To the Editor,
Guanidinoacetic acid (GAA) or glycocyamine, a natural precursor of creatine, was introduced in the field of human nutrition and medicine more than 65 years ago. Seminal studies from the 1950s have pointed to the beneficial effects of GAA in different clinical conditions [1-3]. Nevertheless, in the decades that followed, there was no major research interest shown for this compound. However, a few isolated human studies evaluated supplemental GAA in kidney failure and diabetes . In recent days, a scientific interest for GAA has developed, with a few studies investigating its effectiveness and safety in healthy humans  and in women with chronic fatigue syndrome . It appears that dietary GAA increases methylation demand and provokes mild hyperhomocysteinemia , a risk factor for cardiometabolic diseases. However, the safety effects of GAA on traditional cardiometabolic biomarkers, including high-density lipoprotein (HDL) cholesterol, C-reactive protein, triglycerides, and insulin, have not been investigated so far.
In this open-label trial, we examined the effects of the 10-week supplementation with 3 grams per day of GAA on biomarkers of cardiometabolic risk and inflammation in 20 apparently healthy volunteers (10 men and 10 women; age 22.0 ± 2.3 years). The participants were free to stop using any dietary supplement 4 weeks before study commenced. All were verbally informed about the nature and demands of the study, and everyone gave a written consent to participate in this trial. The study was ethically approved by the local Institutional Review Board in accordance with the Declaration of Helsinki. All participants were assigned to receive GAA for 10 weeks, and were evaluated at baseline and then after 10-week intervention. The primary endpoint of GAA intervention was the change in serum levels of high-sensitivity C-reactive protein (hsCRP) assessed at baseline and at 10-week follow-up. Participants were asked not to change their dietary habits and physical activity routine during the intervention. At each visit to the lab, participants provided a fasting sample of venous blood that was immediately centrifuged with serum analyzed for HDL cholesterol, triglycerides, hsCRP, insulin, ferritin, vitamin B12, folic acid, total homocysteine (tHcy), GAA, creatine, creatinine, and clinical enzymes.
Three participants (1 woman and 2 men) were lost to follow-up due to reasons not connected to the study per se. One female participant (age 21, weight 60 kg) experienced recurrent episodes of gastrointestinal stress (nausea, vomiting) during the first week of the trial, and was excluded from the study at day 8 of the intervention. Other participants reported no adverse events due to GAA treatment. Changes in cardiometabolic markers and clinical enzymes during the study are presented in Table 1. GAA had no significant effects on serum hsCRP, HDL cholesterol, insulin, and triglycerides. Clinical enzymes were mainly unaffected by GAA supplementation. Alkaline phosphatase levels increased after GAA intervention (13.6% corresponding to 9.5 IU/L; p = 0.01), but mean values remained within normal ranges (44–147 IU/L). Supplementation with GAA yielded a statistically significant increase of the mean serum ferritin levels at post-administration (7.5% corresponding to 4.1 µg/L; p = 0.012). Individual and mean changes in markers of GAA-creatine metabolism are presented in Figure 1. Post-administration levels were boosted by 815.4% for GAA, 88.7% for creatine, and 13.7% for creatinine. GAA notably increased serum tHcy (73.5% corresponding to 5.0 µmol/L; p < 0.0001), with 4 participants experiencing clinically high tHcy levels (>15.0 µmol/L) at follow-up. The highest tHcy concentration at post administration (18.2 µmol/L) was reported in a female participant (age 23, weight 73.9 kg). GAA affected serum vitamin B12, with mean levels at post-administration elevated for 13.1% as compared to the baseline (p = 0.01). No effects of the intervention were found for serum folic acid.
We found that supplemental GAA has no significant effects on traditional biomarkers of cardiometabolic risk and inflammation in healthy volunteers. Neither serum hsCRP nor insulin was affected by 10-week GAA administration, with a ratio of triglycerides to HDL cholesterol (an indicator of atherogenic profile) remaining essentially unaffected by the intervention. This implies that there is no major cardiometabolic burden of medium-term GAA intervention in healthy humans. No clinically relevant disturbances in serum enzymes profiles were found, suggesting no considerable stress of the kidney and liver during GAA intake. GAA appeared to elevate serum creatinine, perhaps due to increased creatine utilization after administration , with the escalation remaining mild and clinically marginal throughout the study. Supplemental GAA also affected serum ferritin, an important biomarker of inflammation, with levels significantly increased from 54.8 µg/L at baseline to 58.9 µg/L at follow-up, suggesting a possible proinflammatory effect of GAA in healthy humans. However, mean ferritin values remained within normal ranges (24–300 µg/L) during the trial, with no volunteer having a markedly elevated serum ferritin level after administration associated with inflammatory conditions. This confirms the results from the recent animal study  where authors reported no tissue inflammation (as evaluated by the number of inflammatory cells in the liver, lipogranulomas, or portal triad inflammation) in rats exposed to GAA-rich diet. Nevertheless, dietary GAA should be carefully scrutinized as an experimental dietary additive due to its proven capacity to drive increased homocysteine production , which is again confirmed in the present study. Although GAA positively affected serum markers of bioenergetics, including increased creatine levels at post-administration for up to 104.3%, the intervention also elevated serum tHcy in all participants at 10-week follow-up, with 1 in 4 participants experiencing clinical hyperhomocysteinemia. Since hyperhomocysteinemia is well known as an independent risk factor for several cardiometabolic disorders, a progressive GAA-driven increase in plasma homocysteine should be considered the main adverse effect of the intervention . We also reported minor disturbances in serum vitamin B12 levels at post administration, while folic acid remained unaltered by GAA intake. Since the methylation of GAA to creatine (and concomitant utilization of homocysteine) is controlled by B vitamins, one might expect increased consumption of B vitamins during the intervention . It appears that the current administration protocol with GAA has been either too short or under-dosed to majorly affect selected biomarkers of B vitamin status. This study has several limitations including the lack of a placebo group, an open label design, the limited number of cardiometabolic markers evaluated, and no gender differences were analyzed. Long-term and well-powered placebo-controlled studies are therefore warranted to calculate a net cardiometabolic safety of dietary GAA, linking elevated tHcy during GAA intervention with clinician-reported outcomes of cardiometabolic health.
This project was partly supported by the Serbian Ministry of Education, Science and Technological Development (175037), the Provincial Secretariat for Higher Education and Scientific Research (114-451-710), and the Faculty of Sport and PE (2017 Annual Award).
The authors have no conflicts of interest to disclose.