6450

6450 NITROSAMINES*
6450 A. Introduction
1. Sources and Significance
The chemical N-nitrosodimethylamine (NDMA) has been
identified as a probable human carcinogen by the U.S. Environ-
mental Protection Agency. It is commonly found as a byproduct
of industrial materials, such as liquid rocket fuel, lubricants, and
pesticides. The general public can be exposed to NDMA by
sources including outdoor air, tobacco smoke, rubber products,
cosmetics, and dietary consumption (e.g., cured meats, fish,
cheese, and beer). The average concentrations of NDMA mea-
sured in various foods range from 90 to 100 ng/L for pasteurized
milk, 600 to 1000 ng/kg for fried pork bacon, and 50 to
5900 ng/kg for beer. NDMA also has been detected in recycled
water, wastewater, and potable water, and it may be formed by
chlorine and chloramine disinfection processes.
1,2
The formation
mechanisms that produce NDMA also may form other nitro-
samines. Seven other nitrosamines (see Table 6450:I) that are
structurally related to NDMA and in some instances known to
cause cancer in animals have been included in this method.
2. Selection of Method
Two extraction methods are presented here. Method 6450B is
a solid-phase extraction (SPE) method using a granular carbo-
naceous adsorbent resin. Method 6450C is a micro-liquid-liquid
extraction (MLLE) method. The SPE method has a concentration
factor of 500- to 1000-fold and can typically achieve method
detection levels (MDL) in the 0.5- to 2.0-ng/L range, depending
on the detection system. The MLLE method achieves a maxi-
mum concentration factor of 200-fold with MDL in the 2- to
4-ng/L range. Although the MLLE method generally has a
higher MDL than the SPE method, it may be adequate for many
wastewater samples and formation potential samples, and is
especially useful when sample volume is limited.
A recent report
3
provides additional details on the methods
presented here, as well as other methods potentially useful in
nitrosamine analysis.
The methods presented here are applicable to the analysis of
wastewater, recycled water, and potable water. The methods use
extraction procedures followed by analysis with gas chromatog-
raphy (GC) /chemical ionization (CI) tandem mass spectrometry
(MS/MS) for the separation and detection of eight nitrosamine
compounds. The GC/MS/MS conditions are discussed in detail
in Method 6450B, but are also applicable to the extracts from
Method 6450C. Either acetonitrile or methanol may be used as
the CI reagents.
3. References
1. CHOI, J. & R.L. VALENTINE. 2002. Formation of N-nitrosodimethyl-
amine (NDMA) from reaction of monochloramine: A new disinfec-
tion by-product. Water Res. 36:817.
2. MITCH, W.A. & D.L. SEDLAK. 2002. Formation of N-nitrosodimeth-
ylamine (NDMA) from dimethylamine during chlorination. Environ.
Sci. Technol. 36:588.
3. CHENG, R.C., C. ANDREWS-TATE, C.J. HWANG, Y.C. GUO, S.J. PASTOR,
J.E. GREBEL & I.H. SUFFET. 2005. Alternative Methods for the Anal-
ysis of NDMA and Other Nitrosamines in Water and Wastewater.
WateReuse Foundation, Alexandria, Va.
6450 B. Carbonaceous-Resin Solid-Phase Extraction GC/MS Method
This method is applicable to the determination of N-nitrosodi-
methylamine (NDMA) and seven other nitrosamines at concen-
trations of 2 to 300 ng/L, in raw source water, finished water,
wastewater, and recycled water.
1. General Discussion
a. Principle: This method, based on a previously published
procedure,
1
entails extraction of a sample to which isotopically
labeled surrogates have been added through adsorption onto a
carbonaceous resin, desorption of nitrosamines from the resin
with dichloromethane (DCM or methylene chloride), and anal-
ysis of the resulting solution by gas chromatography (GC)/
tandem mass spectrometry (MS/MS) (GC/MS/MS) with chem-
ical ionization (CI).
Isotope dilution/internal standard quantitation is implemented
with the addition of three isotopically labeled internal standards
(d
6
-NDMA, d
14
-NDPA, and
15
N
2
-NDEA) before extraction to
correct for extraction, as well as instrument variations. A large
injection volume of sample extract (8
␮ L) is recommended for
increased sensitivity, yielding detection levels in the low part-
per-trillion levels.
In chemical ionization tandem mass spectrometry (MS/MS) a
CI reagent (acetonitrile or methanol) is introduced into the ion
trap, ionized, and allowed to react with sample molecules. Re-
actions between the ionized CI reagent and sample molecules
result in various product ions, (M⫹H)
⫹
. The ion corresponding
to the analyte of interest is isolated from other matrix ions in the
trap and is selected as the precursor ion for subsequent fragmen-
tation. CI is a softer ionization technique than electron impact
* Approved by Standard Methods Committee, 2007. Editorial revisions, 2011.
Joint Task Group: Robert C. Cheng (chair), Cordelia J. Hwang, Cecilia O. Lei,
Irwin H. (Mel) Suffet, Vincent Y. Taguchi.
1

ionization, resulting in less sample molecule fragmentation and
consequently a higher concentration of the desired precursor ion.
The fragmentation process is induced by a waveform applied to
the trap, which increases the energy of the isolated precursor ion.
The amplitude of this waveform is called the collision induced
dissociation (CID) excitation amplitude. As the energy of the
precursor ion increases, chemical bonds are broken and product
ions (daughter ions) of lower mass than the precursor ion are
formed. Quantitation is performed by using the product ions
shown in Table 6450:I. Use of the product ions is of particular
importance in the analysis of wastewater samples or matrices
with interfering compounds.
Identification of NDMA and the other nitrosamines is based
on their retention times, parent ion isolation, and fragmentation
patterns. Further information about this method is available
elsewhere.
1,2
b. Interferences: Method interferences may be caused by
contaminants, especially from NDMA in solvents, reagents (in-
cluding reagent water), sample bottles and caps, and other sam-
ple processing hardware that lead to discrete artifacts and/or
elevated baselines in the chromatograms. The samples or ana-
lytical system also may be contaminated by rubber products in
the work area.
Preserve samples properly to avoid inaccurate target analyte
recovery due to formation of nitrosamines caused by the pres-
ence of residual chloramine.
Clean all glassware meticulously by washing with detergent
and tap water, rinsing with tap water, and rinsing again with
reagent water. A final solvent rinse may be needed. In place of
a solvent rinse, nonvolumetric glassware can be heated in a
muffle furnace at 400
o
C for 2 h. Rinse volumetric glassware with
methanol and dry in a fume hood. Store glassware in an inverted
position or capped with aluminum foil to prevent contamination.
Condition and store resin as in 6450B.4dbelow. Proper resin
storage is critical because evidence shows that nitrosamines can
be transmitted to the resin through the gaseous phase.
Coeluting GC peaks with nominal masses equivalent to the
target analytes and internal standards have been observed in
potable water and wastewater samples, requiring MS/MS or
high-resolution MS for quantitation. Surfactants and other or-
ganic contaminants in wastewater may cause emulsions during
extraction. Analyze a reagent-water blank under the same con-
ditions as the samples to demonstrate an absence of interfer-
ences. NDMA has been found in deionized (DI) water at levels
up to 10 ng/L; therefore, use an ultraviolet water purification
system to avoid contamination. The blank concentration must be
equal to, or less than, one-half the minimum reporting level
(MRL) or one and one-half the MDL value.
Carryover may be observed during GC analysis. To avoid
contamination, a solvent injection may be required between
high-concentration and low-concentration sample analyses.
c. Safety: Because DCM and most nitrosamines, including
NDMA, have been identified as animal carcinogens and some as
probable human carcinogens, minimize exposure to these com-
pounds and their isotopically labeled analogs. Keep a reference
file of current material safety data sheets (MSDS), and make it
available to all personnel involved in analyses.
Pass effluents of GC sample splitters and GC/MS vacuum
pumps through a column of activated carbon, bubble effluents
through a trap, or use some other means to remove possible
contamination.
The following precautions for safe handling of NDMA and
other nitrosamines in the laboratory are presented as guidelines
only. Additional information is available elsewhere.
3,4
Use laboratory hood, safety glasses, disposable plastic gloves,
and apron or lab coat. Thoroughly wash hands and forearms after
each sample analysis. If contact of personnel with the chemicals
occurs, scrub the affected area with any mild soap. Wash con-
taminated glassware, tools, and surfaces with detergent and
water. Minimize solvent waste during the analytical process.
Handling the dilute solutions normally used in analytical work
presents no significant inhalation hazards except in case of an
accident.
d. Detection levels: Method detection levels (MDL) (see Sec-
tion 1030C) for the analytes (Table 6450:I) listed using the SPE
method in deionized (DI) water are provided in Table 6450:II.
Prepare at least seven replicates of laboratory-fortified blanks
with nitrosamine standards at a concentration of 1 to 5 ng/L and
extract over a 3-d period. Calculate mean recovery and standard
deviation for each analyte. Multiply the Student tvalue at the
99% confidence level and n-1 degrees of freedom (e.g., 3.143 for
seven replicates) by the standard deviation to yield a statistical
estimate of the MDL.
TABLE 6450:I. TARGET NITROSAMINE ANALYTES:FORMULA,MOLECULAR WEIGHT,INTERNAL STANDARD,AND QUANTITATION ION
Nitrosamine Abbreviation Formula Mol Wt Internal Standard
Quantitation Ion*
amu
N-nitrosodimethylamine NDMA C
2
H
6
N
2
O74 d
6
-NDMA 44
N-nitrosomethylethylamine NMEA C
3
H
8
N
2
O88
15
N
2
-NDEA 61
N-nitrosodiethylamine NDEA C
4
H
10
N
2
O102
15
N
2
-NDEA 75
N-nitrosodi-n-propylamine NDPA C
6
H
14
N
2
O130 d
14
-NDPA 89
N-nitrosomorpholine NMOR C
4
H
8
N
2
O
2
116 d
14
-NDPA 86
87†
N-nitrosopyrrolidine NPYR C
4
H
8
N
2
O100 d
14
-NDPA 55
N-nitrosopiperidine NPIP C
5
H
10
N
2
O114 d
14
-NDPA 69
N-nitrosodi-n-butylamine NDBA C
8
H
18
N
2
O158 d
14
-NDPA 57
103†
* Methanol used as CI reagent; quantitation ions are also used for acetonitrile as CI reagent unless otherwise noted (see below).
† Acetonitrile used as CI reagent.
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method
2
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method

2. Sampling and Storage
This section describes sampling and storage protocols. More
detailed information on sampling and storage can be found in
other reports.
5,6
a. Sample collection: Ensure that sample site is free of auto-
motive exhaust, cigarette smoke, fresh paint, and any other
possible source of contamination. Sample location should pro-
vide for collection of a representative grab sample or composite
(maintained cold and with proper disinfectant residual quench-
ing).
When sampling from a water tap, let tap run until the water
temperature has stabilized (usually about 3 to 5 min). When
sampling from an open body of water, collect sample in a clean
stainless steel bucket with a clean rope. Avoid plastic and rubber
tubing, gaskets, or other equipment that may leach interfering
analytes into the water sample.
Collect samples in pre-cleaned amber glass bottles with TFE-
lined polyethylene caps. Collect sufficient sample volume for the
analysis of the sample, a laboratory-fortified matrix (LFM), and
LFM duplicate for quality assurance purposes, preferably in two
or more bottles. Fill sample bottles to the top, but take care not
to flush out the preservative.
When sampling chlorinated or chloraminated water, quench
the residual at time of sampling. Add solid sodium sulfite or
sodium thiosulfate (approximately 500 mg for wastewater, or 40
to 100 mg for potable water) to a 1-L bottle to minimize
additional nitrosamine formation. For potable water samples,
20 mg of ascorbic acid may be used. Use more quenching agent
if the chloramine residual is greater than 4 mg/L. Do not rinse
sample bottle with sample before collection if bottle has been
prepared with preservatives before sampling.
b. Storage: Ice or refrigerate samples at 6°C or lower (but not
freezing) and maintain at these conditions, away from light, until
extraction. To prevent photo-decomposition, protect samples
from light from the time of collection until extraction; amber-
colored, glass bottles work well for this. Avoid storing samples
under low pH conditions because these conditions have been
observed to produce elevated levels of NDMA in some waste-
water effluents and potable waters. Degradation and formation of
nitrosamines in wastewater matrices can be a complicated pro-
cess and may vary from situation to situation. Therefore, select
appropriate holding times on the basis of case-specific data. As
a general guideline, extract potable water samples within 28 d
and wastewater samples within 14 d.
Store extracts at ⱕ10°C and keep them away from light in
amber glass vials with TFE-lined caps. Under the conditions
used during the method development, archived extracts generally
show minimal NDMA loss over 6 months.
3. Apparatus
a. Sample containers: 1-L amber glass bottles fitted with
PTFE-lined screw caps.
b. Standard solution storage containers: 10- to 20-mL amber
glass vials with PTFE-lined screw caps.
c. Vials: Screw-cap 2.0-mL amber glass autosampler vials
with PTFE-faced septa.
d. Volumetric flasks: Class A, various sizes used for prepara-
tion of standards and samples.
e. Microsyringes, glass with stainless steel needle and plunger
in various sizes for adding solutions and preparing intermediate
solutions.
f. Filters, disposable, used to isolate resin from the water
samples. Use either paper* or glass fiber† filters.
g. Balance, analytical, capable of accurately weighing to
0.0001 g.
h. Aluminum dishes, disposable.
i. Mixing apparatus‡ capable of shaking samples at 50 rpm
and accepting 1-L bottles. Alternatively, an orbital shaker capa-
ble of 200 rpm may be used for the 2-h extraction.
j. Vacuum filtration apparatus.
k. Gas chromatograph: Capillary GC with split/splitless tem-
perature-programmable injector capable of large-volume injec-
tions. GC oven and injector should be able to maintain 35°C.
l. Chromatographic column:§ Several types of capillary col-
umns㛳have been used successfully.
m. Autosampler: The autosampler used during method devel-
opment# was equipped with a side-port needle capable of slow
injection (approximately 0.2
␮ L/s) of 8- ␮ L volumes. Other
autosamplers capable of injecting this volume may be used.
n. Detector: Ultra-trace mass spectrometer, capable of chem-
ical ionization (CI), and tandem mass spectrometry with the
sensitivity to detect low part-per-billion levels of NDMA. One
such system,** using methanol or acetonitrile CI, was used for
development of this method.
4. Reagents
Use chemicals that are reagent-grade or better and high reso-
lution-gas chromatography (HR/GC)-grade solvents. Unless oth-
* Whatman, 55 mm, No.1 (11- ␮ m, Cat. No. 1001 055), No. 4 (20- to 25- ␮ m, Cat.
No. 1004 055), or equivalent.
† Whatman GF/F, 0.7-
␮ m, No. 1825 047, or equivalent.
‡ LE 2002 Heavy Duty, Lab-Line Instruments, Melrose Park, IL, or equivalent.
§ Gas chromatographic methods are extremely sensitive to the materials used.
Mention of trade names by Standard Methods does not preclude the use of other
existing or as-yet-undeveloped products that give demonstrably equivalent results.
㛳HP-VOC (used in method development), 60 m long ⫻0.32-mm ID, 1.8-
␮ m film
thickness; DB1701, 30-m long ⫻0.25-mm ID, 1.0-
␮ m film thickness; DB-VRX,
60 m long ⫻0.32-mm ID, 1.8-
␮ m film thickness.
# Varian 8200cx, Varian, Inc., Palo Alto, CA.
** Varian Saturn 2000, Varian, Inc., Palo Alto, CA.
TABLE 6450:II. METHOD DETECTION LEVELS FOR NITROSAMINES IN
REAGENT WATER,* SOLID-PHASE EXTRACTION
Nitrosamine
Fortification Level
ng/L
MDL
ng/L
NDMA 1.0 0.84
NMEA 1.0 0.45
NDEA 1.0 0.81
NDPA 1.0 1.08
NMOR 1.0 0.62
NPYR 1.0 0.83
NPIP 1.0 0.74
NDBA 1.0 0.71
* For n⫽10, t⫽2.821 at the 99% confidence level. SPE, HP-VOC column,
methanol CI, Ambersorb resin.
Courtesy: WateReuse Foundation.
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method
3
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method

erwise indicated, ensure that all reagents conform to the speci-
fications of the American Chemical Society’s Reagent Chemi-
cals,
7
where such specifications are available.
a. Reagent water: Use purified water that does not contain
target analytes or interfering compounds at levels greater than
one half the MRL for each compound of interest.
b. Methanol,CH
3
OH, high-purity, demonstrated to be free
from analytes or interferences.
c. Dichloromethane (DCM), CH
2
Cl
2
, high-purity, demon-
strated to be free from analytes or interferences. An alkene-
stabilized reagent is preferable to a cyclohexene-stabilized re-
agent because of possible interferences in the chromatogram
during analysis.
d. Carbonaceous adsorbent resin.†† Condition resin in a
shallow tray at a temperature above 250°C (300°C recom-
mended) for 3 h before use and store in a capped amber glass
bottle in a desiccator. To ensure a uniform size distribution of
beads, sieve with a No. 50 ASTM mesh or let the finer particles
settle in the storage container and take beads from the upper
portion only.
e. Helium, ultra-high-purity (UHP), as GC carrier gas.
f. Carbon dioxide, completely dry, with siphon tube, for in-
jector cryogenics.
g. Nitrogen, UHP grade, for autosampler pneumatics.
h. Sodium thiosulfate,Na
2
S
2
O
3,
or sodium sulfite, Na
2
SO
3
, for
use as dechlorinating agent.
i. Nitrosamine stock standard solutions: Prepare from pure
standard materials or purchase as certified solutions, available at
concentrations of 100 to 5000
␮ g/mL. To prepare a stock stan-
dard from a pure material, partially fill a volumetric flask with
methanol. Allow flask to equilibrate, weigh to nearest 0.1 mg,
add desired quantity of pure standard material with a microsy-
ringe, and reweigh. Dilute to volume, stopper, and mix by
inverting three times. Calculate concentration of stock standard
from net gain in weight. When compound purity is assayed to be
96% or greater, use the weight without correction to calculate
concentration of stock standard. Store stock standard solution in
an amber glass container at a temperature below 0°C for up to 3
months.
j. Nitrosamine primary dilution standard (PDS): Prepare a
nitrosamine mix PDS of a suitable concentration by accurately
transferring the appropriate volume of stock standard solution
into a volumetric flask partially filled with methanol. Dilute to
volume, mix thoroughly, transfer to an amber glass vial, and
store at ⱕ6°C. For example, a 1000 ng/mL PDS may be prepared
in 25-mL volumetric flasks and used to make a 100-ng/mL
solution through serial dilutions, which can then be used for
low-level known additions.
k. Nitrosamine internal standard and surrogate primary dilu-
tion standard (ISPDS): Use d
6
-NDMA, d
14
-NDPA, and
15
N
2
-
NDEA‡‡ as internal standards. Prepare an internal standard and
surrogate mix PDS of suitable concentration by accurately trans-
ferring the appropriate volumes of internal standard and surro-
gate stock solutions into a volumetric flask containing methanol
as described in ¶ jabove. For example, 1000 ng/mL PDS of
internal standard and surrogate were prepared in 25-mL volu-
metric flasks and stored in an amber glass vial at ⱕ6°C.
l. Nitrosamine calibration standards: This method uses a
procedural standard calibration curve. Prepare and extract solu-
tion to produce calibration curve with at least five points, as
outlined in 6450B.5c. Fortify a series of 500-mL reagent water
samples contained in 1-L amber bottles with the PDS and the
ISPDS to produce a calibration curve ranging from 2 to 300 ng/L
with internal/surrogate standards at 20 ng/L, as is shown in Table
6450:III.
5. Procedure
a. Sample extraction by solid-phase extraction (SPE): Remove
samples from storage and allow to equilibrate at room temper-
ature. Transfer 500 mL sample with a clean Class A graduated
cylinder into designated 1-L amber glass bottles. QC samples,
including method blanks, calibration standards, continuing check
standards, duplicates, and laboratory-fortified matrices, are pre-
pared in the same manner. If sample is a method standard,
laboratory-fortified blank (LFB), laboratory-fortified sample ma-
trix (LFSM), or LFSM duplicate (LFSMD), add appropriate
volume of intermediate nitrosamine mix (in methanol).
Add a portion of internal standard and surrogate ISPDS that
results in a 20-ng/L final concentration (for example 10
␮ Lofa
1000 ng/mL ISPDS). Place needle of syringe below water sur-
face when adding standards to an aqueous sample. After injec-
tion, cap bottle and invert to allow for mixing. Wipe needle
between samples and after standards to prevent cross-contami-
nation of the ISPDS.
Add 200 mg resin (6450B.4d) to the sample, place bottles in
mixing apparatus (6450B.3i), and rotate for2hat50rpmina
rotator apparatus or 200 rpm in an orbital-type shaker.
Rinse the filter paper and the filter holder with laboratory
water before filtering the sample. Filter the sample through filter
paper (6450B.3f) with the aid of a vacuum system under a hood
to isolate the resin from the water.
To ensure complete transfer of resin, rinse walls of bottle
thoroughly with reagent water (a squeeze bottle can be used for
†† Ambersorb® 572, Supelco P/N: 10432-U.
‡‡ Cambridge Isotopes, Cambridge, MA, or equivalent.
TABLE 6450:III. PROCEDURAL CALIBRATION STANDARDS*
Calibration
Standard
Concentration
ng/L
Volume of
100-ng/mL
Nitrosamine
Mix
␮ L
Volume of
1000-ng/mL
Nitrosamine Mix
␮ L
Volume of
1000-ng/mL
Internal
Standard and
Surrogate Mix
␮ L
15 — 10
210 — 10
525 — 10
10 — 5 10
20 — 10 10
50 — 25 10
100 — 50 10
300 — 150 10
* Add tabulated volumes of analyte and internal standard/surrogate PDS into 500 mL
reagent water to obtain the different concentrations of calibration standards.
Courtesy: WateReuse Foundation.
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method
4
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method

this purpose) and add to collected resin on the filter. Leave resin
in the filtration apparatus under vacuum for approximately 5 min
to remove as much water as possible.
With forceps, transfer the filter paper containing the resin to a
disposable aluminum dish. Air-dry resin for at least 45 min under
the hood or under a gentle stream of dry helium to minimize
contamination. Transfer dry resin to a 2-mL autosampler vial and
cap vial. Store vials containing resin in the refrigerator or freezer
until time of analysis.
Just before analysis, remove vials from refrigerator, immedi-
ately uncap, and add 400
␮ L of DCM with a microsyringe. Heat
is released when the solvent comes into contact with the resin.
To minimize this exothermic reaction, slowly deliver DCM
along the walls of the cold vial.
Cap the vial, gently shake to allow contact between resin and
solvent, and tap vial to expel air bubbles from resin.
Allow a half-hour contact time between resin and DCM to
ensure that desorption equilibrium has been reached.
Load vials onto autosampler tray for analysis, and set for 8-
␮ L
injection (suggested volume). The MS instrument conditions
given in this method are set for an 8-
␮ L injection volume. Adjust
instrument parameters as necessary if other injection volumes
are used.
b. Gas chromatography/mass spectrometry: Operating condi-
tions for several capillary columns are shown in Tables 6450:IV
and 6450:V (injection parameters), and Table 6450:VI (column
parameters).
For the CI analysis, default ionization parameters and a list of
typical ionized forms of various CI reagents and modified ion-
ization parameters for enhanced sensitivity are shown in Table
6450:VII.
Various parameters, including collision–induced dissociation
(CID) values, and precursor and product ions, are shown in
Table 6450:VIII (methanol CI), and Table 6450:IX (acetonitrile
CI). CID values were selected to maximize the transition from
precursor ion to product ions while retaining a minimum of the
precursor ion for confirmation. Because each instrument exhibits
different sensitivity, the percentage of product ion required to
achieve the desired sensitivity will vary. Therefore, follow the
general guideline of maximizing the percentage of product ions
while retaining a minimal amount of the precursor ion for
confirmation. In some instances, the degree of fragmentation has
also been observed to depend on analyte concentration. Other
GC/MS/MS conditions may be used if QC requirements are met.
For each target and surrogate analyte, establish an appropriate
retention-time window and precursor to product ion mass ratio to
facilitate detection and identification in all QC and field samples.
A representative nitrosamine chromatogram is presented in
Figure 6450:1.
c. Calibration: Prepare standards as described in 6450B.4i
through l. Extract and analyze each standard under the same
conditions used for sample extracts, using internal standards
designated in Table 6450:I for quantitation of each nitrosamine.
A relative response factor (RRF) (equal to the relative signal area
ratio of the nitrosamine of interest versus that of the correspond-
ing nitrosamine internal standard) is used in the isotope dilution/
internal standard quantitation technique. The concentration of a
specific nitrosamine in a sample is determined by comparing the
RRF for the nitrosamine in the sample with added internal
standard to the average RRF value determined for the set of
calibration standards. C/MS software can be used to generate a
linear regression or quadratic calibration curve plotting absolute
area ratios (nitrosamine of interest/nitrosamine internal standard)
against known-addition concentration for each nitrosamine. The
coefficient of determination (r
2
) for the plot should round to
0.995 or higher, and curves may be linear from the MRL up to
500 ng/L. A representative NDMA calibration curve is shown in
Figure 6450:2. Verify all initial calibrations with a standard from
a second source.
d. Continuing calibration: To perform continuing calibration
checks, verify the initial calibration by preparing and extracting
a midpoint calibration standard with each batch of 10 samples or
less. Analyze continuing calibration standards at the beginning
and end of each batch of 10 samples or less. If the sample run
contains more than 10 samples, analyze the continuing calibra-
tion standard after every 10 samples in the batch.
TABLE 6450:IV. GAS CHROMATOGRAPH INJECTION PROGRAM
TEMPERATURE CONDITIONS FOR NITROSAMINE ANALYSES
Column
Temperature
°C
Rate
°C/min
Hold Time
min
Total Time
min
HP-VOC 37 0 0.7 0.7
250 200 27.0 28.7
DB-1701 35 0 0.8 0.8
260 200 2.1 4.0
150 200 21.0 25.6
NOTE: Injection volume, 8 ␮ L; plunger inject speed, 0.2 ␮ L/s; post-injection
delay, 99 s.
TABLE 6450:V. GAS CHROMATOGRAPH INJECTION PROGRAM SPLIT
CONDITIONS FOR NITROSAMINE ANALYSES
Time
min Split State Split Ratio
0.0 On 5
0.8 Off —
2.2 On 100
20.0 On 30
Courtesy: WateReuse Foundation.
TABLE 6450:VI. GAS CHROMATOGRAPH COLUMN CONDITIONS FOR
NITROSAMINE ANALYSES
Column
Temperature
°C
Rate
°C/min
Hold Time
min
Total Time
min
HP-VOC 32 0 1.7 1.7
100 15 2 8.2
190 5 0 26.2
270 50 5 30.0*
DB-1701 35 0 4 4
200 15 0 15
240 40 10 26
* May be increased depending on the content of the sample.
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method
5
NITROSAMINES (6450)/Carbonaceous-Resin Solid-Phase Extraction GC/MS Method