1.1. Effect of AOX on Sperm Parameters and Conception Outcomes

Previous systematic reviews and meta-analyses (SRMAs) demonstrated that elevated seminal ROS are associated with sperm dysfunction, SDF, and reduced male reproductive potential, leading to impaired sperm parameters [13, 14].

Recently, an improvement in sperm parameters and spontaneous pregnancy rate has been reported after the administration of AOX therapy to infertile men in a meta-analysis performed on 45 randomized controlled trials (RCTs), including 4332 patients treated with AOX or no treatment/placebo [10]. The authors found that AOX therapy significantly increased the rate of spontaneous pregnancy compared to controls (odd ratio [OR] 1.97, 95% confidence interval [CI] 1.28, 3.04; p  < 0.01) with an absence of significant inter-study heterogeneity (I² = 20%; p = 0.20). Even sperm parameters, including sperm concentration, motility, and normal sperm morphology, improved significantly after AOX therapy, compared to placebo or no treatment. However, this meta-analysis found no significant effect of AOX therapy on live birth or miscarriage rates. Importantly, this study demonstrated that the positive impact of AOX therapy on spontaneous pregnancy and sperm parameters occurred regardless of the presence of varicocele and/or varicocele repair. Building on previous SRMAs, this article is the first in the literature to demonstrate that AOX treatment significantly improves seminal total antioxidant capacity (TAC) and reduces malondialdehyde (MDA) levels compared to controls.

Noegroho et al. [15] conducted a SRMA to evaluate the effect of AOX supplementation on SDF and conventional sperm parameters, including nine studies—seven RCTs and two observational studies. A significant reduction in SDF was reported in four studies, while five found no effect following AOX treatment. However, sperm count, motility, and normal morphology showed significant improvements post-treatment. Among the three studies that assessed pregnancy outcomes, two reported increased pregnancy rates, whereas one found no significant difference.

Zafar et al. [16] recently published a systematic review and network meta-analysis evaluating the effectiveness of nutritional therapies in the treatment of male factor infertility. The analysis included 69 studies encompassing 94 individual study arms, all of which involved placebo or control groups and reported outcomes related to nutritional or dietary supplements, at a minimum, on sperm parameters. Nutritional therapies, particularly L-carnitine alone or in combination with select micronutrients, demonstrated statistically and clinically significant improvements in sperm parameters, along with a notable increase in pregnancy rates.

In subgroup analyses, the authors found that pregnancy rates increased by 8.17 times with vitamin E, 5.88 times with herbal supplements, 4.84 times with AOXs, 3.55 times with L-carnitine plus micronutrients, 4.38 times with selenium, 3.35 times with vitamin D plus calcium, and 3.38 times with folic acid. Among 12 studies that assessed the impact of supplements on SDF, only vitamin and mineral combinations produced a significant reduction. However, this meta-analysis did not evaluate the effects of nutritional therapies on spontaneous or assisted pregnancy outcomes (e.g., in vitro fertilization [IVF]), live birth rates, or IVF-specific results.

In a recent Cochrane systematic review, de Ligny et al. [12 ] evaluated the effectiveness and safety of 20 different oral AOXs for treating male subfertility, analyzing data from 90 RCTs, of which 65 (including 10,303 subfertile men) were meta-analyzed. The findings showed that AOX significantly increased the live birth rate (OR 1.43) and clinical pregnancy rate (OR 1.89) compared to placebo or no treatment. There was no significant difference in miscarriage rates between groups. However, AOX use was associated with a higher risk of mild gastrointestinal discomfort (e.g., nausea or stomachache), with an OR of 2.7 (95% CI: 1.46–4.99).

Notably, 12 of the included studies were small to medium-sized RCTs, contributing to low-certainty evidence. Despite this, the review suggests that AOX supplementation may offer a modest benefit in improving clinical pregnancy and live birth rates among subfertile men attending fertility clinics.

Li et al. [17] included 23 RCTs with 1917 patients receiving 10 types of AOX. They found that L-carnitine, L-carnitine plus L-acetyl carnitine, coenzyme-Q10 (CoQ10), ω-3 fatty acid, and selenium significantly increased sperm quality compared to placebo. However, among the AOXs, L-carnitine provided the highest improvement in sperm motility and sperm morphology, and ω-3 fatty acid provided the highest improvement in sperm concentration. However, none of those AOX significantly increased the pregnancy rate compared with a placebo.

Sharma et al. [18] included 20 RCTs to compare carnitine, coQ10, or selenium or a combination with placebo in men with idiopathic infertility. They found no significant difference in pregnancy and live birth rates, although sperm parameters significantly improved using one of these AOXs.

Su et al. [19] included 18 RCTs with 1790 sub-fertile men receiving eight different types of AOX (folic acid, zinc, vitamin E, carnitine, selenium, CoQ10, N-acetylcysteine [NAC], and vitamin C) or placebo. They found that the most significant improvement in sperm concentration was achieved with CoQ10, in sperm motility with carnitine, and in sperm morphology with vitamin C, compared to the placebo. However, the authors did not include pregnancy outcomes in their study.

Although many SRMAs have reported significant improvements in sperm parameters and spontaneous pregnancy rates following AOX therapy in infertile men, results from a multicenter, double-blind, randomized, and placebo-controlled trial offered a different perspective. In this study, infertile men received a daily AOX formulation (vitamin C, vitamin E, selenium, L-carnitine, zinc, folic acid, and lycopene) or placebo for 3–6 months. After 3 months of treatment, no significant differences were observed in sperm parameters or DNA fragmentation index (DFI). Furthermore, pregnancy and live birth rates did not differ between the groups after 6 months [11].

However, the study has been criticized for several methodological limitations that may have biased the results. These include reliance on a single baseline semen analysis, a short follow-up period, and the inclusion of men with average sperm parameters in more than half of the treatment group [20]. Additionally, all female partners underwent clomiphene citrate treatment with intrauterine insemination (IUI), potentially confounding pregnancy outcomes. The trial has also been considered underpowered to adequately assess the true efficacy of AOX therapy on pregnancy and live birth outcomes [20, 21].

The molecular mechanisms through which AOX exerts their effects still need further elucidation. Along with their study, Agarwal et al. [22] evaluated the sperm proteome of idiopathic infertile men pre- and post-AOX supplementation at 6 months. They found that AOX therapy might improve sperm function at the molecular level by modulating proteins involved in CREM signaling, mitochondrial function, and protein oxidation in men with idiopathic infertility [22]. Further, AOX-dependent activation of the TRiC complex was reported, triggering nuclear compaction, telomere length maintenance, flagella function, and expression of zona pellucida receptors for sperm–oocyte interaction.

One study has suggested that increased ROS may reduce male sex hormone levels and disrupt the hormonal balance, leading to impairment in spermatogenesis [23]. Hormonal alterations to induce spermatogenesis are another advantage of AOX supplements. Specific nutritional supplements improve spermatogenesis by increasing serum total testosterone levels. In a meta-analysis, AOX, consisting of coenzymes, L-carnitine plus micronutrients, L-carnitine/carnitine/L-acetylcarnitine, and minerals, significantly increased serum total testosterone level, compared to placebo or no treatment [16]. ROS may affect endocrine pathways and disrupt normal hormonal secretion and reproductive functions. Therefore, ROS decreases the amount of circulating testosterone. At the same time, during OS, testicular inhibition and estradiol secretion increase and inhibit testosterone release. Decreasing testosterone levels may disrupt the maturation of spermatogenesis [24]. However, further randomized and placebo-controlled studies are needed to support the hypothesis that another advantage of AOX therapy is the improvement of serum testosterone and sperm parameters.

1.2. How to Manage Infertile Men With Elevated Sperm DNA Damage

When analyzing the impact of AOX administration on SDF, although it is still controversial, the majority of the data report a benefit in infertile men, regardless of the type of AOX used. Notably, treatment with NAC for 3 months has reduced SDF, as measured by the TUNEL assay [25]. Similarly, a combination of vitamin C, E, and CoQ10 [26] or CoQ10 alone [27] significantly improved SDF measured by the Sperm Chromatin Structure Assay (SCSA) or the Sperm Chromatin Dispersion (SCD), respectively [28]. On the contrary, a multicenter RCT that administered folic acid and zinc or placebo to 2370 infertile men for 6 months found no improvement in SDF levels in patients compared to controls [29]. In other clinical trials, patients with clinical varicocele did not experience additional benefits from AOX after varicocelectomy [30, 31]. However, in a recent meta-analysis, the positive impact of AOX treatment on spontaneous pregnancy and conventional sperm parameters in infertile men [10]. In addition, seminal levels of TAC were significantly higher, while seminal MDA levels were significantly lower in patients compared to controls. However, the lack of prospective clinical trials limits definitive conclusions on the effect of AOX on the reduction in SDF.

Some professional society guidelines have published conflicting recommendations on the use of AOX therapy in the management of male infertility. The European Association of Urology (EAU) guidelines suggest that no clear recommendation can be made for treating patients with idiopathic infertility using AOX [1]. However, seminal levels of ROS have been negatively associated with Assisted Reproductive Technology (ART) outcomes, and AOX therapy may improve semen parameters [1]. The American Urological Association (AUA) and the American Society for Reproductive Medicine (ASRM) guidelines suggest that clinicians should counsel patients that the benefits of supplements (e.g., AOX and vitamins) are of questionable clinical utility in treating male infertility. Existing data are inadequate to provide recommendations for specific agents for this purpose (Conditional Recommendation; Evidence Level: Grade B) [32].

The guideline recommendations take into account the limitations of existing studies, which are generally based on low-quality evidence, and therefore, do not support the indiscriminate use of AOX therapy. Despite the overall low-quality of evidence, existing SRMAs of RCTs on AOX therapy offer several important take-home messages for clinical practice. First, AOX supplementation should be considered only after addressing underlying causes of infertility—such as varicocele or urogenital infection—to optimize any potential benefit on SDF. Second, while a wide range of compounds and dosages have been studied, no specific AOX formulation has been shown to be superior. Third, to align with the duration of spermatogenesis, a minimum treatment period of 3 months is recommended to allow the AOXs sufficient time to exert their biological effects. In light of these considerations, there is a clear need for well-designed, randomized, and placebo-controlled trials to determine the optimal therapeutic strategy—including the type of compound(s), dosage, and duration—for improving SDF in men with idiopathic infertility or in those whose primary cause has already been treated.

1.3. Clinical Case Scenarios

Recommendations: AOX therapy for 3–9 months might improve SDF and conventional sperm parameters, spontaneous pregnancy rates, and IVF outcomes, leading to increased fertilization rates. AOX therapy would let the couple have the potential for pregnancy spontaneously by improving conventional sperm parameters or down-staging or shifting the level of ART [10, 12, 15–19].