Method for simultaneously determining index components of 21 plant-derived raw materials in toothpaste

Abstract

Compared with the prior art, the method for simultaneously determining the index components of the 21 plant-derived raw materials in the toothpaste has the advantages that liquid chromatography conditions, mass spectrometry parameters and a sample pretreatment method are optimized, and 21 target compounds can be detected within 25 minutes. The method is simple and efficient in pretreatment, short in analysis time, high in sensitivity, strong in matrix interference resistance and accurate and reliable in qualitative and quantitative result, can be used for quality control of related plant source raw materials and evaluation of consistency of labels and components in toothpaste containing plant sources, and provides a foundation for further standardizing label identification of the toothpaste. And the technical support is provided for promoting the standardized and benign competition of toothpaste filing persons and production enterprises and the supervision after the toothpaste is filed and sold on the market.

Description

Technical Field

[0001] The present invention relates to the field of chemical analysis, and in particular to a method for simultaneously determining index components of 21 plant-derived raw materials in toothpaste. Background Art

[0002] With the continuous improvement of living standards, the types of toothpaste products have gradually diversified, and more and more Chinese-style plant extract raw materials are widely used in toothpaste formulas. From December 1, 2023, toothpaste should be registered with the drug regulatory authorities before being put on the market or imported, and it is clear that the toothpaste label should indicate all ingredients. At present, my country's regulations on risk assessment and monitoring of toothpaste products are just beginning. For the quality control of toothpaste containing plant-derived raw materials, on the one hand, there is a lack of current standards to detect whether it contains the raw materials indicated on the label, and on the other hand, there is no reasonable limit set for the relevant functional ingredients. As a result, the concept of plant-derived ingredients has been abused by some companies, and false propaganda about adding plant-derived raw materials, insufficient or no raw materials, and other adverse phenomena often occur. Such behavior not only harms the interests of consumers, but also poses potential risks to the health of the people.

[0003] The appendix to my country's current light industry standard, QB / T2966-2014, "Effective Toothpaste," specifies methods for determining the content of paeonol, Panax notoginseng (Panax notoginseng) saponins (ginsenoside Rg1, ginsenoside Rb1, and Panax notoginseng saponin R1), and the active ingredient of Sarcandra glabra (isofraxinidin) in toothpaste. These methods all utilize high-performance liquid chromatography-ultraviolet (HPLC-UV) detection. Because the amount of botanical ingredients added to toothpaste is generally low, and the content of their indicative ingredients is even lower, the HPLC-UV method struggles to meet the sensitivity requirements for detecting different botanical indicative ingredients in toothpaste products, potentially leading to false-negative results. Furthermore, due to the complex matrix of toothpaste products, the specificity of these methods is also limited, making them inadequate for routine regulatory requirements. Existing methods for detecting botanical ingredients in toothpaste reported in the literature primarily focus on high-performance liquid chromatography, ultra-high-performance liquid chromatography, and gas chromatography. These methods also suffer from low sensitivity, weak specificity, and poor resistance to matrix interference.

[0004] Shui Dian, Xia Zemin, et al. (Simultaneous determination of notoginsenosides and ginsenosides in toothpaste by ultra-performance liquid chromatography-tandem mass spectrometry [J]. Oral Care Products Industry, 2023, 33(5):7-11) used ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS / MS) to simultaneously determine notoginsenosides and ginsenosides in toothpaste. The results showed that some products were labeled as containing notoginseng extracts but no related ingredients were detected. Ye Rongzi, et al. (Determination of isoflurane in toothpaste by ultra-performance liquid chromatography-tandem mass spectrometry [J]. Guangdong Chemical Industry, 2022, 49(24):234-236) used UPLC-MS / MS to determine isoflurane in toothpaste. The results showed that some products were labeled as containing coral extracts but no isoflurane-related ingredients were detected. The above research provides effective technical support for the supervision and quality control of toothpaste products. However, the existing detection method only measures the indicator components of a single plant source, which cannot meet the purpose of rapid screening and quantitative detection of multiple plant sources and multiple indicator components in daily supervision. Summary of the Invention

[0005] The technical problem to be solved by the present invention is: in view of the current technical gap in the qualitative and quantitative quality control of multiple plant-based and multi-index ingredients in toothpaste, the present invention selects plant sources commonly seen on toothpaste labels on the market as research objects, mainly including coral reef, two-sided needle, honeysuckle, Panax notoginseng (Panax notoginseng), Scutellaria baicalensis, Coptis chinensis, Paeonia suffruticosa, Aloe vera, Magnolia officinalis, Scutellaria baicalensis, Tea, Taraxacum mongolicum, Coptis chinensis, Isatis indigotica (Isatis indigotica), etc. A total of 21 index ingredients are selected, and an exclusive, rapid and sensitive qualitative and quantitative detection method is established based on ultra-performance liquid chromatography-tandem mass spectrometry technology.

[0006] The technical solution of the present invention to solve the above technical problems is as follows:

[0007] A method for simultaneously determining 21 index components of plant-derived raw materials in toothpaste comprises the following steps:

[0008] (1) Preparation of reference solution;

[0009] (2) Preparation of test solution;

[0010] (3) Calculate the contents of 21 index components in the sample using the quantitative ion peak area according to the external standard method;

[0011] The method described was detected using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry;

[0012] The ultra-high performance liquid chromatography (UPLC) conditions are as follows: the chromatographic column is filled with octadecylsilane bonded silica gel, with an inner diameter of 2.1 mm and a particle size of 1.7 or 1.8 μm; 0.1% formic acid aqueous solution is used as mobile phase B, acetonitrile is used as mobile phase A, and gradient elution is performed; the injection volume is 2±1 μL; the flow rate is 0.3±0.1 mL / min; and the column temperature is 35±5°C.

[0013] The mass spectrometry conditions are as follows: ion source: electrospray ionization source (ESI), positive and negative ion simultaneous monitoring mode;

[0014] Ion source temperature: 500±50°C; spray voltage: 5500±500V in positive ion mode, 4500±500V in negative ion mode; curtain gas: 30±10psi; nebulizer gas pressure: 55±10psi; auxiliary gas pressure: 55±10psi; scan mode: multiple reaction monitoring (MRM) to detect qualitative and quantitative product ions;

[0015] The indicative components of the 21 plant-derived raw materials are: 1: aloin; 2: syringin; 3: corydaline; 4: nitine chloride; 5: berberine hydrochloride; 6: coptisine hydrochloride; 7: isofraquinone; 8: paeonol; 9: (R,S)-gaoichun; 10: ginsenoside Rb1; 11: notoginseng saponin R1; 12: ginsenoside Rg1; 13: longipenoside; 14: luteolin; 15: baicalin; 16: rosmarinic acid; 17: chlorogenic acid; 18: catechin; 19: magnolol; 20: magnolol; 21: caffeic acid.

[0016] Furthermore, a method for simultaneously determining 21 plant-derived index components in toothpaste comprises the following steps:

[0017] S1, using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry for detection;

[0018] S2. The ultra-high performance liquid chromatography-tandem mass spectrometry method includes parameters: using octadecylsilane bonded silica gel as a filler, with specifications of an inner diameter of 2.1 mm and a particle size of 1.7 or 1.8 μm; specifically, ACQUITY UPLC BEH C 18Chromatographic columns (2.1 mm × 100 mm, 1.7 μm) and Waters ACQUITY UPLC HSS T3 (2.1 mm × 100 mm, 1.8 μm) were used. Acetonitrile was used as mobile phase A; formic acid solution was used as mobile phase B, specifically 0.1% formic acid solution; gradient elution was performed, and the specific gradient elution program was as follows: 0-1 min, 10% A; 1-12 min, 10%-25% A; 12-16 min, 25%-50% A; 16-18 min, 50%-95% A; 18-23 min, 95% A; 23.01-25 min, 10% A; injection volume was 2±1 μL, specifically 2 μL; flow rate was 0.3±0.1 mL / min, specifically 0.3 m / min; column temperature was 35±5°C, specifically 35°C. Under this chromatographic condition, 21 components were detected in positive and negative ion modes. Figure 2-4 .

[0019] S3. A tandem quadrupole mass spectrometer with an electrospray ionization (ESI) source and simultaneous positive and negative ion monitoring mode was used. Source temperature: 500 ± 50°C; spray voltage: 5500 ± 500 V in positive ion mode, 4500 ± 500 V in negative ion mode; curtain gas: 30 ± 10 psi; nebulizer gas pressure: 55 ± 10 psi; auxiliary gas pressure: 55 ± 10 psi; scan mode: multiple reaction monitoring (MRM). The mass spectrometric parameters for the 21 compounds, including parent ions, product ions, declustering potential (DP), and collision energy (CE), are shown in the table below. Ions marked with an "*" are quantification product ions.

[0020]

[0021]

[0022] S4. Preparation of reference substance solution; the specific operation is: weigh appropriate amount of 21 reference substances respectively, add methanol to ultrasonically dissolve and dilute to make single-standard stock solution, accurately measure appropriate amount of single-standard stock solution, add 80% methanol to dilute to make reference substance working solutions of 21 components, specifically, each 1 mL can contain approximately 25 mg / L of reference substance solution, and before use, measure appropriate amount of reference substance working solution to prepare a series of standard curve solutions.

[0023] S5. Preparation of the test solution: Weigh 1 g of toothpaste sample (accurate to 0.0001 g) into a 15 mL stoppered centrifuge tube. Add 10 mL of 80% methanol solution and two glass beads. Vortex thoroughly to evenly disperse the toothpaste. Ultrasonic extraction for 25 minutes, cool to room temperature, and centrifuge at 3000 rpm for 10 minutes. Filter the supernatant through a 0.2 μm filter membrane and use the filtrate as the test solution. Dilute with 80% methanol solution to within the standard curve range, if necessary, based on the detection results, and then inject the sample for measurement.

[0024] S6. Calculate the contents of 21 index components in the sample using the quantitative ion pairing method and the peak area according to the external standard method. Specifically, calculate the contents of 21 index components in the sample using the quantitative ion peak area according to the external standard method.

[0025] This study selected common plant sources on toothpaste labels in the market as research objects, including Coral Grass, Zanthoxylum bungeanum, Honeysuckle, Panax notoginseng (Panax notoginseng), Scutellaria baicalensis, Coptis chinensis, Paeonia suffruticosa, Aloe, Magnolia officinalis, Scutellaria baicalensis, Tea, Taraxacum mongolicum, Coptis chinensis, Isatis indigotica (Isatis indigotica), etc. According to the Chinese Pharmacopoeia Volume 1 (2020 edition, Beijing: China Medical Science and Technology Press), Standard Methods (Ministry of Industry and Information Technology of the People's Republic of China. Light Industry Industry Standard of the People's Republic of China. Functional Toothpaste [S]. China Light Industry Press, QB / T 2966-2014) and references (Zhong Nan, Wei Jiayan, Zhang Guoyu, et al. Application of antibacterial and anti-inflammatory active ingredients of Chinese herbal medicine in functional toothpaste [J]. West China Journal of Pharmaceutical Sciences, 2025, 40(2): 222-226; Liu Zhenjie, Gao Hongwei, Sun Xueping, et al. Determination of the content of multiple index components of Jiubiying based on HPLC-MS [J]. Journal of Pharmaceutical Analysis, 2021, 41(11): 1868-1874), and selected the characteristic or functional components of each plant source as the index components. The 14 plant sources and their corresponding index components are shown in Table 1 below.

[0026] Table 1 14 plant sources and their index components

[0027]

[0028]

[0029] Regarding the test results, referring to the toothpaste label, since the amount of botanical ingredients added to toothpaste is relatively small, the detection of one or more of the labeled ingredients can be considered a preliminary determination that the corresponding botanical ingredient is added to the product. Currently, there are no corresponding standards for botanical ingredients in toothpaste, so this method is also suitable for the qualitative and quantitative detection of indicator components in toothpaste botanical ingredients.

[0030] The response intensities of 21 test components in positive and negative ionization modes were compared. The results showed that the responses of isoflurane, nitine chloride, berberine hydrochloride, coptisine hydrochloride, paeonol, aloin, syringin, corydaline, and (R,S)-coitrin in positive ion mode were significantly higher than those in negative ion mode, making them more suitable for positive ion detection mode. Berberine hydrochloride and coptisine hydrochloride obtained adduct ions [M-HCl+H] + , syringin is [M+Na] + The other 7 compounds all obtained quasi-molecular ions [M+H] + The responses of the 12 index components in negative ion mode were stronger than those in positive ion mode, and they were more suitable for negative ion mode detection. Among them, ginsenoside Rg1, ginsenoside Rb1, notoginsenoside R1, and longipesinol obtained adduct ions [M+HCOO] - , the other 8 analytes obtained quasi-molecular ions [MH] - After obtaining parent ions for the 21 compounds, fragment ions were obtained through daughter ion scanning. Based on the number of fragments, 2 to 4 daughter ions with higher responses were selected as characteristic fragment ions to further optimize the collision energy and declustering voltage. Finally, the two daughter ions with the highest response were selected as qualitative and quantitative ions.

[0031] Considering the target compounds' general solubility in methanol or high-proportion methanol-water solutions, and the fact that toothpaste products disperse well in water, the present invention selected a methanol-water system as the extraction solvent. The extraction effects of different methanol-water ratios (50%, 60%, 70%, 80%, 90%, and pure methanol) on the index components in toothpaste samples were investigated. Toothpaste products contain foaming agents (surfactants, etc.) and thickeners (such as xanthan gum) that are highly water-soluble. Excessive dissolution of these substances in the sample solution can lead to strong matrix effects in mass spectrometry, affecting the accuracy of mass spectrometry. Furthermore, when the methanol ratio is below 70%, the test solution is extremely difficult to filter through the membrane. The study found that when the methanol ratio exceeded 70%, the toothpaste sample extract gradually became clear, making it easier to filter through the membrane. However, when the methanol ratio exceeded 90%, the toothpaste sample tended to clump, making it difficult to disperse, resulting in incomplete extraction of some compounds. Taking into account the polarity of the 21 compounds, 80% methanol was ultimately selected as the extraction solvent. In order to improve the dispersion effect of toothpaste in the extraction solvent, two glass beads were added to assist in the dispersion of the sample.

[0032] Ultrasonic-assisted extraction can effectively shorten the extraction time and improve the extraction efficiency. Different ultrasonic extraction times were investigated in this study. 1g of toothpaste sample (containing Panax notoginseng root extract and Scutellaria baicalensis extract) was weighed and processed according to the preparation of the test solution. Ultrasonic treatment was performed for 5min, 10min, 15min, 20min, 25min and 30min respectively. The sample was cooled to room temperature and centrifuged at 3000rpm for 10min. The supernatant was filtered through a 0.2μm filter membrane and then sampled for determination. As the ultrasonic time increased, the response value of the index component increased. When the ultrasonic time exceeded 25min, the extraction efficiency reached a plateau and the response value of the target compound basically no longer increased. Therefore, 25min was selected as the ultrasonic time.

[0033] The 21 indicative components selected in this patent have significantly different polarities, the toothpaste sample matrix is relatively complex, and when positive and negative ions are scanned simultaneously, co-eluting products will have a certain impact on ionization efficiency. Therefore, good chromatographic separation of each compound is the basis and key to accurate identification and quantification. This study optimized the gradient elution procedure and compared the effects of acetonitrile-water, acetonitrile-0.1% formic acid aqueous solution, methanol-0.1% formic acid aqueous solution, and acetonitrile-5mM ammonium formate solution (containing 0.1% formic acid) mobile phase systems on the separation of target compounds under the same gradient conditions. The results showed that acetonitrile-water resulted in severe peak tailing and broad peaks. Using methanol-0.1% formic acid aqueous solution prolonged the peak elution time and increased peak broadening, hindering rapid detection. Using acetonitrile-5mM ammonium formate solution (containing 0.1% formic acid) resulted in a weak mass spectrometric response, affecting the sensitivity of the method. However, using acetonitrile-0.1% formic acid aqueous solution as the mobile phase resulted in good separation of the compounds, symmetrical peak shapes, and superior mass spectrometric signal response. Therefore, the acetonitrile-0.1% formic acid aqueous solution mobile phase was ultimately selected.

[0034] In the event of any conflict between the Chinese nomenclature and the structural formula of the compounds of the present invention, the structural formula shall prevail.

[0035] The beneficial effects of the present invention are as follows: the present invention uses the above technical solution to establish for the first time an ultra-high performance liquid chromatography-tandem mass spectrometry method for simultaneously detecting 21 index components in 14 plant-derived raw materials in toothpaste, which makes up for the lack and insufficiency of relevant detection methods and standards for the simultaneous determination of multiple plant-derived and multiple index components in toothpaste. The UPLC-MS / MS method used in this application to detect the index components of 21 plant-derived raw materials can detect 21 index components of 14 plant sources at one time under the mode of simultaneous scanning of positive and negative ions. The analysis time of the method is short and the detection efficiency is high, which can meet the purpose of rapid screening and quantitative analysis of multiple plant-derived index components in daily supervision. This method optimizes the liquid chromatography conditions, mass spectrometry parameters and sample pretreatment methods, and completes the detection of 21 target compounds within 25 minutes. The method features simple and efficient pretreatment, short analysis time, high sensitivity, strong resistance to matrix interference, and accurate and reliable qualitative and quantitative results. It can be used for quality control of relevant botanical raw materials and for evaluating the consistency between labeling and ingredients in toothpastes containing botanical ingredients. This method provides technical support for further standardizing toothpaste labeling, promoting standardized and healthy competition among toothpaste registrants and manufacturers, and post-registration and marketing supervision of toothpaste. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 is the total ion current chromatogram (TIC) of the mixed reference solution;

[0037] Figure 2-4 The MRM channel (quantitative ion) chromatogram of 21 compounds in the mixed reference solution;

[0038] Figure 5 is the total ion chromatogram (TIC) of the blank solution;

[0039] Figure 6 is the total ion current chromatogram (TIC) of a test solution;

[0040] Figure 7 This is the MRM channel (quantitative ion) chromatogram of the index component detected in a test solution. DETAILED DESCRIPTION

[0041] The present invention is described below with reference to examples, but is not limited thereto. Simple replacements or improvements made by those skilled in the art to the present invention fall within the technical solutions protected by the present invention.

[0042] Example 1: The chromatographic column is ACQUITY UPLC BEH C 18Chromatographic column (2.1×100 mm, 1.7 μm); mobile phase: acetonitrile (A)-0.1% formic acid solution (B), gradient elution: 0-1 min, 10% A; 1-12 min, 10%-25% A; 12-16 min, 25%-50% A; 16-18 min, 50%-95% A; 18-23 min, 95% A; 23.01-25 min, 10% A. Flow rate: 0.3 mL / min; column temperature: 35°C; injection volume: 2.0 μL.

[0043] Ion source: electrospray ionization (ESI), simultaneous positive and negative ion multiple reaction monitoring (MRM) scan mode, source temperature: 500°C; spray voltage: 5500 V in positive ion mode, 4500 V in negative ion mode; curtain gas: 30 psi; nebulizer gas pressure: 55 psi; auxiliary gas pressure: 55 psi. Scan mode: MRM. Mass spectrometric parameters, including precursor ions, product ions, declustering potential (DP), and collision energy (CE), for the 21 compounds are shown in Table 2 below. Those marked with "*" are quantification product ions.

[0044] Table 2 Mass spectrometry acquisition parameters for 21 index components

[0045]

[0046]

[0047] Preparation of reference solution: Weigh appropriate amount of 21 reference substances, dissolve them in methanol by ultrasonication and dilute them to prepare single standard stock solution. Accurately measure appropriate amount of single standard stock solution and dilute with 80% methanol aqueous solution to prepare reference substance working solution of 21 components. Specifically, each 1mL can contain 25mg / L of reference substance solution. Before use, measure appropriate amount of reference substance working solution to prepare a series of standard curve solutions. The total ion current chromatogram (TIC) of the mixed reference substance solution at the middle concentration point of the standard curve is shown in Figure 1. Figure 1 The MRM channel (quantitative ion) chromatograms of the 21 compounds are shown in Figure 2-4 .

[0048] Preparation of the test solution: Weigh 1g of toothpaste sample (accurate to 0.0001g), place it in a 15mL stoppered centrifuge tube, add 10mL of 80% methanol solution, then add two glass beads, vortex thoroughly to disperse the toothpaste evenly. Ultrasonic extraction for 25 minutes, cool to room temperature, centrifuge at 3000rpm for 10 minutes, take the supernatant and filter it through a 0.2μm filter membrane, and take the filtrate as the test solution. If necessary, dilute it with 80% methanol solution to the standard curve range and then inject the sample for measurement. The total ion current chromatogram (TIC) of a test solution and the MRM channel (quantitative ion) chromatogram of the detected index component are shown in Figure 2. Figure 6 and Figure 7 .

[0049] The specificity test showed that there was no chromatographic peak in the blank solution chromatogram that interfered with the determination of the target substance, and the method was highly specific (see Appendix Figure 5 ).

[0050] Linearity test results showed that each component exhibited a good linear relationship with peak area within the linear range, as shown in Table 3. The limits of detection and quantification (LOQs) for each component were determined using S / N ≥ 3 and S / N ≥ 10, respectively. The LQs for the 21 components were 0.0017 to 0.033 μg / g and 0.0050 to 0.10 μg / g, respectively.

[0051] Table 3 Matrix effects, linear relationships, detection limits, and quantification limits of 21 indicator components

[0052]

[0053]

[0054] Accuracy and precision results showed that blank toothpaste samples were used as spiked matrices. Approximately 1 g of the sample was weighed into a 15 mL stoppered centrifuge tube. Different volumes of the mixed standard solution were added and pretreated according to the test sample preparation method. The spiked sample solutions were prepared at high, medium, and low concentration levels (six replicates were prepared for each concentration level). The samples were injected and measured, and the average recovery and relative standard deviation (RSD, n = 6) of each component were calculated. The average recoveries of the 21 components in the toothpaste matrix ranged from 88.3% to 121.0%, with RSDs ranging from 0.9% to 12.6%. These results demonstrate that the method has good accuracy and precision and can be used for the quantitative determination of 21 plant-derived indicator components in toothpaste.

[0055] Table 4 Recovery and precision results of 21 index components in toothpaste matrix

[0056]

[0057]

[0058] Toothpaste matrix is complex, and it may produce inhibitory or enhanced matrix effects during mass spectrometry detection, thereby affecting the accuracy, precision and sensitivity of the method. The size of the matrix effect is generally evaluated using the formula ME = K2 / K1, where K1 is the slope of the standard curve prepared with the extraction solvent, and K2 is the slope of the standard curve prepared with the toothpaste blank matrix solution. The closer the ME value is to 1, the smaller the impact of the matrix effect. The toothpaste matrix was selected to investigate the influence of the matrix effect of 21 compounds, and the results are shown in Table 4. The results showed that the ME values of the 21 compounds were between 0.68 and 1.60, and some compounds had a certain degree of matrix interference. In order to effectively eliminate the interference of the matrix effect, this study used the toothpaste blank matrix solution to prepare the standard curve to minimize the impact of the matrix effect on the accuracy of the quantitative results of the detected index components.

[0059] The established method was used to conduct qualitative and quantitative testing on 38 batches of toothpaste samples purchased from the market that were labeled as containing single or multiple plant sources. The indicator components of 13 plant sources selected for this study were detected, including coral grass, two-sided needle, honeysuckle, Panax notoginseng (Panax notoginseng), Scutellaria baicalensis, Coptis chinensis, Paeonia suffruticosa, Magnolia officinalis, Scutellaria baicalensis, Tea, Taraxacum mongolicum, Coptis chinensis, Isatis indigotica (Isatis indigotica), but no indicator components of aloe vera were detected (see Table 5 for details). As can be seen from Table 5, among the 5 batches labeled as having plant sources of Zanthoxylum bungeanum, 1 batch had no detected index components; among the 5 batches labeled as having plant sources of Lonicera japonica, 1 batch had no detected index components; among the 8 batches labeled as having plant sources of Panax notoginseng, 1 batch had no detected index components; among the 7 batches labeled as having plant sources of Scutellaria baicalensis, 1 batch had no detected index components; among the 6 batches labeled as having plant sources of Aloe vera, 2 batches had no detected index components; among the 6 batches labeled as having plant sources of Isatis indigotica, 1 batch had no detected index components; and among the toothpastes labeled as having plant sources of Coral Recutita, Paeonia suffruticosa, Coptis chinensis, Magnolia officinalis, Scutellaria baicalensis, Taraxacum mongolicum, and Coptis chinensis, one or more index components were detected in all the toothpastes. In summary, toothpaste labels indicate the addition of plant-derived raw materials, but there are cases where the plant-derived indicator ingredients are not detected. At the same time, the content of the indicator ingredients of the same plant-derived raw materials varies greatly among different manufacturers. For example, the content of notoginseng saponin R1 in toothpaste products labeled as containing Panax notoginseng (Tianqi) plant-derived raw materials by different manufacturers varies by nearly 150 times, and the content of lilacin in toothpaste products labeled as containing Jiubiying plant-derived raw materials by different manufacturers varies by more than 1,000 times. The above results suggest that it is necessary to standardize the standards for plant-derived raw materials in toothpaste to avoid the abuse of the concept of plant-derived ingredients by some companies, such as false advertising of the addition of plant-derived raw materials, insufficient or no addition of raw materials, and other situations that harm the interests of consumers.

[0060] Table 5 Test results of 38 batches of toothpaste samples

[0061]

[0062] The above description is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this field, several variations and improvements can be made without departing from the creative concept of the present invention, which all fall within the scope of protection of the present invention.

Claims

1. A method for simultaneously determining the index components of 21 kinds of plant-derived raw materials in toothpaste, characterized in that: The steps include: (1) Preparation of reference solution; (2) Preparation of test solution; (3) Calculate the contents of 21 index components in the sample using the quantitative ion peak area according to the external standard method; The method described was detected using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry; The ultra-high performance liquid chromatography (UPLC) conditions are as follows: the chromatographic column is filled with octadecylsilane bonded silica gel, with an inner diameter of 2.1 mm and a particle size of 1.7 or 1.8 μm; 0.1% formic acid aqueous solution is used as mobile phase B, acetonitrile is used as mobile phase A, and gradient elution is performed; the injection volume is 2±1 μL; the flow rate is 0.3±0.1 mL / min; and the column temperature is 35±5°C. The mass spectrometry conditions are as follows: ion source: electrospray ionization source (ESI), positive and negative ion simultaneous monitoring mode; Ion source temperature: 500±50°C; spray voltage: 5500±500V in positive ion mode, 4500±500V in negative ion mode; curtain gas: 30±10psi; nebulizer gas pressure: 55±10psi; auxiliary gas pressure: 55±10psi; scan mode: multiple reaction monitoring (MRM) to detect qualitative and quantitative product ions; The index components of the 21 plant-derived raw materials are 1: aloin; 2: syringin; 3: corydaline; 4: nitine chloride; 5: berberine hydrochloride; 6: Coptisine hydrochloride; 7: isoflurane; 8: Paeonol; 9: (R,S)-Gaoitrin; 10: Ginsenoside Rb1; 11: Notoginsenoside R1; 12: Ginsenoside Rg1; 13: Longipenoside; 14: Luteolin; 15: Baicalin; 16: Rosmarinic acid; 17: Chlorogenic acid; 18: Catechin; 19: Honokiol; 20: Magnolol; 21: Caffeic acid.

2. The method according to claim 1, characterized in that The specific operation for preparing the reference substance solution is as follows: weighing appropriate amounts of 21 reference substances respectively, adding methanol to ultrasonically dissolve and dilute to prepare a single-standard stock solution, accurately measuring an appropriate amount of the single-standard stock solution, adding 80% methanol aqueous solution to dilute to prepare a reference substance working solution of 21 components, specifically, each 1 mL can contain 25±10 mg / L of each reference substance solution, and before use, measuring an appropriate amount of the reference substance working solution to prepare a series of standard curve solutions.

3. The method according to claim 1, characterized in that The preparation operation of the test solution is as follows: weighing a toothpaste sample, adding a methanol-water solution, then adding two glass beads, vortexing to evenly disperse the toothpaste, ultrasonically extracting, cooling to room temperature, centrifuging, filtering the supernatant, and taking the filtrate as the test solution.

4. The method according to claim 3, characterized in that The specific operation for preparing the test solution is as follows: weigh 1 g of toothpaste sample, accurate to 0.0001 g, place it in a 15 mL stoppered centrifuge tube, add 10 mL of 80% methanol aqueous solution, then add two glass beads, vortex and oscillate thoroughly to evenly disperse the toothpaste, perform ultrasonic extraction for 25 minutes, cool to room temperature, centrifuge at 3000 rpm for 10 minutes, take the supernatant and filter it through a 0.2 μm filter membrane, and take the filtrate as the test solution.

5. The method according to claim 1, wherein The chromatographic column is ACQUITY UPLC BEH C 18 Chromatographic columns: 2.1 mm × 100 mm, 1.7 μm, Waters ACQUITY UPLC HSS T3 2.1 mm × 100 mm, 1.8 μm.

6. The method according to claim 1, characterized in that The gradient elution has the following specific procedures: 0-1 min, 10% A; 1-12 min, 10%-25% A; 12-16 min, 25%-50% A; 16-18 min, 50%-95% A; 18-23 min, 95% A; 23.01-25 min, 10% A.

7. The method according to claim 1, characterized in that The injection volume is specifically 2 μL, the flow rate is specifically 0.3 mL / min, and the column temperature is specifically 35° C.

8. The method according to claim 1, characterized in that The parent ion, daughter ion, declustering voltage (DP), and collision energy (CE) mass spectrometry parameters of the 21 plant-derived raw material compounds are shown in the following table, where the ions marked with "*" are quantitative daughter ions. 。

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