DE3028569A1 - Bio:physico:chemical sensor - based on conductivity change of enzyme-coated membrane - Google Patents

Abstract

Device whose electrical conductivity changes in response to change in the surrounding conditions comprises an enzyme-coated membrane immersed in an electrically conductive liq. medium contg. a substrate which undergoes an enzyme-catalysed reaction to produce at least one prod. which alters the conductivity of the membrane. The improvement comprises selecting the effective concn. of enzyme mols. on the active surface of the membrane and the effective concn. of substrate mols. (or clusters thereof) in the liq. medium, taking into account the diffusion rate of the substrate and the rate of the enzyme- catalysed reaction, so as to produce pulsed changes in the conductivity of the membrane. The device can be used to monitor enzyme reactions or to measure pH, temp, electric filed strength or flow rate. The device operates with much lower enzyme and substrate concns. than known devices of this type (cf. Experientia, 28, 256, 1972).

Description

Biophysicochemical arrangement, procedures for their

Manufacture as well as sensors with such an arrangement description The invention relates to a biophysicochemical arrangement according to the preamble of claim 1, a method for their production and sensors with such Arrangement.

A biophysicochemical arrangement of the type mentioned at the beginning is described in the journal Experientia, Volume 28 (1972), pages 256 and 257. It reacts to the addition of substrate molecules with a change in conductivity, using electrodes immersed in the liquid medium on both sides of the membrane and a conventional conductivity meter.

The increase in conductivity after adding substrate molecules and the degradation of the increased conductivity takes place in periods of order of magnitude few minutes.

It has now been found that there is a measurable response in conductivity the membrane still receives the reaction catalyzed by enzyme molecules, when one according to claim 1, the number of enzyme molecules on the active area of the Membrane drastically reduced, as far as this is only possible experimentally (concentrations from 10 -10 3 enzyme molecules per molecule of the membrane material can be obtained without experimental Difficulties set), and also the concentration of in the liquid medium existing substrate molecules drastically reduces, which is possible without difficulty is. One then obtains very rapid, pulse-shaped changes in the conductivity of the Membrane, which is directly involved in the catalytic decomposition of a substrate molecule or assigned to a cluster (heap-like agglomeration) of substrate molecules can be.

A biophysicochemical arrangement according to the invention is permitted not just an investigation of the dynamics of catalyzed by surface-bound enzyme molecules Reactions, such biophysicochemical arrangements can also be used directly as inexpensive, Model for nerve membranes that is very easy and quick to produce in large numbers use.

In the case of the biophysicochemical arrangement according to the invention, namely when using the enzymes, substrates, which are also found in living nerve cells and membrane materials in a comparable environment have exactly the same conductivity channels as found in synaptic or axonal membranes in vivo. This means at the examination of the effect of pharmacological agents on nerves is a considerable Simplification of the experiments to be carried out in large numbers and also allows one Investigation of the dynamics of the interaction between active substance and nerve membrane.

Advantageous details in the structure of a biophysicochemical according to the invention Arrangement are specified in subclaims.

In particular, with the development of the invention according to Claim 9 achieves an even closer replica of a natural nerve membrane. A biophysicochemical arrangement according to claim 9 can be achieved by adding substrate molecules just as electrically excite as is the case with nerve membranes.

As a biophysicochemical arrangement according to the present invention Elementary events, namely those catalytically induced by an enzyme molecule Measurement of the reaction of a substrate molecule or a substrate molecule cluster, one has in it generally a very sensitive antenna, which is also on can be used in various ways to implement classic sensors, e.g. in a Experimental arrangement for the investigation of reaction processes of enzymes (cf. claim 12), in a pH meter (cf. claim 14), in a temperature meter (cf. claim 16), in a measuring device for the electric field strength (cf. claim 17) or in a flow meter (cf.

Claim 19).

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

With the development of the invention according to claim 15 it is achieved that the pH value of the liquid should also be determined Substances do not have an effect on the enzyme / substrate system of the biophysicochemical arrangement can.

With the development of the invention according to claims 1E and 24 the sensitivity of the sensor in question is further improved.

The development of the invention according to claim 20 and 21 is in view an economical use of substrate molecules is beneficial and also promotes the sensitivity of the flow meter as its output depends on the difference between the diffusion speed of the substrate molecules perpendicular to the direction of flow and the flow velocity itself depends.

A flow meter according to claim 22 is particularly suitable for Detection of small flow velocities. With him is the supply of substrate molecules and, if necessary, ions in the liquid flow to be measured remain absolutely constant.

In contrast, a flow meter according to claim 23 is particularly suitable good for measuring high flow velocities.

The invention is explained below with the aid of exemplary embodiments Explained in more detail with reference to the accompanying drawing. In this show: figure 1: a vertical section through a biophysicochemical arrangement which is used for Measurement of reactions catalyzed by surface-bound enzymes and for pH measurement is suitable in an intermediate stage of their manufacture; Figure 2: a vertical one Section through the active area of the membrane and an adjacent section a membrane carrier film on a greatly enlarged scale; Figure 3: a schematic Representation of an acetylcholinesterase molecule; Figure 4: a longitudinal section through a flow meter using a biophysicochemical arrangement; FIG. 5: a section through an experimental arrangement for investigation through surface-bound Enzymes of catalyzed substrate reactions; Figure 6: a graphic representation of the Conductivity of the membrane of a biophysicochemical arrangement according to FIG. 1 as a function of the time with very low concentration of the enzyme molecules on the active area the membrane and very low concentration of substrate molecules in the liquid medium; and FIG. 7: a view similar to FIG. 6, but with the The concentration of the enzyme molecules and the substrate molecules is chosen to be somewhat larger.

Figure 1 shows a bowl-shaped container 10, in the bottom Full width slot 12 is provided. In the slot 12 is a partition 14 fluid-tightly displaceable, which with its lateral The edges can also be moved in a fluid-tight manner on the side walls of the container 12 is present. For additional sealing, silicone grease can be provided at the sealing points be. In this way, through the container 10 and the partition 14, there are two compartments 16 and 18, which are each filled with an aqueous salt solution, e.g. with an aqueous, 1-molar (ionic strength) K + Cl or Na + C1 solution, which is buffered to a pH between 7 and 7.5 and to a temperature of 292 - 2960 K is set.

The partition 14 is in its upper portion with one on one Side inclined walls exhibiting the opening 20 provided; which through a thin Foil 22 made of tetrafluoroethylene is covered. The thickness of the film 22 is 25 ii, and in its center is a working opening 24 with a diameter of about 100 A, e.g. pierced with a thin needle.

On the surface of the aqueous in compartments 16 and 18 Solution is in each case a monomolecular layer 26 or 28 of a lipid, preferably L-ob -phophatidylcholine made from soybeans spread.

The lipid layer 26 is also covered with a further layer, which enzyme molecules, in the embodiment under consideration here, acetylcholinesterase, and is obtained by spreading a drop of solution 30.

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

Further details relating to the method known per se for producing membranes are in the journal Proc. Nat.

Acad. Sci. USA, Vol. 69 (1972), pp. 3561'- 3566.

The solution drop 30 contains a total of 10 6g (corresponds to 10 12mol) a tailed native acetylcholinesterase, which is produced in the electrical tissue des eels Electrophorus electricus and isolated by affinity chromatography is brought into a highly pure state. This cleaning procedure is in the magazine Biochemical and Biophysical Research Communications, Volume 79 (1977), pages 640 - 647 described in more detail. 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 section that is in no way to scale through the film 22 in the area of the working opening 24. The two can be seen Lipid molecules 34 built up lipid layers 26 and 28, which are located in the area of Combine the working opening to form the membrane 32. In the lipid layer 26 are additional the acetylcholinesterase enzyme molecules 36 incorporated. For about 10 lipid molecules 34 on average there is one enzyme molecule 36.

Figure 3 shows schematically the structure of an acetylcholinesterase molecule, as stated in the above publication in Biochemical and Biophysical Research Communications is known. Mark a "*" or a "**" different places on the enzyme molecule at which it is split by protolysis can be. The long tail 38 of the enzyme molecule 36 can bind to lipid molecules enter; catalytic subunits 40 are marked with circles, which are in the form of three tetramers.

In the biophysicochemical arrangement shown in FIG are each in one of the two compartments 16, 18 dipping kallomel electrodes 42, 44 are provided, which are connected to the terminals of an operating circuit 46.

This contains a train UB connected to a DC voltage source Series resistor 48 and a DC 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 connected. The output of amplifier 50 is thus the total impedance of the current path formed by the electrodes, the aqueous solution and the membrane assigned. This practically only depends on the impedance of the high-resistance membrane 32 away. Thus, the output signal of the amplifier 50 directly indicates the conductivity of the Membrane 32 again.

If you now also bring a small amount of unstirred into compartment 16 Acetylcholine, e.g. in the form of a 5 x 10-6- olar aqueous solution of acetylcholine + Cl-, discrete conductivity pulses with a Half-value width of 5 x 10-2 sec and one step on the K + Cl concentration of the aqueous Milieu normalized height of about 2 x 10'10 (Ohm x M K + C1 -1, as shown in FIG. 6 is. With an even closer examination of the individual conductivity impulses with higher With a temporal resolution, one can see that this in turn is a rapid succession of very rapid impulses with a half width of a few 10 represent 4 sec. The latter are presumably received when a Acetylcholine molecule comes under the influence of an acetylcholinesterase molecule and then catalytically hydrolyzed, releasing a proton. The pulse train with a total duration of 5 x 10 2 sec, on the other hand, probably corresponds to the successive one Hydrolysis of individual acetylcholine molecules belonging to a cluster, with given Solvent is the number of molecules in the cluster that come together to form a cluster is essentially constant.

Both the conductivity pulses with a half width of about 5 x 10 2 sec as well as the conductivity impulses visible with greater temporal resolution with a half-width of about 10 sec, have not yet been obtained because with large concentrations of enzyme molecules and substrate molecules was worked on.

In order to receive individual conductivity impulses, the frequency with an impact of a substrate molecule or a substrate molecule cluster on an enzyme molecule is obtained, so small that the individual Conductivity impulses in the vast majority of cases do not occur in time overlap. This frequency obviously increases with the concentration of the surface-bound ones Enzyme molecules, with the concentration of the substrate molecules distributed in the aqueous medium, with the size of the active membrane area and with the mobility of the substrate molecules in the respective liquid environment.

First and foremost about the different concentrations, but also on the choice of the temperature of the liquid located in the compartment 16 and on the size of the active membrane area can be determined in individual cases by simple experiments ensure that there are discrete individual conductivity impulses can be obtained, 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 can be easily achieved go so far as, in rare exceptional cases, an impulse already falls into the falling one Edge of a previous pulse falls. A corresponding pulse train is in Figure 7 reproduced in which the units on the coordinate axes are just as large are as in the illustration of FIG. 6.

The arrangement shown in FIG. 1 can also be used directly for measurement of the pH of the liquid in compartment 16, as the speed the hydrolysis of acetylcholine in the presence of acetylcholinesterase from each found pH depends. The frequency output by the operating circuit 46 or the display of the frequency counter 54 is a direct measure of the pH and only needed for larger deviations of the measured pH value from the pH range 7 - 7.5 with the additional use of which can be easily created once through an experiment Correction tables to be modified.

Figure 2 shows a longitudinal section through a flow meter, which has a tubular body 56 which can be switched into a water flow. This is in his Wall provided with an opening 58 which is covered by a film 60. These again has a working opening covered by a membrane, as in FIG 2 shown.

A container 62 is placed on the outside of the pipe.

It contains an aqueous salt solution, e.g. a 1 molar KCl solution into which a kallomel electrode 64 is immersed. A second kallomel electrode 66 is from that Water flow in the interior of the tubular body 56 washes around. As in FIG. 1, the electrodes are connected to an operating circuit 46 whose Output is again connected to the input of a frequency counter 54. This one has a period of the order of a second so that for each reading one large number of conductivity pulses is detected.

Upstream of the opening 58 in the wall of the tubular body 56 is another one Opening 68 is provided, into which a body 70 made of open-pore material is inserted is. The opening 68 is in communication with the inside of an outside of the tubular body 56 tightly fitted storage container 72, which is an aqueous Contains solution of acetylcholine, and possibly also of KCl, if the one to be measured Electrically conducts water very poorly.

The interior of the storage container 72 is via a pressure regulating valve 74 connected to a supply bottle 76 for pressurized inert gas so that a predetermined amount of liquid per unit of time from the storage container 72 in the flow of liquid is released, namely in its outermost peripheral layer.

The flow meter described above works as follows: When The water flowing past in a laminar flow in the tubular body 56 removes its outer layer from the body 70 acetylcholine clusters with. These can now be in the water flow Direction of flow in the perpendicular direction, i.e. from the wall of the pipe body to the pipe body axis diffuse. The slower the flow, the more acetylcholine clusters there are diffused away from the pipe body wall when the boundary layer of the water flow the Slide 60 reached, all the more fewer conductivity impulses are generated by the Frequency counter 54 registered. The reading of the frequency counter 54 is thus direct a measure of the flow velocity of the water.

For compared to the diffusion rate of the acetylcholine cluster very high flow velocities can be used to reverse the device Feeding acetylcholine clusters into the water stream so that the rate of release is assigned to the flow velocity of the water. This can be similar to a motor vehicle carburetor using a Venturi nozzle, which has a feed opening formed in the nozzle wall for the acetylcholine solution, which is connected to the storage container 72. With such a flow meter The number of those arriving at the film 60 per unit of time then also increases Acetylcholine cluster with the flow rate.

On the other hand, if one has compared it with the diffusion rate of the acetylcholine clusters To measure very small velocities of the water flow, one can use the flow meter expand according to Figure 4 by the components shown there in dashed lines, which represent a second biophysicochemical arrangement and already one above correspond to the component described. For this reason, these components are equipped with appropriate, however, additionally provided with a reference number provided with a comma.

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

The count of the frequency counter 54 thus corresponds direct the deviation of the movement of the acetylcholine clusters from the flow velocity Zero obtained completely symmetrical diffusion pattern and is thus directly the small one Assigned drift speed of the water flow.

It can be seen that the sensitivity of the flow meters described above 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 again in frequency modulation which is advantageous for noise-free and rapid signal processing provided.

The flow meter shown in Figure 4 can be with very few Modifications can also be used to measure an electric field, as acetylcholine is usually added in the form of a polar salt, e.g. acetylcholine Cl and the migration of acetylcholine ions through an external electric field in It is influenced in a similar way as by being entrained by a stream of water.

To do this, one only needs on the ends of the tubular body 56 in FIG. 4 Put on cover 78 shown in dash-dotted lines. Then through the applied external electric field again an asymmetry of the diffusion field of the acetylcholine + clusters released by body 70, so that the operating circuits 46 and 46 'emit pulses at different frequencies. A back diffusion to prevent acetylcholine clusters, which run past the foil 60 undecomposed, One can get the inside of the lid 78 with a large number of acetylcholinesterase molecules covering the supporting membrane, so that the acetylcholine clusters arriving there on will be decomposed in any case.

The field strength meter just described is used for both Investigation the direction of the field as well as to determine the field strength, for which one must first use the Tubular body 56 rotates so that as large a reading as possible is obtained on the frequency counter 54 and then note the value of the maximum reading.

With the arrangement described above, the influence is also automatically a slowdown caused by the resulting reaction products of the reaction, since both foils 60 and 60 'in are affected in the same way. After a longer measurement period, the used, acetate-rich Solution inside the tubular body 56 is discarded and replaced by a new solution, for which purpose corresponding connections are provided on the tubular body 56.

FIG. 5 shows a set-up for investigating reactions on surface-bound Enzyme molecules similar to that shown in Figure 1; corresponding parts are again provided with the same reference symbols and do not need to be repeated to be described in detail. The arrangement is ready for operation, thus reproduced after the production of the membranes formed by lipid bilayers, which are otherwise not shown in detail.

The bottom of the container 10 now has a second slot 12 ', in which a second partition 14t is just as displaceably arranged as the Partition 14 in the slot 12. The partition 14 'has an opening 20', which of a film 22 'is covered. In this respect there is complete agreement with the assembly parts already described above.

The working opening of the not shown in FIG foil 22 'is in turn covered by a membrane which is covered by a lipid double layer is formed. However, this membrane in turn carries other enzyme molecules than that Membrane of the film 22, namely those associated with them in their interaction Substrate molecules are to be investigated. These substrate molecules are located also in the aqueous medium of compartment 16. In order to prevent them directly react with the substrate molecules for the enzyme molecules of the membrane of the film 22, and to prevent substrate molecules for the enzyme molecules from the membrane of the Foil 22 reacting with the enzyme molecules of the membrane of foil 22 'is the compartment 16 divided by a wall 80, which only for the reaction products of the catalytic Decomposition of substrate molecules on the membrane of the film 22 'is permeable, e.g.

for protons.

The arrangement shown in Figure 5 thus consists of a left of the Wall 80 lying test part and a measuring part lying to the right of wall 80. Man recognizes that the test arrangement according to Figure 5 has a very simple structure and also the course of reactions of substrate molecules catalyzed by enzymes to be examined if the enzyme molecules are to be carried by membranes, which themselves show no or only a very slight change in conductivity. One can thus, among other things, investigate reactions on molecular single layers.

A temperature meter can also be used with one described above Easily realize the biophysicochemical arrangement, since the rest of them are retained Operating parameters, the mobility of the substrate molecules or - clusters in the liquid Environment depends on the temperature.

In the arrangement according to FIG. 1, this is the case with a fixed concentration from Substrate molecules and enzyme molecules, the same liquid medium and the same geometry the frequency of the conductivity impulses a measure of the temperature of the liquid Milieus in compartment 16, since the enzyme molecules 36 decompose on membrane 32 Substrate molecules through temperature-dependent diffusion from the volume of the liquid Milieus need to be replaced. The compartment 16 therefore needs when using the arrangement according to Figure 1 to be only thermally coupled to the object, its temperature should be measured.

Claims (24)

Claims 1. To changes in the environmental conditions Change in their electrical conductivity in an appealing biophysicochemical arrangement, consisting of a membrane immersed in an electrically conductive liquid medium, whose electrical conductivity depends on the surrounding environment, from the Membrane-borne enzyme molecules and substrate molecules distributed in the liquid environment, which takes part in a reaction in the medium under catalysis by the enzyme molecules participate, which gives at least one reaction product, which the electrical The conductivity of the membrane is influenced, 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 migration speed of the substrate molecules or substrate molecule clusters as well as taking into account the rate of catalytic activity by the enzyme molecules sponsored response are chosen so that individual pulse-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). 3. Arrangement according to claim 2, characterized in that the membrane consists of dipalmitoyl-phosphatidylcholine, sphingomyelin or phcsphatidylserine. 4. Arrangement according to claim 3, characterized in that the membrane consists of L - «- phosphatidylcholine obtained from soybeans. 5. Arrangement according to one of claims 1 - 4, characterized in that that the substrate is acetylcholine and the enzyme is acetylcholinesterase and that the Milieu is an aqueous medium containing ions. 6. Arrangement according to claim 5, characterized in that the acetylcholinesterase is a highly pure, native acetylcholinesterase. 7. Arrangement according to claim 6, characterized in that the acetylcholinesterase by affinity chromatography from the electrical organ of Electrophorüs Electricus is made. 8. Arrangement according to one of claims 1 - 7, characterized in that that the liquid medium additionally AIP, i.e. Adenosine triphosphate and the membrane (32) additionally contains Na + -K -ArPase. 9. Arrangement according to claim 8, characterized in that the concentration of the ATPase is about 10-10 3 molecules per molecule of the membrane material. 10. A method for producing a membrane according to any one of the claims 2 - 9, characterized in that on the surface of the liquid medium to both Sides of a carrier film, which is provided with a working opening, which initially above the free surface of the liquid medium lies, by spreading a monomolecular Lipid layer is made on one of the lipid layers by diluting one side Enzyme solution, the enzyme molecules are applied and then the carrier film with their Working opening is slowly moved into the milieu. 11. The method according to claim 10, characterized in that as Lipid L- «-phosphatidylcholine is used and an aqueous medium to pH 7 - 7.5 buffered aqueous K Cl or Na Cl solution is used, its temperature is between 292 and 296 ° K. 12. Biological test set-up to investigate the reaction processes on enzymes carried by the surface of a membrane, characterized in that that they have a biophysicochemical arrangement according to one of claims 1-9 or one biophysicochemical arrangement which is produced according to claim 10 or 11, which forms a portion of a partition (14) of a container (10) with the liquid medium is filled and its compartments (16, 18) with a conductivity meter (46, 52 or 54) are connected, and that the enzyme to be examined carries Membrane is immersed in that compartment (16) which is connected to the biophysicochemical Arrangement belonging to the enzyme molecules. 13. Experimental arrangement according to claim 12, characterized by a only permeable to the products of the reaction controlled by the enzyme to be examined Partition wall (80), which between the biophysicochemical arrangement and that Milieu is arranged in which the membrane with the enzyme to be examined is immersed. 14. pH meter, characterized in that it is a biophysicochemical Arrangement according to one of claims 1-9 or a biophysicochemical arrangement, which is made according to claim 10 or 11, and one having two compartments (16, 18) Has container (10) for the liquid medium, whose partition delimiting the compartments (14) formed in part by the membrane (32) of the biophysicochemical arrangement is that the compartments (10, 18) of the loading holder (10) with a conductivity meter (46, 52 resp. 54) are connected and that that compartment (16) of the container that communicates with the enzyme molecules of the biophysicochemical arrangement, at the same time is in communication with the liquid whose pH is to be measured. 15. pH meter according to claim 14, characterized in that the compartment (16), which communicates with the enzyme molecules of the biophysicochemical arrangement, with the liquid via another membrane permeable to hydrogen ions communicates whose pH value is to be measured. 16. Temperature meter, characterized in that it is a biophysicochemical Arrangement according to one of claims 1 - 9 or a biophysicochemical arrangement, which is made according to claim 10 or 11, and one having two compartments (16, 18) Has a container (10) for the liquid medium, of which the compartments (16, 18) delimiting Partition (14) partly through the membrane (32) of the biophysicochemical arrangement is formed 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), which communicates with the enzyme molecules of the biophysicochemical assembly, can be thermally coupled to that object whose temperature is being measured target. 17. Electrical field strength meter, characterized in that it a biophysicochemical arrangement according to any one of claims 1-9 or a biophysicochemical one An arrangement made according to claim 10 or 11 and a two compartment (56, 62, 62 ') having container made of dielectric material for the having a liquid medium, the dividing wall delimiting the compartments partly through the membrane of the biophysicochemical arrangement is formed that the compartments of the Container with a conductivity meter (46, 46 ', 54) are connected that at least that compartment (56) of the container which contains the enzyme molecules of the biophysicochemical Arrangement communicates, is movable in the field-filled space, and that the substrate molecules exist in ionized form in a liquid medium. 18. Surveyor according to claim 17, characterized in that two biophysicochemical arrangements (58 - 66; 58 '- 66') symmetrical on both sides a source (68-76) for ionized substrate molecules are arranged and the two Operating circuits (46, 46 ') assigned to biophysicochemical arrangements on the output side are connected to a difference computer (54). 19. Flow meter, characterized in that it is a biophysicochemical Arrangement according to one of claims 1 - 9 or a biophysicochemical arrangement, which is made according to claim 10 or 11, a tubular body (56) for the to be measured Liquid flow and a container placed on the wall of the tubular body (56) (62) has that the membrane of the biophysicochemical arrangement is one in the partition Opening (60) provided between the tubular body (56) and the container (62) at least for the Part that closes the interior of the tubular body (56) and the interior of the container (62) is connected to a conductivity meter (46, 54) and that upstream of the Membrane of the biophysicochemical arrangement a device (68-76) for feeding of substrate molecules and possibly ions provided in the liquid flow to be measured is. 20. Flow meter according to claim 19, characterized in that the feed device (68-76) feeds the substrate molecules and possibly the ions into the Feeds the edge layer of the liquid flow, which is guided in a laminar manner in the tubular body. 21. Flow meter according to claim 20, characterized in that the feed device communicates with the boundary layer of the liquid flow has open-pore body (70) which is connected to the interior of a storage container which is a solution containing substrate molecules and possibly ions contains. 22. Flow meter according to claim 21, characterized by a device (74, 76) for maintaining a constant overpressure in the storage container (72). 23. Flow meter according to claim 20, characterized in that the feed device is traversed by the boundary layer of the liquid stream Has Venturi tube, which one with a storage container for substrate molecules and optionally having ion-communicating opening in the nozzle wall. 24. Flow meter according to one of claims 19-23, characterized in that a second biophysicochemical arrangement upstream of the feed device (58 '- 66') and the outputs of two of the two biophysicochemical Arrangements (58 - 66 or 58 '- 66') of assigned operating circuits (46, 46 ') are connected to the inputs of a differential computer (54).