Quick Hit Summary

Overall, arginine supplements fall far short with respect to enhanced blood flow and NO production. Although one study did show enhanced 2-3x greater muscle blood volume between training sets following acute L-arginine supplementation, various question marks remain in the study. There is some promise with respect to citrulline enhancing blood flow/NO production. One study did report citrulline improving erection hardness in men, possibly related to improved bloodflow. However, there have been few studies that have examined its blood flow enhancing properties in exercising athletes.

Nitric Oxide (NO) Article Series

In this series of articles we're going to discuss the in's and out's of the NO Supplements; covering everything from basic NO physiology to the effects of the NO amino acids in isolation, or as part of propriety blended "NO Boosting" supplements. In case you missed any of the articles, I recommend checking them out.

• Nitric Oxide Supp's Part I – Understanding the Key Players
• Nitric Oxide Supp's Part II – Let's "Pump You Up"… or Not?
• NO Supp's Part III – The "Everything But The Kitchen Sink" NO Supplements
• NO Supp's Part IV – Training Age & Supraphysiological NO Spikes
• Still to Come: NO Supp's Part V – True Potential Benefits of Citrulline Supplementation
• Still to Come: NO Supp's Part VI – True Potential Benefits of Arginine Supplementation

Get Your Swell On!

Figure 1. Are nitric oxide/ "muscle pump" supplements the key to getting that shredded/veiny body you so desire? Image source¹

One of the most popular ergogenics in the neutraceutical industry today is the famous (or, should I say infamous?!?) Nitric Oxide (NO) supplements. According to manufactures, who are – as we all know – always 100% honest in their advertisements (note sarcasm!); these supplements increase blood flow to exercising muscles. As a result, the delivery of vital nutrients as well as removal of metabolic waste products is supposed to be enhanced, thus leading to greater gains in the gym. If that wasn't enough, due to the heightened blood flow, one's muscles will have the veiny "muscle pump" look that's sought after by many hardcore resistance training enthusiast. Now, as awesome at that may sound – it does still beg the question: Are these claims true? Will NO supplements spike muscle blood flow, allowing you to "get your swell on" like never before…. Or, are these statements nothing more than marketing BS? "

In Part 1 of our nitric oxide supplement series we started to answer the above questions by examining their main NO amino acids (arginine, citrulline) and their physiological relationship with nitric oxide. Now that we understand the basics, its time to shift our focus to the question we originally asked ourselves, "Do these supplements actually increase muscle blood flow?"

Effect of Arginine, Arginine Alpha-ketogluterate (AAKG), or Citruline on blood flow and/or Nitric Oxide Production

Arginine

Figure 2. The production of Nitric Oxide. NOS = Nitric Oxide Synthase. Image created by Sean Casey

The amino acid that has been studied to the greatest extent, with respect to nitric oxide production, is arginine. After reading Part 1 of our series, this makes perfect sense. As seen in Figure 2 nitric oxide is directly synthesized out of arginine. It therefore seems reasonable to assume that if we ingest more arginine, more will be available in our bloodstream, leading to greater availability for NO production and enhanced blood flow. [I caution against this kind of simplistic thinking though. After all, we have seen in Part 1 dietary arginine gets processed pretty heavy before it hits general circulation.]

Did you know … There is another way in which arginine influences NO production outside of its direct role depicted in Figure 2. Unbeknownst to many individuals, insulin stimulates NO production.¹³¹⁴. As far back as the 1960's and 70's, scientists have shown that when given via IV, arginine increases blood insulin levels. In 1997, Giugliano et al conducted an interesting study that examined this phenomenon. In a group of healthy individuals, they demonstrated that when administered 30 g of arginine via IV, blood insulin levels spiked and leg blood flow increased by >35%.⁵ However, when the same group of individuals received 30 g of IV arginine plus a few substances that prevented insulin spikes, the enhanced leg bloodflow was only ~20% compared to the "normal arginine IV" condition. Don't be fooled though in thinking that your dietary arginine will necessary spike your insulin levels (and thus muscle blood flow). In a 2002 study, Gannon et al gave 9 healthy individuals between 8.4–12.5 g of arginine (1 mmol/kg lean body mass) and measured the insulin response over the ensuing 2 hours. Results from this study indicated that when taken on its own, dietary arginine does not stimulate insulin secretion.¹⁵

OK – Enough talk about theories, let's look at the studies…

Arginine – Untrained Individuals

In a randomized cross-over designed study, Fahs et al examined the effects of 7g L-Arginine vs. placebo in a group of 18 healthy young men (mean age – 24) on blood pressure, blood flow as well as arm circumference following a 20 minute upper body lifting session². For reference, the lifting session consisted of 4×5 on the bench (80% 1RM) and 4×10 of arm curls (70% 1RM). The arginine/placebo was taken 30 minutes prior to the start of the training session. Study results indicated that arginine supplementation DID NOT increase arm circumference immediately following the training session. Furthermore, arginine DID NOT enhance blood flow when measured within the first 25 minutes post exercise (~50-75 minutes post ingestion).

Similarly, Robinson et al, had 6 individuals ingest an arginine supplement 30 minutes after 1 hour of cycling at 70% peak aerobic capacity or intermittent squatting (25% bodyweight, 20 reps followed by 3 min rest, repeat 15x). Results indicated that arginine DID NOT enhance bloodflow during the subsequent 3 hour rest period during either of the conditions.³ For the record, blood flow measurements were taken every 30 minutes during this time span. In contrast to Fahs et al², Robinson et al's study involved 10 g Arginine + 70 g CHO vs. a "placebo" consisting of 10 g Glycine + 70 g CHO.

As a side note, the study by Robinson et al may not have used the best "placebo". Glycine is not an inert substance in the body. Interestingly enough, it has been shown to enhance blood flow in animal based in-vitro studies⁵. I do point out that in-vitro animal studies are a long way from real world muscle pumps that many trainees seek! That said glycine is also an ingredient of another purported and – contrary to arginine – established NO "Muscle Pump" booster, Glycine Propionyl-L-Carnitine, (GPLC).¹⁸¹⁹ Is this due to the glycine component? Keep that question in the back of your mind, as we'll tackle this question in a future article in our NO series. What's important to remember in the current context is that the choice of an "active" placebo renders the results of this and other studies questionable to a certain degree.

In a 3rd study, involving 8 recreational males, Tang et al sought to find if the addition of 10 g of arginine to a drink already consisting of 10 g of Essential Amino Acids (EAA's) affected muscle blood flow following a bout of resistance training.⁶ In the study, participants completed a ~ 25 minute bout of unilateral leg training that consisted of 5 sets of 8-10 reps on both the leg press as well as leg extensions. This training session was completed 2x, one of which was followed by the arginine based drink and the other by a "placebo" drink that consisted of 10 g EAA's + 14.7g glycine. Final results indicated that the addition of arginine to the EAA drink, DID NOT further enhance muscle blood flow vs. the glycine "placebo" during the 120 minutes that followed. For reference, blood flow measurements were taken at 0,15,30, 60, 90, 120 minutes post ingestion.

Arginine – "Trained" Individuals

One of the few studies examining the affects of arginine supplementation in elite athletes was conducted by Liu et al⁷. In their crossover designed study, 10 elite male judo athletes (mean age – 20 years) took either 6 grams of arginine or a placebo for 3 days. On the morning of the 3rd day, following an overnight fast, athletes took their final dose before engaging in an interval based cycling session that involved 13×20 seconds of max effort bike sprints. Between each interval, participants rested for 15 seconds. Although the research team did not measure muscle blood flow, they did assess NO production via measurements of serum nitrate/nitrite levels. (Both nitrite and nitrate are byproducts of NO metabolism. Thus, increases in blood nitrate/nitrite are generally accepted as measurements for enhanced NO production and collectively abbreviated as NOx). Blood samples were taken after the 9th set, along with 0, 3, 6, 10, 30 and 60 min after exercise (For reference, this corresponds to 60, 65, 70, 73, 76, 80, 100, 130 min post supplementation). At all time points, NO SIGNIFICANT DIFFERENCES were present between trials with respect to serum nitrate/nitrate levels. From a performance standpoint, peak and average power during the intermittent cycling test did not differ from one another.

Figure 3. Ranges in Muscle Blood Volume (ie – difference between minimum and maximum values) after placebo or arginine supplementation. Arginine was significantly different during the 1st and 3rd rest interval vs. placebo. MBV = Muscle Blood Volume. Image created by Sean Casey.

The most recent study to examine the effects of L-Arginine supplementation on muscle blood flow showed surprising results – ARGININE DID INCREASE muscle blood volume. In their crossover designed study, Alvares et al. provided 6 grams of arginine or a placebo to 15 resistance training males (mean training age – 5.9 years).⁸ Changes in muscle blood volume were continually assessed for a 60 minute time period that took place 30-90 minutes post ingestion. At the 80 minute marker, the research team had study participants perform 3×10 of maximal effort arm curls/extensions using an isokinetic machine (Yes, I know, a rather wimpy training protocol ;-). Including the 2 minute rest intervals between sets, the "training session" lasted a total of 10 minutes. During this 2 minute rest intervals, the research team observed 2-3x greater muscle blood volume when taking arginine vs. placebo.

Hmmm, what's the reason for the contrasting results? The research team itself was surprised at the unexpected findings.⁸ However, Alvares et al hypothesized that it may have been related to the actual timing of the measurements as theirs was the first one to actually measure blood flow during exercise vs. after an exercise session. Was the enhanced muscle blood volume caused by spiking NO levels? Well, that's a little harder to assess as NOx was actually the same between both groups leading one to speculate that arginine may not have actually increased NO production. Yet, as stated by the authors, the measurement techniques used by the research team may not have been able to adequately measure local production of NO in the exercised muscle groups.

A final thought on the Alvares et al study… It's important to note that despite enhanced blood volume, no significant differences in performance were noted between the two groups with respect to peak torque, total work, or resistance to fatigue.⁸ Also worth pointing out is that the study's participants only took an acute dose vs. chronic dose which likely affects results.

Supplemental Arginine Alpha-ketogluterate (AAKG)

Although some supplements contain basic L-arginine, if you check the labels of many NO supplements, you'll see that they contain an "improved" form – Arginine alpha-ketogluterate (AAKG). However, is there much research to actually support the idea that AAKG is any better than regular arginine with respect to enhancing blood flow?

The study most often cited by supplement companies in promoting their AAKG spiked NO supplements was completed by Campbell et al.⁹ In their study, 35 resistance trained males (>1 year of training) consumed either 3 g of AAKG (consisting of 1.5 grams of each arginine and AKG) or a placebo 4 times daily in conjunction with a 8 week total body resistance training program. Final results of the trial showed that those consuming a daily dose of 12 g AAKG experienced a significant increase in 1RM bench press vs. the control group (19 lb increase vs. 6 lb increase). Holy cow that is a huge difference in bench press!!! Surely a jump of that magnitude in the bench HAS TO BE the direct results of the "muscle pump" brought on by AAKG supplementation – right? Well…

First – No actual measurements of blood flow and/or NO production were actually measured anywhere in the study. Thus, it provides no actual evidence that AAKG actually induced a "muscle pump". Rather ironic that supplement manufactures often forget this detail when referencing the study!?!?

Second – The thing that strikes me odd about the study is that the training program added 6 lbs to bench press of control group and 19 lbs to that of experimental group WITHOUT any changes between groups in lean tissue gain over the 8 week training period. Although both groups gained ~1.5 lbs in lean body mass, the study did not report if this was or was not statistically significant vs their baseline level. A 19 lbs increase in bench press without a significant increase in lean body mass could be expected with individuals new to resistance training (due to the neural/ "newbie" effect), but all these individuals had been resistance training > 1 year for 4 +/- 1 d/wk (6 + /-3 hr/wk). One could say, "Maybe these lifters hadn’t bench pressed much in their training…" Therefore, it’d be feasible to experience these types of gains based off neural affects. However, how many guys do you know with training experience similar to that reported by the study's participants without spending significant time bench pressing ?!?

Third -The training protocol was by no means a “strength” focused one. During weeks 1-4 it was 3×10; followed by 3×8 for the remaining 4 weeks. If the groups would have been following a program with a Powerlifting emphasis, then yes, MAYBE these changes in bench press 1RM could be expected…

Fourth – It would have been interesting to see what their original bench press 1RM was for each group. In research practice, its standard practice to report baseline levels when reporting on a given variable. Being that they never reported on this value, it makes me wonder if one of the groups had a significantly greater/lower starting point vs. the other group as this may indicate a greater potential for strength gains…. On a similar note, they never actually state the “training age” of each group. All we know is that each participant had > 1 year lifting experience. If the training age was significantly different between groups, then the ability to adapt and improve strength levels to the degree of difference observed in this study would have to be taken into consideration.

Another perspective on the Campbell et al.⁹ study … My friend Adel Moussa from SuppVersity , offered another hypothesis that MAY partially explain the results observed by Campbell et al. The alpha ketogluterate portion of AAKG can replenish glutamate. This latter compound is able to combine with nitrogenous waste produced by exercising muscles such as ammonia to form glutamine [20]. Upon formation, glutamine acts as a "shuttle", carrying the nitrogenous "dirt" from the muscles to the liver for removal via the urea cycle. Additionally the arginine portion of AAKG also plays a role in assisting the urea cycle as described in part 1 of this series. This 1-2 punch of glutamine + arginine may allow for greater removal of metabolic waste products brought on by the higher rep/bodybuilding type of training routine seen in this study. That said, he agrees with me 100% on 2 things…. 1) The gains observed in this study have nothing to do with increased NO production brought about by AAKG supplementation. 2) The fact that original bench press performance was not listed makes the study's results relatively worthless.

Lets move on now to the only study that I'm aware of which actually assessed the effects of AAKG on blood flow/NO production in exercising individuals…

In a study completed by Willoughby et al, researchers compared the affects of AAKG vs. placebo on blood flow, and NOx production in 24 resistance trained men following a bicep training session.¹⁰ Similar to the aforementioned study by Campbell et al⁹, the investigators in this study had participants consume a total of 12 grams of AAKG for 7 days (4 grams, 3x on days 1-6; 6 g, 2x on day 7). On the 7th day, individuals completed an exercise protocol that consisted of 3×15 reps of preacher curls at an intensity of 70-75% 1RM. Between each set, participants rested for 10 seconds. Blood flow and NOx was assessed 2-3 minutes both before and after the preacher curls as well as 30 minutes post exercise. Final results of the study indicated that AAKG did NOT increase either blood flow or NOx production.

Citrulline

Figure 4 The Citrulline-NO cycle. NOS = Nitric Oxide Synthase; ASS = Arginosuccinate synthetase; ASL = Arginosuccinate lyase. Image created by Sean Casey

If our end goal is to increase blood arginine levels, thus giving us more starting material for NO production, one should theoretically turn their focus to citrulline. In contrast to arginine, very little of this amino acid is removed by the liver and the gut before it hits general circulation. Thus, it can later be converted to arginine by the cells that line our blood vessels.¹¹¹² (For greater discussion on this, I refer you to Part 1 of this series as well as Figure 4).

Unfortunately, to my knowledge there has only been a couple of studies that have examined the potential NO enhancing properties that citrulline may have during exercise. The first study worth looking was a 2010 study completed by Sureda et al which examined the effects of 6 g citrulline malate on NO production in a group of 17 semi professional cyclist.¹⁶ Two hours prior to riding a ~ 85 mile (137 km) cycling stage, participants ingested the placebo or citrulline-malate. Upon completion of the exercise session, the research team found that citrulline significantly increased NOx production vs. baseline levels. This was not observed in the placebo group.

Another study of interest, conducted by an Italian research team, provides indirect evidence of potentially heightened penile blood flow in men while having sex; Being that some may consider sex a form of exercise ;-), let's take a closer look at this study …..

The 2 month pilot investigation, conducted by Cormio et al., involved 24 middle aged men (mean age – 56.5 +/- 9.8 years) who reported "mild to moderate reduction of penile rigidity that still allowed some kind of vaginal penetration but not satisfactory penetration and/or completion of successful intercourse."¹⁷ All participants followed the same protocol:

• 1st Month: Placebo

• 2nd Month: 3 g/day of l-citrulline (divided over 2 doses)

Figure 5. Effects of citrulline on erection hardness in men with mild to moderate erectile dysfunction. Differences between those in placebo and citrulline are statistically significant. Image created by Sean Casey

As seen in Figure 5, at the end of the first month, only 2/24 participants (8%) reported having normal erection hardness which was defined as "Penis is completely hard and fully rigid". However, while taking 3 g of l-citrulline per day, 12/24 individuals (50%) reported having normal erection hardness. If one assumes that increased hardness = increased bloodflow, this provides evidence that citrulline may have lead to a very beneficial muscle pump!

The "Kitchen Sink" NO Supplements

I know what you may be thinking right now … Yes, NO supplements contain Arginine, AAKG and Citrulline. However, most of them also contain 30 bazillion other ingredients that are "synergistically blended for enhanced NO production". Surely these supplements are truly able to produce a muscle pump"

Are these supplements, which I like to refer to as "Kitchen Sink" supplements any better at producing the ultimate muscle pump that you desire? With respect to enhanced blood flow, these supplements do not appear to be any more beneficial than the individual amino acids. In Part III of this series, we'll take a closer look at these supplements.

Bottom Line

Overall, both arginine and AAKG fall short with respect to enhanced blood flow and NO production. Although one study did show enhanced blood flow between training sets with L-arginine supplementation, various question marks remain in the study. There is some promise with respect to citrulline enhancing blood flow/NO production. However, there have been few studies that have examined its blood flow enhancing properties in exercising athletes.

For those wondering about the "Kitchen Sink" NO supplements, stay tuned as this will be discussed in Part III of this series!

References

¹ Warren, Branch. December 2011. This file is licensed under the Creative Commons Attribution-Share Alike 2.5 Generic license. Accessed February 15th, 2012 from: http://commons.wikimedia.org/wiki/File:Branch01.jpg

² Fahs CA, Heffernan KS, Fernhall B. .Hemodynamic and vascular response to resistance exercise with L-arginine. Med Sci Sports Exerc. 2009 Apr;41(4):773-9.

³ Robinson TM, Sewell DA, Greenhaff PL. L-arginine ingestion after rest and exercise: effects on glucose disposal. Med Sci Sports Exerc. 2003 Aug;35(8):1309-15.

⁴ Podoprigora GI, Nartsissov YR, Aleksandrov PN. Effect of glycine on microcirculation in pial vessels of rat brain. Bull Exp Biol Med. 2005 Jun;139(6):675-7.

⁵ Giugliano D, Marfella R, Verrazzo G, Acampora R, Coppola L, Cozzolino D, D'Onofrio F. The vascular effects of L-Arginine in humans. The role of endogenous insulin. J Clin Invest. 1997 Feb 1;99(3):433-8.

⁶ Tang JE, Lysecki PJ, Manolakos JJ, MacDonald MJ, Tarnopolsky MA, Phillips SM. Bolus arginine supplementation affects neither muscle blood flow nor muscle protein synthesis in young men at rest or after resistance exercise. J Nutr. 2011 Feb;141(2):195-200. Epub 2010 Dec 29

⁷ Liu TH, Wu CL, Chiang CW, Lo YW, Tseng HF, Chang CK. No effect of short-term arginine supplementation on nitric oxide production, metabolism and performance in intermittent exercise in athletes. J Nutr Biochem. 2009 Jun;20(6):462-8. Epub 2008 Aug 15.

⁸ Alvares TS, Conte CA, Paschoalin VM, Silva JT, Meirelles Cde M, Bhambhani YN, Gomes PS.Acute l-arginine supplementation increases muscle blood volume but not strength performance. Appl Physiol Nutr Metab. 2012 Feb;37(1):115-26. Epub 2012 Jan 17

⁹ Campbell B, Roberts M, Kerksick C, Wilborn C, Marcello B, Taylor L, Nassar E, Leutholtz B, Bowden R, Rasmussen C, Greenwood M, Kreider R. Pharmacokinetics, safety, and effects on exercise performance of L-arginine alpha-ketoglutarate in trained adult men. Nutrition. 2006 Sep;22(9):872-81.

¹⁰ Willoughby DS, Boucher T, Reid J, Skelton G, Clark M. Effects of 7 days of arginine-alpha-ketoglutarate supplementation on blood flow, plasma L-arginine, nitric oxide metabolites, and asymmetric dimethyl arginine after resistance exercise. Int J Sport Nutr Exerc Metab. 2011 Aug;21(4):291-9.

¹¹ Goodwin BL, Solomonson LP and Eichler DC. Argininosuccinate synthase expression is required to maintain nitric oxide production and cell viability in aortic endothelial cells. J Biol Chem 2004; 279: 18353-18360.

¹² Karbach S, Simon A, Slenzka A, Jaenecke I, Habermeier A, Martiné U, Förstermann U, Closs EI. Relative contribution of different l-arginine sources to the substrate supply of endothelial nitric oxide synthase. J Mol Cell Cardiol. 2011 Nov;51(5):855-61. Epub 2011 Aug 2.

¹³ Vincent MA, Montagnani M, Quon MJ. Molecular and physiologic actions of insulin related to production of nitric oxide in vascular endothelium. Curr Diab Rep. 2003 Aug;3(4):279-88.

¹⁴ Tessari P, Coracina A, Puricelli L, Vettore M, Cosma A, Millioni R, Cecchet D, Avogaro A, Tiengo A, Kiwanuka E. Acute effect of insulin on nitric oxide synthesis in humans: a precursor-product isotopic study. Am J Physiol Endocrinol Metab. 2007 Sep;293(3):E776-82. Epub 2007 Jun 5.

¹⁵ Gannon MC, Nuttall JA, Nuttall FQ. Oral arginine does not stimulate an increase in insulin concentration but delays glucose disposal. Am J Clin Nutr. 2002 Nov;76(5):1016-22.

¹⁶ Sureda A, Córdova A, Ferrer MD, Pérez G, Tur JA, Pons A. L-citrulline-malate influence over branched chain amino acid utilization during exercise. Eur J Appl Physiol. 2010 Sep;110(2):341-51. Epub 2010 May 25.

¹⁷ Cormio L, De Siati M, Lorusso F, Selvaggio O, Mirabella L, Sanguedolce F, Carrieri G. Oral L-citrulline supplementation improves erection hardness in men with mild erectile dysfunction. Urology. 2011 Jan;77(1):119-22.

¹⁸ Jacobs PL, Goldstein ER. Long-term glycine propionyl-l-carnitine supplemention and paradoxical effects on repeated anaerobic sprint performance. J Int Soc Sports Nutr. 2010 Oct 28;7:35.

¹⁹ Bloomer RJ, Smith WA, Fisher-Wellman KH. Glycine propionyl-L-carnitine increases plasma nitrate/nitrite in resistance trained men. J Int Soc Sports Nutr. 2007 Dec 3;4:22.

²⁰ Gropper SS, Smith JL, Groff JL. Advanced Nutrition and Human Metabolism. 5th ed. 2009. pg 222-223