Protein ingestion increases muscle protein synthesis in proportion to the essential amino acid (EAA) content of the protein dose. While intact protein ingestion is the most recognizable means for delivering EAA, it is a calorically inefficient means of delivering EAA because they represent 40 to 45% of the total amino acid (AA) content in most high-quality proteins. Furthermore, larger protein intakes result in ureagenesis and excretion of a substantial portion of the ingested AA. On the contrary, provision of only the EAA component presents several advantages over intact proteins.
First, EAA ingestion results in a rapid increase in peripheral AA concentrations and delivery to skeletal muscle, thereby signaling an increase in protein synthesis. Second, ingestion of the EAA only results in an efficient utilization of endogenous nonessential AA, thereby reducing ureagenesis and AA excretion. Third, EAA ingestion maximizes the stimulation of muscle protein synthesis without affecting the metabolic response to subsequent meals and without negatively impacting subsequent caloric intake/substitution. Finally, on the basis of gram of AA uptake per gram of AA ingested, EAA ingestion results in a 2-fold greater accretion of muscle protein.
This article represents a compilation of metabolic studies from our laboratory and others, which used stable isotope methodology to directly determine muscle protein synthesis or net protein balance. When appropriate, functional and body mass outcomes are also discussed.
ESSENTIAL AMINO ACIDS AND MUSCLE ANABOLISM
Development of an EAA formula was originally predicated on the provision of EAA in proportion to their composition in muscle protein. When compared with traditional proteins, such as whey, the anabolic response of muscle to EAA ingestion, per gram intake, is markedly greater (21). Furthermore, when compared with standard clinical meals such as Boost, EAA elicit a much greater response of muscle protein anabolism. A greater response with EAA is evident despite the greater caloric load and carbohydrate component of Boost (20). This response is due in great part to the rapid absorption and subsequent increase in peripheral AA concentrations with EAA (23). Muscle anabolism resulting from AA administration is due entirely to a stimulation of muscle protein synthesis, with no change in protein breakdown (5). Although this mechanism is consistent with intact proteins, ingestion of EAA alone is a more efficient means of stimulating muscle protein synthesis (21).
High-quality proteins, such as whey, casein, and beef, stimulate muscle protein synthesis in proportion to the amount of EAA per dose (26,27). Generally speaking, EAAs make up approximately 40 to 45% of the total AA in high-quality proteins. Thus, given adequate protein ingestion, the EAA component should be sufficient to stimulate muscle anabolism. While ingestion of intact protein is the most common and convenient format, ingestion of free-form EAA has several added advantages. First, on a gram per gram basis, EAAs elicit a greater stimulation of muscle protein synthesis than an intact protein. To demonstrate this point, muscle protein kinetics were calculated before and for 3.5 hours after the bolus oral ingestion of 15 g EAA or 15 g whey protein. Although both supplements stimulated protein synthesis, net protein balance over the post-supplement period was significantly greater for the EAA group compared with the whey group (21).
The greater increase in protein synthesis is due in part to the larger increase in peripheral AA concentrations resulting from free-form AA ingestion (23). By converting AA uptake to milligram of protein, there was a calculated accrual of 4.0 ± 0.4 g of protein/leg for the EAA supplement versus 2.2 ± 0.3 g protein/leg for the whey protein, indicating that a 15-g EAA supplement provides approximately twice the anabolic stimulus as an isocaloric amount of whey protein. Not only was the EAA mixture more effective but also the efficiency of protein utilization (net protein synthesis/protein [i.e., AA] ingestion) was approximately 1.1 for the EAA mixture as opposed to approximately 0.2 for whey protein. The value of 0.2 for whey protein is consistent with the observation of Hegsted (14) that about 20% of nitrogen intake above requirement for balance is retained in the body. The 4-fold higher ratio for the EAA reflects an efficient reutilization of nonessential AA, which would otherwise have been wasted/excreted, to maintain the stimulated protein synthetic rate.
The consolidation of data indicates a dose response of net muscle protein synthesis to EAA administration (7). We continue to elucidate this relationship in the resting state; however, interpretation of the cumulative data to date is confounded at times due to the presence or absence of carbohydrate and/or exercise. The effects of EAA and exercise will be discussed in detail below. Regardless, it appears that as little as 3 to 4 g of EAA results in a stimulation of muscle protein synthesis and that maximal stimulation is achieved at approximately 15 to 18 g. This relationship is more discernable in young subjects, as we have demonstrated that EAA formulation must be altered slightly to account for changes in synthetic capacity that occurs with aging.
EFFECTS OF AGING ON ESSENTIAL AMINO ACID FORMULATION
In order for EAA administration to maintain its anabolic capacity in older individuals, adjustments in formulation are necessary. Katsanos et al. (15) demonstrated that muscle anabolism was similar in young and older subjects when a dose of 15 g of EAA was given. However, at a lower dose of 7 g, the stimulation of muscle protein synthesis was blunted in the older subjects. The synthetic response was blunted despite the fact that the EAA composition was similar to that found in whey protein. The leucine content of that formula (and whey protein) represents approximately 26% of the EAA total (16). However, when the ratio of leucine was increased to approximately 40% of the EAA total, the older subjects experienced a similar increase in muscle protein anabolism as the young (16).
These data are consistent with studies indicating the importance of leucine in the regulation of key initiation factors in skeletal muscle (1,30). In older rats, a greater amount of leucine was required to stimulate key enzymatic activity (p70S6 K) in the initiation pathway, as well as the stimulation of protein synthesis in skeletal muscle (10). With emerging evidence indicating that certain aspects of the initiation pathway are downregulated with aging (Cuthbertson et al. (9) and Mayhew (19)), increased leucine intake is an effective means of targeting specific signal transduction systems (30).
The effects of EAA and carbohydrate on skeletal muscle also vary with aging. In young subjects, the addition of carbohydrate to AA substantially increases muscle anabolism. However, the same formula given to the elderly provides no additional benefit above AA alone (31). These data indicate that the insulin response elicited by the carbohydrate bolus does not enhance protein anabolism in older muscle and supports the premise that older individuals are more resistant to the effects of insulin on muscle protein metabolism. This diminished response of skeletal muscle was evident in subjects with no clinical signs of metabolic syndrome or insulin resistance. This study was performed in resting subjects; however, recent evidence indicates that exercise combined with a protein and carbohydrate supplement in elderly men results in an increase in muscle protein synthesis (17). To our knowledge, however, the combined effects of exercise and EAA plus carbohydrate supplementation have not been directly compared between young and older subjects.
ESSENTIAL AMINO ACIDS AND EXERCISE
Resistance exercise stimulates both protein synthesis and breakdown; however, protein synthesis is stimulated to a greater degree such that the net protein balance across skeletal muscle (synthesis − breakdown) is improved, though still negative (4). On the contrary, AA administration alone, either by infusion (5) or by oral ingestion (23,28), results in a significant increase in muscle protein synthesis such that the net protein balance becomes positive or anabolic. When resistance exercise is performed in conjunction with AA administration, the response is interactive (5), surpassing the simple additive response of each intervention alone (Figure 1). AA administration in conjunction with exercise results in a large increase in peripheral AA delivery to skeletal muscle (arterial AA concentration × blood flow) (5). The increased blood flow to exercising muscle delivers the higher AA concentrations rapidly to the muscle. It is this extracellular signal, or rapid change in arterial AA concentrations, that serves as the signal for increased synthetic rate (6).
The timing of EAA ingestion in relation to exercise also dictates the magnitude of response. A solution containing 6 g of EAA plus 35 g of sucrose was given to young subjects either immediately before or immediately after resistance exercise. Drink administration immediately prior to exercise resulted in a 160% greater increase in muscle anabolism compared with post-exercise drink administration (29). With post-exercise drink ingestion, the anabolic response was only one-third that achieved when the drink was ingested prior to exercise. The larger response of pre-exercise drink ingestion is due to a 3-fold greater delivery of AA to skeletal muscle. AA delivery with pre-exercise ingestion remained 64% greater than post-exercise ingestion up to 1 hour after the conclusion of exercise (29). These results indicate that the ingestion of AA prior to exercise leads to an increased delivery of AA precursors to skeletal muscle via increased blood flow to the exercising muscle. In turn, the increased delivery signals skeletal muscle to upregulate synthetic mechanisms, resulting in greater AA uptake and muscle protein synthesis.
As previously mentioned, the same dose of EAA can be delivered by ingesting approximately 30 g of high-quality protein. However, the ingestion of 30 g of whey entails a larger caloric load, greater digestion time (slower release of AA to the periphery), and increased ureagenesis (Figure 2) as a portion of ingested AA are converted to urea rather than used for protein synthesis. Tipton et al. (27) administered a drink consisting of either 20 g of casein or 20 g of whey protein 1 hour after a bout of leg extension exercise. Although AA delivery was greater with whey protein, each resulted in a similar increase in muscle protein balance and muscle anabolism. Furthermore, each resulted in a similar AA uptake relative to AA ingestion (∼10-15%) (27). EAA administration results in a more efficient AA utilization for protein synthesis when given in conjunction with exercise. When 12 g of EAA was given after resistance exercise, calculations revealed that ∼27% of ingested EAAs were taken up by skeletal muscle for protein synthesis (8). Because protein synthesis involves both essential and nonessential AA, the stimulation of muscle anabolism by EAA is accomplished by using endogenous nonessential AA. Further evidence is provided by the stable urea production after exercise and the decline in alanine concentration and important urea precursors (8) (Figure 3). Thus taken together, the utilization of AA for protein synthesis is approximately twice as efficient, on a gram per gram basis, with EAA versus intact protein (Figure 4).
Until recently, the compelling data with EAA administration have been somewhat esoteric, given that such a supplement was not commercially available. However, based on this compilation of data, a recent proprietary formula of EAA in free and peptide form is available under the brand name BeneVia (authors' disclosure: AAF and RRW have financial interests in this product). A total of 9.5 g of AA, including 7 g of EAA and with a high proportion of leucine, is present in each 6 oz serving. Thus, based on the compilation of data, drinking two 6 oz servings prior to exercise should provide for maximal stimulation of muscle anabolism.
ESSENTIAL AMINO ACIDS AND INJURY REHABILITATION
Another advantage of EAA administration appears to be their ability to maintain skeletal muscle mass and function during periods of compulsory inactivity, similar to that experienced during post-surgical hospitalization or rehabilitation. We have demonstrated the efficacy of EAA supplementation on muscle mass and function during inactivity. An important finding of these studies has been that the acute response of muscle protein anabolism to EAA intake is translatable and sustainable over time in the catabolic condition of bed rest. Paddon-Jones et al. (22) provided 3 daily drinks of 15 g EAA plus 30 g sucrose for 28 days to subjects undergoing strict bed rest. A second group consumed a placebo drink throughout the same period. The addition of carbohydrate may appear beneficial, as the addition of carbohydrate to EAA enhances muscle anabolism in young subjects (31). However, because bed rest is known to induce insulin resistance in skeletal muscle in as little as 7 days (25), the addition of carbohydrate most likely had no additional effect on protein metabolism, although it did serve to improve drink palatability. While the placebo group lost a substantial amount of muscle mass and strength, the EAA group was able to maintain muscle mass and reduce the loss of muscle strength. This was due to the fact that the stimulation of muscle protein synthesis by EAA was unchanged after 28 days of inactivity (22). Furthermore, when net muscle protein synthesis in response to EAA on the first day of bed rest was extrapolated over 28 days, the predicted gain (or ameliorated loss) of muscle was similar to the measured lean mass by dual energy x-ray absorptiometry.
Although muscle strength decreased in both groups, as expected given the absence of muscular activity, the decrease in the EAA group was one-half that of the placebo group (22). The amelioration of strength loss was due to the preservation of single fiber peak force in type I fibers and peak power in type II fibers (12). It appears that the maintenance of muscle protein turnover results in the production of more functional myofibrils, which in turn preserves muscle function. These data are consistent with Balagopal et al. (3) who demonstrated a relationship between myofibrillar protein synthesis and muscle strength.
The impact of EAA on muscle during inactivity may be due amelioration of impaired signaling. Whereas declines in anabolic signaling due to inactivity are not readily apparent (13,18), inactive muscle seems to be resistant to stimulation by AA. Muscle protein synthesis in inactive muscle does not respond to mixed AA (13). Furthermore, intracellular signaling in response to EAA is impaired in elderly muscle, resulting in a reduced response of muscle protein synthesis (9,16). However, EAA with a higher proportion of leucine overcomes this deficiency (9,16,24). Results from studies in old rat muscle are consistent with this interpretation (10,11). In the rat model, signaling deficits after exercise are ameliorated with leucine supplementation, which in turn leads to a restoration of muscle protein synthetic rates (2). Although this concept has yet to be systematically investigated, these data suggest that EAA may be an important consideration during periods of immobilization or muscular inactivity.
In summary, muscle anabolism and/or muscle protein turnover can be facilitated by the proper dose, delivery, and timing of AA ingestion. Free-form AAs configured to mimic the profile of skeletal muscle AA composition afford the greatest stimulation of muscle protein anabolism. In relation to exercise, there is an interactive effect with AA on muscle protein synthesis. This effect is greatest when formulas are given prior to exercise. Finally, older individuals respond differently than their younger counterparts and require a revised AA profile for optimal response.
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