DE3028569C2 - - Google Patents

Description

The invention relates to a biophysicochemical arrangement according to the preamble of claim 1, a method their manufacture and sensors with such an arrangement.

A biophysicochemical arrangement of the above chenen Art is in the journal Experientia, Volume 28 (1972), Pages 256 and 257 described. It reacts to the zu give substrate molecules with a change in the lead ability which is used on both sides of the Electrodes immersed in the liquid medium and a conventional conductivity meter is measured. The increase in conductivity after adding substrate molecules and the degradation of increased conductivity takes place in periods of the order of a few Wed. grooves.

It has now been found that a measurable reaction of the conductivity of the membrane to the reaction catalyzed by enzyme molecules is obtained even if, according to claim 1, the number of enzyme molecules on the active area of the membrane is drastically reduced, insofar as this is only possible experimentally is (concentrations of 10 ^-4 - 10 ^-3 enzyme molecules per molecule of the membrane material can be adjusted without experimental difficulties), and the concentration of the substrate molecules present in the liquid medium drastically decreases, which is possible without difficulty. Very rapid, pulsed changes in the conductivity of the membrane are then obtained, which can be directly assigned to the catalytic decomposition of a substrate molecule or a cluster (cluster-like agglomeration) of substrate molecules.

A biophysicochemical arrangement according to the invention  does not just examine the dynamics of surface-bound enzyme-catalyzed reactions, one can also such biophysicochemical arrangements di rect as an inexpensive, very easy and quick in large pieces use the model to be produced for nerve membranes. In the biophysicochemical arrangement according to the invention namely when using the also in the living ner venzell found enzymes, substrates and membrane materials lien exactly the same conductivity in a comparable environment channels like synaptic or axonal membranes found in vivo. This means when testing the effect of pharmacological agents on nerves a hehebli che simplification of those to be carried out in large numbers Try and also allow an investigation of the dynamics the interaction between the active ingredient and the nerve membrane.

Advantageous details in the construction of an inventive biophysicochemical arrangement are in subclaims given.

In particular, with the further development of the invention according to claim 9 an even closer replica of a reached the natural nerve membrane. A biophysicochemistry cal arrangement according to claim 9 can be by adding Excite substrate molecules just as electrically as this Nerve membranes is the case.

Because a biophysicochemical arrangement according to the present Invention elementary events, namely those caused by an enzyme Molecularly catalyzed reaction of a substrate to measure molecular or a substrate molecule cluster ge , you generally have a very sensitive one in it Feeler, which also reacts in different ways lization of classic sensors, e.g. in a Experimental set-up for the investigation of reaction processes  on enzymes (see claim 12), in a pH meter (see Claim 14), in a temperature meter (see. Claim 16), in a measuring device for the electric field strength (see claim 17) or in a flow meter (see. Claim 19).

With the development of the invention according to claim 13 achieved a direct cross reaction between the to investigating enzyme / substrate system and the enzyme / substrate system of biophysicochemical arrangement is not possible.

With the development of the invention according to claim 15 achieved that in addition the pH of the liquid substances to be determined is not based on the enzyme / Act substrate system of the biophysicochemical arrangement can.

With the development of the invention according to claims 18 and 24 becomes the sensitivity of the sensor in question improved again.

The development of the invention according to claims 20 and 21 is with regard to the economical use of substrate molecules advantageous and also promotes the sensitivity of the flow mungsmessers, because its output signal of the difference between the rate of diffusion of the substrate molecules perpendicular to the direction of flow and the flow rate depends on yourself.

A flow meter according to claim 22 is particularly suitable good for recording small flow velocities. At it is the supply of substrate molecules and possibly ions absolutely constant in the liquid flow to be measured bend.  

A flow meter according to claim 23, however, is suitable particularly good for measuring high flow velocities.

The invention will now be described with reference to embodiments play with reference to the accompanying drawing explained. In this shows

1 is a vertical section through a chemical biophysiko arrangement which functions for measurement by surface-bound enzyme catalyzed reac and is suitable for measuring the pH, at an intermediate stage of their production.

Figure 2 is a vertical section through the active loading area of the membrane and an adjacent section of a membrane support film in a greatly enlarged scale.

Fig. 3 is a schematic representation of a esterasemoleküles acetylcholine;

Fig. 4 is a longitudinal section through a flow meter, which operates with a biophysikochemischen assembly;

Fig. 5 is a sectional catalyzed by an experimental arrangement for investigation by surface-bound enzyme substrate reactions;

. Fig. 6 is a graph of the conductivity of the membrane a biophysikochemischen assembly of Figure 1 in function of time at very ge ringer concentration of the enzyme molecules on the active area of the membrane and very little of the substrate molecules Kon concentration in the liquid medium; and

Fig. 7: a view similar to Fig. 6, but with the concentration of the enzyme molecules and the substrate molecules chosen to be somewhat larger.

Fig. 1 shows a bowl-shaped container 10 , in the bottom of which a slot 12 is provided which extends over the entire width. In the slot 12 , a partition 14 is flow-tightly displaceable, which also rests with its lateral edges in a fluid-tight manner on the side walls of the container 12 . For additional sealing, silicone grease can be provided at the sealing points. In this way, two compartments 16 and 18 are predetermined by the container 10 and the partition 14 , which are each filled with an aqueous salt solution, for. B. with an aqueous, 1-molar (ionic strength) K⁺Cl ^- - or Na⁺Cl⁻ solution, which is manufactured to a pH between 7 and 7.5 and at a temperature of 292 - 296 ° K is set.

The partition 14 is provided in its upper portion with an inclined on one side walls 20 opening, which is covered by a thin film 22 made of tetrafluoroethylene. The thickness of the film 22 is 25 .mu.m, and in the middle is a working opening 24 with a diameter of about 100 .mu.m, z. B. pierced with a thin needle.

A monomolecular layer 26 or 28 of a lipid, preferably L- α- phophatidylcholine made from soybeans, is spread onto the surface of the aqueous solution in compartments 16 and 18 . The lipid layer 26 is also covered with a further layer which contains enzyme molecules, in the exemplary embodiment considered here acetylcholinesterase, and is obtained by spreading a drop of solution 30 .

If you lower the partition 14 very slowly with a speed of about 0.5 mm / min, the adjacent lipid layer 26 or 28 is taken along from the front and the back of the film 22 , and one obtains work opening 24 in the area of the work a molecular double layer which forms a membrane 32 .

Further details regarding the method known per se for membrane production are in the magazine Proc. Nat. Acad. Sci. USA, Volume 69 (1972), pages 3561-3566

The solution drop 30 contains a total of 10 g (corresponds to 10 ^-12 mol) of a tailed native acetylcholinesterase, which is isolated from the electrical tissue of the eel Electrophorus Electricus and is brought into a highly pure state by affinity chromatography. This cleaning process is described in more detail in the journal Biochemical and Biophysical Research Communications, Volume 79 (1977), pages 640-647. When cleaning, care is taken that only the enzyme fractions with the sedimentation coefficients S of 14 and 18 get into the purified product.

Fig. 2 shows schematically a in no way scaled section through the film 22 in the region of the opening 24 . One can see the two lipid molecules 26 and 28 built up from lipid molecules 34 , which combine to form the membrane 32 in the region of the working opening. In the lipid layer 26 , the acetylcholinesterase enzyme molecules 36 are also incorporated. On average, there is one enzyme molecule 36 for every 10 ⁴ lipid molecules 34 .

Fig. 3 shows schematically the structure of an acetylcholine esterase molecule, as is known from the above-mentioned publication in Biochemical and Biophysical Research Communications. A "*" or a "**" indicate different locations of the enzyme molecule at which it can be cleaved by protolysis. The long tail 38 of the enzyme molecule 36 can form compounds with lipid molecules; With circles, catalytic subunits 40 are identified, which are in the form of three tetramers.

In the biophysicochemical arrangement shown in FIG. 1, callomel electrodes 42 , 44 immersed in one of the two compartments 16 , 18 are provided, which are connected to the terminals of an operating circuit 46 . This contains a series resistor 48 connected to a direct voltage source U _B and a direct current amplifier 50 . The output of the DC amplifier 50 is connected to the Y input of a Y t recorder 52 and to the input of a frequency counter 54 . The output signal of the amplifier 50 is thus assigned to the total impedance of the current path formed by the electrodes, the aqueous solution and the membrane. This practically depends only on the impedance of the high-resistance membrane 32 . The output signal of the amplifier 50 thus directly reflects the conductivity of the membrane 32 .

If you now also bring a small amount of non-stirred acetylcholine in compartment 16 , e.g. B. in the form of a 5 × 10 ^-6 -olar aqueous solution of acetylcholine⁺ Cl⁻, so you get 52 discrete conductivity impulses with a half width of 5 × 10 ^-2 sec and one on the K⁺Cl⁻ concentration on the recorder of the aqueous environment normalized height of about 2 × 10 ^-10 (Ohm × MK⁺Cl⁻) ^-1 , as shown in Fig. 6. Upon closer examination of the individual conductivity pulses with a higher temporal resolution, it can be seen that they represent a rapid succession of very rapid pulses with a half-width of a few 10 ^-4 sec. The latter are presumably obtained in each case when an acetylcholine molecule comes under the influence of an acetylcholinesterase molecule and is then catalytically hydrolyzed and thereby releases a proton. The pulse sequence with a total duration of 5 × 10 ^-2 sec, on the other hand, probably corresponds to the successive hydrolysis of individual acetylcholine molecules belonging to a cluster, the number of molecules coming together in a cluster being essentially constant for a given solvent.

Neither the conductivity pulses with a half-value width of about 5 × 10 ^-2 sec nor the conductivity pulses with a half-value width of about 10 ^-4 sec visible at higher temporal resolution have been received so far, since large concentrations of enzyme molecules and substrate molecules were used .

In order to obtain individual conductivity pulses, the frequency with which an impact of a substrate molecule or a substrate molecule cluster is obtained on an enzyme molecule must be kept so small that the individual conductivity pulses do not overlap in the vast majority of cases. This frequency obviously increases with the concentration of the surface-bound enzyme molecules, with the concentration of the substrate molecules distributed in the aqueous environment, with the size of the active membrane area and with the mobility of the substrate molecules in the respective liquid environment. Primarily via the various concentrations, but also via the choice of the temperature of the liquid in the compartment 16 and the size of the active membrane area, it can be ensured in individual cases by simple tests that discrete individual conductivity pulses are obtained on the membrane, on the other hand the mean time interval between the individual conductivity pulses does not become unnecessarily large. When increasing the pulse frequency by increasing the concentration of substrate molecules and / or enzyme molecules, one can go so far until, in rare exceptional cases, a pulse already falls on the falling edge of a previous pulse. A corresponding pulse train is shown in FIG. 7, in which the units on the coordinate axes are the same size as in the representation of FIG. 6.

The arrangement shown in FIG. 1 can also be used directly for measuring the pH of the liquid in compartment 16 , since the rate of hydrolysis of acetylcholine in the presence of acetylcholinesterase depends on the pH found in each case. The frequency output by the operating circuit 46 or the display of the frequency counter 54 is thus a direct measure of the pH value and, if need be, for larger deviations of the measured pH value from the pH range 7-7.5 with additional use of an attempt to modify easy-to-create correction tables.

Fig. 4 shows a longitudinal section through a flow meter, which has a switchable into a water flow tubular body 56 . This is provided in its wall with an opening 58 which is covered by a film 60 . This in turn has a working opening covered by a membrane, as shown in FIG. 2.

A container 62 is placed on the outside of the tube. In it there is an aqueous salt solution, e.g. B. a 1 molar KCl solution, in which a Kallomel electrode 64 is immersed. A second callomel electrode 66 is surrounded by the water flow in the interior of the tubular body 56 . As in FIG. 1, the electrodes are connected to an operating circuit 46 , the output of which is again connected to the input of a frequency counter 54 . This has a period of the order of one second, so that a large number of conductivity pulses are recorded for each reading.

Upstream of the opening 58 , a further opening 68 is provided in the wall of the tubular body 56 , into which a body 70 made of open-pore material is inserted. The opening 68 is in communication with the inside of a tightly placed on the outside of the tubular body 56 Vorratsbehäl age 72 , which contains an aqueous solution of acetylcholine, and possibly additionally of KCl, if the water flow to be measured de conducts electricity very poorly.

The inside of the storage container 72 is connected via a pressure control valve 74 to a storage bottle 76 for pressurized inert gas, so that a given amount of liquid is dispensed from the storage container 72 into the liquid flow per unit of time, namely in its outermost surface layer.

The flow meter described above works as follows measure:

When the water flowing in the tubular body 56 flows in a laminar manner, its edge layer takes 70 acetyl choline clusters out of the body. These can now diffuse in the water flow in a direction perpendicular to the direction of flow, that is to say from the wall of the tubular body to the tubular body axis. The slower the flow, the more acetylcholine clusters are diffused away from the tube body wall when the edge layer of the water flow reaches the film 60 , the less conductivity pulses are registered by the frequency counter 54 . The level of the frequency counter 54 is thus a direct measure of the flow rate of the water sers.

For very high flow rates compared to the diffusion rate of the acetyl choline clusters, the device for feeding acetyl choline clusters into the water flow can be designed so that the discharge rate is assigned to the flow rate of the water. This can be similar to a motor vehicle carburetor using a Venturi nozzle, which has a feed opening for the acetylcholine solution formed in the nozzle wall, which is connected to the storage container 72 . In the case of such a flow meter, the number of acetylcholine clusters arriving at the film 60 per unit of time also increases with the flow rate.

If, on the other hand, you have to measure very low velocities of the water flow compared to the diffusion speed of the acetylcholine cluster, you can expand the flow meter according to Fig. 4 by the parts shown in broken lines there, which represent a second biophysicochemical arrangement and each have a component described above correspond. For this reason, these components are provided with corresponding, but additionally provided with a comma.

The frequency counter 54 is then ausgebil det as an up / down counter, the up counting terminal + with the output of the loading operating circuit 46 and the down counting terminal - is connected to the output of the operating circuit 46 '.

The counter reading of the frequency counter 54 thus corresponds directly to the deviation of the movement of the acetylcholine clusters from the completely symmetrical diffusion pattern obtained at zero flow velocity and is thus directly associated with the low drift velocity of the water flow.

It can be seen that in the flowmeters described above, the sensitivity can be easily changed by adjusting the pressure control valve 74 , while mechanical changes are not necessary. It can also be seen that the measurement result is again provided in frequency modulation which is advantageous for a noise-free and rapid signal processing.

The flow meter shown in Fig. 4 can also be used with very few modifications for measuring an electric field, since the acetylcholine is usually in the form of a polar salt, e.g. B. Acetylcholine⁺ Cl⁻ is added and the migration of acetylcholine⁺ ions through an external electric field is influenced in a similar manner as by the entrainment by a water stream. For this purpose, one need only to the ends of the tubular body 56 in Fig. 4 dash-dot recorded aufzu lid 78 put. Then you get an asymmetry of the diffusion field of the delivered from the body 70 acetylcholine⁺-cluster, so that the operating circuits 46 and 46 'emit pulses with different frequency by the applied external electric field again. In order to prevent a back diffusion of acetylcholine clusters, which run past the film 60 without decomposition, the inside of the cover 78 can be covered with a membrane carrying a large number of acetylcholinesterase molecules, so that the acetylcholine clusters arriving there are definitely decomposed.

The field strength meter just described is used both to determine the direction of the field and to determine the field strength, for which you first turn the tubular body 56 so that the greatest possible reading is obtained on the frequency counter 54 , and then the value of the maximum reading no tiert.

The arrangement described above automatically compensates for the influence of a slowing down of the reaction caused, for example, by the resulting reaction products, since both slides 60 and 60 'are affected in the same way by this slowing down. After a longer measurement period, the used, acetate-rich solution in the interior of the tubular body 56 is discarded and replaced by new solution, for which purpose corresponding connections are provided on the tubular body 56 .

FIG. 5 shows a setup for the investigation of reactions on surface-bound enzyme molecules, which is similar to that shown in FIG. 1; corresponding parts are again provided with the same reference numerals and need not be described again in detail. The arrangement is shown in the operational state, that is, after the manufacture of the membrane formed by lipid bilayers, which are not shown in the rest.

The bottom of the container 10 now has a second slot 12 ', in which a second partition 14 ' is arranged just as ver as the partition 14 in the slot 12th The partition 14 'has an opening 20 ', which is covered by a film 22 '. In this respect, there is complete agreement with the arrangement parts already described above.

The working opening of the film 22 'not shown in FIG. 5 is in turn covered by a membrane which is formed by a lipid double layer. However, this membrane in turn now carries different enzyme molecules than the membrane of the film 22 , namely those which are to be examined in their interaction with substrate molecules assigned to them. These substrate molecules are also in the aqueous environment of compartment 16 . In order to prevent that they react directly with the substrate molecules for the enzyme molecules of the membrane of the film 22, and to prevent substrate molecules for the Enzymmole molecules of the membrane of the film 22 with the enzyme molecules of the membrane of the film 22 'to react, the compartment 16 divided by a wall 80 , which cool only for the reaction products of the catalytic decomposition of substrate moles cool on the membrane of the film 22 ', e.g. B. for protons.

The arrangement shown in FIG. 5 thus consists of a test part to the left of wall 80 and a measuring part to the right of wall 80 . It can be seen that the experimental arrangement according to FIG. 5 has a very simple structure and also allows the course of enzyme-catalyzed reactions of substrate molecules to be investigated if the enzyme molecules are to be carried by membranes which themselves have no or only a very small guide show ability change. Among other things, one can also investigate reactions on molecular single layers.

You can also use a temperature meter with one above described biophysicochemical arrangement easily reali due to the fact that the remaining operating parameters are recorded the mobility of the substrate molecules or clusters in the liquid environment depends on the temperature.

Thus, in the arrangement according to FIG. 1 with a fixed concentration of substrate molecules and enzyme molecules, the same liquid environment and the same geometry, the frequency of the conductivity pulses is a measure of the temperature of the liquid environment in the compartment 16 , since that on the membrane 32 by the enzyme molecules 36 decomposed substrate molecules have to be replaced by temperature-dependent diffusion from the volume of the liquid medium. The compartment 16 therefore only needs to be thermally coupled to the object whose temperature is to be measured when using the arrangement according to FIG. 1.

Claims (24)

1. Biophysical chemical arrangement, consisting of a membrane immersed in an electrically conductive liquid environment, the electrical conductivity of which depends on the environment surrounding it, from enzyme moles supported by the membrane and from molecules distributed in the liquid environment, which molecules are in the Participate milieu under catalysis by the enzyme molecules in a reaction which results in at least one reaction product which influences the electrical conductivity of the membrane, characterized in that the effective concentration of enzyme molecules ( 36 ) on the active surface of the membrane ( 32 ) and the effective concentration of substrate molecules or clusters of substrate molecules taking into account the rate of migration of the substrate molecules or substrate molecule clusters and taking into account the speed of the catalytically promoted reaction by the enzyme molecules are chosen so that individual impuls S-shaped changes in the electrical conductivity of the membrane ( 32 ) can be obtained. 2. Arrangement according to claim 1, characterized in that the membrane ( 32 ) consists of two lipid layers ( 26, 28 ) be. 3. Arrangement according to claim 2, characterized in that the membrane made of dipalmitoyl-phosphatidylcholine, Sphingo myelin or phosphatidylserine. 4. Arrangement according to claim 3, characterized in that the membrane consists of L- α- phosphate dylcholine obtained from soybeans. 5. Arrangement according to one of claims 1-4, thereby indicates that the substrate is acetylcholine and Enzyme is acetylcholinesterase and that the milieu Aqueous environment containing ions. 6. Arrangement according to claim 5, characterized in that the acetylcholinesterase is a highly pure, native acetyl is chlinesterase. 7. Arrangement according to claim 6, characterized in that the acetylcholinesterase by affinity chromatography from the electrical organ of Electrophorus Electricus is made. 8. Arrangement according to one of claims 1-7, characterized in that the liquid medium additionally contains ATP, ie adenosine triphosphate and the membrane ( 32 ) additionally contains Na⁺-K⁺-ATPase. 9. Arrangement according to claim 8, characterized in that the concentration of the ATPase is about 10 ^-4 - 10 ^-3 molecules per molecule of the membrane material. 10. A method for producing a membrane according to one of the Claims 2-9, characterized in that on the Surface of the liquid environment on either side of one Carrier film, which provide a working opening is that initially over the free surface of the river milieus lies, by spreading a monomolecular Lipid layer is made on one of the lipid layers by spreading a dilute enzyme solution Enzyme molecules are applied and the carrier film then slowly opening into the milieu is moved into it. 11. The method according to claim 10, characterized in that L- α -phosphatidylcholine is used as the lipid and an aqueous K⁺Cl⁻ or Na⁺Cl⁻ solution buffered to pH 7-7.5 is used as the aqueous medium, the temperature thereof is between 292 and 296 ° K. 12. Biological test arrangement for examining the reaction sequences on enzymes which are carried by the surface of a membrane, characterized in that use of a biophysicochemical arrangement according to one of claims 1-9 or a biophysicochemical arrangement which is produced according to claim 10 or 11, as it forms a portion of a partition ( 14 ) of a loading container ( 10 ) which is filled with the liquid medium and whose compartments ( 16, 18 ) are connected to a conductivity meter ( 46, 52 or 54 ), and that the immerses the enzyme-carrying membrane to be examined in that compartment ( 16 ) which is connected to the enzyme molecules belonging to the biophysicochemical arrangement. 13. Use according to claim 12, characterized by a partition ( 80 ) which is only permeable to the products of the reaction controlled by the enzyme to be investigated and which is arranged between the biophysicochemical arrangement and the environment into which the membrane with the enzyme to be examined is immersed . 14. pH meter, characterized in that use of a biophysico-chemical arrangement according to one of claims 1-9 or a biophysicochemical arrangement which is produced according to claim 10 or 11 as a two-compartment ( 16, 18 ) container ( 10 ) is provided for the liquid environment, the partition-dividing partition ( 14 ) is partly formed by the membrane ( 32 ) of the biophysicochemical arrangement that the compartments ( 10, 18 ) of the loading container ( 10 ) with a conductivity meter ( 46 , 52 and 54 ) are connected and that that compartment ( 16 ) of the loading container which communicates with the enzyme molecules of the biophysicochromic arrangement is at the same time in connection with the liquid whose pH is to be measured. 15. Use according to claim 14, characterized in that the compartment ( 16 ), which communicates with the enzyme molecules of the biophysicochemical arrangement, communicates with the liquid via a further membrane permeable to hydrogen ions, the pH of which is to be measured by who. 16. Temperature meter, characterized in that use of a biophysicochemical arrangement according to one of claims 1-9 or a biophysicochemical arrangement which is produced according to claim 10 or 11, as a two compartments ( 16, 18 ) having container ( 10 ) for the liquid Milieu is provided, the partition ( 16, 18 ) delimiting the partition ( 14 ) is partly formed by the membrane ( 32 ) of the biophysicochemical arrangement, that the compartments ( 16, 18 ) of the container ( 10 ) with a conductivity meter ( 46, 52 or 54 ) are connected, and that at least that compartment ( 16 ) of the container ( 10 ) that communicates with the enzyme molecules of the biophysicochemical arrangement can be thermally coupled to the object whose temperature is to be measured. 17. Electrical field strength meter, characterized in that use of a biophysicochemical arrangement according to one of claims 1-9 or a biophysicochemical arrangement, which is produced according to claim 10 or 11, as a two compartments ( 56, 62, 62 ') comprising containers Dielectric material is provided for the liquid medium, the dividing wall dividing the compartments is partly formed by the membrane of the biophysicochemical arrangement, that the compartments of the container are connected to a conductivity meter ( 46, 46 ', 54 ) that at least one Compartment ( 56 ) of the container, which communicates with the enzyme molecules of the biophysicochemical arrangement, can be moved into the field-filled space, and that the substrate molecules are present in the liquid medium in ionized form. 18. Use according to claim 17, characterized in that two biophysicochemical arrangements ( 58 - 66; 58 '- 66' ) are arranged symmetrically on both sides of a source ( 68 - 76 ) for ionized substrate molecules and the operating circuits associated with the biophysicochemical arrangements ( 46, 46 ') are connected on the output side to a difference computer ( 54 ). 19. Flow meter, characterized in that use of a biophysicochemical arrangement according to one of claims 1-9 or a biophysicochemical arrangement, which is produced according to claim 10 or 11, as a tubular body ( 56 ) for the liquid flow to be measured and one on the wall, of the tubular body ( 56 ) attached container ( 62 ) are provided that the membrane of the biophysicochemical arrangement at least partially closes an opening ( 60 ) provided in the partition between the tubular body ( 56 ) and the container ( 62 ) that the interior of the tubular body ( 56 ) and the interior of the container (62) having a routing capability knife (46, 54), and in that power means (68 on the membrane of biophysikochemischen arrangement - 76) cules for feeding Substratmole and optionally ions to be measured liquid electricity is provided. 20. Use according to claim 19, characterized in that the feed means (68-76) molecules which Substratmole and optionally the ions in the boundary layer of the laminar body guided within the tube feeding the liquid stream. 21. Use according to claim 20, characterized in that the feed device has an open-pore body ( 70 ) which communicates with the boundary layer of the liquid flow and which is connected to the interior of a storage container before, which contains a substrate and possibly a solution containing ions. 22. Use according to claim 21, characterized. by means ( 74, 76 ) for maintaining a constant overpressure in the storage container ( 72 ). 23. Use according to claim 20, characterized in that that the feed device one of the edge layer of the Fluid flow through venturi tube, which one with a reservoir for substrate moles cool and possibly ion - related opening in the nozzle wall has. 24. Use according to any one of claims 19 - 23, characterized in that a second biophysicochemical arrangement ( 58'-66 ' ) is arranged upstream of the feed device and the outputs of two of the two bio-physicochemical arrangements ( 58-66 and 58'-66 ' ) Associated operating circuits ( 46, 46 ') are connected to the inputs of a differential computer ( 54 ).