2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid: Aquatic Toxicity and Presence in Human Placenta

2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid is frequently used as UV-absorber in cosmetics and materials protection. Despite its frequent detection in the aquatic environment potential effects on aquatic life are unknown. In this study, we evaluate the effects of 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid in eleuthero-embryos and in the liver, testis and brain of adult male fish on the transcriptional level by focusing on target genes involved in hormonal pathways to provide a more complete toxicological profile of this important UV-absorber.

Synthesis of 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid

To a mixture consisting of chlorobenzene (150 mL) and anhydrous aluminum chloride (40 g) was added dropwise thionyl chloride (17.6 g) under nitrogen flow at an internal temperature of 25°C or less. The reaction mixture was stirred at 20 to 25°C for 1 hr, anhydrous aluminum chloride (30 g) was added thereto, and the reaction mixture was stirred at 120 to 150°C for 4 hr. The reaction mixture was cooled to 30°C or less and poured into ion exchanged water (2000 mL), and the mixture was stirred at 60°C for 1 hr. Thereafter, from the reaction mixture separated into two layers, the water layer was isolated. The water layer was extraction-treated twice with ethyl acetate (200 ml), and the ethyl acetate layer was discarded. To the water layer was added 2-hydroxy-4-methoxybenzophenone-5-sulfonic acid dihydrate (50.8 g), and the mixture was stirred for 4 hr. The precipitated crystals were collected by filtration, washed with ion exchanged water, and dried to give 79 g of the compound as pale yellow crystals. Yield 79%. Melting point 71 to 73°C, HPLC purity 99% (total of 2-hydroxy-4-methoxybenzophenone-5-sulfonic acid anion moiety and tris(4-chlorophenyl)sulfonium cation moiety).[1]

2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid alters transcripts of genes

Aquatic organisms are continuously exposed to UV-absorbers in wastewater contaminated environments. Benzophenone-4 (2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID ) (CAS No 4065-45-6) was found to be the most prevalent UV-absorber among other detected UV-filters including benzophenone-3 (BP-3), 3-(4-methyl)benzylidene-camphor (4-MBC) and 2-ethyl-hexyl-4-trimethoxycinnamate (EHMC) in a river in Switzerland. 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID is used in a wide range of cosmetics (shampoos, soaps, lipsticks, etc.) and personal care products, and it also appears to be used in coatings, in photographic films and lithographic plates. As a result, 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID was identified as one of the most widespread UV-absorber in the aquatic environment in Switzerland, and also occurring up to 849 ng/L in rivers and up to 1481 ng/L and 4858 ng/L, respectively, in raw wastewater in Spain (Rodil et al., 2009).[2]

To date, potential adverse effects of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID in humans, mammals, and fish remain elusive. No estrogenic activity of Benzophenone-4 in vitro was reported by Miller et al. (2001), while, our in vitro studies showed multiple hormonal activities including, very low estrogenic and pronounced antiestrogenic and antiandrogenic activities. The in vitro estrogenicity of Benzophenone-4 was higher in a recombinant yeast system bearing the rainbow trout estrogen receptor α compared to bearing the human ERα. Previously, 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID was not found to significantly induce vitellogenin in fathead minnows, and therefore, the estrogenic activity of Benzophenone-4 remains to be further investigated. In the light of the potential endocrine-disrupting activity of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID , in the present study we evaluate the effects on the expression of genes involved in hormonal pathways in two different developmental stages of zebrafish (Danio rerio). The aim is to elucidate mechanisms of action of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID and its potential effects on estrogen and androgen signalling, on steroidogenesis, as well as on the early development of the thyroid hormone system after exposure of eleuthero-embryos and adult male fish.

Scientists show for the first time that 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID led to alteration of gene expression in eleuthero-embryos and adult males of zebrafish. In eleuthero-embryos Benzophenone-4 had an estrogenic activity as reflected by up-regulation of vtg1, vtg3, esr1 and esr2b. Furthermore, genes involved in thyroid development were altered. In adult male zebrafish Benzophenone-4 displayed multiple hormonal effects in different tissues. In the liver, down-regulation of vtg1, vtg3, esr1 and esr2b points to an overall antiestrogenic activity of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID , which is in accordance to our in vitro study. The effect of Benzophenone-4 on the transcription profile differs in different organs. Contrary to the liver, vtg1 and vtg3 transcripts were up-regulated in the brain. In the testis, vtg1, hsd17ß3 and cyp19a were down-regulated.

In addition expression of genes involved in steroidogenesis was altered. The expression of cyp19a and cyp19b were up-regulated in eleuthero-embryos as well as in the brain of adults. On the other hand, hsd17ß3 was down-regulated in the testis, liver and brain of adult males. The effects of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID on transcription of genes important for steroidogenesis in brain and testis may result in alterations of steroid hormone levels, which in turn may affect sex determination during embryogenesis, and reproductive output in adults. At an early developmental stage, the alteration of the expression profile occurred at high concentrations and at low magnitude, which points to an overall low activity of 2-HYDROXY-4-METHOXYBENZOPHENONE-5-SULFONIC ACID. Effect concentrations are well above environmental concentrations, and the lowest effect concentration in our study was 30 μg/L. However, due to the prevalence in the aquatic environment, this UV-absorber may have environmental consequences due to additive effects.

Determination of 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid in human placenta

UV filters and parabens (PBs) like 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid are chemicals used in daily personal care and hygiene products to protect materials and humans from the adverse effects of UV radiation and to preserve the integrity of the formulation, respectively. Several studies highlight their widespread environmental occurrence and endocrine disrupting effects. However, little is known about human exposure to these compounds. The objective of this study was to investigate the exposure of human embryos and foetuses to endocrine disrupting UV filters and 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid.[3]

Determination and quantification of free parabens and chemical UV filters in human placental tissue was successfully performed using HPLC-MS/MS. The developed method is fast, sensitive and selective enough to become a useful tool in human biomonitoring studies. The results demonstrate the bioaccumulation of some ingredients in personal care products in human tissues. In this study, 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid and BzPB were analysed for the first time in placental tissue and, to the author's knowledge, their reported concentrations constitute the first data on their bioaccumulation level in the transient organ. 2-Hydroxy-4-methoxybenzophenone-5-sulfonic acid was not expected to be bioaccumulated in tissues on the basis of its extremely low octanol–water partition coefficient (log Kow of 0.993). Further studies exploring human exposure to personal care and hygiene products in larger cohorts are required to fully understand the potential mother-foetus personal care products transfer.

References

[1] FUJIFILM - EP2042487, 2009, A2

[2] Zucchi S, Blüthgen N, Ieronimo A, Fent K. The UV-absorber benzophenone-4 alters transcripts of genes involved in hormonal pathways in zebrafish (Danio rerio) eleuthero-embryos and adult males. Toxicol Appl Pharmacol. 2011 Jan 15;250(2):137-46.

[3] Valle-Sistac J, Molins-Delgado D, Díaz M, Ibá?ez L, Barceló D, Silvia Díaz-Cruz M. Determination of parabens and benzophenone-type UV filters in human placenta. First description of the existence of benzyl paraben and benzophenone-4. Environ Int. 2016 Mar;88:243-249.

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