Slope spectroscopic measurement without a reference signal

Abstract

The method comprises the steps of determining whether fluctuations in the intensity of the probe radiation meet a stability criterion; when the probe is disposed at a first position, directing the probe radiation to pass through the probe and defining a first path length L1 of the probe radiation through the fluid sample; when the probe is disposed at the first position, measuring a transmission intensity I1 of the probe radiation after passing through the fluid sample; when the probe is disposed at a second position, directing the probe radiation to pass through the probe and defining a second path length L2 of the probe radiation through the fluid sample; when the probe is disposed at the second position, measuring a transmission intensity I2 of the probe radiation after passing through the fluid sample; and, when the stability criterion is met, determining a concentration C of a material in the fluid sample based on L1, I1, L2, and I2.

Description

Technical Field

[0001] [Cross - Reference to Related Applications] This application claims the benefit of priority under 35 U.S.C. § 119 to U.S. Provisional Application No. 63 / 343,357, filed on May 18, 2022, entitled "NO - REF - SIGNAL SLOPE SPECTROSCOPIC MEASUREMENT", the entire disclosure of which is incorporated herein by reference for all purposes.

[0002] [Technical Field] Embodiments of the present disclosure generally relate to spectroscopic analysis, and more specifically, to solution analysis using a light source coupled to a variable path - length measurement system.

Background Art

[0003] Absorption spectroscopy is used to measure the composition and / or properties of materials in any phase, gas, liquid, or solid. For example, the light absorption spectrum of a liquid substance can be measured to determine the concentration or other properties of the species of interest in the liquid medium. The absorption spectrum can provide a distribution of light attenuation (due to absorbance) as a function of the wavelength of light. In a known spectrophotometer, the sample substance under study is positioned within a transparent container, whereby electromagnetic radiation (light) of a known wavelength λ (i.e., ultraviolet, infrared, visible light, etc.) and intensity I can be measured using a suitable detector after passing through the transparent container.

[0004] In known ultraviolet (UV) / visible spectrophotometers, an instrument such as a standard cuvette is utilized, which may have a path length of standard cm, and the incident light is conducted through the liquid containing the substance to be measured over that path length. For a sample consisting of a single homogeneous substance with a concentration c, the light transmitted through the sample will follow the relationship known as Beer's law: A = εCL, where A is the absorbance (also known as the optical density (OD) of the sample at wavelength λ, where OD = -log of the ratio of the transmitted light to the incident light), ε is the absorption coefficient or extinction coefficient (usually constant at a given wavelength), C is the concentration of the sample, and L is the path length of the light through the sample. Thus, in principle, information regarding the concentration of a homogeneous substance can be determined based on the recorded light intensity of the signal passing through the sample's instrument. However, under some circumstances, determination of the concentration in such an apparatus can be difficult. In many cases, the compound of interest in the solution is highly concentrated. For example, certain biological samples such as proteins, DNA, or RNA are often isolated at concentrations that fall outside the linear range of the spectrophotometer when absorbance is measured. Therefore, in many cases, dilution of the sample is required to measure absorbance values that fall within the linear range of the instrument. This results in the need for high-frequency multiple dilutions of the sample, which in turn brings about both dilution errors and the removal of the diluted sample for any downstream applications. Therefore, it is useful to take existing samples without knowing the possible concentrations and measure the absorbance of these samples without dilution. One consequential characteristic common to these known UV / visible spectrophotometers is the ability to accurately determine the path length L, thereby enabling accurate concentration measurements.

[0005] To address these issues, recently, techniques based on variable path length spectrophotometers have been developed. This type of spectroscopic system can generally employ a known light source such as a light source based on an ultraviolet / visible spectrophotometer. The light from the ultraviolet / visible spectrophotometer is then directed to a special probe within an analytical instrument configured to dynamically vary the path length L within a special sample chamber during absorbance measurement. Thus, the intensity of the transmitted radiation generated from the light source of the ultraviolet / visible spectrophotometer is detected after passing through the sample chamber, while the movement of the probe varies the path length L through a plurality of different positions. Therefore, a series of measurements are made that generate different values of A for each different value of l in a manner that does not require knowledge of any particular path length L to determine the concentration C.

[0006] Such variable path length spectroscopy can be carried out through a production system and can be adapted for in-line measurement of samples. However, for example, the equipment required for such measurement scenarios can require significant installation effort and an excessive amount of space. For example, an ultraviolet / visible spectrophotometer system used as a light source can occupy several cubic feet of space and can have a weight of approximately several tens of kilograms. Further, determination of A generally requires multiple measurements of intensity for each sample measurement value taken at a given path length L.

[0007] In view of these and other considerations, the present disclosure is provided.

SUMMARY OF THE INVENTION

[0008] In one embodiment, a method for determining the concentration of a material can comprise determining whether fluctuations in the intensity of probe radiation emitted by a light source of an absorbance spectroscopy system meet a stability criterion. The method defines a first path length L of the probe radiation through a fluid sample when the probe is disposed at a first position and directs the probe radiation to pass through the probe, and the transmitted intensity I of the probe radiation after passing through the fluid sample when the probe is disposed at the first position 1 and defines a first path length L of the probe radiation through a fluid sample when the probe is disposed at a first position and directs the probe radiation to pass through the probe, and the transmitted intensity I of the probe radiation after passing through the fluid sample when the probe is disposed at the first position 1It may further include the step of measuring. When the probe is disposed at the second position, the method directs probe radiation to pass through the probe and defines a second path length L of the probe radiation through the fluid sample, and measures the transmission intensity I of the probe radiation after passing through the fluid sample when the probe is disposed at the second position. When the stability criterion is satisfied, the method may further include the step of determining the concentration C of the material in the fluid sample based on L, I, L, and I. 2 It may also include the step of defining, and measuring the transmission intensity I of the probe radiation after passing through the fluid sample when the probe is disposed at the second position. 2 It may also include the step of measuring. When the stability criterion is satisfied, based on L 1 , I 1 , L 2 , and I 2 It may additionally include the step of determining the concentration C of the material in the fluid sample.

[0009] In another embodiment, it is determined whether the fluctuation of the intensity of the probe radiation emitted by the light source of the absorbance spectroscopy system satisfies the stability criterion; when the probe is disposed at the first position, the light source is directed to direct the probe radiation to pass through the probe to define a first path length L of the probe radiation through the fluid sample; receive the transmission intensity I of the probe radiation after passing through the fluid sample; when the probe is disposed at the second position, the light source is directed to direct the probe radiation to pass through the probe to define a second path length L of the probe radiation through the fluid sample; receive the transmission intensity I of the probe radiation after passing through the fluid sample; and when the intensity fluctuation satisfies the stability criterion, a non-transitory computer-readable storage medium is provided that stores computer-readable program code executable by a processor to determine the concentration C of the material in the fluid sample based on L, I, L, and I. 1 It may also include the step of defining, and receiving the transmission intensity I of the probe radiation after passing through the fluid sample. 1 When the probe is disposed at the second position, the light source is directed to direct the probe radiation to pass through the probe to define a second path length L of the probe radiation through the fluid sample. 2 It may also include the step of receiving the transmission intensity I of the probe radiation after passing through the fluid sample. 2 And when the intensity fluctuation satisfies the stability criterion, based on L 1 , I 1 , L 2 , and I 2 It provides a non-transitory computer-readable storage medium that stores computer-readable program code executable by a processor to determine the concentration C of the material in the fluid sample.

[0010] In a further embodiment, a measuring device is provided that includes a light source for generating a probe signal; and a measuring instrument for receiving the probe signal. The measuring instrument includes a sample container for containing a fluid sample, the sample container including a container wall, and a probe configured to direct the probe signal through the sample container, where the probe is movable along the probe direction relative to the container wall so as to vary the path length L of the probe signal through the fluid sample. The measuring device may also include a detector arranged to receive the probe signal after it has passed through the container wall; and a control system. The control system is configured to determine whether fluctuations in the intensity of the probe radiation emitted by the light source meet a stability criterion; and, if the fluctuations in intensity meet the stability criterion, to calculate the concentration C of a material in the fluid sample based on changes in the intensity of the probe signal measured as a function of the change in the path length L.

Brief Description of the Drawings

[0011] The accompanying drawings illustrate preferred embodiments of the disclosed method, devised heretofore for practical application of its principles.

[0012]

Figure 1A

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Figure 1B

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Figure 1C

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Figure 2

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Figure 3

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Figure 4

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Figure 6

MODE FOR CARRYING OUT THE INVENTION

[0020] According to embodiments of the present disclosure, techniques and apparatuses are provided for improving absorbance measurements based on a variable-pathlength-measurement (VPT) apparatus architecture. These embodiments provide a rational and dynamic approach, in particular, for determining the concentration of a material in a fluid sample. The approach of these embodiments employs a plurality of intensity measurements recorded as radiation passes through the fluid sample while varying the pathlength of the radiation through the fluid sample. As detailed below, and in contrast to known absorbance spectroscopy techniques, these embodiments determine the change in absorbance of the fluid sample, and thus the concentration C of the material in the fluid sample, without the need to perform a reference signal measurement.

[0021] FIG. 1A shows an absorption spectroscopic apparatus shown as system 100 according to an embodiment of the present disclosure. System 100 may include a small light source 102, a measuring device 110 coupled to the small light source 102, and a detector 112 disposed adjacent to the measuring device 110. The small light source 102 may have a light emitting diode (LED) that generates radiation 104 at a target wavelength in the range from UV to IR, and specifically, in the range of 190 nm to 1100 nm, or 250 nm to 1000 nm, etc., according to various non-limiting embodiments. In some examples, the small light source 102 may represent a single LED or an array of LEDs that emits radiation at a single wavelength. In other embodiments, a plurality of LEDs may be provided, where a given LED emits radiation at a wavelength different from that of another LED.

[0022] The measuring device 110 is configured to contain a fluid sample containing the material of the substance to be measured, and the details of the deformation of the measuring device 110 will be discussed below. The detector 112 is configured to detect the intensity I of the radiation that has passed through a given fluid sample contained in the measuring device 110, and the radiation is shown as attenuated radiation 111. According to the Beer Lambert law shown in Equation 1 below, the concentration C of the material in the sample can be determined as A / eL, where A is the absorbance and e is the molar extinction coefficient.

Equation

[0023] Next, A is determined as log 10 (I 0 / I), where I 0 is the intensity of the radiation 104 and I is the intensity of the attenuated radiation 111. To measure the value of I 0 , the system 100 further includes a reference detector 106 for receiving a portion of the radiation 104 before the radiation 104 is conducted through the measuring device 110. This parameter is used to directly calculate the absorbance according to Equation 2, which is the absorbance calculation formula.

Number

[0024] Therefore, in a given measurement instance, detector 112 measures I based on the attenuated radiation 111, while on the other hand, when reference detector 106 measures I 0 is measured, the absorbance A is determined. According to the approach of slope spectrometry, Lambert-Beer's law can be rewritten as A / L = εC and can be further extended to dA / dL = εC, where the entity dA / dL is regarded as the slope parameter m. During operation, system 100 operates according to the principle of slope spectrometry to determine the change in absorbance A as a function of the change in path length L, varying the path length L through which the radiation 104 travels, and thus, when knowledge of ε for a given substance is provided, directly determining the value of C for that substance.

[0025] Details of the modified operation of measurement device 110 are discussed below with respect to FIGS. 2 and 3. Briefly, however, measurement device 110 employs a movable optical probe (as shown in FIG. 2 and discussed in more detail below) to vary the path length L of the radiation 104 passing through a given fluid sample present within measurement device 110 (not shown in FIG. 1A, but refer to fluid sample 211 in FIG. 2). Since the intensity of the attenuated radiation 111 varies in accordance with the change in path length L, the change in absorbance A as a function of the change in path length L can be directly determined using the change in I as a function of the change in path length L. Thus, using the knowledge of I 0 provided by reference detector 106, system 100 is employed to vary the path length of radiation 104 as it passes through measurement device 110 (to determine ΔL) and detect the change in the intensity of the attenuated radiation 111 (to determine ΔA), thereby easily determining the concentration C of the material within the fluid sample.

[0026] FIG. 1C provides a configuration 180 showing details of a geometric arrangement for determining the concentration of a substance according to the principle of slope spectrometry. In configuration 180, a light source such as a small light source 102 directs radiation 104 to pass through a sample 182 such as a fluid sample. The sample 182 attenuates or absorbs a portion of the radiation 104, whereby the attenuated radiation such as attenuated radiation 111 generally exhibits an intensity I 0 lower than the intensity I. The value of I 0 is recorded. A beam splitter 184 or a similar device is provided to direct a portion of the radiation 104 to a reference detector 106 without passing through the sample 182. As described above, within the measuring instrument 110, a movable probe can vary the path length L while simultaneously measuring the change in absorbance A. For each value of L, a measured value of the intensity I of the attenuated radiation 111 is recorded, and a measured value of the intensity I 0 of the radiation 104 is recorded. In this way, for a given change in path length Dl, DA is determined as follows. [Number]

[0027] FIG. 1B shows an absorption spectrophotometer shown as system 150. This system can operate in the same manner as system 100 to determine the concentration C of a substance within the measuring instrument 110, where similar components are labeled with the same labels. The difference is that system 150 employs a light source 152 that can be a wide-spectrum light source such as a known ultraviolet / visible / infrared (UV / vis / IR) absorption spectrophotometer, and the radiation 160 represents light that can be generated at intervals of several seconds or minutes over a wide radiation spectrum. Similar to system 100, the radiation 160 can be directed to a reference detector 156 to measure I 0 , while the attenuated radiation 161 is measured at detector 112 after passing through the fluid sample within the measuring instrument 110.

[0028] In both the embodiments of FIGS. 1A and 1B, a control system 130 is provided to facilitate a reasonable and improved operation of each absorption spectroscopy measurement. Briefly stated, the control system 130 may comprise a non-transitory computer-readable storage medium having instructions that, when executed, such as by using an electronic processor, perform one or more of the operations described below. The control system 130 may comprise various components including a dedicated electronic controller, communication interface routines, or algorithms for controlling the operation of various components of the system 100 or system 150. The control system 130 may control the operation of the system 100 or system 150, including the operation of the system in different operating modes. In the “standard” slope spectroscopy mode, the concentration C is determined by determining DA using the measured values of I and I 0 as described above for each value of L. In the “reference signal-free” mode or NRS slope spectroscopy mode, the system 100 or system 150 may operate to determine the concentration C without the measurement of I 0 as described below. The NRS mode of slope spectroscopy will result in greater flexibility and speed in making measurements, and potentially greater accuracy.

[0029] According to an embodiment of the present disclosure, the NRS slope spectroscopy mode may be used or initiated routinely, or may be initiated when certain stability criteria for operating the absorbance spectroscopy system are met, in which case Equation 3B for determining A may be simplified. The stability criteria may be met, for example, when the variation in the intensity of the absorbance is below a threshold, as further discussed below. According to Equation 3B (see above) outlining the absorbance calculation by a known slope spectroscopy approach, the change in absorbance DA between a first instance t 1 (corresponding to a first path length L 1 ) and a second instance t 2 (corresponding to a second path length L 2 ) is the parameter

Number

[0030] However, the inventors have found that under certain circumstances, item

Number

Number

Number

[0031] When the instability of the incident intensity is low enough to enable measurements using the new NRS slope spectroscopy mode, the determination of the value can be made according to the application. However, generally, in a situation where I 0 varies very slightly over a given time, this variation in the incident intensity can be specified as ±α%. Accordingly, the term alpha is

Number

Number

[0032] Therefore, depending on the application, an upper limit can be set for the maximum value of a to determine when the NRS slope spectroscopy measurement mode should be adopted. In one example, for absorbance measurements regulated under the United States Pharmacopeia (USP) guidelines regarding the operation of ultraviolet-visible spectrophotometers, the USP requires that the absorbance deviation be less than ±0.01. Therefore, for slope spectroscopy measurements performed according to the USP guidelines, 0.01 > log(1 + α), and It means that |α| < 2.33%. Therefore, in some embodiments, the stability criterion can be met when the absorbance deviation is less than a specific value such as less than ±0.03, less than ±0.02, or less than ±0.01. In the latter case, the stability criterion equivalently corresponds to the case where |α| < 2.33%, where a can be defined by the formula described herein.

[0033] In one example, a slope spectroscopic apparatus including an LED light source generally configured according to the embodiment of FIG. 1A was used to determine the light source stability. Over a specific test period, the maximum I max and the minimum I min of the incident intensity were recorded along with the following results.

Number

[0034] Therefore, in the above example, the value of a is far below the upper limit of 2.33% set by the USP, and the use of the NRS slope spectroscopy may be appropriate.

[0035] The use of NRS slope spectroscopy provides advantages for determining the material concentration in a fluid sample, including the ability to measure the concentration more accurately, more quickly, and in a more dynamic manner. FIG. 2 shows an absorption spectroscopic apparatus shown as system 200 according to an embodiment of the present disclosure. System 200 may include a small light source 102, a modification of a measuring device 110 coupled to the small light source 102, and a detector 112 disposed adjacent to the measuring device 110. The small light source 102 may have one or more light-emitting diodes for generating radiation 210 at one or more target wavelengths as discussed above with respect to FIG. 1A.

[0036] In this variation, the measuring device 110 has a movable probe 208 that can be an optical fiber, a fibrette, or a bundle of fibers, configured to conduct radiation 202 into a sample chamber container 215 containing a fluid sample 211 that contains the material of interest whose concentration C is being measured. Radiation 210 enters the movable probe 208 along the probe axis 206 and is directed to pass therethrough. As shown in FIG. 2, the movable probe 208 can translate parallel to the container wall 212 of the sample container 210 so as to vary the radiation path length L. In particular, the path length L represents the distance between the probe tip 208A and the lower portion of the container wall 212. Thus, the value of L corresponds to the distance that the radiation can travel through the sample 182 when the fluid sample is disposed within the sample container 215. Note that a window that is transmissive to radiation 210 is provided to conduct the radiation out of the sample container 215, emerging as attenuated radiation 220, whose intensity I can be detected by a detector 222. Suitable examples of the detector 112 include, among others, a photomultiplier tube, a photodiode, an avalanche photodiode, a charge-coupled device (CCD), and an intensified CCD. Although shown as being disposed in a line-of-sight direction with respect to the probe 172, in various embodiments, the detector 112 can be integrated into the measuring device 110B or positioned remotely by operably coupling the detector 112 to an optical delivery device (not shown) that can carry the electromagnetic radiation traveling through the sample to the detector. The optical delivery device can be fused silica, glass, plastic, or any transmissive material suitable for the wavelength range of the electromagnetic source and the detector. The optical delivery device can be formed of a single fiber or a plurality of fibers, which can have different diameters depending on the use of the measuring device 110. In various non-limiting embodiments, the fiber diameter is in the range from about 0.005 mm to about 20.0 mm.

[0037] To facilitate concentration measurements using an approach where DA / DL is equal to eC, the drive component (not shown separately) can be a motor that translates the probe tip 208A along the probe axis 206. The drive component can provide continuous motion or can be set to vary the path length L in precise steps. In various non-limiting embodiments, suitable examples of the drive component include a stepper motor, a servo motor, a piezo motor, an electric motor, and a magnetic motor, or any device that can be controlled to provide a variable path length L through the sample. In some embodiments of incremental or stepped motion, the movable probe 208 is moved in increments in the range from 0.2 μm to 1 cm relative to the sample container 215, and more specifically, in increments in the range from 1 μm to 50 μm. In other embodiments, the movable probe 208 can be continuously moved to continuously vary L.

[0038] System 200 further includes a reference detector 204, which can function similarly to reference detector 106 to measure the incident intensity I of radiation 210 as generally discussed above. In this embodiment, system 200 can also include a control system 130. Various inputs to control system 130 can include I 0 , L, and I. In one example, information regarding L can be transmitted from component 214, which can be a motor assembly, a sensor, or other component that provides position information. In some implementations, control system 130 can determine that the intensity variations meet stability criteria, whereby system 200 can operate in NRS slope spectroscopy mode where the position of the movable probe changes through a plurality of different positions. Since it is not necessary to record I 0 , at each position of the probe, only the value of L and the value of I of the attenuated radiation 220 are recorded. In this way, the slope parameter m, which is equal to DA / DL or equivalently equal to eC, is m = 0

Number

[0039] FIG. 3 shows an absorption spectroscopic apparatus shown as system 250 according to an embodiment of the present disclosure. System 250 can be considered a variation of system 200 discussed above, and similar components are labeled with the same labels. The difference is that the measuring device 256 includes a sample chamber container 260 having an inlet port 262 for receiving the fluid sample 211; and an outlet port 264 for guiding the fluid sample 211 from the sample chamber container 260. Therefore, the measurement system 110A can be used to couple to the shown processing system 270 to provide a dynamic measurement of the concentration C of the material in the fluid sample as the fluid sample passes through the measuring device 256. Therefore, the processing system 270 can represent any suitable system that generates a fluid sample to be measured, such as a chromatography system, a protein purification system, a filtration system, or other fluid processing system. Thus, system 250 provides an architecture for dynamically measuring the concentration in a fluid sample when the fluid sample being measured is flowing and can vary, such as the concentration C of the material being measured. The advantage provided by this approach is that under conditions where the intensity variation of the light source is below a threshold condition or a threshold meaning a threshold, it is not necessary to measure I 0 to determine C, so it is only necessary to record the measured value of the transmission intensity I in conjunction with the movement of the probe 208 used to vary L, as given by Equation (4).

[0040] To further illustrate the determination of concentration C using the embodiments of the LED light source, FIG. 4 shows an exemplary absorption spectrum according to an embodiment of the present disclosure. In this example, the graph of FIG. 4 shows the radiation intensity detected as a function of wavelength in the near ultraviolet region. Three spectra are shown: spectrum 402, spectrum 404, and spectrum 406, each consisting of a single peak, which represent the detected intensity of the UV light emitted from the LED light source.

[0041] Therefore, spectrum 402 presents the data collected in a first instance when the radiation path length is directed through a probe positioned at a first location and defines a path length L through the fluid sample. 1 Similarly, spectrum 404 presents the data collected in a second instance when the radiation path length is directed through a probe positioned at a second location and defines a path length L through the fluid sample. 2 Spectrum 406 presents the data collected in a third instance when the radiation path length is directed through a probe positioned at a third location and defines a path length L through the fluid sample. For the time frame represented between the first and second instances, assuming that the concentration C is equal to DA / (DLe), C is directly derived by determining the intensity difference between the intensity I of spectrum 404 3 and the intensity I of spectrum 402. This is because DL is simply L 2 -L 1 and DA is simply logI1 - logI2 under the condition that the light source intensity variability is below an acceptable threshold. Similarly, for the time frame represented between the second and third instances, C is directly derived by determining the intensity difference between the intensity I of spectrum 406 2 -L 1 and the intensity I of spectrum 404. 3 and the intensity I of spectrum 404. 2 By determining the intensity difference between them, C is directly derived.

[0042] This NRS slope spectroscopy approach can be easily extended, for example, to record multiple different measured values of I at multiple different probe positions in order to more accurately determine the concentration without measuring I 0 at these positions. In other words, I 1 and L 1 are recorded at a first probe position, I 2 and L 2 are recorded at a second probe position, I 3 and L 3 are recorded at a third probe position, and so on. In some implementations, the determination of C can be performed in the following manner, where, according to Lambert-Beer's law, C = (DA / DL) / e. The intensity data I 1 , I 2 , I 3 is converted to absorbance data A (equivalent to logI) by data for determining logI 1 , logI 2 , logI 3 , etc. Linear regression is performed based on a set of data in which A is plotted as a function of L for three or more probe positions in order to determine a regression line whose slope is proportional to (DA / DL). In this case, DA and DL are determined not from the exact values of, for example, L 1 , I 1 , L n , and logI n , but from the respective logI and L values at both ends of the regression line. In this way, the calculated concentration C can more accurately reflect the true value as compared to the concentration determined from a set of intensity and path length measurements performed at only two probe positions.

[0043] Furthermore, since the measured values of the incident intensity are not recorded for different probe positions, in embodiments of a small LED light source, the overall duration of the set of intensity measurements sufficient to determine C can be reduced to just a few seconds.

[0044] FIG. 5 presents a process flow 500 according to an embodiment of the present disclosure. At block 502, the intensity I 0The variation of [[]] is measured. In some embodiments, the light source can be a known spectrometer, such as an ultraviolet / visible or ultraviolet / visible / infrared spectrometer, configured to generate radiation over a wide range of wavelengths. In other embodiments, the light source can be a small LED light source based on a single LED type that emits radiation in a narrow wavelength range, or a small LED light source based on a plurality of different LEDs that emit radiation in a plurality of different narrow wavelength ranges. The intensity variation can be specified as ±α%, and can be measured over a given period, such as a few seconds or a predetermined interval of up to several minutes.

[0045] In decision block 504, a determination is made as to whether the intensity variation of I 0 is below a threshold. The threshold can be set based on a standard for measuring a given class of materials, a standard for operating a given type or device, or any suitable criterion. In some non-limiting embodiments, the threshold for the variation of I 0 can be ±0.03, ±0.02, or ±0.01. In the latter case, the stability criterion corresponds equivalently to the case where |α| < 2.33%, where a can be defined by the equations described herein.

[0046] If so, the flow proceeds to block 506.

[0047] In block 506, when the probe is in the first position, the probe radiation from the light source is directed to pass through the probe. The probe can be an optical fiber, a fibril, a bundle of fibers, or other suitable structure adapted to conduct the probe radiation. At the first probe position, the probe tip can be placed near or within the fluid sample, and the first probe position acts to define a first path length L 1 of the probe radiation through the fluid sample. In particular, the path length L 1 can represent the distance between the probe tip and the wall of the sample container or other vessel containing the fluid sample.

[0048] At block 508, when the probe is disposed at the first position, a procedure is executed to measure the transmitted intensity I of the probe radiation after the probe radiation has passed through the fluid sample. The transmitted intensity can be measured by any suitable detector, such as an electronic detector adapted to detect radiation over the wavelength range of the probe radiation. 1 At block 510, when the probe is disposed at the second position, the probe radiation from the light source is directed to pass through the probe. The second probe position may define a second path length L of the probe radiation through the fluid sample.

[0049] At block 512, when the probe is disposed at the second position, a procedure is executed to measure the transmitted intensity I of the probe radiation after the probe radiation has passed through the fluid sample. 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L

[0050] At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L

[0051] At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 1 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 1 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 1 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 1 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L 2 At block 514, a procedure is executed to determine the concentration C of the material in the fluid sample based on L, log I, L, and log I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L

[0052] At decision block 504, if it is determined that the variation of I 0 is not below the threshold, the flow proceeds to block 516. At block 516, when the probe is disposed at the first position, an operation is executed to measure the incident intensity I 01 of the probe radiation before passing through the measured fluid sample. This measurement may represent, for example, the first measurement in a series of absorbance measurements for the material in the fluid sample.

[0053] In this situation, the flow proceeds to block 506A, which is generally executed according to block 506 described above.

[0054] After block 506A, the flow proceeds to block 518, where when the probe is placed at the first position, the transmitted intensity I of the probe radiation after passing through the fluid sample is measured. It should be noted that the operations of blocks 516, 518, and 506A can be basically executed simultaneously. 1

[0055] Next, the flow proceeds to block 520, where when the probe is placed at the second position, the incident intensity I of the probe radiation before passing through the fluid sample is measured. 02

[0056] In this situation, the flow proceeds to block 510A, which is generally executed according to block 510 described above. After block 510A, the flow proceeds to block 522, where when the probe is placed at the second position, the transmitted intensity I of the probe radiation after passing through the fluid sample is measured. It should be noted that the operations of blocks 520, 522, and 510A can be basically executed simultaneously. 2

[0057] Next, the flow proceeds to block 524, where an operation to determine the concentration C of the material in the fluid sample is executed based on L, I, L, I, I, I, and I. For example, the concentration C can be calculated as C = (DA / DL) / e according to Lambert-Beer's law, where DL is given by |L - L|, and DA is given by logI - logI or log(I / I). 1 I 1 L 2 I 2 I 01 I 02 1 L 2 1 logI 2.+ log(I 01 / I 02 )

[0058] ​​​​​Figure 6 presents another process flow 600 according to an embodiment of the present disclosure. At block 602, in a first instance, when the probe is disposed at a first position p1, direct the probe radiation to pass through the probe and define a first path length L of the probe radiation through the fluid sample. 1 The operation of defining is performed. The probe radiation can be generated by light, such as a known spectrometer, such as an ultraviolet / visible or ultraviolet / visible / infrared spectrometer, configured to generate radiation over a wide range of wavelengths. In other embodiments, the light source can be a small LED light source based on a single LED type that emits radiation in a narrow wavelength range, or a small LED light source based on a plurality of different LEDs that emit radiation in a plurality of different narrow wavelength ranges.

[0059] At block 604, in a first instance, measure the transmitted intensity I of the probe radiation after passing through the fluid sample. 1 is measured.

[0060] At block 606, in a plurality of additional instances, when the probe is disposed at a plurality of additional positions p2, p3, …, direct the probe radiation to pass through the probe and perform the operation of defining a plurality of additional path lengths L 2 , L 3 of the probe radiation through the fluid sample, respectively.

[0061] At block 608, at a plurality of additional distances, measure the transmitted intensity I 2 , I 3 , ··· of the probe radiation after passing through the fluid sample, respectively.

[0062] At block 610, the operation involves performing a linear regression to determine the slope m of the line of a set of data plotted with A 1 , A 2 , A 3 … as a function of L 1 , L 2 , L 3 , where m is determined from the ratio of the change in absorbance to the change in path length. A 1 , A2 To determine, etc., logI 1 、logI 2 By determining, etc., the intensity data I 1 、I 2 etc. are converted into absorbance data. In other words, linear regression is performed on a set of data constructed from a plurality of data points, where the data points are, for example, A 1 、L 1 ; A 2 、L 2 ; A 3 、L 3 represent; and so on. In particular, the linear regression performed in block 610 is used to define a line that best fits a set of data points of A and L, where the slope of the fit line defines a valid value of m.

[0063] In block 612, an operation is performed to determine the concentration C of the material in the fluid sample, where C = m / e, e is the molar absorptivity of the material, and m is determined in the same manner as in block 610.

[0064] This configuration has been disclosed with reference to specific embodiments, but numerous modifications, alterations, and changes to the described embodiments are possible without departing from the spirit and scope of the disclosed configuration as defined in the appended claims. Accordingly, this configuration is not limited to the described embodiments, and it is intended to have the full scope defined by the language of the following claims and their equivalents.

Claims

1. A method for determining the concentration of a material, comprising: determining whether the fluctuations in the intensity of the probe radiation emitted by the light source of an absorbance spectroscopy system meet a stability criterion; When the probe is disposed at the first position, directing the probe radiation to pass through the probe and defining a first path length L of the probe radiation through a fluid sample containing the material １ ; and When the probe is disposed at the first position, measuring a transmission intensity I of the probe radiation after passing through the fluid sample １ ; When the probe is disposed at the second position, directing the probe radiation so as to pass through the probe and defining a second path length L of the probe radiation passing through the fluid sample ２ ; and When the probe is disposed at the second position, measuring a transmitted intensity I of the probe radiation after passing through the fluid sample; and ２ and When the stability criteria are met, L １ , I １ , L ２ , and I ２ determining the concentration C of the material in the fluid sample based on A method comprising the above.

2. When the stability criterion is met, the concentration C is: It is determined that C = (DA / DL) / e, where e is the molar extinction coefficient of the material and DL is the absolute value of the difference between L １ and L ２ and ΔA = log I1 - log I2. The method according to claim 1

3. The variation in the intensity of the probe radiation is given by a, where a = (I ｍａｘ − I ｍｉｎ ) / I ｍｉｎ , and I ｍａｘ is the maximum value of the intensity of the radiation recorded over a given period, and I ｍｉｎ is the minimum value of the intensity recorded over the given period, the method according to claim 1.

4. The method according to claim 3, wherein the duration of the given period is from 1 second to 100 seconds.

5. The method according to claim 1, wherein when the stability criterion is met, the incident intensity of the probe radiation before passing through the fluid sample is not measured at the first position or the second position.

6. The transmitted intensity I １ and the transmitted intensity I ２ are determined by the measuring instrument of the absorbance spectroscopy system, and the measuring instrument is: It has a sample container for accommodating the fluid sample, the sample container including a container wall, wherein the probe is movable along a probe direction with respect to the container wall so as to change the path length of a probe signal passing through the fluid sample from the first path length L １ to the second path length L ２ ​ The method according to claim 1.

7. The stability criterion includes the state of the light source where the fluctuations in the intensity are below a threshold value. When the fluctuations in the intensity are above the threshold value, the method further comprises: When the probe is disposed at the first position, measuring an incident intensity I of the probe radiation before passing through the fluid sample ０１ ; When the probe is disposed at the second position, measuring an incident intensity I of the probe radiation before passing through the fluid sample ０２ and L １ 、I １ 、L ２ 、I ２ 、I ０１ and I ０２ Based on L, I, L, I, I, and I, determining the concentration C of the material in the fluid sample The method according to claim 1.

8. C = (DA / DL) / e, where DL is given by |L １ - L ２ |, and DA is given by log I １ - log I ２ + log(I ０１ / I ０２ ), the method according to claim 7

9. The probe is arranged at the first position in a first instance and at the second position in a second instance. The method further comprises: For at least one additional instance, when the probe is disposed at at least one additional location, directing the probe radiation to pass through the probe and defining at least one additional path length L for the probe radiation passing through the fluid sample ｎ respectively; In the at least one additional instance, measuring the transmitted intensity I of the probe radiation after the fluid sample has passed through; and ｎ and Execute linear regression for A １ A ２ A ｎ … as L １ L ２ L ｎ Determining the slope m of the line of a set of data plotted as a function of L, where A is equal to logI The method according to claim 1.

10. A computer program for causing a processor to: determine whether the fluctuations in the intensity of the probe radiation emitted by the light source of an absorbance spectroscopy system meet a stability criterion; When the probe is disposed at a first position, directing the probe radiation from a light source to pass through the probe to define a first path length L of the probe radiation through a fluid sample; １ a procedure for defining; Procedure for receiving the transmitted intensity I of the probe radiation after passing through the fluid sample １ ; When the probe is disposed at the second position, direct the probe radiation from the light source to pass through the probe, and define a second path length L of the probe radiation passing through the fluid sample; ２ a procedure for defining; The procedure for receiving the transmitted intensity I of the probe radiation after passing through the fluid sample; and ２ and When the variation in the strength satisfies the stability criterion, L １ , I １ , L ２ , and I ２ A procedure for determining the concentration C of the material in the fluid sample based on A computer program for causing a processor to execute the above.

11. The concentration C is: It is determined that C = (DA / DL) / e, where e is the molar extinction coefficient of the material, and DL is the absolute value of the difference between L １ and L ２ and ΔA = log I1 - log I2. The computer program according to claim 10

12. The variation in the intensity of the probe radiation is given by v, where v = (I ｍａｘ − I ｍｉｎ ) / I ｍｉｎ , I ｍａｘ being the maximum value of the intensity of the radiation recorded over a given period, I ｍｉｎ being the minimum value of the intensity recorded over the given period, the stability criterion being satisfied when v is below a threshold value, computer program according to claim 10.

13. A computer program for causing a processor to: When the variation in the strength satisfies the stability criterion, without receiving a measurement value of the incident intensity I of the probe radiation at the first position before passing through the fluid sample, and without receiving a measurement value of the incident intensity I of the probe radiation at the second position before passing through the fluid sample, cause a procedure for determining the concentration C to be executed. The computer program according to claim 10. ０１ The computer program according to claim 10, which causes a procedure for determining the concentration C to be executed without receiving a measurement value of the incident intensity I of the probe radiation at the first position before passing through the fluid sample and without receiving a measurement value of the incident intensity I of the probe radiation at the second position before passing through the fluid sample. ０２ The computer program according to claim 10, which causes a procedure for determining the concentration C to be executed without receiving a measurement value of the incident intensity I of the probe radiation at the first position before passing through the fluid sample and without receiving a measurement value of the incident intensity I of the probe radiation at the second position before passing through the fluid sample.

14. A computer program for causing a processor to: When the fluctuations in the intensity do not meet the stability criterion: When the probe is disposed at the first position, a procedure for receiving an incident intensity I of the probe radiation before passing through the fluid sample ０１ ; When the probe is disposed at the second position, a procedure for receiving an incident intensity I of the probe radiation before passing through the fluid sample ０２ and L １ 、 I １ 、 L ２ 、 I ２ 、 I ０１ and I ０２ Based on, the procedure for determining the concentration C of the material in the fluid sample The computer program according to claim 10, for causing the processor to execute the above.

15. Cause the processor to execute a procedure for determining C by calculating the change DA in absorbance from the first instance to the second instance, where C = (DA / DL) / e, and DL is given by |L １ - L ２ |, and DA is given by logI １ - logI ２ + log(I ０１ / I ０２ ), the computer program according to claim 14.

16. The probe is arranged at the first position in a first instance and at the second position in a second instance. The computer program causes the processor to: For at least one additional instance, when a probe is disposed at at least one additional location, direct the probe radiation through the probe to the light source so as to define at least one additional path length L for the probe radiation passing through the fluid sample ｎ in each case; In the at least one additional instance, a procedure for measuring the transmitted intensity I of the probe radiation after the fluid sample has passed through; and ｎ and Execute linear regression for A １ A ２ A ｎ … as L １ L ２ L ｎ Procedure for determining the slope m of the line of a set of data plotted as a function of L, where A is equal to logI The computer program according to claim 10, for causing the processor to execute the above.

17. A light source for generating a probe signal; and Measuring equipment for receiving the probe signal A measuring device comprising the above, wherein the measuring equipment: A sample container for containing a fluid sample, the sample container including a container wall; A probe configured to direct the probe signal through the sample container, the probe being movable along the probe direction with respect to the container wall so as to change the path length L of the probe signal through the fluid sample; A detector arranged to receive the probe signal after it has passed through the container wall; and A control system comprising: Determining whether fluctuations in the intensity of the probe radiation emitted by the light source meet a stability criterion; and Calculating the concentration C of the material in the fluid sample based on the change in the intensity of the probe signal measured as a function of the change in the path length L when the fluctuations in the intensity meet the stability criterion A control system configured as such A measuring device having.

18. The control system is: When the probe is disposed at the first position, receive the transmitted intensity I of the probe radiation after passing through the fluid sample; １ ​ When the probe is disposed at the second position, receive the transmitted intensity I of the probe radiation after passing through the fluid sample; and ２ and Calculate C as (DA / DL) / e, where e is the molar extinction coefficient of the material and DL is the absolute value of the difference between L １ and L ２ and ΔA = log I1 - log I2, where L １ is the first path length at the first position and L ２ is the second path length at the second position The measuring device according to claim 17, configured as such.

19. The control system is configured to determine the concentration C without receiving a measurement value of the incident intensity I of the probe radiation at the first position before passing through the fluid sample and without receiving a measurement value of the incident intensity I of the probe radiation at the second position before passing through the fluid sample when the variation in the intensity satisfies the stability criterion. ０１ The measuring device according to claim 18, wherein the measuring device is configured to determine the concentration C without receiving a measurement value of the incident intensity I of the probe radiation at the first position before passing through the fluid sample and without receiving a measurement value of the incident intensity I of the probe radiation at the second position before passing through the fluid sample. ０２ when the variation in the intensity satisfies the stability criterion.

20. The measuring device according to claim 17, wherein the light source includes a light emitting diode (LED) for generating radiation at a target wavelength in the range from ultraviolet to infrared.

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