CN105223182A - A kind of method detecting doped chemical drugs in Chinese patent drug - Google Patents

Abstract

The invention provides a method for detecting doped chemicals in Chinese patent medicines, which adopts the combined method of surface-enhanced Raman spectroscopy and thin-layer chromatography for detection, and performs detection according to the ratio shift value of thin-layer spots and dynamic surface-enhanced Raman spectroscopy. Determine the composition of adulterated chemicals, including three steps: preparation of standard solution and drug solution to be tested, thin plate chromatography, and collection of Raman spectra. In the third step, the surface enhancer uses nano-metal sol with glycerin as the solvent. Compared with the prior art, water is selected as the solvent. Glycerin has high viscosity and low volatility. When it is mixed with metal nanoparticles, it can Wrapped around the metal nanoparticles for a longer period of time, reducing the chance of metal nanoparticles being oxidized and ineffective due to contact with the outside world, reducing the cutting speed of metal nanoparticles, thereby improving the stability and detection sensitivity of Raman signals, and achieving low Better on-site rapid inspection of adulterated Chinese patent medicines.

Description

A method for detecting doped chemicals in Chinese patent medicines

technical field

The invention belongs to the field of drug detection, and in particular relates to a method for detecting doped chemicals in Chinese patent medicines by using a metal nano-sol with a glycerol solution as a solvent and a surface-enhanced Raman spectrum detection method thereof.

Background technique

The phenomenon of doping chemicals in Chinese patent medicines has a long history and is becoming more and more serious. Since the type, quantity, toxicity and other information of the chemical drugs mixed in are not known, the safety of the drug cannot be guaranteed at all, seriously endangering people's health, and disrupting the legitimate and orderly competition in the drug market, which will eventually affect the traditional Chinese medicine industry of sustainable development.

Surface-enhanced Raman spectroscopy is an emerging rapid detection technology in recent years, which has the advantages of high sensitivity, strong characteristics, and short detection time. At present, its technology combined with thin-layer chromatography has been applied to the separation and detection of some complex systems, such as pollutants in the water environment, metabolites in biological urine samples, and chemical drugs illegally added to Chinese patent medicines.

Generally speaking, in order to make Chinese patent medicines quickly achieve obvious therapeutic effects, most illegal pharmaceutical factories often illegally add chemical drugs to them with high content (greater than 1%). These Chinese patent medicines can usually be detected through simple extraction and separation to obtain strong surface-enhanced Raman spectra of illegally added drugs, so it is easy to quickly detect them with a portable Raman spectrometer to determine whether they are adulterated.

However, with the continuous improvement of adulteration technology, some manufacturers try to evade the detection of drug supervision departments by reducing the content of adulterated chemicals and increasing the types of adulterated chemicals. In the prior art, metal nano-sols with water as a solvent are often used as surface enhancers, but when the surface enhancer is dropped on a thin plate, the water in it will evaporate quickly, causing the metal particles wrapped in it to lose their protective layer and quickly lose their protective layer. It is oxidized, so that the surface-enhanced Raman scattering (SERS) signal for detecting the analyte on the thin-layer plate can only last for more than ten seconds, and it is difficult to obtain a lasting SERS signal, which is not conducive to actual detection. For example, when the content of doping chemicals is less than 1%, using metal nano-sol with water as the solvent as the surface enhancer can only collect a few dynamic Raman spectra, the test error is large, and the results are not convincing , even detection is not possible. Therefore, it is often difficult to detect adulterated drugs directly with the previous surface-enhanced Raman spectroscopy.

Contents of the invention

The present invention is carried out in order to solve the above-mentioned problems. It adopts the technology of combining surface-enhanced Raman spectroscopy and thin-layer chromatography to detect doping chemicals in Chinese patent medicines, and replaces the commonly used nano-metal sol with glycerol solution as a solvent. The nano-metal sol as the solvent further improves the sensitivity and stability of the SERS signal. Adopted the following technical solutions:

The invention provides a method for detecting doped chemicals in Chinese patent medicines, which uses surface-enhanced Raman spectroscopy and thin-layer chromatography for detection, according to the ratio shift value of thin-layer spots and dynamic surface-enhanced Raman spectroscopy. Determining the composition of adulterated chemicals includes the following steps:

Step 1, preparation of standard solution and drug solution to be tested: After a certain amount of standard product is dissolved in a solvent, and ultrasonically treated for 10-30 minutes, a standard solution of a certain concentration is obtained; the drug to be tested is ground into powder, After dissolving in the solvent and ultrasonic treatment for 10-30 minutes, centrifuge to take the supernatant to obtain the drug solution to be tested. The solvent used is any one of methanol, ethanol and water. However, in the subsequent steps, compared with water and ethanol, methanol has the best solubility and is the easiest to evaporate to dryness, so that the spot spread is the smallest, so methanol is preferred.

Step 2, chromatography: drop the standard solution and the drug solution to be tested respectively on the thin-layer plate obtained in step 1, after unfolding and drying, use an optical method to compare the ratio shift value of the standard product and the drug to be tested with the standard product The equal points are positioned, and the positioned position is recorded as the point to be tested;

Step 3, collect Raman spectrum: add the surface enhancer dropwise to the site to be tested in step 2, and use a portable Raman spectrometer to dynamically detect the site to be tested for 60s to 240s, and obtain the standard product and the site to be tested respectively Dynamic surface-enhanced Raman spectra of pharmaceuticals. The power of the Raman spectrometer is 150mW-300mW, and the integration time is 5s-10s.

Further, the method for detecting doped chemicals in Chinese patent medicines provided by the present invention also has the following characteristics: the surface enhancer used is nano-metal sol with glycerin solution as the solvent, and the nano-metal sol is preferably nano-silver sol, The volume ratio of the glycerol solution to the nano-silver sol solution is 1:1-3, preferably 1:1 (hereinafter referred to as the glycerin-silver colloid solution).

Further, the method for detecting adulterated chemicals in Chinese patent medicines provided by the present invention also has the following characteristics: the volume fraction of glycerol in the glycerol solution is 10% to 70%, preferably 30% to 70%, most preferably 50% %.

Further, the method for detecting doped chemicals in Chinese patent medicines provided by the present invention also has the following characteristics: the thin-layer plate used in step 2 is a silica gel thin-layer plate; when performing thin-plate chromatography, the sample solution to be tested The dropping volumes of the standard solution and the standard solution were both 1 μL; after the chromatography was completed, the spots were positioned using ultraviolet light with a wavelength of 254 nm.

Furthermore, the method for detecting doped chemicals in Chinese patent medicines provided by the present invention also has the following characteristics: in step 3, the amount of the surface enhancer added is 3 μL to 5 μL, and the excitation of the portable Raman spectrometer used The wavelength is 785nm.

Invention function and effect

The invention provides a method for detecting doped chemicals in Chinese patent medicines, which adopts surface-enhanced Raman spectroscopy combined with thin-layer chromatography for detection, and uses glycerol as a solvent nanometer metal sol for the surface enhancer. Compared with the prior art, choosing water as a solvent, glycerol has a higher viscosity, which makes it less volatile. When glycerin and metal nanoparticles are mixed, they can be wrapped around the metal nanoparticles for a longer period of time, reducing metal nanoparticles and metal nanoparticles. The chance of being oxidized and invalidated by external contact reduces the reduction speed of metal nanoparticles. On the one hand, the stability of the SERS signal is improved, and the signal can be maintained for hundreds of seconds, enabling better on-site rapid detection of low-content adulterated Chinese patent medicines; on the other hand, the detection sensitivity is improved. For example, when using water as the When the metal nanoparticles of the solvent are used as the surface enhancer, the lower limit of the mass fraction of the doping chemicals in the Chinese patent medicine detected by this method is 0.1%, and when the metal nanoparticles with glycerin as the solvent are used as the surface enhancer, the method The lower limit for detecting the mass fraction of doping chemicals in Chinese patent medicines is 0.01%, and the sensitivity is improved by an order of magnitude.

Description of drawings

Fig. 1 is the flowchart of the method for doping chemicals in detection Chinese patent medicine of the present invention;

Fig. 2 (a) is the dynamic surface enhanced Raman spectrogram of the standard product in the embodiment of the present invention;

Fig. 2 (b) is the dynamic surface-enhanced Raman spectrogram of the doping chemicals in the first embodiment of the present invention;

Fig. 3 is a dynamic surface-enhanced Raman spectrum comparison diagram of doping chemicals and their standard products in Example 1 of the present invention;

Fig. 4 (a) is the dynamic surface-enhanced Raman spectrogram of the standard product in the second embodiment of the present invention;

Fig. 4 (b) is the dynamic surface-enhanced Raman spectrogram of the doping chemicals in the second embodiment of the present invention;

Fig. 5 is a dynamic surface-enhanced Raman spectrum comparison diagram of doping chemicals and their standard products in Example 2 of the present invention;

Fig. 6 (a) is the dynamic surface enhanced Raman spectrogram of the standard product in the embodiment three of the present invention;

Fig. 6 (b) is the dynamic surface-enhanced Raman spectrogram of the doping chemicals in the third embodiment of the present invention;

Fig. 7 is a comparison diagram of the dynamic surface-enhanced Raman spectra of the doping chemicals and their standard products in Example 3 of the present invention.

detailed description

The specific implementation manners of the present invention will be described below in conjunction with the accompanying drawings.

Embodiment one

In this embodiment, the method of the present invention is described in detail by taking the detection of diphenhydramine hydrochloride, which is often mixed in Chinese patent medicines for relieving cough and asthma, as an example.

1. Instruments and samples

Instrument: portable Raman spectrometer (i-Raman; B&WTek; USA), excitation wavelength 785nm.

Preparation of silver colloidal solution: Weigh 90 mg of silver nitrate, dissolve it in 500 mL of deionized water and heat to boiling, add 10 ml of 1% sodium citrate, keep boiling for 1 hour, cool to obtain silver colloidal solution.

Preparation of standard solution doped with chemicals: Accurately weigh 1.0 mg of standard product, dissolve the standard product in 1 mL of methanol, and sonicate for 10-30 minutes to obtain a 1 mg/mL standard product solution.

Preparation of the drug solution to be tested: Take a Chinese patent medicine tablet, grind the tablet into a fine powder, dissolve it in an appropriate amount of analytical pure methanol, and ultrasonically treat it for 10-30 minutes, then centrifuge to take the supernatant to obtain the drug solution to be tested .

Wherein, the time of ultrasonic treatment is preferably 20min.

The preparation methods of the above instruments and samples are also applicable to the following examples.

Fig. 1 is a flow chart of the method for detecting adulterated chemicals in Chinese patent medicines in this embodiment.

As shown in Figure 1, the method for detecting diphenhydramine hydrochloride in the Chinese patent medicine for relieving cough and asthma in the present embodiment is as follows:

Step 1, preparing glycerin silver colloid solution: take 1 mL of glycerol solution with a volume fraction of 50% glycerol and mix with 1 mL silver colloid solution to obtain glycerin silver colloid;

Step 2, prepare the standard solution and the drug solution to be tested: prepare according to the above method;

Step 3, thin-plate chromatography separation: absorb 1 μL each of the drug solution to be tested and the standard solution with a capillary tube, drop them on the thin-layer plate, and dry at room temperature or heating temperature after unfolding;

Step 4, use an ultraviolet lamp with a wavelength of 254nm for spot location, determine whether there are spots in the position corresponding to the standard product in the drug to be tested, if there are, record the two spots as the spots to be tested and enter step 5, if not exists, re-select the standard and go to step 2;

Step 5, after dripping 3 μL glycerol silver colloid solution on the site to be tested, use a portable Raman spectrometer to perform dynamic detection. The laser power is 200mw, the integration time is 5s, and the dynamic acquisition time is 240s. Mann spectrogram, as shown in Figure 2;

Step 6: Select a Raman spectrum with the highest peak value for comparison, and then compare the two Raman spectra through spectral segment selection (550cm ^-1 -1600cm ^-1 ), baseline correction (airPLS method), and smoothing (Sgolay method) After preprocessing, compare the difference between the two spectra, as shown in Figure 3. At the same time, combined with the ratio shift value of the site to be tested on the thin-layer plate, it was determined whether such a standard was added to the Chinese patent medicine.

Figure 2(a) is the dynamic surface-enhanced Raman spectrum of the standard product in this example.

Figure 2(b) is a dynamic surface-enhanced Raman spectrum of the doping chemicals in this example.

As shown in Figure 2, in the dynamic surface-enhanced Raman spectrum of the standard product, the Raman spectrum peak intensity is the highest at the 30th second; in the dynamic surface-enhanced Raman spectrum of the drug to be tested, the Raman spectrum peak intensity at the 5th second Highest.

FIG. 3 is a comparison diagram of the dynamic surface-enhanced Raman spectra of the doping chemicals and their standard products in the first embodiment.

As shown in Figure 3, the peak patterns of the Raman spectra of the two groups with the strongest spectral peaks in the standard substance and the drug to be tested are exactly the same, so it can be determined that diphenhydramine hydrochloride is added to the Chinese patent medicine.

Embodiment two:

This embodiment is used for the detection of benproperine phosphate, which is often doped in Chinese patent medicines for relieving cough and asthma, and the detection steps are as follows:

Step 1, preparing glycerin silver colloid solution: take 1 mL of glycerol solution with a volume fraction of 50% glycerol and mix with 1 mL silver colloid solution to obtain glycerin silver colloid;

Step 2, prepare the standard solution and the drug solution to be tested: prepare according to the method in Example 1;

Step 3, thin-plate chromatography separation: absorb 1 μL each of the drug solution to be tested and the standard solution with a capillary tube, drop them on the thin-layer plate, and dry at room temperature or heating temperature after unfolding;

Step 4, use an ultraviolet lamp with a wavelength of 254nm for spot location, determine whether there are spots in the position corresponding to the standard product in the drug to be tested, if there are, record the two spots as the spots to be tested and enter step 5, if not exists, re-select the standard and go to step 2;

Step 5, after dripping 4 μL glycerol silver colloid solution on the site to be tested, use a portable Raman spectrometer to perform dynamic detection. The laser power is 150mw, the integration time is 5s, and the dynamic acquisition time is 240s. Mann spectrogram, as shown in Figure 4;

Step 6: Select a Raman spectrum with the highest peak value for comparison, and then compare the two Raman spectra through spectral segment selection (550cm ^-1 -1600cm ^-1 ), baseline correction (airPLS method), and smoothing (Sgolay method) After preprocessing, compare the difference between the two spectra, as shown in Figure 5. At the same time, combined with the ratio shift value of the site to be tested on the thin-layer plate, it was determined whether such a standard was added to the Chinese patent medicine.

Fig. 4 (a) is the dynamic surface-enhanced Raman spectrum of the benproperine phosphate standard substance in this example.

Figure 4(b) is a dynamic surface-enhanced Raman spectrum of the doping chemicals in this example.

As shown in Figure 4, in the dynamic surface-enhanced Raman spectrum of the benproperine phosphate standard substance, the Raman spectrum peak intensity at the 55th s is the highest; in the dynamic surface-enhanced Raman spectrum of the drug to be tested, at the 30th s The peak intensity of the Raman spectrum is the highest.

FIG. 5 is a comparison diagram of the dynamic surface-enhanced Raman spectra of the doping chemicals and their standard products in the second embodiment.

As shown in Figure 5, the Raman spectra of the two groups with the strongest spectral peaks in the standard benproperine phosphate and the chemical doped in the drug to be tested have the same peak shape, so it can be determined that phosphoric acid is added to the Chinese patent medicine phenproperine.

Embodiment three:

This embodiment is used for the detection of chlorpheniramine maleate, which is often doped in Chinese patent medicines for relieving cough and asthma, and the detection steps are as follows:

Step 1, preparing glycerin silver colloid solution: take 1 mL of glycerol solution with a volume fraction of 50% glycerol and mix with 1 mL silver colloid solution to obtain glycerin silver colloid;

Step 2, prepare the standard solution and the drug solution to be tested: prepare according to the method in Example 1;

Step 3, thin-plate chromatography separation: absorb 1 μL each of the drug solution to be tested and the standard solution with a capillary tube, drop them on the thin-layer plate, and dry at room temperature or heating temperature after unfolding;

Step 4, use an ultraviolet lamp with a wavelength of 254nm for spot location, determine whether there are spots in the position corresponding to the standard product in the drug to be tested, if there are, record the two spots as the spots to be tested and enter step 5, if not exists, re-select the standard and go to step 2;

Step 5, after dripping 5 μL glycerol silver colloid solution on the site to be tested, use a portable Raman spectrometer to perform dynamic detection. The laser power is 300mw, the integration time is 5s, and the dynamic acquisition time is 240s. Mann spectrogram, as shown in Figure 6;

Step 6: Select a Raman spectrum with the highest peak value for comparison, and then compare the two Raman spectra through spectral segment selection (550cm ^-1 -1600cm ^-1 ), baseline correction (airPLS method), and smoothing (Sgolay method) After preprocessing, compare the difference between the two spectra, as shown in Figure 7. At the same time, combined with the ratio shift value of the site to be tested on the thin-layer plate, it is determined whether such chemicals are added to the Chinese patent medicine.

Fig. 6 (a) is the dynamic surface enhanced Raman spectrogram of the chlorpheniramine maleate standard substance in the present embodiment.

Figure 6(b) is a dynamic surface-enhanced Raman spectrum of the doping chemicals in this example.

As shown in Figure 6, in the dynamic surface-enhanced Raman spectrum of chlorpheniramine maleate standard substance, the Raman spectrum peak intensity at the 105th s is the highest; in the dynamic surface-enhanced Raman spectrum of the drug to be tested, the first The peak intensity of the Raman spectrum is the highest at 30s.

FIG. 7 is a comparison diagram of the dynamic surface-enhanced Raman spectra of the doping chemicals and their standard products in the third embodiment.

As shown in Figure 7, the Raman spectra of the two groups with the strongest spectrum peaks in the standard chlorpheniramine maleate and the chemical doped in the drug to be tested have the same peak shape, so it can be determined that the additive in the Chinese patent medicine There is chlorpheniramine maleate.

In the above three embodiments, only one standard product and a sample of the Chinese patent medicine to be tested are selected for control detection each time. In fact, a kind of Chinese patent medicine may be mixed with many kinds of chemicals. Before using the method of this embodiment to detect the ingredients mixed with chemicals in Chinese patent medicine, other methods can be used to roughly judge the possible doped ingredients. Chemical components, and then configure the standard solution of these components and the solution of the Chinese patent medicine to be tested, and spot them on the thin-layer plate at the same time, and then collect the dynamic Raman spectrum.

In the above three embodiments, in order to obtain better detection results, the dynamic detection time of the site to be detected by the portable Raman spectrometer is 240s, but in actual use, it can be flexibly adjusted according to the actual situation, but it is best not to It is lower than 60s, so as not to obtain too few spectra, it is difficult to select the spectrum with the highest intensity, resulting in a large error in the subsequent comparison.

In the above three embodiments, when a portable Raman spectrometer is used for dynamic detection, the integration time used is 5s. In fact, any value within the range of 5s to 10s can be used as the integration time, as long as it can meet the analysis requirements later.

Function and effect of embodiment

This example provides a method for detecting doped chemicals in Chinese patent medicines, using surface-enhanced Raman spectroscopy combined with thin-layer chromatography for detection, and using nano-metal sol with glycerol as a solvent for the surface enhancer. Compared with the prior art, choosing water as a solvent, glycerol has a higher viscosity, which makes it less volatile. When glycerin and metal nanoparticles are mixed, they can be wrapped around the metal nanoparticles for a longer period of time, reducing metal nanoparticles and metal nanoparticles. The chance of being oxidized and invalidated by external contact reduces the reduction speed of metal nanoparticles. On the one hand, the stability of the SERS signal is improved, and the signal can be maintained for hundreds of seconds, enabling better on-site rapid detection of low-content adulterated Chinese patent medicines; on the other hand, the detection sensitivity is improved. For example, when using water as the When the metal nanoparticles of the solvent are used as the surface enhancer, the lower limit of the mass fraction of the doping chemicals in the Chinese patent medicine detected by this method is 0.1%, and when the metal nanoparticles with glycerin as the solvent are used as the surface enhancer, the method The lower limit for detecting the mass fraction of doping chemicals in Chinese patent medicines is 0.01%, and the sensitivity is improved by an order of magnitude.

In the above embodiments, the method of the present invention is described by taking the detection of chemicals doped in Chinese patent medicines for relieving cough and asthma as an example. However, the method of the present invention is equally applicable to the detection of doped chemicals in other types of Chinese patent medicines. At this time, the method of the three steps of sample preparation, chromatography and dynamic Raman spectrum selection should be determined according to the actual situation of the drug to be tested. Make the appropriate changes.

Claims (10)

1. A method for detecting doped chemicals in Chinese patent medicines, which uses surface-enhanced Raman spectroscopy and thin-layer chromatography for detection, and determines the doped chemicals according to the ratio shift value of thin-layer spots and dynamic surface-enhanced Raman spectroscopy. The composition of the heterochemical medicine is characterized in that it comprises the following steps: Step 1, preparation of standard solution and drug solution to be tested After a certain amount of standard substance is dissolved in a solvent, and ultrasonically treated for 10-30 minutes, a certain concentration of standard substance solution is obtained. The drug to be tested is ground into powder in turn, dissolved by the solvent, and ultrasonically treated for 10 to 30 minutes, then centrifuged to take the supernatant to obtain the drug solution to be tested; Step two, thin plate chromatography The standard solution obtained in step 1 and the drug solution to be tested are dripped on the thin-layer plate respectively, and after being unfolded and dried, the standard product and the drug solution to be tested are compared with the standard solution using an optical method. Position the points with the same product ratio shift value, and record the positioning position as the point to be tested; Step 3, collect Raman spectrum Add the surface enhancer dropwise to the site to be tested in step 2, and use a portable Raman spectrometer to dynamically detect the site to be tested for 60s to 240s to obtain the standard product and the drug to be tested respectively The dynamic surface-enhanced Raman spectrum of the Wherein, the solvent is any one of methanol, ethanol and water, The surface enhancer is a nano-metal sol with a glycerin solution as a solvent, The power of the Raman spectrometer is 150mW-300mW, and the integration time is 5s-10s. 2. the method for doping chemicals in the detection Chinese patent medicine according to claim 1, is characterized in that: Wherein, the nano-metal sol is nano-silver sol, and the volume ratio of the glycerol solution to the nano-silver sol solution is 1:1-3. 3. the method for doping chemicals in the detection Chinese patent medicine according to claim 2, is characterized in that: Wherein, the volume ratio of the glycerol solution and the nano-silver sol solution is 1:1. 4. the method for doping chemicals in the detection Chinese patent medicine according to claim 2, is characterized in that: Wherein, the volume fraction of glycerol in the glycerin solution is 10%-70%. 5. the method for doping chemicals in the detection Chinese patent medicine according to claim 4, is characterized in that: Wherein, the volume fraction of glycerol in the glycerin solution is 30%-70%. 6. the method for doping chemicals in the detection Chinese patent medicine according to claim 4, is characterized in that: Wherein, the volume fraction of glycerol in the glycerol solution is 50%. 7. the method for doping chemicals in the detection Chinese patent medicine according to claim 1, is characterized in that: Wherein, in step 2, the thin-layer board is a silica gel thin-layer board. 8. the method for doping chemicals in the detection Chinese patent medicine according to claim 1, is characterized in that: Wherein, in step 2, the dripping amounts of the sample solution to be tested and the standard solution are both 1 μL, In Step 3, the dropwise addition amount of the surface enhancer is 3 μL-5 μL. 9. the method for doping chemicals in the detection Chinese patent medicine according to claim 1, is characterized in that: Wherein, in step 2, ultraviolet light is used for positioning, and the wavelength of said ultraviolet light is 254nm. 10. the method for doping chemicals in the detection Chinese patent medicine according to claim 1, is characterized in that: Wherein, in step 3, the excitation wavelength of the portable Raman spectrometer is 785nm.