Methods of Analysis by the U.S. Geological Survey

National Water Quality Laboratory—Determination of 

Pesticides in Water by Graphitized Carbon-Based SolidPhase Extraction and High-Performance Liquid

Chromatography/Mass Spectrometry

Water-Resources Investigations Report 01–4134 

U.S. Department of the Interior 

U.S. Geological Survey
Methods of Analysis by the U.S. Geological Survey

National Water Quality Laboratory—Determination of 

Pesticides in Water by Graphitized Carbon-Based SolidPhase Extraction and High-Performance Liquid

Chromatography/Mass Spectrometry

By Edward T. Furlong, Bruce D. Anderson, Stephen L. Werner, 

Paul P. Soliven, Laura J. Coffey, and Mark R. Burkhardt 

U.S. Geological Survey

Water-Resources Investigations Report 01–4134 

Denver, Colorado 

2001 
U.S. Department of the Interior

Gale A. Norton, Secretary

U.S. Geological Survey

Charles G. Groat, Director

The use of trade, product, or firm names in this report is for descriptive purposes only and 

does not imply endorsement by the U.S. Government. 

For additional information write to: Copies of this report can be purchased from: 

Chief, National Water Quality Laboratory U.S. Geological Survey

U.S. Geological Survey Branch of Information Services

Box 25046, Mail Stop 407 Box 25286 

Federal Center Federal Center

Denver, CO 80225-0046 Denver, CO 80225-0286 
CONTENTS 

Abstract 1 

Introduction 1 

Analytical Method 3 

1. Application 3 

2. Summary of method 8 

3. Interferences 9 

4. Apparatus and instrumentation 9 

5. Reagents and solutions 11 

6. Safety precautions 19 

7. Procedure 20 

8. Calculations 35 

9. Reporting of results 37 

10. Quality assurance/quality control 37 

11. Method performance 39 

Conclusions 71 

References cited 72 

FIGURES 

1. Chromatogram of a positive ion standard used in this method 26 

2-6. Graphs of recoveries showing:

2. Compounds fortified at three levels in organic-free water and 

determined under positive ionization conditions 59 

3. Compounds fortified at three levels in ground-water samples and 

determined under positive ionization conditions 60 

4. Compounds fortified at three levels in surface-water samples and 

determined under positive ionization conditions 61 

5. Compounds determined under negative ionization conditions 62 

6. Median recoveries of method compounds from 285 laboratory 

reagent spikes determined from October 1, 1999, to September 30, 2000 66 

TABLES 

1. Compound names, uses, pesticide class, Chemical Abstract Service registry 

numbers, and codes 4 

2. Calibration solution composition for compounds determined under 

negative ionization conditions 14 

3. Calibration solution composition for compounds determined under positive 

ionization conditions 15 

4. Surrogate solution composition for compounds determined under positive 

and negative ionization conditions 17 

5. Volume dilutions for method working standards 18 

6. Working standard concentrations and equivalent sample concentrations 

for this method 19 

7. High-performance liquid chromatograph mobile-phase gradient 

for this method 27 

CONTENTS III
8. Mass spectrometer operating conditions during positive and negative 

ionization analyses used in this method 27 

9a. Mass spectrometer time-programmed operating conditions for individual 

compounds determined under positive ionization conditions 28 

9b. Mass spectrometer time-programmed operating conditions for individual 

compounds determined under negative ionization conditions 29 

10. Quantitation and confirmation ions used for the compounds determined 

in this method 30 

11. Autotune parameters for acceptable tuning criteria used in this method 31 

12. Typical minimum sample analysis sequence for high-performance liquid 

chromatography/mass spectrometry analysis in this method 34 

13. Accuracy and precision data from 10 determinations of the method 

compounds fortified at 0.025 microgram per liter in organic-free water, under 

positive ionization conditions 42 

14. Accuracy and precision data from 10 determinations of the method 

compounds fortified at 0.025 microgram per liter in organic-free water, under 

negative ionization conditions 43 

15. Accuracy and precision data from seven determinations of the method 

compounds fortified at 0.10 microgram per liter in organic-free water, under 

positive ionization conditions 43 

16. Accuracy and precision data from seven determinations of the method 

compounds fortified at 0.10 microgram per liter in organic-free water, under 

negative ionization conditions 44 

17. Accuracy and precision data from nine determinations of the method 

compounds fortified at 0.50 microgram per liter in organic-free water, under 

positive ionization conditions 45 

18. Accuracy and precision data from nine determinations of the method 

compounds fortified at 0.50 microgram per liter in organic-free water, under 

negative ionization conditions 46 

19. Accuracy and precision data from 10 determinations of the method 

compounds fortified at 0.025 microgram per liter in ground-water samples, under 

positive ionization conditions 46 

20. Accuracy and precision data from nine determinations of the method 

compounds fortified at 0.025 microgram per liter in ground-water samples, 

under negative ionization conditions 47 

21. Accuracy and precision data from six determinations of the method 

compounds fortified at 0.10 microgram per liter in ground-water samples, 

under positive ionization conditions 48 

22. Accuracy and precision data from six determinations of the method 

compounds fortified at 0.10 microgram per liter in ground-water samples, under 

negative ionization conditions 49 

23. Accuracy and precision data from 11 determinations of the method 

compounds fortified at 0.50 microgram per liter in ground-water samples, under 

positive ionization conditions 49 

24 Accuracy and precision data from 10 determinations of the method 

compounds fortified at 0.50 microgram per liter in ground-water samples, 

under negative ionization conditions 51 

IV CONTENTS
25. Ambient pesticide concentrations measured in South Platte River 

samples during this study 52 

26. Accuracy and precision data from nine determinations of the method 

compounds fortified at 0.05 microgram per liter in surface-water samples, 

under positive ionization conditions 53 

27. Accuracy and precision data from nine determinations of the method 

compounds fortified at 0.05 microgram per liter in surface-water samples, 

under negative ionization conditions 54 

28. Accuracy and precision data from six determinations of the method 

compounds fortified at 0.10 microgram per liter in surface-water samples, 

under positive ionization conditions 54 

29. Accuracy and precision data from six determinations of the method 

compounds fortified at 0.10 microgram per liter in surface-water samples, 

under negative ionization conditions 55 

30. Accuracy and precision data from eight determinations of the method 

compounds fortified at 0.50 microgram per liter in surface-water samples, 

under positive ionization conditions 56 

31. Accuracy and precision data from eight determinations of the method 

compounds fortified at 0.50 microgram per liter in surface-water samples, 

under negative ionization conditions 57 

32. Long-term accuracy and precision calculated from multiple determinations of the 

method compounds fortified in sample-set spikes, with the compounds 

added to organic-free water at 0.25 microgram per liter 64 

33. Detections of compounds in 197 set blanks analyzed with this method 67 

34. Method detection limits for the method described in this study and the 

predecessor method of Werner and others (1996) 69 

CONTENTS V
CONVERSION FACTORS AND ABBREVIATED WATER-QUALITY UNITS

Multiply By To obtain

centimeter (cm) 3.94 x 10-1 inch 

gram (g) 3.53 x 10-2 ounce, avoirdupois 

kilopascal (kPa) 1.45 x 10-1 pounds per square inch 

liter (L) 3.38 x 10-1 ounce 

meter (m) 3.28 x 10 0 foot 

microgram (μg) 3.53 x 10-8 ounce, avoirdupois 

microliter (μL) 3.38 x 10-5 ounce 

micrometer (μm) 3.94 x 10-5 inch 

milligram (mg) 3.53 x 10-5 ounce, avoirdupois 

milliliter (mL) 3.38 x 10-2 ounce 

millimeter (mm) 3.94 x 10-2 inch 

nanogram (ng) 3.53 x 10-11 ounce, avoirdupois 

Degrees Celsius (°C) may be converted to degrees Fahrenheit (°F) using the following equation: 

°F = 9/5 (°C) + 32. 

ABBREVIATED WATER-QUALITY UNITS

g/L gram per liter

g/mL gram per milliliter

kg/m2 kilogram per square meter 

μg/L microgram per liter

μg/μL microgram per microliter

μg/mL microgram per milliliter

μL/L microliter per liter

μL/mL microliter per milliliter

mg/L milligram per liter

mg/mL milligram per milliliter

mL/L milliliter per liter

mL/min milliliter per minute

ng/L nanogram per liter

ng/μg nanogram per microgram 

ng/μL nanogram per microliter

ng/mL nanogram per milliliter

VI CONTENTS
ABBREVIATIONS AND ACRONYMS

amu atomic mass unit 

BDMC 4-bromo-3,5-dimethyl phenyl-n-methylcarbamate 

cat. no. catalog number 

CCB continuing calibration blank 

CCV continuing calibration verification 

DAD photodiode-array detection 

“E” second elution fraction 

FEB field equipment blank 

FMS field matrix spike

GC gas chromatography

GC/MS gas chromatography/mass spectrometry

HPLC high-performance liquid chromatography

HPLC/MS high-performance liquid chromatography/mass spectrometry

kV kilovolt 

LOQ limit of quantitation 

LRB laboratory reagent blank 

LRL laboratory reporting level 

LRS laboratory reagent spike 

“M” first (methanol) elution fraction 

MDL method detection limit 

MHz megahertz 

MS mass spectrometry

m/z mass-to-charge ratio 

NAWQA National Water-Quality Assessment Program 

NWIS National Water Information System

NWQL National Water Quality Laboratory

ODS octadecylsilane

POC polar organic compound 

QA/QC quality assurance/quality control 

QC quality control 

RF response factor 

SPE solid-phase extraction 

TFA trifluoroacetic acid 

USEPA U.S. Environmental Protection Agency

USGS U.S. Geological Survey

UV ultraviolet 

v/v volume per volume 

w/v weight per volume 

≡ identical with, congruent

 equals approximately

± plus or minus 

CONTENTS VII
Methods of Analysis by the U.S. Geological Survey

National Water Quality Laboratory—Determination of 

Pesticides in Water by Graphitized Carbon-Based SolidPhase Extraction and High-Performance Liquid 

Chromatography/Mass Spectrometry

By Edward T. Furlong, Bruce D. Anderson, Stephen L. Werner, 

Paul P. Soliven, Laura J. Coffey, and Mark R. Burkhardt 

Abstract

In 1996, the U.S. Geological Survey

National Water Quality Laboratory (NWQL)

developed and implemented a graphitized

carbon-based solid-phase extraction and highperformance liquid chromatographic 

analytical method to determine polar pesticide 

concentrations in surface- and ground-water 

samples. Subsequently, the NWQL

developed a complementary analysis that uses 

high-performance liquid chromatography/ 

mass spectrometry to detect, identify, and 

quantify polar pesticides and pesticide

metabolites in filtered water at concentrations 

as low as 10 nanograms per liter. This new 

method was designed to improve sensitivity

and selectivity over the prior method, and to 

reduce known interferences from natural 

organic matter.

In this new method, pesticides are 

extracted from filtered water samples by using

a 0.5-gram graphitized carbon-based solidphase extraction cartridge, eluted from the 

cartridge, and concentrations determined by

using high-performance liquid chromatography

with electrospray ionization–mass spectrometry. The upper concentration limit is 1.000 

microgram per liter (μg/L) for most compounds. Single-operator method detection 

limits in organic-free water samples fortified 

with pesticides at a concentration of 0.025 

μg/L ranged from 0.0019 to 0.022 μg/L for all 

compounds in the method. The grand mean 

(mean of mean recoveries for individual 

compounds) recoveries in organic-free water 

samples ranged from 72 to 89 percent, fortified 

with pesticides at three concentrations between

0.025 and 0.5 μg/L. Grand mean recoveries in 

ground- and surface-water samples ranged 

from 46 to 119 percent, also fortified with 

pesticides at three concentrations between 

0.025 and 0.5 μg/L. Long-term recoveries 

from reagent water spikes were used to 

demonstrate that 38 of 65 compounds can be 

reported without qualification of the 

quantitative result across the analytical range of

the method. The remaining 27 are reported 

with qualified estimates of concentration 

because of greater variability of recovery. 

INTRODUCTION 

In 1996, the National Water Quality

Laboratory (NWQL) developed and 

implemented a graphitized carbon-based 

solid-phase extraction and high-performance 

liquid chromatographic analytical method to 

determine pesticide concentrations in surfaceand ground-water samples (Werner and 

others, 1996). The National Water-Quality

Assessment Program (NAWQA) uses this 

method for determining concentrations of 41 

polar pesticides and pesticide metabolites. 

NAWQA used the methods of Werner and 

others (1996) and Zaugg and others (1995) to 

develop a detailed and extensive data set for

nationwide assessment of pesticide presence, 

concentrations, and distribution (Larson and 

others, 1999).

INTRODUCTION 1
The method by Werner and others (1996) 

initially was developed because several 

pesticide classes that might be found in groundwater and surface-water samples are not readily

amenable to analysis by gas chromatography

(GC) or gas chromatography/mass 

spectrometry (GC/MS). Analysis by GC/MS is 

a long-established method for unequivocal 

identification and quantitation of organic

compounds, but these compound classes could 

not be analyzed without derivitization or other 

sample-modifying steps. Examples of these 

pesticide classes include phenylurea herbicides, 

chlorophenoxyacetic acid herbicides, 

methylcarbamate insecticides, sulfonylurea,

sulfonamide, and imidazolinone herbicides, 

and uracil-derivative pesticides. Because these 

pesticides are used on a national scale in large 

quantities, a broad-spectrum, sensitive

analytical method for monitoring selected 

pesticides in these classes was needed. The 

method of Werner and others (1996) initially

fulfilled this requirement, but in routine use,

shortcomings were identified that were inherent 

to the available analytical technologies used in 

the method. Several important polar pesticides 

could not be adequately resolved and identified 

by high-performance liquid chromatography

(HPLC) with photodiode-array detection. In

addition, detection, identification, and 

quantitation of polar pesticides in the presence

of co-isolated and ubiquitous dissolved organic 

matter required a more specific means of 

detection and identification to routinely achieve

the detection limits needed for ambient 

environmental concentrations. 

Concurrent with development of the 

method by Werner and others (1996), 

improvements were made in highperformance liquid chromatography/mass 

spectrometry (HPLC/MS) interfaces so that

sensitive and selective detection of polar

pesticides and pesticide metabolites can be

made routinely in extracts from 1-L water

samples (Ferrer and Barcelo, 1998). These 

newer atmospheric pressure ionization 

interfaces allow unattended and nearly

continuous operation of HPLC/MS systems, 

with detection limits (expressed as a filteredwater concentration in a nominal 1-L water

sample) as low as the tens of nanograms per 

liter (Crescenzi and others, 1997). 

The use of HPLC/MS as a routine 

analytical tool under production conditions 

was tested at the NWQL by Furlong and 

others (2000) for determination of 

sulfonylurea, imidazolinone, and sulfonamide 

herbicides. This study demonstrated that with 

sufficient laboratory and field quality control, 

hundreds of samples could be successfully

analyzed by HPLC/MS, at concentrations in 

the nanogram-per-liter range. The problems 

of analyte coelution and interference from

dissolved organic carbon are substantially

reduced by combining the instrumental

approach of Furlong and others (2000) with 

the solid-phase extraction procedure by

Werner and others (1996). As noted by

Werner and others (1996), the advantages of 

HPLC coupled with solid-phase extraction 

over other methods include use of less 

solvent, rapid extraction, field-extraction 

capabilities, lower solvent exposure to 

technicians, and the ability to automate the

extraction procedure and determine thermally

sensitive compounds. Although the addition 

of mass spectrometry increases the analytical

cost, this cost is outweighed by improvements 

in specificity and detectability. Collectively,

slightly increased analytical costs are 

outweighed by improvements in data quality. 

The method described herein was 

developed by the U.S. Geological Survey

(USGS) for use at the NWQL. It uses 

graphitized carbon-based solid-phase 

extraction coupled with HPLC/MS. The 

method complements the method by Werner 

and others (1996) by increasing the number of 

compounds identified and quantified from 39 

to 65. It provides similar method detection 

limits and enhanced detection of low 

2 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
concentrations of compounds in sample 

matrices with high dissolved organic carbon 

concentrations and other matrix interferences. 

It also supplements other methods of the 

USGS for determination of organic 

substances in water that are described by

Wershaw and others (1987) and by Fishman 

(1993). 

This report provides a detailed 

description of all aspects of the method, 

including equipment, reagents, sample 

extraction and elution techniques, sampling

protocol, tabulated quality-control data, 

calculations, and reporting of results. Bias 

and variability data and method detection 

limits for 65 pesticides are presented. 

The scope of the report includes 

determination of method performance in 

organic-free water samples and in two 

natural-water types—a ground-water sample 

and a surface-water sample from the Denver, 

Colorado, region. Method performance was 

determined at three pesticide concentrations—0.025, 0.10, and 0.50 μg/L—in each 

water type, with the exception of surface 

water, where the lowest concentration was 

0.05 μg/L. Method detection limits (MDLs) 

were determined in organic-free water at a 

concentration of 0.025 μg/L by using the 

method outlined by the U.S. Environmental 

Protection Agency (1997). 

The method described in this report was 

approved in June 1999 as a custom analytical 

method and implemented for routine sample 

analysis as provisional laboratory analytical

schedule LC9060. It was intended primarily

for the analysis of samples associated with the 

NAWQA program. The method has remained 

in use until the present (2001) with only

minor modifications. 

ANALYTICAL METHOD

Organic Compounds and Parameter 

Codes: Pesticides, Dissolved, 

Graphitized Carbon-Based Extraction, 

High-Performance Liquid 

Chromatography/Mass Spectrometry, 

O-2060-01 

1. Application 

This method is suitable for use with 

filtered water samples for the determination of

the pesticides and related degradation products 

that are specified in table 1. Many parent 

compounds, such as 2,4-D, were included 

because of their relative importance in

agriculture in the United States, as indicated 

by the amount applied nationally (Gianessi 

and Anderson, 1996). Other compounds, such 

as aldicarb, have or can have substantial 

impact on human or ecosystem health 

(Gustafson, 1993). Some compounds, such as 

caffeine, are ubiquitously detected and might 

indicate human wastewater contamination 

(Gulyas, 1997). Several compounds, such as 

2-hydroxyatrazine, are important degradation 

products of commonly used pesticides (Kolpin 

and others, 1998). The method is applicable 

for determining pesticides and pesticide

metabolites that are (1) efficiently isolated

from the sample matrix and adsorbed onto a

graphitized carbon black sorbent-filled 

cartridge, and (2) chromatographically

resolved and identified using an HPLC

coupled by an electrospray ionization source 

to a quadrupole mass spectrometer. 

The method has been tested and 

validated for filtered surface water and 

ground water. Although not specifically

tested for other aqueous matrices, it could be 

applied to precipitation, wastewater, tile drain 

discharge, and other sample types as long as

ANALYTICAL METHOD 3
4 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS Table 1. Compound names, uses, pesticide class, Chemical Abstract Service registry numbers, and codes [conc., concentration; Y/N, yes/no; CAS, Chemical Abstract Service; NWIS, National Water Information System; Use codes in alphabetical order; F, Fungicide; H, Herbicide; H-Degr., Herbicide degradant; I, Insecticide; I-Degr., Insecticide degradant. Class codes in alphabetical order: AA, Acylalanine; ACA, Acetamide; B., Biological agent; BI, Benzimidazole; BN, Benzonitrile; BTZ, Benzothiadiazole; BZ, Benzoic acid; C, Carbamate; CN, Chloro-Nicotinyl; CP, Chlorophenoxy Acid; DNA, Dinitroaniline; DNP, Dinitrophenol; DPE, Diphenyl ether; IMI, Imidazolinone; PH, Phthalimide; PU, Phenylurea; PY, Pyridine; PYA, Pyridyloxyacetic acid; PYD, Pyridazine; SA, Sulfonamide; SU, Sulfonylurea; T, Triazine; TC, Thiocarbamate; TDZ, Thiadiazole; TR, Triazole; U, Uracil; n/a, not applicable] Compound name Alternative or common name Reported as an estimated conc.1 (Y/N) Use Class CAS registry number NWIS parameter code Ionization mode for identification and quantitation 2,4,5-T (Negative Ion Surrogate) Trichlorophenoxyacetic acid N H CP 93-76-5 99958 Negative Ion 2,4-D Weedone-2,4-DP N H CP 94-75-7 39732 Negative Ion 2,4-D methyl ester 2,4-Dichlorophenoxyacetic methyl ester 

N

H CP 1928-38-7 50470 Positive Ion

2,4-DB 4-(2,4-dichlorophenoxy) 

butanoic acid 

Y

H CP 94-82-6 38746 Negative Ion

2,4-Dichlorophenylacetic acid (DCAA; negative ion internal standard) 

n/a 19719-28-9 Negative Ion

2-Hydroxyatrazine 4-(ethylamino)-6-[(1-

methylethyl)amino]-1,3,5-Triazin-2(1H)-one 

Y H-Degr.

T 2163-68-0 50355 Positive Ion

3(4-chlorophenyl)-1-methyl urea N'-(4-chlorophenyl)-Nmethylurea 

N H-Degr. PU 5352-88-5 61692 Positive Ion

3-Hydroxycarbofuran 2,3-dihydro-2,2-dimethyl-3,7-

Benzofurandiol, 7-(methylcarbamate) 

N I-Degr.

C 16655-82-6 49308 Positive Ion

3-Ketocarbofuran 3-oxocarbofuran Y I-Degr. C 16709-30-1 50295 Positive Ion Acifluorfen Tackle; Blazer N H DPE 50594-66-6 49315 Negative Ion Aldicarb Temik Y I C 116-06-3 49312 Positive Ion Aldicarb sulfone Aldoxycarb Y I-Degr. C 1646-88-4 49313 Positive Ion Aldicarb sulfoxide Temik Sulfoxide Y I-Degr. C 1646-87-3 49314 Positive Ion Atrazine Atrex; Atratol N H T 1912-24-9 39632 Positive Ion 
Table 1. Compound names, uses, pesticide class, Chemical Abstract Service registry numbers, and codes—Continued 

ANALYTICAL METHOD 5 

Compound 

name

Alternative or

common name

Reported 

as an

estimated

conc.1

(Y/N) 

Use Class CAS registry number NWIS parameter 

code

Ionization 

mode for

identification

and 

quantitation 

Barban (positive ion surrogate) Carbyne; Neoban; Caryne N H C 101-27-9 90640 Positive IonBendiocarb Ficam N I C 22781-23-3 50299 Positive IonBenomyl Benlate N F BI 17804-35-2 50300 Positive IonBensulfuron-methyl Londax N H SU 83055-99-6 61693 Positive IonBentazon Adagio; Galaxy; Storm Y H BTZ 25057-89-0 38711 Negative IonBromacil Bromax; Hyvar; Uragon Y H U 314-40-9 04029 Positive IonBromoxynil Bromanil; Buctril; Torch Y H BN 1689-84-5 49311 Negative IonCaffeine 1,3,7-Trimethylxanthine N 58-08-2 50305 Positive IonCaffeine-C13 (positive ion surrogate) N 99959 Positive IonCarbaryl Sevin N H C 63-25-2 49310 Positive IonCarbofuran Crisfuran; Furadan N H C 1563-66-2 49309 Positive IonChloramben methyl ester Amiben, methyl ester Y H CP 7286-84-2 61188 Positive IonChlorimuron-ethyl Classic N H SU 90982-32-4 50306 Positive IonChlorothalonil Bravo; Forturf Y H PH 1897-45-6 49306 Negative IonClopyralid Stinger; Lontrel N H PY 1702-17-6 49305 Negative IonCycloate Ro-Neet; Marathon Y H TC 1134-23-2 04031 Positive IonDacthal monoacid DCPA Monoacid; Monomethyl

tetrachloroterephtalate 

N H-Degr. CP 887-54-7 49304 Negative Ion

Deethylatrazine 2-Amino-4-isopropylamino-6-

chloro-s-triazine

Y H-Degr. T 6190-65-4 04040 Positive Ion

Deethyldeisopropylatrazine 6-chloro-1,3,5-triazine-2,4-

diamine 

Y H-Degr. T 3397-62-4 04039 Positive Ion

Deisopropylatrazine Amino-2-chloro-6-ethylamino-striazine Y H-Degr. T 1007-28-9 04038 Positive Ion

Dicamba Banval; Marksman; Clarity N H BZ 1918-00-9 38442 Negative IonDichlorprop 2,4-DP,Weedone DP N H CP 120-36-5 49302 Negative IonDinoseb DNPB; Caldon: Dynamite N H DNP 88-85-7 49301 Negative Ion
6 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS Table 1. Compound names, uses, pesticide class, Chemical Abstract Service registry numbers, and codes—Continued Compound name Alternative or common name Reported as an estimated conc.1 (Y/N) Use Class CAS registry number NWIS parameter code Ionization mode for identification and quantitation Diphenamid Dymid; Enide; Rideon; Dyfen N H ACA 957-51-7 04033 Positive Ion Diuron DCMU; Diurex; Aguron N H PU 330-54-1 49300 Positive Ion Fenuron Beet-Klean N H PU 101-42-8 49297 Positive Ion Flumetsulam DE 498; XRD 498 Y H SA 98967-40-9 61694 Positive Ion Fluometuron Cortoran; Lanex; Cottonex N H PU 2164-17-2 38811 Positive Ion Imazaquin Image 1.5LC; Scepter 1.5L Y H IMI 81335-37-7 50356 Positive Ion Imazethapyr Pursuit; Pursuit DG Y H IMI 81335-77-5 50407 Positive Ion Imidacloprid Admire; Gaucho; Merit N I CN 138261-41-3 61695 Positive Ion Linuron Lorox; Afalon; Linurex; Linex N H PU 330-55-2 38478 Positive Ion MCPA Bordermaster; Metaxon N H CP 94-74-6 38482 Negative Ion MCPB Tropotox; Can-Trol; PDQ Y H CP 94-81-5 38487 Negative Ion Metalaxyl Apron; Subdue; Ridomil N F AA 57837-19-1 50359 Positive Ion Methiocarb Mercaptodimethur; Draza; Mesurol 

Y I C 2032-65-7 38501 Positive Ion

Methomyl Nudrin; Lannate; Lanox Y I C 16752-77-5 49296 Positive Ion Methomyl oxime 1-(methylthio)-acetaldehyde 

oxime 

Y I-Degr.

C 13749-94-5 61696 Positive Ion

Metsulfuron-methyl Escort;Gropper; Ally Y H SU 74223-64-6 61697 Positive Ion Monuron (positive ion internal standard) 

Telvar n/a

H PU 150-68-5 Positive Ion

Neburon Granurex; Herbalt; Kloben N H PU 555-37-3 49294 Positive Ion 

Nicosulfuron Accent; Accent DF

N H SU 111991-09-4 50364 Positive Ion

Norflurazon Zorial; Evital; Solicam; Telok Y H PYD 27314-13-2 49293 Positive Ion 

Oryzalin Ryzelan; Surflan; Dirimal N H DNA 19044-88-3 49292 Positive Ion Oxamyl Vydate: Thioxamyl N I C 23135-22-0 38866 Positive Ion 
Table 1. Compound names, uses, pesticide class, Chemical Abstract Service registry numbers, and codes—Continued 

ANALYTICAL METH

Compound 

name

Alternative or

common name

Reported 

as an

estimated

conc.1

(Y/N) 

Use Class CAS registry number NWIS parameter 

code

Ionization 

mode for

identification

and 

quantitation 

Oxamyl oxime 2-(Hydroxyimino)-N,N-dimethyl2-(methylmercapto)acetamide Y I-Degr. C 30558-43-1 50410 Positive IonPicloram Tordon; Amdon; Grazon N H PY 1918-02-1 49291 Negative IonPropham IPC; Chem-Hoe; Premalox N H C 122-42-9 49236 Positive IonPropiconazole Tilt; Orbit; Banner: Proconazole; 

Wocosin

N F TR 60207-90-1 50471 Positive Ion

Propoxur Baygon; PHC; Suncide; Unden N I C 114-26-1 38538 Positive IonSiduron Tupersan; Trey N H PH 1982-49-6 38548 Positive IonSulfometuron-methyl Oust; DPX-T5648 N H SU 74222-97-2 50337 Positive IonTebuthiuron Graslan; Spike; Perflan N H TDZ 34014-18-1 82670 Positive IonTerbacil Sinbar; DPX-D732; Geonter Y H U 5902-51-2 04032 Positive IonTribenuron-methyl Express; tribenuron methyl ester; 

DPX-L5300 

Y H SU 101200-48-0 61159 Positive Ion

Triclopyr Garlon; Curtail; Redeem; 

Remedy

N H PYA 55335-06-3 49235 Negative Ion

1The term "estimated concentration" is used in this electronic file. It was changed from "qualified estimate," which was used in the original published report. 

Tables 13–24 and tables 26–31 also reflect this change. (Modified: February 4, 2002, etf)

OD 7
(1) the samples have been filtered using the

method of Shelton (1994), and (2) the user 

recognizes that performance characteristics of 

new matrices are not tested and any results

would be provisional. 

Two classes of determinations are

reported for samples analyzed by this method. 

Compounds for which the results of the 

determination are reported without 

qualification make up the first class. 

Compounds for which the results of the 

determination are reported as qualified 

estimates, and the concentration qualified 

with an “E”, are in the second class. The 

classification for each compound in the 

method is listed in table 1. 

These classes, and the criteria used to

establish them, are discussed in detail in 

section 11.6. Compounds that are reported 

without qualification are reproducibly wellrecovered using this method, as defined by

median recoveries of long-term laboratory

reagent spikes (LRS) between 60 and 120 

percent and with variation (as indicated by the 

nonparametric statistic F-pseudosigma) less 

than ± 25 percent. Compounds that are 

reported as qualified estimates do not meet

these long-term method performance criteria, 

but are retained in the method because they

are used in substantial quantity and have

important environmental or toxicological 

effects. Although quantitative method 

performance for this class lies outside the 

criteria described above, the criteria for 

qualitative identification of the detected

compounds are the same as that used for 

compounds reported without quantitative 

qualifications. Greater certainty exists in the

quantitative determination of concentration 

for compounds reported without 

qualification—this is the only difference 

between the reported concentrations of the 

two data classes. 

It is important to note, however, that the 

long-term performance criteria for the 

compounds in the method span a continuum 

of performance, rather than corresponding to 

categories. As a consequence, the criteria 

used to discriminate between the two classes 

of data, although consistent with common 

analytical practice, are inherently arbitrary. 

The qualitative identification of the

compounds in both classes is equally reliable. 

Thus, data users should consider the estimated 

concentration as a categorical warning to pay

extra attention to potential use of numerical 

concentrations, but not as a distinct boundary

between good and poor data. 

2. Summary of Method 

This method is designed for the 

determination of 65 pesticides and pesticide

degradation products and caffeine (table 1) in 

filtered natural-water samples. The method is 

applicable to pesticides that are (1) efficiently

partitioned from the water onto a graphitized

carbon-based solid-phase extraction (SPE) 

material, (2) can be quantitatively eluted from 

the SPE material, and (3) can be efficiently

ionized by HPLC/MS with electrospray

interface. 

Pesticides are extracted from prefiltered 

water samples by using disposable 

polypropylene syringe cartridges that contain 

0.5 g of a graphitized carbon sorbent. A 

prefiltered water sample of 1 L is pumped 

through the SPE cartridge at a flow rate of 20 

mL/min. After extraction, the adsorbed 

compounds are eluted from the SPE cartridge

using two sequential elutions of 

(1) 1.5 mL methanol, followed by

(2) 13 mL of an 80-percent methylene 

chloride and 20-percent methanol 

mixture that has been acidified with

trifluoroacetic acid anhydride (0.2 

percent).

8 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
The two fractions are reduced under 

nitrogen to near dryness and then combined. 

The final volume for the extract is 1,000 μL. 

Analytes are chromatographically separated 

by HPLC using a reverse-phase 

octadecylsilane HPLC column, which is 

coupled to an electrospray ionization interface 

and quadrupole mass spectrometer for 

detection, identification, and quantitation. 

Each extract is analyzed twice using separate

ionization modes. The first analysis is for 

those compounds that preferentially form 

positive ions under electrospray conditions, 

and the second is for those compounds that 

preferentially form negative ions. 

The terms "extraction" and "elution" are 

used to define specific actions during sample 

processing. Extraction is the transfer of the 

selected compounds from the sample onto the 

SPE cartridge. Elution is the removal of the 

selected compounds from the SPE cartridge. 

3. Interferences 

Interferences might be caused by

compounds recovered from a sample matrix

that contains one or more of the same ions as 

the selected compound and that cannot be 

chromatographically resolved from the

selected compounds. Compared to optical 

detection methods typically used with HPLC, 

mass spectrometry is less affected by

interferences, but the potential for

interferences remains and requires special

attention to the ratios of the characteristic ions 

of interest to avoid false positive detections. 

In addition, for the sulfonylurea and 

imidazolinone compound classes, sample 

matrix components have been shown 

empirically to result in an apparent increase in

compound concentration (Furlong and others, 

2000). 

4. Apparatus and Instrumentation 

NOTE: During the development and 

implementation of this method, HewlettPackard Corporation, the manufacturer of the 

HPLC/MS instrumentation used in determining

method performance, formed a new company,

Agilent Technologies, from the original 

Hewlett-Packard Chemical Analysis Division 

and other company components. The 

HPLC/MS systems used in this study were 

originally made by Hewlett-Packard, and after 

formation of the new company, by Agilent 

Technologies. The phrase “HewlettPackard/Agilent Technologies” is used in this 

report to describe this identical instrumentation. 

4.1 High-performance liquid 

chromatograph—Hewlett-Packard Model

1090M Series II or Hewlett-Packard/Agilent 

Technologies Model 1100 high-performance 

liquid chromatographic system equipped as 

follows: a binary (Model 1100) or direct-ratio 

(DR5; Model 1090M) ternary-solvent 

delivery system, a photodiode-array

ultraviolet-absorbance detector (optional), a

250-μL automatic syringe sampler, a 100-

position random-access autosampler equipped 

with a cooling module, and thermostated 

column heating. An IBM-compatible

computer workstation also is required, with a 

minimum configuration of an Intel Pentium II

Processor operating at a minimum of 266 

MHz, 96 megabytes of Random Access 

Memory, and a minimum 20-gigabyte hard 

disk. Hewlett-Packard/Agilent Technologies 

LC/MSD ChemStation Revision A.06.03 

software, and Thru-Put Systems Target

Version 3.4 Software, or equivalent, are also 

required. 

4.2 Mass spectrometer—HewlettPackard/Agilent Technologies Series 1100 

LC/MSD mass spectrometer, or equivalent, 

equipped with an electrospray ionization 

source and capable of operating in positive

and negative ionization mode. 

ANALYTICAL METHOD 9
4.3 Analytical columns—

Chromatographic columns, MetaChem 

Technologies, Inc. Inertsil octadecylsilane 

(ODS-3), 5 m; 2.1-mm inside diameter by

15-cm stainless-steel column or equivalent. A 

guard column (MetaChem Technologies, Inc., 

MetaGuard 2.0-mm Inertsil octadecylsilane 

(ODS-3) direct-connect, 3 m, or equivalent, 

also is required. 

4.4 Manual sample extraction 

apparatus 

4.4.1. Solid-phase extraction 

manifold—Supelco, Inc., Visiprep SolidPhase Extraction Vacuum Manifold or 

equivalent. 

4.4.2 Ceramic-piston valveless 

sample pumps—Capable of pumping 0 to 

25 mL/min, Fluid Metering Inc. Model QSY2 CKC or equivalent. 

4.4.3 Sample flow path—All 

Teflon-perfluoralkoxy (PFA) 1/8-inch tubing

(3.18 mm) or equivalent. 

4.4.4 Tefzel-tetrafluoroethylene 

Luer connectors or equivalent. 

4.4.5 Luer stopcock—flow 

control on-off valves—Burdick & Jackson 

Inert PTFE flow-control valves or equivalent. 

4.4.6 Vacuum pump—Must be 

able to draw a vacuum equivalent to at least 

102 kPa. 

4.5 Automated sample extraction 

apparatus 

4.5.1 Zymark AutoTrace 6-place 

SPE workstation or equivalent—Equipped for 

processing 6-mL SPE tubes. 

4.5.2 Laboratory-auto trace 

barrel adapters—Made from 6-mL

polypropylene syringe tubes (structurally

identical to the SPE cartridge without the SPE 

packing), cut to one-half the original length 

(various manufacturers). 

4.5.3 SPE cartridge connectors—

Polyethylene (Restek #26007 or equivalent). 

4.5.4 Evaporative concentrator—

Temperature controlled to 34°C and nitrogen 

gas pressure of 69 kPa Zymark Turbo-Vap or 

equivalent. 

4.6 Liquid-handling apparatus 

4.6.1 Syringes—Hamilton 

Gastight 1750RN, 500 μL (cat. no. 81131); 

Gastight 1001LTN, 1,000 μL (cat. no. 

81317); and Hamilton Microliter 701, 10 μL

(cat. no. 80366) or equivalent. 

4.6.2 Micropipets—Van Waters 

and Rogers (VWR) 10- to 100-μL variable 

volume digital microdispenser (cat. no. 

53506201), VWR 100-μL fixed-volume 

microdispenser (cat. no. 53506675), and 

VWR 100-μL replacement tubes (cat. no. 

53508499) or equivalent. 

4.6.3 Autosampler vials—

National Scientific Company, 2-mL, 

graduated amber glass for use with screw-top 

caps (cat. no. C4000-2W) or equivalent. 

4.6.4 Vial caps and septa—

National Scientific Company, screw-top caps 

with 11-mm dual Teflon-faced silicone septa 

(cat. no. C4000-53B) or equivalent. 

4.6.5 20-mL solution storage 

vials—Eagle-Picher Technologies, LLC, 

20 mL, amber with Teflon-faced silicone 

rubber-lined screw caps (cat. no. 139-20A/CT) 

or equivalent. 

10 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
4.7 Consumables 

4.7.1 Amber-glass bottles—

1,000 mL, baked at 450°C for 2 hours, fitted 

with Teflon-lined screw caps or equivalent. 

4.7.2 Solid-phase extraction 

cartridges—Supelco ENVIRO-Carb 

Graphitized Carbon Black, graphitized 

nonporous carbon, 500 mg, 120/400 mesh, in 

6-mL syringe barrel or equivalent. 

4.7.3 Nitrogen gas for sample

extract concentration, ultrapure. 

4.7.4 Test tubes, graduated 14-

mL polypropylene round-bottom, 17- by 100-

mm style or equivalent. 

4.7.5 Disposable Pasteur pipets

cleaned by ashing at 440C for 2 hours. 

5. Reagents and Solutions 

NOTE: Material Safety Data Sheets for all 

materials described herein need to be read 

prior to using any of these materials to ensure

safe handling and proper disposal. Unless 

otherwise specified, solutions should be 

stored at room temperature and should be 

used for no longer than 6 months. 

5.1 Neat reagents 

5.1.1 Acetonitrile—Burdick and 

Jackson, ultraviolet (UV) grade or equivalent. 

5.1.2 L–(+)–Ascorbic acid—J.T. 

Baker, reagent grade or equivalent. 

5.1.3 Formic acid solution—88 

percent, Mallinckroft AR or equivalent. 

5.1.4 Liquinox, liquid 

detergent—Alconox Inc. or equivalent. 

5.1.5 Methanol—Burdick and 

Jackson, HPLC grade or equivalent. 

5.1.6 Methylene chloride—

Burdick and Jackson, pesticide grade or 

equivalent. 

5.1.7 Trifluoroacetic acid 

anhydride (TFA)—Pierce Chemical, Inc., 

reagent grade or equivalent. 

5.1.8 Water, organic-free—

Deionized and distilled water that is free from 

interfering organic compounds and chlorine. 

5.1.9 Ammonium acetate—J.T.

Baker, reagent grade or equivalent. 

5.1.10 Sodium chloride—EM 

Science, reagent grade or equivalent. 

5.1.11 Sodium hydroxide 

pellets—Reagent grade or equivalent. 

5.1.12 Isopropyl alcohol—

Burdick and Jackson, HPLC grade or 

equivalent. 

5.2 Solutions 

5.2.1 SPE cartridge conditioning

solutions 

5.2.1.1 Solution 1—80-percent 

methylene chloride/20-percent methanol. Mix

400 mL of methylene chloride with 100 mL

of methanol. Store in a calibrated adjustable

dispenser. Replace solution at least weekly. 

5.2.1.2 Solution 2—Ascorbic acid 

solution. Dissolve 10 g of ascorbic acid 

(5.1.2) in 1 L organic-free water and mix. 

Refrigerate at 4°C and replace solution at 

least weekly. 

5.2.2 SPE cartridge elution 

solution—80-percent methylene chloride/20-

percent methanol/0.2-percent trifluoroacetic 

acid anhydride (TFA). Mix 400 mL of 

methylene chloride, 100 mL methanol, and 

ANALYTICAL METHOD 11
1,000 μL of TFA (5.1.7). Prepare solution 

daily and store it in AutoTrace reservoir. 

5.2.3 Eluent neutralization 

solution—10 percent w/v sodium hydroxide 

solution. Weigh 10 g of sodium hydroxide 

pellets (5.1.11). Dissolve sodium hydroxide 

pellets in 100 mL organic-free water and mix. 

5.2.4 AutoTrace workstation 

cleaning solution (Liquinox detergent 

solution)—Dilute four drops of Liquinox

(5.1.4) with 4 L of organic-free water and 

mix. 

5.2.5 Concentrated ammonium 

acetate in organic-free water solution—

Weigh 20 g ammonium acetate (crystalline) 

(5.1.9) into a 1-L flask. Dissolve the 

ammonium acetate with organic-free water 

(5.1.8) and dilute to volume in the flask. 

Filter the solution through a 47-mm, 0.2-μm 

nylon filter membrane into a clean, burned 

filter flask. Transfer solution into a clean and 

burned 1-L bottle. The final concentration is 

20 g/L (0.26 molar). Store at room 

temperature. 

5.2.6 Concentrated ammonium 

acetate in methanol and acetonitrile 

solution—Weigh 20 g ammonium acetate 

(crystalline) (5.1.9) into a tared 1-L flask. 

Dissolve the ammonium acetate in the flask 

with 500 mL of methanol, then add 500 mL

of acetonitrile. Do not premix the methanol

and acetonitrile because ammonium acetate 

does not dissolve easily in the presence of 

acetonitrile. Use a stirring magnet and mixer 

with no heat to fully dissolve all ammonium 

acetate. Once it is dissolved, the ammonium

acetate will stay in solution. The final 

concentration is 20 g/L (0.26 molar). Store at 

room temperature. 

5.3 HPLC mobile phase preparation 

NOTE: The concentrations of formic acid in 

the mobile phase are varied from instrumentto-instrument and column-to-column to 

optimize chromatographic separation. The 

following concentrations are for reference. 

5.3.1 Organic solvent eluent 

(acetonitrile modified with ammonium acetate

solution)—Use a 7-mL class A pipet to 

measure 7 mL of the ammonium acetate in

acetonitrile:methanol solution (5.2.6), and 

transfer to a 500-mL volumetric flask. Dilute 

the solution in the flask to 500 mL with 

HPLC-grade acetonitrile (5.1.1) and mix. 

Filter the solution through a 47-mm, 0.2-μm 

nylon filter membrane into a clean, burned 

filter flask. Transfer to HPLC eluent reservoir

B for use. The final concentration is 0.28 g/L

(3.63 millimolar). 

5.3.2 Aqueous eluent (organicfree water modified with ammonium acetate

solution and formic acid)—Use a 7-mL class 

A pipet to measure 14 mL (in two aliquots) of 

the ammonium acetate in organic-free water 

solution (5.2.5), and place in a 1-L volumetric 

flask. Use a graduated cylinder to add 40 mL

of acetonitrile to the 1-L volumetric flask. 

Use a 500-μL syringe to add 200 μL of 

formic acid (5.1.3) to the solution in the flask. 

Dilute the solution in the flask to 1 L with 

organic-free water (5.1.8) and mix. Filter the 

solution through a 47-mm, 0.2-μm nylon filter 

membrane into a clean, burned filter flask. 

Transfer to HPLC eluent reservoir A for use. 

The final concentration of ammonium acetate 

in the solution is 0.28 g/L (3.63 millimolar). 

The final concentration of formic acid in the 

solution is 0.24 g/L (5.22 millimolar). 

12 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
5.4 Calibration and quality-control 

standard solutions 

In this method, each sample is analyzed 

twice, once for pesticides that preferentially

form negative ions and once for pesticides 

that preferentially form positive ions. 

Although the chromatographic conditions are 

identical for both analyses, it is important that 

the stock solutions used for calibration and 

quality-control standards be prepared

separately rather than in combination. There 

is a substantial possibility that interferences

from co-eluting compounds or crossreactivity among compounds will affect the 

identification and quantitation of method 

analytes if solutions are combined. This

problem also might be exacerbated by the 

high concentrations of the stock solutions. 

All calibration and quality control (QC)

standard solutions are derived from three 

common mixed standards, mixed from the

high-concentration, single-component 

solutions (see 5.4.1 and 5.4.2 below). These 

solutions are referred to as positive mix 1, 

positive mix 2, and negative mix. The purpose

of using these solutions for all subsequent 

calibration and QC standard solutions is to 

reduce potential discrepancies in interpreting

QC results. 

The overall approach is to prepare three 

20,000-μg/L multicomponent solutions that 

together include all compounds in the method 

except for surrogates and internal standards. 

From these solutions, working standards are 

made that contain the appropriate positive and 

negative surrogate compounds. The 

calibration solutions are then derived from the

working standards. The specifications for the 

solutions are contained in the following

sections and should be used if an external 

(commercial) source of standard solutions is 

used for this method. Individual singlecomponent stock solutions (at about 1 to 10 

mg/mL) are prepared from neat material or 

purchased at a known purity from a 

commercial source. After formulation, all 

solutions, except the calibration standards 

made prior to analysis, are stored in a freezer 

in labeled, amber 20-mL glass vials with 

Teflon-faced, silicone rubber-lined screw 

caps. 

5.4.1 Calibration solutions for 

compounds determined under negative 

ionization conditions—For compound 

mixtures determined under negative 

ionization conditions, single compound stock 

solutions and multicomponent stock and 

calibration solutions are prepared with HPLCgrade methanol (5.1.5). Prepare individual 

stock solutions of 10 mg/mL by dissolving 50 

mg of the selected pesticides in a 5-mL

amber-glass volumetric flask and dilute to 

volume using methanol to dissolve each 

compound. For negative ionization analysis, 

prepare a multicomponent stock solution for 

each listed compound by calculating the 

aliquot of each individual stock solution 

necessary to produce a final concentration of 

20.0 ng/μL, calculated as follows: 















 

ss

f

ss C

V

V C f (1) 

where Vss = the stock solution volume 

used, in microliters, 

 typically 200 μL;

Cf = the final solution concentration (for this solution 20.0 

 ng/μL≡20,000 μg/L);

Vf = the final solution volume 

(for this solution, 100 mL 

 100,000 μL); and 

Css = the stock solution concentration (for this solution, 10 

 mg/mL 10,000 ng/L). 

Use a variable-volume microdispenser 

(see 4.6.2) to add the calculated aliquot of each 

compound to a 100-mL amber-glass volumetric 

flask. Dilute the combined compounds to 

volume with methanol. Prepare a new primary

fortified standard solution every 6 months. 

ANALYTICAL METHOD 13
Store all solutions in a refrigerator in cleaned

and burned amber glass vials or bottles with 

Teflon-lined screw caps. For all compounds 

except triclopyr and chlorothalonil, single

component stock solutions are usable for no 

more than 12 months. For triclopyr and 

chlorothalonil, single component stock 

solutions are usable for no more than 3 

months. The components of the negative ion 

solution are listed in table 2. 

5.4.2 Preparation of 

multicomponent standard solutions for 

compounds determined under positive 

ionization conditions—Calibration, 

calibration verification, lab reagent spike 

fortification, and field matrix spike 

fortification solutions are prepared by diluting

high-concentration (20,000 g/L),

multicomponent standard solutions. These 

solutions are prepared from individual 

compound stock solutions, made up in one or 

more solvents, and are listed for each single

compound in table 3. Prepare individual 

stock solutions of 10 mg/mL by dissolving 50 

mg of the selected pesticides in a 5-mL

amber-glass volumetric flask and dilute to 

volume by using methanol or appropriate solvent 

to dissolve the compound (listed in table 3). 

Two multicomponent calibration 

solutions are prepared for compounds 

determined under positive ionization 

conditions. The first solution contains all the 

positive ionization compounds except for 

deethyldeisopropylatrazine and 2-hydroxyatrazine. The second solution contains only

deethyldeisopropylatrazine and 2-hydroxyatrazine. Two separate solutions are prepared 

because deethyldeisopropylatrazine and 2-

hydroxyatrazine require acid to dissolve the 

compounds in the stock solutions, and, if 

combined into the primary solution, would 

have potential for degradation of other 

compounds in the primary solution. There 

also is potential for cross-reactivity with other

compounds in the primary mixed standard 

solution. For positive ionization analysis, 

prepare two multicomponent stock solutions 

by calculating the aliquot of each individual 

stock solution necessary to produce a final 

concentration of 20.0 ng/μL, calculated as 

indicated in equation (1) and repeated below: 

Table 2 Calibration solution composition for compounds determined under negative ionization conditions

[CAS, Chemical Abstract Services] 

Compound name CAS number

Stock solution

solvent

2,4-D 94-75-7 Methanol

2,4-DB 94-82-6 Methanol

Acifluorfen 50594-66-6 Methanol

Bentazon 25057-89-0 Methanol

Bromoxynil 1689-84-5 Methanol

Chlorothalonil 1897-45-6 Methanol

Clopyralid 1702-17-6 Methanol

Dacthal monoacid (Monomethyl tetrachloroterephthalate) 887-54-7 Methanol

Dicamba 1918-00-9 Methanol

Dichloprop 120-36-5 Methanol

Dinoseb 88-85-7 Methanol

MCPA (4-Chloro-o-tolyloxyacetic acid) 94-74-6 Methanol

MCPB 94-81-5 Methanol

Picloram 1918-02-1 Methanol

Triclopyr 55335-06-3 Methanol

14 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 3 Calibration solution composition for compounds determined under positive ionization conditions

[CAS, Chemical Abstract Services; %, percent] 

Compound name CAS number Stock solution solvent 

Positive Solution Mixture 1

2,4-D methyl ester 1928-38-7 Methanol

3(4-chlorophenyl)-1-methyl urea 1897-46-6 Methanol

3-Hydroxycarbofuran 16655-82-6 Methanol

3-Ketocarbofuran 16709-30-1 Acetone

Aldicarb 116-06-3 Methanol

Aldicarb sulfone 1646-88-4 Methanol

Aldicarb sulfoxide 1646-87-3 Acetonitrile

Atrazine 1912-24-9 Methanol

Bendiocarb 22781-23-3 Methanol

Benomyl 17804-35-2 Acetone/Methanol (75:25)

Bensulfuron-methyl 83055-99-6 Dichloromethane

Bromacil 314-40-9 Methanol

Caffeine 58-08-2 Methanol

Carbaryl 63-25-2 Methanol

Carbofuran 1563-66-2 Methanol

Chloramben methyl ester 7286-84-2 Methanol

Chlorimuron-ethyl 90982-32-4 Acetonitrile

Cycloate 1134-23-2 Methanol

Deethylatrazine 6190-65-4 Methanol

Deisopropylatrazine 1007-28-9 Dichloromethane

Diphenamid 957-51-7 Methanol

Diuron 330-54-1 Methanol

Fenuron 101-42-8 Methanol

Flumetsulam 98967-40-9 Methanol/Acetone (50:50)

Fluometuron 2164-17-2 Methanol

Imazaquin 81335-37-7 Dichloromethane/Methanol (60:40)

Imazethapyr (Pursuit) 81335-77-5 Acetone 

Imidacloprid 138261-41-3 Methanol

Linuron 330-55-2 Methanol

Metalaxyl 57837-19-1 Methanol

Methiocarb 2032-65-7 Methanol

Methomyl 16752-77-5 Methanol

Methomyl oxime 13749-94-5 Methanol

Metsulfuron-methyl 74223-64-6 Methanol

Neburon 555-37-3 Methanol

Nicosulfuron (Accent) 111991-09-4 Acetonitrile/Methanol/Acetone (50:25:25) 

Norflurazon 27314-13-2 Methanol

Oryzalin 19044-88-3 Methanol

Oxamyl 23135-22-0 Methanol

Oxamyl oxime 30558-43-1 Dichloromethane/Methanol (60:40) 

Propham 122-42-9 Methanol

Propiconazole (Tilt) 60207-90-1 Methanol 

Propoxur (Baygon) 114-26-1 Methanol

Siduron 1982-49-6 Methanol

Sulfometuron-methyl 74222-97-2 Acetonitrile/Dichloromethane (50:50)

Tebuthiuron 34014-18-1 Acetone

Terbacil 5902-51-2 Methanol

Tribenuron-methyl 101200-48-0 Acetonitrile

Positive Solution Mixture 2 

2-Hydroxyatrazine 2163-68-0 Methanol/Acetone / Hydrochloric Acid (aq) [36%] (49.9:49.9:0.2) 

Deethyldeisopropylatrazine 3397-62-4 Methanol/Acetone / Hydrochloric Acid (aq) [36%] (49.9:49.9:0.2) 

ANALYTICAL METHOD 15














 

ss

f

ss f C

V

V C (2)

where Vss = the stock solution volume

used, in microliters, 

 typically 200 μL;

 Cf = the final solution 

concentration (for this 

solution, 20.0 ng/μL); 

 Vf = the final solution 

volume (for this 

solution, 100 mL

 100,000 μL); and 

 Css = the stock solution 

concentration (for this 

solution, 10 mg/mL 

 10,000 ng/L). 

Prepare each positive and 

negative multicomponent standard solution 

separately by using a variable-volume 

microdispenser (4.6.2) to add the calculated 

aliquot of each compound to a 100-mL

amber-glass volumetric flask. Dilute the

combined compounds to volume with 

methanol. Prepare new primary fortified 

standard solution for each mixture every 6 

months. Store the solutions in a cleaned and 

burned amber-glass bottle with Teflon-lined

screw caps. For all compounds except 3-

hydroxy- carbofuran, single component 

stock solutions are usable for no more than 

12 months. For 3-hydroxycarbofuran, single 

component stock solutions are usable for no 

more than 3 months. The components of the 

two multicomponent positive solutions and 

the appropriate solvents for the single component stock solutions are listed in table 3. 

5.4.3 Surrogate solution—A 

single surrogate solution is used for both 

ionization modes. The surrogates used for 

each analysis are listed in table 4. Individual 

surrogate stock solutions are made up in 

HPLC-grade methanol to a concentration of 

10 mg/mL. The multicomponent surrogate 

solution is made up in HPLC-grade 

methanol at a final concentration of 20 

ng/L of each listed compound. All 

solutions are transferred to and stored in a 

cleaned and burned amber-glass bottle with 

Teflon-lined screw caps. Stock solutions for 

the individual surrogate compounds are 

usable for no more than 12 months. 

5.4.4 Laboratory reagent spike 

fortification and field matrix spike 

fortification solutions—The same solutions 

are used for laboratory reagent and matrix

spikes. Prepare three separate laboratory

reagent spike fortification solutions 

(excluding positive and negative ionization 

surrogates) at a final concentration of 2.5 

ng/μL each. These solutions are prepared 

with a 100-mL volumetric flask, with 

methanol as the dilution solvent. The

aliquot of each individual compound to be 

used is calculated as follows: 





 



 

ss

t

ss C

V V Ct (3) 

where Vss = the aliquot volume required, 

 in microliters; 

Ct = the target concentration 

(in this solution, 2.5 ng/μL); 

Vt = the volume of target solution 

required, in milliliters 

(100 mL for this solution); and 

Css = the compound standard 

solution concentration, in 

 milligram/milliliter 

(1,000 ng/μL for this solution).

16 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 4. Surrogate solution composition for compounds determined under positive and negative 

ionization conditions 

[CAS, Chemical Abstract Services; %, percent] 

Compound name CAS number Stock solution solvent 

2,4,5-T (negative ionization analysis surrogate) 93-76-5 Methanol

Barban (positive ionization analysis surrogate) 101-27-9 Methanol 

13C-Caffeine (Caffeine (2-13C,99%:1,3-15N2,98%+); positive 

ionization analysis surrogate) 

Methanol

Use a variable-volume microdispenser (see 4.6.2) to add each compound to 

the volumetric flask. Bring the final solution 

to volume with methanol. Make three 

solutions, two for compounds determined 

under positive ionization conditions and one 

for compounds determined under negative 

ionization conditions (5.4.1). Addition of 100 

L of each of these solutions to 1.000 L of 

organic-free water will result in a

concentration of 0.25 g/L of each compound 

in the laboratory fortification sample. 

5.4.5 Internal standard 

solution—Prepare an internal standard 

solution of Monuron and dichloroacetic acid 

at concentrations of 0.1 and 1.0 mg/mL, 

respectively, according to the following

formula: 





 



 

ss

t

ss C

V V Ct (4) 

where Vss = the aliquot volume required, 

 in microliters; 

Ct = target concentration, 

typically 100 ng/μL;

Vt = volume of target solution 

required, in milliliters 

(100 mL for this solution); 

 and 

Css = the compound standard 

solution concentration, in 

 milligram/milliliter 

(1,000 ng/μL for this 

 solution). 

5.4.6 Calibration solutions—A 

set of quantitative calibration solutions is 

analyzed and a calibration curve is generated 

at the beginning of analysis to determine the 

concentrations of qualitatively identified 

compounds. To do this, prepare a series of 

working standard solutions, ranging in 

concentration from 100 to 100,000 ng/L, from 

the stock solutions prepared in sections 5.4.1 

and 5.4.2. At least five concentrations of 

calibration standards and a compound-free 

blank solution (system or instrument blank) 

are analyzed. Each concentration is prepared 

as needed by addition of a specified volume 

of the stock multicomponent standards to 

produce standards with concentrations 

equivalent to 0.010, 0.050, 0.10, 0.20, 0.50, 

and 1.00 g/L in water, assuming an injection 

volume of 50 L and a 1.00-L water sample. 

The preparation of these standard solutions is 

described in detail in the following sections. 

5.4.6.1 Working standard 

solutions for calibration standard 

formulation. Mix 25 mL of surrogate solution 

(5.4.3) with 25 mL of positive mix 1 and 

negative mix (5.4.1) in separate 50-mL

volumetric flasks. The concentration of this 

intermediate solution is 10,000 μg/L. Prepare 

the working standards for positive mix 1 and 

negative mix by using the 10,000-μg/L

intermediate solutions and the dilutions listed 

in table 5(A). Prepare working standards for 

positive mix 2 using the 10,000-μg/L

intermediate solution and the dilutions listed 

in table 5(B).

ANALYTICAL METHOD 17
5.4.6.2 Calibration standards. 

Calibration standards are prepared just prior 

to instrumental analysis. Measure 900 μL of 

organic-free water (5.1.8) into the number of 

vials required for the desired number of 

calibration standards. Add 100 μL of a 

working standard solution to each vial. Add 

5 μL of quantitation internal standard

solution (5.4.5), cap, and mix. Note that for 

ease of sample concentration determination, 

the calibration standard concentrations are 

expressed by an equivalent concentration in 

a 1,000-mL water sample (sample 

equivalence). The concentration of the 

calibration standards is calculated by using

the following formulae, in which the

dilution of a 5,000-μg/L working standard to 

a 0.5-μg/L (sample equivalence) calibration 

standard is listed: 

(A) 100 μL of working standard (5,000 μg/L) is added to 900 μL of organic-free water: 

5,000 μg/L x 100 μL x L/106 μL = 0.5 μg

0.5 μg/mL x 1,000 mL/L = 500 μg/L in vial 

(B) On-column mass of standard made in (A), when using a 50-μL injection: 

500 μg/L x 50 μL x L/106 μL = 0.025 μg on column

0.025 μg on column x 1,000 ng/μg = 25 ng on column

(C) Concentration of calibration standard (sample equivalence): 

25 ng x 1,000-μL vial/50 μL injected x 1/1,000 mL = 0.5 ng/mL 

0.5 ng/mL x 1,000 mL/L x 1 μg/1,000 ng = 0.5 μg/L 

(calibration standard's concentration equivalence in a 1-L water sample) 

Table 5. Volume dilutions for method working standards

[mL, milliliter; μg/L, microgram per liter; μL, microliter] 

Volumetric flask used (mL) Volume of 10,000 μg/L

standard used (μL)

Final concentration of 

working standard (μg/L) 

(A) Working standards for positive mix 1 and negative mix using the 10,000-μg/L solution

10 5,000 5,000

25 5,000 2,000

50 5,000 1,000

100 5,000 500

200 2,000 100

(B) Working standard dilutions for positive mix 2 using the 20,000-μg/L solution

10 5,000 10,000

10 2,500 5,000

25 2,500 2,000

100 2,500 500

18 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
5.4.6.3 Concentration of

calibration standards. Working standard 

concentrations and their sample equivalence 

are listed in table 6. 

Table 6. Working standard concentrations and 

equivalent sample concentrations for this method 

[μg/L, microgram per liter] 

Working standard

concentration 

(μg/L) 

Equivalent sample

concentration

(μg/L) 

10,000 1.00

5,000 .50

2,000 .20

1,000 .10

500 .05

100 .01

50 .005

5.4.7 Third-party check 

solution—An independently verified or 

"third-party" standard solution, containing the 

same or a subset of the compounds in the 

calibration standard solutions dissolved in 

methanol, can be used to determine 

acceptable performance of the instrument

calibration. This solution should not contain 

the method internal standards or surrogates. If

available, obtain this solution from a vendor 

that has prepared a solution and has 

independently validated it by instrumental 

analysis. Store this solution in a freezer at 

15°C, being sure to use the solution prior to 

the manufacturer’s expiration date. Select a 

concentration near the midpoint of the method 

calibration range (0.1 to 0.5 g/L). Use this 

concentration to determine the volume of 

solution required to fortify a 1-L water sample

to the chosen concentration. For example, the 

concentration of the components in thirdparty check solution used in this study was 

17.5 g/L. Add 10 L of the third-party

standard to 990 L of water in a 1.5-mL

amber-glass screw-cap autosampler vial and 5 

L of the internal standard solution to 

produce a per-component solution 

concentration of 0.175 μg/L. Then seal the 

vial with a Teflon-lined septum cap, and add 

to the sequence of sample, calibration, and 

quality-control samples. The concentration of

the third-party standard solution should be 

sufficiently high so that the solvent in the

aliquot that is diluted will not affect 

chromatographic separation. 

5.4.8 Instrument quality-control 

standards—The two instrument quality-control 

standards that monitor instrument performance 

over the course of analysis are continuing

calibration verification standards (CCVs) and 

continuing calibration blanks (CCBs). The 

CCVs are interspersed within a set of samples 

and are used to verify that the instrument has 

stayed within analytical calibration over the

course of analysis. The CCBs are monitored to 

determine if sample or standard injections 

caused cross-contamination. 

5.4.8.1 Continuing calibration 

verification (CCV) standards. Prepare a 0.2-

μg/L midcalibration-level-check sample by

adding an appropriate amount of the 

calibration standard for the ionization mode to 

be analyzed into an appropriate amount of 

organic-free water in an autosampler vial. 

Add 5 L of the internal standard (5.4.5), and 

seal with a Teflon-lined septum cap. 

Concentrations other than 0.2 μg/L may be 

used for CCVs if desired, as long as the 

concentration is near the midpoint of the 

calibration range. 

5.4.8.2 Continuing calibration 

blank (CCB) standards. Place 1,000 μL of 

organic-free water in an autosampler vial. 

NOTE: Do not add internal standard solution 

to the CCB. Seal with a Teflon-lined septum 

cap. 

6. Safety Precautions 

6.1 Use a well-vented fume hood for 

all steps involving organic solvents and acids. 

ANALYTICAL METHOD 19
6.2 Wear eye protection and the 

appropriate type of gloves when using any

reagents. 

6.3 Ensure that the electrospray waste 

exhaust tube and the vacuum pump exhaust 

tube of the mass spectrometer are vented out 

of the ambient laboratory atmosphere through 

ventilation ducting expressly specified for that

purpose. 

7. Procedure 

7.1 Sample filtration 

This method is applicable only to 

filtered water samples. All samples should be 

filtered in the field, preferably at the time of

collection. Filtration will reduce the 

likelihood of compound degradation by

removing particulate-associated bacteria. 

Removal of particulates also will prevent 

clogging of the retaining frit and stationary

phase of the solid-phase extraction cartridge, 

thus improving operation and extraction 

efficiency. Sandstrom (1995) describes a 

USGS-approved filtration method appropriate 

for samples analyzed by this method. 

Occasionally, samples are not filtered on site 

or become cloudy (particulate formation 

caused by chemical reactions or nanobacterial 

growth) during transit to the laboratory. Filter

these cloudy samples at the laboratory

according to the procedure outlined by

Sandstrom (1995) by using a 14.2-cm filter 

holder and positive pressure pump. Use a

0.7-μm pore size, 14.2-cm diameter, glassfiber filter, ashed at 440C for 2 hours. Flush 

the filtration apparatus with 100 mL of 

Liquinox solution (5.2.4), 100 mL of organicfree water (5.1.8), 50 mL of methanol (5.1.5), 

and again with 100 mL of organic-free water. 

Repeat this cleaning procedure between 

samples. Use a separate filter for each sample 

to prevent sample cross-contamination. 

7.2 Solid-phase extraction cartridge

cleaning and conditioning

NOTE: The extraction and elution procedure 

used in this method was designed to perform 

equally well by manual operation or by

automated SPE workstations. The same

cleaning and conditioning procedure is used 

for both. 

7.2.1 Prepare, as needed, the 80-

percent methylene chloride/20-percent

methanol (v/v) and 10-g/L aqueous ascorbic 

acid solutions for conditioning the SPE 

cartridges (5.2.1.1 and 5.2.1.2). These SPE 

cartridge conditioning solutions are prepared 

once a week because evaporation might alter 

the solvent composition. Store the ascorbic 

acid solution in the refrigerator at all times to 

prevent premature degradation of the acid. 

7.2.2 The performance of 

Supelco Graphitized carbon-based SPE (6- 

mL) cartridges may vary from lot to lot. Be

sure to use cartridges from a single lot for any

set of environmental and QC samples. Install 

the graphitized carbon-based SPE cartridges 

on the vacuum extraction manifold. The 

vacuum pump is used to pull the first two 

conditioning solutions through the cartridge. 

Do not exceed 20 mm Hg vacuum pressure, 

or the extraction chamber will implode. 

7.2.3 Cartridges are conditioned by

sequentially eluting the cartridge with two 

5-mL aliquots of 80-percent methylene 

chloride/20-percent methanol (v/v) (5.2.1.1) 

with the vacuum pump to facilitate elution. 

This step is followed by elution with 5 mL of 

methanol (5.1.5) through each cartridge using

the vacuum pump. Eluting three 5-mL aliquots 

of the ascorbic acid solution (5.2.1.2) under 

gravity flow completes conditioning. The 

ascorbic acid conditioning step should take 

place slowly to ensure activation of binding

sites in the cartridge bed. Keep the flow rate of 

ascorbic acid solution at no more than 3 

mL/min. Cover conditioned cartridges with foil 

and set aside until ready for use. Cartridges can

be used up to 8 hours after conditioning. 

Cartridges that have not been used within 8 

hours need to be reconditioned. Collect the 

20 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
conditioning solvents in the vacuum manifold; 

place these solvents in a chlorinated waste 

container for proper disposal. 

NOTE: To avoid deactivation of the SPE 

sorbent surface, cartridges should never be 

allowed to dry after conditioning. 

7.3 Solid-phase extraction

NOTE: The following description is for the 

automated SPE workstation method. This 

method can be carried out manually through 

the elution step by using the same conditions 

outlined in the following procedure.

7.3.1 Prior to extraction, the 

approximate pH of the sample is determined

by removing a small (0.05 mL) aliquot with a 

disposable Pasteur pipet (4.7.5) and applying

the volume to pH paper with a range of 0 to 

14, and recording the pH. Do not adjust the 

sample pH. Record the combined sample and 

bottle weight. Note any unusual appearance 

of the sample and record it. Add 100 μL of 

surrogate solution (5.4.3) and 1 g of sodium 

chloride (NaCl) to each sample. Shake 

samples well to dissolve the NaCl and 

uniformly disperse and mix the surrogate. 

7.3.2 Prepare laboratory reagent 

blank (LRB) and laboratory reagent spike 

(LRS) samples. Obtain two cleaned and 

burned 1-L amber bottles. Fill them with 

1,000 mL organic-free water and add 1 g of 

NaCl to each bottle. Shake well to dissolve 

the NaCl. In preparing the fortification 

sample, add 100 μL each of the three 

multicomponent matrix fortification solutions 

to one bottle containing 1,000 mL of reagent 

water. Record the solution code and bottle

preparation date of the fortification solutions 

(section 5.4.4). Add 100 μL of surrogate 

solution to each bottle (5.4.3). This step will 

result in a final concentration in the set 

fortification of 0.25 g/L. Shake the 

fortification and blank vigorously to mix the 

surrogate and NaCl in the water. Laboratory

reagent blank and reagent spike samples must

be prepared with each set of environmental

samples. A set of samples in this procedure

consists of the reagent spike and reagent

blank samples and up to 10 environmental 

samples. The environmental sample total 

may include duplicate field samples or field 

samples that are to be fortified in the

laboratory (laboratory matrix spike samples). 

Most analytes are stable under refrigeration 

for up to 4 days prior to extraction on the 

basis of results from on-going holding-time

studies. 

7.3.3 Clean the AutoTrace 

pumps and tubing prior to use by flushing

each AutoTrace position with sequential 

aliquots of 50 mL of Liquinox detergent 

solution (5.2.4), 50 mL of water (5.1.8), and 

50 mL of methanol (5.1.5). Pass nitrogen 

through the lines at about 103.4 kPa (15 

pounds per square inch) for about 5 minutes 

to ensure that all traces of methanol are 

removed. 

7.3.4 Install six conditioned, 6-

mL graphitized carbon-based SPE cartridges 

(4.7.2) on the AutoTrace SPE workstation. 

Attach one SPE cartridge connector and one 

of the 6-mL polypropylene syringe barrel 

adapters to each cartridge. Lower the 

plunger/cartridge clamp into the adapter, thus 

ensuring that the entire assembly fits snugly

in the apparatus. It is critical that the plunger 

is lowered fully and snugly into the adapter

because this snug fit, and the plunger O-ring, 

are the primary means for sealing the SPE

cartridge into the flow stream for SPE loading

and elution. Leaks caused by poor sealing

will adversely and irreproducibly affect 

method performance. 

7.3.5 Pump water samples 

through the conditioned cartridges by using a 

flow rate of 20 mL/min. Approximate 

extraction time for 1 L of sample is 50 

minutes. The AutoTrace workstation will emit 

an audible signal and suspend operation when 

extraction is complete.

ANALYTICAL METHOD 21
7.3.6 Upon completion of 

extraction, the SPE cartridges require drying. 

Two methods have been used with equal 

success. The first is to pass nitrogen gas 

through the cartridge, followed by chemical

drying of the cartridge with methanol. After

some experimentation, it was found that the 

nitrogen gas step could be omitted. This 

optional nitrogen gas-drying step is described 

in the following section. The methanol 

drying procedure is explained in section 7.3.8. 

7.3.6.1 Optional nitrogen gas 

drying. Place empty plastic test tubes in the 

AutoTrace eluent collection rack. Continue 

AutoTrace operation, and the programmed 

step for nitrogen flow will begin. Nitrogen

will flow at a pressure of 103 kPa for 1 

minute. Remove the empty polyethylene test 

tubes and dispose of the residual water.

Remove cartridges from the AutoTrace and 

attach to nitrogen manifold. Dry for at least 

15 minutes at 550 to 620 kPa. Return to 

appropriate positions on the AutoTrace. 

7.3.7 Weigh the empty sample 

bottle and record the weight. The difference

between this weight and the initial bottle

weight (7.3.1) provides the sample mass in 

grams, which is assumed to be equal to the 

sample volume in milliliters. Note that this 

procedure assumes that the volumetric density

of a typical freshwater sample is 1 g/mL. For

samples collected from saline environments, a 

salinity or density determination should be 

made and a volume correction applied. 

Occasionally a cartridge will clog, 

even if a sample has been filtered. This is 

likely the result of adsorption of coextracted 

natural organic matter onto the cartridge bed. 

In this event, the entire sample mass may not 

have been extracted. Weigh the bottle and 

remaining sample, discard the remaining

sample and re-weigh the empty bottle. 

Record these results and this condition. This 

information is required to accurately

determine sample concentration. 

7.3.8 SPE cartridge elution

7.3.8.1 After sample loading and 

the optional nitrogen gas drying step, 

cartridges retain a small amount ( 0.1 mL) of 

residual water. Chemically dry the cartridge

by eluting the analytes with 1.5 mL of 

methanol. Some compounds are eluted in this 

fraction. This 1.5-mL fraction is collected 

separately in unused polypropylene test tubes 

by placing six tubes positioned in the 

AutoTrace rack and labeled with the 

appropriate sample identifications. In

addition, this label should also include the 

letter “M” to indicate that this is the methanol 

fraction. Start the first step of the AutoTrace 

elution program to begin collection of the 

“M” fraction for all six tubes. When the 

AutoTrace has completed this first part of the 

elution, it will emit an audible signal. The last 

drops of methanol in the cartridge bed are 

removed by passing nitrogen gas through the 

cartridge at 103 kPa for 30 seconds. When 

the AutoTrace has completed this methanoldrying step, it will emit an audible signal. 

Remove the “M” fractions from the 

AutoTrace rack and bring the fraction to a 

volume of 1 mL with organic-free water. 

Refrigerate the extract until completion of

elution. 

7.3.8.2 Most of the analytes are 

collected in the second elution fraction of the 

SPE cartridge. Place unused polypropylene 

test tubes in the elution rack of the AutoTrace 

workstation to collect the second elution 

fraction. Label these tubes with the 

appropriate sample identifications. Indicate 

that these tubes hold the second elution 

fraction by labeling each tube with an “E”. 

Make sure the tubes are arranged in the 

correct order so that they will receive the 

eluent from appropriate SPE cartridges. Elute 

each SPE cartridge with three aliquots of 80-

percent methylene chloride/20-percent

methanol (v/v) with 0.2 percent trifluoroacetic 

acid anhydride (TFA) solution (5.2.2).

22 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
The three aliquot volumes are, 

in sequence, 5, 5, and 3 mL. This elution 

sequence is programmed for automated 

operation by the AutoTrace workstation. 

Eluting with three aliquots improves analyte 

recoveries compared to continuous elution 

with a single 13-mL aliquot. Start the “E” 

elution portion of the AutoTrace sequence. 

Note: If performing this method manually, do 

not let the cartridge beds dry. When the 

elutions of the three aliquots for each sample 

are completed, the AutoTrace will emit an 

audible signal. Remove the “E” extract tubes 

from the AutoTrace rack, cover with foil, and 

set aside in a refrigerator until it is time for

the concentration step. 

7.3.8.3 The fluid-flow paths of the 

AutoTrace workstation consist of 

polytetrafluoroethylene tubing, which may

adsorb nonpolar analytes. A postextraction 

cleaning of the AutoTrace workstation (or 

manual SPE extraction apparatus) is therefore

required and should be performed 

immediately after elution. Discard the SPE 

sample cartridges, and seal the empty adapter 

cartridges into the elution stations. Wash 

each of the six AutoTrace concentration and 

elution stations with 50 mL of Liquinox

detergent solution, 50 mL of water, and 50 

mL of methanol, at a flow rate of 20 mL/min. 

7.3.9 Concentration

7.3.9.1 Before concentration, add 

10 L of a 10-percent w/v sodium hydroxide 

solution (5.2.3) to the “E” fractions of the 

samples. This is added to neutralize the TFA 

in the eluent and reduce potential analyte 

degradation during volume reduction. 

7.3.9.2 The two fractions are 

reduced in volume under a nitrogen gas 

vortex stream. To reduce volume, place the 

“M” and “E” fraction tubes into the TurboVap 

sample concentration apparatus. Samples are

concentrated under a nitrogen gas stream of 

69 kPa (10 lb/in2

) while kept at 34C in a

water bath. The volume of the “E” fraction is 

about 13 mL, which nearly fills the 15-mL

volume of the test tube. The nitrogen gas 

pressure is therefore increased slowly from 0 

to 69 kPa, while monitoring the behavior of 

the fractions in the TurboVap. This prevents 

ejecting a portion of the “E” extract from the 

tube into the TurboVap water bath or into 

other samples, which might cause crosscontamination. When tubes are initially

placed in the TurboVap, they are spaced apart

as much as possible to minimize potential 

cross-contamination. TurboVap nozzles 

should be cleaned between samples. Note 

that the settings for bath temperature and 

nitrogen pressure are optimized for analyte

recovery within a reasonable total time for

extract volume reduction. Exceeding the 

specified temperature or pressure settings will 

adversely affect compound recoveries. 

7.3.9.3 The “M” fraction will take 

30 minutes to concentrate to about 

400 μL because of the volume of water in the 

fraction. Carefully monitor the fraction 

reduction and do not allow the extracts to dry,

or analyte recoveries will be adversely

affected. Take the “M” fraction out of the 

TurboVap after 30 minutes of concentration 

time and cover with foil. Set the “M” fraction 

aside in the refrigerator until the vialing

process is to begin. 

7.3.9.4 The “E” fraction will 

normally take 45 to 60 minutes to 

concentrate. Allow the “E” fraction to

concentrate for 30 minutes along with the 

“M” extract. After the “M” extract has been 

concentrated and removed from the 

TurboVap, continue to concentrate the “E” 

fraction for an additional 15 minutes. 

Concentrate the sample to about 400 μL. 

Carefully monitor the fraction reduction and 

do not allow extract volume to decrease to 

less than 400 μL, or analyte recoveries will be

adversely affected. If the extract has been 

concentrated for 60 minutes and the extracts

ANALYTICAL METHOD 23
are greater than 400 μL, however, there may

be residual methylene chloride in the “E”

extract. If this occurs, go to step 7.3.9.5. If

there is no residual methylene chloride in the

“E” extract, then the concentrated sample is

ready to be combined with the “M” extract

and to be placed in a sample vial for 

instrumental analysis. 

7.3.9.5 If the “E” fraction 

contains methylene chloride, it must be 

removed to avoid altering the chromatographic separation of analytes and decreased 

analyte recoveries. Typically, methylene 

chloride is present as an immiscible layer

underneath the aqueous phase or as small 

bubbles at the bottom of the test tube, or as a 

cloudy or “milky” appearance. As this 

description indicates, the presence of 

methylene chloride in a sample may be barely

perceptible, so each sample must be carefully

examined for the presence of methylene 

chloride. If a test tube is determined to 

contain methylene chloride, evaporate the 

volume in the test tube to about 400 μL. Then 

add about 200 to 300 μL of organic-free water 

and mix with a vortexing mixer. Evaporate 

the liquid in the test tube down to 400 μL 

again. Repeat this process until there is no 

more methylene chloride in the test tube. 

Note for each sample where treatment to

remove residual methylene chloride is 

required. 

7.3.10 Sample transfer into vials 

and preparation prior to analysis

7.3.10.1 Use clean and preashed 1.5-mL amber screw-top autosampler 

vials. 

7.3.10.2 Label each vial with 

appropriate lab identification and set number. 

This information is important for evaluating

individual sample results by comparison to set 

quality-control samples (duplicates, set 

blanks, set reagent fortifications). 

7.3.10.3 Pair the “M” and “E” 

extracts for each sample. Briefly swirl the 

“M” fractions with a vortexing mixer to wash 

the test tube walls. Transfer the contents of 

the “M” extract tube into the “E” extract tube 

by using a silicone-rubber bulb and a separate 

cleaned and burned disposable Pasteur pipet 

(4.7.5) for each sample. If the sample appears 

“cloudy,” methylene chloride may be present 

and the sample will need further

concentration to remove it (section 7.3.9.5). 

Mix the combined “M” and “E” fractions with

a vortexing mixer and transfer the extract into

the appropriate sample vial by using the same 

pipet that was used for combining the two 

fractions. Repeat this procedure for the entire 

sample set. 

7.3.10.4 Ensure that the final 

combined extract volume is about 1,000 μL 

by comparison to a vial with a known volume 

of liquid. An exact volume is not required 

because quantitation is by internal standard. 

If the final sample volume is less than 900 μL,

dilute to volume with organic-free water. Seal

vials with screw cap by ensuring a tight seal 

but not so tight that the Teflon-lined septum is 

puckered or wrinkled, which can result in 

evaporation of the sample. Place the vials in

a vial tray, organized by set. Store sample 

extracts in a freezer at –14oC until analyzed. 

7.3.10.5 Just prior to analysis, 

inject 5 μL of the internal standard solution 

(5.4.5) into each vial through the septa, using

a dedicated 10-μL syringe (4.6.1). The 

internal standard is added to all samples, 

including the LRS and the LRB. The internal 

standard solution contains the internal 

standards for both positive and negative

analysis, and is only added once. The same 

volume of internal standard is added to the 

instrument calibration solutions and 

continuing calibration verification solutions 

analyzed with the environmental samples. 

These samples and the sequence they are 

analyzed in are discussed in section 7.4.2.5.2. 

24 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
7.4 Instrumental analysis 

7.4.1 Instrumental analysis

overview—The analytes contained in sample 

extracts are separated by HPLC with a 

reverse-phase octadecylsilane column and 

water:acetonitrile gradient elution. The 

separated components are transported in a 

flowing stream to the electrospray ionization 

interface. In the interface, compounds, 

solvent, and any coeluting components of the 

sample matrix are nebulized into small 

droplets and desolvated. During desolvation, 

compounds are ionized by charge adduction, 

ion evaporation, or a combination of 

protonating processes (Kebarle and Ho, 

1997). The ionized compounds are separated 

from the neutral compounds by means of a 3 

kV potential difference between the 

nebulizing electrospray needle and the 

aperture leading to the mass analyzer. The 

nebulized neutral compounds do not enter the 

mass spectrometer, but rather they condense 

with the solvent and drain to a waste

reservoir. 

The ions transit through the

capillary aperture, and, as they exit the 

capillary to a reduced (1 to 2 millitorr) 

pressure region, are subject to an accelerating

voltage (called a fragmentor voltage or

capillary exit voltage by some manufacturers). This accelerating voltage provides 

the ions with sufficient momentum to induce

fragmentation by collision with neutral 

nitrogen gas molecules. The ionized 

fragments are swept from this low-pressure

region to the mass spectrometer by

momentum from a pressure difference 

between the low-pressure region and the 

analyzer region. Electrostatic lenses 

collimate the ions, and a quadrupole mass 

analyzer is used selectively to transport ions 

of a specific mass-to-charge ratio to a 

continuous electron multiplier. The current 

induced by the impact of the ion on the 

multiplier surface is amplified and transmitted 

as a voltage signal, with the number of ions 

proportional to the total voltage of the signal. 

Time-programmed, selected-ion monitoring is 

used to maximize detectability of specific

ions and reduce chemical noise from 

coeluting interferences. 

Coordinated, automated 

computerized programming is used to control 

most aspects of chromatographic separation, 

ionization, fragmentation, ion focusing, mass 

analysis, detection, and data handling. A 

typical separation of a standard mixture of the 

compounds determined under positive 

ionization conditions is shown in figure 1. 

Note that coeluting peaks are not

distinguished because this is a reconstructed 

ion chromatogram of selected-ion monitoring

results. These coeluting peaks would be 

separated and identified by using Target 

Software or equivalent automated graphic 

data-handling software. A similar

chromatogram can be produced for 

compounds eluting under negative ionization 

conditions. 

7.4.2 High-performance liquid 

chromatographic separation 

7.4.2.1 Sample vials are placed in 

the autosampler of either a Hewlett-Packard 

1090 Series II HPLC or a HewlettPackard/Agilent Technologies Series 1100 

HPLC. The autosampler temperature is kept 

at 4oC either by recirculating fluid chiller (for

the HP 1090 Series II HPLC; Neslab 

Coolflow CFT-33 or equivalent) or by a 

Peltier cooling unit (Hewlett-Packard/Agilent 

Technologies 1100 Series HPLC

autosampler). A 50-μL aliquot of the sample 

extract is injected into the HPLC eluent 

stream to start separation. 

ANALYTICAL METHOD 25
26 Pest

icides in Water by Graphitized CarbonBased SPE and HPLC/MS

Figure 1. Chromatogram of a positive ion standard used in this method.
7.4.2.2 The analytical separation 

for the method is carried out by using a 

reverse-phase octadecylsilane column 

(MetaChem Technologies Inertsil ODS-3, 

particle size of 5 μm, column dimensions of 

2 mm inside diameter by 150 mm long or 

equivalent). An integral guard column 

(MetaChem Technologies Metaguard 

Inertsil ODS-3, particle size of 5 μm, 2 mm 

inside diameter or equivalent) also is used. 

7.4.2.3 The separation is carried 

out using a binary eluent system of (1) 

organic-free water modified with 

ammonium acetate and formic acid (3.6 

millimolar ammonium acetate; 5.22 

millimolar formic acid; Mobile Phase A; see

section 5.3.2 for preparation), and (2) 

acetonitrile modified with ammonium

acetate (3.6 millimolar; Mobile Phase B; see 

section 5.3.1 for preparation). Initial HPLC

conditions follow: Autosampler, 4°C; 

column oven, 40°C; binary mobile phases 

(Mobile Phases A and B; sections 5.3.1 and 

5.3.2). The combined mobile-phase flow 

rate is constant at 0.20 mL/min. The 

mobile-phase gradient used for both positive

and negative ionization analyses is listed in 

table 7. Each HPLC analysis requires 85 

minutes to complete, including a postanalysis column re-equilibration period of

11 minutes. HPLC separation and mass 

spectrometric (MS) analysis are synchronized by computer control at the start of 

each analysis. For each HPLC/MS system, 

specific elution compositions and times 

are tested iteratively to achieve optimal 

separation, so the specific times and mobilephase compositions listed in table 7 should 

be used as a starting point for developing an 

acceptable separation. 

Table 7. High-performance liquid chromatograph 

mobile-phase gradient for this method 

Time, in 

minutes

Percentage

of mobile

phase A

Percentage 

of mobile 

phase B 

0.00 96.0 4.0

0.50 96.0 4.0

20.50 79.0 21.0

63.50 32.4 67.6

68.00 0.0 100.0

72.00 0.0 100.0

74.00 96.0 4.0

7.4.2.4 Mass spectral analysis 

parameters

7.4.2.4.1 Ionization source 

operating conditions—The mass 

spectrometer ionization source conditions 

for both positive and negative ionization 

analyses are listed in table 8. These 

conditions are held constant during the 

analysis. 

7.4.2.4.2 Programmable conditions 

during positive ion analysis—The computercontrolled aspects of mass spectrometer 

operation during positive ion analysis are listed 

in table 9a. These time-programming conditions 

are synchronized with the HPLC programming

at the start of each analysis. 

Table 8. Mass spectrometer operating conditions during positive and negative ionization analyses used 

in this method 

Nitrogen dry gas temperature 350 degrees Celsius

Drying gas flow rate 10.0 liters per minute 

Nebulizer gas pressure 212 to 414 kilopascals [30 to 60 pounds per square 

inch (gas); optimized for each instrument] 

Potential difference between nebulizer and capillary -3,500 volts (polarity of voltage reverses for negative ions) 

ANALYTICAL METHOD 27
Table 9a. Mass spectrometer time-programmed operating conditions for individual compounds

determined under positive ionization conditions

[m/z, mass-to-charge ratio] 

Time,

in 

minutes

Selected-ion

group1

Selected-ion

monitoring ion 

mass, in m/z

Typical 

electrometer 

gain 

Fragmentor

voltage, 

in volts 

Individual 

ion dwell 

time, in 

milliseconds 

1.00 1 142 2 70 400

7.00 2 106, 108, 132, 133 

163, 164, 185,207 

2 50 48

16.11 3 106, 163, 174, 175 

176, 195, 196, 198 

199, 220, 223, 237 

238, 240 

2 40 27

22.70 4 146, 160, 161, 163 

165, 166, 175, 181 

188, 190, 192, 198 

199, 209, 220, 256 

312, 313, 326, 348 

2 80 18

33.71 5 100, 116, 128, 151 

167, 168, 172, 179 

185, 187, 205, 213 

229, 230, 236, 290 

291, 312, 313, 411 

412, 413 

2 70 16

38.80 6 144, 161, 163, 199 

201, 205, 207, 290 

291, 312 

2 90 43

41.92 7 111, 145, 146, 153 

165, 167, 168, 202 

216, 217, 218, 220 

222, 223, 224, 233 

234, 248, 280, 290 

291, 365, 366 

2 60 15

49.79 8 120, 134, 138, 149 

155, 182, 220, 222 

240, 241, 290, 291 

304, 305, 306, 396 

411 

2 80 22

53.63 9 121, 122, 137, 160 

169, 186, 213, 233 

234, 249, 251, 415 

2 75 31

57.79 10 160, 249, 251, 252 

 253, 254 

2 35 65

60.56 11 114, 143, 160, 178, 

249, 251, 275 

277, 305, 342, 343 

344, 347 

2 85 29

67.03 12 134, 154, 216 2 80 132 

1

Selected-ion group number refers to specific-mass charge ratios monitored in a time interval.

28 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
7.4.2.4.3 Programmable 

conditions during negative ion analysis—

The computer-controlled aspects of mass 

spectrometer operation during negative ion 

analysis are listed in table 9b. These timeprogramming conditions are synchronized 

with the HPLC programming at the start of

each analysis. 

7.4.2.5 Mass spectrometer 

tuning and calibration 

7.4.2.5.1 Mass spectrometer 

autotuning—Prior to any analysis, the mass 

spectrometer is brought to temperature and 

gas pressure equilibrium, then tuned to 

ensure accurate mass assignment and a 

minimum detector response. An automated 

tuning (autotuning) procedure is used, with 

proprietary tuning solutions (provided by the 

instrument manufacturer) for positive and 

negative ion analysis. The autotune 

procedure uses a proprietary algorithm that 

combines the gain and voltage applied to the

instrument electron multiplier to produce the 

minimal acceptable ion current across the 

mass range of the mass spectrometer.

The ions that must be present are 

listed in table 10, and the target peak width 

for acceptable tuning in positive and 

negative ion modes is listed in table 11. 

Note that for both positive and negative ion 

modes, the autotune mass axis calibration 

must be within 0.13 atomic mass units 

(amu). In addition, note that although a 

typical electrometer gain is listed in tables

9a and 9b, the appropriate electrometer gain 

varies with the condition and age of the 

electron multiplier in the mass spectrometer,

and the electrometer gain listed should be 

used as a starting point. 

Table 9b. Mass spectrometer time-programmed operating conditions for individual compounds 

determined under negative ionization conditions

[m/z, mass-to-charge ratio] 

Time, 

in 

minutes

Selectedion 

group1

Selected-ion

monitoring ion 

mass, in m/z

Typical 

electrometer 

gain 

Fragmentor

voltage, 

in volts 

Individual ion 

dwell time, in 

milliseconds 

7.00 1 146, 190, 192 2 45 192 

13.00 2 195, 239, 241 2 45 192 

18.00 3 175, 177 2 45 289

30.00 4 239, 240, 241 2 45 192 

37.00 5 141, 161, 195, 196 

198, 199, 201, 203 

218, 219, 221, 233 

235, 253, 255, 271 

273, 274, 275, 276 

278 

2 45 26

49.00 6 141, 143, 161, 163 

217, 219, 227, 239 

240 

245, 247, 316, 318 

360 

2 45 43

1

Selected-ion group number refers to specific-mass charge ratios monitored in a time interval.

ANALYTICAL METHOD 29
Table 10. Quantitation and confirmation ions used for the compounds determined 

in this method 

[Note that the absence of an ion indicates that fewer than three ions are used for quantitation and 

confirmation.] 

Compound 

Retention 

time, in 

minutes

Quantitation 

ion 

Primary

confirmation 

ion 

Secondary

confirmation 

ion 

Compounds analyzed under positive ion conditions

2,4-D methyl ester 57.47 252 254 

2-Hydroxyatrazine 20.89 198 199

3(4-chlorophenyl)-1-methyl urea 35.85 185 128 187

3-Hydroxycarbofuran 27.88 163 181 220

3-Ketocarbofuran 36.66 179 236 151

Aldicarb 35.51 116 213

Aldicarb sulfone 17.35 240 223 

Aldicarb sulfoxide 13.71 132 207 

Atrazine 45.67 216 218 217

Barban (Internal Std) 60.09 178 143 

Bendiocarb 42.82 167 224

Benomyl 24.57 192 160 161

Bensulfuron-methly 50.75 411 149 182

Bromacil 37.92 205 207

Caffeine 18.67 195 196

Caffeine C13 (internal standard) 18.66 198 

Carbaryl 45.14 145 202 146

Carbofuran 42.80 222 165 223

Chloramben methyl ester 50.19 220 222 

Chlorimuron-ethyl 55.05 415 186

Cycloate 67.94 216 134 154

Deethylatrazine 28.43 188 190 146

Deethyldeisopropylatrazine 5.11 142

Deisopropylatrazine 20.79 174 176 175

Diphenamid 50.94 240 241 134

Diuron 47.35 233 234

Fenuron 26.59 165 166

Flumetsulam 32.67 326 348

Fluometuron 45.56 233 234

Imazaquin 35.02 312 313

Imazethapyr 31.96 290 291

Imidicloprid 28.65 256 175 209

Linuron 54.59 249 251 160

Metalaxyl 46.62 280 248 220

Methiocarb 53.54 169 121 122

Methomyl 18.94 163 106

Methomyl oxime 10.51 106 108 

Metsulfuron-methyl 38.83 167 168

Monuron (surrogate) 38.68 199 201 

Neburon 62.47 275 277 114

Nicosulfuron 36.61 411 213

Norflurazon 48.98 304 306 305

Oryzalin 60.83 347 305

Oxamyl 17.97 237 220 238

Oxamyl oxime 12.06 163 185 164

30 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 10. Quantitation and confirmation ions used for the compounds determined 

in this method—Continued 

Compound 

Retention 

time, in 

minutes

Quantitation 

ion 

Primary

confirmation 

ion 

Secondary

confirmation 

ion 

Compounds analyzed under positive ion conditions—Continued

Propham 49.88 138 120

Propiconazole 61.33 342 344 343

Propoxur 42.10 168 111 153

Siduron 53.06 233 234 137

Sulfometuron-methyl 43.45 365 366

Tebuthiuron 36.47 229 172 230

Terbacil 38.91 161 163 144

Tribenuron-methyl 50.53 155 396

Compounds analyzed under negative ion conditions

2,4,5-T 47.38 253 255 195

2,4-D 42.07 219 221 161

2,4-DB 55.53 161 163

Acifluorfen 56.97 316 360 318

Bentazon 37.02 239 240 241

Bromoxynil 45.95 276 278 274

Chlorothalonil 62.65 245 247

Clopyralid 13.66 190 146 192

Dacthal, monoacid 42.08 273 271 275

DCAA 60.70 217 219

Dicamba 28.59 175 177

Dichlorprop 45.89 233 235 161

Dinoseb 62.17 239 240

MCPA 42.50 199 141 201

MCPB 55.89 141 143

Picloram 19.73 241 239 195

Triclopyr 44.13 196 198 218

Table 11. Autotune parameters for acceptable 

tuning criteria used in this method 

[m/z, mass-to-charge ratio; amu, atomic mass unit] 

Tune ion mass, 

m/z 

Target peak width,

amu

(A) Tuning paramaters for positive ion mode

118.08 0.65

622.03 .65

922.05 .65

1,521.95 .65

2,121.95 .71

(B) Tuning parameters for negative ion mode

112.99 0.65

601.98 .65

1,033.99 .65

1,633.95 .65

2,233.91 .74

The signal intensity of the system is 

further optimized by using the ion at a massto-charge ratio of 622.03 as part of the 

autotune process. A compensated gainvoltage calibration curve is produced, and the 

appropriate gain and electron multiplier

voltages produce a manufacturer-defined 

minimum ion abundance of 5 x 104

 to 1 x 106

(manufacturer’s arbitrary units). This process 

is automatic and is written to the autotune file. 

The autotune file also contains all the 

pertinent instrument settings for mass axis 

calibration and peak width. For other 

manufacturers’ HPLC/MS systems, the 

ANALYTICAL METHOD 31
appropriate manual or automated tuning

procedure is used to meet instrument 

specifications for calibration and minimal ion 

signal. Regardless of the HPLC/MS system 

used, electronic and paper copies of tuning

conditions should be stored chronologically to 

monitor long-term HPLC/MS system

performance, assist in determining if

maintenance or repairs are required, and allow 

comparison of samples analyzed over 

extended (greater than 6 months) periods. 

The mass spectrometer

performance needs to meet manufacturers'

specifications for peak width, mass axis, 

calibration, and minimum acceptable ion 

signal intensity. If it does, proceed to

calibrate quantitatively. If the performance is 

not acceptable, however, then diagnostic, 

preventative or corrective procedures might

be required. Manufacturer-supplied 

diagnostic procedures are used to identify and 

correct any autotune-identified problems. 

Following any corrections, repeat the 

autotune procedure to verify that the 

corrections result in acceptable instrument

performance. When acceptable mass 

spectrometer mass axis and signal intensity

have been achieved, as indicated by a 

successful autotune, the instrument can be 

calibrated for quantitative analysis. 

7.4.2.5.2 Quantitative

calibration—A multiple concentration 

calibration for quantitative analysis is carried

out for all compounds after acceptable mass 

spectrometer tuning is completed. The seven 

concentrations for this calibration are listed in 

table 6 of section 5.4.6.3 and are the same for 

positive and negative ion analysis. The seven 

calibration concentrations are analyzed 

sequentially, and calibration curves are 

produced. A minimum of four quantitation 

levels must be used to determine the

calibration curve, and the curve should have a 

correlation coefficient (r2

) greater than 0.995. 

Most compounds will exhibit linear 

calibration curves with acceptable r2 values, 

but some analytes (typically atrazine, deethylatrazine, deisopropylatrazine, deethyldeisopropylatrazine, and 2-hydroxyatrazine) 

require quadratic curve fitting to achieve

acceptable r2

 values. If one or two of the 

calibration curve standard levels are not used, 

there must be a legitimate reason for dropping

the point from the curve, such as an 

incomplete or inaccurate injection or some

evidence that standard solution quality has 

fallen below acceptable levels. Corrective 

actions, such as preparing new standards, also 

would be required. Note that two analytes, 2-

hydroxyatrazine and deethyldsisopropylatrazine, require separate calibration curves. 

A limited set of three to four calibration 

points is used because both compounds are 

always reported as estimates. This second 

calibration is calculated for positive ion 

analysis only.

If the initial calibration is acceptable, 

the set(s) of environmental samples, set qualitycontrol (QC) samples, and instrument QC 

samples are combined into a batch and 

analyzed. A batch typically consists of up to six

environmental sample sets of 10 samples each, 

the associated set QC samples, and instrumental

QC samples to monitor performance. Note that

analysis of the entire analytical batch can 

require up to a week or more of continuous 

instrument operation. As a result, QC data 

need to be reviewed during the analysis of the 

batch to ensure acceptable instrument operation 

throughout the analysis. 

Instrumental QC samples are

interspersed between environmental and set 

QC samples. The presence of interspersed 

instrumental QC samples allows use of part of 

the data collected in the batch analysis in case

of calibration problems or instrument failure

during the batch sequence. The first 

instrument QC sample type used to monitor

batch performance is the continuing

calibration verification (CCV) sample. The

CCVs ensure ongoing acceptable calibration 

performance during analysis of the batch. 

32 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
When CCV results vary by more than 30 

percent, data are reported as estimates; if the

CCV results vary by more than 40 percent for 

five analytes, a new calibration is required. A 

continuing calibration blank (CCB) follows 

each CCV, and is used to monitor possible 

cross-contamination between injections as a 

result of incomplete injection or insufficient 

injection-needle washing. Typical batch 

analytical sequences for positive and negative

ion analyses are listed in table 12. Note that 

this sequence includes analyses for producing

a calibration curve. The sequence would be 

adjusted if a calibration curve was not

required. Note also that the position of the 

reagent blank in the preparation set is varied 

to monitor for position-specific

contamination. As a result, the position of the 

set reagent blank can vary within the

instrument analytical sequence.

7.4.2.6 Evaluation of instrument 

analysis results

7.4.2.6.1 The QC and 

environmental sample data are reviewed to

ensure that they meet acceptance criteria upon 

completion or partial completion of the

analysis of a batch (partial completion in the

case of a CCV or CCB failure). These data 

are reviewed qualitatively and quantitatively

to ensure that (1) detected compounds are 

correctly identified, and (2) the quantified 

concentrations are correctly calculated. 

7.4.3 Qualitative

determination—Several criteria are used to

determine that a compound identification is 

qualitatively correct. Correct identification is 

based on (1) the time at which the apex of the

chromatographic peak elutes from the HPLC

into the MS, (2) the presence of up to three 

compound-specific ions in the selected-ion 

monitoring mass spectrum (table 10), and (3) 

the relative abundances of these ions in the 

selected-ion monitoring signal and mass 

spectra. These criteria are determined from

analysis of authentic standards, and are 

verified by analysis of standards in each 

batch, to compensate for long-term changes in 

the HPLC/MS system. Compounds are 

qualitatively detected when the following

criteria are met:

Retention time—The intensities of

the characteristic ions of a 

compound are at a maximum that 

should coincide within ± 0.1 

minute of the retention time of

the selected compound. In

addition, the maxima of the 

primary (quantification) ion and 

secondary and tertiary

(qualification) ions should be 

within 0.05 minute of each other. 

Matrix effects and sample-tosample pH variations can have a 

substantial influence on liquid 

chromatographic retention times, 

thereby resulting in substantial 

variations of absolute retention 

time reproducibility, which also 

can be compound-dependent. An 

absolute retention-time criterion 

is therefore evaluated in

comparison to previously

analyzed standards and samples, 

and with cognizance of known 

problems that can result in 

retention-time variations. 

Spectra—The identity of each 

compound is verified by

comparing the selected-ion 

monitoring spectrum of the 

suspected compound with a 

reference selected-ion monitoring

spectrum obtained from a 

standard for that compound 

analyzed in the same batch. Two 

criteria apply. First, the same two 

or three ions must be present in 

the reference and sample spectra, 

although there may be additional 

ions present if ions for additional 

compounds are collected 

ANALYTICAL METHOD 33
Table 12. Typical minimum sample analysis sequence for high-performance liquid chromatography/mass

spectrometry analysis in this method 

[*, indicates quality-control samples analyzed in positive ion analysis only; μg/L, microgram per liter] 

Injection number Quality-control or environmental-sample type

1 Continuing calibration blank (ultrapure solvent) 

2 0.005 μg/L concentration standard 

3 0.01 μg/L concentration standard 

4 0.05 μg/L concentration standard 

5 0.10 μg/L concentration standard 

6 0.20 μg/L concentration standard 

7 0.50 μg/L concentration standard 

8 1.00 μg/L concentration standard 

9 0.05 μg/L concentration atrazine-degradate standard*

10 0.20 μg/L concentration atrazine-degradate standard*

11 0.50 μg/L concentration atrazine-degradate standard*

12 1.00 μg/L concentration atrazine-degradate standard*

13 Continuing calibration blank (ultrapure solvent) 

14 0.175 μg/L concentration third-party check standard 

15 Environmental sample- #1

16 Environmental sample- # 2 

17 Environmental sample- #3

18 Environmental sample- #4

19 Environmental sample- # 5 

20 Environmental sample- #6

21 Environmental sample- #7

22 Environmental sample- #8

23 Environmental sample- #9

24 Environmental sample- #10 

25 Set quality control sample- #11 (typically set reagent blank, but can vary in sequence position)

26 Set quality control sample- #12 (typically set reagent spike) 

27 0.20 μg/L continuing calibration verification standard

28–39 Twelve sequence entries for environmental/quality control samples ( #13–#24) 

40 0.20 μg/L continuing calibration verification standard

41 Continuing calibration blank (ultrapure solvent) 

42–53 Twelve sequence entries for environmental/quality control samples (#25–#36) 

54 0.20 μg/L continuing calibration verification standard

55–66 Twelve sequence entries for environmental/quality control samples (#37–#48) 

67 0.20 μg/L continuing calibration verification standard

68 Continuing calibration blank (ultrapure solvent) 

69-80 Twelve sequence entries for environmental/quality control samples (#49–#60) 

81 0.20 μg/L continuing calibration verification standard

82–93 Twelve sequence entries for environmental/quality control samples (#61–#72) 

94 0.20 μg/L continuing calibration verification standard

95 0.20 μg/L concentration atrazine-degradate standard*

96 0.05 μg/L limit of quantitation standard

97 0.05 μg/L limit of quantitation atrazine-degradate standard*

98 Continuing calibration blank (ultrapure solvent) 

34 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
concurrently. Second, the areas of 

these ions, determined from 

integrated peak areas of mass 

chromatograms, must be within 20 

percent of the absolute ratios 

obtained on injection of a standard 

solution generated using the

conditions of this method. Meeting

these criteria for qualitative

identification requires careful

consideration. One must determine 

whether the abundances in the 

selected-ion profiles are appropriate 

and if the profiles have relative 

intensities that are consistent with 

the reference mass spectrum, or if

there are contributions to the relative

abundances resulting from 

interference. Experience and 

training are necessary to recognize 

the salient features of individual 

mass spectra and potential 

interferences. Exercise careful

judgment in making a qualitative

identification, given the variability

inherent in identifying compounds at 

low concentrations in environmental 

samples. 

7.4.4 Quantitative

determinations—When a compound is 

qualitatively determined to be present, a

quantitative determination of the compound 

concentration can then be made. The 

concentration of that compound will be based 

on the integrated area from the primary

quantitation ion of that compound, the 

regression line fitted to the initial calibration 

curve, the area of the internal standard in the 

sample, and the internal standard response 

factors relative to the internal standard

response factor from the calibration standards. 

This method typically uses linear-fitted 

curves. For atrazine and the atrazine 

degradates measured in this method, a 

quadratic fitted curve is used for more 

accurate concentration determination across 

the method calibration range. In practice, 

quadratic and linear curves provide equally

acceptable results, and a quadratic curve can 

be used for all compounds if found to be more 

practical. 

7.4.5 Analysis of dilutions—

Samples must be analyzed within the range of

the calibration curve. In environmental 

samples, compound responses that exceed the 

response of the highest standard in the 

calibration curve, 1.0 μg/L, should be brought 

within the range of the calibration curve by

diluting the extract, using the initial calculated 

concentration as a guide for determining the

appropriate dilution volume, and then 

reanalyzing. For example, an undiluted 

sample with an initial calculated 

concentration of 5.0 g/L could be diluted to 

10 percent of its original concentration, so a 

predicted concentration of the diluted extract 

should be within the calibration range of the 

method. 

8. Calculations 

In this method, the calculation of a final 

concentration of a polar organic compound 

(POC) in a filtered water sample requires 

multiple calculations, as follows. 

8.1 Calculate the relative response 

factors for each POC from the calibration 

analyses conducted in 7.4.2.5.2 by using a 

best-fit linear regression or quadratic fit 

model. Rearrange the equation of the linear 

form y=mx + b to m = (y-b)/x as follows: 

RRFc = 



































	


















	











is

c

is

c

amt

amt

b

area

area

 (5) 

ANALYTICAL METHOD 35
where RRFc = the relative response 

factor for the polar 

organic compound 

(POC) of interest; 

areac = the integrated peak area 

of the POC of interest; 

 areais = the integrated peak area 

of the positive or 

negative ionization 

internal standard used 

for the POC of interest; 

amtc = the mass of the POC of 

interest, in nanograms; 

amtis = the mass in nanograms

of the POC internal 

standard (see section 

5.4.5) used for the POC of 

interest; and 

b = the y-intercept of the 

best-fit linear regression 

line. 

NOTE: A similar calculation can be made

for fitted quadratic curve calibrations by

rearranging the equation y=ax

2

+bx+c, where 

a, b, and c are experimental constants 

determined from the fitted curve by iterative

mathematical extraction with curve-fitting

software. 

8.2 Calculate the volume of water 

extracted, in liters (Vs):

Vs = (Vi – Vf)/1,000 (6) 

where Vi = initial weight of sample 

and sample bottle, in 

grams (mL; 7.3.7); 

Vf = final weight of sample and 

sample bottle, in grams 

(mL; 7.3.7); and 

1,000 = conversion factor for 

milliliters to liters.

NOTE: This procedure assumes that the 

volumetric density of a typical freshwater 

sample is 1 g/mL. For samples collected from 

saline environments, a salinity or density

determination should be made and a volume 

correction applied (see 7.3.7). 

8.3 Calculate sample polar organic 

compound concentrations 

If the compound of interest has met the 

qualitative identification criteria listed in 

7.4.3, calculate the compound concentration 

in the sample as follows: 

C = 















c is s

is c

RRF A V

amt A

 

 

x x

 x (7) 

where C = the concentration of the 

compound of interest in 

the sample, in micrograms 

per liter; 

 amtis = the mass of internal standard 

added to the sample, in 

micrograms;

 Ac = the area of the quantitation 

ion for the compound of 

interest; 

 RRFc = the relative response factor 

for the compound of interest, 

calculated above in 8.1; 

 Ais = the area of the quantitation 

ion for the internal standard; and 

Vs = the volume of sample extracted, 

in liters, calculated in 8.2 

(equation 6).

8.4 Calculate the percentage recovery

of the surrogate compounds in each sample 

by using

Ra = 



 





Ca Va Vs

Cs

( x ) /

x 100 (8)

36 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
where Ra = recovery of surrogate in 

sample, in percent; 

Cs = concentration of surrogate 

in sample, in micrograms 

per liter, calculated 

by using equation 7; 

Ca = concentration of compound 

in the surrogate solution 

added to the sample, in 

micrograms per microliter 

(5.4.3); 

Va = volume of POC surrogate 

solution added to the sample, 

typically 100 μL (7.3.1); and 

Vs = volume of sample, in liters 

(calculated in 8.2). 

8.5 Calculate the percentage recovery

of compounds in set reagent spike sample 

by using

Rb = 

















b b s

s

C V V

C

( x ) /

x 100 (9) 

where Rb = recovery of fortified compound 

in the set POC fortification 

sample, in percent;

Cs = concentration of compound in 

set reagent spike sample, in 

micrograms per liter, 

calculated using equation 7; 

 Cb = concentration of compound in 

reagent spike fortification 

solution added to sample, in 

micrograms per microliter 

(5.4.4); 

Vb = volume of reagent spike 

fortification solution added 

to the sample, typically 

100 μL (7.3.2); and 

Vs = Set reagent spike sample volume, 

in liters (calculated in 8.2). 

9. Reporting of Results 

9.1 Reporting units—Report 

compound concentrations for field samples in 

micrograms per liter. Report compound 

concentrations for field samples to 4 decimal 

places, but no more than 3 significant figures. 

Report data for compounds reported as 

qualified estimates to 4 decimal places, but no 

more than 2 significant figures. Report 

surrogate data for each sample type as percent

recovered, and report to 1 decimal place 

(tenths of a percent), but no more than 3 

significant figures. Report data for the set 

fortification sample as percent recovered, and 

report to 1 decimal place (tenths of a percent), 

but no more than 3 significant figures. 

Compounds quantified in the set blank sample 

are reported in micrograms per liter, and are

reported to 4 decimal places, but no more than 

3 significant figures. 

9.2 Reporting limits and levels—

Method detection limits (MDLs) that use the 

procedures outlined by the U.S. 

Environmental Protection Agency (1997) 

have been calculated for this method and are 

discussed further in section 11.8. The interim 

laboratory reporting level (LRL) for each 

compound determined using this method is 

calculated according to Childress and others 

(1999) and is twice the method detection 

limit. Report qualitatively identified 

compound concentrations (those POCs that 

are identified from relative retention time and 

MS spectral fit) that are less than the MDL or

less than the lowest calibration standard as

estimated concentrations. Compounds that 

are not detected are reported as less than the 

interim LRL.

10. Quality Assurance/Quality Control 

Laboratory extraction samples are 

formed into sets, each consisting of 10 

environmental samples, a set reagent spike

and set reagent blank, for a total of 12 

ANALYTICAL METHOD 37
samples. Field equipment blanks and 

laboratory matrix spikes, whose frequency is 

determined by the method user, provide 

additional quality assurance/quality control 

(QA/QC). The frequency of analysis of these 

QA/QC samples and the aspects of the 

analytical process they monitor are described. 

10.1 Surrogate. Surrogates are organic 

compounds that are placed into all filtered 

water samples prior to extraction on the SPE 

cartridge. Surrogates are expected to behave 

similarly to selected compounds for SPE 

recovery and are not expected to be present in 

the environment. Three surrogates are used in

this method, two (13C-caffeine and Barban) 

for compounds determined under positive 

ionization conditions, and one (2,4,5-T), for 

compounds determined under negative 

ionization conditions. The herbicide 2,4,5-T 

had been previously used as a pesticide, 

although registrations for all uses in the 

United States have been canceled since 2 

January 1985 (U.S. Environmental Protection 

Agency, 2000). In 12,064 surface- and 

ground-water samples, 2,4,5-T was detected 

68 times using a similar analytical method 

(Werner and others, 1996) Given the limited 

number of appropriate negative ionization 

surrogates, 2,4,5-T was used as a surrogate 

because the frequency of environmental

presence was very low, and the frequency of 

detections in the future is likely to decrease

further. 

Examination of surrogate recovery for 

individual samples provides insight into 

overall method performance for that particular 

sample. Control limits, determined by using

statistical process control techniques and an 

extended sequence of laboratory reagent spike 

and reagent blank surrogate recoveries, are 

used to evaluate surrogate recoveries of 

individual samples. 

10.2 Laboratory reagent spike (LRS). 

A 1-L organic-free water sample is fortified at

0.25 μg/L for all compounds determined in 

this method. This sample then is included 

with each sample set and is carried through 

the entire extraction, elution, and analytical 

procedure. The LRS recoveries reflect 

method performance in the absence of any

environmental sample matrix. These results 

are used to determine if overall set recoveries

are acceptable, or if there was a gross change

in method performance in the set. 

Acceptability is defined from analysis of a 

series of LRS samples, typically 30 or more, 

processed by multiple operators, who used

different instruments. Statistical process 

control analysis is used with these data to 

develop acceptance criteria. 

10.3 Laboratory reagent blank (LRB). 

A 1-L organic-free water sample is fortified 

with method surrogates only. One LRB is 

included with each sample set and is carried 

through the entire extraction, elution, and 

analysis procedure. The LRB is used to 

monitor for impurities and contamination, 

and, because it follows the LRS in the sample 

instrumental analysis sequence, it also 

monitors for carryover between sample

injections. 

10.4 Continuing calibration verification 

(CCV). For each analysis type (positive 

ionization or negative ionization), a 0.25-μg/L

calibration standard that contains all of the 

selected compounds, including surrogates and 

internal standards, is inserted in an

autosampler vial and placed between every 12 

environmental and set QA/QC samples 

throughout the HPLC/MS analysis. These 

CCV samples are used to ensure that the 

calibration of the HPLC/MS system is within 

acceptable limits, typically 30 percent. If

the control limits are exceeded, environmental 

samples that follow the last acceptable CCV

are reanalyzed. Control limits for the CCV 

rarely were exceeded during the course of this

study. 

10.5 Continuing calibration blank

(CCB). A sample of organic-free water is

38 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
placed in an autosampler vial behind a CCV, 

with a typical frequency of one every third set 

of samples. The CCB follows the CCV, and 

thus monitors for potential injection-toinjection carryover, as well as instrumental

contamination. 

10.6 Limit-of-quantitation (LOQ)

standard. The LOQ standard is an aliquot of 

the lowest concentration calibration solution 

used to develop the calibration curve 

(typically 0.005 μg/L; see section 5.4.6.3). 

The LOQ is analyzed at the end of a sample

analytical sequence to verify that sufficient

instrument sensitivity has been maintained

throughout the sequence. 

10.7 Field equipment blank (FEB). A 

volume of organic-free water is processed 

exactly as environmental samples by using all 

appropriate on-site sampling equipment and 

techniques. This process includes bottles, 

compositing, splitting, and filtering. The FEB

is processed at the start of sampling and then

about every 15 to 20 samples. The FEB

monitors for contamination or carryover, or 

both, resulting from field sampling and 

equipment cleaning techniques that could 

cause equipment contamination of 

environmental samples. 

10.8 Field matrix spike (FMS). The 

FMS is a duplicate environmental sample that

is fortified at 0.25 μg/L for all compounds 

determined in this method. The unfortified 

duplicate is used to determine naturally

present concentrations of any compounds 

measured in the sample. If concentrations of 

method compounds are determined, they are 

subtracted from the measured concentrations 

in the fortified sample. The corrected

recoveries of method compounds are 

determined from the background 

concentration-subtracted results. The FMS 

measures the effects of the sample matrix on 

the recovery of method compounds. Several 

effects are possible, including matrixenhanced compound degradation, matrixintroduced coeluting interferences, and 

matrix enhancement of compound 

concentration. The frequency of FMS 

analyses is determined by data-quality

objectives. 

11. Method Performance 

This method was originally developed 

as a custom analytical method by the USGS 

and was put into routine use in June 1999 at 

the NWQL. As part of the initial custom 

method testing, compound recovery was 

determined in June 1999 in three water types 

at a fortified concentration of 0.100 g/L. 

The three matrices were laboratory-produced, 

pesticide-free, organic-free water; a groundwater sample from a private domestic well; 

and a surface-water sample collected from the 

South Platte River in Denver, Colorado. The 

surface- and ground-water samples were 

collected following the precautions suggested 

in Shelton (1994) for avoiding sample 

contamination. 

Upon acceptance of these results, the 

method was applied to surface- and groundwater samples collected across the United 

States from March 1999 to July 2000. 

Additional method validation samples were 

analyzed in February 2000 using the same 

water types and fortified at nominal

concentrations of 0.025 and 0.50 g/L during 

this period when the method was being used 

routinely for custom analysis. One 

compound, dichlorprop, which was not 

present in the initial 0.100 fortifications, was 

added to the spiking solutions after a suitable

standard was obtained. The combined 

performance studies at fortifications of 0.025, 

0.100, and 0.50 g/L are used to evaluate

method performance. Provisional MDLs

were determined using the initial organic-free 

water data, fortified at 0.100 g/L. Later

MDL determinations were made using the

results from organic-free water fortified at

0.025 g/L for organic-free water and ground 

water, and at 0.050 g/L for surface water. 

ANALYTICAL METHOD 39
As of July 2000, about 3,300 surfaceand ground-water samples were analyzed with 

this method. The results for set qualitycontrol samples (197 laboratory reagent 

blanks and 285 laboratory reagent spikes) 

were analyzed concurrently with these 

environmental samples, aggregated, and the 

data evaluated for long-term (multiple

instrument, multiple operator) method 

performance. 

11.1 Recoveries from fortified water 

samples 

11.1.1 The performance of this 

method for the extraction and analysis of 

POCs was evaluated by adding aliquots of 

standard solutions to a minimum of six water

samples and processing the fortified samples 

through the entire method. Two unfortified 

water samples were processed with each set

of fortified samples to determine the

concentrations of any POCs present in the 

water prior to spiking. Reagent spikes and 

unfortified laboratory reagent blank samples 

also were processed with each set. Three 

water types were used for evaluating method 

performance and are described below. 

11.1.2 The ground-water sample 

was collected from a single-family domestic

supply well near Evergreen, Colorado. Water 

was collected from the well after a sustained 

period of domestic use to minimize contributions of water that had been stored in a 

lined pressurization tank. The well penetrates 

85 m into a fractured rock aquifer with 

minimal overlying soil. This well was part of

a cooperative U.S. Geological SurveyJefferson County, Colorado, ground-water 

monitoring program near Evergreen, 

Colorado (Schwartz, 1997). Water was 

collected into a precleaned, 40-L stainlesssteel container and filtered in the laboratory

using the procedure described in Sandstrom 

(1995). Samples were collected sequentially

into individual, pre-ashed 1-L amber bottles 

for analysis. Water was collected several

times from this site over the course of the

study. Dissolved organic carbon (DOC) was 

not measured each time a water sample was 

collected at this site; instead, DOC 

concentrations were measured as part of the 

ground-water monitoring program described 

by Schwartz (1997). 

11.1.3 The surface-water samples 

were collected from the South Platte River as 

it passes through metropolitan Denver,

Colorado. The water quality of the South 

Platte River has been extensively studied; 

Litke and Kimbrough (1998) provide an 

overview. Grab samples of South Platte 

River water were collected in stainless-steel

containers of either 10- or 40-L capacity that 

had been washed with soap and water and 

sequentially rinsed with water and solvent. 

The water samples were filtered using the

procedure described in Sandstrom (1995), and 

split into individual 1-L aliquots in pre-ashed 

1-L amber bottles for analysis. Over the

course of the study, water was collected 

several times from two sites, one upstream

and one downstream of Denver. The DOC 

concentrations were not measured on the 

individual samples used in this study. Median 

DOC concentrations for the South Platte 

River from 1993–1995 were 5.2 mg/L at 

Denver and 7.0 mg/L at Henderson, Colorado, 

on the basis of measurements made as part of 

the Survey's National Water-Quality

Assessment (NAWQA) program. These data 

are summarized in Litke and Kimbrough 

(1998). 

11.1.4 The pesticide-free, organicfree water was produced at the NWQL using

a Solution 2000 water purification system 

(Model 2002AL, Solution Consultants, Inc., 

Jasper, Ga.). The following method was used. 

Laboratory-distilled water was introduced 

into a 1-μm activated carbon prefilter, passed 

through a series of ion exchange resin beds to 

remove dissolved inorganic constituents, 

followed by high-intensity UV radiation 

40 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
oxidation to remove dissolved organic carbon, 

and then filtered through a 0.22-μm sterile

filter. The organic-free water was dispensed 

into pre-ashed 1-L amber bottles for analysis. 

This water also was used for extraction set 

QC samples (laboratory reagent spikes and 

blanks). In these water samples, DOC was 

not determined, but routine monitoring of 

DOC in the water produced by this system did 

not detect DOC concentrations at the longterm method detection level of 0.016 mg/L. 

11.2 Organic-free water fortification 

recovery results

11.2.1 Sets of 10 samples of 

organic-free water were fortified with a 

laboratory reagent spike solution (5.4.4) at 

concentrations of 0.025, 0.100, and 0.500 

g/L. The results are listed in tables 13 to 18. 

The mean, median, and the standard deviation 

of recovery for all compounds in the tables 

were calculated from the data for mean

recoveries of individual compounds. No 

method compounds were detected in unspiked 

reagent water. 

11.2.2 The mean positive 

ionization recovery results for compounds 

reported without qualification (tables 13, 15, 

and 17) were 103.5  11.5, 92  10.4, and 

93.1  9.0 percent at fortifications of 0.025, 

0.100, and 0.500 g/L, respectively. The 

median recoveries were similar to the means, 

99.5, 88.0, and 93.0 percent, respectively. The 

mean positive ionization recovery results for

compounds reported as qualified estimates 

were 87.9 ± 31.6, 89.3 ± 42.3, and 82.0 ± 26.1 

percent at fortifications of 0.025, 0.100, and 

0.500 g/L, respectively. The median 

recoveries for qualified estimates also were

similar, at 94.0, 88.0, and 88.5 percent, 

respectively.

11.2.3 Mean recoveries for 

compounds determined by negative ionization 

and reported without qualification (tables 14, 

16, and 18) averaged slightly less than the 

positive ionization results, and were 76.9  5.4, 

84.7  6.4, and 75.8  17.4 percent, at 

fortifications of 0.025, 0.100, and 0.500 g/L,

respectively. The corresponding median 

recoveries were similar to the means, at 78.0, 

85.0, and 78.0 percent, respectively. For 

negative ionization compounds reported as 

qualified estimates, mean recoveries decreased

and were more variable, at 64.6  10.7, 59.2 

27.5, and 64.4  39.8 percent, respectively,

with corresponding median recoveries of 65.0, 

69.0, and 62 percent. 

11.2.4 Average recoveries of all 

compounds reported without qualification at 

all three fortification levels varied about 10.3 

percent for compounds determined by positive 

ionization and 9.7 percent for compounds 

determined by negative ionization. This 

result, however, does not reflect the analytical

precision, which is better reflected by the 

relative standard deviation of individual 

compound recoveries in organic-free water, 

particularly those that are reported without 

quantitation qualification. Under positive 

ionization conditions, the mean relative 

standard deviations for compounds reported 

without qualification in organic-free water 

were 8.9  4.8, 11  3.4, and 6.3  3.6 percent 

for 0.025-, 0.100-, and 0.500-μg/L fortifications, respectively. The corresponding

relative standard deviations for negative 

ionization results reported without qualification were 10.4  3.7, 13.1  3.6, and 13.5 

10.5 percent for 0.025-, 0.100-, and 0.500-

μg/L fortifications, respectively.

11.3 Ground-water fortification 

recovery results

11.3.1 Sets of 10 individual 

ground-water samples were fortified at 

concentrations of 0.025, 0.100, and 0.500 g/L

(tables 19 through 24). The mean, median, and 

the standard deviation of recovery for all 

compounds in the tables were calculated

ANALYTICAL METHOD 41
Table 13. Accuracy and precision data from 10 determinations of the method compounds fortified 

at 0.025 microgram per liter in organic-free water, under positive ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

2,4-D methyl ester N 0.0279 0.00136 5 111 0.0043 

2-Hydroxyatrazine Y .0241 .00126 5 96 .0040

3(4-chlorophenyl)-1-methyl urea N .0283 .00382 14 113 .0121 

3-Hydroxycarbofuran N .0236 .00092 4 94 .0029

3-Ketocarbofuran Y .0156 .00224 14 62 .0071

Aldicarb Y .0233 .00625 27 93 .0198

Aldicarb sulfone Y .0227 .00309 14 91 .0098 

Aldicarb sulfoxide Y .0231 .00130 6 92 .0041 

Atrazine N .0243 .00143 6 97 .0045

Bendiocarb N .0235 .00398 17 94 .0126

Benomyl N .0240 .00060 2 96 .0019

Bensulfuron-methyl N .0324 .00251 8 130 .0079

Bromacil Y .0265 .00514 19 106 .0163

Caffeine N .0236 .00150 6 95 .0048

Carbaryl N .0274 .00447 16 110 .0142

Carbofuran N .0235 .00090 4 94 .0028

Chloramben methyl ester Y .0130 .00282 22 52 .0089 

Chlorimuron-ethyl N .0293 .00153 5 117 .0048

Cycloate Y .0220 .00206 9 88 .0065

Deethylatrazine Y .0288 .00444 15 115 .0141

Deethyldeisopropylatrazine Y .0436 .00702 16 175 .0222

Deisopropylatrazine Y .0241 .00164 7 96 .0052

Diphenamid N .0220 .00418 19 88 .0132

Diuron N .0253 .00235 9 101 .0075

Fenuron N .0275 .00498 18 110 .0158

Flumetsulam Y .0242 .00180 7 97 .0057

Fluometuron N .0282 .00488 17 113 .0155

Imazaquin Y .0236 .00247 10 94 .0078

Imazethapyr Y .0244 .00265 11 98 .0084

Imidicloprid N .0242 .00107 4 97 .0034

Linuron N .0277 .00228 8 111 .0072

Metalaxyl N .0241 .00315 13 96 .0100

Methiocarb Y .0242 .00127 5 97 .0040

Methomyl Y .0234 .00071 3 94 .0022

Methomyl oxime Y .0088 .00166 19 35 .0053 

Metsulfuron-methyl Y .0237 .00387 16 95 .0123

Neburon N .0251 .00189 8 101 .0060

Nicosulfuron N .0244 .00205 8 98 .0065

Norflurazon Y .0263 .00259 10 105 .0082

Oryzalin N .0299 .00277 9 120 .0088

Oxamyl N .0222 .00194 9 89 .0061

Oxamyl oxime Y .0070 .00211 30 28 .0067 

Propham N .0260 .00153 6 104 .0048

Propiconazole N .0318 .00332 10 127 .0105

Propoxur N .0223 .00127 6 89 .0040

Siduron N .0283 .00266 9 113 .0084

Sulfometuron-methyl N .0233 .00140 6 93 .0044

Tebuthiuron N .0241 .00098 4 97 .0031

Terbacil Y .0231 .00156 7 92 .0049

Tribenuron-methyl Y .0079 .00140 18 32 .0044

42 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 14. Accuracy and precision data from 10 determinations of the method compounds fortified 

at 0.025 microgram per liter in organic-free water, under negative ionization 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

2,4-D N 0.0239 0.00334 14 96 0.0109

2,4-DB Y .0207 .00245 12 83 .0080

Acifluorfen N .0232 .00101 4 93 .0033

Bentazon Y .0153 .00170 11 61 .0055

Bromoxynil Y .0151 .00260 17 60 .0085

Chlorothalonil Y .0147 .00531 36 59 .0173

Clopyralid N .0258 .00213 8 103 .0069

Dacthal monoacid N .0257 .00179 7 103 .0058 

Dicamba N .0244 .00197 8 97 .0064

Dichlorprop N .0258 .00213 8 103 .0069

Dinoseb N .0064 .00184 29 26 .0060

MCPA N .0230 .00250 11 92 .0081

MCPB Y .0194 .00236 12 78 .0077

Picloram N .0230 .00303 13 92 .0099

Triclopyr N .0249 .00344 14 100 .0112

Table 15. Accuracy and precision data from seven determinations of the method compounds fortified at 

0.10 microgram per liter in organic-free water, under positive ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D methyl ester N 0.0800 0.01376 17 80 

2-Hydroxyatrazine Y .1131 .03065 27 113

3(4-chlorophenyl)-1-methyl urea N .1003 .01455 15 100 

3-Hydroxycarbofuran N .0924 .00991 11 92

3-Ketocarbofuran Y .0822 .01150 14 82

Aldicarb Y .0619 .01297 21 62

Aldicarb sulfone Y .0672 .02544 38 67 

Aldicarb sulfoxide Y .0765 .00431 6 77 

Atrazine N .0940 .01172 12 94

Bendiocarb N .0860 .00973 11 86

Benomyl N .0820 .00349 4 82

Bensulfuron-methyl N .0939 .00767 8 94

Bromacil Y .0944 .01283 14 94

Caffeine N .1094 .01280 12 109

Carbaryl N .0885 .00999 11 89

Carbofuran N .0850 .00901 11 85

Chloramben methyl ester Y .0939 .01812 19 94 

Chlorimuron-ethyl N .0864 .00584 7 86

Cycloate Y .0720 .00864 12 72

Deethylatrazine Y .1089 .01381 13 109

Deethyldeiopropylatrazine Y .0882 .00953 11 88

Deisopropylatrazine Y .1063 .01173 11 106

Diphenamid N .0847 .00925 11 85

Diuron N .0918 .01262 14 92

Fenuron N .0863 .01169 14 86

Flumetsulam Y .1341 .01377 10 134

ANALYTICAL METHOD 43
Table 15. Accuracy and precision data from seven determinations of the method compounds fortified at 

0.10 microgram per liter in organic-free water, under positive ionization conditions—Continued 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

Fluometuron N 0.0873 0.00982 11 87

Imazaquin Y .1664 .01639 10 166

Imazethapyr Y .2007 .01399 7 201

Imidicloprid N .0987 .01688 17 99

Linuron N .0872 .01105 13 87

Metalaxyl N .0840 .00909 11 84

Methiocarb Y .0884 .01265 14 88

Methomyl Y .0810 .01222 15 81

Methomyl oxime Y .0121 .00162 13 12 

Metsulfuron-methyl Y .0759 .01810 24 76

Neburon N .0901 .01188 13 90

Nicosulfuron N .1295 .01039 8 130

Norflurazon Y .0925 .01232 13 93

Oryzalin N .0874 .01131 13 87

Oxamyl N .0836 .00255 3 84

Oxamyl oxime Y .0259 .01025 40 26 

Propham N .0843 .01141 14 84

Propiconazole N .1000 .01024 10 100

Propoxur N .0854 .00945 11 85

Siduron N .0960 .01484 15 96

Sulfometuron-methyl N .0982 .00618 6 98

Tebuthiuron N .1062 .01220 11 106

Terbacil Y .0966 .01517 16 97

Tribenuron-methyl Y .0268 .01078 40 27

Table 16. Accuracy and precision data from seven determinations of the method compounds fortified at 

0.10 microgram per liter in organic-free water, under negative ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter; nd, not determined] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D N 0.0722 0.01231 17 72

2,4-DB Y .0705 .00856 12 71

Acifluorfen N .0847 .00990 12 85

Bentazon Y .0612 .00307 5 61

Bromoxynil Y .0827 .00909 11 83

Chlorothalonil Y .0123 .01182 96 12

Clopyralid N .0862 .00654 8 86

Dacthal monoacid N .0851 .01148 13 85 

Dicamba N .0918 .01527 17 92

Dichlorprop N nd nd nd nd

Dinoseb N .0823 .00683 8 82

MCPA N .0786 .00931 12 79

MCPB Y .0690 .00994 14 69

Picloram N .0904 .01133 13 90

Triclopyr N .0907 .01604 18 91

44 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
ANALYTICAL METHOD 45

Table 17. Accuracy and precision data from nine determinations of the method compounds fortified at 

0.50 microgram per liter in organic-free water, under positive ionization conditions 

[conc., concentration; Y/N, yes/no; g/L, microgram per liter] 

Compound 

Reported 

as an 

estimated 

conc. 

(Y/N) 

Mean 

observed 

conc. 

(μg/L) 

Standard 

deviation 

(μg/L) 

Relative 

standard 

deviation 

(percent) 

Mean 

accuracy 

(percent of 

true conc.) 

2,4-D methyl ester N 0.6060 0.04205 7 121 

2-Hydroxyatrazine Y .5415 .09270 17 108 

3(4-chlorophenyl)-1-methyl urea N .5098 .02956 6 102 

3-Hydroxycarbofuran N .4725 .04037 9 95 

3-Ketocarbofuran Y .2774 .1582 57 55 

Aldicarb Y .3883 .05049 13 78 

Aldicarb sulfone Y .3783 .04853 13 76 

Aldicarb sulfoxide Y .3747 .04017 11 75 

Atrazine N .5250 .01725 3 105 

Bendiocarb N .4330 .02770 6 87 

Benomyl N .4329 .04982 12 87 

Bensulfuron-methyl N .4756 .02557 5 95 

Bromacil Y .4304 .05635 13 86 

Caffeine N .4857 .04800 10 97 

Carbaryl N .4523 .05996 13 90 

Carbofuran N .4954 .02407 5 99 

Chloramben methyl ester Y .4370 .05847 13 87 

Chlorimuron-ethyl N .4794 .04320 9 96 

Cycloate Y .3198 .05180 16 64 

Deethylatrazine Y .4836 .02359 5 97 

Deethyldeiopropylatrazine Y .4199 .03419 8 84 

Deisopropylatrazine Y .4416 .05683 13 88 

Diphenamid N .4554 .02355 5 91 

Diuron N .4639 .00897 2 93 

Fenuron N .4364 .05450 12 87 

Flumetsulam Y .4383 .02062 5 88 

Fluometuron N .4581 .01034 2 92 

Imazaquin Y .5825 .02403 4 117 

Imazethapyr Y .5589 .02199 4 112 

Imidicloprid N .3844 .05725 15 77 

Linuron N .4911 .01474 3 98 

Metalaxyl N .4808 .01269 3 96 

Methiocarb Y .4505 .03728 8 90 

Methomyl Y .3890 .03807 10 78 

Methomyl oxime Y .0757 .02830 37 15 

Metsulfuron-methyl Y .6061 .08198 14 121 

Neburon N .4703 .01215 3 94 

Nicosulfuron N .4886 .01786 4 98 

Norflurazon Y .4721 .02388 5 94 

Oryzalin N .4175 .01890 5 84 

Oxamyl N .3890 .03985 10 78 

Oxamyl oxime Y .1049 .05437 52 21 

Propham N .4630 .01057 2 93 

Propiconazole N .4655 .03045 7 93 

Propoxur N .4466 .02265 5 89 

Siduron N .4446 .02779 6 89 

Sulfometuron-methyl N .3896 .01961 5 78 

Tebuthiuron N .5213 .01444 3 104 

Terbacil Y .4191 .05142 12 84 

Tribenuron-methyl Y .4235 .07477 18 85 
Table 18. Accuracy and precision data from nine determinations of the method compounds fortified at 

0.50 microgram per liter in organic-free water, under negative ionization conditions 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Mean

accuracy

(percent of

true conc.) 

2,4-D N 0.4431 0.05134 89

2,4-DB Y .3078 .02720 62

Acifluorfen N .3918 .03392 78

Bentazon Y .2891 .04762 58

Bromoxynil Y .3734 .03328 75

Chlorothalonil Y .0384 .02494 8

Clopyralid N .3147 .04925 63

Dacthal monoacid N .1600 .06604 32 

Dicamba N .3894 .02944 78

Dichlorprop N .4239 .02782 85

Dinoseb N .3922 .04564 78

MCPA N .4662 .03263 93

MCPB Y .5971 .05247

Relative

standard

deviation 

(percent)

12

9

9

16

9

65

16

41 

8

7

12

7

9 119

Picloram N .3828 .07625 20 77

N .4256 .03132 7 85

Table 19. Accuracy and precision data from 10 determinations of the method compounds fortified at 

0.025 microgram per liter in ground-water samples, under positive ionization conditions

[conc., concentration; Y/N, yes/no; g/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

2,4-D methyl ester 0.0228 0.00153 7 91 0.0049 

Y

Triclopyr

Mean

observed 

conc. 

(μg/L) 

N

2-Hydroxyatrazine .0230 .00142 6 92 .0045

3(4-chlorophenyl)-1-methyl urea N .0226 .00106 5 90 .0034 

3-Hydroxycarbofuran N .0294 .00233 8 118 .0074

3-Ketocarbofuran Y .0099 .00035 4 40 .0011

Aldicarb Y .0218 .00112 5 87 .0036

Aldicarb sulfone Y .0229 .00088 4 92 .0028 

Aldicarb sulfoxide Y .0232 .00072 3 93 .0023 

Atrazine N .0245 .00088 4 98 .0028

Bendiocarb N .0238 .00111 5 95 .0035

Benomyl N .0231 .00149 6 93 .0047

Bensulfuron-methyl N .0386 .00340 9 154 .0108

Bromacil Y .0225 .00105 5 90 .0033

Caffeine N .0238 .00061 3 95 .0019

Carbaryl N .0237 .00062 3 95 .0020

Carbofuran N .0238 .00080 3 95 .0025

Chloramben methyl ester Y .0210 .00101 5 84 .0032 

Chlorimuron-ethyl N .0239 .00150 6 95 .0048

Cycloate Y .0240 .00307 13 96 .0097

Deethylatrazine Y .0229 .00092 4 92 .0029

Deethyldeiopropylatrazine Y .0243 .00061 3 97 .0019

Deisopropylatrazine Y .0229 .00091 4 92 .0029

Diphenamid N .0255 .00132 5 102 .0042

Diuron N .0244 .00056 2 98 .0018

46 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 19. Accuracy and precision data from 10 determinations of the method compounds fortified at 

0.025 microgram per liter in ground-water samples, under positive ionization conditions—Continued 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

Fenuron N 0.0223 0.00101 5 89 0.0032

Flumetsulam Y .0383 .00304 8 153 .0096

Fluometuron N .0244 .00065 3 97 .0021

Imazaquin Y .0527 .00965 18 211 .0306

Imazethapyr Y .0350 .00191 5 140 .0060

Imidicloprid N .0382 .00320 8 153 .0101

Linuron N .0240 .00106 4 96 .0034

Metalaxyl N .0239 .00093 4 95 .0029

Methiocarb Y .0239 .00097 4 96 .0031

Methomyl Y .0235 .00113 5 94 .0036

Methomyl oxime Y .0091 .00097 11 37 .0031 

Metsulfuron-methyl Y .0396 .00543 14 159 .0172

Neburon N .0237 .00100 4 95 .0032

Nicosulfuron N .0429 .00327 8 171 .0104

Norflurazon Y .0242 .00065 3 97 .0021

Oryzalin N .0225 .00172 8 90 .0054

Oxamyl N .0238 .00139 6 95 .0044

Oxamyl oxime Y .0071 .00498 70 29 .0158 

Propham N .0238 .00065 3 95 .0021

Propiconazole N .0221 .00158 7 88 .0050

Propoxur N .0237 .00072 3 95 .0023

Siduron N .0236 .00090 4 94 .0029

Sulfometuron-methyl N .0388 .00292 8 155 .0092

Tebuthiuron N .0268 .00272 10 107 .0086

Terbacil Y .0229 .00062 3 92 .0020

Tribenuron-methyl Y .0157 .00203 13 63 .0064

Table 20. Accuracy and precision data from nine determinations of the method compounds fortified at 

0.025 microgram per liter in ground-water samples, under negative ionization conditions

[conc., concentration; Y/N, yes/no; g/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

2,4-D N 0.0239 0.00175 7 96 0.0057

2,4-DB Y .0227 .00169 7 91 .0055

Acifluorfen N .0236 .00132 6 94 .0043

Bentazon Y .0207 .00099 5 83 .0032

Bromoxynil Y .0163 .00185 11 65 .0060

Chlorothalonil Y .0148 .00272 18 59 .0088

Clopyralid N .0233 .00104 4 93 .0034

Dacthal monoacid N .0238 .00090 4 95 .0029

Dicamba N .0232 .00082 4 93 .0027

Dichlorprop N .0238 .00159 7 95 .0052

Dinoseb N .0062 .00085 14 25 .0028

MCPA N .0236 .00098 4 94 .0032

MCPB Y .0205 .00099 5 82 .0032

Picloram N .0234 .00102 4 94 .0033

Triclopyr N .0242 .00115 5 97 .0038

ANALYTICAL METHOD 47
Table 21. Accuracy and precision data from six determinations of the method compounds fortified at 0.10 

microgram per liter in ground-water samples, under positive ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D methyl ester N 0.0826 0.01318 16 83 

2-Hydroxyatrazine Y .1226 .02965 24 123

3(4-chlorophenyl)-1-methyl urea N .0918 .01323 14 92 

3-Hydroxycarbofuran N .0822 .00729 9 82

3-Ketocarbofuran Y .0554 .01110 20 55

Aldicarb Y .0585 .01442 25 59

Aldicarb sulfone Y .0473 .02192 46 47 

Aldicarb sulfoxide Y .0735 .00649 9 73 

Atrazine N .0827 .01322 16 83

Bendiocarb N .0653 .00282 4 65

Benomyl N .0748 .00470 6 75

Bensulfuron-methyl N .1260 .00945 8 126

Bromacil Y .0768 .01486 19 77

Caffeine N .0991 .00840 8 99

Carbaryl N .0845 .01063 13 84

Carbofuran N .0844 .00944 11 84

Chloramben methyl ester Y .0811 .00521 6 81 

Chlorimuron-ethyl N .0982 .01716 17 98

Cycloate Y .0724 .00824 11 72

Deethylatrazine Y .0702 .01419 20 70

Deethyldeiopropylatrazine Y .0825 .00639 8 82

Deisopropylatrazine Y .0651 .01304 20 65

Diphenamid N .0842 .01009 12 84

Diuron N .0900 .01335 15 90

Fenuron N .0796 .01081 14 80

Flumetsulam Y .1725 .00761 4 172

Fluometuron N .0863 .01027 12 86

Imazaquin Y .1455 .01904 13 145

Imazethapyr Y .1807 .05831 32 181

Imidicloprid N .1453 .01455 10 145

Linuron N .0887 .01014 11 89

Metalaxyl N .0872 .00980 11 87

Methiocarb Y .0891 .01143 13 89

Methomyl Y .0785 .01676 21 78

Methomyl oxime Y .0111 .00100 9 11 

Metsulfuron-methyl Y .1436 .03049 21 144

Neburon N .0860 .01542 18 86

Nicosulfuron N .1740 .01222 7 174

Norflurazon Y .0930 .01094 12 93

Oryzalin N .0905 .01251 14 90

Oxamyl N .0700 .00724 10 70

Oxamyl oxime Y .0238 .01125 47 24 

Propham N .0803 .01164 14 80

Propiconazole N .0972 .00808 8 97

Propoxur N .0807 .00926 11 81

Siduron N .0996 .01481 15 100

Sulfometuron-methyl N .1546 .01200 8 155

Tebuthiuron N .0961 .01555 16 96

Terbacil Y .0795 .01433 18 80

Tribenuron-methyl Y .0434 .02271 52 43

48 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 22. Accuracy and precision data from six determinations of the method compounds fortified at 0.10 

microgram per liter in ground-water samples, under negative ionization conditions

[conc., concentration; Y/N; yes/no; g/L, microgram per liter; nd, not determined] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D Y 0.0646 0.00468 7 65

2,4-DB N .0748 .00654 9 75

Acifluorfen N .0780 .01180 15 78

Bentazon N .0600 .00282 5 60

Bromoxynil Y .0738 .00531 7 74

Chlorothalonil Y .0486 .01359 28 49

Clopyralid N .0755 .00975 13 75

Dacthal monoacid N .0777 .00770 10 78 

Dicamba Y .0786 .00959 12 79

Dichlorprop Y nd nd nd nd

Dinoseb N .0792 .00390 5 79

MCPA Y .0725 .00556 8 73

MCPB N .0651 .00614 9 65

Picloram N .0821 .00625 8 82

Triclopyr Y .0835 .01317 16 83

Table 23. Accuracy and precision data from 11 determinations of the method compounds fortified at 0.50 

microgram per liter in ground-water samples, under positive ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D methyl ester N 0.6056 0.03970 7 121 

2-Hydroxyatrazine Y .5503 .09067 16 110

3(4-chlorophenyl)-1-methyl urea N .5286 .07316 14 106 

3-Hydroxycarbofuran N .4740 .03803 8 95

3-Ketocarbofuran Y .2994 .03088 10 60

Aldicarb Y .2440 .15406 63 49

Aldicarb sulfone Y .2038 .16250 80 41 

Aldicarb sulfoxide Y .2546 .11533 45 51 

Atrazine N .5031 .07281 14 101

Bendiocarb N .6663 .16411 25 133

Benomyl N .4042 .10879 27 81

Bensulfuron-methyl N .4340 .13865 32 87

Bromacil Y .3884 .09041 23 78

Caffeine N .3404 .21684 64 68

Carbaryl N .4448 .10781 24 89

Carbofuran N .4703 .09763 21 94

Chloramben methyl ester Y .3528 .14311 41 71 

Chlorimuron-ethyl N .4784 .03925 8 96

ANALYTICAL METHOD 49
Table 23. Accuracy and precision data from 11 determinations of the method compounds fortified at 

0.50 microgram per liter in ground-water samples, under positive ionization conditions—Continued 

Compound 

Reported as

an estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

Cycloate Y 0.2102 0.13361 64 42

Deethylatrazine Y .4838 .02207 5 97

Deethyldeiopropylatrazine Y .4176 .18583 44 84

Deisopropylatrazine Y .3387 .19134 56 68

Diphenamid N .4130 .13814 33 83

Diuron N .4515 .04772 11 90

Fenuron N .4367 .05099 12 87

Flumetsulam Y .4407 .02060 5 88

Fluometuron N .4563 .01552 3 91

Imazaquin Y .5570 .09995 18 111

Imazethapyr Y .5188 .08684 17 104

Imidicloprid N .3771 .06654 18 75

Linuron N .4922 .01486 3 98

Metalaxyl N .4376 .14267 33 88

Methiocarb Y .2805 .19175 68 56

Methomyl Y .2756 .14205 52 55

Methomyl oxime Y .0641 .03562 56 13 

Metsulfuron-methyl Y .6685 .16345 24 134

Neburon N .4716 .01157 2 94

Nicosulfuron N .4627 .08009 17 93

Norflurazon Y .3877 .12897 33 78

Oryzalin N .3836 .11295 29 77

Oxamyl N .3199 .09500 30 64

Oxamyl oxime Y .0887 .04992 56 18 

Propham N .4312 .12601 29 86

Propiconazole N .4241 .13628 32 85

Propoxur N .4251 .07483 18 85

Siduron N .4064 .10948 27 81

Sulfometuron-methyl N .3587 .10319 29 72

Tebuthiuron N .5225 .01363 3 105

Terbacil Y .4115 .05046 12 82

Tribenuron-methyl Y .3196 .11716 37 64

from the data for mean recoveries of 

individual compounds. No method 

compounds were detected in the groundwater samples, so no background correction 

was required. 

11.3.2 The mean positive 

ionization recoveries for compounds from 

the ground-water samples reported without 

qualification (tables 19, 21, and 23) were 

104.8  23.1, 95.0  25.1, and 90.2  14.6 

percent at fortifications of 0.025, 0.100, and 

0.500 g/L, respectively. The median 

recoveries were similar to the means, at

95.0, 86.5, and 88.5 percent, respectively. 

Mean recoveries of compounds reported as 

qualified estimates were more variable, at

96.6  40.8, 84.7  43.9, and 70.6  30.1 

percent, with corresponding medians of 

92.0, 77.5, and 69.5 percent, respectively.

11.3.3 Mean recoveries 

determined by negative ionization and 

reported without qualification in the groundwater samples (tables 20, 22, and 24)

50 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 24. Accuracy and precision data from 10 determinations of the method compounds fortified at 0.50 

microgram per liter in ground-water samples, under negative ionization conditions

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter; nd, not determined] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. (μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D N 0.0722 0.01231 17 72

2,4-DB N .0705 .00856 12 71

Acifluorfen Y .0847 .00990 12 85

Bentazon N .0612 .00307 5 61

Bromoxynil N .0827 .00909 11 83

Chlorothalonil N .0123 .01182 96 12

Clopyralid N .0862 .00654 8 86

Dacthal monoacid N .0851 .01148 13 85 

Dicamba N .0918 .01527 17 92

Dichlorprop Y nd nd nd nd

Dinoseb Y .0823 .00683 8 82

MCPA N .0786 .00931 12 79

MCPB Y .0690 .00994 14 69

Picloram N .0904 .01133 13 90

Triclopyr Y .0907 .01604 18 91

averaged slightly lower, at 87.6  22.0, 74.0 

 7.6 and 73.1  23.5 percent, at fortifications 

of 0.025, 0.100, and 0.500 g/L, respectively. 

The median recoveries were similar to the

means, at 94.0, 76.5, and 81.0 percent, 

respectively. In this matrix, mean recoveries

of compounds reported as qualified estimates 

were similar to compounds reported without 

qualification, at 76.0  13.4, 70.5  12.2, and 

81.8  9.3 percent, with corresponding

medians of 92.0, 77.5, and 69.5 percent, 

respectively.

11.3.4 In the ground-water 

samples, average recoveries of all compounds 

reported without qualification at all three 

fortification levels varied about 20.9 percent 

for compounds determined by positive 

ionization and 17.7 percent for compounds 

determined by negative ionization. This 

result, however, does not reflect the analytical

precision, which is better reflected by the 

relative standard deviation of individual 

compound recoveries in the ground-water 

samples, particularly those that are reported 

without quantitation qualification. Under 

positive ionization conditions, the mean 

relative standard deviations for compounds 

reported without qualification in the groundwater samples were 5.4  2.2, 11.7  3.6, and 

20.5  13.5 percent for 0.025-, 0.100-, and 

0.500-μg/L fortifications, respectively. The

corresponding relative standard deviations for 

negative ionization results reported without 

qualification in the ground-water samples 

were 5.9  3.1, 9.2  3.5, and 20.4  26.8 

percent for 0.025-, 0.100-, and 0.500-μg/L

fortifications, respectively. The greater 

variability in the relative standard deviation 

for compounds determined by negative 

ionization in the 0.500-μg/L fortification was 

the result of particularly variable recovery of 

chlorothalonil. If this result is excluded, then

the mean relative standard deviation was 12 

± 3.8 percent, similar to the mean relative 

standard deviations reported for the other 

fortifications. 

11.4 Surface-water fortification 

recovery results

ANALYTICAL METHOD 51
11.4.1 In contrast to the other 

matrices tested, one set of 10 surface-water 

samples was fortified at 0.050 g/L as the 

lowest fortification concentration rather than

0.025 g/L, as was the case with the reagentwater and ground-water fortification 

experiments. This change was made because 

of nonspecific matrix interferences. 

Concentrations of dissolved organic carbon, 

the likely source of these interferences in the 

South Platte River, typically ranged between 

3 and 30 mg/L (Litke and Kimbrough, 1998), 

although median concentrations were 5.2 and 

7.0 mg/L in the South Platte River at Denver 

and Henderson, Colorado, respectively. The 

remaining two sets of samples were fortified 

at 0.100 and 0.500 g/L, identical to the

organic-free and ground-water fortifications. 

The surface-water fortification experiments at

0.100 g/L were conducted 8 months prior to 

the 0.05 and 0.5 fortifications. All results are 

listed in tables 25 through 31. The mean, 

median, and the standard deviation of 

recovery for all compounds in the tables were 

calculated from the data for mean recoveries 

of individual compounds. In comparison to 

the reagent-water and ground-water samples, 

measurable concentrations of a few method 

analytes were detected. Caffeine, 2-hydroxyatrazine, atrazine, diuron, 2,4-D, and MCPA 

were detected in at least two of three replicate 

unspiked South Platte surface-water samples. 

The mean concentrations are listed in table 

25. Spiked South Platte River surface-water 

results were corrected for ambient

concentrations prior to calculation of 

recoveries. 

11.4.2 Compound recoveries from 

the surface-water samples were higher, often 

substantially greater than 100 percent at the 

0.05- and 0.5-g/L fortifications, and more 

variable between compounds than in either 

the organic-free water or the ground-water 

samples. The mean positive ionization 

recovery results for compounds reported 

without qualification in the surface-water 

samples were 131  66.3, 90.8  10.0, and 

125  93.5 percent at fortifications of 0.050, 

0.100, and 0.500 g/L, respectively. The 

median recoveries were similar to the means 

at 119.0, 87.0, and 108.0 percent, 

respectively. For those compounds 

determined under positive ion conditions and 

Table 25. Ambient pesticide concentrations measured in South Platte River samples during this study

[μg/L, microgram per liter] 

Compound 

Mean detected

concentrations, 

in μg/L 

Standard deviation of 

mean detected

concentrations, 

in μg/L 

Number of 

detections/number of 

determinations

2,4-D 0.1925 0.1025 2/2

2-Hydroxyatrazine .0645 .0021 2/3

Atrazine .0065 .0007 3/3

Caffeine .1033 .0112 3/3

Diuron .0380 .0066 3/3

MCPA .0250 .0156 2/2

52 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 26. Accuracy and precision data from nine determinations of the method compounds fortified at 

0.05 microgram per liter in surface-water samples, under positive ionization conditions. Recoveries

corrected for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.)

Method 

detection 

limit 

(μg/L) 

2,4-D methyl ester N 0.0173 0.00543 31 35 0.0177 

2-Hydroxyatrazine Y .0362 .00449 12 72 .0146

3(4-chlorophenyl)-1-methyl urea N .0442 .00380 9 88 .0124 

3-Hydroxycarbofuran N .0501 .00525 10 100 .0171

3-Ketocarbofuran Y .0032 .00045 14 6 .0015

Aldicarb Y .0439 .00668 15 88 .0217

Aldicarb sulfone Y .0363 .00700 19 73 .0227 

Aldicarb sulfoxide Y .0247 .00173 7 49 .0056 

Atrazine N .0536 .00416 8 107 .0135

Bendiocarb N .0124 .00453 36 25 .0147

Benomyl N .0459 .00417 9 92 .0135

Bensulfuron-methyl N .1446 .00757 5 289 .0246

Bromacil Y .0289 .00314 11 58 .0102

Caffeine N .0541 .01065 20 108 .0346

Carbaryl N .0412 .00474 11 82 .0154

Carbofuran N .0484 .00317 7 97 .0103

Chloramben methyl ester Y .0737 .00433 6 147 .0141 

Chlorimuron-ethyl N .1143 .00953 8 229 .0320

Cycloate Y .0236 .00609 26 47 .0198

Deethylatrazine Y .0466 .00305 7 93 .0099

Deethyldeiopropylatrazine Y .0256 .00277 11 51 .0093

Deisopropylatrazine Y .0386 .00309 8 77 .0100

Diphenamid N .0676 .00388 6 135 .0126

Diuron N .0606 .00604 10 121 .0196

Fenuron N .0519 .00688 13 104 .0224

Flumetsulam Y .1003 .00873 9 201 .0284

Fluometuron N .0623 .00346 6 125 .0113

Imazaquin Y .1069 .00643 6 214 .0209

Imazethapyr Y .2210 .01377 6 442 .0447

Imidicloprid N .0729 .01179 16 146 .0383

Linuron N .0683 .00632 9 137 .0206

Metalaxyl N .0663 .00374 6 133 .0122

Methiocarb Y .0396 .00515 13 79 .0167

Methomyl Y .0654 .00543 8 131 .0177

Methomyl oxime Y .0038 .00158 42 8 .0053 

Metsulfuron-methyl Y .0124 .00450 36 25 .0146

Neburon N .0583 .00450 8 117 .0146

Nicosulfuron N .1563 .01168 7 313 .0380

Norflurazon Y .0671 .00369 5 134 .0120

Oryzalin N .0606 .01500 25 121 .0488

Oxamyl N .0487 .01150 24 97 .0374

Oxamyl oxime Y .0241 .00326 14 48 .0106 

Propham N .0457 .00566 12 91 .0184

Propiconazole N .0637 .00502 8 127 .0163

Propoxur N .0516 .00240 5 103 .0078

Siduron N .0744 .00439 6 149 .0143

Sulfometuron-methyl N .1291 .00645 5 258 .0210

Tebuthiuron N .0638 .00432 7 128 .0141

Terbacil Y .0236 .00334 14 47 .0112

Tribenuron-methyl Y .0974 .00665 7 195 .0216

ANALYTICAL METHOD 53
Table 27. Accuracy and precision data from nine determinations of the method compounds fortified at 

0.05 microgram per liter in surface-water samples, under negative ionization conditions. Recoveries

corrected for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; g/L, microgram per liter; nd, not determined] 

Compound 

Reported 

as an

estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

Method 

detection 

limit 

(μg/L) 

2,4-D N 0.0226 0.00477 21 45 0.0155

2,4-DB Y .0252 .00295 12 50 .0096

Acifluorfen N .0261 .00112 4 52 .0036

Bentazon Y .0249 .00289 12 50 .0094

Bromoxynil Y .0260 .00132 5 52 .0043

Chlorothalonil Y .0258 .00311 12 52 .0101

Clopyralid N .3057 .01425 5 611 .0463

Dacthal monoacid N .0159 .00401 25 32 .0130 

Dicamba N .0251 .00280 11 50 .0091

Dichlorprop N .0134 .00837 62 27 .0272

Dinoseb N .0591 .00337 6 118 .0110

MCPA N nd nd nd nd nd

MCPB Y .0263 .00187 7 53 .0061

Picloram N .0261 .00302 12 52 .0098

Triclopyr N .0234 .00113 5 47 .0037

Table 28. Accuracy and precision data from six determinations of the method compounds fortified at 0.10 

microgram per liter in surface-water samples, under positive ionization conditions. Recoveries corrected 

for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter; nd, not determined; na, not available] 

Compound 

Reported as

an estimated

conc. 

(Y/N) 

Mean

observed 

conc. 

(μg/L) 

Standard

deviation 

(μg/L) 

Relative

standard

deviation 

(percent)

Mean

accuracy

(percent of

true conc.) 

2,4-D methyl ester N 0.0780 0.01388 18 78 

2-Hydroxyatrazine Y .1086 .03100 29 109

3(4-chlorophenyl)-1-methyl urea N .0997 .01584 16 100 

3-Hydroxycarbofuran N .0916 .01059 12 92

3-Ketocarbofuran Y .0822 .01260 15 82

Aldicarb Y .0622 .01417 23 62

Aldicarb sulfone Y .0649 .02707 42 65 

Aldicarb sulfoxide Y .0758 .00418 6 76 

Atrazine N .0925 .01202 13 92

Bendiocarb N .0852 .01039 12 85

Benomyl N .0809 .00212 3 81

Bensulfuron-methyl N .0921 .00657 7 92

Bromacil Y .0928 .01322 14 93

Caffeine N .1085 .01380 13 109

Carbaryl N .0877 .01070 12 88

Carbofuran N .0846 .00981 12 85

Chloramben methyl ester Y .0928 .01962 21 93 

Chlorimuron-ethyl N .0848 .00448 5 85

Cycloate Y .0705 .00838 12 71

Deethylatrazine Y .1080 .01490 14 108

Deethyldeiopropylatrazine Y .0877 .01034 12 88

Deisopropylatrazine Y .1055 .01265 12 105

Diphenamid N .0834 .00939 11 83

Diuron N .0899 .01267 14 90

54 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
ANALYTICAL METHOD 55

Table 28. Accuracy and precision data from six determinations of the method compounds fortified at 0.10 

microgram per liter in surface-water samples, under positive ionization conditions. Recoveries corrected 

for ambient compound concentrations listed in table 25—Continued 

Compound 

Reported as 

an estimated 

conc. 

(Y/N) 

Mean 

observed 

conc. 

(μg/L) 

Standard 

deviation 

(μg/L) 

Relative 

standard 

deviation 

(percent) 

Mean 

accuracy 

(percent of 

true conc.) 

Fenuron N 0.0856 0.01263 15 86 

Flumetsulam Y .1302 .01003 8 130 

Fluometuron N .0865 .01046 12 86 

Imazaquin Y .1620 .01256 8 162 

Imazethapyr Y nd nd nd nd 

Imidicloprid N .0956 .01620 17 96 

Linuron N .0864 .01184 14 86 

Metalaxyl N .0835 .00985 12 83 

Methiocarb Y .0870 .01324 15 87 

Methomyl Y .0796 .01282 16 80 

Methomyl oxime Y .0129 .00008 1 13 

Metsulfuron-methyl Y .0746 .01945 26 75 

Neburon N .0886 .01225 14 89 

Nicosulfuron N .1261 .00532 4 126 

Norflurazon Y .0918 .01335 15 92 

Oryzalin N .0860 .01175 na 86 

Oxamyl N .0841 .00244 3 84 

Oxamyl oxime Y .0256 .01120 44 26 

Propham N .0824 .01114 14 82 

Propiconazole N .0978 .00927 9 98 

Propoxur N .0846 .01011 12 85 

Siduron N .0939 .01512 16 94 

Sulfometuron-methyl N .0962 .00355 4 96 

Tebuthiuron N .1052 .01302 12 105 

Terbacil Y .0952 .01614 17 95 

Tribenuron-methyl Y .0278 .01141 41 28 

Table 29. Accuracy and precision data from six determinations of the method compounds fortified at 0.10 

microgram per liter in surface-water samples, under negative ionization conditions. Recoveries corrected 

for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter; nd, not determined] 

Compound 

Reported as 

an estimated 

conc. 

(Y/N) 

Mean 

observed 

conc. 

(μg/L) 

Standard 

deviation 

(μg/L) 

Relative 

standard 

deviation 

(percent) 

Mean 

accuracy 

(percent of 

true conc.) 

2,4-D N 0.0692 0.00858 12 69 

2,4-DB N .0672 .00855 13 67 

Acifluorfen Y .0732 .00775 11 73 

Bentazon N .0573 .00425 7 57 

Bromoxynil N .0649 .00613 9 65 

Chlorothalonil N .0764 .01028 13 76 

Clopyralid N .0501 .01623 32 50 

Dacthal monoacid N .0744 .00804 11 74 

Dicamba N .0733 .00890 12 73 

Dichlorprop Y nd nd nd nd 

Dinoseb Y .0743 .00440 6 74 

MCPA N .0751 .00535 7 75 

MCPB Y .0651 .00921 14 65 

Picloram N .0871 .00765 9 87 

Triclopyr Y .0789 .00898 11 79 
56 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS 

Table 30. Accuracy and precision data from eight determinations of the method compounds fortified at 

0.50 microgram per liter in surface-water samples, under positive ionization conditions. Recoveries 

corrected for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter] 

Compound 

Reported as 

an estimated 

conc. 

(Y/N) 

Mean 

observed 

conc. 

(μg/L) 

Standard 

deviation 

(μg/L) 

Relative 

standard 

deviation 

(percent) 

Mean 

accuracy 

(percent of 

true conc.) 

2,4-D methyl ester N 0.0123 0.01210 98 2 

2-Hydroxyatrazine Y .4449 .16531 37 89 

3(4-chlorophenyl)-1-methyl urea N .4648 .04682 10 93 

3-Hydroxycarbofuran N .1589 .10825 68 32 

3-Ketocarbofuran Y .0058 .00096 17 1 

Aldicarb Y .3174 .07261 23 63 

Aldicarb sulfone Y .2719 .07615 28 54 

Aldicarb sulfoxide Y .5699 .15729 28 114 

Atrazine N .5258 .07168 14 105 

Bendiocarb N .0069 .00544 79 1 

Benomyl N .4231 .04948 12 85 

Bensulfuron-methyl N 1.9925 .30680 15 399 

Bromacil Y .2469 .01911 8 49 

Caffeine N .6533 .06538 10 131 

Carbaryl N .0839 .07925 94 17 

Carbofuran N .2638 .14014 53 53 

Chloramben methyl ester Y .8634 .07006 8 173 

Chlorimuron-ethyl N 1.4456 .10097 7 289 

Cycloate Y .1804 .08792 49 36 

Deethylatrazine Y .4493 .06076 14 90 

Deethyldeiopropylatrazine Y .0443 .01743 39 9 

Deisopropylatrazine Y .3958 .05684 14 79 

Diphenamid N .6759 .07794 12 135 

Diuron N .6533 .06118 9 131 

Fenuron N .5284 .04383 8 106 

Flumetsulam Y 1.0458 .09840 9 209 

Fluometuron N .6446 .06166 10 129 

Imazaquin Y 1.3143 .14009 11 263 

Imazethapyr Y 1.5173 .04774 3 303 

Imidicloprid N .8706 .08315 10 174 

Linuron N .5155 .07320 14 103 

Metalaxyl N .7384 .08215 11 148 

Methiocarb Y .1597 .02914 18 32 

Methomyl Y .6341 .12760 20 127 

Methomyl oxime Y .0399 .04037 101 8 

Metsulfuron-methyl Y .0071 .00569 80 1 

Neburon N .5493 .12373 23 110 

Nicosulfuron N 1.5246 .11032 7 305 

Norflurazon Y .7425 .06540 9 149 

Oryzalin N .4705 .06710 14 94 

Oxamyl N .0676 .06188 92 14 

Oxamyl oxime Y .4521 .06752 15 90 

Propham N .4011 .10534 26 80 

Propiconazole N .8118 .16649 21 162 

Propoxur N .3371 .13514 40 67 

Siduron N .7349 .06842 9 147 

Sulfometuron methyl N 1.2940 .10436 8 259 

Tebuthiuron N .6379 .05940 9 128 

Terbacil Y .3790 .04253 11 76 

Tribenuron-methyl Y 1.4160 .00000 0 283 
ANALYTICAL METHOD 57

Table 31. Accuracy and precision data from eight determinations of the method compounds fortified at 

0.50 microgram per liter in surface-water samples, under negative ionization conditions. Recoveries 

corrected for ambient compound concentrations listed in table 25 

[conc., concentration; Y/N, yes/no; μg/L, microgram per liter; nd, not determined] 

Compound 

Reported as 

an estimated 

conc. 

(Y/N) 

Mean 

observed 

conc. 

(μg/L) 

Standard 

deviation 

(μg/L) 

Relative 

standard 

deviation 

(percent) 

Mean 

accuracy 

(percent of 

true conc.) 

2,4-D N 0.4874 0.32028 66 97 

2,4-DB N .2254 .15642 69 45 

Acifluorfen Y .2168 .16552 76 43 

Bentazon N .2014 .12745 63 40 

Bromoxynil N .2520 .17770 71 50 

Chlorothalonil N nd nd nd nd 

Clopyralid N .1158 .08788 76 23 

Dacthal monoacid N .1428 .11184 78 29 

Dicamba N .1913 .13510 71 38 

Dichlorprop Y .2376 .16962 71 48 

Dinoseb Y .3043 .21623 71 61 

MCPA N .2393 .16637 70 48 

MCPB Y .2320 .16347 70 46 

Picloram N .1385 .08582 62 28 

Triclopyr Y .2076 .14622 70 42 

reported as qualified estimates, the 

corresponding mean recoveries were 103.9 

96.1, 82.9  33.8, and 104.5  91.0 percent at 

fortifications of 0.050, 0.100, and 0.500 g/L, 

respectively. The corresponding medians 

were 75.0, 87.0, and 84.0 percent. 

Furlong and others (2000) have 

previously noted apparent matrix 

enhancement of concentration observed in the 

0.05- and 0.5-g/L fortifications in this study. 

The sample used for these fortifications was 

collected about 12 months after the 0.1-g/L 

fortification. The apparent matrix enhancement observed in the surface-water samples 

also appears to be compound class-specific 

and is further discussed in a subsequent 

section (11.5.3) of this report. The fact that 

the South Platte River water used for these 

fortifications was collected at a different time 

and location may have a substantial role in 

determining the type and extent of matrix 

effects. 

11.4.3 Mean recoveries for 

compounds reported without qualification and 

determined by negative ionization in the 

surface-water samples were lower than in 

positive ionization, at 115  188, 69.3  10.4, 

and 44.2  21.9 percent, at fortifications of 

0.05, 0.100, and 0.500 g/L, respectively. 

The corresponding median recoveries were 

50.0, 71.0, and 40.0 percent, and reflect the 

higher variablity observed among compounds 

in negative ionization analysis. The mean 

recoveries of compounds reported as qualified 

estimates and determined by negative 

ionization in surface water were lower but 

less variable, at 51.4  1.3, 72.8  5.8, and 

48.0  7.6 percent at fortifications of 0.05, 

0.100, and 0.500 g/L, respectively. The 

corresponding median recoveries were 52.0, 

73.5, and 46.0 percent. The determination of 

whether a compound was reported as a 

qualified estimate was a function of long-term 

recoveries in reagent water (11.6.2). In this 

case, matrix effects may have resulted in 

lower but less variable recoveries. 

11.4.4 Average surface-water 

recoveries of all compounds at all three 

fortification levels varied about 65 percent 
for compounds determined by positive 

ionization and 59 percent for compounds 

determined by negative ionization. This 

variation indicates that the accuracy of the 

method in the surface-water samples is 

compound dependent, and might reflect 

compound-specific bias resulting from the 

surface-water matrix. Understanding

analytical precision is equally important to 

using any analytical method, however, and 

analytical precision is better reflected by the 

relative standard deviation of individual 

compound recoveries from the surface-water 

samples. Under positive ionization 

conditions, the mean relative standard 

deviations for all compounds in the surfacewater samples were 12  9, 15  9, and 27 

27 percent for 0.050-, 0.100-, and 0.500-μg/L

fortifications, respectively. The

corresponding relative standard deviations for 

negative ionization results were 14  15, 12 

6, and 70  5 percent for 0.050-, 0.100-, and 

0.500-μg/L fortifications, respectively. 

Surface-water matrix effects, particularly the 

higher DOC concentration in the South Platte 

River, might be responsible for the 

substantially higher variability observed in the

0.500-g/L fortifications for both positive and 

negative ionization compounds than in the 

ground-water samples and the organic-freewater samples (11.2 and 11.3). 

11.5 Compound-specific recovery

results 

11.5.1 The method validation 

results by class for all three tested matrices

are shown in figures 2, 3, 4, and 5. In figures 

2, 3, and 4, each figure contains the analytical

results under positive ionization for one 

matrix at three concentrations, subdivided by

compound class. The four compound class 

groupings were as follows: (A) carbamate 

insecticides; (B) triazine and phenylurea

herbicides; (C) sulfonylurea, sulfonamide,

and imidazolinone herbicides; and (D) 

miscellaneous herbicides and other 

compounds. Pesticides determined by

negative ionization HPLC/MS are grouped by

matrix in figure 5. 

11.5.2 A review of the positive 

ionization results for each matrix (figs. 2 

through 4) reveals consistent similarities 

across matrices. Two carbamate degradation 

products, oxamyl oxime and methomyl

oxime, have poor recoveries (averaged only

23, 22, and 29 percent in organic-free water, 

ground-water and surface-water samples, 

respectively) compared to the remaining

carbamate insecticides in which recoveries

averaged 87, 82, and 66 percent, respectively. 

In the other groupings, tribenuron was lower 

than the average recoveries for other 

miscellaneous compounds in the organic-free 

water and in the ground-water samples. 

Tribenuron in the organic-free water and in 

the ground-water samples averaged 48 and 57 

percent, respectively, but in contrast, recovery

was apparently enhanced in the surface-water 

samples, at 178 percent, although recovery in 

surface water varied between fortification 

levels, with the greatest variation measured at

the highest level (fig. 4D). 

11.5.3 When compared with the 

organic-free water and the ground-water 

samples (figs. 2 and 3), fortified surface-water 

recoveries were more variable, which 

indicated that several compound classes 

experienced apparent enhancement of 

recovery. As noted above, this finding applied 

to tribenuron, but also was observed for the 

sulfonylurea, sulfonamide, and imidazolinone 

herbicides. This result is consistent with 

apparent matrix enhancement of recovery for 

sulfonylurea, sulfonamide, and imidazolinone 

herbicides observed in a range of naturalwater samples and South Platte River water 

(Furlong and others, 2000). The results shown 

in figure 4 suggest that enhancement varies by

compound class. The carbamate insecticides 

(fig. 4A) appear to experience the least

enhancement and, for 3-ketocarbofuran, 

perhaps some suppression of recovery. The 

recoveries of the triazine and phenylurea 

herbicides (fig. 4B) are consistent with 

recoveries in the organic-free water and in the 

ground-water samples (figs. 2B and 3B), 

58 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
A B 

C D 

ANALYTICALMETHOD 59

Figure 2. Recoveries, in percent, for compounds fortified at three levels in organic-free water and determined under positive ionization conditions. (A) N-methylcarbamate and related compounds; (B) phenylurea and triazine compounds; (C) sulfonylurea, imidazolinone and related compounds; and (D) other classes of compounds. Error bars indicate ±1 standard deviation.
60 Pesti

cides in Water by Graphitized Carbon-Based SPE and HPLC/MS

A B 

D C 

Figure 3. Recoveries, in percent, for compounds fortified at three levels in ground-water samples and determined under positive ionization conditions. (A) N-methylcarbamate and related compounds; (B) phenylurea and triazine compounds; (C) sulfonylurea, imidazolinone and related compounds; and (D) other classes of compounds. Error bars indicate ±1 standard deviation. 
B A 

C D 

ANALYTICALMETHOD 61

Figure 4. Recoveries, in percent, for compounds fortified at three levels in surface-water samples and determined under positive ionization conditions. (A)N-methylcarbamate and related compounds; (B) phenylurea and triazine compounds; (C) sulfonylurea, imidazolinone and related compounds; and (D) otherclasses of compounds. Error bars indicate ±1 standard deviation. 
62 Pesti

cides in Water by Graphitized Carbon-Based SPE and HPLC/MS

A A B B 

C 

Figure 5. Recoveries, in percent, for compounds determined under negative ionization conditions. (A) Compounds fortified at three levels in organic-free water; (B) compounds fortified at three levels in ground-water samples; and (C) compounds fortified at three levels in surface-water samples. Error bars indicate ±1 standard deviation. 
although they are somewhat more variable in 

the surface-water samples. Recoveries of the 

sulfonylurea, sulfonamide, and imidazolinone 

herbicides (fig 4C) are the most enhanced of 

all the compound classes in the surface-water 

samples and averaged 214  35 percent 

compared to 108  9 and 134  14 percent in 

the organic-free and in the ground-water 

samples, respectively. The miscellaneous 

compounds are variable, with the most 

enhancement measured in the recoveries of 

tribenuron, but with significant enhancement 

in the recoveries of imadacloprid, metalaxyl, 

norflurazon, and propiconazole (fig. 4D). 

Concentrations of compounds 

determined under negative ionization 

conditions (fig. 5) suggest no matrix

enhancement. Recoveries were typically less 

than 100 percent. Recoveries for all 

compounds at all fortifications averaged 75.2 

± 9.3, 80.0 ± 6.5, and 45.8 ± 18.8 percent in 

samples of organic-free water, ground water, 

and surface water, respectively. Chlorothalonil was variably and poorly recovered in 

all matrices and was a substantial influence on 

the observed recoveries and standard 

deviations in all matrices (fig. 5). Recoveries 

also were lower and more variable in the 

surface-water samples and may reflect either 

matrix interference or suppression of 

recovery. 

11.6 Long-term laboratory reagent 

spike results 

11.6.1 As noted earlier, fortified 

organic-free water or laboratory reagent spike 

(LRS) samples were analyzed as part of the 

quality control for each environmental sample 

set. These LRS samples were made up from 

the same pesticide-free water used for

reagent-water fortification recovery

experiments and were at concentrations of 

0.250 μg/L for each compound. The LRS

provides set-specific quality-control 

information for interpretation of individual 

samples in the set. When aggregated 

(table 32), LRS data also provide long-term, 

multiple instrument, and multiple operator

performance data. For the purposes of 

determining long-term method performance, 

results from fiscal year 2000 were aggregated 

and summary statistics were produced. The 

aggregation was made for fiscal year 2000 so 

that the data reflect the performance of the 

method after implementation and training of 

the chemists who perform the analyses. A 

total of 285 reagent spikes was determined in 

fiscal year 2000. 

11.6.2 Long-term LRS recoveries 

averaged 73.8 percent for all compounds, 

with an average standard deviation of 24.2 

percent. The average median recovery for all

compounds was similar to the mean, at 73.4 

percent, and the average f-pseudosigma of 

recovery was 21.4 percent. All compounds in 

the method ranked according to increasing

mean recoveries are shown in figure 6. 

Two statistics were used to 

determine if the reported concentration of any

compound required quantitation qualification. 

Median recoveries calculated from long-term

LRS data are used to estimate the accuracy of 

concentration. A nonparametric statistic, fpseudosigma (Hoaglin, 1983) is calculated to 

determine the variation of LRS recoveries. 

These statistics are used instead of the mean 

and standard deviation because they minimize 

the effects of outlier values, and are thus more

representative of long-term method 

performance under multiple operators and 

instruments. Median recoveries had to fall 

within 60 and 120 percent for a compound to 

be considered reportable without qualification. 

In addition, the f-pseudosigma statistic had to 

be less than 25 percent for a compound to be 

reported without qualification. 

Median recoveries and the fpseudosigma statistics for those recoveries were 

calculated for set LRS data determined between 

October 1, 1999, and September 31, 2000. This 

data period was used because it was considered 

representative of method performance after the

ANALYTICAL METHOD 63
Table 32. Long-term accuracy and precision calculated from multiple determinations of the method compounds 

fortified in sample-set spikes with the compounds added to organic-free water at 0.25 microgram per liter

[N, number of determinations] 

Compound N

Number 

of 

missing

values 

Mean

recovery

(percent)

Standard

deviation 

of 

recovery

(percent)

Median

recovery

(percent)

Fpseudosigma

of recovery

(percent)

Minimum 

recovery

(percent)

Maximum 

recovery

(percent)

Compounds reported without qualification (Median Recovery between 60 and 120 percent and f-pseudosigma 

less than 25 percent)

2,4-D 281 4 101.8 43.6 102.0 16.3 0.1 754

2,4-D methyl ester 282 3 83.5 19.3 84.5 15.4 0.1 150 

3(4-chlorophenyl)-1-methyl urea 282 3 95.5 14.8 96.0 9.5 0.1 155 

3-Hydroxycarbofuran 282 3 91.7 18.7 92.0 12.6 0.1 178

Acifluorfen 280 5 84.0 20.2 86.0 18.5 0.0 134

Atrazine 282 3 74.4 19.3 77.0 16.3 0.1 135

Bendiocarb 282 3 75.8 21.1 79.0 12.6 0.1 154

Benomyl 280 5 89.0 22.9 90.0 13.5 0.0 169

Bensulfuron-methyl 282 3 97.2 22.0 95.0 13.3 0.0 177

Caffeine 282 3 101.2 23.5 103.5 20.6 0.1 172

Carbaryl 282 3 87.7 17.1 89.0 9.5 0.0 173

Carbofuran 282 3 90.7 16.1 90.0 9.6 0.1 166

Chlorimuron-ethyl 282 3 102.6 26.3 101.5 19.1 0.0 179

Clopyralid 281 4 78.8 17.9 80.0 14.8 0.0 117

Dacthal monoacid 279 6 82.1 14.3 83.0 14.1 0.1 115 

Dicamba 281 4 88.8 19.6 89.0 17.8 0.0 144

Dichlorprop 280 5 75.5 15.0 78.0 14.8 0.1 106

Dinoseb 281 4 80.0 21.6 79.0 21.5 0.0 139

Diphenamid 282 3 83.4 16.2 84.0 11.7 0.1 158

Diuron 282 3 89.8 15.5 90.0 9.6 0.1 160

Fenuron 282 3 73.6 19.5 75.0 21.3 0.0 145

Fluometuron 282 3 85.7 14.9 86.0 11.1 0.1 155

Imidacloprid 282 3 106.2 22.7 106.5 19.3 0.1 178

Linuron 282 3 89.5 16.7 91.0 11.1 0.1 154

MCPA 281 4 70.5 18.6 72.0 14.8 0.1 156

Metalaxyl 282 3 85.8 39.5 84.0 9.6 0.1 701

Neburon 281 4 89.8 18.2 92.0 9.6 0.1 160

Nicosulfuron 282 3 103.9 27.7 103.5 22.2 0.1 205

Oryzalin 282 3 80.7 17.9 81.0 14.1 0.1 150

Oxamyl 282 3 64.1 19.4 66.0 16.9 0.0 144

Picloram 281 4 88.7 17.4 89.0 15.6 0.0 130

Propham 282 3 90.9 16.3 90.0 13.3 0.1 161

Propiconazole 281 4 102.4 20.2 105.0 14.1 0.1 157

Propoxur 282 3 83.6 14.2 85.0 11.1 0.1 126

Siduron 282 3 80.3 21.0 83.0 23.0 0.1 148

Sulfometuron-methyl 282 3 85.5 21.6 88.0 15.6 0.0 148

Tebuthiuron 282 3 100.4 16.1 101.0 11.1 0.1 172

Triclopyr 281 4 83.2 16.7 85.0 14.1 0.1 136

64 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 32. Long-term accuracy and precision calculated from multiple determinations of the method compounds fortified in 

sample-set spikes with the compounds added to organic-free water at 0.25 microgram per liter—Continued 

Compound N

Number 

of 

missing

values 

Mean

recovery

(percent)

Standard

deviation 

of 

recovery

(percent)

Median

recovery

(percent)

Fpseudosigma

of recovery

(percent)

Minimum 

recovery

(percent)

Maximum 

recovery

(percent)

Compounds qualified with an E-code because bias was outside acceptable range 

(median recovery less than 60 percent or greater than 120 percent)

2,4-DB 281 4 53.1 15.0 55.0 12.6 0.1 99

3-Ketocarbofuran 282 3 44.7 28.5 41.5 32.6 0.0 112

Aldicarb 281 4 11.4 15.5 4.0 13.3 0.0 66

Aldicarb sulfone 279 6 55.2 18.9 59.0 17.0 0.0 109 

Aldicarb sulfoxide 271 14 30.5 27.8 26.0 37.8 0.0 113 

Bentazon 278 7 38.6 35.4 25.0 43.6 0.0 121

Bromoxynil 281 4 60.3 61.4 52.0 51.9 0.1 890

Chloramben methyl ester 275 10 56.2 30.7 57.0 37.1 0.0 157 

Chlorothalonil 281 4 20.1 21.2 17.0 25.2 0.0 80

Cycloate 277 8 38.2 30.7 31.0 40.8 0.0 123

Flumetsulam 282 3 130.0 34.8 128.5 34.8 0.1 212

Imazaquin 277 8 123.0 36.9 124.0 26.7 0.1 259

MCPB 281 4 55.9 42.8 56.0 13.3 0.1 725

Methiocarb 282 3 41.6 36.6 30.0 53.9 0.0 159

Methomyl 282 3 43.2 32.9 39.0 46.0 0.0 152

Methomyl oxime 281 4 3.3 4.0 3.0 4.4 0.0 24 

Metsulfuron-methyl 282 3 25.3 30.7 11.0 31.9 0.0 105

Oxamyl oxime 271 14 10.3 9.9 10.0 11.9 0.0 65 

Tribenuron-methyl 278 7 6.1 12.2 0.0 5.9 0.0 117

Compounds qualified with an E-code because variation was outside acceptable range 

(f-pseudosigma greater than 25 percent)

2-Hydroxyatrazine 282 3 102.6 33.1 100.0 28.2 0.2 213

Bromacil 282 3 67.7 47.5 68.0 36.1 0.1 691

Deethylatrazine 282 3 69.2 27.7 69.0 30.9 0.1 132

Deethyldeisopropylatrazine 282 3 64.7 26.5 62.5 31.1 0.1 179

Deisopropylatrazine 282 3 67.3 29.2 67.0 30.4 0.1 149

Imazethapyr 229 56 113.5 54.3 115.0 35.6 0.0 336

Norflurazon 282 3 69.9 24.4 73.0 28.9 0.1 154

Terbacil 282 3 65.7 31.7 70.5 40.8 0.1 141

method had been implemented and the operators 

of the method experienced in its use. The results 

of these calculations are listed in table 32. 

Median recoveries for all compounds are plotted 

in figure 6 to provide a reference set of typically

acceptable performance. Median recovery and fpseudosigma limits are used as a guide, thereby

resulting in 27 compounds reported as qualified 

concentration estimates. 

Of these 27 compounds, nine are 

pesticide degradates. Two compounds, 

flumetsulam and imazaquin, had median 

recoveries greater than 120 percent. These 

two compounds are known to experience 

matrix enhancement of concentration when 

determined with a completely different 

SPE isolation method and by positive ion 

electrospray HPLC/MS (Furlong and 

others, 2000). Although there should be no 

matrix present in these set fortification 

samples, it is possible that matrix

carryover, or minor amounts of matrix

contamination that result from leaching

from the Carbopack SPE phase, might 

result in matrix enhancement. 

ANALYTICAL METHOD 65
66 Pesti

cides in Water by Graphitized Carbon-Based SPE and HPLC/MS

Figure 6. Median recoveries of method compounds from 285 laboratory reagent spikes determined from October 1, 1999 to September 30, 2000. Error bars are from the f-pseudosigma statistic.
For these 27 compounds, recoveries averaged 

55.5 percent for all compounds, with an 

average standard deviation of 29.7 percent. 

The average median recovery for all compounds was similar to the mean, at 53.5 

percent, and the average f-pseudosigma of 

recovery was 30.4 percent. 

The remaining 38 compounds met 

recovery performance criteria and were 

reported without qualification. For these 

compounds, recoveries averaged 87.3 percent 

for all compounds, with an average standard 

deviation of 20.1 percent. The average 

median recovery for all compounds was 

similar to the mean, at 88.2 percent, and the 

average f-pseudosigma of recovery was 14.7 

percent.

11.7 Laboratory reagent blanks

A total of 498 LRBs was analyzed while 

this method was used as a provisional custom 

method. A subset of 197 LRBs was reviewed in 

detail to determine the frequencies and 

concentrations of possible blank contamination. 

These results are listed in table 33. Twentyeight of 65 compounds measured in this method 

were detected in the set blanks. Except for

Table 33. Detections of compounds in 197 set blanks analyzed with this method 

[μg/L, microgram per liter] 

Compound Number of 

detections

Mean

concentration, 

in μg/L 

Median

concentration, 

in μg/L 

2,4-D methyl ester 1 0.0166 0.0166 

2-Hydroxyatrazine 1 .041 .0041

3(4-chlorophenyl)-1-methyl urea 1 .0018 .0018 

3-Hydroxycarbofuran 2 .0014 .0014

Atrazine 4 .0016 .0014

Bendiocarb 1 .0004 .0004

Benomyl 1 .0018 .0018

Bromacil 1 .0089 .0089

Caffeine 3 .0059 .0057

Carbaryl 2 .0023 .0023

Cycloate 4 .0109 .0106

Deethylatrazine 2 .0011 .0011

Deethyldeisopropylatrazine 1 .0015 .0015

Dinoseb 5 .0011 .0011

Diphenamid 1 .0007 .0007

Diuron 6 .0109 .0047

Fenuron 38 .0116 .0059

Fluometuron 4 .0018 .0020

Imazaquin 7 .0054 .0073

Imazethapyr 3 .0295 .0298

Metalaxyl 2 .0006 .0006

Methiocarb 2 .0275 .0275

Neburon 3 .0013 .0011

Norflurazon 5 .0018 .0014

Propiconazole 12 .0011 .0011

Siduron 1 .0014 .0014

Sulfometuron-methyl 5 .0036 .0041

Tebuthiuron 6 .0016 .0015

METHOD ANALYSIS 67
fenuron, the frequency of detections ranged 

from 1 to 12 in 197 blanks. The median 

frequency of detection in all blanks, except 

for fenuron, was 2.5 detections. Detections 

were clustered in 38 blank samples. In total, 

93 detections of all compounds were 

measured, a rate of 0.73 percent, calculated 

on the basis of 12,805 individual compound 

determinations made for 65 compounds in 

197 blank samples. 

One compound, imazethapyr, was 

detected one time at a concentration greater

than its median method laboratory reporting

level (LRL is calculated as twice the method 

detection limit; see section 11.8). Only 11 of 

the method analytes detected were at 

concentrations greater than 0.0025 μg/L. For 

these 11 compounds (sulfometuron-methyl, 

norflurazon, methiocarb, imazethapyr, 

imazaquin, diuron, cycloate, caffeine, 

bromoxynil, 2-hydroxyatrazine, 2,4-D), the 

median detected concentration was 0.0018 

μg/L. 

Fenuron was determined in 38 of 197 

blanks, a rate of 19 percent, and was the only

compound identified frequently enough to be 

considered a chronic contaminant for the 

method. It is unclear whether this chronic 

contamination was a result of actual fenuron 

contamination or the result of a coeluting

interference that contributed ions that were 

identical to those used for the selected-ion 

monitoring determination of fenuron. For 

chronic contamination of this type, the 

statistics of the distribution should be 

calculated, the 95th percentile of the

distribution determined, and the result used to 

qualify detections in field samples. 

11.8 Method detection limits 

11.8.1 Method detection limits 

(MDLs) were determined for this method 

using the procedures of the U.S. 

Environmental Protection Agency (1997). 

Data for the MDL determination were for the

lowest concentration of fortification used to 

determine method recoveries (0.025 g/L for 

the organic-free samples and the groundwater samples, and 0.05 g/L for the surfacewater samples). The calculated MDLs are 

listed in table 34. The MDLs calculated for 

the predecessor method of Werner and others 

(1996) are also listed in table 34. The 

extraction part of this method is based on that 

report. There are 30 compounds in the 

present method that are not included in the 

method by Werner and others; the latter 

contains six compounds not in the present 

method. These exceptions are listed 

separately at the bottom of table 34. 

11.8.2 On average, the MDLs for 

both methods were comparable. The mean 

MDLs for all compounds in the present 

method were 0.019 ± 0.090, 0.007 ± 0.012, 

and 0.017 ± 0.010 μg/L for the organic-free 

water, ground-water, and surface-water 

samples, respectively. The mean MDLs for 

all compounds in the method by Werner and 

others (1996) were 0.014 ± 0.006, 0.018 

± 0.009, and 0.029 ± 0.023 μg/L for organicfree water, ground-water, and surface-water 

samples, respectively. The median MDLs for

the present method were 0.007, 0.003, and 

0.014 μg/L for the organic-free water, 

ground-water, and surface-water samples, 

respectively. These MDLs compare 

favorably with median MDLs of 0.007, 0.009, 

and 0.026 μg/L for organic-free water, 

ground-water, and surface-water samples, 

respectively, in the method by Werner and 

others (1996). The mean MDL for all 

analytes in the present method, regardless of 

long-term performance, was 0.033 g/L.

11.8.3 If the comparison of MDLs is 

limited to the 35 compounds common to both 

methods, the data suggest that this new 

method offers improved detection at lower 

concentrations. In the new method, the mean 

MDLs for the 35 compounds were 0.008 

0.004, 0.004 0.002, and 0.016 0.011 g/L

for organic-free water, ground-water, and 

surface-water samples, respectively.

68 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
Table 34. Method detection limits for the method described in this study and the predecessor method of

Werner and others (1996)

[Concentrations in microgram per liter (μg/L). MDL, method detection limit; nd, not determined] 

Compound 

This

method, 

MDL in

organicfree 

water

Werner

method, 

MDL in

organicfree 

water

1

This

method, 

MDL in

groundwater

samples 

Werner

method, 

MDL in

groundwater

samples1

This

method, 

MDL in

surfacewater

samples 

Werner

method, 

MDL in

surfacewater

samples1

2,4-D 0.0109 0.013 0.0057 0.023 0.0463 0.021

2,4-DB .0080 .013 .0055 .015 .0091 .012

3-Hydroxycarbofuran .0029 .014 .0074 .015 .0171 .026

Acifluorfen .0033 .008 .0043 .021 .0094 .022

Aldicarb .0198 .016 .0036 .014 .0217 .020

Aldicarb sulfone .0098 .021 .0028 .016 .0227 .037 

Aldicarb sulfoxide .0041 .016 .0023 .019 .0056 .043 

Bentazon .0055 .014 .0032 .022 .0130 .029

Bromacil .0163 .011 .0033 .040 .0102 .050

Bromoxynil .0085 .012 .0060 .011 .0155 .011

Carbaryl .0142 .008 .0020 .018 .0154 .016

Carbofuran .0028 .028 .0025 .028 .0103 .044

Chloramben methyl ester .0089 .011 .0032 .011 .0141 nd

Chlorothalonil .0173 .007 .0088 .012 nd .021

Clopyralid .0069 .018 .0034 .007 .0037 .023

Dacthal monoacid .0058 .017 .0029 .035 .0272 .024 

Dicamba .0064 .011 .0027 .025 .0036 .021

Dichlorprop .0069 .032 .0052 .018 .0096 .043

Dinoseb .0060 .010 .0028 .032 .0061 .013

Diuron .0075 .012 .0018 .012 .0196 .040

Fenuron .0158 .013 .0032 .017 .0224 .067

Fluometuron .0155 .010 .0021 .010 .0113 .038

Linuron .0072 .006 .0034 .016 .0206 .024

MCPA .0081 .014 .0032 .009 .0110 .016

MCPB .0077 .010 .0032 .009 .0101 .038

Methiocarb .0040 .026 .0031 .015 .0167 .058

Methomyl .0022 .017 .0036 .008 .0177 .047

Neburon .0060 .015 .0032 .015 .0146 .044

Norflurazon .0082 .024 .0021 .014 .0120 .024

Oryzalin .0088 .019 .0054 .008 .0488 .042

Oxamyl .0061 .018 .0044 .008 .0374 .042

Picloram .0099 .004 .0033 .011 .0098 .022

Propham .0048 .011 .0021 .011 .0184 .021

Propoxur .0040 .008 .0023 .033 .0078 .027

Triclopyr .0112 .010 .0038 .007 .0043 .021

ANALYTICAL METHOD 69
Table 34. Method detection limits for the method described in this study and the predecessor method of

Werner and others (1996)—Continued 

Compound 

This

method,

MDL in

organicfree water

Werner

method, 

MDL in

organicfree 

water

1

This

method, 

MDL in

groundwater

samples 

Werner

method, 

MDL in

groundwater

samples1

This

method, 

MDL in

surfacewater

samples 

Werner

method, 

MDL in

surfacewater

samples1

Compounds analyzed in the present method only

2,4-D methyl ester 0.0043 0.0049 0.0177 

2-Hydroxyatrazine .0040 .0045 .0146

3(4-chlorophenyl)-1-methyl urea .0121 .0034 .0124 

3-Ketocarbofuran .0071 .0964 .0015

Atrazine .0045 .0028 .0135

Bendiocarb .0126 .0035 .0147

Benomyl .0019 .0047 .0135

Bensulfuron-methyl .0079 .0108 .0246

Caffeine .0048 .0019 .0346

Chlorimuron-ethyl .0048 .0048 .0320

Cycloate .0065 .0097 .0198

Deethylatrazine .0141 .0029 .0099

Deethyldeisopropylatrazine .0222 .0019 .0093

Deisopropylatrazine .0052 .0029 .0100

Diphenamid .0132 .0042 .0126

Flumetsulam .0057 .0096 .0284

Imazaquin .0078 .0306 .0209

Imazethapyr .0084 .0060 .0447

Imidicloprid .0034 .0101 .0383

Metalaxyl .0100 .0029 .0122

Methomyl oxime .0053 .0031 .0053 

Metsulfuron-methyl .0123 .0172 .0146

Nicosulfuron .0065 .0104 .0380

Oxamyl oxime .0067 .0158 .0106 

Propiconazole .0105 .0050 .0163

Siduron .0084 .0029 .0143

Sulfometuron-methyl .0044 .0092 .0210

Tebuthiuron .0031 .0086 .0141

Terbacil .0049 .0020 .0112

Tribenuron-methyl .0044 .0064 .0216

Compounds analyzed in the method of Werner and others (1996) but not determined in 

the present method 

1-Naphthol 0.007 0.023 0.027

2,4,5-T .010 .037 .027

Dichlobenil .012 .12 .16

DNOC .006 .015 .011

Esfenvalerate .019 .033 .039

Silvex .021 .021 .015

1

Werner, S.L., Burkhardt, M.R., and DeRusseau, S.N., 1996, Methods of analysis by the U.S. Geological Survey

National Water Quality Laboratory—Determination of pesticides in water by Carbopak-B solid-phase extraction and 

high-performance liquid chromatography: U.S. Geological Survey Open-File Report 96-216, 42 p. 

70 Pesticides in Water by Graphitized Carbon-Based SPE and HPLC/MS
The mean MDLs for the same compounds in 

the method by Werner and others (1996) were 

0.014 0.006, 0.017 0.009, and 0.031 

0.014 g/L for organic-free water, groundwater, and surface-water samples, 

respectively. The median MDLs of the 

common compounds for the present method 

were 0.008, 0.003, and 0.014 g/L for the 

organic-free water, ground-water, and 

surface-water samples, respectively. These 

compare favorably with median MDLs of 

0.006, 0.009, and 0.026 g/L for organic-free 

water, ground-water, and surface-water 

samples, respectively, in the method by

Werner and others (1996). 

The difference between the mean 

and standard deviations for all compounds in 

relation to mean and standard deviations for 

the compounds common to both methods is 

likely the result of the additional compounds 

in the present method being more difficult to 

isolate, more subject to degradation during

sample preparation, or more difficult to 

resolve chromatographically. These effects 

would likely introduce additional variation to 

the concentrations measured. The MDLs for 

these difficult compounds would be greater 

than for the compounds common to both 

methods because the MDL determination is a 

function of standard deviation. The method 

documented in this report is likely to result in 

more frequent detections at comparable or 

lower concentrations than the method of 

Werner and others (1996), given that 

HPLC/MS offers more specific detection in

the presence of matrix interferences than does 

HPLC with ultraviolet detection. 

CONCLUSIONS

Solid-phase extraction coupled with 

high-performance liquid chromatography/ 

mass spectrometry (HPLC/MS) analysis can 

be used to determine and quantify 65 polar 

pesticides and pesticide degradates in surfaceand ground-water samples, at concentrations 

as low as 0.002 g/L (microgram per liter) in 

environmental water samples. This method 

has comparable or better method detection 

limits than the existing complementary

method of Werner and others (1996), with the 

additional advantage that 65 compounds from 

25 pesticide classes are measured compared 

to 40 compounds in 15 pesticide classes in the 

Werner method. The single-operator mean 

standard deviation at 0.025 μg/L in organicfree water samples is 11 percent. Recoveries 

in organic-free water samples ranged from 28 

to175 percent, with elevated recoveries 

reflecting apparent matrix enhancement, 

likely from elevated dissolved organic carbon 

concentrations. The multiple operator (five),

multiple instrument (five) mean relative

standard deviation is 24.2 percent for all 

compounds in 285 organic-free water samples 

fortified at 0.25 μg/L and analyzed for 1 year 

from October 1999 through September 2000. 

Mean recoveries in these same samples were

73.8 percent for all compounds. The mean 

long-term set fortification recovery of the 38 

compounds in the method reported without 

qualification ranged between 60 and 120 

percent. An interference corresponding to the 

chromatographic and mass spectrometric

characteristics of fenuron was identified as a 

chronic contaminant and required qualified 

reporting of this compound. Eleven 

compounds were detected sporadically in 

blanks at concentrations greater than 0.025 

g/L. The single operator, single instrument 

validation data and the long-term qualitycontrol data reported here provide strong

evidence for the application of HPLC/MS to

large-scale, routine monitoring programs for 

pesticides in surface and ground water at 

concentrations as low as 10 parts per trillion 

(0.010 g/L or 10 ng/L).

CONCLUSIONS 71
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