Test Material: Cyproconazole
MRID: 49863306
Cyproconazole - Analytical Method GRM033.01 A for the
yjtle. Determination of Cyproconazole, 1,2,4-Triazole and Triazole Acetic
Acid in Soil Using Liquid Chromatography-Electrospray Ionization
Tandem Mass Spectrometry (Including Validation Data) - Method
MRID: 49863305
Cyproconazole - Cyproconazole - Independent Laboratory Validation of
the Analytical Method GRM033.01A, Determination of Cyproconazole,
Title: 1,2,4-Triazole and Triazole Acetic Acid in Soil Using Liquid
Chromatography-Electrospray Ionization Tandem Mass Spectrometry -
Final Report
EPA PC Code: 128993
OCSPP Guideline: 850 6100
For CDM/CSS-Dynamac JV
Primary Reviewer: Lisa Muto Signature
Date: 10/13/16
Secondary Reviewer: Kathleen Ferguson Signature:
Date: 10/13/16
Quality Assurance Manager: Joan Gaidos Signature:
Date: 10/13/16
This Data Evaluation Record may have been altered by the Environmental Fate and Effects
Division subsequent to signing by CDM/CSS-Dynamac JV personnel.
-------
Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
Analytical method for cyproconazole, and its metabolites, 1,2,4-triazole and triazole acetic
acid, in soil
Reports: ECM: EPA MRID No.: 49863306. Huang, S-B. 2007. Cyproconazole -
Analytical Method GRM033.01 A for the Determination of Cyproconazole,
1,2,4-Triazole and Triazole Acetic Acid in Soil Using Liquid
Chromatography-Electrospray Ionization Tandem Mass Spectrometry
(Including Validation Data) - Method. Syngenta Report and Task No.
T004701-06. Report prepared, sponsored and submitted by Syngenta Crop
Protection, LLC, Greensboro, North Carolina; 41 pages. Final report issued
August 7, 2007.
ILV: EPA MRID No. 49863305. Thomas, C.A. 2009. Cyproconazole-
Cyproconazole - Independent Laboratory Validation of the Analytical
Method GRM033.01A, Determination of Cyproconazole, 1,2,4-Triazole and
Triazole Acetic Acid in Soil Using Liquid Chromatography-Electrospray
Ionization Tandem Mass Spectrometry - Final Report. Syngenta Report and
Task No. T004702-06. NCL Study No.: 110.029. Report prepared by North
Coast Laboratories, Ltd. (NCL), Areata, California, sponsored and submitted
by Syngenta Crop Protection, LLC, Greensboro, North Carolina; 155 pages.
Final report issued February 5, 2009.
Document No.: MRIDs 49863306 & 49863305
Guideline: 850.6100
Statements: ECM: The study was conducted in accordance with the USEPA Good
Laboratory Practice (GLP) standards (40 CFRPart 160; p. 3 of MRID
49863306). Signed and dated No Data Confidentiality, Quality Assurance
and GLP statements were provided (pp. 2-4). A certification of authenticity
was not included.
Classification:
PC Code:
Reviewer:
ILV: The study was conducted in accordance with the USEPA FIFRA GLP
standards (40 CFR Part 160), except for a few instances of late data entries,
which have been noted in the study report (p. 3 of MRID 49863305). Signed
and dated No Data Confidentiality, GLP and Quality Assurance statements
were provided (pp. 2-4). A certification of authenticity was not included.
This analytical method is classified as Acceptable. In the ECM, no samples
were fortified at lOxLOQ. The purities of the test materials were not
reported in the ECM. ^
128993
^ 14'48'18-04 00
Jerrett Fowler, Physical Scientist Signature: 1/
Date: 9/24/2018
2018.09.24
Stephen P. Wente, Senior Scientist
STEPHEN
Signature: WENTE
Date: 9/24/2018
Digitally signed by STEPHEN
WENTE
Date: 2018.09.2414:16:36
-04W
Page 2 of 13
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
Executive Summary
This analytical method, Syngenta Residue Method GRM033.01 A, is designed for the
quantitative determination of cyproconazole and its metabolites, 1,2,4-triazole and triazole acetic
acid, in soil using LC/MS/MS. The method is quantitative for all three analytes at the stated LOQ
of 1.0 ng/g (1.0 ppb). The LOQ is less than the lowest toxicological level of concern in soil for
all three analytes. The ECM validated the method using sandy loam soil. The ILV validated the
method using sandy loam soil with the first trial with minor modifications to the analytical
method; however, the ILV study author requested optimization of a few steps of the method. The
sources for the soil matrices of the ECM and ILV appeared to be the same. One ion transition
was monitored in the ECM and ILV; a confirmatory method is not usually required when
GC/MS or LC/MS is the primary identification method. In the ECM, no samples were fortified at
lOxLOQ, and the purities of the test materials were not reported. In the ECM and ILV, the
interferences were <50% of the LOD; sample recoveries were only corrected in the ECM.
Table 1. Analytical Method Summary
Analyte(s) by
Pesticide1
MRID
EPA
Review
Matrix
Method Date
(dd/mm/yyyy)
Registrant
Analysis
Limit of
Quantitation
(LOQ)
Environmental
Chemistry
Method
Independent
Laboratory
Validation
Cyproconazole
1,2,4-Triazole
Triazole acetic
acid
49863306
49863305
Soil2,3
07/08/2007
Syngenta
Crop
Protection,
LLC
LC/MS/MS
1.0 ng/g
(1.0 ppb)
1 Cyproconazole = (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-l-(lH-l,2,4-triazol-l-yl)butan-2-ol; 2-
(4-Chloro-phenyl)-3 -cyclopropyl-1 -[ 1,2,4]triazol-1 -yl-butan-2-ol.
2 In the ECM, the sandy loam soil matrix (47% sand, 30% silt, 23% clay, pH 7.5, percent organic matter 3.4) was
collected from Iowa obtained from an on-going Syngenta field dissipation study (USDA soil textural
classification; p. 20 of MRID 49863306). The soil characterization was performed at Agvise Laboratories located
in Northwood, North Dakota.
3 In the ILV, the sandy loam soil matrix (54% sand, 26% silt, 20% clay, pH 7.3, percent organic matter 2.6) was
collected from Greene County, Iowa, and obtained from Syngenta (USDA soil textural classification; Sample ID
RIEN00707-0002; p. 16 of MRID 49863305). The soil characterization was performed at Agvise Laboratories
located in Northwood, North Dakota. The soil sample appeared to be sourced from the same bulk control soil
sample which was used in the ECM.
Page 3 of 13
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Cvproconazole (PC 128993)
MRIDs 49863306 / 49863305
I. Principle of the Method
The method contained the following precautions due to the low detection limit of the method: 1)
new plastic-ware/glassware should be used for each batch; 2) each solvent should be checked to
verify that it is free from contamination (if contamination is suspected); and 3) existing
glassware should be solvent rinsed, after washing and before use in the method (p. 11 of MRID
49863306).
Samples of soil (10 ± 0.1 g) were transferred to 50-mL disposable plastic centrifuge tubes and
fortified, as necessary (pp. 12-13; Appendix 5, p. 41 of MRID 49863306). After five minutes of
equilibration with the fortification solution, the samples were extracted twice with 25 mL of
methanol:water (80:20, v:v) via shaking on a mechanical shaker at a unspecified speed (a speed
which provided visible agitation) for a minimum of 20 minutes; samples should be in a flat or
horizontal orientation. After centrifugation at 5000 rpm for five minutes, the supernatant was
decanted and collected in a 250-mL flat bottom flask via a Whatman 2V filter paper lined funnel.
After the second extraction supernatant was collected, the filter paper was rinsed with 10 mL of
methanol which was combined with the supernatants. The combined supernatants and rinse were
concentrated to aqueous (ca. 3 mL) under vacuum with a rotary evaporator at a bath temperature
of 35°C. The residue was transferred to a clean 15-mL plastic centrifuge tube using de-ionized or
HPLC grade water to rinse the concentration flask. The volume of the residue was adjusted to 10
mL using de-ionized or HPLC grade water. The method noted that this point was a method
stopping point. The analytes were isolated from the soil extract via two different solid phase
extraction (SPE) procedures. During the SPE clean-ups, the cartridges were to remain moist, the
solvents were to be added after the prior solvent had completely entered cartridge, and the flow
rate should be less than 20 drops per minute, with the aid of vacuum if necessary.
To isolate triazole acetic acid (TAA), the soil extract was purified via Waters Oasis MAX SPE
cartridge (150 mg/6-mL, 60 jam; the method noted that no substitution was allowed; p. 13;
Appendix 1, p. 25; Appendix 5, p. 41 of MRID 49863306). The SPE column was pre-
conditioned with methanol (one full cartridge amount x 1), 2% formic acid in methanol (5 mL x
1), de-ionized or HPLC grade water (5 mL x 2), 1% ammonium hydroxide in methanol (freshly
prepared, 5 mL x 2), 1% ammonium hydroxide in water (freshly prepared, 3 mL x 2) and 1%
ammonium hydroxide in water (400 |iL). After 2.0-mL of the soil extract was loaded onto the
column by gravity, the cartridge was washed with de-ionized or HPLC grade water (2 mL x 2),
de-ionized or HPLC grade water (5 mL x 1), methanol:water (50:50, v:v; 5 mL x 1), 1%
ammonium hydroxide in methanol (freshly prepared, 2 mL x 3) and methanol (2 mL x 1). The
analyte was eluted with 2% formic acid in methanol (2 mL x 4) into a clean 15-mL plastic
centrifuge tube. The solvent was evaporated to dryness under a gentle stream of nitrogen at a
bath temperature of 40°C. The residue was reconstituted to a final volume of 2 mL (or higher
volumes for higher concentrations) with methanol:water (5:95, v:v). After vortex mixing, the
sample was analyzed via LC/MS/MS.
To isolate cyproconazole (CCZ) and 1,2,4-triazole (T), the soil extract was purified via Varian
Bond Elut Certify SPE cartridge (300 mg/3-mL; the method noted that no substitution was
allowed; pp. 13-14; Appendix 1, p. 25; Appendix 5, p. 41 of MRID 49863306). The SPE column
was pre-conditioned with methanol (one full cartridge amount x 2), 0.5% ammonium hydroxide
Page 4 of 13
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
in methanol:water (90:10, v:v; freshly prepared, 2 mL x 2), de-ionized or HPLC grade water (2
mL x 2), 5% formic acid in methanol (2 mL x 2) and 2% formic acid in methanol (2 mL x 1).
After 2.0-mL of the soil extract was loaded onto the column by gravity, the cartridge was washed
with de-ionized or HPLC grade water (2 mL x 2) and methanol (2 mL x 2). The analyte was
eluted with 0.5% ammonium hydroxide in methanol:water (90:10, v:v; freshly prepared, 2 mL x
3) into a clean 15-mL plastic centrifuge tube. The solvent was evaporated to aqueous (< 0.6 mL)
under a gentle stream of nitrogen at a bath temperature of 40°C. The residue was reconstituted to
a final volume of 2 mL (or higher volumes for higher concentrations) with methanol:water (5:95,
v:v). After vortex mixing, the sample was analyzed via LC/MS/MS.
Samples were analyzed for cyproconazole (CCZ), 1,2,4-triazole (T) and triazole acetic acid
(TAA) using a Thermo Electron Surveyor Plus LC (pp. 15-17 of MRID 49863306). The
following LC conditions were used: Zorbax SB-Aq column (75 x 4.6 mm, 3.5 |im; column
temperature 25°C), ColumnSaver column filter, mobile phase of (A) 0.1% formic acid in HPLC
grade water and (B) 0.1% formic acid in HPLC grade methanol [TAA: percent A:B (v:v) at 0.0-
5.0 min. 98:2; CCZ and T: percent A:B (v:v) at 0.0-2.0 min. 98:2, 3.0-8.0 min. 10:90, 8.1-11.0
min. 98:2], and injection volume of 50 |iL. The following MS/MS conditions were used: ESI
negative ion polarity and multiple reaction monitoring (MRM) for TAA and ESI positive ion
polarity and multiple reaction monitoring (MRM) for CCZ and T. One ion pair transition was
monitored for each analyte: m/z 126.1 —~ 82.2 for TAA, m/z 70.1 —~ 43.2 for T and m/z 292.1 —~
125.0 for CCZ. Expected retention times were ca. 2.8, 2.6 and 6.0 minutes for TAA, T and CCZ,
respectively.
In the ILV, the method was performed as written (pp. 15, 18-20; Appendix 4, pp. 109-153 of
MTRD 49863305). An Applied Biosciences/MDS Sciex API 4000 LC/MS/MS triple quadrupole
mass spectrometer was used for all analyses. The following analytical parameters were used in
the ILV: an Agilent ZORBAX SB-Aq Rapid Resolution column (4.6 x 75 mm, 3.5 |im; column
temperature unreported), Phenomenex MAX-RP "Security Guard" cartridge guard column,
mobile phase of (A) 0.1% formic acid in water and (B) 0.1% formic acid in methanol [TAA:
percent A:B (v:v) at 3.51 min. 98:2, 3.60 min. 2:98, 6.70 min. 98:2; CCZ and T: percent A:B
(v:v) at 2.00 min. 98:2, 2.50 min. 10:90, 8.11 min. 98:2], and injection volume of 60 |iL. One ion
pair transition was monitored for each analyte: m/z 125.8 —~ 82.2 for TAA, m/z 70.1 —~ 43.1 for
T and m/z 292.0 —~ 125.1 for CCZ. Expected retention times were ca. 2.56, 3.00 and 4.47
minutes for TAA, T and CCZ, respectively.
The Limit of Quantification (LOQ) for cyproconazole, 1,2,4-triazole and triazole acetic acid was
reported as 1.0 ng/g (1.0 ppb) in the ECM and the ILV (pp. 8, 21-22; Figure 2, p. 33 of MRID
49863306; pp. 12, 17, 29 of MRID 49863305). The Limit of Detection (LOD) was reported as 25
pg on-column based on a 50-|iL LC injection volume in the ECM, which was equivalent to the
0.5 ppb calibration standard. The LOD was not reported in the ILV, but appeared to be 0.5 ppb
based on the data in the recovery tables.
Page 5 of 13
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
II. Recovery Findings
ECM (MRID 49863306): Mean recoveries and relative standard deviations (RSD) were within
guideline requirements (mean 70-120%; RSD <20%) for analysis of cyproconazole, 1,2,4-
triazole and triazole acetic acid at the LOQ (1.0 ppb, 1.0 ng/g), 50>
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
Table 2. Initial Validation Method Recoveries for Cyproconazole, 1,2,4-Triazole and
Triazole Acetic Acid in Soil
Analyte
Fortification
Level (ppb)
Number
of Tests
Recovery
Range (%)
Mean
Recovery (%)
Standard
Deviation (%)
Relative Standard
Deviation (%)
Sandy Loam Soil1'2
Cyproconazole3
1.0 (LOQ)
5
99.0-106
102
3.0
3.0
50
5
103-108
105
2.2
2.1
100
5
99.6-102
101
0.9
0.89
1,2,4-Triazole
1.0 (LOQ)
3
84.5-90.7
86.7
2.6
3.0
50
5
78.9-82.7
80.5
1.7
2.1
100
5
77.6-83.0
80.3
2.1
2.6
Triazole acetic acid
1.0 (LOQ)
3
81.6-91.8
86.3
4.6
5.3
50
3
79.6-81.9
80.7
0.9
1.1
100
5
77.9-82.8
79.7
1.8
2.3
Data (recovery results were corrected when residues were quantified in the controls; pp. 18-19) were obtained from
p. 21; Appendix 3, Tables 1-4, pp. 28-31 of MRID 49863306.
1 One ion pair transition was monitored for eachanalyte: m/z 126.1 —> 82.2 for triazole acetic acid, mz 70.1 —>43.2
for 1,2,4-triazole and m/z 292.1 —> 125.0 for cyproconazole (pp. 15-17).
2 The sandy loam soil matrix (47% sand, 30% silt, 23% clay, pH 7.5, percent organic matter 3.4) was collected from
Iowa obtained from an on-going Syngenta field dissipation study (USDA soil textural classification; p. 20). The
soil characterization was performed at Agvise Laboratories located in Northwood, North Dakota.
3 Cyproconazole = (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-l-(lH-l,2,4-triazol-l-yl)butan-2-ol; 2-
(4-Chloro-phenyl)-3 -cyclopropyl-1 -[ 1,2,4]triazol-1 -yl-butan-2-ol.
Table 3. Independent Validation Method Recoveries for Cyproconazole, 1,2,4-Triazole and
Triazole Acetic Acid in Soil
Analyte
Fortification
Number
Recovery
Mean
Standard
Relative Standard
Level (ppb)
of Tests
Range (%)
Recovery (%)
Deviation (%)
Deviation (%)
Surface Soil1'2
Cyproconazole3
1.0 (LOQ)
5
78.3-85.6
82.3
2.6
3.2
10
5
79.3-88.5
83.6
3.5
4.2
1,2,4-Triazole
1.0 (LOQ)
5
96.3-106
101
3.5
3.5
10
5
82.0-91.0
86.4
3.4
3.9
Triazole acetic acid
1.0 (LOQ)
5
97.7-108
104
4.8
4.6
10
5
86.4-95.1
90.2
3.8
4.2
Data (uncorrected recovery results, pp. 26-27) were obtained from Table 1, pp. 31-32 of MRID 49863305.
1 One ion pair transition was monitored for eachanalyte: m/z 125.8 —> 82.2 for triazole acetic acid, m/z 70.1 —>43.1
for 1,2,4-triazole and/w/z 292.0 —> 125.1 for cyproconazole (pp. 18-20).
2 The sandy loam soil matrix (54% sand, 26% silt, 20% clay, pH 7.3, percent organic matter 2.6) was collected from
Greene County, Iowa, and obtained from Syngenta (USDA soil textural classification; Sample ID RIEN00707-
0002; p. 16). The soil characterization was performed at Agvise Laboratories located in Northwood, North
Dakota. The soil sample appeared to be sourced from the same bulk control soil sample which was used in the
ECM.
3 Cyproconazole = (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-l-(lH-l,2,4-triazol-l-yl)butan-2-ol; 2-
(4-Chloro-phenyl)-3 -cyclopropyl-1 -[ 1,2,4]triazol-1 -yl-butan-2-ol.
Page 7 of 13
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Cvproconazole (PC 128993)
MRIDs 49863306 / 49863305
III. Method Characteristics
The LOQ for cyproconazole, 1,2,4-triazole and triazole acetic acid was reported as 1.0 ng/g (1.0
ppb) in the ECM and the ILV (pp. 8, 21-22; Figure 2, p. 33 of MRID 49863306; pp. 12, 17, 29;
Table 1, pp. 31-32 of MRID 49863305). In the ECM, the LOQ was defined as the lowest analyte
concentration in a sample at which the methodology has been validated, i.e. which yielded a
mean recovery of 70-120% and relative standard deviation of <20%. No LOQ calculations were
provided in the ECM. No justifications of the LOQ were provided in the ILV. The LOD was
reported as 25 pg on-column based on a 50-|iL LC injection volume in the ECM, which was
equivalent to the 0.5 ppb calibration standard. The LOD was not reported in the ILV, but
appeared to be 0.5 ppb based on the data in the recovery tables. In the ECM, the LOD was
defined as the smallest standard amount injected during the chromatographic run and typically
corresponds to an amount of analyte equivalent to ca. one-half of the theoretical amount for a
recovery sample at the method LOQ. The ECM study author noted that the LOD may vary from
instrument to instrument depending on the injection volume and concentrations needed to obtain
adequate analyte response.
Page 8 of 13
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
Table 4. Method Characteristics
Analyte
Cyproconazole1
1,2,4-Triazole
Triazole acetic acid
Limit of Quantitation (LOQ)
1.0 ng/g (1.0 ppb)
Limit of Detection (LOD)
ECM
0.5 ppb
(25 pg on-column based on a 50-|iL LC injection volume)
ILV
0.5 ppb2
Linearity (calibration curve r2 and
ECM
r2 = 0.9996
r2 = 0.9994
r2 = 0.9997
concentration range)
(0.5-60 pg/(iL)
ILV3
r2 = 0.9998
r2= 1.0000
r2= 1.0000
(0.5-20 ng/g)
Repeatable
ECM4
Yes at LOQ, 50/LOQ and lOOxLOQ, but no samples were
prepared at lOxLOQ.
ILV5
Yes at LOQ and lOxLOQ.
Reproducible
Yes at LOQ and lOxLOQ.
Specific
ECM
Yes, no matrix
Yes. Matrix
Yes, no matrix
interferences were
interferences were
interferences were
observed.
<10% of the LOQ.
observed.
ILV
Yes, matrix
Yes. No matrix
interferences were
<5% of the LOQ.
Yes, matrix
interferences were
12-14% of the LOQ.6
interferences were
13-15% of the LOQ.7
Non-ideal peak
shapes were
observed at the LOQ.
Data were obtained from pp. 8, 21-22; Appendix 3, Tables 1-4, pp. 28-31 (recovery results); Appendix 4, Figures 1-
6, pp. 32-37 (chromatograms); Appendix 4, Figures 7-9, pp. 38-40 (calibration curves) of MRID 49863306; pp. 12,
17, 29; Table 1, pp. 31-32 (recovery results); Figures 1-45, pp. 35-79 (calibration curves and chromatograms) of
MRID 49863305; DER Attachment 2.
1 Cyproconazole = (2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-l-(lH-l,2,4-triazol-l-yl)butan-2-ol; 2-
(4-Chloro-phenyl)-3 -cyclopropyl-1 -[ 1,2,4]triazol-1 -yl-butan-2-ol.
2 The LOD was not reported in the ILV, but appeared to be 0.5 ppb based on the data in the recovery tables (pp. 12,
17, 29; Table 1, pp. 31-32 of MRID 49863305).
3 Reported correlation coefficients were reviewer-calculated from r values reported in the study report (Figure 1, p.
35; Figure 16, p. 50; Figure 31, p. 65 of MRID 49863305; see DER Attachment 2).
4 In the ECM, the sandy loam soil matrix (47% sand, 30% silt, 23% clay, pH 7.5, percent organic matter 3.4) was
collected from Iowa obtained from an on-going Syngenta field dissipation study (USD A soil textural
classification; p. 20 of MRID 49863306). The soil characterization was performed at Agvise Laboratories located
in Northwood, North Dakota.
5 In the ILV, the sandy loam soil matrix (54% sand, 26% silt, 20% clay, pH 7.3, percent organic matter 2.6) was
collected from Greene County, Iowa, and obtained from Syngenta (USDA soil textural classification; Sample ID
RIEN00707-0002; p. 16 of MRID 49863305). The soil characterization was performed at Agvise Laboratories
located in Northwood, North Dakota. The soil sample appeared to be sourced from the same bulk control soil
sample which was used in the ECM.
6 Based on Figures 24-27, pp. 58-61 of MRID 49863305.
7 Based on Figures 39-42, pp. 73-76 of MRID 49863305.
A confirmatory method is not usually required when LC/MS and GC/MS is the primary method.
Page 9 of 13
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Cyproconazole (PC 128993)
MRIDs 49863306 / 49863305
IV. Method Deficiencies and Reviewer's Comments
1. In the ECM analysis, no samples were fortified at lOxLOQ (Tables 1-4, pp. 28-31 of
MRID 49863306). OCSPP guidelines recommend that a minimum of five spiked
replicates were analyzed at each concentration (i.e., minimally, the LOQ and 10x LOQ)
for each analyte.
2. The purities of the test materials were not reported in the ECM (pp. 8-9 of MRID
49863306).
In the EMC, the reviewer noted that the CAS # of cyproconazole was reported as 94361-
06-5, which corresponds to the (2RS, 3RS)-isomers (p. 8 of MRID 49863306). (p. 17 of
MRID 49863305). In the ILV, the reviewer noted that the CAS #s of cyproconazole were
reported as 94361-06-5, see above, and 94361-07-6, which corresponds to the (2RS,
3SR)-isomers.
3. The ECM soil matrix was sandy loam (47% sand, 30% silt, 23% clay, pH 7.5, percent
organic matter 3.4) was collected from Iowa obtained from an on-going Syngenta field
dissipation study (USDA soil textural classification; p. 20 of MRID 49863306), and ILV
soil matrix was sandy loam (54% sand, 26% silt, 20% clay, pH 7.3, percent organic
matter 2.6) was collected from Greene County, Iowa, and obtained from Syngenta
(USDA soil textural classification; Sample ID RIEN00707-0002; p. 16 of MRID
49863305). The ILV soil sample appeared to be sourced from the same bulk control soil
sample which was used in the ECM; however, more details about the soil sources would
need to be provided in order to make an accurate judgement.
4. The estimations of the LOQ and LOD in ECM and ILV were not based on scientifically
acceptable procedures as defined in 40 CFR Part 136 (pp. 21-22; Figure 2, p. 33 of MRID
49863306; pp. 12, 17, 29; Table 1, pp. 31-32 of MRID 49863305). In the ECM, the LOQ
was defined as the lowest analyte concentration in a sample at which the methodology
has been validated, i.e. which yielded a mean recovery of 70-120%) and relative standard
deviation of <20%. No LOQ calculations were provided in the ECM or ILV. No
justifications of the LOQ were provided in the ILV. In the ECM, the LOD was defined as
the smallest standard amount injected during the chromatographic run and typically
corresponds to an amount of analyte equivalent to ca. one-half of the theoretical amount
for a recovery sample at the method LOQ. The ECM study author noted that the LOD
may vary from instrument to instrument depending on the injection volume and
concentrations needed to obtain adequate analyte response). No LOD calculations were
reported in ECM or ILV.
Additionally, the lowest toxicological level of concern in soil for the analytes was not
reported in the ECM and ILV. An LOQ above toxicological levels of concern results in
an unacceptable method classification.
5. The ILV study author noted the following suggestions to improve the method: 1) more
details about the evaporation time and conditions should be provided for concentration of
Page 10 of 13
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Cvproconazole (PC 128993)
MRIDs 49863306 / 49863305
the extracts after each SPE column clean-up; and 2) the SPE clean-up flow rate of less
than 20 drops per minute was considered time consuming and difficult to achieve, and
more guidance was requested to discuss the flow rate optimization (p. 23 of MRID
49863305).
6. The ECM method was validated by the ILV with the second analysis of the extracts of
the first trial (pp. 12, 23-24 of MRID 49863305). For the analysis of the first extracts, the
ILV study author used an acetonitrile-based organic mobile phase, which was a sponsor-
approved modification from the methanol-based organic mobile phase specified in the
method. The ILV study author noted that this modification "appeared to be allowed by
the flexibility of the method as written" (p. 24). The ILV first analysis yielded shaper
peaks for the analytes, but unacceptable results. When the ILV used a methanol-based
organic mobile phase for the second analysis of the extracts of the first trial, acceptable
results were achieved. Therefore, the reviewer concluded that the methanol-based organic
phase for the analytical method cannot be modified.
7. In the ECM, the method calculations allowed for recoveries to be corrected for residues
quantified in the controls; however, residues were only identified in the control samples
of 1,2,4-triazole (1.840-2.660 ppb, ca. 4-6% of the LOQ; pp. 18-19; Tables 2-4, pp. 29-
31 of MRID 49863306).
In the ILV, the study author noted that analyte interference peaks were observed in the
controls for 1,2,4-triazole and triazole acetic acid (pp. 12, 22 of MRID 49863305). The
interferences were considered negligible because the area count was <50% of the LOD.
No values for the control residues were reported in the recovery data tables in the study
report (Table 1, pp. 31-32 of MRID 49863305).
8. In ECM and ILV representative LOQ and lOxLOQ chromatograms showed minor matrix
interferences (<15% of the LOQ) in the controls at the retention times of the analytes
(Appendix 4, Figures 1-6, pp. 32-37 of MRID 49863306; Figures 1-45, pp. 35-79 of
MRID 49863305).
9. The ILV study author provided communication details between the ILV laboratory
personnel and the Study Sponsor (pp. 23-24 of MRID 49863305). These communications
included typical questions about the laboratory materials for the method and method
clarifications.
10. It was reported for the ILV that a sample set consisting of approximately 13 samples
required ca. 12 hours {ca. 1.5 person days) to complete (p. 22 of MRID 49863305).
Page 11 of 13
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Cvproconazole (PC 128993)
MRIDs 49863306 / 49863305
V. References
U.S. Environmental Protection Agency. 2012. Ecological Effects Test Guidelines, OCSPP
850.6100, Environmental Chemistry Methods and Associated Independent Laboratory
Validation. Office of Chemical Safety and Pollution Prevention, Washington, DC. EPA
712-C-001.
40 CFR Part 136. Appendix B. Definition and Procedure for the Determination of the Method
Detection Limit-Revision 1.11, pp. 317-319.
Page 12 of 13
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Cvproconazole (PC 128993) MRIDs 49863306 / 49863305
Attachment 1: Chemical Names and Structures
Cyproconazole (SAN619; CCZ)
IUPAC Name:
CAS Name:
CAS Number:
SMILES String:
(2RS,3RS;2RS,3SR)-2-(4-chlorophenyl)-3-cyclopropyl-l-(lH-l,2,4-
triazol-l-yl)butan-2-ol.
2-(4-Chloro-phenyl)-3 -cyclopropyl-1 -[ 1,2,4]triazol-1 -yl-butan-2-ol.
a-(4-Chlorophenyl)-a-(l -cyclopropyl ethyl)- 1H-1,2,4-triazole-1 -ethanol.
94361-06-5 (2RS, 3RS)-isomers.
94361-07-6 (2RS, 3SR)-isomers.
113096-99-4 (unstated stereo chemistry).
Clclccc(ccl)C(0)(C(ClCCl)C)Cnlncncl (ISIS v2.3/Universal
SMILES).
c 1 cc(Cl)ccc 1 C(0)(C(C)C2CC2)Cn3ncnc3 (EPI Suite, v3.12 SMILES).
OH f ^
°H N—N
CI
1,2,4-Triazole (CGA 71019; T)
IUPAC Name:
CAS Name:
CAS Number:
SMILES String:
4H-[l,2,4]Triazole.
4H-1,2,4-Triazole.
lH-l,2,4-Triazole.
288-88-0.
nlcnncl (ISIS v2.3/Universal SMILES),
nlncncl (EPI Suite, v3.12).
N
\
N
JJ
Triazole Acetic Acid (CGA-142856; CSAA131731; TAA)
IUPAC Name: [l,2,4]Triazol-l-yl-acetic acid.
CAS Name: lH-l,2,4-Triazole-l-acetic acid.
CAS Number: 28711 -29-7
SMILES String: Not found
N^\
HO
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