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Anal. Chem.

1994,66, 1416-1423

Determination of Acidic and Basic/Neutral Pesticides in


Water with a New Microliter Flow Rate LWMS Particle
Beam Interface
Achlile Cappiello,' Giorgio Famlglini, and Fabrirlo Bruner
Istituto di Scienze Chimiche, Universith di Urbino, Piazza Rinascimento 6, 6 1029 Urbino, Italy

A new method for the determination of 32 baseheutral and mass spectrometer. Furthermore, HPLC samples are either
13 acidic pesticides in water, which has been proven to be thermally unstable or nonvolatile and for these characteristics
particularly amenable to high-performanceliquid chromatog- soft ionization techniques are usually employed.
raphy, is discussed. A liquid/solid extraction procedure for The thermospray interface,' introduced in the mid-l980s,
the isolation of pesticides from aqueous samples is employed. represented a significant breakthrough and gave decisive
The LC/MS analysis is carried out with a LC-18 reversed- impetus for a worldwide diffusion of LC/MS instrumentation.
phase packed capillary column coupled with a Hewlett-Packard Reversed-phase conventional columns, employing high water
5989A quadrupole mass spectrometer using a new microliter content mobile phases and volatile buffers, were easily coupled
flow rate particle beam interface. Mobile-phase flow rates, by use of the new interface. Bellar and Budde2 reported an
ranging between 1 and 5 ML/min, pass into the electron impact interesting evaluation of the thermospray performance in the
ion source of the mass spectrometer. The aerosol generated analysis of numerous organic compounds in environmental
by the reduced solvent imput allows improved signal response samples. Although having undoubted advantages over pre-
for high water content mobile phases with better chromato- vious attempts, thermospray started to show some limitations,
graphic performance during gradient analyses. Reliable and these included necessity of different temperature settings for
reproducible results for positive identificationand quantitation different experimental conditions, ion abundance unstability,
of target analytes are presented. The very low contamination and lack of structurally significant fragmentation.
of the instrument satisfies the continuous use demands of routine Electrospray, which has captured considerable attention
analysis. for managing delicate high molecular weight compound^,^*^
has been employed successfully for the determination of
The use of high-performance liquid chromatography environmental polar pollutant^.^ The absolute absence of
(HPLC) methods for the analysis of pesticides in various fragment ions requires the use of collision-induced decom-
matrices has become necessary since the introduction of a position (CID) in order to collect useful structural information
large series of highly effective and easily degradable polar for undoubted identification or confirmation. Recently, high-
compounds. However, the lack of specificity of the usual flow ion spray, which also employs CID for structural
HPLC detectors, and the large number of compounds to be confirmation, has been used for the determination of some
identified and quantitatively analyzed, push toward the use pesticides.6
of LC/MS techniques. These techniques can provide high The particle beam interface, developed by Browner and
confidence analyte identification in complex environmental c o - w o r k e r ~is, becoming
~~~ a viable alternative for the analysis
mixtures. GC/MS-based analytical methods for nonvolatile of small molecules with chemical properties which are not
organic compounds are less attractive because of their suitable for GC/MS. Thousands of biologically and envi-
complexity and quantitative uncertainty, as well as the ronmentally important compounds can be included in this
additionai time and cost when derivatization processes are category. Particle beam is fully compatible with a preexisting
involved. electron impact (EI)/CI ion source and may give, for target
compounds, the advantages of both worlds: HPLC compat-
In the last few years an increasing number of publications ibility with the variety of structurally significant fragment
have contributed to the development of very effective LC/ ions characteristic of conventional electron impact ionization
MS techniques for the analysis of various classes of organic mass spectrometry.
compounds. The main advantage over consolidated GC/MS Since the early reports about a monodisperse aerosol-based
alternatives is the possibility of injecting the extract directly interface (MAGIC), a big effort has been devoted to the
into the system without further analyte derivatization, allowing
for consistent time saving in routine analyses. (1) Vestal, M. L.; Fergusson, G. J. Anal. Chem. 1985, 57, 2373-2378.
(2) Bellar, T. A.; Budde, W. L. Anal. Chem. 1988, 60, 2076-2083.
Although LC/MS coupling techniques have reached a high (3) Whitehouse, C. M.; Dreyes, R. N.; Yamashita, M.; Fenn, J. B. Anal. Chem.
1985, 57, 6 7 5 4 7 9 .
level of development, each one uses a radically different (4) Fenn, J . B.; Mann, M.; Mong, C. K.; Wong, S . F.; Whitehouse, C. M. Mass
approach to introduce the chromatographic effluent into the Specrrom. R ~ G1990,. 9, 37-70.
(5) Lin. H . ; Voyksner, R. D. Anal. Chem. 1993, 65, 4 5 1 4 5 6 .
ion source and to promote sample ionization. The enormous (6) Hopfgartner, G.; Wachs, T.; Bean, K.; Henion, J. Anal. Chem. 1993, 65,
difference in chemical properties and molecular weight among 439446.
molecules suitable for HPLC requires different approaches (7) Willoughby, R. C.; Browner, R. F. A n d . Chem. 1984, 56, 2626-2631.
(8) Winkler, P. C.; Perkins. D. D.: William, W. K.; Browner, R. F. Anal. Chem.
to transfer the sample from the HPLC environment into the 1988, 60. 489,

1410 Analytical Chemistry. Vol. 66, No. 9, May 1, 1994 0003-2700/94/0366-14 18$04.50/0
0 1994 American Chemical Society
development of a more reliable interface. Most of the criticism Table 1. Selected Pertlcldes
was addressed to its lower level of sensitivity compared with pesticide ClaSSU CAS RN*
other LC/MS interfaces, lower response with high water
Baaic/NeutralPesticides
content mobile phases, and unreliable quantitative results. (1) oxamyl carbamate (I) 23135-22-0
Bellar et al.9J0 have investigated the transport mechanism (2) methomyl carbamate (I) 16752-77-5
involved in the particle beam interface and pointed out a (3) chloridazon pyridazinone (H) 1698-60-8
(4) dimethoate phoaphorodithioate (A) 60-51-5
relationship between enhanced ion abundance and the use of (5) aldicarb carbamate (I) 116-06-3
a given carrier. Calibration curves for quantitative analyses (6) metoxuron phenylurea (H) 19937-59-8
(7) bromacil uracil (H) 314-40-9
have benefited from a complete understanding of these (8)cyanazine triazine (H) 21725-46-2
processes, thus clearing up any perplexity over quantitation (9) monuron phenylurea (H) 150-68-5
results. (10)metribuzin triazine (H) 21087-64-9
(11) carbofuran carbamate (I) 1563-66-2
Many research groups have successfullyemployed particle (12) chlortoluron phenylurea (H) 15545-48-9
(13) carbaryl carbamate (I) 63-25-2
beam interfaces for the analysis of some pesticides and other (14) fluometuron phenylurea (H) 2164-17-2
compounds of environmental interest."-l7 The methods (15) atrazine triazine (H) 1912-24-9
presented can be considered a valuable improvement in the (16) isoproturon phenylurea (H) 34123-59-6
(17) paraoxon organophosphate (I) 311-45-5
analysis of nonvolatile pollutants. (18) difenoxuron phenylurea (H) 14214-32-5
Recently, our research group presented a radical modifi- (19) monolinuron phenylurea (H) 1746-81-2
(20) diuron phenylurea (H) 330-54-1
cation of the nebulization process of a particle beam interface (21) propachlor acetanilide (H) 1918-16-7
which allows the introduction of a much lower mobile-phase (22) r" carbamate (I) 122-42-9
(23) propanil propionalide(H) 709-98-8
flow rate, on the order of I pL/min,18 into the mass (24) linuron phenylurea (H) 330-55-2
spectrometer ion source. The new coupling device, which is (25) chloroxuron phenylurea (H) 1982-47-4
fully compatible with a commercially available desolvation (26) chlorbromuron phenylurea (H) 13360-45-7
(27) chlorpropham carbamate (H) 101-21- 3
chamber and momentum separator of a Hewlett-Packard (28) fenitrothion phosphorothioate(I) 122-75-5
59980B particle beam unit, shows several advantages compared (29) azinphos-ethyl phosphorothioate(I) 2642-71-9
(30) parathion-ethyl phosphorothioate(I) 56-38-2
to the conventional device. Benefits include drastic reduction (31) coumaphos phosphorothioate(I) 56-72-4
in solvent consumption with negligible contamination by (32) phoxim phosphorothioate(I) 14816-18-3
solvent vapors of the pumping system, the ion source, and the Acidic Pesticides
analyzer. These characteristics widen the choice of HPLC (1) dicamba methoxybenzoic(H) 1918-00-9
(2) bentazone thiadiazinone (H) 25057-89-0
buffers or mobile phases to those considered potentially (3) 2,4-D phenoxy acid (H) 94-75-7
harmful to the instrument. Better signal response for high (4) MCPA phenoxy acid (H) 94-74-6
(5) bromoxynil phenol (H) 1689-84-5
water content mobile phases with improved sensitivity and (6) dichlorprop phenoxy acid (H) 120-36-5
chromatographic performance during gradient analysis were (7) mecoprop phenoxy acid (H) 7085-19-0
also noticed. (8) 2,4,5-T phenoxy acid (H) 93-76-5
(9) 2,4-DB phenoxy acid (H) 94-82-6
The purpose of this work was to evaluate the possibility of (10) MCPB phenoxy acid (H) 94-81-5
the new device to contribute positively to the analysis of a (11) 2,4,5-TP phenoxy acid (H) 93-72-1
(12) dinoseb phenol (H) 88-85-7
large number of basic/neutral and acidic pesticides which are (13) dinoterb phenol (H) 1420-07-1
particularly amenable with HPLC. The method employs a
Key: A, acaricide;H, herbicide;I, insecticide. b ChemicalAbatract
(I
reversed-phase packed capillary c o l ~ m n ' ~for - ~chromato-
~ Service registry number.
graphic separations and SPE cartridges for the extraction
p r ~ c e d u r e . ~The
~ - ~detection
~ limits were generally in the
range of 0.1-30 ppb as required for environmental analysis.
all compound identification and accurate quantitative results
This method provides sufficient qualitative information for
in the range of concentrations considered. Moreover, it does
(9) Bellar, T. A.; Behymer, T. D.; Budde, W . L. J. Am. Soc. Mass Specrrom.
not require costly instrumentation and, although unique, is
1990.
-. 1. 92-98.
. -,-, -- - - particularly oriented for routine applications.
(10) Ho,J. S.;Behymer, T. D.; Budde, W. L.; Bcllar, T. A. J . Am. Soc. Mass
Specrrom. 1992, 3, 662671.
(11) Bchymer, T. D.; Bellar, T. A.; Budde, W. L. Anal. Chem. 1990, 62, 1686. EXPER I MENTAL SECTION
(12) Jones,T. L.; Betowski, L. D.;Lwnik, B.; Chiang,T. C.;Teberg, J. E. Enuiron.
Sci. Techno/. 1991, 25, 1880-1884. Particle Beam Interface and Mass Spectrometer. A
(13) Jones, G. G.; Pauls, R. E.; Willoughby, R. C. Anal. Chem. 1991,63,460463.
(14) Incorvia Mattina, M. J. J . Chromatogr. 1991, 542, 385-395. modified Hewlett-Packard 59980B particle beam interface,
(1 5) Julien-Larose, C.; Voiron, P.; Mas-Chamberlin, C.; Dufour, A. J . Chromnrogr. coupled with a Hewlett-Packard 5989A quadrupole mass
1991, 562. 39-45.
(16) Kim, I. S.;Sasinos, F. I.; Stephens, R. D.; Wang, J.; Brown, M. A. Anal. spectrometer, was used in this work. The original supplied
Chem. 1991,63, 819-823. nebulizer was replaced with a laboratory-made microneubu-
(17) Betowski, L. D.; Pace, C. M.; Roby, M. R. J . Am. Soc. MussSpectrom. 1992,
3, 823-830. lizer which has been described in a previous paper.'s This
(18) Cappiello, A.; Bruner, F. Anal. Chem. 1993, 65, 1281-1287. new device does not require modifications in the desolvation
(19) Cappiello, A.; Palma, P.; Mangani, F. Chromnrographia 1991,32,389-391.
(20) Crescenthi, G.; Mastrogiacomo, A. R. J . Microcolumn Sep. 1991, 3, 539- chamber, in the momentum separator, or in their original
545. assembly. Nozzle and skimmer holes and their mutual
(21) Crescenthi, G.; Bruner, F.; Mangani, F.; Yafeng, G. Anal. Chem. 1988.60,
1659. distance were left in their original settings. The final transfer
(22) Palma, P.; Cappiello, A. Ann. Chim. 1992, 82, 371-377. tube of the interface was set in fully retracted position, thus
(23) Di Corcia, A.; Marchetti, M.Anal. Chem. 1991, 63, 580-585.
(24) Di Corcia, A.; Marchetti, M. J. Chromatogr. 1991, 541, 365-373. leaving -5 mm of free space prior to the ion source. This
( 2 5 ) Di Corcia, A.; Samperi, R. A n a l . C h e m . 1993, 65, 907-912. gap allows the addition of a third pumping stage for sample

AnaiyticalChemlstry, Vol. 66, No. 9,May 1, 1994 1417


a

Abundance

20 30 40
Time (min.)

Flguro 1. UV profile (a) and TIC profile (b) of a chromatographic separation of a standard mixture of 32 basic/neutrai pesticides.

enrichment, operated by the roughing pump of the mass automatically using perfluorotributylamine (PFTBA) as the
spectrometer. A 50-pm-i.d., 180-pm-0.d. fused-silica capillary reference compound. The repeller potential was adjusted
tubing was used as the nebulizer tip and to connect the manually, monitoring fragment ions with m / z close to sample
chromatographic column at the opposite end. The mobile- values. Mobile phase flowed into the ion source while the
phase in-line filter was removed from the liquid path. The calibration procedure was operating. The mass spectrometer
nebulizing gas, helium (5.6 purity grade), was obtained from was scanning in the 50-400 amu range with a threshold of 50
SOL (Milano, Italy). The helium flow rate was -0.2 L/min
when maximum signal response was monitored by the ion
source with a mobile-phase flow rate of 2 pL/min. This value
-
counts when operated in TIC mode. The scan speed was 1.2
scan/s, which gave a mean of 10 acquisition samples for
each HPLC peak. Peak area values were calculated with
corresponds to a gas pressure of 30 psi and to a linear velocity automatic integration. The electron energy was set at 70 eV
at the nebulizer tip of 200 m/s. Because of a redesigned gas in positive ion mode.
delivery tubing inside the nebulizer, a different relation between Interface PerformanceOptimization. Signal optimization
gas pressure and gas flow was found. The gas temperature with the new interface was less critical and consistently easier
was ambient while the desolvation chamber temperature was than with the conventional one. A specific combination of
40 OC for all the experiments. The pressure was reduced to the position of the fused-silica capillary inside the coaxial gas
-0.5 atm in the desolvation chamber, 0.3 Torr in the second tubing and helium pressure offers the highest signal intensity
stage of the momentum separator, and (5-8) X le5Torr in for basic/neutral and acidic pesticides. No further adjust-
the manifold of the ion source. The temperature in the ion ments are usually required as the two species are exchanged.
source was set at 250 OC, while the analyzer was 120 "C.The The optimization procedure was carried out in accordance
mass spectrometer tuning and calibration was operated with the flow injection analysis technique (FIA) for prelimi-

1410 AnalyticalChemistry, Vol. $6,No. 9, May 1, 1994


nary adjustment, followed by HPLC/MS under required Table 2. SIM Acquldtkn Paramelere Used for Barlc/Neutral
chromatographic conditions, with proper buffers and solvents. end Acldk Pestkhles
A unique advantage of the new interface is its excellent time (min) program compound mlz dwell(ms)
performance even with a high concentration of water in the Basic/Neutral Pesticides
mobile phase. The signal response remains nearly constant 0 1 oxamyl 72,98 300
over a wide range of mobile phase relative concentrations. methomyl 88,105 300
chloridazon 77,105,221 300
Tuning was not affected by the water content in the mobile dimethoate 87,125 300
phase, thus simplifying the overall procedure. All the tuning 22 2 aldicarb 115,144 120
tests were conducted with a 50% water/acetonitrile solution metoxuron 72 120
bromacil 205 120
added to the proper amount of the final buffer. A vernier dial cyanazine 198,212 120
allows shifts of the nebulizer capillary tip, and the monuron 72,198 120
most appropriate position was with the tip end aligned with metribuzin 144,198 120
carbofuran 164 120
the sharp restriction of the gas tubing or slightly withdrawn. chlortoluron 72,132,212 120
Liquid Chromatography. The packed capillary column used carbaryl 115,144 120
fluometuron 72,187,232 120
in this work was made in our laboratory from '/Isin.-o.d., atrazine 200 120
250-pm4.d. poly(ether ether ketone) (PEEK) tubing (Alltech isoproturon 72 120
Associates Inc., Deerfield, IL) and packed with C18 reversed- paraoxon 109 120
difenoxuron 72,241 120
phase 5-pm particle size purchased from Phase Sep (Queens- monolinuron 61 120

-
ferry, UK). The 25-cm-long column had an efficiency of
15 000 theorical plates measured at 1 pL/min flow rate.
The liquid chromatography was carried out with a Kontron
diuron
propachlor
propham
72,187,232
93,120
93,172
120
120
120
propanil 161 120
Instrument 420 dual-pump binary-gradient conventional linuron 61,187 120
HPLC system (Kontron Instrument, Milano, Italy). Micro- chloroxuron 72 120
chlorbromuron 61 120
liter flow rates were obtained with a laboratory-made splitter chlorpropham 127 120
that was placed between the pumping system and the injector. fenitrothion 109 120
The splitter allows accurate and stable microliter flow rates azinphos-ethyl 132 120
40 3 parathion-ethyl 97,109,291 300
and rapid delivery of solvent concentration changes for reliable coumaphos 109,226,362 300
and reproducible gradients.22 A motor-assisted solvent mixer phoxim 77,103 300
was placed after the pumps and before the splitter device. For Acidic Pesticides
sample injection a zero-volume Valco injector equipped with 0 1 dicamba 173,220 150
bentazone 119,198 150
a 60-nL internal loop was employed (Valco, Houston, TX). 2,4-D 162,164,220 150
Larger loops are not advisable for flow rates lower than 5 MCPA 107,141,200 150
pL/min because of their consistent loss of chromatographic bromoxynil 88,277 150
dichlorprop 162,164,234 150
efficiency. mecoprop 107,142,214 150
The 50-pm-i.d., 180-pm-0.d. fused-silica capillary tubing 2,4,5-T 162,196,254 150
used in the nebulizer was purchased from Polymicro Tech- 2,4-DB 162,164,198 150
MCPB 107,142 150
nologies (Phoenix, AZ). 2,4,5-TP 162,196,198 150
Extraction Procedure. A liquid/solid extraction procedure 24 2 dinoseb 163,211 500
dinoterb 177,225 500
for the isolation of pesticides from aqueous samples was
employed. A cartridge was filled with 250 mg of graphitized
carbon black (Carbograph 1) and was capable of sampling up pesticides were collected in a second extraction percolating
2 L of water. By taking advantage of the presence of positively through the trap, drop by drop, 6 mL of methylene chloride/
charged active centers on the Carbograph surface, a stepwise methanol (60:40 by volume) alkalinized with 0.016 M KOH.
elution system allowed the complete separation of basic/ Prior to the concentration step, the extract containing the
neutral pesticides from acidic ones. acidic pesticides was acidified by adding 0.35 mL of 2% (v/v)
The extraction cartridge was made using a polypropylene trifluoracetic acid (TFA) in water. The extracts were then
tube, 6.5 X 1.4 cm i.d., packed with 250 mg of Carbograph concentrated toobtain a finalvolume of 1OOpL. For a detailed
1,120-400 mesh (Alltech, Deerfield, IL). Polyethylene frits, discussion of the extraction procedure see ref 23,
20-pm pore size, were located above and below the sorbent Reagents. All solvents were HPLC grade from Farmitalia
bed. Before the water sample was extracted, the cartridge Carlo Erba (Milano, Italy) and were filtered and degassed
was washed with 5 mL of methylene chloride/methanol (80: before use. Pesticides were purchased from Riedel-De Ha&n
20 by volume) followed by 2 mL of methanol and 15 mL of (Hannover, Germany). Ammonium acetate and trifluoracetic
10 g/L ascorbic acid in HC1-acidified water (pH 2). The acid were purchased from Sigma Scientific (St. Louis, MO).
water sample was forced to pass through the trap at a flow Ascorbic acid was supplied by Carlo Erba. Reagent water
rate of 150-160 mL/min using a vacuum apparatus placed was obtained from a Milli-Q water purification system
below the cartridge. Distilled water (7 mL) was added to the (Millipore Corp., Bedford, MA).
empty trap after all the sample had passed through.
Basic/neutral pesticides were eluted by passing 1 mL of RESULTS AND DISCUSSION
methanol, drop by drop, through the trap followed by 6 mL For this study we have selected 32 basic/neutral and 13
of methylene chloride/methanol (80:20 by volume). Acidic acidic pesticides according to the criteria of poor volatility

Analytical Chemistry, Vol. 66,No. 9, May I , 1994 1419


a

Abundance
9
4OooO - IO'
b

30000 -
61
7

2ooM)-

10000 -

Flgurr 2. uv pestlckles.

and thermal stability and wide use in agriculture (Table 1). mance and overall particle beam carrier process, with evident
Most of them are not suitable for GC/MS methods or result advantages in sensitivity and response linearity. A 0.1 M
in distorted GC peak shapes, poor sensitivity,or multiple peaks solution of ammonium acetate in water was used for the
or show other evidence of partial thermal decomposition. separation of base/neutral pesticides while 0.05% TFA in
Conventional HPLC analysis and detection techniques are water and 0.025% TFA in acetonitrile were used for the acidic
feasible for detecting a few analytes in well-known or ones. TFA is also utilized to lower the pH, as required for
noncomplex samples, e.g., production control, but fail in acidic pesticide separation using C18 reversed-phase chro-
complex environmental mixtures. matograph. A 25-cm-long, 250-pm4.d. packed capillary
A mixture of water and acetonitrile was chosen as mobile column was employed for this work. The column is laboratory
phase for the separation of both pesticide species. Although made using PEEK tubing. This polymer is rigid but not fragile,
methanol gives higher signal response when used at high has great resistance to chemical and physical agents, and comes
concentration with the particle beam interface, acetonitrile in a standard l/la-in.-o.d. size. These features can extend the
offers considerably less viscosity and allows more favorable column lifetime and facilitate its connections and the overall
column pressure. Moreover, as already discussed in a previous use of HPLC capillary columns. The packing method is also
paper,'* the performance of the new interface is only slightly rapid, effective, and allows the preparation of very efficient
affected by variations in the organic solvent/water mobile- columns. The use of capillary HPLC hardware does not
phase composition, thus giving homogeneous responses when require particular skills or specifically trained personnel.
gradient analyses are performed. As demonstrated by Bellar Simple liquid chromatography expertise is often enough to
and co-~orkers,~JO the addition of certain substances to the successfully employ microliter liquid flow rates. HPLC
mobile phase can enhance both the chromatographic perfor- gradients were optimized with a conventional UV detector

1420 Analyticel Chemlstty, Vol. 66, No. 9, May 1, 1994


a

6000.0-
5400.0 --
4800.0 --
4200.0 --
3600.0-
3000.0--
2400.0 --
1800.0 --
1200.0-r
600.01 /
0 . o w ; I
0 5 10 15 20 25 30 compoundi
b
"9
Flgure 3. Linear regression of concentration calibration data for
chlortokuon. Meanstandard deviationf8.6 % (averageof five injections
for each concentration).
peak

1.6E4- 20

1.4E4 --

1.2E4 --

1.OE4--
/ 1a
8000.0_-

/
° t L
1 2 3 4 5 6 7
6000.0--
compounds
Figure 5. Instrument detection limits of basic/neutrai (a) and acidic
(b) pesticides. The limits are estimated by SIM detection with a signal:
noise ratio of 5:l.
0.0
0 5 10 15 20 25 30 35 40 45 50 55 60
detection are shown in Table 2. Three different ion programs
ng are used for basic/neutral detection while two programs are
Flgure 4. Linear regression of concentration calibration data for used for acidic analysis. High dwell time is used according
bentazone. Mean standard deviationf9.6% (average of five injections to the number of ions and chromatographic peak sampling.
for each Concentration). Figure 2a,b shows the chromatographic separation of a
standard mixture of 13 acidic pesticides with UV detector at
fitted with a down-scaled microflow cell. Figure la,b shows 225 nm (a) and with the particle beam inteface in the SIM
the chromatographic profile of a standard mixture of 32 basic/ mode (b). The amount injected is 45 ng for each compound
neutral pesticides obtained at 230 nm with the UV detector in a 60-nL loop volume, with a mobile-phase gradient from
(a) and with the PB-MS in the TIC mode (b). The amount 0% to 40% acetonitrile in 5 min and then to 80% in 25 min.
injected is 20 ng for each compound in a 60-nL loop volume, TFA buffer was used in the mobile phase as described above.
with a mobile-phase gradient from 20% to 80% acetonitrile The evident peak tailing shown by most acidic pesticides during
over 45 min. Because of solvent variation delay, 5 min of mass spectrometric analysis is due mainly to extra chromato-
initial isocratic elution should be also considered in the solvent graphic processes and, in particular, to the thermal decom-
program. It can be clearly noticed that no appreciable position phenomena activated by the electron impact source
differences in the peak shape or width can be discerned when environment. An extented discussion by Betowsky et al. is
the two profiles are compared. No band broadening takes given in ref 17.
place for extracolumn dead volumes. A 50-cm-long, 50-pm- A calibration curve and an evaluation of result reproduc-
i.d. fused-silica tubing connects the capillary column end and ibility using the new interface were performed. A well-
acts like a nebulizer tip on the other side. The delay between dectected compound for each pesticide species was chosen for

-
the injection of the compound and its appearance as signal
peak in the mass spectrometer is 2 min at 2 pL/min mobile-
phase flow rate, without considering its chromatographic
the test. A 60-nL loop of a solution containing a given amount
of pesticide was injected five times for each concentration.
Each sample was introduced without the column directly into
retention. All HPLC connections are standard ' / l a in. with the mass spectrometer via the particle beam interface with a
PEEK nuts and ferrules. Methomil, propachlor, and propham mobile phase composed of equal concentrations of water and
gave insufficient response with TIC detection. Totally acetonitrile. Specific buffers were used for basiclneutral and
overlapped peaks, like those obtained from aldicarb and acidic pesticides. The linear regression of concentration
metoxuron or carbaryl and fluometuron, are detected sepa- calibration data is reported in Figure 3 for chlortoluron and
rately by selecting only specific ions. Selected ions for SIM in Figure 4 for bentazone. The mean standard deviations,

AnalytcalChemlstry, Vol. 66, No. 9, May 1, 1994 1421


,.'**

CL
,
30000 1 on

I
I
II 2,,,]o i I
-0 0
I
1" 20 30 40 10 20 30
TIme(min) T,,"~")

Figure 6. Baslc/neutrai pesticides extracted from 1 L of river water. Figure 7. Acidic pesticides extracted from 1 L of river water. Ion
Ion chromatogram of chlortoluron (1.3 pg/L). Asterisks Indicate the chromatogram of bromoxynii (4.4 pg/L). Asterisks indlcate the
correspondent mlr value for each Ion chromatogram. correspondent m/r value for each Ion chromatogram.

calculated using the average of the peak area values for each concentration in the mobile phase is only slightly affected by
concentration experiment, are f8.6% and f9.6%, respectively. solvent composition.
The mass spectrometer was operating in SIM a t m / z 212 for Detection limits for the analytes considered in this work
chlortoluron and m / z 198 for bentazone. The mobile-phase were calculated by injecting diluted concentrations of the
flow rate was kept at 2 pL/min for all experiments. Excellent standard mixture in operative chromatographic conditions.
linearity can be found in all calibration plots due to the The mass spectrometer was operating in SIM mode according
beneficial buffer effect on the interface carrier process. In to the ion program shown in Table 2. The lowest amount was
fact, we have noticed that at very high concentrations of organic determined for a signal-to-noise ratio of 5 1 . Figure 5a,b shows
solvent in the mobile phase, used for basic/neutral pesticide the estimated instrument detection limits for basic/neutral
separation (>80%), the lowest concentrations of chlortoluron (a) and acidic pesticides (b). Assuming a 100% extraction
were poorly detected. Ammonium acetate, which is water recovery for all pesticides from a 2-L water sample, with a
soluble, is decreased in concentration in the mobile phase when concentration factor of 2000 and an injection volume of 60
the aceonitrile component is increased. When over 80% nL, the instrument detection limits correspond to a method
acetonitrile is present in the mobile phase, ammonium acetate detection limit of 0.2-30 ppb. This range satisfies the detection
concentration is insufficient to positively influence carrier requirements for environmental pollutants in water and
mechanisms. The calibration curve is thus affected, and wastewater. Actual recoveries for most pesticides are reported
quantitation results are unsatisfactory. Thegradient employed in ref 23.
in this work for basic/neutral separation does not surpass A sample was collected from a river near Urbino, and the
80% acetonitrile until after 45 min of linear slope, plus 5 min pesticide contents were determined as shown in in Figures 6
of isocratic delay, when the last compound is eluted in 43 min. and 7. The river flows through a hilly rural area, with scattered
This ensures reliable quantitation results for all analytes grain, hay, and sunflower cultures. The sample was collected
considered. Trifluoroacetic acid shows characteristics similar after an early spring thunderstorm. The spectra reported in
to ammonium acetate in its influence on carrier processes. the figures were obtained from standard solutions and were
Acidic pesticides are eluted before 25 min when the acetonitrile injected separately. The peak abundances in the ion chro-
concentration is well below 8076,and it should be kept in mind matograms match the relative abundances of the same ions
that TFA is also dissolved in the organic solvent and that its in the mass spectra.

1422 Analytical Chemistty, Vol. 06, No. 9, May I , 1994


The data obtained in this work may contribute to a better withstand stringent instrumentation requirements when
promising future for liquid chromatography/mass spectrom- routine analyses are employed.
etry-based methods in environmental applications. The
ACKNOWLEDGMENT
particle beam interface with reduced flow rates offers several The authors are particularly indebted to Pierangela Palma
advantages in terms of sensitivity and performance with and Arnaldo Berloni for having supplied HPLC capillary
complex HPLC gradients for improved separations. Fur- columns.
thermore, the availability of precise retention times and high-
quality library-searchable mass spectra with linear calibration Received for review August 3, 1993. Accepted February 15,
plots allows unequivocal identification and quantitation of 1994.'
target and unexpected compounds. The new interface can 0 Abstract published in Aduonce ACS Abstracts, March 15, 1994.

Ana&ticalChemistry, Vol. 66, No. 9, May 1, 1994 1423