《美国FDA农残分析手册》（第一卷英文版）Chapter 3 Multiclass MRMS

Pesticide Analytical Manual Vol. I SECTION 301 Experiences with such effects are usually noted in a method so use of particular solvents can be avoided Polarity. Increasing the polarity of an extraction solvent may improve a methods ability to extract particular residues, but it usually also increases the amount of co- extractives. The presence of polar solvents may also affect subsequent cleanup steps, so residues may need to be transferred to a different solvent before the next step of the method is performed Boiling Point. Solvents with a low boiling point are preferred, if evaporation to accommodate detector compatibility or appropriate polarity is necessary. In some temperature if an azeotrope is fi nigh boiling point can be evaporated at a lower cases, a solvent with a relatively h formed by addition of another solvent. Several types of evaporation apparatus exist(Section 202 C), and choice of which to use is often related to the boiling point of a particular solvent Toxicity. Solvents vary in toxicity, and laboratories should choose the least toxic among equivalent choices. Certain solvents(benzene, carbon tetrachloride) should no longer be used in residue analysis Concentration and evaporation steps must be performed in an adequately ventilated hood, and other standard safety precau- tions must be followed (Section 207) Extraction The necessity of using water-miscible solvents to extract pesticide residues from high moisture products has long been established, as has the necessity of a"blend ing type"extraction process [1-4]. Acetone (Section 302), acetonitrile(Section 03), and methanol (Sections 401, 403) are used in PAM I multiclass and selective MRMs to extract nonionic residues from fruits and vegetables. Variations in polar ity may affect the degree to which each can extract any particular residue [5-8] Because extraction capabilities of these solvents are similar, other characteristics affect which solvent a developer chooses to use in a method. For example, devel- tile (Section 303)because it is less toxic, has a lower boiling point(57 C us 82 C), does not affect detectors adversely, and does not form a two-phase system with water during analysis of fruit, as acetonitrile does [9] Liquid-liquid partitioning of residues from initial extractant to nonaqueous sol- vent is a step common to most MRMs. Nature of the solvent(s)used in this step affects the degree of transfer of both residues and co-extractives. For example, in Section 302 El, petroleum ether is included in the separator with aqueous acetone and methylene chloride to reduce the amount of polar plant constituents that partition into the organic phase. However, in a method variation targeted at the highly polar methamidophos, petroleum ether is replaced with acetone to im- prove partitioning of methamidophos from the aqueous to the organic layer [10j Any MRM is applied with the understanding that certain residues are particularly difficult to extract, e. g, the polar residue methamidophos, above. In such cases notation of partial recovery is made in the table(s) of data that accompany the method description. Tentative identification of a residue known to be incom- pletely extracted by the method in use should then lead to re-analysis by another method or variation 301-5

Pesticide Analytical Manual Vol. I SECTION 301 301–5 Transmittal No. 94-1 (1/94) Form FDA 2905a (6/92) Experiences with such effects are usually noted in a method so use of particular solvents can be avoided. Polarity. Increasing the polarity of an extraction solvent may improve a method’s ability to extract particular residues, but it usually also increases the amount of co￾extractives. The presence of polar solvents may also affect subsequent cleanup steps, so residues may need to be transferred to a different solvent before the next step of the method is performed. Boiling Point. Solvents with a low boiling point are preferred, if evaporation to accommodate detector compatibility or appropriate polarity is necessary. In some cases, a solvent with a relatively high boiling point can be evaporated at a lower temperature if an azeotrope is first formed by addition of another solvent. Several types of evaporation apparatus exist (Section 202 C), and choice of which to use is often related to the boiling point of a particular solvent. Toxicity. Solvents vary in toxicity, and laboratories should choose the least toxic among equivalent choices. Certain solvents (benzene, carbon tetrachloride) should no longer be used in residue analysis. Concentration and evaporation steps must be performed in an adequately ventilated hood, and other standard safety precau￾tions must be followed (Section 207). Extraction The necessity of using water-miscible solvents to extract pesticide residues from high moisture products has long been established, as has the necessity of a “blend￾ing type” extraction process [1-4]. Acetone (Section 302), acetonitrile (Section 303), and methanol (Sections 401, 403) are used in PAM I multiclass and selective MRMs to extract nonionic residues from fruits and vegetables. Variations in polar￾ity may affect the degree to which each can extract any particular residue [5-8]. Because extraction capabilities of these solvents are similar, other characteristics affect which solvent a developer chooses to use in a method. For example, devel￾opers of the method in Section 302 used acetone as extractant instead of acetoni￾trile (Section 303) because it is less toxic, has a lower boiling point (57° C vs. 82° C), does not affect detectors adversely, and does not form a two-phase system with water during analysis of fruit, as acetonitrile does [9]. Liquid-liquid partitioning of residues from initial extractant to nonaqueous sol￾vent is a step common to most MRMs. Nature of the solvent(s) used in this step affects the degree of transfer of both residues and co-extractives. For example, in Section 302 E1, petroleum ether is included in the separator with aqueous acetone and methylene chloride to reduce the amount of polar plant constituents that partition into the organic phase. However, in a method variation targeted at the highly polar methamidophos, petroleum ether is replaced with acetone to im￾prove partitioning of methamidophos from the aqueous to the organic layer [10]. Any MRM is applied with the understanding that certain residues are particularly difficult to extract, e.g., the polar residue methamidophos, above. In such cases, notation of partial recovery is made in the table(s) of data that accompany the method description. Tentative identification of a residue known to be incom￾pletely extracted by the method in use should then lead to re-analysis by another method or variation

SECTION 301 Pesticide Analytical Manual Vol I Certain commodities also present greater challenges to the extraction process, and methods may include special steps as an accommodation. Dry products are ex- tracted with combinations of organic solvent and water to make up for the absence of water in the commodity itself. Several studies support the use of water/aceton- trile(Section 303 E3) for this purpose [11-13]. Water/acetone( Section 302 E4)is also used but has been found in some cases to extract less residue than water/ acetonitrile [14, 15 ]; the two methods should be used to check one another when a residue has been identified that can be determined by both methods Extraction of residues from fatty products (e.g, Section 304 El-E5) has tradition ally been aimed at nonpolar, lipophilic residues, which are readily extracted from the product when the fat itself is extracted. Currently, no method is available this manual for quantitative determination of polar residues in fatty products Some residues absorbed from soil by plants, e.g., dieldrin in potatoes, have been shown to be incompletely extracted by methods such as Section 303 [18]; other root-absorbed residues(e.g, dieldrin and DDT in carrots) have been extracted completely by the same procedure [16, 17]. Laboratories analyzing root crops must be aware that the method may not be extracting all the residue present Other, more exhaustive processes, such as use of a Soxhlet extractor [18], may be necessary if the residue or commodity warrants Cleanup Cleanup steps are designed to purify extracts to permit more definitive identifica- tion of residues at lower limits of quantitation, and to minimize adverse effects on determinative step instrumentation. However, almost all cleanup steps adsorb, destroy, or otherwise remove at least some residues from the extract. Thus, cleanup may reduce the number of detectable residues in the final extract. Schemes for multiclass MRMs attempt to determine as many residues as possible by examining uncleaned extracts with selective detectors, e.g., flame photometric and electrolytic conductivity(GLC)and fluorescence(HPLC). Cleanup can subse quently be performed on the extract to permit determination with less selective detectors, e.g., electron capture (GLC) or UV(HPLC). Use of several cleanup steps, each on a separate aliquot of extract, permits examination of each aliquot with a different determinative step. This approach provides coverage for the maxi- mum number of residues, excluding only those not recovered from any cleanup step and also not determined by initial selective detectors Residues can often be detected but not reliably quantitated in an uncleaned ex- tract; itation be possible once the extract is cleaned nique known to recover the particular residue. Other residues can be quantita- tively measured only by re-analysis with a different extraction step. Tables of recov ery data for each method provide the analyst with information to guide the choice of an appropriate cleanup technique or alternative method Many cleanup steps involve chromatography of the extract solution on a column or cartridge. Choices of the column/cartridge material and eluting solvent(s) dictate what chemicals can be recovered; e. g, columns of the adsorbent Florisil provide suitable cleanup of relatively nonpolar residues(Sections 302 Cl, 303 CI etc. ). Increasing the polarity of the eluant permits recovery of more polar residues but decreases the degree of cleanup, because more co-extractives are also eluted Transmittal No 94-1(1/94) Form FDA 2905a(6/92

301–6 Transmittal No. 94-1 (1/94) Form FDA 2905a (6/92) SECTION 301 Pesticide Analytical Manual Vol. I Certain commodities also present greater challenges to the extraction process, and methods may include special steps as an accommodation. Dry products are ex￾tracted with combinations of organic solvent and water to make up for the absence of water in the commodity itself. Several studies support the use of water/acetoni￾trile (Section 303 E3) for this purpose [11-13]. Water/acetone (Section 302 E4) is also used but has been found in some cases to extract less residue than water/ acetonitrile [14, 15]; the two methods should be used to check one another when a residue has been identified that can be determined by both methods. Extraction of residues from fatty products (e.g., Section 304 E1-E5) has tradition￾ally been aimed at nonpolar, lipophilic residues, which are readily extracted from the product when the fat itself is extracted. Currently, no method is available in this manual for quantitative determination of polar residues in fatty products. Some residues absorbed from soil by plants, e.g., dieldrin in potatoes, have been shown to be incompletely extracted by methods such as Section 303 [13]; other root-absorbed residues (e.g., dieldrin and DDT in carrots) have been extracted completely by the same procedure [16, 17]. Laboratories analyzing root crops must be aware that the method may not be extracting all the residue present. Other, more exhaustive processes, such as use of a Soxhlet extractor [18], may be necessary if the residue or commodity warrants. Cleanup Cleanup steps are designed to purify extracts to permit more definitive identifica￾tion of residues at lower limits of quantitation, and to minimize adverse effects on determinative step instrumentation. However, almost all cleanup steps adsorb, destroy, or otherwise remove at least some residues from the extract. Thus, cleanup may reduce the number of detectable residues in the final extract. Schemes for multiclass MRMs attempt to determine as many residues as possible by examining uncleaned extracts with selective detectors, e.g., flame photometric and electrolytic conductivity (GLC) and fluorescence (HPLC). Cleanup can subse￾quently be performed on the extract to permit determination with less selective detectors, e.g., electron capture (GLC) or UV (HPLC). Use of several cleanup steps, each on a separate aliquot of extract, permits examination of each aliquot with a different determinative step. This approach provides coverage for the maxi￾mum number of residues, excluding only those not recovered from any cleanup step and also not determined by initial selective detectors. Residues can often be detected but not reliably quantitated in an uncleaned ex￾tract; quantitation may be possible once the extract is cleaned up using a tech￾nique known to recover the particular residue. Other residues can be quantita￾tively measured only by re-analysis with a different extraction step. Tables of recov￾ery data for each method provide the analyst with information to guide the choice of an appropriate cleanup technique or alternative method. Many cleanup steps involve chromatography of the extract solution on a column or cartridge. Choices of the column/cartridge material and eluting solvent(s) dictate what chemicals can be recovered; e.g., columns of the adsorbent Florisil provide suitable cleanup of relatively nonpolar residues (Sections 302 C1, 303 C1, etc.). Increasing the polarity of the eluant permits recovery of more polar residues but decreases the degree of cleanup, because more co-extractives are also eluted

Pesticide Analytical Manual Vol. I SECTION 301 Very polar residues usually cannot be eluted from Florisil no matter how polar an eluant is used. Instead, charcoal columns are often used for cleaning up extracts containing polar residues, e.g., Sections 302 C2 and C3 The nature of the solvent in which the extract is dissolved when placed on a cleanup column may affect which residues elute from the column. Recovery data associated with a method are valid only when the extract is in the specified solvent When combining method modules, the extract added to a cleanup column may be in a solvent different from that originally specified; in such cases, recovery data may not be applicable. To make use of existing tables of data related to chemicals recovered through a method, it may be necessary to change the extract solvent by evaporation or azeotroping Determinative Steps Use of minimal cleanup in an MRM reduces analysis time and reagent costs, but it can jeopardize determinative step reliability by introducing co-extractives that in- terfere with the determination or cause physical damage to the system. Presence of materials to which the detector responds can cause (1) false reports of residues not actually present, (2)inaccurate quantitation of residues, or(3)complete mask ing of residues. Risk of chromatographic degradation is increased by repetitive injection of an uncleaned extract. The analyst using methods from PAM I is responsible for ensuring that extract injected into any determinative step system does not contain potential interfer- ences or materials that adversely affect chromatographic performance. Sections 501 C and 601 E provide recommendations related to determinations with GlC and HPLC systems, respectively. Analytical accuracy and minimal disruption of laboratory operations will result if reasonable use of cleanup steps and regular maintenance of instruments are both employed In certain cases, special precautions are needed to detect particular residues. For example, thiometon is known to break down while standing in the extract solution of Section 302 El; examination of the extract soon after its preparation perm determination of thiometon residues that would not otherwise be detectable Notes are included in the method tables of data to provide such advice. References [1] Klein, AK(1958) Assoc Of Agric. Chem. 41, 551-555 2] Klein, AK, et al.(1959)/. Assoc. Off. Agric. Chem. 42, 539-544 [8] Burke, J.A., and Porter, M L.(1966)/. Assoc. Off. Anal. Chem. 49, 1157-1162 4] Porter, M.L., and Burke, J.A.(1968)/. Assoc. Off. Anal. Chem. 51, 63-64 [5] Watts,R.R.(1971)/. Assoc. Off. Anal. Chem. 54, 953-958 [6] Wheeler, W.B., et al.(1978)/. Agric. Food Chem. 26, 1833-138 [7] Sonobe, H, et al.(1982)J. Agric. Food Chem. 30, 696-70 301-7

Pesticide Analytical Manual Vol. I SECTION 301 301–7 Transmittal No. 94-1 (1/94) Form FDA 2905a (6/92) Very polar residues usually cannot be eluted from Florisil no matter how polar an eluant is used. Instead, charcoal columns are often used for cleaning up extracts containing polar residues, e.g., Sections 302 C2 and C3. The nature of the solvent in which the extract is dissolved when placed on a cleanup column may affect which residues elute from the column. Recovery data associated with a method are valid only when the extract is in the specified solvent. When combining method modules, the extract added to a cleanup column may be in a solvent different from that originally specified; in such cases, recovery data may not be applicable. To make use of existing tables of data related to chemicals recovered through a method, it may be necessary to change the extract solvent by evaporation or azeotroping. Determinative Steps Use of minimal cleanup in an MRM reduces analysis time and reagent costs, but it can jeopardize determinative step reliability by introducing co-extractives that in￾terfere with the determination or cause physical damage to the system. Presence of materials to which the detector responds can cause (1) false reports of residues not actually present, (2) inaccurate quantitation of residues, or (3) complete mask￾ing of residues. Risk of chromatographic degradation is increased by repetitive injection of an uncleaned extract. The analyst using methods from PAM I is responsible for ensuring that extract injected into any determinative step system does not contain potential interfer￾ences or materials that adversely affect chromatographic performance. Sections 501 C and 601 E provide recommendations related to determinations with GLC and HPLC systems, respectively. Analytical accuracy and minimal disruption of laboratory operations will result if reasonable use of cleanup steps and regular maintenance of instruments are both employed. In certain cases, special precautions are needed to detect particular residues. For example, thiometon is known to break down while standing in the extract solution of Section 302 E1; examination of the extract soon after its preparation permits determination of thiometon residues that would not otherwise be detectable. Notes are included in the method tables of data to provide such advice. References [1] Klein, A.K. (1958) J. Assoc. Off. Agric. Chem. 41, 551-555 [2] Klein, A.K., et al. (1959) J. Assoc. Off. Agric. Chem. 42, 539-544 [3] Burke, J.A., and Porter, M.L. (1966) J. Assoc. Off. Anal. Chem. 49, 1157-1162 [4] Porter, M.L., and Burke, J.A. (1968) J. Assoc. Off. Anal. Chem. 51, 63-64 [5] Watts, R.R. (1971) J. Assoc. Off. Anal. Chem. 54, 953-958 [6] Wheeler, W.B., et al. (1978) J. Agric. Food Chem. 26, 1333-1337 [7] Sonobe, H., et al. (1982) J. Agric. Food Chem. 30, 696-702

SECTION 301 Pesticide Analytical Manual Vol I [8] Sonobe, H, et aL.(1988)/. Agric. Food Chem. 31, 96-104 [9] Luke, M.A., et aL. (1975)/. Assoc Off. Anal. Chem. 58, 1020-1026 [10 Luke, M.A., and Doose, G M.(1988) Bull. Environ. Contam. Toxicol. 30, 110- 16 [11 Bertuzzi, P F, et aL.(1967) .Assoc. Of. Anal. Chem. 50, 629-627 [12] Burke, J.A., and Porter, ML.(1967)/. Assoc Off. Anal. Chem. 50, 1260-1262 [13] Burke, J.A., et al. (1971)/. Assoc. Off. Anal. Chem. 54, 142-146 [14] Cox, B, and Uribe, M.(Nov. 1984)"Comparison of Acetone and Acetoni tile for Extraction of Endrin from Mung Beans, " LIB 2881, FDA, Rockville [15] Weishaar, J-A.(Nov. 1989)"Comparison of Acetone-Water and Acetonitrile- Water for Extraction of Pirimiphos Methyl from Pasta Products, "LIB 3397, FDA. Rockville MD [16] Porter, M.L., et al. (Dec. 1970)FDA Private communication [17] Luke, MA(Feb. 1971)FDA private communication [18 Mumma, RO, et al.(1966)Science 152, 530-531 Transmittal No 94-1(1/94) Form FDA 2905a(6/92

301–8 Transmittal No. 94-1 (1/94) Form FDA 2905a (6/92) SECTION 301 Pesticide Analytical Manual Vol. I [8] Sonobe, H., et al. (1983) J. Agric. Food Chem. 31, 96-104 [9] Luke, M.A., et al. (1975) J. Assoc. Off. Anal. Chem. 58, 1020-1026 [10] Luke, M.A., and Doose, G.M. (1983) Bull. Environ. Contam. Toxicol. 30, 110- 116 [11] Bertuzzi, P.F., et al. (1967) J. Assoc. Off. Anal. Chem. 50, 623-627 [12] Burke, J.A., and Porter, M.L. (1967) J. Assoc. Off. Anal. Chem. 50, 1260-1262 [13] Burke, J.A., et al. (1971) J. Assoc. Off. Anal. Chem. 54, 142-146 [14] Cox, B., and Uribe, M. (Nov. 1984) “Comparison of Acetone and Acetoni￾trile for Extraction of Endrin from Mung Beans,” LIB 2881, FDA, Rockville, MD [15] Weishaar, J.A. (Nov. 1989) “Comparison of Acetone-Water and Acetonitrile￾Water for Extraction of Pirimiphos Methyl from Pasta Products,” LIB 3397, FDA, Rockville, MD [16] Porter, M.L., et al. (Dec. 1970) FDA private communication [17] Luke, M.A. (Feb. 1971) FDA private communication [18] Mumma, R.O., et al. (1966) Science 152, 530-531

Pesticide Analytical Manual Vol. I SECTION 302 302: METHODI FOR NONFATTY FOODS BASIC REFERENCES Luke, M.A., et al.(1975)/. Assoc. Of. Anal. Chem. 58, 1020-1026 Luke, M.A., et al.(1981)/ Assoc. Of. Anal. Chem. 64, 1187-1195 GENERAL PRINCIPLES Residues are extracted from nonfatty foods by blending with acetone or water/ acetone, then transferred from the filtered aqueous extractinto organic solvent. The extract is cleaned up if necessary and examined by various determinative steps; the amount of cleanup necessary is dictated by the determinative step(s)to be used and by the type of commodity being analyzed APPLICABILITY Consult Guide to PAM I for additional information pertinent to the appropriate application of multiresidue methodology Method is applicable to nonionic residues in nonfatty foods. Cleanup steps may be needed for particularly dirty extracts or for examination by less selective detectors; some residues may be lost during cleanup Extract is amenable to examination by many determinative steps, and the residues covered by a particular analysis are dependent on the number of different determinative steps used. See Tables 302-a and 302-b, following the method description, for results of recovery tests METHOD MODULES Choose from these method modules, using Figure 302-a for guidance Recommended use El(P. 302-7) Extraction with acetone, liquid-liquid nonfatty, high moisture partitioning with petroleum ether/ commodities methylene chloride 2(P. 302-9) Extraction with acetone, removal of nonfatty, high moisture water with 40 g Hydromatrix (p 302-11) Extraction with acetone, removal of alternative to E2 for water with 25 g Hydromatrix luction in solvent use E4 (p. 302-13) Extraction with water/acetone, nonfatty, low moisture liquid-liquid partitioning wit commodities petroleum ether/methylene chloride E5(P. 302-15) Extraction with acetone alternative to el liquid-liquid partitioning with for relatively polar acetone/methylene chloride E6 (P. 302-16) Extraction with water/acetone alternative to e4 for liquid-liquid partitioning wit acetone/methylene chloride eques E7 (P. 302-17) Extraction with acetone and nonfatty, high moisture solid phase extraction cartridges commodities for liquid-liquid partitioning relatively polar residues

Transmittal No. 2000-1 (10/1999) Form FDA 2905a (6/92) 302–1 Pesticide Analytical Manual Vol. I SECTION 302 302: METHOD I FOR NONFATTY FOODS BASIC REFERENCES Luke, M.A., et al. (1975) J. Assoc. Off. Anal. Chem. 58, 1020-1026 Luke, M.A., et al. (1981) J. Assoc. Off. Anal. Chem. 64, 1187-1195 GENERAL PRINCIPLES Residues are extracted from nonfatty foods by blending with acetone or water/ acetone, then transferred from the filtered aqueous extract into organic solvent. The extract is cleaned up if necessary and examined by various determinative steps; the amount of cleanup necessary is dictated by the determinative step(s) to be used and by the type of commodity being analyzed. APPLICABILITY Consult Guide to PAM I for additional information pertinent to the appropriate application of multiresidue methodology. Method is applicable to nonionic residues in nonfatty foods. Cleanup steps may be needed for particularly dirty extracts or for examination by less selective detectors; some residues may be lost during cleanup. Extract is amenable to examination by many determinative steps, and the residues covered by a particular analysis are dependent on the number of different determinative steps used. See Tables 302-a and 302-b, following the method description, for results of recovery tests. METHOD MODULES Choose from these method modules, using Figure 302-a for guidance: Extraction (E) Recommended Use E1 (p. 302-7) Extraction with acetone, liquid-liquid nonfatty, high moisture partitioning with petroleum ether/ commodities methylene chloride E2 (p. 302-9) Extraction with acetone, removal of nonfatty, high moisture water with 40 g Hydromatrix commodities E3 (p. 302-11) Extraction with acetone, removal of alternative to E2 for water with 25 g Hydromatrix reduction in solvent use E4 (p. 302-13) Extraction with water/acetone, nonfatty, low moisture liquid-liquid partitioning with commodities petroleum ether/methylene chloride E5 (p. 302-15) Extraction with acetone, alternative to E1 liquid-liquid partitioning with for relatively polar acetone/methylene chloride residues E6 (p. 302-16) Extraction with water/acetone, alternative to E4 for liquid-liquid partitioning with relatively polar acetone/methylene chloride residues E7 (p. 302-17) Extraction with acetone and nonfatty, high moisture solid phase extraction cartridges, commodities for liquid-liquid partitioning relatively polar residues