Arginine

Arginine is an amino acid that is important in the body for a few reasons. It is one of the three substrates to form creatine which is a vital nutrient (deficiency induces mental retardation) and is also used to form agmatine, a signalling molecule in the body. Arginine is an intermediate in both the urea cyle (with L-ornithine, L-citrulline, and arginosuccinate) and the nitric oxide cycle (with ornithine and arginosuccinate),^[5] and vicariously through ornithine it produces polyamine structures which can regulate cellular function.^[6]

It is a fairly popular sports supplement, with one survey noting that 5% and 8% of females and males of NCAA level supplemented L-Arginine.^[7]

Arginine is an amino acid involved in two regulatory cycles (urea and nitric oxide) and can be converted into a few other bioactive molecules such as creatine or agmatine to regulate the body. It is known as one of the more versatile amino acids in the human body

Arginine is known as a conditionally essential amino acid (elevated to 'essential amino acid' status under certain conditions) due to, in otherwise healthy persons, arginine homeostasis being somewhat maintained following prolonged dietary deprivation.^[8]

An arginine deficiency (which can be induced by increasing the activity of the arginase enzyme^[9] that converts arginine to ornithine) impairs B-cell function (immunity)^[10] and impairment of both hair and muscle growth.^[9] Neuromuscular function may also be impaired.^[11]

Protein structures (such as enzymes) that require arginine^[12] or are otherwise rich in arginine^[9] do not appear to be significantly modified.

Arginase is known to be overexpressed in type II diabetics and is a risk factor for development of cardiovascular problems in this population cohort^[13][14] and may be reduced in persons with renal failure.^[15]

Arginine is essential in function, but not necessarily for dietary intake as it is regulated during deprivation. A deficiency can be induced experimentally, and a subclinical deficiency state may exist in renal failure and diabetics (which would increase the need for either arginine of citrulline supplementation)

Arginine is first met with intestinal and splanchic metabolism, in which a degree of arginine is consumed by enterocytes or interconverted into L-citrulline or L-ornithine. Aside from the liver utilizing arginine to a high degree, intestinal uptake of arginine is also poor under normal conditions and increased during various diseases.^[16] It seems that a minimal amount of L-Arginine gets to systemic tissue in relation to other amino acids in the Urea cycle, as supplemental L-Ornithine achieves twice the serum concentration relative to L-Arginine and L-Citrulline 9.3 times the amount. This seems directly related to the degree of hepatic and intestinal metabolism.^[17][18][19]

Dietary arginine accounts for 40-60% of serum arginine, as evidenced by an equivalent drop during arginine free periods. The rate of conversion of L-citrulline into L-arginine seems unaffected by dietary intake.^[20]

Arginine uptake from the diet is somewhat regulated in intestinal cells (enterocytes), and dietary deprivation of L-arginine does not necessarily impair the bodily functions of arginine

Arginine serves a vital role in both the urea cycle and in the nitric oxide cycle (amongst other things; metabolism into creatine, polyamines, or agmatine is not pictured above)

The urea cycle is a metabolic pathway involving the three amino acids Arginine, Ornithine, and Citrulline as well as an intermediate known as Arginosuccinate. This pathway is intimately involved in nitrogen detoxification (reducing ammonia in the body) and producing urea as a byproduct.^[21]

Arginine is converted into L-Ornithine via the arginase enzyme (giving off urea as a cofactor)^[22][23] and from there ornithine (using carbamoyl phosphate as a cofactor) is subject to the Ornithine carbamoyltransferase enzyme to produce L-Citrulline. In this sense, the metabolic pathway from arginine towards citrulline (via ornithine) causes an increase in urea and a concomitant decrease in ammonia, which was used by the Carbamoyl phosphate synthase enzyme to create carbamoyl phosphate.^[21] If need be, arginine can directly be converted into L-citrulline via a Arginine deiminase enzyme to produce, rather than require, ammonia.^[24]

The cycle is formed as citrulline then binds with L-aspartate (related to D-Aspartic acid as its isomer) to form arginosuccinate via the arginosuccinate synthase enzyme, and then the arginosuccinate lysase enzyme degrades arginosuccinate into free arginine and fumarate; arginine then reenters the urea cycle anew.^[21] Fumarate can simply enter the TCA (Krebs) cycle as an energy intermediate.^[24]

The urea cycle is the metabolic pathway that links Arginine, Ornithine, and Citrulline (alongside Arginosuccinate) to one another and regulates bodily concentrations of ammonia and urea

Arginine also has a simple conversion into L-Citrulline via the nitric oxide synthase (NOS) enzymes, of which there are endothelial forms (eNOS) and neuronal specific (nNOS) forms as well as an inducible form (iNOS) that responds to inflammatory signals. The conversion of arginine via NOS enzymes produces nitric oxide as the most relevant byproduct, and citrulline is seen as a byproduct.^[25] Citrulline can then convert back into L-arginine via arginosuccinate, but L-ornithine is not involved in the nitric oxide pathway.

Arginine can convert directly to citrulline to give off nitric oxide as a byproduct. Nitric oxide mediates a fair bit of the effects of arginine supplementation, and at least for these purposes NOS inhibitors abolish the effects

Arginine can be converted to the molecule 4-(aminobutyl)guanidine, commonly known as Agmatine, via decarboxylation^[26] from the arginine decarboxylase (ADC) mitochondrial enzyme.^[27] Despite this, tissue concentrations of the ADC enzyme are not highly correlated with tissue concentrations of agmatine which are fairly widespread^[28][29] likely due to uptake of agmatine from other sources such as bacterial synthesis or the diet.^[26]

This molecule is a signalling molecule which is a ligand at α2-adrenoceptors and imidazoline receptors with enough affinity on the former to displace the reference drug clonidine (an α2-adrenoceptor agonist).^[30]

Agmatine can be derived from L-Arginine, and itself acts as a novel signalling molecule in the brain (mostly interacted with cognition and pain perception)

6g L-Arginine appears to have a bioavailability of around 68+/-9% in humans^[31] while higher doses (10g) have been noted to have lower bioavailabilities at around 20%.^[32]

Both studies have noted a degree of variability, with absolute ranges of 51–87%^[31] and 5-50%.^[32]

Arginine is absorbed in the intestines, and appears to be well absorbed at low oral doses but with gradually higher oral doses the overall percentage absorbed is reduced. The intestinal stage of absorption appears to be a regulatory step

Arginine is absorbed in the intestines by a collection of transporters such as system Y⁺ (handles lysine, ornithine, and arginine preferentially and is sodium independent)^[33][34] of the cationic amino acid transporter (CAT) family,^[35][36] system Y⁺L with the transport protein y⁺LAT1 (handles the same amino acids and is also sodium independent, but requires neutral amino acids for cooperation),^[37][38][39] system B^0,+ (lysine, arginine, valine, alanine and is sodium dependent),^[39] and system b^0,+ with the transport b0,⁺AT (lysine, arginine, and leucine).^[39]

A collection of cationic amino acid transporters handle arginine transport across the intestinal barrier

At rest, serum levels of L-Arginine (prior to supplementation) in a fasted state are 71+/-4μmol/L^[31] or 15.1+/-2.6μg/mL.^[32]

Oral supplementation of 5g Arginine in a fasted state is able to increase the AUC of L-arginine (for 5 hours) by 64%^[40] and increasing the dose to 9g is able to further increase AUC by 181% (relative to placebo); interesting, a bolus of 13g which causes intestinal distress has failed to increase serum levels significantly.^[40]

6g Arginine is able to increase peak plasma arginine by 336% (C_max of 310+/-152μmol/L)^[31]

10g Arginine has been noted to increase plasma arginine from 15.1+/-2.6μg/mL to 50.0+/-13.4μg/mL (331%) at a T_max of 1 hour post oral ingestion (fasted state).^[32]

Supplemental arginine can increase serum arginine acutely which is most evidence by the C_max value (peak concentration in the blood), within an hour or so. There may be a limit cap at around 300% increased serum arginine levels, as higher doses have gradually less absorption

The half-life appears to be dose-dependent, with the most relevant data being a half-life of 76+/-9min following oral intake of 6g Arginine (infusions of 6g or 30g that reach higher concentrations have quicker half lives).^[31]

The half life of arginine with normal supplemental dose is just over an hour (75 minutes or so) and if you somehow manage to get a higher serum concentration (tad difficult since intestinal absorption of arginine is limited and conversion of citrulline into arginine has a rate limit as well) then the half-life will shorten somewhat

Despite oral L-Citrulline increasing plasma L-arginine, supplemental L-arginine does not appear to influence plasma citrulline significantly in otherwise healthy individuals.^[31] In hypertensives, however, such as effect has been noted with 2-4g of L-arginine taken thrice daily.^[41]

At least one study using 3g L-arginine in persons with peripheral arterial resistance noted that supplemental arginine was able to increase plasma ornithine concentrations to a similar level (about 10nmol/L more than baseline).^[42] This is thought to be due to the arginase enzyme (converts arginine to ornithine) having a high K_m value^[43] and is likely not saturated at physiological concentrations of arginine, which would mean anything that increases plasma arginine can increase plasma ornithine.

Long term potentiation (LTP) is a form of synaptic plasticity in which synaptic transmission is enhanced for hours or days in response to repetitive stimulation of presynaptic terminals due to a postsynaptic calcium influx via NMDA receptors from stimulatory neurotransmitters.^[44][45][46] LTP appears to be dependent on the nitric oxide synthase enzyme^[47] and due to hemoglobin also sequestering LTP (hemoglobin cannot enter a cell and sequesters nitric oxide) an extracellular release of NO was determined to be vital.^[46][47] Nitric oxide is released from post-synaptic neurons and returns to the presynaptic neuron to induce LTP^[48] and this may occur directly in hippocampal cells.^[49]

Long term potentiation (LTP; involved in memory formation) involves repeated stimulation of a neuron, and nitric oxide appears to play an important role in this regard as it stimulates the presynaptic (first) neuron to continously stimulate the post-synaptic neuron. This is known as a retrograde messenger

Surprisingly, the neuronal NOS enzyme (nNOS) does not appear to be critical for LTP.^[50] The eNOS enzyme (present in hippocampal cells^[51]) also does not appear to be critical.^[52] However, abolishing both nNOS and eNOS reduces LTP by about 50% and this was not further reduced with NOS inhibitors.^[52]

It has been noted that inhibiting myristoylation (the addition of a myristic acid to eNOS to facilitate its actions^[53]), which inhibits extracellular release of nitric oxide via eNOS, is sufficient to abolish LTP ex vivo.^[54]

There is mixed evidence as to whether eNOS is exclusively required for production of NO involved in LTP or whether nNOS can compensate somewhat for a lack of eNOS activity. Regardless, eNOS and production of nitric oxide from arginine within the cell appears crucial

L-Arginine and L-Citrulline form a cycle with nitric oxide where arginine converts to citrulline via giving off a nitric oxide molecule, and the citrulline may convert to arginine via sequential metabolism by argininosuccinate synthase and argininosuccinate lyase (with L-arginosuccinate as an intermediate).^[55][56] This cycle is thought to be relevant to neurogenesis as the arginosuccinate synthase enzyme (rate-limiting for nitric oxide in many tissues^[57]) appears to be upregulated in the developing brain.^[58]

Nitric oxide itself has been found to regulate neuronal stem cells^[59] and is known to influence synapses in a similar way to brain-derived neurotrophic factor (BDNF)^[60][61] while negatively regulating BDNF release,^[62] suggesting overlap between their roles. In fact, the reduction in neural differentiation seen with inhibiting nitric oxide synthase (and impairing the actions of nitric oxide) is rescued by BDNF.^[63]

Nitric oxide appears to be a regulatory of neurogenesis, and may act on similar mechanisms as BDNF does. However, there is currently no evidence to support the usage of citrulline over arginine or whether supplementation of either of these precursors promotes neurogenesis in a living system

In subjects with high baseline anxiety, a supplemental mixture of Lysine and Arginine (3g each) for 10 days prior to a psychosocial stress test based on public speaking and the trier test^[64] was able to slightly reduce the increase in state anxiety induced by the stress despite increasing levels of catecholamines and cortisol;^[65] the neuroendocrine response to stress is normally impaired in persons with high state anxiety^[66] so this suggests normalization.^[65] A subsequent study using slightly lower doses (2.64g each) in otherwise healthy adults subject was able to reduce resting state anxiety (11%) and prevented the increase in anxiety seen with stress testing.^[67]

The mixture tends to be used since a deficiency in Lysine promote anxiety^[68] and normalizing a lysine deficient diet improves anxiety symptoms in humans;^[69] lysine itself can act upon benzodiazepine receptors^[70] and is a partial serotonin 5-HT4 receptor antagonist^[71][72] (which is useful in stress-induced anxiety).

When studies mention L-arginine, it is mentioned^[73] that nitric oxide modulates ACTH and cortisol and that stress could induce an arginine deficiency. However, due to nitric oxide being a positive modulator of stress-induced anxiety^[74] and no studies linking either amino acid in isolation towards a reduction in anxiety it is uncertain what role arginine plays in anxiety.

A lysine and arginine mixture appears to reduce stress-induced anxiety at reasonable supplemental dosages, but this may simply be a role of lysine supplementation and not arginine (since lysine has some plausible mechanisms and arginine just seems to be semi-useless filler in this regard)

Nitric oxide signalling (and nitroxidative stress) appears to have a role in aging,^[75][76] usually due to excessive inflammatory signalling causing an upregulation of iNOS and then the excessive concentrations of oxidative metabolites (such as peroxynitrate) causing neurotoxicity.^[77] Otherwise, normal physiological concentrations of NO are neuroprotective^[77] and thus NO serves a modulatory role.

When comparing aged rats to young rats, there are reduced levels of L-arginine and its related structures and metabolites (ornithine, citrulline, agmatine, putrescine, spermidine, and spermine) as well as glutmate but not GABA in the hippocampal and prefrontal cortex regions^[78] and less nitric oxide containing enzymes have also been detected in aged neuronal cells in these regions,^[79] these regions are known to be subactive or impaired during aging that is associated with impaired learning.^[80][81]

Nitric oxide signalling appears to be important for cognitive aging, where low levels are neuroprotective but high levels are neurotoxic (the high levels are usually just one of the mechanisms by which excessive inflammation can exert neurotoxicity, may not be a concern with supplementation). The aging brain appears to have reduced concentrations of nitric oxide intermediates and the enzymes involved in this pathway

The main mechanism by which arginine supplementation (and by extension, L-Citrulline supplementation) influences blood health is through being the substrate for nitric oxide synthase (NOS) enzymes to produce nitric oxide from, which then signals via soluble cyclic guanylyl receptors to produce cGMP. The production of nitric oxide and subsequent production of intracellular cGMP underlie a good deal of arginine's benefits.

NOS enzymes come in three main isoforms: ^[82][83] inducible NOS (iNOS), which is created in response to inflammatory stressors, neuronal NOS (nNOS), which was first discovered in neurons and is also at the motor endplates in skeletal muscles, and endothelial NOS (eNOS), which was initially thought to be only in the endothelium but is quite widespread^[84] including brain tissue.^[52][50] 

NOS enzymes work in dimers that are jointed head to head and the catalytic mechanisms is dependent on this dimerization as well as heme, tetrahydrobiopterin, calmodulin, NADPH (as electron donor) and both FMN and FAD.^[85][86][87] As such, NOS enzymes (all three isoforms) are NADPH-requiring flavoproteins.^[88][89][90] Their structure and function is complex (reviewed here^[91]) but there is a basic binding site for arginine and the electrons donated from NADPH cause arginine to convert to citrulline giving off nitric oxide as a byproduct; iNOS exclusively uses and eNOS may also use a free radical intermediate called N^ω-hydroxy-L-arginine (L-NOHA), which degrades to citrulline and nitric oxide in the presence of H₂O₂.^[82][92]

Nitric Oxide Synthase (NOS) enzymes use arginine to produce nitric oxide, and nitric oxide (which produces cGMP) is known to mediate a wide variety of actions associated with arginine supplementation. Although arginine supplementation is thought to increase activity of NOS enzymes (by providing more substrate), simply stimulating the activity of these enzymes is also a feasible option to increase nitric oxide

Increased nitric oxide (usually measured by plasma nitrate/nitrite, citrulline, or urinary cGMP concentrations) appears to be increased with L-arginine in persons with essential hypertension,^[93] artherothrombosis,^[94] and type II diabetes.^[95] Studies in otherwise healthy athletes taking L-arginine are quite mixed; there are instances where biomarkers of nitric oxide metabolism are increased^[96][96] while other studies fail to note any modification.^[97][98][99] Not surprisingly, the benefits associated with nitric oxide do not occur when nitric oxide biomarkers are not increased.

The unreliability of arginine increasing nitric oxide may be due to the physiological concentrations of arginine (40-100µM in extracellular space^[100] and possibly reaching up to 800µM intracellularly^[101]) being enough to saturate endothelial nitric oxide synthase (eNOS) inherently (Usually stated to be a K_m of 3µM^[102][103] but at times is measured as high as 29μM^[104]). This implies that the enzyme is already at maximal efficacy, and that further supplementation does not increase conversion rate (due to a backlog of arginine in serum); the observations that arginine still increases nitric oxide at times (albeit unreliably) is referred to as the L-arginine paradox.^[105][106]

This theory is in line with observations that sometimes nitric oxide metabolism is unaffected despite up to 300% increases in plasma arginine.^[99][107]

Arginine is the substrate from which nitric oxide is derived (supplemental citrulline can also increase nitric oxide as a consequence of increased plasma arginine levels), a signaling molecule that mediates an increase in cGMP, leading to a cascade of reactions that lead to vascular relaxation and blood vessel dilation. Importantly, the proposed mechanism for increased production of nitric oxide from an increased level of serum or intracellular arginine levels, is flawed when enzyme kinetics are taken into consideration. Substrate availability (arginine) is not typically a limiting factor (in normal physiological conditions) in the production of nitric oxide. As such, an excess of arginine, achieved through supplementation, would not yield higher levels of nitric oxide.

One study has noted a transient increase in nitric oxide production that appears to be more like that of an agonist than a substrate^[108] and later it was found that arginine has the ability to active the α2-adrenergic receptors,^[109] which can directly stimulate nitric oxide without requiring converion into citrulline via NOS. However, arginine was fairly weak (outperformed by agmatine)^[109] but this mechanism has not been ruled out yet.

Furthermore, extracellular arginine does appear to be a determinate of nitric oxide release^[104] (the CAT1 transporter that shuttles arginine is highly associated with eNOS^[110] and inhibiting extracellular influx prevents eNOS activation^[111]) while intracellular arginine concentration does not appear to be associated.^[106] Due to the transport being required but intracellular arginine not being per se required, it is thought that the colocalization of CAT1 with eNOS^[110][112] may also play a role in stimulation of eNOS activity.

Theoretically possible that it can directly stimulate nitric oxide independent of being a substrate, but the mechanisms are either not overly potent (α2-adrenergic receptors) or they are currently unexplored in depth (CAT1 transportation)

Asymmetrical dimethylarginine (ADMA) is a methylated arginine derivative and is a competitive inhibitor of all three subsets of the NOS enzyme^[113] and appears to be elevated in instances of chronic renal failure^[113] and intermittent claudication.^[114] It correlates well with severity^[115] and occurrence^[115][116] of cardiovascular disease, and is seen as a negative stressor for blood flow (as infusions of ADMA into people immediately cause peripheral resistance and a reduction in blood flow^[117][118]). In regards to the aformentioned 'arginine paradox', it is thought that the ratio of arginine to ADMA can explain some of the effects of arginine on nitric oxide metabolism.^[105]

Monomethylarginine is another methylated derivative that is also derived from arginine which is a competitive NOS inhibitor, and symmetric (rather than asymmetrical) dimethylarginine does not appear to be an inhibitor.^[119]

It is metabolized by the enzyme dimethylarginine dimethylaminohydrolase (DDAH, two tissue-specific isoforms^[120]) and instances where this enzyme is reduced cause an increase in ADMA production^[119] while overexpression reduces ADMA and causes vasodilating actions similar to supplemental L-arginine.^[121] Stressors that impair DDAH include homocysteine,^[122] elevated blood cholesterol and triglycerides,^[123][124] and elevated blood glucose^[125] all vicariously through increased oxidative stress.

ADMA is a methylated metabolite of arginine, and appears to act in opposition to arginine by inhibiting the actions of the NOS enzyme and subsequent nitric oxide production. Excessive levels of ADMA can occur from oxidative stressors decreasing the activity of the enzyme that degrades it, and reducing ADMA causes vasodilation due to nitric oxide production

Studies that assess the influence of supplemental arginine on plasma ADMA fail to find an influence of 3g arginine over 6 months (persons with peripheral artery disease^[42]) nor 7 days of 12g arginine AAKG^[126] or 3g L-arginine^[127] in otherwise healthy persons. Citrulline also appears to be implicated in not increasing ADMA,^[128][127] although a lone study has noted that in persons with mild hypertension an increase in ADMA was noted after 28 days of 6-12g L-arginine.^[41]

It should be noted that increasing the arginine:ADMA ratio has failed to be associated with blood flow improvement^[127] yet one study where negative effects were seen with prolonged L-arginine supplementation it was not determined to be associated with ADMA either.^[42]

Although most evidence suggests that ADMA is not increasd with L-arginine supplementation (these studies noting that the Arginine:ADMA ratio is enhanced due to increases in plasma arginine), there is limited evidence to suggest an increase that requires further investigation.

Blood pressure and peripheral resistance have been noted to be decreased in healthy individuals following a 30g infusion of arginine (reaching a serum concentration of 6223+/-407μmol/L) by 4.4+/-1.4% and 10.4+/-3.6% respectively, but oral intake of 6g arginine (822+/-59μmol/L) has failed to do anything.^[31]

Oral supplementation of a combination supplement of 4g citrulline with 2g arginine in hypertensives has been noted to, over 6 weeks, reduce both systolic and diastolic blood pressure (as well as mean arterial pressure) in both the ankle and brachial measurements without affecting heart rate.^[1]

Oral supplementation of arginine (citrulline also applies here due to increasing plasma arginine) is able to increase blood flow in persons with impaired blood flow, and although it has the potential to reduce blood pressure it seems a tad unreliable and may only occur in hypertensives

In peripheral artery disease, an infusion of arginine (30g over 30 minutes) is able to double blood flow to extremities and performed equally to prostaglandin E1, a vasodilating drug, and was associated with an increase in nitric oxide and cGMP;^[129] these benefits were later noted with oral supplementation of 8g arginine twice a day (total dose of 16g) which improved pain-free and total walking distance in persons with intermittent claudication by 230+/-63% and 155+/-48% respectively (outperforming 80mcg PGE1).^[114]

In persons with intermittent claudition, oral L-arginine supplementation (3g) for 6 months has actually resulted in a reduction of flow-mediated vasodilation and impairment of endurance on a treadmill test (11.5% improvement in the arginine group, 28.3% in placebo).^[42] It was not sure why this occurred, but the authors suggested development of a tolerance to arginine^[42] (based on the notion that nitric oxide donors cause nitrate tolerance^[130]) and another letter^[43] suggested that the increase in ornithine could have produced polyamines which induced vascular remodelling (less vessel elasticity results in less responsiveness to vasodilating stimuli^[131][132]).

There is mixed evidence as to the effects of arginine supplementation on blood flow in persons with peripheral resistance or intermittent cladication, with short term studies noting a benefit and longer term studies noting an impairment

Arginine supplementation appears to have protective effects on pancreatic β-cells as assessed by animal models of alloxan toxicity^{[133][134][135]} associated with β-cell neogenesis.^[133] These protective effects are associated with nitric oxide metabolism (as the protection against alloxan is mimicked by sodium nitroprusside and abolished by NOS inhibitors^[135]) despite excessive nitric oxide levels being known as cytotoxic.^[136][137]

Furthermore, arginine (via nitric oxide^[138]) can directly stimulate insulin release. Due to the combined protective effect on the pancrease and acting as a secretogogue, arginine is thought to be somewhat protective against diabetes.

Arginine supplementation is protective of pancreatic β-cells and is an insulin secretagogue (causes secretion of insulin from β-cells)

Improved glucose handling has been found as assessed by post-meal insulin release (30 minutes) with a concomitant decrease in postprandial insulin, but no alterations in fasting glucose, seen over 18 months supplementation of 6.4g arginine.^[139]

At least in persons with impaired glucose tolerance, arginine supplementation can restore insulin secretion over a prolonged periods of time

The arginase enzyme has been found to be increased in diabetic persons associated with cardiovascular impairment^[13][14] and is thought to be a reason for the reduced global arginine bioavailability ratio ('Arginine bioavailability ratio' refers to the total left after dividing arginine concentrations by the sum of ornithine and citrulline; it is a diagnostic marker for cardiovascular disease^[140][141]) as increased arginase activity can redirect arginine away from the nitric oxide pathway and towards the urea cycle.

As reduced nitric oxide bioavailability is causative of endothelial dysfunction, it is thought that supplementing this deficiency of nitric oxide with supplemental arginine can confer a cardioprotective effect in diabetics.

Diabetics at increased risk for cardiovascular disease appear to have less arginine in their body relative to citrulline and ornithine, due to increased activity of an enzyme that directs arginine away from nitric oxide production and towards the urea cycle. This causes a drop in nitric oxide, and potentially increases the risk for cardiovascular diseases

Supplementation of 6g L-arginine daily for 2 months in persons with both diabetes and peripheral artery disease (impaired blood flow) has failed to influence HbA1c and glucose despite improving total antioxidant capacity and nitric oxide.^[142]

21 days of arginine supplementation (8.3g daily) in type II diabetics who are already on a hypocaloric diet and an exercise routine noted that supplementation had added benefits on endothelial function and oxidative stress^[95] and usage of 6.4g for 18 months is associated with a greater chance for persons with impaired glucose tolerance to regress into normal glucose tolerance (however, it is not associated with reduced diabetes risk);^[139] this latter longterm study did not confirm benefits to blood flow at rest.^[139]

Arginine appears to either inherently improve diabetic status when supplemented or to augment the benefits of a diet and exercise routine

Arginine has been noted to reduce the oxygen cost of exercise as assessed by cycle ergometry, which is associated with an increase in time to exhaustion.^[96] 

For studies that assess blood flow or pressure during exercise, there are either no significant differences^[97] or a reduction in blood pressure may occur.^[96]

L-Arginine is marketed as being able to (secondary to nitric oxide) enhance muscular blood flow,^[143] studies investigating this have noted that despite plasma arginine increasing 300% following an oral dose of 10g there was no significant influence on nitric oxide metabolism nor blood flow^[98] and elsewhere 6g arginine has been noted to increase muscular blood flow (but not oxygenation) which was correlated with nitric oxide production.^[99]

1mM L-arginine in isolated muscle cells is able to enhance myotube and nuclei density as well as nuclear fusion, which was mediated via nitric oxide.^[144]

In a study where nitric oxide was not influenced (despite plasma arginine increasing 300%), there was no significant influence on muscle protein synthesis.^[98]

Supplementation of arginine to rats is able to increase post-exercise urinary nitrate, indicative of nitric oxide production.^[145] Increases in nitric oxide production (either urinary or serum nitrate) have also been confirmed in humans following either oral intake^[96] or intravenous infusion.^[146]

Increases in nitric oxide production are not always seen (even despite increases in plasma arginine^[107]), suggesting NOS enzyme activity could be a limiting factor.

Increased nitric oxide production during exercise does appear to occur with L-arginine supplementation, although it is not 100% reliable

For acute studies (taking a single dose of L-arginine prior to exercise) 3g of arginine (as AAKG) has failed to benefit weight training,^[147] 6g L-arginine for 3 days has failed to modify cycle ergometer results in judo athletes^[107] while a similar protocol in trained cyclists has noted improved time to exhaustion (25.8%).^[96] 

Some studies have used a form of arginine known as GAKIC (Glycine L-Arginine α-Ketoisocaproic acid) and have noted an increase in mean power output during 10s sprints on a cycle ergometer (with 11.2g GAKIC)^[3] and 10.5+/-0.8% increase in work volume and increased fatigue resistance (28%) on a knee extension test.^[4] These studies, however, are confounded with both the inclusion of glycine and that of the leucine metabolite α-Ketoisocaproic acid.

Unreliable benefits with L-arginine and acute exercise, as it appears that some studies fail to find a benefit with supplementation over 1-3 days (although these studies also fail to note an increase in nitric oxide production despite an increase in arginine concentrations)

For more chronic studies, supplementation of L-arginine (as asparate) conferring 2.8g or 5.7g arginine daily for 4 weeks has failed to modify performance or other biomarkers^[148] with a previous study over 2 weeks with similar methodology also failing.^[149]

Limited studies have been conducted on longer supplementation periods of L-arginine with exercise, they similarly show a lack of overall effects with dietary supplementation

Overall, when looking at reviews or meta-analyses on the topic of L-arginine and sports performance it is noted that it is promising but lacks enough consensus to recommend as an ergogenic.^[143][150]

Arginine does have promise, but due to its unreliability there are differences between persons on whether or not it is recommended

During exercise, although one study using 3g L-arginine (confounded with 2,200mg L-ornithine and 12mg Vitamin B12) for 3 weeks noted a 35.7% increase in exercise-induced growth hormone secretion (which normalized within an hour)^[152] other studies note the opposite; arginine supplementation has been noted to result in a lower growth hormone spike during exercise relative to exercise alone^[153][154] that although it seems to affect youth more than older persons^[155] it is said to influence both age groups.^[153] The magnitude of suppression (assuming 100% is baseline) has been noted to be around a 300-500% increase seen with exercise being attenuated to 200%.^[153]

It is possible that an excessive increase in growth hormone stimulates autonegative feedback, which would explain how older individuals are less sensitive to this suppression as they inherently have less GH spikes from exercise relative to youth.^[156] Finally, due to the spikes in growth hormone normalizing within a few hours anyways^[152][156] it is unsure exactly how much of a concern this suppression is (since 24-hour growth hormone concentrations are more relevant).

Although the evidence is not unanimous, it appears that supplemental L-arginine prior to physical exercise is able to suppress the exercise-induced spike in growth hormone concentrations somewhat. It is not sure exactly how significant this information is, since despite being suppression it is still a short spike

At rest, supplementation of 5-9g L-arginine is able to cause an increase in peak growth hormone concentrations (34.4-120% increase) while 13g was ineffective due to intestinal distress preventing absorption of the L-arginine.^[40]

For studies measuring 24 hour GH secretion, there have been no significant alterations with twice daily dosing of 2g^[157] or acute doses of 5g.^[155] This is potentially related to a known phenomena of growth hormone autonegative feedback,^{[153][158][159]} and a similar modulatory effect on growth hormone is also seen during sleep restriction (where a reduction of sleep-induced growth hormone release is compensated for during daylight hours).

For studies not using arginine alongside exercise, it appears that arginine may be able to acutely increase growth hormone spikes but doesn't appear to effective at increasing whole-day GH levels. Due to whole day levels being more important than acute for the majority of purposes, the importance of arginine supplementation is questionable

High dose arginine (250mg/kg of arginine aspartate daily, around 17.5g arginine) has been noted to enhance the slow-wave sleep GH pulse by approximately 60%, despite not having sufficient influence on waking GH concentrations.^[160] It is unsure how this large increase affects whole-day growth hormone concentrations.

Arginine prior to sleep is able to enhance the sleep-induced GH release, but the long-term significance of this is not known (it is possible that it is still subject to autonegative feedback)

When 3g of Arginine paired with 2.2g Ornithine is taken prior to exercise, no augmentation of the exercise-induced increase in testosterone is seen in weight trained males.^[152]

When 3g of Arginine paired with 2.2g Ornithine is taken prior to exercise, no influence on circulating insulin levels are seen in weight trained males.^[152]

When 3g of Arginine paired with 2.2g Ornithine is taken prior to exercise, no augmentation of the exercise-induced increase in cortisol is seen in weight trained males.^[152]

Arginine has been observed to accelerate lympocyte (immune cell) proliferation and wound healing rates.^[161][162] This mechanism appears to be mediated via activation of NO-synthase^[163][164] and has been seen with another intermediate of Arginine metabolism, ornithine.^[165]

Arginine seems to have increased intestinal uptake when paired with a salt such as alpha-ketoglutarate.^[166]

The mechanism by which it works should extend to the chloride salt (Arginine hydrochloride) as well as the salts aspartate, pyroglutamate, and malate as evidenced by experiments with L-ornithine.^[167]

Supplemental L-Citrulline has been touted to be an alternative to L-arginine due to it circumventing the poor absorption and then converting to L-arginine in the kidneys. L-Citrulline technically follows dose-dependent increases in serum L-arginine up to 15g, but the higher oral dose of Citrulline taken has continually less returns (ie. for every 5g more citrulline you ingested you will add less arginine to serum).^[168]

Oral citrulline at 0.18g/kg has been noted to approximately double plasma arginine (100% increase)^[169][170] or slightly higher (123%) with 0.08g/kg.^[171] As higher doses of L-citrulline have less conversion into arginine^[168] it is unlikely that supplemental L-Citrulline can be used to outperform arginine for acute increases in serum arginine.

The studies that have directly compared L-arginine against L-citrulline have noted both increase the C_max to comparable levels at similar oral doses (C_max of 79+/-8μM for 3g citrulline and 84+/-9μM for arginine) yet citrulline results in a greater overall AUC (48.7% more than arginine).^[127] This may be due to how, even up to 15g citrulline ingestion, no significant upregulation of citrulline excretion occurring.^[168] No increase in the elimination of L-citrulline from blood despite supplementation would allow a pool of L-citrulline to be available for on-demand conversion to L-arginine.

While Arginine still appears to outperform citrulline in regards to causing a peak increase in plasma arginine concentrations, it appears that Citrulline is more effective at increasing overall bodily exposure to arginine (assessed via the area under curve or AUC)

For this reason, supplemental L-arginine may be useful for instances where only an acute increase in required (trying to increase a sleep-induced growth hormone pulse or before exercise) yet if higher serum arginine levels are wanted throughout the day (erectile dysfunction or cardiovascular health) citrulline would be preferrable

L-Lysine is an essential amino acid, and appears to be synergistic with L-Arginine for increasing growth hormone.

1200mg of lysine (as hydrochloride) paired with 1200mg arginine (as 2-pyrrolidone-5-carboxylate) was able to increase secretion of insulin and somatotropin which promoted growth hormone secretion (peak value was 90 minute after ingestion and reached 7.94-fold baseline levels; there was no placebo for comparison), it was said to be synergistic as either agent alone at the same dose failed to promote growth hormone secretion, and measurements lasted 120 minutes.^[172] 

A later study using 3g of L-arginine paired with 3g L-lysine twice daily (12g total) for 12 days in both youth and elderly men has failed to find appreciable increases in 24 hour AUC measurements of growth hormone^[2] and this failure has been replicated in a study using 2g of the three amino acids L-Arginine, L-Ornithine, and L-Lysine in weight trained men.^[157]

Although technically they appear to be synergistic (based on one study without a placebo control, so it has its flaws), this synergism is only acute and has failed to manifest in day-long GH measurements. Despite the synergism, the combination still may not be an appreciable way to increase growth hormone.

The observed safety limit, the highest dose in which one can be relatively assured that no side effects will occur over a lifetime, has been suggested at being 20g of arginine a day in supplemental form (above food intake).^[173] Higher doses have been tested and well tolerated, but no evidence exists to suggest their safety in all populations across a lifetime.

L-arginine has a fairly poor gastrointestinal uptake rate. It also may act as an absorbagogue, releasing water and electrolytes into the gut lumen via nitric oxide stimulation and inducing gastric upset and diarrhea.^[16] This is known as osmolitic diarrhea, and tends to occur at oral doses above 10g or so when taken as a bolus.^[40]

This is thought to occur via stimulating nitric oxide production, as D-Arginine (unable to produce NO) does not produce diarrhea^[174] and nitric oxide itself is known to be a mechanism by which many osmolytic laxatives work.^[175]

Single boluses of 5-9g L-arginine without food do not appear to cause intestinal distress like doses above 10g do,^[40] suggesting that at least for an empty stomach dosing 9g is an upper limit.