Hypoxia & Hyperoxia
REVIEW
Cobalt Misuse in Sports

Cobalt Misuse in Sports

Kobaltmissbrauch im Sport

Summary

Cobalt is a heavy metal, that wasused between the 1940s and 1980s as a therapeutic agent to treat anemic diseases. Similar to hypoxia, cobalt stabilizes the HIFαsubunits and stimulates the renal production of erythropoietin.

For several decades, it has been suggested that cobalt is also used in sports to optimize oxygen transport via increased hemoglobin mass. In 2015, WADA put it in the list of banned substances. In various dietary supplements purporting to increase performance, cobalt was detected whilst incorrectly or not declared by the manufacturers. Our research suggests that an oral dose of 5mg/day exceeds the erythropoietic threshold. As a result, a 3-week supplementation period at this dose leads to an increase in Hb mass of 2% and to a tendency of higher performance, which corresponds to the effects of a training camp lasting200h at 2000m. About 10% of the administered cobalt is absorbed by the body and excreted in urine with a half-life of 4-12 hours.

In order to detect doping with cobalt,a reference limit of 14ng/ml in urine has been suggested, which is 4 standard deviations higher than the normal urine cobalt concentration. This value is clearly exceeded during and until one week after a 3-week supplementation period at 5 mg/day. Since it is possible that athletes use higher amounts of cobalt for doping purposes in hopes of achieving greater gains in performance, WADA must set appropriate reference limits and introduce regular test procedures to avoid serious health risks to these athletes.

KEY WORDS: Hemoglobin Mass, Performance, Erythropoietin, Nutritional Supplement, Blood Manipulation

Zusammenfassung

Kobalt ist ein Schwermetall, das in den 1940-iger bis 1980-iger Jahren als Therapeutikum bei Anämieerkrankungen eingesetzt wurde, wobei schwerwiegende Nebenwirkungen beobachtet wurden. Ähnlich wie Hypoxie stabilisiert Kobalt die HIFα-Untereinheiten und stimuliert somit u. a. die renale Bildung von Erythropoietin.

Seit mehreren Jahrzehnten wird vermutet, dass Kobalt auch im Sport eingesetzt wird, um über eine erhöhte Hämoglobinmasse den Sauerstofftransport zu optimieren, sodass es 2015 von der WADA auf die Liste der verbotenen Substanzen gesetzt wurde. In Nahrungsergänzungsmitteln wird Kobalt als sogenannter Performance-Booster eingesetzt, wobei es zumeist nicht oder falsch deklariert wird. Bei einer oralen Dosis von 5mg/Tag wird eine erythropoietische Schwelle überschritten und verstärkt Erythropoietin gebildet, was bei 3-wöchiger Anwendung zu einem Anstieg der Hb-Masse von 2% führt und tendenziell die Leistungsfähigkeit steigert, was den Effekten eines 200h langen Trainingslagers auf ca. 2000m Höhe entspricht. Etwa 10% des verabreichten Kobalts wird vom Körper aufgenommen und mit einer Halbwertzeit von 4-12h Stunden über den Urin ausgeschieden.

Um Doping mit Kobalt nachweisen zu können,wurde ein Grenzwert von 14ng/ml im Urin vorgeschlagen, was der 4-fachen Standardabweichung der normalen Kobaltkonzentration im Urin entspricht. Dieser Wert wird während und eine Woche nach einer 3-wöchigen Supplementation mit 5mg/Tag klar überschritten. Da zu erwarten ist, dass Sportler auch höhere Kobaltmengen zu Dopingzwecken missbrauchen und dadurch ihre Leistung deutlich steigern dürften, aber damit auch schwerwiegende Gesundheitsrisiken in Kauf nehmen, muss die WADA entsprechende Grenzwerte festlegen und regulär danach testen.

SCHLÜSSELWÖRTER: Hämoglobinmasse, Leistung, Erythropoietin, Nahrungsergänzungsmittel, Blutmanipulation

Introduction

Maximum oxygen uptake as a measure of endurance performance is closely related to the absolute amount of circulating hemoglobin. After phlebotomy or after application of recombinant human erythropoietin (rhEPO), a change of 1 g of hemoglobin causes a change in VO2max of approx. 4ml/min (29). Therefore, altitude training is used as an allowed method and blood manipulations, either by blood transfusions or by erythropoiesis stimulating agents (ESAs), are assumed to occur frequently as a malpractice among elite endurance athletes (30, 36). The following review provides an overview of oral cobalt ingestion as a possible doping measure to stimulate erythropoietic activity. Key data and opinions are mostly based on two original publications from Hoffmeister et al. (15, 16).

Co2+ mimics the effects of hypoxic exposure by stabilizing the HIF 1αand 2αsubunits. HIF 1 stimulates the synthesis of the vascular endothelial growth factor (VEGF), while HIF 2 augments the production of endogenous erythropoietin (18). Actual data indicate that Co2+ may bind to HIF 1α and 2α preventing the interaction with the von-Hippel-Landau tumor suppressor protein (pVHL) (18). The application of cobaltous ions is therefore one possibility to stimulate endogenous EPO production resulting in an increase in hemoglobin mass and improved endurance performance.

In equestrian sports, doping with cobalt has become an important issue. National and international equine organisations have discussed upper reference limits for cobalt in blood and urine and responsible persons have already been sanctioned (5, 14, 24).

Although there are no scientific data available, scientists have also warned of its abuse in human sports (22, 23) and in 2015 WADA has put cobalt on the list of prohibited substances and methods (35). In accordance to the anti-doping regulations, doping cases have already been reported (19).

Erythropoietic Effect of Co2+ Administration

From the mid-1940s until the 1980s, cobalt ions (Co2+) were used for patients suffering from anemia from various causes (8, 18). Co2+acts similarly as a hypoxic stimulus; it stabilizes HIF-1αand HIF-2αand thereby induces increased renal erythropoietin production (for a review see (18)). Before the development of recombinant erythropoietin, one unit of EPO was defined as the erythropoietic effect achieved by 5µmol of Co2+ (17). Because of the severe side effects of high-dose Co2+ treatment, it was replaced by anabolic steroids and later by recombinant erythropoietin (8). In clinical practice, daily doses between 11 to 135 mg Co2+ were administered for several months (for review, see (8, 18)). While higher doses consistently improved conditions of anemia, doses of 11-25 mg Co2+ (e.g. (3, 7)) also increased hemoglobin concentration ([Hb]), hematocrit (Hct), and reticulocytes. Unfortunately, high dosing in anemic patients (>25 mg Co2+/day (4, 11)), was associated with a high risk for intolerability, i.e., acute poisoning and organ injury, gastro-intestinal sickness, thyroidal dysfunction, myocardial effects, and neural and sensory disturbances (8). Because of this high toxicity and the frequent side effects, cobalt therapy was abandoned in the late 1970s, and anabolic steroids were used until recombinant erythropoietin became available (8).

In healthy subjects, low dosages (0.45mg Co2+/day for 31 days (12); 1mg Co2+/day for 90 consecutive days (33)) did not have any effect on [Hb], Hct or red blood cell count (RBC), nor showed any side effects. Oral application of high dosages, i.e. 70mg Co2+, however, increased [Hb] by 20% and the reticulocyte count from 1.1% to 2.7% within 7-22 days. After cessation, baseline values were reached within 15 days (6).

Cobaltous Supplements

Although there are no scientific data available, many manufacturers of nutritional supplements claim erythropoietic stimulatory effects of their cobalt-containing products and suggest a subsequent enhancement of endurance performance. To manipulate their blood, cheating athletes may therefore use cobalt salts offered for chemical purposes or veterinarian nutrition, or they may use dietary supplements that can be purchased over the counter without any restriction. The Co2+ concentrations in these mostly liquid supplements are between 0.03mg/ml and 0.5mg/ml, and the recommended daily dose is between 0.15 and 1.0mg/day (31). This is far below the therapeutic dosages formerly used for treatment of anemia but well above the amounts required as the central component of vitamin B12 (0.1µg/day (32)). These recommendations are within the range classified as harmless by two international health and nutritional organizations indicating upper limits as 0.009mg/kg/day (9) and 0.02mg/kg/day (10), corresponding to 0.6mg and 1.4mg for a 70kg subject. The oral reference dose (RfD), i.e., the maximum daily amount of cobalt ingestion considered not to cause adverse non-cancer health effects during a lifetime, has been indicated to 2.1mg/day for a 70kg subject (0.03mg/kg) (11).

Recently, Thevis et al. analyzed 19 commercially available dietary supplements advertised as endurance performance enhancers (31). Eleven of them contained up to 2.6mg Co2+/ml, which was only declared in two of these supplements. The Co2+ concentration of the supplements we used in two recent studies (15, 16) was declared to be 0.2mg/ml. However, in almost half of the units used for supplementation, the real concentration was more than 3 times higher. This data suggests that ‘performance enhancing’ dietary supplements have to be used with caution to avoid any side effects caused by undeclared and/or falsely declared Co2+ content.

Erythropoietic Effect of Low Dose Co2+ Administration

To evaluate the effects of cobaltous supplements, we (16) determined the minimum doses of oral cobalt administration exerting any erythropoietic effect. While Co2+ dosages of 1mg/day or 2mg/day did not exert any hematological changes, a single dose of 10mg of Co2+ clearly increased plasma [EPO] and 5 mg showed a strong tendency for elevated [EPO] suggesting that this dosage might be a cobalt threshold for EPO stimulation (Fig. 1). Similar to hypoxic exposure, [EPO] increased after a delay of 3-5h (28) which was explained by similar effects of Co2+ and of hypoxia on HIF stabilization (18). A 5-day Co2+administration of 10 mg/day clearly passed the erythropoietic threshold, as indicated by increased plasma [EPO], a higher percentage of the immature reticulocyte fraction (IRF), and decreased plasma ferritin concentration proving the effectiveness of this dosage (16). In a follow up study (15) we administered 5mg of Co2+/day for three weeks which produced clear erythropoietic stimulation, resulting in a 2% increase in Hbmass (Fig. 2), which is similar to what has been observed during hypoxic training camps lasting 200h at an altitude of ~2,000m (13).

Although suppliers of nutritional supplements claim enhanced endurance performance from Co2+ supplementation, this hypothesis has not yet been verified in the scientific literature. In a meta-analysis of 145 elite athletes, Saunders et al. (27) found a 3% increase in Hbmass and VO2max after hypoxic exposure (“live high – train high” and “live high – train low” protocols). The correlation between these changes was, however, weak, and it explained less than one-sixth of the variation, indicating that, apart from increased Hbmass, other factors must also be important in causing the elevated VO2max after altitude training (27). Like after altitude training measures, also after the 3-week Co2+ administration period described above VO2max tended to increase and there was a strong tendency for a relationship (r=0.40, p=0.1) for changes in Hbmassand changes in VO2max (15). We, therefore, concluded the occurrence of stronger effects on VO2max in case of higher Co2+ dosages.

Urinary Cobalt Concentration and Possible Reference Limits

The normal [Co2+] in urine is relatively low and shows a small scattering (0.34ng/ml, 95% confidence limits (CL) 0.31-0.37ng/ml, (34)). A recently conducted study from the anti-doping laboratory in Cologne / Germany yielded slightly, but significantly higher values in endurance trained athletes than in untrained subjects (0.36ng/ml vs 0.28ng/ml, (20)), while acute exercise did not change urinary Co2+ concentration neither in trained nor in untrained subjects (25). Exposure to cobalt containing environments, however, massively increases urinary [Co2+]. People living closer to mines than 3 km show in the mean approx. 50-fold increased [Co2+] (2). Similarily, hard metal factory workers are characterized by extremely high values reaching up to 2000 ng/ml (26). Another source of cobalteous ions are endoprosthetics. Metal-on-metal hip implants deliver the highest systemic and urinary Co2+concentrations which range normally between 0.2 and 10ng/ml, but may rise in state of malfunction up to 1085ng/ml (21).

When Co2+ is ingested most of it is excreted in the feces (1). Urinary excretion accounts for only 10-27% of ingested Co2+ (1, 16) and the elimination half-life after low-dose oral ingestion is between 4h and 12h (1, 32). The time course of urinary Co2+ excretion after a single oral application of 5 mg and 10mg Co2+ shows maximum values 3-7h (increase from 0.8ng/ml to 150 ng/ml and 340ng/ml, respectively; Fig. 3)) after ingestion and still 50-fold elevated values after 24h. There exists also a relatively high inter-individual variation of urine Co2+ which may be attributable to differences in gastrointestinal Co2+ absorption as well as an intra-individual variation due to different diuresis.During a 3-week Co2+ application period (5 mg/day), we (15) found the highest urine Co2+ concentrations at the end of the administration period (310 ng/ml after one week; 460ng/ml after 3 weeks), which also exceeded those values measured after a single or 5-day application of the equal Co2+ dosage. We therefore assumed that Co2+ accumulates in body fluids, which may lead to a stronger erythropoietic response than expected from short-term administration. After cessation of the administration, urine Co2+ concentration dropped by approx. 80% within two days, which is consistent with the elimination half-life of Co2+ after low-dose oral ingestion (1, 32). Compared to baseline, urine [Co2+] was still elevated until two weeks after cessation, which has implications for establishing threshold limits to combat its misuse. Krug et al. (20) calculated 14ng/ml as a tentative reference limit for urine Co2+ concentration, which was determined by multiplying the reference population (n=100 subjects) standard deviation by four. In our study (15), all participants except one individual exceeded this threshold one week after cessation (mean=43.1±47.1ng/ml), and some individuals exceeded it even after 2 or 3 weeks. Therefore, using the reference limit proposed by Krug et al. (20), it should be possible to detect cheating athletes during the Co2+ingestion period and in most cases for at least one week after cessation.

Another way to establish reference limits may be the determination of Co2+ in erythrocytes. Co2+ is rapidly cleared from the plasma (33) but it is stored in the red cells until they are destroyed after a maximum lifespan of approximately 120 days.

Detection of cheating athletes, may therefore even easier by measuring Co2+ in red cells. Until now, however, a routine detection method has not yet been established.

Conclusions

Cobalt is a strong erythropoiesis stimulating agent which can easily be acquired by cheating athletes. In 2015, WADA put cobalt on the list of prohibited substances; but until today no detection procedure has been introduced. Cobalt can be used by athletes to mimic the effects of hypoxic training measures (live high – train high, or live high – train low) or to prolong the effects of a preceding altitude training camp. Because cobalt acts in a dose dependent manner and because many athletes may try to get the maximum “benefit” from a cobalt supplementation, there exists the reasonable assumption that some athletes will use very high dosages which are associated with severe side effects. WADA has, therefore, to implement reasonable reference limits which we suggest being 14ng/ml in urine to keep sports as clean as possible and to protect the cheating athletes. WADA should also conduct re-tests of stored urine samples and it will be interesting to know whether and in which prevalence cobalt has been used by cheating athletes in the past.

Conflict of Interest
The authors have no conflict of interest.

References

  1. AYALA-FIERRO F, FIRRIOLO JM, CARTER DE. Disposition, toxicity,and intestinal absorption of cobaltous chloride in maleFischer 344 rats. J Toxicol Environ Health A. 1999; 56: 571-591.
  2. BANZA CL, NAWROT TS, HAUFROID V, DECRÉE S, DE PUTTER T,SMOLDERS E, KABYLA BI, LUBOYA ON, ILUNGA AN, MUTOMBO AM,NEMERY B. High human exposure to cobalt and other metals inKatanga, a mining area of the Democratic Republic of Congo.Environ Res. 2009; 109: 745-752.
  3. BOWIE EA, HURLEY PJ. Cobalt chloride in the treatmentof refractory anaemia in patients undergoing longtermhaemodialysis. Aust N Z J Med. 1975; 5: 306-313.
  4. BRADBERRY S, SABATTA M, VALE J. Cobalt Chloride. Updated 2016 [10th February 2019].
  5. COOK C. Robin Bastiman banned for three years by BHA forinjecting horse. Updated 2019 [10th February 2019].
  6. DAVIS JE, FIELDS JP. Experimental production of polycythemia inhumans by administration of cobalt chloride. Proc Soc Exp BiolMed. 1958; 99: 493-495.
  7. DUCKHAM JM, LEE HA. The treatment of refractory anaemia ofchronic renal failure with cobalt chloride. Q J Med. 1976; 45:277-294.
  8. EBERT B, JELKMANN W.
  9. EUROPEAN FOOD SAFETY AUTHORITY (EFSA). Scientific Opinion on theuse of cobalt compounds as additives in animal nutrition. EFSAJ. 2009; 7: 1383.
  10. EXPERT GROUP ON VITAMINS AND MINERALS. 2003. Safe Upper Levelsfor Vitamins and Minerals. Updated 2003 [26th April 2019].
  11. FINLEY BL, MONNOT AD, PAUSTENBACH DJ, GAFFNEY SH. Derivation of achronic oral reference dose for cobalt. Regul Toxicol Pharmacol.2012; 64: 491-503.
  12. FINLEY BL, UNICE KM, KERGER BD, OTANI JM, PAUSTENBACH DJ,GALBRAITH DA, TVERMOES BE. 31-day study of cobalt(II) chlorideingestion in humans: pharmacokinetics and clinical effects. JToxicol Environ Health A. 2013; 76: 1210-1224.
  13. GARVICAN-LEWIS LA, SHARPE K, GORE CJ. Time for a new metric forhypoxic dose? J Appl Physiol (1985). 2016; 352-355.
  14. HO ENM, CHAN GHM, WAN TSM, CURL P, RIGGS CM, HURLEY MJ, SYKES D. Controlling the misuse of cobalt in horses. Drug Test Anal. 2015;7: 21-30.
  15. HOFFMEISTER T, SCHWENKE D, KRUG O, WACHSMUTH N, GEYER H,THEVIS M, BYRNES WC, SCHMIDT WFJ. Effects of 3 weeks of oral lowdosecobalt on hemoglobin mass and aerobic Performance. FrontPhysiol. 2018; 9: 1289.
  16. HOFFMEISTER T, SCHWENKE D, WACHSMUTH N, KRUG O, THEVIS M,BYRNES WC, SCHMIDT WFJ. Erythropoietic effects of low-dosecobalt application. Drug Test Anal. 2019; 11: 200-207.
  17. JELKMANN W. Efficacy of recombinant erythropoietins: is thereunity of international units? Nephrol Dial Transplant. 2009; 24:1366-1368.
  18. JELKMANN W. The disparate roles of cobalt in erythropoiesis,and doping relevance. Open J Hematol. 2012; 3-6.
  19. KREUZER HP. Kobalt: Effektiv und gefährlich. Updated 2016 [10th February 2019].
  20. KRUG O, KUTSCHER D, PIPER T, GEYER H, SCHÄNZER W, THEVIS M. Quantifying cobalt in doping control urine samples—a pilotstudy. Drug Test Anal. 2014; 6: 1186-1190.
  21. LEYSSENS L, VINCK B, VAN DER STRAETEN C, WUYTS F, MAES L. Cobalttoxicity in humans-A review of the potential sources andsystemic health effects. Toxicology. 2017; 387: 43-56.
  22. LIPPI G, FRANCHINI M, GUIDI GC. Cobalt chloride administrationin athletes: a new perspective in blood doping? Br J Sports Med.2005; 39: 872-873.
  23. LIPPI G, FRANCHINI M, GUIDI GC. Blood doping by cobalt. Shouldwe measure cobalt in athletes? J Occup Med Toxicol. 2006; 1: 18.
  24. MOBASHERI A, PROUDMAN CJ. Cobalt chloride doping in racehorses:Concerns over a potentially lethal practice. Vet J. 2015; 205: 335-338.
  25. MUNOZ D, MAYNAR M, BARRIENTOS G, SIQUIER-COLL J, BARTOLOME I,GRIJOTA FJ, ROBLES MC. Effect of an Acute Exercise UntilExhaustion on the Serum and Urinary Concentrations of Cobalt,Copper, and Manganese Among Well-Trained Athletes. BiolTrace Elem Res. 2018.
  26. PRINCIVALLE A, IAVICOLI I, CERPELLONI M, FRANCESCHI A, MANNO M,PERBELLINI L. Biological monitoring of cobalt in hard metalfactory workers. Int Arch Occup Environ Health. 2017; 90: 243-254.
  27. SAUNDERS PU, GARVICAN-LEWIS LA, SCHMIDT WF, GORE CJ. Relationship between changes in haemoglobin mass andmaximal oxygen uptake after hypoxic exposure. Br J Sports Med.2013; 47: i26-i30.
  28. SCHMIDT W, ECKARDT KU, HILGENDORF A, STRAUCH S, BAUER C. Effectsof maximal and submaximal exercise under normoxic andhypoxic conditions on serum erythropoietin level. Int J SportsMed. 1991; 12: 457-461.
  29. SCHMIDT W, PROMMER N. Impact of alterations in total hemoglobinmass on VO2max. Exerc Sport Sci Rev. 2010; 38: 68-75.
  30. SOTTAS PE, ROBINSON N, FISCHETTO G, DOLLE G, ALONSO JM, SAUGY M. Prevalence of blood doping in samples collected from elite trackand field athletes. Clin Chem. 2011; 57: 762-769.
  31. THEVIS M, KRUG O, PIPER T, GEYER H, SCHÄNZER W. Solutionsadvertised as erythropoiesis-stimulating products were found tocontain undeclared cobalt and nickel species. Int J Sports Med.2016; 37: 82-84.
  32. TVERMOES BE, FINLEY BL, UNICE KM, OTANI JM, PAUSTENBACH DJ,GALBRAITH DA. Cobalt whole blood concentrations in healthyadult male volunteers following two-weeks of ingesting a cobaltsupplement. Food Chem Toxicol. 2013; 53: 432-439.
  33. TVERMOES BE, UNICE KM, PAUSTENBACH DJ, FINLEY BL, OTANI JM,GALBRAITH DA. Effects and blood concentrations of cobalt afteringestion of 1 mg/d by human volunteers for 90 d. Am J Clin Nutr.2014; 99: 632-646.
  34. U.S. DEPARTMENT OF HEALTH & HUMAN SERVICES. Fourth nationalreport on human exposure to environmental chemicalsUpdated Tables, January 2019, Volume 1. Updated 2019 [10th February 2019].
  35. WADA. List of prohibited substances and methods 2019. Updated 2019 [18th February 2019].
  36. WINTERMANTEL J, WACHSMUTH N, SCHMIDT W. Doping Cases amongElite Athletes from 2000 to 2013. Dtsch Z Sportmed. 2016; 67:263-269.
Prof. Dr. Walter Schmidt
Head of Department, Dept. Sports Medicine/
Sports Physiology, University of Bayreuth
Universitätsstrasse 30
95440 Bayreuth, Germany
walter.schmidt@uni-bayreuth.de