Modular Conductivity System MSC-200 User's Manual

Version 1.0 January 2006

MCS-200 Hardware user's manual Table of Contents 1 WARRANTY.................................................................................................................................... 3 2 Specifications.................................................................................................................................... 4 3 General Description of Instrument.................................................................................................... 5 3.1 Principle of operation.................................................................................................................5 3.2 Schematics of Components of the MCS-200.............................................................................5 3.3 Description of conductimetric observation head....................................................................... 5 3.4 Description of MSC-200 amplifier............................................................................................ 7 4 Installation of instruments and cabling instructions.......................................................................... 7 4.1 Installation of mechanical components......................................................................................7 4.2 Cabling instructions................................................................................................................... 8 4.3 Networking of the MCS-200..................................................................................................... 8 4.4 Configuration and initialisation in BioKine.............................................................................10 5 Practical guide for experiments....................................................................................................... 12 5.1 Recording of Impedance spectra..............................................................................................12 5.2 Recording of Rapid kinetics in conductivity mode................................................................. 14 5.3 Determination of experimental electrical cell constant........................................................... 18

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MCS-200 Hardware user's manual 1 WARRANTY BIO-LOGIC WARRANTS EACH INSTRUMENT IT MANUFACTURES TO BE FREE FROM DEFECTS IN MATERIAL AND WORKMANSHIP UNDER NORMAL USE AND SERVICE FOR THE PERIOD OF ONE YEAR FROM DATE OF PURCHASE. THIS WARRENTY EXTENDS ONLY TO THE ORIGINAL PURCHASER. THIS WARRANTY SHALL NOT APPLY TO FUSES OR ANY PRODUCT OR PARTS WHICH HAVE BEEN SUBJECT TO MISUSE, NEGLECT, ACCIDENT, OR ABNORMAL CONDITIONS OF OPERATION. IN THE EVENT OF FAILURE OF A PRODUCT COVERED BY THIS WARRENTY, THE PRODUCT MUST BE RETURNED TO AN AUTHORIZED SERVICE FACILITY FOR REPAIR AND CALIBRATION AND TO VALIDATE THE WARRANTY. THE WARRANTOR MAY, AT THEIR DISCRETION, REPLACE THE PRODCUT IN PLACE OF REPAIR. WITH REGARD TO ANY INSTRUMENT RETURNED BECAUSE OF DEFECT DURING THE WARRENTY PERIOD, ALL REPAIRS OR REPLACEMENTS WILL BE MADE WITHOUT CHARGE. IF THE FAULT HAS BEEN CAUSED BY MISUSE, NEGLECT, ACCIDENT, OR ABNORMAL CONSITIONS OF OPERATION, REPAIRS WILL BE BILLED AT NORMAL COST. IN SUCH CASES, AN ESTIMATE WILL BE SUBMITTED BEFORE WORK IS STARTED. IN CASE ANY FAULT OCCURS: NOTIFY BIO-LOGIC OR NEAREST SERVICE FACILITY, GIVING FULL DETAILS OF THE DIFFICULTY, AND INCLUDE THE MODEL NUMBER, TYPE NUMBER, AND SERIAL NUMBER. UPON RECEIPT OF THIS INFORMATION, SERVICE OR SHIPPING INSTRUCTIONS WILL BE FORWARDED TO YOU.

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MCS-200 Hardware user's manual

2 Specifications The general technical specifications of the MSC-200 are: Detector:

Rapid Impedance spectrometer with Fast-Fourier Transform (FFT) Technology Excitation frequency: 1 Hz to 200 kHz Excitation amplitude: 1 to 500 mV or 1000 mVpp Accuracy of detection: 1 % Highest sampling rate: 300 s/point Number of recorded data: 3 (real part and imaginary part of impedance, frequency) Power supply: 220 V/16 A (110 V on demand)

Observation cuvette with conductivity cell:

Standard Stopped-flow cuvette modified with special electrodes In impedance mode and optical absorbance mode (with appropriate spectrometer unit) Combination with all SFM models Connection to detector: banana plugs at the observation head Connection to hardstop: via PEEK tubings and standard HPLC fittings Electrical cell constant: 8 cm-1 Optical path length of cuvette: 1.5 mm Minimal dead time: 2.4 ms (in combination with SFM-20) Electrode material: Glassy carbon Other materials in contact with the solutions: Quartz, PEEK, Teflon, Viton (can be repalced by EPDM or ISOLAST on demand)

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MCS-200 Hardware user's manual

3 General Description of Instrument 3.1 Principle of operation The Modular Conductivity System, MCS-200, is a rapid impedance spectrometer. It allows (i) to measure steady-state spectra of the impedance of a solution and (ii) to follow rapid changes in the solution's impedance in the course of a chemical reaction (time course measurements). The principle of the detection technique is to impose a high-frequency electrical voltage in the solution between two electrodes and to follow the time course of the resulting current. By means of a FastFourier-Transform (FFT) technique, the two components of the complex impedance (real part and imaginary part) can be computed at sampling rates up to 200 s per data point. The real part of the complex impedance represents the electrical resistance of the solution in the cuvette provided that the imaginary part is sufficiently small. The spectral mode of the MCS-200 (measurement of real and imaginary part as a function of the excitation frequency) allows to determine the optimal frequency at which the imaginary part is minimal and the real part does not show a strong variation with the frequency (plateau state, see Fig. 11). 3.2 Schematics of Components of the MCS-200 The MCS-200 consists of an impedance spectrometer unit and a special observation head with an observation cuvette that contains two electrodes made from glassy carbon. For rapid kinetics experiments, the observation head needs to be coupled to a Stopped-Flow Mixer unit (SFM-20, SFM-300 or -400) to allow a rapid mixing of solutions prior to the conductimetric detection. Handling of the SFM unit is supposed to be known and will not be explained further on in this manual (see respective manuals). The spectrometer unit is connected to the observation cell via three cables that serve to apply the high-frequency excitation voltage and to detect the resulting current; a third cable connects the observation head (stainless steel) to the common electrical ground, since the observation head serves also as shielding against external electrical voltages (Faraday cage). The MCS-200 is connected to a PC computer workstation via an Ethernet link (standard network connection); therefore the PC needs to be equipped with a network card. The MCS-200 has its own IP address and can be identified by this number. All data transfer between PC and MCS-200 passes across the network cable, which can be either connected directly to the MCS-200 or via an existing PC network. The control of the stopped-flow mixer, the MSC-200, the data acquisition, the recording and the analysis of data is done entirely by the software BioKine32 (for detailed information on BioKine32 refer to the appropriate manual). 3.3 Description of conductimetric observation head In Fig. 1, a schematic view of the observation head for conductimetric detection is shown. Two adaptors on both sides allow to fix the cables from the MCS-200 module. A drilling hole on the foot of the head is provided to set the head chassis to the same ground as the measuring device.

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Figure 1: Schematic view of the special observation head. Two adapters on the sides allow to fix the cables coming from the MCS-200. The connection to the hardstop is made via PEEK tubings that are fixed by means of a HPLC fitting screwed onto the cuvette holder (top).

A more detailed view of the cuvette with the two electrodes from glassy carbon can be seen in Fig. 2. The cuvette is fixed to a support made from Teflon and is inserted into the observation head. The top screw tightens the cuvette into the head. By means of PEEK tubings and an HPLC fitting, the support can be connected to the hardstop of the stopped-flow module. Please note that due to electrical interferences, the hardstop can not be connected directly onto the observation cuvette as in standard optical stopped-flow experiments, but it has to be placed further away onto a special hardstop fixation.

Figure 2: Schematic view of the cuvette for conductimetric detection. Electrodes are made from glassy carbon, a material which is more inert than Platinum and which avoids adhesion of air bubbles on the surface. The cuvette can also be used for optical absorbance with a path length of 1.5 mm.

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MCS-200 Hardware user's manual 3.4 Description of MSC-200 amplifier A schematic view of the front and rear faces of the MCS-200 unit is given in Fig. 3. Several cables provide connections to the different devices. An Ethernet (TCP/IP) cable is used to establish communication between the MCS-200 and the controlling PC workstation (Communication card). The Impedance Card is connected directly to the SFM connector card via a 25-pin cable. A trigger cable from the MPS power supply of the stopped-flow module can be connected to start the acquisition via an external trigger pulse.

Figure 3: Front (top) and rear (bottom) view of the MCS-200 module. The front view shows three parts: the communication card, the impedance card, and the SFM connector. The communication card is connected to the network via an Ethernet cable. The impedance card has an entry for a trigger cable (from the MPS unit). The Sfm connector card has a cable that is connected to a small blue interface card with cables to the impedance cell at the observation head.

4 Installation of instruments and cabling instructions 4.1 Installation of mechanical components To install the conductimetric detection accessory on an existing SFM model, remove the standard observation head by loosening the four screws. Replace the head with the conductimetric observation head that contains the impedance cell (Fig. 1). Fix the special hardstop support and the support for the small blue interface board on two opposite sides of the upper part of the SFM by means of two screws each; in case of a SFM-20 model, please use the shorter faces. Take the hardstop electro-valve device and unscrew the beige connection that is used with the standard support of optical cuvettes; in case you have an additional hardstop without this connector, just replace the two hardstops. Please take care that the MPS unit is switched off before you change the double-pin BNC cable (high voltage, damage)! Fix an appropriate PEEK tube with HPLC connectors to the hardstop and to the cell support with is already inside the observation head. Please do not remove the conductivity cell to avoid a damage of the electrical connections!
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MCS-200 Hardware user's manual Place the small blue interface box from the MCS-200 unit onto the corresponding support that you have previously fixed onto the SFM. 4.2 Cabling instructions In order to have a correct cabling and mass shielding an appropriate mass connection has to be assured for all power cables of the MCS-200 and the stopped-flow device (in particular the MPS unit). Several cables have to be connected: A From the small blue interface box to observation head:

Two red cables connected together go to one electrode connector on one side of the observation head. The two blue cables and the white cable are connected together and go to the second electrode connector on the opposite side of the observation head. The thin black cable is connected to the black connection of the interface box and to a hole on the bottom of the observation head; this cable serves as electrical mass. A network cable; for details about the networking see next chapter. A power cable The small interface box is already inserted into the corresponding slot of the MCS-200.

B From the MCS-200

4.3 Networking of the MCS-200 The MCS-200 must be connected to the PC via a network connection; all communication (configuration, initialisation and data transfer) use an Ethernet protocol. Both the PC and the MCS200 need their personal IP addresses. The MCS-200 can either be connected directly to the network card of the PC via an inverted Ethernet cable; alternatively, the MCS-200 and the PC can be connected to the same Ethernet network via an Ethernet Hub (or similar) and a direct Ethernet cable (in most of the cases); please note that both types (inverted and direct) of Ethernet cables are delivered with the MCS-200. To configure the IP address of the MCS-200, a special software tool, Ipsettings.exe, is provided. Start this software and a configuration window appears (Fig. 4). Click on Search to find present instruments (Fig. 5). Choose the instrument that shows MCS-200 as Device type. You can now edit the new IP number, Netmask and Gateway. Activate new settings by clicking on Change param. A successful is indicated in a green box (Fig. 6).

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Figure 4: IP settings for the MCS-200. To view present instruments click on Search.

Figure 5: Select the number with Device type MCS200. You can add the new IP setting and activate them by clicking on Change param.

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MCS-200 Hardware user's manual 4.4 Configuration and initialisation in BioKine After correct connection of all cables, the instruments and devices have to be configured in BioKine32. The actual version is CMI.1.02. To configure devices, go to Install and Device ... . Check the box Conductimeter and eventually Use stopped-flow (see Fig. 7). Please note that no acquisition bard has to be detected for the work with the MCS-200.

Figure 7: Configuration of the Device Installation menu for work with the MCS-200.

Click on the button to connect to the MCS-200 module. The MCS-200 control menu opens that gives access to the status control and configuration of the IP address of the MCS-200. In case a Channel failure (0) error occurs (red box on the right side of Fig. 8), edit the correct IP address, click on the Connect button and reload eventually the firmware. A green box with Channel ready shows that the channel is ready for measurements.

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Figure 8: Control status and configuration menu for MCS-200. On the right side, a wrong IP address has been given resulting in a Channel failure (0) error (red box). On the left side, the correct IP address has been edited and the Channel ready box appears in green.

Click OK to leave the MCS-200 control menu.

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5 Practical guide for experiments 5.1 Recording of Impedance spectra To record an impedance spectrum of the actual solution inside the observation cell, open Device and Transient Recorder (kinetics). Click on Acquisition setup menu to program the spectral settings of the MCS-200. Check Impedance Spectrum in the Conductimeter measurement menu (see Fig. 9). The different parameters and their meanings are:

Initial frequency (Hz): the excitation frequency at which the spectral recording should start; between 1 Hz and 200 kHz. Final frequency (Hz): the excitation frequency up to which the impedance spectrum should be recorded; between 1 Hz and 200 kHz. Linear spacing: Check this box if you want a linear spacing of frequencies between the initial and the final frequency; if not, a logarithmic spacing is chosen automatically. Number of points: number of measured frequencies between the initial and the final value. Amplitude (mV): amplitude of the high-frequency excitation electrical voltage in mV; maximal value is 500 mV;an amplitude of 50 mV is usually sufficient for spectral recordings. Average Na: number of repetitions for each frequency value in order to get an average value. With I Range: current shunt, leave this on auto range for spectral recordings. Bandwidth: a parameter that helps that the calculation converges to a stable value; default setting is 5 (medium speed). Advanced: these parameters can be used to impose a static electrical voltage for a certain time period prior to the spectral recording; for standard experiments, leave Figure 9: Configuration for recordings of impedance spectra (details see text). these settings in default configuration.

Click on the button in order to start the acquisition of the impedance spectrum. The button allows to interrupt the recording at any moment. A typical impedance spectrum (Z(f)) with the above given settings and 2 mM Kcl in the cuvette is shown in Fig. 10. Two modes of representation can be chosen:

Nyquist: The negative imaginary part of Z is plotted as a function of the real part of Z (Fig. 1). This is a representation that many electro-chemists prefer. It allows to determine best the point where the imaginary part of Z is minimal (Fig. 10).
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ReZ/ImZ: Both real and imaginary part of Z are plotted as a function of the excitation frequency. In this representation, the excitation frequency where the imaginary part is minimal can be determined directly from the plot (Fig. 11).

Figure 10: Typical impedance (Z) spectrum for a 2 mM Kcl solution in Nyquist representation (x-axis: real part of Z in Ohm, ReZ(ohm); y-axis: imaginary part of Z in Ohm, -ImZ(ohm)). Measurement settings are as given above. The cursor indicates the point where the imaginary part is minimal, which would be optimal for rapid kinetics recordings.

A practical and easy procedure to determine the excitation frequency where the imaginary part is minimal is first to choose a Nyquist representation and determine the minimum with the cursor function of BioKine. Then switching to a ReZ/ImZ representation will leave the cursor at the same point, and the optimal frequency can be read from the cursor information. If the resolution of frequency values is too low in the region of the minimum, you can record a new spectrum with a restrained settings of the frequency range but the same number of measured frequency values.

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Figure 11: Typical impedance (Z) spectrum for a 2 mM KCl solution in a classic representation (x-axis: frequency in logarithmic scaling; y-axis: real (yellow, top) and imaginary part (green, bottom) of Z in Ohm (ReZ(ohm), -ImZ(ohm)). Measurement settings are as in Fig. Xxx. The cursor indicates the point where the imaginary part is minimal, which would be optimal for rapid kinetics recordings (optimal frequency here: f = 22 kHz).

5.2 Recording of Rapid kinetics in conductivity mode To record a rapid kinetics time-course of the actual solution inside the observation cell, open Device and Transient Recorder (kinetics). Click on acquisition setup menu to program the spectral settings of the MCS-200. Check Kinetics in the Conductimeter measurement menu (see Fig. 12). The different parameters and their meanings are:

Sinus frequency (Hz): frequency of the electrical excitation voltage between the two electrodes; this should have been determined as described above; between 10 Hz and 200.kHz, please note that too low setting can result in electro-chemical reactions on and pollution of the electrode surfaces Sinus amplitude (mV): amplitude of the high-frequency electrical voltage; between 1 and 500 mV; please note, the higher this setting the better the signal-to-noise ratio; however, high values
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MCS-200 Hardware user's manual can also result in electro-chemical degradation of the solution and deposition of compounds on the electrode surfaces.

Repeat: Number of data points recorded for the kinetics; from 1 to 8.000. Number of FFT points: Sets the number of points that are used for the FFT calculation for each recorded data point; multiples of 2, to be chosen with the two arrow buttons. Non stationary correction: Check this option for FFT points lower than 64 to improve the calculation of impedance for very fast recordings. With I Range: current shunt; this value has to be chosen according to the stationary impedance value of the solution (see below). Bandwidth: a parameter that helps to prevent instabilities during the measurement; default setting is 5 (medium speed). Advanced: these parameters can be used to impose a static electrical DC potential additionally to the highfrequency voltage; for standard experiments, leave these settings in default configuration (Set DC potential (V): 0.0; Vs OC unchecked) since a static voltage can induce electro-chemical degradation of the solutions and pollution of electrode surfaces. Display in S/cm: Check this box if you want the measured values to be recorded in specific Figure 12: Configuration of recordings of conductivity (Siemens/cm) rather than in resistance impedance kinetics (details see text). (Ohm). The value k denominates the electrical cell constant that serves to convert resistances into specific conductivities (see below).

Click OK to accept the setting. The measurement can be started in the transient recorder as usual. In case, no trigger signal is connected to the MCS-200, the acquisition will start automatically and will be repeated until acquisition is stopped manually by clicking on the Now or End button. A typical recording of a time course in impedance mode (2 mM KCl, no kinetics) can be seen in Figs. 13 and 14. The measured data will be plotted and the display screen updated only at the end of the entire acquisition. Display of data is either in resistance values (R in Ohm) or in specific conductances (L in Siemens/cm). The electrical cell constant k (in 1/cm) is used to convert data automatically according to the following formula: L = k/R The desired units must be chosen prior to data acquisition in the Conductimetry measurement menu. Automatic change of units is not possible after data acquisition. For determination of k see the respective chapter.
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Figure 13: Rapid time-course measurement in impedance mode for a 2 mM KCl solution with the above given settings. A number of 1000 points results in an acquisition time of 0.9 s, thus, an effective sampling of 0.9 ms per data points has been achieved with the settings. Real part in yellow (top) and imaginary part in green (bottom) are plotted as resistances in kOhms.

The exact sampling rate (ms/point) can not be set directly in the Conductimetry measurement menu. Due to the principle of measurement, the rate depends on the combination of two other settings, i.e. the Sinus frequency and the Number of FFT points (see above for detailed description). As a rule, the sampling rate increases with increasing Sinus Frequency and with decreasing Number of FFT points. As the excitation Sinus Frequency is determined by the minimum of the imaginary part of the impedance for the solution under investigation (see above), the sampling rate can only be increased by decreasing the Number of FFT point. However, a smaller Number of FFT points will also result in a decreased signal-to-noise ratio. The total time of sampling depends on the experimental sampling rate and the total number of points.

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Figure 14: Rapid time-course measurement in impedance mode for a 2 mM KCl solution with the above given settings. Same recording as in Fig. 13, a detailed view of the real part is shown in kOhms. Under the given conditions, the noise is lower than 0.2 %.

The current range parameter (With I range, see above) has to be carefully chosen for rapid kinetics recordings, since it can not be adjusted automatically at each measurement. The optimal range depends on the value of the real part of the impedance of the solution under investigation. As a rule, the division of 1 V by the actual value of ReZ must be in the same order of magnitude as the current range value, but inferior. For instance in Fig. 14, the resistance ReZ = 28.4 kOhm, the ratio of 1 V/28.4 kOhm yields 3.57 A; therefore a current range of 10 A has been selected for this recording (see settings above).

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5.3 Determination of experimental electrical cell constant To determine the experimental cell constant, impedance spectra of solutions with known values of the specific conductance (L in S/cm) have to be recorded. The corresponding resistance of the solution (R in Ohm) can be measured at the frequency where the imaginary part of the impedance is minimal. A plot of L as a function of 1/R yields a straight line; the slope of the linear regression denotes the cell constant, which has been determined to k = 8.0 cm-1. The corresponding measurement is shown in Fig. 15.
1250 L = k1/R
-1

theoretical specfic conductivity,

1000

k = 7.9886 cm R = 0.9986
2

L (Scm )

-1

750

500

250

0 0 50 100 150

measured conductance, 1/R (S)

Figure 15: Determination of nominal cell constant. A plot of the measured conductance (1/R in S, real part at frequency where imaginary part is minimal) versus the theoretical specific conductance (L in S/cm) of the respective concentration of KCl yields a straight line (intercept A=0.9004); from the slope, the cell constant can be calculated to k = 8.0 cm-1. Measurements have been conducted at 25 C.

The following values of the specific conductance of different concentrations of KCl at 25 C have been used in the calculations: Concentration of KCl (mol/l) 0.0002 0.0005 0.001 0.002 0.004 0.008 Specific conductivity L (Scm-1) 29.9 73.7 145.6 287.7 568.6 1123.8

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