JPS6140342B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a device for measuring chloride ions in biological fluids. At present, there are hardly any suitable analytical methods that can directly measure chloride ions in whole blood. Methods currently used to measure chloride ions in biological fluids include coulometric analysis and ion electrode methods. Coulometric analysis requires coulometric analysis to be performed after diluting blood sludge or serum, in which blood cell components have been separated from whole blood, with an electrode solution, and Cl ions cannot be directly measured in whole blood. The ion electrode method uses an ion electrode that selectively responds to Cl ions, specifically a liquid membrane electrode or a silver chloride electrode. Liquid film electrodes are made by impregnating a porous material with an appropriate solution to form a liquid film electrode using surface tension, so it has insufficient mechanical strength and cannot withstand cleaning.
It also has poor selectivity for Cl ions and has a short lifetime.
Furthermore, since silver chloride electrodes are solid, they are strong enough and have good selectivity for Cl ions, but they are affected by proteins and other coexisting substances that coexist in biological fluids, making accurate analysis of Cl ions impossible. This method is difficult to use when directly measuring Cl ions in whole blood for practical purposes.

Therefore, the present invention allows whole blood to be used as a sample without being affected by coexisting substances other than Cl ions in biological fluids.
Practical method for measuring Cl ions in biological fluids
This was done with the purpose of providing an ion measuring device. Of course, it can also be applied to serum, urine, or other biological fluids, and is extremely useful in clinical tests.

First, the principle of the present invention will be explained. The present invention is based on an electrochemical analysis method called potentiometry. Since the purpose of the present invention is to measure Cl ions in biological fluids, it belongs to potentiometry using an ion-selective electrode, and as an ion-selective electrode, it is necessary to use an ion electrode that responds only to Cl ions. Become. However, the monovalent anion (anion) in biological fluids is represented by Cl ^- , and although other anions do not exist,
Anions with divalent or higher valences and anions with properties significantly different from Cl ^- (e.g. H ₂ PO ₄ ^- , COO ^- , HCO ₃ ^- ,
OH ^-, etc.), the characteristic of selectively responding only to Cl ^- is not necessary. In other words, even among monovalent anions, NO ₃ ^- , F ^- , Br ^- , I ^-, etc. do not exist in normal biological fluids, and even if they do exist, the amount is extremely small compared to Cl ^- .
This is one of the principles of the present invention, which allows for a wider range of ion electrode choices. From the standpoint of the present invention
Although the electrode has considerable selectivity for Cl ions, it is rather undesirable to have an electrode that responds considerably to divalent anions. This is because biological fluids contain a considerable amount of divalent anions such as carbonate groups and phosphate groups. Another principle of the present invention is the use of an ion exchange membrane with a homogeneous membrane structure. This point will be discussed next.

Ion exchange membranes are synthetic resin solid membranes that have high permeability selectively to anions or cations.Cation exchange membranes have significantly higher permeability to cations than anions; Anion exchange membranes have these properties. These ion exchange membranes are used as diaphragms in electrolytic cells to prevent electrolytically produced substances from mixing with electrolytic stock solutions and lowering electrolytic efficiency, or as ion filters in seawater concentration and desalination. These methods involve passing an electric current through the membrane to drive the ions and sieving them out, and the quality of the ion exchange membrane is evaluated by the method of electrical conduction through the membrane. What happens when such an ion-exchange membrane is placed at the boundary between solutions with different concentrations of ions permeable to the membrane and the membrane potential is precisely observed without passing any current? In this case, a potential related to specific ions in the solution is generated at the interface between the membrane and the solution. Moreover, the generation of this potential changes extremely quickly even in response to changes in the ionic activity in the solution (for example, the over-liquid response speed is less than 100 ms), and as long as the ionic activity in the solution is constant, the electron It has been found that it is extremely stable as long as no movement is allowed (i.e., no current is applied). The magnitude E of the generated potential corresponds well to Nernst's theoretical formula, and is given by E=E0+0.0591logai (volts), where ai is the ratio of the ion activities of both solutions. The coefficient of 0.0591 is the value of the field of monovalent ions at 25°C, and if the ion activity on one side of the membrane changes by a factor of 10, a potential change of E≈59mV appears. Therefore, if the ion activity of the solution on one side of the ion exchange membrane is kept constant, the ion exchange membrane can be used as an ion electrode membrane. This is the second principle of the invention.

There are two types of ion exchange membranes: anion exchange membranes and cation exchange membranes, but although these membranes usually have selectivity for anions and cations, they have a high selectivity for monovalent, divalent, etc. ions among ions of the same sign. is generally scarce. However, ion exchange membranes that are selective to the valence of ions have been made using various special manufacturing methods (for example, Japanese Patent Publication No. 8515-1983 and Japanese Patent Publication No. 36-1982).
15258), several types of ion exchange membranes for monovalent anions are currently on the market, and by slightly changing the composition of the ion exchange groups that constitute them, more specific ion exchange membranes for monovalent anions are available. This type of membrane was originally manufactured by focusing on the selective permeability of monovalent anions during electrolysis, as described above, but this type of membrane has a selectivity for ion groups. The exchange membrane can constitute an ion electrode that responds to a specific ion group, and according to the above-described first principle of the present invention, it can be used as an ion electrode for measuring Cl ions in biological fluids. can.

FIG. 1 shows the basic structure of an apparatus that uses the above-mentioned ion exchange membrane to form an ion electrode and measures the ion concentration of a solution. 1 is an indicator electrode tank, 2 is a reference electrode tank, and 3 is a sample solution whose ion concentration is to be measured. An ion exchange membrane 4 is placed on the bottom of the indicator electrode tank 1, and an electrode internal solution 5 is placed in the indicator electrode tank 1. so that it forms the boundary between
The electrode 6 is inserted into the electrode internal liquid 5. In this case, the electrode solution 5 is a solution containing a certain amount of chlorine ions, and a certain interfacial potential is generated between the electrode 6 and the electrode internal liquid 5. For example, a solution of KCl at a constant concentration is used as the electrode internal solution 5, and a silver wire with a AgCl coating formed on the surface is inserted into this solution as the electrode 6. AgCl
generates an electrode potential in response to Cl ions,
Since the KCl concentration in the tank 1 is constant, the electrode potential is constant. The reference electrode tank 2 contains the same solution as the solution 5 in the indicator electrode tank 1 as an electrode internal solution 7, and the tank 2 is provided with a liquid junction 8 between the solutions 3 and 7. Reference electrode 9 has the same structure as electrode 6, so the potential difference between electrode 9 and electrode solution 7 is equal to the potential difference between electrode 6 and electrode solution 5, and the potential difference between electrodes 6 and 9 is equal to the potential difference between electrode 6 and electrode solution 5. When measuring , the potential differences generated by the electrodes 6 and 9 with respect to the electrode solutions 5 and 7 are canceled out and do not appear in the measurement. Since the electrode solution 7 is electrically connected to the solution 3 through the liquid junction 8, both solutions have the same potential (strictly speaking, a liquid-liquid interface potential occurs, but it is small and can be ignored), and the membrane A potential difference is generated on both sides of 4, which is proportional to the logarithm of the ratio of the monovalent anion activities of the solution 3 and the electrode solution 5.
This potential difference is measured as the potential difference between electrodes 6 and 9.

FIG. 2 shows an example in which the characteristics of an anion exchange membrane as an ion electrode were measured using an apparatus configured as described above. A 0.1 MoI/l KCl solution was used as the electrode internal solution, and the measurement solution contained 0.001 MoI/l acetic acid and KCl.
Solutions with various concentrations were used. Dotted line A is a straight line in the range of 10 ^-2 to 10 ^-1 MoI/l due to the relationship between Cl ion activity and electrode potential difference when a normal monovalent anion-permeable ion exchange membrane is used as the ion-selective electrode membrane of the present invention. There is a relationship, and a change of 10 times the Cl ion activity results in an electrode potential of approximately 58 mV, which is approximately in agreement with Nernst's theoretical formula. Solid line B shows the relationship between the Cl ion activity and the electrode potential difference when a cation exchange group is introduced in addition to an anion exchange group into the membrane, and the amount is set to ^the optimum ^condition . l has a good linear relationship up to the C range. In curve A, the influence of acetate ions (COO) ^- can be ignored in the range where Cl ion activity is high, but in areas where Cl ion activity is low, the influence of acetate ions becomes effective and the electrode potential becomes approximately constant. However, the effect of such coexisting ions can be improved by introducing an appropriate amount of cation exchange groups in addition to anion exchange groups (COO) ^-, almost all divalent or higher anions, HCO ₃ ^- , H ₂ PO ₄ ^- ,
The effects of experiments using monovalent anions such as Br ^- and F ^- were observed. In addition, a membrane in which a thin film layer formed by polymerizing amino groups, aldehyde groups, methylol groups, etc. on a general anion exchange membrane as described in claim 3 also improves the selectivity between anions and straightens the membrane. Improved range.

This completes the basic explanation of the present invention, and the present invention will be explained below using examples. FIG. 3 shows an embodiment of the present invention. Reference numeral 10 denotes a main body block through which a measuring liquid channel 11 passes through from left to right. A recess 12 is formed in the upper surface of the main body 10, and a part of the flow path 11 has an opening 13 at the bottom of the recess 12. An anion-selective electrode membrane 4 is stretched over the bottom of the recess 12 to cover the opening 13, and a plug 14 is inserted and screwed into the recess 12. The plug 14 has a vertical through hole 1 formed at a position facing the central opening 13, and this corresponds to the indicator electrode layer 1 in FIG. The lower end surface of the plug 14 presses the anion-selective electrode membrane 4 against the bottom of the recess 12, so that the electrode membrane 4 is connected to the through hole 1 and the channel 11.
It also serves as a gasket to provide a liquid-tight seal between the two. A 0.1 mol/l KCl solution is placed in the through hole as an electrode internal solution 5, and an indicator electrode 6 is inserted into the through hole. The electrode has an AgCl layer formed on the surface of the silver wire. Reference numeral 15 denotes a lid for the through hole 1 of the plug 14. A hole 16 is bored to the left of the recess 12 and perpendicular to the channel 11, and a solution 7 containing 0.1 mol/l of KCl, which is the same as the electrode solution 5, is poured into the channel 11 through this hole. The measurement liquid flows into the flow path 11 from the right side, merges with the KCl solution flowing in from the hole 16, and flows out from the left end of the flow path 11. A reference electrode 9 is inserted into the hole 16 from the side. This electrode is the indicator electrode 6
Similarly to , this is a silver wire with an AgCl layer formed on it.
The liquid flowing through the hole 11 is a reference electrode liquid, and the reference electrode 9 is contained in this electrode liquid and does not come into contact with the measuring liquid. Reference numeral 17 denotes a pump for feeding the measurement liquid, and a syringe needle 18 is attached to the suction side of the pump.
Reference numeral 19 is a container having rubber membranes 20, 20' stretched on both upper and lower surfaces and communicates with the reference liquid reservoir 21. Rubber membrane 2
A straight cut is made at 0 and 20', and these cuts are held together by the elasticity of the rubber, so that the liquid inside will not leak. The tip of the injection needle 18 usually has a rubber membrane 2 as shown in the figure.
The pump 17 passes through the notch 0 and is located between the rubber membranes 20 and 20', and the pump 17 sucks the reference liquid from the reference liquid reservoir 21 and sends it toward the channel 11. Standard solution is 0.1
It is a KCl solution with a concentration of 0.1 mol/l, and while this liquid is flowing through the channel 11, both the ion-selective electrode membrane 4 and the reference electrode 9 are in contact with the KCl solution with a concentration of 0.1 mol/l, so the electrode 6 , 9 is 0, and the concentration indicator meter should read 0.1 (mol/l), so adjust the meter during this time.
A specimen 23 contained in a container is sent below the container 19. The specimen may be whole blood taken from the subject, blood plasma from which blood cell components have been removed, serum from which fibrin has been removed, or cerebrospinal fluid. Lower the injection needle 18 and remove the lower rubber membrane 20'.
When the tip is inserted into the removed body 23 through the cut, the pump 17 aspirates the specimen 23. Thereafter, the injection needle 18 is returned to its original position. Then, temporarily sample 23
is sent to the flow path 11, and then the standard solution starts flowing through the flow path 11 again. While the specimen 23 is in contact with the membrane 4, the meter indicates the Cl ion concentration of the specimen. Since samples are sent one after another, the above operation is repeated each time. While the sample is not being sent to the flow path 11, a standard solution is flowing through the flow path 11 to wash the membrane 4 and remove residual deposits from the sample from the previous measurement. Since the pump 17 sucks air and sends it to the channel 11 while the injection needle 18 passes through the rubber membrane 20' and reaches the specimen 23 and returns, the pump is stopped during this period. Or syringe needle 1
When the membrane 8 penetrates the rubber membrane 20', air is sucked in for a short period of time, so that air bubbles pass through the flow path 11, and the cleaning effect of the standard solution on the membrane 4 is enhanced.

FIG. 4 shows an experimental example for confirming the performance of the above-mentioned device, in which the serum Cl ion concentration was compared with the results measured by coulometric analysis, which is one of the conventional serum Cl ion concentration measurement methods. The horizontal axis shows the coulometric values measured by the analytical method (coulometry), and the vertical axis shows the measurement results according to the present invention.

Both values are distributed on a straight line at 45° C., indicating that the device of the present invention operates correctly.

Finally, the effects of the present invention will be described. 4th above
As is clear from the results shown in the figure, regarding the Cl ion concentration in serum, the measurements made by the device of the present invention and the results obtained by the conventional coulometric analysis method are in exact agreement. On the other hand, in the conventional coulometric analysis method, serum is diluted with an electrolytic solution and analyzed, so it is not possible to directly measure whole blood, and the measurement lacks rapidity. In contrast, the device of the present invention can also directly measure whole blood, and the measurement can be performed with high efficiency. Furthermore, when the ion electrode using an ion exchange membrane in the apparatus of the present invention is compared with the conventional silver chloride electrode having Cl ion selectivity, results as shown in FIG. 5 are obtained. The solid line in Figure 5 is an electrode using a monovalent anion exchange membrane, and the dotted line is an electrode using a monovalent anion exchange membrane.
The difference in reproducibility between AgCl solid membrane electrodes is shown.
The AgCl solid membrane electrode is a solid membrane electrode made by compacting and sintering powders of 9 parts AgCl and 1 part AgCl. The test solution is human serum. Using the equipment configuration shown in Figure 3, the same test solution was sent at one-minute intervals, and the Cl concentration was measured as the standard solution for calibration.
NaCl aqueous solutions of 100 meq/l and 140 meq/l were used. When according to the present invention, baseline (indication of Cl ion concentration of 100 meq/l NaCl aqueous solution)
is constant, and the serum Cl ion concentration shows a constant value every time. On the other hand, when AgCl solid membrane electrodes were used, both the baseline and the pulse height, which indicates the serum Cl ion concentration, gradually decreased each time, indicating that the cleaning effect of the standard solution was not sufficiently exerted. There is. This is thought to be because proteins in the biological fluid adhere to the electrode surface. The same phenomenon as in the AgCl solid membrane electrode is observed in the conventional liquid membrane type ion electrode. In addition, 2 with different Cl ^- concentrations
When the sample solution of the seed was replaced instantaneously and changes in the indication response speed were automatically recorded, it was found that the method of the present invention with the electrode structure shown in Figure 3 caused a stable indication change in about 1 second as shown in Experiment A. Admitted. In the same way, in the case of AgCl electrode, about 5
The response time was seconds, and it was relatively unstable. On the other hand, the response speed of the ion-selective electrode according to the present invention can be increased by changing the structure and making the sample liquid sufficiently fast.
It has also been experimentally confirmed that it is less than 10mssec.

As mentioned above, the present invention enables direct measurement of Cl ions in whole blood, has excellent reproducibility of measurement, is quick,
In these respects, it is possible to provide a biological fluid Cl ion measuring device that is superior to any conventional biological fluid Cl ion measuring method.

[Brief explanation of the drawing]

Fig. 1 is a schematic side view of an apparatus for explaining the principle of the present invention, Fig. 2 is a graph showing the characteristics of an ion exchange membrane electrode, Fig. 3 is a longitudinal side view of an embodiment of the apparatus of the present invention, and Fig. 4 is Graph showing agreement of measurement results obtained by the device of the present invention and by coulometric analysis, No. 5
The figure is a graph showing measurement records by the apparatus of the present invention, and FIG. 6 is a graph showing the transient response speed of the electrode of the present invention and the conventional electrode.

Claims (1)

[Claims] 1. A monovalent anion exchange resin membrane with a homogeneous membrane structure is used as a boundary membrane, and one side of the membrane contains an internal solution with a constant chloride ion activity, and the other side contains a biological sample solution with an unknown chloride ion activity. By inserting a reference electrode with a constant unipolar potential into each solution and measuring the potential difference between both reference electrodes and detecting the potential difference between both sides of the limiting membrane, the chloride ion activity in the biological sample solution is determined. A device for measuring chlorine ions in biological fluids, which measures the amount of chlorine ions in biological fluids. 2. The device for measuring chloride ions in a biological fluid according to claim 1, which uses a membrane mainly composed of anion exchange groups and further introduced cation exchange groups through cross-linking as the boundary membrane. 3. The bioreactor according to claim 1, which uses an anion exchange resin membrane having an anion exchange group as a base and a thin polymer layer formed on the surface thereof to improve permselectivity between anions as a boundary membrane. Chlorine ion measuring device in body fluids.