lable at ScienceDirect

Food and Chemical Toxicology 84 (2015) S110eS121

Contents lists avai

Food and Chemical Toxicology

journal homepage: www.elsevier .com/locate/ foodchemtox

Short review

RIFM fragrance ingredient safety assessment, isoamyl salicylate, CASregistry number 87-20-7

A.M. Api a, *, D. Belsito b, S. Bhatia a, M. Bruze c, P. Calow d, M.L. Dagli e, W. Dekant f,A.D. Fryer g, L. Kromidas a, S. La Cava a, J.F. Lalko a, A. Lapczynski a, D.C. Liebler h,Y. Miyachi i, V.T. Politano a, G. Ritacco a, D. Salvito a, T.W. Schultz j, J. Shen a, I.G. Sipes k,B. Wall a, D.K. Wilcox a

a Research Institute for Fragrance Materials, Inc., 50 Tice Boulevard, Woodcliff Lake, NJ 07677, USAb Columbia University Medical Center, Department of Dermatology, 161 Fort Washington Ave., New York, NY 10032, USAc Malmo University Hospital, Department of Occupational & Environmental Dermatology, Sodra Forstadsgatan 101, Entrance 47, Malmo SE-20502, Swedend University of Nebraska Lincoln, 230 Whittier Research Center, Lincoln, NE 68583-0857, USAe University of Sao Paulo, School of Veterinary Medicine and Animal Science, Department of Pathology, Av. Prof. Dr. Orlando Marques de Paiva, 87, Sao PauloCEP 05508-900, Brazilf University of Wuerzburg, Department of Toxicology, Versbacher Str. 9, 97078 Würzburg, Germanyg Oregon Health Science University, 3181 SW Sam Jackson Park Rd., Portland, OR 97239, USAh Vanderbilt University School of Medicine, Department of Biochemistry, Center in Molecular Toxicology, 638 Robinson Research Building, 2200 PierceAvenue, Nashville, TN 37232-0146, USAi Department of Dermatology, Kyoto University Graduate School of Medicine, 54 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japanj The University of Tennessee, College of Veterinary Medicine, Department of Comparative Medicine, 2407 River Dr., Knoxville, TN 37996-4500, USAk Department of Pharmacology, University of Arizona, College of Medicine, 1501 North Campbell Avenue, P.O. Box 245050, Tucson, AZ 85724-5050, USA

a r t i c l e i n f o

Article history:Received 23 July 2015Accepted 22 September 2015Available online 28 September 2015

Keywords:GenotoxicityRepeated dose, developmental andreproductive toxicitySkin sensitizationPhototoxicity/photoallergenicityLocal respiratory toxicityEnvironmental safety

* Corresponding author.E-mail address: AApi@rifm.org (A.M. Api).

http://dx.doi.org/10.1016/j.fct.2015.09.0140278-6915/© 2015 Elsevier Ltd. All rights reserved.

a b s t r a c t

The use of this material under current use conditions is supported by the existing information. Thismaterial was evaluated for genotoxicity, repeated dose toxicity, developmental toxicity, reproductivetoxicity, local respiratory toxicity, phototoxicity, skin sensitization potential, as well as, environmentalsafety. Repeated dose toxicity was determined using to have the most conservative systemic exposurederived NOAEL of 47 mg/kg/day. A dietary 13-week subchronic toxicity study conducted in rats on asuitable read across analog resulted in a MOE of 2350 while considering 10.3% absorption from skincontact and 100% from inhalation. A MOE of >100 is deemed acceptable.

© 2015 Elsevier Ltd. All rights reserved.

Version: 071615. This version replaces any previous versions.Name: Isoamyl salicylateCAS Registry Number: 87-20-7

RIFM's Expert Panel* concludes that this material is safe under the limits described in this safety assessment.This safety assessment is based on RIFM's Criteria Document (Api et al., 2015) and should be referred to for clarifications.Each endpoint discussed in this safety assessment reviews the relevant data that were available at the time of writing (version number in the top box is indicative of

the date of approval based on a two digit month/day/year), both in the RIFM database (consisting of publicly available and proprietary data) and through publiclyavailable information sources (i.e., SciFinder and PubMed). Studies selected for this safety assessment were based on appropriate test criteria such as, acceptableguidelines, sample size, study duration, route of exposure, relevant animal species, most relevant testing endpoints, etc. A key study for each endpoint was selectedbased on the most conservative end-point value (e.g., PNEC, NOAEL, LOEL, and NESIL).

*RIFM's Expert Panel is an independent body that selects its own members and establishes its own operating procedures. The Expert Panel is comprised ofinternationally known scientists that provide RIFM guidance relevant to human health and environmental protection.

Summary: The use of this material under current use conditions is supported by the existing information.This material was evaluated for genotoxicity, repeated dose toxicity, developmental toxicity, reproductive toxicity, local respiratory toxicity, phototoxicity, skin

sensitization potential, as well as, environmental safety. Repeated dose toxicity was determined using to have the most conservative systemic exposure derivedNOAEL of 47mg/kg/day. A dietary 13-week subchronic toxicity study conducted in rats on a suitable read across analog resulted in aMOE of 2350 while considering10.3% absorption from skin contact and 100% from inhalation. A MOE of >100 is deemed acceptable.

Human health safety assessmentGenotoxicity: Not Genotoxic (RIFM, 1999a)Repeated dose toxicity: NOAEL ¼ 47 mg/kg/day (Belsito et al., 2007)Developmental and reproductive toxicity: NOAEL ¼ 75 mg/kg/day (Collins et al., 1971)Skin sensitization: Not sensitizing (Ishihara et al., 1986; RIFM, 1970)Phototoxicity/Photoallergenicity: Not phototoxic/photoallergenic (UV spectra, RIFM database)Respiratory toxicity: No NOAEC available. Exposure below TTC.Environmental safety assessmentHazard assessment:Persistence: Critical measured value: 86% based on read-across to Amyl Salicylate (RIFM, 1996a, b)Bioaccumulation: Screening level:: 429 L/Kg (EPISUITE ver 4.1)

Abbreviation list

2-Box Model a RIFM, Inc. proprietary in silico tool used tocalculate fragrance air exposure concentration

97.5th percentile The concentration of the fragrance ingredientis obtained from examination of severalthousand commercial fine fragranceformulations. The upper 97.5th percentileconcentration is calculated from these dataand is then used to estimate the dermalsystemic exposure in ten types of the mostfrequently used personal care and cosmeticproducts. The dermal route is the major routein assessing the safety of fragranceingredients. Further explanation of how thedata were obtained and of how exposureswere determined has been previouslyreported by Cadby et al. (2002) and Ford et al.(2000).

AF Assessment FactorBCF Bioconcentration factorDEREK Derek nexus is an in silico tool used to identify

structural alertsDST Dermal Sensitization ThresholdECHA European Chemicals AgencyEU Europe/European UnionGLP Good Laboratory PracticeIFRA The International Fragrance Association

LOEL Lowest Observable Effect LevelMOE Margin of ExposureMPPD Multiple-Path Particle Dosimetry. An in silicomodel for

inhaled vapors used to simulate fragrance lungdeposition

NA North AmericaNESIL No Expected Sensitization Induction LevelNOAEC No Observed Adverse Effect ConcentrationNOAEL No Observed Adverse Effect LevelNOEC No Observed Effect ConcentrationOECD Organisation for Economic Co-operation and

DevelopmentOECD TG Organisation for Economic Co-operation and

Development Testing GuidelinesPBT Persistent, Bioaccumulative, and ToxicPEC/PNEC Predicted Environmental Concentration/Predicted No

Effect ConcentrationQRA quantitative risk assessmentREACH Registration, Evaluation, Authorisation, and Restriction

of ChemicalsRIFM Research Institute for Fragrance MaterialsRQ Risk QuotientTTC Threshold of Toxicological ConcernUV/Vis. Spectra Ultra Violet/Visible spectraVCF Volatile Compounds in FoodVoU Volume of UsevPvB (very) Persistent, (very) BioaccumulativeWOE Weight of Evidence

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121 S111

Ecotoxicity: Critical ecotoxicity endpoint: based on read across to cis-E-hexenyl salicylate (CAS# 65405-77-8)Daphnia magna 48 h EC50: 0.28 mg/l (RIFM, 2010)Conclusion: Not PBT or vPvB as per IFRA Environmental Standards

Risk assessment:Screening-level: PEC/PNEC (North America and Europe) > 1 (Salvito et al., 2002)Critical Ecotoxicity Endpoint: Based on read across to cis-E-hexenyl salicylate (CAS# 65405-77-8), Daphnia magna 48 h EC50: 0.28 mg/l (RIFM, 2010)RIFM PNEC is 0.28 mg/L

� Revised PEC/PNECs (2011 IFRA VoU): North America and Europe <1

Expert judgment Toxtree (v 2.6.0) OECD QSAR Toolbox (v. 3.2)I I I

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121S112

1. Identification

1. Chemical Name: Isoamyl salicylate2. CAS Registry Number: 87-20-73. Synonyms: Amyl(iso) salicylate, Benzoic acid, 2-hydroxy-, 3-

methylbutyl ester, Isoamyl o-hydroxybenzoate, isoamyl salicy-late, Isopentyl salicylate, 3-Methylbutyl salicylate, 3-Methylbutyl o-hydroxybenzoate, (C ¼ 1～22)

4. Molecular formula: C12H16O35. Molecular weight: 208.266. RIFM number: 102

2. Physical data

1. Boiling point: >200 �C [FMA database], (calculated) 306.01 �C[EPI Suite]

2. Flash point: >200 �F; CC [FMA database]3. Log Kow: 4.49 [EPI Suite]4. Melting point: 82.45 �C [EPI Suite]5. Water solubility: 21.89 mg/L [EPI Suite]6. Specific gravity: 1.049 [FMA database]7. Vapor pressure: 0.000338 mm Hg @ 20 �C [EPI Suite 4.0],

0.000651 mm Hg @ 25 �C [EPI Suite]8. UV Spectra:Minor absorbance between 290 and 200 nm; molar

absorption coefficient below the benchmark (1000 L/mol cm�1)9. Appearance/Organoleptic: Orchidaceous scent. Colorless liquid.

ColorNo.4maximum.Sweetherbaceous-green, slightlyfloralodor.Very tenacious, somewhat woody-earthy, sweet dry out notes.

3. Exposure

1. Volume of Use (worldwide band): 100e1000 metrictons per year

[IFRA, 2011]

2. Average Maximum Concentration in Hydroalcoholics:2.19%

[IFRA, 2002]

3. 97.5th Percentile: 4.09% [IFRA, 2002]4. Dermal Exposure*: 0.1042 mg/kg/day [IFRA, 2002]5. Oral Exposure: Not available6. Inhalation Exposures**: 0.009 mg/kg/day [IFRA, 2002]7. Total Systemic Exposure (Dermal þ Inhalation):

(0.1042 mg/kg/day X 10.3%) þ 0.009 mg/kg/day ¼ 0.020mg/kg/day

*Calculated using the reported 97.5th percentile concentration based on the levels ofthe same fragrance ingredient in ten of the most frequently used personal care andcosmetic products (i.e., anti-perspirant, bath products, body lotion, eau de toilette,face cream, fragrance cream, hair spray, shampoo, shower gel, and toilet soap)(Cadby et al., 2002; Ford et al., 2000).**Combined (fine fragrances, hair sprays, antiperspirants/deodorants, candles,aerosol air fresheners, and reed diffusers/heated oil plug-ins) result calculated usingRIFM's 2-Box/MPPD in silico models, based on the IFRA survey results for the 97.5thpercentile use in hydroalcoholics for a 60 kg individual.

1 VCF Volatile Compounds in Food: database/Nijssen, L.M.; Ingen-Visscher, C.A.van; Donders, J.J.H. [eds]. e Version 15.1 e Zeist (The Netherlands): TNO Triskelion,1963e2014. A continually updated database, contains information on publishedvolatile compounds which have been found in natural (processed) food products.

4. Derivation of systemic absorption

1. Dermal: 10.3% (Read-across from amyl salicylate (CAS # 2050-08-0))

RIFM (1984a, b, c, d): The penetration of radiolabeled isomer

amyl salicylate (CAS # 2050-08-0) through intact pig skin underin vitro conditions using a glass penetration chamber was evaluatedat 1, 6, and 16 h after application. It was concluded that 10.3% ofamyl salicylate was absorbed.

2. Oral: Data not available e not considered.3. Inhalation: Assumed 100%4. Total Assume Dermal (10.3%) þ Inhalation (100%) absorbed¼

(0.1042mg/kg/day X 10.3%)þ 0.009mg/kg/day¼ 0.20mg/kg/day

5. Computational toxicology evaluation

1. Cramer classification: Class I, low

2. Analogues Selected:a. Genotoxicity: Ethyl hexyl salicylate (CAS # 118-60-5)b. Repeated dose toxicity: Amyl salicylate (CAS # 2050-08-0)c. Developmental and reproductive toxicity: Amyl salicylate

(CAS # 2050-08-0); methyl salicylate (CAS # 119-36-8)d. Skin sensitization: Nonee. Phototoxicity/Photoallergenicity: Nonef. Local respiratory toxicity: Noneg. Environmental toxicity: Salicylates SAG

3. Read across justification: See Appendix below

6. Natural occurrence (discrete chemical) or composition(NCS)

Isoamyl salicylate is reported to occur in the following foods1:Grape (Vitis species).Tea.

7. IFRA standard

None.

8. REACH dossier

Pre-registered for 2010; No dossier available as of 7/16/2015.

Includes FEMA GRAS and EU-Flavis data.

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121 S113

9. Summary

1. Human Health Endpoint Summaries:

9.1. Genotoxicity

Based on the current existing data and use levels, isoamyl sa-licylate does not present a concern for genetic toxicity.

9.2. Risk assessment

Isoamyl salicylate was determined not to be genotoxic with orwithout metabolic activation in the BlueScreen assay (RIFM,2013). The mutagenic activity of isoamyl salicylate was assessedin an Ames study conducted in accordance with OECD TG 471using the standard plate incorporation method. Salmonellatyphimurium TA1535, TA1537, TA98, TA100, and TA102 weretreated with isoamyl salicylate in DMSO (dimethyl sulfoxide) atthe concentrations 0,5, 15, 50, 150, 500 and 1500 mg/plate in theabsence of exogenous metabolic activation and 0, 5, 15, 50, 150,500, 1500 and 5000 mg/plate presence and absence of metabolicactivation. No increase in the number of revertant coloniesoutside the range of the vehicle control was observed in any ofthe tester strains at any concentration (RIFM, 1999a, b, c). Underthe conditions of the study, isoamyl salicylate was considered notmutagenic in the Ames test.

There are no studies assessing the clastogenicity of isoamylsalicylate. The clastogenic activity of read across material,ethyl hexyl salicylate (CAS # 118-60-5; see Section V), wasassessed in an in vivo micronucleus study conducted incompliance with GLP regulations and in accordance withOECD TG 474. Groups of male and female NMRI mice wereadministered a single dose of ethyl hexyl salicylate in 10 ml/kg b.w. arachis oil via oral gavage at the concentration of2000 mg/kg b.w. Animals were euthanized at 24, 48 and 72ours post administration and bone marrow smears prepared.There were no increases in the number of micronucleatedpolychromatic erythrocytes in treated samples compared tonegative controls (RIFM, 1989). Under the conditions of thestudy, ethyl hexyl salicylate was considered unable to inducechromosomal damage or damage to the mitotic apparatus inthe bone marrow cells of mice and this can be extended tothe target material, isobutyl salicylate.

Based on the available data, isoamyl salicylate does not present aconcern for genotoxic potential.

Additional references: RIFM, 2013.Literature search and risk assessment completed on: 07/12/13.

9.3. Repeated dose toxicity

The margin of exposure for the repeated dose toxicity endpointis 2350.

9.4. Risk assessment

A dietary 13-week subchronic toxicity study was conductedwith isoamyl salicylate (Drake et al., 1975). RIFM's Expert Panel2

reviewed the study and concluded the NOAEL to be 47 mg/kg/daysince the only finding at this dose was increased relative kidneyweights in females that had no histopathological correlates

2 RIFM's Expert Panel is composed of scientific and technical experts in theirrespective fields. This group provides advice and guidance.

(Belsito et al., 2007). Therefore, the MOE is equal to the NOAELin mg/kg/day divided by the total systemic exposure, 47/0.020or 2350.

Additional references: Lapczynski et al., 2007a; Jimbo, 1983;Novikov et al., 1992; Webb and Hansen, 1963; Cross et al., 1998;Lapczynski et al., 2007b; Webb and Hansen, 1962; Giroux et al.,1954; RIFM, 1978; Hruban et al., 1966; Gage, 1970; Stoner et al.,1973; NTP, 1984; Harrisson et al., 1963; Packman et al., 1961; Barand Griepentrog, 1967; Burdette and Strong, 1941; RIFM, 1984d;Morrissey et al., 1989; Chapin and Sloane, 1997; Pyun, 1970;Warkany and Takacs, 1959; Infurna et al., 1990; Overman andWhite, 1978, 1983; Kavlock et al., 1982; Daston et al., 1988;Bertone and Monie, 1965; Overman, 1979; Woo and Hoar, 1972;Lamb et al., 1997a, 1997b; Gross et al., 1970; Miller et al., 2001;Fang et al., 2003; Hanzlik and Wetzel, 1920; Robinson andWilliams, 1956; Bohnlein et al., 1994; Williams, 1938; Davisonet al., 1961; RIFM, 1984a; RIFM, 1979a; Yano et al., 1986; ScottDuncan et al., 2002; Riviere et al., 2001; Klecak, 1985; Martinet al., 2004; Pratzel et al., 1990; Loveday, 1961; Behrendt andKampffmeyer, 1989; Siddiqi and Ritschel, 1972; Boehnlein et al.,1994; Davis et al., 1981; Danon et al., 1986; Watkinson et al.,1992; Riviere et al., 2000; Cross et al., 1997; Megwa et al., 1995;Higo et al., 1995; Kasting et al., 1987; Brown and Scott, 1934a,1934b; Bliss, 1935; Meyer, 1965; Beutner et al., 1943; Lapczynskiet al., 2007c;RIFM, 1979b; RIFM, 1984b; RIFM, 1995a; RIFM, 1996a,b; RIFM, 1995b; Treffel and Gabard, 1996; RIFM, 2002a;Lapczynski et al., 2007d; Morohoshi et al., 2005; RIFM, 2002b;RIFM, 2002c, RIFM, 1995c.

Literature search and risk assessment completed on: 07/12/13.

9.5. Developmental and reproductive toxicity

Themargin of exposure for the developmental and reproductivetoxicity endpoints is 3750.

9.6. Risk assessment

There are no developmental or reproductive toxicity data onisoamyl salicylate. Read across material methyl salicylate (CAS #119-36-8; see Section V) has a dietary 3-generation reproductivetoxicity study in rats, which concluded a NOAEL of 75 mg/kg/daybased on litter parameters, weanling weights, and fertility indices(Collins et al., 1971). Therefore, the MOE is equal to the NOAEL inmg/kg/day divided by the total systemic exposure, 75/0.020 or3750.

Additional references: Lapczynski et al., 2007a; Jimbo, 1983;Novikov et al., 1992; Webb and Hansen, 1963; Cross et al., 1998;Lapczynski et al., 2007b; Webb and Hansen, 1962; Giroux et al.,1954; RIFM, 1978; Hruban et al., 1966; Gage, 1970; Stoner et al.,1973; NTP, 1984; Harrisson et al., 1963; Packman et al., 1961; Barand Griepentrog, 1967; Burdette and Strong, 1941; RIFM, 1984a, b,c, d; Morrissey et al., 1989; Chapin and Sloane, 1997; Pyun, 1970;Warkany and Takacs, 1959; Infurna et al., 1990; Overman andWhite, 1978, 1983; Kavlock et al., 1982; Daston et al., 1988;Bertone and Monie, 1965; Overman, 1979; Woo and Hoar, 1972;Lamb et al., 1997a, 1997b; Gross et al., 1970; Miller et al., 2001;Fang et al., 2003; Hanzlik and Wetzel, 1920; Robinson andWilliams, 1956; Bohnlein et al., 1994; Williams, 1938; Davisonet al., 1961; RIFM, 1984a; RIFM, 1979a; Yano et al., 1986; ScottDuncan et al., 2002; Riviere et al., 2001; Klecak, 1985; Martinet al., 2004; Pratzel et al., 1990; Loveday, 1961; Behrendt andKampffmeyer, 1989; Siddiqi and Ritschel, 1972; Boehnlein et al.,1994; Davis et al., 1981; Danon et al., 1986; Watkinson et al.,1992; Riviere et al., 2000; Cross et al., 1997; Megwa et al., 1995;

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121S114

Higo et al., 1995; Kasting et al., 1987; Brown and Scott, 1934a,1934b; Bliss, 1935; Meyer, 1965; Beutner et al., 1943; Lapczynskiet al., 2007c;RIFM, 1979b; RIFM, 1984b; RIFM, 1995a; RIFM, 1996a,b; RIFM, 1995b; Treffel and Gabard, 1996; RIFM, 2002a;Lapczynski et al., 2007d; Morohoshi et al., 2005; RIFM, 2002b;RIFM, 2002c, RIFM, 1995c.

Literature search and risk assessment completed on: 07/12/13.

9.7. Skin sensitization

Based on the existing data, isoamyl salicylate does not present aconcern for skin sensitization.

9.8. Risk assessment

Based on the existing data, isoamyl salicylate does not present aconcern for skin sensitization. It is predicted to be non-reactive toskin proteins and therefore would not be expected to act as a skinsensitizer (Roberts et al., 2007; Toxtree 2.5.0; OECD Toolbox v3.1).Also, based on available guinea pig test data, isoamyl salicylate isnot a skin sensitizer (Ishihara et al., 1986; RIFM, 1970).

RIFM's Expert Panel reviewed the available data and concludedthat isoamyl salicylate does not present a concern for skin sensiti-zation (Belsito et al., 2007).

Additional references: None.Literature search and risk assessment completed on: 07/12/13.

9.9. Phototoxicity/Photoallergenicity

Based on UV/Vis absorption spectra, isoamyl salicylate wouldnot be expected to present a concern for phototoxicity orphotoallergenicity.

9.10. Risk assessment

There are no phototoxicity studies available for isoamyl salicy-late in experimental models. UV/Vis absorption spectra indicateminor absorbance between 290 and 700 nm. The molar absorptioncoefficient is below the benchmark (1000 L/mol ∙ cm�1) of concernfor phototoxic effects (Henry et al., 2009). Based on lack of signif-icant absorbance in the critical range, isoamyl salicylate does notpresent a concern for phototoxicity or photoallergenicity.

Additional references: None.Literature search and risk assessment completed on: 07/12/13.

9.11. Local respiratory toxicity

The isoamyl salicylate exposure level is below the inhalation TTCCramer Class I limit for local effects.

9.12. Risk assessment

There are no inhalation data available in the database. Based onthe IFRA survey results for hydroalcoholics, the 97.5th percentilewas reported to be 4.09%. Assuming the same amount is used in allproduct types (fine fragrances, hair sprays, antiperspirants/de-odorants, candles, aerosol air fresheners, and reed diffusers/heatedoil plug-ins) the inhalation aggregate exposure would be 0.54 mg/day, as calculated by RIFM's 2-Box Model and further refined usingthe Multiple Path Particle Deposition Model, using the 97.5thpercentile IFRA survey hydroalcoholic use value. This value is belowthe Cramer Class I TTC level of 1.4 mg/day (based on human lungweight of 650 g; Carthew et al., 2009) and is deemed safe for use atthe reported use level.

Additional references: None.

Literature search and risk assessment completed on: 07/12/13.

2. Environmental endpoint summary

9.13. Screening-level assessment

A screening level risk assessment of isoamyl salicylate wasperformed following the RIFM Environmental Framework (Salvitoet al., 2002) which provides for 3 levels of screening for aquaticrisk. In Tier 1, only the material's volume of use in a region, its logKow and molecular weight are needed to estimate a conservativerisk quotient (RQ; Predicted Environmental Concentration/Pre-dicted No Effect Concentration or PEC/PNEC). In Tier 1, a generalQSAR for fish toxicity is used with a high uncertainty factor asdiscussed in Salvito et al. (2002). At Tier 2, the model ECOSAR(providing chemical class specific ecotoxicity estimates) is used anda lower uncertainty factor is applied. Finally, if needed, at Tier 3,measured biodegradation and ecotoxicity data are used to refinethe RQ (again, with lower uncertainty factors applied to calculatethe PNEC). Following the RIFM Environmental Framework, isoamylsalicylate was identified as a fragrance material with the potentialto present a possible risk to the aquatic environment (i.e., itsscreening level PEC/PNEC >1).

A screening-level hazard assessment using EPISUITE ver 4.1 didnot identify isoamyl salicylate as either being possibly persistentnor bioaccumulative based on its structure and physicalechemicalproperties. This screening level hazard assessment is a weight ofevidence review of a material's physicalechemical properties,available data on environmental fate (e.g., OECD Guidelinebiodegradation studies or die-away studies) and fish bio-accumulation, and review of model outputs (e.g., US EPA's BIOWINand BCFBAF found in EPISUITE ver.4.1). Specific key data onbiodegradation and fate and bioaccumulation are reported belowand summarized in the Environmental Safety Assessment sectionprior to Section I.

9.14. Risk assessment

Based on current VoU (as of 2011) isoamyl salicylate presents arisk to the aquatic compartment.

9.15. Key studies

Biodegradation: No data available. Please see “other availabledata” section for read across data

9.15.1. EcotoxicityRIFM,1999b: The acute toxicity in Daphnia magnawas evaluated

using a static system. The EC0 at 48 h was 2 mg/l; the EC100 was4 mg/l. The geometric mean determined at 48 h was 2.8 mg/l.

RIFM, 1983: A 24 h acute toxicity test with D. magna was con-ducted with test material. The EC50 at 24 h was 4.5 mg/l.

RIFM, 2000a, b, c: An acute toxicity study was conducted for96 h in freshwater fish (Danio rerio) according to the OECD 203guidelines. The LC50 was reported to be between 10 and 100 mg/l.

Other available data: Isoamyl salicylate has been pre-registeredfor REACH with no additional data. For the summary of read-acrossdata, see Appendix below.

9.16. Risk assessment refinement

Ecotoxicological data and PNEC derivation (all endpoints re-ported in mg/L; PNECs in mg/L) Endpoints used to calculate PNECare underlined.

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121 S115

Exposure information and PEC calculation (following RIFMEnvironmental Framework: Salvito et al., 2002).

Exposure Europe (EU) North America (NA)

Log Kow used 4.49 4.49Biodegradation Factor Used 1 1Dilution Factor 3 3Regional Volume of Use Tonnage Band 100e1000 10e100

Risk Characterization: PEC/PNEC <1 <1

Based on read across, the RQs for these materials are <1. No further assessment isnecessary.

3 Information sources outside of RIFM's database are noted as appropriate in the

The RIFM PNEC is 0.28 mg/L. The revised PEC/PNECs for EU andNA are <1 and therefore, do not present a risk to the aquaticenvironment at the current reported volumes of use.

Literature search and risk assessment Completed on: 07/12/13.

10. Literature search3

� RIFM database: Target, Fragrance Structure Activity Groupmaterials, other references, JECFA, CIR, SIDS

� ECHA: http://echa.europa.eu/� NTP: http://tools.niehs.nih.gov/ntp_tox/index.cfm� OECD toolbox� SciFinder: https://scifinder.cas.org/scifinder/view/scifinder/scifinderExplore.jsf

� PUBMED: http://www.ncbi.nlm.nih.gov/pubmed� TOXNET: http://toxnet.nlm.nih.gov/� IARC: (http://monographs.iarc.fr)� OECD SIDS: http://www.chem.unep.ch/irptc/sids/oecdsids/sidspub.html

� EPA Actor: http://actor.epa.gov/actor/faces/ACToRHome.jsp;jsessionid¼0EF5C212B7906229F477472A9A4D05B7

� US EPA HPVIS: http://www.epa.gov/hpv/hpvis/index.html

safety assessment.

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121S116

� US EPA Robust Summary: http://cfpub.epa.gov/hpv-s/� Japanese NITE: http://www.safe.nite.go.jp/english/db.html� Japan Existing Chemical DataBase: http://dra4.nihs.go.jp/mhlw_data/jsp/SearchPageENG.jsp

� Google: https://www.google.com/webhp?tab¼ww&ei¼KMSoUpiQK-arsQS324GwBg&ved¼0CBQQ1S4

This is not an exhaustive list.

Appendix

Principal name Target material Read across materials

Isoamyl salicylate Ethyl hexyl salicylate

CAS No. 87-20-7 118-60-5Structure

3D structure http://www.thegoodscentscompany.com/opl/87-20-7.html

http://www.thegoodsccompany.com/opl/118-60-5.html

Read-across endpoint � Genotoxicity

Molecular formula C12H16O3 C15H22O3

Molecular weight 208.26 250.34Melting point

(�C, EPISUITE)82.45 108.87

Boiling point(�C, EPISUITE)

306.01 344.94

Vapor pressure(Pa @ 25 �C,EPISUITE)

0.08679 0.0009506

Log Kow (KOWWINv1.68 in EPISUITE)

4.49 5.97

Water solubility(mg/L, @ 25 �C,WSKOW v1.42in EPISUITE)

21.89 0.7171

Jmax (mg/cm2/h, SAM) 41.58888349 5.518241291Henry's Law

(Pa$m3/mol,Bond method,EPISUITE)

1.430709 3.347778

Similarity (Tanimotoscore)1

71%

Skin absorptionSkin absorption

percentage(SAM)

80%

GenotoxicityDNA binding

(OASIS v1.1)� No alert found � No alert found

DNA binding(OECD)

� No alert found � No alert found

Carcinogenicity(genotox andnon-genotox)alerts (ISS)

� No alert found � Structural alert fornongenotoxiccarcinogenicity

� Substitutedn-alkylcarboxylicacids (Nongenotox)

DNA alerts forAmes, MN,CA (OASIS v1.1)

� No alert found � No alert found

In vitro mutagenicity(Ames test) alerts(ISS)

� No alert found � No alert found

In vivo mutagenicity(Micronucleus)alerts (ISS)

� H-acceptor-path3-H-acceptor � H-acceptor-path3-H-

Summary

There are insufficient toxicity data on isoamyl salicylate (RIFM #102, CAS # 87-20-7). Hence, in silico evaluation was conducted todetermine suitable read-across material. Based on structural simi-larity, reactivity, metabolism data, physicochemical properties andexpert judgment, the above shown read-across materials wereidentifiedasproper readacross for their respective toxicityendpoints.

Amyl salicylate Methyl salicylate

2050-08-0 119-36-8

ents http://www.thegoodscentscompany.com/opl/2050-08-0.html

http://www.thegoodscentscompany.com/opl/119-36-8.html

� Skin Absorption� Repeated Dose� Devel/Repro

�Devel/Repro

C12H16O3 C8H8O3

208.26 152.1590.74 51.47

314.48 252.18

0.1123 7.119

4.57 2.60

18.94 1875

36.62002466 292.14132951.430709 0.460725

79% 66%

80%

acceptor

(continued )

Principal name Target material Read across materials

Isoamyl salicylate Ethyl hexyl salicylate Amyl salicylate Methyl salicylate

Oncologicclassification(OECD)

� Phenol Type Compounds � Phenol Type Compounds

Repeated dosetoxicity

Repeated dose (HESS) Not categorized Not categorizedDevelopmental and reproductive toxicityER binding (OECD) Strong binder, OH group Strong binder, OH group Weak binder, OH groupDevelopmental toxicity

model (CAESAR v2.1.6)Toxicant (moderate reliability) NON-Toxicant (moderate reliability) NON-Toxicant (low reliability)

MetabolismRat liver S9 metabolism

simulator (OECD)See Supplemental data 1 See Supplemental data 2 See Supplemental data 3 See Supplemental data 4

1Values calculated using J. Chem with FCFP4 1024 bits fingerprint (Rogers and Hahn, 2010).

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121 S117

Methods

� The identified read-across analogs were confirmed by usingexpert judgment

� The physicochemical properties of target and analogs werecalculated using EPI Suite™ v4.11 developed by US EPA (USEPA,2012)

� The Jmax were calculated using RIFM skin absorption model(SAM), the parameters were calculated using consensus model(Shen et al., 2014)

� DNA binding, mutagenicity, genotoxicity alerts and oncologicclassification were estimated using OECD QSAR Toolbox (v3.1)(OECD, 2012)

� ER binding and repeat dose categorizationwere estimated usingOECD QSAR Toolbox (v3.1) (OECD, 2012)

� Developmental toxicity and skin sensitization were estimatedusing CAESAR (v.2.1.6) (Cassano et al., 2010)

� The major metabolites for the target and read-across analogswere determined and evaluated using OECD QSAR Toolbox(v3.1) (OECD, 2012)

Conclusion/Rationale

� Ethyl hexyl salicylate (analog) was used as a read-across forisoamyl salicylate (target) based on:^ The target and analog both belong to the generic class of ar-omatic esters, specifically salicylates.

^ The key differences are that the target has an isopentyl chain,while the analog has a 2-ethyl hexyl chain. These differencesdo not materially change the physicochemical properties norraise any additional structural alerts and therefore, the geno-toxicity profiles are expected to be similar.

^ Both the target and the analog show similar alerts for DNAbinding and mutagenicity. The analog shows additional alertsfor non genotoxicity carcinogenicity. However, such alerts donot have genotoxicity concerns.

^ As per the OECD Toolbox both the materials are predicted tohave similar metabolites.

� Amyl salicylate (analog) was used as a read-across for isoamylsalicylate (target) based on:^ The target and analog both belong to the generic class of ar-omatic esters, specifically salicylates.

^ Botharestructural isomers. Thekeydifferencesare that the targethas an isopentyl chain,while the analog has a 2 ethyl hexyl chain.These differences do not materially change the physicochemicalpropertiesnor raise anyadditional structural alerts and therefore,the repeated dose toxicity profiles are expected to be similar.

^ They both also show similar alerts for Repeated Dose (HESS)Categorization.

^ Bothmaterials are expected to bemetabolized similarly. As perthe OECD Toolbox both the materials are predicted to havesimilar metabolites.

� Methyl salicylate (analog) was used as a read-across for isoamylsalicylate (target) based on:^ The target and analog belong to the generic class of aromaticesters, specifically salicylates.

^ The only difference between the target andmethyl salicylate isthat they have different alcohol parts. These differences do notmaterially change the physicochemical properties nor raiseany additional structural alerts and therefore, the reproductiveand developmental toxicity profiles are expected to be similar.

^ They both also show similar alerts for Repeated Dose (HESS)Categorization and ER Binding. ER Binding is molecular initi-ating event. ER binding is not necessarily predictive of endo-crine disruption given the complex pre- and post-receptorevents that determine activity.

^ As per the OECD Toolbox both the materials are predicted tohave similar metabolites.

Environmental toxicity justification

Within the RIFM Database there are a number of salicylatematerials that are structurally related. Some of thesematerials havemeasured data available which can be useful in assessing the safetyof other “data poor” materials. For details see below.

Ecotoxcity

There are a number of studies that were conducted for multiplesalicylates; unfortunately most of these studies did not followgenerally acceptable guidelines (ex. OECD) or the reported resultswere not supported by appropriate analytical analyses. Therefore,to support the most conservative approach for safety for this classof materials, a 72 h EC50 of 0.28 mg/l for an Algae Inhibition Studyconducted with cis-E-hexenyl salicylate was selected as the key

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121S118

study/endpoint in deriving the PNEC for this class of chemicals. Thisvalue was derived based on standard methodology (OECD 201guidelines) following GLP requirements. The shorter chain lengthsalicylates would be less toxic; therefore using the higher chainlength measured value with an assessment factor of 1000 providesa conservatively derived PNEC for this class of materials.

Biodegradation

There are a number of biodegradation studies, using different

Name CAS Structure Kow/Watersolubility

Biodegradation (and hazard asses

Amyl salicylate 2050-08-0 4.4 and 4.5(measured)

301B: 81.3% - RIF86% - RIFM, 199692/69/EEC C.4: 879/831/EVG V.C:

Benzoic acid, 2-hydroxy-, 2-methylbutylester

51115-63-0 4.4 and 4.5(measured)/21.89 mg/l

N/A

Butyl salicylate 2052-14-4 4.08/19.78 mg/l

N/A

1,3-Dimethyl-3-butenylsalicylate

80118-10-1 4.91/8.43 mg/l

N/A

Ethyl hexylsalicylate

118-60-5 5.97/0.71 mg/l

84/449 EEC: 89%

Hexyl salicylate 6259-76-3 5.5(measured)/6.084 mg/l

OECD 301F: 91%OECD 301B: 89.9

OECD guidelines that demonstrate similar degradation rates acrossthe class, which indicates that a read e across between these ma-terials is appropriate.

For details on Ecotoxicity and biodegradation studies availablein RIFM Database see table below.

In addition, 1 hexyl salicylate (CAS# 6259-76-3) has beenregistered under REACH and the following additional data isavailable:

Hexyl salicylate: OECD 202: Daphnia magna: 48 h EC50:0.357 mg/l.

all useful for safetysment)

Ecotoxicity

Study Usefulness/Notes

M, 1994b; 301F:a4% - RIFM, 2001a29% - RIFM, 2001b

96 h Fish LC50: 1.34 mg/l (LC0/LC100) (RIFM,1993)96 h Danio rerio OECD 203: LC50: 10e100 mg/l(RIFM, 2000c, http://rifmdatabase.rifm.org/RifmDatabase/Studies/42556)

Useful withlimitations

N/A N/A

N/A N/A

N/A N/A

(RIFM, 1992) 48 h Daphnia magna 67/548/EEC C.2: EC50:between 10 and 31 mg/l (RIFM, 1992)96 h Brachydanio rerio acute: LC50: 613 mg/l(RIFM, 1992)

Useful withlimitations

(RIFM, 1995a)% (RIFM, 1994a)

48 h acute Daphnia Magna: EC0/EC100:0.39 mg/l (RIFM, 1999c)24 h Acute Daphnia magna EC50: 1.5 mg/l(RIFM, 1983)96 h Danio rerio Acute OECD 203; LC50:>100 mg/l (RIFM, 2000b)

Useful withlimitations

(continued )

Name CAS Structure Kow/Watersolubility

Biodegradation (all useful for safetyand hazard assessment)

Ecotoxicity

Study Usefulness/Notes

Isoamyl salicylate 87-20-7 4.49/21.89 mg/l

N/A 24 h Acute Daphnia magna EC50: 4.5 mg/l(RIFM, 1983)48 h acute Daphnia Magna: EC0/EC100: 2.8 mg/l(RIFM, 1999a)96 h Danio rerio Acute OECD 203; LC50: 10e100 mg/l (RIFM, 2000a)

Useful withlimitations

Isobutyl salicylate 87-19-4 4.0/67.83 mg/l

N/A N/A N/A

cis-E-Hexenylsalicylate

65405-77-8 4.6/9.518 mg/l

OECD 301F: 89% (RIFM, 1995e)OECD 301B: 56% (RIFM, 1995d)http://rifmdatabase.rifm.org/RifmDatabase/Studies/34959

48 h Daphnia Magna OECD 202: EC50: 2.7 mg/l;72 h Algae OECD 201: 0.28 mg/l (RIFM, 2010)

Useful withlimitations

A.M. Api et al. / Food and Chemical Toxicology 84 (2015) S110eS121 S119

Appendix A. Supplementary data

Supplementary data related to this article can be found at http://dx.doi.org/10.1016/j.fct.2015.09.014.

Transparency document

Transparency document related to this article can be foundonline at http://dx.doi.org/10.1016/j.fct.2015.09.014.

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