Etridiazole (PC 084701)
MRIDs 50534503/50584601
Analytical method for etridiazole and its metabolites etridiazole acid and 3-DCMT (DCE) in
water
Reports: ECM: EPA MRID No. 50534503. DeVellis, S.R. 2017. Validation of the
Analytical Method for the Determination of Etridiazole and its Metabolites
in Surface and Ground Water by LC-MS/MS and GC-MS. Smithers Viscient
Study No.: 14088.6157. Report prepared by Smithers Viscient, Wareham,
Massachusetts; sponsored and submitted by MacDermid Agricultural
Solutions, Inc. c/o Arysta LifeScience North America, LLC, Cary, North
Carolina; 124 pages. Final report issued December 21, 2017.
ILV: EPA MRID No. 50584601. Cashmore, A. 2018. Independent
Laboratory Validation of: Etridiazole - Independent Laboratory Validation
of Analytical Method 14088.6157 for the Determination of Etridiazole and
its metabolites Etridiazole acid and DCE in Water. Smithers Viscient (ESG)
Ltd. Study No.: 3202058 and Document No.: 2017-356. Report prepared by
Smithers Viscient (ESG) Ltd., North Yorkshire, United Kingdom; sponsored
and submitted by MacDermid Agricultural Solutions, Inc. c/o Arysta
LifeScience North America, LLC, Cary, North Carolina; 100 pages. Final
report issued April 24, 2018 (pp. 2-5).
Document No.: MRIDs 50534503 & 50584601
Guideline: 850.6100
Statements: ECM: The study was conducted in accordance with the USEPA FIFRA (40
CFR Part 160) and OECD Good Laboratory Practices (GLP; p. 3 of MRID
50534503). Signed and dated No Data Confidentiality, GLP and Quality
Assurance statements were provided (pp. 2-4). A certification of the
authenticity of the report was included with the QA statement.
Classification:
PC Code:
EFED Final
Reviewer:
CDM/CSS-
Dynamac JV
Reviewers:
ILV: The study was conducted in accordance with the UK GLP and OECD
GLP and was in compliance with the GLP regulation and are suitable for
submission to the US FDA/EPA/Japanese regulatory authorities (p. 3 of
MRID 50584601). Signed and dated No Data Confidentiality, GLP,
Authenticity, and Quality Assurance statements were provided (pp. 2-5).
This analytical method is classified as supplemental. ECM and ILV linearity
were not satisfactory for etridiazole analysis. ILV linearity was not
satisfactory for 3-Carb-T (etridiazole acid) in surface water. The specificity
of the method was not supported for 3-DCMT and 3-Carb-T based on ILV
representative chromatograms. The ECM representative chromatogram did
not support the specificity for 3-Carb-T in surface water at the LOQ. ECM
matrices were not well-characterized.
084701
O * Digitally signed by CHERYL
Signature: cherylsutton ^
O Date: 2018.10.30 09:38:14-04'00'
Date: October 24, 2018
Signature:
Date: 9/2S/18
Signature:
Date: *9/28/18
Cheryl Sutton, Ph.D.
Environmental Scientist
TeresaNelis, M.S.,
Environmental Scientist
Lisa Muto, M.S.,
Environmental Scientist
Page 1 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
This Data Evaluation Record may have been altered by the Environmental Fate and Effects
Division subsequent to signing by CDM/CSS-Dynamac JVpersonnel. The CDM/CSS-Dynamac
Joint Venture role does not include establishing Agency policies.
Executive Summary
This analytical method, Smithers Viscient Study No. 14088.6157, is designed for the quantitative
determination of etridiazole and its metabolite 3-DCMT (DCE) in water at the LOQ of 0.100 |ig/L
using GC-MS/EI, and metabolite 3-Carb-T (etridiazole acid) in water at the LOQ of 0.100 |ig/L
using LC-MS/MS. The ECM and ILV validated the method using ground and surface water
matrices; however, the ECM matrices were not well-characterized. In the ECM and ILV, three ions
were monitored for etridiazole and 3-DCMT, and two ion transitions were monitored for 3-Carb-T
analysis. The ILV validated the ECM method for the quantitation and confirmation analyses of
etridiazole, 3-DCMT, and 3-Carb-T in ground water and surface water with insignificant
modifications to the analytical instruments. The ILV validation was completed in the first trial for
all analyses, except for etridiazole and 3-DCMT in ground water at the LOQ which passed in the
second trial without additional modification of the ECM. All ECM and ILV data were satisfactory
regarding repeatability, accuracy, and precision for all analytes. All ECM and ILV data were
satisfactory regarding specificity for etridiazole. The specificity of the method was not supported
for 3-DCMT and 3-Carb-T based on ILV representative chromatograms due to significant baseline
noise which interfered with accurate identification and integration of the quantitation analysis LOQ
peak in both matrices. Matrix interference was also observed in the ECM chromatogram for 3-Carb-
T in surface water at the LOQ. The reviewer believed that additional clean-up of the matrix or
matrix-matched standards may be required. ECM and ILV linearity was satisfactory for 3-DCMT
analyses, but was not satisfactory for etridiazole analysis. ECM and ILV linearity was satisfactory
for 3-Carb-T analyses, except for ILV analysis in surface water.
Page 2 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
Table 1. Analytical Method Summary.
Analyte(s)
by
Pesticide
MRID
EPA
Review
Matrix
Method
Date
Registrant
Analysis
Limit of
Quantitation
(LOQ)
Environmental
Chemistry
Method
Independent
Laboratory
Validation
Etridiazole
50534503
50584601
Water1,2
12/21/2017
MacDermid
Agricultural
Solutions,
Inc.
c/o Arysta
LifeScience
North
America,
LLC
GC/MS
0.100 ng/L
3-DCMT
(DCE)
3-Carb-T
LC/MS/MS
1 In the ECM, ground water was obtained from a 100-meter bedrock well, location not reported, and filtered (method
not specified) prior to use; and surface water (pH 6.18, 5.92 mg/L dissolved oxygen content) was obtained from
Weweantic River, Wareham, Massachusetts. Ground water characterization was absent; surface water
characterization was performed by Smithers Viscient, Wareham, Massachusetts (pp. 16-17 of MRID 50534503).
2 In the ILV, ground water (pH 7.6, 176 mg/L total hardness (as CaCCh)- 4 mg/L suspended solids, 0.0 mg/L dissolved
organic carbon, and 467 |iS/cm conductivity) was obtained from AgroChemex, well location not reported; and
surface water (pH 7.9, 200 mg/L total hardness (as CaCCh)- 15 mg/L suspended solids, 4.94 mg/L dissolved organic
carbon, and 49 |iS/cm conductivity] was obtained from Fountains Abbey, Ripon, UK (Appendix 2, pp. 96-97). Water
characterization was performed by Smithers Viscient, Wareham, Massachusetts (p. 15, Appendix 2, pp. 96-97 of
MRID 50584601).
I. Principle of the Method
Samples (ca. 60.0 mL) were fortified with 0.0600 mL or 0.600 mL of the fortification solutions
(0.100 mg/L) of etridiazole and 3-DCMT in acetonitrile to prepare LOQ (0.100 |ig/L) and lOxLOQ
(1.00 |ig/L) fortified samples, respectively; samples {ca. 25.0 or 8.00 mL) were fortified with
0.0250 mL or 0.0800 mL, respectively, of the fortification solutions (0.100 mg/L) of 3-Carb-T in
acetonitrile to prepare LOQ and lOxLOQ fortified samples (pp. 18, 21-23 of MRID 50534503).
The water samples fortified with etridiazole and 3-DCMT were extracted with 2.00 mL of iso-
octane using 60 mL glass vials with PTFE lined caps, placed on a shaker table for 30 minutes at 250
rpm, settled for 10 minutes, and the entire extraction layer, including residual water and any
emulsion, was transferred using a transfer pipet into disposable conical glass vials (pp. 23-25 of
MRID 50534503). The etridiazole and 3-DCMT samples were centrifuged at 1200 rpm for 20
minutes to complete sufficient phase separation. The lOxLOQ-level extracts were further diluted
into the calibration range with the prepared matrix matched blank before analysis via GC-MS/EI.
The water samples fortified with 3-Carb-T received ammonium hydroxide (5 [xL to the 25.0 mL
sample and 1 [xL to the 8.00 mL sample), were loaded onto Oasis Mixed-Mode Strong Anion
Exchange (MAX) SPE columns (60 mg, 3 mL, pre-conditioned with two column volumes of
methanol and two column volumes of purified reagent water), and allowed to flow through under
vacuum (1 drop/sec. flow) (pp. 25-27 of MRID 50534503). Each sample vessel and column were
rinsed with 5.00 mL of purified reagent water, the rinsate loaded on the column, and allowed to
flow under vacuum at 1 drop/sec., and then rinsed in a similar manner with 5.00 mL of methanol,
and the rinsates discarded. The columns were quickly dried under full vacuum, and the test
substance was eluted from the SPE columns with 3.00 mL of 2% trifluoroacetic acid in methanol
Page 3 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
under vacuum at 1 drop/sec. and collected into glass conical vials. The samples were concentrated
to incipient dryness under a gentle stream of nitrogen at 50.0°C. The residue was reconstituted to
5.00 mL with 20:80:0.1 acetonitrile:purified reagent watentrifluoroacetic acid (v:v:v) via mixing
and sonication for five minutes before analysis via LC-MS/MS.
Etridiazole and 3-DCMT sample extracts were analyzed via GC-MS/EI using an Agilent 6890
series GC and Agilent DB-5MS column (15 m x 0.25 mm x 0.25 |im) using a column temperature
program (initial 65°C and held for 2.00 minutes to post 150°C held for 0.00 minutes); injection
volume of 2.00 |iL; and helium carrier gas coupled with MS detection (300°C transfer line and
230°C source temperature) (pp. 16, 27-28 of MRID 50534503). Three ions were monitored as
follows (quantitation, confirmation 1, and confirmation 2, respectively): m/z 211.00, 185.00, and
183.00 for etridiazole, and m/z 149.00, 184.00, and 186.00 for 3-DCMT. Reported retention times
for etridiazole and 3-DCMT were ca. 12.9 and 11.0 minutes, respectively.
3-Carb-T sample extracts were analyzed via LC-MS/MS using an Agilent column and MDS Sciex
API 5000 mass spectrometer (pp. 28-29 of MRID 50534503). The LC conditions consisted of a
Agilent Poroshell 120 EC-C8 column (2.7 |im x 3.0 mm x 50 mm column, column temperature
35°C), a mobile phase of (A) 0.1% trifluoroacetic acid in water and (B) 0.1% trifluoroacetic acid in
acetonitrile [percent A:B (v:v) at 0.01 min. 98.0:2.0, 0.50 min. 98.0:2.0, 3.00 min. 0:100, 4.00 min.
0:100, 4.10 min. 98.0:2.0, and 5.00 min. 98.0:2.0] and MS/MS detection in positive ion mode
(ionization temperature 550°C). Injection volume was 50 |xL. Two ion transitions were monitored
(quantitation and confirmation, respectively) as follows: m/z 175.16—>147.10 and m/z
175.16—>-129.00. Retention time was ca. 2.2 minutes.
In the ILV, the ECM was performed as written, except for insignificant modifications of the
analytical instrumentation and parameters, and differences in calibration standards used and matrix
effect assessments. The ILV reported that a Thermo Trace 1300 Gas Chromatograph with ISQ LT
single quadrupole mass spectrometer detector was used for etridiazole and 3-DCMT (similar GC-
MS parameters; retention times for etridiazole and 3-DCMT were 11.8 and 9.9 min., respectively)
and a Shimadzu Nexera UPLC system coupled with an AB Sciex API 5000 MS/MS detector
(similar LC-MS/MS parameters; same ionization temperature, 550°C) was used for 3-Carb-T (pp.
15, 22-23 of MRID 50584601). A 10 [xL injection volume was used for the first attempt of the
matrix assessment for 3-Carb-T by LC-MS/MS, but sensitivity and precision were insufficient at
this volume and the ILV determined a 50 [xL injection volume was necessary for the matrix
assessment and validation; the ECM also used a 50 [xL injection volume (pp. 23, 28 of MRID
50584601).
In the ECM and ILV, the method LOQs in water were 0.100 |xg/L for etridiazole, 3-DCMT, and 3-
Carb-T (pp. 32-33, 35-40 of MRID 50534503; pp. 12, 28 of MRID 50584601). In the ECM, the
method LODs were 0.01 |xg/L (Q) and 0.03-0.05 |xg/L (Cl/2) in ground water and 0.03 |xg/L (Q,
Cl/2) in surface water for etridiazole; 0.04 |xg/L (Q) and 0.03-0.05 |xg/L (Cl/2) in ground water and
0.03 |xg/L (Q) and 0.02-0.03 |xg/L (Cl/2) in surface water for 3-DCMT; and 0.009 |xg/L (Q) and
0.002 |xg/L (C) in ground water and 0.05 |xg/L (Q) and 0.01 |xg/L (C) in surface water for 3-Carb-T
(pp. 35-40 of MRID 50534503). In the ILV, the method LODs were 0.0252 |xg/L (Q) and 0.0379-
0.0599 |xg/L (Cl/2) in ground water and 0.0123 |xg/L (Q) and 0.0218-0.0383 |xg/L (Cl/2) in surface
water for etridiazole; 0.0636 |xg/L (Q) and 0.0133-0.0134 |xg/L (Cl/2) in ground water and 0.0152
|xg/L (Q) and 0.009-0.0106 |xg/L (Cl/2) in surface water for 3-DCMT; and 0.0304 |xg/L (Q) and
Page 4 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
0.0268 |ig/L (C) in ground water and 0.962 |ig/L (Q) and 0.0506 |ig/L (C) in surface water for 3-
Carb-T (pp. 26-27 of MRID 50584601).
II. Recovery Findings
ECM (MRID 50534503): Mean recoveries and relative standard deviations (RSD) from GC-MS/EI
analysis were within guideline requirements (mean 70-120%; RSD <20%) for etridiazole and its
metabolite 3-DCMT at 0.100 |ig/L (LOQ) and 1.00 |ig/L (lOxLOQ) in two water matrices (Tables
1-12, pp. 43-54). Mean recoveries and relative standard deviations from LC-MS/MS analysis were
within guideline requirements for 3-Carb-T at 0.100 |ig/L (LOQ) and 1.00 |ig/L (lOxLOQ) in two
water matrices (Tables 13-16, pp. 55-58). Performance data (recovery results) from primary and
confirmatory analyses were comparable, except for etridiazole at the LOQ in both matrices and 3-
DCMT at the LOQ in ground water (based on RSD values). The ground water was obtained from a
100-meter bedrock well, location not reported, and filtered (method not specified) prior to use; and
surface water (pH 6.18, 5.92 mg/L dissolved oxygen content) was obtained from Weweantic River,
Wareham, Massachusetts (pp. 16-17). Ground water characterization was absent; surface water was
poorly characterized and was performed by Smithers Viscient, Wareham, Massachusetts.
ILV (MRID 50584601): Mean recoveries and RSDs from GC-MS analysis were within guideline
requirements for etridiazole and its metabolite 3-DCMT at 0.10 |ig/L (LOQ) and 1.0 |ig/L
(lOxLOQ) in two water matrices (Tables 1-6, pp. 33-38, Tables 9-14, pp. 41-46 of MRID
50584601). Mean recoveries and relative standard deviations from LC-MS/MS analysis were within
guideline requirements for 3-Carb-T at 0.10 |ig/L (LOQ) and 1.0 |ig/L (lOxLOQ) in two water
matrices (Tables 7-8, pp. 39-40, Tables 15-16, pp. 47-48 of MRID 50584601). Performance data
(recovery results) from primary and confirmatory analyses were comparable. Ground water (pH 7.6,
176 mg/L total hardness (as CaC03), 4 mg/L suspended solids, 0.0 mg/L dissolved organic carbon,
and 467 |iS/cm conductivity) was obtained from AgroChemex, well location not reported; and
surface water (pH 7.9, 200 mg/L total hardness (as CaC03), 15 mg/L suspended solids, 4.94 mg/L
dissolved organic carbon, and 49 |iS/cm conductivity] was obtained from Fountains Abbey, Ripon,
UK (Appendix 2, pp. 96-97). Water characterization was performed by Smithers Viscient,
Wareham, Massachusetts (p. 15, Appendix 2, pp. 96-97). The ILV validated the ECM method for
the quantitation and confirmation analyses of etridiazole, 3-DCMT, and 3-Carb-T in ground water
and surface water with insignificant modifications to the analytical instruments (pp. 15, 22-23). The
ILV validation was completed in the first trial for all analyses, except for etridiazole and 3-DCMT
in ground water at the LOQ which passed in the second trial without additional modification of the
ECM (pp. 28-29).
Page 5 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
Table 2. Initial Validation Method Recoveries for Etridiazole and Metabolites 3- DCMT and
3-Carb-T in Wal
er.1,2
Analyte
Fortification
Level (jig/L)
Number
of Tests
Recovery
Range (%)
Mean
Recovery (%)
Standard
Deviation (%)
Relative Standard
Deviation (%)
Ground (Well) Water
Quantitation ion
Etridiazole
0.100 (LOQ)
5
88.0-109
92.9
11.3
12.2
1.00
5
90.8-99.8
95.1
3.65
3.83
3-DCMT
0.100 (LOQ)
5
88.9-107
98.5
7.99
8.11
1.00
5
97.3-99.3
98.1
0.751
0.766
3-Carb-T
0.100 (LOQ)
5
102-113
108
4.08
3.78
1.00
5
84.8-91.7
88.1
2.95
3.35
Confirmation ion 1
Etridiazole
0.100 (LOQ)
5
90.5-105
99.1
6.33
6.39
1.00
5
90.6-93.7
92.1
1.15
1.24
3-DCMT
0.100 (LOQ)
5
98.6-104
100
2.65
2.64
1.00
5
90.9-103
96.8
4.37
4.51
3-Carb-T
0.100 (LOQ)
5
105-112
109
3.25
2.99
1.00
5
83.4-93.7
88.7
4.35
4.90
Confirmation ion 2
Etridiazole
0.100 (LOQ)
5
98.0-104
101
2.59
2.55
1.00
5
93.7-101
96.9
2.65
2.74
3-DCMT
0.100 (LOQ)
53
68.1-104
95
15
16
1.00
5
91.4-101
96.2
4.78
4.97
Surface (River) Water
Quantitation ion
Etridiazole
0.100 (LOQ)
5
93.2-102
97.0
3.40
3.51
1.00
5
95.2-102
98.8
3.31
3.35
3-DCMT
0.100 (LOQ)
5
95.0-104
98.2
3.52
3.59
1.00
5
95.3-102
98.0
2.61
2.66
3-Carb-T
0.100 (LOQ)
5
95.9-109
101
5.67
5.60
1.00
5
85.0-102
93.0
5.85
6.30
Confirmation ion 1
Etridiazole
0.100 (LOQ)
5
75.5-106
97.3
12.7
13.1
1.00
5
90.9-99.8
97.4
3.74
3.84
3-DCMT
0.100 (LOQ)
5
92.3-97.8
93.9
2.30
2.45
1.00
5
94.0-103
98.0
3.34
3.41
3-Carb-T
0.100 (LOQ)
5
101-115
106
5.48
5.19
1.00
5
93.3-98.2
96.4
1.94
2.02
Confirmation ion 2
Etridiazole
0.100 (LOQ)
5
86.5-104
94.0
6.27
6.67
1.00
5
93.1-98.7
95.7
2.00
2.09
3-DCMT
0.100 (LOQ)
5
88.0-111
102
8.60
8.42
1.00
5
93.0-101
96.8
3.02
3.12
Data (uncorrected recovery results) were obtained from Tables 1-16, pp. 43-58, of MRID 50534503; DER Attachment
2.
1 The ground water was obtained from a 100-meter bedrock well, location not reported, and filtered (method not
specified) prior to use; and surface water (pH 6.18, 5.92 mg/L dissolved oxygen content) was obtained from
Weweantic River, Wareham, Massachusetts (pp. 16-17). Ground water characterization was absent; surface water
characterization was performed by Smithers Viscient, Wareham, Massachusetts.
Page 6 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
2 For GC-MS/EI: three ions were monitored as follows (quantitation, confirmation 1, and confirmation 2, respectively):
m/z 211.00, 185.00, and 183.00 for etridiazole, and m/z 149.00, 184.00, and 186.00 for 3-DCMT. For LC-MS/MS:
two ion transitions were monitored (quantitation and confirmatory, respectively) as follows: m/z 175.16—>147.10 and
m/z 175.16—>129.00 for 3-Carb-T.
3 Mean, standard deviation, and relative standard deviation were reviewer-calculated based on all five recovery values
in the study report; the study author omitted one recovery value from the statistics since it was deemed to be an
outlier (DER Attachment 2). Rules of significant figures were followed.
Page 7 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
Table 3. Independent Validation Method Recoveries for Etridiazole and Metabolites 3-
)CMT and 3-Carb-T in Water.1'2'3
Analyte
Fortification
Number
Recovery
Mean
Standard
Relative Standard
Level (jig/L)
of Tests
Range (%)
Recovery (%)
Deviation (%)
Deviation (%)
Ground (Well) Water
Quantitation ion
Etridiazole
0.100 (LOQ)
5
91-114
105
8.9
8.5
1.00
5
108-116
112
3.6
3.2
3-DCMT
0.100 (LOQ)
5
91-95
93
1.5
1.6
1.00
5
91-98
95
2.9
3.0
3-Carb-T
0.100 (LOQ)
5
71-98
83
12.0
14.5
1.00
5
85-93
88
3.2
3.7
Confirmation ion 1
Etridiazole
0.100 (LOQ)
5
103-121
111
8.8
7.9
1.00
5
106-120
111
5.8
5.2
3-DCMT
0.100 (LOQ)
5
102-115
109
5.3
4.9
1.00
5
93-102
98
3.5
3.6
3-Carb-T
0.100 (LOQ)
5
87-102
97
6.1
6.3
1.00
5
85-99
92
6.1
6.7
Confirmation ion 2
Etridiazole
0.100 (LOQ)
5
94-109
103
5.7
5.5
1.00
5
110-118
113
3.4
3.0
3-DCMT
0.100 (LOQ)
5
95-106
101
4.8
4.7
1.00
5
96-101
98
2.2
2.2
Surface (River) Water
Quantitation ion
Etridiazole
0.100 (LOQ)
5
81-113
94
14.2
15.0
1.00
5
90-104
99
5.9
6.0
3-DCMT
0.100 (LOQ)
5
80-89
86
4.3
5.0
1.00
5
76-93
87
6.5
7.5
3-Carb-T
0.100 (LOQ)
5
96-123
110
11.0
10.0
1.00
5
79-92
85
4.8
5.6
Confirmation ion 1
Etridiazole
0.100 (LOQ)
5
85-115
100
14.7
14.7
1.00
5
95-105
101
4.1
4.0
3-DCMT
0.100 (LOQ)
5
67-100
82
15.3
18.7
1.00
5
78-90
86
4.8
5.6
3-Carb-T
0.100 (LOQ)
5
77-117
92
15.0
16.4
1.00
5
81-97
87
6.3
7.2
Confirmation ion 2
Etridiazole
0.100 (LOQ)
5
85-116
97
13.4
13.8
1.00
5
92-107
101
6.0
5.9
3-DCMT
0.100 (LOQ)
5
67-100
83
14.9
17.9
1.00
5
78-91
86
4.9
5.7
Data (uncorrected recovery results) were obtained from Tables 1-16, pp. 33-48, of MRID 50584601.
1 3-DCMT synonym DCE and 3-Carb-T synonym etridiazole acid used in ILV.
2 The ground water (pH 7.6, 176 mg/L total hardness (as CaCCh). 4 mg/L suspended solids, 0.0 mg/L dissolved organic
carbon, and 467 |iS/cm conductivity) was obtained from AgroChemex, well location not reported; and surface water
(pH 7.9, 200 mg/L total hardness (as CaCCh). 15 mg/L suspended solids, 4.94 mg/L dissolved organic carbon, and 49
|iS/cm conductivity] was obtained from The Lake, Ripon, UK (Fountains Abbey). Water characterization was
Page 8 of 15
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
performed by Smithers Viscient, Wareham, Massachusetts.
3 For GC-MS: three ions were monitored as follows (quantitation, confirmation 1, and confirmation 2, respectively):
m/z 211, 185, and 183 for etridiazole, and m/z 149, 184, and 186 for 3-DCMT (DCE). For LC-MS/MS: two ion
transitions were monitored (quantitation and confirmatory, respectively) as follows: m/z 174.9—>146.9 and m/z
174.9—>129 for 3-Carb-T (etridiazole acid).
III. Method Characteristics
In the ECM and ILV, the method LOQs in water were 0.100 |ig/L for etridiazole, 3-DCMT, and 3-
Carb-T (pp. 32-33, 35-40 of MRID 50534503; pp. 12, 28 of MRID 50584601). In the ECM and
ILV, the LOQ was defined as the lowest fortification level, and it was noted that blank values
should not exceed 30% of the LOQ (p. 30 of MRID 50534503; p. 24 of MRID 50584601). For the
ECM, all blank values were < 30% for all analytes in ground water and surface water (pp. 35-40 of
MRID 50534503). For the ILV, the ILV criteria for specificity of < 50% of the analytes in the
blanks at their respective LOD and < 30% of the analytes in their blanks at their respective LOQ
were met for all blanks, except one blank for etridiazole at m/z 183 (confirmation ion 2) in ground
water, which showed an uncharacteristic noisy baseline (p. 27, Figure 13, p. 62 of MRID
50584601). No calculations or comparisons to background levels were reported to justify the LOQ
for the method in the ECM. In the ILV, the LOQ was reported from the ECM without justification.
In the ECM, the method LODs were 0.01 |ig/L (Q) and 0.03-0.05 |ig/L (Cl/2) in ground water and
0.03 |ig/L (Q, Cl/2) in surface water for etridiazole; 0.04 |ig/L (Q) and 0.03-0.05 |ig/L (Cl/2) in
ground water and 0.03 |ig/L (Q) and 0.02-0.03 |ig/L (Cl/2) in surface water for 3-DCMT; and 0.009
|ig/L (Q) and 0.002 |ig/L (C) in ground water and 0.05 |ig/L (Q) and 0.01 |ig/L (C) in surface water
for 3-Carb-T (pp. 35-40 of MRID 50534503). In the ILV, the method LODs were 0.0252 |ig/L (Q)
and 0.0379-0.0599 |ig/L (Cl/2) in ground water and 0.0123 |ig/L (Q) and 0.0218-0.0383 |ig/L
(Cl/2) in surface water for etridiazole; 0.0636 |ig/L (Q) and 0.0133-0.0134 |ig/L (Cl/2) in ground
water and 0.0152 |ig/L (Q) and 0.009-0.0106 |ig/L (Cl/2) in surface water for 3-DCMT; and 0.0304
|ig/L (Q) and 0.0268 |ig/L (C) in ground water and 0.962 |ig/L (Q) and 0.0506 |ig/L (C) in surface
water for 3-Carb-T (pp. 26-27 of MRID 50584601).
The LOD was calculated in the ECM using the following equation:
LOD = (3x(SNcti)/(RespLs) x ConcLs
Where, LOD is the limit of detection of the analysis, SNcti is the mean signal to noise in height of
the control samples (or blanks), RespLs is the mean response in height of the two low calibration
standards, and ConcLs is the concentration of the low calibration standard (p. 31 of MRID
50534503). The ILV LOD is calculated from 3 x height of control baseline noise x control dilution
factor x calibration standard concentration ([j,g/mL) / height of calibration standard peak (p. 24 of
MRID 50584601).
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
Table 4. Method Characteristics for Etridiazole, 3-DCMT, and 3-Carb-T in Water.
Analyte1
Etridiazole
3-DCMT
3-Carb-T
Analysis
GC/MS
LC/MS/MS
Limit of
Quantitation
(LOQ)
ECM
ILV
0.100 ng/L
0.100 ng/L
0.100 ng/L
Limit of
Detection
(LOD)
ECM
(Calc)
Ground
0.01 (ig/L (Q)
0.05 (ig/L (CI)
0.03 (ig/L (C2)
0.04 (ig/L (Q)
0.03 (ig/L (CI)
0.05 (ig/L (C2)
0.009 (ig/L (Q)
0.002 (ig/L (C)
Surface
0.03 (ig/L (Q/C1/C2)
0.03 (ig/L (Q)
0.02 (ig/L (CI)
0.03 (ig/L (C2)
0.05 (ig/L (Q)
0.01 (ig/L (C)
ILV
(Calc)
Ground
0.0252 (ig/L (Q)
0.0379 (ig/L (CI)
0.0599 (ig/L (C2)
0.0636 (ig/L (Q)
0.0134 (ig/L (CI)
0.0133 (ig/L (C2)
0.0304 (ig/L (Q)
0.0268 (ig/L (C)
Surface
0.0123 (ig/L (Q)
0.0218 (ig/L (CI)
0.0383 (ig/L (C2)
0.0152 (ig/L (Q)
0.009 (ig/L (CI)
0.0106 (ig/L (C2)
0.962 (ig/L (Q)
0.0506 (ig/L (C)
Linearity
(calibration
curve r2 and
concentration
range)
ECM
Ground
r2 = 0.99760 (Q)
r2 = 0.99565 (CI)
r2 = 0.99769 (C2)
r2 = 0.99768 (Q)
r2 = 0.99687 (CI)
r2 = 0.99773 (C2)
r2 = 0.99582 (Q)
r2 = 0.99761 (C)
Surface
r2 = 0.99385 (Q)
r2 = 0.99353 (CI)
r2 = 0.99262 (C2)
r2 = 0.99642 (Q)
r2 = 0.99493 (CI)
r2 = 0.99605 (C2)
r2 = 0.99836 (Q)
r2 = 0.99639 (C)
ILV2
Ground
r2 = 0.9936/0.9981 (Q)
r2 = 0.9935/0.9977 (CI)
r2 = 0.9936/0.9967 (C2)
r2 = 0.9964 (Q)
r2 = 0.9992 (CI)
r2 = 0.9986 (C2)
r2 = 0.9970 (Q)
r2 = 0.9944 (C)
Surface
r2 = 0.9926 (Q)
r2 = 0.9888 (CI)
r2 = 0.9882 (C2)
r2 = 0.9989/0.9968 (Q)
r2 = 0.9976 (CI)
r2 = 0.9986 (C2)
r2 = 0.9930 (Q)
r2 = 0.9851 (C)
Range
2-20 ng/L
0.250-2.50 (ig/L
Repeatable
ECM3
Yes at LOQ and lOxLOQ in one poorly characterized surface water and one
uncharacterized ground water matrices.
ILV4
Yes at LOQ and lOxLOQ in one well-characterized surface water and one well-
characterized ground water matrices.
Reproducible
Yes at LOQ and lOxLOQ
Specific
ECM5
Yes, matrix interferences
were <20% of the LOQ
(based on peak area).
Yes, no matrix
interferences were
detected in the controls.
Yes, matrix interferences
were <14% of the LOQ
(based on peak area);
however, Q LOQ
chromatogram for surface
water showed significant
baseline noise.6
ILV
Yes, matrix effects were
insignificant for ground
water and surface water,
but minor baseline noise
interfered with Q/Cl LOQ
peak attenuation.
No, matrix effects at the
analyte retention time
were insignificant;
however, the Q LOQ peak
integration and attenuation
was significantly altered
by baseline noise in
ground water and the Q
LOQ peak co-eluted with
overwhelming baseline
No, matrix effects at the
analyte retention time
were insignificant;
however, the Q LOQ peak
was only distinguishable
from the baseline noise by
retention time, and the Q
LOQ peak was very small
compared to contaminant
peaks (ca. 80% of the
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
Analyte1
Etridiazole
3-DCMT
3-Carb-T
Analysis
GC/MS
LC/MS/MS
noise in surface water.7
LOQ peak ht.).8
Data were obtained from pp. 32-33, 35-40 (LOQ/LOD); Tables 1-16, pp. 43-58 (recovery results); Figures 1-56, pp. 59-
114 (chromatograms and calibration curves) of MRID 50534503; pp. 12, 28 (LOQ/LOD); Tables 1-16, pp. 33-48
(recovery results); pp. 26-27, Figures 1-72, pp. 56-91 (chromatograms and calibration curves); Tables 17-22, pp. 49-54
(matrix effects) of MRID 50584601; DER Attachment 2. Analytes were identified using one quantitation ion and two
confirmation ions for etridiazole and 3-DCMT and one quantitation and one confirmatory ion transition for 3-Carb-T. Q
= Quantitation ion/ion transition; CI = Confirmation ion 1; C2 = Confirmation ion 2; C = Confirmation ion transition.
1 3-DCMT synonym DCE and 3-Carb-T synonym etridiazole acid used in ILV.
2 Two correlation coefficients (r2) values for etridiazole in ground water and 3-DCMT in surface water include the
repeat of the validation at the LOQ (0.10 |ig/L) for etridiazole and the re-injection of 3-DCMT at the LOQ (0.10
|ig/L) to achieve acceptable precision and/or accuracy (pp. 26-27; Appendix 4, p. 99 of MRID 50584601). 3-Carb-T
correlation coefficients (r2) values were reviewer-calculated from rvalues provided in the study report (pp. 26-27;
DER Attachment 2).
3 In the ECM, ground water was obtained from a 100-meter bedrock well, location not reported, and filtered (method
not specified) prior to use; and surface water (pH 6.18, 5.92 mg/L dissolved oxygen content) was obtained from
Weweantic River, Wareham, Massachusetts (pp. 16-17 of MRID 50534503). Ground water characterization was
absent; surface water characterization was performed by Smithers Viscient, Wareham, Massachusetts.
4 In the ILV, ground water (pH 7.6, 176 mg/L total hardness (as CaCOs,). 4 mg/L suspended solids, 0.0 mg/L dissolved
organic carbon, and 467 |iS/cm conductivity) was obtained from AgroChemex, well location not reported; and
surface water (pH 7.9, 200 mg/L total hardness (as CaCOs), 15 mg/L suspended solids, 4.94 mg/L dissolved organic
carbon, and 49 |iS/cm conductivity] was obtained from Fountains Abbey, Ripon, UK (Appendix 2, pp. 96-97). Water
characterization was performed by Smithers Viscient, Wareham, Massachusetts (p. 15, Appendix 2, pp. 96-97 of
MRID 50584601)..
5 In the ECM, matrix interferences based on representative peak areas of the control and the LOQ were <20%, 0%, and
<14% for etridiazole, 3-DCMT, and 3-Carbon-T, respectively. Matrix effects assessment performed by the study
author determined matrix effects were minimal for etridiazole and 3-Carb-T ground water and surface water but were
significant for 3-DCMT in ground water and surface water (pp. 35-40 of MRID 50534503); results of the matrix
assessment were not provided; the study author used matrix-matched standards for the method validation.
6 The chromatogram for 3-Carb-T for surface water primary quantitation for at the LOQ (0.100 |ig/L) showed
significant baseline noise the LOQ (0.100 |ig/L) (Figure 47, p. 105 of MRID 50534503). Additional clean-up may be
required to reduce the baseline noise or matrix-matched calibration standards should be used.
7 Based on Figure 32, p. 71 and Figure 35, p. 73 of MRID 50584601. Similar issues were not observed in the
confirmation ion chromatograms.
8 Based on Figure 59, p. 85 and Figure 62, p. 86 of MRID 50584601. Similar issues were noted in the confirmation ion
chromatograms.
Linearity is satisfactory when r2 > 0.995.
IV. Method Deficiencies and Reviewer's Comments
1. For the quantitation analysis, ILV linearity was not satisfactory for etridiazole in ground
water for the analyses at 0.10 |ig/L (the LOQ), r2 = 0.9936, or in surface water r2 = 0.9926
and for 3-Carb-T in surface water (r2 = 0.9930; pp. 26-27, Figure 1, p. 56 of MRID
50584601; DER Attachment 2).
For the quantitation analysis, ECM linearity was not satisfactory for etridiazole in surface
water, r2 = 0.99385 (p. 36, Figure 34, p. 92 of MRID 50534503).
For the confirmation analysis, ILV linearity was not satisfactory for etridiazole in ground
water for the analyses at 0.10 |ig/L (the LOQ), r2 = 0.9935 (CI) and 0.9936 (C2), or in
surface water r2 = 0.98888 (CI) and 0.9882 (C2) and for 3-Carb-T [r2 = 0.9944 (ground) and
0.9851 (surface); pp. 26-27 of MRID 50584601], For the confirmation analysis, ECM
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
linearity was not satisfactory for etridiazole in surface water, r2 = 0.99353 (CI) and 0.99262
(C2) and for 3-DCMT in surface water [r2 = 0.99493 (CI); pp. 36, 38 of MRID 50534503],
In the case of the confirmation analyses, the reviewer did not consider this guideline
deviation to be significant since a confirmatory method is not typically required where
GC/MS and/or LC/MS methods are used as the primary method(s) to generate study data.
Linearity is satisfactory when r2 >0.995.
2. The specificity of the method was not supported for 3-DCMT and 3-Carb-T based on ILV
representative chromatograms. For both analytes, matrix effects at the analyte retention time
were insignificant; however, significant baseline noise interfered with accurate identification
and integration of the quantitation analysis LOQ peak in both matrices. For 3-DCMT, the
quantitation LOQ peak integration and attenuation was significantly altered by baseline
noise in ground water and the quantitation LOQ peak co-eluted with overwhelming baseline
noise in surface water (Figure 32, p. 71 and Figure 35, p. 73 of MRID 50584601). Similar
issues were not observed in the confirmation ion chromatograms. For 3-Carb-T, the
quantitation LOQ peak was only distinguishable from the baseline noise by retention time,
and the quantitation LOQ peak was very small compared to contaminant peaks (ca. 80% of
the LOQ peak ht.; Figure 59, p. 85 and Figure 62, p. 86). Similar issues were noted in the
confirmation ion chromatograms.
3. In the ECM, the chromatograms for etridiazole and 3-DCMT analysis by GC/MS in ground
water and surface water were acceptable, even though the LOQ peak was broad (Figures 1-
5, 9-13, 29-33, 37-41, pp. 59-63, 67-71, 87-91, 95-99 of MRID 50534503). The
chromatogram for 3-Carb-T analysis by LC-MS/MS for surface water primary quantitation
at the LOQ (0.100 |ig/L) showed significant baseline noise, which indicates additional
clean-up of the matrix or matrix-matched standards may be required (Figure 47, p. 105 of
MRID 50534503). The reviewer did not fault the support of the specificity of the method
since representative ground water chromatograms were acceptable.
4. In the ECM, ground water characterization was absent; and surface water was poorly
characterized (p. 16 of MRID 50534503).
5. For the ILV, the first validation attempt for etridiazole and 3-DCMT in ground water
required re-injection due to precision and accuracy failure for etridiazole at the LOQ (0.10
|ig/L). Etridiazole at 10 x LOQ and 3-DCMT at the LOQ and 10 x LOQ passed on the first
attempt. Appendix 4, p. 99 of MRID 50584601 indicates that the validation was repeated,
without modification to the method, and was successful on the fourth attempt; however, no
explanation was given why the two re-injections failed.
6. In the ILV, matrix-matched standards were not used for calibration; matrix effects data
indicated there were no matrix interferences (Tables 17-22, pp. 49-54 of MRID 50584601).
The chromatogram for the etridiazole control in ground water for confirmation ion two
showed appreciable noise, indicating clean-up of the matrix or matrix-matched standards
may be required (Figure 13, p. 62 of MRID 50584601).
In the ECM, matrix-matched standards were used for the calibration of etridiazole and 3-
DCMT; matrix-matched standards were not used for 3-Carb-T calibration. The ECM
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
reported matrix effects were significant for 3-DCMT in ground water and surface water and
were insignificant for etridiazole and 3-Carb-T in both waters (pp. 35-40 of MRID
50534503); however, no matrix effects data were provided.
The ILV reported matrix effects as: (mean measured concentration with solvent - mean
measured concentration with matrix)/(measured concentration with solvent) x 100 (p. 17,
Tables 17-22, pp. 49-54 of MRID 50584601). The ILV matrix assessment differed from the
ECM in concentration of matrix matched standards [10 |ig/L etridiazole and 3-DCMT and
1.0 |ig/L for 3-Carb-T in the ILV (p. 18 of MRID 50584601); and 3.0 |ig/L for etridiazole
and 3-DCMT and 0.5 |ig/L for 3-Carb-T in the ECM (pp. 20-21 of MRID 50534503)]. All
monitored ion transitions were the same as those of the ECM.
7. The ECM laboratory is Smithers Viscient Laboratory in Wareham, Massachusetts, and the
ILV laboratory is also a Smithers Viscient Laboratory, but located in Harrogate, North
Yorkshire, United Kingdom. One communication was documented between the two
laboratories, and that was to confirm the SPE cartridge particle size. The other documented
communications were between the Harrogate laboratory and the sponsor MacDermid
Agricultural Solutions, Inc., concerning schedule and approval to repeat of etridiazole at
0.10 |ig/L in ground water (Appendix 5, p. 100 of MRID 50584601).
Note, if the laboratory that conducted the validation belonged to the same organization as
the originating laboratory, the analysts, study director, equipment, instruments, and supplies
of the two laboratories must have been distinct and operated separately and without
collusion. The analysts, study director, equipment, instruments, and supplies of the two
laboratories were different and operated separately. Furthermore, the analysts and study
director of the ILV must have been unfamiliar with the method both in its development and
subsequent use in field studies; the unfamiliarity of the ILV with the method and its
development was not stated in the ILV, but the report stated the method was independently
validated (p. 30 of MRID 50584601).
8. The reviewer noted that the ILV reported that a single calibration standard run after the
sample analysis for etridiazole and 3-DCMT on ground water was used to verify that the
instrument had not drifted significantly after analysis of samples because the GC-MS
stopped after the first calibration standard run post sample analysis (p. 17 of MRID
50584601). For 3-Carb-T, the LC-MS/MS stopped before the last three standards were run
after the samples were analyzed; the first three calibration standards were accepted and any
re-analyzed samples were analyzed with the initial full set of calibration standards so the
calibration and sample extracts were equally aged (quality of aged calibration standards was
verified by acceptable correlation coefficients.
9. The time required to complete the method for a sample set was not reported in the ECM or
ILV.
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
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.
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Etridiazole (PC 084701)
MRIDs 50534503/50584601
)ER ATTACHMENT 1. Etridiazole and Its Environmental Transformation Products.
Code Name/ Synonym
Chemical Name
Chemical Structure
Study Type
MRID
Maximum
%AR (day)
PARENT
Etridiazole
IUPAC: Ethyl 3-trichloromethyl-
l,2,4-thiadiazol-5-yl ether
CAS: 5-Ethoxy-3-(trichloromethyl)-
1,2,4-thiadiazole
CAS No.: 2593-15-9
Formula: C5H5CI3N2OS
MW: 247.5 g/mol
SMILES:
CCOcl nc(ns 1 )C(C1)(C1)C1
H 3 C C O
H ,
850.6100
ECM water
850.6100
ILV water
50534503
50584601
NA
Etridiazole acid (3-
Carb-T)
IUPAC: 5-Ethoxy-l,2,4-thiadiazole-
3-carboxylic acid
CAS No.: 67472-43-9
Formula: C5H6N2O3S
MW: 174.17 g/mol
SMILES: CCOclnc(nsl)C(=0)0
850.6100
ECM water
50534503
NA
850.6100
ILV water
50584601
DCE (3-DCMT; T-03)
IUPAC: 5-Ethoxy-3-
dichloromethy 1-1,2,4-thiadiazole
Formula: C5H6CI2N2OS
MW: 213.08 g/mol
SMILES: CCOclnc(nsl)C(Cl)Cl
850.6100
ECM water
50534503
NA
850.6100
ILV water
50584601
A AR means "applied radioactivity". MW means "molecular weight". NA means "not applicable". ECM means "environmental chemical methods". ILV means "independent laboratory
validation".
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