The impact of zinc and folic acid supplementation on sperm DNA methylation: results from the folic acid and zinc supplementation randomized clinical trial (FAZST)

A randomized, placebo-controlled trial was conducted to assess the impact of folic acid and zinc supplementation on sperm DNA methylation signatures. No significant alterations to DNA methylation patterns were identified.

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VOLUME 117, ISSUE 1, P75-85


Tim Jenkins, Ph.D., Kenneth Aston, Ph.D., Douglas Carrell, Ph.D., Elizabeth DeVilbiss, Ph.D., Lindsey Sjaarda, Ph.D., Neil Perkins, Ph.D., James L. Mills, M.D., Zhen Chen, Ph.D., Amy Sparks, Ph.D., Traci Clemons, Ph.D. Kayla Chaney, B.A., C. Matthew Peterson, M.D., Benjamin Emery, M.Phil., Jim Hotaling, M.D., Erica Johnstone, M.D., Enrique Schisterman, Ph.D., Sunni L. Mumford, Ph.D. 



To determine if 6-month folic acid (5 mg) and zinc (30 mg) supplementation impacts sperm DNA methylation patterns.


A multicenter, double-blind, block randomized, placebo-controlled trial titled “The Folic Acid and Zinc Supplementation Trial (FAZST).”


Infertility care centers.


Male partners (18 years and older) from heterosexual couples (female partners aged 18–45 years) seeking fertility treatment were recruited.


Men were randomized 1:1 to receive folic acid (5 mg) and elemental zinc (30 mg) (n = 713) or a matching placebo (n = 757) daily for 6 months.

Main Outcome Measure(s)

Sperm DNA methylation was analyzed using the EPIC methylation array (Illumina) at 6 months. Differential sperm DNA methylation was assessed at multiple levels (regional, single cytosine phosphate guanine, etc.). We additionally assessed the impact of supplementation on epigenetic age.


No significant differences were identified between the treatment and placebo groups although some trends appeared to be present. To determine if these trends were noteworthy, we implemented various permutations and found that the patterns we identified were no more than would be expected by random chance.


The data presented here strongly suggest that this supplementation regimen is not effective at altering sperm DNA methylation. These data comport well with previous findings from the FAZST study that found no impact of supplementation on basic semen analysis parameters or live birth.

Clinical Trial Registration Number Identifier: NCT01857310.

Fertility and Sterility

Editorial Office, American Society for Reproductive Medicine

Fertility and Sterility® is an international journal for obstetricians, gynecologists, reproductive endocrinologists, urologists, basic scientists and others who treat and investigate problems of infertility and human reproductive disorders.