CN116242941A

CN116242941A - Analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics based on electrospray technology

Info

Publication number: CN116242941A
Application number: CN202310242767.4A
Authority: CN
Inventors: 吴映璇; 邵琳智; 陈思敏; 欧阳少伦; 董洁; 蓝草; 邹游
Original assignee: Guangzhou Customs Technology Center
Current assignee: Guangzhou Customs Technology Center
Priority date: 2023-03-13
Filing date: 2023-03-13
Publication date: 2023-06-09

Abstract

An analytical method for measuring 1,3, 7-trimethyl xanthine in cosmetics is established by adopting a high performance liquid chromatography-mass spectrometry/mass spectrometer. The method has the advantages of simple operation, low cost, high sensitivity, strong anti-interference capability and accurate qualitative and quantitative analysis. The method can provide technical support for monitoring 1,3, 7-trimethyl xanthine in cosmetics, is helpful for enhancing the detection technology in China, provides technical support for regulatory management market of regulatory departments, and provides technical support for protecting the health of consumers and guaranteeing the safety of cosmetics.

Description

Analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics based on electrospray technology

Technical Field

The invention belongs to the technical field of cosmetic detection, and particularly relates to a method for measuring the content of 1,3, 7-trimethyl xanthine in cosmetics.

Background

1,3, 7-trimethylxanthine is also known as Caffeine (Caffeine, molecular formula: C) ₈ H ₁₀ N ₄ O ₂ ) Is a central nervous stimulant, can temporarily expel drowsiness and restore energy, and is clinically used for treating neurasthenia and coma resuscitation. 1,3, 7-trimethylxanthine is also the most commonly used psychotropic drug in the world. The world health organization International cancer research institute lists 1,3, 7-trimethylxanthine in the 3-class carcinogen list, which is also listed in China as a "psychotropic" control.

At present, a plurality of beauty and skin care products are added with 1,3, 7-trimethyl xanthine for improving microcirculation to reduce eye bags and dark circles, such as eye cream; in hair care products, to regulate hair growth and to help slow down hair loss; the skin-activating and color-increasing composition is also used for improving the appearance of the skin, enabling the appearance of the skin to be smoother, and the like. The cosmetic can be in direct contact with the skin, hair, lips, oral cavity and other parts of a human body for a long time in the use process, so that the safety problem of the cosmetic becomes a focus of attention of all parties, and the use of part of components is always a focus of research at home and abroad. In China, the technical Specification for cosmetic safety (2015 edition) does not make a limit regulation on 1,3, 7-trimethyl xanthine components in cosmetics, and 1,3, 7-trimethyl xanthine components in import and export cosmetics are not monitored.

Disclosure of Invention

In view of the technical problems of the existing cosmetic detection, the invention provides an analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics based on an electrospray technology. In order to solve the technical problems, the invention adopts the following technical scheme:

an analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics based on an electrospray technology, wherein a sample treatment method of the analysis method comprises the following steps: about 0.5g of the sample is weighed, placed in a 50mL centrifuge tube, added with 20mL of normal hexane, well dispersed by vortex, accurately added with 20mL of methanol-water solution, extracted by ultrasonic for 20min, and centrifuged at 2500 r/min for 5min. Discarding the upper n-hexane layer, accurately sucking 0.1mL of the purified sample extract into a 1.5mL centrifuge tube, adding 0.9mL of methanol-water solution, performing vortex oscillation for 30s, centrifuging at 10000r/min for 5min, sucking the intermediate supernatant, filtering with a filter membrane, and measuring by a high performance liquid chromatography-mass spectrometer.

The high performance liquid chromatography conditions of the analysis method are as follows: chromatographic column: waters Atlantis T3 column, 2.1mm (inner diameter). Times.100 mm,3 μm; mobile phase: acetonitrile-5 mmol/L formic acid solution, and gradient elution; flow rate: 0.25mL/min; column temperature: 40 ℃; sample injection amount: 5. Mu.L; the gradient elution conditions are as follows: 0min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;0.5min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;4min, acetonitrile% -5mmol/L formic acid solution% = 95% -5%;5.5min, acetonitrile% -5mmol/L formic acid solution% = 95% -5%;5.51min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;10min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%.

The mass spectrum conditions of the analysis method are as follows: ion source: an electrospray ion source; scanning mode: a positive ion; the detection mode is as follows: multiple Reaction Monitoring (MRM); mass spectrum/mass spectrum reference conditions: curtain air pressure (CUR): 172kPa; electrospray voltage: 5500V; ion source Temperature (TEM): 650 ℃; atomizer pressure (GAS 1): 379kPa; auxiliary Gas pressure (Gas 2): 379kPa; the conditions for monitoring the multiple reactions are as follows: ion pair m/z:195/138, declustering voltage: DP/V49, collision cell inlet voltage: EP/V8, collision cell outlet voltage CXP/V:10, collision energy CE/V:25, a step of selecting a specific type of material; ion pair m/z:195/110, declustering voltage: DP/V49, collision cell inlet voltage: EP/V8, collision cell outlet voltage CXP/V:10, collision energy CE/V:23.", is a quantitative ion.

The liquid chromatography-mass spectrometry/mass spectrometry determination and confirmation method of the analysis method comprises the following steps: and measuring 2.0 mug/L, 5.0 mug/L, 10 mug/L, 20 mug/L and 50 mug/L of series standard working solutions according to the conditions of high performance liquid chromatography-mass spectrometry/mass spectrometry, drawing a standard working curve by taking the standard solution concentration as an abscissa and the corresponding peak area as an ordinate, and quantifying according to the solution concentration on the standard working curve corresponding to the peak area. The content of the to-be-detected object in the sample solution is within the linear range of the standard curve, and the to-be-detected object is diluted and then analyzed when the content exceeds the linear range. Under the condition, the multi-reaction monitoring total ion flow diagram and the secondary mass spectrum of the 1,3, 7-trimethyl xanthine are shown in the accompanying drawings 1-2 of the specification.

The sample processing solution is measured according to the conditions of liquid chromatography-mass spectrometry/mass spectrometry, if the retention time of chromatographic peaks of substances to be detected in the sample is consistent with that of a standard solution (the deviation is within +/-2.5%), the relative abundance of qualitative ion pairs is expressed by the intensity percentage relative to the abundance of the strongest ions, the relative abundance should be consistent with that of a working solution with a concentration equivalent to that of the standard, and the allowable deviation range of the relative abundance is as follows: the relative ion abundance is more than 50 percent, and the allowable relative deviation is +/-20 percent; the relative ion abundance is more than 20-50%, and the allowable relative deviation is +/-25%; the relative ion abundance is more than 10-20%, and the allowable relative deviation is +/-30%; the relative ion abundance is less than or equal to 10 percent, and the allowable relative deviation is +/-50 percent, so that the existence of the corresponding object to be detected in the sample can be judged.

The result of the analysis method is calculated and expressed as:

calculating the content of 1,3, 7-trimethylxanthine in the sample by using a chromatographic data processor or according to formula (1), and subtracting a blank value from the calculated result:

in formula 1:

X _i -the content of the component to be measured in micrograms per kilogram (mg/kg) in the sample;

c-the concentration of the measured component in the sample solution obtained by the standard curve is given in nanograms per milliliter (mug/mL);

c ₀ -the concentration of the component to be tested in nanograms per milliliter (μg/mL) in a blank test obtained from a standard curve;

v-final constant volume of sample solution in milliliters (mL);

m-the mass of the sample represented by the final sample solution in grams (g).

The result of the calculation retains three significant digits.

The linear relation and the detection limit of the analysis method are as follows: under the experimental conditions determined by the standard method, a series of standard solutions with different concentrations are taken, the concentration of 1,3, 7-trimethylxanthine is linearly regressed by the response peak area of an instrument, and the result shows that when the concentration of 1,3, 7-trimethylxanthine is in the range of 2.0 mug/L to 50 mug/L, the linear relation is good, the regression equation y=16362.77865x+2845.75302 and the correlation coefficient r= 0.99998. When the 1,3, 7-trimethylxanthine concentration in the sample exceeds this linear range, the dilution factor of the sample can be appropriately increased. The detection limit and the quantitative limit are calculated by the signal to noise ratio of 3 times and the signal to noise ratio of 10 times respectively, and the detection limit of the obtained 1,3, 7-trimethyl xanthine is 0.06mg/kg and the quantitative limit is 0.2mg/kg.

The sample analyzed by the analysis method comprises: toner, lotion, eye cream, shampoo, eye shadow, blush, and lipstick.

The invention has the following beneficial effects in cosmetics:

the invention establishes an analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics by adopting a high performance liquid chromatograph for the first time, and fills up the technical blank in the field. The method has the advantages of simple operation, short analysis time, low detection cost, good precision and high accuracy. The n-hexane-methanol-water solution is used as an extraction solvent, so that the key technical problem of sample dispersion is solved, the extraction efficiency of 1,3, 7-trimethyl xanthine is improved, and the effect of sample purification is achieved. Such technical effects are unexpected to those skilled in the art, having outstanding substantive features and significant technical advances. The invention can be applied to daily detection work of 1,3, 7-trimethyl xanthine in cosmetics, and provides technical support for regulatory management market of regulatory departments.

The patent is different from the research method of the former, wherein most of the research of the former is concentrated on matrixes such as tea, beverage, health care products and the like, the matrixes related to cosmetics are few, and the matrixes applicable to the detection method only detect water, dew, milk or cream only, and relevant reports of cosmetics such as inclusion water, dew, milk, cream, eye shadow, blush, lipstick and the like are fresh; the previous researches mostly adopt methanol, acetonitrile or water-methanol-acetonitrile solution as extraction solvent, as most of samples contain a large amount of lipid, the dispersion is difficult to be uniform, target analytes are coated and difficult to be extracted completely, the recovery rate is 67.4% -127.6%, n-hexane-methanol-water solution is not adopted as extraction solvent, and in order to solve the problems, 4 solvents with better lipophilicity are selected in the patent: n-hexane, dichloromethane, diethyl ether and ethyl acetate were studied, as can be seen from the lipid partition coefficients (ACD/LogP) of the 4 solvents: ACD/LogP of n-hexane, dichloromethane, diethyl ether and ethyl acetate were 3.94, 1.19, 0.98 and 0.71 respectively, while the lipid water partition coefficient (ACD/LogP) of 1,3, 7-trimethylxanthine was-0.13, according to the principle of similar miscibility, ethyl acetate was more similar to that of 1,3, 7-trimethylxanthine, not only lipid substances in the sample, but also 1,3, 7-trimethylxanthine in the sample was dissolved, diethyl ether was inferior, dichloromethane was inferior, and n-hexane was worst. Thus, it was preliminarily predicted that ethyl acetate was most effective for dissolving a sample containing a large amount of lipid substances and 1,3, 7-trimethylxanthine. Subsequent comparative studies of the extraction test of n-hexane, dichloromethane, ethyl acetate and diethyl ether on the samples revealed that: the methylene dichloride, ethyl acetate and diethyl ether can only partially disperse and dissolve the lipstick and other samples, the extraction is incomplete, and the normal hexane can completely disperse the lipstick and other samples containing a large amount of lipids, so as to achieve the maximum extraction of target components (see Table 1 for details). This result is inconsistent with the predicted conclusion of the lipid partition coefficient. Then according to the lipid water distribution coefficient (ACD/LogP: -0.13) of the 1,3, 7-trimethyl xanthine, methanol (ACD/LogP: -0.72) -water (ACD/LogP: -1.38) solution with the lipid water distribution coefficient as small as possible is selected to extract the target analyte from the normal hexane solution, a large amount of lipid substances still remain in the normal hexane solution, the extraction effect is obvious, and the recovery rate is 95% -108%; the former research (Liao Hao, et al in the cosmetics published in the "health research" of 2016, 1) needs to use a water bath heating mode to assist in sample extraction, and the patent can complete sample extraction without adding an additional device, thereby reducing detection cost; the previous research does not see that any purification mode is adopted for direct detection, and the liquid-liquid extraction purification mode is adopted when the sample is extracted, so that the pretreatment time is short, and the cost is low; previous studies performed chromatographic separations using isocratic elution, and no gradient elution was seen. The patent finds that the actual sample is subjected to chromatographic separation by adopting an isocratic elution mode, impurities still remain on the chromatographic column until the next sample is eluted from the chromatographic column together during separation, and the chromatographic separation of the next sample is seriously interfered. The chromatographic separation of the actual sample by adopting the gradient elution mode can ensure that the target analyte and impurities in the secondary sample can be eluted without influencing the chromatographic separation of the next sample. The former research adopts liquid chromatography to detect, is easily interfered by complex matrixes in a sample, cannot accurately quantify, does not adopt high performance liquid chromatography-mass spectrometry/mass spectrometry to detect, can effectively avoid the interference of impurities in the sample extraction process, improves the detection accuracy, and has higher selectivity and higher sensitivity. In addition, the patent also adopts a dilution method, simplifies the pretreatment process, avoids the loss of the target in the complex pretreatment process, and weakens the matrix effect. Thereby obviously improving the accuracy and repeatability of the method and improving the precision of the method. The method is simple and convenient to operate, adopts the technologies of high performance liquid chromatography-tandem quadrupole mass spectrometry combined electrospray positive ion monitoring and the like, has high sensitivity and strong anti-interference capability, is accurate in qualitative and quantitative, can provide technical support for monitoring work of 1,3, 7-trimethyl xanthine in cosmetics, is beneficial to enhancing the detection technology in China, and provides technical support for protecting the physical health of consumers and guaranteeing the safety of the cosmetics.

Drawings

FIG. 1,3, 7-trimethylxanthine standard total ion flow graph (5.0. Mu.g/L);

FIG. 2, secondary mass spectrum of 3, 7-trimethylxanthine;

FIG. 3 reagent blank total ion flow diagram;

FIG. 4 is a total ion flow diagram of 1,3, 7-trimethylxanthine in a toner;

FIG. 5 is a total ion flow diagram of 1,3, 7-trimethylxanthine in a standard addition sample of toner (addition level 2.0 mg/kg);

FIG. 6 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in a lotion;

FIG. 7 total ion flow diagram of 1,3, 7-trimethylxanthine in standard addition sample of lotion (addition level 2.0 mg/kg);

FIG. 8 is a total ion flow diagram of 1,3, 7-trimethylxanthine in an eye cream;

FIG. 9 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in a standard addition sample of eye cream (addition level 2.0 mg/kg);

FIG. 10 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in a shampoo;

FIG. 11 total ion flow diagram of 1,3, 7-trimethylxanthine in shampoo standard addition sample (addition level 2.0 mg/kg);

FIG. 12 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in an eye shadow;

FIG. 13 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in a standard addition sample (addition level 2.0 mg/kg);

FIG. 14 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in blush;

FIG. 15 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in a blush standard addition sample (addition level 2.0 mg/kg);

FIG. 16 is a total ion flow diagram of 1,3, 7-trimethylxanthine in lipstick;

FIG. 17 total ion flow diagram of 1,3, 7-trimethylxanthine in lipstick standard addition sample (addition level 2.0 mg/kg);

FIG. 18 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in toner A;

FIG. 19 is a total ion flow diagram of 1,3, 7-trimethylxanthine in toner B;

fig. 20 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in lotion a;

fig. 21 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in lotion B;

FIG. 22 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in shampoo A;

FIG. 23 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in shampoo B;

FIG. 24 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in eye cream A;

FIG. 25 shows a total ion flow diagram for 1,3, 7-trimethylxanthine in eye cream B;

FIG. 26 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in eye shadow A;

FIG. 27 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in eye shadow B;

FIG. 28 shows a total ion flow diagram for 1,3, 7-trimethylxanthine in blush A;

FIG. 29 is a total ion flow diagram of 1,3, 7-trimethylxanthine in lipstick A;

FIG. 30 shows a total ion flow diagram of 1,3, 7-trimethylxanthine in lipstick B.

Detailed Description

In order to more clearly illustrate the technical scheme of the present invention, the present invention will be described in detail with reference to specific embodiments and drawings, but the scope of the present invention is not limited to the following embodiments.

Example 1: comparative experiments with different extraction solvents

TABLE 1 residue of samples after lipstick extraction with n-hexane, ethyl acetate, dichloromethane and diethyl ether

The same comparative experiments were performed on lotions and lotions, shampoos, eye creams, eye shadows, blushes, and the results were similar to lipsticks.

Example 2: method for establishing

1.1 major materials, reagents and instruments

5 parts of toner, 5 parts of skin lotion, 4 parts of shampoo, 6 parts of eye cream, 5 parts of eye shadow, 3 parts of blush and 7 parts of lipstick samples, which are commercially available in Guangzhou.

UFLC LC-20A ultra-high performance liquid chromatograph, shimadzu corporation; API 4000QTRAP triple quadrupole tandem mass spectrometer distribution spray ion source AB Sciex company, america; vortex oscillator, IKA company, germany; a type 3-16K high-speed centrifuge, sigma, germany; ultrasonic cleaner, bo Xuan Co.

1,3, 7-trimethylxanthine standard solution (10016.9 mg/L), man Ha Ge, china; methanol, chromatographic purity, tedia company, usa; n-hexane, analytically pure, national drug group chemical agents limited; the laboratory water was Milli-Q ultra-pure water.

1.2 preparation of solutions

Methanol-water solution: 50mL of methanol and 50mL of water are measured and placed in a triangular flask with a stopper, and are shaken well for later use.

1,3, 7-trimethylxanthine standard stock solution (500 mg/L): accurately sucking a proper amount of 1,3, 7-trimethyl xanthine standard solution, placing the solution into a 10mL volumetric flask, fixing the volume to a scale with methanol, and uniformly mixing. The solution is preserved at 0-4 ℃.

1,3, 7-trimethylxanthine standard intermediate (10 mg/L): accurately sucking a proper amount of 1,3, 7-trimethylxanthine standard solution (500 mg/L), placing into a 100mL volumetric flask, metering to scale with methanol, and mixing. The solution is preserved at 0-4 ℃.

Standard use solution of 1,3, 7-trimethylxanthine (500. Mu.g/L): accurately transferring the 1,3, 7-trimethylxanthine standard intermediate (10 mg/L) into a volumetric flask of 2.5mL to 50mL, diluting to a scale with methanol, and shaking uniformly. The solution is preserved at 0-4 ℃.

1,3, 7-trimethyl xanthine standard working solution: respectively and accurately transferring 0.2mL, 0.5mL, 1.0mL, 2.0mL and 5.0mL to 50mL of standard use solution (500 mug/L) of 1,3, 7-trimethylxanthine, diluting to a scale with methanol, and shaking uniformly to obtain a series of standard working solutions of 2.0 mug/L, 5.0 mug/L, 10 mug/L, 20 mug/L and 50 mug/L, and preparing for use.

1.3 sample handling

About 0.5g (accurate to 0.0001 g) of the sample is weighed, placed in a 50mL centrifuge tube, 20mL of normal hexane is added, after vortex dispersion is uniform, 20mL of methanol-water solution is accurately added, ultrasonic extraction is carried out for 20min, and centrifugation is carried out for 5min at 3500r/min. Discarding the upper n-hexane layer, accurately sucking 0.1mL of the purified sample extract into a 1.5mL centrifuge tube, adding 0.9mL of methanol-water solution, performing vortex oscillation for 30s, centrifuging at 10000r/min for 5min, sucking the intermediate supernatant, filtering with a filter membrane, and measuring by a high performance liquid chromatography-mass spectrometer.

1.4 chromatographic conditions

The conditions of high performance liquid chromatography-mass spectrometry/mass spectrometry are as follows:

chromatographic column: waters Atlantis T3 column, 2.1mm (inner diameter) ×100mm,3 μm, or equivalent;

mobile phase: acetonitrile-5 mmol/L formic acid solution, gradient elution, see Table 1;

flow rate: 0.25mL/min;

column temperature: 40 ℃;

sample injection amount: 5. Mu.L;

ion source: an electrospray ion source;

scanning mode: a positive ion;

the detection mode is as follows: multiple Reaction Monitoring (MRM);

mass spectrum/mass spectrum reference conditions: curtain air pressure (CUR): 172kPa; electrospray voltage: 5500V; ion source Temperature (TEM): 650 ℃; atomizer pressure (GAS 1): 379kPa; auxiliary Gas pressure (Gas 2): 379kPa; the conditions for multiple reaction monitoring are shown in Table 2:

TABLE 1 gradient elution procedure for mobile phases

Gradient time/min	Acetonitrile/%	5mmol/L formic acid solution/%
			0	10	90
0.5	10	90
			4	95	5
5.5	95	5
			5.51	10	90
10	10	90

TABLE 2 Multi-reaction monitoring (MRM) conditions for 1,3, 7-trimethylxanthine

1.5 liquid chromatography-Mass Spectrometry/Mass Spectrometry determination and validation

A series of standard working solutions of 2.0 mug/L, 5.0 mug/L, 10 mug/L, 20 mug/L and 50 mug/L are measured according to the condition of high performance liquid chromatography-mass spectrum/mass spectrum (1.4), the standard solution concentration is taken as an abscissa, the corresponding peak area is taken as an ordinate, a standard working curve is drawn, and the quantification is carried out according to the solution concentration on the standard working curve corresponding to the peak area. The content of the to-be-detected object in the sample solution is within the linear range of the standard curve, and the to-be-detected object is diluted and then analyzed when the content exceeds the linear range. Under the condition, the multi-reaction monitoring total ion flow diagram and the secondary mass spectrum of the 1,3, 7-trimethyl xanthine are shown in figures 1-2.

The sample processing solution is measured according to the conditions of liquid chromatography-mass spectrometry/mass spectrometry, if the retention time of the chromatographic peak of the substance to be detected in the sample is consistent with that of the standard solution (the deviation is within +/-2.5%), the relative abundance of the qualitative ion pair is expressed by the intensity percentage relative to the abundance of the strongest ion, the relative abundance should be consistent with that of the standard working solution with the equivalent concentration, and the allowable deviation of the relative abundance does not exceed the range specified in table 3, the existence of the corresponding substance to be detected in the sample can be judged.

1.6 liquid chromatography-Mass Spectrometry/Mass Spectrometry determination and validation

A series of standard working solutions of 2.0 mug/L, 5.0 mug/L, 10 mug/L, 20 mug/L and 50 mug/L are measured according to the condition of high performance liquid chromatography-mass spectrum/mass spectrum (1.4), the standard solution concentration is taken as an abscissa, the corresponding peak area is taken as an ordinate, a standard working curve is drawn, and the quantification is carried out according to the solution concentration on the standard working curve corresponding to the peak area. The content of the to-be-detected object in the sample solution is within the linear range of the standard curve, and the to-be-detected object is diluted and then analyzed when the content exceeds the linear range. Under the condition, the multi-reaction monitoring total ion flow diagram and the secondary mass spectrum of the 1,3, 7-trimethyl xanthine are shown in the accompanying drawings 1-2 of the specification.

TABLE 3 maximum allowable deviation of relative ion abundance in qualitative validation

Relative ion abundance	＞50％	＞20％～50％	＞10％～20％	≤10％
					Allowable relative deviation	±20％	±25％	±30％	±50％

1.7 blank test

The procedure was followed except that no sample was added.

1.8 results calculation and presentation

wherein:

c ₀ -concentration of the tested component in nanograms per milliliter in a blank test obtained from a standard curve

(μg/mL)；

V-final constant volume of sample solution in milliliters (mL);

m-the mass of the sample represented by the final sample solution in grams (g).

The result of the calculation retains three significant digits.

1.9 Linear relationship and detection Limit

Under the experimental conditions determined by the standard method, a series of standard solutions with different concentrations are taken, the concentration of 1,3, 7-trimethylxanthine is linearly regressed by the response peak area of an instrument, and the result shows that when the concentration of 1,3, 7-trimethylxanthine is in the range of 2.0 mug/L to 50 mug/L, the linear relation is good, the regression equation y=16362.77865x+2845.75302 and the correlation coefficient r= 0.99998. When the 1,3, 7-trimethylxanthine concentration in the sample exceeds this linear range, the dilution factor of the sample can be appropriately increased. The detection limit and the quantitative limit are calculated by 3 times signal to noise ratio and 10 times signal to noise ratio respectively, the detection limit of the obtained 1,3, 7-trimethyl xanthine is 0.06mg/kg, and the quantitative limit is 0.2mg/kg

1.10 method recovery and precision

Three level addition recovery experiments were performed on cosmetic samples such as toner, skin lotion, eye cream, shampoo, eye shadow, blush, and lipstick, each addition level was repeated 6 times, the average recovery and relative standard deviation data results of each addition level are detailed in table 4, and the corresponding liquid chromatogram is shown in fig. 3-17 of the specification.

Table 4 recovery and precision of 1,3, 7-trimethylxanthine (n=6) in cosmetic products

Example 3: actual sample measurement

The method is used for measuring 5 parts of commercially available toner, 5 parts of skin lotion, 4 parts of shampoo, 6 parts of eye cream, 5 parts of eye shadow, 3 parts of blush and 7 parts of lipstick samples, and the details are shown in figures 18-30 of the specification.

Claims

1. An analysis method for measuring 1,3, 7-trimethyl xanthine in cosmetics based on an electrospray technology is characterized in that the sample treatment method of the analysis method comprises the following steps: weighing about 0.5g of a sample, placing the sample into a 50mL centrifuge tube, adding 20mL of normal hexane, accurately adding 20mL of methanol-water solution after vortex dispersion is uniform, performing ultrasonic extraction for 20min, and centrifuging for 5min at 2500 r/min; discarding the upper n-hexane, accurately sucking 0.1mL of the purified sample extract into a 1.5mL centrifuge tube, adding 0.9mL of methanol-water solution, performing vortex oscillation for 30s, centrifuging at 10000r/min for 5min, sucking the middle clarified liquid, filtering by a filter membrane, and measuring by a high performance liquid chromatography-mass spectrometer;

the high performance liquid chromatography conditions are as follows: chromatographic column: waters Atlantis T3 column, 2.1mm×100mm,3 μm; mobile phase: acetonitrile-5 mmol/L formic acid solution, and gradient elution; flow rate: 0.25mL/min; column temperature: 40 ℃; sample injection amount: 5. Mu.L; the gradient elution conditions are as follows: 0min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;0.5min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;4min, acetonitrile% -5mmol/L formic acid solution% = 95% -5%;5.5min, acetonitrile% -5mmol/L formic acid solution% = 95% -5%;5.51min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;10min, acetonitrile% -5mmol/L formic acid solution% = 10% -90%;

the mass spectrum conditions are as follows: ion source: an electrospray ion source; scanning mode: a positive ion; the detection mode is as follows: monitoring multiple reactions; mass spectrum/mass spectrum reference conditions: air curtain air pressure: 172kPa; electrospray voltage: 5500V; ion source temperature: 650 ℃; atomizer pressure: 379kPa; auxiliary air pressure: 379kPa; the conditions for monitoring the multiple reactions are as follows: ion pair m/z:195/138, declustering voltage: DP/V49, collision cell inlet voltage: EP/V8, collision cell outlet voltage CXP/V:10, collision energy CE/V:25, a step of selecting a specific type of material; ion pair m/z:195/110, declustering voltage: DP/V49, collision cell inlet voltage: EP/V8, collision cell outlet voltage CXP/V:10, collision energy CE/V:23.

2. the analytical method according to claim 1, wherein the liquid chromatography-mass spectrometry/mass spectrometry assay and validation method is as follows: measuring 2.0 mug/L, 5.0 mug/L, 10 mug/L, 20 mug/L and 50 mug/L of serial standard working solutions according to the conditions of high performance liquid chromatography-mass spectrometry/mass spectrometry, drawing a standard working curve by taking the standard solution concentration as an abscissa and the corresponding peak area as an ordinate, and quantifying according to the solution concentration on the standard working curve corresponding to the peak area; the content of the to-be-detected object in the sample solution is in the linear range of the standard curve, and the to-be-detected object is diluted and then analyzed when the content exceeds the linear range; determining a sample processing solution according to the conditions of liquid chromatography-mass spectrometry/mass spectrometry, wherein if the retention time of a chromatographic peak of a substance to be detected in the sample is consistent with that of a standard solution, the deviation is within +/-2.5%, the relative abundance of a qualitative ion pair is expressed by the intensity percentage relative to the abundance of the strongest ion, the relative abundance is consistent with that of a working solution with the concentration equivalent to the standard, and the allowable deviation range of the relative abundance is as follows: the relative ion abundance is more than 50 percent, and the allowable relative deviation is +/-20 percent; the relative ion abundance is more than 20-50%, and the allowable relative deviation is +/-25%; the relative ion abundance is more than 10-20%, and the allowable relative deviation is +/-30%; the relative ion abundance is less than or equal to 10 percent, and the allowable relative deviation is +/-50 percent, so that the existence of the corresponding object to be detected in the sample can be judged.

3. The method of claim 1, wherein the results of the method are calculated and expressed as: calculating the content of 1,3, 7-trimethylxanthine in the sample by using a chromatographic data processor or the following formula 1, and subtracting a blank value from the calculated result:

in formula 1:

X _i the content of the detected component in the sample is expressed in micrograms per kilogram;

c, the concentration of the tested component in the sample liquid obtained by the standard curve is in nanograms per milliliter;

c ₀ -the concentration of the component to be tested in nanograms per milliliter in a blank test obtained from a standard curve;

v-final constant volume of sample solution in milliliters;

m-the mass of the sample represented by the final sample liquid, the unit is gram, and the calculated result retains three valid figures.

4. The method of claim 1, wherein the linear relationship and detection limit of the method is: under the experimental conditions determined by a standard method, a series of standard solutions with different concentrations are taken, the concentration of 1,3, 7-trimethylxanthine is linearly regressed by the response peak area of an instrument, and the result shows that when the concentration of 1,3, 7-trimethylxanthine is in the range of 2.0 mug/L to 50 mug/L, the linear relation is good, the regression equation y=16362.77865x+2845.75302 and the correlation coefficient r= 0.99998; when the concentration of 1,3, 7-trimethylxanthine in the sample exceeds the linear range, the dilution factor of the sample can be properly increased; the detection limit and the quantitative limit are calculated by the signal to noise ratio of 3 times and the signal to noise ratio of 10 times respectively, and the detection limit of the obtained 1,3, 7-trimethyl xanthine is 0.06mg/kg and the quantitative limit is 0.2mg/kg.

5. The assay method of claim 1, wherein the cosmetic comprises: toner, lotion, eye cream, shampoo, eye shadow, blush, and lipstick.