Solutions that meet your demands for food testing & agriculture Our measure is your success. Excellent choices for food & agriculture applications products I applications I software I services Agilent Technologies Consumer Products Toys, jewelry, clothing, and other products are frequently recalled due to the presence of unsafe levels of substances such as lead from paint and phthal- ates from product polymers and packaging. Whether your perspective is to guarantee your products are free of contaminants or you are screening for harmful contaminants in a wide variety of consumer products, Agilent Tech- nologies provides the tools you need to detect and measure these and other harmful contaminants. > Search entire document Agilent 1290 Infinity LC with Agilent Poroshell columns for simultaneous determination of eight organic UV filters in under two minutes Application Note Consumer Products Authors Siji Joseph Agilent Technologies India Pvt. Ltd. mAU Amino benzoic acid Bangalore, India 2 Oxybenzone 1.5 4-Methyl benzylidene camphor Dioxybenzone Avobenzone Michael Woodman 1 Octyl methoxycinnamate 0.5 Octocrylene Agilent Technologies, Inc. Octyl salicylate 2850 Centerville Road 0 0 0.5 1 1.5 2 min Wilmington DE 19808 USA Abstract Levels of UV filters in personal care products are regulated by the FDA and European Pharmacopeia (EP). Liquid chromatographic (LC) methods are widely accepted analyt- ical techniques for the qualitative and quantitative analysis of these UV filters. Most of these traditional LC methods require about 25–50 minutes. In this Application Note, the Agilent 1290 Infinity LC, in combination with Agilent Poroshell columns, were used for development of a short, sensitive, robust and well resolved separation of eight FDA/EP approved active UV filter ingredients in 99 seconds. Standard deviation (SD) and relative standard deviation (RSD) values of retention time for replicate injec- tions confirmed the excellent performance of the Agilent 1290 Infinity Binary Pump. Exceptional performance of the Agilent 1290 Infinity Diode Array Detector was estab- lished by minimum area RSD values and a wide linear range with standard organic UV filters in amounts from 0.25 ng to 200 ng on-column. In addition, the method was effectively used to identify active UV filters extracted from six international personal care products. Introduction Sl No: Component Wavelength (nm) Effects of over exposure on skin The wavelength of UV rays lies 1 UV-A I 340-400 Can cause tanning but has minimal erythemal effect. between X-rays and visible light Can cause long term damage. Penetrates deeply. it can (~ 10 – 400 nm)1. Production of contribute to skin cancer via indirect DNA damage. melanin, a pigment that causes the 2 UV-A II 320-340 Slightly erythemal contribution darkening of skin, is a natural defense of the human body against UV radia- 3 UV-B 290-320 Causes sunburn and is a major contributor to skin cancer tion. Melanin absorbs UV radiation and development. dissipates the energy as harmless heat, 4 UV-C 100-290 Very energetic radiation. Absorbed by the ozone layer. Direct DNA damage though the response to UV radiation and production of melanin pigmentation Table 1 depends on skin color and other genetic Major classifications of UV radiation. factors 2, 3. The intensity of UV radiation and length of exposure are the main parameters involved in sunburn, irre- major concerns. Some sunscreen ingre- and a diode array detector, was used for spective of skin tone and ability of the dients have been shown to have car- data collection. The injection volume skin to produce melanin. The major cinogenic properties. Additionally, older was set to 1 µL and the needle wash classifications of UV light are presented and more widespread sunscreen chemi- was enabled using acetonitrile for three in Table 14. cals cannot dissipate the energy of the seconds. The sample thermostat was excited state as efficiently as melanin, set at 5 °C, while the columns were Sunscreens protecting the skin against therefore the penetration of these sun- operated at 55 °C. The binary pump was sunburn contain one or more of the fol- screen ingredients into the lower layers operated at a flow rate of 1 mL/min. lowing types of active ingredients: of the skin may increase the amount of The detector was programmed for three free radicals and reactive oxygen different wavelengths (288, 304 and 358 • Organic chemicals that absorb species6. Therefore, extensive testing nm) and operated at a sampling acqui- UV light. of sunscreens is advisable to reveal the sition rate of 80 Hz (response time • Inorganic particulates that efficacy of the ingredients. This 0.062 seconds, >0.003 min). An Agilent reflect, scatter, and absorb UV Application Note discusses a short LC Poroshell 120 EC-C18 column (75 mm × light. method to separate eight widely used 2.1 mm, 2.7 µm) was used for the chro- UV filters within 99 seconds. Cosmetic matographic separation. • Organic particulates that have all the manufacturers can adopt this method above features. to simplify the analysis of sunscreen Chemicals and standards raw materials and personal care prod- All eight UV filter standards and acetic Organic UV filters are usually aromatic ucts in product development, regulatory acid (mobile phase modifier) were pur- compounds conjugated with carbonyl compliance, and quality control to chased from Aldrich (India). Super gra- groups. The FDA has approved seven increase the efficiency of analysis. dient grade acetonitrile (ACN) was pur- UV-A filter compounds and nine UV-B chased from Lab-Scan (Bangkok, filter compounds for sunscreen formu- Experimental Thailand). HPLC grade water was fresh- lations in the United States, while the ly taken from a Milli-Q water purifica- European Commission has approved Instrument configuration tion system. Six different brands of the use of ten additional UV filters in An Agilent 1290 Infinity LC, controlled international sunscreen formulations European countries5. by ChemStation (Version B.04.02) and were purchased locally. The details of equipped with a binary pump with inte- organic UV filter standards used in this Though several approved UV filters are grated vacuum degasser, autosampler, study are tabulated in Table 2. available in the market, extensive use thermostatted column compartment of these UV filters may have several 2 Compound number Compound Structure UV Spectra COOH 1 Name: Aminobenzoic acid mAU 288 CAS No: 150-13-0 500 Mol Formula: C7H7NO2 300 Mol Wt: 137.14 100 0 Detection: 288 nm 250 300 350 nm NH2 OH O OH mAU 2 Name: Dioxybenzone 400 284 CAS No: 131-53-3 300 200 Mol Formula: C14H12O4 Mol Wt: 244.24 100 H3OC 0 Detection: 288 nm 250 300 350 nm OH O 288 3 Name: Oxybenzone mAU CAS No: 131-57-7 600 Mol Formula: C H O 400 14 12 3 200 Mol Wt: 228.24 O 0 Detection: 288 nm 250 300 350 nm 4 Name: 4-Methyl benzylidene camphor 300 CAS No: 36861-47-9 mAU 400 Mol Formula: C18H22O 300 Mol Wt: 254.37 200 100 Detection: 304 nm 0 250 300 350 nm O O O 5 Name: Avobenzone 358 mAU CAS No: 70356-09-1 500 Mol Formula: C20H22O3 Mol Wt: 310.39 CH 300 H C 3 Detection: 358 nm 3 100 O 0 CH3 250 300 350 nm H3C 308 6 Name: Octylmethoxycinnamate O CAS No: 5466-77-3 mAU 500 Mol Formula: C18H26O3 O Mol Wt: 290.397 300 Detection: 304 nm 100 H3OC 0 250 300 350 nm 7 Name: Octocrylene CAS No: 6197-30-4 N mAU Mol Formula: C H NO 24 27 2 500 304 Mol Wt: 361.48 300 Detection: 304 nm O 100 0 O 250 300 350 nm 8 Name: Octyl salicylate CAS No: 118-60-5 mAU Mol Formula: C H O O O 15 22 3 500 Mol Wt: 250.33 304 300 Detection: 304 nm OH 100 0 250 300 350 nm Table 2 Detailed list of organic UV filter standards used in this study. 3 LC parameters equal volumes of 100 ppm standard mix ingredients, without chromatographic Premixed solutions of 0.1% acetic acid solution and extracted sample stock blank interference. The last peak of the in water and acetonitrile in the ratio solution. This spiked sample was used standard mix (octyl salicylate) eluted at 90:10 (A) and 10:90 (B) were used as for the confirmation of peak identity in 1.62 minutes. A chromatographic rep- mobile phase. The gradient used for the extracted samples by means of reten- resentation of the standard mix is as study is presented in Table 3. A post run tion time and UV spectra. shown in Figure 1. An unknown peak was observed at approximately time of 1 minute was set for column re- Procedure equilibration. 0.65 minute, which is an impurity pre- A blank injection was performed in all sent in the avobenzone standard. A Time (min) B (%) trials to check the chromatographic peak purity check by spectral scanning 050interference in the resolution. Standard in the range of 200 to 400 nm revealed 0.1 70 mix, linearity levels, diluted extracted that all eight compounds eluted with- 285samples and spiked diluted extracted out co-elution of any detectable impu- Table 3 samples were also injected. The reten- rities. Three different wavelengths Gradient used for experiment. tion time of each standard was con- were selected for detection as the firmed by individual standard injections. maximum absorbance values vary for Standard mix individual components. The peak width A premixed solution of acetonitrile and Results and Discussion (half height), peak symmetry, USP tail- 0.1% acetic acid in the ratio 50:50 was LC chromatogram of standard ing factor, and resolution values confirm used as the diluent. A stock solution of mixture the baseline separation of all the stan- each standard was prepared individual- dard analytes in 99 seconds using the ly at a concentration of 1000 ppm The results showed excellent baseline Agilent Poroshell 120 EC-C18 column (1000 ng/µL).