A computer-controlled potentiometric/spectrophotometric titrator

A laboratory computer controlled potentiometric titrator interfaced to a diode array spectrophotometer is described. The titrator consists of widely used, commercially available components; therefore, major attention is given to modes of interconnection and software implementation in data format and system control. Replicate potentiometric titrations of glycines gave a relative standard deviation in titre of 1.035% and a relative standard deviation in pH of 0.745%. Replicate spectrophotometric titrations of bromophenol blue were analysed at three wavelengths to yield pKa= 3.898 ± 0.075 (1.9% rsd). Methods of data presentation and manipulation are presented.


Introduction
The technique of titrimetry lends itself well to automation. Two general methods for controlling the titration operation have been employed: the older hardware control (i.e. the use of dedicated discrete logic circuits), and more recently, software control (i.e. the use of programmable microprocessors or microcomputers). Both methods have been successfully applied to potentiometric titrations [1][2][3][4][5][6][7][8] and to spectrophotometric/ potentiometric titrations [9][10][11][12][13]. Various approaches to the implementation of these techniques are delineated in figure 1.

AUTOMATIC TITRATORS
The two broad categories of control each have advantages and disadvantages. Hardware control generally is less expensive, but severely limits flexibility. Software control utilizing dedicated microprocessors yields much greater flexibility at the expense ofgreater instrumental complexity. A substantial portion of this complexity can be eliminated by use of a self-contained computer. While such an approach may require the development of interfaces for use between general purpose or personal computers and external equipment, laboratory computers equipped with a wide variety of integrated interfaces and systems drivers callable by high-level programming languages have become widespread.
Additional information and wider scope of operation are available by the incorporation of spectrophotometric analysis of the titrand. This can be realized simply as endpoint determination of a chromophoric titrand at fixed wavelengths [8 and 14], or extensive analysis and data gathering from a complex equilibrium system involving the recording of the entire absorption spectrum [10 and 11]. The latter approach can avail recently devised matrix formalism for manipulating large amounts of data generated by spectrophotometric titrations for the determination of wavelength optima for the analysis of multicomponent systems 14 [8, 12, 13 and 14] and the use of a microprocessor-controlled scanning spectrophotometer [11]. More recently [10], a photodiode array detector was employed in a dedicated microprocessor-controlled titrator which afforded reduced data collection times. This paper presents a discussion of the construction of a spectrophotometric titration apparatus employing commercially available and widely used equipment. Hard-copy graphics were generated on a Hewlett-Packard HP9872C eight-pen flat bed digital plotter driven by the MINC IEEE-488 interface bus. The programming language used throughout was MINC BASIC V1.2.
Titration system: The titration system is comprised of a titrant delivery device and a digital pH meter. The pH meter used was the Orion model 801A digital pH/mV meter. Data from the meter is encoded as parallel BCD. On this particular unit (and all those with serial numbers lower than 7300) the voltage of a logical is +1.5 V, which is too low to ensure recognition by standard TTL circuitry. A simple NPN transistor switch (2N4001) for each BCD bit was employed, which produces an inverted output. The inversion was compensated by setting the data invert switch on the computer's digital input unit. The titrant delivery system was similar to the one previously described [6]. The Gilmont micrometer burette, 2"5 ml total capacity, is driven directly by a Superior Electric Slo-Syn HS25 stepping motor. The stepping motor is driven by a Slo-Syn model STM103 translator module wired according to the manufacturer's 96 specifications. The module translates serial pulses from the computer into the proper motor coil energization sequence. One motor step (0.9 , or 0.25 1 using a 2.5 ml Gilmont syringe) is produced from each pulse generated by one bit of the computer's digital output unit with the translator set to half-step mode.
The solution being titrated is circulated from a jacketted vessel (about 20 ml total capacity, equipped with an efficient stirrer) through a spectrophotometer flow cell (Fisher) using a Cole-Parmer Masterflex peristaltic pump with viton tubing. The speed of the pump is set to provide complete exchange of the flow cell contents with the titration vessel in 5 s. The burette tip is immersed in the solution continuously. A block diagram of the titrator is shown in figure 2.

Software
Data structure and communicalion Preliminary consideration: The system drivers called by MINC BASIC to operate the RS232 interface ports will transmit and receive only serial ASCII, while the HP8450A is capable of sending data in serial eight-bit binary. Such data will always by interpreted as seven-bit ASCII by MINC, thereby losing the most significant of the eight binary bits. It was therefore necessary to instruct the 8450A to send all data as serial ASCII, a much slower process than sending eight-bit binary. ASCII data sent by the HP8450A contain a notation for the wavelength in nm, as, for example, L 201, followed by the absorbance at that wavelength. Hence, records arriving after the first one will begin with LF. It was found that the very first record transmitted after measuring a spectrum and sending it to the computer will begin with the characters 0 (zero, ASCII 48), or occasionally with a LF, 0 (ASCII 10, ASCII 48) as indicated in table 2, alternate structure. It is therefore necessary to execute software which will examine each At this point the deuterium and tungsten lamps are turned on by the computer and allowed to warm up for 25 min. After warm-up, another 'lamp on' command is issued to allow correct diode amplifier gain settings to be established. A balance measurement is then made by the computer (with or without cells in sample and reference compartments, at the operator's discretion).
Setting spectral parameters It is often desirable to alter the default spectral width parameters (200-800 nm). The time required for transmission of a spectrum increases with the width of the spectrum, and attention is often confined to a particular spectral region. It is also sometimes necessary to set fixed upper and lower absorbance limits. These limits are entered into the computer as character strings and must be translated into the correct key-code command strings. The character strings are stored as elements of a string array. Each element of the array is 'decomposed' by the SEG$ operator, which is set to return a single character from the argument string, in order from first to last. Each single character substring is then translated into the corresponding key-code by reference to a translation table. The individual component key-code values are stored sequentially in a numeric string. Thus, the first three elements of this array contain the key-code equivalents for each of the three digits of the lower wavelength limit. The next three elements contain the upper limit, the next three the lower absorbance limit, and the last three the upper absorbance limit. Each key-code number is for a given value converted to a character (with the CHR$ operator) and concatenated to form the key-code character strings of the lower and upper wavelengths for a spectrum. The command string to set the HP8450A to these values would be:

Results and discussion
The major purpose of the titrator described here was to obtain a set of absorption spectra corresponding to a series of pH or other potentiometric measurements in machine-readable form in the quickest, simplest and most reliable manner available to us. The accuracy is pH Figure 3. Data (+) and calculated best fit for the titrations of bromophenol blue at A" 588 nm; B: 309 nm; C: 436 nm. concentration of 7 x 10-3M plus 0"48-0"50 x 10-3M HC1 provide information on the repeatability of the titrant delivery system and the pH measurement system. The average titre for the first end-point was 499.0 + 1"6 t,1 (rsd 0"3%), and the average titre for the second end-point was 1211"6 + 13"0 ml (rsd 1-1%). The average pH at the first end-point was 6"023 + 90"127 (rsd 2"1%) and at the second point it was 10 The results of these calculations are shown in table 3, which gives pKa values as a function of experiment and wavelength. The average value for all experiments at a particular wavelength was 3"898. + 0"062 (1'6%) and the average value for all experiments at all wavelengths was 3"898 + 0.075 (1"9%). The data and simulated curves are shown in figure 3.
A particular advantage of machine-readable data is the flexibility imparted to data manipulation, analysis, and presentation. As an example, a single pH titration of bromophenol blue is presented in figure 4 in conventional format, and in two three-dimensional formats. The latter produce a titration surface, which in the case of more complex systems would be capable of revealing additional information which might be obscured in more conventional representations. Aside from using other source languages, the major difference expected in using other computers would be in the RS232 drivers. A more flexible system which allows reception of eight-bit binary would be more efficient in terms of transmission time, but more rigid in that the entire spectrum (1664 bytes) is sent in binary mode, regardless of the requested spectral width parameters. Transmission by the computer in RS232 would be facilitated by other drivers which allow inclusion of the actual output data in the subroutine argument without need for the generation of cumbersome command strings, as well as the mode in which it is to be sent. The actual transmission format as presented here is completely general, however.