this analysis (9-11). It seems probable t h a t other such examples will appear as the number of practical applications of atomic fluorescence increases. In summary, it has been shown theoretically and experimentally t h a t atomic fluorescence spectroscopy enjoys a considerable advantage over atomic absorption spectroscopy insofar as the error associated with matrix molecular absorption is concerned. Since the error values tabulated in Table I are thought t o be in the range of many analytical applications of atomic fluorescence spectroscopy, it is hoped t h a t Table I will find use with atomic fluorescence
spectroscopists for making rough estimates as t o the magnitude of the concentration error to be expected for a given degree of matrix molecular absorption interference. C . David West Department of Chemistry Occidental College Los Angeles, Calif. 90041 Received for review October 19, 1972. Accepted January 17, 1974.
I AIDS FOR ANALYTICAL CHEMISTS Simple and Inexpensive Temperature Controlled Spectrophotometric Cell Holder R. L. Wilson and J. D. Ingle, J r . l Department ofChemistry, Oregon State University. Corvallis, Ore. 9733 1
The temperature of the solutions in the sample cells of molecular absorption or fluorescence spectrometers must be carefully controlled to make precise and accurate equilibrium- or kinetics-based measurements. Temperature regulation in the range of * O . l to k0.01 "C is often required ( I ) . Most commercial spectrometer manufacturers provide controlled temperature sample cell holders as accessories and a number of authors (1-4) have described the construction of thermostatable cell holders. In this paper. the construction and performance of a new simple and inexpensive controlled temperature cell holder is discussed. This cell holder is constructed to be part of a fluorometric reaction rate monitoring system, although it is easily adapted to other applications. Temperature stability is particularly critical in fluorometric reaction rate measurements because of the significant temperature dependence both of rate constants and of fluorescence parameters such as the quantum efficiency. The temperature stability of solutions and the time required for them t o reach eyuilibrium are comparable to recently described cell holders (1-5). Compared to previous thermostatable cell holders. the described holder has the following advantages: simple and inexpensive construction, immediate capability for use in fluorescence or absorbance measurements, and ability t o remove and replace the sample cell with ease.
GENERAL COXSIDERATIONS A temperature controlled cell holder is basically a block through which is circulated water from a thermostated constant temperature bath. The cell holder must have To whom correspondence should be addressed. P. D. Feil, D . G Kubler. and D . J. Wells, Jr.. Ana/. Chem.. 41, 1908 (19 6 9 ) . Theodore Weichselbaum, Raymond E. Adams, and Harry E. Mark, Jr , A n a / . Chem.. 4 1 , 1913 (1969) Harry L. Pardue and Pedro A. Rodriguez, A n a / . Chem.. 39, 901 (1967). Paul H . Bell and C. R. Stryker. Science. 105,415 (1947). J. D . Ingle. Jr.. Ph.D. Thesis, Michigan State University. East Lansing, Mich.. 1971
provision for securely holding a spectrophotometric cell. The ability of the holder to bring solutions rapidly to a desired temperature and to maintain a constant temperature is dependent on the thermal contact between the holder and the cell and the flow rate and temperature stability of the thermostated water. Two basic approaches have been used for construct ion of spectrophotometric cell holders. In one approach. the sample cell is readily removed from the holder. while the other approach involves sealing the sample cell in the holder. The former approach allows easy cleaning of the cell and use of the cell in other applications. but often has poor thermal contact. The latter approach provides good thermal contact but does not possess the convenience of the first design approach. One variation of the first approach used by one of us (5) is basically a hollow metal block with a square well in the center for insertion from the top of a standard 1-cm square spectrophotometric cell. Appropriate windows are located in the sides of the block for absorbance or fluorescence measurements. The square well must be larger than the sample cell for easy insertion and to prevent scratching. Because of this. the cell position is not completely secure and the thermal contact between the glass cell walls and metal lilock is not good. It was also noted that the outside dimensions of standard 1-cm cells varied among different manufacturers and even among cells of the same manufacturer. Some cells w o i ~ l d not fit into the hole. while others were somewhat loose. Many of the temperature controlled holder> provided hy manufacturers of spectrophotometers are basically t h e same design discussed above, except the well in which the cell is inserted is made larger and springs are placed on the inside walls of the well. The springs securely hold cells of slightly varying outside dimensions. The disadvantage is that the contact area of the glass cell walls with the thermostatable metal block walls is considerably diminished. Recently. two temperature controlled cell holders for absorption measurements have been reported that PO,.yqess excellent characteristics. The designs involve sealing the sample cell into a block in a way that provides efficient A N A L Y T I C A L C H E M I S T R Y , VOL. 46,
NO. 6.
M A Y 1974
799
, flT;-l
lure 3. Photograph of cell holder and mounting plate d
Figure 1. Construction of cell holder
THERMISTOR
~~
T
RECORDER
o.,
YOLT&I(IE
OFFSET
3t.
SOURCE
Figure 4. Temperature measurement system 0
I I i V
0
I
O-rings and a gasket three modified 1-cm T stoppered cells in a holder with the circulating water in direct contact with the cell walls for efficient heat transfer. They reported the stability as +0.005 "C and a response time of 1.5 minutes for methanol initially 5 "C below the equilihrium bath temperature. Weichselbaum, Adams, and Mark (21, reported a cell holder in which the cell is sealed in the holder with fine copper powder in contact with the cell for efficient heat transfer. They reported a stability of better than +0.01 "C and a response time of less than 2 minutes for water initially 7 "C below the equilibrium temperature.
CONSTRUCTION OF THE THERMOSTATABLE HOLDER holder for a standard 1-cm square spectrophotometer cell is illustrated in Figures 1-3. The cell basically consists of four symmetrical corner metal blocks which are mounted to a metal plate. Figures 1 and 2 indicate the ease with which the four corner blocks are machined, Aluminum was used because it was less expensive and easier to machine than brass. For circulation of water, a hole K6 inch in diameter and inch long is drilled from the top into each corner block. Then, a %6 inch in diameter hole is drilled in the side of the corner block to meet the hole from the top. 800
ANALYTICAL CHEMISTRY, VOL. 46, NO. 6 , MAY 1974
These side holes are drilled in the left face for two of the corners and in the right face for the other two corners (Figure 2a). A hole is drilled and tapped in the bottom of each comer block for the mounting screws. The corner blocks are anodized black to minimize reflection and scattering off the cell holder. To make ports, aluminum tuhing sections (%-inch i d . , %-inch wall, and %-inch length) are epoxied into the holes. The aluminum mounting plate was prepared as shown in Figure 26. The corner blocks are mounted to the plate with screws from beneath the plate. Note that one corner block is mounted permanently while the other three corner blocks are movable. Connections are made between the four side ports and between two of the top ports of the individual corner blocks with 2-inch sections of %inch i.d., %-inch wall, Tygon tubing (see Figure 3). The inlet and outlet tubing to the water bath are connected to the two remaining unconnected top ports. Note that the resulting flow pattern allows constant circulation of water through all four corner blocks with essentially no dead volume. The sample cell is pasitioned in the holder and the three corner blocks are moved in and the screws tightened to grip the cell tightly in place. The mounting plate is. then attached in a proper position in the sample module which in our case was to the top of a motor driven magnetic stirrer (Waco Midget Model 86335) in a fluorescence sample compartment. Note that the stationary corner block provides a means to reproducibly align the cell, although the three mobile corners allow quick and easy removal of the cell. This method of providing, tight contact between the cell and the holder prevents accidental scratching of the cell during removal or placement, yet provides good thermal contact between the cell wall and each of the four corner blocks, even for cells of slightly varied sizes.
~~~
Voltage source Operational amplifier Potentiometric amplifier DC offset Recorder Thermistor
Heath, Model EU-80A Analog Devices, Model 415 Heath, Model EU-200-01 Heath, Model EU-200-02 Heath, Model EU-20-V Victory Engineering Corp, Model T35A7 Nominal resistance a t 24 "C =
4.8K
Temperature coefficient 3.8 70, ' "C
~
~~~
Table 11. T e m p e r a t u r e M e a s u r e m e n t s
T a b l e I. T e m p e r a t u r e M e a s u r e m e n t E q u i p m e n t
=
The dimensions of the cell holder provide 8.5 X 15-mm windows on each side of the cell holder. This window size was chosen so that the walls of the cell are not directly visible to a light source illuminating a window or to a photomultiplier observing fluorescence or transmitted radiation from a cell window. This is particularly critical in fluorescence work if the fluorescence or scattering of the cell walls is significant. A spectrophotometric cell magnetic stirrer (Bel-Art F37150) was placed in the sample cell. The position of the cell holder window is well above the stirring bar. An overhead stirring rod could be used although the windows of the cell holder would have to be lowered. TEMPERATURE MEASUREMENT Instrumentation. The stability, reproducibility, and response time of the temperature controlled cell were measured with the temperature measurement system shown in Figure 4 and the equipment listed in Table I. A 12-inch X 12-inch borosilicate glass jar plus a constant temperature circulator (Techne TU-12) served as the constant temperature bath. The thermostated water from the bath was circulated through the temperature controlled cell holder with a Jabsco (Model Pl-M6) Pump a t a measured flow rate of 1.5 l./min. The temperature controller has control specifications of *0.01 "C which should limit the expected stability of the temperature of the solution within the cell. A thermistor was used for temperature measurements as shown in Figure 4. Sormally temperature measurements with thermistors employ the use of conventional bridge circuits. The configuration presented in Figure 4 with the thermistor as the resistive feedback element of the operational amplifier (OA) is convenient because the output of the OA is directly proportional to the thermistor resistance. The output of the OA changed about 0.1 mV per 0.01 "C. This voltage output is amplified by a factor of ten with the potentiometric amplifier and displayed on the 10-mV scale of the recorder. The d.c. offset module was needed for suppression to allow small changes of less than 1 mV or 0.01 "C to be observed on the recorder. With a readout resolution of 0.1 mV and a peak-to-peak noise of less than 1 mV, temperature changes of 0.01 "C could easily be observed. The current that passed through the thermistor was low enough to prevent error from electrical heating. The thermistor was calibrated over a temperature range of 15-32 "C with an immersion thermometer accurate to *0.02 "C. The thermometer was read to 0.01 "C with a cathetometer. Measurement Procedure. For all measurements, the thermistor was placed in the middle of the cell and the stirrer was on. With stirring, the temperature throughout the cell varied less than 0.01 "C. Without stirring, the sta-
Response time to
Equil. temp, O C
AT, 'Ca
0.1 OC, min
15.88 20.60 25.38 30.27
2.31 2.67 5.38 7.17
0.5 2.3 2.7
a
1.6
0.01 O C , min
Reproducibility, OC
2.7 1.7 3.9 4.1
