By: Karl Hoffman at http://www.scivation.com/?pageID=26
Citrulline malate (CM) is a combination of two compounds that occur naturally in the human body. Malate is an intermediate in the so-called tricarboxylic acid cycle (TCA). ATP, which the body uses as a source of energy, is produced via the TCA when oxygen is abundant. (In reality, no ATP is produced directly from the TCA, although this statement is often heard. Rather, reduced coenzymes, NADH, are used to generate ATP in electron transport chain powered oxidative phosphorylation.) This is so called aerobic energy production.
Tricarboxylic acid cycle (TCA) at a glance. Each kind of major fuel is converted to acetyl groups, which are handled by attachment to a particular coenzyme known as coenzyme A. Ultimately ATP is produced from NADH generated by the TCA.
Malate is dehydrogenated in the TCA cycle to oxaloacetate, the concentration of which is one of the most critical controls of the rate of aerobic ATP production. During prolonged aerobic activity, and in patients suffering from malate deficiency, malate becomes depleted and the TCA is unable to produce ATP fast enough to meet the demands of working muscle. One classic disease characterized by malate deficiency is fibromyalgia. When patients suffering with this disease are given malate, their energy levels improve dramatically (1).
Not only patients suffering from malate deficiency benefit from malate supplementation. As mentioned above, strenuous, prolonged aerobic activity depletes the body’s malate stores. One recent study looked at the effects of CM supplementation in 18 otherwise healthy men who complained of easy fatigability. (2) The subjects were administered 6 gm/day of CM for 15 days. To quote from the results of the study,
"CM ingestion resulted in a significant reduction in the sensation of fatigue, a 34% increase in the rate of oxidative ATP production during exercise, and a 20% increase in the rate of phosphocreatine recovery after exercise, indicating a larger contribution of oxidative ATP synthesis to energy production… The expansion of the TCA intermediate pool [through malate supplementation] can therefore be regarded as a means of attaining higher rates of aerobic energy production, in agreement with our results showing that malate supplementation promotes a greater contribution of aerobic ATP production to total energy production. These results suggest that this hyperactivation of aerobic ATP production coupled to a reduction in anaerobic energy supply may contribute to the reduction in fatigue sensation reported by the subjects."So not only were objective measures of energy production increased, but the study participants felt a subjective improvement in energy levels as well.
Thus far we have only addressed the role of malate in enhancing ATP production during aerobic metabolism. What about citrulline? Citrulline is a non-essential amino acid produced from glutamine in the body. Citrulline is involved in the so-called urea cycle, which is responsible for the removal of excess nitrogen from the breakdown of amino acids. Were excessive levels of nitrogen to accumulate in the body, ammonia toxicity would develop. Besides stimulating hepatic ureogenesis , citrulline also promotes the renal reabsorption of bicarbonates. The latter acts as a buffer against lactic acidosis, which also helps to stave off fatigue. In fact there has been some debate over the years whether citrulline or malate is primarily responsible of prolonging endurance (3). The consensus now seems to be that the two compounds work in concert, with malate maintaining TCA intermediates and allowing for increased ATP production, and citrulline buffering against lactic acid and ammonia buildup.
So we have seen that citrulline malate seems to be a worthwhile adjunct to any supplement protocol, especially where aerobic performance and fatigue resistance are important.
The Role of Branch-Chain Amino Acids in Fatigue Resistance
The branched-chain amino acids isoleucine, leucine, and valine are widely used among athletes for their protein sparing effect.
L-leucine is also known as 2-amino-4-methylvaleric acid, alpha-aminoisocaproic acid and (S)-2-amino-4-methylpentanoic acid. It is abbreviated as Leu or by its one letter abbreviation L. Its molecular formula is C6H13NO2, and its molecular weight is 131.17 daltons.
L-isoleucine is also known as 2-amino-3-methylvaleric acid, alpha-amino-beta-methylvaleric acid and (2S, 3S)-2-amino-3-methylpentanoic acid. It is abbreviated as Ile or by its one letter abbreviation I. Its molecular formula is C 6H13NO2, and its molecular weight is 131.17 daltons.
L-valine is also known as 2-aminoisovaleric acid, 2-amino-3-methylbutyric acid, alpha-aminoisovaleric acid and (S)-2-amino-3-methylbutanoic acid. It is abbreviated as Val, and its one letter abbreviation is V. Its molecular formula is C5H11NO2, and its molecular weight is 117.15 daltons.
A number of studies have shown that branched chain amino acids exert both an anabolic and ergogenic effect.
For example, one study showed that BCAA administration post exercise resulted in an approximately 30% decrease amino acid efflux from skeletal muscle. The authors concluded that BCAAs exert a post training protein-sparing effect on muscle tissue (4). These results have been verified in numerous other studies. It is now believed that BCAAs act through a specific pathway, the so-called signal transduction p70(S6k) pathway in skeletal muscle (5). p70(S6k) is believed to control growth-related protein synthesis (5). There is also some evidence that branched chain amino acids are preferentially broken down for fuel during exercise, arguing for BCAA supplementation to offset this effect. If this is the case, this might be one mechanism where BCAA supplementation would hold off fatigue. Leucine seems particularly critical in stimulating overall protein synthesis. Leucine mediated signaling results in a stimulation of initiation of mRNA translation and involves increases in the phosphorylation status of the translational repression 4E-BP1 and the ribosomal protein S6 kinase S6K1mentioned above. It also requires sustained activation of the mammalian target of rapamycin (mTOR) protein kinase, a field of active research. Leucine, however, also signals to stimulate protein synthesis in skeletal muscle by a mammalian target of rapamycin protein kinase independent (i.e. rapamycin insensitive) pathway, suggesting that the amino acid may signal for protein synthesis through multiple pathways.
Interestingly, insulin is believed to exert at least part of its anabolic effect by activating the same p70(Sk6) pathway as leucine, but via different upstream channels. This argues for an additive role between elevated insulin levels and elevated BCAA levels in the promotion of anabolism.
There is another mechanism whereby BCAAs might prevent fatigue. We mentioned that BCAAs are used for fuel during exercise. As these amino acids become depleted, the ratio of tryptophan to BCAAs in the plasma rises (6). It turns out that tryptophan and BCAAs compete for the same amino acid transporter into the brain. The excess tryptophan in the brain is converted to serotonin, which induces a feeling of lethargy and fatigue. (Recall that tryptophan was widely used as a sleep aid.)
Interestingly men may have a greater need for BCAA supplementation than women. It is well established that women rely more on fat oxidation and less on glycogen and amino oxidation that do men during exercise. The BCAA leucine seems to be preferentially used among the amino acids as a fuel substrate in men (7).
TNF-Alpha and BCAA's
Tumor necrosis factor alpha (TNF-alpha) is a cytokine produced by immune cells in the body called monocytes and macrophages. TNF exerts a number of deleterious effects on the body, including muscle wasting, and locally produced igf-1 suppression, and general fatigue. While usually associated with illness, TNF-alpha levels are also high in hypogonadal patients and overtrained athletes. In one study, lipopolysachharide, a bacterial toxin that elevates TNF-alpha, was administered to rats. One group of rats was fed citrulline malate, and one group served as controls. The citrulline malate group performed much better on treadmill tests and exhibited less overall fatigue that did the controls (8).
These results may be of significance to overtrained athletes in whom TNF-alpha is elevated, and in anabolic steroid using athletes who are essentially hypogonadal post cycle. Citrulline malate may help alleviate the fatigue associated with both these conditions.
BCAAs may suppress TNF-alpha and its damaging effects on muscle tissue as well. In one study in animals, TNF-alpha was administered and diaphragm tissue was examined post mortem. Chronic TNF-alpha treatment produced a significant decline in the synthesis of all types of myofibrillar proteins, namely heavy chain myosin, light chain myosin and G-actin. TNF-alpha impaired peptide-chain initiation in diaphragm muscle was reversed by the branched-chain amino acids (BCAA) therapy of TNF-alpha treated rats. The authors concluded that
"These findings indicate a significant [inhibitory] role for TNF-alpha in the translational regulation of protein synthesis in skeletal muscle [which is reversed by BCAA administration ] (9) We see here a potential additive or even synergistic effect between citrulline malate and BCAAs in fighting muscle loss due to cytokines like TNF-alpha which are associated with illness, overtraining, and post anabolic steroid use.
In addition to blocking the muscle wasting catabolic effects of pro-inflammatory cytokines, BCAAs also seem to fight a tug of war with catabolic glucocorticoids. This suggests that under conditions such as stress or overtraining, BCAAs might help alleviate the catabolic effects of cortisol (10).
Darkhorse on 08-31-2005, 05:20 PM
Leucine Activates system A Amino Acid Transport in Skeletal Muscle Cells
Just as glucose uptake into cells is dependent upon a family of so called GLUT (Glucose Transporters), amino acids are transported into cells by a distinct family of transporters. One main transport system is the so-called System A. In vitro studies have shown that leucine upregulates System A transporters, allowing for greater entry of a number of amino acids into muscle cells (11). It should be noted that one of the anabolic effects of IGF-1 is believed to be upregulation of this same transport system. So in this sense leucine may share at least one of the anabolic effects associated with insulin like growth factor.
So we have seen that amino acids, and leucine in particular are capable of activating signal transduction pathways in an almost hormone like manner. They are much more than simple nutrients in this regard. To quote from one recent review,
"The protein kinase mTOR is a common intermediate in both nutrient and hormone signal transduction pathways. Signaling through mTOR is enhanced by nutrients and anabolic hormones, such as insulin or IGF-I and repressed by elevation of cAMP or activation of AMPK suggesting that one function of mTOR is to integrate the anabolic response to nutrients and insulin and the catabolic response to counter-regulatory hormones, such as glucagon...Although other amino acids have been shown to increase signaling through mTOR, leucine is arguably the most potent of the amino acids in activating the pathway." (12)
The importance of leucine to post resistance exercise recovery is highlighted in another recently published study where participants undertook a bout of resistance exercise and were fed either carbohydrates alone, carbohydrates plus whey protein, or a combination of carbs, whey and leucine (13). Subjects received a beverage volume of 3 ml.kg-1 every 30 minutes to ensure a given dose of 0.3 g carbohydrate.kg-1 (50% as glucose and 50% as maltodextrin) and 0.2 g.kg-1 of a protein hydrolysate [whey] every h, with or without the addition of 0.1 g.kg-1.h-1 leucine. To quote from the report,
"Mixed muscle [protein synthesis rate], measured over a 6h period of post-exercise recovery, was significantly greater in the CHO+PRO+leu trial compared to the CHO, with intermediate values observed in the CHO+PRO trial . We conclude that the co-ingestion of protein and leucine stimulates muscle protein synthesis and optimizes whole-body protein balance when compared to the intake of carbohydrate only."
Before leaving the subject of BCAAs and citrulline, it might be worthwhile to briefly discuss the role of another amino acid, glutamine that is both quite popular and controversial as a supplement. The bulk of the research suggests that under normal circumstances the body is capable of making adequate amounts of glutamine from the branched chain amino acids discussed above. However, when the body is taxed by illness, sepsis, cancer, cachexia, and trauma, and perhaps chronic overtraining, the body may become depleted in BCAAs. The BCAAs released to circulation may be used for protein synthesis or synthesis of alanine and glutamine. Glutamine and/or alanine infusion has an inhibitory effect on the breakdown of body proteins and decreases BCAA catabolism in postabsorptive control, endotoxemic, and irradiated rats. This helps preserve muscle mass under these conditions (14). So while glutamine supplementation might not be absolutely necessary with an adequate intake of amino acids, especially BCAAS, it’s hard to see how glutamine supplementation could hurt, and might actually be beneficial.
So in summary we can obtain an additive anti-fatigue effect by combining citrulline malate with branched-chain amino acids. The former replenishes TCA intermediates allowing for enhanced ATP production, while the latter preserves vital muscle tissue BCAAs and blocks the entry of fatigue inducing tryptophan into the brain. Additional glutamine supplementation might be beneficial should the body become depleted in BCAAs during strenuous prolonged exercise or illness.
Dosage of Citrulline Malate
Concerning dosage of citrulline malate in humans, based on the work in (2), 6 grams per day of citrulline malate taken in two divided doses during the day may be the optimal dose. None of the studies in humans suggested any serious side effects, and the drug appears to be well tolerated.
Russell IJ, Michalek JE, Flechas JD, Abraham GE. Treatment of fibromyalgia syndrome with Super Malic: a randomized, double blind, placebo controlled, crossover pilot study. J Rheumatol. 1995 May;22(5):953-8.
Bendahan D, Mattei JP, Ghattas B, Confort-Gouny S, Le Guern ME, Cozzone PJ. Citrulline/malate promotes aerobic energy production in human exercising muscle. Br J Sports Med 2002 Aug;36(4):282-9
Callis A, Magnan de Bornier B, Serrano JJ, Bellet H, Saumade R. Activity of citrulline malate on acid-base balance and blood ammonia and amino acid levels. Study in the animal and in man. Arzneimittelforschung 1991 Jun;41(6):660-3.
Blomstrand E, Saltin B. BCAA intake affects protein metabolism in muscle after but not during exercise in humans. Am J Physiol Endocrinol Metab 2001 Aug;281(2):E365-74
Karlsson HK, Nilsson PA, Nilsson J, Chibalin AV, Zierath JR, Blomstrand E. Branched-chain amino acids increase p70S6k phosphorylation in human skeletal muscle after resistance exercise. Am J Physiol Endocrinol Metab. 2004 Jul;287(1):E1-7
Blomstrand E, Celsing F, Newsholme EA. Changes in plasma concentrations of aromatic and branched-chain amino acids during sustained exercise in man and their possible role in fatigue. Acta Physiol Scand 1988 May;133(1):115-21
Lamont LS, McCullough AJ, Kalhan SC. Gender differences in the regulation of amino acid metabolism. J Appl Physiol. 2003 Sep;95(3):1259-65
Verleye M, Heulard I, Stephens JR, Levy RH, Gillardin JM. Effects of citrulline malate on bacterial lipopolysaccharide induced endotoxemia in rats. Arzneimittelforschung 1995 Jun;45(6):712-5
Cheema IR, Hermann C, Postell S, Barnes P. Effect of chronic excess of tumour necrosis factor-alpha on contractile proteins in rat skeletal muscle. Cytobios. 2000;103(404):169-76.
Liu Z, Jahn LA, Long W, Fryburg DA, Wei L, Barrett EJ. Branched chain amino acids activate messenger ribonucleic acid translation regulatory proteins in human skeletal muscle, and glucocorticoids blunt this action. J Clin Endocrinol Metab 2001 May;86(5):2136-43.
McDowell HE, Christie GR, Stenhouse G, Hundal HS. Leucine activates system A amino acid transport in L6 rat skeletal muscle cells. Am J Physiol. 1995 Nov;269(5 Pt 1):C1287-94.
Kimball SR, Jefferson LS. Amino acids as regulators of gene expression. Nutr Metab (Lond). 2004 Aug 17;1(1):3
Koopman R, Wagenmakers AJ, Manders RJ, Zorenc AH, Senden JM, Gorselink M, Keizer HA, van Loon LJ. The combined ingestion of protein and free leucine with carbohydrate increases post-exercise muscle protein synthesis in vivo in male subjects. Am J Physiol Endocrinol Metab. 2004 Nov 23
Holecek M. Relation between glutamine, branched-chain amino acids, and protein metabolism. Nutrition. 2002 Feb;18(2):130-3
Nakata M, Yada T. Endocrinology: nitric oxide-mediated insulin secretion in response to citrulline in islet beta-cells. Pancreas. 2003 Oct;27(3):209-13