Method for testing blood concentration of cyclopofol based on thermal desorption-photo ionization ion mobility spectrometry

Abstract

The invention discloses a method for testing blood concentration of cyclopofol based on thermal desorption-photoionization ion mobility spectrometry, which comprises the following steps: detecting the concentration of cyclopofol by using a thermal desorption sampler and a photoionization ion mobility spectrometer, and taking acetone or butanone as a photoionization dopant; the method comprises the following steps: selecting a combined solvent in a thermal desorption sample injector for online extraction and separation of cyclopofol in a to-be-detected sample, preparing a cyclopofol standard substance with gradient concentration, and carrying out qualitative and quantitative analysis on the cyclopofol standard substance in a negative ion high-pressure mode of a thermal desorption-photoionization ion mobility spectrometer; the method comprises the following steps: respectively and continuously accumulating and recording cyclopofol spectrum peak ion mobility spectrometry spectrograms acquired by different cyclopofol concentrations, and fitting a standard curve equation between a target sample concentration and a cyclopofol monomer spectrum peak double-peak signal peak area to realize high-sensitivity and accurate analysis of cyclopofol in blood. The method is simple, rapid and sensitive, automatic analysis of an analyzer can be realized, and the ion mobility spectrometry technology can be more accurately used for measuring the blood concentration of cyclopofol.

Description

Technical Field

[0001] The present invention relates to the technical field of separation, analysis and detection, and more particularly to a method for testing the blood concentration of puromycin based on thermal desorption-photoionization ion mobility spectrometry. Background Art

[0002] Ion mobility spectrometry (IMS) is a rapid separation and detection technology that emerged in the 1970s. Compared to traditional mass spectrometry and chromatography instruments, it boasts a simple structure, high sensitivity, rapid analysis, and reliable results. The IMS we are researching has been widely used in fields such as drug detection, environmental monitoring, food monitoring, and life sciences. Ion mobility spectrometry is a normal-pressure analytical chemistry method that detects and identifies different substances based on the mobility constants or mobilities of gas-phase ions in a weak electric field. It operates under ambient pressure and is particularly well-suited for trace detection of volatile organic compounds (VOCs). The core component of an IMS system is the ion transfer tube, which consists of an ionization region and a migration region, separated by an ion gate. After being heated and vaporized in the injector, the sample is carried into the ionization region of the ion transfer tube by a carrier gas. Under the influence of the ionization source, carrier gas molecules and sample molecules undergo a series of ionization and ion-molecule reactions, forming various product ions. Under the influence of the electric field, these product ions enter the migration region through the periodically opened ion gate. On the one hand, they gain energy from the electric field for directional drift; on the other hand, they lose energy through continuous collisions with neutral migrating gas molecules flowing in the opposite direction. Because these product ions have different masses, charges, collision cross-sections, and spatial configurations, their migration times and mobilities in the electric field vary. This causes different ions to arrive at the detector at different times and be separated. The calculated differences in mobility constants can be used to qualitatively distinguish ions and molecules of different components. Different sample concentrations correspond to different peak heights or peak areas, thus enabling quantitative analysis.

[0003] Wang Xin et al. invented a quantitative analysis method for ion mobility spectrometry (Patent No. ZL201510229629.8), which discloses an accurate quantitative analysis method for rapid sample detection using ion mobility spectrometry. This method, based on qualitative analysis using ion mobility spectrometry, records the entire thermal desorption process of the sample under test. The recorded curve is defined as the tracking trend line of the sample's thermal desorption. The invention proposes a quantitative analysis method for ion mobility spectrometry, which is a method for quantitatively studying single monomer peaks.

[0004] Wang Xin and others invented an online detector for propofol in blood and its application (patent number ZL201510736154.1). The invention mainly focuses on the detection device, and the invention content uses ion mobility spectrometry technology as the basic detection technology, combined with sample thermal desorption and headspace sampling technology. The blood sample on the sampling chip can be continuously injected and thermally desorbed, and then carried by the carrier gas into the detector for online detection and analysis. Continuous online detection of propofol in human blood during surgery is realized. The aforementioned invention is the basis of the work, and in recent years there have been new progress in hardware integration prototypes and quantitative analysis.

[0005] The chemical name of propofol is 2-[(1R)-1-cyclopropylethyl]-6-isopropylphenol, and its molecular formula is C 14 H 20 O1, molecular weight 204.31. my country's innovative Class I drug, propofol injection, was approved for marketing in December 2020. It is based on the established foreign drug propofol (approved by the FDA in 1989). It boasts significantly higher efficacy and improved safety than propofol. The approval of propofol for marketing will significantly enhance the competitiveness and market share of Chinese anesthetics in both domestic and international markets.

[0006] Propofol injection is a white or off-white, homogeneous, emulsified suspension formulated with soybean oil (for injection), medium-chain triglycerides, refined egg yolk lecithin, sodium oleate, glycerol (for injection), disodium edetate, sodium hydroxide, and water for injection. It is suitable for sedation and induction of general anesthesia during gastrointestinal endoscopy. Propofol is primarily distributed in the extracellular fraction of the blood (with a blood / plasma ratio between 0.5 and 0.6) and is highly bound to human plasma proteins (approximately 95% binding) at concentrations of 80-1200 ng / ml. Phase I clinical trials demonstrated that in healthy Chinese adult subjects, peak plasma concentrations were reached approximately 2 minutes after intravenous injection. Propofol plasma concentrations exhibit a three-phase elimination pattern, with half-lives (α, β, and γ) of 2.0 minutes, 34.9 minutes, and 6.2 hours, respectively. Propofol is primarily metabolized in the liver through oxidation, glucuronidation, and sulfate conjugation. Its metabolites are inactive and are primarily excreted via the kidneys (approximately 84.59%), with a small amount excreted in the feces (approximately 2.65%). Following a single dose, approximately 99% of propofol and its metabolites are excreted within one week. The pharmacokinetic characteristics of propofol are characterized by a short half-life and high clearance.

[0007] The blood concentration of propofol is a crucial indicator for anesthesiologists to monitor and adjust the depth of anesthesia during surgery. However, because propofol is a new drug currently undergoing clinical trials, no methods for measuring its blood concentration have been reported in domestic or international literature. Even if analytical methods such as separation, enrichment, liquid chromatography, and mass spectrometry were developed, the entire process would be time-consuming, hindering rapid perioperative blood concentration measurement. Summary of the Invention

[0008] The purpose of the present invention is to overcome the above-mentioned defects of the prior art and provide a method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry. The method is simple, rapid and sensitive, can realize automatic analysis by an analyzer, and more accurately realizes the use of ion mobility spectrometry technology for the determination of the blood concentration of propofol.

[0009] To achieve the above object, the technical solution of the present invention is as follows:

[0010] A method for testing the blood concentration of puromycin based on thermal desorption-photoionization ion mobility spectrometry is disclosed. The method uses a thermal desorption injector and a photoionization ion mobility spectrometer to detect the puromycin concentration. Acetone or butanone is used as a photoionization dopant. A combined solvent is selected in the thermal desorption injector to online extract and separate the puromycin in a sample to be tested to enhance the thermal desorption effect and shorten the analysis time. A puromycin blood sample standard with a concentration of 1-10 μg / ml is prepared. The puromycin standard is qualitatively analyzed in the negative ion high-pressure mode of the thermal desorption-photoionization ion mobility spectrometer. After the qualitative analysis, puromycin peak ion mobility spectra obtained at different puromycin concentrations are continuously accumulated and recorded. A standard curve equation is fitted between the target sample concentration and the peak area of the double-peak signal of the puromycin monomer peak. The peak area of the sample to be tested with an unknown concentration is substituted into the standard curve equation to obtain the puromycin concentration value in the sample to be tested.

[0011] Optionally, the blood sample includes whole blood, plasma or serum, and can be directly injected for analysis without complicated sample pre-treatment techniques.

[0012] Optionally, a single blood sample should be vortexed for 2-5 seconds or manually shaken before testing to maintain sample homogeneity.

[0013] Optionally, the combined solvent comprises alcohols and ethers in a ratio of (1-3):2.

[0014] Optionally, the alcohol is methanol; the ether is anisole.

[0015] Optionally, the thermal desorption temperature is 140-180°C.

[0016] Optionally, when the method uses a thermal desorption injector to inject samples, 2-20 μL of the propofol sample is dropped onto the surface of a blood collection card for thermal desorption injection.

[0017] Optionally, the ion mobility spectrum desorption time of the entire thermal desorption process starts from the data acquisition and ends within 1-3 minutes, and the software automatically records the data and outputs the analysis and detection results.

[0018] Optionally, the photoionization ion mobility spectrometer includes an ion mobility tube, which includes an ionization region and a drift region. A drift gas inlet 4 is provided at the tail end of the ion mobility tube. The ionization region is provided with a gas outlet 1, a dopant carrier gas inlet 2, and a sample carrier gas inlet 3 in sequence in a direction perpendicular to the central axis of the ion mobility tube. Both the carrier gas and the drift gas are clean air sources. The photoionization dopant enters the ionization region through dual channels of the dopant carrier gas inlet 2 and the sample carrier gas inlet 3 to enhance the ionization effect.

[0019] The specific steps are (taking propofol as an example):

[0020] Step 1: Set the test conditions of the medical blood drug concentration analyzer according to the standard method determined in the preliminary experiment. The test conditions of qualitative analysis, quantitative analysis and marking equation are consistent.

[0021] Step 2: First, a single drug standard or blank blood sample with a concentration range of 0-10 ng / μl is measured and dropped into a blood collection card under the set conditions. The peak migration time of the puromycin drug to be tested is determined by comparing the peak migration time of the puromycin detection signal in the blank blood with that in the blood, thereby completing the qualitative analysis.

[0022] Step 3: Record the continuously collected ion mobility spectra of the two monomer peaks of propofol (within 1-3 minutes), average the data 10-20 times, and record the entire spectrum of the entire drug desorption process;

[0023] Step 4: The fitted relationship between the peak area of the propofol blood sample monomer and the blood drug concentration of the sample is the standard curve equation generated by fitting.

[0024] Step 5: Measure the same volume of blood sample with unknown blood drug concentration, and analyze it using a medical blood drug concentration analyzer based on ion mobility spectrometry technology according to the same conditions and methods as above; based on the standard curve equation, the unknown blood drug concentration of propofol can be calculated, or a blood sample with an unknown blood drug concentration that is different in volume from the standard sample can be measured, and the multiple relationship is fitted to convert it into a volume-consistent analysis.

[0025] Step 6: The medical blood drug concentration analyzer automatically saves the blood drug concentration value of propofol and can output a printed report.

[0026] Optionally, the method for obtaining the two monomer peak signals of the cyclopentane photoionization product ion is: regulating the photoionization to produce CO3 - (H2O)n and CO4 - (H2O)n two reagent ions;

[0027] The two monomer peaks of propofol products are (MH) - and (M-O2) - ; Regulate photoionization to produce CO3 -(H2O)n and CO4 - The method of using two reagent ions (H2O)n is that the dopant dopant enters the ion transfer tube in two ways, one way directly enters the ion transfer tube, and the other way the dopant dopant is added to the sample carrier gas.

[0028] The successful development of this quantitative analysis method for puromycin blood concentration will undoubtedly guide the widespread application of ion mobility spectrometry in clinical drug analysis, enabling rapid and accurate quantitative analysis. This method achieves highly sensitive and precise quantitative analysis of puromycin blood concentration. The method has a simple algorithm, enables automatic fitting of data processing, shortens analysis time, and allows for simultaneous monitoring of puromycin blood concentration during surgery.

[0029] The implementation of the present invention will have the following beneficial effects:

[0030] 1. The present invention provides a highly sensitive detection method for online monitoring of the blood concentration of propofol during surgery, which is conducive to the promotion and application of the domestically produced new drug propofol in clinical practice.

[0031] 2. The method of adding two dopants into the gas path in the technical solution of the present invention enhances the ionization efficiency of the target by regulating the high concentration of dopants in the ionization zone, which is beneficial to improving the detection sensitivity of propofol.

[0032] 3. In combination with the quantitative analysis method established by the present invention, three reagents are added to the injection zone to enhance the thermal desorption of the target drug, which is beneficial to improve the detection sensitivity of propofol and shorten the analysis time.

[0033] 4. This measurement method is simple, fast, easy to correct, has high data accuracy, and has broad application prospects in clinical practice. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is the product spectrum of the ion mobility spectrometry detection of acetone air reagent ion peak of the present invention.

[0035] Figure 2 This is the product spectrum of two monomer peaks of propylparaben detected by ion mobility spectrometry of the present invention.

[0036] Figure 3 This is a standard curve diagram of different concentrations of puromycin detected by ion mobility spectrometry of the present invention.

[0037] Figure 4 Schematic diagram of the ion mobility spectrometer structure of the present invention.

[0038] Among them, 1. gas outlet, 2. dopant carrier gas inlet, 3. sample carrier gas inlet, 4. float gas inlet. DETAILED DESCRIPTION

[0039] The present invention will be further described below with reference to specific examples, but the present invention is not limited thereto in any way.

[0040] The ion mobility spectrometer described in the embodiment uses a VUV light ionization source and acetone as a dopant. The medical blood drug concentration analyzer primarily comprises a sample injection device, an ionization source, a reaction zone, an ion gate, a migration zone, a signal receiving and detection system, and a gas purification system. The sample injection device primarily comprises a thermal desorption injector, a blood collection card, and a carrier gas delivery pipeline.

[0041] Blood samples containing propofol were accurately prepared at concentrations of 1, 2.5, 5, and 10 ng / μl. Blank whole blood was provided by the hospital. During the experiment, the migration tube temperature was maintained at 120°C, the injector temperature at 160°C, and the carrier gas (purified air), float gas (purified air), and dopant gas (purified air) flows were 300 mL / min, 600 mL / min, and 100 mL / min, respectively. A 10 μl blood sample was injected, and a combination reagent of methanol and anisole was added in a 3:2 ratio, for a total volume of 3 μl. The standard curve equation was used. Three replicates were run for each concentration.

[0042] Example 1

[0043] like Figure 1 As shown in Figure 2, the medical blood drug concentration analyzer based on ion mobility spectrometry technology detects the acetone air reagent ion peak product. Acetone photoionization produces two reagent ion peaks at 3.0ms and 3.2ms, which are the photoionization products CO3 - (H2O)n and CO4 - (H2O)n. Under the action of two photoionization reagent ions, (MH) can be produced - and (M-O2) - Two monomeric peaks of propofol.

[0044] Example 2

[0045] According to the conditions set in the embodiment, 10 ng / μl of purocycline in whole blood was injected and analyzed. Figure 2 As shown, propylparaben (MH) - and (M-O2) - The two monomer peaks are located at 5.4ms and 5.7ms respectively. The software sets the data tracking range to 5.3-5.5ms and 5.6-5.8ms.

[0046] Example 3

[0047] When the experimental conditions and instrument parameters are the same as those in the above embodiment, the ion mobility spectrometry signal peak intensity change tracking curve of each monomer peak is continuously measured. The quantitative analysis equation of propofol is calculated based on the cumulative peak area sum of the two monomer peaks.

[0048] Example 4

[0049] When the experimental conditions and instrument parameters remain the same as those in the above implementation method, Figure 3 The experimental fitting equation of the standard curve of 1-10 ng / μl propofol blood concentration and the sum of the double monomer peak signal is Y=-2064+3110*X, and the correlation coefficient R 2 >0.9991.

[0050] Example 5

[0051] When the experimental conditions and instrument parameters were consistent with the above embodiment, a test was conducted on a blood sample containing an unknown concentration of propofol. The analyzer automatically detected a thermal desorption area of 20626 mV. The analyzer software automatically subtracted the background signal from the equation and calculated the drug concentration to be 5.0 ng / μl.

[0052] Based on the above results, the present invention controls acetone to form a double reagent ion and then controls puromycin to form a double product ion peak. High dopant concentration is then controlled to improve ionization efficiency, and the combined solvent accelerates the thermal desorption efficiency of puromycin. The puromycin blood concentration analyzer based on ion mobility spectrometry demonstrates stable and repeatable testing and analysis methods with good reliability, making it suitable for further clinical application.

[0053] The above-described embodiments merely represent several implementation methods of the present invention. While the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the patent application. It should be noted that a person skilled in the art would be able to make various modifications and improvements without departing from the spirit of the present invention, all of which fall within the scope of protection of the present invention. Therefore, the scope of protection of the patent for this invention shall be determined by the appended claims.

Claims

1. A method for measuring the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry, characterized in that: The method uses a thermal desorption injector and a photoionization ion mobility spectrometer to detect the blood concentration of puromycin, uses acetone or butanone as a photoionization dopant, selects a combined solvent in the thermal desorption injector to extract and separate puromycin in a sample to be tested online, prepares a puromycin standard with a concentration of 1-10 μg / ml, and uses the negative ion high-pressure mode of the thermal desorption-photoionization ion mobility spectrometer to perform qualitative analysis on the puromycin standard. After the qualitative analysis, ion mobility spectra of puromycin peaks obtained at different puromycin concentrations are continuously accumulated and recorded, a standard curve equation is fitted between the target sample concentration and the peak area of the double-peak signal of the puromycin monomer peak, and the peak area of the sample to be tested with an unknown concentration is substituted into the standard curve equation to obtain the puromycin concentration value in the sample to be tested.

2. The method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: Blood samples include whole blood, plasma or serum, and can be directly injected and analyzed without complicated sample pretreatment techniques.

3. The method for testing the blood concentration of puromycin based on thermal desorption-photoionization ion mobility spectrometry according to claim 2, characterized in that: Before testing a single blood sample, vortex for 2s-5s or shake it manually to maintain sample homogeneity.

4. The method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: The combined solvent includes alcohols and ethers in a ratio of (1-3):

2.

5. The method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry according to claim 4, characterized in that: The alcohol is methanol; the ether is anisole.

6. The method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: The thermal desorption temperature is 140-180°C.

7. The method for testing the blood concentration of propofol based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: When the method uses a thermal desorption injector to inject samples, 2-20 μL of the propofol sample is dropped onto the surface of a blood collection card for thermal desorption injection.

8. The method for testing the blood concentration of puromycin based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: The desorption time of the ion mobility spectrum of the entire thermal desorption process starts from data acquisition and ends within 1-3 minutes. The software automatically records the data and outputs the analysis and detection results.

9. The method for testing the blood concentration of puromycin based on thermal desorption-photoionization ion mobility spectrometry according to claim 1, characterized in that: The photoionization ion mobility spectrometer comprises an ion transfer tube, the ion transfer tube comprises an ionization region and a drift region, a drift gas inlet (4) is provided at the tail end of the ion transfer tube, and the ionization region is sequentially provided with a gas outlet (1), a dopant carrier gas inlet (2), and a sample carrier gas inlet (3) in a direction perpendicular to the central axis of the ion transfer tube; Both carrier gas and drift gas are clean air sources; The photoionized dopant enters the ionization region through the dual channels of the dopant carrier gas inlet (2) and the sample carrier gas inlet (3).

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