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OCR for page 65
4
Sampling and Chemical Chamctenzation
This chapter presents a set of general guidelines and strategies to consider
when designing a scheme for sampling and analyzing complex mixtures. No
protocol or group of protocols for sampling or analysis will be applicable or
appropriate for all types of mixtures. What follows is not intended to constitute
a handbook of sampling and analytic methods for complex mixtures. Rather,
researchers should design or modify methods as appropriate in each particular
instance.
However, some general rules can be followed. All of them are based on the
approach in which sampling of a complex mixture has three components: the
relevance of the material collected to the human situation, the use to which the
material will be put, and the potential for human exposure, including routes
and extent of exposure and bioavailability.
CLASSIFICATION
It is important to know as much as possible about the chemical composition
of a mixture before any useful toxicologic prediction can be undertaken (toxi-
cologic testing can be performed on a mixture in the absence of chemical
knowledge if prediction is not the objective). Information on the chemical and
physical properties of a mixture can have a direct impact on the selection of
testing strategies. Once the mixture has been partially defined, sampling tech-
niques, bioavailability, and chemical characterization can be considered.
In light of the understanding that complex mixtures can vary widely from
combustion and distillation products of fossil or synthetic fuels, to pyrolysis
products of tobacco or synthetic materials in buildings, to components leaking
from waste-dump sites some criteria must be identified for the mixture, re-
65
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66
COMPLEX MIXTURES
gardless of origin. The first criterion, physical state, must be determined. One
can encounter such complexity as a mixed-state situation, a slurp of particu-
late matter with a liquid or a gas or vapors and gases, which differs substan-
tially from each fraction and is not toxicologically comparable with the "mix."
The second, chemical-class diversity and component multiplicity, will have to
be evaluated; this is difficult, in that a continuum of complexity can be encoun-
tered, and whether the components are known or unknown will influence the
predictability of the composition and the biohazard. The third criterion, stabil-
ity of the mixture, will be important, because chemical instability (i.e., the
potential for chemical interactions) will seriously affect the reliability of the
sample and the reproducibility of the toxicologic experiment to evaluate the
mixture.
Complex mixtures can be divided according to whether they are derived
from combustion or distillation products or from noncombusted materials. Ex-
amples of the first case are fossil and synthetic fuels and vegetable and syn-
thetic materials. Examples of the second are food, water, and drugs; hazardous
waste; municipal and sewage waste. If one knows the origin of a sample, one
can deduce qualitative and quantitative information about the mixture in ques-
tion, even if physical characterization and chemical characterization are mini-
mal. Extensive information on chemical characterization of a series of fossil-
fuel-related materials has been published (e.g., Wright and Dauble, 19861.
That information makes it possible to predict chemical classes to be found in
mixtures derived from these sources. A detailed discussion of the various ori-
gins of complex mixtures is found in Appendix A.
SAMPLING
Consideration of bioavailability (relevant to human exposure) and of ulti-
mate sample use must be incorporated into the sampling strategy, if it is to be
effective in producing materials for assay that will provide data relevant to
human health. It is important also to consider the preservation of sample integ-
rity; sample alterations must be minimized to ensure that the substance assayed
is the substance presented to the human environment.
COLLECTION STRATEGY
Exposure
In designing a sampling protocol, input from several kinds of specialists is
necessary. Toxicologists, industrial hygienists, or other experts in human
health effects should be consulted regarding the most likely routes of exposure
to the mixture in question. Exposure of humans via inhalation or ingestion of
water or food is most common. In occupational settings, absorption through
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SAMPLING AND CHEMICAL CHARACTERIZATION
67
the skin can also be important. Exposure to a mixture via a combination of
routes is not unusual. For example, particles might be inhaled and later swal-
lowed after being cleared from the lungs by mucocilia~y action. It is usually
practical to consider only the presumed primary route of exposure in designing
a sampling protocol.
Meteorology
Seasonal or meteorologic considerations are particularly germane to the de-
sign of protocols for sampling environmental mixtures. Increases in air tem-
perature can result in loss of benzoLa~pyrene (BaP), a carcinogenic polycyclic
aromatic hydrocarbon (PAH), from particles collected during high-volume at-
mospheric sampling (De Wiest and Rondia, 19761. Seasonal variation can also
affect surface areas and density of atmospheric particles (Corn et al., 1971;
Flessel et al., 19841. Wastewater, soil, and waste-dump sample composition
can be expected to vain seasonally. Changes in temperature, freezing condi-
tions, and rainfall can all modify the water table and influence environmental
sample composition.
Temporal Factors
Industrial hygienists and process engineers should be consulted as to tempo-
ral considerations of sampling. For industrial samples, the time course of pro-
duction is important. One must consider whether to sample only at the end of a
particular batch process, at the middle of a run, or throughout the production of
the material in question. For example, it has been shown that the composition
of coal-liquefaction recycle oils changes markedly during the first few days
after startup of a process, but is relatively stable after that (Burke et al., 19841.
In the case of air sampling at a manufacturing site, both the time of day and the
point in the workweek are important. The variability of samples during a work-
week has been illustrated in the monitoring of oncology nurses handling cyto-
static drugs; the mutagenicity of material extracted from their urine increased
toward the end of the week and decreased during the weekend (Nguyen et al.,
1982).
The duration of sampling is important, particularly for emission materials. If
the substances of interest are in low concentration, the sampling period must be
long enough to collect material required for the assays proposed. This state-
ment of the obvious must be balanced by two other considerations. First, it can
take some preliminary investigation to ascertain the likely concentration and to
determine whether the concentration of the material is relatively stable or likely
to fluctuate. Second, the nature of the collected material might change as the
capacity ofthe collection device is approached. Gorse et al. (1982) showed that
the biologic activity of diesel exhaust particles changed as the collecting filter
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68
COMPLEX MIXTURES
became loaded: the percentage of extractable material, the fluorescence of
high-pressure liquid chromatography (HPLC) fractions, and the mutagenicity
with respect to Salmonella typhimurium all increased linearly, presumably be-
cause the loaded filter became a more efficient collector of mutagenic vapors.
Spatial Considerations
Inclusion of spatial considerations in the sampling design requires the advice
of experts, including industrial hygienists, engineers, and biostatisticians. The
primary consideration should be the relevance of the sample site to potential
human exposure. If the material is from a manufacturing or refining process,
the design should ignore materials in closed systems in favor of open points of
maintenance, ingredient addition, product removal, and so forth. Waste sites
should be studied as to the availability of deposited materials. Personnel access
to the site and the potential for materials to enter water supplies, soil, and food
chains are important. In the case of emission, the nature of the research or
regulatory questions asked will define whether sampling should focus on the
point of discharge, some distant site, or both.
A testing program will probably require consideration of multiple testing
sites for comparative studies. Whether sampling is to be purposive or of a
random statistical design will be determined largely by the nature of the poten-
tial for human exposure.
Purposive sampling requires selection of a set of sites for generating samples
of a particular type. In a study of coal-liquefaction biohazard potential, sam-
pling sites would be at various process points: the coal-slur~ying step, the reac-
tion vessel, points of refining and separation, discharge of wastewater, final
products, recycle materials, and waste. Similarly, sampling could be done in a
waste-dump site with a history of material escape or near an emission source
and at designated distances from it.
Random statistical designs seek to provide an overall picture of an area by
sampling at a statistically determined number of places within that area. In a
series of publications prepared for the Department of Energy, Gilbert (1983,
1984) described a number of sampling designs, generally termed "probability
sampling" (see Table 4-1~. In simple random sampling (see Figure 4-1), eveIy
unit of a target sample population (or geographic area) has an equal probability
of being collected for assay. The units are numbered from 1 to N. and samples
are chosen by drawing n numbers (n < N) from a random-number table or by
use of a computer. The units designated by the selected numbers are sampled.
This mode of sampling is best applied to a homogeneous population with no
dominant spatial or temporal trends. Gilbert noted that most statistical inferen-
tial procedures assume that data were collected through simple random sam-
pling. Sites of likely sampling of complex mixtures might not be homogeneous
with regard to composition or bioavailability of components (e.g., waste
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SAMPLING AND CHEMICAL CHARACTERIZATION
TABLE 4-1 Summary of Sampling Designs and When They Are Most
Useful
69
Sampling Design
Haphazard sampling
Most Useful When
Population homogeneous over time and space essential;
method not recommended, because of difficulty in verifying
assumption of homogeneity
Target population well defined and homogeneous, so
sample-selection bias is not a problem; or specific
environmental samples selected for unique value and
interest, rather than for making inferences to wider
population
Judgment sampling
Probability sampling
Simple random sampling
Stratified random sampling
Systematic sampling
Multistage sampling
Cluster sampling
Double sampling
Homogeneous, i.e., no dominant trends or patterns
Homogeneous within strata (subregions); might want to
consider strata as domains of study
Trends over time or space must be quantified or strictly random
methods are impractical
Target population large and homogeneous; simple random
sampling used to select contiguous groups of population units
Population units cluster (schools of fish, clumps of plants,
etc.); ideally, cluster means are similar in value, but
concentrations within clusters should vary widely
Must be strong linear relation between variable of interest and
less expensive or more easily measured variable
dumps). In addition, practical considerations in the collection of field samples
can limit the use of simple random samples.
Another type of random sampling is stratified sampling. This assumes a
number of nonoverlapping populations that differ from each other, but are
internally homogeneous. The population or study site is divided into as many
strata as appropriate, and simple random sampling is applied to each stratum.
Another technique is multistage subsampling, which involves division of the
site or population into primary units (Cochran, 19771. A set of primary units is
chosen by simple random sampling, and samples are collected within each of
these, under the direction of simple random sampling. This type of design is
particularly applicable to geographic areas with variations in terrain, habitat,
or other factors that require comparison. It is also useful for a relatively homo-
geneous, but large, study site, if the number of samples that can be collected is
limited. In cluster sampling, which is related, population units are grouped into
clusters, a number of clusters are randomly selected, and all units within the
selected clusters are sampled.
Distinct from the random designs are the methods of systematic sampling.
Gilbert (1984) indicated that these methods often find use in environmental
monitoring, because they are generally easier to implement under field condi-
tions. In addition, data from statistical investigations have indicated that sys-
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70
SIMPLE RANDOM
(a)
(c)
(e)
(g)
COMPLEX MIXTURES
STRATIFIED RANDOM
~ -
(b)
· · ~ -
· /- ~
TWO-STAGE
SAMPLING
I A. I . I . |
. .
PRIMARY
UNITS — (d) · | · | · |
\m -- 1 1
_ . . _
SYSTEMATIC GRID
RANDOM WITHIN
BLOCKS
.. 1 . .. ...
. I 1. 1- (flu .~.~.~-e
1. 1.1- .-- ·. A.
SYSTEMATIC
UNALIGNED
_
AI B I C I D
·E I F |.G |.H
_~1 J it ;
(h)
SYSTEMATIC RADII
CLUSTER SAMPLING
6~9
CLUSTERS
FIGURE 4-1 Some probability field sampling designs. Reprinted with permission from Gilbert
(1983).
tematic sampling can be superior to simple or stratified random sampling for
some types of environmental variables. In general, systematic sampling begins
with random selection of one unit of the study population. This serves as the
starting point of a sampling pattern according to some established spatial or
temporal frequency. A problem common to systematic designs is that, if the
variable being measured is subject to periodicity or cycles, misleading results
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SAMPLING AND CHEMICAL CHARACTERIZATION
71
will be obtained. Gilbert cited another difficulty: the accuracy of estimates of
sampling errors and other statistics depends on the study population's being
random.
In the aligned-square grid design, a location (or other unit) is chosen at
random. The location of each sample is then chosen by applying a grid of fixed
dimensions over the chosen area, and two random coordinate numbers are
drawn to fix the location of the original point. Each sample site is then fixed in
relation to this original point. Variations include the use of a triangular grid and
the unaligned-grid-pattern design. In the latter (Figure 4-2), a point A is chosen
randomly, and X and Y coordinates are established. To set points B. C, and D,
one uses the X coordinate of A and three new randomly chosen Y coordinates.
To set points E through I, the point A coordinate Y and random X coordinates
are used.
One can also design systematic sampling methods based on lines. Figure 4-1
illustrates periodic sampling of radii from a central point, such as a known
discharge source.
Aligned Square Grid
·
I ~ I I J I
-HA my- ~ -- T.--
(a)
(c)
Central Aligned
Square Grid
_ —_ - - · — 1_ - -
I1 1
1
2- it- 1~- r.
(b)
:
l 1 , 1 1 1 1
- . - 1 _ - _ _ —
1 1
t_~-
_ ~ ~ ~ I
, 1 1 1
Unaligned Grid
~ .BI I .D| ~—Hi— Hi'—- '—~1
·E | I G I ·H 1 (d) ) \\ >/ \ A\ ,~ \ I
~ I ~ K .~ | ~ / \|\ / ~ / ~ / \
Triangular Grid
FIGURE 4-2 Some systematic designs for sampling in space. Reprinted with permission from Gilbert
(1984).
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72
COMPLEX MIXTURES
COLLECTION PROCEDURE
Designing the collection process requires the input of a toxicologist to pro-
vide information on the nature of the assays and of a chemist and an engineer to
produce a protocol that will efficiently provide a relevant sample material. One
type of information to be supplied by a toxicologist is the amount of material
that needs to be collected. Testing protocols geared to chemical analysis gener-
ally require smaller samples than more biologically oriented approaches. In
vitro tests are more sparing of sample than are whole-animal studies.
The collection procedure depends largely on the source of the material and
its physical state. Environmental samples are likely to be mixtures of materials
in various physical states. They might include mixtures of gases, aerosolized
liquids, and suspended particles; combinations of liquid and solid waste; bio-
logic tissues, which are largely water; and so forth. We present here examples
of sample types and methods deemed appropriate for their collection. Many of
these were described at greater length in a recent EPA document (U. S. EPA,
1985), and some useful techniques were also reviewed by Alfheim et al.
(19841.
Gases and Aerosols
Vapor-phase organic chemicals have been collected from ambient air, com-
bustion exhaust gases, cigarette smoke, and indoor spaces for chemical charac-
terization and bioassay (Hanson et al., 1981 , 1984; Griest et al., 1982; Higgins
et al., 1983; Pellizzari et al., 1976; Pellizzari, 1982; Hughes et al., 1980;
Krost et al., 1982~. Chemical transformations and perturbations of the original
chemical mixture can occur during the sampling step (Berkley and Pellizzari,
1978; Pellizzari et al., 1984; Pellizzari and Krost, 19841.
Air particles are a common source of mixtures that have environmental or
health significance. Alfheim et al. (1984) recommended that particle-size frac-
tionation be included in the collection scheme to increase the biologic rele-
vance of a particulate sample. The International Standards Organization (ISO,
1983) and the American Conference of Governmental Industrial Hygienists
(Lippmann, 1985) have established cut sizes appropriate to fractionation. They
depend on the toxicity at deposition sites in the airways or at sites along the
clearance pathways or storage sites. Inspirable particles are those which can be
aspirated by the nose or mouth. Thoracic particles are those which can enter the
thorax. Respirable particles are those which penetrate the lung's conductive
airways. Another reason for size separation is that the chemical composition
and biologic activities are related to particle size.
Among urban air particles, small size is associated with the presence of
acidic compounds, and larger sizes with basic groups (Miller et al., 19791. In
comparative assays, most mutagenic activity has been associated with smaller
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SAMPLING AND CHEMICAL CHARACTERIZATION
73
particles, less than 2.5,um in diameter (Talcott and Harger, 1980; Preidecker,
1980; Chrisp and Fisher, 19801. Most of the total PAH content of air particles
is found in particles less than 5 ,um in diameter. It has been recommended,
however, that results of assays of particles collected with size fractionation be
compared with results of assays of particles collected without fractionation.
Artifacts of size separation reportedly have reduced activity in mutagenicity
assays (Alfl~eim et al., 1983), whereas other chemical transformations report-
edly have increased mutagenicity (Clark et al., 19811.
Point-source sampling for air particles includes use of impactors (Cheng and
Yeh, 1979), cyclones, electrostatic precipitators, and filters (U.S. EPA, 1985~.
Sampling methods have been described by Brusick and Young (1982) and
Lentzen et al. (19781. Collection of particles from ambient air generally uses
high-volume samplers (U.S. EPA, 1971), so-called massive-volume samplers
(Henry et al., 1978; Cheng et al., 1984), and medium-, low-, or ultrahigh-
volume samplers (Fitz et al., 19831. It has been recommended (U.S. EPA,
1985) that preparation of particulate samples for bacterial-mutagenicity moni-
toring begin with collection of standard high-volume samples with inert filter-
collection media. For collection of larger amounts of material or when circum-
stances dictate a shorter collection period, use of an ultrahigh-volume sampler
would be appropriate (U.S. EPA, 19851.
If gaseous emission is collected at high temperature, there will be less oppor-
tunity for condensation of materials on particles. To collect a gaseous mixture
that adequately reflects the material being produced or emitted to the human
environment, the possible presence of volatile materials must be considered in
the collection scheme. When it is feasible, a gas can be cooled before introduc-
tion to an adsorbent material. Alfheim et al. (1984) indicated that XAD-2 resin
was most commonly used for this purpose. Gaseous samples can thus be re-
duced to collected particles, extracts from absorbents, and condensates. The
testing of volatile materials not captured that way is a subject for research and is
not generally applicable to routine environmental monitoring or widely used in
bioassays.
Aqueous Materials
Aqueous samples may be collected in a relatively homogeneous state or may
contain suspended solids. The most common method of sampling water is
manual-grab collection of the required volume. Water generally has low con-
centrations of biologically active materials, which must be concentrated for
application in most tests. Concentration protocols are therefore identified as
the major part of the collection procedure. Methods for concentration of water
with less than 5 % associated solids, particularly drinking water, have been well
described (Kopfler, 1980; Jolley, 1981; NRC, 19821. Aqueous solutions can
be concentrated by removing the water via freeze concentration, lyophiliza-
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74
COMPLEX MIXTURES
lion, vacuum evaporation, and reverse osmosis (Shapiro, 1961; Baker, 1970;
Dawson and Mopper, 1978; Crathorne et al., 1979; Jolley et al., 1975; Kopfler
et al., 19771. These methods concentrate organic and inorganic materials to-
gether, a procedure that Kopfler (1980) claimed was a disadvantage if the bio-
assay system to be used could not tolerate high concentrations of inorganic
materials.
Another scheme involves selective concentration of organic materials from
drinking water by adsorption of the contaminants on activated carbon or XAD
resin columns (Middleton et al., 1956; Kopfler, 1980; McGuire and Suffet,
1983; U.S. EPA, 1985~. A variety of solvents can be used for elusion of or-
ganic substances from the activated-carbon columns. Supercritical liquid car-
bon dioxide is an efficient elusion solvent (Modell et al., 1978~. This method
can be used for large-scale processing of water samples and results in the re-
cove~y of gram quantities of contaminants. The limitations of these techniques
include the nonextraction or nonrecove~y of inorganic materials and highly
polar materials, ionic organic species, or volatile low-molecular-weight or-
ganic compounds. Methods that might be suitable for collection of these sub-
stances were cited by EPA (19851. Kopfler (1980) noted that organic acids and
bases might be amenable to collection on resin columns after adjustment of the
water pH to suppress ionization. Altering the pH of a mixture of materials,
however, also results in changing its chemical composition or biologic activity.
Aqueous samples with large amounts of suspended solids are generally best
prepared by separation ofthe phases. Gravity partitioning is done by storing the
sample at 4°C for 24 hours. The solid phase can then be removed and pro-
cessed separately for assay. Alternatively, high-pressure filtration or high-
speed centrifugation can be used for phase separation. The liquid portion can
be separated into aqueous and nonaqueous components, concentrated if neces-
saty, and assayed. Some samples with suspended solids are amenable to pro-
cessing by liquid-liquid extraction methods. Details of some of these protocols
were described by EPA (19851.
Nonaqueous Materials
Many environmental and indust~y-related samples are nonaqueous liquids,
including organic liquids, light and heavy oils, and some tars. Samples can
contain mixtures of volatile (b.p., 36-100°C), moderately volatile (b.p., 100-
300°C), and nonvolatile (b.p., over 300°C) materials, and there might be
associated solids. Generally, the composition of such materials is poorly char-
acterized and subject to variation after sampling. Much of the information on
samples of this sort has been derived from research on fossil-fuel materials
(Wright and Dauble, 19861. Sampling itself is usually not difficult; grab sam-
pling often suffices. The samples are generally concentrated enough for assay
of undiluted or unprocessed material. The preparatory issue with these materi-
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SAMPLING AND CHEMICAL CHARACTERIZATION
75
als therefore is not sampling, but rasher treatment to make the sampled material
compatible with the bioassay in question. Some of these issues are discussed
later.
Solids and Sediments
Solid materials appear to be amenable to simple sampling techniques. How-
ever, the entire mass of material collected might not constitute the sample of
interest. Soils and sediments are examples of materials wherein an unknown or
small concentration of a substance of biologic interest is associated with a large
quantity of matrix. The protocol for these materials must therefore include
extraction and concentration. Specific recommendations for treatment of soils
were made by EPA (19851.
Efforts should first be made to provide a homogeneous sample. Aggregates
can be broken by crushing or cutting. To avoid a particle-size bias, the material
can be quartered. This is done by spreading out the material on a clean surface
and mechanically dividing the sample into four parts; three parts are returned to
the storage vessel, and the remainder is quartered until the desired amount is
obtained.
Sediments might contain water, which can be removed as described previ-
ously. The next step should be extraction in a Soxhlet apparatus with highly
pure organic solvents. The nature of material to be recovered will depend
largely on the solvents used. Solvents used for extracting materials from soil
matrices for chemical analysis have included the following: benzene, ethyl
acetate, and benzene-methanol-acetone (2: 1: 1) for humic substances (Ogner
and Schnitzer, 1970; Cifrulak, 19691; n-pentane, carbon tetrachloride, and
methylene chloride for oil (Jobson et al., 1974; Jensen, 1975; McGill and
Rowell, 1980~; and hexane-isopropanol (3:1) for polychlorinated biphenyls
(Carey and Gowen, 19781.
Other extracting solvents have been used specifically to produce samples
that would be amenable to biologic testing. Mutagenic materials were obtained
from agricultural soils extracted with hexane-acetone (2:1) and dichloro-
methane (Goggelmann and Spitzauer, 1982; Brown et al., 19851. Likewise,
mutagenic sediment extracts have been obtained through sequential extraction
with diethyl ether and methanol (Kinae et al., 198 11. Ether has also been used
on sediments (Sato et al., 19831. The choice of solvents can likely be dictated
solely by the nature of the material to be recovered, if the solvent can be evapo-
rated or otherwise removed before bioassay. EPA (1985) has recommended
rotary evaporation. The pH of the soil or sediment will be a major influence on
the types of compounds extracted.
For most other types of solid samples, the primary preparative consideration
will be to constitute a material compatible with the bioassay selected. The
sample must be representative of the environmental or human exposure poten-
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88
COMPLEX MIXTURES
TABLE 4-4 Conventional Instrumental Techniques for Chemical Analysis
Instrumental
Technique
Ultraviolet and visible
spectroscopy
Vibrational spectroscopy
(infrared and Raman)
Information
Content
Conjugation
Selected
References
Brown, 1980; Silverstein and
Bassler, 1963
Borwn, 1980; Strommen and
Nakamoto, 1984; Gans,
1980; Case and Fately,
1980; Clerc et al., 1981
Penzer, 1980
Chemical functionalities and
assembly of atoms
Molecular emission
spectroscopy
Mass spectrometer
Nuclear magnetic resonance
spectrometer
Atomic absorption
spectroscopy and
inductively coupled argon
. . .
plasma emission
Electron paramagnetic spin
resonance
Selective detection
Molecular weight, chemical
functionalities, overall
structure, and assembly of
atoms
Anangement of atoms in
molecule
Elements
Free-radical formation Brown, 1980
Watson and Throck, 1985;
Wilson et al., 1981; Smith
and Udseth, 1983
Jones, 1980; Jackrnan and
Sternhell, 1969
Wood, 1980; Cantle, 1982
used to identify the chemicals responsible for toxicity. Extreme caution should
be exercised when one is comparing chemical data bases derived from different
complex mixtures, because the separation techniques introduce uncertainty
as to whether structural elucidation is unequivocal or merely postulative or
tentative.
For structural analysis, these modern methods are used most efficiently if
they are combined, because they can provide complementary information that
increases their overall effectiveness. Guidelines for the combined application
of spectroscopic methods are available (Clerc et al., 1981; Silverstein and
Bassler, 19631. The most generally applied methods for the characterization of
organic substances are ultraviolet spectroscopy, infrared spectroscopy, proton
and carbon-13 nuclear magnetic resonance, and mass spectrometer (Clerc et
al., 19811. Specialized techniques such as magnetic resonance of other nu-
clei, Raman spectroscopy, and optical rotary dispersion are used less often.
Gas chromatography/Fourier-transform infrared (GC/FT-IR) spectroscopy is
now being developed for routine use in identifying analyses in complex mix-
tures with the aid of software programs for the automatic interpretation of
spectra (Growths, 1980; Gurka, 1985~. Infrared, Raman, visible, ultraviolet,
electron-spin resonance, and magnetic circular dichroism spectroscopies have
all been used in conjunction with matrix isolation (Barnes and Orville-Thomas,
19801. Matrix isolation traps isolated molecules of the species to be studied in a
OCR for page 89
SAMPLING AND CHEMICAL CHARACTERIZATION
89
large excess of an inert material by rapid condensation at a low temperature, so
that the diluent forms a rigid cage or matrix (Barnes and Orville-Thomas,
1980~. The noble gases (primarily argon) and nitrogen are most widely used for
matrix materials. More recent uses of matrix isolation have been in combina-
tion with GC effluents in which high-resolution gas chromatography (HRGC)
has been coupled with FT-IR spectroscopy. Several HPLC/FT-IR systems
have been described (Growths, 19801. The exquisite sensitivity of GC/FT-IR
with matrix isolation rivals that of mass spectrometer; these two methods can
also be combined to provide simultaneous and complementary information
during chromatography of a complex mixture.
The gas chromatograph is the most commonly used inlet system for analysis
of complex mixtures with mass spectrometry (Watson and Throck, 19851. It is
rarely possible to isolate in pure form each of the hundreds of individual sub-
stances present. However, HRGC combined with mass spectrometer (GC/
MS) can often serve as the final purification step by resolving the various
components and presenting them one at a time to the mass spectrometer.
The mass spectrometer can also serve as a universal detector for SFC and
HPLC (Smith and Udseth, 19831. Several types of devices are under develop-
ment (e.g., moving wire or moving belt, direct inlet, and thermospray) to
introduce relatively nonvolatile materials after separation into the ion source of
the mass spectrometer. A fast-atom-bombardment (FAB) ionization technique
combined with a moving belt exploits the attributes of FAB ionization of non-
volatile analyses and allows analysis in a manner very similar to that of GC/MS
(Smith and Udseth, 19831.
An alternative to resolving components chromatographically is separating
them by mass spectrometric methods the so-called MS/MS methods (Watson
and Throck, 1985; Smith and Udseth, 1983; Henderson et al., 1982, 1983,
19841. The complex mixture is ionized, usually gently, to produce charactens-
tic ions from each component. The ions suspected to arise from the component
of interest are selected by mass analysis in the first stage of a double mass
spectrometer, and their identity is confirmed by mass analysis in the second
stage on the basis of the fragment ions arising from collision-induced associa-
tion of the selected ions.
REFERENCES
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Representative terms from entire chapter:
chemical characterization
