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Key value for chemical safety assessment

Effects on fertility

Description of key information
While some in vitro endocrine disruptor assays have demonstrated that BBP shows weak oestrogenic activity, there is little evidence of such an effect in vivo, either for BBP itself or for its two major metabolites, MBuP and MBeP.
Effect on fertility: via oral route
Dose descriptor:
NOAEL
250 mg/kg bw/day
Additional information

Studies in laboratory animals

Effects of BBP on fertility and reproductive function have been studied in rodents, primarily rats. Of the studies that attempted to establish a no-observed-adverse-effect level (NOAEL) for reproductive effects, those by Tyl et al. (2002), Nagao et al. (2000) and the National Toxicology Program (NTP, 1997f) are the most important.

 

In the study by Tyl et al. (2002), a reliable two-generation reproductive toxicity study, groups of 30 male and 30 female rats received Santicizer 160 (BBP) in the diet at 0, 50, 250 or 750 mg/kg bw/day for 10 weeks prior to mating and during the 2-week mating period. The males were then killed, while the females continued with the same treatment throughout gestation, lactation and weaning, until sacrifice on postnatal day (PND) 21. The F1 offspring (reduced to 10 pups/litter on PND 4) were reared by their dams to PND 21, at which point at least one pup/sex/litter was randomly selected to produce the second (F2) generation and continued under the same exposure regime as the parents. Systemic toxicity was seen in all three generations at the top dose, which also produced reproductive toxicity. In the F0 generation this was limited to the females (with reductions in the relative weights of the ovary and uterus), while in the F1 generation both males and females showed a reduction in mating and fertility indices. In the F1 and F2 offspring, growth was impaired and sexual maturation delayed at 750 mg/kg bw/day. The no-observed-adverse-effect level (NOAEL) for reproductive and systemic toxicity was 250 mg/kg bw/day.

 

In an earlier, reliable, two-generation study, groups of 20-24 male and female rats were given BBP at 0, 20, 100 or 500 mg/kg bw/day by gavage, for 12 weeks (males) and 2 weeks (females) prior to mating and during the 2-week mating period. The males and any non-pregnant females were then killed, while the pregnant females continued on the same treatment throughout pregnancy, lactation and weaning. The F1 offspring (reduced to 8 pups/litter on PND 4) were reared by their dams until PND 21, when at least one pup/sex/litter was randomly selected to produce the second (F2) generation. Ten F1 weanlings of each sex (not selected for mating) in the control and high-dose groups were examined microscopically for changes in the reproductive organs on postnatal day 22. Reproductive ability was unaffected in either the parental or F1 generations, while systemic toxicity (changes in organ weights) was evident in both generations at 100 mg/kg bw/day and above. In addition, high-dose F1 male weanlings showed a significant reduction in testis and epididymis weights, accompanied by testicular damage, while high-dose F1 females had a decrease in ovary weight and an increase in uterine weight. A decrease in testosterone levels was evident in high-dose F0 and F1 males. In the F2 generation, there were no significant treatment-related effects by postnatal day 21. [See Developmental toxicity section below for effects on pup development.] The NOAEL for reproductive toxicity was considered to be 100 mg/kg bw/day, while that for systemic toxicity was 20 mg/kg bw/day (Nagao et al., 2000). [This NOAEL of 20 mg/kg bw/day for systemic toxicity was not used for DNEL derivation, as the effects at the next dose up (100 mg/kg bw/day) were limited to organ weight changes, and the spacing of the doses in this study was poor.]

 

The National Toxicology Program conducted a 10-week feeding study, in which groups of 15 male rats were given dietary levels of 0, 300, 2800 or 25000 ppm (equivalent to around 0, 20, 200 or 2200 mg/kg bw/day), and then each male was mated with two untreated females for 7 days. Blood was collected for haematological examination from the males at study termination and a comprehensive range of tissues from the 25000 ppm and control groups were examined microscopically. The testis, epididymis, seminal vesicle and prostate were also examined in the other treated groups. A necropsy was performed on the dams on day 13 of gestation, or 13 days after the end of the mating period for virgin females, and the uterus and ovaries were grossly examined. At study termination, the high-dose males showed a lower mean body weight than the controls (30% less), a minimal, macrocytic anaemia, slight increases in the relative heart, liver, lung and thymus weights (P <0.01), marked reductions in the absolute weights of the testis, cauda and epididymis (P <0.01), increased incidences of necrosis and atrophy of the seminiferous tubules and hypospermia in the epididymis (P < 0.01), and a huge reduction in the concentration of sperm in the cauda epididymal tissue (to about 0.2% that in the control males). There was a less marked reduction in sperm concentration in the mid-dose males (P <0.05). Effects of the test substance on other tissues and organs were not included in the report. Although 10 of the 30 females mated with the high-dose males were initially sperm positive, none showed any evidence of pregnancy when examined on gestation day 13. The numbers of pregnancies resulting from mating with the low- and mid-dose groups were comparable to the control group, and there were no differences in litter size or in the numbers of dead foetuses, implantations or resorptions (NTP, 1997f).

 

Thus, in the above NTP study, dietary administration of BBP to male rats at 25000 ppm (equivalent to around 2200 mg/kg bw/day) for 10 weeks prior to mating with (unexposed) virgin females resulted in no pregnancies (despite 10 of the 30 females being initially sperm positive). This dose level caused degenerative changes to the testis and epididymis, and the sperm concentration in the cauda epididymis was about 0.2% of the level in the control group. A 30% reduction in sperm concentration was also observed in the mid-dose group given 2800 ppm in the diet (around 200 mg/kg bw/day), although this had no effect on the number of pregnancies. The NOAEL in this study was therefore 300 ppm (20 mg/kg bw/day) (NTP, 1997f). However, this value is not carried forward as the lowest NOAEL for reproductive toxicity, as the sperm effects observed at 200 mg/kg bw/day in this study were not reproducible at 550 mg/kg bw/day in a subsequent 26-week feeding study by the NTP (see next paragraph).

 

The 26-week feeding study referred to above involved groups of 15 male rats given 0, 300, 900, 2800, 8300 or 25000 ppm BBP in their diet, corresponding to around 0, 30, 60, 180, 550 or 1660 mg/kg bw/day (NTP, 1997b). These males were not mated, but the effects of treatment on the testis, epididymis, seminal vesicle and prostate were assessed in all groups, and epididymal spermatozoal parameters were examined in the 0, 300, 8300 and 25000 ppm groups. (Although a comprehensive range of tissues and organs from the high-dose and control groups were examined microscopically on study termination, only the effects on the reproductive organs were discussed.) Blood was collected for haematological examination at various intervals throughout the study. At study termination, the mean body weight of the 25000 ppm group was 30% less than that of the control group. A minimal to mild macrocytic anaemia was detected at 30 days in the high-dose group and continued throughout the study. The absolute weights of the testis, cauda and epididymis were reduced (P <0.01) in this group, as was the absolute prostate weight, while relative liver and kidney weights were increased (P <0.01). Incidences of hypospermia and atrophy of the seminiferous tubules and hypospermia in the epididymis were significantly greater (P < 0.01) in the high-dose group, and the concentration of sperm in the cauda epididymal tissue was less than 1% of that in the control group. Although this 26-week study revealed degenerative changes to the testis and epididymis at 25000 ppm (equivalent to around 1660 mg/kg bw/day), no such effects were evident at 8300 ppm (550 mg/kg bw/day). The latter is therefore the NOAEL for reproductive toxicity in this study. Thus, the observation of decreased caudal epididymal spermatozoa concentration at 200 mg/kg bw/day in the NTP’s 10‑week feeding study could not be replicated in their 26-week feeding study conducted in the same strain of rat (F344/N) at higher dose levels.

 

In a combined reproductive/developmental screening test (OECD Guideline No. 421), groups of 10 male and 10 female rats were treated orally (by gavage) with 0, 250, 500 or 1000 mg/kg bw/day for 2 weeks prior to mating and throughout the mating period. Treatment then continued, for a total dosing period of 29 days in the males, and throughout pregnancy up until postnatal day 6 in the females, followed by necropsy examination of the parental animals. Effects were observed only at the top dose, with reductions in body weight, pregnancy rate, and testicular and epididymal weights, and evidence of testicular degeneration. The NOAEL for reproductive toxicity was 500 mg/kg bw/day (Piersma et al., 1995).

A one-generation reproductive toxicity study (GLP-compliant and in accordance with OECD Guideline No. 415) was conducted in which groups of 12 male and 24 female rats were given 0, 0.2, 0.4 or 0.8% BBP in the diet (providing around 0, 108, 206 or 418 mg/kg bw/day for males and 0, 106, 217 or 446 mg/kg bw/day for females). Males were treated for a 10-week period prior to mating, females for 2 weeks prior to mating, and treatment continued for the production of two litters. Fertility and reproductive performance were evaluated, and there was microscopic examination of male and female reproductive organs (ovaries, uterus, vagina, testis, epididymis, seminal vesicle, prostate, coagulating gland) and of the pituitary. Females in the high-dose group showed a reduction in mean body weight and body-weight change during gestation and lactation, and an increase in relative liver weight. The NOAEL for parental animals was therefore established at 0.4% in the diet (217 mg/kg bw/day). There was no evidence of adverse effects on fertility, pregnancy or offspring development in either of the two litters that were produced, and the NOAEL for reproductive effects was therefore 0.8% (around 430 mg/kg bw/day) (Monsanto, 1993, cited in EU RAR, 2007).

 

In an earlier study in which male rats were treated daily by gastric intubation at dose levels of 0, 160, 480 or 1600 mg/kg bw/day for 14 days, absolute testis weights (and relative liver weights) were reduced at the top dose and severe testicular atrophy was evident in these males. One of three males given 480 mg/kg bw/day also showed less severe testicular lesions, so the NOEL was determined as 160 mg/kg bw/day (Lake et al., 1978). The same investigators also conducted a 4-day oral study in rats, comparing the testicular effects of BBP and its metabolites, monobutyl phthalate (MBuP) and monobenzyl phthalate (MBeP). BBP induced atrophic changes in 3/6 and 5/6 males at 800 and 1600 mg/kg bw/day respectively, while all males treated with MBuP and MBeP (at doses equimolar to 1200 mg/kg bw/day of the diester) showed testicular atrophy. MBuP produced more severe effects than did MBeP (Lake et al., 1978).

 

Human Experience

In two studies that evaluated phthalate exposure and semen quality in 168 and 463 men attending fertility clinics (Duty et al., 2003; Hauser et al., 2006), there was an association between high levels of MBeP and/or MBuP in the urine (measured only in a single spot urine sample) and semen concentration, motility and morphology. However, no conclusions can be drawn on any possible relationship between semen quality and BBP exposure due to the mixed phthalate exposures, other environmental and nutritional exposures, the small group sizes, and the unknown impact of underlying reproductive pathology.

 

 


Short description of key information:
Many rodent studies have been conducted to assess the reproductive toxicity of BBP. The compound affects the male reproductive organs, causing testicular damage and reduced sperm count at doses of around 500 mg/kg bw/day and above. The highest NOAEL is 250 mg/kg bw/day. Impaired fertility has been observed at 750 mg/kg bw/day.

Effects on developmental toxicity

Description of key information
In a reliable 2-generation rat study involving oral administration of BBP at up to 750 mg/kg bw/day, F1 and F2 male pups showed a reduced anogenital distance (an indication of foetal androgen action) from 250 mg/kg bw/day, providing a no-observed-effect level (NOEL) for developmental toxicity of 50 mg/kg bw/day. Two other 2-generation rat studies provided no-observed-adverse-effect levels (NOAELs) for developmental toxicity of <100 and 100 mg/kg bw/day, while higher NOAELs have been obtained in shorter-term developmental toxicity assays. 
Effect on developmental toxicity: via oral route
Dose descriptor:
NOAEL
50 mg/kg bw/day
Additional information

Studies in laboratory animals

Numerous studies have been conducted in rodents to assess the developmental toxicity of BBP and its major metabolites (monobutyl and monobenzyl phthalates), and have demonstrated prenatal mortality, reduced foetal weight and foetal malformations, generally at doses that were also toxic to the dams (reduced body-weight gain, increased liver or kidney weights).

 

In a reliable two-generation reproductive toxicity study by Tyl et al. (2002), Santicizer 160 (BBP) was given in the diet of rats (30/sex/group) at concentrations of 0, 750, 3750 or 11250 ppm (corresponding to around 0, 50, 250 or 750 mg/kg bw/day). Treatment was for 10 weeks prior to mating and throughout the 2-week mating period, after which the males were killed while the females continued with the same treatment throughout gestation, lactation and weaning, until sacrifice on postnatal day (PND) 21. The F1 offspring (reduced to 10 pups/litter on PND 4) were reared by their dams to PND 21, at which point at least one pup/sex/litter was randomly selected to produce the second (F2) generation and continued under the same exposure regime as the parents. The NOAEL for developmental toxicity was 50 mg/kg bw/day, as there was a dose-related reduction in anogenital distance (AGD) in F1 and F2 male offspring from 250 mg/kg bw/day, statistically significant compared to the control group even after adjusting for individual body weights. (A reduction in AGD at birth is considered one of the most sensitive indicators of anti-androgenic activity.) Other developmental effects observed in the F1 and/or F2 generations at the top dose of 750 mg/kg bw/day included a significant increase in male pups with one or more nipples and/or areolae, and effects on the male reproductive organs (reduced weights and gross or histopathological changes). The NOAEL for maternal toxicity was 250 mg/kg bw/day based on organ weight changes (liver and kidney) and histopathological lesions graded as minimal in the liver at 750 mg/kg bw/day.

 

In an earlier, reliable, two-generation study by Nagao et al. (2000), BBP was administered to groups of 20-25 rats by gavage (0, 20, 100 or 500 mg/kg bw/day) for 12 weeks (males) and 2 weeks (females) prior to mating and during the 2-week mating period. The males and any non-pregnant females were then killed, while the pregnant females continued on the same treatment throughout pregnancy, lactation and weaning. The F1 offspring (reduced to 8 pups/litter on post-natal day 4) were reared by their dams until post-natal day 21, when at least one pup/sex/litter was randomly selected to produce the second (F2) generation. Ten F1 weanlings of each sex (not selected for mating) in the control and high-dose groups were examined microscopically for changes in the reproductive organs on postnatal day 22. There was a reduction in body weight at birth in the F1 offspring from 100 mg/kg bw/day, and at the top dose the weaned pups (the males in particular) remained lighter in weight throughout the study in comparison with controls. Systemic toxicity (changes in organ weights) was evident in the parental and F1 generations at 100 mg/kg bw/day and above. In addition, high-dose F1 females had a decrease in ovary weight and an increase in uterine weight, while high-dose F1 male weanlings showed a significant reduction in testis, epididymis and seminal vesicle weights, accompanied by testicular damage, as well as preputial separation delay and a significant reduction in anogenital distance (AGD, an indication of androgenic action). Reproductive outcome, growth, development and viability of the F2 pups were unaffected by treatment with BBP. The reduction in F1 maleAGDat PND 0 was not statistically different from control; moreover, since the impact of an adjusted pup body weight onAGDwas not provided, it is not clear if the decreasedAGDwas a co-variate of reduced pup body weight or a frank effect of treatment. The NOAEL for developmental toxicity was considered to be 100 mg/kg bw/day, while that for systemic toxicity was 20 mg/kg bw/day.

 

In another two-generation rat study (not reported in the EU RAR), Aso et al. (2005) gave BBP by oral gavage to rats at doses of 0, 100, 200 or 400 mg/kg bw/day. Systemic toxicity was noted in mid- and high-dose F0 males and females (increased relative kidney and liver weights, decreased relative uterus weight in mid-dose females). Reproductive endpoints for F0 animals were unaffected by treatment. Data on pup body weights were not provided, but AGDs were said to correlate with pup body weight. On postnatal day 4, F1 (but not F2) female pups at all treatment levels showed an increase in AGD that was statistically-significantly different from controls when adjusted for body weight, although there was no dose-response relationship. F1 male pups did not demonstrate a change in AGD on postnatal day 4, whereas F2 male pups at all treatment levels showed a significant, dose-related, decrease in AGD at the same time point (when adjusted for body weight). F1 male adults had testicular changes (weight and histopathology) in the high-dose group. The NOAEL for developmental toxicity in this study was <100 mg/kg bw/day (Aso et al., 2005).

 

A reliable teratogenicity study was performed by the NTP (Field, 1989), in which groups of 30 pregnant rats were given diets containing 0, 0.5, 1.25 or 2% BBP (providing 0, 420, 1100 or 1640 mg/kg bw/day respectively) on days 6-15 of pregnancy, and the dams were killed on day 20 of pregnancy. The uterus, liver and kidney were weighed, and some livers from the high-dose and control groups were examined microscopically. The uterine contents were examined for implantations and resorptions. Viable foetuses were weighed and evaluated for external, soft tissue and skeletal malformations. At 1.25% in the diet, the mothers showed reduced weight gain (despite increased food intake) and increased relative liver weight. At this dose there was a slight (but not statistically significant) increase in the numbers of litters with external, visceral and skeletal malformations. Effects at 2% in the diet (a dose that produced clear maternal toxicity including lethargy, muscular weakness and abnormal gait) included increased resorptions, reduced foetal body weights, and a marked increase in foetal malformations (53% per litter, compared to 2% in controls); the urogenital system, eye, heart and axial skeleton were most frequently affected. The NOAEL for maternal and foetal effects was 0.5% (420 mg/kg bw/day).

 

A brief overview of other relevant studies is provided below, based on information provided in the EU RAR.

 

A number of other developmental toxicity studies in rats have demonstrated effects on the reproductive system of male offspring at doses of around 270 mg/kg bw/day (Piersma et al., 2000) or 500-1000 mg/kg bw/day (Ema et al., 2002; Gray et al., 2000; Parks et al., 1999), in some cases in the absence of maternal toxicity. In the study by Piersma et al. (2000), in which female rats were exposed to BBP on days 6-15 or 6-20 of pregnancy and necropsied on day 21, the longer exposure resulted in reduced relative testicular weight in the offspring at all dose levels (i.e. 270 mg/kg bw/day and above). Maternal toxicity was also evident at all dose levels, with increases in the relative weights of the liver and kidney. The investigators went on to estimate, using benchmark dose techniques, that a dose of 95 mg/kg bw/day was associated with a 1% increase in abnormal testis location, the most sensitive indicator of the development of the male reproductive tract.

 

In a developmental toxicity study in mice, groups of 28-30 females were given BBP in the diet at 0, 0.1, 0.5 or 1.25% (approximately 0, 182, 910 or 2330 mg/kg bw/day) on days 6-15 of pregnancy. Maternal toxicity (decreased weight gain) was observed from 910 mg/kg bw/day, together with foetal toxicity. Effects observed in both the mid- and high-dose groups included a decrease in the number of live foetuses per litter, and increases in the percent of non-live implants per litter, malformed foetuses per litter and litters with malformed foetuses. Increased resorptions were observed at the top dose. The NOAEL for maternal and developmental toxicity was 182 mg/kg bw/day (NTP, 1990).

 

The EU RAR describes a number of low-dose exposure studies in which rats were given BBP in the drinking water during gestation and early postnatal life and the reproductive performance of the offspring was evaluated (Ashby et al., 1997; Bayer, 1998; Sharpe et al., 1995; TNO, 1998a,b). The EU RAR notes that there is a problem with these studies, the instability of BBP in drinking water. In the study by Sharpe et al. (1995), when female rats received BBP at 1 mg/l in drinking water (corresponding to 0.13 to 0.37 mg/kg bw/day), for 2 weeks prior to mating and throughout pregnancy and lactation, the male offspring showed reductions in absolute testis weight and daily sperm production. However, the investigators considered that biological variance had a greater influence on the test results than did the identity of the test compound. In a similar (but not identical) study performed by Ashby et al. (1997), no effects were observed on testis weight or spermatogenesis in the offspring. Three further studies involving doses of 0.01 to 0.67 mg/kg bw/day (TNO, 1998a,b) and 0.09 to 0.80 mg/kg bw/day (Bayer, 1998) were unable to duplicate the Sharpe findings. Overall, these studies indicate that there is no impairment of reproductive development in the offspring of rats exposed to very low concentrations of BBP during gestation and lactation.

 

The EU RAR’s risk characterisation for developmental effects uses the , which was based on a dose-related significant reduction in absolute and adjusted AGD in both F1 and F2 offspring from 250 mg/kg bw/day in the absence of maternal toxicity. However, the point is made that, in the rat study by Nagao et al. (2000), there was a decrease in absolute AGD at 500 mg/kg bw/day, but no adjustment was made for the reduction in body weight (evident from 100 mg/kg bw/day); it is unclear whether such an analysis would have shown a reduction in adjusted AGD at 100 mg/kg bw/day [which would result in an NOAEL of 20 mg/kg bw/day].

In determining the various DNELs for systemic effects of long-term oral exposure of the general population, the NOAEL of 50 mg/kg bw/day from the Tyl 2-generation rat study provided the lowest DNEL (of 0.5 mg/kg bw/day).

 

Human Experience

The 2007 EU RAR on BBP describes a study by Swan et al. (2005), in which an association was reported between maternal exposure to BBP and other phthalates (based on the concentrations of nine phthalate monoesters in the mother’s urine during pregnancy) and decreased anogenital distance (AGD) in their sons. The analysis was based on 85 of 134 mother-son pairs with AGD measurements for which urine samples were available. The measurement of AGD in human neonates, particularly males, is not precise, and the investigators used a modification of the measurement, the anogenital index (AGI), as a weight-normalized index of AGD. Comparing boys of mothers with a prenatal urine concentration of monobenzyl phthalate (MBeP, reflecting exposure to BBP) in the highest compared with the lowest quartiles, the odds ratio (OR) for a shorter than expected AGI was 3.8 (p<0.05). For the other monoester phthalates the ORs were 10.2 for monobutyl phthalate (MBuP), 4.7 for monoethyl phthalate (MEP), and 9.1 for monoisobutyl phthalate (MiBP) (all p-values < 0.05). Swan et al. conclude that the finding of reduced AGI supports the hypothesis that prenatal phthalate exposure at environmental levels can adversely affect male reproductive development. However, they do not address the biological significance of these changes in human AGD or AGI. The absence of an accepted value for normal AGD or AGI in males of this age, as well as our incomplete understanding of normal variability in neonatal AGD, adds uncertainty to the conclusions offered by Swan et al. These uncertainties, as well as the small sample size and number of data points in the study, indicate that the data should be interpreted with caution. 

 

The EU RAR also describes a study by Main et al. (2005), in which a significant association was found between intake of breast milk contaminated with phthalates (MEP, MBuP, MMP, MINP) and postnatal surge of reproductive hormones (sex hormone-binding globulin, luteinizing hormone, testosterone and inhibin B) in 130 newborn boys. A similar tendency was observed with the monoester metabolite of BBP, MBeP, although the effect was not statistically significant. No association was found between the levels of these phthalate monoesters in breast milk and cryptorchidism in newborn boys.

 

In considering the studies by Swan et al. (2005) and Main et al. (2005), the EU RAR states that “further studies with larger sample size have to be performed before clear conclusions can be drawn from these studies.”

Toxicity to reproduction: other studies

Additional information

In a reliable uterotrophic assay, groups of 10 ovariectomised immature rats were dosed orally with BBP at 20, 200 or 2000 mg/kg bw/day for 4 days. Two further groups of 10 animals received sesame oil only (vehicle control) and ethynyl oestradiol at 1 mg/kg bw/day (positive control). Uterine weights were recorded at necropsy on the day after the final dose, and mean absolute wet weight and wet weight relative to body weight were calculated. The assay was performed twice (experiments 1 and 2). Groups of 10 ovariectomised mature rats were dosed similarly, and vaginal lavages performed prior to dosing, on each day of dosing and on the day after dosing. The number of positive smears and the proportion of smears with vaginal cell cornification were recorded. There was no reproducible effect of BBP on uterine weight or vaginal cell cornification, whereas ethynyl oestradiol produced a strong positive response in both assays (Zacharewski et al., 1998).

In other uterine growth tests involving daily administration to rats by gavage or subcutaneous injection for 3-4 days, BBP and its two key metabolites, monobutyl phthalate (MBuP) and monobenzyl phthalate (MBeP), gave no indication of oestrogenic activity; maximum doses were, by gavage, 2240 mg BBP/kg bw/day (Monsanto, 1996b), 1000 mg MBuP/kg bw/day (Monsanto, 1996c) and 1500 mg MBeP/kg bw/day (Monsanto, 1996d), and by subcutaneous injection, 5000 mg BBP/kg bw/day (Monsanto, 1996a). In other in vivo assays, BBP showed only weak oestrogenic activity in rats (Funabashi et al., 2001), and no activity in mice (Coldham et al., 1997; Milligan et al., 1998).

 

Weak positive results were reported in three in vitro assays for oestrogenic activity, one for competitive ligand-binding to rat uterine oestrogen receptors, one assessing gene expression in recombinant receptor/reporter gene assays in mammalian cells, and one evaluating viability in an oestrogen-dependent strain of yeast (Zacharewski et al., 1998). Other investigators have also reported weak oestrogenic activity in vitro with BBP (Coldham et al., 1997; Harris et al., 1997; Jobling et al., 1995;Sohoni and Sumpter, 1998;Soto et al., 1995 – all cited in EU RAR), whereas MBuP and MBeP showed no such in vitro activity (Harris et al., 1997).

Justification for classification or non-classification

Butyl benzyl phthalate is a reproductive and developmental toxin. Under Directive 67/548/EEC it is classified as Repr. Cat. 3; R62 (possible risk of impaired fertility) and Repr. Cat. 2; R61 (may cause harm to the unborn child). The equivalent classifications under the EU CLP regulations (Regulation (EC) No. 1272/2008) are, respectively, Repr. 2 ("suspected human reproductive toxicant" - based on animal data indicating a possible adverse effect on sexual function and fertility) and Repr. 1B ("presumed human reproductive toxicant" - based on animal data providing clear evidence of an adverse effect on development in the absence of other toxic effects).