JP6276949B2 - Large algae growth diagnosis apparatus and large algae growth diagnosis method - Google Patents

Description

  The present invention relates to a macroalgae growth diagnostic apparatus and a macroalgae growth diagnostic method.

  In Patent Document 1, a sample of large algae is irradiated with excitation light, the fluorescence intensity is measured, and the obtained measurement value is collated with a preset fluorescence intensity-quality test curve to evaluate the quality. The technique of the quality evaluation method of the macroalgae characterized by these is described.

JP-A-61-133842

“Possibility for growth diagnosis of Susabiori using fluorescence lifetime”, 2012 (45th) National Congress of Lighting Society, 12-4, September 6-8, 2012

  The technique described in Patent Document 1 obtains the correlation between the amount of photosynthetic oxygen generation and the fluorescence intensity of the dye, and evaluates the quality based on the obtained correlation. With the technique described in Patent Document 1, it is possible to easily carry out non-destructive growth diagnosis of large algae. However, large algae may cause a phenomenon in which the fluorescence intensity increases due to disease, growth (nurturing) environment stress, and the like. For this reason, although the technique of patent document 1 can evaluate normal macroalgae, the technique which evaluates macroalgae which received the disease, the stress of the growth environment, etc. is not established.

  Large-scale algae farming uses the sea as a place for production, so the effects of weather and sea conditions are greatly affected. Further, in the cultivation of macroalgae, there is a demand to suppress the deterioration of the growth state of macroalgae due to the influence of the interrelationship of ecosystem organisms, the water quality used for macroalgae culture, the disease of macroalgae, and the like. The fishermen who measure the cultivation of macroalgae, that is, the measurers, visually judged the deterioration of the growth state of macroalgae. By the way, the deterioration of the growth state of macroalgae is difficult to be visually determined at the initial stage. If the deterioration of the growth state of the large algae can be diagnosed at the initial stage, it is possible to take measures to eliminate the cause of the deterioration of the growth state of the large algae, and it is possible to improve the productivity of the cultivation of the large algae. Therefore, the present inventor has studied a method for diagnosing the growth of large algae using a fluorescence lifetime measurement method, but at the time described in Non-Patent Document 1, the deterioration of the growth state of the large algae is diagnosed at an early stage. I didn't get any knowledge.

  This invention is made | formed in view of the above, Comprising: It aims at providing the growth diagnostic apparatus of the large algae which can diagnose the deterioration of the growth state of a large algae at an early stage, and the growth diagnostic method of a large algae.

  In order to solve the above-mentioned problems and achieve the object, the macroalgae growth diagnostic apparatus comprises a pulsed light source that irradiates a sample of macroalgae with pulsed excitation light including a wavelength near the absorption peak wavelength of phycoerythrin, and Among the fluorescence emitted from the sample of macroalgae irradiated with pulsed excitation light, a detector for detecting fluorescence at a wavelength of 580 nm, storage means for storing threshold information regarding fluorescence lifetime, and output from the detector Based on the output signal, the analyzing means for calculating the average fluorescence lifetime of the fluorescence having the wavelength of 580 nm and the fluorescence having the wavelength of 580 nm calculated by the analyzing means in comparison with the threshold value stored in the storage means And when the average fluorescence lifetime is greater than or equal to the threshold value, a growth diagnosis result output means for outputting a diagnosis result indicating that the macroalgae sample is abnormal is provided.

  According to this macroalgae growth diagnostic apparatus, since the deterioration of the growth state of the large algae can be diagnosed at an early stage, it is possible to take measures to eliminate the cause of the deterioration of the growth state of the large algae, and Productivity can also be improved.

  As a desirable mode, the threshold is preferably 0.3 ns. Since the fluorescence lifetime of fluorescence at a wavelength of 580 nm is prolonged even in the initial stage when the growth state of macroalgae is degraded, the macroalgae growth diagnostic device provides a clear standard for determining the degradation of macroalgae growth state In addition, it is possible to suppress the possibility that differences in the evaluation of seaweed will occur depending on the measurer. In addition, since the macroalgae growth diagnostic apparatus only needs to calculate and evaluate the fluorescence lifetime of the fluorescence having a wavelength of 580 nm, the measurement and evaluation can be performed quickly, resulting in a simple measurement system, and the apparatus itself is low in cost. .

  As a desirable mode, the growth diagnosis result output means, when the average fluorescence lifetime of the fluorescence of the wavelength of 580 nm calculated by the analysis means is less than the threshold, compared with the threshold value stored in the storage means, It is preferable to output a diagnosis result indicating that the macroalgae sample is normal. As a result, the device for diagnosing the growth of large algae provides a clear standard for determining the deterioration of the growth state of large algae, distinguishes healthy large algae as normal, and prevents large-scale algae in the initial stage where it is difficult to make a visual judgment. It can be distinguished from macroalgae whose growth state has deteriorated.

  As a desirable mode, the storage means stores information on the correlation between the average fluorescence lifetime of the wavelength of 580 nm and the degree of deterioration of the growth state of macroalgae, and the growth diagnosis result output means is the abnormality. The degree of deterioration of the growth state of the macroalgae is output based on the average fluorescence lifetime of the wavelength of 580 nm calculated by the analysis means and the information on the correlation of the macroalgae sample from which the diagnosis result is output. It is preferable. Thereby, it is possible to quantitatively evaluate the degree of deterioration of the growth state of the large algae with respect to the sample of the large algae diagnosed as abnormal by the apparatus for diagnosing the growth of the large algae. For example, depending on the degree of deterioration of the growth state of the large algae, the fishermen can take measures to remove the cause of the deterioration of the growth state of the large algae.

  As a desirable mode, the detector detects fluorescence at the wavelength of chlorophyll fluorescence, and the storage means has a ratio of the fluorescence intensity at the wavelength of 580 nm to the fluorescence intensity at the wavelength of the chlorophyll fluorescence and the growth state of macroalgae. Information on the correlation with the degree of deterioration of the chlorophyll fluorescence of the macroalgae sample for which the diagnosis result is output as abnormal, and the analysis means It is preferable that the ratio of the fluorescence intensity of the wavelength is calculated, and the growth diagnosis result output means outputs the degree of deterioration of the growth state of the large algae from the fluorescence intensity ratio calculated by the analysis means and the correlation information. Thereby, it is possible to quantitatively evaluate the degree of deterioration of the growth state of the large algae with respect to the sample of the large algae diagnosed as abnormal by the apparatus for diagnosing the growth of the large algae. For example, depending on the degree of deterioration of the growth state of the large algae, the fishermen can take measures to remove the cause of the deterioration of the growth state of the large algae.

  In order to solve the above-described problems and achieve the object, the macroalgae growth diagnosis method irradiates a sample of macroalgae with pulsed excitation light including a wavelength near the absorption peak wavelength of phycoerythrin, and the pulsed excitation light Receiving fluorescence emitted from the sample of the macroalgae irradiated with, and diagnosing that the sample of the macroalgae is abnormal when the fluorescence lifetime of the fluorescence at 580 nm is 0.3 ns or more. And

  According to this method for diagnosing macroalgae growth, the deterioration of the growth state of the macroalgae can be diagnosed at an early stage, so it is possible to take measures to eliminate the cause of the degradation of the macroalgae growth state, Productivity can also be improved.

Desirable embodiments, the sample of the macroalgae diagnosed as abnormal based on the average fluorescence lifetime of the wavelength of 580 nm, and diagnose child degree of deterioration of the state of growth of macroalgae is preferred. Thereby, the growth diagnostic method for macroalgae can quantitatively evaluate the degree of deterioration of the growth state of macroalgae for the macroalgae samples diagnosed as abnormal. As a result, the number of diagnoses can be reduced, and the labor for diagnosing the growth of large algae can be suppressed. For example, depending on the degree of deterioration of the growth state of the large algae, the fishermen can take measures to remove the cause of the deterioration of the growth state of the large algae.

  As a desirable embodiment, the method for diagnosing macroalgae growth is based on a ratio of the fluorescence intensity of the fluorescence at 580 nm to the fluorescence intensity of the fluorescence chlorophyll fluorescence for the macroalgae sample diagnosed as abnormal. It is preferable to diagnose the degree of deterioration of the growth state of macroalgae. Thereby, the growth diagnostic method for macroalgae can quantitatively evaluate the degree of deterioration of the growth state of macroalgae for the macroalgae samples diagnosed as abnormal. As a result, the number of diagnoses can be reduced, and the labor for diagnosing the growth of large algae can be suppressed. For example, depending on the degree of deterioration of the growth state of the large algae, the fishermen can take measures to remove the cause of the deterioration of the growth state of the large algae.

  ADVANTAGE OF THE INVENTION According to this invention, the growth diagnostic apparatus of the large algae which can diagnose the deterioration of the growth state of a large algae at an initial stage, and the growth diagnostic method of a large algae can be provided.

FIG. 1 is a block diagram showing a schematic configuration of a macroalgae growth diagnostic apparatus. FIG. 2 is a flowchart showing processing steps of the macroalgae growth diagnosis method. FIG. 3 is an explanatory diagram showing the fluorescence spectrum of normal Susabiori. FIG. 4 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to a normal sausobiori reference sample 1. FIG. 5 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to a normal sausobiori reference sample 1. FIG. 6 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak at 580 nm measured for the second time with respect to the normal reference sample 1 of Susavino. FIG. 7 is an explanatory diagram showing an appearance example showing the progress of disease progression of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4. FIG. 8 is an explanatory diagram showing fluorescence spectra of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4 shown in FIG. FIG. 9 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 1. FIG. 10 is an explanatory diagram showing a fluorescence decay curve at a wavelength peak of 720 nm with respect to Evaluation Example 1. FIG. 11 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 1. FIG. FIG. 12 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 2. FIG. 13 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 2. FIG. 14 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 2. FIG. 15 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 3. FIG. 16 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 3. FIG. 17 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 3. FIG. 18 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 4. FIG. 19 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 4. FIG. 20 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 4. FIG. 21 is an explanatory diagram showing fluorescence decay curves of a wavelength peak of 580 nm with respect to a normal sausobiori reference sample 1 and evaluation example 4. FIG. 22 shows the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ and the average fluorescence lifetime of the wavelength of 580 nm and the wavelength of 720 nm for the reference sample 1, evaluation example 1, evaluation example 2, evaluation example 3 and evaluation example 4. It is explanatory drawing explaining a relationship. FIG. 23 is an explanatory diagram illustrating energy transfer in macroalgae. FIG. 24 is an explanatory diagram for explaining an absorption spectrum of a photosynthetic pigment. FIG. 25 is an explanatory diagram showing the fluorescence spectrum of Susabiori soaked in fresh water for 15 minutes. FIG. 26 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 5. FIG. 27 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 5. FIG. 28 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 5. FIG. 29 is an explanatory diagram for explaining the relationship between the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ and the average fluorescence lifetime of the wavelength of 580 nm and the wavelength of 720 nm in the evaluation example 5 in addition to the evaluation of FIG.

  Hereinafter, the present invention will be described in detail with reference to the drawings. The present invention is not limited by the following modes for carrying out the invention (hereinafter referred to as embodiments). In addition, constituent elements in the following embodiments include those that can be easily assumed by those skilled in the art, those that are substantially the same, and those in a so-called equivalent range. Furthermore, the constituent elements disclosed in the following embodiments can be appropriately combined.

  FIG. 1 is a block diagram showing a schematic configuration of a macroalgae growth diagnostic apparatus. The large algae growth diagnostic apparatus 1 includes a control device 2, a pulse light source 3, a detector 4, a spectroscope 5, a display means 6, and a measurement circuit 8, and applies pulse excitation light to a sample 7 of the large algae. To receive the fluorescence emitted from the macroalgae sample 7.

  The control device 2 is a computer such as a personal computer (PC), and includes a central processing unit (CPU), a read only memory (ROM), a random access memory (RAM), a storage device, an input / output interface, and a pulse. A control circuit for controlling the light source 3 and the detector 4 is included. The control device 2 stores programs such as BIOS (Basic Input Output System) in the ROM. The storage device is storage means 23, for example, an HDD (Hard Disk Drive), a flash memory, etc., and stores information on threshold values of the average fluorescence lifetime, which will be described later, and also stores operating system programs and application programs. ing. The CPU is a calculation means, and executes various programs such as the analysis means 21 and the growth diagnosis result output means 22 by executing a program stored in the ROM and the storage means 23 while using the RAM as a work area. Can be realized. The control device 2 is connected to the measurement circuit 8 and the display means 6 via an input / output interface. The measurement circuit 8 is connected to the pulse light source 3 and the detector 4. The display means 6 is a device that displays an image, such as a CRT (Cathode Ray Tube) or a liquid crystal display.

  FIG. 2 is a flowchart showing processing steps of the macroalgae growth diagnosis method. As shown in FIG. 2, the analysis means 21 can instruct the measurement circuit 8 that controls the pulse light source 3 and the detector 4 by the CPU to synchronize the start of measurement and start measurement (step S <b> 1).

  The pulsed light source 3 may be a light emitting diode (LED) or laser diode having a wavelength near the absorption peak of phycoerythrin and having a wavelength of excited light of 450 nm to 550 nm. In the following embodiment, a fluorescence spectrum measurement in which the pulse light source 3 is an excitation light source of a laser diode will be described. The fluorescence spectrum measurement is a method of detecting fluorescence emitted when the sample 7 is irradiated with excitation light to excite electrons in the photosynthetic dye and return to the original energy state. This method is non-destructive, quick and efficient measurement compared to conventional chemical analysis by destructive inspection, and in addition, remote measurement is possible.

  As the macroalgae sample 7, green algae such as Aonori and Aosa, brown algae such as kombu and wakame, red algae such as Susabiori and Tengusa can be used. As the macroalgae sample 7, it is more preferable to use red algae such as Susabinori, Asakusa Nori, Proboscis and Ogonori. The macroalgae sample 7 of the present embodiment uses Susabinori.

  The fluorescence emitted from the macroalgae sample 7 irradiated with the pulsed excitation light enters the detector 4 via the spectroscope 5. The spectroscope 5 is, for example, an interference filter or the like, and can mainly transmit the measurement wavelength component of fluorescence and allow the detector 4 to detect it. The detector 4 detects fluorescent photons, converts them into electrical signals via the measurement circuit 8, and sends them to the control device 2.

In the control device 2, the analysis means 21 calculates the average fluorescence lifetime of fluorescence having a wavelength of 580 nm (step S2). Of the plurality of components of the fluorescence lifetime (τ ₁ , τ ₂ ... Τ _i ), for example, when the fluorescence lifetime is 2 components (τ ₁ , τ ₂ ... Τ _i ), the following equation (1) is used. A fluorescence decay curve that is a function of time t is calculated by fitting (approximate) the detection electric signal of the detector 4 described above and performing deconvolution processing. Here, A ₁ and A ₂ are coefficients determined by fitting (approximation). The fluorescence lifetime calculated by the analysis means 21 may be one component or three or more components.

  Next, in the control device 2, the analysis means 21 calculates the average fluorescence lifetime τ using the following formula (2).

  Next, the growth diagnosis result output unit 22 compares the average fluorescence lifetime τ of the fluorescence having a wavelength of 580 nm calculated by the analysis unit 21 in step S2 with the threshold value stored in the storage unit 23, and when it is equal to or greater than the threshold value (step S21). (S3, Yes), the growth diagnosis result output means 22 outputs an abnormality to the display means 6 (step S4). For this reason, the display means 6 can inform the measurer of the abnormality. The means by which the growth diagnosis result output means 22 outputs an abnormality is not limited to the display means 6, and may be any means that can be perceived by the measurer, such as sound, vibration, and printing on paper.

  In the present embodiment, the threshold value stored in the storage unit 23 is 0.3 ns. The threshold value stored in the storage unit 23 may be 0.5 ns, but the threshold value stored in the storage unit 23 is 0.3 ns, so the deterioration of the growth state of the large algae can be diagnosed at an early stage. It is possible to take measures to eliminate the cause of deterioration of the growth state of the large algae, and it is possible to improve the productivity of the cultivation of the large algae.

  The value of the average fluorescence lifetime at a wavelength of 580 nm increases according to the degree of deterioration of the growth state of macroalgae. For this reason, when the storage unit 23 stores information on the correlation between the value of the average fluorescence lifetime at a wavelength of 580 nm and the degree of deterioration of the growth state of the large algae, the large-sized diagnostic result is output as abnormal. Based on the average fluorescence lifetime of the wavelength of 580 nm calculated by the analysis means 21 of the algae sample 7 and the correlation information described above, the degree of deterioration of the growth state of the large algae is calculated (step S5). Next, the growth diagnosis result output means 22 outputs the degree of deterioration of the growth state of the large algae calculated in step S5 to the display means 6 (step S6). For this reason, the display means 6 can inform the measurer of the degree of deterioration of the growth state of the macroalgae. The means by which the growth diagnosis result output means 22 outputs the degree of deterioration of the growth state of the large algae is not limited to the display means 6, and may be any means that can be perceived by the measurer, such as sound, vibration, and printing on paper.

  Thereby, the growth diagnostic apparatus 1 for large algae according to the present embodiment can quantitatively evaluate the degree of deterioration of the growth state of the large algae for the large algae sample diagnosed as abnormal in step S4 described above. . As a result, it is not necessary to quantitatively evaluate the degree of deterioration of the growth state of the large algae for all the samples, and the large algae growth diagnostic apparatus 1 reduces the calculation load and obtains the results for all the samples. Will improve. After the growth diagnosis result output means 22 processes step S6, the process ends. In addition, when there is no need to output the degree of deterioration of the growth state of the large algae, the processing may be terminated after the growth diagnosis result output means 22 processes step S4.

  As described above, the growth diagnosis result output unit 22 compares the average fluorescence lifetime τ of the fluorescence having a wavelength of 580 nm calculated in step S2 shown in FIG. 2 with the threshold value stored in the storage unit 23, and is less than the threshold value. (Step S3, No), the growth diagnosis result output means 22 outputs normality to the display means 6 (Step S7). For this reason, the display means 6 can inform the measurer of normality. Accordingly, the macroalgae growth diagnosis apparatus 1 provides a clear standard for judging the deterioration of the growth state of the macroalgae, identifies healthy macroalgae as normal, and the macroalgae in the initial stage where visual judgment is difficult to make. It can be distinguished from large algae whose growth state has deteriorated. Note that the means by which the growth diagnosis result output means 22 outputs normality is not limited to the display means 6 and may be any means that can be perceived by the measurer, such as sound, vibration, and printing on paper. Step S7 in which the growth diagnosis result output means 22 outputs normality can be omitted.

(Modification)
When the storage means 23 stores information on the correlation between the ratio of the fluorescence intensity of the wavelength of 580 nm to the fluorescence intensity of the wavelength of chlorophyll fluorescence and the degree of deterioration of the growth state of the large algae, the growth diagnosis result output means 22 You may perform the process of step S5 and step S6 shown in FIG.

  The macroalgae sample 7 expresses chlorophyll fluorescence in the range of 670 nm to 750 nm, for example. For example, it has been found that there is a correlation between the ratio of the fluorescence intensity at a wavelength of 580 nm to the fluorescence intensity at a wavelength of 720 nm or 685 nm and the degree of deterioration of the growth state of macroalgae. Therefore, the storage means 23 stores in advance information on the correlation between the ratio of the fluorescence intensity of the wavelength of 580 nm to the fluorescence intensity of the wavelength of chlorophyll fluorescence in the range of 670 nm to 750 nm and the degree of deterioration of the growth state of the large algae. Keep it. Then, the ratio of the fluorescence intensity of the wavelength of 580 nm to the intensity of chlorophyll fluorescence in the range of 670 nm to 750 nm of the macroalgae sample 7 from which the diagnostic result indicating that the growth diagnosis result output means 22 is abnormal is output (step S5). ). Next, based on the ratio of the fluorescence intensity of the wavelength of 580 nm to the intensity of chlorophyll fluorescence in the range of 670 nm to 750 nm calculated by the analysis means 21, the growth diagnosis result output means 22 stores 670 nm to 750 nm stored in the storage means 23. From the correlation between the ratio of the fluorescence intensity at the wavelength of 580 nm to the intensity of chlorophyll fluorescence in the range of γ and the degree of deterioration of the growth state of the large algae, the degree of deterioration of the growth state of the corresponding large algae is calculated (step S6). Thereby, the growth diagnostic apparatus 1 for large algae according to the present embodiment can quantitatively evaluate the degree of deterioration of the growth state of the large algae for the large algae sample diagnosed as abnormal in step S4 described above. . As a result, it is not necessary to quantitatively evaluate the degree of deterioration of the growth state of the large algae for all the samples, and the large algae growth diagnostic apparatus 1 reduces the calculation load and obtains the results for all the samples. Will improve. After the growth diagnosis result output means 22 processes step S6, the process ends.

  As described above, the macroalgae growth diagnosis method according to the embodiment irradiates the macroalgae sample 1 with pulsed excitation light including a wavelength near the absorption peak wavelength of phycoerythrin, and is emitted from the macroalgae sample 7. When the average fluorescence lifetime at 580 nm is 0.3 ns or more, it is diagnosed that the macroalgae sample is abnormal. According to this method for diagnosing macroalgae growth, the deterioration of the growth state of the macroalgae can be diagnosed at an early stage, so it is possible to take measures to eliminate the cause of the degradation of the macroalgae growth state, Productivity can also be improved.

  The method for diagnosing macroalgae is based on the ratio of the fluorescence intensity at 580 nm to the chlorophyll fluorescence intensity in the range of 670 nm to 750 nm of the macroalgae sample 7 diagnosed as abnormal. The degree of deterioration can be diagnosed. Thereby, the growth diagnostic method for macroalgae can quantitatively evaluate the degree of deterioration of the growth state of macroalgae for the macroalgae samples diagnosed as abnormal. As a result, the number of diagnoses can be reduced, and the labor for diagnosing the growth of large algae can be suppressed. For example, depending on the degree of deterioration of the growth state of the large algae, the fishermen can take measures to remove the cause of the deterioration of the growth state of the large algae.

(Evaluation)
An evaluation example of the macroalgae growth diagnosis method and the macroalgae growth diagnosis apparatus 1 of the present embodiment will be described with reference to FIGS. FIG. 3 is an explanatory diagram showing the fluorescence spectrum of normal Susabiori. The reference sample 1 and the reference sample 2 are sasabinori cultivated by the Shin Futtsu Fishery Cooperative in December 2012. The wavelength of the excitation light was measured using a 470 nm excitation light in the vicinity of the absorption peak wavelength of phycoerythrin, which is an excitation wavelength suitable for the growth diagnosis of Susabiori. The fluorescence wavelengths measured were 580 nm of the fluorescence wavelength considered to be attributable to phycoerythrin, 660 nm of the fluorescence wavelength believed to be attributable to allophycocyanin, and 685 nm and 720 nm of the fluorescence wavelengths believed to be attributable to chlorophyll a.

  As shown in FIG. 3, both the reference sample 1 and the reference sample 2 have the same fluorescence spectrum. Next, for the reference sample 1, the macroalgae growth diagnosis apparatus 1 of the present embodiment processed the macroalgae growth diagnosis method. FIG. 4 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to a normal sausobiori reference sample 1. FIG. 5 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to a normal sausobiori reference sample 1. FIG. 6 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak at 580 nm measured for the second time with respect to the normal reference sample 1 of Susavino. 4 to 6 show an apparatus response function (IRF) when the fluorescence lifetime is measured with respect to a normal sausobiori reference sample 1. FIG. As shown in FIGS. 4 and 6, the 580 nm peak was measured twice for confirmation, but it was confirmed that there was no significant difference in the measurement results and there was no alteration of the sample. Reference sample 2 was similarly evaluated.

  From the above-mentioned formulas (1) and (2), in the case of the fluorescence peak at 580 nm as shown in FIGS. 4 and 6, the average fluorescence lifetime τ of normal Susavino was 0.12 ns or more and 0.2 ns or less. From the above-mentioned formulas (1) and (2), in the case of a fluorescence peak at 720 nm as shown in FIG. 5, the average fluorescence lifetime τ of normal Susavino was 0.12 ns or more and 0.13 ns or less.

  Next, as an example of the deterioration of the growth state of macroalgae, Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4 of Susabiori infected with scabbed disease were prepared. FIG. 7 is an explanatory diagram showing an appearance example showing the progress of disease progression of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4. As shown in FIG. 7, the measurer performs sensory evaluation of the appearance of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3 and Evaluation Example 4, and evaluation example 1 and evaluation example are as indicated by arrows indicating the disease progression state. 2. It has been confirmed that the disease is progressed in the order of Evaluation Example 3 and Evaluation Example 4. Red-spotted disease can cause red rust spots due to bacterial infestation, which can cause deterioration of the leaves of Susabiori.

  FIG. 8 is an explanatory diagram showing fluorescence spectra of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4 shown in FIG. For comparison, FIG. 8 also shows a normal fluorescence spectrum (reference sample 1). The excitation light source is a laser diode with a wavelength of 470 nm used for fluorescence lifetime measurement. As shown in FIG. 8, the intensity of the peak considered to be caused by phycoerythrin near 580 nm is relatively increased when infected with scabbed disease. As shown in FIG. 8, the peaks considered to be caused by phycoerythrin near 580 nm increase in the order of reference sample 1, evaluation example 1, evaluation example 2, evaluation example 3, and evaluation example 4.

  FIG. 9 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 1. FIG. 10 is an explanatory diagram showing a fluorescence decay curve at a wavelength peak of 720 nm with respect to Evaluation Example 1. FIG. 11 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 1. FIG. 9 to 11 show an apparatus response function (IRF) when the fluorescence lifetime is measured for Evaluation Example 1. FIG.

  FIG. 12 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 2. FIG. 13 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 2. FIG. 14 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 2. FIGS. 12 to 14 show the apparatus response function (IRF) when the fluorescence lifetime is measured for the evaluation example 2. FIG.

  FIG. 15 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 3. FIG. 16 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 3. FIG. 17 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 3. 15 to 17 show an apparatus response function (IRF) when the fluorescence lifetime is measured in the evaluation example 3.

  FIG. 18 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 4. FIG. 19 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 4. FIG. 20 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 4. 18 to 20 show an apparatus response function (IRF) when the fluorescence lifetime is measured for Evaluation Example 4. FIG.

For Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4, the macroalgae growth diagnosis apparatus 1 of this embodiment processed the macroalgae growth diagnosis method. Then, for the evaluation example 1, the evaluation example 2, the evaluation example 3 and the evaluation example 4, fitting is performed by assuming the fluorescence lifetime of two components from the above formulas (1) and (2), and the average fluorescence lifetime is calculated. did. FIG. 21 is an explanatory diagram showing fluorescence decay curves of a wavelength peak of 580 nm with respect to a normal sausobiori reference sample 1 and evaluation example 4. As shown in FIG. 21, it can be seen that the approximation of the fluorescence lifetime of the formula (1) is an appropriate fitting with respect to the normal Susavinori reference sample 1 and the evaluation example 4. The evaluation results are shown in FIG. FIG. 22 shows the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ and the average fluorescence lifetime of the wavelength of 580 nm and the wavelength of 720 nm for the reference sample 1, evaluation example 1, evaluation example 2, evaluation example 3 and evaluation example 4. It is explanatory drawing explaining a relationship. The solid triangle of the reference sample 1 is the average fluorescence lifetime at a wavelength of 580 nm, and the dashed triangle of the reference sample 1 indicates the average fluorescence lifetime at a wavelength of 720 nm. The solid circle in Evaluation Example 1 is the average fluorescence lifetime at a wavelength of 580 nm, and the dashed circle in Evaluation Example 1 indicates the average fluorescence lifetime at a wavelength of 720 nm. The solid-line square in Evaluation Example 2 is the average fluorescence lifetime at a wavelength of 580 nm, and the dashed-line square in Evaluation Example 2 indicates the average fluorescence lifetime at a wavelength of 720 nm. The solid diamond in Evaluation Example 3 is the average fluorescence lifetime at a wavelength of 580 nm, and the dashed diamond in Evaluation Example 3 is the average fluorescence lifetime at a wavelength of 720 nm. The solid pentagon in Evaluation Example 4 is the average fluorescence lifetime at a wavelength of 580 nm, and the dashed pentagon in Evaluation Example 4 indicates the average fluorescence lifetime at a wavelength of 720 nm.

When infected with scarlet disease, the peak intensity I ₅₈₀ near 580 nm relatively increases. That is, when the scab is progressed, the intensity ratio I ₅₈₀ / I _{720 of the} fluorescence peak increases. The ratio of the fluorescence intensity at the wavelength of 580 nm to the fluorescence intensity at the wavelength of 720 nm, that is, the peak intensity ratio I ₅₈₀ / I ₇₂₀ is shown in Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4 as shown in FIG. It can be seen that there is a correlation that is arranged in the order in which the disease is progressed and the disease progresses. First, focusing on the fluorescence lifetime at 580 nm, the average fluorescence lifetime becomes longer as the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ increases. From this, it is understood that the diagnosis of scabbed disease is possible by measuring the fluorescence lifetime. The fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ correlates with the degree of deterioration of the growth state of macroalgae, and the storage means 23 previously stores the ratio of the fluorescence intensity at the wavelength of 580 nm to the fluorescence intensity at the wavelength of 720 nm, and Information of correlation with the degree of deterioration of the growth state of macroalgae can be stored. However, the average fluorescence lifetime at 580 nm is greatly changed even in the case of mild scab that hardly changes at the intensity ratio I ₅₈₀ / I ₇₂₀ . The macroalgae growth diagnosis method according to the embodiment irradiates the macroalgae sample 7 with pulsed excitation light including a wavelength near the absorption peak wavelength of phycoerythrin, and receives fluorescence emitted from the macroalgae sample 7. When the average fluorescence lifetime at 580 nm is 0.3 ns or more, it is diagnosed that the sample of macroalgae is abnormal. Accordingly, the method for diagnosing macroalgae according to the embodiment can perform measurement with higher sensitivity in the early scabbed disease by comparing the average fluorescence lifetime of 580 nm with a threshold value.

  According to the macroalgae growth diagnostic apparatus and macroalgae growth diagnostic method according to the present embodiment, the deterioration of the growth state of the macroalgae can be diagnosed at an early stage, so that the cause of the deterioration of the macroalgae growth state can be removed. It is also possible to improve the productivity of large algae culture.

The average fluorescence lifetime of 580 nm of the reference sample 1 is in the range of 0.3 ns or less, and the average fluorescence lifetime of 580 nm of the evaluation example 1 increases more rapidly than 0.3 ns. As shown in FIG. 22, the average fluorescence lifetime of 580 nm has a correlation that increases in the order of evaluation and progression of disease in the order of evaluation example 1, evaluation example 2, evaluation example 3, and evaluation example 4. For this reason, the value of the average fluorescence lifetime of 580 nm of Evaluation Example 1, Evaluation Example 2, Evaluation Example 3, and Evaluation Example 4 increases according to the degree of deterioration of the growth state of the macroalgae. On the other hand, the average fluorescence lifetime at 720 nm is not significantly different between the values of the reference sample 1 and the evaluation example 1 (evaluation example 2), and increases almost in proportion to the intensity ratio I ₅₈₀ / I ₇₂₀ .

  FIG. 23 is an explanatory diagram illustrating energy transfer in macroalgae. FIG. 24 is an explanatory diagram for explaining an absorption spectrum of a photosynthetic pigment. As can be seen from FIGS. 23 and 24, the light having a wavelength of 470 nm used for the excitation light is mainly absorbed by phycoerythrin. The energy of light absorbed by phycoerythrin is transferred to chlorophyll a, which is a photosynthetic reaction center, through various processes. For this reason, both fluorescence at 580 nm which is considered to be attributable to phycoerythrin and fluorescence at 685 nm and 720 nm which are considered to be attributable to chlorophyll a are observed. In the case of normal sausobiori, the energy of light absorbed by phycoerythrin is smoothly transferred, and the energy transferred to chlorophyll a is smoothly used for photosynthesis. For this reason, it is considered that the lifetime of fluorescence at 580 nm, which is considered to be caused by phycoerythrin, and fluorescence of 685 nm or 720 nm, which is considered to be caused by chlorophyll a, is shortened to 0.1 ns or more and 0.2 ns or less. On the other hand, when infected with a scab, the transfer of energy absorbed by phycoerythrin to chlorophyll a is first inhibited at an early stage, so that the average fluorescence lifetime at 580 nm is considered to be long. Furthermore, when the disease is progressed, utilization of the energy transferred to chlorophyll a for photosynthesis is also inhibited, so that it is considered that the average fluorescence lifetime at 720 nm is prolonged. Such a reaction is different from the reaction of plants mainly composed of chlorophyll.

  As can be seen from FIG. 24, it is considered that the same evaluation can be performed by exciting with excitation light in the vicinity of the absorption peak of allophycocyanin and evaluating it with the average fluorescence lifetime of allophycocyanin fluorescence (for example, 660 nm). However, the absorption range of allophycocyanin is in the range of 550 nm to 700 nm, which overlaps the entire fluorescence wavelength range (about 550 nm to 800 nm), and the range of 550 nm to 600 nm overlaps with the absorption of phycoerythrin. Yes. For this reason, the excitation including the wavelength near the absorption peak wavelength of phycoerythrin (450 nm or more and 550 nm or less) than the excitation with the excitation light near the absorption peak of allophycocyanin and evaluating the fluorescence lifetime with the fluorescence of allophycocyanin. More accurate evaluation is possible by exciting with light and evaluating with the average fluorescence lifetime in the fluorescence of phycoerythrin. Similarly, the absorption range of phycocyanin overlaps with that of phycoerythrin and allophycocyanin, rather than being excited by excitation light near the absorption peak of phycocyanin and evaluated by the average fluorescence lifetime in the fluorescence of phycocyanin. More accurate evaluation is possible by exciting with excitation light including a wavelength in the vicinity of the absorption peak wavelength of phycoerythrin (450 nm or more and 550 nm or less) and evaluating with the average fluorescence lifetime in the fluorescence of phycoerythrin.

  As shown in FIG. 22, if the threshold value stored in the storage means 23 is 0.3 ns, the average fluorescence lifetime of 580 nm is prolonged due to the deterioration of the growth state of the large algae, and the deterioration of the growth state of the large algae is in the initial stage. Diagnose with As shown in FIG. 22, if the threshold value stored in the storage means 23 is 0.3 ns, the average fluorescence lifetime of 720 nm is not high in the initial stage, so the threshold value of 0.3 ns is set as the average fluorescence lifetime of 720 nm. There is a possibility of making a misjudgment if judged by contrast.

  FIG. 25 is an explanatory diagram showing the fluorescence spectrum of Susabiori soaked in fresh water for 15 minutes. In Evaluation Example 5, the effect of fresh water on the average fluorescence lifetime was examined in the case where Susabiori was soaked in fresh water for 15 minutes. The fluorescence spectrum also changes when soaking the water in fresh water, and the peak intensity at 580 nm increases in about several tens to 100 seconds after soaking in fresh water. FIG. 26 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm with respect to Evaluation Example 5. FIG. 27 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 720 nm with respect to Evaluation Example 5. FIG. 28 is an explanatory diagram showing a fluorescence decay curve of a wavelength peak of 580 nm measured for the second time in Evaluation Example 5. 26 to 27 show the apparatus response function (IRF) when the fluorescence lifetime is measured for Evaluation Example 5. FIG.

For Evaluation Example 5, the macroalgae growth diagnostic apparatus 1 of the present embodiment processed the macroalgae growth diagnostic method. And about the evaluation example 5, it assumed that the fluorescence lifetime of 2 components was assumed from the formula (1) and Formula (2) mentioned above, and the average fluorescence lifetime was calculated. FIG. 29 is an explanatory diagram for explaining the relationship between the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ and the average fluorescence lifetime of the wavelength of 580 nm and the wavelength of 720 nm in the evaluation example 5 in addition to the evaluation of FIG. The solid black hexagon in Evaluation Example 5 has an average fluorescence lifetime at a wavelength of 580 nm, and the broken hexagon in Evaluation Example 5 indicates an average fluorescence lifetime at a wavelength of 720 nm.

Susavinori has a relatively increased peak intensity I ₅₈₀ near 580 nm immersed in fresh water. That is, Susabinori increases the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ in response to stress such as fresh water. First, focusing on the fluorescence lifetime at 580 nm, the average fluorescence lifetime becomes longer as the fluorescence peak intensity ratio I ₅₈₀ / I ₇₂₀ increases. From this, it can be seen that according to the macroalgae growth diagnostic apparatus and macroalgae growth diagnostic method according to the present embodiment, it is possible to diagnose the deterioration of the growing environment such as water quality used for the culture of macroalgae. As a result, a fisherman engaged in the cultivation of macroalgae can take measures to eliminate the cause of the deterioration of the growth state of macroalgae, and the productivity of macroalgae culture can be improved.

DESCRIPTION OF SYMBOLS 1 Large algae growth diagnostic apparatus 2 Control apparatus 3 Pulsed light source 4 Detector 5 Spectrometer 6 Display means 7 Sample 8 Measuring circuit 21 Analyzing means 22 Growth diagnostic result output means 23 Storage means

Claims (7)

A pulsed light source that irradiates a sample of macroalgae with pulsed excitation light including a wavelength near the absorption peak wavelength of phycoerythrin;
A detector for detecting fluorescence having a wavelength of 580 nm among fluorescence emitted from the sample of the macroalgae irradiated with the pulsed excitation light;
Storage means for storing threshold information regarding fluorescence lifetime;
Based on an output signal output from the detector, an analysis means for calculating an average fluorescence lifetime of the fluorescence having the wavelength of 580 nm,
Compared with the threshold value stored in the storage means, when the average fluorescence lifetime of the fluorescence of the wavelength of 580 nm calculated by the analysis means is equal to or greater than the threshold value, a diagnosis result that the sample of the macroalgae is abnormal A diagnostic diagnosis result output means for outputting;
With
The growth diagnostic apparatus for macroalgae , wherein the threshold is 0.3 ns .   The growth diagnosis result output means compares the threshold value stored in the storage means with the sample of the macroalgae when the average fluorescence lifetime of the fluorescence having the wavelength of 580 nm calculated by the analysis means is less than the threshold value. The apparatus for diagnosing macroalgae growth according to claim 1, which outputs a diagnosis result of normal. The storage means stores information on the correlation between the average fluorescence lifetime of the wavelength of 580 nm and the degree of deterioration of the growth state of macroalgae,
The growth diagnosis result output means is based on the average fluorescence lifetime of the wavelength of 580 nm calculated by the analysis means and the correlation information of the macroalgae sample whose diagnosis result is output as abnormal. The growth diagnostic apparatus for macroalgae according to claim 1 or 2 , which outputs a degree of deterioration of the growth state of macroalgae. The detector detects fluorescence of the wavelength of chlorophyll fluorescence,
The storage means stores information on the correlation between the ratio of the fluorescence intensity at the wavelength of 580 nm to the fluorescence intensity at the wavelength of the chlorophyll fluorescence and the degree of deterioration of the growth state of the macroalgae,
The analysis means calculates a ratio of the fluorescence intensity of the wavelength of 580 nm to the fluorescence intensity of the wavelength of the chlorophyll fluorescence of the macroalgae sample whose diagnostic result is output as the abnormality,
3. The growth of the large algae according to claim 1 or 2 , wherein the growth diagnosis result output means outputs a degree of deterioration of the growth state of the large algae from the fluorescence intensity ratio calculated by the analysis means and the correlation information. Diagnostic device. Irradiate a sample of macroalgae with pulsed excitation light containing a wavelength near the absorption peak wavelength of phycoerythrin,
Receiving fluorescence emitted from the macroalgae sample irradiated with the pulsed excitation light;
A method for diagnosing the growth of a large algae, comprising diagnosing that the sample of the large algae is abnormal when the fluorescence lifetime of the fluorescence at 580 nm is 0.3 ns or longer. 6. The diagnosis of growth of macroalgae according to claim 5 , wherein the degree of deterioration of the growth state of macroalgae is diagnosed based on an average fluorescence lifetime of a wavelength of 580 nm for the macroalgae sample diagnosed as abnormal. Method. Diagnosing the degree of deterioration of the growth state of macroalgae based on the ratio of the fluorescence intensity of 580 nm of the fluorescence to the fluorescence intensity of the fluorescence chlorophyll fluorescence for the macroalgae sample diagnosed as abnormal, The method for diagnosing the growth of macroalgae according to claim 5 .