Within-person changes in reproductive hormones and cognition in the menopause transition.

OBJECTIVES: Most women complain of cognitive deficits in the menopause transition, though the cause is unclear. The current study investigated the role that within-person changes in reproductive hormones, particularly estradiol, may play in triggering such perimenopausal cognitive difficulties. STUDY DESIGN: Participants were 43 women aged 45-55 years and currently in the menopause transition. Once a week for 12 weeks, participants provided a urine sample for the measurement of estrone glucuronide (E1G), a urinary metabolite of estradiol. Every three weeks across the 12-week period, participants also underwent cognitive testing, including assessments of immediate and delayed memory, attention, and executive function, and completed questionnaires assessing subjective cognitive performance. Potential confounding variables including sleep quality, vasomotor symptoms, and depressive symptoms were also assessed. RESULTS: Within-person E1G was positively associated with objective measures of attention, particularly the ability to passively register auditory information on the first pass, as well as subjective measures of memory, specifically relating to a lower frequency of forgetting things in everyday life. Perimenopausal women with lower estimated levels of intellectual functioning furthermore exhibited a stronger relationship between E1G changes and objective cognitive performance. While depressive mood, poor sleep, and vasomotor symptoms were all negatively associated with at least one aspect of cognitive function, the E1G-cognition relationship was not explained by these factors. CONCLUSIONS: This study provides evidence that validates perimenopausal women's cognitive complaints but also suggests that cognitive deficits are generally mild and transient.

Short-term serum and urinary changes in sex hormones of healthy pre-pubertal children after the consumption of commercially available whole milk powder: a randomized, two-level, controlled-intervention trial in China.

Currently, commercial milk may contain abundant pregnancy-related hormones, the regular consumption of which puts children at a risk of precocious puberty and sex-hormone-associated tumors in adulthood. In this intervention trial, 51 healthy prepubescent children were randomly assigned to the intervention or control arms at a ratio of 3 : 1 to receive 250 or 600 mL m-2 (body surface area) of milk intervention or matching equienergetic sugar water as the control. On testing cow's milk, progesterone was detected, while estrone, estradiol (E2), and testosterone (T2) were not. Cow's milk ingestion did not significantly influence the serum FSH, E2, PRL, LH, and T2 levels (P > 0.05) of pre-pubertal children 3 h after the intervention, while it increased their serum progesterone levels (P < 0.05) when compared with that in the control arm. Regarding the urinary hormone levels, cow's milk ingestion increased the urinary pregnanediol level within 4 h (P < 0.05), but not significantly when compared with that of the control (P > 0.05). The level of pregnanediol and E2 in the morning urine for three consecutive days showed no significant difference between the two arms (P > 0.05). Drinking commercial milk with progesterone influenced the progesterone levels of pre-pubertal children in hours but not days and did not affect other sex hormone levels of pre-pubertal children.

CYP3A7*1C allele: linking premenopausal oestrone and progesterone levels with risk of hormone receptor-positive breast cancers.

BACKGROUND: Epidemiological studies provide strong evidence for a role of endogenous sex hormones in the aetiology of breast cancer. The aim of this analysis was to identify genetic variants that are associated with urinary sex-hormone levels and breast cancer risk. METHODS: We carried out a genome-wide association study of urinary oestrone-3-glucuronide and pregnanediol-3-glucuronide levels in 560 premenopausal women, with additional analysis of progesterone levels in 298 premenopausal women. To test for the association with breast cancer risk, we carried out follow-up genotyping in 90,916 cases and 89,893 controls from the Breast Cancer Association Consortium. All women were of European ancestry. RESULTS: For pregnanediol-3-glucuronide, there were no genome-wide significant associations; for oestrone-3-glucuronide, we identified a single peak mapping to the CYP3A locus, annotated by rs45446698. The minor rs45446698-C allele was associated with lower oestrone-3-glucuronide (-49.2%, 95% CI -56.1% to -41.1%, P = 3.1 × 10-18); in follow-up analyses, rs45446698-C was also associated with lower progesterone (-26.7%, 95% CI -39.4% to -11.6%, P = 0.001) and reduced risk of oestrogen and progesterone receptor-positive breast cancer (OR = 0.86, 95% CI 0.82-0.91, P = 6.9 × 10-8). CONCLUSIONS: The CYP3A7*1C allele is associated with reduced risk of hormone receptor-positive breast cancer possibly mediated via an effect on the metabolism of endogenous sex hormones in premenopausal women.

Mood sensitivity to estradiol predicts depressive symptoms in the menopause transition.

BACKGROUND: The risk for depression markedly rises during the 5-6 years leading up to the cessation of menstruation, known as the menopause transition. Exposure to extreme estradiol levels may help explain this increase but few studies have examined individual sensitivity to estradiol in predicting perimenopausal depression. METHOD: The current study recruited 101 perimenopausal women. During Phase 1, we quantified each woman's sensitivity to changes in estradiol using 12 weekly measures of estrone-3-glucuronide (E1G), a urinary metabolite of estradiol, and concurrent depressive symptoms. The weekly cortisol awakening response was measured to examine the hypothalamic-pituitary-adrenal (HPA) axis in mediating mood sensitivity to estradiol. In Phase 2, depressive symptoms and major depression diagnoses were assessed monthly for 9 months. The relationship between Phase 1 E1G sensitivity and Phase 2 depressive symptoms and major depressive episodes was examined. Several baseline characteristics were examined as potential moderators of this relationship. RESULTS: The within-person correlation between weekly E1G and mood varied greatly from woman to woman, both in strength and direction. Phase 1 E1G mood sensitivity predicted the occurrence of clinically significant depressive symptoms in Phase 2 among certain subsets of women: those without a prior history of depression, reporting a low number of baseline stressful life events, and reporting fewer months since their last menstrual period. HPA axis sensitivity to estradiol fluctuation did not predict Phase 2 outcomes. CONCLUSION: Mood sensitivity to estradiol predicts risk for perimenopausal depression, particularly among women who are otherwise at low risk and among those who are early in the transition.

Expanding the applicability of magnetic ionic liquids for multiclass determination in biological matrices based on dispersive liquid-liquid microextraction and HPLC with diode array detector analysis.

Monitoring biological samples at trace levels of chemicals from anthropogenic actions such as pesticides, pharmaceuticals, and hormones has become a very important subject. This work describes a method for the determination of eight compounds of different chemical classes in human urine samples. Dispersive liquid-liquid microextraction based on magnetic ionic liquids was used as the sample preparation procedure. The main parameters of the method, such as sample dilution, type, and volume of disperser solvent, amount of magnetic ionic liquids, extraction time, and pH were optimized by univariate and multivariate procedures. Validation was performed using a urine sample of a male volunteer in order to obtain a calibration curve and the main analytical parameters of merit such as limits of detection and quantification. Values varied from 3.0 to 7.5 µg/L and from 10 to 25 µg/L, respectively. Satisfactory precisions of 21% for intraday (n = 3) and 16% for interday (n = 9) were achieved. Accuracy was evaluated by relative recovery assays using different urine samples and ranged from 75 to 130%. Robustness was assured by the Lenth method. The validated procedure was applied to five urine samples from different volunteers and the hormone estrone was found in one sample.

Hormonal Profiles of Menstrual Bleeding Patterns During the Luteal-Follicular Transition.

CONTEXT: Menstrual cycle function is determined by a complex endocrine axis that controls the ovaries and endometrium. While the late luteal phase is characterized by declining progesterone and estrogen, how these hormonal profiles relate to menstrual bleeding patterns is not well understood. OBJECTIVE: Characterize associations between luteal phase hormonal profiles and subsequent menstrual bleeding patterns, specifically spotting before bleeding. DESIGN, SETTING, AND PARTICIPANTS: We examined creatinine-adjusted urinary estrone 3-glucuronide (E13G) and pregnanediol 3-glucuronide (Pd3G) levels in relation to spotting in 116 premenopausal women (ages 20-47) who kept daily menstrual diaries and collected first morning urine samples for ≥ 2 consecutive cycles or 1 luteal-follicular transition (n = 283 transitions). We used linear mixed models to estimate associations between luteal phase hormone levels and spotting before bleeding. MAIN OUTCOME MEASURE(S) AND RESULTS: Transitions with ≥ 1 days of spotting before menstrual bleeding (n = 118) had greater luteal phase Pd3G levels vs nonspotting transitions (n = 165). Differences in Pd3G between spotting and nonspotting transitions were largest at menses onset (34.8%, 95% confidence interval, 18.9%, 52.7%). Pd3G levels for spotting transitions dropped to similar levels as nonspotting transitions an average of 1 day later, which aligned with the first day of bleeding for transitions with contiguous spotting. Spotting transitions were preceded by slower rates of Pd3G decline than nonspotting transitions, whereas E13G declines were similar. CONCLUSIONS: Self-reported bleeding patterns may provide insight into luteal phase Pd3G levels. First bleed appears to be the best choice for defining the end of the luteal phase and achieving hormonal consistency across transitions.

Trimester-specific, gender-specific, and low-dose effects associated with non-monotonic relationships of bisphenol A on estrone, 17ß-estradiol and estriol.

BACKGROUND: Bisphenol A (BPA) may cause some adverse effects on human health by mimicking estrogen activities. In vitro andanimalstudies have observed the non-monotonic associations between BPA and natural estrogens, but the evidence in human study is lacking, particularly at multiple points in time during pregnancy. OBJECTIVE: We aimed to examine the relationships between BPA and estrogens in the three trimesters among Chinese pregnant women and their gender variations. METHODS: This study included 851 participants from a birth cohort conducted in Wuhan, China between 2014 and 2015. We measured concentrations of BPA and three estrogens (estrone (E1), 17ß-estradiol (E2) and estriol (E3)) in urine samples collected in the three trimesters of pregnancy (mean for each visit: 13.0, 23.6, and 35.9 weeks' gestation). We calculated the estimated daily intakes using urinary BPA concentrations and compared them with the tolerable intake value to assess potential health risks. We used multivariate linear regression models stratified by trimester and gender to explore trimester-specific and gender-specific associations of BPA with E1, E2, and E3. RESULTS: We found the decreased levels of estrogens (ß < 0, P < 0.05) in the upper BPA quartiles over three trimesters, except for the elevated levels of E3 (ß = 0.20, 95% CI: 0.02, 0.38) in the highest BPA quartile in the 2nd trimester. There were significant non-linear associations (overall associations P < 0.05, non-linear associations P < 0.05) between BPA and E3 in the three trimesters. In the gender-stratified analysis, we observed significant negative relationships (ß < 0, P < 0.05) between BPA and E2 among mothers carrying male fetuses in the 1st trimester and significant associations between BPA and E3 among mothers carrying female fetuses in the 2nd trimester. However, we found no significant relationship between BPA and E2 among mothers carrying female fetuses over three trimesters. CONCLUSIONS: Our findings support experimental evidence of non-monotonic relationships between BPA and three major estrogens, even at low doses of BPA. Mothers delivering male fetuses may be more sensitive to E2 at early pregnancy, and those delivering female fetuses may be more susceptive to E3 at mid-pregnancy.

Variation of urinary follicle-stimulating hormone level after menopause†:†From the results of Japan Nurses' Health Study.

The change in follicle-stimulating hormone (FSH) during the menopausal transition and associations of FSH with various diseases have been assessed by using blood samples. We examined cross-sectionally the variation of FSH levels, associations of estrone and estradiol with FSH, and associations of BMI with these hormones by using urinary samples from peri- and postmenopausal women in Japan. Of 4472 participants in the Urinary Isoflavone Concentration Survey of the Japan Nurses' Health Study, we analyzed urinary levels of estrone, estradiol and FSH in 547 women aged from 45 to 54 years. Urinary FSH levels varied widely in postmenopausal women and the pattern of change in urinary FSH levels seems to be similar to that in blood FSH levels in previous studies. There were no significant differences in age, body mass index (BMI), estradiol, estrone and estradiol/estrone ratio among three groups according to the tertile of FSH. In postmenopausal women, there were significant associations of BMI with levels of estrone and estradiol, but there was no significant association of BMI with FSH. Studies using urinary samples will allow us to establish a study project as a large-scale population-based study to determine associations between FSH and various diseases after menopause. J. Med. Invest. 66 : 297-302, August, 2019.

Association between FSH, E1, and E2 levels in urine and serum in premenopausal and postmenopausal women.

OBJECTIVE: We aimed to establish correlations for the levels of follicle-stimulating hormone (FSH), estrone (E1) and estradiol (E2) between urine and serum in premenopausal and postmenopausal women using immunoassays. METHODS: In this study of 92 women (61 postmenopausal, 31 premenopausal), both urine and blood specimens were collected on the same day and stored at 4 °C for analysis by chemiluminescent immunoassay, radioimmunoassay and/or electrochemiluminescent immunoassay. RESULTS: There were correlations in the levels of FSH, E1 and E2 between urine and serum in both postmenopausal (r = 0.96 for FSH, r = 0.91 for E1, r = 0.80 for E2) and premenopausal (r = 0.98 for FSH, r = 0.92 for E1, r = 0.90 for E2) women. It is indicated that the correlations were stronger in the premenopausal group compared with the postmenopausal group, especially for FSH. CONCLUSION: The levels of FSH, E1 and E2 in urine correlated with those in the serum in premenopausal and postmenopausal women. Urine samples could be used instead of serum samples to measure hormone levels, which would reduce the difficulty of conducting large survey studies.

Home ovulation test use and stress during subfertility evaluation: Subarm of a randomized controlled trial.

OBJECTIVES: A prospective, randomized controlled trial in women seeking to conceive examined the impact of using ovulation tests on self-reported levels of stress, psychological well-being, and quality of life in women with unexplained infertility. METHOD: The test group used a home ovulation test to detect the day of ovulation, whereas the control group were provided with a predicted day of ovulation based on the average length of menstrual cycle reported during study recruitment. Volunteers collected their first morning urine samples to evaluate biochemical levels of stress (urinary cortisol and estrone-3-glucouronide) and completed questionnaires over two complete menstrual cycles. RESULTS: Overall, the use of digital ovulation tests by sub-fertile women under medical care had negligible negative effects and no detectable positive benefit on psychological well-being, according to multiple measurements of stress by questionnaire and biochemical markers. No significant differences were found between groups for all stress measures at the various study time points, except in relation to "couple concordance" where the test group scored much higher than the control group (mean difference at end of study was 21.25 (95% confidence interval: 9.25, 33.25; P = 0.0015)). The maximum difference in log cortisol: creatinine ratio between the test and control groups was -0.28 (95% confidence interval: -0.69, 0.13). CONCLUSIONS: These results do not support propositions that using digital ovulation tests can cause stress in women trying to conceive.

The association of urinary estrogen levels with urinary isoflavone levels: Difference between premenopausal women and postmenopausal women.

Results of studies on the associations of soy food intake with urinary estrogen levels in premenopausal women and in postmenopausal women have been inconsistent. We examined the associations of urinary isoflavone levels as well as soy food intake with estrone (E1) and estradiol (E2) in pre- and postmenopausal women. In addition, we compared the levels of isoflavones, E1 and E2 across current hormone users such as those receiving hormone replacement therapy and those using oral contraceptives and non-users among both pre- and postmenopausal women. Urinary levels of isoflavones, E1 and E2 in 498 women (36 hormone users and 462 non-users) were analyzed. Premenopausal women with a higher frequency of soy food intake had higher urinary isoflavone levels, but there were no significant associations between E1 and E2 levels and urinary isoflavone levels. Levels of E1 and E2 in hormone users were significantly lower than those in hormone non-users among premenopausal women, but levels of E1 and E2 in hormone users were significantly higher than those in hormone non-users among postmenopausal women. Postmenopausal women with a higher frequency of soy food intake had higher urinary isoflavone levels, and postmenopausal women with high urinary isoflavone levels had significantly higher E1 and E2 levels. In conclusion, the associations of urinary isoflavone levels with urinary estrogen levels differed with menopausal status. Urinary levels of E1 and E2 were high in postmenopausal women with high urinary isoflavone levels but not in premenopausal women with high urinary isoflavone levels.

Urinary excretion of sex steroid hormone metabolites after consumption of cow milk: a randomized crossover intervention trial.

Background: Current cow milk production practices introduce considerable levels of pregnancy hormones into the milk. Humans are exposed to these hormones when cow milk is consumed, and this may explain the observed association between cow milk consumption and several hormone-sensitive cancers. Objectives: The aim of the study was to evaluate whether cow milk consumption is associated with an increase in urinary excretion of sex steroid hormones and their metabolites in humans. Methods: We conducted a randomized crossover intervention feeding experiment. A total of 109 postmenopausal women consumed 1 L of semiskimmed milk (1.5% fat) per day for 4 d and 1 L of whole milk (3.5% fat) per day for 4 d, intersected by 4-d wash-out periods. Sex steroid hormone levels were measured in 24-h urine samples collected at the end of each intervention and wash-out period. Results: Estrogens, androgens, and progesterone were detected in the examined milk samples used for our intervention. Although a very high proportion of the estrogens were conjugated, only small proportions of the androgens and progesterone were conjugated. Milk consumption resulted in a significant increase in urinary estrone (E1) excretion, whereas estradiol (E2), estriol (E3), and 16ketoE2 excretion only increased after semiskimmed milk consumption. Urinary pregnanediol glucuronide excretion was not significantly affected. Conclusion: Cow milk consumption increases urinary excretion of E1 in humans. Ingestion of semiskimmed milk appears also to raise E2, E3, and 16ketoE2 excretion, but future studies need to confirm these associations. This trial was registered at https://www.drks.de as DRKS00003377.

Salivary and urinary metabolome analysis for pre-puberty-related biomarkers identification in porcine.

Estrus synchronization is important for optimal management of gilt reproduction in pig farms. Hormonal treatments, such as synthetic progestogens, are used on a routine basis, but there is a growing demand for non-hormonal alternative breeding tools. Before puberty, gilts exhibit a 'waiting period,' related to the ovarian development and gonadotrophin secretions, during which external stimulations, such as boar exposure, could induce and synchronize first ovulation. Practical non-invasive tools for identification of this period in farms are lacking. During this period, urinary oestrone levels are high, but urine sampling is difficult in group-housed females. The aim of this work was to search for specific biomarkers of the 'waiting period' in saliva and urine. In total, nine 144- to 147-day-old Large White gilts were subjected to trans-abdominal ultrasonography three times a week for 5 weeks until puberty detection (week -5 to week -1 before puberty). Urine and saliva samples were collected for oestrone assay to detect the 'waiting period' and for metabolome analysis using 1H-nuclear magnetic resonance spectroscopy to detect potential biomarkers of the 'waiting period.' Gilts were slaughtered 7 days after puberty detection for puberty confirmation. Results were consistent with ultrasonography data for six gilts. Urine and saliva samples from these six gilts were analyzed. Urinary estrone concentration significantly increased 2 weeks before puberty detection. Metabolome analysis of urine samples allowed the identification of 78 spectral bins, among them, 42 low-molecular-weight metabolites were identified. Metabolome analysis of salivary samples allowed the identification of 59 spectral bins, among them, 23 low-molecular-weight metabolites were detected and 17 were identified. No potential biomarker was identified in urinary samples. In saliva, butyrate and 2HOvalerate, 5.79 ppm (putatively uridine), formate, malonate and propionate could be biomarker candidates to ascertain the pre-puberty period in gilt reproduction. These results confirm that non-invasive salivary samples could allow the identification of the physiological status of the gilts and presumably the optimal time for application of the boar effect. This could contribute to synchronize puberty onset and hence to develop non-hormonal breeding tools.

Impaired vascular function in exercising anovulatory premenopausal women is associated with low bone mineral density.

In estrogen-deficient post-menopausal women, osteoporosis shares a common link with cardiovascular disease risk, including endothelial dysfunction. The current study sought to examine associations between bone mineral density (BMD) and endothelial function in estrogen-deficient premenopausal women with exercise-associated menstrual disturbances. Recreationally trained women (24.3 ± 0.8 years; overall mean ± SEM) who were estrogen deficient (amenorrheic or eumenorrheic anovulatory cycles; E2Def; n = 13) or estrogen replete (eumenorrheic ovulatory cycles; E2Rep; n = 14) were studied. Total body and lumbar BMD (L1-L4) were determined using dual-energy X-ray absorptiometry. Serum markers of oxidative stress (oxidized low-density lipoprotein; OxLDL), energy deficiency (triiodothyronine), and bone turnover (osteocalcin, c-telopeptide X, P1NP) were assessed. Estrogen exposure was determined by assessing daily urinary estrone-3-glucuronide (E1G) across a monitoring period. Calf blood flow (CBF), an index of endothelial function, was measured using strain-gauge plethysmography. CBF, total body and L1-L4 BMD, triiodothyronine and E1G were lower (P < 0.05), and c-telopeptide crosslinks higher (P < 0.05) in E2Def. Osteocalcin and OxLDL did not differ (P > 0.05) between groups. L1-L4 BMD, osteocalcin, and E1G were the strongest predictors of CBF (R2 =0.615, P < 0.001). CBF was the strongest predictor of L1-L4 BMD (R2 =0.478, P < 0.001). L1-L4 (r = 0.558, P = 0.008) and CBF (r = 0.534, P = 0.004) were independently correlated with E1G. In young recreationally trained premenopausal women with anovulatory menstrual disturbances, low CBF predicts decreased lumbar BMD, suggesting impaired peripheral endothelial function may predict early unfavorable changes in bone metabolism. This finding may be of relevance in the early detection of cardiovascular and bone health decrements in otherwise healthy estrogen-deficient premenopausal women.

Determination of Five Sex Hormones in Urine Samples for Early Evaluation of Male Reproductive Toxicity Induced by Phthalate Esters in Rats.

In this work, male rats were exposed to multiple phthalate esters (MIXPs) in a long-term low-dose model for the early evaluation of reproductive toxicity. An ananlysis method with better sensitivity, accuracy and precision was established to determine the five sex hormones (androstenedione, testosterone, dehydroepiandrosterone, dihydrotestosterone, and estrone) in collected urine samples. The results showed that all the analytes in the MIXPs treated group changed in a time-dependent manner. Specifically, estrone significantly decreased from the 30th day and the other four changed from the 30th day and then significantly increased on the 60th day, while no obvious changes were found in the control group. Therefore, a possible way was provided for the early evaluation of male reproductive toxicity induced by Phthalate esters (PEs) . The reliability of judgment was improved by observing the changes of five target hormones simultaneously. Furthermore, good compliance was predicted for the practical application due to the noninvasive and convenient urine sample collection.

Urine and Serum Sex Steroid Profile in Testosterone-Treated Transgender and Hypogonadal and Healthy Control Men.

Background: The impact of testosterone (T) treatment on antidoping detection tests in female-to-male (F2M) transgender men is unknown. We investigated urine and serum sex steroid and luteinizing hormone (LH) profiles in T-treated F2M men to determine whether and, if so, how they differed from hypogonadal and healthy control men. Method: Healthy transgender (n = 23) and hypogonadal (n = 24) men aged 18 to 50 years treated with 1000 mg injectable T undecanoate provided trough urine and blood samples and an additional earlier postinjection sample (n = 21). Healthy control men (n = 20) provided a single blood and urine sample. Steroids were measured by mass spectrometry-based methods in urine and serum, LH by immunoassay, and uridine 5'-diphospho-glucuronosyltransferase 2B17 genotype by polymerase chain reaction. Results: Urine LH, human chorionic gonadotropin, T, epitestosterone (EpiT), androsterone (A), etiocholanolone (Etio), A/Etio ratio, dehydroepiandrosterone (DHEA), dihydrotestosterone (DHT), and 5α,3α- and 5ß,3α-androstanediols did not differ between groups or by time since last T injection. Urine T/EpiT ratio was <4 in all controls and 12/68 (18%) samples from T-treated men, but there was no difference between T-treated groups. Serum estradiol, estrone, and DHEA were higher in transgender men, and serum T and DHT were higher in earlier compared with trough blood samples, but serum LH, follicle-stimulating hormone, and 3α- and 3ß,5α-diols did not differ between groups. Conclusion: Urine antidoping detection tests in T-treated transgender men can be interpreted like those of T-treated hypogonadal men and are unaffected by time since last T dose. Serum steroids are more sensitive to detect exogenous T administration early but not later after the last T dose.

Aged Black-and-Gold Howler Monkey Female (Alouatta caraya): A Sign of Reproductive Senescence?

Reproductive senescence patterns have been scarcely studied in Neotropical primates. The few studies available on the hormonal profiles of aging female monkeys indicate that the decline of ovarian function in nonhuman primates may resemble the hormonal events associated with the perimenopause in women. In this study, we explore a reproductive hormone profile of an aged black-and-gold howler monkey female (Alouatta caraya) from a wild population in northeastern Argentina and compare this profile with that of a cycling female in the same population. As part of a larger study, we recorded sociosexual behaviors in adult and subadult females belonging to two groups, and we collected urine (n = 877) to determine the sex hormone profile of each female. These samples were analyzed using enzyme immunoassays for estrone conjugates and pregnanediol-3-glucuronide (PdG). We found differences in mean values of PdG between the younger (cycling) and the older female. These hormone values were lower in the older female, and she did not show any signs of cyclicity for either reproductive hormone. Our results show that the aging female in this wild population shows signs of ovarian senescence, indicated by low, acyclic levels of progesterone metabolites.

Magnetic ionic liquids as versatile extraction phases for the rapid determination of estrogens in human urine by dispersive liquid-liquid microextraction coupled with high-performance liquid chromatography-diode array detection.

In this study, a rapid and straightforward approach based on magnetic ionic liquids (MIL) as extraction phases and dispersive liquid-liquid microextraction (DLLME) was developed to analyze the hormones estriol, 17-ß-estradiol, 17-α-ethynylestradiol, and estrone in human urine samples. This is the first report of an application of manganese-based MILs compatible with HPLC to extract compounds of biological interest from urine samples. The hydrophobic MILs trihexyltetradecylphosphonium tetrachloromanganate (II) ([P6,6,6,14+]2[MnCl42-]) and aliquat tetrachloromanganate (II) ([Aliquat+]2[MnCl42-]) were employed and the optimized extraction conditions were comprised of 5 mg of MIL ([P6,6,6,14+]2[MnCl42-]), 5 µL of methanol (MeOH) as disperser solvent, and an extraction time of 90 s at sample pH 6. The analytical parameters of merit were determined under optimized conditions and very satisfactory results were achieved, with LODs of 2 ng mL-1 for all analytes, determination coefficients (R2) ranging from 0.9949 for 17-ß-estradiol to 0.9998 for estrone. In addition, good results of method precision were achieved with the intraday (n = 3) varying from 4.7% for 17-ß-estradiol to 19.5% for estriol (both at 5 ng mL-1) and interday precision (evaluated at 100 ng mL-1) ranging from 11.4% for estrone to 17.7% for 17-α-ethynylestradiol and analyte relative recovery evaluated in three real samples ranged from 67.5 to 115.6%. The proposed DLLME/MIL-based approach allowed for a reliable, environmentally friendly and high-throughput methodology with no need for a centrifugation step. Graphical abstract An overview of the rapid and straightforward extraction procedure using DLLME/MIL-based approach.

Menstrual Disruption with Exercise Is Not Linked to an Energy Availability Threshold.

INTRODUCTION: Chronic reductions in energy availability (EA) suppress reproductive function. A particular calculation of EA quantifies the dietary energy remaining after exercise for all physiological functions. Reductions in luteinizing hormone pulse frequency have been demonstrated when EA using this calculation is <30 kcal·kg·fat-free mass (ffm)·d. PURPOSE: We determined whether menstrual disturbances (MD) are induced when EA is <30 kcal·kg ffm·d. METHODS: Thirty-five sedentary, ovulatory women age 18 to 24 yr (weight, 59.0 ± 0.8 kg; body mass index, 21.8 ± 0.4 kg·m) completed a diet and exercise intervention over three menstrual cycles. Participants were randomized to groups that varied in the magnitude of negative energy balance created by the combination of exercise and energy restriction. Menstrual disturbances were determined using daily urinary estrone-1-glucuronide and pregnanediol glucuronide, midcycle luteinizing hormone, and menstrual calendars. In a secondary analysis, we calculated EA from energy balance data and tested the association of EA with MD. RESULTS: A generalized linear mixed-effects model showed that the likelihood of a MD decreased by 9% for each unit increase in EA (odds ratio, 0.91; 95% confidence interval, 0.84-0.98; P = 0.010). No specific value of EA emerged as a threshold below which MD were induced. When participants were partitioned into EA tertile groups (low EA, 23.4-34.1; n = 11; moderate EA, 34.9-40.7; n = 12, and high EA, 41.2-50.1; n = 12 [kcal·kg ffm·d]), estrone-1-glucuronide (P < 0.001), pregnanediol glucuronide (P < 0.001), and luteal phase length (P = 0.031) decreased significantly, independent of tertile. CONCLUSIONS: These findings do not support that a threshold of EA exists below which MD are induced but do suggest that MD increase linearly as EA decreases. Menstrual disturbances can likely be prevented by monitoring EA using a simplified assessment of metabolic status.

A pilot study on the utility of reduced urine collection frequency protocols for the assessment of reproductive hormones in adolescent girls.

BACKGROUND: The objectives of this study were to assess the feasibility of and compliance to collecting urine samples in pre- and postmenarcheal girls and to determine if a less than daily collection frequency was sufficient for assessing ovarian function. METHODS: Twenty-five postmenarcheal girls (11-17 years) collected samples using either a two or a three samples/week protocol during one menstrual cycle. Exposure and mean estrone-1-glucuronide (E1G) and pregnanediol glucuronide concentrations were calculated, and evidence of luteal activity (ELA) was evaluated. Sixteen premenarcheal girls (8-11 years) collected one sample/month for six consecutive months. Samples were analyzed for E1G concentration. Participant compliance was calculated using dates on the urine samples and paper calendars. RESULTS: Participants collecting three samples/week were more compliant to the protocol than those collecting two samples/week (83.6%±2.6% vs. 66.8%±6.6%; p=0.034). There were no differences (p>0.10) regarding paper calendar return (81.8%±12.2% vs. 92.9%±7.1%), recording menses (55.6%±17.6% vs. 92.3%±7.7%) or sample collection (88.9%±11.1% vs. 84.6%±10.4%) between the two protocols. The average cycle length was 30.5±1.3 days and 32% of cycles had ELA. The premenarcheal girls were 100% compliant to the protocol. Only 68.8% of participants returned the paper calendar and 81.8% of those participants recorded sample collection. The average E1G concentration was 15.9±3.8 ng/mL. CONCLUSIONS: Use of a less than daily collection frequency during one menstrual cycle in postmenarcheal, adolescent girls is feasible and provides informative data about ovarian function. Collection of one sample/month in premenarcheal girls is feasible and detects the expected low E1G concentrations. Alternate strategies to the use of a paper calendar should be considered.