CN111474087A - Method and device for online quantitative monitoring of plasma liquid phase active particle space-time distribution - Google Patents

Abstract

The invention provides a method and a device for online quantitative monitoring of the spatiotemporal distribution of plasma liquid phase active particles. The principle of the invention is as follows: the liquid-phase active particles react with the chemical color-developing reagent to generate color-developing products, and the color-developing distribution can be observed in the solution; Absorption spectrum, from which the absorption spectrum curve with wavelength (λ) as the abscissa and absorbance (A) as the ordinate can be drawn; according to the Beer-Lambert law, according to the calibration of a certain liquid phase active particle at the maximum absorption wavelength Absorbance-concentration standard curve, the absorbance of the treated aqueous solution can be obtained under the same detection conditions, and the concentration of the active particles in the liquid phase can be obtained by comparing the standard curve. The invention can realize the time-space distribution measurement of the plasma activation liquid particles more efficiently, accurately and in real time, the device is simple and easy to operate, and can be applied to various practical application occasions.

Description

Method and device for online quantitative monitoring of temporal and spatial distribution of plasma liquid active particles

technical field

The invention relates to the quantitative real-time online monitoring of the penetration distribution of the plasma liquid phase long-lived active particles, promotes the further deepening of the plasma technology in the field of biomedicine, and has great significance in the field of clinical application of plasma biomedicine.

Background technique

Plasma will generate a large number of neutral active particles in the gas phase, such as H, O, O ₃ , OH, NO, NO ₂ , N ₂ O, H ₂ O ₂ , N ₂ O ₅ , etc. After the particles come into contact with the liquid, a variety of liquid-phase active particles will be generated in the liquid phase. The liquid-phase active particles will participate in a large number of biochemical reactions and are indispensable participants in the physiological process, which will produce anti-cancer, sterilization, etc. role in biomedicine.

With the development of the world's big cities and the continuous increase of population density, the large-scale spread and pollution of various bacteria and viruses is a major challenge in the field of public health today. Various medical facilities and devices are indispensable in large-scale diagnosis and treatment of diseases such as hospitals and clinics. Therefore, the requirements for sterilization and disinfection brought by instruments and workplaces to kill microorganisms continue to be consistent. Traditional sterilization methods include ultraviolet radiation sterilization, ethylene oxide, high temperature and high pressure steam and so on. However, ultraviolet rays and ethylene oxide may cause harm to the human body, and high temperature and high pressure steam may cause damage to precision instruments. The highly chemically active free radicals generated by the plasma liquid phase have become a new type of sterilization and disinfection technology, which has the advantages of safe and simple operation, high efficiency, low cost, and no drug residues.

In terms of cancer treatment, plasma activation solution can induce apoptosis of cancer cells. Cancer cells maintain the integrity of the cell membrane structure during the process of death, and will not release intracellular enzymes and cell debris products into the surrounding environment of cells, thereby preventing It will affect the surrounding cells and cause an inflammatory response in the human body. Many studies have pointed out that the systemic inflammatory response caused by a large number of cancer cells dying in a short period of time in cancer patients is an important reason for the death of cancer patients.

On the other hand, the plasma activation solution showed selectivity for the inactivation of cancer cells. That is, an appropriate dose of plasma can remove cancerous cells attached to the body without causing damage to normal cells in the body. Although traditional cancer treatment methods such as radiotherapy can kill tumor cells in patients, an obvious defect is that while radiation inactivates tumor cells, it can also cause damage to normal cells in the body. Therefore, this treatment method has non-negligible side effects, such as the patient's immune system will be destroyed during the treatment.

Long-lived active particles have a long existence time due to their stable chemical properties. Common long-lived particles are: H ⁺ , O ₃ , H ₂ O ₂ , NO ₂ ^- . A large number of studies have shown that these active particles have important biomedical effects. The spatiotemporal distribution of active particles is closely related to their infiltration process, which is a necessary link in the application of plasma medicine. At present, although the simulation of the spatial distribution of active particles can be achieved, there is still a lack of effective experimental detection methods to confirm. Therefore, accurate qualitative and quantitative diagnosis of long-lived active particles in the liquid phase is of great significance for the effective regulation of the dose of key plasma active particles.

At present, there are various experimental detection methods for the concentration of active particle components generated during the interaction between plasma and solution, including photochemical probe method, colorimetric method, electron spin resonance spectroscopy, etc. However, the current measurement method has the following shortcomings:

1. Only the average concentration in the activation solution can be measured;

2. It is impossible to measure the spatial penetration distribution of a specific active particle in real time;

3. It is impossible to measure the time distribution of a particle at the same location in real time;

4. The detection equipment is cumbersome and not portable, the operation is complicated, and it takes a long time;

Off-line measurement of active particle concentration means that the conditions for active particle generation cannot be adjusted in time during the application process, that is, it cannot be controlled according to the requirements of the active particle dosage or type during use, which will make the development of plasma in the biomedical field. constrained. Therefore, the development of plasma biomedicine urgently requires the realization of online measurement technology for the spatial distribution of active particles.

SUMMARY OF THE INVENTION

The purpose of this application is to realize real-time online monitoring of the temporal and/or spatial distribution of active particles in the plasma liquid phase.

The inventive concept of the present application is as follows:

According to ultraviolet absorption spectroscopy, ultraviolet light is incident into the plasma activation liquid, and the intensity change after the light is absorbed can be measured, which can qualitatively and quantitatively analyze some specific particles.

Liquid-phase active particles such as H ⁺ , O ₃ , H ₂ O ₂ , and NO ₂ ^- react with chemical color-developing reagents to generate color-developing products, and color-developing distribution can be observed in the solution. The molecular structures of different active particles are different, so that the corresponding color products have different absorption capabilities for different wavelengths of light. Therefore, there is a maximum absorption wavelength of selective absorption characteristics, forming a maximum absorption peak, and then generating a unique absorption spectrum. The concentration of the same active particle is different, and the degree of light absorption is also different. For example, taking the wavelength (λ) as the abscissa and the absorbance (A) as the ordinate, the absorption spectrum curve of the particle can be drawn, and the active particle can be qualitatively or quantitatively analyzed by the curve.

According to the Beer-Lambert law, the absorbance of the solution A=a×b×c, where a is the absorption coefficient, b is the thickness of the light passing through the solution layer, and c is the concentration of active particles in the solution.

Therefore, the A-c standard curve of a certain liquid-phase active particle is firstly made at the maximum absorption wavelength, and then the absorbance of the treated aqueous solution is obtained under the same detection conditions, and the concentration of the liquid-phase active particle can be obtained by comparing the standard curve.

The present invention specifically provides the following technical solutions:

1. A method for online quantitative monitoring of the time distribution of active particles in the plasma liquid phase, including:

1) Build a test system

Take a transparent vessel to hold the solution to be treated, and add a color-developing reagent corresponding to the active particles to be tested in the solution to be treated;

2) Test for setting sampling points

Turn on the plasma generator to generate liquid-phase active particles in the solution to be treated; turn on the ultraviolet light source and the spectrometer at the same time, so that the ultraviolet band beam is incident from one side of the transparent vessel, passes through the sampling position of the solution to be treated, and starts from the other side of the transparent vessel. Exit, carry out spectral analysis on the outgoing light, and obtain the absorbance at the maximum absorption wavelength of the active particle to be measured; according to the Beer-Lambert law, compare the absorbance at the calibrated maximum absorption wavelength-concentration curve of the active particle to be measured , the concentration of the active particle to be tested at the sampling location is obtained;

The plasma generator, the ultraviolet light source and the spectrometer continue to work, and by continuously measuring the concentration of the active particles to be measured at the same sampling position during the penetration of the active particles, the time distribution of the active particles in the plasma liquid phase at this position is obtained.

2. The method for online quantitative monitoring of the spatial distribution of active particles in the plasma liquid phase, including:

1) Build a test system

Take a transparent vessel to hold the solution to be treated, and add a color-developing reagent corresponding to the active particles to be tested in the solution to be treated; the plasma generator generates plasma diffusion to generate liquid-phase active particles in the solution to be treated;

2) A sampling point test

The ultraviolet band beam is incident from one side of the transparent vessel, passes through the sampling position of the solution to be treated, and exits from the other side of the transparent vessel, and performs spectral analysis on the emitted light to obtain the absorbance at the maximum absorption wavelength of the active particle to be tested; according to Bill -Lambert (Beer-Lambert) law, compare the absorbance at the calibrated maximum absorption wavelength-concentration curve of the active particle to be measured, and obtain the concentration of the active particle to be measured at the sampling position;

3) Testing at other sampling points

Repeat steps 1) and 2), in which each time, the solution to be treated is re-supplied, and according to the set next sampling position, the incident position and the exit position of the ultraviolet band beam are synchronously adjusted, and other conditions remain unchanged; The concentration of the active particles to be tested at the sampling position is obtained; and the spatial distribution of the active particles in the plasma liquid phase is obtained by summarizing.

Further, the absorbance at the calibrated maximum absorption wavelength-concentration curve of the active particles to be measured, the specific calibration process is as follows: prepare a plurality of standard solutions of active particles to be measured with concentration gradients, and then add the corresponding standard solutions to each standard solution respectively. The color reagent is fully reacted, and then the absorbance at the maximum absorption wavelength of the corresponding reaction product is detected by a spectrometer. The measured result matches the concentration of the active particles to be tested to make a standard curve.

Further, the discharge form for generating plasma-activated water is creeping discharge, plasma jet or plasma brush.

Further, the solution to be treated is PBS solution, deionized water, physiological saline or other medical aqueous solutions.

Further, the transparent vessel and the solution to be treated are replaced with biological tissue, and correspondingly, a color-developing reagent corresponding to the active particles to be tested is injected into the biological tissue.

Further, the active particles to be tested are H ⁺ , O ₃ , NO ₂ ^- or H ₂ O ₂ ; the corresponding color reagents are:

Methyl orange, the characteristic wavelength of the maximum absorption spectrum of methyl orange and H ⁺ reaction product is λ=462nm;

Indigo reagent, the characteristic wavelength of the maximum absorption spectrum of the reaction product of indigo reagent and O ₃ is λ=600nm;

Gries reagent, the characteristic wavelength of the maximum absorption spectrum of Gries reagent and NO ₂ ^-reaction product is λ=540nm;

Ammonium metavanadate, the characteristic wavelength of the maximum absorption spectrum of its reaction product with H ₂ O ₂ is λ=450nm, or acidify the treated solution with sulfuric acid in advance, add sodium azide to remove NO ₂ ^- particles in the solution , titanyl sulfate is detected, and the characteristic wavelength of the maximum absorption spectrum of its reaction product with H ₂ O ₂ is λ=407 nm.

3. A device for on-line quantitative monitoring of the temporal and spatial distribution of plasma liquid-phase active particles, including a transparent vessel, a plasma generator, an ultraviolet light source, an optical fiber probe holder, a spectrometer and a computer; the transparent vessel contains the solution to be treated, and the solution to be treated contains The color reagent corresponding to the active particles to be tested; the plasma generator keeps a constant distance from the transparent vessel, so that the plasma can generate liquid-phase active particles in the solution to be treated; the two sides of the transparent vessel are provided with pairs of The optical fiber probes are respectively connected with the ultraviolet light source and the spectrometer through the optical fiber; the paired optical fiber probes are installed on the optical fiber probe bracket and their positions are synchronously adjustable; the spectral information output end of the spectrometer is connected with the data input end of the computer.

Further, the transparent vessel adopts a quartz cuvette.

Further, the optical fiber probe bracket is fixedly installed on the three-dimensional platform, so as to realize the position adjustment of the optical fiber probe in the vertical direction and the horizontal direction.

Further, the computer also stores the absorbance-to-be-measured active particle concentration curve at the calibrated maximum absorption wavelength and a control program, and the control program realizes the following functions when running:

controlling and recording the positions of the paired fiber optic probes;

Record the type of active particles to be tested;

Control the working status of plasma generator, ultraviolet light source and spectrometer and record working time;

Record real-time absorption spectrum curve;

According to the Beer-Lambert law, the absorbance at the calibrated maximum absorption wavelength-concentration curve of the active particle to be measured is compared to obtain the active particle to be measured at different sampling positions (the positions of the paired fiber probes) concentration;

Summarizes the temporal and/or spatial distribution of the plasma liquid-phase active particles.

The present invention can realize the time-space distribution measurement of plasma activation liquid particles in a more efficient, accurate and real-time manner, and has the following advantages:

(1) The absorption spectra of different substances are different, that is, the method can achieve extremely high selectivity and can realize qualitative detection;

(2) The response speed of the spectrum is extremely fast, which can realize continuous online monitoring of active particles;

(3) Ultraviolet absorption spectroscopy is to detect the optical signal, and the position of the optical signal can be adjusted by the optical fiber probe, which can realize the measurement in space.

(4) Fix the optical fiber probe to measure the concentration of active particles at the same position at different times, which can realize time measurement.

(5) The device is simple and easy to operate and can be applied to a variety of practical applications.

(6) The system monitoring activation solution has low cost.

Description of drawings

FIG. 1 is a schematic diagram of the principle of an apparatus according to an embodiment of the present invention;

Figure 2 shows the reaction process of Griess reagent and NO ₂ ^- .

Description of reference numbers:

1-UV light source; 2-Quartz cuvette; 3-Three-dimensional platform; 4-Fiber probe holder; 5-Spectrometer; 6-Computer; 7-Creep plasma generator; 701-High voltage electrode; 702-Dielectric plate; 703 - Ground electrode; 8 - High voltage AC power supply.

Detailed ways

The present invention will be further described below in conjunction with the accompanying drawings and embodiments. It should be understood that these examples are only used to illustrate the present invention and not to limit the scope of the present invention.

As shown in FIG. 1 , the real-time online monitoring device for quantifying the penetration distribution of plasma liquid phase long-lived active particles in the present embodiment mainly includes an ultraviolet light source 1, a quartz cuvette 2, a three-dimensional platform 3, a fiber optic probe holder 4, and a spectrometer 5 , Computer 6 , Creeping Plasma Generator 7 .

In this embodiment, the time distribution and spatial penetration distribution of particles can be measured by ultraviolet absorption spectroscopy. _{The quartz cuvette} is filled with deionized water as the solution to be treated, ^and the position ^of the quartz _cuvette is fixed. The color-developing reagent reacts to generate a color-developing product, that is, the color-developing distribution can be observed in the solution. At the same time, the ultraviolet light generated by the ultraviolet light source is transmitted through the optical fiber and passes through the color-developing plasma activation solution in the quartz cuvette. The spectrometer receives it, and the spectrometer converts it into an electrical signal to display the spectrum by the computer, so as to complete the measurement of the ultraviolet absorption spectrum of the long-lived active particles such as H ⁺ , O ₃ , H ₂ O ₂ , NO ₂ ^- and so on.

By moving the three-dimensional platform in the vertical direction or the horizontal direction, the collection of ultraviolet absorption spectra at different positions during the penetration process of the active particles is realized. Using the Beer-Lambert law, the liquid phase concentration can be calculated from the absorbance, thereby obtaining Temporal and spatial distribution of active particles.

Optionally, the treatment solution may be a medical aqueous solution, physiological saline, PBS solution, deionized water and other solutions.

Optionally, the discharge form for generating plasma-activated water may be different types of atmospheric pressure-cooled plasma, such as creeping discharge, plasma jet, plasma brush, and the like.

Optionally, the three-dimensional platform can be moved in the vertical direction or the horizontal direction to realize the collection of ultraviolet absorption spectra at different positions during the penetration process of the active particles.

Optionally, the spectrometer can be replaced with a spectrophotometer.

Optionally, quartz cuvettes can be replaced with other transparent vessels that do not reduce light penetration.

Optionally, the penetration distribution measurement of active particles is not limited to plasma-activated water, and the penetration of active particles into skin tissue or other biological tissues can also be measured.

Optionally, the developer for measuring long-lived particles H ⁺ is 0.5% (w/v) methyl orange (chemical formula: C ₁₄ H ₁₄ N ₃ SO ₃ Na, manufacturer: Sinopharm Chemical Reagent Co., Ltd) , or other chromogenic reagents that react with H ⁺ .

Optionally, the chromogenic reagent for measuring the long-lived particle O ₃ is indigo reagent or other chromogenic reagent that reacts with O ₃ .

Optionally, the chromogenic reagent for measuring NO ₂ ^- of long-lived particles is Griess reagent, or other chromogenic reagent for reacting with NO ₂ -.

Optionally, the color-developing reagent for measuring the long-lived particles H ₂ O ₂ is ammonium metavanadate or titanyl sulfate or other color-developing reagents that react with H ₂ O ₂ .

The monitoring of the spatial distribution and the temporal distribution is introduced by taking the monitoring of liquid-phase active particles NO ₂ ^- as an example.

The absorbance ^- NO ₂ -concentration curve at the maximum absorption wavelength was calibrated as follows:

Different concentrations of NO ₂ -standard solutions were prepared, and the concentration gradients adopted were: 1, 2, 5, 10, 20, 50, 100, 200, 500, 1000 μM. Then add Griess reagent to the standard solution to make it fully react, and then use a spectrometer to detect the absorbance at the maximum absorption wavelength λ=540nm, the measured result matches the NO ₂ ^{-concentration} to make a standard curve, and it is fitted to obtain draw a standard curve. The reaction process is shown in Figure 2. The calibration methods for other particles are similar.

The quartz cuvette 200 is filled with deionized water as a solution to be treated, Griess reagent is added as a chemical color developing reagent for NO ₂ ^- , and the position of the quartz cuvette 200 is fixed. The plasma device is discharged for 5 minutes, and the ultraviolet light generated by the ultraviolet light source 100 is transmitted through the optical fiber and then passes through the plasma-activated water in the quartz cuvette 200 and is received by the spectrometer 400. In order to complete the measurement of the UV absorption spectrum of the long-lived active particles NO ₂ ^- , the absorbance was converted into the particle concentration according to the Beer-Lambert law against the above-mentioned standard curve. By moving the three-dimensional platform 300 in the vertical direction and the horizontal direction, the concentration of the active particles at different positions during the penetration process can be measured, so as to obtain the spatial distribution of the liquid particles when the plasma is discharged for 5 minutes.

The quartz cuvette 200 is filled with deionized water as a solution to be treated, Griess reagent is added as a chemical color developing reagent for NO ₂ ^- , and the position of the quartz cuvette 200 is fixed. The three-dimensional platform 300 is fixed in the middle of the petri dish in the vertical direction. After the plasma device is discharged, the ultraviolet light generated by the ultraviolet light source 100 is transmitted through the optical fiber and then passes through the plasma-activated water in the quartz cuvette 200 and is received by the spectrometer 400. The spectrometer 400 After converting it into an electrical signal, the spectrum is displayed by the computer 500, so as to complete the measurement of the ultraviolet absorption spectrum of the long-lived active particle NO ₂ ^- , according to the above-mentioned standard curve, according to the Beer-Lambert (Beer-Lambert) law. Converted to particle concentration. By continuously measuring the concentration of the active particles at the same position during the penetration process of the active particles, the time distribution of the active particles in the plasma discharge activation solution at this position is obtained.

Claims (11)

1. The method for online quantitative monitoring of the time distribution of the plasma liquid-phase active particles is characterized by comprising the following steps:

1) building a test System

Taking a transparent vessel to contain a solution to be treated, and adding a color reagent corresponding to active particles to be detected into the solution to be treated;

2) testing with set sampling points

Starting a plasma generator to generate liquid-phase active particles in a solution to be treated, simultaneously starting an ultraviolet light source and a spectrometer to enable ultraviolet waveband light beams to enter from one side of a transparent vessel, penetrate through a sampling position of the solution to be treated and exit from the other side of the transparent vessel, and carrying out spectrum analysis on emergent light to obtain the absorbance at the maximum absorption wavelength of the active particles to be detected;

the plasma generator, the ultraviolet light source and the spectrometer continue to work, and the time distribution of the plasma liquid-phase active particles at the position is obtained by continuously measuring the concentration of the active particles to be measured at the same sampling position in the active particle permeation process.

2. The method for online quantitative monitoring of the space distribution of the plasma liquid-phase active particles is characterized by comprising the following steps:

1) building a test System

Taking a transparent vessel to contain a solution to be treated, and adding a color reagent corresponding to active particles to be detected into the solution to be treated; the plasma generator generates plasma to diffuse in the solution to be treated to generate liquid-phase active particles;

2) testing of a sampling point

According to the Beer-lambert (Beer-L ambert) law, the concentration curve of the active particles to be detected at the calibrated maximum absorption wavelength is contrasted to obtain the concentration of the active particles to be detected at the sampling position;

3) testing of other sampling points

Repeatedly executing the steps 1) and 2), wherein in each execution, the solution to be processed is provided again, the incident position and the emergent position of the ultraviolet band light beam are synchronously adjusted according to the set next sampling position, and other conditions are unchanged; finally obtaining the concentration of the active particles to be detected at the sampling position; further summarizing the space distribution of the plasma liquid phase active particles.

3. The method according to claim 1 or 2, wherein the calibration of the absorbance-concentration curve of the active particles to be measured at the maximum absorption wavelength is carried out by the following specific calibration process: preparing a plurality of concentration gradient standard solutions of active particles to be detected, adding corresponding color developing reagents into the standard solutions respectively to enable the solutions to react fully, detecting the absorbance at the maximum absorption wavelength of corresponding reaction products by using a spectrometer, matching the detected result with the concentration of the active particles to be detected to form a standard curve, and fitting the standard curve to obtain the standard curve.

4. The method according to claim 1 or 2, wherein the form of the discharge that generates plasma-activated water is a creeping discharge, a plasma jet or a plasma brush.

5. The method according to claim 1 or 2, wherein the solution to be treated is a PBS solution, deionized water, physiological saline or other medical water solution.

6. Method according to claim 1 or 2, characterized in that the transparent vessel and the solution to be treated are replaced by biological tissue, into which, in response, a chromogenic reagent corresponding to the active particles to be detected is injected.

7. The method according to claim 1 or 2,

the active particles to be detected are H⁺、O₃、NO₂ ^-Or H₂O₂(ii) a The corresponding color developing reagents are respectively:

methyl orange, methyl orange and H⁺The characteristic wavelength of the maximum absorption spectrum of the reaction product is lambda-462 nm;

indigo reagent, indigo reagent and O₃The characteristic wavelength of the maximum absorption spectrum of the reaction product is lambda which is 600 nm;

grilis reagent, grilis reagent and NO₂ ^-The characteristic wavelength of the maximum absorption spectrum of the reaction product is lambda 540 nm;

ammonium metavanadate, its reaction with H₂O₂The characteristic wavelength of the maximum absorption spectrum of the reaction product is lambda-450 nm, or the treated solution is acidified by sulfuric acid in advance, and sodium azide is added to remove NO in the solution₂ ^-After the particles, titanyl sulfate was detected, which was in contact with H₂O₂The characteristic wavelength of the maximum absorption spectrum of the reaction product was λ 407 nm.

8. The device for online quantitative monitoring of the time-space distribution of the plasma liquid-phase active particles is characterized by comprising a transparent vessel, a plasma generator, an ultraviolet light source, an optical fiber probe bracket, a spectrometer and a computer; the transparent vessel is filled with a solution to be treated, and the solution to be treated contains a color reagent corresponding to active particles to be treated; the plasma generator keeps a constant distance from the transparent vessel, and liquid-phase active particles are generated in the solution to be treated; the two sides of the transparent vessel are provided with paired optical fiber probes which are respectively connected with an ultraviolet light source and a spectrometer through optical fibers; the paired optical fiber probes are arranged on the optical fiber probe bracket and the positions of the optical fiber probes are synchronously adjustable; the spectrum information output end of the spectrometer is connected with the data input end of the computer.

9. The apparatus of claim 8, wherein the transparent vessel is a quartz cuvette.

10. The device of claim 8, wherein the fiber optic probe holder is fixedly mounted on the three-dimensional platform to enable the position adjustment of the fiber optic probe in the vertical direction and the horizontal direction.

11. The apparatus of claim 8, wherein the computer further stores a calibrated absorbance-active particle concentration profile at a maximum absorption wavelength and a control program, the control program when executed performs the following functions:

controlling and recording the positions of the paired fiber probes;

recording the kind of active particles to be detected;

controlling the working states of the plasma generator, the ultraviolet light source and the spectrometer and recording the working time;

recording a real-time absorption spectrum curve;

according to the Beer-lambert (Beer-L ambert) law, contrasting the absorbance at the calibrated maximum absorption wavelength-concentration curve of the active particles to be detected, and obtaining the concentration of the active particles to be detected at the sampling position;

and summarizing to obtain the time distribution and/or the space distribution of the plasma liquid phase active particles.

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