JPH0762652B2 - Fluorescence analysis method and apparatus - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to a fluorescence analysis method for a liquid sample and an analysis apparatus therefor, and more particularly, to in-line measurement of a trace amount of an analysis substance in a liquid sample containing a coexisting substance. The present invention relates to a fluorescence analysis method and apparatus suitable for measurement in.

[Conventional technology]

The fluorescence analysis method is widely used as a highly sensitive analysis method for trace substances. Since this analysis method uses light for input and output,
In-situ analysis is possible through the optical window, and there is an advantage that the optical fiber can be used for remote analysis at a position apart from the main body of the analyzer.

Since it has high sensitivity and in-situ analysis is possible, it can be used as a process monitor for various chemical plants. Furthermore, since remote analysis is possible, it also has the advantage that process monitoring can be performed even when a person cannot approach the analysis site or the analyzer body cannot be installed. This is an advantage because light is used for input and output. For this reason, the possibility of using it as an in-line analyzer in solution at a nuclear fuel reprocessing plant is being investigated. (Fuel Cycles for the 80's,
DOE CONF-800943)).

Especially in reprocessing, it is indispensable to shift from the current batch operation to continuous dissolution operation to achieve higher operating rates and lower costs. In that case, in-line analyzers for uranium and plutonium concentrations as process monitors are required. Is an essential technology. In particular, on the waste liquid side such as the solution on the FP (fission product) side after the co-decontamination process, it is necessary to establish in-line analysis technology from the viewpoint of safeguards.

[Problems to be solved by the invention]

However, in a system containing a large amount of coexisting substances, such as a nuclear fuel reprocessing solution, the fluorescence condensing efficiency decreases due to absorption / attenuation of excitation light / fluorescence by the coexisting substances and fluorescence quenching due to coexisting substances. However, there is a problem that the analysis sensitivity cannot be sufficiently obtained. Further, fluctuations in the concentration of coexisting substances cause a decrease in analysis accuracy due to the above mechanism. Further, when the fluorescence of the analyte is temperature dependent, the temperature fluctuation also causes a decrease in analysis accuracy.

For this reason, conventionally used fluorescence analyzers, such as Kyanpen and Beckmann, have been developed by W.Campen and K.Bachman; Mikrochimica Acta, p.
159-170, 1979 II), it was difficult to perform highly sensitive and accurate fluorescence analysis even if it was applied to a system containing a large amount of coexisting substances such as a nuclear fuel reprocessing solution. .

An object of the present invention is to provide a fluorescence analysis method and an apparatus therefor capable of enhancing the detection sensitivity for the concentration of a trace amount of an analyte in a liquid sample containing a coexisting substance.

Another object of the present invention is to provide a fluorescence analysis method capable of reducing the influence of a change in light transmittance over time.

[Means for solving problems]

A feature of the present invention that achieves the above object is to irradiate a liquid sample diluted with a cooled diluent with excitation light, and to disperse the fluorescence emitted from the substance to be analyzed in the liquid sample by this irradiation. The purpose is to detect the intensity of the fluorescent light that has been spectrally separated and obtain the concentration of the substance to be analyzed based on the intensity of the fluorescent light.

A feature of the present invention that achieves the above-mentioned other object is that the analysis based on the ratio of the intensity of the fluorescence detected by the feature to the measured intensity of Raman light emitted from the solvent in the liquid sample. To determine the concentration of the substance to be.

[Action]

By diluting a liquid sample containing a coexisting substance with a diluent, absorption and attenuation of excitation light and fluorescence due to the coexisting substance can be reduced, and fluorescence quenching due to the coexisting substance can be suppressed, thus increasing the fluorescence yield. be able to. Further, since the diluent is cooled, the temperature of the liquid sample is lowered and the temperature quenching of fluorescence is suppressed, so that the fluorescence yield can be further increased. Therefore, it is possible to increase the detection sensitivity for the concentration of a trace amount of the analysis substance in the liquid sample containing a large amount of the coexisting substance, and accordingly improve the detection accuracy.

Furthermore, another feature of the present invention is to obtain the concentration of the substance to be analyzed based on the ratio between the intensity of the fluorescence and the intensity of the Raman light emitted from the solvent in the liquid sample, while producing the above-mentioned action. Thereby, it is possible to reduce the influence of the change of the light transmittance with time and to increase the detection sensitivity with respect to the concentration of the substance to be analyzed, and to improve the detection accuracy accordingly.

〔Example〕

Next, an embodiment of the present invention will be described with reference to FIGS. FIG. 1 is an elevation view of this embodiment, and FIG. 2 is a side view. First, the device configuration will be described.

A laser 1, which is an excitation light source, a spectroscope 2 that disperses fluorescence from a sample, a photodetector 3 that detects the dispersed light, and a data processing device 4 that quantifies the concentration of an analyte based on the photodetection intensity.
The analyzer body such as is placed in the analysis room and is isolated from the environment around the analysis site by the wall 5. When the analysis site is a high radiation field such as a nuclear fuel reprocessing plant, the wall 5 serves to protect the analysis operator and the analyzer body. The laser 1 and the spectroscope 2 are connected to a light transmitting fiber 6 and a light receiving fiber 7, respectively, and a fiber connector 8
It is connected to the dark box 9 via. Inside the dark box 9, the sample
There is a quartz cell 11 containing ten. The excitation light 12 transmitted by the light transmitting fiber 6 is reflected by the reflecting mirror 13, passes through the quartz of the quartz cell 11 and excites the analyte in the sample 10. The generated fluorescent light 14 is condensed on the end surface of the light receiving fiber 7 by the condenser mirror 15. In the vicinity of the end face of the fiber 6, 7 on the side of the dark box 9
When 6 is arranged and stray light or the sample 10 emits radiation, the radiation is reduced.

Next, FIG. 2 will be described. The quartz cell 11 in the dark box 9 is connected to the process pipe 18 by a sampling pipe 17. The dark box 9 can be replaced by the valve 19. A certain amount of the sampled process solution is taken in by the fractionator 20, and the dilution water supply pipe 21 is used to introduce the dilution water.
It is mixed with the dilution water introduced through 22 and sent to 11 parts of the quartz cell. The dilution water is cooled by the cooler 24 provided in the dilution water supplier 21. In the figure, a thick white arrow represents a process solution, a thin white arrow represents a sampled process solution, and a thin black arrow represents dilution water. When the process solution emits radiation, the process pipe 18 is provided with a hanger 23 to protect the fiber.

Next, the analysis procedure will be described.

First, the process solution is taken into the fractionator 20 from the sampling pipe 17. In the dispenser 20, after a fixed amount of the process solution is dispensed using a flow meter, a predetermined amount of the cooled dilution water from the dilution water supplier 21 is dispensed in the same manner, and these liquids are respectively supplied to the quartz cell 11 Next, the diluted sample 10 is irradiated with excitation light 12 from the laser 1, the generated fluorescence 14 is separated and detected, and data processing is performed to obtain the concentration of the analyte. After the analysis is completed, the inside of the fractionator 20, the sampling pipe 17 and the inside of the quartz cell 11 below are washed using the diluting water as the washing water to prepare for the next sampling.

If the process solution does not contain large amounts of coexisting substances, diluting the sample as described above will reduce the concentration of the analyte, so unless there is a high concentration of analyte where self-absorption of the analyte is a problem. It is meaningless in analysis. However, such as the analysis of uranium in the nuclear fuel reprocessing solution, it contains a large amount of coexisting substances, and therefore, the absorption / attenuation of excitation light / fluorescence due to the decrease in the transparency of the sample and the fluorescence absorption due to the quenching action of coexisting substances. When a decrease in the rate occurs, it is possible to achieve higher sensitivity by diluting.
The principle will be described below.

First, considering the optical path length of the quartz cell 11 as shown in FIG. 3, the excitation light 12 passes through the center of the cell width l, and the generated fluorescence 14 is focused from the side surface of the quartz cell 11, The fluorescence intensity I _f is given by the following equation.

Where I _i : incident light intensity φ _f : fluorescence yield of analyte ε: absorption coefficient of analyte C: concentration of analyte μ: absorbance of sample λ: wavelength Subscripts f and e are fluorescence and excitation, respectively Represents light.

Therefore, the fluorescence intensity before and after dilution was
_{Given f1} and I _f2 , the ratio R of these is It becomes like a formula. Here, k represents the dilution rate.

Here, for example, μ2 is used as in the nuclear fuel reprocessing solution,
If k10, then R0.5. That is, R <1
(Or R5 / k). This is because absorption / attenuation by the coexisting substance of the excitation light and the fluorescence is reduced. However, this effect alone would rather reduce the sensitivity. The effect of dilution contributes to the suppression of the quenching mechanism as well as the reduction of absorption and attenuation. That is, when a coexisting substance is present, this excites the excitation energy by the interaction with the excited analyte, and the analyte returns to the ground state without emitting fluorescence. Since this quenching mechanism depends on the concentration of the coexisting substance, the quenching can be suppressed by dilution and the fluorescence yield can be increased. For example, in the case of a nuclear fuel reprocessing solution, as a substance that quenches the fluorescence of uranium, there is a corrosion product such as iron ion. For iron ion, the quenching suppression effect is 3 to 10 at a dilution ratio of 10 times. It can be improved nearly twice. Therefore, when diluted, the absorption / attenuation of excitation light / fluorescence is reduced, and the fluorescence quenching is suppressed, whereby the fluorescence detection efficiency is improved and the sensitivity of analysis can be increased. Taking the nuclear fuel reprocessing solution as an example, the sensitivity becomes 1.5 to 5 times higher.

Furthermore, by diluting, the fluctuations of the coexisting substance concentration and temperature of the process solution can be suppressed to 1 / (k) of the dilution rate, reducing the measurement error and improving the accuracy of analysis. it can.

In this embodiment, high sensitivity and high accuracy can be achieved by diluting the sample with diluting water and then performing the fluorescence analysis. Furthermore, in this method, the concentration of the sample existing in the quartz cell is lower than that when it is not diluted, so when replacing the dark box / quartz cell, like reprocessing solution, if you want to reduce internal contamination as much as possible, It will have advantages over the undiluted case. Further, it also has an effect of cleaning and removing the dirt of the quartz cell such as the adhered cladding. In addition, since the radioactivity concentration of the process solution also decreases due to the dilution, various effects can be expected at the same time, such as suppressing the progress of deterioration of the fiber and the quartz cell due to the radiation from the sample.

As the diluting liquid, water is generally one of the suitable ones, since mixing in the process solution causes no problems in the process. Since the process solution contains a large amount of coexisting substances, it is not necessary to increase the purity of the dilution water, and general tap water is sufficient. The appropriate dilution rate depends on the transparency of the process solution and the concentration dependence of the quencher, but in the case of the nuclear fuel reprocessing solution, the dilution rate k = about 5 to 20 is appropriate.

In particular, diluting the process solution with cooled dilution water as described above is effective when the fluorescence yield of the analyte shows a negative temperature dependence. Cooling the liquid sample suppresses thermal motion of all substances in the liquid sample and reduces collisions between the analyte and coexisting substances, as well as collisions between the analyte and solvent. Therefore, the energy loss of the analyte due to collision can be reduced, and the quenching of fluorescence can be suppressed. That is, temperature quenching can be suppressed and the fluorescence yield can be increased. For example, as a result of examining the temperature dependence of the fluorescence intensity I _f of uranium in the nuclear fuel reprocessing solution, it was found that it was given by the following equation.

I _f ∝C · exp (E / RT) (4) E: Activation energy (35.7 kJ / mol) R: Gas constant T: Temperature (K) Therefore, when the temperature rises from 30 ℃ to 40 ℃, The light intensity will be reduced by about 40%. Therefore, the present embodiment will be described below by taking the nuclear fuel reprocessing solution as an example. First, the process solution at a temperature of 30 to 40 ° C. is sampled by the fractionator 20. Next, the dilution water cooled to about 0 ° C. is introduced through the dilution water introduction pipe 22. Here, if the dilution ratio k = 10, the temperature of the diluted sample will be about 3 to 4 ° C., and the fluorescence intensity _If will increase by about 4.8 times from the equation (4). Therefore, absorption of excitation light and fluorescence by coexisting substances obtained by dilution
When evaluated together with the reduction of attenuation and the suppression effect of fluorescence quenching by the coexisting substance, 7.2% was obtained when the nuclear fuel reprocessing solution having an absorbance of μ2 and a temperature of 30 ° C to 40 ° C was diluted by a cooling decay of about 0 ° C.
~ 24 times higher sensitivity. As a result, the detection accuracy for the concentration of the analyte is further increased.

Here, the structure of the precipitator 20 does not have to be of the type controlled by the flow meter described above, and may be a cut-out type as shown in FIG. That is, first, the process solution (white arrow) is dispensed from the sampling pipe 17 into the process solution dispensing hole 24 (Fig. 4A), and then one side of the two disks constituting the dispenser 20 is rotated to separate the solution. The intake hole 24 and the introduction hole 25 are connected to each other, and at the same time, the dilution water is introduced from the dilution water introduction pipe 22, and the sample is transferred to the quartz cell portion downstream while diluting (FIG. 4B). With this mechanism, the sorting work can be performed only by rotating the disc.

As described above, according to the present example, by diluting the sample with cooling water, the absorption attenuation of the excitation light / fluorescence due to the coexisting substance is reduced, and the fluorescence quenching due to the coexisting substance is suppressed. Quenching can also be suppressed, and the detection sensitivity for the concentration of the analyte can be increased, which can also increase the detection accuracy. Regarding the temperature quenching alone, there is also a means for cooling the quartz cell portion from the outside, but it takes time to cool, and it is also easily affected by fluctuations in the process temperature. On the other hand, when the combined use of the dilution and the cooling is performed as in the present embodiment, the cooling can be performed quickly, and the dilution is less susceptible to the fluctuation of the process temperature. Also, in the case of external cooling, stain removal of the quartz cell, reduction of the amount of contamination inside the quartz cell,
Of course, additional effects such as reduction of radiation from the sample cannot be obtained. A temperature control device may be added to the quartz cell portion in order to improve the analysis accuracy of this embodiment.

Next, another embodiment of the present invention will be described.

As shown in FIG. 5, the device configuration of the present embodiment is almost the same as the device configuration shown in FIG. 2, except that a stirring tank 27 is installed downstream of the fractionator 20. In the present embodiment, first, the process solution is fractionated by the fractionator 20 and cooling / diluting water is introduced with the valve 19 at the bottom of the stirring tank 27 closed, and these mixed liquids are once put into the stirring tank 27. Here, even if the stirring tank 27 is a simple cavity, the mixing of the process solution and the dilution water is promoted by the drop of the water flow. Immediately thereafter, the valve 19 is opened to transfer the uniformly diluted and mixed sample into the quartz cell 11, and fluorescence analysis is performed.
If the sample stays in the stirring tank 27 for a long time, the sample temperature rises toward room temperature, but the stirring time may be about 1 second, and the fluorescence spectroscopic / detection time after transfer to the quartz cell 11 is the light detection. If you use a multi-channel detector
Since it is 1 ms or less, it is not necessary to consider the effect of temperature rise. However, it goes without saying that a temperature control mechanism may be added to the stirring tank 27.

As described above, according to the present embodiment, by adding the stirring tank, it is possible to mix the process solution with the dilution water.
There is an advantage that homogenization can be achieved more quickly.

Another embodiment of the present invention will be described below.

The device configuration of this embodiment is basically the same as the other embodiments described so far, and can be combined to exert the effect. The feature of this embodiment is that a pulse oscillation laser is used as a laser of an excitation light source and fluorescence is observed by time resolution. That is, it is the size of the background that determines the detection limit of the fluorescence analysis, but one of the factors is that the temporal behavior of scattered light and fluorescence is different as shown in FIG. The aim is to improve the S / N ratio by temporally discriminating the scattered light generated at the same time as the excitation light irradiation from the fluorescence light that decays at a certain life after the excitation light irradiation. Most laser pulses have a pulse width of several nanoseconds. Therefore, it is desirable that the fluorescence lifetime of the analyte of interest in this example be several tens of nanoseconds or more. Fluorescence lifetime measurement results of uranium in nuclear fuel reprocessing solution are several tens.
It is suitable because it is 0 μs or more. In the case of uranium in the reprocessed solution, it has an absorption peak near the wavelength of 340 nm, so for example N
_{If two} lasers (wavelength 337 nm) are used as the pulsed laser, the above-mentioned time analysis measurement can be performed. In this case, a time-resolving device such as a multi-channel detector is used for spectroscopy and photodetection. Considering that the transmission loss due to the radiation of the optical fiber strongly appears on the short wavelength side, it is also a good idea to use the wavelength near 410 nm which is the second absorption peak as the excitation wavelength of uranium in the reprocessing solution. is there. In this case, a pulsed dye laser is used.

As described above, when the pulsed laser is used as the excitation light source and the fluorescence that is the signal and the scattered light that is the noise are time-resolved and discriminated, the S / N ratio can be improved and the detection limit can be widened. . Cooling the sample generally prolongs the fluorescence life, so that dilution / cooling is also advantageous in this respect.

Another embodiment of the present invention will be described below.

This embodiment relates to data processing, and can be applied to the device configurations of the embodiments described so far without any restrictions.

This example is the fluorescence intensity of the analyte when the change over time in the light transmittance outside the sample liquid cannot be ignored in the analysis due to the adhesion of the cladding, the coloring of quartz due to radiation damage, etc., as in the case of the nuclear fuel reprocessing solution. By taking the ratio of I _f and the Raman light intensity I _r of the solvent, the effect of the fluctuation of the above light transmittance is offset. The principle will be described below.

(1) from equation, the fluorescent intensity I _f can be rewritten as the following equation.

Here, μt _q : Equivalent absorption amount by quartz cell and adhered dirt Similarly Raman light intensity I _r of the solvent is given by the following equation.

Here, k _r : constant peculiar to the solvent C _S : solvent concentration The subscript r represents Raman light.

Therefore, the ratio R _fr between the fluorescence intensity I _f of the analyte and the Raman light intensity I _r of the solvent is Was Although connexion (1) fluorescent wavelength lambda _f, Raman wavelength lambda _r in the expression and absorbance μ of the sample by the exciting light wavelength lambda _e, and change in the light absorption Myuti _q by quartz cell and fouling ([Delta] [mu
_{If fr} , Δμ _er , Δμ t _q , _fr ) is small, the equation (6) can be written as follows.

R _fr ≒ K ・ C (8) That is, the analyte concentration C can be known from R _fr .

As shown above, after diluting the sample, by measuring the ratio of the fluorescence intensity of the analyte and the Raman intensity of the solvent, the absorption / attenuation of the excitation light / fluorescence by the sample and the fluorescence quenching are reduced. In addition, it is possible to cancel the influence of the time-dependent change in the transmittance of excitation light / fluorescence due to the radiation damage of the quartz cell and the adhesion of the cladding, which has the advantage of achieving higher sensitivity and higher accuracy in the fluorescence analysis. is there. Not only the wavelength dispersion method but also the method in which the pulse laser is used as the excitation light source and the time-resolved photometry is effective for the discrimination between the fluorescence light and the Raman light.

Another embodiment of the present invention will be described below.

FIG. 7 shows an elevation view of this embodiment. The excitation light 12 from the laser 1 passes through the excitation light passage hole 28 and is incident on the diluted sample in the quartz cell 11. The sample dilution procedure is the same as in the other examples. After passing through the sample, the excitation light 12 is reflected by a reflecting mirror arranged on the back surface of the quartz cell 11 and returns to the sample again,
Increase excitation efficiency. Fluorescence 14 emitted from the analyte is a condenser mirror
After condensing at 15, the light is focused into parallel rays by the reflecting mirror 13 and the focusing mirror 29, passes through the fluorescence passage hole 30 and enters the spectroscope 2, and after being spectroscopically detected by the photodetector 3, the data processing device. Four
Gives the analyte concentration. A reflecting mirror 13 is installed on the side of the quartz cell opposite to the condensing mirror 15, and the fluorescence emitted to these sides is collected by the condensing mirror 15 to improve the condensing efficiency of the fluorescence. The shiya plate 31 is installed to reduce stray light.
In FIG. 7, the excitation light passage hole 28 and the fluorescence light passage hole 30 are linear, but it is also possible to dispose a reflecting mirror in the middle to bend the optical path. In addition, since time-resolved photometry can be performed using a pulsed laser as an excitation light source, scattered light and fluorescent light can be discriminated.
In this case, the excitation light passage hole 28 and the optical plate 31 are not always necessary.

As described above, according to the present embodiment, the pumping light and the fluorescent light can be transmitted without using the light transmitting and light receiving fibers, so that there is an advantage that it is not necessary to consider the transmission loss due to the radiation damage of the fiber.

〔The invention's effect〕

According to the present invention, by diluting and cooling a liquid sample containing a coexisting substance, absorption / attenuation of excitation light and fluorescence by the coexisting substance is reduced, suppression of fluorescence quenching by the coexisting substance, and temperature quenching of fluorescence. As a result, the detection sensitivity for the concentration of a trace amount of the analyte in the liquid sample can be increased, and the detection accuracy can be improved accordingly.

Furthermore, according to another feature of the present invention, in addition to the above effects, the concentration of the substance to be analyzed can be determined based on the ratio of the intensity of fluorescence to the intensity of malan light emitted from the solvent in the liquid sample. Since it is obtained, it is possible to reduce the influence of the change over time of the light transmittance.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is an elevation view of an embodiment of the present invention, FIG. 2 is a side view thereof, FIG. 3 is a sectional view of a quartz cell, and FIG. 5 is a side view of another embodiment of the present invention, FIG. 6 is a view showing the fluorescence spectrum and its change with time, and FIG. 7 is another embodiment of the present invention. FIG. 6 is an elevational view of an example. 1 ... Laser, 2 ... Spectrometer, 3 ... Photodetector, 4 ...
Data processing device, 6 ... Destination fiber, 7 ... Receiving fiber, 10 ... Sample, 11 ... Quartz cell, 12 ... Excitation light, 14
...... Fluorescence, 17 …… Sampling pipe, 18 …… Process pipe, 20 …… Preparation device, 21 …… Diluting water supply device, 22 …… Diluting water introducing pipe, 24 …… Cooler, 27 …… Agitating Tank.

Claims (6)

[Claims] 1. A liquid sample diluted with a cooled diluent is irradiated with excitation light, and the fluorescence emitted from a substance to be analyzed in the liquid sample is dispersed by this irradiation. And a fluorescence analysis method for determining the concentration of the substance to be analyzed based on the intensity of the fluorescence. 2. The method according to claim 1, wherein the diluent is water.
Fluorescence analysis method described in paragraph. 3. A liquid sample diluted with a cooled diluent is irradiated with excitation light, and the fluorescence emitted from the substance to be analyzed in the liquid sample is separated by this irradiation, and the separated fluorescence is obtained. The intensity of Raman light emitted from the solvent in the liquid sample is detected, and the concentration of the substance to be analyzed is determined based on the ratio between the intensity of the fluorescence and the intensity of the Raman light. Fluorescence analysis method. 4. A means for diluting a liquid sample containing a substance to be analyzed with a cooled diluent, a cooling means for cooling the diluent, an analysis cell for containing the diluted liquid sample, and the analysis. Means for irradiating the liquid sample in the cell with excitation light, a spectroscope for separating fluorescence emitted from the substance to be analyzed in the liquid sample irradiated with the excitation light, and spectroscopic by the spectroscope A fluorescence analysis device comprising: a detector for detecting the intensity of the emitted fluorescence light; and a data processing device for obtaining the concentration of the substance to be analyzed based on the intensity of the fluorescence light. 5. The fluorescence according to claim 4, wherein the means for irradiating the excitation light is a pulse transmission laser device, the spectroscope is a time-resolved type, and the detector is a time-resolved type. Analysis equipment. 6. The fluorescence analyzer according to claim 5, wherein said diluting means includes a mixing means for mixing the liquid sample containing the substance to be analyzed with the cooled diluting liquid.