Professional Documents
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6, 2007
Trends
1. Introduction
This article reviews gas chromatographymass spectrometry (GC-MS) and liquid
chromatography-mass spectrometry (LCMS)-based analytical methods used for the
determination of bioactive pharmaceuticals and personal-care products (PPCPs) in
the environment. MS methods reviewed
may use single-quadrupole MS, magnetic
sector tandem MS (MS2), triple-quadrupole (QqQ) MS2, ion-trap MS (IT-MS),
0165-9936/$ - see front matter Crown Copyright 2007 Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.trac.2007.02.011
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time-of-flight MS (TOF-MS), or hybrid quadrupole-TOFMS (qTOF-MS) in the analysis. PPCPs discussed in this
article refer to human and veterinary drugs and medical
diagnostic agents, as well as ingredients in personal-care
products (e.g., cosmetics, fragrances, food supplements,
shampoos, toothpastes, natural and synthetic compounds with endocrine-disrupting potency), and their
respective metabolites and transformation products.
Categorized as emerging organic pollutants (EOPs),
PPCPs have been the focus of global environmental
researchers attention since the late 1990s. PPCPs have
caused widespread concerns due to their extensive human and veterinary consumption, excretion or washing
off their hosts after their intended usages, and their entry
into the environment through effluents of wastewatertreatment plants (WWTPs), as well as surface-water runoffs and soil leaching after agricultural applications of
manure or treated sludge. As PPCPs were designed to
correct, enhance, or protect a specific physiological or
endocrine condition, their target effects in humans and/
or farm stocks are well understood and documented.
However, there is limited knowledge about their unintended effects in the environment. The fact that PPCPs
are in the environment and the lack of understanding in
their combined effects on non-target hosts is at the root
of the concerns that they cause.
To address the occurrence, the distribution and the
fate of PPCPs in the environment, efficient and reliable
analytical methods are needed. The relatively low concentration, high polarity, and thermal lability of PPCPs,
together with their interaction with a host of complex
environmental matrices, make their analysis challenging. Sample preparation followed by GC or HPLC separation, and qualitative and quantitative analysis using
various detectors has become the standard approach.
Compared to non-specific detectors (e.g., electron capture, ultraviolet or fluorescence), MS has the inherent
advantages of high selectivity, specificity, and sensitivity
and has been the technique of choice for PPCP analysis.
GC-MS, GC-MS2, LC-MS and LC-MS2 have become
indispensable tools for investigating PPCPs in environmental matrices. MS also allows the use of isotope-labeled compounds (ILCs), if available, to correct for matrix
effects and the uncertainty introduced during sample
preparation and analysis, further enhancing data
quality.
Since the first GC-MS determination of clofibric acid, a
metabolite of lipid-regulator clofibrate, in WWTP effluents [1], environmental scientists have reported the
occurrence of more than 80 PPCPs in the environment,
including soil, sediment, sludge (biosolid), manure,
treated and untreated sewage, surface water, groundwater, and drinking water. These PPCPs included
anti-inflammatory drugs, pain-killing drugs, cholesterollowering drugs, antibiotics, antiepileptic, anticonvulsants,
hormones, beta-blockers, lipid regulators, hypnotics,
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2. Sample preparation
Environmental concentrations of PPCPs are mostly in
the ng/Llg/L or pg/gng/g ranges, depending on
sample matrices. Analytes in samples therefore need to
be extracted and concentrated prior to instrumental
analysis. For aqueous sample matrices, solid-phase
extraction (SPE) has replaced traditional liquid-liquid
extraction (LLE) and become the sample-preparation
technique used most, as SPE allows sample extraction
and clean-up to be conducted at the same time. SPE
cartridges packed by different sorbents (e.g., C-18, nonpolar phase [2], ion-exchange phase [3], and polymeric
phase [4]) have been used to extract target PPCPs.
Waters Oasis HLB (Hydrophilic-Lipophilic Balanced)
cartridge has been the cartridge of choice for the preconcentration of both polar and non-polar compounds
using the same extraction conditions, a pre-requisite for
multi-residue analysis of different PPCPs. Novel polymer
sorbents have also been used in solid-phase microextraction (SPME), where smaller sample volumes and
shorter extraction times are required [5,6].
On-line, automated SPE procedures coupled with LCMS or LC-MS2 have been widely used in drug-discovery
laboratories to enhance data quality and operational
efficiency, and have been applied to the analysis of
PPCPs in aquatic environmental samples. This was done
by interfacing an automated SPE directly to LC-MS or
LC-MS2 to analyze low-ng/L levels of antibiotics in surface water, including sulfonamides [7], fluoroquinolones
and penicillins [8]. In addition, a new SPE-LC-MS2 system was also reported for the determination of estrogens
in natural waters with limits of detection (LODs) at the
ng/L levels [9]. On-line SPE eliminates time-consuming
and costly off-line SPE, enables higher throughput for
routine PPCP analysis, reduces human interactions with
the sample, and can be a useful tool to produce data
consistently with high quality.
For solid matrices (e.g., soil, sediment, sludge and fish
tissue), antibiotics are the most analyzed compound
class, although there are a few reports about other drugs
and endocrine-disrupting compounds (EDCs). Various
methods have been selected for the extraction of antibiotics and EDCs from solid matrices. These include initial
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Table 1. Summary of GC-MS and GC-MS2 methods for the quantitative analysis of PPCPs
Class/compound
Sample matrix
Derivatization agents
MS system
Environmental
concentrations (ng/L)
Ref.
Estrogens
Surface
water,wastewater
BSTFA
pptppb
[10]
Estrogens
Environmental water
0.165
[23]
Wastewater
Surface & treated
water
Wastewater
Sewage water
1356
10107
[53]
[54]
17313
3036,530
[55]
[56]
75400 ng/g
[21]
502,500
[32]
5560
[31]
1925,700
[28]
0.1135
[26]
0.229
[27]
53500
[57]
101100
[58]
2.98059
[59]
30420
[60]
NSAIDs
Estrogens, acidic drugs,
personal-care products
Triclosan & related
compounds
Phenazone drugs
Pharmaceuticals
Estrogens
Estrogens
Estrogens
Acidic/neutral drugs
Acidic drugs
Acidic drugs/caffeine
Acidic/neutral drugs
MSTFA
MTBSTFA
Acetic anhydride/
MTBSTFA
PFBBr
Groundwater, swine
lagoon
Surface & drinking
waters
surface & sewage
water
Surface & drinking
waters & wastewater
Groundwater
PFBBr/TMSI
Wastewater &
receiving water
Environmental water
PFBCl
Diazomethane
Methanol/BF3
Tetrabutylammonium
(TBA) salts
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ability allowed the collection of high-sensitivity, fullscan MS spectra, high MS resolution, and the ability to
perform both MS and MS2 analysis functions. Following
IT-MS is the single quadrupole mass-selective detector
(MSD) because of its low cost and high availability in
environmental laboratories. Although GC-TOF-MS is
commercially available and has been used for environmental studies, it has not been applied for the
analysis of PPCPs. On the contrary, the application of
TOF-MS started to boom in the LC-MS and LC-MS2
analysis of PPCPs, especially after the advent of new
qTOF-MS.
In general, GC-MS analysis acquires data in full-scan
and/or SIM modes for identification and quantitation
purposes, respectively [23,31,32]. By carefully choosing
characteristic fragment ions with high intensity, unique
m/z, and low background (i.e., not being interfered by
column bleedings), GC-MS SIM analysis can provide
good sensitivity and selectivity for quantitative PPCP
analysis. However, when dealing with complex matrices
(e.g., sewage and sludge) where co-extracted components from the matrix may cause difficulty in GC-MS
quantitative analysis, GC-MS2 becomes the preferred
Class/compound
Sample matrix
Additive(s) in LC mobile
phase
MS system
Environmental
concentrations (ng/L)
Ref.
Formic acid
178,800
[39]
Wastewater, sludge
Acetic acid,
triethylamine
[10]
3,100138,000
(water)1,3008,500
ng/g (sludge)
3.7198.7 ng/g
[11]
10270 ng/g
[12]
0.0822.8 ng/g
[19]
N/A
[20]
0.7 ng/g
[51]
580600
[38]
[47]
0.71,572
[44]
5295
[2]
10100
[3]
0.220.68
[9]
665,533
[36]
60170
1001300
[41]
[4]
103000
[7]
200450 ng/g
[14]
12197 ng/g
[15]
1.91200
[46]
Tetracyclines, tylosin
Sulfonamides & metabolites,
trimethoprim
Estrogens, progestogens
River sediment
Soil
Tetracyclines, fluoroquinolones
Acetic acid
Estrogens
Wastewater
Estrogens, bisphenol A
Ammonium acetate,
formic acid
Various PPCPs
River sediment
Ammonium acetate
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Tri-n-butylamine
Tetracyclines, sulfonamides
Sulfonamides, trimethoprim,
fluoroquinolone, tetracyclines,
macrolides, caffeine
Sulfonamides, acetyl metabolites
Surface water
Surface water, groundwater
Formic acid
Formic acid
Surface water
Sludge
Sewage sludge
Formic acid
Wastewater
Ammonium acetate
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Estrogens
Table 2. Summary of LC-MS and LC-MS2 methods for the quantitative analysis of PPCPs
574
[49]
[58]
10122,000
181860
251100
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Ammonium acetate
[48]
46215
Formic acid
[40]
530 ng/g
1105400
Thermo Electron TSQ triple quadrupole, ESI(+), MRM
Waters q-TOF-Micro, ESI(+), ESI(), MS, MS2
Aged agricultural soil
Wastewater
Acetic acid
Ammonium acetate,
acetic acid
Formic acid
[18]
[37]
[16]
[17]
0.0680.227 ng/g
155700 ng/g
Sulfonamides, penicillins
Tetracyclines, sulfonamides,
macrolides
Sulfonamides & metabolites
Various PPCPs
Formic acid
Formic acid
[35]
27215,500
Formic acid
[8]
54
Formic acid
Sample matrix
Class/compound
Table 2 (continued)
Additive(s) in LC mobile
phase
Ref.
Environmental
concentrations (ng/L)
MS system
Trends
The technology using an atmospheric pressure ionization source (API) as an interface between an LC and an
MS or MS2 (i.e. the modern LC-MS or LC-MS2) has
gained great popularity due to its compatibility with
polar, non-volatile and thermally labile PPCPs since the
late 1980s. Over the past decade, various mass spectrometers have all been interfaced to liquid chromatographs, and applied to analyze PPCPs extracted from
aqueous and solid environmental matrices. Each MS
analyzer possesses its own specificity and applicability in
the qualitative confirmation and quantitative determination of the PPCP-target analytes; however, QqQ-based
LC-MS2 systems have the unique ability of doing SRM
with high SNR, have attractive price/performance ratio,
are relatively easy to maintain, and have become the
norm in PPCP analysis. Table 2 lists MS2 operational
parameters, matrix types, LC mobile phase, target analytes and their environmental concentrations in 30 LCMS and LC-MS2 studies.
Although MS provides a second dimension for the
confirmation of target analytes, LC separation is needed
to improve detection performance, especially when
dealing with isomeric chemicals. Reversed-phase (e.g.,
C18 and C8) analytical columns are most commonly used
in the separation of PPCPs. The inner diameter of 2 mm
is usually selected for analytical columns being used for
LC-MS or LC-MS2. The pH of the aqueous mobile phase is
normally adjusted with formic acid, acetic acid, ammonium hydroxide, ammonium formate and/or ammonium acetate, while the organic mobile phase employs
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5. Matrix effect
The matrix effect exists in analysis by GC-MS, GC-MS2,
LC-MS and LC-MS2, and it affects the data quality of
target-compound analysis. It could be caused by those
co-eluted, interfering components in the sample extract
that have similar ions in the MS or MS2 experiment. It
might also arise from the interaction between the target
analytes and those co-extracted matrix components
(organic or inorganic) during sample preparation and in
the ionization chamber. The former is common in GC-MS
and GC-MS2 analysis, and might be encountered rarely
in LC-MS and LC-MS2 analysis. It can be resolved using a
non-interfered fragment ion or, where there are isomers
or chiral compounds, when MS or MS2 separation is
difficult to achieve, improving the GC or LC separation
would resolve the matrix effect problem.
The matrix effect, in the form of analyte and matrix
component interactions, is unique to ESI- and APCIbased LC-MS or LC-MS2 instrumentation. It can appear
as signal suppression or enhancement (when compared
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Table 3. Terms, definitions, and numerical values of low level data qualifier and QC samples used in the method
Terms
Target analytes
Quality control
Definition
Ref.
Spikes
10 SD of lowest calibration
standard; SNR of 10
N/A
Serial dilution to SNR of 3
SNR of 10
SNR of 10
lowest calibration point
9 SD of blank noise
US EPA protocol
3 noise height
[10]
[23]
[54]
[33]
[21]
[32]
[31]
[28]
[26]
0.030.2
110
3.39.6
0.120
18
3 noise height
US EPA protocol
DIN 32645
US EPA protocol
SNR P 10
[27]
[57]
[58]
[59]
[60]
N/A
556
SNR of 3
[39]
Spikes
SNR of 10
[10]
Spikes
Spikes
SNR of 3
SNR of 3
[11]
[12]
Spikes
Spikes
Spikes
Blanks, spikes
Spikes
Spikes
Spikes
Spikes
Spikes
Spikes
Spikes
Spikes
0.041.00 ng/g
0.420 ng/g
0.21.6 ng/g
46
12
0.84.8 pg
0.5314.79
525
0.010.38
6198
3050
30190
[19]
[20]
[51]
[38]
[47]
[44]
[2]
[3]
[9]
[36]
[41]
[4]
Reagent blank,
blanks, spikes
Spikes
15
SNR of 3
2nd lowest calibration standard
SNR of 3
SNR of 3
SNR of 3
SNR of 3
SNR of 3
SNR of 10
SNR of 3
SNR of 3
SNR of 6, US EPA protocol
Concentration corresponding to
signal at the y-intercept plus
three times its SD
SNR of 3
2050 ng/g
[14]
Spikes
341 ng/g
Spikes
Blanks, spikes
Spikes
Spikes
Spikes
0.635
0.44.3
86,000
0.0010.27 ng/g
2.732.1 ng/g
[46]
[8]
[35]
[16]
[17]
515 ng/g
SNR of 3
[18]
10500
2070
[37]
[40]
3.847
775
1.013
SNR of 3
Concentration corresponding to
signal at the y-intercept plus
three times its SD
SNR of 3
SNR of 3
DIN 32645
Estrogens
Various PPCPs
Estrogens
Triclosan & related compounds
Phenazone drugs
Pharmaceuticals
Estrogens
Estrogens
LOD
MDL
LOD
MDL
LOQ
Estrogens
Acidic/neutral drugs
Acid drugs
Acidic drugs/caffeine
Acidic/neutral drugs
Spikes
Spikes
Spikes
Spikes
Spikes
Spikes, blanks
Spikes
Spikes, blanks,
reagent blanks
Spikes, blanks
Spikes
Spikes, blanks
Spikes
Spikes
LOD
MDL
LOD
Various PPCPs
Sulfonamides
Reagent blank,
blanks, spikes
N/A
N/A
MDL
LOD
LOD
Various PPCPs
NSAIDs
Various PPCPs
Spikes
Spikes
Spikes
LOQ
LOQ
LOD
LOQ
LOD
LOD
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[7]
[15]
[48]
[49]
[58]
Trends
[2]
[3]
[4]
[5]
[6]
[7]
[8]
[9]
[10]
[11]
7. Conclusions
The quality of data of MS methods developed to deal with
PPCPs in aqueous and solid environmental matrices has
been improving since the 1990s. Target analytes are
detectable down to the sub-ng/L level by GC-MS, GCMS2, LC-MS and LC-MS2.
PPCP studies to date have covered removal from
WWTPs, mobility in the environment, and several surveillance programs focused on a limited number of
compound classes.
The matrix effect has been a difficult problem to resolve. In the future, we hope to see more practical approaches to resolve the problem of the matrix effect to
alleviate the uncertainty in reported data.
We hope to see more large-scale surveillance projects,
such as that carried out by the US Geological Survey, to
gain further understanding on the occurrence of not
only parent products but also transformation products to
allow the full assessment of environmental impact of
these EOPs. This will require environmental chemists to
make the best use of available resources and collaborative initiatives.
The need to have an agreed protocol for the development and the use of low-level data qualifiers, as well as
the use of sufficient QC samples with each reported study
to maintain the confidence level of environmental data,
should be viewed as an obligation instead of a resource
issue. This resolve will remove any concerns that we
might have on the data and augment our ability to
understand the eco-toxicological effects of both parent
compounds and their metabolites in the environment.
[12]
[13]
[14]
[15]
[16]
[17]
[18]
[19]
[20]
[21]
[22]
[23]
[24]
[25]
[26]
[27]
[28]
[29]
[30]
[31]
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