Classifications

• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
  • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
  • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
  • G01N21/64—Fluorescence; Phosphorescence
  • G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
  • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
  • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
  • G01N21/64—Fluorescence; Phosphorescence
  • G01N21/6402—Atomic fluorescence; Laser induced fluorescence
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
  • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
  • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
  • G01N21/64—Fluorescence; Phosphorescence
  • G01N21/645—Specially adapted constructive features of fluorimeters
  • G01N21/6456—Spatial resolved fluorescence measurements; Imaging
• • G—PHYSICS
  • G01—MEASURING; TESTING
  • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
  • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
  • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
  • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
  • G01N21/64—Fluorescence; Phosphorescence
  • G01N2021/6417—Spectrofluorimetric devices
  • G01N2021/6419—Excitation at two or more wavelengths

Definitions

• Device for non-destructive analysis of plants and vehicle comprising such an on-board device,
• the present invention relates to the field of the study of vegetation, more particularly plants, in the forestry and agricultural field, and relates to a device for the non-destructive analysis of plants, as well as a vehicle for analysis comprising such an on-board device.
• a device for the non-destructive analysis of plants as well as a vehicle for analysis comprising such an on-board device.
• the principle of fluorescence measurement is known, which consists in optically acquiring the image of the fluorescence of the vegetation, in particular of the leaves, induced by a brief light excitation, coming from a laser beam projected on the latter. .
• Multi-Color Fluorescence Imaging ", JONAS JOHANSSON & al., A device for remote measurement by laser fluorescence and for collecting images produced by fluorescence at different wavelengths.
• fluorescence is measured globally that is the area of interest in the image by the interposition of a spectrophotometer which scans the different wavelengths successively and a second channel allows the acquisition of two simultaneous images at two different frequencies.
• the aforementioned known devices do not provide enough information to allow precise analysis and establishment of a diagnosis after a single measurement session, and none of these known devices is suitable for carrying out, with the same device. , a global measurement in situ and a measurement on a particular sampled element.
• the subject of the invention is a device for the non-destructive analysis of plants, and more generally of plants, by measuring the fluorescence induced under laser excitation, comprising a unit for emitting a laser excitation beam.
• the transmitting unit provides at least two wavelengths of laser excitation and in that the measurement and image taking unit comprises means for forming, from the same fluorescence beam emitted by the plant (s), simultaneously or successively, at least two secondary beams, each having its own wavelength and each constituting a fluorescence image at the level of a matrix sensor, on the entire surface of the latter (successive acquisition of images) or on a part of the latter's surface, distinct for each secondary beam (simultaneous acquisition of images).
• the subject of the invention is also a vehicle for in situ analysis of plants and, more generally of plants, characterized in that it comprises at least one analysis device as mentioned above, mounted in an orientable and tiltable manner. on a telescopic stem.
• FIG. 1 is a schematic view of an analysis device according to the invention
• Figure 2 is a schematic view of the synchronization loop forming part of the device shown in Figure 1
• Figure 3 is a schematic view showing the various components of the device according to the invention
• FIGS. 4A and 4B are views in side elevation of an analysis vehicle according to the invention, with the analysis device respectively in an external analysis position and in an analysis position of a sample at l vehicle interior also forming part of the invention
• FIGS. 5 to 11 represent images obtained by means of the analysis device according to the invention by fluorescence induced in parts of plants
• FIGS. 12 to 14 represent examples of flowcharts of algorithms for executing certain functions of the analysis device according to the invention.
• the device for the non-destructive analysis of plants, and more generally plants, by measuring the fluorescence induced under laser excitation comprises a unit 3 for emitting a beam d laser excitation 6 calibrated, a unit 4 for measuring and taking images of the fluorescence emitted by the irradiated plant (s) 2 and a digital processing, storage and editing or display unit of images collected, associated with a computer unit for controlling and managing the operation of the device 1.
• the emission unit 3 supplies at least two wavelengths of laser excitation and the measurement and image taking unit 4 comprises means 7, 8, 9 for forming, at from the same fluorescence beam 6 ′ emitted by the plant (s) 2, simultaneously or successively, at least two secondary beams, each having its own wavelength and each constituting a fluorescence image at the level of a matrix sensor 10, over the entire surface of the latter (successive acquisition of images) or over a part of the surface of the latter, distinct for each secondary beam (simultaneous acquisition of images).
• the means 8 of FIG. 1 comprises several interference filters (at least two and preferably four) mounted on a disc-shaped filter holder whose motorized rotation movement, controlled by the 'control and management computer unit, brings said filters into the light path of the fluorescence beam 6'.
• the means 8 comprises elements for dividing the fluorescence beam 6 'into at least two, preferably four, secondary beams of distinct wavelengths each of which is directed to an area of the matrix sensor 10 which is specially assigned to it.
• each treated by one of the interference filters 8 and the images collected by the aforementioned unit 4 are, for example, taken at four different wavelengths which correspond to the maxima of blue fluorescence.
• green at 440 nm and 520 nm (F440 and F520) and chlorophyll fluorescence at 690 nm and 740 nm (F690 and F740) the beam 6 of the excitation laser advantageously emitting in the ultraviolet preferably from 380 nm to 390 nm, and in the green, preferably from 560 nm to 600 nm.
• the excitation in the green will allow, on the one hand, a greater depth of penetration and, on the other hand, a more direct excitation of the chlorophyll, while the blue excitation will not only provide information on the blue fluorescence and green but also on the efficiency of energy transfer to chloroplasts.
• the durations of the opening sequences of the assembly 9 [light shutter / intensifier] forming part of the unit 4 for measuring and taking images and the durations of the laser pulses of the emission unit 3 are correlated with each other and that the operations of the matrix sensor 10, of the laser device 3 ′ of the emission unit 3 and of the assembly 9 shutter / light intensifier are synchronized with each other via a corresponding loop control circuit.
• This arrangement makes it possible to overcome the electromagnetic disturbances usually induced at the level of optical matrix sensors 10 of CCD type (charge coupled circuit) by the switching edges of the voltage pulses controlling the light intensifier tube adjacent to said sensors.
• the opening time of the shutter / light intensifier assembly can advantageously be around 30 nanoseconds for a width at half-height of the laser pulses of 10 nanoseconds.
• the synchronization or loop control circuit consists, firstly, by a burst signal generator 11, triggered by the CCD camera forming the matrix sensor 10 and controlled by the computer control and management unit, on the other hand, by an adjustable delay line 12 ensuring the transmission of said burst signals, in particular towards the trigger input of the laser device 3 'and, finally, a module 13 triggering of the assembly 9 [shutter / intensifier of light], receiving the burst signals transmitted by the delay line 12 with a time offset determined with respect to the laser device 3 ', said offset being a function of the distance between the device 1 and the plant (s) or area of vegetation to be analyzed, possibly measured using a range finder.
• the triggering of the burst signal generator 11 is advantageously operated by the signal delivered by the CCIR composite video output of the CCD camera forming a matrix sensor 512 x 512 pixels, for example, divided into four image zones of 256 x 256 pixels, authorizing the accumulation, digitization and memorization of the fluorescence induced at the level of the vegetation to be analyzed at four different wavelengths, under the same conditions of excitation and illumination.
• the fluorescence images intended to be used are obtained by summing a plurality of temporary images, each of which results from the subtraction of two raw fluorescence images obtained during two consecutive opening phases of the 'set 9, a first with laser excitation and a second without laser excitation.
• the first raw fluorescence image is obtained by the fluorescent emission resulting from laser excitation and from excitation by sunlight and the second raw fluorescence image is obtained by the fluorescent emission resulting from the only excitation. by sunlight, each intermediate temporary image constituting a fluorescence image resulting solely from excitation by the laser.
• FIGS. 12 to 14 illustrate certain procedures for configuring and initializing the analysis device according to the invention, all of these procedures being able to be executed by the operator at the level of the computer unit by means of screen, keyboard, "mouse” or similar interfaces.
• the various executable software may, for example, be accessible by a selection program or "main menu" allowing access, by validation, to various options corresponding in particular to the possibilities of choice of the type of acquisition, of configuration of the acquisition , review and display of previous acquisitions, orientation and pointing of the analysis and launching of a parameterized acquisition.
• FIG. 12 of the appended drawings illustrates the possibilities and the configuration sequence in the case of a single acquisition without archiving
• FIG. 13 illustrates the possibilities and the configuration sequence in the case of multiple acquisitions or in sequences with archiving
• figure 14 illustrates the configuration sequence for physical initialization and adjustment of the analysis device 1.
• the present invention also relates to a vehicle for in situ analysis of plants and, more generally of plants, characterized in that it comprises at least one analysis device as described above, mounted in an orientable and tiltable manner. on a telescopic bracket 15 (FIGS. 4A and
• the installation of the analysis device 1 on a vehicle 14 makes it mobile and transportable.
• the interior space of this laboratory vehicle 14 is advantageously divided into two compartments, one of which is reserved for the operator and the associated digital processing, storage and editing and / or visualization unit. to the computer control and management unit, the other of which is fitted out for the storage of measurement equipment (interior measurements and transport).
• a mobile generator also transported by the vehicle
• stabilization jacks 16 can ensure the stalling of the vehicle 14 during the measurement operations.
• the different elements making up the emission unit 3 (laser device 3 ', beam enlarger 3 ") and the measurement and image taking unit 4 (objective 7, possible beam splitter, interference filters 8, together 9 [shutter / light intensifier], CCD camera 10) are preferably small and able to withstand strong accelerations without damage.
• said components of the transmission unit 3 and of the measurement and image-taking unit 4 are mounted on a support platform or orientable nacelle 17, forming two parallel structural assemblies in lines located in an air-conditioned enclosure 18.
• the latter is provided on its front face with an entry window and is provided with a connection cord to the vehicle 14 for the circulation of fluids and electrical energy towards the nacelle 17, the pumping of the laser by optical fiber and the transfer in return of the positioning information and digital images collected by the CCD camera 10.
• the size of the laser excitation beam 6 can be increased by a beam enlarger 3 "provided, for example, to cover a circle with a diameter of 0.30 m at a distance of 30 meters.
• the CCD 10 camera and the device Image intensifiers are oriented so as to reproduce an image of fluorescence of identical surface, confused with the irradiated zone. ''
• An optical range finder can supplement the aforementioned equipment so as to be able to accurately measure the distance between the vegetation and the equipment of the measuring head (instrumentation). It is from this information that the optical adjustments are made as well as the electronic delay adjustment.
• the vehicle 14 is provided with a hatch and the nacelle 17 carried by the bracket 15 is movable and orientable so that the analysis device 1 can be moved between a position in situ analysis of plants 2 arranged around the vehicle 14 and a position for analyzing a sample of plant 2 taken from the surrounding medium and placed in a sample holder inside the vehicle 14.
• the preferred laser excitation wavelengths are, on the one hand, between 380 nm and 390 nm and, on the other hand, between 560 nm and 600 nm, the inventors used to carry out their analyzes of the lengths 355 nm and 532 nm excitation waves (material available).
• the right leaf is healthy and the left leaf is contaminated with a pathogen (in this case red spider - invisible to the naked eye).
• a pathogen in this case red spider - invisible to the naked eye.
• FIG. 6 represents fluorescence images obtained from a sheet of Digitalis Purpurea, subjected to a treatment with herbicide (DCMU: 3 - (3,4 Dichlorophenyl - 1,1 - dimethylurea) by root.
• DCMU 3 - (3,4 Dichlorophenyl - 1,1 - dimethylurea
• the four images of this figure were collected at a wavelength of 690 nm under laser excitation at 355 nm and at different time intervals after watering the plant with a dilute DCMU solution (10 ' ⁇ M).
• FIG. 7 represents images obtained by making the pixel-to-pixel ratio of two fluorescence images from maize leaves, respectively, in good health (references), deficient in iron and deficient in zinc.
• FIG. 8 represents images of fluorescence obtained from vine leaves, respectively healthy, deficient in potassium, deficient in magnesium and affected by calcareous chlorosis. These images were collected at 690 nm, under laser excitation at
• potassium deficiency can be characterized by the appearance of very irregular fluorescence spots and of different intensities and that magnesium deficiency can be characterized by a greater fluorescence at the center of the leaf and rapidly decreasing in intensity towards the edge thereof.
• FIG. 9 represents fluorescence images obtained from tobacco leaves, respectively, in good health and exhibiting water stress. These images were collected at 740 nm, under laser excitation at 355 nm. J
• FIG. 11 represents fluorescence images obtained from a tobacco leaf subjected to a herbicide treatment (DCMU: 3 - (3,4 Dichlorophenyl - 1,1 - dimethylurea) by surface spraying. The three images of this figure were collected at 690 nm, under laser excitation of 355 nm.
• DCMU herbicide treatment
• Images a, b and c respectively represent a healthy leaf before treatment, the same leaf treated by spraying on its left half on the rear face and analyzed 10 minutes, then 30 minutes, after application of the herbicide.

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• Health & Medical Sciences (AREA)
• Physics & Mathematics (AREA)
• Life Sciences & Earth Sciences (AREA)
• Immunology (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Chemical & Material Sciences (AREA)
• Analytical Chemistry (AREA)
• Biochemistry (AREA)
• General Health & Medical Sciences (AREA)
• General Physics & Mathematics (AREA)
• Pathology (AREA)
• Molecular Biology (AREA)
• Biomedical Technology (AREA)
• Engineering & Computer Science (AREA)
• Optics & Photonics (AREA)
• Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)

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• 1997
  • 1997-12-22 LU LU90186A patent/LU90186B1/fr active
• 1998
  • 1998-12-21 EP EP98962535A patent/EP1040339A1/de not_active Withdrawn
  • 1998-12-21 JP JP2000525745A patent/JP2001527213A/ja active Pending
  • 1998-12-21 US US09/582,104 patent/US6573512B1/en not_active Expired - Fee Related
  • 1998-12-21 WO PCT/FR1998/002811 patent/WO1999032876A1/fr not_active Application Discontinuation
  • 1998-12-21 CA CA002315357A patent/CA2315357A1/fr not_active Abandoned
• 2000
  • 2000-04-28 NO NO20002263A patent/NO20002263L/no not_active Application Discontinuation

Non-Patent Citations (1)

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