Effects of betaine on body composition, performance, and homocysteine thiolactone

Betaine (trimethylglycine) is an organic osmolyte found in many foods, including spinach, beets, and whole grains[1]. Administration of supplemental betaine for 10–15 days has enhanced performance in several studies but with varying results: Lee et al.[2] reported increased power output and force production, whereas others[3, 4] reported improvements in muscular endurance but not power. On the other hand, Del Favero et al.[5] reported no improvements in power output, strength, or body composition with 10 days of betaine treatment; however, subjects were instructed to avoid training and supplementation was ceased 5 days prior to performance testing.

To the author’s knowledge, only two studies have examined the effects of betaine on body composition and hypertrophy in humans. Betaine did not improve body composition in obese, sedentary subjects on a 500 kcal/day caloric deficit following 12 weeks of supplementation[6]. Similarly, 10 days of betaine supplementation did not improve body composition in sedentary young male subjects[5]. Though research is limited in humans, chronic betaine supplementation has been shown to reduce adipose mass and increase muscle mass in animals[7–9]. Greater improvements in body composition with betaine supplementation were observed when pigs were given extra pen space to move and exercise[9], suggesting that betaine may exert the most influential effects on growth under conditions of metabolic or nutritional stress. Because the subjects in Schwab et al.[6] and Del Favero et al.[5] were instructed not to exercise, the absence of a metabolic stressor may have compromised the effects of betaine. Given the enhanced effects of movement in pigs and the ineffectiveness reported in sedentary, non-exercising humans, we hypothesize that the effects of betaine on body composition, strength and power may be most apparent when supplementation occurs over several weeks accompanied by a resistance training program.

By donating a methyl group to transmethylate Hcy back to methionine (Met), betaine increases Hcy metabolism and the availability of the universal methyl donor, S-adenosylmethionine (SAM)[10]. We hypothesize betaine supplementation may enhance protein synthesis and thus improve body composition by reducing Hcy and homocysteine thiolactone (HCTL). Hcy directly impairs insulin signaling by reducing insulin receptor stubstrate-1 (IRS-1) activation and thus inhibiting Akt-phosphorylation[11]. Moreover, excess dietary Met is metabolized to form Hcy and both high dietary Met consumption and the resultant increase in plasma Hcy contributes to elevated HCTL[12]. A short (10 min) HCTL treatment inhibits insulin signaling, including insulin-mediated mRNA expression and protein synthesis[13]. This suggests that HCTL is more effective than Hcy in promoting insulin resistance. Additionally, HCTL has been shown to modify protein lysine residues, which causes protein aggregation, and inactivates enzymes associated with protein synthesis[14].

Concentrations of plasma Hcy or HCTL levels in strength athletes have yet to be reported. Given that transmethylation capacity is dependent upon plasma folate and betaine[15] and because weight trainers regularly consume excess Met and inadequate folate and betaine[16], Hcy transmethylation may be impaired resulting in excess HCTL generation. Thus, by decreasing insulin receptor signaling[11], elevated HCTL in weight lifters may compromise body composition directly by inhibiting mRNA expression and protein synthesis.

In healthy adults the ingestion of 500 mg of betaine decreased fasting plasma Hcy and attenuated Hcy rise for 24 hr following a Met load[11], and betaine treatment lowers HCTL in patients with genetically compromised transmethylation capacities[12]; however, to date there are no published reports investigating the effects of betaine ingestion on HCTL in healthy subjects. We hypothesize that by increasing transmethylation capacity betaine supplementation reduces plasma Hcy and may thus decrease HCTL generation, resulting in improved insulin signaling and myofibril protein synthesis, and ultimately enhancing muscle and strength gains. Therefore, the purpose of this study was to investigate the sub-chronic effects of betaine on strength, power, and body composition during resistance training in experienced strength trained males. Additionally, urine HCTL was measured to determine if betaine affects performance by reducing plasma HCTL. We hypothesized that betaine supplementation would improve strength, vertical jump, limb CSA, and body composition between the 1^st week and 6^th week over placebo. We also hypothesized that betaine supplementation would reduce urinary HCTL over the course of 6 weeks.

We hypothesized body composition would improve with 6 weeks of betaine supplementation. This hypothesis was supported by significant increases in lean mass, and decreases in fat mass and body fat percentage with betaine compared to placebo. Increases in arm CSA were found to be greater with betaine than placebo; however, thigh CSA did not increase in either group. We also expected strength and power performance to improve with betaine supplementation. While back squat 1 RM increased for both groups, there were no differences in improvement between betaine and placebo. There was a trend (p = .07) for greater vertical jump power with betaine versus placebo, however there were no increases in bench press 1 RM.

The improvements in lean mass, fat mass and body fat percentage with betaine supplementation contrast previous investigations[5, 6]. Differences in methodology may explain these discrepancies: subjects in the previous studies were both sedentary and instructed not to exercise, whereas the subjects in the present study were currently training and given a structured exercise program. Betaine has been suggested to act as a nutrient partitioner and thereby accelerate lean mass gains in pigs. By increasing Hcy transmethylation, betaine spares Met, allows for more efficient use of dietary protein, and increases nitrogen retention[7]. Due to the inclusion of resistance training in this study but not previous studies[5, 6], the demand for Met in the initiation of translation in protein synthesis was likely elevated, thereby leading to a greater utilization of elevated Met, and thus improvements in lean mass. Therefore, the results from the present study lend support to the hypothesis that the action of betaine to improve body composition in humans may be most effective when accompanied by exercise.

The increase in arm CSA in the betaine group compared to placebo was accompanied by an improvement in bench press work capacity. The greatest improvements in volume over placebo occurred during the first and third training micro-cycles, where subjects were instructed to perform 3 sets of 12–15 repetitions with 90 sec rest periods and 3 sets of 8–10 repetitions with 120 sec rest periods, respectively. Given the relationship between training volume and hypertrophy[29], betaine may have positively impacted muscle growth by promoting a greater training load over a series of subsequent workouts.

The improvements in bench press work capacity differ from previous studies where betaine did not improve single-set repetitions to fatigue at 75%[3] or 3 sets of repetitions to fatigue at 85% 1 RM[2]. In contrast, betaine improved work capacity for 10 sets of repetitions to fatigue at 50% 1 RM[4]. Given improved work capacity with higher volume resistance training prescriptions, and the lack of improvement during micro-cycle 2 which imposed less of a metabolic demand (4 sets of 4–6 repetitions with 3 min rest), it is likely that betaine poses the most ergogenic potential in resistance training exercise protocols that impose higher metabolic demands. Betaine is actively taken up by skeletal muscle during periods of stress, and may be ergogenic as an osmolyte by protecting sensitive metabolic pathways against cellular hypertonicity such as protein turnover, amino acid and ammonia metabolism, pH regulation, and gene expression[30]. Specifically, betaine maintains cellular hydration to protect myosin ATPase and myosin heavy chain proteins against denaturation by urea[31]. Moreover, the affinity of troponin for Ca²⁺_, and thus force production, is negatively affected by reductions in protein hydration[32].

Contrary to the changes in arm CSA, no differences in leg CSA were found between groups. Similar results have been reported in animal studies investigating the effects of betaine supplementation on carcass cuts where betaine supplementation improved shoulder and butt, but not ham meat yield[9]. Additionally, changes in upper body muscle thickness occur at a greater magnitude and earlier than do the lower extremities[33]. Therefore, it is possible that changes in thigh CSA may have occurred with a longer study period.

Although the back squat requires recruitment of the quadriceps femoris, it also has a high gluteal/hip requirement. Increases in muscle mass may have occurred predominantly in the gluteals as seen in animal studies, or the adaptations leading to greater back squat volume and 1 RM occurred separately from increased muscle CSA. Back squat work capacity increased for each group at each training micro-cycle; however, the betaine group improved nearly two-fold compared to placebo during micro-cycle three (4 sets of 4–6 repetitions with 3 min rest) which posed a higher neural and lower metabolic demand than the previous micro-cycles. These improvements in back squat work capacity contrasts previous results[34] whereby betaine did not improve mean or peak isokinetic power during 5 sets of 6 repetitions at 80% peak force. The improvements in work capacity at micro-cycle three but not micro-cycle one or two also contradict our hypothesis that betaine may be most ergogenic when combined with exercise protocols producing higher levels of metabolic stress. Given the improvement in bench press work capacity that also occurred at micro-cycle three but not two, and the lack of improvement with only 2 weeks of supplementation[2, 4], it may also be that the effects of increased intramuscular betaine manifest over a longer period of time, and therefore require at least a 4–6 week ingestion period.

There were no differences between groups for back squat 1 RM improvements, and despite increases in bench press training volume with betaine, bench press 1 RM did not improve. This contrasts previous reports[2], and may be partially explained by difference in subject training status. Lee et al. employed recreationally trained subjects, whereas subjects in the present study averaged 4.8 years of training experience. The ability to make large performance gains, termed the “window of adaptation”[35], decreases with training experience. The “window of adaptation” was likely smaller for the subjects in the present study, thus reducing the ability to detect changes in strength. Finally, the primary aim of this study was to evaluate the effects of betaine on muscle growth; thus, the training program utilized was selected because it provided the greatest stimulation for hypertrophy. Given the high training status of the subjects, a strength concentrated program (i.e.: 4–6 sets of 1–3 repetitions) may have been needed to induce further improvements in bench press and back squat 1 RM with betaine supplementation.

There was a trend (p = .07) toward an increased vertical jump with betaine supplementation. The positive trend in the present study and improvements reported by Lee et al.[2] differs from the results reported by other researchers where vertical jump did not increase with betaine[3, 4]. Variances in training prescription may account for these discrepancies. In Lee et al. and the present study subjects were assigned standardized training between testing sessions, whereas subjects in Hoffman et al.[4] and Trepanowski et al.[3] were not. Because detections in power improvements are compromised when power movements are not a regular part of training[34], future researchers should include exercises that train muscular contractile velocity when investigating the effects of betaine supplementation on power output.

We hypothesized that subjects would have high urinary HCTL values due to reduced Hcy transmethylational capacity; however, the results did not support this hypothesis. The normal range for urinary HCTL is .011-.473 nmol/mL[24]. Mean pretreatment HCTL was .028 nmol/mL (± .02 nnmol/mL), which suggests that the subjects began the study with low HCTL levels. Betaine supplementation attenuated the rise in HCTL observed in placebo at weeks 2 and 4, but did not appear to reduce HCTL values. Many subjects moved from the campus dormitories to live with their parents for the summer. It is possible that subjects had access to foods higher in protein quality and richer in fats and cholesterol than when living on campus, and this led to the increase in HCTL. Increases in dietary fat and cholesterol have been shown to increase plasma Hcy[36] as 3 Hcy are produced during the methylation of phosphatidylethanolamine in very low density lipoprotein synthesis. Thus, higher methionine and fat intakes may have increased Hcy generation, leading to higher levels of HCTL. Given the ability of betaine to increase Hcy transmethylation, it is possible that betaine supplementation attenuated the dietary induced rise in HCTL.

HCTL decreased in both groups between week 4 and week 6, although there was a trend for a reduction in HCTL when comparing week 6 to baseline with betaine and not placebo. While subjects were instructed to maintain the same diet throughout the study, many foods rich in betaine and folate come into season in June including spinach (0.3 mg/cup folate) and collard greens (0.2 mg/cup folate), and the consumption of two-three servings of folate rich food per day will reduce Hcy by 20%[37]. Because the start of June corresponded with week 4 of the study, it is possible that the consumption of local greens and the resultant increase in folate consumption may have reduced HCTL values in week 6.

Based on the small differences in HCTL changes, betaine supplementation may have impacted body composition via other mechanisms. Betaine has been shown to elevate plasma GH and IGF-1, and increase Akt phosphorylation in human skeletal muscle[38]. In mice betaine improves insulin sensitivity by restoring activation of IRS1 and the subsequent phosphorylation of PI3K/Akt by 50-100% in a concentration-dependent manner[39]. Thus, it is possible that by elevating anabolic hormones and enhancing downstream cellular signaling, betaine may have improved muscle protein synthesis, thus leading to an increase in lean mass. Finally, because betaine is a powerful osomylte, the increases in lean mass may have been due to cellular swelling without an appreciable increase in myofibril protein accretion.

In summary, the major findings of the present study are that 6 weeks of betaine supplementation improved body composition, muscle size, work capacity, attenuated a rise in HCTL, tended to improve power, but not strength in resistance trained men. Further work is warranted to confirm any role of HCTL on body composition compared to other mechanisms like lipogenic enzymatic activity, growth hormones, cellular signaling, or gene expression. Betaine attenuated an increase in urinary HCTL; however, because strength trained men in this study had low baseline HCTL betaine likely affected body composition via another mechanism. HCTL has been implicated in vascular disease[40], insulin resistance[13], diabetic retinopathy[41], seizures, and Alzheimer’s disease[42]. Thus, future investigations are needed to evaluate the clinical ability of betaine to reduce HCTL in at risk populations with elevated Hcy.