trends in analytical chemistry, vol. 3, no. 8, 1984 191

Computer Conw

Package for on-line data acquisition and processing with microcomputers Professor Ducauze and his co-workers from the Institut National Agronomique Paris-Grignoninterfaced their CBM 8032 microcomputer with several chromatographs and developed a software package for the data acquisition and processing of the chromatograms. A specific option of the package is the correction of peak positions for long term drift by a comparison with reference chromatograms stored on disk. _

B. G. M. Vandeginste

D. N. Rutledge, J.-L. Excoffier and C. J. Ducauze

Because of the increasing quantity of information produced in the Analytical Chemistry Laboratory, computerized data acquisition and processing has become widespread. In order to treat and store in- formation obtained from a UV detector for high-per- formance liquid chromatography (HPLC), a flame ionisation detector for capillary gas chromatography (GC) and a polarometer, we have developed a mi- crocomputer system capable of carrying out up to three parallel data acquisitions from different instru- ments, and reducing this data to a directly usable form.

The hardware consists of a Commodore Business Machines CBM 8032 microcomputer (6502 micro-

Fig. 1. Data aquisition and processing system.

processor, 32 Kb RAM, equipped with a graphics card) connected to a CBM 8050 double floppy-disk drive (2x512 Kb); a +/- 5 V, 12 bit, 8 channel A/D converter-amplifier, CAN-12-2 (Sysmod-Eristel), with a maximum frequency of 5 kHz and a software controlled signal amplification of between 1 and 1024 for each channel.

The ,converter receives the analog signal via a shielded cable and a 25-pin connector (DB25) while address, data, and control signals are transferred be- tween the CAN-12-2 and the CBM 8032 via an inter- face PROM and bus (Sysmod-Eristel) connected to two of the microprocessor’s memory extension ports. The data are accessed directly by reading the contents of the corresponding memory addresses.

Hardcopies are produced on a CBM 8023P printer and/or a Hewlett-Packard 7470A two dimensional plotter. The system is shown in Fig. 1.

192 trends in analytical chemistry, vol. 3, np. 8, 1984

Data acquisition A multichannel data-acquisition program,

MULACQ, was developed in BASIC and ASSEM- BLER. With this program, up to 90,000 points per analysis can be stored with a sampling period of be- tween 0.060 and 15.3 s. Subroutines are inserted into the microprocessor’s interrupt routine, which nor- mally interrupts all other CPU tasks to allow for scanning of the keyboard, reading and writing to cas- settes etc., as well as for updating of the 50Hz inter- nal clock. The added subroutines, which are also ex- ecuted every 1/50th of a second, control the data ac- quisition.

For each channel, data points, obtained by aver- aging overthe chosen period, are temporarily stored in one of two buffers in the screen memory. A full buffer is transferred between two acquisitions to a relative file on the floppy-disk. For instance, with a period of 250 ms it is possible to carry out analyses lasting more than 6 h and store more than 18 h of data on a single floppy-disk.

For those situations where a higher resolution or a higher signal-to-noise ratio is needed, we also devel- oped a rapid acquisition program, RAPACQ, with a base frequency of 1 kHz. In this case, the mi- croprocessor is programmed to generate and emit l- kHz pulses through the user port by pin CB2, to be detected directly at pin CA1 on the 6522 VIA (versa- tile interface adaptor).

The binary data are read in the addresses allo- cated to CAN-12-2 and summed over a period of be- tween 1 ms and 2 s. The averaged points are all stored in the memory and transferred to the floppy- disk at the end of the analysis. This limits the number of averaged measurements to about 8000 per analy- sis.

Data pre-processing The processing is carried out in two steps: firstly

peak detection (program PEAK), then peak quantification (program SURF).

Peaks are detected using the pseudoderivative method 1, in which the variation between two points with a given time-lag is calculated. Interactive pa- rameter inputs aid the operator in choosing the best threshold values for each detector and signal-to- noise ratio. Optional time-outs (for example, the maximum number of -points between the apparent beginning of a peak and the detection of a summit) are included to reduce the effect of spurious points and changes in baseline. It is also possible to plot any portion or all of the data in order to verify the correct detection of the peaks. At the end of the treatment, a sequential file is created containing the approximate

positions of the beginning, the summit and the end of each peak.

Peak quantification The values obtained in the previous step are used

in the second program, SURF, to determine more precisely the peak position and to calculate the area of up to 250 peaks as well as other parameters such as half-height width or third and fourth order moments.

In the case of overlapping peaks three methods of calculating the peak area are available:

0 tangential - the surface under overlapping peaks is simply neglected;

0 dropline - the surface under each peak is con- sidered as belonging only to that peak;

0 pro rata - the total surface under the group of overlapping peaks is divided among them in propor- tion to their ‘tangentiai’ surface.

During the processing, graphics are provided on the monitor screen and/or on hardcopy for checking. A report is printed out containing peak number, po- sition, relative and absolute areas as well as an indi- cator of converter saturation. This information is stored in a new sequential file. In order to facilitate the treatment of a large number of similar raw data files, a master-program was developed to control the sequence of programs described above.

Correction of peak drift Many analytical systems, such as HPLC and capil-

lary GC, are liable to introduce a drift in peak posi- tion as a result of prolonged use or changes in equip- ment. This problem has been largely solved by means of a third program (CORRECT) which al- lows the operator to readjust the positions contained in the reduced file in relation to those of a reference file.

After displaying peak patterns for each file the op- erator introduces, if necessary, the numbers of those pairs of peaks he considers equivalent and all the peak positions of the second file are recalculated by interpolation. The process is iterated until the oper- ator considers the correspondence satisfactory, at which time a corrected file is created. After each iteration a hardcopy can be obtained, and at the end of the process a checkup-list of proposed peak corre- spondences, based on proximity, are printed out with the associated area ratios.

This data acquisition and processing system is cur- rently used in our laboratory for the collection and interpretation of raw data. In some instances, files created by the program are transmitted by a modem to a main-frame computer for further multivariate statistical analyses such as pattern recognition.

016%9936/84/$02.00. 0 Elsevier Science Publishers B.V.

trends in anplytical chemistry, vol. 3, no. 8, 1984 193

As it would in most cases be possible to transpose this software into other microcomputers, we believe it could be of use to many other analytical chemists. For further information, write to: D. N. Rutledge, Laboratoire de Chimie Analytique, Institut National Agronomique Paris-Grignon, 16, Rue Claude-Ber- nard, 75005 Paris, France.

References 1 J. L. Excoffier and G. Guiochon, Chromatogruphia, 1.5 (1982)

543-W..

Computer Corner - Contributions

Contributions of between 400 and 900 words are welcome in the categories described below and should be sent to the appropriate contributing editor.

Information on hardware, general software, software tips, and interfacing should be sent to:

TrAC Computer Corner B. G. M. Vandeginste, Depart- ment of Analytical Chemistry, University of Nijmegen, Toernooiveld, 6525 ED Nijmegen, The Netherlands.

Information on chemical applications, software and math- ematical tools for improving information content should be sent to:

TrAC Computer Corner, D. L. Massart, L. Kaufman, Vrije Universiteit Brussel, Fakulteit der Geneeskunde en der Farmacie, Farmaceutisch Scheikunde, Laarbeeklaan 103, B-1090 Brussels, Belgium.

trends

Micellar enhanced analytical fluorimetry

Willie L. Hinze, H. N. Singh, Yoshimi Baba and Noel G. Harvey Winston-Salem, NC, USA

Fluorescence measurements can be enhanced by the judi- cious use of micelles in the analytical scheme. This new ap- proach to jluorescence provides increased sensitivity, reduc- es the number of potential interferences, and offers greater experimental convenience.

Quantitative spectrofluorimetric methods are very useful in chemical analysis which is principally due to their extreme sensitivity and inherent selectivity. Recent advances have almost exclusively revolved around development of sophisticated instrumental and computerized techniques to further improve de- tection and specificity. It seems that considerations of solution chemistry and specific fluorophore-en- vironmental effects to achieve the same goals have been largely neglected. This article reviews a rela- tively new approach to facilitate fluorescence assays, based on the formation and manipulation of molecu- lar association between the fluorescent species and appropriate aqueous surfactant micelles in solution.

Micellar systems Surfactants, also called detergents, are amphiphil-

ic molecules where a polar headgroup attached to a

0165-9936/84/$02.00.

long non-polar tail provides it with distinct hydro- philic and hydrophobic regions. A surfactant can be classified as cationic, non-ionic, zwitterionic, or an- ionic, depending upon the nature of the polar head- group. Above the Kraft temperaturei, upon reach- ing a characteristic concentration called the critical micelle concentration (CMC), surfactant molecules dynamically associate to form roughly spherical ag- gregates of colloidal dimensions termed micelles (Fig. 1) in equilibrium with a concentration (CMC) of dissolved surfactant. The number of surfactant molecules comprising a micelle is its aggregation number (ZV). Table I lists the name, acronym, struc- ture, and micellar parameters (i.e. CMC and N> of some common micelle-forming surfactants that have been employed in analytical fluorescence measure- ments.

Although aqueous micellar systems continue to be the focus of intensive study in many disciplinesi they have had few practical applications in analytical chemistrys. The crucial factor in all successful appli- cations of micelles lies in the ability of solutes to as- sociate and bind with the micellar assembly (Fig. 1). The nature, i.e. charge-type and carbon chain length, of both the solute and surfactant are impor- tant factors influencing the strength of association between different solutes and micelles because these factors determine the net hydrophobic and/or

0 Elsevier Science Publishers B.V.