Anal. Chem. 1990, 62, 2237-2241
2237
Photometric Determination of Acidity Constants by the Flow Gradient Technique without pH Measurements Juliana Marcos, Angel
os, and Miguel Valclrcel*
Department of Analytical Chemistry, Faculty of Sciences, University of Cdrdoba, Cdrdoba, Spain
A slmpie photometric method for the determination of acidity constants without pH measurements is proposed. I t is based on the establishment of a pH gradient in a flow system which Is created by means of a flow gradient. The proposed method thus avoids other less accurate procedures for the estabiishment of pH gradients. Large pH gradients can be maintained over long periods. The acidity constants of various compounds were determined automatically from different parameters of the absorbance-time recordings. The proposed method allows more than one acidlty constant for the same compound to be determined. The accuracy of the method is between 0.9% and 5.3%, in terms of reiatlve standard deviation, and fO.l and f0.4 in terms of confidence intervals.
INTRODUCTION Since its inception in 1975 (1, 2), flow injection analysis (FIA) has evolved a t a vertiginous speed and has proved to have a great potential in different fields of analytical chemistry (3, 4). FIA, and unsegmented flow methods in general, are among the most interesting approaches to automatic methods of analysis (5). Different operational modes and technological innovations have increased their possibilities even further in the last few years (3,4). One such mode is that of gradient techniques, which rely on measurements made before or after the residence time of the FIA peak. All these techniques (FIA titrations, gradient dilution, electronic calibration, etc.) are based on concentration gradients rather than on modified hydrodynamic features. In addition to affecting hydrodynamic aspects of unsegmented flow methods flow gradients result in concomitant reagent or analyte concentration gradients. Thus, the flow rate gradient, Q, can be expressed as the change in flow rate, q, over an interval of time, t
where qo, q and to, t are the initial and final flow rates and times, respectively. Although programmed-flow systems have long been used in high-performance liquid chromatography, there are only two references to their use with unsegmented flow methods (6, 7). Both report on the influence of flow rate gradients on the typical FIA parameters and on the stoichiometries of various complexes, which were determined by increasing the flow rate of one of the pumps stepwise while having the other pump deliver a constant flow rate (8). Obviously, the potential analytical applications of flow rate gradients in unsegmented flow methods are greater, providing the establishment of concentration gradients in an easy, fast, and accurate way. Moreover, there is no need to resort to the internal coupling of the injection valves (9,lO)or other procedures such as those involving the injection of very large sample volume at a very different pH from that of the carrier stream (11,12). These procedures are less accurate than the proposed method.
Flow rate gradients were used in this work to create accurate pH gradients, which in turn allowed the photometric determination of acidity constants of compounds active in the UV-vis spectral region with no pH measurements. The method thus developed is very simple and requires little human participation. All previous references to the determination of acidity constants in flow systems were based on absorbance-pH curves, so they required two detectors (photometric and potentiometric) (13, 14).
EXPERIMENTAL SECTION Reagents. Aqueous solutions of HC1 (Merck) of pH 3.2 and NaOH (Merck) of pH 10.5 were used. Aqueous solutions of the following compounds (10.0 /.tg mL-’) in acid (pH 3.2) and alkaline media (pH 10.5) were also used: bromocresol greeen (Merck);bromothymol blue (Merck); thymol blue (Merck);p-cresol (Merck); phenol (Merck);phenolphthalein (Aldrich);p-chlorophenol (Merck);p-nitrophenol (Merck);methyl red (Sigma); 8-hydroxyquinoline (oxine) (Merck);morin (Merck); L-histidine (Sigma); L-aspartic acid (Sigma); pyrocatechol violet (Aldrich); and quinine sulfate (Aldrich). Apparatus. A Hewlett-Packard 8415A diode array spectrophotometer furnished with an HP-9121 floppy disk drive, an HP-98155A keyboard, and an HP-7470A plotter was used. A Gilson Minipuls-3 peristaltic pump controlled by a Commodore-64 microcomputer through a home-made interface that allowed the continuous change of speed between a minimum and a maximum value at a preset rate was employed to establish the flow rate gradients. An Ismatec peristaltic pump was employed to ensure a constant flow rate. Finally, a Hellma 178.12 QS flow-cell (inner volume 18 kL) was also used. Creation of Flow Rate Gradients. The program controlling the functioning of the peristaltic pump through the interface was written in BASIC. Such control was effected through two bytes, DFOO and DF02. The interface and the direction of these bytes were activated from the program through POKE instructions and the continuous change of the pump speed was achieved through two consecutive FOR ...NEXT loops. To obtain different slopes of the flow rate-time function (flow rate gradient),it was necessary to introduce a STEP instruction in each loop (the same in both cases),and thus, the STEP instruction was directly related to the flow rate gradient. Performance of the Flow System. The configuration used to establish flow gradients for the photometric determination of acidity constants is depicted in Figure 1. It includes two channels that merge at confluence point P. One of them is fitted to the conventional pump (C.P.), which is always on and provides the initial flow rate (qo). The other channel is fitted to the programable pump (P.P.). It is started through the computer as each experiment is started and allows the flow rate through the channel to be changed from an initial qo/ = 0 to-‘4 (the maximum flow rate). The gradient value (Q) is given by Q(mL m i n 3 = [(qo + q’”) - qOI/(te - t o ) = q’max/te (1) where t , is the final time of the experiment and to is the initial time, which is equal to 0 in this case (all times are given in minutes). When solution A in Figure 1is acidic [pH = (pH),] and solution B is alkaline [pH = (pH)b], one can establish a pH gradient between (pH)b(at t = to)and (pH)’, (at t = te),from point P and the flow cell. If qe and q b are the flow rate of the acidic and alkaline solution, respectively (in this case q b is kept constant),
0003-2700/90/0362-2237$02.50/00 1990 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 62, NO. 20, OCTOBER 15, 1990
2238
,
pH
I
I00 SOLUTION
80
60
B SOLUT
Figure 1. Manifold used to create flow gradients in unsegmented flow systems: P.P.. programable pump; C.P., conventional pump (nonprogramable); P, confluence point; R, reactor; W, waste.
equations*
regression coeff Q,mL min-2
+ 0.84 + 0.84 = 0.840t + 0.84 = 0.960t + 0.84
0.999 0.998 0.998 0.998
0.12 0.60 0.84 0.96
+ 0.83 = 2.40t + 0.84 = 4.80t + 0.81 = 5.823 + 0.87
0.998 0.998 0.999 0.997
1.20 2.40 4.80 5.82
0.001 0.005 0.0075 0.008
(7 q q q
= 0.118t = 0.600t
0.01 0.02 0.04 0.05
q q q q
= 1.20t
"STEP values in the computer program. bFlow rate (4) in mL/min; time ( t ) in minutes.
and caand Cb their initial concentrations, total flow rate (qt) will be given by Pt = 9 a + q b = Qt +
qb
for pH >7 PH = 14 + log
[(cbqb
- caqa)/qtl
( 2)
A similar expression can be obtained for pH HA > H2A+ the absorbance-time curve shows two successive decreases and the derivative curve shows two negative peaks. This is the case with thorin in Figure 6a. When the absorbance sequence is
A- < HA > HzA+
> HA < H2A+
the recordings are like those in Figure 6c (oxine), with a minimum (PK,) followed by a maximum (pKJ in the derivative curve. Table IV lists the pK, values obtained for oxine, morin, pyrocatechol violet, thorin, quinine, and histidine. The results are all satisfactory and the proposed method allows the two pK, values of these compounds to be satisfactorily discriminated. Two amino acids (histidine and aspartic acid) were included in this study. Note that they required the acid-base reaction to be monitored a t 200 nm; in any case, the pK, values obtained are consistent with their literature counterparts. There are some limitations of this method for the diprotic acids when their pK1 and pK2 constants are quite different values (lo5or larger difference),because of the deviation from linearity in the calibration curve used (parameter pK, plot) at its ends. In these cases, the solution involves running two calibration curves by using acid and basic solutions of different concentrations. This allows one to obtain a calibration line suitable for the determination of each dissociation constant. If monitoring the system absorbance at one wavelength results in decreased sensitivity to one of the species in the system (second acid-basic equilibrium), it is possible to monitor two wavelengths simultaneously with the detector used (diode array), so this is no constraint to the method.
CONCLUSIONS A simple, rapid, and automatic method is proposed for the photometric determination of acidity constants without pH measurements, which is based on the establishment of pH gradients in a flow system through flow-rate changes. It is applicable to compounds absorbing in the UV-vis region provided the spectra of their acid and basic forms are different. The method should lend itself to other applications involving pH gradients or flow rate gradients on which, our team is currently conducting research. LITERATURE CITED (1) Ruzicka, J.; Hansen, E. H. Anal. Chim. Acta 1975, 7 8 , 145. (2) Stewart, K. K.; Beecher, G.R.; Hare, P. A. Anal. Eiochem. 1978, 70, 167. (3) Ruzicka, J.; Hansen, E. H. Flow Injection Analysis; Wiley: New York, 1988. (4) Valdrcel, M.; Luque de Castro, M. D. Now Injection Analysis. Principles and Applications; Ellis Horwood: Chichester, 1987. (5) Valdrcel, M.; Luque de Castro, M. D. Automatic Methods of Analysis ; ELsevier: Amsterdam, 1988. (6) Rlos. A.; Luaue de Castro. M. D.; Valclrcel, M. Talanta 1985, 32, 845. (7) Toei, J. Talanta 1988, 35, 425. (8) Martinez-Calatayud, J.; Campins, P.; Pascual, M. C. Analyst 1986, 1 1 1. .. 1317. (9) Rks, A.; Luque de Castro, M. D.; Valclrcel, M. Anal. Chim. 1988, 58, 663. (IO) Rks, A,; Luque de Castro, M. D.; Valclrcel, M. Anal. Chim. Acta 1987, 787, Y39. (11) Betteridge, D.; Fields, B. Anal. Chem. 1978, 50, 654. (12) Betteridge, D.; Fields, B. Anal. Chim. Acta 1981, 732,139. (13) Ramsing, A.; Ruzicka, J.; Hansen, E. H. Anal. Chim. Acta 1980, 774, 195. (14) Rlos, A.; Luque de Castro, M. D.; Valclrcel. M. Anal. Chim. Acta 1985, 777, 303.
RECEIVED for review April 4, 1990. Accepted June 29, 1990. The CICYT is acknowledged for financial support (Grant No. PA86/0146). J. Marcos is also grateful to the Bask Government for financial support received through a personal fellowship.
