Sunday, September 17, 2017

2.5g/d Betaine Double Fat Loss, Improve Lean Mass Gains | Citrulline & Glutathione Lack Effect on Body Composition

Betaine or citrulline? Choose now.
Article #2 of this series is a bit more "anabolic" than the first serving. Based on the abstracts to studies by Hudson et al, Cicholski et al and Hwang et al it explores the effects of betaine supplementation on the adaptational response to resistance training in female rookies and the longitudinal effect of the NOx-boosting combination of citrulline + glutathione (see previous article) on the body composition of resistance-trained male subjects.

As usual, the discussion of the studies is infused with plenty of information from previous research... and for those who want nothing but the gist, there are the practical implications in the bottom line.
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  • 9 weeks "on 2.5g/d betaine" significantly improve the body composition of untrained young women - You will remember that I've covered the potential beneficial effects of betaine, aka N,N,N-trimethylglycine or glycine-betaine (note: that is not betaine HCL!), an amino acid from sugar beets, for lifters (learn more).

    There's was also preliminary evidence that betaine can significantly increase IGF-1 (x2) and help you shed body fat. The study Hudson et al. (2017) presented at #ISSN17 expands on the existing evidence by providing evidence that the daily ingestion of 2.5g/kg betaine over 9 weeks can, when it is combined with resistance training, significantly improve the body composition of young women (N=23; 21.0±1.4 years, 165.9±6.4 cm, 68.6±11.8 kg, 32.7±7.6% body fat).

    To reduce confounding due to baseline inter-group differences, the women had been pair-matched based on body composition and squat strength before they were then randomly assigned to a placebo (n=12) or a betaine (2.5 g/day, BetaPower®, Finnfeeds Oy, Finland; n=11) group in a double-blind fashion. Furthermore, all participants followed the same lower/upper body split, performing the two workouts on non-consecutive days for two 4 week blocks with 1 week of active rest between blocks. Body composition data you see in Figure 1 was measured using BodPod device (air displacement plethysmography), the thickness of the rectus femoris (only right leg) was measured using B-mode ultrasound. The effects on performance were reported in a separate presentation.
    Figure 1: Relative and absolute reduction in body fat (left), effect sizes (right | Hudson 2017).
    What is worth mentioning is that both treatment significantly reduced total body mass and body fat mass while increasing fat-free mass and muscle size. However, the betaine group had a significant edge in three out of four of these variables: The subjects in the betaine group lost an extra 1.6% body fat (-3.3+1.9%) compared to the placebo (1.7+1.6%) group, the saw greater reductions in total body fat and more pronounced increases in fat-free mass (45.4+6.1 vs. 47.9+5.9 kg), with the effect sizes for BF% (-.45 vs. -.22), FM (-.22 vs. -.08), and FFM (.48 vs. .31) telling you that only the betaine group scratched the value of d=0.5 that would signify a medium effect size.
Warning - the studies discussed in this series are not yet peer-reviewed and published! Since the write-up is based on abstracts, only. I cannot discuss and scrutinize the results with the same degree of detail and healthy skepticism you're used to from other SuppVersity articles.
  • In contrast to the changes in body composition, the performance gains that were part of an individual presentation (Chicholsky 2017) did not show significant inter-group differences: Vertical jump (39.7±6.2 vs. 44.4±7.0 cm), 1RM back squat (60.1±16.4 vs. 78.2±17.0 kg), and 1RM bench press (34.9±7.2 vs. 39.0±8.0 kg) increased sign. without inter-group differences and the effect sizes (Cohen’s d) were similar for betaine vs. placebo - a noteworthy difference was observed only for the 1RM bench press (d = .46 vs. 69), where the effect sizes seem to favor the placebo over betaine.

    Figure 2: Betaine supplementation promotes an anabolic millieu (Apecilla 2013).
    The inclusion of the separately reported performance data is also interesting because they seem to suggest a potential sex-difference. After all, previous trials in male subjects showed significant inter-group differences favoring betaine over placebo in terms of both, body composition and performance. Future studies will thus have to (a) clarify the effect of sex, investigate (b) the influence of training status and assess (c) the interaction with staple supplements most of you will already be taking (e.g. whey/other protein shake and creatine).

    Maybe these studies will also be able to elucidate betaine's mechanism of action. As previously hinted at, the lean mass increases could be related to increases in IGF-1. Additional candidates are the significant improvements in the mTOR-targets Akt (Ser473) and the "protein-pump" p70S6k (Thr389), Apecilla et al. (2013) observed alongside increases in GH and reductions in cortisol in their 2013 trial in twelve trained young men (see Figure 2).

    The latter, i.e. the reduction in cortisol and increase in GH could, alongside the reduction in lipogenic (FAS, LPL, FABP) mRNA expression that has been observed in animal studies before, also explain the highly significant body fat reductions Hudson et al. observed in their untrained female subjects. 
  • Glutathione + citrulline has anabolic effects that go beyond the previously established increase in "pump" (NOx) -- You will remember my 2015 article about the nitric oxide boosting effects of a combination of citrulline and glutathione. And if you do, you will probably remember being disappointed that the acute phase study lacked data on the potential longitudinal effects of citrulline + glutathione on muscle and strength gains... well, Paul Whang et al. (2017) have now investigated whether this long-term effect on your gains does indeed exist.

    Can citrulline supplementation prevent you from hitting a catabolic wall, when you are dieting? And is it more potent than leucine? Find out!
    In that, it is noteworthy that they start with the hypothesis that any beneficial effects of the two supplements would, in fact, be mediated by increases in plasma nitric oxide metabolites (NOx) and cyclic guanosine monophosphate (cGMP), of which they speculate that they "may play a role in muscle protein synthesis" (Hwang 2017). A far more obvious reason, i.e. the well-known m-TOR promoting effect of citrulline (Le Plénier 2012; Cynober 2013), on the other hand, is not mentioned in the abstract.

    What? Oh, yes. You're less interested in the mechanism than the study design and outcome. Ok, here's the obviously limited information from the abstract (would be nice to have information about the subjects' diet and whether they were using other supplements).

    Well, Hwang et al. conducted randomized, double-blind, placebo-controlled study with, believe it or not, 75 resistance-trained males were randomly assigned to ingest 2 g/day of L-citrulline + 200 mg/day of GSH (CIT+GSH), 2 g/day of L-citrulline-malate (CIT), or 2 g/day of cellulose placebo (PLC). Over the course of the 8-week study period, all participants followed identical resistance training protocols and body composition and muscle performance were assessed before and after 4 and 8 weeks of RT and supplementation.
    Figure 3: The only measurable inter-group difference were the augmented lean mass gains in week 4/8.
    Neither RT nor supplementation had any significant effects on total body mass, total body water, fat mass, muscular strength, and endurance, or any of the blood clinical chemistry variables (p > 0.05) - remember: we're talking about resistance-trained subjects, in whom changes in body composition take place at much slower rates, compared to rookies as the female subjects in the previously discussed betaine study.

    After 4 weeks, the scientists did, however, observe a significant advantage of the GSH+CIT and CIT groups compared to PLC - an advantage that was lost after 8 weeks (p > 0.05), though. In view of the fact that the results, i.e. both glutathione+citrulline and citrulline, alone, yielded measurable advantages point towards citrulline as being the main motor of the lean mass increases in week 4, it is a pity that we don't know more about the study yet - if the subjects were, for example, allowed to consume protein shakes and/or followed high protein diets, the previously referenced effects on mTOR may have been overridden by their supplemental or dietary protein intake.

    What I can say without having the full paper in front of me, though, is that the benefits of citrulline and, even more so, those of glutathione are probably very limited for experienced trainees striving to improve their body composition.
As SuppVersity reader you've known about betaine's IGF boosting effects for more than 3 years | learn more
What does that mean for you? Although the data from Hudson's betaine study looks quite exciting. It would be preliminary to head over to the bulk-supplier of your choice to order a few lbs of betaine (remember: that's glycine-betaine, not betaine HCL). After all, the average SuppVersity reader is not exactly untrained and (that's at least what I assume) consumes a high(er)-protein diet and/or supplemental protein shakes. Unless the effect of these confounding variables is assessed it's hard to see if trained subjects with increased protein intakes will benefit to the same extent as the untrained young women, who usually consume low(er) protein diets (e.g. 75g | Cooper 1996).

The previously phrased reservations make the study by Hwang et al the perfect complement to the Hudson study. Hwang et al had exactly what the Hudson study didn't have: trained individuals and (at least most likely) high(er) protein intakes (than the average 21-year-old untrained woman)... and guess what: even though the protein anabolic effects of citrulline have been well established in rodent models, the provision of 2 g/day of L-citrulline had, if anything, an intermediate effect on the subjects' body composition... ah, in case you're asking yourself why I don't address the glutathione content of the supplement? Well, it had absolutely zero effect and is useless for anyone supplementing with a glutathione booster like whey protein (Ha 2003; Middleton 2004).

What does this teach us? Yes, individuality and context matter: What works in untrained individuals with (purportedly) suboptimal protein intakes, doesn't necessarily work in athletes who pound several g/kg body weight of protein. In all fairness, it should be said, though, that the reverse applies to the failure of citrulline in Hwang's study: less trained individuals or people with lower baseline protein intakes (I simply assume that the "resistance-trained" subjects got more than 0.8g/kg) could well have seen body composition benefits from citrulline | Comment on Facebook!
  • Apicella, Jenna M., et al. "Betaine supplementation enhances anabolic endocrine and Akt signaling in response to acute bouts of exercise." European journal of applied physiology 113.3 (2013): 793-802.
  • Cooper, C., et al. "Dietary protein intake and bone mass in women." Calcified tissue international 58.5 (1996): 320-325.
  • Cynober, Luc, Jean-Pascal de Bandt, and Christophe Moinard. "Leucine and citrulline: two major regulators of protein turnover." Nutrition in Intensive Care Medicine: Beyond Physiology. Vol. 105. Karger Publishers, 2013. 97-105.
  • Ha, Ewan, and Michael B. Zemel. "Functional properties of whey, whey components, and essential amino acids: mechanisms underlying health benefits for active people." The Journal of nutritional biochemistry 14.5 (2003): 251-258.
  • Le Plénier, Servane, et al. "Effects of leucine and citrulline versus non-essential amino acids on muscle protein synthesis in fasted rat: a common activation pathway?." Amino acids 43.3 (2012): 1171-1178.
  • Middleton, N., P. Jelen, and G. Bell. "Whole blood and mononuclear cell glutathione response to dietary whey protein supplementation in sedentary and trained male human subjects." International journal of food sciences and nutrition 55.2 (2004): 131-141.