Role of Vitamin B₆ in Testosterone Synthesis

Abstract

Boosting testosterone production and maintaining its adequate concentrations in males is still a critical area of the ongoing research. Vitamin B₆ (VB6), also named pyridoxine, is an essential nutrient and biochemically an organic cofactor in various enzymatic reactions that activate several catabolic and anabolic processes, including energy production, neurotransmitter synthesis, immune system function. While various studies have revealed an indirect association between testosterone and VB6, a collective and comprehensive review of its role in testosterone synthesis has been lacking. Here, we review and summarize the involvement of VB6 in testosterone production and regulation. To achieve this, a thorough search of the PubMed and Scopus databases was conducted, identifying English-language articles published from August 1956 to the present using the keywords “vitamin B₆” and “pyridoxine” versus testosterone. Relevant studies contributing to a mechanistic understanding of this relationship were also included. In summary, VB6 is essential for testosterone production and regulation through various indirect mechanisms. These include its impact on hormonal signaling pathways like GnRH and prolactin, its role in activating enzymatic processes that affect testosterone synthesis, its modulation of androgen receptor sensitivity, and its protection against oxidative stress and homocysteine toxicity. Such critical role of VB6 in regulating testosterone levels in men is considered a strong incentive for researchers to conduct interventional studies.

INTRODUCTION

Testosterone not only supports fertility and sexual function in men [1] but also affects their overall health. Several recent studies have linked low testosterone levels to a number of chronic diseases, especially those associated with aging [1]. For instance, low levels of testosterone in men has been found to be associated with diabetes, particularly type 2 diabetes [2,3,4,5], cancers [6,7], cardiovascular diseases [8,9], Alzheimer’s disease [10,11,12], Parkinson’s disease [13,14], osteoporosis [15,16], constant fatigue [17,18], and depression [19,20]. Above and beyond, recent clinical interventions have considered testosterone as one of the main therapeutic strategies to manage such diseases [4,8,13,21].

Vitamin B₆ (VB6), also known as pyridoxine, and its active form pyridoxal 5′-phosphate (PLP) (Fig. 1), is a water-soluble vitamin that belongs to B-vitamin family. PLP is involved in several aspects of macronutrient metabolism, histamine synthesis, neurotransmitter synthesis, hemoglobin synthesis, and gene expression [22,23]. Biochemically, PLP serves as a cofactor (coenzyme) for various enzymatic reactions including transamination, decarboxylation, elimination, racemization, beta-group interconversion, and replacement [24].

Fig. 1. Chemical structure of pyridoxal 5′-phosphate; the active form of vitamin B₆.

VB6 plays a multifaceted role in testosterone synthesis and overall hormone regulation. Male rats with a VB6 deficiency exhibited a reduced rate of testosterone synthesis [25]. Also, male rats subjected to a disrupted daily rhythm and administered normal amounts of VB6 with vitamin B1 and pantothenic acid, experienced a reduction in the suppression of gonadal development [26]. On the other hand, VB6 was found not to exhibit an androgenic activity [27]. In addition, intraperitoneal injection of Wistar rats with megadoses of pyridoxine (500 mg/kg or 1,000 mg/kg per day, for 2 to 6 weeks [28]) did not significantly alter the serum levels of testosterone [29].

Although several published studies, however indirectly, have linked VB6 with testosterone, no collective and comprehensive review has yet examined the role of VB6 in testosterone synthesis. This review explores and summarizes the involvement of VB6 in testosterone synthesis and regulation. To do so, we conducted a thorough search of the Scopus and PubMed databases for articles published in English from August 1956 through September 2024, using the keywords “VB6” and “pyridoxine” in conjunction with “testosterone.” Additionally, we included supporting articles that enhance and substantiate the mechanistic discussion.

TESTOSTERONE SYNTHESIS

Testosterone synthesis is a complex, tightly regulated process primarily occurring in the Leydig cells of the testes in males, with smaller contributions from the ovaries in females and the adrenal glands in both sexes [30]. The process begins with the transport of cholesterol into the mitochondria, where it is converted to pregnenolone by the enzyme cytochrome P450 sidechain cleavage enzyme [31]. Pregnenolone is then converted into testosterone through a series of enzymatic reactions involving 3β-hydroxysteroid dehydrogenase, 17α-hydroxylase, and 17,20-lyase [31]. The final and critical step in testosterone synthesis is catalyzed by 17β-hydroxysteroid dehydrogenase, which converts androstenedione to testosterone [31].

Testosterone synthesis is principally regulated by luteinizing hormone, a glycoprotein hormone secreted from the anterior pituitary gland [1]. Luteinizing hormone is secreted in response to gonadotropin-releasing hormone (GnRH) from the hypothalamus [32]. It binds to its receptors on Leydig cells, triggering a cascade of intracellular signaling events [30]. This signal transduction stimulates the expression and activity of enzymes like 17β -hydroxysteroid dehydrogenase, hence promoting testosterone synthesis [31]. In addition, follicle-stimulating hormone, which is also a glycoprotein secreted in response to GnRH, plays a supportive role by stimulating the Sertoli cells, and in consequence provide essential factors that support Leydig cell function and testosterone production [30,31].

Testosterone synthesis is further modulated by feedback mechanisms, where elevated testosterone levels inhibit the release of luteinizing hormone and GnRH, thus maintaining hormonal balance [33]. This intricate interplay of hormones ensures the precise regulation of testosterone concentrations that are vital for building muscle mass, enhancing reproductive function, supporting bone density, and other physiologic factors that affect overall health [34].

VB6 AND TESTOSTERONE: IN VIVO SYSTEM STUDIES

To date, direct studies linking VB6 to testosterone regulation and production remain limited, with most focusing primarily on in vivo models. A study conducted in 1984 revealed that male rats with a VB6 deficiency exhibited a reduced rate of testosterone synthesis compared to control [25]. In addition, male rats subjected to a disrupted daily rhythm and administered normal amounts of VB6 and vitamin B₁, along with high amounts of pantothenic acid, experienced a reduction in the suppression of gonadal development [26].

Alternatively, administering megadoses of pyridoxine at 500 mg/kg or 1,000 mg/kg daily, which corresponds in humans to 83.3 mg/kg and 166.7 mg/kg, respectively [28], injected intraperitoneally into Wistar rats five days a week for 2 or 6 weeks, affected spermatogenesis and reduced the weights of reproductive organs; however, it did not significantly alter serum testosterone levels in male rats [29]. In addition, an earlier study reported that VB6 does not exhibit an androgenic activity [27]. In fact, this evidence provides a support to the implication that the involvement of VB6 in testosterone production is not direct; instead, it supports several key steps and processes within the endocrine system that contribute to testosterone production. However, further direct animal studies seem to be highly desirable to filter the link between VB6 and testosterone.

MECHANISMS OF VB6 IN TESTOSTERONE SYNTHESIS

1. Regulation of hormonal signaling pathways

1) Modulation of prolactin levels

VB6 affects the level of prolactin, which is a hormone, when elevated, can negatively affect testosterone production (Fig. 2) [35]. VB6 is crucial for the synthesis of key neurotransmitters such as dopamine and serotonin. In particular, dopamine has an inhibitory effect on prolactin release; in which, increased availability of VB6 can enhance dopamine production via augmenting the activity of aromatic L-amino acid decarboxylase [36,37]. This enzyme increases the conversion of L-DOPA to dopamine, potentially leading to reduced prolactin level, and consequently affects testosterone synthesis [37]. Alternatively, high prolactin levels inhibit GnRH release via interfering with the action of kisspeptin peptide [38], a key hypothalamic regulator of GnRH release leading to decreased luteinizing hormone and subsequently reduce synthesis of testosterone [39,40,41].

Fig. 2. Pyridoxal 5′-phosphate (PLP), the active form of vitamin B₆, activates the conversion of L-DOPA, synthesized from tyrosine, into dopamine via activating L-DOPA decarboxylase (L-DD). Dopamine, in turn, inhibits the release of prolactin from anterior pituitary, a process that supports the production of testosterone. Generated using BioRender.com/PPT.

Moreover, chronic stress elevates cortisol levels, which can stimulate prolactin secretion [42]. Since VB6 plays a role in managing stress responses, adequate levels may help mitigate cortisol-induced prolactin elevation, which consequently affect testosterone production.

2) GnRH secretion

VB6 is crucial in the synthesis of serotonin and gamma-aminobutyric acid, which modulate the hypothalamic secretion of GnRH [43]. GnRH stimulates the pituitary gland to release luteinizing hormone, which in turn stimulates Leydig cells in the testes to produce testosterone [38]. In particular, pyridoxal-5′-phosphate, the active form of VB6, acts as a coenzyme for aromatic L-amino acid decarboxylase, facilitating the decarboxylation of 5-hydroxytryptophan to serotonin [44]. Whereas pyridoxal-5′-phosphate is essential for the activity of glutamate decarboxylase, which catalyzes the decarboxylation of glutamate to produce gamma-aminobutyric acid (Table 1) [45].

Table 1. The main enzymes and their functions that require PLP as a cofactor for their activity.

Enzyme	Function
Serine hydroxymethyltransferase	Converts serine to glycine and generates 5,10-methylenetetrahydrofolate, involved in one-carbon metabolism.
Cystathionine β-synthase	Catalyzes the condensation of homocysteine and serine to form cystathionine, a key step in the transsulfuration pathway.
Cystathionine γ-lyase	Converts cystathionine into cysteine, ammonia, and α-ketobutyrate in the transsulfuration pathway.
Glutamate decarboxylase	Converts glutamate to γ-aminobutyric acid (GABA), a major inhibitory neurotransmitter in the brain.
Aromatic L-amino acid decarboxylase	Catalyzes the decarboxylation of L-DOPA to dopamine and 5-hydroxytryptophan to serotonin, key neurotransmitters.
Aspartate aminotransferase	Catalyzes the reversible conversion of aspartate and α-ketoglutarate to oxaloacetate and glutamate, involved in amino acid metabolism.
Alanine aminotransferase	Catalyzes the reversible transfer of an amino group from alanine to α-ketoglutarate, producing pyruvate and glutamate, crucial in gluconeogenesis.
Glycogen phosphorylase	Breaks down glycogen into glucose-1-phosphate, important for energy production in muscle and liver cells.
Histidine decarboxylase	Catalyzes the decarboxylation of histidine to form histamine, which is involved in immune response and gastric acid secretion.
Threonine aldolase	Catalyzes the cleavage of threonine to glycine and acetaldehyde, involved in amino acid catabolism.

PLP: pyridoxal 5′-phosphate.

2. Enzymatic cofactor in steroidogenesis

1) Steroidogenic enzymes

It is known that VB6 plays a vital role as a coenzyme in numerous biochemical reactions involved in amino acid, glucose, and lipid metabolism [46]. One important specific route in such co-enzymatic action, particularly in lipid metabolism, is steroidogenesis, a process by which cholesterol is converted into steroid hormones [47]. This conversion occurs primarily in the adrenal glands and gonads and involves multiple enzymatic steps facilitated by cytochrome P450 enzymes. VB6 has been found to enhance several enzymes involved in steroidogenesis [48]. In particular, pyridoxine hydrochloride was found to significantly enhance the activity of 5 alpha-reductase, 3 alpha- and 17 beta-hydroxysteroid dehydrogenase [48]. Adequate levels of VB6 are essential for the optimal activity of these enzymes, ensuring efficient hormone synthesis. Above and beyond, VB6 deficiency has been associated with increased and prolonged nuclear uptake of steroid hormones, including testosterone, in target tissues. In VB6 deficient rats, there is increased and prolonged nuclear accumulation of estradiol in the uterus and testosterone in the prostate [49]. As mentioned above, supporting the biosynthesis of steroid hormones, particularly testosterone, by VB6 contributes not only to reproductive health but also to the modulation of numerous physiological processes influenced by androgens, such as muscle and bone development, mood regulation, and metabolic function.

Moreover, VB6 is involved in the metabolism of glycogen through its role in glycogen phosphorylase, which helps release glucose from stored glycogen, contributing to energy homeostasis (Table 1) [50,51]. It is now well-established that energy homeostasis and adequate testosterone levels are interconnected, with testosterone playing a vital function in metabolism, fat distribution, muscle function, and glucose regulation [52].

2) Alpha-reductase activity

VB6, in the form of pyridoxine hydrochloride, is believed to play a role in modulating the activity of the enzyme 5-alpha reductase, which converts testosterone into its more potent form, dihydrotestosterone [48]. While dihydrotestosterone is crucial for certain androgenic functions, its overproduction can lead to conditions like benign prostatic hyperplasia and androgenetic alopecia [53,54,55]. A study by Stamatiadis et al [56] (1988) investigated the effects of zinc and azelaic acid on 5α-reductase activity in human skin. The researchers found that zinc was a potent inhibitor of 5α-reductase, and this inhibitory effect was enhanced when combined with VB6 [56]. Specifically, VB6 potentiated the inhibitory effect of zinc, suggesting a synergistic interaction between the two in reducing 5α-reductase activity.

This finding indicates that VB6 may indirectly influence testosterone metabolism by modulating the activity of 5α-reductase, thereby affecting the conversion rate of testosterone to dihydrotestosterone. By inhibiting 5α-reductase activity, VB6 could potentially help maintain higher levels of testosterone relative to dihydrotestosterone, which may be beneficial in managing conditions linked to excessive dihydrotestosterone production.

It is important to note that while these findings are promising, further research is needed to fully elucidate the mechanisms by which VB6 influences 5α-reductase activity and to determine the clinical significance of these interactions in testosterone metabolism and related health conditions.

3. Modulation of androgen receptor activity

Androgen receptors are found in several bodily tissues such as bones, muscles, and the brain [57]. Principally, they modulate the actions of androgens by binding to testosterone and initiating a cascade of biochemical responses [57,58]. The appropriate action of testosterone is highly affected by the regulation of androgen receptors expression and sensitivity [58]. Alterations in these main receptors may lead to disorders such as hypogonadism or androgen insensitivity syndrome [59].

It has been shown that VB6 may play a significant role in modulating the sensitivity and expression of androgen receptors [60]. As a cofactor for various enzymes involved in amino acid metabolism (Table 1), VB6 is crucial for the synthesis of neurotransmitters, and hence the maintenance of hormonal balance [46]. Adequate levels of VB6 may enhance androgen receptor expression and sensitivity, leading to improved testosterone signaling. However, evidence from Allgood and Cidlowski (1992) [60] indicates that elevated VB6 levels reduce transcriptional activation of androgen and progesterone receptors by approximately 35% to 40%. This finding suggests that VB6 does not uniformly enhance androgen receptor activity but instead modulates it in a more complex manner, with potential reductions under certain conditions. These results highlight the importance of maintaining appropriate VB6 levels for optimal hormonal function, as both deficiencies and excessive concentrations could have distinct and potentially adverse effects. Additionally, VB6 deficiency has been linked to an increase in nuclear uptake and prolonged retention of steroid hormones, resulting in heightened sensitivity of target tissues to hormonal actions [61]. These findings underscore the importance of maintaining adequate VB6 levels for the effective regulation of androgen receptor-mediated gene expression and the corresponding cellular responses [61]. This interplay between VB6 and androgen receptor activity underscores the importance of nutritional factors in hormone regulation. It also suggests that deficiencies in VB6, in particular, may negatively affect the action of testosterone, with implications for metabolic health and mood regulation, thus overall well-being, particularly in males. Here, further research is desired to reveal the precise mechanisms by which VB6 impacts dynamics of androgen receptors, thereby investigating the therapeutic potential of VB6 supplementation in androgen-related diseases.

4. Antioxidant properties and protection against oxidative stress

VB6 has been identified as having a potent antioxidant activity [62]. It plays a vital function in maintaining cellular redox balance, as it is a cofactor for various enzymes involved in amino acid metabolism and the production of antioxidant molecules like glutathione (Fig. 3) [63]. When VB6 levels are deficient, there is a marked reduction in the synthesis of these antioxidants, leading to an accumulation of reactive oxygen species and consequent oxidative stress. This oxidative environment can damage cellular components, including lipids, proteins, and DNA [64,65]. Recent study suggested that pyridoxine deficiency may significantly alter the glutathione antioxidant defense system in the testes, leading to asthenozoospermia [65].

Fig. 3. PLP, the active form of vitamin B₆, enhances the conversion of homocysteine into cysteine, which in turn supports the production of glutathione. As a potent antioxidant, glutathione helps reduce oxidative stress in the testes, thereby contributing to the maintenance of testosterone production. CBS: cystathionine-β-synthase, CSE: cystathionine γ-lyase, PLP: pyridoxal 5′-phosphate, GCL: glutamate cysteine ligase, GSS: glutathione synthetase. Modified from de Queiroz et al [34] using BioRender/PPT.

However, this evidence is not direct, in the context of testosterone synthesis, oxidative stress particularly affects the Leydig cells in the testes, which are responsible for producing testosterone [66,67]. The high metabolic activity of Leydig cells makes them especially vulnerable to oxidative damage. Damage to these cells disrupts the enzymes and signaling pathways necessary for testosterone synthesis, leading to reduced hormone levels. Thus, VB6 deficiency indirectly impairs testosterone production by fostering an oxidative environment that compromises Leydig cell function and overall testicular. In 2020, a clinical intervention demonstrated that VB6, when combined with other antioxidants, increased serum testosterone levels in men with idiopathic infertility [68]. Future research should focus on generating direct evidence through targeted in vitro and in vivo studies examining VB6’s effects on oxidative stress markers and cellular health in Leydig cells. These efforts will help clarify VB6’s role and potential therapeutic applications in oxidative stress-related conditions.

5. Homocysteine metabolism

Homocysteine (2-amino-4-sulfanylbutanoic acid) is a homologue of cysteine. High levels of homocysteine in the blood are termed hyperhomocysteinemia, a toxic condition linked to various health problems, mainly cardiovascular diseases and cancers [69,70]. Studies have revealed that hyperhomocysteinemia is associated with reduced testosterone level in males, potentially influences its consequent metabolic action [71]. In addition, homocysteine accelerates oxidative injury and endothelial dysfunction [65], which negatively impacts the function of Leydig cells. The testes require a precise balance of reactive oxygen species and antioxidants, and excessive homocysteine disrupts this balance, leading to oxidative stress [65], and subsequently impairs synthesis of testosterone.

In addition, homocysteine-induced endothelial dysfunction can reduce blood flow to the testes, exacerbating this issue by creating a hypoxic environment that further hampers Leydig cell function [72]. In particular, hyperhomocysteinemia leads to quenching nitric oxide by oxidative stress and disrupts the uncoupling of nitric oxide synthase activity, leading to a reduction in nitric oxide bioavailability [72]. Decreased levels of nitric oxide can lead to decreased blood flow and further exacerbate the oxidative stress state within the testes, which negatively affects the Leydig cells’ function to produce testosterone. In addition, reduced levels of nitric oxide were found to influence the signaling pathways of luteinizing hormone that stimulates testosterone synthesis [73].

Moreover, VB6 is involved in the transsulfuration pathway, where it helps converting homocysteine to cysteine via activating cystathionine β-synthase and cystathionine γ-lyase (Fig. 3). Such metabolic coaction will reduces the accumulation of homocysteine, and hence indirectly enhances the synthesis of testosterone [62]. Furthermore, homocysteine may also disrupt the hypothalamic-pituitary-gonadal axis by interfering with the release of GnRH and luteinizing hormone, which are crucial for stimulating testosterone production [74].

CONCLUSIONS

VB6 plays a vital role in testosterone regulation and synthesis. The mechanisms by which this occurs are indirect via regulation of hormonal signaling pathways such as prolactin and GnRH, acting as a cofactor in enzymatic processes that affects testosterone regulation and production, modulating of androgen receptor sensitivity, protecting against oxidative stress, and reducing homocysteine toxicity (Fig. 4). However, a key limitation of this review is its reliance on existing literature, which primarily consists of indirect associations between VB6 and testosterone synthesis. While the review provides a comprehensive summary, the lack of direct in vivo evidence limits the strength of these conclusions.

Fig. 4. Mechanisms of VB6 in testosterone synthesis and function. The mechanisms through which VB6 influences testosterone include regulating hormonal signaling pathways such as prolactin and GnRH, modulating AR sensitivity, enhancing glucose homeostasis, and reducing oxidative stress and homocysteine toxicity. Testosterone influences reproduction, bone development, muscle development, mood regulation, and metabolic function. Generated using BioRender.com/PPT. VB6: vitamin B₆, GnRH: gonadotropin-releasing hormone, AR: androgen receptor.

Finally, the evident role for VB6 in maintaining testosterone levels in men serves as a compelling incentive for researchers to pursue further interventional studies to explore the potential of VB6 as a therapeutic tool to manage various medical conditions, particularly idiopathic infertility and reduced libido in males. Additionally, further research is warranted to establish a clear recommendation for daily VB6 supplementation, particularly through well-designed in vivo studies that assess its direct impact on testosterone levels.

Data Sharing Statement

All data generated or analyzed during this study are included in this published article.