Live Birth, Pregnancy, Spontaneous Abortion
Conditions
Keywords
Folic Acid, Zinc, Semen, In vitro fertilization, Assisted reproductive technology, Ovulation induction, Intrauterine insemination, Pregnancy, Live Birth, Abortion, spontaneous
Brief summary
The overarching goal of this trial is to determine if an intervention comprising folic acid and zinc dietary supplementation improves semen quality and indirectly fertility outcomes (i.e., live birth rate) among couples trying to conceive and seeking assisted reproduction. The following study objectives underlie successful attainment of the overarching research goal: 1. To estimate the effect of folic acid and zinc dietary supplementation on semen quality parameters, including but not limited to concentration, motility, morphology, and sperm DNA integrity, relative to the placebo group. 2. To estimate the effect of folic acid and zinc dietary supplementation on fertility treatment outcomes \[fertilization, embryo quality, implantation/human Chorionic Gonadotropin (hCG) confirmed pregnancy, clinical pregnancy, live birth\], relative to the placebo group. 3. To estimate the association between semen quality parameters, sperm DNA integrity and fertility treatment outcomes (fertilization, embryo quality, clinical pregnancy, live birth) and to identify the best combination of semen quality parameters for prediction of clinical pregnancy and live birth. 4. To estimate the effect of folic acid and zinc dietary supplementation on fertilization rates among couples undergoing assisted reproductive technology procedures, relative to the placebo group. 5. To estimate the effect of folic acid and zinc dietary supplementation on embryonic quality among couples undergoing assisted reproductive technology procedures, relative to the placebo group.
Detailed description
Two micronutrients fundamental to the process of spermatogenesis, folic acid (folate) and zinc, are of particular interest for fertility as they are of low cost and wide availability. Though the evidence has been inconsistent, small randomized trials and observational studies show that folate and zinc have biologically plausible effects on spermatogenesis and improved semen parameters. These results support the potential benefits of folate on spermatogenesis and suggest that dietary supplementation with folate and zinc may help maintain and improve semen quality, and perhaps, fertility rates. The Epidemiology Branch of the Eunice Kennedy Shriver National Institute of Child Health and Human Development intends to conduct a multi-site double-blind, randomized controlled clinical trial to evaluate the effect of folic acid and zinc dietary supplementation on semen quality and conception rates among male partners of couples seeking assisted reproduction. Randomization will be stratified (with random sequences of block sizes) by site and assisted reproduction technique (IVF, non-IVF receiving fertility treatment at a study site, and non-IVF receiving fertility treatment at a nonstudy site) to ensure that balance between the treatment groups is maintained within site and within fertility treatment type over the enrollment period. The study is designed with a sample size of 2,400 randomized participants based on obtaining adequate power to detect meaningful differences in the live birth rate between cohorts. Since the comparison of sperm parameters are differences between continuous assay measurements, this sample size will be more than sufficient for the primary sperm parameter comparisons. Additionally, calculations were done to demonstrate adequate statistical power when stratified analysis is to be performed (i.e., sample size distributions among the strata and their corresponding live birth RRs detected at 80% statistical power, with an alpha level of 0.05 and a total sample size of 2400 couples divided among the folic acid/zinc and placebo arms of the trial). Data collection will include screening male and female partners for eligibility, administering baseline questionnaires, and collecting biospecimens in both partners of the couple, body measurements for both partners, daily journal reporting for male partners, medical record abstraction related to required treatment and outcome data, and semen quality of four samples collected at baseline, two, four, and six months following study enrollment. A data coordinating center (DCC) will support the trial. The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two cohorts based on the randomized assignment, both overall and by treatment strata (IVF, non-IVF receiving fertility treatment at a study site, and non-IVF receiving fertility treatment at a nonstudy site).This approach will be applied to the two primary endpoints (semen parameters and live birth rate) as well as designated secondary endpoints (number of follicles, number and proportion of oocytes fertilized). The DCC will perform periodic safety analyses and present interim reports to the Data and Safety Monitoring Board (DSMB) as requested, during the recruitment phases of the trial. It is anticipated that safety analyses will be performed every 6-12 months. The final analysis will be performed upon completion of data collection and editing in the follow-up and close-out phase of the trial. Also one full formal interim analysis is planned and the power calculations with considerations for the choice of optimal time for the analysis have been conducted.
Interventions
DIETARY_SUPPLEMENT5 mg folic acid and 30 mg elemental zinc
Sponsors
Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD)
Study design
Allocation
RANDOMIZED
Intervention model
PARALLEL
Primary purpose
TREATMENT
Masking
QUADRUPLE (Subject, Caregiver, Investigator, Outcomes Assessor)
Eligibility
Sex/Gender
ALL
Age
18 Years to 45 Years
Healthy volunteers
Yes
Inclusion criteria
Couples Inclusion Criteria: 1. Heterosexual couples in a committed relationship with a female partner aged 18-45 years and male partner aged 18 years and older attempting to conceive and seeking assisted reproduction at participating fertility clinics. 2. Couples actively trying to conceive. 3. Couples who are planning ovulation induction (OI), natural fertility optimization methods, or intrauterine insemination (IUI) should be willing to be on the study dietary supplement for at least 3 weeks before starting the next assisted reproduction cycle.Women with regular periods may initiate their fertility therapy at the start of the woman's menstrual cycle following randomization if randomization occurred within the first 10 days of the cycle, but must wait one menstrual cycle if the visit occurred after day 10 of the cycle). For women with irregular periods or amenorrhea, the male must be on the study supplement for 3 weeks prior to initiation of any ovulation induction medication (e.g., clomid, letrozole, gonadotropins). Couples
Exclusion criteria
1. Female partner unwilling to participate (e.g., no abstraction of her assisted fertility treatment record or unwilling to complete baseline visit). 2. Couples using donor, cryopreserved sperm, or sperm obtained via microsurgical or percutaneous epididymal sperm aspiration. 3. Couples attempting to conceive with a gestational carrier (surrogate). 4. Positive urine pregnancy test at screening. Male Inclusion Criteria: 1. Willing to provide semen samples according to the proposed schedule at baseline, 2, 4, and 6 months of follow-up. 2. Able to complete regular study questionnaires and daily journals aimed at capturing ejaculation, sexual intercourse and lifestyle factors considered to affect male fecundity (e.g., cigarette smoking, fever, high temperature environment and other environmental exposures) and other data collection instruments (e.g., physical activity, food frequency questionnaire, stress). Male
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Total Motile Sperm Count | 6 months | Calculated as semen volume (mL) \* sperm concentration (10\^6 spermatozoa/mL) \* motility (% motile) |
| Live Birth | At delivery | Based on hospital delivery records |
| Semen Volume | 6 months | Volume of the ejaculate, mL Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010. |
| Sperm Concentration | 6 months | Number of spermatozoa per unit of volume of semen Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010. |
| Sperm Motility | 6 months | % motile (including percentage of progressive motile sperm and percentage of nonprogressive motile sperm) Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010. |
| Sperm Morphology | 6 months | % normal morphology Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010. |
| DNA Fragmentation Index | 6 months | Comet assay used to measure sperm DNA integrity based on excess DNA strand breaks Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010. |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Cesarean Delivery | Delivery | Abstracted from hospital records and medical charts |
| Preterm Delivery | Delivery | Abstracted from hospital records and medical charts |
| Small for Gestational Age | Delivery | Abstracted from hospital records and medical charts |
| Gestational Age | Delivery | Abstracted from hospital records and medical charts |
| Birth Weight | Delivery | Abstracted from hospital records and medical charts |
| Stillbirth | Delivery | Loss at or after 20 weeks gestation. Determined based on hospital records and medical chart abstraction. |
| Neonatal Mortality | Delivery | Abstracted from hospital records and medical charts |
| Major Neonatal Complications | Delivery | Abstracted from hospital records and medical charts: includes bronchopulmonary dysplasia, necrotizing enterocolitis, severe intraventricular hemorrhage, periventricular leukomalacia, and retinopathy of prematurity |
| Structural Malformations | Delivery | Abstracted from birth record: includes major (n = 21; 6 with known genetic cause), minor (n = 6), and unclassified (n = 2) defects Structural birth defects: includes hydronephrosis/ureteropelvic junction obstruction, transposition of the great arteries, renal agenesis, cleft lip, club feet, multicystic/dysplastic kidney, tetralogy of fallot, gastroschisis, atrioventricular septal defects, other oral-facial defects, other cardiovascular defects, other CNS defects, other eye defects, other oral-facial defects, other anomalies, other syndromes |
| Severe Maternal Morbidity | Delivery | Abstracted from delivery record: including postpartum hemorrhage, anemia requiring transfusion, sepsis, seizure, HELLP syndrome or preeclampsia with pulmonary edema |
| Fertilization Rate Per Cycle, % | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum Oocytes will be assessed 16-18 hours after insemination or microinjection to determine whether fertilization occurred. Fertilization will be considered normal if two pronuclei and two polar bodies are identified. Oocytes without visible pronuclei will be considered unfertilized. Oocytes with more than two pronuclei will be considered abnormally fertilized, and will thus be discarded. |
| Number of Good Quality Embryos on Day 5 Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria. |
| Number of Embryos Transferred Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum |
| Number of Embryos Cryopreserved Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum |
| Sperm Penetration Per Cycle, % | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum |
| Cells on Day 3 Per Embryo Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum |
| Cells on Day 3 Per Embryo Per Cycle, Categorical | Up to 9 months of fertility treatment post-randomization | Number of cells per embryo among women in the IVF stratum |
| Cells on Day 5 Per Embryo Per Cycle, Categorical | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum |
| Embryo Morphology on Day 3 Per Cycle, Categorical | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum Embryos will be scored three days after fertilization according to the size and shape of blastomeres and to their degree of fragmentation. Veeck LL. Oocyte assessment and biological performance. Ann N Y Acad Sci 1988;541:259-74.:259-74. |
| Embryo Morphology on Day 5 Per Cycle, Categorical | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria. |
| Method of Fertilization Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the in vitro fertilization (IVF) stratum: method of fertilization classified into intracytoplasmic sperm injection (ICSI) and other |
| Quality of Embryos Transferred Per Cycle, Categorical | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum Embryonic grading based on Society for Assisted Reproductive Technologies (SART) morphology criteria |
| Chromosomal Complement of Embryo Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum Chromosomal complement in the embryo assessed using methodology cited by Rubio et al. Rubio C, Rodrigo L, Mir P et al. Use of array comparative genomic hybridization (array-CGH) for embryo assessment: clinical results. Fertil Steril 2013 March 15;99(4):1044-8. |
| Percentage of Good Quality Embryos on Day 5 Per Cycle | Up to 9 months of fertility treatment post-randomization | Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria. |
| Clinical Intrauterine Pregnancy | approximately 6.5 weeks gestation | Visualized gestational sac in the uterus on ultrasound |
| Human Chorionic Gonadotropin (hCG) Detected Pregnancy (Implantation) | For IVF, 12 days post embryo transfer for day 5 embryo transfers, and 14 days post embryo transfer for day 3 embryo transfers; for couples undergoing OI/IUI, after self-report of positive pregnancy test | A quantitative hCG evaluation in serum \> 5 milli-international units per milliliter (mIU/ml) |
| Ectopic Pregnancy | approximately 6.5 weeks gestation | Either visualization of no gestational sac in the uterus with a suspicious mass in the adnexa on ultrasound, an hCG level more than 1500 mIU/ml without visualization of an intrauterine gestational sac on ultrasound, or a slowly rising or plateauing serum hCG level without visualization of an intrauterine gestation on ultrasound. |
| Early Pregnancy Loss | hcG-detected pregnancy until 20 weeks of pregnancy | hCG pregnancy loss will be defined as a serum hCG \> 5 mIU/ml followed by a decline. Clinically recognized pregnancy losses will be defined as visualization of an intrauterine gestational sac followed by a loss prior to 20 weeks gestation. |
| Preeclampsia or Gestational Hypertension | Delivery | Abstracted from hospital records and medical charts |
| Gestational Diabetes | Delivery | Abstracted from hospital records and medical charts |
Other
| Measure | Time frame | Description |
|---|---|---|
| Reproductive Hormones and Other Measured Biomarkers | 4 or 6 months | Urinary, serum, and salivary concentrations of reproductive hormones, particularly androgens, proteomic analysis of human sperm and cardiometabolic risk factors and markers of oxidative stress, as well as measures of trace elements in toenails (collected at month 4 clinic visit). Biospecimens have been collected but laboratory analysis still needs to be done to be able to evaluate these endpoints. |
Countries
United States
Participant flow
Pre-assignment details
Only the male partners in couples attempting to conceive were enrolled and assigned to treatment arms.
Participants by arm
| Arm | Count |
|---|---|
| Folic Acid and Zinc Supplementation 5 mg folic acid and 30 mg elemental zinc, taken orally, daily for 6 months.
5 mg folic acid and 30 mg elemental zinc | 2,370 |
| Folic Acid and Zinc Supplementation 5 mg folic acid and 30 mg elemental zinc, taken orally, daily for 6 months.
5 mg folic acid and 30 mg elemental zinc | 1,185 |
| Placebo Matching placebo, taken orally daily for 6 months.
Placebo Comparator: Placebo | 2,370 |
| Placebo Matching placebo, taken orally daily for 6 months.
Placebo Comparator: Placebo | 1,185 |
| Total | 7,110 |
Withdrawals & dropouts
| Period | Reason | FG000 | FG001 |
|---|---|---|---|
| Secondary Analysis: Semen Quality at 6m | Lost to Follow-up | 315 | 282 |
Baseline characteristics
| Characteristic | Folic Acid and Zinc Supplementation | Placebo | Total |
|---|---|---|---|
| Age, Continuous | 30.6 years STANDARD_DEVIATION 5 | 30.8 years STANDARD_DEVIATION 5.2 | 30.7 years STANDARD_DEVIATION 5.1 |
| Annual household income $100,000 and greater | 252 Male-female couples | 278 Male-female couples | 530 Male-female couples |
| Annual household income <$40,000 | 176 Male-female couples | 157 Male-female couples | 333 Male-female couples |
| Annual household income $40,000-$74,999 | 422 Male-female couples | 456 Male-female couples | 878 Male-female couples |
| Annual household income $75,000-$99,999 | 261 Male-female couples | 232 Male-female couples | 493 Male-female couples |
| Annual household income Do not wish to provide | 74 Male-female couples | 62 Male-female couples | 136 Male-female couples |
| Body mass index (BMI) | 30.1 kg/m^2 STANDARD_DEVIATION 6.7 | 29.6 kg/m^2 STANDARD_DEVIATION 6.7 | 29.8 kg/m^2 STANDARD_DEVIATION 6.7 |
| Diastolic blood pressure | 78.4 mmHg STANDARD_DEVIATION 10.8 | 78.0 mmHg STANDARD_DEVIATION 10.9 | 78.2 mmHg STANDARD_DEVIATION 10.9 |
| Education Bachelor's degree | 335 Participants | 389 Participants | 724 Participants |
| Education Do not wish to provide | 12 Participants | 17 Participants | 29 Participants |
| Education High school or less | 198 Participants | 173 Participants | 371 Participants |
| Education Master's degree or higher | 212 Participants | 220 Participants | 432 Participants |
| Education Some college | 428 Participants | 386 Participants | 814 Participants |
| Employment status Employed full-time | 802 Participants | 798 Participants | 1600 Participants |
| Employment status Employed part-time | 58 Participants | 53 Participants | 111 Participants |
| Employment status Full-time student | 87 Participants | 96 Participants | 183 Participants |
| Employment status Not employed | 149 Participants | 148 Participants | 297 Participants |
| Male factor infertility diagnosis No | 598 Participants | 594 Participants | 1192 Participants |
| Male factor infertility diagnosis Yes | 160 Participants | 165 Participants | 325 Participants |
| Male health insurance No | 65 Participants | 54 Participants | 119 Participants |
| Male health insurance Yes | 1108 Participants | 1117 Participants | 2225 Participants |
| Male infertility insurance Do not know | 316 Participants | 355 Participants | 671 Participants |
| Male infertility insurance No | 493 Participants | 495 Participants | 988 Participants |
| Male infertility insurance Yes | 297 Participants | 266 Participants | 563 Participants |
| Marital status Do not wish to provide | 1 Male-female couples | 2 Male-female couples | 3 Male-female couples |
| Marital status Married/living with partner | 1180 Male-female couples | 1179 Male-female couples | 2359 Male-female couples |
| Marital status Single/other | 4 Male-female couples | 4 Male-female couples | 8 Male-female couples |
| Race/Ethnicity, Customized Asian | 43 Participants | 70 Participants | 131 Participants |
| Race/Ethnicity, Customized Do not wish to provide | 5 Participants | 7 Participants | 10 Participants |
| Race/Ethnicity, Customized Hispanic or Latino | 70 Participants | 75 Participants | 127 Participants |
| Race/Ethnicity, Customized Non-Hispanic black | 22 Participants | 21 Participants | 43 Participants |
| Race/Ethnicity, Customized Non-Hispanic white | 974 Participants | 962 Participants | 1956 Participants |
| Race/Ethnicity, Customized Other racial/ethnic groups | 51 Participants | 68 Participants | 142 Participants |
| Sex: Female, Male Female | 1185 Participants | 1185 Participants | 2370 Participants |
| Sex: Female, Male Male | 1185 Participants | 1185 Participants | 2370 Participants |
| Systolic blood pressure | 126.8 mmHg STANDARD_DEVIATION 12.8 | 126.5 mmHg STANDARD_DEVIATION 13.9 | 126.7 mmHg STANDARD_DEVIATION 13.4 |
| Taking multivitamin within past 3 mo No | 453 Participants | 468 Participants | 921 Participants |
| Taking multivitamin within past 3 mo Yes | 298 Participants | 284 Participants | 582 Participants |
| Time trying to conceive | 19 months | 18 months | 18 months |
Adverse events
| Event type | EG000 affected / at risk | EG001 affected / at risk |
|---|---|---|
| deaths Total, all-cause mortality | 0 / 1,185 | 0 / 1,185 |
| other Total, other adverse events | 189 / 1,185 | 162 / 1,185 |
| serious Total, serious adverse events | 7 / 1,185 | 5 / 1,185 |
Outcome results
Primary
DNA Fragmentation Index
Comet assay used to measure sperm DNA integrity based on excess DNA strand breaks Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010.
Time frame: 6 months
Population: 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit. Additionally, 44 men in the active arm and 55 men in the placebo arm had insufficient quantity or quality to assess DFI.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | DNA Fragmentation Index | Overall | 29.7 % breakage | Standard Deviation 20.5 |
| Folic Acid and Zinc Supplementation | DNA Fragmentation Index | IVF stratum | 27.1 % breakage | Standard Deviation 19.6 |
| Folic Acid and Zinc Supplementation | DNA Fragmentation Index | Other treatment onsite | 30.0 % breakage | Standard Deviation 20.3 |
| Folic Acid and Zinc Supplementation | DNA Fragmentation Index | Other treatment offsite | 30.7 % breakage | Standard Deviation 22.7 |
| Placebo | DNA Fragmentation Index | Other treatment offsite | 28.5 % breakage | Standard Deviation 20.5 |
| Placebo | DNA Fragmentation Index | Overall | 27.2 % breakage | Standard Deviation 17.8 |
| Placebo | DNA Fragmentation Index | Other treatment onsite | 27.0 % breakage | Standard Deviation 16.8 |
| Placebo | DNA Fragmentation Index | IVF stratum | 26.8 % breakage | Standard Deviation 19.6 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. DNA fragmentation will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.5, 4.4]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. DNA fragmentation will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.3, 4.3]
Primary
Live Birth
Based on hospital delivery records
Time frame: At delivery
Population: Complete randomized cohort
| Arm | Measure | Group | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Live Birth | Overall | 404 Participants |
| Folic Acid and Zinc Supplementation | Live Birth | IVF stratum | 97 Participants |
| Folic Acid and Zinc Supplementation | Live Birth | Other treatment onsite | 264 Participants |
| Folic Acid and Zinc Supplementation | Live Birth | Other treatment offsite | 43 Participants |
| Placebo | Live Birth | Other treatment offsite | 48 Participants |
| Placebo | Live Birth | Overall | 416 Participants |
| Placebo | Live Birth | Other treatment onsite | 277 Participants |
| Placebo | Live Birth | IVF stratum | 91 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Live birth will be compared using two sided tests conducted at the 0.05 level.95% CI: [-4.7, 2.8]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Live birth will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.88, 1.09]
Primary
Semen Volume
Volume of the ejaculate, mL Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010.
Time frame: 6 months
Population: Analyzed overall and by fertility treatment stratum (subgroup) 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | Semen Volume | Other treatment onsite | 3.5 mL | Standard Deviation 1.7 |
| Folic Acid and Zinc Supplementation | Semen Volume | Overall | 3.5 mL | Standard Deviation 1.7 |
| Folic Acid and Zinc Supplementation | Semen Volume | IVF stratum | 3.5 mL | Standard Deviation 1.5 |
| Folic Acid and Zinc Supplementation | Semen Volume | Other treatment offsite | 3.5 mL | Standard Deviation 1.7 |
| Placebo | Semen Volume | Other treatment offsite | 3.4 mL | Standard Deviation 1.7 |
| Placebo | Semen Volume | Other treatment onsite | 3.5 mL | Standard Deviation 1.8 |
| Placebo | Semen Volume | IVF stratum | 3.5 mL | Standard Deviation 1.7 |
| Placebo | Semen Volume | Overall | 3.5 mL | Standard Deviation 1.8 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Semen volume will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.2, 0.2]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Semen volume will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.1, 0.2]
Primary
Sperm Concentration
Number of spermatozoa per unit of volume of semen Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010.
Time frame: 6 months
Population: 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit. Additionally, one male participant in the placebo arm had insufficient quantity or quality to assess concentration.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | Sperm Concentration | Overall | 84.8 10^6 spermatozoa/mL | Standard Deviation 85.2 |
| Folic Acid and Zinc Supplementation | Sperm Concentration | IVF stratum | 81.8 10^6 spermatozoa/mL | Standard Deviation 96.5 |
| Folic Acid and Zinc Supplementation | Sperm Concentration | Other treatment onsite | 85.0 10^6 spermatozoa/mL | Standard Deviation 83.1 |
| Folic Acid and Zinc Supplementation | Sperm Concentration | Other treatment offsite | 87.2 10^6 spermatozoa/mL | Standard Deviation 83 |
| Placebo | Sperm Concentration | Other treatment offsite | 87.7 10^6 spermatozoa/mL | Standard Deviation 92.5 |
| Placebo | Sperm Concentration | Overall | 89.0 10^6 spermatozoa/mL | Standard Deviation 85 |
| Placebo | Sperm Concentration | Other treatment onsite | 92.2 10^6 spermatozoa/mL | Standard Deviation 84.8 |
| Placebo | Sperm Concentration | IVF stratum | 76.1 10^6 spermatozoa/mL | Standard Deviation 78.6 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm concentration represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-12.5, 3.9]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm concentration represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-13.6, 3.1]
Primary
Sperm Morphology
% normal morphology Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010.
Time frame: 6 months
Population: 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit. Additionally, 19 men in the active arm and 17 men in the placebo arm had insufficient quantity or quality to assess morphology.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | Sperm Morphology | Overall | 5.7 % normal | Standard Deviation 4.2 |
| Folic Acid and Zinc Supplementation | Sperm Morphology | IVF stratum | 5.2 % normal | Standard Deviation 4.3 |
| Folic Acid and Zinc Supplementation | Sperm Morphology | Other treatment onsite | 5.6 % normal | Standard Deviation 4 |
| Folic Acid and Zinc Supplementation | Sperm Morphology | Other treatment offsite | 6.7 % normal | Standard Deviation 4.9 |
| Placebo | Sperm Morphology | Other treatment offsite | 5.6 % normal | Standard Deviation 4.3 |
| Placebo | Sperm Morphology | Overall | 6.0 % normal | Standard Deviation 4.8 |
| Placebo | Sperm Morphology | Other treatment onsite | 6.2 % normal | Standard Deviation 4.9 |
| Placebo | Sperm Morphology | IVF stratum | 5.4 % normal | Standard Deviation 4.7 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm morphology represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-0.8, 0.1]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm morphology represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-0.9, 0]
Primary
Sperm Motility
% motile (including percentage of progressive motile sperm and percentage of nonprogressive motile sperm) Assessed utilizing the World Health Organization (WHO) semen analysis procedure 5th edition World Health Organization. WHO laboratory manual for the Examination and processing of human semen. 5th Edition ed. Switzerland: 2010.
Time frame: 6 months
Population: 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit. Additionally, one male participant in the placebo arm had insufficient quantity or quality to assess motility.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | Sperm Motility | Overall | 52.7 % motile | Standard Deviation 21.2 |
| Folic Acid and Zinc Supplementation | Sperm Motility | IVF stratum | 51.7 % motile | Standard Deviation 21.9 |
| Folic Acid and Zinc Supplementation | Sperm Motility | Other treatment onsite | 52.5 % motile | Standard Deviation 21.1 |
| Folic Acid and Zinc Supplementation | Sperm Motility | Other treatment offsite | 55.0 % motile | Standard Deviation 20.9 |
| Placebo | Sperm Motility | Other treatment offsite | 51.5 % motile | Standard Deviation 22.8 |
| Placebo | Sperm Motility | Overall | 53.2 % motile | Standard Deviation 20.1 |
| Placebo | Sperm Motility | Other treatment onsite | 53.9 % motile | Standard Deviation 19.5 |
| Placebo | Sperm Motility | IVF stratum | 51.7 % motile | Standard Deviation 20 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm motility represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-2.5, 1.5]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. The confidence interval for sperm motility represents 95.1% coverage to properly account for the alpha spent in the interim analysis.95.1% CI: [-2.7, 1.4]
Primary
Total Motile Sperm Count
Calculated as semen volume (mL) \* sperm concentration (10\^6 spermatozoa/mL) \* motility (% motile)
Time frame: 6 months
Population: 870 and 903 men in the active and placebo treatment arms, respectively, returned for the 6-month study visit. 76 men in the active arm and 67 men in the placebo arm did not provide semen samples at this visit. Additionally, 1 man in the active arm and 2 men in the placebo arm had insufficient quantity or quality to assess total motile count.
| Arm | Measure | Group | Value (MEAN) | Dispersion |
|---|---|---|---|---|
| Folic Acid and Zinc Supplementation | Total Motile Sperm Count | Overall | 183 million motile sperm | Standard Deviation 226 |
| Folic Acid and Zinc Supplementation | Total Motile Sperm Count | IVF stratum | 165 million motile sperm | Standard Deviation 221 |
| Folic Acid and Zinc Supplementation | Total Motile Sperm Count | Other treatment onsite | 186 million motile sperm | Standard Deviation 226 |
| Folic Acid and Zinc Supplementation | Total Motile Sperm Count | Other treatment offsite | 192 million motile sperm | Standard Deviation 233 |
| Placebo | Total Motile Sperm Count | Other treatment offsite | 184 million motile sperm | Standard Deviation 262 |
| Placebo | Total Motile Sperm Count | Overall | 182 million motile sperm | Standard Deviation 212 |
| Placebo | Total Motile Sperm Count | Other treatment onsite | 188 million motile sperm | Standard Deviation 207 |
| Placebo | Total Motile Sperm Count | IVF stratum | 152 million motile sperm | Standard Deviation 188 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Total motile sperm count will be compared using two sided tests conducted at the 0.05 level.95% CI: [-19.7, 22.5]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Total motile sperm count will be compared using two sided tests conducted at the 0.05 level.95% CI: [-20.9, 21.4]
Secondary
Birth Weight
Abstracted from hospital records and medical charts
Time frame: Delivery
Population: Among participants with live birth
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Birth Weight | 3062 grams | Standard Deviation 731 |
| Placebo | Birth Weight | 3133 grams | Standard Deviation 654 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Birth weight will be compared using two sided tests conducted at the 0.05 level.95% CI: [-156, 26.4]
Secondary
Cells on Day 3 Per Embryo Per Cycle
Among participants in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 1215 embryos among cycles among women in the FA/Zn arm; 1129 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively~\# of cells on day 3 unavailable in medical records for 25 and 36 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Cells on Day 3 Per Embryo Per Cycle | 5.60 cells per embryo | Standard Deviation 0.2 |
| Placebo | Cells on Day 3 Per Embryo Per Cycle | 5.98 cells per embryo | Standard Deviation 0.19 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of cells on day 3 will be compared with Poisson regression using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [-0.16, 0.03]
Secondary
Cells on Day 3 Per Embryo Per Cycle, Categorical
Number of cells per embryo among women in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 1222 embryos among cycles among women in the FA/Zn arm; 1140 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively~\# of cells on day 3 (categorical) unavailable in medical records for 25 and 36 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Cells on Day 3 Per Embryo Per Cycle, Categorical | Fewer than 4 cells | 0.27 predicted probability |
| Folic Acid and Zinc Supplementation | Cells on Day 3 Per Embryo Per Cycle, Categorical | 4 cells or greater | 0.73 predicted probability |
| Placebo | Cells on Day 3 Per Embryo Per Cycle, Categorical | Fewer than 4 cells | 0.22 predicted probability |
| Placebo | Cells on Day 3 Per Embryo Per Cycle, Categorical | 4 cells or greater | 0.78 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of cells on day 3 will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.88, 1.61]
Secondary
Cells on Day 5 Per Embryo Per Cycle, Categorical
Among participants in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 1160 embryos among cycles among women in the FA/Zn arm and 1205 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively~\# of cells on day 5 (categorical) unavailable in medical records for 5 and 14 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Cells on Day 5 Per Embryo Per Cycle, Categorical | Fewer than 8 cells | 0.20 predicted probability |
| Folic Acid and Zinc Supplementation | Cells on Day 5 Per Embryo Per Cycle, Categorical | 8 cells or greater | 0.80 predicted probability |
| Placebo | Cells on Day 5 Per Embryo Per Cycle, Categorical | Fewer than 8 cells | 0.19 predicted probability |
| Placebo | Cells on Day 5 Per Embryo Per Cycle, Categorical | 8 cells or greater | 0.81 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of cells on day 5 will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.77, 1.43]
Secondary
Cesarean Delivery
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Cesarean Delivery | 143 Participants |
| Placebo | Cesarean Delivery | 129 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Cesarean delivery will be compared using two sided tests conducted at the 0.05 level.95% CI: [-1.4, 3.8]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Cesarean delivery will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.89, 1.39]
Secondary
Chromosomal Complement of Embryo Per Cycle
Among participants in the IVF stratum Chromosomal complement in the embryo assessed using methodology cited by Rubio et al. Rubio C, Rodrigo L, Mir P et al. Use of array comparative genomic hybridization (array-CGH) for embryo assessment: clinical results. Fertil Steril 2013 March 15;99(4):1044-8.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Chromosomal complement of embryo unavailable in medical records for 121 and 124 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Chromosomal Complement of Embryo Per Cycle | Abnormal | 0.75 predicted probability |
| Folic Acid and Zinc Supplementation | Chromosomal Complement of Embryo Per Cycle | Normal | 0.25 predicted probability |
| Placebo | Chromosomal Complement of Embryo Per Cycle | Abnormal | 0.32 predicted probability |
| Placebo | Chromosomal Complement of Embryo Per Cycle | Normal | 0.68 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Chromosomal complement will be compared using generalized estimating equations accounting for multiple cycles per couple.95% CI: [0.74, 7.43]
Secondary
Clinical Intrauterine Pregnancy
Visualized gestational sac in the uterus on ultrasound
Time frame: approximately 6.5 weeks gestation
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Clinical Intrauterine Pregnancy | 449 Participants |
| Placebo | Clinical Intrauterine Pregnancy | 462 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Clinical intrauterine pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [-4.9, 2.8]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Clinical intrauterine pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.89, 1.08]
Secondary
Early Pregnancy Loss
hCG pregnancy loss will be defined as a serum hCG \> 5 mIU/ml followed by a decline. Clinically recognized pregnancy losses will be defined as visualization of an intrauterine gestational sac followed by a loss prior to 20 weeks gestation.
Time frame: hcG-detected pregnancy until 20 weeks of pregnancy
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Early Pregnancy Loss | 137 Participants |
| Placebo | Early Pregnancy Loss | 150 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Early pregnancy loss will be compared using two sided tests conducted at the 0.05 level.95% CI: [-3.7, 1.5]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Early pregnancy loss will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.75, 1.15]
Secondary
Ectopic Pregnancy
Either visualization of no gestational sac in the uterus with a suspicious mass in the adnexa on ultrasound, an hCG level more than 1500 mIU/ml without visualization of an intrauterine gestational sac on ultrasound, or a slowly rising or plateauing serum hCG level without visualization of an intrauterine gestation on ultrasound.
Time frame: approximately 6.5 weeks gestation
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Ectopic Pregnancy | 6 Participants |
| Placebo | Ectopic Pregnancy | 5 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Ectopic pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.5, 0.6]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Ectopic pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.37, 3.97]
Secondary
Embryo Morphology on Day 3 Per Cycle, Categorical
Among participants in the IVF stratum Embryos will be scored three days after fertilization according to the size and shape of blastomeres and to their degree of fragmentation. Veeck LL. Oocyte assessment and biological performance. Ann N Y Acad Sci 1988;541:259-74.:259-74.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 961 embryos among cycles among women in the FA/Zn arm; 931 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively Embryo morphology on day 3 (categorical) unavailable in medical records for 27 and 37 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Embryo Morphology on Day 3 Per Cycle, Categorical | Excellent or good | 0.66 predicted probability |
| Folic Acid and Zinc Supplementation | Embryo Morphology on Day 3 Per Cycle, Categorical | Fair or poor | 0.34 predicted probability |
| Placebo | Embryo Morphology on Day 3 Per Cycle, Categorical | Excellent or good | 0.68 predicted probability |
| Placebo | Embryo Morphology on Day 3 Per Cycle, Categorical | Fair or poor | 0.32 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Embryo morphology on day 3 will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.87, 1.09]
Secondary
Embryo Morphology on Day 5 Per Cycle, Categorical
Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 973 embryos among cycles among women in the FA/Zn arm; 1009 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively Embryo morphology on day 5 (categorical) unavailable in medical records for 10 and 19 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Embryo Morphology on Day 5 Per Cycle, Categorical | Excellent or good | 0.28 predicted probability |
| Folic Acid and Zinc Supplementation | Embryo Morphology on Day 5 Per Cycle, Categorical | Fair or poor | 0.72 predicted probability |
| Placebo | Embryo Morphology on Day 5 Per Cycle, Categorical | Excellent or good | 0.35 predicted probability |
| Placebo | Embryo Morphology on Day 5 Per Cycle, Categorical | Fair or poor | 0.65 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Embryo morphology on day 5 will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.63, 1.06]
Secondary
Fertilization Rate Per Cycle, %
Among participants in the IVF stratum Oocytes will be assessed 16-18 hours after insemination or microinjection to determine whether fertilization occurred. Fertilization will be considered normal if two pronuclei and two polar bodies are identified. Oocytes without visible pronuclei will be considered unfertilized. Oocytes with more than two pronuclei will be considered abnormally fertilized, and will thus be discarded.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Fertilization rate unavailable in medical records for 17 and 28 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Fertilization Rate Per Cycle, % | 75.3 percent per cycle | Standard Deviation 2.25 |
| Placebo | Fertilization Rate Per Cycle, % | 77.7 percent per cycle | Standard Deviation 1.74 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Fertilization rate will be compared using generalized estimating equations accounting for multiple cycles per couple.95% CI: [-7.9, 3.23]
Secondary
Gestational Age
Abstracted from hospital records and medical charts
Time frame: Delivery
Population: Among participants with live birth
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Gestational Age | 38.6 weeks | Standard Deviation 2.5 |
| Placebo | Gestational Age | 38.8 weeks | Standard Deviation 2.2 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Gestational age will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.5, 0.1]
Secondary
Gestational Diabetes
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Gestational Diabetes | 26 Participants |
| Placebo | Gestational Diabetes | 34 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Gestational diabetes will be compared using two sided tests conducted at the 0.05 level.95% CI: [-1.9, 0.6]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Gestational diabetes will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.47, 1.27]
Secondary
Human Chorionic Gonadotropin (hCG) Detected Pregnancy (Implantation)
A quantitative hCG evaluation in serum \> 5 milli-international units per milliliter (mIU/ml)
Time frame: For IVF, 12 days post embryo transfer for day 5 embryo transfers, and 14 days post embryo transfer for day 3 embryo transfers; for couples undergoing OI/IUI, after self-report of positive pregnancy test
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Human Chorionic Gonadotropin (hCG) Detected Pregnancy (Implantation) | 479 Participants |
| Placebo | Human Chorionic Gonadotropin (hCG) Detected Pregnancy (Implantation) | 490 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. hCG detected pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [-4.7, 3]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. hCG detected pregnancy will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.91, 1.09]
Secondary
Major Neonatal Complications
Abstracted from hospital records and medical charts: includes bronchopulmonary dysplasia, necrotizing enterocolitis, severe intraventricular hemorrhage, periventricular leukomalacia, and retinopathy of prematurity
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Major Neonatal Complications | 2 Participants |
| Placebo | Major Neonatal Complications | 1 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Major neonatal complications will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.2, 0.4]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Major neonatal complications will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.18, 21.9]
Secondary
Method of Fertilization Per Cycle
Among participants in the in vitro fertilization (IVF) stratum: method of fertilization classified into intracytoplasmic sperm injection (ICSI) and other
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 1689 embryos among cycles among women in the active treatment arm; 1800 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively Method of fertilization unavailable in medical records for 2 and 5 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Method of Fertilization Per Cycle | ICSI | 0.74 predicted probability |
| Folic Acid and Zinc Supplementation | Method of Fertilization Per Cycle | Other | 0.26 predicted probability |
| Placebo | Method of Fertilization Per Cycle | ICSI | 0.79 predicted probability |
| Placebo | Method of Fertilization Per Cycle | Other | 0.21 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Method of fertilization will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.8, 1.09]
Secondary
Neonatal Mortality
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Neonatal Mortality | 3 Participants |
| Placebo | Neonatal Mortality | 2 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Neonatal mortality will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.3, 0.5]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Neonatal mortality will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.25, 8.95]
Secondary
Number of Embryos Cryopreserved Per Cycle
Among participants in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Number of embryos cryopreserved unavailable in medical records for 40 and 35 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Number of Embryos Cryopreserved Per Cycle | 4.22 embryos per cycle | Standard Deviation 0.32 |
| Placebo | Number of Embryos Cryopreserved Per Cycle | 4.32 embryos per cycle | Standard Deviation 0.31 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of embryos cryopreserved will be compared using Poisson regression with generalized estimating equations accounting for multiple cycles per couple.95% CI: [-0.23, 0.18]
Secondary
Number of Embryos Transferred Per Cycle
Among participants in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Proportion day 5 good quality embryos unavailable in medical records for 7 and 9 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Number of Embryos Transferred Per Cycle | 1.50 embryos per cycle | Standard Deviation 0.06 |
| Placebo | Number of Embryos Transferred Per Cycle | 1.51 embryos per cycle | Standard Deviation 0.05 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of embryos transferred will be compared using Poisson regression with generalized estimating equations accounting for multiple cycles per couple.95% CI: [-0.11, 0.1]
Secondary
Number of Good Quality Embryos on Day 5 Per Cycle
Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Day 5 good quality embryos unavailable in medical records for 56 and 64 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Number of Good Quality Embryos on Day 5 Per Cycle | 2.66 embryos per cycle | Standard Deviation 0.23 |
| Placebo | Number of Good Quality Embryos on Day 5 Per Cycle | 2.98 embryos per cycle | Standard Deviation 0.21 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Number of good quality embryos will be compared using Poisson regression with generalized estimating equations accounting for multiple cycles per couple.95% CI: [-0.33, 0.11]
Secondary
Percentage of Good Quality Embryos on Day 5 Per Cycle
Among participants in the IVF stratum For couples who meet criteria for blastocyst culture, embryos will be graded 5 days after fertilization based on Society for Assisted Reproductive Technologies (SART) morphology criteria.
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Proportion day 5 good quality embryos unavailable in medical records for 17 and 28 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Percentage of Good Quality Embryos on Day 5 Per Cycle | 17.2 percent per cycle | Standard Deviation 2.05 |
| Placebo | Percentage of Good Quality Embryos on Day 5 Per Cycle | 18.5 percent per cycle | Standard Deviation 1.81 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Proportion of good quality embryos on day 5 will be compared using generalized estimating equations accounting for multiple cycles per couple.95% CI: [-6.66, 4.05]
Secondary
Preeclampsia or Gestational Hypertension
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Preeclampsia or Gestational Hypertension | 47 Participants |
| Placebo | Preeclampsia or Gestational Hypertension | 51 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Preeclampsia/gestational hypertension will be compared using two sided tests conducted at the 0.05 level.95% CI: [-1.9, 1.3]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Preeclampsia/gestational hypertension will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.63, 1.36]
Secondary
Preterm Delivery
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Preterm Delivery | 67 Participants |
| Placebo | Preterm Delivery | 45 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Preterm delivery will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.2, 3.6]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Preterm delivery will be compared using two sided tests conducted at the 0.05 level.95% CI: [1.04, 2.16]
Secondary
Quality of Embryos Transferred Per Cycle, Categorical
Among participants in the IVF stratum Embryonic grading based on Society for Assisted Reproductive Technologies (SART) morphology criteria
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 172 embryos among cycles among women in the FA/Zn arm; 187 embryos among cycles among women in the placebo arm 124 and 135 participants randomized to active and placebo arms, respectively Quality of embryos transferred (categorical) unavailable in medical records for 21 and 30 participants in the active and placebo arms, respectively
| Arm | Measure | Group | Value (NUMBER) |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Quality of Embryos Transferred Per Cycle, Categorical | Excellent or good | 0.75 predicted probability |
| Folic Acid and Zinc Supplementation | Quality of Embryos Transferred Per Cycle, Categorical | Fair or poor | 0.25 predicted probability |
| Placebo | Quality of Embryos Transferred Per Cycle, Categorical | Excellent or good | 0.73 predicted probability |
| Placebo | Quality of Embryos Transferred Per Cycle, Categorical | Fair or poor | 0.27 predicted probability |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Quality of embryos transferred will be compared using generalized estimating equations accounting for multiple embryos per cycle and multiple cycles per couple.95% CI: [0.88, 1.21]
Secondary
Severe Maternal Morbidity
Abstracted from delivery record: including postpartum hemorrhage, anemia requiring transfusion, sepsis, seizure, HELLP syndrome or preeclampsia with pulmonary edema
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Severe Maternal Morbidity | 15 Participants |
| Placebo | Severe Maternal Morbidity | 10 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Severe maternal morbidity will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.4, 1.3]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Severe maternal morbidity will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.68, 3.32]
Secondary
Small for Gestational Age
Abstracted from hospital records and medical charts
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Small for Gestational Age | 62 Participants |
| Placebo | Small for Gestational Age | 59 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Small for gestational age will be compared using two sided tests conducted at the 0.05 level.95% CI: [-1.5, 2]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Small for gestational age will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.75, 1.49]
Secondary
Sperm Penetration Per Cycle, %
Among participants in the IVF stratum
Time frame: Up to 9 months of fertility treatment post-randomization
Population: 124 and 135 participants randomized to active and placebo arms, respectively Sperm penetration \& unavailable in medical records for 114 and 121 participants in the active and placebo arms, respectively
| Arm | Measure | Value (MEAN) | Dispersion |
|---|---|---|---|
| Folic Acid and Zinc Supplementation | Sperm Penetration Per Cycle, % | 62.7 percent penetration | Standard Deviation 13 |
| Placebo | Sperm Penetration Per Cycle, % | 74.8 percent penetration | Standard Deviation 11 |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Sperm penetration percentage will be compared using generalized estimating equations accounting for multiple cycles per couple.95% CI: [-45.4, 21.2]
Secondary
Stillbirth
Loss at or after 20 weeks gestation. Determined based on hospital records and medical chart abstraction.
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Stillbirth | 1 Participants |
| Placebo | Stillbirth | 4 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Stillbirth will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.6, 0.1]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Stillbirth will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.03, 2.24]
Secondary
Structural Malformations
Abstracted from birth record: includes major (n = 21; 6 with known genetic cause), minor (n = 6), and unclassified (n = 2) defects Structural birth defects: includes hydronephrosis/ureteropelvic junction obstruction, transposition of the great arteries, renal agenesis, cleft lip, club feet, multicystic/dysplastic kidney, tetralogy of fallot, gastroschisis, atrioventricular septal defects, other oral-facial defects, other cardiovascular defects, other CNS defects, other eye defects, other oral-facial defects, other anomalies, other syndromes
Time frame: Delivery
| Arm | Measure | Value (COUNT_OF_PARTICIPANTS) |
|---|---|---|
| Folic Acid and Zinc Supplementation | Structural Malformations | 15 Participants |
| Placebo | Structural Malformations | 14 Participants |
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Structural malformations will be compared using two sided tests conducted at the 0.05 level.95% CI: [-0.8, 1]
Comparison: The primary analysis plan is based on an intention-to-treat (ITT) approach comparing the two treatment arms based on the randomized assignment. The common null hypothesis states that the effect of folic acid/zinc on the outcomes is null compared to the placebo. Structural malformations will be compared using two sided tests conducted at the 0.05 level.95% CI: [0.53, 2.21]
Other Pre-specified
Reproductive Hormones and Other Measured Biomarkers
Urinary, serum, and salivary concentrations of reproductive hormones, particularly androgens, proteomic analysis of human sperm and cardiometabolic risk factors and markers of oxidative stress, as well as measures of trace elements in toenails (collected at month 4 clinic visit). Biospecimens have been collected but laboratory analysis still needs to be done to be able to evaluate these endpoints.
Time frame: 4 or 6 months