Interrupted-Sweep Voltammetry for the Identification of Polychlorinated Biphenyls and Naphthalenes S. 0. Farwell,’
F. A.
Beland, and R. D. Geer
Department of Chemistry, Montana State University, Bozeman, MT 597 15
Interrupted-sweep voltammetry has been applied to the identification of polychlorinated biphenyls (PCB’s) and polychlorinated naphthalenes (PCN’s). Characteristic voltammetric “fingerprints” and reductlon potential data were obtained for 31 PCB’s and 37 PCN’s. Only two of these chlorinated compounds could not be positively identified using this technique. The theoretical basis, design, and function of the inexpensive, interrupted-sweep instrument that was developed for this investigation are described. This instrument was specifically designed for Obtaining accurate and reproducible voltammetric data of multireducible compounds exhibiting irreversible electrochemical reactions. Experimentally, reduction peaks separated by less than 60 mV were resolved, and this particular instrument has a potential resolution of 43 mV for overlapping reductlon peaks.
Presently, only relatively complex and expensive methods are available for the structural identification of such organochlorine compounds as the polychlorinated biphenyls (PCB’s) and the polychlorinated naphthalenes (PCN’s). For example, Sissons and Welti ( I ) have characterized the major constituent PCB’s in Aroclor 1254 by 220 MHz nuclear magnetic resonance spectroscopy and gas chromatographic retention indices; Webb and McCall (2) have identified 27 P C B positional isomers in various commercial Aroclors by matching their gas chromatographic retention times and infrared spectra with known P C B compounds, and Safe et al. (3) have demonstrated t h a t the ion kinetic energy (IKE) spectra of the P C B isomers are different and may be of some use in P C B analysis. Although normal electron-impact mass spect,rometry can be used as a method for classification in chlorinated hydrocarbon analysis, it cannot distinguish between most P C B or P C N positional isomers because randomization of chlorine atoms occurs prior to fragmentation 14). Therefore, no simple procedure was available for identifying the individual isomers of the PCB’s, PCN’s, and other polychlorinated hydrocarbons in the environment. In our laboratory, we are particularly interested in the systematic identification of environmental pollutants, and it occurred to us t h a t voltammetry might prove useful for the qualitative identification of polychlorinated hydrocarbons ( 5 ) . Polyhaloaromatics are known to be irreversibly reduced a t a mercury electrode, usually in a stepwise manner (6-8), and it also seemed likely t h a t the voltammetric reduction potentials of the various carbon-chlorine bonds would depend upon their structural positions on the parent aromatic molecule. This last hypothesis was partially suggested by the results of Zavada et al. ( 9 ) , who showed t h a t polarography could be used in the determination of the stereochemistry and structure of dibromo compounds. Our preliminary work also indicated that voltammetry could be used to identify certain chlorinated hydrocarbons based To whom all correspondence should be sent. Present address, Air Pollution Research, Engineering Research Division, Washington State University, F’ullman, WA 99163
upon determining both the number of chlorines and their structural positions from their voltammetric reduction patterns (5, 10). Since polychlorinated hydrocarbons are reduced stepwise, it is essential to employ a voltammetric method that will minimize the interference due to overlapping reduction peaks in order to obtain accurate and reproducible reduction data for these compounds. A normal voltammetric curve for the noncomplex reduction of an electroactive substance a t a stationary electrode reaches a maximum peak height, i,, after which the current decreases as a function of l / f i ( w h e r e t is time) as the cell potential is continuously increased. As discussed by Perone et al. ( I I ) , this “current tailing” severely complicates the polarographic analysis of mixtures of electroactive species because the reduction tail of one component may distort or conceal the next reduction signal of another constituent. Likewise, “current tailing” can be a problem in the voltammetric identification and analysis of multireducible polyhalogenated compounds. Perone et al. (11, 12) have successfully demonstrated the effectiveness of interrupted-sweep stationary electrode polarography for the analysis of several inorganic mixtures. They first employed an on-line minicomputer and subsequently a hardware device to obtain quantitative resolution of overlapping reduction waves where the peak potentials were separated by a t least 150 mV. Since then, Gutknecht and Perone (13)have improved the resolution of waves for inorganic mixtures to 40 mV by using a minicomputer for numerical deconvolution of overlapping polarographic curves. Until now, there has been no evaluation of the interrupted-sweep technique for the resolution of irreversible voltammetric waves for multireducible compounds. This paper demonstrates the utility of interrupted-sweep voltammetry for the identification of PCB’s and PCN’s by characteristic current-voltage patterns (or “fingerprints”) and interrupt-potential data. The inexpensive instrumentation developed for this identification technique is particularly advantageous in obtaining rapid and reproducible voltammetric data when the chlorinated hydrocarbon reduction waves overlap. The theoretical basis, design, and operation of this instrument, in addition to its “fingerprint” capability, are reported.
EXPERIMENTAL Cell and Electrodes. The electrochemical cell used was the Sargent S-29405 small-volume H-form cell. The stationary electrode was a mercury-coated platinum (Hg-Pt) electrode with a working surface of 42 mm2 and was prepared according to the procedure described by Ramaley et al. (14). Between samples, the Hg-Pt electrode was rinsed with double-distilled water and thoroughly cleaned with concentrated nitric acid before being replated with fresh, triple-distilled mercury. The counter electrode was a 20-gauge, coiled platinum wire and the reference electrode was a Sargent S-30080-25saturated calomel electrode (SCE). Reagents. Reagent-grade dimethylsulfoxide (DMSO) from the J. T. Baker Company was used as the solvent, with 0.1 M tetraethylammonium bromide (TEABr) (reagent grade, J. T. Baker Co.) as the supporting electrolyte. The cell temperature was maintained for all experiments at 26.0 f 0.1 O C . All solutions were deaerated ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, MAY 1975
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operational amplifier, and by the values of the integrator input resistor and/or feedback capacitor. A conventional voltage divider with a 20-turn potentiometer for selection of the output voltage is 1 connected to a +5 V regulated power supply for the ramp rate control of Figure 1. When the integrator input resistance is 1 MR and the feedback capacitance is 5 pF, an input voltage of +0.825 V gives the 142 mV per second sweep rate used in this work. The ramp output from the integrator is controlled by S-1,which is one of the four switches available in the RCA CD4016A integrated circuit quad bilateral switch, and by Q4, which is a Motorola 2N5464 P-channel field effect transistor (FET). The excellent switching characteristics of S-1 and Q4 are such that, during the interrupt or hold operation, the cell potential is limited to a maximum change of 1 mV per minute. The appropriate manipulation of S-1 and Q4 generates the waveform used in this study and is discussed in the following paragraphs. The momentary-contact reset switch in Figure 1 is used to reset flip-flops F1, F2, F3, and F4. This reset operation returns control of the ramp generator to the master control switch and/or S-1. 1 I Flip-flops F1 through F4 are units in the RCA CD4027A dual J-K %5flip-flops with setheset capability. The master control switch provides for manual control of the ramp generator. When the master Figure 1. Schematic diagram of interrupted-sweep instrumentation control switch goes from +15 V (logical 1 or high) to ground (logiCircuit components: comparators 1, 2, 3 pA710C (Fairchild);Q1, Q2, Q3, cal 0 or low), the output of G4 must go to a logical 1, thereby setting the Q output of F4 to a logical 1. A high at the gate terminal of 2N2368 (Motorola); Q4. 2N5464 (Motorola); integrator, Zeltex 148; C1, Q4 turns off the FET and allows the integrator to be controlled by 0.001, 0.01, 1.0, 1.5,5.0 pF; C2, C3, 1.0 pF; C4, C5, 0.1 pF; F1, F2, F3, F4, S-1. When the output of NAND gate G5 is high, the control input CD4027A (RCA);S-1.S-2. 5-3, CD4016A (RCA);G1, G2, G3, G4, G5, G11. of analog switch S-1 is also high. Thus, S-1 is now “ON” (or effecCD4011A (RCA); G6, G7, G9, G10, CD4001A (RCA); G8, G12, CD4002A tively closed) and connects a reference voltage (+0.825 V) to the (RCA); R1. R5, R9. 10 kR, 5%; R3. R 7 , R11, 330R2,5%; R 2 , R6, R10, 22 integrator input, which subsequently starts the voltage ramp. To kR, 5%; R 4 , R8. R12, 10 kR, 20-turn potentiometer;R13, R14, R15. 1 MR, interrupt or hold the ramp at a constant voltage, G5 must go to a 20-turn potentiometer; Master Control Switch, SPDT; Reset Swtich, SPST, logical 0, which turns S-1 “OFF” (or effectively open). Typical momentary contact “ON” resistance for the CD4016A switch is 300R and typical “OFF” resistance is 10’2R. A new C-MOS quad bilateral switch, with high-purity nitrogen that was further purified by passage the CD4066A, which is pin-for-pin replaceable for the C4016A and through two gas-washing bottles containing vanadous chloride soexhibits a typical “ON” resistance of only l O O R is now available lutions and then through a third gas-bubbler containing DMSO. from RCA (19). All solution concentrations were 5 X lo-* M in the electroactive Differential voltage comparators 1, 2, and 3 are Fairchild pA710 species and a sample volume of 3 ml was used. integrated circuits. It was necessary to use external positive feedThe naphthalene was reagent grade from Allied Chemical and back to produce hysteresis in the transfer characteristics of these Dye Corporation. The biphenyl and the two monochlorocomparators (20). The npn switching transistors 91, Q2, and Q3 naphthalenes were Baker grade from the 3. T. Baker Company. are used for logic level interfacing between the pA710 comparator and 2,3,4,5,6-pentachlorobiphenyl outputs and the inputs of the C-MOS J-K flip-flops. Since conThe 2,3,5,6-tetrachlorobiphenyl were 99% pure compounds from Analabs, Incorporated. The 2,3,6struction of this instrument, new types of comparators have betrichlorobiphenyl was synthesized by the procedure described by come commercially available which would be more compatible Tas and Kleipool (15). Decachlorobiphenyl was prepared by the with the power requirements and logic levels of the C-MOS logic method of Hutzinger et al. (16). The remaining chlorinated bifamily (21, 22). phenyls, in addition to the 1,8-, 2,3-, and 1,2-dichloroThe inverting input of comparator 1 is connected to the secondnaphthalenes, were synthesized as we have described elsewhere (5, derivative signal of the electrochemical cell current, and the strobe 17). The rest of the PCN’s were obtained from Julius Hyman, is maintained at +5 V to enable this comparator. A 20-turn potenBerkeley, CA; Imperial Chemical Industries, Ltd., England; and tiometer, R4, is used to set the threshold voltage for comparator 1. Dusan Hadzi and Ludvik Cencelj, University Institute of Boris KiSince the second derivative signal is quite sensitive to minor flucdrie, Ljubljana, Yugoslavia, to whom we are indebted. The identity tuations in the cell current, which are due to electrical noise or of the PCB and PCN compounds was confirmed by mass spectrace contaminants in the cell, it was found that false triggering of trometry on a Varian CH5 mass spectrometer and by melting comparator 1 was avoided by setting its threshold potential at +80 point comparisons when the corresponding literature values were mV. Under normal conditions, the second derivative of the cell available. The purity of each chlorinated compound was detercurrent is changing so rapidly as it passes through zero when demined to be at least 95% by gas chromatography. Two chromatotecting a reduction peak that this slight offset from zero volts has a graphs were employed in this purity analysis: a Varian Aerograph negligible effect on the expected interrupt potential. When the Series 1200 with a tritium electron-capture detector and an F&M positive-going second-derivative signal, which is inverted because Model 400 with a flame ionization detector. Three different 10-ft of the differentiation network (18),passes through zero and crossx lh-in. silanized glass columns were used: a 10% Carbowax 20M es the threshold potential, the output of comparator 1 goes from on 60/80-mesh Chromosorb W, a mixed 10% OV-101/5% Bentone high to low. Transistor Q1 inverts this logic transition and toggles 34 on 100/120-mesh Supelcoport, and a 3% OV-101 on 100/120Q of F1 from low to high. During the ramp cycle of the instrument, mesh Chromosorb W. the input of G5 that is connected to Q of F1 is low and the input of G5 that is connected to Q of F2 is high. When Q of F1 goes from INSTRUMENTATION low to high, then both inputs to the gate G5 are high and the output of G5 is forced from high to low. This low at the control input A schematic diagram of the logic control and ramp generator cirof C-MOS switch S - 1 causes the interrupt, or hold, in the ramp cuits is presented in Figure 1. The potentiostat, integrator, difvoltage. ferentiator, and current measurement circuits are the same as preIn addition to producing the interrupt in the sweep, the low-toviously described (18).The initial cell potential is set by a potentihigh transition in Q of F1 provides two other functions. First, when ometer located in the potentiostat module. The f 1 5 V power the F1 input to G1 goes high, the output of G1 goes low and comsupplies required for the circuit were constructed on separate pels the output at inverter G2 to go high. This high output at G2 printed circuit boards from Fairchild pA723 voltage regulators closes S-3, which connects the current-to-voltage converter of the with RCA 40389 and 40410 external pass transistors. All of the digpotentiostat to a Heath EU-800-HB V/F card. The output freital-logic integrated circuits used in the construction of the interquency of the VIF card is summed by a Heath EU-805-AA Univerrupted-sweep instrument were complementary-symmetry metal sal Digital Instrument operating in the events-counter mode. Secoxide semiconductors (C-MOS). ond, the Q transition of F1 closes S-2. The closure of S-2 applies The ramp rate is determined by the voltage applied to the input +5 V rather than ground to the strobe of comparator 2, thereby of the integrator, which is a Zeltex Model 148 chopper-stabilized 896
ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, MAY 1975
enabling comparator 2. The cell current during the hold operation is monitored by this comparator and when the voltage analog of the cell current falls below the voltage selected by potentiometer R8, in this case +0.500 V, the output of comparator 2 changes from high to low. Transistor $2 inverts this transition and toggles F2. The resulting high-to-low transition in Q of F2 is responsible for three actions. First, it places a low at the input to control NAND gate G5. The output of G5 then goes high and subsequently closes S-1. The closure of S-1 terminates the hold operation and the ramp continues on to locate the next reduction peak or, in the case of no more peaks, the cathodic sweep limit. Second, the Q transition of F2 sets a low on the input to G1, thus forcing S-3 to open. When S-3 opens, a ground is placed at the V/F input, which stops the oscillator and the events counter. The accumulated count on the counter is proportional to the integrated cell current during the hold on a particular peak. Third, the Q transition of F2 triggers a monostable multivibrator composed of NOR gates G6, G7, G9 and G10 (23). This portion of the circuit could be simplified by using the RCA CD4047A monostable multivibrator unit (24), which is now available. The monostable multivibrator is necessary to reset and disable F1 during the restarting of the ramp and to reset F2. If F1 is not disabled during this restart, the second-derivative charging spike associated with this abrupt change in the ramp rate will erroneously toggle comparator 1, which in turn will interrupt the ramp. The monostable delay time, T D ,was 0.3 sec; and, since the sweep rate was 142 mV per sec, a T Dof 0.3 sec produces a “dead zone’’ of approximately 43 mV. During this T D ,the voltage ramp cannot be stopped even if a reduction peak is detected by the instrument. Therefore, this 43-mV “dead zone” established the minimum possible peak resolution for this instrument. The final potential at which the voltage sweep is terminated and returned to the initial cell potential is set by the threshold voltage applied to comparator 3. This comparator output changes from high to low when the cell potential reaches this threshold voltage limit. Transistor Q3 inverts this logic transition and toggles F3, thereby changing the Q output of F3 from low to high, which in turn initiates the following actions. The Q output of F4 is reset to a low, causing the FET $4 to short capacitor C1 of the integrator. In addition, the 4 output of F2 is reset to a high via gates G8 and G3. The Q output of F1 is also set high and this action has the same function as when F1 was toggled by the second-derivative signal, which has already been described. At this point, pressing the reset switch will return control to the master control switch for the next run. In addition to the interrupted-sweep mode, this instrument can be converted readily to a single-cycle sawtooth waveform generator for normal polarographic investigations by disconnecting the second-derivative signal from the input of comparator 1. The circuit of Figure 1 was constructed on modular printed circuit boards. A Heath EU-805AA digital voltmeter was used to display and read out the interrupt or Ezd voltages to four significant figures. A Tektronix 564 storage oscilloscope with a Tektronix C-27 camera system was used to record the analog displays of the voltammetric curves.
RESULTS A N D D I S C U S S I O N We have previously described a signal generator (18, 25) that has essentially t h e same voltage ramp and interrupted-sweep capabilities as one recently reported by Pool e t al. (26). However, our original programmable waveform generator was comparatively inefficient when it was programmed for ramp-and-hold waveforms, since t h e operator had t o preset t h e hold potentials and t h e durations of t h e hold times. Consequently, t h e original waveform generator proved t o be inadequate for our electrochemical studies of chlorinated compounds and was replaced by t h e interrupted-sweep instrument described in this paper. This interrupted-sweep instrument is similar in one respect t o Perone’s (11, 12); i t provides a voltage ramp that is controlled by t h e sample and therefore does not require t h e operator t o preset hold potentials and hold times. However, t h e logic schemes used in sensing t h e hold potentials (E2d) a n d controlling t h e hold times are different from those of Perone’s instruments. T h e C-MOS logic circuitry used in our instrument offers several advantageous features (e.g., excellent noise immunity, low power dissipation, longer rise
a n d fall times, compatibility with analog signal levels, etc.) that make them very suitable for use in chemical instrumentation (24). Theory. Qualitatively, t h e theory behind t h e design of our technique is similar t o that presented by Perone a n d coworkers (11, 12) for their computer-controlled a n d hardware-controlled interrupt, fast-sweep derivative polarography. They showed that interrupted-sweep polarography a t a stationary electrode may be used t o resolve a n d quantify reversible electrochemical reduction waves for mixtures of electroactive inorganic cations. Their technique should also work for irreversible electrochemical processes and with multireducible compounds. For reversible systems, t h e procedure requires holding t h e cell potential at values sufficiently cathodic of E112 t o ensure “pseudocomplete” depletion of t h e oxidized form of t h e electroactive species around t h e electrode. This has limited t h e resolution of adjacent waves for 2 e- reductions t o about 150 mV, including t h e “dead zone” t h a t is d u e to t h e charging spike from the restart of t h e voltage sweep after a hold operation. However, with irreversible waves, a potential held at a n y point on t h e wave where t h e electron transfer process is much faster than t h e diffusion of t h e electroactive species will cause its depletion around t h e stationary working electrode. Under these latter conditions, t h e diffusion current, id, will decrease according t o t h e Cottrell equation (27) until nondiffusion processes become significant, which may be about 30 seconds in some cases (28). Since t h e voltage sweep can be interrupted earlier for irreversible waves, t h e resolution of adjacent irreversible waves should be improved over that for reversible systems a n d may be limited by the “dead zone” caused by t h e sweep-restart current spike. This problem of t h e “dead zone” limitation on t h e resolution of overlapping reduction waves could be eliminated by first backing u p t h e sweep voltage sufficiently so that t h e restart “dead zone” is over before a new portion of t h e voltammogram is scanned. I n this case, t h e waveform applied t o t h e cell would be described by ramp/hold/step back/ramp/etc. T h e first zero-crossing of t h e second derivative of t h e cell current was chosen as t h e interrupt potential (E2d) on t h e reduction wave, since i t is readily determined experimentally a n d is sufficiently anodic of t h e peak potential (E,) t o provide good resolution between waves. T h e quantitative relationships between E2d, E,, Ep/2,a n d Eo may be established by reference t o t h e current function d a t a a n d equations given in t h e theoretical paper of Nicholson and Shain (29). For example, by plotting t h e second derivative of their current function v%x(bt) us. potential for irreversible charge transfer (Case 11), a value of 33.25 mV is obtained for t h e first zero crossing of t h e second derivative on their potential scale. This value may be used t o express E2d in terms of Eo a n d other electrochemical parameters as defined in t h e paper by Nicholson and Shain (29); Le.,
+
( R T / F ) In m / k , = 33.25 mV (1) which can be rearranged t o obtain (E2d - EO)crti,
Eed = E o - (RT/cuiz,F)(-0.729
+
In
-
In k,)
(2) where b = an,Fu/RT; a is t h e transfer coefficient; n, is t h e number of electrons transferred in t h e rate-limiting step; DO is t h e diffusion coefficient of t h e reducible species; u is t h e rate of potential scan; k , is t h e rate constant; a n d F, R, and T have their usual significance. Equation 2 for E2d may be compared with t h e equivalent equations derived for E , a n d Ep/2by Nicholson a n d Shain where E , = -5.34 mV and Ep/2 = 42.36 mV: ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, M A Y 1975
897
E P = E o - ( R T / a n 8 ( 0 . 7 8 1 + In $%v and
E P l 2 = E o - (RT/mzaF)(-1.08
+
In
- In k,)
vm
(3)
- In k,)
(4) Thus,
Ep - Ep/Z = -1.86(RT/~~tz,F) =
-47.7 mV CY t 2 ,
(5)
and
For this case of a totally irreversible reduction, theoretical Equations 5 and 6 can be reduced to:
Therefore, the E 2 d value is 9.0 mV/cYn, more cathodic than
E,/% The second derivative interrupt potential is also a function of the rate of potential scan (v)
(E2d)2 - (E2d)l = ( R T / a a F )1nUj/U2
(8)
and thus, just as for the peak potential and the half-peak potential, there is a cathodic shift of about 30/an, mV for each tenfold increase in the potential scan rate. The other cases presented by Nicholson and Shain (29); e.g., reversible charge transfer and chemical reactions coupled to the charge transfer reaction, can be treated in the same manner to relate E 2 d to E,, E p / 2 , and E o for those particular cases. The equation for the diffusion current a t the second-derivative interrupt potential can be derived for Case I1 from the value of the current function a t the interrupt potential and the relation
i = ~~~~~*>,.;rD,bx(ht)
(9)
where Co* is the bulk concentration of the reducible species, A is the electrode area, and the other parameters have been defined previously. For Case 11, the diffusion current equation a t the interrupt potential is:
Figure 2. Voltammograms of 1,2,3,4-tetrachloronaphthalene Voltage range: -0.700 V to -2.600 V vs. SCE Upper trace 2A: Normal 0.5 V/div and 1000 pA/full scale. Middle trace 28: Second-derivative scan at 1 0 V/div. and 1000 pA/full scale. Lower trace 2C: Interruptedsweep fingerprint at 0.5 V/div and 1000 pA/full scale
iPd = 6.41 x I O - ~ Z F A C ~ * ~ ~ D ~ D(~1 0Z) , U scan at Equation 10 shows that, analogous to the peak current (ip), the diffusion current a t the start of the voltage interrupt should be directly proportional to the bulk concentration of the reducible species (C,*) and could be used for quantitative analysis. However, there are several problems which arise with multireducible compounds that limit the utilization of the quantitative aspects of the interrupt diffusion current. In the case of severely overlapping waves, the initial base of the second wave may contribute to the reduction current of the first wave, thereby causing i 2 d of the first wave to be proportionally greater than i2d of the second wave. In addition, since our instrument requires a current-limit setting to restart the voltage sweep, there is a current pedestal after the initial wave which includes both the residual diffusion current of preceding waves and the background current. This latter characteristic increases the i z d of all reduction waves except the first wave. For identification purposes the qualitative aspects of the various i 2 d assist the recorded E 2 d data in producing a characteristic fingerprint. Analytical Data. The application of voltammetry to the identification of aromatic chlorine compounds has been 898
ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, M A Y 1975
quite limited (8). Although the polarographic reductions of various halogenated compounds have been investigated and reviewed (6, 7, 30, 3 1 ) , little of this prior work has been done systematically on individual groups of chlorine compounds. Illustrative reduction voltammograms of 1,2,3,4-PCN and 1,2,3,5,7-PCN are shown in Figure 2 and Figure 3, respectively. The most cathodic peak in Figures 2 and 3 corresponds to the reduction of naphthalene and the remaining reduction peaks represent the stepwise removal of the chlorines. Since the reduction signal for the removal of the first chlorine may distort the succeeding reduction peak, as illustrated in the normal single-sweep voltammograms in Figures 2A and 3A, the voltage ramp must be stopped a t a potential cathodic enough to deplete the more easily reducible chlorinated species from the solution layer surrounding the working electrode. The zero-crossing of the second-derivative curve (Figure 2 B ) controls the E 2 d interrupt voltages for the fingerprint pattern (Figure 2C). Thus, a series of ramps and holds are required t o examine a polychlorinated compound.
Figure 4. Interrupted-sweep fingerprint of 2,3,5,6-tetrachlorobiphenyl Voltage range: -0.700 V to -2.600 V vs. SCE. Sensitivity: 0.5 V/div. and 1000 KA/full scale
Figure 3.Voltammograms of 1,2,3,5,7-pentachloronaphthalene Voltage range: -0.700 V to -2.600 V vs. SCE. Upper trace 3A: Normal scan at 0.65 V/div. and 1000 wA/full scale. Lower trace 38: Interruptedsweep fingerprint at 0.5 V/div. and 1000 FA/full scale
In order to use the ramp-and-hold voltammetric data for qualitative identification, the experimental E 2 d values must be reproducible. Table I lists the E z d data and the corresponding standard deviations for naphthalene and 37 chlorinated naphthalenes. The data summarized in Table I were collected by performing a number of runs over a period of several days. A 20-mV variation in the day-to-day experimental values of E 2 d was observed and the source of this variation was finally found to be due to fluctuations in the contact resistance of a switch on the potentiostat. However, each reduction peak varied by the same magnitude and the relative difference in volts between the individual E 2 d potentials (reported as a 2 d ) was constant. Furthermore, all of the P C N voltammograms contain the reduction peak for naphthalene ( E 2 d of -2.326 V vs. SCE) which can be used as an internal reduction standard. T h e E 2 d data for the P C N were corrected by employing the internal reduction standard of naphthalene. T h e precision data in Table I show t h a t the E 2 d values are very reproducible (within &5 mV) and, except for 2,6P C N and 2,7-PCN, the EZd fingerprints are distinctive for the individual PCN’s and can be used for identification. Notice t h a t the S z d for 1,2,7-PCN is 57 mV, which approaches the calculated resolution limit of 43 mV for this interrupted-sweep instrument. The first reduction peak for naphthalene a t -2.197 V did not appear in any of the P C N interrupted-sweep voltammograms; however, inflections on several of the more sensitive second-derivative voltammograms suggested the presence of a trace of this electroactive species. T h e reason for the unanticipated disappearance of this naphthalene peak is not known, but is being investigated. Hoijtink and Van Schooten (32) have calculated that naphthalene should show two 1-electron reduction waves with Ellz’s t h a t are separated by approximately 100 mV, and this value com-
pares favorably to our -2d of 129 mV. Von Stackelberg and Stracke (6) report the only previous polarographic results for a chloronaphthalene. They published Ellz’s for 1ch1oronapht’:alene of -2.10 V and -2.38 V vs. SCE in a solvent system of 0.05M TEABr in 75% dioxane, which indicates t h a t they also did not see two reduction peaks for naphthalene. Figure 4 shows an interrupted-scan fingerprint for 2,3,5,6-PCB, which is a representative PCB voltammogram, and Table I1 lists the E 2 d data for biphenyl, all the PCB’s with chlorines on one ring, and some of the PCB’s with chlorines on both rings. In a manner analogous to the PCN’s, the PCB’s E 2 d values can be corrected to an internal reduction standard, which in this case is the biphenyl E 2 d of -2.411 V vs. SCE. T h e E 2 d for the PCB’s were also very reproducible and, with the exceptions of 2,5-PCB and 2,3,6-PCB, the E 2 d values are characteristic for each PCB. However, in the case of 2,5-PCB and 2,3,6-PCB, the relative magnitude of the i2d for the first reduction of 2,3,6PCB a t -1.942 V is approximately twice the magnitude of the ipd for the first reduction of 2,5-PCB a t -1.936 V. Thus, the use of the qualitative i2d values permits identification in this instance when the E 2 d are essentially indistinguishable, and is an example showing the benefit of recording both the E 2 d and i2d values. The scientific literature contains no previous reports concerning the reduction of chlorinated biphenyls, but the oxidative electrochemistry of various PCB’s currently is being investigated by S t u art et al. (33). Because the reported interrupt voltages for the PCB’s and PCN’s are referenced to the final reduction waves for biphenyl and naphthalene, respectively, serious shifts in the E 2 d values were not expected to appear with changes in their concentration. This expectation was maintained over a 100-fold change in concentration, from 10-3M to 10-jM. An apparent anodic shift in the interrupt potentials relative to the last redaction wave was observed for concentrations of reducible species less than 10-5M and may be due to a cathodic shift in the last wave caused by the higher sensitivity used a t these lower concentrations and the relatively greater contribution from electrolyte decomposition. An apparent cathodic shift was observed a t concentrations greater than 10-3M and may be due to the characteristics of the differentiator circuits when the rate of current change exceeds their design limit and they start to integrate the signal. This action will cause a delay in their response to the inflection point in’the diffusion current signal. Since the dimethylsulfoxide used as the solvent was taken directly from the reagent bottle and is quite hygroscopic, the effect of water on E 2 d was investigated. Water ANALYTICAL CHEMISTRY, VOL.
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Table I. Reduction Potentials for the Chlorinated Naphthalenes No. Sample
No.
Corrected, mean
of
EZd vs. SCE, V
runs
Std dev,V
SCE, V
A E z d vs.
Naphthalene
-2.197 -2.326
50
k0.003 *0.001
-0.129
1-
-1.940 -2.326
15
k0.003 *0.001
-0.386
2-
-1.975 -2.326
18
*0.002 *0.001
-0.351
1,2-
-1.726 -1.957 -2.326
10
*0.002 *0.001 *0.001
-0.231 -0.369
-1.752 -1.965 -2.326
10
1,4-
-1.751 -1.944 -2.326
1,5-
1,6-
1,3-
10.001 *0.001 10.001
-0.213 -0.361
11
*0.001 *0.001 *0.004
-0.193 -0.382
-1.765 -1.933 -2.326
10
*0.001 1.0.001 10.002
-0.168 -0.393
-1.802 -1.948 -2.326
10
*0.001 10.001 10.001
-0.146 -0.378
-1.793 -1.944 -2.326
10
*0.001 rO.OO1 *0.001
-0.151 -0.382
1,8-
-1.704 -1.938 -2.326
10
10.001 *0.001 10.001
-0.234 -0.388
2,3-
-1.769 -1.960 -2.326
10
+0.004 +0.003 *0.002
-0.191 -0.366
2,6-
-1.844 -1.957 -2.326
10
*0.003 *0.001 *0.001
-0.113 -0.369
2,7-
-1.838 -1.959 -2.326
10
10.001 10.001 *0.001
-0,121 -0.367
1,2,3-
-1.554 -1.761 -1.975 -2.326
10
*0.002 *0.001 rO.OO1 *0.002
-0.207 -0.214 -0.351
-1.565 -1.752 -1.939 -2.326
12
*0.001 10.001 10.001 10.001
-0.187 -0.187 -0.387
-1.581 -1.783 -1.942 -2.326
10
*0.002 *0.001 *0.001 *0.001
-0.202 -0.159 -0.384
-1.620 -1.827 -1.955 -2.326
10
*0.001 *0.001 *0.001 10.001
-0.207 -0.128 -0.371
-1.613 -1.898 -1.955 -2.326
9
*0.002 *0.001 1-0.004 *0.001
-0.285 -0.057 -0.371
1.7-
1,2,4-
1,2,5-
1,2,6-
1,2,7-
900
Sample
of
EZd VS. SCE, V
rum
Std dev, V
SCE, V
A E Z d vs.
1,2,8-
-1.512 -1.785 -1.941 -2.326
10
*0.001 *0.001 10.001 10.001
-0.273 -0.156 -0.385
1,3,5-
-1.578 -1.781 -1.938 -2.326
9
10.002 *0.002 *0.001 10.002
-0.203 -0.157 4.388
1,3,6-
-1.634 -1.824 -1.958 -2.326
12
*0.001 10.002
-1.635 -1.853 -1.974 -2,326
10
1,3,8-
-1.541 -1.806 -1.960 -2.326
10
10.002 10.002 *0.001 *0.001
-0.265 -0.154 -0.366
1,4,5-
-1.540 -1.758 -1.932 -2.326
10
k0.004 *0.001 *0.001 *0.001
-0.218 -0.174 -0.394
.1,4,6-
-1.618 -1.776 -1.941 -2.326
12
*0.001 -0.004 *0.001 *0.001
-0.158 -0.165 -0.385
1,6,7-
-1.599 -1.778 -1.938 -2.326
10
*0.002 *0.001 *0.003 *0.001
-0.179 -0.160 -0.388
2,3,6-
-1.657 -1.820 -1.956 -2.326
15
*0.001 *0.001 *0.001 *0.002
-0.163 -0.136 -0.370
1,2,3,4-
-1.393 -1.563 -1.751 -1.946 -2.326
10
1,2,3,5-
-1.411 -1.591 -1.784 -1.948 -2.326
10
*0.002 *0.002 *0.001 *0.001 *0.001
1,2,3,7-
-1.445 -1.627 -1.832 -1.959 -2.326 -1.445 -1.609 -1.783 -1,954 -2.326
10
10.001 10.001 *0.001 *0.001 *0.001 10.001 *0.001 10.002 10.001 *0.001
1,3,7-
ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, MAY 1975
Corrected,mean
1,2,4,6-
*0.001 *0.001
*o.ooo *0.002 *0.001 10.002
*0.001
*o.ooo *o.ooo *0.001 10.001
10
-0.190 -0.134 -0.368 -0.218 -0.121 -0.352
-0.170 -0.188 -0.195 -0.380 -0.180 -0.193 -0.164 -0.378
-0.182 -0.205 -0.127 -0.367 -0.164 -0.174 -0.171 -0.372
Table I. (Continued) Sample
1,3,5,7-
1,3,5,8-
1,3,6,7-
1,4,5,8-
No. of runs
Std dev,V
ABZd vs. SCE,V
-1.444 -1.615 -1.836 -1.960 -2.326
10
*0.001
-0.171
-1.373 -1.616 -1.783 -1.946 -2.326 -1.490 -1.607 -1.782 -1.955 -2.326
10
-1.345 -1.535 -1.755 -1.929 -2.326
10
Corrected,mean VI. SCE, V
io*oo1
*O*Ool
*o*ool io.001
*o.ool
*o'ool
*0.001
10
1,2,3,5,7-0.167 -0.163 -0.380
*0.001
-o.117
*o'ool
-0.175 -0,173 -0.371
*O*Oo2 10.001 *0.002 *0.001
*o.oo1 io.001 *O*Oo3 *0.002
1,4,6,7-
-0.221 -0.124 -0.366
10.001
*o'oo2
Sample
1,2,3,4,5,6,7,8-
-o.190 -0.220 -0.174 -0,397
No. of runs
Std dev,V
&EZd vs. SCE,V
-1.431 -1.576 -1.729 -1.957 -2.326
10
*0.001
-o.145
-1.340 -1.506 -1.639 -1.833 -1.985 -2.326
14
-0.940 -1.081 -1.298 -1.411 -1.600 -1.706 -1.840 -2.028 -2.326
30
Corrected, m e a n EZd vs. SCE, V
*o*oo2 *O*Oo3 *0'004 *0.002
-0.153 -0.228 -0.369
*0.003
-o.166
*o'ool
-0,133 -0.194 -0.152 -0.341
io.001 *0.001 *O'Oo3 *0.003 10.002 *O.OO'
*o.002 *0*004 *0*009 *0.011 *0'004 10.007
-0.141 -0.2 17 -0.11 3 -0.189 -0.106 -0.134 -0.188 -0.298
Table 11. Reduction Potentials for the Chlorinated Biphenyls Sample
Corrected, m e a n
30. of
V E Z ~ V SSCE, .
W
S
NO.
Std dev,V
-2.41 1
80
*0.001
2-
-2.097 -2,411
25
*0.002 10.002
3-
-2.108 -2.41 1
15
h0.003 *0.002
4-
-2.056 -2.41 1
15
10.003
-1.956 -2.103 -2.411
15
2,4-
-1.983 -2.070 -2.41 1
15
10.002 io.001 *0.001
-0.087 -0.341
2,5-
-1.942 -2.099 -2.41 1
15
*0.001 10.003 *0.002
-0.157 -0.312
2,6-
-2.107 -2.41 1
15
k0.003 *o, 002
-0.304
3,4-
-1.871 -2.107 -2.411
15
io.001 *0.002 *0.001
-0.236 -0.304
3,5-
-1.897 -2.095 -2.411
15
k0.003
-1.852 -2.091 -2.41 1
26
-1.783 -1.891 -2.090 -2.41 1
15
2,3,4-
2,3,5-
of
E ~ ~ vSCE, s .V
runs
Std dev, V
-1.937 -2.091 -2.411
15
*0.001
-1.837 -1.955 -2.106 -2.411
15
io.001
*O .003
2,4,6-
-!.966 -2.070 -2.41 1
15
i0.002 *0.002 i0.002
-0.104 -0.341
10.003 10.004
3,4,5-
-1.696 -1.894 -2.092 -2.411
15
io.001 *0.001 10.001 *0.002
-0.198 -0.198 -0.319
2,3,4,5-
-1.679 -1.819 -1.913 -2.096 -2.41 1
39
*0.001 10.003 r0.002 to.001 10.001
-0.140 -0.094 -0.183 -0.315
2,3,4,6-
-1.784 -1.944 -2.089 -2.411
25
10.003 10.002 10.003 10.002
-0.160 -0.145 -0.322
*0.001
2,3,5,6-
-1.787 -1.903 -2.098 -2.41 1
15
10.001 10.001 10.004
10.002 i0.003 *0.001 10.001
-1.566 -1,784 -1.917 -2.091 -2.41 1
43
10.001
*o.ooo *0.001
*o .002
-2.126
17
Sample
SCE, V
Biphenyl
2,3 -
Corrected,mean
A EZd vs.
2,3,6-0.314 2,4,5-0.303 -0.355 -0.147 -0.308
-0.198 -0.316 -0.239 -0.320 -0.108 -0.199 -0.321
2,2'-
A EZd
10.002 io.001 *0.001
*0.002
*o.ooo rto.001
*0.001
10.002 *0.001
k0.005 k0.003 *0.002
YE.
SCE, V
-0.154 -0.320 -0.118 -0.151 -0.305
-0.116 -0.195 -0.313 -0.218 -0.133 -0.174 -0.320
*0.002
ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, M A Y 1975
901
Table 11. (Continued) NO.
Sample
Collected, mean
of
Ezd vs. SCE, V
m s
-2.411 3,3'-
SO.
A E 2 d vs.
Std dev,V
io.001
-2.030 -2.128 -2.411
20
4,4'-
-2 .ooo -2.411
20
10.002 10.002
2,4'-
-2.042 -2.41 1
17
*0.001
-2.123 -2.41 1 -1 .goo -2.01 7 -2.119 -2.41 1
10
-1.764 -2.001 -2.112 -2.411
15
-1.720 -1.807 -1.988
10
2,6,2',6'2,5,2',5'-
3,4,3',4'-
3,5,3',5'-
*0.001
10.001 10.001
io.001
15
10.001 io.001 10.002 10.002 10.002 10.005
SCE,V
-0.285 -0.098 -0.283
-0.369
2,5,2',4',5'-
2,4,5,2',4',5'-
-0.288 -0.117
-0.102 -0.292
*0.002 *0.002 *o ,002 10.003
-0.237 -0.111 -0.299
10.002 r0.002 10.001
-0.087 -0.181
CONCLUSIONS There are several important features of the interruptedsweep technique t h a t make it especially effective in the fingerprint identification of chlorinated hydrocarbons. Since the applied waveform is automatically controlled by the reduction behavior of the sample, reduction data ( E 2 d and i2d) are easily collected. Another feature of this particular instrument is its resolution capability. Whereas normal polarographic measurements have resolution limits of about 150 mV, the interrupted-sweep instrument can resolve reduction peaks whose E 2 d values are separated by less than 60 mV. In addition, the E 2 d are very reproducible and therefore permit the values to be reported to the nearest millivolt. A final advantage is the relatively low cost of the integrated circuit instrumentation, which is approximately $150 for the logic circuitry of Figure 1 and the necessary differentiator and integrator circuitry. The required power supplies were constructed for about $25 each. Using the described equipment, the voltammetric fingerprint technique requires a t least 9 pg of relatively pure compound in order to obtain positive identification. However, it may be possible to improve the sensitivity by optimization of the electrode and cell size, and by employing interrupted-sweep staircase voltammetry. A current problem is that the Hg-Pt working electrode must be cleaned and replated between samples, but preliminary results with a vitreous carbon electrode from Beckwith Carbon Corp. suggest that the carbon electrode can be used to replace the Hg-Pt electrode. As mentioned, our present system requires relatively pure compounds and in this manner is similar to infrared analysis. Both methods require that the components of a complex mixture first be separated by a chromatographic technique before the components can be ANALYTICAL CHEMISTRY, VOL. 47, NO. 6 , MAY 1975
of
E ~ ~ v SCE, s . V
runs
-2.103 -2.411
-0.411
was found to have no significant effect on the E 2 d data if the water content did not exceed 10%by volume. Additional water beyond 10% caused the E 2 d to shift to more negative potentials.
902
Sample
Corrected, m e a n
2,4,6,2',4',6'-
Decac hloro -
AEZd V I . Std dev, V
10.002 10.002
-1 .?71 -1.871 -1.981 -2.093 -2.411
20
-1.764 -1.902 -2.023 -2.127 -2.411
10
-1.908 -2.047 -2.411
10
-1.406 -1.687 -1.814 -1.997 -2.111 -2.411
15
*0.002 10.003 10.002 *0.003 10.004
io.002 *0.001
10.002 =0.001
10.001 10.002 *0.002 *0.001
10.002 10.002 10.003 10.002 10.003 i0.004
SCE, 1 '
-0.115 -0.308 -0.100 -0.110 -0.112 -0.318
-0.138 -0.121 -0.104 -0.284 -0.139 -0.364
-0.281 -0.127 -0.183 -0.114 -0.300
identified by comparing the unknown fingerprint with a known standard fingerprint. Controlled-potential coulometry and electrolysis, including chromatographic product analysis, have been performed on the polychlorinated compounds in order to identify the reduction pathways. These results will be reported in future papers ( 3 4 ) . ACKNOWLEDGMENT The authors are indebted to Robert Fifield for his technical assistance and Tektronix, Inc. for the loan of the Tektronix 564 storage oscilloscope. LITERATURE CITED D. Sissons and D. Welti, J . Chromatogr., 60, 15 (1971). R. G. Webb and A. C. McCall, J . Assoc. Off. Anal. Chem., 55, 746 (1972). S. Safe, 0. Hutzinger, and W. D. Jamieson. Org. Mass Spectrom., 7, 169 (1973). 0. Hutzinger and S. Safe, J . Chem. SOC.,Perkin Trans. 1, 5, 686 (1962). S.0. Farwell, F. A. Beland, and R. D. Geer, Bull. fnviron. Contam. Toxicol.. 10, 157 (1973). M. Von Stackelberg and W. Stracke, Z.Electrochem., 53, 118 (1949). C. L. Perrin in "Organic Polarography", P. Zuman, Ed., Interscience Publishers, New York, NY. 1969, pp 256-271. C. K. Mann and K. K. Barnes, "Electrochemical Reactions in Nonaqueous Systems," Marcel Dekker, Inc., New York, NY, 1970, pp 202-21 1. J. Zavada, J. Krupicka, and J. Sicher, Collect. Czech. Chem. Ccmmun., 28, 1664 (1963). S. 0. Farwell. F. A. Beland, and R. D. Geer, "Polarographic Identification of PCB Isomers," presented at the 26th Northwest Regional Meeting of the American Chemical Society, Bozeman, MT, June 1971. S. P. Perone, D. 0.Jones, and W. F. Gutknecht, Anal. Chem., 41, 1154 (1969). D. 0.Jones and S. P. Perone. Anal. Chem., 42, 1151 (1970). W. F. Gutknecht and S. P. Perone, Anal. Chem., 42, 906 (1970). L. Ramaley, R. L. Brubaker, and C. G. Enke, Anal. Chem., 35, 1088 (1963). A. C. Tas and R. J. C. Kleipool, Bull. fnviron. Contam. Toxicol., 8, 32 (1972). 0. Hutzinger. S. Safe, and V. Zitko, Bull. fnviron. Contam. Toxicol., 6, 209 (1971). F. A. Beland and R. D. Geer. J. Chromatogr., 84, 59 (1973). S.0. Farwell and R. D. Geer, Chem. Instrum., 5, 199 (1974). "RCA Preliminary CD4066A Data Sheet," RCA, Somerville, NJ, March 1973.
(20) R. J. Widlar, "Fairchild Application Bulletin App-116," Fairchild Semiconductor, Mountain View, CA. Feb. 1966. (21) M. K. Vander Kooi, "L144 Programmable Micro-Power Triple Op Amp, Siliconix Application Note." Siliconix Inc., Santa Clara, CA, Dec. 1973. (22) "Fairchild kA735 1 Data Sheet," Fairchild Semiconductor, Mountain View, CA, May 1973. (23) J. A. Dean and J. P. Rupley, "RCA ApDlication Note ICAN-6267," RCA, Harrison, NJ, March 1971. (24) "COS/MOS Digital Integrated Circuits," RCA Solid State Databook Series, Somerville, NJ, 1973. (25) S. 0. Farwell and R. D. Geer, "A Voltammetry Waveform Generator Controlled by Digital Logic," presented at the 26th Northwest Regional Meeting of the American Chemical Society, Bozeman, MT, June 1971. (26) K. H.Pool, C. J. Johnson, G. I. Connor, and G. H.Boehme, Anal. Cbem., 45, 437 (1973). (27) F. G. Cottrell. 2.Pbys, Cbem., 42, 385 (1902). (28) R. N. Adams. "Electrochemistry of Solid Electrodes", Marcel Dekker, Inc., New York, NY. (29) R. S.Nicholson and I. Shain, Anal. Cbem., 36,706 (1964). (30) L. Eberson and H. Schafer, "Organic Electrochemistry", Springer-Verlag, New York. NY, 1969, pp 256-271. (31) P. T. Allen in "Analytical Metods for Pesticides, Plant Growth Regulators, and Food Additives", Vol. V, G. Zweig, Ed., Academic Press, New York, NY, 1967, Chapter 3.
(32) G. J. Hoijtink and J. Van Schooten. Rec. Trav. Cbem., fays-Bas, 71, 1089 (1952). (33) J. D. Stuart, R. R. Kennan, R. J. Fenn, R. G. Jensen, and W. J. Pudelkievicz, "Degradation of Polychlorinated Biphenyls as Studied by Oxidative Electrochemistry", presented at the PCB Symposium of the American Chemical Society, Division of Water, Air and Waste Chemistry, Aug. 1972. (34) S. 0. Farwell, F. A. Beland and R. D. Geer, "Reduction Pathways of Organohalogen Compounds, Parts I and II", submitted for publication in J. Nectroanal. Chem.
RECEIVEDfor review February 4, 1974. Accepted January 20, 1975. Contribution from Agricultural Experiment Station, Montana State University, Bozeman, and published as Journal Series No. 490. Work supported in part by regional research funds, Project W-45. Taken from the Ph.D. thesis of S. 0. Farwell, Montana State University, August 1973.
Dye-Sensitized Continuous Photochemical Analysis: Identification and Relative Importance of Key Experimental Parameters R. White'
V.
and J. M. Fitzgerald2
Department of Chemistry, University of Houston, Houston, TX 77004
Important operating conditions for a new analytical technique, based on dye-sensitized reactions carried out in flowing streams, are evaluated. The photoredox reaction used is a form of triplet energy transfer wherein a colored organic dye absorbs light to generate the excited state triplet, which then abstracts electrons from a suitable substrate. The products of the reaction are colorless reduced dye and substrate oxidation products; spectrophotometric measurement of dye photobleaching is employed. The rate and stoichiometry of photoredox reactions are dramatically affected by the structures of dye and substrate used, dissolved oxygen, and reaction pH. Other important parameters, largely associated with the apparatus used, are dyecarrier flow rate, photolysis source power, and reaction temperature. Data for DCPA measurements, using Methylene Blue and Rose Bengal, are reported for the model substrates ascorbic acid, epinephrine, nicotine, and caffeine. Appropriate operating conditions yield low micromolar limits-of-detection and 1 YO precision.
Continuous flow analysis has been extensively applied in clinical chemistry with great success; a recent review details state-of-the-art developments ( I ). New continuous flow methods are often developed by adapting well-established manual procedures to machine execution. A second approach to continuous flow analysis involves development of new methods designed expressly for automation. Analytical methods based on photochemical reactions have been recently reviewed ( 2 ) . The theories of photochemical and Present address, M e d i c a l Laboratory Associates, 1025 S. 1 8 t h St., B i r m i n g h a m , A L 35205. Address r e p r i n t requests t o t h i s author.
*
related processes which an excited state molecule can undergo have been developed in detail (3-8). Intensely colored dyes are used as the photoactive reagent for the present work. Thus, the extent of the photochemical reaction can be measured spectrophotometrically, and the modest energies (40-50 kcal/mole) of visible light are well suited for use with biochemical substrates. We have previously reported the continuous determination of ascorbic acid (AA) by sensitized photooxidation with Methylene Blue (MB) (9). There exists an extensive literature detailing similar dye-sensitized reactions (6, 1015); most experiments were carried out batchwise in Warburg apparatus. Nevertheless, this literature provides a base for development of new analytical methods employing dye-sensitized photochemical reactions carried out in flowing streams. This technique will be referred to as "DyeSensitized Continuous Photochemical Analysis", and abbreviated hereafter as "DCPA". Although a study of the MBIAA system showed DCPA to have analytical potential ( 9 ) , it became clear that better understanding of the interactions between the photochemistry and operating conditions for DCPA was badly needed. T h e following objectives were selected for the present work: first, identify and evaluate those experimental parameters which have a significant effect on rate and extent of dye photobleaching reactions; second, determine whether experimental parameters for one given dye-sensitizer vary with the substrate photooxidized; third, develop guidelines for future study of dye-sensitized reactions to be tested for DCPA. Numerous dyes are potentially applicable as photosensitizers for DCPA; the exceptional paper by Gomyo et al. ( 1 1 ) served as a screening study. Based on their data, M B (a phenothiazine dye), and Rose Bengal, R B (a xantheine dye), were chosen because they have different wavelengths ANALYTICAL CHEMISTRY, VOL. 47, NO. 6, M A Y 1975
9
903
