Method for detecting dimethyl sulfone from hair

Abstract

The invention belongs to the technical field of detection, and particularly relates to a method for detecting dimethyl sulfone from hair. The method comprises the following steps: (1) cutting a collected hair sample into pieces to obtain broken hair; (2) adding a digestion solution into the chopped hair, carrying out ultrasonic-assisted extraction, and standing after the ultrasonic-assisted extraction is finished, so as to obtain a digestion extracting solution; (3) adding a mixed solution of ethyl acetate and acetonitrile into the digested extracting solution, and oscillating and cleaning to obtain an oscillating and cleaned extracting solution; (4) centrifuging the extracting solution subjected to oscillation cleaning, then taking an organic layer, blow-drying, taking residues, and dissolving the residues with acetonitrile to obtain a solution to be detected; and (5) carrying out dimethyl sulfone component detection on the liquid to be detected by adopting a liquid chromatography-tandem mass spectrometry method.

Description

Technical Field

[0001] The invention belongs to the field of detection technology, and in particular relates to a method for detecting dimethyl sulfone from hair. Background Art

[0002] Dimethyl sulfone, chemically known as methylsulfonylmethane (Methyl sulfone), is an organic sulfide that appears as white needle-shaped crystals at room temperature and has a certain smell. Dimethyl sulfone is not only a necessary substance for the synthesis of human collagen, but also contained in human skin, hair, nails, bones, muscles and organs. In the medical field, it is widely used to improve joint health, anti-inflammatory, antioxidant and skin repair, and as a sulfur supplement to support keratin and glutathione synthesis. In the field of daily chemicals, its moisturizing, repairing and anti-aging properties are used in skin care products to enhance barrier function and reduce wrinkles. At the same time, it is used in hair care products to strengthen hair quality and reduce breakage, and it is both mild and effective. Some well-known brands will add dimethyl sulfone as an active ingredient to their product formulas.

[0003] Adding dimethyl sulfone to shampoo has attracted more and more attention from manufacturers. After adding dimethyl sulfone to shampoo, the content of dimethyl sulfone in the hair itself is extremely low, but the content of dimethyl sulfone in the hair will increase after using shampoo containing dimethyl sulfone. Detecting dimethyl sulfone in hair after shampooing is one of the research and development work of studying shampoo with added dimethyl sulfone, because it can understand the adhesion of dimethyl sulfone on the hair in shampoo and evaluate the effectiveness of dimethyl sulfone in shampoo formula; by measuring the content of dimethyl sulfone in hair, it can be judged that its role in promoting hair health, improving hair quality, etc., and its safety for long-term use on hair and scalp can be evaluated to prevent possible adverse reactions. At the same time, according to the test results, the amount of dimethyl sulfone added to the shampoo is adjusted, the product formula is optimized, and the effect and quality of the shampoo are improved. Through scientific testing, the actual effect of dimethyl sulfone in shampoo is verified, the publicity of the product is supported, and the trust of consumers is enhanced.

[0004] Through these tests, we can fully understand the effect and safety of dimethyl sulfone in shampoo, and provide a scientific basis for product improvement and optimization. However, the detection of dimethyl sulfone in hair in the current laboratory research and development process is mainly extracted by keratinase, but the extraction is not complete, the impurities have a great impact, and it is difficult to accurately evaluate the accumulation of dimethyl sulfone. Therefore, a method for detecting dimethyl sulfone from hair is urgently needed. Summary of the invention

[0005] The object of the present invention is to provide a method for detecting dimethyl sulfone from hair.

[0006] In order to achieve the above object, the present invention provides the following technical solutions: A method for detecting dimethyl sulfone from hair, comprising the following steps: (1) Cutting the collected hair sample into pieces to obtain broken hair; (2) adding a digestion solution to the broken hair to perform ultrasonic-assisted extraction, and after completion, allowing the extraction to stand to obtain a digestion extraction solution; the digestion solution comprises: 7000-8000 U / L of ficin, 1000-2000 U / L of neutral protease, 3000-4000 U / L of keratinase, 30-35 g / L of Na2HPO4 and 4-6 g / L of NaCl, with the remainder being water; (3) adding a mixture of ethyl acetate and acetonitrile to the digestion extract, and washing by shaking to obtain an extract after shaking and washing; (4) Centrifuge the extract after shaking and washing, then take the organic layer and blow dry it, take the residue and dissolve it in acetonitrile to obtain the test solution; (5) Using liquid chromatography-tandem mass spectrometry to detect the dimethyl sulfone component in the test liquid.

[0007] The present invention uses an extract with a specific composition. Different types of enzymes have different substrate specificities and action sites. They can work synergistically to more effectively degrade complex protein structures, thereby releasing dimethyl sulfone originally wrapped or adsorbed in the protein network, thereby releasing more dimethyl sulfone.

[0008] Furthermore, in the step (1), the collected hair sample is cut into pieces less than 5 mm using medical scissors.

[0009] Furthermore, the conditions for the shaking cleaning in step (3) are an shaking frequency of 20kHz-40kHz, a temperature of 40-50°C, and a time of 30-40min.

[0010] Furthermore, in the step (3), the volume ratio of ethyl acetate to acetonitrile is (1.3-1.6):1.

[0011] The mixed solution of the present invention using a specific ratio can adjust its overall polarity to a certain extent, making it closer to the optimal dissolution condition of dimethyl sulfone and reducing the co-extraction of impurities.

[0012] Furthermore, the centrifugation condition in step (4) is 900-1000 rpm for 3-5 min.

[0013] Furthermore, the conditions for ultrasound-assisted extraction in step (2) are: microwave power 150-250 W, temperature 40-45° C., and extraction time 15-20 min.

[0014] Furthermore, in the step (4), the organic layer is dried with nitrogen at 55°C-60°C.

[0015] Furthermore, in step (2), the solid-liquid ratio of the shredded hair to the digestion solution is 15 mg: 40-50 mL.

[0016] Furthermore, in step (3), the volume ratio of the digestion extract to the mixed solution is 1:(1-3).

[0017] Furthermore, the digestion solution comprises: 7600U / L ficin, 1500U / L neutral protease, 3200U / L keratinase, 31g / L Na2HPO4 and 5.4g / L NaCl, and the balance is water.

[0018] Further, chromatographic column: C18 column, 5 μm, 150 mm × 4.6 mm; Furthermore, the detection conditions of the liquid chromatography tandem mass spectrometry method are: Mobile phase: 0.1% formic acid aqueous solution: acetonitrile = 70:30 (v / v) Flow rate: 1.0 mL / min; Detection wavelength: 210nm; Injection volume: 10 μL; Column temperature: 30℃.

[0019] Compared with the prior art, the advantages and beneficial effects of the present invention are: 1. The present invention uses an extract of specific composition. Different types of enzymes have different substrate specificities and action sites. They can work synergistically to more effectively degrade complex protein structures, thereby releasing dimethyl sulfone originally wrapped or adsorbed in the protein network, thereby releasing more dimethyl sulfone.

[0020] 2. The mixed solution of the specific ratio used in the present invention can adjust its overall polarity to a certain extent, making it closer to the optimal dissolution condition of dimethyl sulfone, and reduce the co-extraction of impurities and improve the detection limit. DETAILED DESCRIPTION

[0021] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0022] Keratinase, enzyme activity 70,000 u / g. Shanghai Yuanye Biotechnology Co., Ltd.

[0023] Neutral protease, enzyme activity 100u / mg. Shanghai Yuanye Biotechnology Co., Ltd.

[0024] Fig enzyme, enzyme activity 100u / mg. Shanghai Yuanye Biotechnology Co., Ltd.

[0025] Hair sample 1: from volunteers who used Korean brand LYU shampoo for 3 months.

[0026] Hair sample 2 was from a volunteer who used the German brand Schwarzkopf shampoo for 3 months.

[0027] Example 1 This embodiment provides a method for detecting dimethyl sulfone from hair, comprising the following steps: (1) Cut the collected hair sample into pieces less than 5 mm using medical scissors to obtain broken hair; (2) adding a digestion solution to the broken hair, wherein the solid-liquid ratio of the broken hair to the digestion solution is 15 mg:45 mL; performing ultrasonic-assisted extraction, wherein the conditions for the ultrasonic-assisted extraction are: microwave power of 200 W, temperature of 42° C., and extraction time of 18 min; after the extraction, standing the mixture to obtain a digestion extract; the digestion solution comprises: 7600 U / L of fig protease, 1500 U / L of neutral protease, 3200 U / L of keratinase, 31 g / L of Na2HPO4, and 5.4 g / L of NaCl, with the remainder being water.

[0028] (3) Add a mixed solution of ethyl acetate and acetonitrile to the digestion extract, the volume ratio of the digestion extract to the mixed solution is 1:2; the volume ratio of ethyl acetate to acetonitrile is 1.5:1; shake and wash to obtain an extract after shaking and washing; the shaking and washing conditions are an shaking frequency of 30 kHz, a temperature of 45°C, and a time of 35 min.

[0029] (4) Centrifuge the extract after shaking and washing at 950 rpm for 4 min; then take the organic layer and blow dry it with nitrogen at 58°C, take the residue and dissolve it with acetonitrile to obtain the test solution; (5) Using liquid chromatography-tandem mass spectrometry to detect the dimethyl sulfone component in the test liquid.

[0030] The detection conditions of the liquid chromatography-tandem mass spectrometry method were: Chromatographic column: C18 column, 5μm, 150mm×4.6mm; Mobile phase: 0.1% formic acid aqueous solution: acetonitrile = 70:30 (v / v) Flow rate: 1.0 mL / min; Detection wavelength: 210nm; Injection volume: 10 μL; Column temperature: 30℃.

[0031] Example 2 This embodiment provides a method for detecting dimethyl sulfone from hair, comprising the following steps: (1) Cut the collected hair sample into pieces less than 5 mm using medical scissors to obtain broken hair; (2) adding a digestion solution to the broken hair, wherein the solid-liquid ratio of the broken hair to the digestion solution is 15 mg:45 mL; performing ultrasonic-assisted extraction, wherein the conditions for the ultrasonic-assisted extraction are: microwave power 200 W, temperature 42° C., and extraction time 18 min; after completion, standing the mixture to obtain a digestion extract; the digestion solution comprises: 8000 U / L of fig protease, 1000 U / L of neutral protease, 4000 U / L of keratinase, 30 g / L of Na2HPO4, and 6 g / L of NaCl, with the remainder being water; (3) Add a mixed solution of ethyl acetate and acetonitrile to the digestion extract, the volume ratio of the digestion extract to the mixed solution is 1:2; the volume ratio of ethyl acetate to acetonitrile is 1.5:1; shake and wash to obtain an extract after shaking and washing; the shaking and washing conditions are an shaking frequency of 30 kHz, a temperature of 45°C, and a time of 35 min.

[0032] (4) Centrifuge the extract after shaking and washing at 950 rpm for 4 min; then take the organic layer and blow dry it with nitrogen at 58°C, take the residue and dissolve it with acetonitrile to obtain the test solution; (5) Using liquid chromatography-tandem mass spectrometry to detect the dimethyl sulfone component in the test liquid.

[0033] The detection conditions of the liquid chromatography-tandem mass spectrometry method were: Chromatographic column: C18 column, 5μm, 150mm×4.6mm; Mobile phase: 0.1% formic acid aqueous solution: acetonitrile = 70:30 (v / v) Flow rate: 1.0 mL / min; Detection wavelength: 210nm; Injection volume: 10 μL; Column temperature: 30℃.

[0034] Example 3 This embodiment provides a method for detecting dimethyl sulfone from hair, comprising the following steps: (1) Cut the collected hair sample into pieces less than 5 mm using medical scissors to obtain broken hair; (2) adding a digestion solution to the broken hair, wherein the solid-liquid ratio of the broken hair to the digestion solution is 15 mg:45 mL; performing ultrasonic-assisted extraction, wherein the conditions for the ultrasonic-assisted extraction are: microwave power 200 W, temperature 42° C., and extraction time 18 min; after completion, allowing the extraction to stand to obtain a digestion extract; the digestion solution comprises: 7000 U / L of fig protease, 2000 U / L of neutral protease, 3000 U / L of keratinase, 35 g / L of Na2HPO4, and 4 g / L of NaCl, with the remainder being water; (3) Add a mixed solution of ethyl acetate and acetonitrile to the digestion extract, the volume ratio of the digestion extract to the mixed solution is 1:2; the volume ratio of ethyl acetate to acetonitrile is 1.5:1; shake and wash to obtain an extract after shaking and washing; the shaking and washing conditions are an shaking frequency of 30 kHz, a temperature of 45°C, and a time of 35 min.

[0035] (4) Centrifuge the extract after shaking and washing at 950 rpm for 4 min; then take the organic layer and blow dry it with nitrogen at 58°C, take the residue and dissolve it with acetonitrile to obtain the test solution; (5) Using liquid chromatography-tandem mass spectrometry to detect the dimethyl sulfone component in the test liquid.

[0036] The detection conditions of the liquid chromatography-tandem mass spectrometry method were: Chromatographic column: C18 column, 5μm, 150mm×4.6mm; Mobile phase: 0.1% formic acid aqueous solution: acetonitrile = 70:30 (v / v) Flow rate: 1.0 mL / min; Detection wavelength: 210nm; Injection volume: 10 μL; Column temperature: 30℃.

[0037] Comparative Example 1 The difference between this comparative example and Example 1 is that the digestion solution includes: 12300 U / L keratinase, 31 g / L Na2HPO4 and 5.4 g / L NaCl, and the balance is water.

[0038] Comparative Example 2 The difference between this comparative example and Example 1 is that the digestion solution includes: 9100 U / L of ficin, 3200 U / L of keratinase, 31 g / L of Na2HPO4 and 5.4 g / L of NaCl, and the balance is water.

[0039] Comparative Example 3 The difference between this comparative example and Example 1 is that the digestion solution includes: 4100U / L ficin, 4100U / L neutral protease, 4100U / L keratinase, 31g / LNa2HPO4 and 5.4g / LNaCl, and the balance is water. Comparative Example 4 The difference between this comparative example and Example 1 is that the ultrasonic-assisted extraction in step (2) is replaced by static extraction at 25° C. for 18 min.

[0040] Comparative Example 5 The difference between this comparative example and Example 1 is that the step (3) is to add ethyl acetate to the digestion extract, and the volume ratio of the digestion extract to ethyl acetate is 1:2. Performance Testing Using dimethyl sulfone (analytical grade), a standard curve of dimethyl sulfone was prepared according to the methods of Example 1 and Comparative Examples 1-5, and the detection limit of dimethyl sulfone was determined. The concentration of dimethyl sulfone in the hair sample was detected.

[0041] The results are shown in Table 1.

[0042] Table 1 Performance test results

[0043] It can be seen from the above performance test results that the dimethyl sulfone detected in Examples 1-3 in the same sample is at a higher concentration.

[0044] The different enzyme compositions in the digestion solutions in Comparative Examples 1-3 resulted in a decrease in the detected concentration of dimethyl sulfone.

[0045] In Comparative Example 4, the ultrasonic-assisted extraction was replaced by static extraction at 25° C. for 18 min, resulting in a decrease in the detected concentration of dimethyl sulfone.

[0046] In comparative example 5, step (3) is to add ethyl acetate to the digestion extract, resulting in a decrease in the detected concentration of dimethyl sulfone.

[0047] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for detecting dimethyl sulfone from hair, characterized in that: The following steps are involved: (1) Cutting the collected hair sample into pieces to obtain broken hair; (2) adding a digestion solution to the broken hair to perform ultrasonic-assisted extraction, and after completion, allowing the extraction to stand to obtain a digestion extraction solution; the digestion solution comprises: 7000-8000 U / L of ficin, 1000-2000 U / L of neutral protease, 3000-4000 U / L of keratinase, 30-35 g / L of Na2HPO4 and 4-6 g / L of NaCl, with the remainder being water; (3) adding a mixture of ethyl acetate and acetonitrile to the digestion extract, and washing by shaking to obtain an extract after shaking and washing; (4) Centrifuge the extract after shaking and washing, take the organic layer and blow dry, take the residue and dissolve it in acetonitrile to obtain the test solution; (5) Using liquid chromatography-tandem mass spectrometry to detect the dimethyl sulfone component in the test liquid.

2. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: In the step (1), the collected hair sample is cut into pieces less than 5 mm using medical scissors.

3. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: The conditions for the shaking cleaning in step (3) are an shaking frequency of 20 kHz-40 kHz, a temperature of 40-50° C., and a time of 30-40 min.

4. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: The volume ratio of ethyl acetate to acetonitrile in step (3) is (1.3-1.6):

1.

5. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: The centrifugation condition in step (4) is 900-1000 rpm for 3-5 min.

6. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: The conditions for ultrasound-assisted extraction in step (2) are: microwave power 150-250W, temperature 40-45°C, and extraction time 15-20min.

7. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: In the step (4), the organic layer is dried by blowing with nitrogen at 55°C-60°C.

8. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: In step (2), the solid-liquid ratio of the shredded hair to the digestion solution is 15 mg: 40-50 mL.

9. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: In the step (3), the volume ratio of the digestion extract to the mixed solution is 1:(1-3).

10. The method for detecting dimethyl sulfone from hair according to claim 1, characterized in that: The digestion solution comprises: 7600U / L of ficin, 1500U / L of neutral protease, 3200U / L of keratinase, 31g / L of Na2HPO4 and 5.4g / L of NaCl, and the balance is water.

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