Anal. Chem. 2004, 76, 6187-6196
Laser Desorption and Matrix-Assisted Laser
Desorption/Ionization Mass Spectrometry of
29-kDa Au:SR Cluster Compounds
T. Gregory Schaaff*
Chemical Sciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831-6131
Positive and negative ions generated by laser-based
ionization methods from three gold:thiolate cluster compounds are mass analyzed by time-of-flight mass spectrometry. The three compounds have similar inorganic
core masses (∼29 kDa, ∼145 Au atoms) but different
n-alkanethiolate ligands associated with each cluster
compound (Au:SR, R ) butane, hexane, dodecane).
Irradiation of neat films (laser desorption/ionization) and
films generated by dilution of the cluster compounds in
an organic acid matrix (matrix-assisted laser desorption/
ionization) with a nitrogen laser (337 nm) produced
distinct ion abundances that are relevant to different
structural aspects of the cluster compound. Laser desorption/ionization of neat Au:SR compound films produces ions consistent with the inorganic core mass (i.e.,
devoid of original hydrocarbon content). Matrix-assisted
laser desorption/ionization produces either ions with m/z
values consistent with the core mass of the cluster
compounds or ions with m/z values consistent with the
approximate molecular weight of the cluster compounds,
depending on ionization conditions. The ion abundances,
and ionization conditions under which they are detected,
provide insight into desorption/ionization processes for
these unique cluster compounds as well as other analytes
typically studied by matrix-assisted laser desorption/
ionization.
Gold:thiolate (Au:SR) cluster compounds (or monolayer protected cluster compounds) constitute a special subset of metallic
nanostructures that have been the subject of numerous studies
in recent years.1 Conceptually, these compounds consist of a dense
metallic gold core surrounded by a shell of thiolate ligands (See
Chart 1.). The measured optical properties are dominated by the
electronic structure associated with metallic bonding within the
inorganic core, while the gross chemical properties are derived
with the organic (or biologic) ligands attached to that core. Similar
metallic and semiconductor cluster compounds are finding applications in biologic imaging, such as strongly scattering centers
* Phone: (865) 574-2297. Fax: (865) 574-9771. E-mail: [email protected].
Current address: BWXT Y-12, P.O. Box 2009, MS 8189, Oak Ridge, TN 378218189.
(1) Templeton, A. C.; Wuelfing, M. P.; Murray, R. W. Acc. Chem. Res. 2000,
33, 27-36.
10.1021/ac0353482 CCC: $27.50
Published on Web 10/06/2004
© 2004 American Chemical Society
for electron microscopy2,3 or bright fluorescent microscopy4-6
probes. Since the optical and electronic properties of cluster and
nanocrystal compounds are inexorably linked to the size of the
inorganic core, many future applications rely on the ability to
synthesis and isolate compounds with either a narrow core size
distribution or those that are molecularly pure (i.e., one single
structure). Giant (nanometer-scale) cluster compounds have been
isolated for selected systems recently (e.g., Pd145 metallic clusters7
and semiconductor Ag2S cluster compounds8). However, due to
their inherent compositional and structural complexity, these types
of compounds represent a significant challenge for routine
analytical chemical techniques, even those developed for other
macromolecular systems.
Because chemical properties are derived from the organic
ligands, one of the unique aspects of the Au:SR compounds
is the ability to isolate and accumulate cluster compounds
with distinct inorganic core sizes through fractionation,9-11
chromatography,11-13 supercritical extraction,14 and electrophoretic
methods.15 As a result of such separations, it was possible to map
the evolution of optical properties from bulklike (broad plasmon
resonance excitation)10 to molecular-like (discrete electronic
transitions)11,15 metallic electronic structure. In addition, other
interesting properties have been discovered for these cluster
(2) Hainfeld, J. F.; Furuya, F. R. J. Histochem. Cytochem. 1992, 40, 177-184.
(3) Powell, R. D.; Halsey, C. M. R.; Hainfeld, J. F. Microsc. Res. Technol. 1998,
42, 2-12.
(4) Dubertret, B.; Skourides, P.; Norris, D. J.; Noireaux, V.; Brivanlou, A. H.;
Libchaber, A. Science 2002, 298, 1759-1762.
(5) Bruchez, M.; Moronne, M.; Gin, P.; Weiss, S.; Alivisatos, A. P. Science 1998,
281, 2013-2016.
(6) Chan, W. C. W.; Nie, S. M. Science 1998, 281, 2016-2018.
(7) Tran, N. T.; Powell, D. R.; Dahl, L. F. Angew. Chem., Int. Ed. 2000, 39,
4121-4125.
(8) Wang, X. J.; Langetepe, T.; Persau, C.; Kang, B. S.; Sheldrick, G. M.; Fenske,
D. Angew. Chem., Int. Ed. 2002, 41, 3818-3822.
(9) Whetten, R. L.; Khoury, J. T.; Alvarez, M. M.; Murthy, S.; Vezmar, I.; Wang,
Z. L.; Stephens, P. W.; Cleveland, C. L.; Luedtke, W. D.; Landman, U. Adv.
Mater. 1996, 8, 428-433.
(10) Alvarez, M. M.; Khoury, J. T.; Schaaff, T. G.; Shafigullin, M. N.; Vezmar, I.;
Whetten, R. L. J. Phys. Chem. B 1997, 101, 3706-3712.
(11) Schaaff, T. G.; Shafigullin, M. N.; Khoury, J. T.; Vezmar, I.; Whetten, R. L.;
Cullen, W. G.; First, P. N.; GutierrezWing, C.; Ascensio, J.; JoseYacaman,
M. J. J. Phys. Chem. B 1997, 101, 7885-7891.
(12) Song, Y.; Jimenez, V.; McKinney, C.; Donkers, R.; Murray, R. W. Anal. Chem.
2003, 75, 5088-5096.
(13) Jimenez, V. L.; Leopold, M. C.; Mazzitelli, C.; Jorgenson, J. W.; Murray, R.
W. Anal. Chem. 2003, 75, 199-206.
(14) Clarke, N. Z.; Waters, C.; Johnson, K. A.; Satherley, J.; Schiffrin, D. J.
Langmuir 2001, 17, 6048-6050.
(15) Schaaff, T. G.; Whetten, R. L. J. Phys. Chem. B 2000, 104, 2630-2641.
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004 6187
Chart 1
compounds, which also show strong size dependencies or
quantum size effects (e.g., electrochemical charging of the cluster
core,16,17 chiroptical effects in gold clusters with biologically
derived ligands,15,18 and solid-state molecular crystal structure19).
Laser desorption/ionization mass spectrometry has been
highly efficient for analyzing the core mass of the cluster
compounds to provide both rapid monitoring of size separation
techniques and optimization of reaction parameters to produce
specific cluster compounds in high yield and the qualitative
observations regarding further reactions of the cluster compounds.20,21 Owing to the central role of laser desorption/ionization
(LDI)-MS, many reports have differentiated the separated cluster
compounds by their respective core masses. For example, 29kDa Au:SC4 refers to a gold cluster compound that, upon
irradiation with UV irradiation, produces a group of ions centered
at m/z 29 000 and has an associated ligand shell composed of
butanethiolate. While LDI-MS mass spectrometry has served a
central role in isolating Au:SR cluster compounds, implementation
of low-fragmentation ionization methods (e.g., electrospray ionization and matrix-assisted laser desorption/ionization, MALDI) has
only achieved sporadic success.
While gold:thiolate cluster compounds have received increased
attention in recent years due to the ease of preparation, the first
analysis of gold cluster compounds by mass spectrometry was
performed on gold:phosphane cluster compounds by McNeal and
co-workers using 252Cf-plasma desorption mass spectrometry.22
(16) Chen, S. W.; Ingram, R. S.; Hostetler, M. J.; Pietron, J. J.; Murray, R. W.;
Schaaff, T. G.; Khoury, J. T.; Alvarez, M. M.; Whetten, R. L. Science 1998,
280, 2098-2101.
(17) Chen, S. W.; Murray, R. W. J. Phys. Chem. B 1999, 103, 9996-10000.
(18) Schaaff, T. G.; Knight, G.; Shafigullin, M. N.; Borkman, R. F.; Whetten, R.
L. J. Phys. Chem. B 1998, 102, 10643-10646.
(19) Whetten, R. L.; Shafigullin, M. N.; Khoury, J. T.; Schaaff, T. G.; Vezmar, I.;
Alvarez, M. M.; Wilkinson, A. Acc. Chem. Res. 1999, 32, 397-406.
(20) Alvarez, M. M.; Khoury, J. T.; Schaaff, T. G.; Shafigullin, M.; Vezmar, I.;
Whetten, R. L. Chem. Phys. Lett. 1997, 266, 91-98.
(21) Schaaff, T. G.; Shafigullin, M. N.; Khoury, J. T.; Vezmar, I.; Whetten, R. L.
J. Phys. Chem. B 2001, 105, 8785-8796.
(22) McNeal, C. J.; Winpenny, R. E. P.; Hughes, J. M.; MacFarlane, R. D.;
Pignolet, L. H.; Nelson, L. T. J.; Gardner, T. G.; Irgens, L. H.; Vigh, G.; J. P.
Fackler, J. Inorg. Chem. 1993, 32, 5582-5590.
6188
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
From these studies, three to four molecular-like ions were
observed for a class of gold-phosphane cluster compounds, which
had previously been thought to contain a single component.
Recently, one such compound has been separated in high purity
by low-pressure chromatographic techniques and analyzed by
matrix-assisted laser desorption/ionization mass spectrometry.23
In addition, other metallic-phosphane cluster compounds have
recently been studied by electrospray ionization mass spectrometry.24 Despite many refinements in the methods for preparation
and isolation (or separation) of the gold-thiolato metal cluster
compound materials, they have never been conclusively demonstrated to exist as molecularly defined substances (e.g., singlecrystal structure determination). However, previous studies allude
to a molecular system composed of cluster species with nearly
identical composition and molecular weight. Clearly, an advanced
mass spectrometry investigation, utilizing ultrasensitive and ultrasoft (gentle) ionization methods, should enable a direct, rapid
determination of the distribution of assemblies present after
synthesis and isolation.
This report presents (among other things) progress toward
this goal and illuminates some of the mechanisms and challenges
inherent to the (MA)LDI processes, as applied to such materials.
Results from both LDI and MALDI mass spectrometry are
presented for three gold cluster compounds with the same Au
core mass: 29-kDa Au:SC4, Au:SC6, and Au:SC12. The abundance
of structurally relevant ions under both LDI and MALDI conditions
was dependent on both the irradiance delivered (on a single-shot
basis) and the total power delivered to the sample. Unlike many
other macromolecular structures, structurally relevant high m/z
ions were produced from Au:SR cluster compounds under many
ionization conditions (i.e., desorption from neat films or from
clusters diluted in organic matrixes). The ability to detect these
ions under many ionization conditions provides insight into likely
(23) Gutierrez, E.; Powell, R. D.; Furuya, F. R.; Hainfeld, J. F.; Schaaff, T. G.;
Shafigullin, M. N.; Stephens, P. W.; Whetten, R. L. Eur. Phys. J. D 1999, 9,
647-651.
(24) Kawano, M.; Bacon, J. W.; Campana, C. F.; Winger, B. E.; Dudek, J. D.;
Sirchio, S. A.; Scruggs, S. L.; Geiser, U.; Dahl, L. F. Inorg. Chem. 2001, 40,
2554-2569.
desorption/ionization processes for the gold cluster compounds.
In addition, this unique system may provide further detail
concerning proposed mechanisms for MALDI of typical biologic
analytes.
EXPERIMENTAL SECTION
The 29-kDa Au:SR cluster compounds used for these studies
were synthesized and separated using procedures described in
detail elsewhere.21 Briefly, the cluster compounds were prepared
by a procedure based on methods described by Brust et al.,25
which are optimized to yield (in high abundance) the compound
that produces ions under LDI conditions centered at m/z 29 000
for gold:butanethiolate, gold:hexanethiolate, and gold:dodecanethiolate clusters (Au:SC4, Au:SC6, and Au:SC12, respectively). The
compounds were separated by fractional crystallization, which is
effected by slow addition of acetone to a concentrated toluene
solution. Neat films of the mixture and separated cluster compounds (for LDI-MS) were prepared by pipeting 1 µL of a
concentrated solution (5 mg/mL in toluene) onto a sample plate
and allowing the solution dry under ambient conditions. Separation
of the compounds was monitored by LDI-MS, as shown below.
As described by many reports dealing with matrix-assisted
laser desorption/ionization, the ability to cocrystallize the analyte
in a matrix crystal is highly advantageous, requiring a common
solvent. For this reason, methylene chloride (HPLC grade, Baker
Scientific) was used as a common solvent for the cluster
compounds and the organic matrixes. Four matrixes (SigmaAldrich) were tested: 3,5-dihydroxybenzoic acid (DHB), 3,5dimethoxy-4-hydroxycinnamic acid (sinapinic acid), 1,8,9-anthracenetriol (dithranol), and trans-3-indoleacrylic acid. Films of
the matrix-diluted cluster compounds were generated by diluting
20 µL of a 10 µg/mL methylene chloride solution of Au:SR cluster
compounds 1:1 (v/v) with a saturated matrix solution in methylene
chloride. Of the four matrix molecules, sinapinic and trans-3indoleacrylic acid produced comparable mass spectra, while DHB
and dithranol did not seem to reduce fragmentation of the Au:SR
cluster compounds as well (i.e., most spectra were similar mass
spectra obtained by irradiating neat films). MALDI mass spectra
shown in this report are derived from ions produced by irradiation
of cluster compounds diluted in a sinapinic acid matrix.
Laser desorption and matrix-assisted laser desorption/ionization mass spectra were obtained using Perseptive Biosystems
Voyager DE linear time-of-flight mass spectrometer operating in
delayed extraction mode with an accelerating voltage of 20 kV.
The postdesorption delay time and space-focusing conditions were
optimized using laser-desorbed cations and anions from the neat
film by resolving the m/z 32 spacing in the ions detected at m/z
values centered at 29 000. While these ions could not be baseline
resolved, the periodic spacing of m/z 32 could still be used to
ensure optimum resolution conditions and the calibration was
correct under different ionization conditions (e.g., LDI vs MALDI)
for this region of the mass spectrum.
The Voyager DE mass spectrometer is equipped with a
VSL-337 ND pulsed N2 laser (Laser Science, Newton, MA) with a
4-ns pulse width at 337 nm. To determine the magnitude of the
laser irradiance delivered to the sample under different computer(25) Brust, M.; Walker, M.; Bethell, D.; Schiffrin, D. J.; Whyman, R. J. Chem.
Soc., Chem. Commun. 1994, 801-802.
Figure 1. Low m/z (a) and high m/z (b) regions of negative ion
laser desorption ionization mass spectrum obtained by irradiating a
neat film of a mixture of gold:thiolate cluster compounds with N2 laser
at 2.4 MW/cm2.
controlled settings, the beam was diverted outside the vacuum
chamber with a 90% transmission prism into a J3-09 pyroelectric/
silicon Joulemeter (Molectron). The laser power settings in the
instrument control software were changed, and the output from
the Joulemeter (fluence) was monitored with a digital storage
oscilloscope. The irradiance values listed below were calculated
for a focus of 200 µm (diameter) on the sample plate and corrected
for additional transmission losses due to the focusing lens and
quartz window installed on the vacuum chamber. Reported values
were obtained by averaging the measured fluence from 20
individual laser shots.
RESULTS
The Au:SR cluster compounds (where R ) butane, C4; hexane,
C6; and dodecane, C12), produced different types of high m/z
ions upon irradiation with a pulsed UV laser (N2, 337 nm) under
various ionization conditions. To differentiate these two conditions
in the results to follow, LDI refers to irradiation of neat (thick)
films generated by depositing 1-2 µL of a concentrated (10-20
mg/mL) solution of Au:SR cluster compounds and MALDI refers
to irradiation of cluster compounds dispersed within a sinapinic
acid matrix.
LDI mass spectra of gold:thiolate were obtained by irradiating
neat films of the Au:SR cluster compounds, typically at an
irradiance of 2-3 MW/cm2. Figure 1 shows different regions of
the negative ion LDI mass spectrum generated from a mixture of
Au:SR cluster compounds (R ) C12). Two distinct types of anions
are produced when neat Au:SR films are irradiated. Ions in the
low m/z region (Figure 1a) are extremely intense compared to
those at higher m/z values (on the order of 10-100 times signal
intensity) and can produce large background signals (due to
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
6189
detector relaxation) that extends to m/z ∼5000. In addition to the
m/z 197 [Au]- anion, groups of anions are detected with nominal
differences in m/z of 229. The lowest m/z ions in each group
corresponds to the “bare” [AuNSM]- anion (see labels in Figure
1a). Higher m/z ions in each group are detected at m/z values
that are separated by either m/z 12 or 13, which presumably
corresponds to addition of elemental carbon or the combination
of carbon and hydrogen. The relative intensity of these ions
increases with increasing laser irradiance and are invariant with
respect to ligand composition. Therefore, the remainder of this
report concentrates on the higher m/z ions found at lower relative
abundance that have a direct relation to the structure of the cluster
compound as it exists in the condensed phase.
Similar to previous reports,9,11,20 the higher m/z region of LDI
mass spectra shown in Figure 1b is composed of peaks corresponding to negative ions in distinct regions centered about m/z
8000, 15 000, 22 000, and 29 000. The spectrum represents the
average of 512 individual mass spectra obtained at an irradiance
of 4.1 MW/cm2. To lower the contribution of noise arising from
the low m/z ions, a low-mass cutoff was used to blank the mass
spectrum below m/z 4000. The mass spectrometer resolution (e.g.,
space focus and delayed extraction) was optimized for the mean
m/z shown in the Figure (m/z ∼15 000) to obtain the best
resolution for the entire window. The effect of delayed extraction
on mass resolution and measured ion abundances is well documented26 and has been found to produce LDI mass spectra from
mixtures of Au:SR’s that do not always reflect the true abundances
of components in the mixture being analyzed.27 Thus, when
monitoring separations or optimizing reaction conditions, LDI
mass spectra are acquired with static acceleration and fixed space
focusing to obtain a qualitative, but more accurate representation
of component abundance.
When mixtures of Au:SR cluster compounds are irradiated,
the negative ions (and positive ions, not shown) detected have
periodic spacing from m/z ∼6000 until it is impossible to resolve
the spacing due to instrumental resolution limitations. The ions
detected can be described as having major and minor m/z spacing
consistent with the composition of the inorganic cluster core. The
minor spacing in each group of ions corresponds to an m/z
difference of 32 (S). The major m/z spacing between the most
abundant ion in each adjacent group corresponds to a difference
of either 197 (Au) or 229 (AuS). The N, M labels in the inset
correspond to ions having the general formula [AuNSM]. Ions of
the same general AuNSM- composition were observed by Arnold
and Reilly for the separated compound that produces negative
ions centered at m/z 15 000.28 Abundant ions follow this general
progression from approximately m/z 6000 until the m/z 32 spacing
cannot be resolved. While Au is monoisotopic, S has four natural
isotopes: 32S (95.02), 33S (0.75), 34S (4.21), and 36S (0.02). The
resolution of the mass spectrometer is not sufficient to resolve
the isotopic abundances; thus assignments and calibration were
made using the exact mass of 197Au (196.966 55) and the average
mass of S (32.065).
(26) Reilly, J. P.; Colby, S. M. Anal. Chem. 1996, 68, 1419-1428.
(27) Schaaff, T. G.Preparation and Characterization of Thioaurite Cluster
Compounds. Ph.D. Dissertation, School of Chemistry and Biochemistry,
Georgia Institute of Technology, 1998.
(28) Arnold, R. J.; Reilly, J. P. J. Am. Chem. Soc. 1998, 120, 1528-1532.
6190 Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
Figure 2. Positive (a) and negative (b) ion laser desorption/
ionization mass spectra (at 2.4 MW/cm2) from a Au:SC4 cluster
compound after separation from smaller and larger size cluster
compounds, as seen in Figure 1. The insets in (a) and (b) show the
narrower m/z range centered at m/z 29 000. The peaks detected at
higher m/z values correspond to dimer, trimer, etc., ions of the base
ion at m/z 29 000.
After fractionation and isolation, the 29-kDa (core mass) Au:
SC4 cluster compound was separated from its other compounds
having disparate core masses. Figure 2 shows the negative and
positive ion LDI mass spectra for this compound obtained with
an irradiance of 4.1 MW/cm2. The spectra shown are the average
of mass spectra obtained from 32 laser shots. The acquisition of
32 shots has been observed to aid in the ability to resolve the
m/z 32 spacing. When a larger set of spectra are averaged, the
m/z 197 and 229 spacing is resolved well (as seen in Figure 1b),
but the m/z 32 spacing cannot be resolved in the m/z 30 000
region of the mass spectrum. This is likely due to slight changes
in laser power, electronic jitter, or both, which can become more
pronounced when averaging larger sets of spectra. Comparison
of the positive and negative ion mass spectra indicates that similar
groups of ions are produced at what seems to be approximately
the same intensity in both positive and negative ion modes. As
can be seen in the insets of Figure 2, the same general m/z 197
and 229 spacing is prevalent. The distribution of ion abundances
starts as an abrupt rise at m/z ∼26 500, and ions at lower m/z
values with the characteristic m/z 32 and 197 spacing are not
detected below that cutoff. The distribution of ions reaches a
maximum at m/z ∼29 000 and decreases until ions with the
characteristic m/z 32 and 197 spacing are not detected (m/z
32 000). Peaks detected at higher m/z values correspond to
multimers of the m/z 29 000 ions (i.e., 2 × 29 000, 3 × 29 000,
etc.). These multimer ions have been observed up to 10 × 29 000
under negative ion operation and up to 8 × 29 000 under positive
ion operation.
Threshold irradiance for detection of positive ions from the
Au:SC4 cluster compound was measured at 1.5 MW/cm2, which
Figure 3. Positive ion laser desorption/ionization mass spectra
obtained from the separated cluster compound shown in Figure 2 at
increasing irradiance (from top to bottom). The numbers above each
mass spectrum correspond to irradiance in MW/cm2.
produces a broad, featureless peak (similar to the top spectrum
in Figure 3) at a signal-to-noise level of 2. Figure 3 illustrates the
changes in positive ion abundance as a function of irradiance. At
slightly above threshold for production of positive ions from neat
films, no m/z 32 or 197 spacing is observed in the ion abundances
(1.7 MW/cm2, top spectrum). In addition, at this lower laser
power, the peak apex is centered at m/z ∼30 000. As irradiance
is increased, not only do the ion abundances start to show the
characteristic m/z 32 and 197 spacing, but the apex of the ion
abundances shifts to lower m/z values (∼29 000). At high
irradiance (>5 MW/cm2), the m/z 32 spacing cannot be resolved
and the peak apex has shifted still lower, to m/z ∼28 500. A similar
dependence of ion abundances on irradiance was observed
negative ions generated from neat films of the 29-kDa Au:SC4
cluster compounds. Similar to high-fragmentation conditions for
MALDI, shown below, it should be noted that the abundance of
ions detected at m/z ∼29 000 were relatively invariant with respect
to chainlength of the thiolate ligand. For example, similar ion
abundances were detected for both LDI-generated positive and
negative ions from Au:SC4, Au:SC6, and Au:SC12 cluster compounds with core masses of 29 kDa.
MALDI has been used to mitigate fragmentation of the Au:
SR cluster compounds during ionization repeatedly with a number
of typical organic matrix molecules. To date, the best matrix has
been determined to be sinapinic acid. However, the use of trans3-indoleacrylic acid (typically used for MALDI of organic polymers) was found to have performance comparable to that of
sinapinic acid. Unlike many other MALDI analytes, it appears that
two desorption/ionization regimes exist for MALDI of Au:SR
cluster compounds (high and low fragmentation), which are
dependent on the incident irradiance and total irradiance delivered
to the matrix/cluster mixture. Figure 4 shows the dependence of
Figure 4. Positive ion matrix-assisted laser desorption/ionization
mass spectra obtained from the separated Au:SC4 cluster compound
shown in Figure 2 at increasing irradiance (from top to bottom). The
numbers above each mass spectrum correspond to irradiance in MW/
cm2. The * above spectra at 2.1-3.0 MW/cm2 corresponds to m/z
28 400.
mass spectra on the irradiance when the 29-kDa (core mass)
Au:SC4 cluster compound is diluted in a matrix of sinapinic acid.
Because it is well known that MALDI matrixes can produce socalled “hot spots” (where the signal is much stronger in specific
areas of the sample), the mass spectra shown in Figure 4 were
obtained by continuously repositioning the laser spot over the
entire sample surface while acquiring data.
At lower irradiance (below 3 MW/cm2), the abundances of
ions centered at m/z 29 000 wwere similar to those measured from
LDI-MS of neat films (top 3 spectra, Figure 4), though the ions
detected correspond to slightly lower m/z values. At nearthreshold irradiance, the ions detected correspond to a broad,
featureless peak with an apex at m/z 29 200, similar to the nearthreshold LDI from neat films but at lower m/z values. At slightly
higher irradiance (2-3 MW/cm2), the m/z spacing is resolved,
also consistent with resolution of these characteristic ions under
LDI conditions at higher irradiance. While the m/z 32 spacing is
not resolved to the degree found in LDI from neat films, the m/z
197 spacing is partially resolved for approximately three or four
groups of ions. The first group of ions detected (i.e., the first peak
in the progresses of m/z 197 spacing, denoted by * in Figure 4)
corresponds to m/z 28 400.
At 3 MW/cm2, the ions corresponding to m/z values between
28 000 and 30 000 remain relatively unchanged, while a distribution of ions corresponding to the broad featureless peak at m/z
32 000 increases in relative abundance. Above 3.0 MW/cm2, the
low m/z peaks again appear featureless and the ions corresponding to m/z 32 000 are detected at much higher relative abundance.
The absolute intensity of the ions detected does not change
appreciably, but the relative signal-to-noise ratio increases slightly.
Reproducible features are partially resolved, superimposed on the
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
6191
Figure 5. Positive ion matrix-assisted laser desorptionionization
mass spectra obtained at 2.1 MW/cm2 from the Au:SC4, Au:SC6,
and Au:SC12 cluster compounds (from top to bottom, respectively).
Again, the * above the spectra corresponds to m/z 28 400, common
among all three compounds.
broad peak centered at m/z 32 000, but resolution at this m/z
range is not sufficient to make qualified assignments of ion
composition, nor do the m/z values contain recognizable spacing
(e.g., consistent with loss of whole ligand molecules from the ion).
Ion abundances detected for the 29-kDa Au:SC6 and Au:SC12
cluster compounds showed similar irradiance-induced effects.
Figure 5 shows the average of 32 mass spectra obtained at low
irradiance (2.4 MW/cm2) from the three 29-kDa cluster compounds (top to bottom): Au:SC4, Au:SC6, and Au:SC12. With only
slight differences, the three compounds produce remarkably
similar groups of ions with m/z 197 spacing, but the m/z 32
spacing was not resolved for these compounds. While mass
resolution limitations preclude unequivocal assignment of the ions
detected, it is clear that the position of the first abundant group
of ions (with the characteristic m/z 197 spacing) is measured at
approximately the same m/z (28 400) for all three cluster
compounds (denoted by * in Figures 4 and 5). The distribution
of ions in this region is narrower than those detected with LDI.
As seen in Figure 5, the rise in abundance under this ionization
condition occurs at m/z 27 500, compared to m/z 27 000 for LDI
(inset, Figure 1a), and the ion abundance returns to near baseline
at m/z ∼31 000 (compared to m/z ∼32 000 for LDI).
Upon repeated experiments to evaluate changes in ion abundances as a function of irradiance for the three compounds, it
was apparent that the relative ion abundances were also affected
by the total power delivered to the sample. Mass spectra obtained
from repeated pulses delivered to dense sample areas (i.e., areas
with large crystalline matrix structures) changed with subsequent
laser shots, while those obtained from sparse or thin areas on
the sample were consistently similar to those seen in Figure 5.
Focusing the laser on an area in the matrix/cluster sample region,
6192 Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
Figure 6. Evolution of positive ion matrix-assisted laser desorption/
ionization mass spectra obtained from the Au:SC4 cluster compound
at irradiance of 4.0 MW/cm2. The numbers above each mass
spectrum correspond to the number of laser shots delivered to the
matrix/analyte sample.
which appeared optically dense, and triggering the laser (off and
on) manually generated mass spectra such as those shown in
Figure 6. The irradiance used for the spectra in Figure 6 was 4.0
MW/cm2. The mass spectrum obtained in the first few laser shots
is similar to both low-irradiance MALDI (Figures 4 and 5) and
LDI from neat films. During these first few shots, the distribution
of ions has an apex at m/z ∼28 000, but the characteristic m/z
197 spacing is only partially resolved across the distribution of
ion abundances detected. After the first 10-20 laser pulses, the
ion abundances centered at m/z 29 000 decreases and the higher
m/z ions are detected at an increased relative abundance centered
at m/z 32 000. At ∼30 laser shots, ions centered at m/z 29 000
and 32 000 are approximately the same abundance. Finally, at ∼75
laser shots, the ions at m/z 29 000 are completely suppressed and
the ions detected are centered at m/z 32 000. In addition, initial
mass spectra (first 10-20 shots) had ion abundances centered at
m/z 58 000, 87 000, etc., corresponding to the multimers of the
ions centered at m/z 29 000 as seen in LDI mass spectra obtained
from irradiating neat films. The relative abundance of multimers
remained constant with respect to the abundance of the ions at
m/z 29 000. However, no ion abundances were detected at the
m/z values corresponding to multimers of the ions centered at
m/z 32 000 (see discussion below).
After determining the changes in the abundance of desorbed
ions under different laser conditions for the 29-kDa Au:SC4
compound, the other two 29-kDa cluster compounds were
investigated using similar ionization conditions. Figure 7 shows
the low-fragmentation MALDI mass spectra from three different
29-kDa cluster compounds (from top to bottom): Au:SC4, Au:
SC6, and Au:SC12. The position of the peak apex and tailing edge
changes consistent with the longer chain length ligands of the
different cluster compounds. It is also interesting to note that the
Figure 7. Positive ion matrix-assisted laser desorption/ionization
mass spectra obtained by irradiating the matrix/sample area at an
irradiance of 4.0 MW/cm2 for 60-70 laser pulses and then acquiring
mass spectra for 32 additional laser pulses. The mass spectra (from
top to bottom) correspond to ions generated from the Au:SC4, Au:
SC6, and Au:SC12 cluster compounds, which produced m/z ∼28 400
ions shown in Figure 5. The arrows in each mass spectrum correspond to the expected m/z for a molecular-type ion from an intact
cluster compound with an Au:SR ratio of 2.57:1, as determined by
elemental analysis.
full width half-maximum (fwhm) of the distribution of ions does
not change appreciably with different chain lengths. A previously
isolated 29-kDa Au:SC12 cluster compound was ionized by MALDI
with 3,5-dihydroxybenzoic acid as a matrix.21 However, in that
case, the ion distribution extended from m/z 28 000 to 40 000.
In summary, the three 29-kDa Au:SR compounds (R ) C4,
C6, and C12) produced nearly identical mass spectra both at low
irradiance and during the initial irradiation of the matrix/cluster
sample. The ions produced under this particular ionization
condition were centered at m/z 29 000, with the first detected
(resolved) ion being m/z 28 400 for all three compounds. For the
Au:SC4, Au:SC6, and Au:SC12, higher irradiance or additional laser
pulses produced higher m/z ions that were centered at m/z 32 000,
33 200, and 37 400, respectively. In previous reports, the Au:S ratio
for the 29-kDa (core mass) Au:SR compounds was determined
by elemental analysis to be 2.57:1.21 The arrows in Figure 7 denote
the m/z value that would correspond to an intact ion assuming
the peak at m/z 28 400 corresponds to the number of gold atoms
in the cluster core.
DISCUSSION
While matrix-assisted laser desorption/ionization has proven
extremely useful for the analysis of thermally labile macromolecules, the implementation of this “soft” ionization method to
inorganic cluster compounds still faces significant challenges.
Considering the structure of the cluster compounds (i.e., inorganic
core surrounded by a hydrophobic monolayer), there are similarities to typical MALDI analytes (similar in size to proteins, chemical
properties similar to hydrophobic polymers). As would be expected for a complex molecular structure, many other subtle
properties must be considered for application of MALDI to metallic
cluster compounds. For example, the near-IR, visible, and ultraviolet optical properties are quite different from that of typical
MALDI analytes. The extinction coefficient (at 337 nm) for the
29-kDa cluster compounds is on the order of that for the sinapinic
matrix molecules in which they are dispersed. In addition, the
hydrophobicity of the cluster compounds may preclude their
dispersal into various matrixes. Overcoming these differences and
understanding specific ionization processes in these complex
compounds are required before soft ionization techniques can be
applied to this class of cluster compounds, or more importantly,
used to develop similar techniques for other types of cluster
compounds (e.g., semiconductor and other metallic cluster
compounds).
The lack of mass spectrometry-based studies of cluster
compounds is likely due to a number of factors, with one of the
most important being sample purity of available cluster compounds. To draw on the biologic MALDI analogy, understanding
and applying MALDI to metallic cluster compounds before
isolation by cluster size would be similar to developing MALDI
for proteins by starting with an extract of all proteins from a whole
cell without any prior chemical separations. As shown in Figure
S1 (Supporting Information), the changing abundances of ions
as a function of laser irradiance and number of laser shots produce
mass spectra with peaks superimposed from different size nanocrystal cores or, more detrimental, at m/z values that normally
correspond to ions generated from LDI of neat films. Thus, without
separations, it is impossible to determine which ions are produced
under which conditions. Of the gold:thiolate cluster compounds,
the compound that produces ions centered at m/z 29 000 by LDI
is the best understood compound (structurally and electronically),21 which is why this compound (with different ligand
molecules) was chosen for these studies.
The measurement of ion abundances corresponding to the
approximate core mass of the cluster compound is consistent with
other analytical techniques more commonly used to analyze
cluster sizes, e.g., X-ray diffraction,19 electron microscopy,11 and
scanning probe microscopy.29,30 Under LDI conditions, the fwhm
of the distribution of ions centered at m/z 29 000 is ∼2500. This
in turn translates into a core diameter dispersion (typically
determined by high-resolution electron microscopic analysis) of
1.67 ( 0.02 nm using the formula
Deq )
(
NAu
(π/6)d
)
(1/3)
(1)
which is based on the density (d ) 59 atoms/nm3) of gold in its
native fcc structure. While the relative error associated with the
mass spectrally derived diameter (0.02 nm) clearly cannot translate
into a physical parameter, it does illustrate the precision associated
with the measurement, which cannot be achieved by either
electron microscopy or powder X-ray diffraction measurements.
(29) Harrell, L. E.; Bigioni, T. P.; Cullen, W. G.; Whetten, R. L.; First, P. N. J.
Vac. Sci. Technol. B 1999, 17, 2411-2416.
(30) Bigioni, T. P.; Harrell, L. E.; Cullen, W. G.; Guthrie, D. E.; Whetten, R. L.;
First, P. N. Eur. Phys. J., D 1999, 6, 355-364.
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
6193
From the seminal work by Tanaka and co-workers31 and others
to follow,32,33 it is well documented that metallic colloids can effect
liberation of large molecular biologic ions from a glycerol matrix
to produce “fragmentation-free” mass spectra similar to those
obtained routinely with organic matrixes. Bulk gold and silver also
share similar photochemically driven reactions that occur under
intense UV irradiationsselective cleavage of the S-C bond.34,35
The propensity to selectively cleave the S-C bond under UV
irradiation provides a mechanism similar to those proposed for
the irradiation of organic matrix crystals (i.e., violent disruption
of a specific crystal structure into an expanding plume) to liberate
molecular ions from biopolymers. Presumably, this violent disruption of the Au:SR cluster compounds can allow liberation of “intact”
cluster cores. Thus, the detection of high m/z ions such as those
shown in Figure 1 may be the consequence of the Au:SR clusters
acting both as a poor matrix and an analyte.
The mechanism for desorption and ionization of these large
structurally relevant ions from neat films of Au:SR cluster
compounds is likely similar to those proposed for MALDI.
Presumably, the high m/z ions are generated in a dense,
expanding plume populated by both high m/z ions and low m/z
ions. Both fragmentation of these high m/z ions and aggregation
with themselves (e.g., multimers in Figure 1b) and with low m/z
ions (as shown in Figure 1a) likely occurs within this dense plume
to add to the distribution of ions detected in the final LDI mass
spectrum. Reactions within a dense plume (under LDI conditions)
would be consistent with the narrower distribution of ions detected
under certain MALDI conditions (plume is likely not as dense
with MALDI prepared samples).
The gold:thiolate cluster compounds are unique (compared
to other thermally labile macromolecules) in that they produce
structurally relevant high m/z ions under all ionization conditions.
Many have reported on a phenomenon in MALDI in which it takes
a few laser pulses to observe ions being produced from the
matrix/analyte sample.36-38 This has been previously attributed
to a so-called “cleaning off” effect, where protein molecules not
incorporated in the matrix crystal or amorphous material is ablated
from the face of well-cocrystallized matrix/analyte samples and
protein ions are not formed. While this effect is difficult to observe
directly in the analysis of proteins (because they form either
metastable ions or no ions at all during this process), this effect
is observed more clearly in MALDI of the cluster compounds.
Considering the changes in mass spectra under different
ionization conditions, it is likely that gold cluster compounds are
present both as a surface (or amorphous) layer and cocrystallized
within the matrix crystals. Even though the cluster compounds
(31) Tanaka, K.; Waki, H.; Ido, Y.; Akita, S.; Yoshida, Y.; Yoshida, T. Rapid
Commun. Mass Spectrom. 1988, 2, 151-156.
(32) Lai, E. P. C.; Owega, S.; Kulczycki, R. J. Mass Spectrom. 1998, 33, 554564.
(33) Schurenberg, M.; Dreisewerd, K.; Hillenkamp, F. Anal. Chem. 1999, 71,
221-229.
(34) Lewis, M.; Tarlov, M.; Carron, K. J. Am. Chem. Soc. 1995, 117, 95749575.
(35) Rieley, H.; Price, N. J.; Smith, T. L.; Yang, S. H. J. Chem. Soc., Faraday
Trans. 1996, 92, 3629-3634.
(36) Bolbach, G.; Riahi, K.; Spiro, M.; Brunot, A.; Breton, F.; Blais, J. C. Analusis
1993, 21, 383-387.
(37) Perera, I. K.; Kantartzoglou, S.; Dyer, P. E. Int. J. Mass Spectrom. Ion Processes
1996, 156, 151-172.
(38) Pittenauer, E.; Schmid, E. R.; Allmaier, G.; Puchinger, L.; Kienzl, E. Eur.
Mass Spectrom. 1996, 2, 247-262.
6194 Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
Figure 8. Illustration of ion formation of Au:SR cluster compounds
dispersed in an organic matrix under high- and low-fragmentation
conditions (left and right images, respectively). The left image
corresponds to the processes that occur at low irradiance or initial
irradiation at high-irradiance levels to produce spectra similar to those
generated from neat films due to either a neat film on the matrix/
analyte crystal or an amorphous layer, which allows for extensive
fragmentation. The right illustrates a situation in which the amorphous
(or neat) layer has been ablated and the cluster compounds are
ejected (or liberated) from the matrix/analyte crystal similar to other
macromolecules.
and the matrix molecules are both soluble in methylene chloride,
the differences in hydrophobicity can likely still cause a high
degree of segregation during the evaporation of the methylene
chloride solvent. For example, the Au:SR cluster compounds with
R ) C4, C6, and C12 are only soluble in nonpolar solvents or
slightly polar solvents. On a microscopic scale, cocrystallization
with a highly polar molecule (e.g., an organic acid) would
presumably produce either a highly amorphous solid or a neat
film of the cluster compounds on the matrix crystal (See Figure
8.). At low irradiance, erosion of the amorphous or neat layer
would presumably require many shots to finally reach a portion
of the matrix/analyte crystal in which the clusters are adequately
dispersed within the crystal structure. This cleaning-off period
would be substantially less at higher irradiance, which is consis-
Figure 9. Positive ion matrix-assisted laser desorption/ionization
mass spectra for the Au:SC4 cluster compound obtained by irradiating
the matrix/sample area at an irradiance of 4.0 MW/cm2 for the initial
64 laser pulses (a) and the average of mass spectra from 32
subsequent laser pulses (b). The dimer ion of the ions centered at
m/z 29 000 is detected at m/z 58 000 in the first 64 laser pulses. Upon
suppression of the m/z 29 000 ions, a distribution of ions at m/z 16 000
is detected in the mass spectrum corresponding to final 32 laser
pulses, presumably the doubly charged species of the group of ions
producing the distribution centered at m/z 32 000.
tent with the observation that, under low irradiance (and the first
few laser shots under high irradiance), the mass spectra obtained
from the matrix/analyte sample is similar to that obtained from
neat films. After the ablation of the neat film or amorphous
material, the ions are instead generated from a well-cocrystallized
matrix/analyte structure in which the cluster compounds are
highly diluted to produce molecular-like ions.
Because ions are formed under many ionization conditions, it
is possible to probe the transition from neat film (or amorphous
matrix/cluster crystallinity) with studies such as those shown in
Figure 6. Several aspects of the complete mass spectra obtained
in this intermediate stage provide additional information in support
of ablation of a neat or amorphous film. Figure 9 shows an
extended mass spectrum obtained at 4.0 MW/cm2 during the
changeover from high- to low-fragmentation regimes. In addition
to the two distributions of ions centered at m/z 29 000 (high
fragmentation) and 32 000 (low fragmentation), two additional
distributions of ions are detected centered at m/z 16 000 and
58 000. The ion at m/z 58 000 is the characteristic dimer ion
produced under LDI conditions, but the m/z 32 000 ion does not
produce a corresponding dimer at m/z 64 000. The peak at m/z
16 000 presumably corresponds to the doubly charged species of
the ions that contribute to the peak detected at m/z 32 000. The
detection of the two additional peaks centered at m/z 58 000 and
16 000 are consistent with the ablation of an amorphous or neat
layer (Figure 8a) followed by desorption and ionization of
desorbed molecules ions from a diluted state within the organic
matrix (Figure 8b).
Complete mitigation of fragmentation during ionization still
seems elusive for this class of macromolecular compounds, but
the combination of different ionization conditions provides a
consistent description of the molecular structure of the Au:SR
cluster compounds. Considering the plume density is likely lowest
for the high-fragmentation MALDI conditions (less propensity for
broadening the distribution of ions due to aggregation with low
m/z ions), this method would provide the more accurate determination for the core mass of the cluster compound at m/z 28 400
(though this number likely includes a small contribution from
remaining sulfur). In addition to sharing a remarkably similar core
mass, the number of ligands is also similar. Assuming the m/z
28 400 represents the lowest fragment of the inorganic core, the
core size is estimated at 144 gold atoms. With this assumption,
the number of ligands associated with all three compounds (i.e.,
mass difference between 28 400 and arrows shown in Figure 7)
suggests all three compounds have ∼53-56 ligand molecules
associated with the condensed-phase structure. While it is still
not possible to unequivocally assign a true “molecular weight”,
the approximate molecular weights of the Au:SC4, Au:SC6 and
Au:SC12 cluster compounds is determined to be ∼33 500, 35 000,
and 39 000, respectively.
Of course, an alternative interpretation considers the mass
distribution of ions produced by MALDI under both high and low
fragmentations. The fwhm of ions produced under all MALDI
conditions is approximately the same for all three compounds.
That is, a similar distribution of ions is detected regardless of
ligand or ionization conditions. Considering the distribution of ions
in this manner would lead to the assumption that an “island of
stability” exists for gold cluster compounds, which is centered at
∼145 atoms. This is also a plausible explanation, considering
different theoretical models that have predicted a number of gold
cluster structures of exceptional stability between 140 and 150
atoms.39,40 From this interpretation, the peak positions and widths
can be treated statistically to determine level of “purity” or size/
molecular weight dispersion, somewhat analogous to a polymer
system. Fitting the distribution of ion abundances under different
ionization to a typical Gaussian distribution function can provide
statistical information regarding constraints on the assembly
composition. For this set of compounds, the core mass is 29 100
( 800 Da, corresponding to a Au core number of 147 ( 4.
Considering some sulfur content is still present, the apex estimate
is likely skewed toward slightly higher core mass, and the
distribution is likely larger than that in the intact cluster compound. The molecular weights and associated distributions of the
Au:SC4, Au:SC6, and Au:SC6 compounds obtained by the same
type of fitting are determined to be 32 000 ( 700, 33 000 ( 1100,
and 37 000 ( 1000, respectively.
Regardless of the interpretation (e.g., based on assumption of
single core and fragmentation or a statistical treatment of the
distribution of ion abundances), it is clear the overall molecular
assembly is highly uniform in structure when the masses of the
respective components are considered. (i.e., gold 197 Da and
(39) Cleveland, C. L.; Luedtke, W. D.; Landman, U. Phys. Rev. Lett. 1998, 81,
2036-2039.
(40) Cleveland, C. L.; Landman, U.; Schaaff, T. G.; Shafigullin, M. N.; Stephens,
P. W.; Whetten, R. L. Phys. Rev. Lett. 1997, 79, 1873-1876.
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
6195
thiolate ligands C4 89 Da, C6 117 Da, and C12 201 Da). The
similarity of the MW distributions to the core mass distribution
though, brings into question this type of treatment. For example,
if MALDI is producing intact ions from compounds with a
distribution of core sizes, there is no (or only slight) statistical
deviation in the number of ligand molecules associated with the
different core sizes. It should also be noted that the approximate
compositions extracted from the statistical approach (Au143-151SC433,
Au143-151SC632-34, Au143-151SC1238-40) are both inconsistent with
elemental analysis and counterintuitive (e.g., more SC12 ligands
associated with assembly than SC6 or SC4). The single core and
fragmentation discussed in the previous paragraph are both
consistent with elemental analysis and intuitively attractive,
considering the isolation of the Pd145 cluster compound by Tran
and co-workers.7
CONCLUSIONS
Cluster compounds are an increasingly important class of
macromolecular structures. In addition to providing excellent
model systems for understanding fundamental quantum effects
in nanostructures, many applications could be potentially impacted
by their further development: catalysis, sensor development,
biologic tags, and molecular-scale electronics. Because the optical
and electronic properties of clusters and nanocrystal compounds
are inexorably linked to the core size of the compound, realization
of these proposed applications requires that methods and techniques be developed to accurately synthesize and isolate the
compounds with distinct inorganic core sizes. Accompanying this
need for isolation of molecular-like structures on the nanometer
scale, high-throughput analytical methods need to address separations and determination of purity, in much the same way as they
are used for other macromolecular structures across chemistry
and biology.
The ability to quickly “size” the gold:thiolate cluster through
monitoring ions generated by LDI-MS of neat films allowed for
efficient optimization of synthetic parameters, as well as a
convenient technique for monitoring size separations. There are
two advantages to the mass spectrometry approach over traditional
inorganic materials analytical methods (e.g., X-ray diffraction and
TEM) for analysis of these nanostructured materials: speed and
statistics. The typical time required to analyze the cluster
compound by MALDI (or LDI) mass spectrometry is on the order
of 2-5 min from sample preparation to data collection and analysis.
6196
Analytical Chemistry, Vol. 76, No. 21, November 1, 2004
The data obtained by mass spectrometry represent the statistical
average of the complete ensemble of cluster compounds. While
X-ray diffraction represents the statistical average of the compounds, it cannot provide information regarding the relative
abundance of different cluster sizes in mixtures or offer qualitative
information regarding the sample purity. For high-resolution
electron microscopy, typical electron micrographs (and histograms
generated from them) represent only a few hundred clusters,
which could be considered a statistically valid representation of
the entire ensemble on cluster compounds. However, the time in
which the structurally relevant information is extracted from these
two methods is usually on the order of hours, not minutes.
With the increasing interest in nanostructured materials, there
is a distinct need for efficient and reliable analytical tools for their
analysis. Mass spectrometry, combined with ionization techniques
developed for other macromolecules, is promising because of its
high sensitivity and ever-increasing mass range and mass resolution. While it is still problematic to obtain completely fragmentation-free MALDI mass spectra for the gold:thiolate and gold:
phosphine cluster compounds, these compounds still constitute
the bulk of any studies involving mass spectrometry and metallic
clusters. Advances in both ionization of the cluster compounds
and application of these advances to other inorganic cluster
compounds will rely on further studies of ionization processes
and optimization of ionization conditions for these selected types
of cluster compounds and other related nanocrystals.
The author thanks Robert L. Whetten and Robert L. Hettich
for their advice and suggestions regarding the drafting of the
manuscript and Gregory B. Hurst for use of the time-of-flight
instrumentation. Research was supported by the Division of
Chemical Sciences, Geosciences and Biosciences, Office of Basic
Energy Sciences, U.S. Department of Energy at Oak Ridge
National Laboratory, managed and operated by UT-Battelle, LLC
under Contract DE-AC05-00OR22725.
SUPPORTING INFORMATION AVAILABLE
Additional information as noted in text. This material is
available free of charge via the Internet at http://pubs.acs.org.
Received for review November 14, 2003. Accepted April
23, 2004.
AC0353482