in Protocol C for chromatography of acid/phenol ref erence standard. Report results on separate copy of Reporting Form C Methylate 100 ug acid/phenol reference standard in acetone solution according to procedure described in Section 402 CIb Dilute as needed with hexane. If any other methylation procedure is used, describe pro- cedure on Reporting Form B Follow directions in Protocol C for chromatography of methylated ref- erence standard Report results on separate copy of Reporting Form C (Note: if ester/ether reference standard is available, these results should Form FDA 2905a(6/92) ppendix -9Pesticide Analytical Manual Vol. I APPENDIX II/PROTOCOL B Appendix II–9 Transmittal No. 94-1 (1/94) Form FDA 2905a (6/92) PROTOCOL B: PROCEDURE FOR TESTING CHEMICALS THROUGH SECTION 402 BACKGROUND Method: Section 402 Chemical Type: Applicable to chemicals with acid or phenol structure. Chemicals are methylated before determination by GLC, because esters and ethers are more easily chromatographed than acids and phenols. Commodity Type: Applicable to wide variety of commodities, both fatty and nonfatty. Different extraction steps are used depending on commodity. PAM I Tables: Table 402-a DATA DEVELOPMENT Gas Chromatography Because the method being tested includes methylation of the analyte, GLC char￾acteristics of both the acid/phenol and its methylation product are collected. Perform the following operations: • Dissolve methyl ester/ether reference standard, if available, in 10% acetone/isooctane (v/v) to prepare stock standard solution. Dilute with isooctane. • Follow directions in Protocol C for chromatography of methyl ester/ ether reference standard. Report results on Reporting Form C. • Stop work if methyl ester/ether does not cause response on any detector when chromatographed on any appropriate column. • Dissolve acid/phenol reference standard in acetone to prepare stock standard solution. Dilute as needed with acetone. Also dilute with 50% methylene chloride/hexane to prepare solution suitable for testing re￾covery through gel permeation chromatography (GPC). (Note: 2,4,5-T will not dissolve in 50% methylene chloride/hexane, so solutions in that solvent mixture must be prepared by diluting acetone stock solution.) • Follow directions in Protocol C for chromatography of acid/phenol ref￾erence standard. Report results on separate copy of Reporting Form C. • Methylate 100 µg acid/phenol reference standard in acetone solution according to procedure described in Section 402 C1b. Dilute as needed with hexane. If any other methylation procedure is used, describe pro￾cedure on Reporting Form B. • Follow directions in Protocol C for chromatography of methylated ref￾erence standard. Report results on separate copy of Reporting Form C (Note: if ester/ether reference standard is available, these results should