Essential metals, vitamins and antioxidant enzyme activities in COVID-19 patients and their potential associations with the disease severity.

The role of micronutrient deficiency in the pathogenesis of COVID-19 has been reviewed in the literature; however, the data are limited and conflicting. This study investigated the association between the status of essential metals, vitamins, and antioxidant enzyme activities in COVID-19 patients and disease severity. We recruited 155 patients, who were grouped into four classes based on the Adults guideline for the Management of Coronavirus Disease 2019 at King Faisal Specialist & Research Centre (KFSH&RC): asymptomatic (N = 16), mild (N = 49), moderate (N = 68), and severe (N = 22). We measured serum levels of copper (Cu), zinc (Zn), selenium (Se), vitamin D3, vitamin A, vitamin E, total antioxidant capacity, and superoxide dismutase (SOD). Among the patients, 30%, 25%, 37%, and 68% were deficient in Se (< 70.08 µg/L), Zn (< 0.693 µg/mL), vitamin A (< 0.343 µg/mL), and vitamin D3 (< 20.05 µg/L), respectively, and SOD activity was low. Among the patients, 28% had elevated Cu levels (> 1.401 µg/mL, KFSH&RC upper reference limit). Multiple regression analysis revealed an 18% decrease in Se levels in patients with severe symptoms, which increased to 30% after adjusting the model for inflammatory markers. Regardless of inflammation, Se was independently associated with COVID-19 severity. In contrast, a 50% increase in Cu levels was associated with disease severity only after adjusting for C-reactive protein, reflecting its possible inflammatory and pro-oxidant role in COVID-19 pathogenesis. We noted an imbalance in the ratio between Cu and Zn, with ~ 83% of patients having a Cu/Zn ratio > 1, which is an indicator of inflammation. Cu-to-Zn ratio increased to 45% in patients with mild symptoms and 34%-36% in patients with moderate symptoms compared to asymptomatic patients. These relationships were only obtained when one of the laboratory parameters (lymphocyte or monocyte) or inflammatory markers (neutrophil-to-lymphocyte ratio) was included in the regression model. These findings suggest that Cu/Zn might further exacerbate inflammation in COVID-19 patients and might be synergistically associated with disease severity. A 23% decrease in vitamin A was seen in patients with severe symptoms, which disappeared after adjusting for inflammatory markers. This finding may highlight the potential role of inflammation in mediating the relationship between COVID-19 severity and vitamin A levels. Despite our patients' low status of Zn, vitamin D3, and antioxidant enzyme (SOD), there is no evidence of their role in COVID-19 progression. Our findings reinforce that deficiency or excess of certain micronutrients plays a role in the pathogenesis of COVID-19. More studies are required to support our results.

Potential health risks of maternal phthalate exposure during the first trimester - The Saudi Early Autism and Environment Study (SEAES).

Phthalates are the most ubiquitous contaminants that we are exposed to daily due to their wide use as plasticizers in various consumer products. A few studies have suggested that in utero exposure to phthalates can disturb fetal growth and development in humans, because phthalates can interfere with endocrine function. We collected spot urine samples from 291 pregnant women in their first trimester (9.8 ± 2.3 gestational weeks) recruited in an ongoing prospective cohort study in Saudi Arabia. A second urine sample was collected within 1-7 d after enrollment. The aims of this study were to: (1) assess the extent of exposure to phthalates during the first trimester and (2) estimate the risk from single and cumulative exposures to phthalates. Most phthalate metabolites' urinary levels were high, several-fold higher than those reported in relevant studies from other countries. The highest median levels of monoethyl phthalate, mono-n-butyl phthalate (MnBP), mono-iso-butyl phthalate (MiBP), and mono-(2-ethylhexyl) phthalate (MEHP) in µg/l (µg/g creatinine) were 245.62 (197.23), 114.26 (99.45), 39.59 (34.02), and 23.51 (19.92), respectively. The MEHP levels were highest among three di (2-ethylhexyl) phthalate (DEHP) metabolites. %MEHP4, the ratio of MEHP to four di (2-ethylhexyl) phthalate metabolites (∑4DEHP), was 44%, indicating interindividual differences in metabolism and excretion. The hazard quotient (HQ) of individual phthalates estimated based on the reference dose (RfD) of the U.S. Environmental Protection Agency indicated that 58% (volume-based) and 37% (creatinine-based) of the women were at risk of exposure to ∑4DEHP (HQ > 1). Based on the tolerable daily intake (TDI) from the European Food Safety Authority, 35/12% (volume-/creatinine-based data) of the women were at risk of exposure to two dibutyl phthalate (∑DBP) metabolites (MiBP and MnBP). The cumulative risk was assessed using the hazard index (HI), the sum of HQs of all phthalates. The percentages of women (volume-/creatinine-based data) at health risks with an HI > 1 were 64/40% and 42/22% based on RfD and TDI, respectively. In view of these indices for assessing risk, our results for the anti-androgenic effects of exposing pregnant women to ∑4DEHP and ∑DBP early during pregnancy are alarming.

Selenium status in lactating mothers-infants and its potential protective role against the neurotoxicity of methylmercury, lead, manganese, and DDT.

A total of 206 lactating mothers and their infants (3-12 months) were included in this study to evaluate postnatal exposure to neurotoxic pollutants such as methylmercury (MeHg), lead (Pb), manganese (Mn), dichlorodiphenyltrichloroethane (DDT) and its metabolites [dichlorodiphenyldichloroethane (DDD), and dichlorodiphenyldichloroethylene (DDE)] and their association with delayed neurological development and to explore the protective role of selenium (Se) against chemical neurotoxicity. Neurodevelopmental performance was evaluated using Denver Developmental Screening Test II and Parents' Evaluation of Developmental Status (PEDS). Multivariate log-binomial regression modeling was applied for both single and multiple exposures to chemicals using a principal component analysis that generated six principal components. Both mothers and their infants had been exposed to metals and DDT metabolites, with some exceeding the accepted permissible limits. The geometric means of MeHg, Pb, Mn, DDD, DDE and DDT levels in breast milk were 1.333, 45.327, 15.576, 0.069, 0.542 and 1.08 µg/l, respectively. A single-exposure model identified a high risk of reduced PEDS performance significantly associated with DDD in breast milk [relative risk (RR) = 1.484; 95% confidence interval (95%CI) = 1.091-2.019] and marginally significantly associated with Pb in the mothers' blood (RR = 2.164; 95%CI = 0.87-5.382). We did not find a protective role of Se in neurodevelopment due to its high levels in the mothers. Models of multi-chemical exposure indicated that Mn in blood and breast milk, Se in blood and Pb in the mothers' urine were marginally significantly associated with a high risk of reduced PEDS performance (RR = 0.424; 95%CI = 0.176-1.022). The use of multi-chemical exposure approach in early life risk assessments is important because it indicates real-world exposure. Our results were not conclusive because the sample size was small, so future studies examining the implications to health of the impact of prenatal/postnatal exposure to a mixture of chemicals in the Saudi population are merited.

Exposure to phthalates in couples undergoing in vitro fertilization treatment and its association with oxidative stress and DNA damage.

This prospective study of 599 couples seeking fertility treatment and who were recruited between 2015 and 2017 was conducted to (a) explore the associations between phthalate exposure and in vitro fertilization (IVF) outcomes; and (b) examine the implication of oxidative stress as a mediator of these. We measured eight phthalate metabolites in two spot urine samples; oxidative stress biomarkers such as malondialdehyde, 8-hydroxy-2-deoxyguanosine, hydrogen peroxide, catalase (CAT), and total antioxidant capacity in follicular fluid and seminal plasma. We also examined DNA damage in sperm and granulosa cells. Couples were exposed to a broad range of phthalate compounds and seven metabolites were detected in over 94% of the urine samples, whereas monobenzyl phthalate was found in only 24% of women and 26% of men. Our results showed high levels of seven urinary phthalate metabolites (except monobenzyl phthalate) and a notable increase in many oxidative stress markers in both follicular fluid and seminal plasma. However, their associations with exposure were rather limited. Multivariate binomial regression modeling showed higher levels of follicular CAT levels reduced the probability of fertilization rate (≤ 50%) [Adjusted relative risk (RRadj) = 0.52, p = 0.005] and unsuccessful live birth (RRadj = 0.592, p = 0.023). We observed a 46% decrease in the probability of clinical pregnancy in association with an elevated percentage of DNA in the tail (RRadj = 0.536, p = 0.04). There was a 32% and 22% increase in the probability of clinical pregnancy and unsuccessful live birth associated with higher levels of mono-(2-ethylhexyl) phthalate (RRadj = 1.32, p = 0.049) and monoethyl phthalate (RRadj = 1.22, p = 0.032) in women, respectively. In contrast, the probability of clinical pregnancy reduced by 20% with higher levels of mono-(2-ethyl-5-carboxypentyl) phthalate (RRadj = 0.797, p = 0.037) and 19.6% with mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) (RRadj = 0.804, p = 0.041) in men. Other oxidative stress biomarkers or urinary phthalate metabolites showed suggestive relationships with certain IVF outcomes. Lastly, our results demonstrated that elevated levels of CAT in follicular fluid might have a positive impact on fertilization rate ≥ 50% and successful live birth. CAT seems to play a potential role in mediating the relationship between the risk of poor fertilization rate and MEOHP and mono-isobutyl phthalate. Additional data are required to understand the clinical implications of oxidative stress and its contribution to the reproductive toxicity of phthalate exposure.

The extent and predictors of phthalate exposure among couples undergoing in vitro fertilization treatment.

Phthalates are chemicals used as plasticizers and solvents in many consumer products but are suspected of disrupting the endocrine system and are known for their reproductive/developmental health risks. This study examined the extent and predictors of phthalate exposure among 599 couples undergoing in vitro fertilization. A questionnaire was administered to obtain sociodemographic, health, and lifestyle data, and two spot urine samples were collected from the couples to analyze eight phthalate metabolites, cotinine (COT) as a smoking index, and creatinine to adjust for urine dilution. Seven phthalate metabolites were detected in > 94% of the urine samples, and monobenzyl phthalate (MBzP) was found in 24% of the women and 26% of their male partners. Median phthalate levels were highest for monoethyl phthalate (MEP), at 333.26 µg/l in women and 290 µg/l in male partners, and lowest for MBzP, at 1.17 µg/l in women and 1.14 µg/l in male partners. Correlation coefficients of ≥ 0.4 between the women and their male partners for the eight urinary phthalate metabolites may indicate a shared source of exposure. A multivariate regression model was used to assess the association between predictors and each urinary phthalate metabolite. Several potential predictors for the variations in specific urinary phthalate metabolites were identified, including the body mass index, age, socioeconomic status, and regional distribution for both women and their male partners but with slightly different patterns. Women with a history of breastfeeding, using bottled water for cooking and storing food in plastic bags had lower MEP (8.7%), mono-(2-ethyl-5-carboxypentyl) phthalate (MECPP) (9.2%), and both mono-iso-butyl phthalate and MECPP (8.2 and 8.1%). A history of contraceptive use was associated with an increase in MECPP (8.7%), mono-(2-ethyl-5-hydroxyhexyl) phthalate (11.4%), mono-(2-ethyl-5-oxohexyl) phthalate (7.6%), and the molar sum of bis (2-ethylhexyl) phthalate metabolites (8.9%). Urinary COT levels were associated with an increase of 10-16% in all urinary metabolites in women but of only 10.5% in mono-(2-ethylhexyl) phthalate in male partners. More than 95% of the couples reported the use of cosmetics, perfumes, and personal-care products, but we were not able to find associations with urinary phthalate metabolites, perhaps due to their short half-lives. MEP levels associated with the use of household cleaning products were 11.2% higher in male partners. Our levels were generally higher than those reported elsewhere, perhaps due to different lifestyles, cultural practices, dietary habits, use of personal-care products, and governmental legislation.

The extent of mercury (Hg) exposure among Saudi mothers and their respective infants.

A total of 1016 healthy Saudi mothers and their respective infants (aged 3-12 months) were recruited from 57 Primary Health Care Centers (PHCCs) in Riyadh, Saudi Arabia, to evaluate the extent of mercury (Hg) exposure and predict its sources in the healthy Saudi population. Total Hg levels were measured in maternal urine, breast milk, blood, and hair and in the infants' urine and hair. Only 1.9% of the mothers had urinary Hg (UHg)>10 µg/l, the limit for asymptomatic adults recommended by the World Health Organization, but the median (0.99 µg/l) was higher than in other countries. Also, 49.3% of the mothers had UHg>1 µg/l, the German reference value for adults. Median infant UHg was 0.729 µg/l, and 77 and 93 % of the infants had levels higher than 0.4 and 0.1 µg/l, the reference values of the Centers for Disease Control and Prevention and for Germany, respectively. The median Hg level in breast milk was 0.884 µg/l. Even though 43.2% of the milk samples were above the background level for Hg in human milk (1 µg/l), our results were lower than those reported from other countries. Median maternal total Hg in blood was 0.637 µg/l, and only 0.4 and 6.9% of samples were higher than the Hg reference levels of 5.8 µg/l of the Environmental Protection Agency (EPA) and of 2 µg/l for Germany, respectively. Total Hg levels in hair (HHg) varied widely among mothers and infants, but only 3.9% of the mothers and 2.8% of the infants had HHg>1 µg/g (the EPA reference level). Median HHg values were 0.117 µg/g dry weight in mothers and 0.1 µg/g dry weight in infants; both were lower than in other countries. The Hg levels in mothers and their respective infants were relatively low, but our results were consistent with other studies indicating that dental amalgam fillings and fish consumption were the main predictors of maternal Hg exposure. Among the several biomarkers of Hg exposure, Hg levels in maternal hair and urine were the strongest predictors of infant exposure. The lack of an association between Hg in breast milk and Hg in infant urine and hair suggested that the infants were exposed to Hg predominately during pregnancy rather than during breastfeeding. We expect that our data can serve as a baseline for further biomonitoring and follow-up studies, particularly of the long-term impact of Hg on childhood neurodevelopment.

The cumulative risk assessment of phthalates exposure in preterm neonates.

Phthalates are widely used plasticizers in various consumer products and medical devices, with some reporting as having estrogenic and anti-androgenic endocrine-disrupting effects. Premature neonates may be exposed to high levels of specific phthalates during hospitalization in the neonatal intensive care unit (NICU) because of reliance on multiple medical procedures that pose a possible health risk. The present study utilized seven urinary phthalate metabolites of dibutyl phthalate isomers [(di-n-butyl phthalate (DnBP) and diisobutyl phthalate (DiBP)], butylbenzyl phthalate (BBzP), and di(2-ethylhexyl) phthalate (DEHP) that had been previously measured in 33 preterm neonates sampled at hospital admission (N = 23) and daily during their NICU stay (N = 260). We aimed to perform: (1) cumulative risk assessment (CRA) using the volume and creatinine-adjusted models; (2) examine the temporal variability of CRA from repeated measures and (3) estimate the risk of cumulative exposure to phthalates based on their anti-androgenic and/or estrogenic properties. We multiplied the relative activity of individual phthalates exhibiting estrogenic or anti-androgenic effects by daily intake. For each preterm neonate, CRA was assessed based on the hazard index (HI) metric [the sum of hazard quotients] based on three reference doses for anti-androgenicity: the tolerable daily intake (TDI) from the European Food Safety Authority, the reference dose (RfD-AA) published in 2010 and newly revised published in 2020 (NRfD-AA). The metabolites of BBzP and DEHP were 2-23 fold higher in preterm neonates during their NICU stay. Median HIs increased in the order of HINRfDAA > HIRfDAA > HITDI. In the creatinine-based model, 87% (92%), 87% (96%), and 100% (100%) of preterm neonates at admission (during NICU stay) showed HITDI, HIRfD-AA, and HINRfD-AA exceeding 1, respectively with DEHP the most prevalent. The temporal reproducibility of HI (based on three reference doses) during preterm neonate stay in the NICU was high, with intra-class correlation coefficients ranging between 0.77 and 0.97, suggesting persistent exposure to phthalates. The four phthalates that preterm neonates were exposed to in the NICU exhibited estrogenic binding and anti-androgenic effects with median values (creatinine-based) of 98.7 and 56.9 µg/kg body weight/day, respectively. This was especially true for DEHP. The results indicate that preterm neonates in this NICU setting are probably at high risk of cumulative phthalate exposure with anti-androgenic properties that may have long-term adverse reproductive and developmental effects.

Exposure of preterm neonates receiving total parenteral nutrition to phthalates and its impact on neurodevelopment at the age of 2 months.

This prospective study assessed the exposure to phthalates of preterm neonates who received total parenteral nutrition (TPN) during their stay in the neonatal intensive care unit (NICU) and the risk of neurodevelopment delays at the age of 2 months. Our study recruited 33 preterm neonates who required TPN upon NICU admission. Urine samples for analyzing phthalate metabolites were obtained at admission and then daily until the last day of receiving TPN. Phthalates in the daily TPN received by the preterm neonates were analyzed. The neurodevelopment of the neonates was assessed using the Ages and Stages Questionnaire Edition 3 (ASQ-3). Diethyl phthalate and butyl benzyl phthalate were found in all TPN samples, while 27% and 83% contained dibutyl phthalate and di-(2-ethylhexyl) phthalate (DEHP), respectively. Yet, the daily dose of each phthalate that our preterm neonates received from TPN was much lower than the recommended tolerable limit. Urinary levels of monobenzyl phthalate and four metabolites of DEHP [i.e., mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate, mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), and mono-(2-ethyl-5-carboxypentyl) phthalate (MECPP)] and the sum of four DEHP metabolites (∑4DEHP) increased significantly in preterm neonates before discharge. However, these levels were not correlated with their phthalate parent compounds in TPN, suggesting other sources of exposure in the NICU. At 2 months, we found that urinary levels of mono-iso-butyl phthalate (MiBP), MECPP, MEHP, and ∑4DEHP were inversely related to fine motor skills. After adjusting for head circumference, the inverse relationships remained significant, suggesting direct effects from phthalates. Given the extreme vulnerability of our population, it is critical to minimize exposure to phthalates during their NICU stay.

Exposure of preterm neonates to toxic metals during their stay in the Neonatal Intensive Care Unit and its impact on neurodevelopment at 2 months of age.

BACKGROUND: Premature neonates might be exposed to toxic metals during their stay in the neonatal intensive care unit (NICU), which could adversely affect neurodevelopment; however, limited evidence is available. The present study was therefore designed to assess the exposure to mercury, lead, cadmium, arsenic, and manganese of preterm neonates who received total parenteral nutrition (TPN) and/or red blood cell (RBC) transfusions during their NICU stay and the risk of neurodevelopment delay at the age of 2 months. METHODS: We recruited 33 preterm neonates who required TPN during their NICU admission. Blood samples were collected for metal analysis at two different time points (admission and before discharge). Metals in the daily TPN received by preterm neonates were analyzed. Neurodevelopment was assessed using the Ages and Stages Questionnaire Edition 3 (ASQ-3). RESULTS: All samples of TPN had metal contamination: 96% exceeded the critical arsenic limit (0.3 µg/kg body weight/day); daily manganese intake from TPN for preterm neonates exceeded the recommended dose (1 µg/kg body weight) as it was added intentionally to TPN solutions, raising potential safety concerns. All samples of RBC transfusions exceeded the estimated intravenous reference dose for lead (0.19 µg/kg body weight). Levels of mercury, lead and manganese in preterm neonates at discharge decreased 0.867 µg/L (95% CI, 0.76, 0.988), 0.831 (95%CI, 0.779, 0.886) and 0.847 µg/L (95% CI, 0.775, 0.926), respectively. A decrease in ASQ-3-problem solving scores was associated with higher levels of blood lead in preterm neonates taken at admission (ß = -0.405, 95%CI = -0.655, -0.014), and with plasma manganese (ß = -0.562, 95%CI = -0.995, -0.172). We also observed an association between decreased personal social domain scores with higher blood lead levels of preterm neonates before discharge (ß = -0.537, 95%CI = -0.905, -0.045). CONCLUSION: Our findings provide evidence to suggest negative impacts on the neurodevelopment at 2 months of preterm infants exposed to certain metals, possibly related to TPN intake and/or blood transfusions received during their NICU stay. Preterm neonates may be exposed to levels of metals in utero.

The relationship between urinary phthalate metabolites and polycystic ovary syndrome in women undergoing in vitro fertilization: Nested case-control study.

Certain endocrine disruptor chemicals are involved in the pathogenesis of polycystic ovary syndrome (PCOS), a hormonal disease related to infertility in women. Phthalates are the most common plasticizers found in several consumer products. Experimental and epidemiologic evidence suggests that some phthalates disrupt endocrine functions in reproductive mechanisms and development. We previously measured the levels of eight phthalate metabolites in the urine of 599 Saudi women who underwent in vitro fertilization (IVF) treatment and were enrolled in a prospective study (2015-2017). The current nested case-control study aimed to determine the association between urinary levels of phthalate metabolites and PCOS. Overall, 441 women from the IVF study were identified as eligible for this study. Women in the case group included those diagnosed with PCOS (N = 82). The control group comprised those unable to conceive due to male azoospermia or who underwent preimplantation genetic diagnosis (N = 359). Most urinary phthalate metabolite levels were several-fold higher than those reported in national surveys from other countries. The ratio of luteinizing hormone to follicle-stimulating hormone, an index of PCOS, was significantly higher in the case than in the control group, with no indication of its association with phthalate metabolites. The logistic regression model was applied after adjusting for confounders to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for each metabolite modeled as a natural logarithm (ln). For each ln-unit increase in the sum of the four di (2-ethylhexyl) phthalate (∑4DEHP) metabolites as well as two individual metabolites, mono-(2-ethyl-5-oxohexyl) phthalate and mono-(2-ethyl-5-carboxypentyl) phthalate, the odds of PCOS increased by 40.5% [OR = 1.405 (95% CI: 1.025, 1.925)], 41.1% [OR = 1.055 (95% CI: 1.055, 1.885)], and 38.6% [OR = 1.386 (95% CI: 1.033, 1.86)], respectively. In contrast, the % odds of PCOS decreased marginally significantly by 44% [OR = 0.560 (95% CI: 0.313, 1.002)] with an ln-unit increase of %MEHP4, the ratio of mono-(2-ethylhexyl) phthalate to ∑4DEHP. These findings suggest that DEHP may contribute to PCOS, and further investigation is required to understand the underlying mechanisms.

Health Risk Assessment of Trace Metals Through Breast Milk Consumption in Saudi Arabia.

We previously measured the levels of inorganic mercury, methylmercury, lead, cadmium, and manganese in the breast milk of 203 healthy Saudi mothers who participated in a cross-sectional study between 2011 and 2013. The current study aimed to (1) calculate reference values (RVs) for these metals in breast milk based on the 95th percentile of the metal and the corresponding 95% confidence interval following the approach of the German Human Biomonitoring Commission, and (2) assess the health risk associated with these metals (except lead) by determining the hazard quotient (HQ) and hazard index (HI) for breastfed infants. The risk characterization for the lead was applied using the margin of exposure (MOE) approach. Moreover, the cancer risk (CR) associated with lead was calculated. The RV95s (percentage of samples for which the value was higher than the set value) for inorganic mercury, methylmercury, total mercury, cadmium, lead, and manganese in breast milk (µg/L) were 1.5 (7.9%), 1.5 (5.4%), 2.8 (8.9%), 2.5 (8.4%), 53 (11.3%), and 22.3 (11.8%) µg/L, respectively. The methylmercury, lead, and manganese levels in the present study were higher than those reported previously. The HQ for methylmercury greater than 1 was found in 68.5% of the samples, indicating there may be a potential non-carcinogenic health risk of infant exposure to the toxic metal via breast milk consumption. Despite the high cadmium and manganese levels in breast milk, our results suggested no health risk (HQ < 1). The HI representing the combined non-carcinogenic health risk of four metals was > 1, with methylmercury (74%) being the major contributor. The estimated MOE mean value of 0.134, less than 1, indicates that our breastfed infants may be at increased risk of neurodevelopmental impairments. The CR for lead in two infants was higher than the acceptable level of 1 × 10-4. Although our results may suggest potential carcinogenic and non-carcinogenic risks of infant exposure to toxic metals through breast milk consumption, the benefits of breastfeeding are well recognized and outweigh the potential risks.

Oxidative stress and DNA damage status in couples undergoing in vitro fertilization treatment.

This study examined the status of oxidative stress in 599 couples undertaking in vitro fertilization (IVF) treatment and its association with reproductive hormones, smoking, and outcomes. Oxidative stress biomarkers such as malondialdehyde, 8-hydroxy-2-deoxyguanosine, hydrogen peroxide (H2O2), catalase (CAT), and total antioxidant capacity (TAC) were determined in follicular fluid and seminal plasma. Tail moment (TM) was used to evaluate DNA damage in the sperm and granulosa cells. Reproductive hormones in serum and cotinine (COT) in urine, follicular fluid, and seminal plasma samples were determined. Separate multivariate linear regression was used to assess associations between levels of each oxidative stress biomarker and each hormone and smoking parameter (modeled as natural log-transformed). The findings indicate that some oxidative stress and DNA damage biomarkers played a role in disrupting certain reproductive hormones in women and their male partners either by overproducing reactive oxygen species or reducing antioxidant defense capacity. Although women were nonsmokers, COT levels > 50 and 10 µg/L in urine and follicular were observed in 5.7 and 1.7%, respectively. Levels of follicular fluid COT were positively associated with H2O2 and TM. We used log-binomial multivariate regression to estimate relative risks for the association between oxidative stress/DNA damage and IVF binary outcomes (fertilization rate > 50%, biochemical pregnancy, clinical pregnancy, and live birth). An increase in the CAT levels of follicular fluid was associated with a 48 and 41% decrease in the risk of poor fertilization rate (≤50%) and unsuccessful live birth, respectively. After the models were adjusted for hormonal factors, the associations remained the same, except that the elevated TAC in follicular fluid became significantly associated with a decrease of 42% in the risk of poor fertilization rate (≤50%). The higher antioxidant activity (CAT and TAC) in follicular fluid might positively impact specific IVF outcomes. LAY SUMMARY: Oxidative stress occurs when antioxidant molecules are insufficient in the body to destroy free radicals that can damage the cells, proteins and DNA, causing different health conditions, including infertility. The role of oxidative stress in female infertility has not received as much attention as male infertility, and research is still limited. This study explored whether the overproduction of free radicals can impact the success of in vitro fertilization (IVF) treatment using several biological markers such as hydrogen peroxide, catalase, and total antioxidant capacity. Our findings revealed that the high antioxidant levels in the fluid surrounding the egg were linked with a high fertilization rate. Additionally, oxidative stress status in couples was associated negatively with several reproductive hormones and smoking status. Biomarkers of oxidative stress and DNA damage might have potential applications in evaluating IVF patients' clinical characteristics such as causes of infertility, hormonal profile, fertilization rate, implantation and live birth.

The effect of caffeine consumption on the success rate of pregnancy as well various performance parameters of in-vitro fertilization treatment.

BACKGROUND: This study evaluated the effect of caffeine consumption on the success rate of pregnancy and various in vitro fertilization (IVF) performance parameters. MATERIAL/METHODS: Serum and follicular fluid samples were collected from 619 women undergoing IVF treatment (2002-2003). Caffeine assessment was based on measuring the levels of caffeine in serum and follicular fluid and on the number of coffee or tea or caffeinated drinks consumed per day. RESULTS: A total of 97.3% of participants reported the consumption of caffeinated drinks such as coffee, tea and soft drinks. Their average caffeine consumption was 455.82 mg/day (range: 3.71-3561 mg/day). Coffee was the primary source of caffeine intake. The average caffeine levels in serum (0.913 µg/ml) were significantly higher than in follicular fluid (0.701 µg/ml). After controlling for various potential confounding variables, no association was found between coffee or tea consumption and the success rate of pregnancy. Looking at the effect of caffeine consumption on the IVF performance parameters, we found that the number of eggs decreased as the caffeine serum levels increased (P=0.011). An increase in coffee consumption was positively associated with the number of aborted pregnancy (P=0.007), while the number of good embryo decreased with high tea consumption (P=0.015). CONCLUSIONS: Though no association was seen between coffee or tea consumption and pregnancy rate, this study is the first to report that caffeine can reach the follicular fluid and there is a suggestive evidence of its possible harmful role on the consequences of reproductive process. This clearly warrants further investigation.

Levels of DNA adducts in the blood and follicular fluid of women undergoing in vitro fertilization treatment and its correlation with the pregnancy outcome.

This study is designed to investigate the impact of DNA damage on pregnancy and fertilization rate outcome in a sub-sample of women undergoing IVF treatment. Blood and follicular fluid samples (n = 60) were analyzed for DNA adducts. While no BPDE-DNA adducts were detected, other unknown lipophilic adducts were seen in blood and follicular fluid. Women who failed to achieve pregnancy had higher DNA adducts in follicular fluid than those who succeeded (p < 0.05). Follicular fluid cotinine levels were associated with DNA adduct levels in blood and follicular fluid (p < 0.05). Evaluation of DNA damage resulting from oxidative stress could have a role in predicting IVF success rate.

Reference values for heavy metals in the urine and blood of Saudi women derived from two human biomonitoring studies.

We previously assessed the exposure of Saudi women to mercury, cadmium and lead based on reference values (RVs) established for other populations experiencing differences in environmental exposure, diet and lifestyle as an indicator of background exposure to pollutants. The present study aimed to (1) calculate RV95 for mercury, cadmium and lead in blood and urine from Saudi women based on the 95th percentile of the metal and its corresponding 95% confidence interval (95%CI) as defined by the German Human Biomonitoring Commission, and (2) compare with RV95s established in other countries. RV95s were derived using data from two different human biomonitoring studies measured three metals in the blood (2005-2006), and both urine and blood (2011-2013) from healthy non-smokers and non-occupationally exposed women living in Al-Kharj and Riyadh. RV95s for mercury, cadmium and lead in Al-Kharj (Riyadh) blood were 5.9 (1.6) µg/l, 1.4 (1.9) µg/l and 4.3 (4.8) µg/dl, respectively. RV95s for urinary mercury, cadmium and lead in Riyadh samples expressed in µg/l (µg/g creatinine) were 2.5 (1.9), 1.2 (0.96) and 14 (10.8), respectively. Values in both matrices remained high even when potential factors such as secondhand smoking, dental amalgam, and/or fish consumption excluded. RV95s for metals in blood were generally higher than in other countries, except for mercury in Riyadh samples, which was also 4-fold lower than in Al-Kharj. However, due to the time lag between the two studies, and since the Al-Kharj study was conducted 13 years ago, the most recent RV95s derived from the Riyadh study are recommended for Saudi women living in the Central region. The RV95s for urinary mercury and cadmium were comparable to other countries, but lead requires closer attention. To our knowledge, this study is the first to propose RV95s for mercury, cadmium, and lead in blood and urine of Saudi women of childbearing age, a population that is highly susceptible to the adverse health effects of these metals.

Effects of early and recent mercury and lead exposure on the neurodevelopment of children with elevated mercury and/or developmental delays during lactation: A follow-up study.

This follow-up study of 82 children investigated the potential impact of early and recent exposure to mercury and lead on their neurodevelopmental performance at 5-8 years of age (2017-2018). Early exposure of these children to mercury, methylmercury, and lead was assessed during lactation at 3-12 months old, as well as their mother's exposure using measurements from a cross-sectional study (2011-2013). Only infants who failed to pass the neurodevelopment screening tools and/or had elevated mercury were included in this study. Urine and hair were sampled during the follow-up study to assess the children's recent exposure to mercury, methylmercury, and lead. Their cognitive performance and visual-motor integration were also measured using the Test of Non-Verbal Intelligence (TONI) and the Beery-Visual-Motor Integration (Beery VMI), respectively. The association between alterations in urinary porphyrins excretion and exposure to metals was analyzed and their influence on the children's neurodevelopment was explored. Linear regression models revealed a significant negative association between the infants' mercury exposure during lactation and the TONI Quotient (ß = -0.298, 95%CI = -4.677, -0.414) and Beery VMI Age Equivalent scores at age 5-8 (ß = -0.437, 95%CI = -6.383, -1.844). The mothers' blood methylmercury was inversely and significantly associated with their children's TONI Quotient (ß = -0.231, 95%CI = -8.184, -0.331). In contrast, the children's Beery VMI Age Equivalent scores were positively and significantly associated with the hair methylmercury of the mothers (ß = 0.214, 95%CI = 0.088, 3.899) and their infants (ß = 0.256, 95%CI = 0.396, 4.488). These relationships suggest the presence of negative confounding that we did not take into account. Unlike mercury, there was some evidence that lead in breast milk had an inverse relationship with the children's visual-motor coordination skills. Our study did not show a clear association between children's recent exposure to metals and neurodevelopment. However, a significant inverse association was observed between the TONI Quotient and the interaction of hair methylmercury × ∑porphyrins (ß = -0.224, 95%CI = -0.86, -0.049), implying that porphyrins are a sensitive measure of low body-mercury burden. Although lead induced higher ∑porphyrins excretion in urine (ß = 0.347, 95%CI = 0.107, 0.525), their interaction did not influence children's neurodevelopmental scores. The interactions between metals and porphyrins might provide insights into their potential contributory role in the pathogenesis associated with neurological disorders or other diseases. Despite the small sample size of the present study, its findings about the association between toxic metal exposure and the high risk of poor neurodevelopmental performance are worrying, particularly at an early age, and additional research is needed using larger sample sizes.

Cytotoxic and genotoxic effects of e-liquids and their potential associations with nicotine, menthol and phthalate esters.

In this study, we determined DNA damage and chromosome breakage (indicators of genotoxicity) and cell viability (an indicator of cytotoxicity) in human lymphoblastoid TK6 and Chinese hamster ovary (CHO) cells treated with 33 e-liquids using in vitro single cell gel (comet), micronucleus (MN), and trypan blue assays, respectively. We also measured the contents of nicotine, five phthalate esters, and DL-menthol in the e-liquids to examine their effects on DNA damage, chromosome breakage, and cell viability. Our chemical analyses showed that: (1) six e-liquids had nicotine ≥2-fold higher than the manufacture's label claim (2-3.5 mg); (2) both dimethyl- and dibutyl-phthalate levels were >0.1 µg/g, i.e., their threshold limits as additives in cosmetics; and (3) the DL-menthol contents ranged from 0.0003 to 85757.2 µg/g, with those of two e-liquids being >1 mg/g, the threshold limit for trigging sensory irritation. Though all the e-liquids induced DNA damage in TK6 cells, 20 resulted in cell viabilities ≤75%, indicating cytotoxicity, yet the inverse relationship between cell viability and DNA damage (r = -0.628, p = 0.003) might reflect their role as pro-apoptotic and DNA damage inducers. Fifteen e-liquids induced MN% in TK6 cells ≥3-fold that of untreated cells. Some of the increase in %MN might be false due to high cytotoxicity, yet six brands showed acceptable cell viabilities (59-71%), indicating chromosome damage. DNA damage and %MN increased when the TK6 cells were exposed to metabolic activation. The CHO cells were less sensitive to the genotoxic effects of the e-liquids than the TK6 cells. DL-menthol was found to be associated with decreased cell viability and increased DNA damage, even at low levels. We cannot dismiss the presence of other ingredients in e-liquids with cytotoxic/genotoxic properties since out of the 63 different flavors, 47 induced DNA damage (≥3-folds), and 26 reduced cell viability (≤75%) in TK6 cells.

Outcome of in-vitro fertilization treatment and DDT levels in serum and follicular fluid.

BACKGROUND: Although p,p'-dichlorodiphenyltrichloroethane (DDT) is banned for agricultural purpose in Saudi Arabia, it is occasionally used to control vector-borne diseases in certain regions of the country. MATERIAL/METHODS: A case-control study was designed to investigate the possible effects of DDT and its metabolites on pregnancy and fertilization rate outcome. The study population was composed of 619 Saudi women (age 19-50 years) who sought in-vitro fertilization (IVF) treatment between 2002 and 2003. RESULTS: p,p'-DDE, the main metabolite of DDT, was the most frequently detected residue in serum or follicular fluid, with mean values of 1.646 microg/L and 0.407 microg/L, respectively. After controlling for many potential confounding variables, multiple logistic regression analysis revealed no association between pregnancy outcome or fertilization rate and p,p'-DDE levels in serum or follicular fluid. CONCLUSIONS: The inability to identify an effect may be related to the comparatively low concentrations of DDE in our population. But because p,p'-DDE was detected in the serum of 77.7% our participants, it should be considered as a matter of public heath concern. Currently there is no active source of DDT in our region; therefore, further studies are needed to identify sources in order to develop preventive measures because we can not exclude its potential reproductive toxicity.

Assessment of lead in cosmetic products.

There have been a number of recent reports in the media and on the internet about the presence of lead in brand-names lipsticks. This has drawn our attention to assess the safety of various cheap brands of cosmetics sold at 2-riyals stores in Saudi market that are imported from countries where safety regulations are poorly enforced as well as they lack perfect conditions for manufacturing. Lead contents were determined in 26 and eight different brands of lipsticks and eye shadows using the Zeeman atomic absorption spectrophotometer coupled to graphite tube atomizer after an acid digestion procedure. Lead was detected in all the studied samples. The median (25th-75th percentile) lead content in 72 lipsticks samples was 0.73 (0.49-1.793) PPM wet wt. in the range of 0.27-3760 PPM wet wt. There were four brands of lipsticks with lead content above the FDA lead limit as impurities in color additives (20 PPM). The FDA does not set a limit for lead in lipstick. Three of them were extremely high points and considered outliers. The median (25th-75th percentile) lead contents in pressed powder eye shadow was 1.38 (0.944-1.854) PPM wet wt. (n=22) in the range of 0.42-58.7 PPM wet wt. One brand was above 20 PPM the US FDA's lead limit as impurities. The overall results indicate that lead in lipsticks and eye shadows are below the FDA lead limit as impurities and, thus, probably have no significant toxicological effects. Nevertheless, few brands had lead content above 20 PPM that might put consumers at the risk of lead poisoning. Lead is a cumulative, and applying lead-containing cosmetics several times a day or every day, can potentially add up to significant exposure levels. Pregnant and nursing mothers are vulnerable population because lead passes through placenta and human milk and affect fetus or infant's developments. Our findings call for an immediate mandatory regular testing program to check lead and other toxic metals in lipsticks and other cosmetic products imported to Saudi Arabia in order to curtail their excess and safeguard consumer health.

Couples exposure to phthalates and its influence on in vitro fertilization outcomes.

This prospective study examined the associations between the levels of eight urinary phthalate metabolites in 599 couples and in vitro fertilization (IVF) outcomes. We used log-binomial multivariate regression to estimate relative risks (RR) for the association between phthalate concentration and IVF binary outcomes (fertilization rate >50%, biochemical pregnancy, clinical pregnancy and live birth) for each woman after adjusting the model for the concentration in a male partner and each relevant confounders. RR was expressed per unit increase in log-transformed urinary metabolite concentration. The percentage of bis-2-ethylhexyl phthalate (DEHP) metabolites excreted as mono-2-ethylhexyl phthalate (MEHP) was calculated as %MEHP. Urinary MEHP in women was associated with an increased risk of biochemical pregnancy (RR = 1.35; p = 0.04), failed clinical pregnancy (RR = 1.56; p = 0.006) and live birth (RR = 1.54; p = 0.011). An increase in monoethyl phthalate was associated with a high risk of failed clinical pregnancy (RR = 1.25; p = 0.03) and live birth (RR = 1.35; p = 0.006). An increase in %MEHP was associated with an increase in the risk of biochemical pregnancy (RR = 1.55; p = 0.05), failed clinical pregnancy (RR = 1.73; p = 0.02) and live birth (RR = 1.65; p = 0.046). Our results demonstrated that exposure to some phthalates may adversely affect IVF outcomes, particularly when couples' exposure was jointly modeled, emphasizing the importance of a couple-based approach in assessing fertility outcomes. The associations between IVF outcomes and DEHP metabolites were stronger in women whose %MEHP was >75th percentile which may be due to their less efficient metabolism and excretion of DEHP and/or MEHP.