Assessment of environmental chemicals affecting puberty gland development begins at about week 4 and continues through week 10 of gestation. The primary mammary bud is formed during this period. From weeks 10 through 15 of ges- tation, formation of secondary buds, the nip- ple, and the areola occurs. The critical window of development for the external genitalia is from weeks 7 through 12 of gestation. Because it is important to assess for exposure during this early period of gestation, it is recom- mended to recruit study participants before conception to ensure that this critical period during gestation is not missed. Once the sexual organs are formed, their activation for reproduction is determined by hormonal (gonadotropins, estrogens, andro- gens) regulation. In females this is largely deter- mined by estrogenic activity and in males, by androgens. In the prepubertal female, estrogen levels are low and stable until puberty, when a surge in estrogen level occurs, resulting from gonadotropin stimulation. The pituitary gland begins to secrete luteinizing hormone at approximately 1–3 years before the onset of puberty. The prepubertal period is an impor- tant time to assess for environmental chemical exposure because of several reported cases of girls developing premature thelarche after being exposed to estrogen compounds (Teilmann et al. 2002). Thus, the sampling would include in utero (gestation), postnatal (for breast-feeding infants), before puberty (pre- and peripuberty), and postpuberty (or the completion) periods. It is important to monitor puberty to its com- pletion because the time of onset of puberty and the duration for its completion may vary independently. Further deﬁnitions of the fre- quency for monitoring during the prepubescent period may be gained from reviewing longitudi- nal studies on puberty (Biro et al. 1995; Lee 1980; Roche et al. 1995). Populations with High Exposures Certain populations that may be susceptible or vulnerable to environmental chemical exposure will need to be considered in the study design. The reasons are variable and include genetic predisposition, nutrition, and socioeconomic factors. For example, persons frequently ingest- ing ﬁsh or whale meals are likely to have ele- vated levels of PCBs, polychlorinated dibenzo-p-dioxins (PCDDs), and polychlori- nated dibenzofurans (PCDFs). This was observed in persons residing near the Great Lakes region of the United States and members of the Inuit tribe in Canada. Children residing in homes built before 1946 were found to have approximately a 5-fold increased incidence of blood lead levels ≥ 10 µg/dL compared with children residing in homes built after 1973 (Pirkle et al. 1998). Low-income status and African-American race/ethnicity were associated with elevated blood lead levels (≥ 10 µg/dL) in families living in homes built before 1946. Environmental Chemicals Many environmental chemicals are observed to have endocrine or hormonal activity (National Research Council 1999). However, only a few of these chemicals are associated with effects on the age at puberty in humans, including lead (Selevan et al. 2003; Wu et al. 2003), PBBs (Blanck et al. 2000), PCBs (Den Hond et al. 2002; Gladen et al. 2000), PCDDs/PCDFs (Den Hond et al. 2002), p,p´-DDT/p,p´-DDE (Gladen et al. 2000; Krstevska-Konstantinova et al. 2001), endo- sulfan (Saiyed et al. 2003), phthalates (Colon et al. 2000), and phytoestrogens (Freni- Titulaer et al. 1986). To encompass all these chemicals into the assessment strategy, the chemicals can be categorized as either biologi- cally persistent or nonpersistent (Table 2). Generally, persistent chemicals have long half- lives (months to years) and are measured in the blood. Although this may seem advanta- geous from a sampling perspective (i.e., wide window of opportunity), it does pose a chal- lenge when critical windows of exposure are of concern and when subacute exposures may have occurred. Thus, if exposure occurs dur- ing critical periods of a child’s development, then specimens should be obtained proximate to those periods. Questionnaires and environ- mental assessment would be necessary if the pathways of exposure are important, because biologic monitoring would not be able to identify the exposure pathway or the exposure location. For example, the presence of a persis- tent chemical in the blood of a person who recently moved to a new location would most likely represent an exposure from the former location. However, this would not be known by the blood test alone. If documentation of exposure of a persistent chemical is all that is necessary, then biologic monitoring should be adequate. Nonpersistent chemicals have short half-lives (hours to days) and are usually meas- ured in the urine. For nonpersistent organic chemicals, increased reliance is needed on questionnaires and environmental samples to document exposure. This is because of the relatively short time that these chemicals exist in the body. Measurements Exposure assessment. The choice of the biologic matrix to measure for the chemicals is largely determined by the properties of the chemical (e.g., lipid solubility, protein-binding capacity, and ionic charge), which governs the distribu- tion of the chemical in the body. Blood and urine are the standard specimens used for the measurement of chemicals, and water-soluble chemicals are commonly quantiﬁed in urine. In addition, the target organ or organism needs to be considered. For example, if the interest is in fetal exposure, then cord blood is the pre- ferred specimen. If the route of exposure is Environmental Health Perspectives • VOLUME 113 | NUMBER 8 | August 2005 important, for example, identify either the source of exposure or the degree to which one source may be more important in determining a health outcome. If two sources are involved, then several matrices will have to be sampled. For example, an infant’s exposure can occur either during the in utero period through the placenta or during breast-feeding. However, neither the amount of chemical exposure nor the determined health effect is consistent by either of these routes of exposure. In the case of PCB exposure, it was determined that breast- feeding contributed to a larger amount of expo- sure to the infant than did placental transfer; however, the latter was deemed more conse- quential to delayed neurodevelopment (Jacobson and Jacobson 1996). If the extent to which a chemical partitions into various bio- logic matrices is known, then alternative speci- mens may be used if the desired specimen is not adequate. For example, maternal blood, cord blood, and maternal milk may be used to assess for fetal exposure to persistent organic chemi- cals. The assessment of the infant to exposure to chemicals from human milk requires multiple milk samplings during the breast-feeding period because of the expected decline in the level of persistent chemicals in human milk from maternal depuration. Additional consideration is needed to characterize the difference between infants who are exclusively breast-fed and those who are partially breast-fed because the volume of milk intake will vary by these practices of breast-feeding. Meconium is a biologic matrix that is currently under investigation as an indi- cator of chemical exposure. The interest in this matrix is because of its potential to reflect chemical exposures to the fetus as early as the second trimester. Researchers may need to customize ques- tionnaires and environmental sampling based on the use patterns of particular classes of chemicals. Phthalates, for example, are Table 3. Health effect measures to assess for puber- tal development.a Onset of puberty Sexual maturity rating (Tanner staging) Performed by observer or by self-reporting Frequency: annually Stature Hormones [luteinizing hormone, estradiol, testosterone, dehydroepiandrostendione-sulfate (DHEA-S)] Salivary hormone levels Thyroid-stimulating hormone Later stages of puberty Voice pattern change (males) Menarche Menstrual history Dual-energy X-ray absorptiometry At age 6 and 12 years for females and at age 6 and 14 years for males aDiscussed at the NCS inter-work group meeting “Obesity and Physical Development.” This meeting was co-chaired by the Nutrition, Growth, and Pubertal Development and the Exposure to Chemical Agents working groups in Baltimore, Maryland (USA), on 17–18 December 2002. 1105