CN111812054A - Optical fiber sensing microfluidic chip propofol online derivatization detection system and method - Google Patents

Abstract

The invention discloses the technical field of rapid drug detection, in particular to an optical fiber sensing microfluidic chip propofol online derivatization detection system and method. The fluidic mixing chip performs online derivatization of samples and derivatized reagents, and can precisely control the flow rate and flow of the reagents, so that the detection can be completed with a small amount of samples and reagents; the optical fiber sensing technology is used for online detection, which is transmitted through optical fibers. Optical signal, simplifies the structure of the instrument, reduces the size of the instrument, makes the entire analysis system miniaturized and portable, and is suitable for the needs of rapid on-site detection; online derivatization technology is used for rapid detection of biological samples, and the detection sensitivity and accuracy are improved through derivatization , to solve the problem that spectral detection is easily interfered by other impurities in the sample.

Description

Optical fiber sensing microfluidic chip propofol online derivatization detection system and method

technical field

The invention relates to the technical field of rapid drug detection, in particular to an optical fiber sensing microfluidic chip propofol online derivatization detection system and method.

Background technique

Real-time detection of anesthetic drugs is a long-term clinical need. Overdose of anesthesia can cause circulatory and respiratory depression, while insufficient drugs can cause patients to perceive pain during surgery. At present, the control and adjustment of drug doses mainly rely on clinical experience and the monitoring of patient signs, and the drug metabolism varies greatly from individual to individual. Timely and individualized feedback of drug in vivo behavior, in vivo drug concentration is the most important auxiliary information. Due to the limitations of the rigid structure and other characteristics of the analytical instrument, the current methods for analyzing drug concentration in vivo are the analysis and detection of offline sampling, and the process is complicated and time-consuming, which is not conducive to obtaining the analysis results in time. The control lacks a better objective evaluation.

New technologies and new methods for the instant detection of drug components in vivo are the hotspots of research at home and abroad. At present, there is still a lack of feasible instant detection methods for anesthetic drugs. An optical fiber sensing microfluidic chip for online derivatization detection of propofol is proposed. system and method.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide the detection system and method of the most widely used modern intravenous anesthetic propofol at home and abroad, through the combination of microfluidic chip technology, optical fiber chemical sensing online process monitoring technology, and online derivatization technology , build a miniaturized, online, on-site measurement analysis system, establish a new method for real-time online rapid detection of anesthetic drug concentration, and evaluate the in vivo behavior of the detection drug, thereby improving the safety and effectiveness of clinical anesthetic drugs, and at the same time for faster research. , Real-time, online high-sensitivity drug analysis new technologies and new methods open up new ideas.

In order to achieve the above purpose, the present invention provides the following technical solutions: an optical fiber sensing microfluidic chip propofol online derivatization detection system, including a syringe pump, the outlet end of the syringe pump is connected to the inlet end of the microfluidic mixing chip , the outlet end of the microfluidic mixing chip is connected with the liquid inlet of the Z-type flow cell, the liquid outlet of the Z-type flow cell is connected with the waste liquid container through a connecting pipe, and the Z-type flow cell is connected with the waste liquid container. The pool is respectively provided with an optical path entrance and an optical path exit, the optical path entrance is connected with the light source through the input optical fiber, the optical path exit is connected with the optical fiber spectrometer through the output optical fiber, the optical fiber spectrometer is electrically connected with the computer, and the computer is installed with Spectral acquisition and processing software.

Preferably, the Z-shaped flow cell is provided with a Z-shaped pipeline, and the two ports of the Z-shaped pipeline are the liquid inlet and the liquid outlet of the Z-shaped flow cell, respectively. are the optical path entrance and the optical path exit respectively.

Preferably, the optical fiber sensing microfluidic chip propofol online derivatization detection system, its working process is as follows: the derivatization reagent and the propofol sample reagent enter the microfluidic mixing chip through the syringe pump for online derivatization After the derivatization product is generated, the Z-type flow cell is used for real-time detection of ultraviolet-visible spectrum, and the optical fiber sensor is used to transmit the optical signal, and the light of the light source is introduced into the Z-type flow cell through the input fiber. The output optical fiber is introduced into the optical fiber spectrometer for spectral detection, the optical fiber spectrometer transmits the signal to the computer, and the spectrum acquisition and processing software obtains the real-time dynamic UV-visible spectrum of the derivatization reaction.

Preferably, the syringe pump is an independently controllable multi-channel syringe pump, which can provide multiple channels to accurately control the flow rate and flow of the detected sample and derivatized reagent. The multi-channel syringe pump can be a TS-1B syringe pump, TS-1B syringe pump is a controller of a four-channel multi-function syringe pump, which can set different working parameters for the four channels respectively. It can be installed with standard syringes of various specifications from 5μL to 60mL. The range of linear speed (7.9μm/min-79.4mm/min) can meet the needs of different experiments.

Preferably, the chip of the microfluidic mixing chip is formed by bonding upper and lower substrates, wherein the material of the upper substrate is polydimethylsiloxane, the material of the lower substrate is glass, and the microchannels of the microfluidic mixing chip are The depth and width are both 50-500μm. The mixing chip consists of three inlets and one outlet. The mixing pipe adopts an S-shaped pipe design. The mixing chip fully mixes the sample test solution and derivatization reagent in the microchannel to generate sample derivatization. product, realize online derivatization of samples.

Preferably, the light source is a deuterium-tungsten halogen combined light source, which can provide stable and continuous spectral output in the ultraviolet-visible spectral region, and the deuterium-tungsten halogen combined light source can be a DH-2000-BAL deuterium-tungsten halogen combined light source, Produces stable and continuous spectral output at 215nm-2000nm.

Preferably, the optical fiber spectrometer uses a back-illuminated CCD detector to measure the absorbance change of the derivatized product of the sample in real time, and the back-illuminated CCD detector can use a Maya 2000PRO optical fiber spectrometer to detect the absorbance in the wavelength range of 200nm-900nm.

Preferably, both the input optical fiber (8) and the output optical fiber (10) use ultraviolet-visible optical fiber for transmission, and both the input optical fiber and the output optical fiber can use QP400-2-UV-VIS optical fiber, and the optical fiber length is 2m , with a diameter of 400 μm, suitable for UV and visible light detection.

Preferably, the spectral acquisition and processing software collects and records the spectral signals of the sample derivatized products generated online on the microfluidic chip in real time to form a visualized real-time dynamic change signal curve, and the spectral acquisition and processing software can use SpectraSuite spectrum The workstation is used to collect the real-time spectrum of the sample measurement.

An optical fiber sensing microfluidic chip propofol on-line derivatization detection method, the method includes the following steps: adopting Gibbs' derivatization method, detecting propofol using Gibbs' reaction derivatization method, and modifying the structure of the derivatization reagent to make The detection wavelength of the derivatized product is red-shifted to the visible light region, eliminating the interference of endogenous substances and exogenous substances in the blood, thereby improving the sensitivity and selectivity of the UV-Vis spectral detection of propofol.

Gibbs' reaction principle: Gibbs' reagent generates quinoneimine under alkaline conditions provided by tetramethyl ammonium hydroxide solution, and quinoneimine and the active hydrogen at the para position of the phenolic hydroxyl group of propofol synthesize a blue compound. The reaction can be completed instantaneously at room temperature, which is suitable for the requirement of rapid online derivatization and detection of propofol.

The reaction equation is as follows:

The concentration of Gibbs' reagent was 0.15 mg/mL, the concentration of tetramethylammonium hydroxide solution was 0.05%, and the injection flow rate of both reagent and sample was 75 μL/min. After 3 min of derivatization reaction, a stable derivatized product was obtained. The linear range for the detection of propofol biological samples was 1.0-18.0 μg/mL.

Compared with the prior art, the beneficial effects of the present invention are: optical fiber sensing microfluidic chip propofol online derivatization detection system and method: it has the following advantages:

1. Using a microfluidic mixing chip for online derivatization of samples and derivatization reagents, the flow rate and flow of the reagents can be precisely controlled, so that the detection can be completed with a small amount of samples and reagents.

2. The optical fiber sensing technology is used for online detection, and the optical signal is transmitted through the optical fiber, which simplifies the structure of the instrument, reduces the size of the instrument, and makes the entire analysis system miniaturized and portable, which is suitable for the needs of rapid detection on site.

3. Use online derivatization technology for rapid detection of biological samples, improve detection sensitivity and accuracy through derivatization, and solve the problem that spectral detection is easily interfered by other impurities in the sample.

4. The optical fiber sensing microfluidic chip on-line derivatization detection system established by the present invention can be used in the rapid detection of ultraviolet-visible spectrum of food, medicine, clinical biological samples, toxic and harmful components in the environment and other substances, and has a wide range of applications. application prospects.

Description of drawings

Fig. 1 is the detection standard curve of propofol derivatization in the embodiment of the present invention;

Fig. 2 is the ultraviolet-visible spectrogram of the propofol reference substance and the biological sample after derivatization in the embodiment of the present invention, wherein 1 is a blank plasma sample, 2 is the plasma added with the propofol standard solution, and 3 is the propofol reference substance ;

FIG. 3 is a schematic structural diagram of an on-line derivatization detection system for propofol on an optical fiber sensing microfluidic chip according to an embodiment of the present invention.

In the figure: 1-syringe pump, 2-microfluidic mixing chip, 3-Z-type flow cell, 4-connecting tube, 5-waste container, 6-optical path inlet, 7-optical path outlet, 8-input fiber, 9 - Light source, 10- output fiber, 11- fiber spectrometer, 12- computer.

Detailed ways

The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

Example: see Figures 1-3,

Step 1: Pretreatment of Propofol in Biological Samples:

Precisely aspirate 0.5 mL of the propofol-containing plasma sample, dilute the sample with the same volume of 4% phosphoric acid, activate the HLB solid-phase extraction cartridge with 1 mL of methanol, equilibrate with 1 mL of water, and then add 1 mL of phosphoric acid-acidified plasma sample to the solid-phase extraction column. Phase extraction cartridge, rinsed once with 1 mL of water, rinsed twice with 1 mL of acetonitrile-water (1:9) containing 1% KHCO ₃ , and finally eluted twice with 0.5 mL of methanol, and collected the eluates together.

Step 2: Examination of the linear range:

Precisely measure 5 parts of 490 μL propofol blank plasma, and add 10 μL of propofol series concentration standard solution respectively to make the plasma concentration of 1.0, 3.0, 6.0, 9.0, 12.0, 15.0, 18.0 μg/mL. Use the processing method of step 1 for sample pretreatment, and then conduct online derivatization and detection through the optical fiber sensing microfluidic chip propofol online derivatization detection system, and measure the absorbance of the sample. The drug plasma concentration (μg/mL) is the abscissa X, the absorbance is the ordinate Y, and the linear regression is performed. The standard curve equation is Y=0.0283X+0.0812, r=0.9960, indicating that the linearity is within the concentration range of 1.0～18.0μg/mL good relationship.

Propofol-containing plasma concentration and absorbance

Step 3: Recovery and Precision

Precisely measure 9 parts of 490 μL of propofol blank plasma, add 10 μL of standard solution of propofol series concentration respectively to make the blood concentration of 5.0, 10.0, 15.0 μg/mL, prepare 3 copies of each concentration sample in parallel, and use step 1 to process The recovery rate of the samples was determined after sample pretreatment.

Recovery and Precision Measurement Results (n=3)

Step 4: Simulate Biological Sample Assay

Precisely measure 9 copies of 490 μL of propofol blank plasma, add propofol injection emulsion respectively, and calculate according to the labeled amount to make the plasma concentration of 4.0, 8.0, and 12.0 μg/mL, and prepare 3 copies of each concentration in parallel, using step 1 The processing method is to conduct sample pretreatment, and then conduct online derivatization and detection through the optical fiber sensing microfluidic chip propofol online derivatization detection system, and calculate the propofol concentration according to the standard curve equation.

Simulation results of biological samples (n=3)

Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, and substitutions can be made in these embodiments without departing from the principle and spirit of the invention and modifications, the scope of the present invention is defined by the appended claims and their equivalents.

Claims (10)

1. The optical fiber sensing microfluidic chip propofol online derivatization detection system is characterized in that: comprising a syringe pump (1), the outlet end of the syringe pump (1) and the inlet of the microfluidic mixing chip (2) The ends are connected, the outlet end of the microfluidic mixing chip (2) is connected with the liquid inlet of the Z-type flow cell (3), and the liquid outlet of the Z-type flow cell (3) is connected by a connecting pipe (4) Connected to the waste liquid container (5), the Z-shaped flow cell (3) is respectively provided with an optical path inlet (6) and an optical path outlet (7), and the optical path inlet (6) passes through the input optical fiber (8). ) is connected with the light source (9), the optical path outlet (7) is connected with the optical fiber spectrometer (11) through the output optical fiber (10), the optical fiber spectrometer (11) is electrically connected with the computer (12), and the computer ( 12) Spectral acquisition and processing software is installed inside. 2. The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 1, characterized in that: the Z-shaped flow cell (3) is provided with a Z-shaped pipeline, and the Z-shaped pipeline The two ports of the Z-shaped flow cell (3) are the liquid inlet and the liquid outlet respectively, and the bends of the Z-shaped pipe are respectively the optical path inlet (6) and the optical path outlet (7). ). 3. The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 1 or 2, wherein the working process is: the derivatization reagent and the propofol sample reagent pass through the syringe pump (1) Entering into the microfluidic mixing chip (2) to carry out on-line derivatization reaction, after generating derivatized products, the Z-shaped flow cell (3) is used for real-time detection of ultraviolet-visible spectrum, and optical fiber sensors are used to transmit optical signals , the light of the light source (9) is introduced into the Z-type flow cell (3) through the input optical fiber (8), and into the optical fiber spectrometer (11) through the output optical fiber (10) for spectral detection. The optical fiber spectrometer (11) transmits the signal to the computer (12), and the spectrum acquisition and processing software obtains the real-time dynamic change ultraviolet-visible spectrogram of the derivatization reaction. 4 . The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3 , wherein the syringe pump ( 1 ) is an independently controllable multi-channel syringe pump. 5 . 5. The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3, wherein the chip of the microfluidic hybrid chip (2) is formed by bonding the upper and lower substrates, wherein The material of the upper substrate is polydimethylsiloxane, the material of the lower substrate is glass, the microchannel depth and width of the microfluidic mixing chip (2) are both 50-500 μm, and the mixing chip consists of three inlets and one The outlet is composed, and the mixing pipeline adopts S-shaped pipeline. 6. The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3, wherein the light source (9) is a deuterium-tungsten halogen combined light source. 7 . The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3 , wherein the optical fiber spectrometer ( 11 ) adopts a back-illuminated CCD detector. 8 . 8. The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3, characterized in that: both the input optical fiber (8) and the output optical fiber (10) use ultraviolet-visible optical fibers transmission. 9 . The optical fiber sensing microfluidic chip propofol online derivatization detection system according to claim 3 , wherein the spectrum acquisition and processing software is to the spectrum of the sample derivatized product generated online on the microfluidic chip. 10 . The signal is collected and recorded in real time to form a visualized real-time dynamically changing signal curve. 10. An optical fiber sensing microfluidic chip on-line derivatization detection method for propofol, characterized in that: the method comprises the following steps: using Gibbs' derivatization method, the Gibbs' reagent concentration is 0.15 mg/mL, tetramethyl hydroxide The concentration of the amine solution is 0.05%, the injection flow rate of the reagent and the sample is 75 μL/min, and the stable derivatized product is obtained after the derivatization reaction for 3 minutes, which is detected at 615 nm, and the linear range of the detection of propofol biological samples is 1.0～ 18.0 μg/mL.

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