Advertisement

A systematic review of the effect of oral antioxidants on male infertility

  • C. Ross
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK
    Search for articles by this author
  • A. Morriss
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK
    Search for articles by this author
  • M. Khairy
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK
    Search for articles by this author
  • Y. Khalaf
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK
    Search for articles by this author
  • P. Braude
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK

    Division of Reproduction and Endocrinology, King’s College London, London, UK
    Search for articles by this author
  • A. Coomarasamy
    Affiliations
    School of Clinical and Experimental Medicine, University of Birmingham, Birmingham, UK
    Search for articles by this author
  • T. El-Toukhy
    Correspondence
    Corresponding author.
    Affiliations
    Assisted Conception Unit, Guy’s and St. Thomas’ Hospital NHS Foundation Trust, 11th Floor Tower Wing, Guy’s Hospital, St. Thomas Street, London SE1 9RT, UK
    Search for articles by this author
Published:April 09, 2010DOI:https://doi.org/10.1016/j.rbmo.2010.03.008

      Abstract

      The use of antioxidants in treatment of infertile men has been suggested, although the evidence base for this practice is unclear. A systematic review of randomized studies was conducted to evaluate the effects of oral antioxidants (vitamins C and E, zinc, selenium, folate, carnitine and carotenoids) on sperm quality and pregnancy rate in infertile men. MEDLINE, EMBASE, Cochrane Library and CINAHL were searched for relevant trials published from respective database inception dates to May 2009. Study selection, quality appraisal and data extraction were performed independently and in duplicate. Seventeen randomized trials, including a total of 1665 men, were identified, which differed in the populations studied and type, dosage and duration of antioxidants used. Only two-thirds of the studies (11/17) reported using allocation concealment and three studies (18%) used intention-to-treat analysis. Despite the methodological and clinical heterogeneity, 14 of the 17 (82%) trials showed an improvement in either sperm quality or pregnancy rate after antioxidant therapy. Ten trials examined pregnancy rate and six showed a significant improvement after antioxidant therapy. The use of oral antioxidants in infertile men could improve sperm quality and pregnancy rates. Adequately powered robust trials of individual and combinations of antioxidants are needed to guide clinical practice.

      Keywords

      Introduction

      Infertility affects about 15% of couples of reproductive age (
      • Sharlip I.D.
      • Jarow J.P.
      • Belker A.M.
      • et al.
      Best practice policies for male infertility.
      ) and impaired semen parameters are implicated in up to 50% of infertile couples (

      World Health Organization, 1987. WHO Laboratory Manual for the Examination of Human Semen and Semen-Cervical Mucus Interaction, second ed., Cambridge University Press, Cambridge.

      ). The commonest aetiology of male infertility is idiopathic oligoasthenoteratozoospermia (
      • Hirsch A.
      ABC of subfertility: male subfertility.
      ), for which a specific treatment remains elusive. Intra-cytoplasmic sperm injection (ICSI) enables this form of infertility to be circumvented mechanically (
      • Tarlatzis B.
      • Bili H.
      Intracytoplasmic sperm injection. Survey of world results.
      ) but does not tackle the fundamental reasons behind male infertility (
      • Georgiou I.
      • Syrrou M.
      • Pardalidis N.
      • et al.
      Genetic and epigenetic risks of intracytoplasmic sperm injection method.
      ,
      • Johnson M.D.
      Genetic risks of intracytoplasmic sperm injection in the treatment of male infertility: recommendations for genetic counselling and screening.
      ,
      • Pauer H.U.
      • Hinney B.
      • Michelmann H.W.
      • et al.
      Relevance of genetic counselling in couples prior to intracytoplasmic sperm injection.
      ,
      • Varghese A.C.
      • Goldberg E.
      • Agarwal A.
      Current and future perspectives on intracytoplasmic sperm injection: a critical commentary.
      ). As a result, there is growing interest in identifying reversible causes of male factor infertility.
      Considerable evidence points towards a significant role of oxidative stress in causing male infertility (
      • Agarwal A.
      • Sharma R.K.
      • Desai N.R.
      • et al.
      Role of oxidative stress in pathogenesis of varicocele and infertility.
      ,
      • Kefer J.C.
      • Agarwal A.
      • Sabanegh E.
      Role of antioxidants in the treatment of male infertility.
      ; Lewis and Agbaje, 2007;
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ). Reactive oxygen species (ROS) include hydroxyl radicals, superoxide anions and hydrogen peroxide (
      • Combelles C.
      • Gupta S.
      • Agrawal A.
      Could oxidative stress influence the in-vitro maturation of oocytes?.
      ,
      • Sharma R.K.
      • Agarwal A.
      Role of reactive oxygen species in male infertility.
      ,
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ), the principal sources of which are leukocytes and sperm cytoplasm (
      • Aitken R.J.
      • Harkiss D.
      • Knox W.
      • et al.
      A novel signal transduction cascade in capacitating human spermatozoa characterised by a redox-regulated, cAMP-mediated induction of tyrosine phosphorylation.
      ). Mature and morphologically normal spermatozoa produce relatively less ROS compared with immature teratozoospermic forms, which have a greater amount of cytoplasm (
      • Henkel R.
      • Kierspel E.
      • Stalf T.
      • et al.
      Effect of reactive oxygen species produced by spermatozoa and leukocytes on sperm functions in non-leukocytospermic patients.
      ). Spermatozoa and seminal plasma possess an abundance of antioxidant activity, both enzymatic (such as superoxide dismutase, glutathione peroxidase and catalase) and non-enzymatic (vitamins C and E, glutathione, carnitine and carotenoids) (
      • Alvares J.
      • Storey B.
      Evidence for increased lipid perioxidative damage and loss of superoxide dismutase activity as a mode of sublethal cryodamage to human sperm durino cryopreservation.
      ,
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ).
      Under normal conditions, the balance between the production of ROS and antioxidant activity is maintained. When ROS are present in excessive amounts either due to increased generation or impaired clearance, they can cause extensive sperm DNA structural damage (
      • Davies K.J.A.
      Protein damage and degradation by oxygen radicals 1 – general aspects.
      ,
      • Lopes S.
      • Jurisicova A.
      • Sun J.G.
      • et al.
      Reactive oxygen species: potential cause for DNA fragmentation in human spermatozoa.
      ), reduced sperm motility (
      • Jones R.
      • Mann T.
      • Sherins R.J.
      Peroxidative breakdown of phospholipids in human spermatozoa, spermicidal properties of fatty acid peroxides, and protective action of seminal plasma.
      ,
      • Kao S.H.
      • Chao H.T.
      • Chen H.-W.
      • et al.
      Increase in oxidative stress in human sperm with lower motility.
      ,
      • MacLeod J.
      The role of oxygen in the metabolism and motility of human spermatozoa.
      ) and defective sperm membrane integrity via lipid peroxidation (
      • Agarwal A.
      • Saleh R.A.
      • Bedaiwy M.A.
      Role of reactive oxygen species in the pathophysiology of human reproduction.
      ,
      • Aitken R.J.
      • Clarkson J.S.
      • Fishel S.
      Generation of reactive oxygen species, lipid peroxidation, and human sperm function.
      ), all of which are important mechanisms behind sperm dysfunction (
      • Davies K.J.A.
      Protein damage and degradation by oxygen radicals 1 – general aspects.
      ,
      • Kefer J.C.
      • Agarwal A.
      • Sabanegh E.
      Role of antioxidants in the treatment of male infertility.
      ,
      • Lopes S.
      • Jurisicova A.
      • Sun J.G.
      • et al.
      Reactive oxygen species: potential cause for DNA fragmentation in human spermatozoa.
      ,
      • Sharma R.K.
      • Agarwal A.
      Role of reactive oxygen species in male infertility.
      ,
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ). Furthermore, the antioxidant capacity of semen from infertile men is less effective than that from fertile men (
      • Fraga C.G.
      • Motchnik P.A.
      • Wyrobek A.J.
      • et al.
      Smoking and low antioxidant levels increase oxidative damage to sperm DNA.
      ,
      • Lewis S.
      • Sterling E.
      • Young I.
      • et al.
      Total anti-oxidant capacity of seminal plasma is different in fertile and infertile men.
      ,
      • Lewis S.
      • Boyle P.
      • McKinney M.
      • et al.
      Comparison of individual antioxidants of sperm and seminal plasma in fertile and infertile men.
      ,
      • Saleh R.A.
      • Agarwal A.
      • Kandirali E.
      • et al.
      Leukocytospermia is associated with increased reactive oxygen species production by human spermatozoa.
      ,
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ).
      Antioxidants (such as vitamins C and E, folate, zinc, selenium, carnitine and carotenoids) are scavengers of ROS and their use has been studied as a treatment to reverse the adverse impact of high ROS concentrations on semen parameters. Observational studies have shown that men with high dietary intake of antioxidants have a lower frequency of sperm aneuploidy and improved semen quality compared with men with lower intake (
      • Silver E.W.
      • Eskenazi B.
      • Evenson D.P.
      • et al.
      Effect of antioxidant intake on sperm chromatin stability in healthy nonsmoking men.
      ,
      • Young S.S.
      • Eskenazi B.
      • Marchetti F.M.
      • et al.
      The association of folate, zinc, and antioxidant intake with sperm aneuploidy in healthy non-smoking men.
      ).
      • Aitken R.J.
      • Clarkson J.S.
      • Fishel S.
      Generation of reactive oxygen species, lipid peroxidation, and human sperm function.
      demonstrated a dose-dependent reduction in the ability of human spermatozoa to fuse with oocytes with increased oxidative stress, which could be reversed by the inclusion of vitamin E. Likewise,
      • Hughes C.
      • Lewis S.
      • McKelvey-Martin V.
      • et al.
      The effects of antioxidant supplementation during Percoll preparation on human sperm DNA integrity.
      reported a beneficial effect on sperm DNA integrity during Percoll sperm preparation for assisted reproduction by the addition of antioxidants. However, most of these studies were uncontrolled, included fertile men and rarely used pregnancy as an outcome measure (
      • Kefer J.C.
      • Agarwal A.
      • Sabanegh E.
      Role of antioxidants in the treatment of male infertility.
      ,
      • Lewis S.
      • Agbaje I.
      Using the alkaline comet assay in prognostic tests for male infertility and assisted reproductive technology outcomes.
      ).
      This systematic review sought to rigorously evaluate current evidence from randomized trials on the effect of oral antioxidants on sperm parameters and the likelihood of spontaneous and treatment-related pregnancy in infertile men.

      Materials and methods

      Identification of oral antioxidants and their mode of action

      This study reviewed substances with recognized antioxidant properties. These were: (i) vitamin C (ascorbic acid), a water-soluble potent ROS scavenger that can also influence the expression of genes involved in the intracellular redox pathways (
      • Duarte T.
      • Lunex J.
      Review: when is an antioxidant not an antioxidant? A review of novel actions and reactions of vitamin C.
      ,
      • Gershoff S.N.
      Vitamin C (ascorbic acid): new roles, new requirements?.
      ); (ii) vitamin E (α-tocopherol), a lipid-soluble antioxidant whose primary role is to protect the integrity of the phospholipid bi-layer of the cell membrane and mitochondrial sheath by interrupting the chain reactions involved in lipid peroxidation and enhancing the production of scavenger antioxidant enzymes (
      • Fukuzawa K.
      • Takase S.
      • Tsukatani H.
      • et al.
      The effect of concentration on the antioxidant effectiveness of alpha-tocopherol in lipid peroxidation induced by superoxide free radicals.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ,
      • Traber M.
      • Atkinson J.
      Vitamin E, antioxidant and nothing more.
      ); (iii) folic acid (folate), which acts as an antioxidant by reducing homocysteine concentrations and via its free ROS scavenging properties (
      • Alvares Delfino V.D.
      • de Andrade Vianna A.C.
      • Mocelin A.J.
      • et al.
      Folic acid therapy reduces plasma homocysteine levels and improves plasma antioxidant capacity in hemodialysis patients.
      ,
      • Joshi R.
      • Adhikari S.
      • Patro B.S.
      • et al.
      Free radical scavenging behaviour of folic acid: evidence of possible anti-oxidant activity.
      ); (iv) zinc, an antioxidant trace element which has a membrane stabilizing activity by inhibiting membrane-bound oxidative enzymes such as NADP oxidase (
      • Powell S.R.
      The antioxidant properties of zinc.
      ,
      • Prasad A.
      Clinical, immunological, anti-inflammatory and antioxidant roles of zinc.
      ) and may also have an immunological function (
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ); (v) selenium, which plays a pivotal role in increasing glutathione peroxidase-1 expression and activity, which in turn destroys hydrogen peroxide molecules (
      • Rotruck J.T.
      • Pope A.L.
      • Ganther H.E.
      • et al.
      Selenium: biochemical role as a component of glutathione peroxidase.
      ,
      • Schnabel R.
      • Lubos E.
      • Messow C.M.
      • et al.
      Selenium supplementation improves antioxidant capacity in vitro and in vivo in patients with artery disease: The Selenium Therapy in Coronary Artery disease Patients (SETCAP) Study.
      ); (vi) carnitines, which are involved in transport of long chain fatty acids into the mitochondrial matrix for beta-oxidation and exert antioxidant activity via increasing expression of antioxidant enzymes such as haeme oxygenase-1 and endothelial nitric oxide synthetase (
      • Arduini A.
      Carnitine and its acyl esters as secondary antioxidants?.
      ,
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Calò L.A.
      • Pagnina E.
      • Davisb P.A.
      • et al.
      Antioxidant effect of l-carnitine and its short chain esters: relevance for the protection from oxidative stress related cardiovascular damage.
      ,
      • Jeulin C.
      • Lewin L.M.
      Role of free l-carnitine and acetyl l-carnitine in post-gonadal maturation of mammalian spermatozoa.
      ) and buffering acetyl-CoA pools (
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ); (vii) N-acetyl cysteine, an amino acid precursor of glutathione and that acts as an antioxidant by increasing the availability of intracellular glutathione and by acting directly as a free oxygen radical scavenger (
      • Circu M.L.
      • Moyer M.P.
      • Harrison L.
      • et al.
      Contribution of glutathione status to oxidant-induced mitochondrial DNA damage in colonic epithelial cells.
      ); and (viii) astaxanthin, a carotenoid extracted from algae Haematococcus pluvialis with a high number of conjugated double bonds rendering it a more potent antioxidant than either vitamin E or carnitine (
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ,
      • Rao A.V.
      • Agarwal S.
      Role of antioxidant lycopene in cancer and heart disease.
      ,

      Wolf, A., Asoh, S., Hiranuma, H. et al., 2009. Astaxanthin protects mitochondrial redox state and functional integrity against oxidative stress. J. Nutr. Biochem. – Epub ahead of print, May 2009.

      ).

      Literature search methodology

      A systematic review was conducted on randomized controlled studies evaluating the effects of the oral antioxidants (vitamins C and E, folate, zinc, selenium, carnitine, N-acetyl cysteine and carotenoids) on semen parameters and spontaneous or treatment-related pregnancy rate in infertile men. MEDLINE, EMBASE, Cochrane Library and CINAHL were searched for relevant trials published from respective database inception dates up to 31 May 2009 with no language restrictions.
      Two subsets of search terms were used, one describing the antioxidants (oral antioxidants, antioxidant therapy, folate, folic acid, selenium, zinc, vitamin, vitamin C, vitamin E, ascorbic acid, ascorbate, tocopherol, carnitine, cysteine, N-acetyl cysteine, beta carotene, carotenoid and trace elements) and the other describing spermatozoa, infertility and pregnancy (semen, sperm, spermatozoa, oligozoospermia, athenozoospermia, teratozoospermia, male subfertility, male infertility, in-vitro fertilization, intra-cytoplasmic sperm injection conception, pregnancy, embryo transfer and assisted reproduction techniques, which were combined in the search using the ‘OR’ operator). The two subsets of terms were then combined using the ‘AND’ operator.

      Study selection

      Studies were selected if they had a randomized design, the target population was infertile men and the therapeutic intervention was oral antioxidant(s) compared with placebo or no treatment. Outcome measures were basic semen parameters (sperm concentration, motility and morphology) and/or occurrence of pregnancy. Randomized studies with a cross-over design were excluded to avoid any possible residual effect from an inadequate wash out period (
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ). Studies involving the use of antimicrobial or hormonal medication were also excluded.
      The titles and abstracts retrieved from the electronic searches were scrutinised and full manuscripts of relevant citations that definitely or possibly met the search predefined selection criteria were obtained. Final study inclusion was made on examination of the full manuscripts. The reference lists of these studies and those of related reviews were further searched to ensure all relevant randomized studies were included. Study selection, quality appraisal and data extraction were performed independently and in duplicate by two authors (CR and AM) with a third author (TE) acting as an arbitrator to resolve uncertainty or disagreement about study inclusion.

      Data extraction and quality assessment

      The selected studies were assessed for methodological quality by using the components of study design that are related to internal validity (
      • Jadad A.R.
      • Moore R.A.
      • Carroll D.
      • et al.
      Assesssing the quality of reports of randomised clinical trials. Is blinding necessary?.
      ), including method of randomization, allocation concealment, double blinding, intention-to-treat analysis and follow up (or drop-out) rate.
      Study characteristics, participant features, study inclusion and exclusion criteria, nature of intervention (type and dose of antioxidant(s) used and duration of treatment) were extracted from each study. Exploration of clinical heterogeneity was conducted using variation in features of the population, intervention and study quality.

      Statistical analysis

      Data entry and analysis were performed using a Statview software package (Abacus Concepts, Berkeley, California, USA). Analysis of the study outcome measures and associated clinical variables was performed using a two-sample t-test (for continuous variables) and chi-squared test (for categorical variables). The odds ratio (OR) and 95% confidence interval (CI) were calculated for the spontaneous pregnancy outcome. A P-value <0.05 was considered statistically significant.

      Results

      The initial search identified 3740 citations, of which 3702 were excluded following screening of the titles and abstracts and 38 manuscripts were retrieved and reviewed in full, including one article published in Polish and translated into the English language (Figure 1). Of the 38 manuscripts examined in full, 17 original randomized controlled studies including 1665 men met the inclusion criteria (Figure 1). Sixteen studies (including 1605 randomized men) evaluated the effect of oral antioxidants on semen parameters and 10 studies (including 783 randomized men) reported the effect of oral antioxidants on pregnancy rate, including one study reporting the effect of oral antioxidants on pregnancy rate after assisted conception treatment (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ). All 17 studies were published in full in the English language.

      Methodological quality of included studies

      The methodological quality of the trials included in this review was variable (Table 1). Less than half (41%, 7/17) of the studies described a recognized method of randomization, and less than two-thirds (59%, 10/17) reported using allocation concealment during group assignment. The majority of studies were placebo-controlled (88%, 15/17) and had a double-blind design (82%, 14/17), but only three studies (18%) analysed their results according to the principles of intention-to-treat (ITT) analysis. The follow up rate varied from 37% to 100%, but was greater than 75% in 15 of the 17 studies (88%). Only two studies (12%) satisfied all five quality assessment criteria (
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ).
      Table 1Quality assessment of included trials.
      StudyRandomization methodAllocation concealmentBlindingITT analysisFollow-up rate (%)
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      Not clearNot clearDoubleNo79
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      Random no. tablesAdequateDoubleNo93
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      Not clearNot clearNoNo97
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      Random no. tablesAdequateDoubleNo94
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      Computer generatedAdequateDoubleNo92
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      Random no. tablesAdequateDoubleNo37
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      Not clearNot clearDoubleNo93
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      Not clearAdequateDoubleNo86
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      Not clearNot clearDoubleNot clear98
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      Not clearNot clearDoubleYes100
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      Not clearAdequateDoubleNot clear100
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      Not clearNot clearDoubleNot clear76
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      Computer generated tablesAdequateDoubleNot clear87
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      Computer generated tablesAdequateDoubleYes100
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      Not clearNot clearNot clearNot clearNot clear
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      Computer generated tablesAdequateDoubleNo89.7
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      Not clearAdequateSingleYes100
      ITT = intention to treat.

      Study characteristics

      Study population

      All studies were performed as single-centre studies, except that of
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      , which was conducted across two fertility centres. Nine of the 17 studies (53%) were performed in Europe (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ), five (29%) were performed in Asia (
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ), one (6%) in Africa (
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ), one in the USA (
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ) and one in Australia (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ).
      The number of men randomized per study varied widely and ranged between 26 and 468 (median 60), with 11of the 17 studies (65%) randomizing less than 100 men per study. Only four studies (24%,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ) described an a-priori power calculation for estimation of sample size.
      Three studies (18%) recruited men with unclassified infertility (
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      ;
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      );
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ;
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ), eight (47%) recruited infertile men with asthenozoospermia (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ), one (6%) recruited infertile men with oligozoospermia (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      );
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ) and two studies (12%) recruited infertile men with oligoasthenozoospermia (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ). Only two studies (
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ) recruited infertile men based on actual sperm DNA damage level as measured by the TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) assay using more than 15% and 25% sperm DNA fragmentation as their inclusion threshold, respectively (Table 2). For the study of
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      , the group comprised of fertile men was excluded from this review.
      Table 2Study characteristics in all included studies.
      StudyNo. of randomized participants and characteristicsIntervention (daily dose)Control groupDuration (weeks)
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      110; asthenozoospermiaVit E 300 mgPlacebo26
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      100; asthenozoospermiaZnSO4 500 mgNone13
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      69; asthenozoospermiaVit A 1 mg, vit C 10 mg, vit E 15 mg and selenium 100 μgPlacebo13
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      33; asthenozoospermiaVit C 1000 mg and vit E 800 mgPlacebo8
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      103; unclassified subfertilityFolic acid 5 mg and/or ZnSO4 66 mg and folic acid 5 mg/ZnSO4 66 mgPlacebo26
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      54; unclassified subfertilityVit E 400 mg and selenium 225 μgVit B13
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      60; asthenozoospermiaCarnitine 2 g and acetyl l-carnitine 1 gPlacebo26
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      219; oligoasthenozoospermia ± varicoceleCarnitine 2 g and acetyl l-carnitine 1 g/carnitine 2 g plus acetyl l-carnitine 1 gPlacebo26
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      60; asthenozoospermial-Carnitine 3 g or acetyl l-carnitine 3 g or l-carnitine 2 g and acetyl l-carnitine 1 gPlacebo26
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      30; unclassified subfertilityAstaxanthin 16 mgPlacebo13
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      64; >15% of sperm fragmented DNAVit C 1 g and vit E 1 gPlacebo9
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      26; asthenozoospermial-Carnitine 1 g and acetyl l-carnitine 500 mgPlacebo16
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      42; oligospermia 6/12 after varicocele embolisationVit A 0.06 IU/kg, vit C 3 mg/kg, vit E 0.2 mg/kg, N-acetyl cysteine 10 mg/kg, zinc 0.01 mg/kg and others
      Thiamine 0.4mg/kg/day, riboXavin 0.1mg/kg/day, piridoxin 0.2mg/kg/day, nicotinamide 1mg/kg/day, pantothenate 0.2mg/kg/day, biotin 0.04mg/kg/day, cyanocobalamin 0.1mg/kg/day, ergocalciferol 8IU/kg/day, calcium 1mg/kg/day, magnesium 0.35mg/kg/day, phosphate 0.45mg/kg/day, iron 0.2mg/kg/day, manganese 0.01mg/kg/day, copper 0.02mg/kg/day.
      Placebo13
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      60; poor morphology, motility or membrane integrity and >25% of spermatozoa showing significant DNA fragmentation
      >25% of spermatozoa were microscopic TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling assay positive.
      Menevit one capsule
      Menevit, a commercially available supplement, contains vitamin C 100mg, vitamin E 400IU, folate 500μg, garlic 1000mg, lycopene 6mg, selenium 26μg and zinc 25mg.vt=vitamin.
      Placebo13
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      45; asthenozoospermiaVit C 10 mg, vit E 20 mg with ZnSO4 400 mg, vit E 20 mg with ZnSO4 400 mg or ZnSO4 400 mgNone13
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      468; idiopathic oligoasthenoteratozoospermiaN-acetyl cysteine 600 mg/N-acetyl cysteine 600 mg and selenium 200 μg/selenium 200 μgPlacebo26
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      120; idiopathic infertilityN-acetyl cysteine 600 mgPlacebo13
      a Thiamine 0.4 mg/kg/day, riboXavin 0.1 mg/kg/day, piridoxin 0.2 mg/kg/day, nicotinamide 1 mg/kg/day, pantothenate 0.2 mg/kg/day, biotin 0.04 mg/kg/day, cyanocobalamin 0.1 mg/kg/day, ergocalciferol 8 IU/kg/day, calcium 1 mg/kg/day, magnesium 0.35 mg/kg/day, phosphate 0.45 mg/kg/day, iron 0.2 mg/kg/day, manganese 0.01 mg/kg/day, copper 0.02 mg/kg/day.
      b >25% of spermatozoa were microscopic TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling assay positive.
      c Menevit, a commercially available supplement, contains vitamin C 100 mg, vitamin E 400 IU, folate 500 μg, garlic 1000 mg, lycopene 6 mg, selenium 26 μg and zinc 25 mg.vt = vitamin.
      In the eight studies which recruited athenozoospermic men, the threshold for study inclusion also varied. Whereas asthenozoospermia was defined as sperm motility below 50% in five studies (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ), it was defined as the presence of 40% or more immotile spermatozoa in the studies of
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      and
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      and was undefined in the study of
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      .

      Study exclusion criteria

      Nine studies (53%) excluded infertile men with oligozoospermia, defined as sperm concentration of: less than 20 × 106 spermatozoa/ml in the studies of
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      and
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ; less than 10 × 106 spermatozoa/ml in the study of
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ; less than 7 × 106 spermatozoa/ml in the study of
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ; and less than 5 × 106 sperm/ml in the studies of
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      and
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      .
      Only eight studies (47%) excluded men with genital infection (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ), six studies (35%) excluded smokers (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ) and five studies (29%) excluded men with varicocele (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ).

      Intervention

      The studies varied considerably in the nature and dose of antioxidant(s) used and duration of treatment (Table 2). Eight studies (47%) used a single antioxidant, while nine studies (53%) used a combination of antioxidants. The antioxidants used were vitamins C and E, zinc, selenium, folic acid, carnitine, N-acetyl cysteine, astaxanthin (a strong natural antioxidant) and Menevit (a commercially available combination of vitamins E and C, zinc, selenium, folate, lycopene and garlic). The antioxidant studied most as a single agent was carnitine (four studies:
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ), while vitamin C is the only antioxidant that has not been studied as a single agent among the studies selected for this review. Treatment duration lasted on average 18 weeks and ranged from 8 to 26 weeks (median = 13 weeks).

      Outcomes

      All studies reported outcome in terms of semen variables, pregnancy outcome or both (Table 2, Table 3). Overall, 14 of the 17 studies (82%) showed improvement in at least one of these outcome measures in the treatment group. In two of the three studies in which no improvement occurred in the treatment group (
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ), the antioxidants were used for only 8 and 9 weeks, respectively.
      Table 3Effect of antioxidants on sperm parameters in the included studies.
      StudyAntioxidant usedStudy outcome measuresImprovements
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      Vitamin EMotility, level of lipid peroxidationMotility in asthenozoospermic men and reduction in level of lipid peroxidation
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ZnSO4Concentration, motility, membrane integrity, serum zinc, cadmium, antisperm antibodies, FSH, LH, prolactin, TNFα and IL-4 concentrationsCount, progressive motility, membrane integrity and reduction in antisperm antibodies
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      Vitamins A, C, E, seleniumConcentration, motility, serum seleniumMotility
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      Vitamins C and EConcentration, motility, morphology, volume, 24-h sperm survivalNo improvement
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      Folic acid and/or ZnSO4Concentration, motility, morphology, seminal plasma, blood plasma and erythrocyte folate and zincConcentration and total normal sperm count
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      Vitamin E and seleniumConcentration, motility, morphology, volume, viability, semen MDA, serum vitamin E and cholesterolMotility
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      CarnitineConcentration, motility, morphology, volumeNumber of total motile sperm and forward motile sperm
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      CarnitineConcentration, motility, morphology, testicular volumeConcentration, motility and morphology, except for grade IV and V varicocoeles, where there was no difference.
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      CarnitineConcentration, motility, morphology, volume, semen total oxyradical scavenging capacityStraight progressive velocity and total oxyradicalscavenging capacity
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      AstaxanthinConcentration, motility, morphology, volume, pregnancy rate, serum FSH, LH, testosterone, inhibin B, ROS generation,
      Measured by chemoluminescence in high quality spermatozoa after centrifugation.
      a, zona-free hamster oocyte test, seminal γ-glutamyltransferase, gluconidase.
      No improvement.
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      Vitamins C and EConcentration, motility, morphology, sperm DNA fragmentation
      Measured using the TUNEL test.
      DNA fragmentation.
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      CarnitineVolume, motility, concentration, forward progression, seminal plasma and semen carnitineNo improvement.
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      Vitamins A, C, E, N-acetyl cysteine and zincConcentration, motility, morphology, volumeOdds of having a normal sperm count.
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      Vitamins C and E with ZnSO4, vitamin E with ZnSO4 or ZnSO4 aloneConcentration, motility, morphology, volume, sperm chromatin integrity and apoptosis,
      Chromatin integrity determined by acid denaturation by acridine orange and apoptosis by light and electron microscopy.
      sperm membrane integrity, serum and seminal plasma antioxidants, cadmium, magnesium, selenium, zinc, cytokines, oxidants, antisperm antibodies antibody levels and Bcl2 and Bax expression
      Sperm motility, fertilizing capacity (in vivo) and DNA fragmentation index (in vitro).
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      N-acetyl cysteine,

      N-acetyl cysteine and/or selenium
      Concentration, motility, morphology, serum testosterone, inhibin B, LH, FSHCount, motility, morphology.
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      N-acetyl cysteineConcentration, motility, morphology, volume, viscosity, liquefaction time, oxidative statusMotility, volume, viscosity, liquefaction time, oxidative status (total antioxidant capacity, total peroxide and oxidative stress index).
      IL = interleukin; ROS = reactive oxygen species; TNF = tumour necrosis factor.
      a Measured by chemoluminescence in high quality spermatozoa after centrifugation.
      b Measured using the TUNEL test.
      c Chromatin integrity determined by acid denaturation by acridine orange and apoptosis by light and electron microscopy.
      In addition, seven studies used a measure of sperm oxidative stress to assess treatment effect. This was performed either directly by measuring ROS concentration (
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ) or indirectly by measuring the degree of sperm lipid peroxidation (via determining the malondialdehyde concentration), DNA fragmentation (using the TUNEL assay) or total antioxidant capacity (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ). All seven studies reported evidence of reduction in oxidative stress in the treatment group compared with controls.

      Effect of antioxidants on semen variables

      Of the 16 trials examining the effect of oral antioxidants on semen variables, 12 (75%) showed an improvement in at least one sperm parameter compared with placebo or no treatment (Table 3 and Figure 2).
      Figure thumbnail gr2
      Figure 2Effect of antioxidants on sperm parameters, pregnancy rate and oxidative stress. Numbers indicate the number of studies that showed an improvement or no improvement.
      Ten out of the 16 studies (63%) showed a significant improvement in sperm motility compared with placebo (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      ,
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ), five out of 15 studies (33%) showed an improvement in sperm concentration (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ) and two out of 12 studies (17%) showed an improvement in sperm morphology (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ).

      Effect of antioxidants on pregnancy rate

      Ten studies including 783 randomized men examined the effect of oral antioxidants on pregnancy rate and reported data on 745 men who completed the studies (Table 4). Nine of the 10 studies reported results for spontaneous conception (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ) and, overall, showed a significantly higher pregnancy rate in the treatment group compared with controls (19% (69/368) versus 3% (9/317), OR = 7.9, 95% CI 3.9–16.1, P < 0.0001). One study (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ) reported results after ICSI treatment and reported a significantly higher viable pregnancy rate per embryo transfer in the treatment group (39% (20/52) versus 16% (4/25), P = 0.046).
      Table 4Effect of antioxidants on spontaneous or treatment-related pregnancy rate in the included studies.
      StudyAntioxidantEffect of antioxidants on pregnancy rate
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      Vitamin E21% (11/52) versus 0% (0/35), P = 0.003
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      Live birth rate 17% (9/52) versus 0% (0/35), P = 0.009
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      Zinc sulphate22% (11/49) versus 4% (2/48), P < 0.03
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      Selenium, vitamins A, C and E11% (5/46) versus 0% (0/18), P = 0.15
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      Vitamins C and E0% (0/15) versus 0% (0/16), P = 1
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      Carnitine13% (4/30) versus 0% (0/26), P = 0.04
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      Acetyl l-carnitine and carnitine22% (22/101) versus 2% (2/118), P < 0.01
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      l-Carnitine and acetyl l-carnitine20% (9/44) versus 20% (3/15), P = 1
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      Astaxanthin55% (6/11) versus 11% (2/19), P = 0.028
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      Vitamins A, C and E, N-acetyl

      cysteine and zinc
      5% (1/20) versus 0% (0/22), P = 0.95
      Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      Pregnancy rates per embryo transfer.
      Menevit64% (23/36) versus 38% (6/16), P = 0.077
      Viable pregnancy rate at 13 weeks (38% (20/52) versus 16% (4/25), P = 0.046
      a Pregnancy rates are for spontaneous pregnancies that occurred during the study period.
      b Pregnancy rates per embryo transfer.
      Individually, six of the 10 studies reported a significantly higher pregnancy rate following antioxidant treatment compared with controls (Figure 2). In two of the four remaining studies (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ), there was a higher pregnancy rate in the treatment group but the difference did not reach statistical significance. Interestingly, four of the nine studies reporting data on spontaneous pregnancy rates during the study period failed to provide sufficient confirmation that female partners of the randomized men had been objectively investigated for female causes of infertility (
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ).
      As shown in Table 3, in five of the six studies in which there was a significantly higher pregnancy rate in the treatment group, sperm quality was also examined and four of these showed an improvement in at least one sperm parameter, with all four studies demonstrating improved motility (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ).

      Effect of individual antioxidants on semen variables and pregnancy rate

      In this review, the different antioxidants studied have been used either as a single therapy or in combination with other antioxidants. Overall, all the antioxidants encountered in this review, except vitamin C, have been studied individually and resulted in improvement in at least one semen variable in one or more studies. However, these results were not consistent and varied considerably between the studies (Table 2, Table 3, Table 4).

      Vitamin C

      Sole use: Vitamin C has not been evaluated as the sole antioxidant in any of the 17 studies included in this review and therefore its effectiveness as a single oral antioxidant agent in infertile men is unknown.
      Combined use: Vitamin C has been used in conjunction with other antioxidants in six studies (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ; and one arm of
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ). Sperm motility improved in two out of five studies (40%,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ), sperm concentration improved in one out of five studies (20%,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ) and sperm DNA fragmentation index was reduced in two out of two studies (
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ). However, sperm morphology did not improve in any of the five studies. Pregnancy rate after ICSI was significantly improved in the treatment group in one study (P = 0.046;
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ).

      Vitamin E

      Sole use: Vitamin E has been studied as the sole antioxidant supplement in one study (
      • Suleiman S.A.
      • Elamin Ali M.
      • Zaki Z.M.S.
      • et al.
      Lipid peroxidation and human sperm motility: protective role of vitamin E.
      ) at a daily dose of 300 mg for 26 weeks. This study reported improvement in both sperm motility and pregnancy rate.
      Combined use: Vitamin E has been used in conjunction with other antioxidants in seven studies (
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ). Sperm motility improved in three out of six studies (50%,
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ), sperm concentration improved in one out of six studies (17%,
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ) and sperm DNA fragmentation index in two out of two studies (
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ). However, sperm morphology did not improve in any of the five studies where this parameter was evaluated (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Greco E.
      • Iacobelli M.
      • Rienzi L.
      • et al.
      Reduction of the incidence of sperm DNA fragmentation by oral antioxidant treatment.
      ;
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ;
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Rolf A.
      • Cooper T.G.
      • Yeong C.H.
      • et al.
      Antioxidant treatment of patients with asthenozoospermia or moderate oligoasthenozoopermia with high-dose vitamin C and vitamin E: a randomized, placebo-controlled, double-blind study.
      ). Pregnancy rate was higher in only one out of four studies where this was considered (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ).

      Zinc

      Sole use: Zinc sulphate has been studied as a sole antioxidant in three studies (
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ). Motility improved in two studies (
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ), concentration improved in two studies (
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ), but morphology did not improve in any of the three studies. Pregnancy rate improved in the only study where this was examined (
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.
      ).
      Combined use: Zinc sulphate has been used in conjunction with other antioxidants in four studies (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ). Motility improved in one out of three studies (
      • Omu A.E.
      • Al-Azemi M.K.
      • Kehinde E.O.
      • et al.
      Indication of the mechanisms involved in improved sperm parameters by zinc therapy.
      ), concentration improved in two out of three studies (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ), but morphology did not improve in any of the three studies. Pregnancy rate improved in one out of two studies (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ).

      Selenium

      Sole use: Selenium has been used as the sole antioxidant in only one study (
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ), which reported an improvement in all three sperm variables; concentration, motility and morphology.
      Combined use: Selenium has been used in conjunction with other antioxidants in four studies (
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ;
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ,
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ). Sperm motility was improved in three out of three studies (
      • Keskes-Ammar L.
      • Feki-Chakroun N.
      • Rebai T.
      • et al.
      Sperm oxidative stress and the effect of an oral vitamin E and selenium supplement on semen quality in infertile men.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ,
      • Scott R.
      • MacPherson A.
      • Yates R.W.S.
      • et al.
      The effect of oral selenium supplementation on human sperm motility.
      ), concentration improved in one out of three studies (
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ), morphology improved in one out of two studies (
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ) and pregnancy rate improved in one out of two studies (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ).

      Folic acid

      Sole use: Folic acid has been studied as the sole antioxidant in one study (
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      , at a daily dose of 5 mg for 26 weeks). This study reported improvement in sperm concentration, but not in sperm motility or morphology.
      Combined use: Folic acid has been used in conjunction with other antioxidants in two studies (
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      ,
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      ). Only sperm concentration, but not motility or morphology, was improved in the study of
      • Wong W.Y.
      • Merkus H.M.W.M.
      • Thomas C.M.G.
      • et al.
      Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial.
      and pregnancy rate after assisted conception treatment was significantly improved in the study of
      • Tremellen K.
      • Miari G.
      • Froiland D.
      • et al.
      A randomized control trial of an antioxidant (Menevit) on pregnancy outcome during IVF-ICSI treatment.
      .

      Carnitine

      Sole use:l-Carnitine and acetyl l-carnitine have been used as sole antioxidants in four studies (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Sigman M.
      • Glass S.
      • Campagnone J.
      • et al.
      Carnitine for the treatment of idiopathic asthenospermia: a randomized, double-blind, placebo-controlled trial.
      ). Sperm motility improved in three out of four studies (
      • Balercia G.
      • Regoli F.
      • Armeni T.
      • et al.
      Placebo-controlled double-blind randomized trial on the use of l-carnitine, l-acetylcarnitine, or combined l-carnitine and l-acetylcarnitine in men with idiopathic asthenozoospermia.
      ,
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ), concentration improved in one out of four studies one (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ) and morphology in one out of three studies (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ). Pregnancy rate improved in two out of three studies (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ).

      Astaxanthin

      Astaxanthin has only been used in one study as the sole antioxidant in the study of
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      . Only the pregnancy rate was improved in this study, but none of the three main semen variables showed an improvement after treatment.

      N-acetyl cysteine

      Sole use:N-Acetyl cysteine has been used in two studies as a sole antioxidant:
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      (600 mg daily for 26 weeks) and
      • Ciftci H.
      • Verit A.
      • Savas M.
      • et al.
      Effects of N-acetylcysteine on semen parameters and oxidative/antioxidant status.
      (600 mg daily for 13 weeks). Sperm motility improved in both studies, but concentration and morphology improved only in the study of
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      .
      Combined use:N-acetyl cysteine has been used in combination with other antioxidants in two studies (
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      ,
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ). Sperm motility improved in one study (
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ), whereas concentration improved in both studies and morphology improved in only one study (
      • Safarinejad M.R.
      • Safarinejad S.
      Efficacy of selenium and/or N-acetyl-cysteine for improving semen parameters in infertile men: a double-blind, placebo controlled, randomized study.
      ). Pregnancy rate did not show an improvement in the study of
      • Galatioto G.P.
      • Gravina G.L.
      • Angelozzi G.
      • et al.
      May antioxidant therapy improve sperm parameters of men with persistent oligospermia after retrograde embolization for varicocele?.
      .

      Discussion

      High ROS level and oxidative stress have been implicated in the pathophysiology of male infertility (
      • Tremellen K.
      Oxidative stress and male infertility–a clinical perspective.
      ) and correlated with sperm DNA damage, impaired fertilization and embryo development, low rates of implantation and occurrence of miscarriage (
      • Agarwal A.
      • Sharma R.K.
      • Nallella K.P.
      • et al.
      Reactive oxygen species as an independent marker of male factor infertility.
      ,
      • Aitken R.
      • Baker M.
      Oxidative stress and male reproductive biology.
      ,
      • Aitken R.J.
      • Clarkson J.S.
      • Fishel S.
      Generation of reactive oxygen species, lipid peroxidation, and human sperm function.
      ,
      • Carrell D.T.
      • Liu L.
      • Peterson C.M.
      • et al.
      Sperm DNA fragmentation is increased in couples with unexplained recurrent pregnancy loss.
      ,
      • Lewis S.E.
      • Aitken R.J.
      DNA damage to spermatozoa has impacts on fertilisation and pregnancy.
      ,
      • Loft S.
      • Kold-Jensen T.
      • Hjollund N.H.
      • et al.
      Oxidative DNA damage in human sperm influences time to pregnancy.
      ,
      • Morris I.
      • Ilott S.
      • Dixon L.
      • et al.
      The spectrum of DNA damage in human sperm assessed by single cell gel electrophoresis (COMET assay) and its relationship to fertilization and embryo development.
      ,
      • Seli E.
      • Gardner D.
      • Schoolcraft W.
      • et al.
      Extent of nuclear DNA damage in ejaculated spermatozoa impacts on blastocyst development after in vitro fertilization.
      ). This systematic review of a large number of randomized trials shows that treatment of infertile men with oral antioxidants reduces seminal oxidative stress and could improve sperm motility, but has a less predictable impact on sperm concentration and morphology (Figure 2).
      These results concord with evidence from other randomized studies not included in this review. Two randomized studies examined the effect of oral antioxidants on sperm parameters in fertile men.
      • Dawson E.B.
      • Harris W.A.
      • Rankin W.E.
      • et al.
      treated 30 men with sperm agglutination of over 25% with vitamin C 200 mg or 1000 mg daily or placebo. The authors found that motility, viability and morphology significantly increased after 4 weeks of treatment compared with baseline and more so in the 1000 mg group than the 200 mg group, but not in the placebo group. The same group (
      • Dawson E.B.
      • Harris W.A.
      • Teter M.C.
      • et al.
      Effect of ascorbic acid supplementation on the sperm quality of smokers.
      ) supplemented 75 heavy smokers with normal reproductive function with vitamin C at a daily dose of either 200 mg or 1000 mg of vitamin C or placebo. The authors reported that the control group showed no improvement in sperm quality whereas the vitamin C supplemented groups had improved sperm agglutination, 24-h viability and morphology.
      This review also excluded two randomized studies with a cross-over design.
      • Kessopoulou E.
      • Powers H.J.
      • Sharma K.K.
      • et al.
      A double-blind randomized placebo crossover controlled trial using the antioxidant vitamin E to treat reactive oxygen species associated male infertility.
      studied the effect of daily administration of 600 mg of vitamin E versus placebo for 3 months followed by a 1-month washout period and then cross-over to the other treatment on 30 infertile men with high levels of ROS generation. The authors reported a significant improvement in the zona-binding test for both orders of treatment, but that a significant carry-over effect was only evident in the group receiving vitamin E first.
      Likewise,
      • Lenzi A.
      • Lombardo F.
      • Sgro P.
      • et al.
      Use of carnitine therapy in selected cases of male factor infertility: a double-blind crossover trial.
      treated 100 infertile men with l-carnitine 2 g/day or placebo for 2 months followed by a 2-month washout period and then 2 months of the other treatment. They found that total motile spermatozoa and forward motile spermatozoa were significantly improved following carnitine treatment and that all the eight pregnancies which occurred during the trial were in the carnitine therapy period. Furthermore, the greatest increases in the number of motile spermatozoa were found in those subjects with the lowest baseline number of motile spermatozoa/ml of ejaculate.
      More relevant clinically, this review shows that oral antioxidant therapy was associated with a significant improvement in spontaneous and assisted conception pregnancy rates in six of the 10 randomized studies identified in the database search (
      • Cavallini G.
      • Ferraretti A.P.
      • Gianarolli L.
      • et al.
      Cinnoxicam and l-carnitine/acetyl-l-carnitine treatment for idiopathic and varicocoele-associated oligoasthenospermia.
      ,
      • Comhaire F.H.
      • El Garem Y.
      • Mahmoud A.
      • et al.
      Combined conventional/ antioxidant ‘Astaxanthin’ treatment for male infertility: a double blind, randomized trial.
      ,
      • Lenzi A.
      • Sgro P.
      • Salacone P.
      • et al.
      A placebo-controlled double-blind randomized trial in the use of combined l-carnitine and l-acetyl-carnitine treatment in men with asthenozoospermia.
      ,
      • Omu A.E.
      • Dahti H.
      • Al-Othman S.
      Treatment of asthenozoospermia with zinc sulphate: andrological, immunological and obstetric outcome.