Study procedures
ADVERSE EVENTS
The patients randomized for the treatment arm also received instructions on how to assume the supplement and how to report on adverse events. They assumed one tablet per day of a supplement containing essential or conditionally essential micronutrients in support to the OCM including: betaine 200 mg, l-cystine 200 mg, chelated zinc 10 mg, niacin (vit. B3) 16 mg, pyridoxine (vit. B6) 1.4 mg, riboflavin (vit. B2) 1.4 mg, 5-methyl-tetrahydrofolate (5MTHF) 400 μg, and methylcobalamin (CHAll patients underwent COH according to the standard practices of the study site for the same patients without any study-intended adjustments. Briefly, patients received 150 IU of rFSH daily until day 5; thereafter, the FSH dose was adjusted according to the patient’s response. When the leading group of follicles reached 13 mm, a GnRH antagonist was commenced to inhibit premature LH surge. Once at least 3 follicles reached 18 mm diameter, a bolus of rhCG was injected to induce final maturation. The oocyte pick-up was performed 35–36 h post hCG injection by US-guided transvaginal needle aspiration. The fluid contained in each punctured follicle was collected immediately after aspiration and placed on ice until centrifugation to eliminate any cells and frozen at − 20. Mature oocytes were injected with the partner sperm and fertilization was confirmed by the appearance of two pronuclei. The zygotes were cultured up to blastocyst stage. The best embryo based on morphological score was selected for the fresh embryo transfer. All patients received luteal support with micronized progesterone, 600 mg per day, vaginal route.The follicular fluid samples corresponding to the oocytes that generated the blastocysts selected for transfer were thawed and analyzed for Hcy content. The concentration of Hcy in follicular fluids was tested by a commercially available ELISA kit (Sunred Bio, China) with a sensitivity range of 0 to 155 μmol/L, an intra-assay variability < 10%, and inter-assay variability < 12%. The follicular fluid samples of concern were thawed and tested at a single laboratory all together after the completion of the follow-up of the last enrolled patient.