Anal. Chem. 1985, 57, 1075-1079
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Sample Preparation and System Calibration for Proton-Induced X-ray Emission Analysis of Hair from Occupationally Exposed Workers Eric Clayton* Applied Physics Division, Australian Atomic Energy Commission Research Establishment, Lucas Heights Laboratories, Private Mail Bag, Sutherland, New South Wales 2232, Australia
K. K. Wooller Department of Industrial Relations, Division of Occupational Health, P.O. Box 163,Lidcombe, New South Wales 2141, Australia
Blood and urine collections are commonly used to monitor trace element concentration in the body. Hair may also be used. The concentration of many elements is much higher in hair than in either blood or urine and may provide a ready record of a period of exposure to heavy metals. Proton-induced X-ray emission (PIXE) has been used to study trace elements in hair. A method of preparing samples by charring hair and mixing with yttrium-splked graphite has been developed. Thick targets suitable for PIXE analysis are made. The trace elements K, Ca, Ti, Mn, Fe, Ni, Cu, Zn, Pb, Br, Rb, and Sr are routinely measured in most samples and Cr, Co, As, Bi, $e, Zr, and Cd can be measured in occupationally exposed workers. Helium backscattering and elastic recoil detection are used to determine the malor components H,C, N, 0, and S.
The collection of blood and urine for analysis of trace element concentrations is standard practice. Hair may also be useful as its collection is noninvasive, it can be stored, and its analysis for trace elements is reasonably easy. Moreover, the concentration of many trace elements is much higher in hair than in blood or urine and hair may provide a ready record of a much longer period of exposure to heavy metals than either. Proton-induced X-ray emission (PIXE) has been used here in a survey of trace element concentrations in hair. One virtue of PIXE analysis is that it can routinely analyze such elements as Pb, a difficult task for neutron activation analysis, or the nonmetallic elements such as C1 or Br which cannot be analyzed by atomic spectrometry. The survey had two aims. First to establish a data base of concentration data from control groups to compare with occupationally exposed groups and second to correlate hair concentrations with the appropriate blood or urine levels. There are two control groups with approximately 100 samples in the group from the country and 55 in the group from metropolitan Sydney (population 3.5 million). Approximately 100 samples were taken from occupationally exposed workers. Because they are exposed to a variety of elements-Pb, Cd, and Zn in smelters, P b and Cd in battery factories, and As in chemical plants-a multielement technique capable of sensitivity to the Mg/g level is desirable, hence the selection of PIXE. This paper reports on the techniques used in sample preparation and analysis. Previous work on hair analysis by PIXE has been undertaken in two ways: probe analysis, either across (I,2) or along (3)the hair shaft or root (4),and preparation of a homogeneous sample. The latter approach may involve ashing (4),dissolution in acid (6),or preparation at liquid nitrogen temper0003-2700/85/0357-1075$01.50/0
atures (7). Whitehead (8)has discussed some of the problems arising in both probe and homogeneous sample analyses. For this survey, a preparation technique involving charring of approximately 100-mg samples was developed. Methods used to estimate trace element concentrations are given. Helium ions were used to provide values for the major components of hair, viz., H, C, N, 0, and S. Backscattering and elastic recoil detection methods to determine these components are mentioned. EXPERIMENTAL SECTION PIXE spectra were measured by using a 2.5-MeV proton beam in vacuum with currents between 30 and 40 nA (beam size 2 mm). An accumulated charge of 50 pC was collected, taking some 20-25 min per sample. X-rays were detected with a 4 mm diameter Si(Li) detector (fwhm 140 eV at 5.9 keV) set at an angle of 135’ to the incident beam and 62 mm from the target. A 0.526 mm thick Perspex filter was used to attenuate low-energy X-rays so that only potassium and heavier elements could be analyzed by this system. The X-ray spectra were analyzed off line by using the programs described by Clayton et al. (9-11). Helium elastic recoil spectra were measured with a very simple arrangement. A beam of 2.6-MeV 4He+ions is incident at a glancing angle to the sample (loo)and elastically scattered in the sample. Hydrogen ions recoil only in the forward direction and are detected by a surface barrier detector placed at an angle of loo to the surface. Simple kinematics (12)show that the recoil H ions have a maximum energy of 1.47 MeV. A 16 Mm thick aluminum foil placed in front of the detector is sufficient to stop the He ions scattered from the sample while allowing the recoil H ions from the surface through with an energy of 913 keV. Helium Backscattering. The use of backscattering in elemental analysis is based both on the energy loss of the projectile when scattered from a target nucleus and the charge of the target nucleus. Several elements mixed homogeneously in a thick target give a backscatter spectrum consisting of a number of steps of different heights. The position of each step is determined by the mass of the scattering element and its height is a measure of the concentration of that element in the sample (13,14). Spectra were measured with a beam of 1.4 MeV 4He+ions incident normally to the sample surface, at a target current of 50 nA through a 1-mm beam. Two hundred microcoulombs of charge was collected. The backscattered particles were detected with a silicon surface barrier detector set at 1 3 5 O to the beam. Sample Collection. Hair samples were obtained by clipping close to the scalp with scissors at the back of the head (vertico posterior). Approximately 60 to 100 mg of hair was taken from each volunteer. Samples were stored in a sealed, numbered, low density clear polythene bag. Strands up to 10 cm long were accepted, those over this length being cut and the excess being stored separately, but not analyzed. During sample collection, 13 volunteers (5% of the group) had duplicate samples taken. This involved splitting the sample and storing it as two separate samples which were to be analyzed “blind”. From each donor, 0 1985 Arnerlcan Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985 l 0 F
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Table I. Concentrations, Count Rates, and Minimum Detection Limits for a 50 pC Measurement of an Illustrative Sample Encountered during Routine Analysis element
K Ca
Ti
Mn Fe Ni
cu
2.0
6.0
10.0 14.0 X-RRY ENERGY ( K E V I .
18.0
Figure 1. PIXE spectrum from a typical metropolitan control group sample. A reduced x2 of 3.9 was obtalned In this analysis. a complete medical, occupational, and residential history was taken. This included the date of the last hair wash, the frequency of washing, and information on hair preparation used. As this work is concerned with occupational exposure, the survey took data only from males of working age, thus avoiding problems of trace element difference between the sexes. The question of whether to wash the samples is very complex with the work of many proponents of both positions appearing in the literature. We decided to follow the view of Chittleborough (15) who proposed an holistic approach of leaving samples unwashed. Thus each sample will be a unique record of possible environmental exposure. Sample Preparation. Direct irradiation of hair by a proton beam has been discussed by several authors (1-6). This procedure has problems with electrostatic charging of the sample and determining absolute concentrations for trace elements. Direct analysis has, however, been successful for profiling concentrations along the hair shaft (3), showing the time response to exposure. As this survey is concerned with the effects of occupational exposure, with the implication of long-term exposure, it was decided to homogenize the sample before analysis. Thus the entire sample with ita information of many months growth (possibly up to a year) could be used. It was decided to powder the samples rather than dissolve them in acid to make thin targets (16). To powder the samples, the hair was charred in an oven for 2 h at 160 "C, the maximum temperature available. A small fraction of the samples (less than 3%) required a longer period of charring. The charring would be more efficient if higher temperatures can be used. Pyrex beakers were used to hold the hair. The average weight loss during charring was 30%. After charring, the samples were stored in a desiccator until the PIXE targets were made. As indicated by an increase in measured elemental X-ray intensities, biological samples tend to be unstable under irradiation in vacuum (17). To overcome the problem of stability,the charred samples were thoroughly mixed with spectroscopic grade graphite in an electrical shaker. Mixing with graphite allows higher beam currents to be used (18)and also prevents electrostatic charging. The final target composition was four parts by weight of charred sample to one part by weight of graphite. This mixture was shaken for 20 min in a plastic bottle with 10-mm Plexiglas balls to homogenize the sample. After shaking, the mixture was pressed into 12 mm diameter aluminum caps at a pressure of 35 MPa. It was easy to spike the graphite with 820 pg/g of yttrium so an internal standard could be obtained. Yttrium was chosen as it does not appear naturally in hair.
DISCUSSION In Figure 1 an X-ray spectrum from the hair of one member of the metropolitan control group demonstrates the multielement capability of PIXE. The trace elements K, Ca, Ti, Mn, Fe, Ni, Cu, Zn, Pb, Br, and Sr were routinely determined in most samples and Co, As, Bi, Se, Zr, and Cd can be measured in occupationally exposed workers. If the 0.526-mm
Zn Pb Br Sr Y Cd
concn, pglg
counts per 50 pC
200 700 10 2 60 2 50 240 15 20 3 160 (in C)
1100 15500 900 320 9700 270 5500 20200 250 550 50 1700
MDL, pg/g 20 10
2 0.8 0.6 0.5 0.5 0.5 2 0.7 1.0
1.0 20
Perspex filter is removed, the additional elements S and C1 can also be measured. The experimental conditions used resulted in the typical count rates and minimum detection limits (MDL) (10)given in Table I. The concentrations shown are representative of the survey values. Before sample preparation by charring and mixing with graphite can be applied, it is necessary to check on two potential problems: loss of trace elements during charring and contamination during the process. A large (3 g) sample of hair was subsampled and analyzed to monitor sample preparation steps. Analyses were performed by neutron activation analysis and flame atomic spectrometry on (i) the original sample, (ii) the charred powder, and (iii) the powder mixed with graphite. Subsample (iii) was analyzed by PIXE also. As well as checking for element loss or contamination, the results presented in Table I1 allow us to verify the accuracy of our PIXE measurements by comparison with other techniques. Although not all elements could be measured by all three methods, only C1 and Hg suffered significant loss during sample preparation. The presence of some rare earths in the graphite a t submicrogram per gram levels was also noticed. This is not significant for this analysis as our PIXE measurements were not sensitive enough to determine these low levels of the rare earths. Elements such as Ti and Ni which were given only in the PIXE measurement are presented for information illustrating hair concentrations for these elements. Overall, this table shows that the preparation technique does not distort trace element concentrations in hair; furthermore, the good agreement between the three analysis methods gives some confidence in the PIXE method. Replicate Analysis. After analysis the trace element concentration data of the 13 duplicate samples were assessed to provide an indication of the technique's reproducibility under realistic working conditions. The ratio of the trace element concentration of the sample to its duplicate has been averaged over all the pairs (Figure 2). Not shown is the ratio for P b of 1.05 0.15. Overall, there is good agreement for most elements. This subsample has not revealed any systematic errors or change in experimental conditions during the measurements. The results indicate that the standard deviations were heavily weighted by the discrepant values for one or two of the 13 pairs (say a ratio of 0.5 or 2.0 for instance). For all samples in which an element was discrepant, the other elements gave acceptable ratios. Following the arguments of Gibbons Natrella (19) on discrepant results, the Dixon criterion for rejecting observations was applied to the data. Two pairs were deleted from the K, Ti, and Fe set and one from the Ca data. Averaging over the reduced set gave the results shown in Figure 2. In summary, this shows that the reproducibility of the technique is acceptable and illustrates the
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 6, MAY 1985
1077
Table 11. Concentration Data from Three Subsamples of Hair: (i) the Original Sample, (ii) the Sample after Charring, and (iii) the Charred Sample with Added Graphitea concentrations, u d p ~~
316 17 1104 150 342
323 19 1248 166
Na A1
c1
K Ca Ti Mn Fe Ni
(iii)
328 19 1098 162
0.32 32
0.35 29
PIXE
FAS
NAA (ii)
(9
element
(9
(ii)
(iii)
av NAA
av FAS
(iii)
143 343
123 398
181 415
159 342
161 385
140 363 4.2
0.33 28
37
30
2.1
cu
Zn Hg Pb Se Br Rb Sr
163 3.0
160 1.2
164 1.2
0.4 262
0.4 253
0.4 257
27 124
25 134
19 153
8
9
11
162
257
24 137
27 150
9
11
310 10 2.1
OValues shown are given in terms of the original hair sample by compensating for the effects of weight loss during charring and graphite addition. NAA and flame atomic spectrometry measurements are given together with their averages and a PIXE measurement.
t 0.6
1
-
I
1
1
To check the accuracy of these calculations, two standard reference materials (SRM), NBS1571 Orchard Leaves and NBS1577 Bovine Liver, have been analyzed by the methods reported here. They were mixed with graphite, and included with the hair samples for analysis under the same routine conditions as the survey samples. Four measurements were made of each. Their major component concentrations are taken from Gladney (20). Trace element concentrations given by calculated yields (Table 111) indicate that an accurate estimate can be given by the techniques reported here. Determining the Major Components of Hair. An initial estimate for the major components in hair is given by the ICRP reference man (21): H, 7.56%; C , 46.27%; N, 13.69%; 0, 28.33%; and S, 4.15%. For eq 2 an estimate of the composition after charring is required. A sample having a dry/fresh weight ratio nearly that of the average of all the samples (70%) was selected for an analysis of the major components. Carbon, nitrogen, oxygen, and sulfur can be determined by helium backscattering and the hydrogen concentration by helium elastic recoil detection (22, 23). Hydrogen Measurement. Two amino acids, cystine (CsH12N204Sz) and glycine (C2H5N0J,were used to estimate the hydrogen atom density. Both were mixed with graphite as in the sample preparation step. After mixing, the hydrogen atom density is 38.71 at. % for cystine and 43.24 at. 70 for glycine. Figure 3 shows the recoil spectrum from the hair sample. Data were analyzed by summing the counts in channels 150 to 200. This is below the surface energy which occurred at channel 250 for this geometry but was chosen to exclude the possibility of analyzing adsorbed hydrogen on the surface of the sample. Ten microcoulombs of charge was accumulated with the beam current being kept below 10 nA to reduce the problem of beam heating of the sample. Multiple runs were necessary for both acids and hair sample as individual measurements had quite a large spread owing to a number of experimental difficulties. These included beam heating, sample roughness, imprecision in the geometry, and charge collection. Final results were 17 612 f 7000 counts for glycine and 16 344 f 5000 for cystine. The ratio of counts was 1.08 which is in reasonably good agreement with the expected atom density ratio of 1.12. For the hair sample, summing gave 16 504 f 6000 counts, which corresponds to an atom density of 39.8 i 14.5 at. %. Although there is a large error on the
I
0.4 15.0
20.0
30.0 ELEflENT 2
25.0
5.0
40.0
Figure 2. Result of blind dupllcate analysls performed on 13 pairs of samples. The error bars shown encompass the mean f l u values.
natural variation that must be expected when dnalyzing biological systems which, after all, are not as rigidly controlled as a standard reference material. Determining the Trace Element Yield Curve. Given the major element composition for a sample, it is possible to calculate the X-ray yield for a given element in a thick target by modeling proton slowing down and estimating X-ray absorption in the sample (9,lO). As the graphite has been spiked with yttrium (820 pg/g) it is only necessary to calculate the yield relative to Y. This removes uncertainties about absolute charge collection and solid angle. As
N1 c M ~ (1) where N1is the number of X-rays detected, c is the concentration of the element, and Ml is the calculated yield of X-rays per unit concentration, then given an internal standard, it follows that c = N~c,/N~R (2)
R = MI/M2
(3) where c2 is the concentration of the standard, N2 its X-ray counts, and R is the ratio of the X-ray yield per unit concentration for sample and standard (9, 10).
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ANALYTICAL CHEMISTRY, VOL. 57, NO. 0, MAY 1985
Table 111. Trace Element Concentrations for Two Standards element
PIXE
K (%I
orchard leavesa ref 20
1.41 f 0.09 1.69 f 0.05 21.9 f 3.0 35.9 f 7.0 2.4 f 1.1 98.9 f 11.0 266 i 21 1.5 f 0.7 12.4 f 1.4 24.4 f 0.9 15.7 f 5.0 46.1 f 7.0
Ca (%) Ti Ba Cr Mn Fe Ni
cu
Zn As
Pb
bovine liverb
1.46 & 0.06 2.02 f 0.09 20 i 2 45 f 8 2.48 f 0.34 85 f 5 280 f 50 1.7 f 1.0 11.7 f 1.4 25 f 3 11.5 f 2.1 46 f 4
ratio
PIXE
ref 20
ratio
0.97 0.84 1.10 0.80 0.98
0.85 f 0.02 143 f 19
0.943 f 0.09 116 f 18
0.90 1.23
1.11
9.7 f 0.3 269 f 9
10.2 f 0.9 263 f 13
0.95 1.02
191 f 9 135 f 5
189 f 21 132 f 10
1.01 1.02
1.24 f 0.3
1.08 i 0.07 8.7 f 1.7 19.5 f 3.6
1.15 1.10 0.98
0.95 0.88 1.06 0.98 1.37
1.00
Se
Br 9.8 f 1.1 9.3 f 1.2 1.05 9.6 f 0.7 Rb 12.8 f 0.7 11.6 f 1.2 1.10 19.1 i 0.8 Sr 36.3 f 1.8 36 f 4 1.01 aAverage ratio PIXE/SRM, 1.01 f 0.14. bAverage ratio PIXE/SRM, 1.04 f 0.10.
0.0 0. 0
100 0
200.0
300.0
IO ‘
410 0
1. 0
hydrogen measurement, this has little effect on the PIXE yield calculation of eq 2. Varying the hydrogen content between 25 and 55 at. % changes the calculated yield by 5% for K and by less than 1% for elements heavier than Ti. C, N,0 , and S Measurement. Figure 4 shows the backscattering spectrum from the hair sample used for major element determination. Steps arising from C, N, 0, and S are readily apparent. The ratio of the heights of steps corresponding to elements A and B is given in ref 13
HB
- NAaA(EO)[ ~ ] B m i x NBaB(Et))[ €]Amix
120.0
CHWNEL NO.
Flgure 3. Helium eiastlc recoil spectrum used to determine the hydrogen content of hair.
H _A
m. n
49.0
CHfiNNEL NO.
Flgure 4. Rutherford backscatterlng spectrum for ‘Het ions incident on a pressed halr sample. Steps from C, N, 0, and S are clearly visible.
0
(4)
where HAis the step height (counts) for element A, N A is the number of atoms of type A in the mixture, UA is the Rutherford scattering cross section for A, and [cIAmh is the stopping cross section factor for A; similar definitions hold for element B. Eo is the incident energy (1.4 MeV). If [e] values are required, the composition of the sample has to be known. Fortunately, the ratio of the stopping cross section factors in eq 4 can initially be set to unity, as this approximation is sufficiently accurate to provide reasonable estimates for an iterative calculation in which the values for atom numbers from the “zerothniteration can be inserted into eq 4 and the process repeated. The ratio in most cases will be within 10% of unity. Using this calculation for the spectrum of Figure 4 resulted in the following composition of the charred sample: C, 72.21%; N, 13.56%; 0, 5.36%; S, 2.60%; and H, 6.27%. S was lost
0.85 18.0
24.0
30.0
36.0
42.0
ELENENT 2
Flgure 5. Ratlo of the calculated yield R from eq 2 for different standards relative to the halr value. Half is represented by the solid ilne, graphite by the short dashed Ilne, orchard leaves by the long dashed ilne, and bovine liver by the long and short dashed line.
during charring. A zero filter PIXE analysis confirmed this result. The above composition was used to calculate the PIXE yield curve needed to determine trace element concentrations in hair. It can be noted that uncertainties in these values are
Anal. Chem. 1985, 57, 1079-1083
not reflected in the yield as PIXE is relatively insensitive to changes in the sample major components. This is shown in Figure 5 which displays R from eq 3 for hair, carbon, orchard leaves, and bovine liver, normalized to the hair ratio. Although these four samples have different compositions, their yield ratios are quite similar. The ICRP reference man concentrations for hair gave a yield curve lying between the orchard leaves and bovine liver lines, showing a maximum 10% change in calculated concentrations. Of course, the charred sample composition had to be measured before this comparison, which yields an estimate on the final uncertainties, could be made.
ACKNOWLEDGMENT We thank J. Goulding who performed the neutron activation analysis and J. F. Chapman who supplied the atomic spectrometry results. We also thank the operating staff of the AAEC 3 MeV Van de Graaff accelerator for their assistance, in particular L. Russell whose work on the target chamber has made measurements so much easier. We thank L. Dale for his useful advice on powder sample preparation methods. Registry No. K, 7440-09-7;Ca, 7440-70-2;Fe, 7439-89-6;Ni, 7440-02-0; Cu, 7440-50-8; Zn, 7440-66-6; Pb, 7439-92-1; Brz, 7726-95-6; Mn, 7439-96-5; Co, 7440-48-4; Cd, 7440-43-9; Zr, 7440-67-7; Se, 7782-49-2;Bi, 7440-69-9; As, 7440-38-2; Cr, 744047-3; Ti, 7440-32-6; Sr, 7440-24-6; Rb, 7440-17-7.
LITERATURE CITED (1) Cookson, J. A.; Pilling, F. D. Phys. Med. Bbl. 1975, 20, 1015-1020. (2) Houtman, J. P. W.; Bos, A.; Vis, R.; Cookson, J. A.; Tjioe, P. S. J . Radionanal. Chem. 1982, 7 0 , 191-208.
1079
Horowitz, P.; Aronson, M.; Grodzins, L.; Ryan, J.; Merriam, G.; Lechene, C. Science 1976, 194, 1162-1 165. Henley, E. C.; Kassouny, M. E.; Nelson, J. W. Science 1977, 197,
277-278.
Chen, J. X.; Guo, Y. 2.; Li, H. K.; Ren, C. G.; Tang, G. H.; Wang, X. D.; Yang, F. C.; Yao, H. Y. Nucl. Instrum. Methods 1981. 181, 269-273. Baptista, G. B.; Montenegro, E. C.; Paschoa, A. S.;Barros Leite, C. V. Nucl. Instrum. Methods 1981, 181, 263-267. Pillay, A. E.; Pelsach, M. J . Radloandl. Chem. 1981, 63, 85-95. Whitehead, N. E. Nucl. Instrum. Methods 1979, 164, 381-380. Clayton, E.; Cohen, D. D.; Duerden, P. Nucl. Insfrum. Methods 1981, 180, 541-548. Clayton, E. Nucl. Instrum. Methods 1981, l g l , 567-572. Clayton, E. Nucl. Instrum. Methods 1983, 218, 221-224. Mayer, J. W., and Rimlni, E., Eds. “Ion Beam Handbook for Material Analysis”; Academic Press: New York, 1979; p 228. Chu, W. K.; Mayer, J. W.; Nicolet, M. A. “Backscattering Spectrometry”; Academic Press: New York, 1977. Hyvonen-Dabek, M.; Riihonen, M.; Dabek, J. T. Phys. Med. Biol. 1979, 2 4 , 988-998. Chittleborough, G. Sci. Total Environ. 1980, 14, 53-75. Montenegro, E. C.; Baptista, G. B.; De Castro Faria, L. U.; Paschoa, A. S. Nucl. Instrum. Methods 1980, 168. 479-483. Campbell, J.; Faiq, S.;Gibson, R. S.;Russell, S. B.; Schulte, C. W. Anal. Chem. l W l , 53, 1249-1253. Berti, M.; Buso, G.; Colautti, P.; Moschini, G.; Stievano, B. M.; Tregnaghi, C. Anal. Chem. 1977, 49, 1313-1315. Gibbons Natrelia, M. “Experimental Statistics”; National Bureau of Standards, Washington, DC. 1963; National Bureau of Standards Handbook 91. Giadney, E. S. Anal. Chim. Acta 1980, 118, 356-396. “Report on the Task Group on Reference Man (ICRP 23)”; Internatlonal Commission on Radiological Protection; Pergamon Press: Oxford, 1974. Doyle, 0. L.; Peercy, P. S . Appl. Phys. Left. 1979, 3 4 , 811-813. Sofieid, C. J.; Bridweli, L. B.; Wright, C. J. Nucl. Instrum. Methods 1981, 191, 379-382.
RECEIVEDfor review July 13,1983. Resubmitted and accepted December 26, 1984.
Saturation Effects in Gas-Phase Photothermal Deflection Spectrophotometry George R. Long and Stephen E. Bialkowski* Department of Chemistry and Biochemistry, UMC 03, Utah State University, Logan, Utah 84322
This paper descrlbes some effects of optlcal saturatlon on a photothermal deflectlon signal and presents a slmple theory to describe these effects. These effects Increase the sensltlvlty while decreaslng the relatlve error of the method as the lntenstty exceeds the saturatlon Intensity. Detectlon lknb of 1.3 ppbv for chlorodlfluoromethane, 2 ppbv for dlchlorodlfluoromethane, and 3 ppmv for sulfur dioxide, In 13.3 kPa of argon, are found. These detectlon limits extrapolate to atmospheric detectlon llmlts of 170 pptv for chlorodlfluoromethane and 260 pptv for dlchlorodlfluoromethane. The correspondlng mass detectlon llmlts In the Infrared laser Irradlated volume are 55 fg for chlorodlfluoromethane and 70 fg for dlchlorodlfluoromethane.
Photothermal deflection spectrophotometry (PDS) is one of a class of several techniques that is used to measure small absorbances by probing the refractive index gradient created in the sample when absorption of radiation and subsequent thermal relaxation occurs ( 1 , 2 ) . PDS employs a pump laser operating on a frequency resonant with an absorption of the analyte to produce a temperature gradient in the sample. The 0003-2700/85/0357-1079$0 1.50/0
temperature gradient results in a refractive index gradient which subsequently deflects a probe laser beam. The angle of deflection is proportional to the absorbance of the sample. Theories that describe PDS have been discussed by Jackson et al. (3))and the technique has been successfully applied to a number of analytical problems ( 4 , 5 ) . One interesting fact presented in the theories that describe photothermal spectroscopy is that the observed signal is directly proportional to the energy of the excitation laser pulse, implying that high-energy excitation pulses should provide the optimal analytical sensitivity. Many commercially available lasers have energies exceeding 1 J/pulse. However, at high laser energies optical saturation of the analyte begins to occur and at this point these theories are no longer applicable. Recently we have described some effects of optical saturation in thermal lensing spectrophotometry (TLS) (6). One of these effects was the observation of a PDS signal greater than that of a TLS signal. This is a result of the fact that the PDS signal is proportional to the temperature gradient while the TLS signal is proportional to the curvature of the temperature profile (3,7). As the intensity of the pump laser exceeds the saturation intensity, the temperature profile in the sample begins to resemble a top-hat profile (8). The 0 1985 American Chemical Soclety
