JPS606639B2 - Method and device for measuring uric acid - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method and apparatus for measuring uric acid that indirectly measures the concentration of uric acid contained in a sample such as blood from the amount of change in conductivity of an absorption liquid.

In recent years, advances in biomedical measurements have enabled continuous telemetry of physiological variables such as body temperature, gastrointestinal pressure, blood pressure, rate of respiration, and biological potential, as well as in vivo p02, pC02. , blood pH and electrolytes,
In addition, measurement methods and devices for continuously measuring the pH of the stomach have been developed.

As one such biomedical measurement, uricase enzyme is applied to uric acid in the specimen to generate the enzymatic reaction of the following formula {1], and the uric acid concentration in the specimen is determined from the change in the amount of ultraviolet light absorbed by uric acid. There is a so-called uricase ultraviolet absorption method that measures **. Uric acid + 2 days 20102 uricase 10C○
212th 2○2. ．． …． [1} However, in spectrophotometric methods such as the above-mentioned uricase ultraviolet absorption method, if the sample contains serum proteins, it is necessary to remove the proteins, and when the blood is whole blood, The disadvantage was that it was impossible to measure. Another drawback is that it takes a long time to develop color, making it impossible to quickly measure uric acid in a specimen. The present invention was made in view of these drawbacks, and its purpose is to quickly and accurately measure uric acid contained in specimens such as whole blood without requiring pretreatment operations such as protein removal. An object of the present invention is to provide a method and device for measuring uric acid.

Hereinafter, the present invention will be explained in detail using figures.

FIG. 1 is a configuration explanatory diagram showing an embodiment of the present invention. In the figure, 1 is a flow cell, and a B high film 2 made of, for example, PTFE or silicone rubber thin film, which does not allow ions to pass through but allows gas to pass therethrough. The first chamber 3 and the second chamber 4 are adjacent to each other. Also, the first
The chamber 3 is provided with an inlet 5 and an outlet 5', and the second chamber 4 is provided with an inlet 6 and an outlet 6'. Furthermore, 21
21 is an inlet into which a carrier made of a solution containing boric acid buffer, NaCl, ETDA benzoic acid, etc. is introduced; 23 is an outlet for the carrier, etc.;
A sample injector 27 for injecting the test substance and an immobilized enzyme 22 in which uricase enzyme is immobilized on a carrier such as glass beads or a nylon tube are located in the middle of the flow path from the flow path to the inlet 6 of the second chamber 4. is provided. Furthermore, a reagent addition port 28 is provided in the middle of the flow path from the immobilized enzyme 22 to the introduction port 6 of the second chamber 4 for adding an acidic reagent such as a 0.1N solution of HCI or EN03. It is being On the other hand, 24 is an absorption liquid introduction port,
It is installed so that the absorption liquid 26 (for example, an alkaline solution made of 0.00 state KOH or the like) in the container 25 can be sucked. Reference numeral 10 denotes an absorption liquid discharge port, and in the middle of the flow path from the outlet 5' of the first chamber 3 to the absorption liquid discharge row 01, there is an electrode 7 connected to, for example, a power source 8 and a galvanometer 9. ,7′
For example, it is attached to a pipe with a diameter of 1 skin/m, and the outlet port 5'
It is provided nearby to detect the conductivity of the absorbing liquid discharged from the outlet 5'. In the embodiment of the present invention consisting of the above composition, the carrier introduced from the introduction port 21 includes the immobilized enzyme 22, the introduction port 6, the second chamber 4,
It flows through the outlet 6' and is discharged from the outlet 23.

Further, the absorption liquid 26 sucked from the inlet 24 flows through the inlet 5, the first chamber 3, and the outlet 5', and flows through the outlet 1
Ejected from 0. By the way, sample injector 2
When the analyte is injected into the carrier from 7, the analyte is carried by the carrier and reaches the immobilized enzyme 22.
) It undergoes an enzymatic reaction as shown in the formula to produce C02 and stop02. At this time, the pH of the boric acid buffer in the carrier is 8.5.
Therefore, the carbonic acid generated from the above C02 becomes bicarbonate ion HC03-. However, reagent addition port 2
When the acidic reagent from 8 is added into the carrier and the pH of the mixed liquid containing the carrier etc. becomes 1 or less, most of the bicarbonate ions HCO3- become CO2 gas. In this way, the uric acid contained in the subject is converted into CO2 gas, which is transported by the carrier from the inlet 6 to the second chamber 4 in the flow cell 1. In the second chamber 4, the C02 gas in the carrier passes through the bulkhead 2 in proportion to the partial pressure, reaches the first chamber 3, and reacts with the absorption liquid in the first chamber 3 as shown in the following formula (2). Due to this reaction, the conductivity of the absorption rate changes, and the conductivity changes from C02 12KOH→K2C03 10days 20...
- [21 changes are detected by the electrodes 7, 7'.

From the amount of conductivity change detected by the electrodes 7, 7' in this manner, uric acid in the subject is indirectly quantified through predetermined signal processing (not shown). 2 is an exploded perspective view of a flow cell according to an embodiment of the present invention, and FIG. 3 is an assembled perspective view of FIG. 2. In FIGS. 2 and 3, 11 and 17 are blocks, each having an inlet 11a.
, 17a and a lead-out roll lb, 17b are provided.
2 and 15 are gaskets for the first chamber and the second chamber, respectively.
Cavities 13, 16 forming chambers are provided. Further, 14 is a diaphragm that does not allow ions to pass through but allows gas to pass through. In the flow cell having the above-mentioned configuration, the carrier containing the analyte etc. is introduced from the inlet 17a, passes through the cavity 16 and is led out from the outlet 17b, and the absorption liquid is introduced from the inlet 11a and passes through the cavity 13. It is derived from the derivation unit 11b. In addition, C02 contained in the carrier flowing through the cavity 16
The gas passes through the diaphragm 14 and reaches the cavity 13.
Reacts with the absorption liquid flowing through 3. According to the embodiments of the present invention as described in detail above, there is an advantage that uric acid contained in a subject can be measured quickly and accurately without requiring pretreatment operations such as protein removal.

Furthermore, FIG. 4 is a configuration explanatory diagram showing another embodiment of the present invention. In the figure, reference numeral 29 denotes an absorption liquid inlet port, and Mikawama sample injector 27 injects the specimen into the respective carriers. This is a sample switching valve that supplies the sample by switching the flow path.

In addition, in Figure 4, symbols with the same numbers as in Figure 1 (for example, 5'a, 5'b are symbols with the same numbers as 5') are used with the same meaning, and duplicate explanations will be given here. is omitted. In another embodiment of the present invention having the above configuration, a part of the carrier introduced from the introduction port 21 is connected to the introduction port 6a,
It is discharged from the outlet 23a via the second chamber 4a of the flow cell la and the outlet 6'a, and the rest is the immobilized enzyme 22, the inlet 6b, the second chamber 4b of the flow cell lb,
It is then discharged from the discharge port 23b via the discharge port 6'b. In addition, a part of the absorption liquid 26 in the container 25 is introduced from the absorption liquid introduction port 24, and the absorption liquid 26 is introduced from the introduction port 5b and the flow cell lb.
The liquid flows through the first chamber 3b of the flow cell la and the outflow port 5'b, and is discharged from the discharge port 10b, while the other part is introduced from the absorption liquid inlet 29, flows through the inlet 5a, and the first chamber 3a of the flow cell la. , and flows through the outlet 5'a, and the outlet 10
It is discharged from a. By the way, sample injector 2
A predetermined amount of the analyte injected from step 7 is released. - When the analyte is supplied to the first carrier channel leading to the cell la via the sample sliding valve 30, the analyte is carried by the carrier, but when an acidic reagent is added from the reagent addition port 28a,
If carbonate ions or bicarbonate ions are contained in the above-mentioned subject, these ions are converted to CO2 gas. Thereafter, the C02 gas and the analyte are carried by the carrier to the second chamber 4a in the flow cell la, and only the C02 gas passes through the diaphragm 2a in proportion to the partial pressure and reaches the first chamber 3a.
It reacts with the absorption liquid as shown in the formula (①) above. The reaction causes a change in the conductivity of the absorption liquid in the first chamber 3a, and this change in conductivity is detected by the electrodes 7a, 7'a. Furthermore, after carbonate ions and bicarbonate ions are removed from the specimen in the second chamber 4a of the flow cell la as described above, the specimen is transported to the carrier again and is discharged from the discharge port 23a via the discharge port 6'a. Ru. On the other hand, when a predetermined amount of the analyte injected from the sample injector 27 is supplied to the second carrier channel leading to the immobilized enzyme 22 via the sample switching valve 30, the analyte is carried by the carrier. Immobilized enzyme 22
is reached, and undergoes an enzymatic reaction as shown in formula '1' above, resulting in C0
2 or day 202. At this time, since the pH of the boric acid buffer in the carrier is 8.5, the carbonic acid generated from the above C02 becomes bicarbonate ion HC03-. Further, when an acidic reagent is added into the carrier from the reagent addition port 28b and the pH of the mixed liquid containing the carrier etc. becomes 1 or less, most of the bicarbonate ions HC03- become C02 gas. Therefore, the urea contained in the sample is C0
The gas is converted into two gases and transported by a carrier from the inlet 6b to the second chamber 4b in the flow cell lb. The second chamber 4b
In this case, the C02 gas in the carrier permeates through the diaphragm 2 in proportion to the partial pressure, reaches the first chamber 3b, and reacts with the absorption liquid as shown in Equation 21 above. Due to this reaction, the conductivity of the absorption liquid in the first chamber 3b changes, and this change in conductivity causes the electrodes 7b, 7'b to
detected by. Therefore, from the amount of change in conductivity of the absorption liquid detected by the electrodes 7b and 7'b, the electrode 7a
, 7'M When a predetermined calculation process (not shown) is performed to subtract the amount of change in conductivity of the absorbent detected, it is determined that carbonate ions and bicarbonate ions are contained in the carrier and the sample. uric acid in the sample is measured regardless of these ions. According to other embodiments of the present invention as described in detail above, even when the analyte contains carbonate ions or bicarbonate ions, or even when the carrier contains carbonate ions or bicarbonate ions, This method has the advantage that uric acid in a subject can be measured quickly and accurately without being interfered with by these ions.

[Brief explanation of drawings]

FIG. 1 is a configuration explanatory diagram showing an embodiment of the present invention, FIGS. 2 and 3 are exploded perspective views and assembled perspective views of a flow cell according to an embodiment of the present invention, and FIG. 4 is another embodiment of the present invention. FIG. 1, la, lb...flow cell, 2, 2a, 2b...
...Diaphragm, 3, 3a, 3b...First chamber, 4,
4a, 4b... 2nd chamber, 5, 5a, 5b, 6,
6a, 6b, 21, 24, 29...Inlet, 5', 5'
a, 5'b, 6', 6'a, 6'b...outlet, 7, 7
a, 7b, 7', 7'a, 7'b... Electrode, 10, 10
a, 10b, 23...discharge port, 25...container, 26...・
...Absorption liquid, 27...Sample injector,
28, 28a, 28b... Reagent addition port, 30...
...Sample switching valve. ＾ z ′Nyuuta J Diagram 71 Corner 4th Box

Claims (1)

[Scope of Claims] 1. A means for causing an enzymatic reaction by causing uricase to act on uric acid contained in an object to be detected; means for converting the carbon dioxide gas into a basic absorption liquid by passing the carbon dioxide gas through a diaphragm that does not allow ions to pass through but allows the gas to pass through the membrane, and detecting the amount of change in conductivity of the basic absorption liquid. A method for measuring uric acid, comprising: indirectly measuring uric acid in a subject from the amount of change in conductivity of the absorption liquid. 2. A first chamber that is provided with an inlet and an outlet through which a basic absorption liquid that absorbs carbon dioxide gas is introduced and taken out, respectively, and constitutes a flow path on one side of the flow cell, and a carrier containing a sample is introduced and taken out, respectively. A second chamber is provided with an inlet and an outlet, and is separated from the first chamber via a diaphragm that does not allow ions to pass through but allows gas to pass therethrough, and constitutes a flow path on the other side within the flow cell; An immobilized enzyme having immobilized uricase enzyme that acts on uric acid in a sample to cause an enzymatic reaction, and a reagent addition port provided in the middle of a flow path from the immobilized enzyme to the inlet of the second chamber. and a sample injector provided near the inlet of the immobilized enzyme to inject a predetermined amount of the analyte, and an electrode provided near the outlet of the first chamber to detect the conductivity of the absorption liquid. . A uric acid measuring device that indirectly measures uric acid in a subject from the amount of change in conductivity of the absorption liquid. 3. A first chamber that is provided with an inlet and an outlet through which a basic absorption liquid that absorbs carbon dioxide gas is introduced and extracted, respectively, and constitutes a flow path on one side of the flow cell, and a first chamber in the vicinity of the outlet of the first chamber. An electrode is provided to detect the conductivity of the absorption liquid, an inlet and an outlet are provided through which a carrier containing the analyte is introduced and taken out, respectively, and the first chamber is separated from the first chamber by a diaphragm that does not allow ions to pass through but allows gas to pass through. A second chamber that is separated by a second chamber and constitutes a flow path on the other side of the flow cell, and a reagent addition port provided near an inlet of the second chamber are disposed upstream of the immobilized enzyme, and A sample switching valve is added to supply the analyte injected from the sample injector to each carrier channel by channel switching, and the absorption detected by two electrodes upstream and downstream of the immobilized enzyme is The uric acid measuring device according to claim 2, which measures uric acid in a subject based on the difference in the amount of change in conductivity of the liquid.