JP6191477B2 - Particle size measuring apparatus and particle size measuring method - Google Patents

Description

  The present invention relates to a particle size measuring apparatus, a particle size measuring method, and a particle size measuring program for measuring the particle size of a sample.

  Conventionally, in order to measure the particle diameter of a sample, a particle diameter measuring apparatus such as a so-called particle counter has been used (for example, see Patent Document 1 below). In this type of particle size measuring device, the particle size can be measured based on the obtained scattered light intensity by irradiating the particles in the sample with laser light and receiving the scattered light from the particles with a detector. it can.

  The particles in the sample pass through the measurement region in the cell one by one. The laser light is focused on the measurement region and scattered by each particle passing through the measurement region one by one, and the scattered light intensity corresponding to the particle diameter of each particle is detected by the detector.

Japanese Patent No. 2899359

  In a particle diameter measuring apparatus such as a particle counter, as described above, the intensity of scattered light must be detected by irradiating each particle with one laser beam. For this reason, it may be difficult to separate each particle, and if two or more particles are simultaneously irradiated with laser light, there is a problem that the particle size cannot be measured accurately.

  The lower limit of the particle diameter that can be measured with a particle counter is generally about 250 nm. Therefore, for example, it is difficult to measure the particle diameter of fine particles of 100 nm or less, and there is a problem that the measurement range is limited.

  The present invention has been made in view of the above circumstances, and an object thereof is to provide a particle size measuring apparatus, a particle size measuring method, and a particle size measuring program capable of accurately measuring a wider range of particle sizes. .

The particle size measurement apparatus according to the present invention includes a particle separation unit, a particle concentration measurement unit, a scattered light intensity measurement unit, a measurement reference information storage unit, and a particle size calculation unit. The particle separation unit separates the sample into particle groups for each particle diameter. The particle concentration measurement unit measures the particle concentration for each particle group separated by the particle separation unit. The scattered light intensity measurement unit measures the scattered light intensity for each particle group separated by the particle separation unit. The measurement reference information storage unit stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement reference information. The particle diameter calculation unit, particle concentration measurement by the particle concentration measurement unit, and, based on the scattered light intensity measured by the scattered light intensity measuring unit calculates the scattered light intensity per unit concentration, the Based on the value and the measurement reference information, the particle size of the particle group separated by the particle separation unit is calculated.
The particle size measurement apparatus according to the present invention includes a particle separation unit, a particle concentration measurement unit, a scattered light intensity measurement unit, a measurement reference information storage unit, and a particle size calculation unit. The particle separation unit separates the sample into particle groups for each particle diameter. The particle concentration measurement unit measures the absorbance for each particle group separated by the particle separation unit, and measures the particle concentration from the absorbance. The scattered light intensity measurement unit measures the scattered light intensity for each particle group separated by the particle separation unit. The measurement reference information storage unit stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement reference information. The particle size calculation unit is a peak area of absorbance corresponding to each particle group measured by the particle concentration measurement unit, a peak area of scattered light intensity corresponding to each particle group measured by the scattered light intensity measurement unit, And based on the said measurement reference information, the particle diameter of the particle group isolate | separated by the said particle | grain separation part is calculated.

  According to such a configuration, it is possible to calculate the particle diameter of each particle group after separating the sample into particle groups for each particle diameter. At this time, by using the measurement reference information representing the relationship between the scattered light intensity per unit concentration and the particle diameter, the measured particle concentration and scattered light of each particle group without being separated into individual particles. The particle diameter can be accurately measured based on the strength.

  Further, since the scattered light intensity is measured in units of particle groups, it is easier to discriminate changes in the scattered light intensity depending on the difference in particle diameter than in a configuration in which the scattered light intensity is measured in units of particles. Therefore, for example, it is possible to measure the particle diameter even with extremely fine particles such as 100 nm or less, and a wider range of particle diameters can be measured.

  The scattered light intensity measurement unit may include one detector that receives scattered light from the sample. In this case, the particle size calculation unit may calculate the particle size of the particle group separated by the particle separation unit based on the scattered light intensity measured by the one detector.

  According to such a configuration, since the particle diameter can be measured using one detector, a simpler and cheaper configuration can be achieved. In this case, if the position of the detector is set according to the particle diameter to be measured and the angle at which scattered light from the particle group is received is changed, the particle diameter can be accurately measured over a wide range of, for example, several nanometers to several micrometers. can do.

  The particle concentration measurement unit may include a light source that irradiates light to the particle group separated by the particle separation unit. In this case, the one detector may receive the scattered light of the light irradiated to the particle group from the light source.

  According to such a configuration, not only can the particle concentration of the particle group be measured using light emitted from the light source of the particle concentration measurement unit, but also the scattered light is received by the detector. The scattered light intensity of the particle group can also be measured. In this case, the particle diameter can be measured with a simple configuration in which only one detector is added to the existing particle concentration measurement unit.

The particle size measurement method according to the present invention includes a particle separation step, a particle concentration measurement step, a scattered light intensity measurement step, a measurement reference information reading step, and a particle size calculation step. In the particle separation step, the sample is separated into particle groups for each particle diameter. In the particle concentration measurement step, the particle concentration is measured for each particle group separated in the particle separation step. In the scattered light intensity measurement step, the scattered light intensity is measured for each particle group separated in the particle separation step. In the measurement standard information reading step, the measurement standard information is read from a measurement standard information storage unit that stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement standard information. Wherein the particle diameter calculating step, the particle concentration measured measured particle concentration in step, and, on the basis of the measured scattered light intensity in the scattered light intensity measuring step to calculate the scattered light intensity per unit concentration, the Based on the value and the measurement reference information, the particle size of the particle group separated in the particle separation step is calculated.
The particle size measurement method according to the present invention includes a particle separation step, a particle concentration measurement step, a scattered light intensity measurement step, a measurement reference information reading step, and a particle size calculation step. In the particle separation step, the sample is separated into particle groups for each particle diameter. In the particle concentration measurement step, the absorbance is measured for each particle group separated in the particle separation step, and the particle concentration is measured from the absorbance. In the scattered light intensity measurement step, the scattered light intensity is measured for each particle group separated in the particle separation step. In the measurement standard information reading step, the measurement standard information is read from a measurement standard information storage unit that stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement standard information. In the particle diameter calculation step, the peak area of absorbance corresponding to each particle group measured in the particle concentration measurement step, the peak area of scattered light intensity corresponding to each particle group measured in the scattered light intensity measurement step, And based on the said measurement reference information, the particle diameter of the particle group isolate | separated by the said particle separation step is calculated.

  The particle size measurement program according to the present invention causes a computer to execute the particle separation step, the particle concentration measurement step, the scattered light intensity measurement step, the measurement reference information reading step, and the particle size calculation step. .

  According to the present invention, the particle size can be accurately determined by separating the sample into particle groups for each particle size and measuring the particle concentration and scattered light intensity of each particle group without separating the particles into individual particles. Can be measured. Further, according to the present invention, since the scattered light intensity is measured in units of particle groups, it is possible to measure the particle diameter even with extremely fine particles, and a wider range of particle diameters can be measured.

It is the schematic which showed the structural example of the particle diameter measuring apparatus which concerns on one Embodiment of this invention. It is the schematic which showed an example of the specific structure of a scattered light intensity | strength measurement part. It is the block diagram which showed an example of the specific structure of a control apparatus. It is a figure for demonstrating an example of measurement reference information. It is the figure which showed an example of the light absorbency A and the scattered light intensity | strength I at the time of calculating a particle diameter with time passage. It is the flowchart which showed an example of the process by the control part at the time of measuring the particle diameter of a sample. It is the schematic which showed the modification of the particle concentration measurement part.

  FIG. 1 is a schematic diagram showing a configuration example of a particle size measuring apparatus according to an embodiment of the present invention. This particle size measuring device is a device for measuring the particle size of a sample such as protein, and includes an introduction unit 1, a particle separation unit 2, a particle concentration measurement unit 3, a scattered light intensity measurement unit 4, a control device 5, and the like. I have.

  The sample is introduced from the introduction unit 1 to the particle separation unit 2. The particle separation unit 2 is configured by a liquid chromatograph such as a size exclusion chromatograph, for example, and separates a sample into particle groups for each particle diameter. That is, by configuring the particle separation unit 2 using a mechanism that changes the flow rate of particles according to the particle diameter, particles having similar particle diameters can be eluted as particle groups in the same time zone. However, the particle separation unit 2 is not limited to a liquid chromatograph, and is configured to be able to separate a sample into particles for each particle diameter by other methods such as FFF (Field Flow Fractionation). Also good.

  The particle concentration measurement unit 3 includes a light source 31 and a detector 32, for example, and measures the particle concentration for each particle group separated by the particle separation unit 2. Specifically, the measurement light is irradiated from the light source 31 to the particle groups having the respective particle diameters separated by the particle separation unit 2 and sequentially guided to the particle concentration measurement unit 3. Then, the absorbance can be measured by detecting the measurement light passing through the particle group with the detector 32, and the concentration of the particles can be calculated from the absorbance.

  When calculating the particle concentration from the absorbance, for example, a calibration curve obtained by measuring the absorbance of a particle group having a known concentration can be used. As the detector 32, for example, various detectors such as a UV detector (ultraviolet detector), an RI detector (differential refractive index detector), and an RF detector (fluorescence detector) can be used.

  The scattered light intensity measurement unit 4 is for measuring the scattered light intensity for each particle group separated by the particle separation unit 2. In this example, the particle group having each particle diameter that has passed through the particle concentration measurement unit 3. Are sequentially guided to the scattered light intensity measurement unit 4. The particle groups of each particle diameter sequentially guided to the scattered light intensity measurement unit 4 are irradiated with the measurement light from the light source 41, and the scattered light emitted when the measurement light is scattered by the particle group is detected by the detector 42. The

  As the light source 41, various light sources, such as a lamp, LED (Light Emitting Diode), or a laser light source, can be used, for example. Moreover, as the detector 42, various detectors, such as a photodiode or PMT (photomultiplier tube), can be used.

  The control device 5 controls the operation of each unit such as the introduction unit 1, the particle separation unit 2, the particle concentration measurement unit 3, and the scattered light intensity measurement unit 4, and performs calculations based on the detection results by the detectors 32 and 42. . In the present embodiment, the particle diameter of each particle group separated by the particle separation unit 2 using the particle concentration measured by the particle concentration measurement unit 3 and the scattered light intensity measured by the scattered light intensity measurement unit 4. Is calculated.

  FIG. 2 is a schematic diagram illustrating an example of a specific configuration of the scattered light intensity measurement unit 4. As shown in FIG. 2, the measurement light from the light source 41 is irradiated to the particle group P for each particle diameter sequentially guided into the cell 43, and the scattered light generated at this time is detected by the detector 42. .

  In this example, the detector 42 is constituted by one side sensor arranged on the side of the cell 43 as viewed from the light source 41 side. The cell 43 is formed of, for example, a thin hollow member, and is arranged so that the thickness direction D thereof is parallel to the optical axis L of the measurement light incident from the light source 41. The side sensor as the detector 42 is arranged with respect to the cell 43 in a direction orthogonal to the thickness direction D, for example.

  FIG. 3 is a block diagram illustrating an example of a specific configuration of the control device 5. The control device 5 is configured by a computer, for example, and includes a control unit 51 and a storage unit 52. The control unit 51 includes, for example, a CPU (Central Processing Unit), and functions as various functional units such as the particle size calculation unit 511 when the CPU executes a program. The storage unit 52 can be configured by, for example, a ROM (Read Only Memory), a RAM (Random Access Memory), a hard disk, and the like.

  The particle size calculation unit 511 is a particle group particle separated by the particle separation unit 2 based on the particle concentration measured by the particle concentration measurement unit 3 and the scattered light intensity measured by the scattered light intensity measurement unit 4. A process for calculating the diameter is performed. The storage unit 52 is assigned a measurement reference information storage unit 521 that stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement reference information. The particle diameter calculation unit 511 stores the measurement reference information. Use to calculate the particle size of the particle group.

  The particle diameter calculated by the particle diameter calculation unit 511 is stored in the particle diameter storage unit 522 allocated to the storage unit 52 in association with each particle group. About the particle diameter of each particle group memorize | stored in the particle diameter memory | storage part 522, it reads as a measurement result and can perform the process of displaying on a display part (not shown).

  FIG. 4 is a diagram for explaining an example of the measurement reference information. As shown in FIG. 4, the particle concentration (C) measured by the particle concentration measuring unit 3 and the scattered light intensity (I) measured by the scattered light intensity measuring unit 4 are proportional to each other. Depending on the particle diameter (d), the scattered light intensity per unit concentration (“tilt” in FIG. 4) changes. The measurement standard information representing the relationship between the particle concentration (C), the scattered light intensity (I), and the particle diameter (d) can be obtained by using, for example, a calculation formula based on the Mie scattering theory or the like. A calibration curve can also be obtained by measuring the particle concentration and scattered light intensity of a particle group having a particle diameter.

  Based on the particle concentration (C) measured by the particle concentration measuring unit 3 and the scattered light intensity (I) measured by the scattered light intensity measuring unit 4, the particle diameter calculating unit 511 is configured to scatter light per unit concentration. By calculating the intensity (slope) and substituting the slope into a function corresponding to the measurement reference information as shown in FIG. 4, the particle diameter (d) of each particle group is specified. For example, when the particle concentration is C = C1 and the scattered light intensity is I = I1, the particle diameter of each particle in the particle group is d = 50 nm.

  Thus, even if the particle concentration (C) of the particle group separated by the particle separation unit 2 is obtained, the particle diameter (d) is specified unless the scattered light intensity (I) of the particle group is obtained. It is not possible. Similarly, even if the scattered light intensity (I) of the particle group separated by the particle separation unit 2 is obtained, the particle diameter (d) is specified unless the particle concentration (C) of the particle group is obtained. I can't. That is, only when both the particle concentration (C) and the scattered light intensity (I) of the particle group separated by the particle separation unit 2 are obtained, the particle diameter ( d) can be specified.

  FIG. 5 is a diagram showing an example of the absorbance A and scattered light intensity I when calculating the particle diameter over time. In this example, the correspondence between the timing at which the absorbance A is measured by the particle concentration measuring unit 3 and the timing at which the scattered light intensity I is measured by the scattered light intensity measuring unit 4 is accurately obtained for each particle group. Will be described.

  For example, as shown in FIG. 5, for the particle group separated by the particle separation unit 2, it is assumed that the absorbance at a certain timing is A = A2, and the scattered light intensity at the corresponding timing is I = I2. In this case, the scattered light intensity (I2 / C2) per unit concentration is calculated by dividing the scattered light intensity I2 by the particle concentration C2 calculated from the absorbance A2. By substituting the scattered light intensity (I2 / C2) per unit concentration calculated in this way into a function representing the relationship between the scattered light intensity per unit concentration and the particle diameter, for example, the particle diameter of the particle group Can be calculated.

  In this example, each particle is based on the instantaneous value (A2) of the absorbance A measured by the particle concentration measuring unit 3 and the instantaneous value (I2) of the scattered light intensity I measured by the scattered light intensity measuring unit 4. The configuration for calculating the particle size of the group has been described. However, for example, the particle diameter of each particle group may be calculated based on the peak areas of absorbance A and scattered light intensity I corresponding to each particle group.

  In this case, for the peak P1 of absorbance A and the peak P2 of scattered light intensity I corresponding to the same particle group, respective integrated values may be obtained to calculate the scattered light intensity per unit concentration. As described above, when the integrated value is used instead of the instantaneous value, the timing when the absorbance A is measured by the particle concentration measuring unit 3 and the timing when the scattered light intensity I is measured by the scattered light intensity measuring unit 4. Even if the correspondence relationship cannot be obtained accurately, if the peaks P1 and P2 of the absorbance A and scattered light intensity I corresponding to the same particle group can be discriminated, the scattered light intensity per unit concentration can be accurately calculated. be able to.

  FIG. 6 is a flowchart showing an example of processing by the control unit 51 when measuring the particle diameter of the sample. When measuring the particle size of a sample, first, the sample is introduced into the particle separation unit 2 from the introduction unit 1 at a predetermined flow rate so that the sample is separated into particle groups for each particle size in the particle separation unit 2. Is performed (step S101: particle separation step).

  The particle groups separated for each particle group in this manner are sequentially guided to the particle concentration measurement unit 3, and the particle concentration measurement unit 3 performs a process for measuring the particle concentration for each particle group (step S102). : Particle concentration measurement step). Each particle group is sequentially guided from the particle concentration measuring unit 3 to the scattered light intensity measuring unit 4, and the scattered light intensity measuring unit 4 performs processing for measuring the scattered light intensity for each particle group (step S103). : Scattered light intensity measurement step).

  Thereafter, measurement reference information is read from the measurement reference information storage unit 521 (step S104: measurement reference information reading step), and based on the measurement reference information, the particle concentration and the scattered light intensity measured in advance, Processing for calculating the particle diameter of the particle group is performed (step S105: particle diameter calculating step). Such processing of steps S102 to S105 is sequentially executed for each particle group separated in the particle separation unit 2.

  In this embodiment, after separating a sample into particle groups for each particle diameter, the particle diameter of each particle group can be calculated. At this time, by using the measurement standard information indicating the relationship between the scattered light intensity per unit concentration and the particle diameter as shown in FIG. 4, each measured particle group without being separated into individual particles. The particle diameter can be accurately measured based on the particle concentration and scattered light intensity.

  Further, since the scattered light intensity is measured in units of particle groups, it is easier to discriminate changes in the scattered light intensity depending on the difference in particle diameter than in a configuration in which the scattered light intensity is measured in units of particles. Therefore, as shown in FIG. 4, it is possible to measure the particle diameter even with extremely fine particles such as 100 nm or less, and a wider range of particle diameters can be measured.

  In particular, in the present embodiment, the scattered light from the sample is received by one detector 42 provided in the scattered light intensity measurement unit 4 (see FIG. 2). Then, based on the scattered light intensity measured by the one detector 42, the particle size calculation unit 511 calculates the particle size of the particle group separated by the particle separation unit 2.

  Thus, since the particle diameter can be measured using one detector 42, a simpler and cheaper configuration can be achieved. In this case, if the position of the detector 42 is set according to the particle diameter to be measured and the angle at which the scattered light from the particle group is received is changed, the particle diameter can be accurately adjusted over a wide range of, for example, several nm to several μm. Can be measured.

  For example, as shown in FIG. 2, when the detector 42 is configured to receive scattered light scattered in a 90 ° direction with respect to the optical axis L of the measurement light, a particle diameter of about several nm to 100 nm can be measured. it can. In the case where the detector 42 receives scattered light scattered from the cell 43 toward the light source 41 (rear), a smaller particle diameter can be measured. On the other hand, when the detector 42 receives scattered light scattered from the cell 43 to the side opposite to the light source 41 (forward), a larger particle diameter can be measured.

  FIG. 7 is a schematic view showing a modification of the particle concentration measuring unit 3. In this modification, the scattered light intensity measurement unit 4 is integrated with the particle concentration measurement unit 3. That is, light (for example, ultraviolet light) is irradiated from the light source 31 to the particle group P for each particle diameter sequentially guided into the cell 33 provided in the particle concentration measuring unit 3, and the absorbance is measured by the detector 32. In addition, the scattered light intensity is measured by the detector 42.

  At this time, since the wavelength region where the absorption occurs differs depending on the type of the sample, the absorbance can be satisfactorily measured by measuring the absorbance based on the received light intensity of the detector 32 at the wavelength where the absorption occurs. On the other hand, if the scattered light intensity is measured based on the received light intensity of the detector 42 at a wavelength at which light absorption does not occur or light absorption is difficult to occur, the scattered light intensity can be measured satisfactorily.

  The detector 32 for measuring the absorbance is preferably arranged on the opposite side (front) of the cell 33 to the light source 31 side. On the other hand, the detector 42 for detecting the scattered light intensity can be arranged at an arbitrary position according to the particle diameter to be measured as described above.

  As described above, in the configuration as shown in FIG. 7, not only the particle concentration of the particle group can be measured using the light irradiated to the particle group from the light source 31 of the particle concentration measuring unit 3, but also the scattered light is detected. The scattered light intensity of the particle group can be measured by receiving the light with the device 42. In this case, the particle diameter can be measured with a simple configuration in which only one detector 42 is added to the existing particle concentration measurement unit 3.

  In the above embodiment, each particle group separated by the particle separation unit 2 is guided to the particle concentration measurement unit 3 and the particle concentration is measured, and then guided to the scattered light intensity measurement unit 4 to determine the scattered light intensity. The configuration to be measured has been described. However, the present invention is not limited to this configuration, and each particle group separated by the particle separation unit 2 is guided to the scattered light intensity measurement unit 4 and the scattered light intensity is measured, and then guided to the particle concentration measurement unit 3. The particle concentration may be measured.

  In this case, steps S102 and S103 in FIG. 6 may be executed in a reversed order. In the case where the particle concentration measuring unit 3 and the scattered light intensity measuring unit 4 are integrated as shown in FIG. 7, steps S102 and S103 of FIG. 6 may be executed simultaneously.

  In the above embodiment, the configuration in which the control unit 51 automatically measures the particle diameter of the sample by executing the process illustrated in FIG. 6 has been described. However, the configuration is not limited to such a configuration, and a configuration in which at least one of the processes illustrated in FIG. 6 is manually performed by an operator may be employed.

  Further, as in the above-described embodiment, not only can a particle size measuring device for measuring the particle size of a sample be provided, but also a program (particles) for causing a computer to execute the processing illustrated in FIG. It is also possible to provide a diameter measurement program. In this case, the program may be configured to be provided in a state stored in a storage medium, or may be configured to provide the program itself via wired communication or wireless communication. .

DESCRIPTION OF SYMBOLS 1 Introduction part 2 Particle separation part 3 Particle concentration measurement part 4 Scattered light intensity measurement part 5 Control apparatus 31 Light source 32 Detector 33 Cell 41 Light source 42 Detector 43 Cell 51 Control part 52 Memory | storage part 511 Particle diameter calculation part 521 Measurement reference information Storage unit 522 Particle size storage unit

Claims (2)

A particle separation unit for separating the sample into particles for each particle size;
Measuring the absorbance for each particle group separated by the particle separation unit, a particle concentration measurement unit for measuring the particle concentration from the absorbance,
A scattered light intensity measurement unit for measuring the scattered light intensity for each particle group separated by the particle separation unit;
A measurement standard information storage unit for storing the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement standard information;
In the peak area of absorbance corresponding to each particle group measured by the particle concentration measuring unit, the peak area of scattered light intensity corresponding to each particle group measured by the scattered light intensity measuring unit, and the measurement reference information A particle size measuring apparatus comprising: a particle size calculating unit that calculates a particle size of the particle group separated by the particle separating unit. A particle separation step for separating the sample into particles for each particle size;
Measuring the absorbance for each particle group separated in the particle separation step, measuring the particle concentration from the absorbance, a particle concentration measurement step,
A scattered light intensity measurement step for measuring the scattered light intensity for each particle group separated in the particle separation step;
A measurement reference information reading step for reading the measurement reference information from a measurement reference information storage unit that stores the relationship between the scattered light intensity per unit concentration and the particle diameter as measurement reference information;
The absorbance peak area corresponding to each particle group measured in the particle concentration measurement step, the scattered light intensity peak area corresponding to each particle group measured in the scattered light intensity measurement step, and the measurement reference information And a particle size calculating step for calculating a particle size of the particle group separated in the particle separation step.

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