Method for detecting content of glycosylglycerol in cosmetics by using high performance ion chromatography

Abstract

The invention belongs to the technical field of cosmetic detection, and relates to a method for detecting the content of glycosylglycerol in cosmetics by using high performance ion chromatography. The invention discloses a method for detecting the content of glycosylglycerol in cosmetics by using high performance ion chromatography. The method comprises the following steps: step 1, preparing a standard solution; step 2, pre-treating a sample; step 3, performing high performance ion chromatography-electrochemical testing; the method has the advantages of being high in sensitivity, good in selectivity, low in detection cost, environmentally friendly, safe, simple in pretreatment and the like, and the ion chromatography is a common detection instrument which is commonly equipped in a basic laboratory and is easy to popularize in actual production quality detection. The method disclosed by the invention is suitable for determining the glycosylglycerol in the cosmetics with relatively complex matrixes, has a good linear relation, is high in accuracy, good in repeatability and strong in specificity, and provides technical support for research and supervision of the glycosylglycerol in the cosmetics.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic testing technology, specifically relating to a method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography. Background Technology

[0002] Glucosylglycerol is a natural osmotic pressure-resistant molecule composed of glycerol and glucose molecules linked by glycosidic bonds. It protects the skin, possesses water-locking power similar to hyaluronic acid, and has a smaller molecular weight, resulting in better absorption and a non-sticky feel. In the cosmetics industry, it is commonly used as a moisturizer in skincare products such as masks, toners, lotions, and cleansers. In recent years, the number of cosmetic products containing glutosylglycerol has been rapidly increasing.

[0003] The main synthetic methods for glycerol glucosides include chemical synthesis, fermentation, and enzymatic catalysis. Chemical synthesis results in low conversion rates and numerous byproducts, leading to more impurities and fewer active ingredients. Fermentation offers the advantage of producing bioactive components, but its technological barriers lie in the screening of bacterial strains and subsequent separation and purification. Enzymatic catalysis may leave substrate residues such as glycerol, fructose byproducts, and phosphate impurities, but its high catalytic efficiency and low catalytic temperature make it a promising method. Currently, the sources of glycerol glucosides on the market are complex, quality control is unreliable, and the use of inferior or counterfeit products is rampant. For example, sensory-similar glycerol is used as a raw material for glycerol glucosides. This results in significant discrepancies between the actual and labeled content of products, highlighting the urgent need for quality improvement. Therefore, developing accurate, efficient, and specific methods for detecting glycerol glucoside content is imperative to improve product quality control, enhance the scientific nature of regulation, protect consumer rights, and promote the upgrading of the cosmetics industry.

[0004] Chinese patent CN 118759092A uses high-performance liquid chromatography-mass spectrometry (HPLC-MS / MS) to detect glycerol glucosides in cosmetics. While this method boasts short detection time and high sensitivity, it requires a costly mass spectrometer, making its widespread application in routine testing difficult. High-performance ion chromatography (HPLC) is one of the effective methods for carbohydrate detection, often referred to as a "carbohydrate expert." However, the literature (Liang Wenhui, Fa Yun, Wang Minglin. Determination of sucrose and glycerol glucosides in cyanobacterial cell culture medium by high-performance anion exchange chromatography-integrated pulse amperometry. Chemical Analysis and Metrology, 2014, 23(Z1):9-12.) uses HPLC to detect glycerol glucosides, with cyanobacterial cell culture medium as the sample matrix, and the sample pretreatment is relatively complex. Therefore, a simplified sample pretreatment method using HPLC to detect glycerol glucoside content in cosmetics is needed. Summary of the Invention

[0005] In view of the above-mentioned technical problems, the present invention provides a method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography. This method has the advantages of high sensitivity, good selectivity, low detection cost, environmental safety, and simple pretreatment, providing diversity for the detection of glycerol glucoside content in cosmetics.

[0006] A method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography includes the following steps: Step 1: Preparation of standard solution; Step 2, Sample pretreatment; Step 3, High Performance Ion Chromatography-Electrochemical Testing: Column: Dionex CarboPac MA1IC column (4mm × 250mm) equipped with Dionex CarboPac MA1Guard column (4mm × 50mm); Eluent: 200mmol / L sodium hydroxide solution, flow rate 0.4mL / min; Injection volume: 25μL; Column temperature: 30℃; Detection chamber temperature: 30℃; Integrating pulse amperometric detection; Gold working electrode; Ag / AgCl reference electrode; Step 4: Results Statistics and Analysis.

[0007] Further, step 1 specifically involves: accurately weighing 0.01 g of glycerol glucoside standard, placing it in a 10 mL volumetric flask, dissolving it in ultrapure water and making up to volume to prepare a 1000 mg / L glycerol glucoside standard stock solution; then diluting it sequentially with ultrapure water to prepare a series of standard solutions with mass concentrations of 0.5, 1, 5, 10, 20, 50, and 100 mg / L.

[0008] Further, step 2 specifically involves: accurately weighing 0.2 g of the mixed sample, adding 3 mL of water, vortexing to mix, diluting to 10 mL with water, sonicating for 20 min, adding 1 mL of dichloromethane, gently shaking, vortexing to mix, letting stand for 10 min, centrifuging at 10000 rpm for 15 min, filtering through a 0.22 μm organic membrane, and taking the filtrate as the test solution.

[0009] Furthermore, in step 4, standard series solutions and test sample solutions are measured and injected into the ion chromatograph, the chromatograms are recorded, and the peak areas are calculated using the standard curve method. The calculation formula is as follows: ω=ρ×V×D×100% / 1000000 / m In the formula: ω—the content of glycerol glucoside in cosmetics, %; ρ—The mass concentration of glycerol glucoside obtained from the standard curve, in μg / mL; V—Sample volume at final volume, mL; D—Sample dilution factor (1 if undiluted); m—sample amount, in grams.

[0010] Compared with the prior art, the beneficial effects of the present invention are as follows.

[0011] 1. High sensitivity: This invention can detect glycerol glucoside in cosmetics with a detection concentration of 7.5 ug / g and a quantitative concentration of 25 ug / g. In addition to being used for the quantitative detection of glycerol glucoside, it can also be used for the trace detection of glycerol glucoside.

[0012] 2. High selectivity: The electrochemical detector used in this invention only responds to substances with electroactive groups, such as sugars and alcohols, and can effectively eliminate interference from non-electroactive substances, such as salts, amino acids, and pigments, making it particularly suitable for the detection of cosmetics with complex matrices.

[0013] 3. Low testing cost and environmentally friendly and safe: The extraction solvent of this invention is water and the rinsing solution is an aqueous solution of sodium hydroxide, which reduces the use of organic reagents and the discharge of harmful waste liquid, thus not only reducing the testing cost but also making it environmentally friendly and safe.

[0014] 4. Simple pretreatment: The pretreatment of this invention uses water as the extraction solvent. After ultrasonication, centrifugation and filtration, it can be detected. The operation is simple and quick, without the need for complicated pretreatment methods, which greatly improves the efficiency of production quality detection.

[0015] 5. Easy to promote: The ion chromatography method used in this invention is a commonly used detection instrument that is widely equipped in grassroots laboratories, and it is easy to promote in actual production quality testing.

[0016] 6. Providing technical support: The method of this invention is applicable to the determination of glycerol glucosides in cosmetics with complex matrices. It has good linearity, high accuracy, and good repeatability, providing technical support for the research and regulation of glycerol glucosides in cosmetics. Attached Figure Description

[0017] Figure 1 A graph showing the relationship between eluent concentration and theoretical plate number.

[0018] Figure 2 Chromatograms of glycerol and glycerol glucoside.

[0019] Figure 3 Ultrasonic time-content change curve.

[0020] Figure 4 Chromatogram of an aqueous emulsion sample solution.

[0021] Figure 5 Chromatogram of a cream or ointment sample solution. Detailed Implementation

[0022] The claims of the present invention will be further described in detail below with reference to specific embodiments, but this does not constitute any limitation on the present invention. Any limited modifications made by any person within the scope of the claims of the present invention shall still be within the scope of protection of the claims of the present invention.

[0023] A method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography includes the following steps: Step 1: Preparation of standard solutions: Accurately weigh 0.01 g of glycerol glucoside standard, place it in a 10 mL volumetric flask, dissolve it in ultrapure water and dilute to volume to prepare a 1000 mg / L glycerol glucoside standard stock solution; then dilute it with ultrapure water to prepare a series of standard solutions with mass concentrations of 0.5, 1, 5, 10, 20, 50 and 100 mg / L respectively. Step 2, Sample pretreatment: Accurately weigh 0.2g of the mixed sample, add 3mL of water, vortex to mix, dilute to 10mL with water, sonicate for 20min, add 1mL of dichloromethane, gently shake, vortex to mix, let stand for 10min, centrifuge at 10000rpm for 15min, filter through a 0.22μm organic membrane, and take the filtrate as the test solution; Step 3, High Performance Ion Chromatography-Electrochemical Testing: Column: Dionex CarboPac MA1IC column (4mm × 250mm) equipped with Dionex CarboPac MA1Guard column (4mm × 50mm); Eluent: 200mmol / L sodium hydroxide solution, flow rate 0.4mL / min; Injection volume: 25μL; Column temperature: 30℃; Detection chamber temperature: 30℃; Integrating pulse amperometric detection; Gold working electrode; Ag / AgCl reference electrode; Step 4: Result Statistics and Analysis. Measure out standard series solutions and test sample solutions respectively, inject them into the ion chromatograph, record the chromatograms, and calculate the results based on peak area using the standard curve method. The calculation formula is: ω=ρ×V×D×100% / 1000000 / m In the formula: ω—the content of glycerol glucoside in cosmetics, %; ρ—The mass concentration of glycerol glucoside obtained from the standard curve, in μg / mL; V—Sample volume at final volume, mL; D—Sample dilution factor (1 if undiluted); m—sample amount, in grams.

[0024] Example 1: Optimization of chromatographic conditions.

[0025] Currently, there are instances in the market where glycerol, with its similar sensory properties, is used as a substitute for glyceryl glucoside in cosmetics, and glycerol is frequently added to moisturizing cosmetics. Therefore, this study investigated the peak elution effects of sodium hydroxide solutions with eluent concentrations of 50, 100, 200, 400, 600, 800, and 1000 mmol / L, and also examined the separation of glycerol and glyceryl glucoside. The bar chart is shown below, with eluent concentration on the x-axis and the theoretical plate number of the glyceryl glucoside peak on the y-axis. Figure 1 As shown. By Figure 1 It can be seen that when the eluent concentration is 200 mmol / L, the theoretical plate number of the glycerol glucoside peak is the highest (7209), and glycerol and glycerol glucoside can be separated within 20 min, with good separation between the two peaks (10.79). The separation of glycerol and glycerol glucoside is as follows: Figure 2 .

[0026] Example 2: Sample pretreatment optimization.

[0027] 1. Selection of extraction method and solvent Method 1: Accurately weigh 0.2 g of the mixed sample, add 3 mL of water, vortex to mix, dilute to 10 mL with water, sonicate for 20 min, centrifuge (10000 rpm, 15 min), filter (organic membrane, 0.22 μm), and take the filtrate as the test solution. Method 2: Accurately weigh 0.2 g of the mixed sample, add 2 mL of water and 1 mL of saturated sodium chloride, vortex to mix, dilute to 10 mL with water, sonicate for 20 min, add 1 mL of dichloromethane, gently shake, vortex to mix, let stand for 10 min, centrifuge (10000 rpm, 15 min), filter (organic membrane, 0.22 μm), and take the filtrate as the test solution; Method 3: Accurately weigh 0.2 g of the mixed sample, add 1 mL of n-hexane, vortex to mix, dilute to 10 mL with water, sonicate for 20 min, let stand for 10 min, centrifuge (10000 rpm, 15 min), filter (organic membrane, 0.22 μm), and take the filtrate as the test solution. Method 4: Accurately weigh 0.2 g of the mixed sample, add 3 mL of water, vortex to mix, dilute to 10 mL with water, sonicate for 20 min, add 1 mL of dichloromethane, gently shake, vortex to mix, let stand for 10 min, centrifuge (10000 rpm, 15 min), filter (organic membrane, 0.22 μm), and take the filtrate as the test solution.

[0028] After extraction using Method 1, the precipitate is distributed at the bottom of the centrifuge tube, and the solution is clear. Liquid samples are easy to filter, while emulsions and pastes are more difficult to filter. After extraction using Method 2, the precipitate is distributed at the bottom of the solution, and flocculent matter is produced, increasing the difficulty of filtration. After extraction using Method 3, the solution separates into layers: the aqueous phase is at the bottom and is relatively clear, with impurities precipitated at the bottom of the centrifuge tube; the organic phase (n-hexane) is at the top, making it difficult to aspirate and filter the solution. After extraction using Method 4, the solution separates into layers: the aqueous phase is at the top and the solution is clear; the organic phase (dichloromethane) is at the bottom, with the precipitate distributed at the bottom of the centrifuge tube, making the solution easier to aspirate during filtration. Compared to the previous three methods, the aqueous phase solution is the clearest and easiest to filter. In conclusion, Method 4, using water as the extraction solvent and dichloromethane to remove impurities, is the optimal extraction method. The role of dichloromethane is twofold: firstly, after interfering substances such as emulsifiers and lipids enter the organic phase dichloromethane layer, their influence can be eliminated, and the aqueous phase becomes clearer; secondly, the aqueous phase is distributed in the upper layer and the organic phase dichloromethane is distributed in the lower layer, which is more conducive to the absorption of sample solution.

[0029] 2. Selection of centrifugation time Five 0.2g portions of sample were accurately weighed and prepared according to step 2. The centrifugation times were set to 5, 10, 15, 20, and 25 min, respectively. After centrifugation, it was observed that the amount of precipitate no longer increased after 15 min. Therefore, 15 min was selected as the optimal centrifugation time.

[0030] 3. Selection of extraction time Accurately weigh 0.2g of sample into 5 portions, prepare samples according to step 2, and set the ultrasonic time to 5, 10, 20, 30, and 40 min respectively. Determine the content under the chromatographic conditions of step 3. Plot an ultrasonic time-content curve with ultrasonic time (min) on the x-axis and content (%) on the y-axis, as shown below. Figure 3 As shown. From Figure 3 As can be seen, the sample content no longer changes when the ultrasonic time reaches 20 minutes. Therefore, the ultrasonic time of 20 minutes is selected.

[0031] Example 3: Methodology Establishment.

[0032] 1. Examining the linear range 25 μL of each glycerol glucoside standard solution was injected sequentially, and the chromatograms were recorded. A linear equation was fitted using the mass concentration of glycerol glucoside (X) and the peak area of ​​glycerol glucoside (Y). The linear equation was Y = 2.1921X + 1.5514, with a correlation coefficient of 0.9998, which is greater than 0.999, indicating a good linear relationship.

[0033] 2. Limit of detection and limit of quantitation Accurately weigh 0.2 g of blank sample, add 2 mL of water, and accurately add an appropriate amount of standard solution. Vortex to mix well, and process according to the sample preparation method in step 2. Determine the signal-to-noise ratio (SNR) under the chromatographic conditions in step 3. Using the mass concentration of glycerol glucoside at a SNR (S / N) of 3 as the detection concentration, the detection concentration of this method is determined to be 7.5 μg / g.

[0034] As described above, the mass concentration of glycerol glucoside with a signal-to-noise ratio (S / N) of 10 was used as the quantitative concentration, and the quantitative concentration of this method was determined to be 25 μg / g.

[0035] 4. Precision A 5 μg / mL glycerol glucoside standard solution was injected six times consecutively, and the peak area RSD (n=6) was 0.7%, indicating that the instrument has good precision.

[0036] 5. Recovery rate and repeatability Six 0.2 g portions of liquid (emulsion) and six 0.2 g portions of solid (cream) samples were accurately weighed. An appropriate amount of glycerol glucoside standard solution was added to each sample, and the mixture was vortexed and mixed thoroughly. The samples were then processed according to step 2, and the chromatographic conditions of step 3 were used for determination. The recovery rate and RSD values ​​were calculated and shown in Table 1. Table 1 shows that the method has high accuracy and good repeatability.

[0037] Table 1 Spike recoveries and their RSD values ​​(n=6).

[0038] 6. Stability test A blank sample was added to a standard solution to achieve a concentration of 10 mg / L. The sample was then processed according to step 2 and allowed to stand at room temperature for 0, 2, 4, 8, 12, 16, 24, 36, and 48 hours. The peak area was then measured by injection according to the method. The relative standard deviation (RSD) of the peak area within 48 hours was calculated to be 0.6%, indicating that the test sample solution remained stable within 48 hours.

[0039] Example 4: Determination of glycerol glucoside in commercially available samples.

[0040] Fourteen batches of commercially available cosmetics, all labeled as containing glycerol glucoside, were tested using the method described above. The results are shown in Table 2. All purchased samples were labeled as containing glycerol glucoside; glycerol glucoside was detected in 9 batches, while it was not detected in the remaining 5 batches. Typical chromatograms are shown below. Figure 4 (Emulsions) Figure 5 (Ointments and creams).

[0041] Table 2. Results of glycerol glucoside content determination in 14 commercially available samples Note: ND: Not detected.

[0042] This study tested products on the market that were labeled as containing glycerol glucoside. The results showed that the actual content of glycerol glucoside in some products was significantly different from the labeled value, fully demonstrating the importance of the detection method of this invention in the industry. The method of this invention has been validated, and the results obtained all meet the requirements of the "Technical Specification for Validation of Detection Methods for Prohibited and Restricted Substances in Cosmetics" and the "Management Procedures for Supplementary Testing Methods for Cosmetics".

Claims

1. A method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography, characterized in that, Includes the following steps: Step 1: Preparation of standard solution; Step 2, Sample pretreatment; Step 3, High Performance Ion Chromatography-Electrochemical Testing: Column: Dionex CarboPac MA1IC column equipped with Dionex CarboPac MA1Guard column; Eluent: 200 mmol / L sodium hydroxide solution, flow rate 0.4 mL / min; Injection volume: 25 μL; Column temperature: 30℃; Detection chamber temperature: 30℃; Integrated pulse amperometric detection; Gold working electrode; Ag / AgCl reference electrode; Step 4: Results Statistics and Analysis.

2. The method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography as described in claim 1, characterized in that, Step 1 specifically involves: accurately weighing 0.01 g of glycerol glucoside standard, placing it in a 10 mL volumetric flask, dissolving it in ultrapure water and making up to volume to prepare a 1000 mg / L glycerol glucoside standard stock solution; then diluting it sequentially with ultrapure water to prepare a series of standard solutions with mass concentrations of 0.5, 1, 5, 10, 20, 50, and 100 mg / L.

3. The method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography as described in claim 1, characterized in that, Step 2 specifically involves: accurately weighing 0.2g of the mixed sample, adding 3mL of water, vortexing to mix, diluting to 10mL with water, sonicating for 20min, adding 1mL of dichloromethane, shaking, vortexing to mix, letting stand for 10min, centrifuging at 10000rpm for 15min, filtering through a 0.22μm organic membrane, and taking the filtrate as the test solution.

4. The method for detecting the content of glycerol glucoside in cosmetics using high-performance ion chromatography as described in claim 1, characterized in that, In step 4, standard series solutions and test sample solutions are measured and injected into the liquid chromatograph, chromatograms are recorded, and peak areas are calculated using the standard curve method. The calculation formula is as follows: ω=ρ×V×D×100% / 1000000 / m In the formula: ω—the content of glycerol glucoside in cosmetics, %; ρ—The mass concentration of glycerol glucoside obtained from the standard curve, in μg / mL; V—Sample final volume, mL; D—sample dilution factor; 1 if undiluted. m—sample amount, in grams.

Images