- Email: [email protected]
Simultaneous determination of seven prohibited substances in cosmetic products by liquid chromatography–tandem mass spectrometry
Chinese Chemical Letters 24 (2013) 509–511
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters journal homepage: www.elsevier.com/locate/cclet
Original article
Simultaneous determination of seven prohibited substances in cosmetic products by liquid chromatography–tandem mass spectrometry Cai-Sheng Wu, Ying Jin, Jin-Lan Zhang *, Yan Ren, Zhi-Xin Jia Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China
A R T I C L E I N F O
A B S T R A C T
Article history: Received 27 January 2013 Received in revised form 1 March 2013 Accepted 8 March 2013 Available online 18 April 2013
In this paper, we developed and validated a simple, sensitive, and selective high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method to identify and measure the following prohibited substances that may be found in cosmetic products: minoxidil, hydrocortisone, spironolactone, estrone, canrenone, triamcinolone acetonide and progesterone. Chromatographic separation was performed on a Waters Symmetry C18 (100 mm 2.1 mm, 3.5 mm particle size) with a gradient elution system composed of 0.2% (v/v) formic acid aqueous solution and methanol containing 0.2% (v/v) formic acid at a flow rate of 0.3 mL/min. The substances were detected using a triple quadrupole mass spectrometer in the multiple reaction monitoring mode with an electrospray ionization source. All of the calibration curves showed good linearity (r > 0.999) within the tested concentration ranges. The limit of detection was <25 pg. The relative standard deviations for intraday precision for each of the prohibited substances were <3.5% at two concentration levels (2 mg/g, 10 mg/g). The relative recovery rate for each of the prohibited substances ranged from 91.8% to 111% at three concentration levels (0.1 mg/g, 2 mg/g, 10 mg/g), including the limit of quantification. In conclusion, we have developed and validated a method that can identify seven prohibited substances in cosmetic products. ß 2013 Jin-Lan Zhang. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: High-performance liquid chromatography– tandem mass spectrometry (HPLC–MS/MS) Cosmetic products Prohibited substances Minoxidil Steroids
1. Introduction Baldness, alopecia, and acne are common skin and hair diseases that may adversely influence the patient’s social relationships [1–3]. For reasons of safety and ease of use, cosmetic products are widely used to manage these conditions. However, for the sake of profit and to enhance short-term efficacy, some manufacturers illegally add synthetic drugs to their cosmetic products. According to previous reports [3–6], minoxidil, hydrocortisone, spironolactone, estrone, canrenone, triamcinolone acetonide, and progesterone are among the most common topically active anti-alopecia, anti-baldness, and anti-acne drugs. Therefore, we focused on these substances in this study. Up to now, many articles have reported methods to detect minoxidil or steroids in drug formulations [7,8], environmental samples [8], food products [9], and biological samples [10–12]. However, few procedures for the detection of minoxidil or steroids in cosmetic products have been reported to date [3,13–16]. Furthermore, minoxidil was detected in a pure Chinese herbal medicine shampoo (Zhangguang 101) in 2010. Therefore, methods that can rapidly and simultaneously screen for,
* Corresponding author. E-mail address: [email protected] (J.-L. Zhang).
and quantify, prohibited substances in cosmetic products are urgently required. In this paper, we established and validated a simple and rapid high-performance liquid chromatography– tandem mass spectrometry (HPLC–MS/MS) method capable of simultaneously determining seven prohibited substances in cosmetic products. We applied this method to analyze 37 different cosmetic products used for prophylaxis and treatment of hair loss and acne. 2. Experimental 2.1. Instruments and materials The assay was performed on an Agilent 6410A triple quadrupole LC–MS system (Agilent Technologies, Santa Clara, CA, USA) consisting of an Agilent 1200 RRLC system (Agilent Corporation) connected to a triple quadrupole MS analyzer with an electrospray ionization (ESI) interface that could be used in either positive or negative ionization modes. A MassHunter workstation was used to control the LC–MS and for data acquisition (Agilent Corporation). LC/MS reagent grade acetonitrile was obtained from Mallinckrodt Baker, Inc. (Phillipsburg, NJ, USA). Deionized water was purified using a Milli-Q1 academic water purification system (Merck Millipore, Billerica, MA, USA). Analytical grade formic acid
1001-8417/$ – see front matter ß 2013 Jin-Lan Zhang. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. http://dx.doi.org/10.1016/j.cclet.2013.03.037
[(Fig._1)TD$IG]
C.-S. Wu et al. / Chinese Chemical Letters 24 (2013) 509–511
510
Fig. 1. Chemical structures of seven prohibited substances.
was obtained from Merck Inc. (Darmstadt, Germany). Analytical grade methanol was purchased from Beijing Chemical Corp. (Beijing, China). Reference standards of the seven prohibited substances were obtained from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). Their structures are shown in Fig. 1. Thirty-seven different cosmetic products used for prophylaxis and treatment of hair loss and acne were obtained from two cosmetic shops and an internet-based supplier (www.taobao.com). 2.2. LC–ESI-MS analytical conditions Chromatographic separation was performed using a Waters Symmetry C18 (100 mm 2.1 mm, 3.5 mm particle size) column. The mobile phase, consisting of solvent A (0.2% (v/v) formic acid aqueous solution) and solvent B (methanol containing 0.2% (v/v) formic acid), was delivered at a flow rate of 0.3 mL/min. The gradient elution was started with 5% B, increased to 25% B in 2 min, then to 55% B in 2 min, to 80% B in 7 min, then to 90% B in 4 min. Then the column was flushed with 5% B for 8 min. The column temperature was maintained at 30 8C and the injection volume was 5 mL. The optimum operating parameters of the ESI interface in the positive mode were as follows: nebulizer, 40 psi; dry gas, 8 L/min; dry temp, 325 8C, capillary voltage, 4000 V; and Delta EMV, 300 V. The LC eluent flow from 0 to 3.5 min was not introduced to the MS for data acquisition. Each substance was detected in the multiple reaction monitoring (MRM) mode, using the parameters listed in Table 1. 2.3. Sample preparation A sample of lotion (1 mL) or cream (1 g) was accurately weighed into a 10 mL colorimetric cylinder. Then, 1 mL of a saturated
solution of sodium chloride was added to avoid emulsification and the mixture was vortexed for 30 s. Acetonitrile was added followed by vortexing. The cosmetic extracts were ultrasonicated for 30 min at 25 8C and vortexed for 60 s. After centrifugation at 4500 rpm for 10 min, an aliquot of the supernatant was filtered through a 0.45 mm filter and injected into the HPLC–MS/MS system. 3. Results and discussion Optimization of the sample preparation method was essential to obtain a good accuracy and reliability for quantification. According to the previous reports [3,8–16], several methods, including solid-phase extraction (SPE) and liquid–liquid extraction (LLE), were used to prepare samples for analysis of minoxidil or steroids. For cosmetic products, there are different formulas and the ingredients of same formula from different factories. After tried, it was difficult to find a solid phase of SPE that was suitable for all kind products. In addition, some ingredients of cosmetic products may block the SPE cartridge. The LLE method was also tried. However, emulsion was readily formed and the accuracy and reliability of quantification was not satisfied. After optimizing the pretreatment conditions, the same amount of saturated sodium chloride solution was used to break the emulsification of cosmetic products and to reduce extraction process emulsification effects. Finally, by comparing the processing of more than 20 different products, we found that methanol was more likely to cause emulsification than acetonitrile, therefore acetonitrile was used as the extraction solvent to simplify the procedure and maintain its accuracy. The total run time for each sample was just 26 min, and good separation of the seven prohibited substances was achieved under the specified gradient HPLC–MS/MS conditions. There was no interference of the retention time for any of the prohibited
Table 1 Optimized mass parameters for the seven prohibited substances. No.
Analyte
Precursor ion (m/z)
Fragmentor (V)
1
Minoxidil
210.0
110 110
2
Hydrocortisone
363.1
125 125
3
Spironolactone
341.1
147 165
107.1a 90.9
35 65
4
Estrone
271.2
105 105
253.1a 132.8
5 22
5
Canrenone
341.1
147 165
107.1a 90.9
35 65
6
Triamcinolone acetonide
477.1
100 100
457.2a 439.1
5 10
7
Progesterone
315.2
140 140
109 97a
25 23
a
Quantified ion.
Product ion (m/z) 193.2 164.1a 327 121a
Collision energy (V) 10 25 10 25
[(Fig._2)TD$IG]
C.-S. Wu et al. / Chinese Chemical Letters 24 (2013) 509–511
511
Fig. 2. Representative chromatograms (quantifier transition) obtained in the multiple reaction monitoring (MRM) mode for the seven prohibited substances (0.5 mg/mL). The numbers assigned to the peaks are the same as those in Fig. 1. The optimized MS parameters for each substance are listed in Table 1.
substances. The chromatograms obtained in the MRM mode for the seven prohibited substances (0.5 mg/mL) are shown in Fig. 2. All of the calibration curves showed good linearity (r > 0.999) within the tested concentration ranges (10–1500 ng/mL). The limit of detection was <25 pg for each substance. The absolute matrix effects for the seven prohibited substances were calculated by comparing the mean peak areas of the analytes at two concentration levels spiked in post-extracted negative samples with those of the pure standards prepared in methanol. The ratios of the peak areas were within the acceptable range (90.6%–118.0%) in two different matrices (lotion and cream). These results indicate that co-eluting substances in cosmetic products did not affect the ionization of the seven prohibited substances under these analytical conditions. The relative standard deviations for the intraday precision for each substance were <3.5% at both concentration levels (2 mg/g, 10 mg/g). The relative recovery rate for each substance ranged from 91.8% to 111.1% at three concentration levels (0.1 mg/g, 2 mg/g, 10 mg/g). In addition, to assess post-preparation stability, the seven prohibited substances were found to be stable after being stored at room temperature (23 8C) for 24 h, or at low temperature (4 8C) for 6 days. This method was successfully used to detect and quantify seven prohibited substances present in cosmetic products. The seven prohibited substances were detected and confirmed by comparing their retention time and MS data with those of reference substances. None of the prohibited substances were in 23 different cosmetic products purchased from two cosmetics shops. However, at least one of the prohibited substances was found in each of the 14 cosmetic products purchased from an internet-based supplier. In particular, minoxidil was found in eight products, minoxidil and spironolactone were simultaneously detected in one product, and progesterone was found in one cosmetic product. An earlier study used HPLC with diode array detection and single quadrupole mass spectrometry to detect seven prohibited substances in cosmetic products [8]. Compared with our proposed method, the analysis time of the method developed in that study was 50 min with the poorer separation observed for spironolactone and canrenone. 4. Conclusion We established a gradient HPLC–MS/MS method for screening and identifying seven prohibited substances contained in some cosmetic products. The results suggest that HPLC–MS/MS is a valuable analytical tool for the identification and determination of prohibited substances in cosmetic products. Of note, some of these
prohibited substances were present in cosmetic products purchased from an internet-based supplier. Therefore, we believe improved the supervision of internet-based marketing of cosmetics is necessary. Acknowledgment We thank the State Food and Drug Administration of the People’s Republic of China for financially supporting this work.
References [1] V.M. Meidan, E. Touitou, Treatments for androgenetic alopecia and alopecia areata: current options and future prospects, Drugs 61 (2001) 53–69. [2] J. Ayer, N. Burrows, Acne: more than skin deep, Postgrad. Med. J. 82 (2006) 500–506. [3] D. De Orsi, M. Pellegrini, S. Pichini, et al., High-performance liquid chromatography-diode array and electrospray-mass spectrometry analysis of non-allowed substances in cosmetic products for preventing hair loss and other hormonedependent skin diseases, J. Pharm. Biomed. Anal. 48 (2008) 641–648. [4] D. Wasserman, D.A. Guzman-Sanchez, K. Scott, A. Mcmichael, Alopecia areata, Int. J. Dermatol. 46 (2007) 121–131. [5] N. Otberg, A.M. Finner, J. Shapiro, Androgenetic alopecia, Endocrinol. Metab. Clin. North Am. 36 (2007) 379–398. [6] Q.Q. Dinh, R. Sinclair, Female pattern hair loss: current treatment concepts, Clin. Interv. Aging 2 (2007) 189. [7] S.C. Patterson, T. Ramstad, K.A. Mills, Development and validation of a procedure for the determination of minoxidil in hair-regrowth formulations using two variants of capillary zone electrophoresis, Farmaco 60 (2005) 547–554. [8] S. Gorog, Advances in the analysis of steroid hormone drugs in pharmaceuticals and environmental samples (2004–2010), J. Pharm. Biomed. Anal. 55 (2011) 728–743. [9] J.P. Scarth, J. Kay, P. Teale, et al., A review of analytical strategies for the detection of ‘endogenous’ steroid abuse in food production, Drug Test. Anal. 4 (1 Suppl.) (2012) 40–49. [10] F.Z. Stanczyk, N.J. Clarke, Advantages and challenges of mass spectrometry assays for steroid hormones, J. Steroid Biochem. Mol. Biol. 121 (2010) 491–495. [11] V.M. Carvalho, The coming of age of liquid chromatography coupled to tandem mass spectrometry in the endocrinology laboratory, J. Chromatogr. B: Analyt. Technol. Biomed. Life Sci. 883–884 (2012) 50–58. [12] F. Gosetti, E. Mazzucco, M.C. Gennaro, E. Marengo, Ultra high performance liquid chromatography tandem mass spectrometry determination and profiling of prohibited steroids in human biological matrices. A review. J. Chromatogr. B: Analyt. Technol. Biomed. Life Sci., in press, http://dx.doi.org/10.1016/ j.jchromb.2012.12.003. [13] Hygienic Standard for Cosmetics. Issued by the Ministry of Public Health of the People’s Republic of China, 2007 version. [14] Standard Method of Determination Minoxidil in Cosmetics. Issued by the State Food and Drug Administration of the People’s Republic of China, August 2010. [15] Y.S. Nam, I.K. Kwon, K.B. Lee, Monitoring of clobetasol propionate and betamethasone dipropionate as undeclared steroids in cosmetic products manufactured in Korea, Forensic Sci. Int. 210 (2011) 144–148. [16] A. Panusa, M. Ottaviani, M. Picardo, et al., Analysis of corticosteroids by high performance liquid chromatography–electrospray mass spectrometry, Analyst 129 (2004) 719–723.
Contents lists available at SciVerse ScienceDirect
Chinese Chemical Letters journal homepage: www.elsevier.com/locate/cclet
Original article
Simultaneous determination of seven prohibited substances in cosmetic products by liquid chromatography–tandem mass spectrometry Cai-Sheng Wu, Ying Jin, Jin-Lan Zhang *, Yan Ren, Zhi-Xin Jia Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China
A R T I C L E I N F O
A B S T R A C T
Article history: Received 27 January 2013 Received in revised form 1 March 2013 Accepted 8 March 2013 Available online 18 April 2013
In this paper, we developed and validated a simple, sensitive, and selective high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method to identify and measure the following prohibited substances that may be found in cosmetic products: minoxidil, hydrocortisone, spironolactone, estrone, canrenone, triamcinolone acetonide and progesterone. Chromatographic separation was performed on a Waters Symmetry C18 (100 mm 2.1 mm, 3.5 mm particle size) with a gradient elution system composed of 0.2% (v/v) formic acid aqueous solution and methanol containing 0.2% (v/v) formic acid at a flow rate of 0.3 mL/min. The substances were detected using a triple quadrupole mass spectrometer in the multiple reaction monitoring mode with an electrospray ionization source. All of the calibration curves showed good linearity (r > 0.999) within the tested concentration ranges. The limit of detection was <25 pg. The relative standard deviations for intraday precision for each of the prohibited substances were <3.5% at two concentration levels (2 mg/g, 10 mg/g). The relative recovery rate for each of the prohibited substances ranged from 91.8% to 111% at three concentration levels (0.1 mg/g, 2 mg/g, 10 mg/g), including the limit of quantification. In conclusion, we have developed and validated a method that can identify seven prohibited substances in cosmetic products. ß 2013 Jin-Lan Zhang. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords: High-performance liquid chromatography– tandem mass spectrometry (HPLC–MS/MS) Cosmetic products Prohibited substances Minoxidil Steroids
1. Introduction Baldness, alopecia, and acne are common skin and hair diseases that may adversely influence the patient’s social relationships [1–3]. For reasons of safety and ease of use, cosmetic products are widely used to manage these conditions. However, for the sake of profit and to enhance short-term efficacy, some manufacturers illegally add synthetic drugs to their cosmetic products. According to previous reports [3–6], minoxidil, hydrocortisone, spironolactone, estrone, canrenone, triamcinolone acetonide, and progesterone are among the most common topically active anti-alopecia, anti-baldness, and anti-acne drugs. Therefore, we focused on these substances in this study. Up to now, many articles have reported methods to detect minoxidil or steroids in drug formulations [7,8], environmental samples [8], food products [9], and biological samples [10–12]. However, few procedures for the detection of minoxidil or steroids in cosmetic products have been reported to date [3,13–16]. Furthermore, minoxidil was detected in a pure Chinese herbal medicine shampoo (Zhangguang 101) in 2010. Therefore, methods that can rapidly and simultaneously screen for,
* Corresponding author. E-mail address: [email protected] (J.-L. Zhang).
and quantify, prohibited substances in cosmetic products are urgently required. In this paper, we established and validated a simple and rapid high-performance liquid chromatography– tandem mass spectrometry (HPLC–MS/MS) method capable of simultaneously determining seven prohibited substances in cosmetic products. We applied this method to analyze 37 different cosmetic products used for prophylaxis and treatment of hair loss and acne. 2. Experimental 2.1. Instruments and materials The assay was performed on an Agilent 6410A triple quadrupole LC–MS system (Agilent Technologies, Santa Clara, CA, USA) consisting of an Agilent 1200 RRLC system (Agilent Corporation) connected to a triple quadrupole MS analyzer with an electrospray ionization (ESI) interface that could be used in either positive or negative ionization modes. A MassHunter workstation was used to control the LC–MS and for data acquisition (Agilent Corporation). LC/MS reagent grade acetonitrile was obtained from Mallinckrodt Baker, Inc. (Phillipsburg, NJ, USA). Deionized water was purified using a Milli-Q1 academic water purification system (Merck Millipore, Billerica, MA, USA). Analytical grade formic acid
1001-8417/$ – see front matter ß 2013 Jin-Lan Zhang. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. http://dx.doi.org/10.1016/j.cclet.2013.03.037
[(Fig._1)TD$IG]
C.-S. Wu et al. / Chinese Chemical Letters 24 (2013) 509–511
510
Fig. 1. Chemical structures of seven prohibited substances.
was obtained from Merck Inc. (Darmstadt, Germany). Analytical grade methanol was purchased from Beijing Chemical Corp. (Beijing, China). Reference standards of the seven prohibited substances were obtained from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). Their structures are shown in Fig. 1. Thirty-seven different cosmetic products used for prophylaxis and treatment of hair loss and acne were obtained from two cosmetic shops and an internet-based supplier (www.taobao.com). 2.2. LC–ESI-MS analytical conditions Chromatographic separation was performed using a Waters Symmetry C18 (100 mm 2.1 mm, 3.5 mm particle size) column. The mobile phase, consisting of solvent A (0.2% (v/v) formic acid aqueous solution) and solvent B (methanol containing 0.2% (v/v) formic acid), was delivered at a flow rate of 0.3 mL/min. The gradient elution was started with 5% B, increased to 25% B in 2 min, then to 55% B in 2 min, to 80% B in 7 min, then to 90% B in 4 min. Then the column was flushed with 5% B for 8 min. The column temperature was maintained at 30 8C and the injection volume was 5 mL. The optimum operating parameters of the ESI interface in the positive mode were as follows: nebulizer, 40 psi; dry gas, 8 L/min; dry temp, 325 8C, capillary voltage, 4000 V; and Delta EMV, 300 V. The LC eluent flow from 0 to 3.5 min was not introduced to the MS for data acquisition. Each substance was detected in the multiple reaction monitoring (MRM) mode, using the parameters listed in Table 1. 2.3. Sample preparation A sample of lotion (1 mL) or cream (1 g) was accurately weighed into a 10 mL colorimetric cylinder. Then, 1 mL of a saturated
solution of sodium chloride was added to avoid emulsification and the mixture was vortexed for 30 s. Acetonitrile was added followed by vortexing. The cosmetic extracts were ultrasonicated for 30 min at 25 8C and vortexed for 60 s. After centrifugation at 4500 rpm for 10 min, an aliquot of the supernatant was filtered through a 0.45 mm filter and injected into the HPLC–MS/MS system. 3. Results and discussion Optimization of the sample preparation method was essential to obtain a good accuracy and reliability for quantification. According to the previous reports [3,8–16], several methods, including solid-phase extraction (SPE) and liquid–liquid extraction (LLE), were used to prepare samples for analysis of minoxidil or steroids. For cosmetic products, there are different formulas and the ingredients of same formula from different factories. After tried, it was difficult to find a solid phase of SPE that was suitable for all kind products. In addition, some ingredients of cosmetic products may block the SPE cartridge. The LLE method was also tried. However, emulsion was readily formed and the accuracy and reliability of quantification was not satisfied. After optimizing the pretreatment conditions, the same amount of saturated sodium chloride solution was used to break the emulsification of cosmetic products and to reduce extraction process emulsification effects. Finally, by comparing the processing of more than 20 different products, we found that methanol was more likely to cause emulsification than acetonitrile, therefore acetonitrile was used as the extraction solvent to simplify the procedure and maintain its accuracy. The total run time for each sample was just 26 min, and good separation of the seven prohibited substances was achieved under the specified gradient HPLC–MS/MS conditions. There was no interference of the retention time for any of the prohibited
Table 1 Optimized mass parameters for the seven prohibited substances. No.
Analyte
Precursor ion (m/z)
Fragmentor (V)
1
Minoxidil
210.0
110 110
2
Hydrocortisone
363.1
125 125
3
Spironolactone
341.1
147 165
107.1a 90.9
35 65
4
Estrone
271.2
105 105
253.1a 132.8
5 22
5
Canrenone
341.1
147 165
107.1a 90.9
35 65
6
Triamcinolone acetonide
477.1
100 100
457.2a 439.1
5 10
7
Progesterone
315.2
140 140
109 97a
25 23
a
Quantified ion.
Product ion (m/z) 193.2 164.1a 327 121a
Collision energy (V) 10 25 10 25
[(Fig._2)TD$IG]
C.-S. Wu et al. / Chinese Chemical Letters 24 (2013) 509–511
511
Fig. 2. Representative chromatograms (quantifier transition) obtained in the multiple reaction monitoring (MRM) mode for the seven prohibited substances (0.5 mg/mL). The numbers assigned to the peaks are the same as those in Fig. 1. The optimized MS parameters for each substance are listed in Table 1.
substances. The chromatograms obtained in the MRM mode for the seven prohibited substances (0.5 mg/mL) are shown in Fig. 2. All of the calibration curves showed good linearity (r > 0.999) within the tested concentration ranges (10–1500 ng/mL). The limit of detection was <25 pg for each substance. The absolute matrix effects for the seven prohibited substances were calculated by comparing the mean peak areas of the analytes at two concentration levels spiked in post-extracted negative samples with those of the pure standards prepared in methanol. The ratios of the peak areas were within the acceptable range (90.6%–118.0%) in two different matrices (lotion and cream). These results indicate that co-eluting substances in cosmetic products did not affect the ionization of the seven prohibited substances under these analytical conditions. The relative standard deviations for the intraday precision for each substance were <3.5% at both concentration levels (2 mg/g, 10 mg/g). The relative recovery rate for each substance ranged from 91.8% to 111.1% at three concentration levels (0.1 mg/g, 2 mg/g, 10 mg/g). In addition, to assess post-preparation stability, the seven prohibited substances were found to be stable after being stored at room temperature (23 8C) for 24 h, or at low temperature (4 8C) for 6 days. This method was successfully used to detect and quantify seven prohibited substances present in cosmetic products. The seven prohibited substances were detected and confirmed by comparing their retention time and MS data with those of reference substances. None of the prohibited substances were in 23 different cosmetic products purchased from two cosmetics shops. However, at least one of the prohibited substances was found in each of the 14 cosmetic products purchased from an internet-based supplier. In particular, minoxidil was found in eight products, minoxidil and spironolactone were simultaneously detected in one product, and progesterone was found in one cosmetic product. An earlier study used HPLC with diode array detection and single quadrupole mass spectrometry to detect seven prohibited substances in cosmetic products [8]. Compared with our proposed method, the analysis time of the method developed in that study was 50 min with the poorer separation observed for spironolactone and canrenone. 4. Conclusion We established a gradient HPLC–MS/MS method for screening and identifying seven prohibited substances contained in some cosmetic products. The results suggest that HPLC–MS/MS is a valuable analytical tool for the identification and determination of prohibited substances in cosmetic products. Of note, some of these
prohibited substances were present in cosmetic products purchased from an internet-based supplier. Therefore, we believe improved the supervision of internet-based marketing of cosmetics is necessary. Acknowledgment We thank the State Food and Drug Administration of the People’s Republic of China for financially supporting this work.
References [1] V.M. Meidan, E. Touitou, Treatments for androgenetic alopecia and alopecia areata: current options and future prospects, Drugs 61 (2001) 53–69. [2] J. Ayer, N. Burrows, Acne: more than skin deep, Postgrad. Med. J. 82 (2006) 500–506. [3] D. De Orsi, M. Pellegrini, S. Pichini, et al., High-performance liquid chromatography-diode array and electrospray-mass spectrometry analysis of non-allowed substances in cosmetic products for preventing hair loss and other hormonedependent skin diseases, J. Pharm. Biomed. Anal. 48 (2008) 641–648. [4] D. Wasserman, D.A. Guzman-Sanchez, K. Scott, A. Mcmichael, Alopecia areata, Int. J. Dermatol. 46 (2007) 121–131. [5] N. Otberg, A.M. Finner, J. Shapiro, Androgenetic alopecia, Endocrinol. Metab. Clin. North Am. 36 (2007) 379–398. [6] Q.Q. Dinh, R. Sinclair, Female pattern hair loss: current treatment concepts, Clin. Interv. Aging 2 (2007) 189. [7] S.C. Patterson, T. Ramstad, K.A. Mills, Development and validation of a procedure for the determination of minoxidil in hair-regrowth formulations using two variants of capillary zone electrophoresis, Farmaco 60 (2005) 547–554. [8] S. Gorog, Advances in the analysis of steroid hormone drugs in pharmaceuticals and environmental samples (2004–2010), J. Pharm. Biomed. Anal. 55 (2011) 728–743. [9] J.P. Scarth, J. Kay, P. Teale, et al., A review of analytical strategies for the detection of ‘endogenous’ steroid abuse in food production, Drug Test. Anal. 4 (1 Suppl.) (2012) 40–49. [10] F.Z. Stanczyk, N.J. Clarke, Advantages and challenges of mass spectrometry assays for steroid hormones, J. Steroid Biochem. Mol. Biol. 121 (2010) 491–495. [11] V.M. Carvalho, The coming of age of liquid chromatography coupled to tandem mass spectrometry in the endocrinology laboratory, J. Chromatogr. B: Analyt. Technol. Biomed. Life Sci. 883–884 (2012) 50–58. [12] F. Gosetti, E. Mazzucco, M.C. Gennaro, E. Marengo, Ultra high performance liquid chromatography tandem mass spectrometry determination and profiling of prohibited steroids in human biological matrices. A review. J. Chromatogr. B: Analyt. Technol. Biomed. Life Sci., in press, http://dx.doi.org/10.1016/ j.jchromb.2012.12.003. [13] Hygienic Standard for Cosmetics. Issued by the Ministry of Public Health of the People’s Republic of China, 2007 version. [14] Standard Method of Determination Minoxidil in Cosmetics. Issued by the State Food and Drug Administration of the People’s Republic of China, August 2010. [15] Y.S. Nam, I.K. Kwon, K.B. Lee, Monitoring of clobetasol propionate and betamethasone dipropionate as undeclared steroids in cosmetic products manufactured in Korea, Forensic Sci. Int. 210 (2011) 144–148. [16] A. Panusa, M. Ottaviani, M. Picardo, et al., Analysis of corticosteroids by high performance liquid chromatography–electrospray mass spectrometry, Analyst 129 (2004) 719–723.