Figure 2.
Measurements taken with sensor 1.4 cm from rotating
band. Upper curve: Output of monostable multivibrator (1 V/div]: the lower dashed line is the 1-msec ouiput. LOW curye: input to monostable (0.5 V/div)-base line is + o s v. Time base, 1 rnEec/cm: rotation rate, 5400 rpm
from the phototransistor. In normal use, much better waveforms me ohtained when the sensor is about 3 mm from the hand: however, this photograph shows more dramatically hou, the waveform is improved by the multivibrator. The spikes from the phototransistor shown in Figure 2 do not trigger the multivibrator. Good output waveforms were obtained at rotation rates from less than 0.1 to 5000 rpm. The latter speed is faster than would normally be used in electrochemical studies. At slow speeds where only a few pulses per second are observed, an event timer can be used to measure the rotation rate. At the slower rotation rates, the output of the phototransistor was not sharp enough to trigger the frequency counter. If a n event timer is to be used, then one stripe could be painted on the Eflective tape and smaller values of CT and RT (Figure l) can be to Obtain a pulse' This also be desirable for computer interfacing. If a sine wave generator and power amplifier are used to Dower a ." svnchronous motor. hichlv stahle rotation rates can be obtained. However, we have found that the system described here conveniently provides a high degree of accuracy for the value of the rotation rate. The speed controller alone provided a constant rotation rate, but it was not highly reproducible. Another advantage of this system is that the uniform pulse shape is easily interfaced with a computer so that the rotation rate could be stored with electrochemical variablesif desired. I
normal output in which a 5-V output is triggered from
a base line output is and an inverted output in which triggered from B 5-v base line. We have arbitrarily chosen to use the inverting m t to measure the frequency. -i n. .~
about
RESULTS AND DISCUSSION Figure 2 shows the resulting waveform from the sensor when the sensor is 1.4 cm from the rotating striped hand, and also the output from the multivihrator. An inverted pulse is delivered from the monostable multivibrator which is triggered on the declining portion of the signal
1
"
Received for review June 25, 1973. Accepted August 15, 1973.
Device for the Accurate Electronic Measurement of Microliter Sample Volumes L. R. Layman and G. M. Hieftje Department of Chemistry, Indiana University, Bloomington, Ind. 47401 In several analytical techniques, such a nonflame atomic absorption spectrometry, small (microliter) volumes of sample solution must be accurately measured and evaporated before a determination can be made. The measurement and transfer of these small volumes by micropipet or syringe often contributes an important fraction of the total error in the determination ( I ) . In this report, a method is described that minimizes these difficulties by indirectly measuring the mass of liquid after it is placed in the apparatus. This measurement is based on a determination of the time required to evaporate the liquid using a constant rate of heat input. To determine this time, a suitable support for the liquid (a wire filament in this case) is heated by a constant current, with changes in the support temperature monitored as a drop of sample is placed on the support and evaporates. The simple apparatus necessary for this measurement is shown in Figure 1 and consists of a 0.010-inch diameter tungsten or platinum wire filament (or other heated evaporator) enclosed in a glass tube and surrounded by a controlled atmosphere, a constant current supply (OAl and the booster amplifier), a n operational amplifier derivative circuit (OA2 and OA3), and either a digital timer or strip chart recorder. In the procedure, the filament is heated by a 1- to 2ampere constant current to a temperature below the boil-
(1)
322
C. F. Ernanuel,Anal. Chem., 45,1568 (19731
ing point of the solvent. A microliter sample placed on the warm filament then produces a rapid drop in filament temperature which is reflected in a corresponding decrease in filament resistance. This reduction in resistance is registered as a drop in voltage and converted to a negative voltage peak by the operational amplifier derivative circuit, as shown in Figure 2. As the liquid evaporates, the filament remains a t this lower temperature, allowing the voltage derivative to return to zero. When the last of the liquid has evaporated, the filament quickly returns to its original temperature, causing the filament voltage to rise and producing a positive peak in the derivative circuit. The time between the negative and positive derivative peaks is measured either from a strip chart recorder output, or directly on an electronic timer. The volume of the sample is then directly proportional to the measured time. Of course, for this proportionality to hold, the gas composition and flow rate, the current, and the nature of the solvent must be held constant. In this method, the greatest precision was obtained using an automatically triggered electronic timer. This timing system, shown in Figure 1, consists of two comparators and an electronic clock. The comparator reference voltages are set such that the first comparator triggers on a negative voltage level, while the second responds to a positive voltage. The derivative circuit output is therefore converted to fast-rising square pulses which are used to start and stop the clock.
ANALYTICAL CHEMISTRY, VOL. 46. NO. 2, FEBRUARY 1974
A Tempnature of Iha Ftlornant, ob0 Voltoge OCrO#s
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Figure 1. Schematic of drop volume measuring system showing constant current supply, heated filament, derivative circuit, and time measurement devices. Voltage waveforms are shown at several points in the circuit If a strip chart recorder is used for timing, the distance between the leading edge of the negative peak (the moment the sample is applied) and the top of the positive peak (the moment evaporation is complete) is the value directly proportional to sample volume. The measurement points are shown in Figure 2 . This method has been shown to be very accurate and linear over a wide range of volumes and under a variety of conditions. Both tungsten and platinum wire filaments have been successfully used in still and flowing air or argon atmospheres. The precision obtained with 15 trials at 1-p1 volume was 1.83% relative standard deviation, most of which could be ascribed to syringe errors. The precision of the syringe was ascertained by weighing 20 samples of 2 p1 each on a microbalance. After correction for evaporation, the relative standard deviation was 1.93%, of which the balance contributed approximately 0.1%. Therefore, the error in the apparatus presented herein is significantly less than that of the syringe. As seen in Figure 3, the linearity of the system was also well within the syringe error over the entire measured range of 0.5 to 5.0 pl. With the present filament size, samples larger than 5.0 pl could not be measured because they would not adhere to the filament; volumes smaller than 0.5 1 1 could not be accurately tested because of limitations in the reproducibility of the 5-pl syringe. This technique for the measurement of microliter samples should be applicable to a number of other evaporative systems now employed in chemical analysis, such as the rod, boat, and filament type of atomizers becoming popular in atomic absorption spectrometry. In addition, it could be applied to gas chromatography inlet systems, if modification to allow very fast evaporation were made. This measurement device is presently being used in conjunction with a microwave-excited atomic emission source, which is operated under complete computer con-
i '-Tim# Meaaured with-' Racorder
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Figure 2. Waveforms generated at the atomizing filament and by the derivative circuit. Timing points are shown in 8. ( A ) 1-pI sample size, (8)2 4 sample size
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Figure 3. Relationship between evaporation time and sample volume. Line shown is a least squares fit to the points. Hash marks above and below each point represent the f95% confidence limits for that point trol. The real time clock in the computer provides the necessary timing so that the volume of each sample is automatically and accurately determined. Thus, the entire system is automated for maximum convenience and precision in analysis of samples of, for example, clinical importance. Received for review August 13, 1973. Accepted September 19, 1973. The authors gratefully acknowledge support of this work by Public Health Services Grant GM 17904-02.
ANALYTICAL CHEMISTRY, VOL. 46, NO. 2. FEBRUARY 1974
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