ES2948924B2 - EQUIPMENT AND PROCEDURE FOR PERFORMING SPECTROPHOTOMETRY AND MOLECULAR FLUORIMETRY USING LED TECHNOLOGY - Google Patents

Description

DESCRIPTION

EQUIPMENT AND PROCEDURE FOR CARRYING OUT

SPECTROPHOTOMETRY AND MOLECULAR FLUORIMETRY USING

LED TECHNOLOGY

OBJECT OF THE INVENTION

It is the object of the present invention, just as the title of the invention establishes equipment and a procedure for carrying out a spectrofluorimetric analysis simultaneously with the same equipment.

The present invention is characterized by the special design and configuration of each and every one of the elements that the equipment has in such a way that it allows the simultaneous measurement and analysis with the same equipment of the spectrophotometric and fluorimetric response with respect to a light received from some LEDs, in addition to the equipment not being conditioned by the geometry of the sample specimens used.

The equipment also allows the possibility of varying the intensity or brightness level of each incoming or excitation LED light depending on the water load. That is, depending on the load of the sample we can vary the reference brightness level of the LED, in order to have a better characterization of the study parameter of the samples.

Therefore, the present invention is limited within the scope of devices or equipment for spectrophotometric and fluorimetric characterization on water samples, specifically, in the present invention, on wastewater samples.

BACKGROUND OF THE INVENTION

Spectrophotometry is one of the most used experimental techniques for the specific detection of molecules. It is characterized by its precision, sensitivity and its applicability to molecules of different nature (pollutants, biomolecules, etc.) and state of aggregation (solid, liquid, gas).

In the state of the art, the Utility Model ES1273069U is known, which discloses equipment for the characterization of the contaminant load present in wastewater that comprises a rotating wavelength selector disc that internally comprises a disc with a plurality of diodes, where this The disk comprises a self-positioning system, it also comprises a vertical displacement mechanism, and a tower that includes a vertical cavity, a circular channel where at one end it has an opening coinciding with a through hole of the circular support that houses a diode.

Fluorimetry is an analysis method used to measure the intensity of the radiation emitted by a material when excited mainly with ultraviolet light as well as with other light close to the ultraviolet region and including visible light.

There is equipment that allows spectrophotometry and fluorimetry to be carried out, however, to carry out fluorimetry they use a CCD matrix, which is a charge-coupled device, also known as CCD, which is an integrated circuit that contains a certain number of linked or coupled capacitors. Under the control of an internal circuit, each capacitor can transfer its electrical charge to one or more of the capacitors next to it on the printed circuit; in short, it is a sensor with tiny photoelectric cells that record the image.

The state-of-the-art fluorimetry embodiment requires more complex electronics and greater electrical consumption.

Furthermore, on the other hand, the spectrophotometry and fluorimetry equipment on the market is only valid for a certain type of specimen geometry, so its functionality is limited.

Therefore, it is the object of the present invention to overcome the noted drawbacks of performing fluorimetry with electronic means, where said equipment can also be used for a multiplicity of different geometries of specimens, developing equipment such as the one described below and is collected in its essentiality in the first claim.

DESCRIPTION OF THE INVENTION

The object of the present invention is included in its essentiality in the independent claim and the different embodiments are included in the dependent claims.

The purpose of the present invention is a device that analyzes both the spectral and fluorimetric response of the samples manually introduced into the device, being able to perform both analyzes simultaneously. The team works in the range of wavelengths known as ultraviolet and visible UV-Vis.

The equipment is controlled by a computer, through a USB connection.

The sample is introduced by the user through the upper part of the equipment, which is isolated from outside light by means of a black folding lid, in order to prevent it from causing alterations in the analysis.

The equipment object of the invention can adapt to samples of different path lengths automatically, as well as samples of different heights. The path length is understood as the distance that the light traverses through the sample to end up emitting a light intensity, direct or reflected, and which is recorded. That is, the path length is the width of the sample specimen to be crossed.

The equipment object of the invention comprises:

- A generator block for visible and infrared wavelengths.

- An independent UV light generation module using LED technology - A sample containment and spectrophotometric analysis block with the capacity to support cuvettes of different dimensions or path lengths. This block houses the electronics necessary to generate wavelengths belonging to the Ultraviolet, visible and infrared region of the spectrum.

- A fluorimetric analysis block.

Where the generating block of visible and infrared wavelengths comprises a rotating disk that contains multiple concentrically shaped LED diodes, which aligns with the sample with the combination of two movements: a first rotation movement and a second vertical displacement movement. The light generated by this block is transmitted by means of a fiber optic cable (sheathed with anti-reflective and with a black plastic covering) to the sample containment block, where, after passing through the sample, it hits a first sensor, which is in the same direction of the beam (180°).

With this configuration, the spectrophotometric analysis is carried out, which is carried out by aligning each LED with the fiber, which in turn is connected to the fluorimetric analysis block. Each LED emits multiple wavelengths simultaneously, and through mathematical models, we can "select" one of the specific wavelengths, which allows multiple wavelengths to be used with very few LEDs.

Given that the wavelengths belonging to the ultraviolet region of the spectrum, especially UVc, cannot be transported by optical fiber, the equipment object of the invention has an independent UV light generation module using LED technology, placed adjacently. at the optical fiber input, arranged in such a way that the UV LEDs surround the input fiber without hindering the passage of the visible and infrared wavelengths, thus being able to irradiate the samples under study with UV wavelengths, Visible and Infrared.

To carry out the fluorimetric analysis, the diffuse or emitted light must be captured at 90° to the main beam, so that the amount of light captured is that emitted by the sample itself and not by the main beam. This is done through a second sensor.

To do this, use is made of the fluorimetric analysis block, which consists of a rotating disk made up of a gear that houses various optical filters of different spectral width, which are aligned with the light beam at 90° and with the second sensor ( with adequate gain to adequately capture the reflected light).

Fluorimetry consists broadly in that when the sample is irradiated by a certain wavelength, due to its physicochemical properties, this sample emits light at other different wavelengths. This light emitted by the sample is measured at 90° to the main beam, to avoid detecting the wavelengths irradiated on the sample, that is, those coming from the wavelength generating block.

To carry out the analysis, a different process is carried out depending on whether the wavelengths to be irradiated belong to the ultraviolet region of the spectrum or, on the contrary, to the visible-infrared region.

In the first case, given that the UV LEDs are fixedly placed on a printed circuit, arranged concentrically to the fiber optic beam, so that they do not obstruct it, the equipment object of the invention turns on each one in isolation. one of the UV LEDs, so that they affect the sample under study. The visible and infrared wavelength block remains inactive during the time in which the UV block works in order not to interfere with the measurement.

In the case of visible and infrared wavelengths, for each LED of the visible and infrared wavelength generating block, each and every one of the different optical filters of the fluorimetric analysis block are aligned, in order to determine, not only the amount of light emitted by the sample, but also what wavelengths it is emitting. The latter is achieved through optical filters. If, for example, we irradiate the sample with a wavelength of 500nm, and the sample, due to its properties, emits wavelengths of 400 and 440nm, to know the emission intensity of each of them, we will align the 400nm filter, which will that the 440nm wavelengths do not reach the sensor, so that all the light received by it is a result of the 400nm wavelength. In this way, each optical filter is tested to know what wavelengths are emitted, which will be those that coincide with the passband of the filter, and of these wavelengths that manage to pass, we measure the amount of light that affects on the second sensor.

The visible and infrared wavelength generator block comprises an LED support plate that has a circular shape and where the LEDs are preferably arranged concentrically, said plate being housed in a wheel provided with a series of perforations where the LEDs are housed. LEDs and through which the light radiation emitted by each LED is allowed to pass once activated, the previous assembly being housed in a support piece.

This visible and infrared wavelength generator block has two movements, a first rotation movement of the wheel where the LEDs are housed, carried out by means of a first motor placed at the back of the wheel, and a second rotation movement. raising and lowering of the assembly carried out along cylindrical rods using a spindle driven by a gear train driven by a second motor.

The LED support plate is connected by means of a Slip Ring that allows the cables to rotate without tangling. This disc is connected to the main motor shaft and rotates freely by the action of axial bearings. To position the disk during the initialization process, the LED support disk (represented as the red disk) has positioning means, which in a possible non-limiting embodiment has two magnets on its back. of neodymium located at 90° to each other, where through a circuit board that analyzes the variations in the magnetic field produced by the magnets, precise positioning of the disc is possible

This visible and infrared wavelength generator block has the possibility of varying the intensity or brightness level of each incoming or excitation LED light depending on the water load. That is, depending on the sample load we can vary the reference brightness level of the LED. This is done based on the initial value of the recorded transmittance. Starting below a transmittance threshold, the brightness level is modified by increasing it, in order to have a better characterization of the study parameter of the samples. As for low levels, it is done the other way around.

The sample container block serves to contain both the sample under analysis and the spectrophotometry sensor or first sensor and the optical fiber through which the wavelengths of the visible and infrared region of the spectrum are transmitted, as well as the generation block. of UV wavelengths.

This block is integral with the fluorimetry block. The optical fiber is joined by means of couplings that start from the rear of the wavelength generating block and connect to the sample generating block.

This block allows the introduction of specimens with a variable excitation light passage width for which the light passage tunnel is adapted. This represents an important contribution since it allows correcting the possible hiding of molecules from the passage of light in cases where there is a high number of molecules.

In addition, the sample container block has a mechanism that allows it to be adjusted to samples with different path lengths or dimensions, which translates into the possibility of using analysis containers of different dimensions, which is crucial when carrying carry out their analysis.

This mechanism consists of two movements, one of horizontal displacement of the walls surrounding the sample, which allows the sensors and the fiber/UV wavelength generator to be moved closer or further away from it; and on the other hand, it has a vertical movement, which allows the height of the inserted test piece to be automatically adjusted, in order to adapt to different configurations.

The first movement is achieved by rotating a toothed wheel with a certain number of teeth, which has 4 curved grooves equidistant from each other inside, which serve as a guide for rectangular claws arranged in the shape of a crosshead, which They move thanks to a cylindrical stud that moves through the different grooves, converting the rotation movement of the toothed wheel into a horizontal displacement of the walls connected integrally to each of the claws. Each of the walls has the necessary elements to house the main sensor, as well as the UV wavelength generation block and the fiber optic socket, respectively.

This toothed wheel is driven by a servomotor through a train of straight and conical gears. The servomotor allows you to precisely control the degree of opening or closing of the sample container block, and therefore, the proximity of each wall (sensors/input wavelengths) to it.

Since the displacement of the walls that make up the sample containing block generates gaps during its displacement that would affect the light insulation conditions to which the sample must be subjected, the block has four black bands of a flexible material. , which connect each of the mobile faces to each other. These bands are deformed to adjust to the arrangement of the walls, thus allowing the sample to be isolated from possible light interference coming from inside the equipment.

On the other hand, to control the vertical displacement of the introduced sample, the equipment object of the invention has a second servomotor arranged adjacent to the mobile wall that houses the main sensor. By means of a tilting lever, activated by the servomotor through a set of articulated arms connected to each other, so that it transforms the rotation of the servomotor into a vertical displacement of the sample.

Finally, the fluorimetric analysis block comprises a toothed wheel provided with a series of preferably circular perforations in a circular arrangement, where in each of the perforations an optical filter is housed where each of these optical filters allows the passage of a very small portion of the spectrum, where these filters are secured by means of a circular cover, the toothed wheel where the filters are housed is driven by means of a pinion driven by a motor.

One of the holes in the disc does not have any filter, in order to be used for the equipment calibration process.

For the correct positioning of the disk during initialization, it has positioning means, which in a possible embodiment, the filter containing wheel has a system of magnets placed at 90° to each other, similar to that of the generator block. wavelengths described above, which are used together with a circuit board connected to the main wall, which detects changes in the magnetic field during the movement of the disk.

The set has a protective casing, which has two tabs at its base to connect it to the equipment and has an interchangeable support to house the second sensor used for fluorimetry.

The procedure for spectrophotometry and fluorimetry analysis with the equipment described above includes the steps of:

- Introduce a sample into the equipment from the top and close the lid to prevent external light from affecting the analysis.

- Control of the equipment through a PC application, and through a USB connection.

- Automatic adjustment of the sample container block according to its path length, as well as automatic regulation of the height of the introduced sample.

- Beginning of an initialization process that includes

o Initialization of the UV wavelength selection block. o Initialization of the visible and infrared wavelength selection block.

o Initialization of the fluorimetric analysis block

- Determination of the brightness level

o Brightness determination for spectrophotometric analysis

o Brightness determination for fluorimetric analysis

- Equipment calibration

- Beginning of the spectrophotometric and fluorimetric analysis process:

The sample container block Initialization process:

Given that the equipment object of the invention can analyze samples with different path lengths, it must adjust the block that houses the sample to its dimensions, so that both the main sensor and the wavelength input ( optical fiber for visible and infrared, and UV generation block), are as close as possible to the sample, without leaving gaps. The procedure is the next:

- A sample of a certain path length is introduced through the top of the equipment.

- Using a servomotor located in the lower part of the block, a toothed wheel is driven by a train of gears, the rotation of which causes the horizontal displacement of 3 mobile claws simultaneously, which have the three mobile walls of the assembly attached, where only Two of them have the main sensor and the wavelength input.

- The servomotor reduces the distance between the walls until the equipment object of the invention detects a change in the current demanded by the servomotor, which indicates that there is an opposition to its movement, and, therefore, that it has already been adjusted. to the dimensions of the sample. - At that moment, the walls of the sample are separated by a sufficient distance to allow its vertical displacement, but ensuring the maximum proximity of the components to it.

- Immediately afterwards, a second servomotor controls the vertical position of the sample in order to ensure the closure of the walls.

Where the initialization process of the visible and infrared wavelength selection block comprises:

- The wavelength generating block must first position the LED precisely with the optical fiber so that all the emitted light reaches the samples. To do this, start by rotating the disc clockwise until the neodymium magnets located at 90° to each other on the back of the disc align perfectly with the hall effect sensors also located at 90°, which are located on the body. main disk holder. - The way of positioning varies depending on what position the disc is in before starting its movement:

1. If none of the hall effect sensors detect any of the magnets, the disc rotates clockwise until both hall effect sensors detect their respective magnet, which is the one placed in the same position. After that, an automatic adjustment is made to detect the position in which the magnetic field detected by each of the sensors is strongest.

2. If one or both magnets are detected, the disc rotates clockwise until neither sensor detects. After that, rotate a little more and then repeat the process described in the previous step.

- Once the disc has been adjusted radially, it is positioned vertically. To do this, the block is lowered by means of the gear train located at the base, until one of the limit switches is activated, which indicates that the block has completely descended. After this, the block rises a certain distance to be perfectly aligned with the optical fiber.

- The UV wavelength generation block does not require any initialization process as it does not have moving parts.

The Initialization process of the fluorimetric analysis block includes:

The disc rotates on its axis by the action of the pinion until the first hole of the disc (which does not have an optical filter) is aligned with the sample. To do this, like the wavelength generating block, it has two neodymium magnets located at 90° to each other, located on the back, which are detected using the same process described in the previous step.

The brightness determination process for spectrophotometric analysis includes the following stages:

b. A sample of distilled water is introduced into the equipment

c. Since the test tube can have different dimensions, the team secures it using a system of retractable clamps that adjust to the geometry of the test tube, ensuring that it remains aligned with the light beam, as well as with the filters and sensors.

d. Once the sample is aligned and secured, another mechanism adjusts the height of the cuvette so that it does not protrude from the equipment at the top and that the analysis is carried out in the center of it (half of its height).

and. It connects to the computer via USB and connects to the electrical network through a voltage transformer.

F. Once the equipment has been initialized (positioning all its elements in the correct positions), the first LED is turned on at its minimum brightness capacity.

g. The main sensor (Sensor 1) waits 1.5 seconds before starting to measure, so that the amount of light that manages to pass through the sample is homogeneous.

h. The sensor measures the amount of light that manages to pass through the distilled water sample, at different time intervals, in order to make an average.

Yo. If the average value of light that manages to pass through the sample is greater than a certain predefined threshold, the brightness level is considered adequate and is displayed on the screen.

j. Otherwise, the brightness is increased by a certain amount and the process is repeated.

k. The threshold is an empirical value, which should be high enough to take advantage of the sensor resolution, but not too high to avoid saturation.

l. Once the LED brightness determination is complete, the assembly is rotated and moved vertically to align the next LED, and the process is repeated until all of them are completed.

m. To determine the brightness of the UV LEDs present in the UV wavelength generation block, each LED is turned on in isolation, and steps f-i described above are repeated.

The process for determining gloss for fluorimetric analysis includes:

to. A sample of distilled water is introduced into the equipment.

b. Since the test tube can have different dimensions, the team secures it using a system of retractable clamps that adjust to the geometry of the test tube, ensuring that it remains aligned with the light beam as well as with the filters and sensors.

c. Once the sample is aligned and secured, another mechanism adjusts the height of the cuvette so that it does not protrude from the equipment at the top and that the analysis is carried out in the center of it (half of its height).

d. It connects to the computer via USB and connects to the electrical network through a voltage transformer.

and. With the fluorimetry block positioned so that the unfiltered hole is aligned with the sample at 90° to the main beam, the process begins.

F. Once the equipment has been initialized (positioning all its elements in the correct positions), the first LED is turned on at its minimum brightness capacity.

g. The secondary sensor (Sensor 2) waits a few seconds before taking data.

h. If the selected captured light value is greater than a certain predefined threshold, the brightness level is considered adequate and is displayed on the screen.

Yo. Otherwise, the LED brightness is increased by a certain amount and the above process is repeated.

j. Once the LED brightness determination is complete, the block 1 assembly is rotated and moved vertically to align the next LED, and the process is repeated until all LEDs are completed. k. To determine the brightness of the UV LEDs present in the UV wavelength generation block, each LED is turned on in isolation, and steps g-i described above are repeated.

Furthermore, depending on the responses received from the analyzed samples, it is possible to vary the intensity or brightness level of each incoming or excitation LED light depending on the water load. That is, depending on the sample load we can vary the reference brightness level of the LED. This is done based on the initial value of the recorded transmittance. Starting below a transmittance threshold, the brightness level is modified by increasing it, in order to have a better characterization of the study parameter of the samples. As for low levels, it is done the other way around.

The equipment calibration process includes the stages of:

This process is carried out once the optimal brightness level has been determined for both spectrophotometry and fluorimetry.

to. The sample is introduced into a cuvette through the top of the equipment.

b. Since the test tube can have different dimensions, the team secures it using a system of retractable clamps that adjust to the geometry of the test tube, ensuring that it remains aligned with the light beam as well as with the filters and sensors.

c. Once the sample is aligned and secured, another mechanism adjusts the height of the cuvette so that it does not protrude from the equipment at the top and that the analysis is carried out in the center of it (half of its height).

d. Once the equipment is initialized, the first LED is turned on at the selected brightness level, whose light travels through the optical fiber until it reaches the sample under study.

and. Depending on its physicochemical properties, more or less light will pass through, which will be captured by a sensor located in the same direction of the beam (180°).

F. The main sensor (Sensor 1) waits 1.5 seconds before starting to measure, so that the amount of light that manages to pass through the sample is homogeneous.

g. The sensor measures the amount of light that manages to pass through the distilled water sample, at different time intervals, in order to make an average.

h. This average value (/0, spectrophotometry) is stored in the internal memory of the equipment (EEPROM) in a specific position determined by the LED used.

Yo. Next, the value of diffuse light that sensor 2 manages to capture is stored, without the use of an optical filter:

- The secondary sensor (Sensor 2) waits a few seconds before taking data.

- The LED turns on at maximum brightness.

- The captured value (/0fluorimetry) is stored in another position of the EEPROM for that LED (different from the other value of I0 registered previously)

j. After the previous step, the next LED is aligned and the process is repeated.

k. Once the analysis of the visible and infrared wavelengths arranged on the rotating disk is completed, the UV wavelengths are calibrated, by turning them on isolatedly, and following the steps f-i described above.

Both the calibration and analysis processes carry out identical steps, with the difference that during the calibration a sample of distilled water is used and the amount of light detected by the sensor that manages to pass through the sample at each wavelength (/ 0), is stored in the internal memory of the device.

The spectrophotometric and fluorimetric analysis process includes the stages of:

The equipment can carry out only the spectrophotometric analysis or perform it in conjunction with the fluorimetric analysis. Next, we are going to describe this last process, given that it implies the other:

to. The sample is introduced into a cuvette through the top of the equipment. b. Since the test tube can have different dimensions, the team secures it using a system of retractable clamps that adjust to the geometry of the test tube, ensuring that it remains aligned with the light beam as well as with the filters and sensors.

c. Once the sample is aligned and secured, another mechanism adjusts the height of the cuvette so that it does not protrude from the equipment at the top and that the analysis is carried out in the center of it (half of its height).

d. Once the equipment is initialized, the first LED is turned on at the selected brightness level, whose light travels through the optical fiber until it reaches the sample under study.

and. Depending on its physicochemical properties, more or less light will pass through, which will be captured by a sensor located in the same direction of the beam (180°).

F. The main sensor (Sensor 1) waits 1.5 seconds before starting to measure, so that the amount of light that manages to pass through the sample is homogeneous.

g. The sensor measures the amount of light that manages to pass through the sample under analysis, at different time intervals, in order to make an average.

h. This average value represents in the expression of TransmittanceT =—io, the value of 7”, which is divided by the value ofI0 that the equipment stored for that LED in the calibration process, thus calculating giving the value of “gross transmittance”. .

Each LED emits multiple wavelengths simultaneously following a normal distribution of brightness intensities. The gross transmittance value corresponds to the value recorded by the effect of all those wavelengths that simultaneously pass through the samples. This means that the values detected by the equipment, even though it has the same shape as those recorded by commercial equipment based on incandescent lamps, are higher.

Yo. To eliminate this effect of multiple wavelengths and be able to isolate the effect of a single one of them without the need for optical elements, a series of mathematical models preloaded in the equipment are used, which allow modeling, based on the gross transmittance value, the transmittance value that would be obtained if only one of the wavelengths among the multiple emitted ones were activated. This not only allows us to solve the problem of deviation of the results, but also allows us to extend the number of supported wavelengths.

j. For each LED, the transmittance values produced by each of the wavelengths emitted by each LED are displayed simultaneously on the screen, which speeds up the analysis.

k. Once the spectral response has been analyzed for the region of wavelengths emitted by the current LED, the fluorimetric analysis is carried out.

l. To do this, the first optical filter is aligned with the sample, and the LED is turned on in progress at short time intervals, at the brightness level selected in step 4.

m. The secondary sensor (Sensor 2) waits a few seconds before taking data.

n. The amount of light captured by sensor 2 is recorded.

either. This process is repeated several times to take an average value.

p. That selected reflected light value represents the value of I in the relationship T= —, which is divided by the value of /0 (the one recorded during the period

fluorescence calibration, not spectrophotometry), stored for that LED during the calibration phase. This allows obtaining a gross transmittance value.

q. By means of various mathematical models, the transmittance values recorded from the diffuse light (light measured at 90°) produced by each of the lengths are calculated from the transmittance value measured by each LED (gross transmittance). of waves emitted by the LED. r. Each of these values are sent to the computer to be graphed, indicating the measured transmittance value, excitation wavelength and wavelength emitted by the sample. In this way, the heat map that makes up the fluorimetric response is generated. s. This process is repeated with each and every optical filter installed.

t. When this process ends, block 3 is initialized again, to leave it ready for the next fluorimetric analysis with the next LED.

or. The next LED is aligned with the sample and the process described is repeated until each and every one of them is completed.

v. Once the analysis of the visible and infrared wavelengths arranged on the rotating disk is completed, the UV wavelengths are calibrated, by turning them on isolatedly, and following the steps f-t described above.

w. Once the spectrophotometric and fluorimetric analysis is completed, the equipment repositions block 1 and 3, being prepared for the next analysis.

x. At that moment, the PC application used to control the equipment notifies the user that the analysis has been completed and indicates whether they want to send the results to the cloud, repeat the analysis or discard it.

Unless otherwise indicated, all technical and scientific elements used herein have the meaning normally understood by a person skilled in the art to which this invention belongs. Procedures and materials similar or equivalent to those described herein may be used in the practice of the present invention.

Throughout the description and claims the word "comprises" and its variants are not intended to exclude other technical characteristics, additives, components or steps. For those skilled in the art, other objects, advantages and characteristics of the invention will emerge in part of the description and part of the practice of the invention.

EXPLANATION OF THE FIGURES

To complement the description that is being made and in order to help a better understanding of the characteristics of the invention, in accordance with a preferred example of its practical implementation, a set of drawings is attached as an integral part of said description. where, for illustrative and non-limiting purposes, the following has been represented.

In Figure 1, we can see a perspective representation of the equipment that is the object of the invention.

In figure 2, we can see a first perspective representation of the interior of the equipment.

Figure 3 shows a second perspective representation of the interior of the equipment.

Figure 4 shows the wavelength generating block in an exploded manner.

Figure 5 shows a view that allows us to appreciate the means used to raise and lower the wavelength generating block.

Figure 6 shows a front view of the interior of the wavelength generating block.

Figure 7 shows the interior of the fluorimetric analysis block.

Figure 8 shows the rear face of the filter container wheel.

Figure 9 shows schematically where the light emitted by the LEDs circulates, and the relative positioning of the sensors to carry out the spectrophotometric and fluorimetric analysis.

Figure 10 shows a detailed view of the sample container block adjustable to different path lengths, as well as the UV wavelength generation block.

Figure 11 shows an exploded view of the sample container block adjustable to different path lengths, as well as the UV wavelength generation block, where all its elements can be seen.

Figure 12 shows a detailed view of the mechanism for varying the path length of the sample containing block.

Figure 13 shows a detailed view of the UV wavelength generation block as well as its arrangement with respect to the fiber optic input for visible and infrared wavelengths.

PREFERRED EMBODIMENT OF THE INVENTION

In view of the figures, a preferred embodiment of the proposed invention is described below.

In Figure 1 we can see that the equipment comprises a casing (1) under which a base (3) is placed resting on a piece of floor (4), leaving a cover (2) on the upper part of the casing (1). ).

In Figure 2 it can be seen that inside the equipment there is a platform (5) on which a wavelength generator/selector block (6) is fixed together with a sample containment block with variable path length (7 ) and spectrophotometric analysis by means of an optical fiber (9), where the sample containment block with variable path length (7) is integrally linked with a fluorimetric analysis block (8).

In Figures 3 and 4 it can be seen that the wavelength generating block (6) comprises a wheel (37) provided with a series of perforations (38) where some LEDs (35) are housed mounted on a support plate ( 11) of the LEDs (35), where said wheel (37) is mounted on a wheel support (12). A first motor (10) for rotating the wheel (37) is fixed to this wheel support (12).

This visible and infrared wavelength generator block (6) has a lifting and lowering movement along vertical guides (14), with an anti-twisting support for cables (31) provided with a rolling ring or "slip ring" (19); with a second motor (15) for lifting and lowering the block, and which transmits said drive through a gear train (16) (figure 5) where the last gear (17) is attached a spindle (13) that threads on the anti-twist cable support (31) and that transforms a circular movement into a lifting and lowering movement. In addition, it has some stops or limit switches (18).

In Figure 6 you can see an embodiment of the means for the correct alignment of the wheel (37), which consists of the arrangement of magnets (34), preferably at 90°, and a support plate for a hall effect sensor. (33) that analyzes the variations in the magnetic field produced by the magnets, precise positioning of the disc is possible

The fluorimetric analysis block (8) comprises, as can be seen in Figures 3 and 7, a series of filters (29) housed within a filter container (25) and covered by a lid (26), housing the previous assembly inside a housing formed by a support piece (21) and a cover (22) (figure 3).

The filter container (25) takes the shape of a toothed wheel and meshes with a pinion (24) driven by a third motor (23). In addition, the fluorimetric analysis block (8) has a fluorescence sensor support (27) on which a photodiode (28) is mounted.

Figure 8 shows the means used for the correct positioning of the filter container (25), which, like the wavelength generating block, has magnets (36) and a support plate for a hall effect sensor. (30) that analyzes the variations in the magnetic field produced by the magnets, precise positioning of the disc is possible.

The sample containment block with adjustable path length (7) comprises means for housing a first sensor (20) responsible for spectrophotometry measurement, as well as means for regulating the height (32) of the inserted specimens. , and means necessary to vary the arrangement of the walls of the container to bring them closer or further away from the sample depending on its dimensions, and therefore, adjusting the position of the first sensor (20), as well as a wavelength generation block UV (41) and fiber optic input. Said means of variation of the walls allow the container with variable path length (7) to allow the container to adjust to the geometry of the specimen, ensuring that it remains aligned with the light beam, as well as with the filters and the sensors without leaving gaps.

Figure 9 shows in detail the flow of the light generated by the wavelength generation block (6) that is conducted by the optical fiber (9) to the sample containment block with variable path length. (7) where the first sensor is arranged aligned 180° with the light emitted and passing through the sample, while the light emitted perpendicularly as the beam passes through the optical fiber is received by the fluorimetry analysis block (8) where it is houses the second sensor.

In this figure 9 it is observed that the optical fiber (9) is connected to the visible and infrared wavelength generating block (6) through a fiber support (39), and with the sample containment block (7) through a coupling piece (40) of the optical fiber. Between the optical fiber (9) and the sample containment block with variable path length (7), the UV wavelength generation block (41) is located.

As seen in Figure 10 and Figure 11, the sample container with variable path length (7) comprises four walls (42, 43, 44 and 45) of which the walls (42, 43 and 44) They are mobile walls that move simultaneously and are separated from each other by means of elastic elements (46) which stretch and/or contract during the movement of the walls, isolating a sample housed in the receptacle (47) from any unwanted light source

Furthermore, the sample container with variable path length (7) has means to modify the positioning of the height (32) of the sample introduced automatically and thus provide its path length.

The UV wavelength generation block (41) is arranged on the wall (44), while the first sensor (20) is arranged on the facing wall (42).

The remaining wall (45), which is a fixed wall, is used to hold the assembly with the filter container (25).

To carry out the horizontal movement of the mobile walls (42, 43 and 44), the block has a first toothed wheel (48) provided with curved grooves (49) through which lugs (50) linked with claws move. rectangular (51) to which the mobile walls (42, 43 and 44) are integrally connected.

The first toothed wheel (48) is driven by a servomotor (52), whose movement is transmitted by a train of straight and conical gears (53) and (54) respectively, which allows controlling the degree of opening or closing of the assembly.

The means for regulating the height (32) of the sample introduced into the cavity (47) comprise a system of articulated arms (55) and (56) actuated through a pivoting lever (57) by a second servomotor (58).

The UV wavelength generation block (41) which, according to Figure 13, consists of a printed circuit board (59) that has a hole (60) through which the optical fiber (9) runs without obstacle, which is surrounded by UV LEDs (61) arranged radially, so that the sample under study can be irradiated by both the wavelengths generated by the visible and infrared wavelength generation block (6) that enter the sample containment block with variable path length (7) through the optical fiber (9), as well as by the UV wavelength generation block (41) directly without the need to travel by fiber optics.

Having sufficiently described the nature of the present invention, as well as the way of putting it into practice, it is stated that, within its essentiality, it may be put into practice in other embodiments that differ in detail from that indicated by way of example. , and to which the protection sought will also reach, as long as it does not alter, change or modify its fundamental principle.

Claims (11)

1.- Equipment for performing spectrophotometry and fluorimetry characterized because it includes: - A visible and infrared wavelength generator block (6) - A UV wavelength generation block (41) - A sample containment block with variable path length (7), and analysis. - A fluorimetric analysis block (8). Where the visible and infrared wavelength generating block (6) is linked to the UV wavelength generation block (41) by means of an optical fiber (9), and the UV wavelength generation block ( 41) is mounted on the sample containment block with variable path length (7) and this is integrally connected to a fluorimetric analysis block (8), where the visible and infrared wavelength generator block (6) It comprises a rotating disk (37) containing multiple LED diodes (35) arranged concentrically, which aligns with the sample with the combination of two movements: a first rotation movement by means of a first motor (10) and a second rotation movement. vertical displacement by means of a second motor (15), it also has means for the correct positioning of the rotating disk (37), where the sample containment block with variable path length (7) comprises means to house a first sensor ( 20) responsible for spectrophotometry measurement, where the sample containment block with variable path length (7) comprises - means for housing a first sensor (20) responsible for spectrophotometry measurement - mobile walls (42, 43 and 44) that can be moved simultaneously - means for regulating the height (32) of the sample where the fluorimetric analysis block (8) comprises a series of filters (29) housed within a filter container (25) that can rotate by the action of a third motor (23), it also has means for correct positioning of the filter container (25) and with means for supporting a second sensor. 2. - Equipment for carrying out spectrophotometry and fluorimetry according to claim 1 characterized in that the generating block of visible and infrared wavelengths (6) for carrying out the vertical movement of elevation and descent comprises vertical guides (14), a second motor (15) for lifting and lowering the block, and which transmits said drive by means of a gear train (16) where a spindle (13) is attached to the last gear (17) that threads on an anti-twist cable support. (31) and that transforms a circular movement into a lifting and lowering movement, in addition, it has some stops or limit switches (18). 3. - Equipment for performing spectrophotometry and fluorimetry according to claim 1 or 2, characterized in that the wavelength generating block (6) has an anti-twist support for cables (31) provided with a rolling ring or "slip ring" ( 19) 4. - Equipment for performing spectrophotometry and fluorimetry according to claim 1 or 2 or 3 characterized in that the means for correct alignment of the wheel (37) consists of the arrangement of magnets (34), and a support plate of a hall effect sensor (33) that analyzes the variations in the magnetic field produced by the magnets. 5. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that the means for regulating the height (32) of the introduced test tubes consists of a system of retractable clamps. 6. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that the filter container (25) is covered by a lid (26), the previous assembly being housed within a housing formed by a support piece ( 21) and a cover (22). 7. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that the means used for the correct positioning of the filter container (25) are magnets (36) and a support plate for a hall effect sensor. (30) which analyzes the variations in the magnetic field produced by magnets. 8. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that the mobile walls (42, 43 and 44) are movable simultaneously thanks to a first toothed wheel (48) provided with curved grooves (49). through which lugs (50) move, linked to rectangular claws (51) to which the mobile walls (42, 43 and 44) are connected integrally, where the first toothed wheel (48) is driven by a servomotor (52), 9. - Equipment for carrying out spectrophotometry and fluorimetry according to claim 8 characterized in that the drive of the servomotor (52) is transmitted through a train of straight and conical gears (53) and (54) respectively, which allows controlling the degree of opening. or closure of the set. 10. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that to regulate the height (32) of the sample introduced into the cavity (47) the equipment comprises a system of articulated arms (55) and (56) actuated through a pivoting lever (57) by a second servomotor (58) 11. - Equipment for carrying out spectrophotometry and fluorimetry according to any of the previous claims, characterized in that the UV wavelength generation block (41) consists of a printed circuit board (59) that has a hole (60) through which that the optical fiber (9) runs without obstacle, which is surrounded by UV LEDs (61) arranged radially, so that the sample under study can be irradiated both by the wavelengths generated by the generation block of visible and infrared wavelengths (6) that enter the sample containment block with variable path length (7) through the optical fiber (9), as well as by the UV wavelength generation block ( 41) directly without having to travel through optical fiber. to. The main sensor (Sensor 1) waits 1.5 seconds before starting to measure, so that the amount of light that manages to pass through the sample is homogeneous. b. The sensor measures the amount of light that manages to pass through the sample under analysis, at different time intervals, in order to make an average. c. This average value represents in the Transmittance expression T =—,io the value of 7”, which is divided by the value of I0 that the equipment stored for that LED in the calibration process, thus calculating giving the gross transmittance value. Each LED emits multiple wavelengths simultaneously following a normal distribution of brightness intensities. The gross transmittance value corresponds to the value recorded by the effect of all those wavelengths that simultaneously pass through the samples. This means that the values detected by the equipment, even though it has the same shape as those recorded by commercial equipment based on incandescent lamps, are higher. d. To eliminate this effect of multiple wavelengths and be able to isolate the effect of a single one of them without the need for optical elements, a series of mathematical models preloaded in the equipment are used, which allow modeling, based on the gross transmittance value, the transmittance value that would be obtained if only one of the wavelengths among the multiple emitted ones were activated. This not only allows us to solve the problem of deviation of the results, but also allows us to extend the number of supported wavelengths. and. For each LED, the transmittance values produced by each of the wavelengths emitted by each LED are displayed simultaneously on the screen, which speeds up the analysis. F. Once the spectral response has been analyzed for the region of wavelengths emitted by the current LED, the fluorimetric analysis is carried out. g. To do this, the first optical filter is aligned with the sample, and the LED is turned on in progress at short time intervals, at the brightness level selected in step 4. h. The secondary sensor (Sensor 2) waits a few seconds before taking data. Yo. The amount of light captured by sensor 2 is recorded. j. This process is repeated several times to take an average value. k. That selected reflected light value represents the value of I in the relationship T = —io, which is divided by the value of I0 (the one recorded during the fluorescence calibration, not spectrophotometry), stored for that LED during the calibration phase. This allows obtaining a gross transmittance value. l. By means of various mathematical models, the transmittance values recorded from the diffuse light (light measured at 90°) produced by each of the lengths are calculated from the transmittance value measured by each LED (gross transmittance). of waves emitted by the LED. m. Each of these values are sent to the computer to be graphed, indicating the measured transmittance value, excitation wavelength and wavelength emitted by the sample. In this way, the heat map that makes up the fluorimetric response is generated. n. This process is repeated with each and every optical filter installed. either. When this process ends, block 3 is initialized again, to leave it ready for the next fluorimetric analysis with the next LED. p. The next LED is aligned with the sample and the process described is repeated until each and every one of them is completed. q. Once the analysis of the visible and infrared wavelengths arranged on the rotating disk is completed, the UV wavelengths are calibrated, by turning them on isolatedly, and following the steps f-t described above. r. Once the spectrophotometric and fluorimetric analysis is completed, the equipment repositions block 1 and 3, being prepared for the next analysis. s. At that moment, the PC application used to control the equipment notifies the user that the analysis has been completed and indicates whether they want to send the results to the cloud, repeat the analysis or discard it.