The TC02 Testing Conundrum

Off The Record with Charl Pretorius

The shock withdrawal of Main Defender from the IOS Gr2 Drill Hall Stakes on Saturday has once again opened the ongoing debate about the merits and demerits of TC02 testing.

Charl Pretorius writes that the South African racing industry is divided in leadership, opinions, and future vision, but there are two fundamental aspects that garner widespread agreement: the protection of the thoroughbred racehorse and professional control and policing of the sport. 

‘Milkshaking’:The administration of bi-carb via a naso-gastric tube (Pic – 4Racing)

Measures have been instituted on the horse protection side, but are we getting the policing right? Are our resources devoted to the right channels? What is fact, and what is conjecture?

The National Horseracing Authority, mandated and under increased pressure to identify trainers who use illegal methods to gain an advantage, last month introduced a new specimen analysis procedure for TCO2 (total carbon dioxide) measurement on a race day.

These are tests performed on the blood samples of racehorses to combat the practice of administering alkalizing agents for the purpose of enhancing performance.

For those who are still uncertain of how this works: Blood specimens are taken from selected horses approximately 50 minutes prior to race time, on any given race day. The specimens are immediately analysed using handheld blood gas analysers.

When a sample taken is found to have an elevated pre-race blood TCO2 level above of 36 millimoles per litre (mmol/L) – the threshold set by the International Horse Racing Federation (IFHA) and prescribed to by the NHA – a second blood specimen is collected, between 10 and 15 minutes later. If the TCO2 concentration of the second specimen is more than 36.0 mmol/L, the horse is withdrawn and then detained for additional specimen collection.

During the additional tests, screening can be performed for other illegal substances that may or may not be present in the blood sample.

Dr Matthew Bawden (Pic – 4Racing)

The administration of sodium bicarbonate (bi-carb) in the hours prior to racing, is viewed by the IHFA as performance enhancing. It is performed using a practice widely known as ‘milkshaking’ – commonly via the insertion of a naso-gastric tube between 3 to 5 hours prior to a race.

This is intended to elevate TCO2 and therefore reduce lactate. Three horses have been withdrawn from races due to elevated TCO2 levels since the NHA implemented testing for this violation in recent weeks – two on the Highveld and one in Durban.

The introduction of TC02 testing has drawn active interest from the public from as far as the USA, with most commentators in favour of the procedures.

Team Valor’s Barry Irwin, a prominent anti-drug activist with significant South African racing interests, wrote on a Sporting Post forum: “Welcome to the real world of horse racing. It is about time. This technology has been around for decades.

This updated protocol will go a long way in levelling the playing field on the Highveld. Targeting trainers whose horses improve is a proven formula used by regulators worldwide. Not to follow this practice would be wasteful and foolish.”

Michael Jacobs, responding to a posting about the potential additional operating costs involved in additional TRC02 testing, said: “Why, if the regulator puts measures in place to combat and detect doping practices, would these steps be questioned? Are we trying to protect the dopers or the betting public?”

Geoff Logan argued that TCO2 tests will “prove very disruptive to say the least”, and a few others, including ‘Joe King’, suggested that TC02 testing is a result of pressure on the NHA from powerful industry players and that tests are now aggressively performed with focus on specific stables (in reference to Irwin’s suggestion that the Highveld is where milkshaking is giving an unfair advantage to certain trainers).

Graham Niven commented: “What protocol has been put in place to prohibit targeting particular trainers? There have been rumours and certainly evidence of late that trainers whose horses show a marked improvement are suddenly deemed ‘doped’ horses. May I suggest that every trainer with horses on the day draws a chip and either a colour or number is associated to that chip and those are randomly drawn by an independent to minimise targeting.”

TC02 testing with hand-held analysers (Pic – 4Racing)

Dr. Matthew Bawden, the NHA’s Chief Veterinarian, remarked: “The main goal of measuring TCO2 before racing is to detect horses that may have received an alkalizing agent. Alkalinization of a horse or the use of an alkalizing agent is prohibited on race day, and a level exceeding the international threshold constitutes prima facie evidence of alkalinization. The administration of an alkalizing agent results in a measurable increase in the TCO2 over time with an intent to achieve maximal buffering capacity at the time of the race.”

The implication here is that a horse could pass the TC02 test before the race, but could possibly fail it afterwards, which presents an interesting conundrum.

The NHA categorizes TCO2 as a Class 2 Prohibited substance, alongside substances such as Ketamine and Morphine. According to the guidelines on the NHA’s website, first-time offenders face a fine of R250,000 and a six-month suspension, while second-time offenders receive a warning off. Interestingly, the NHA does not conduct post-race testing for high TCO2 levels. This is a peculiar situation: There is significant anticipation and warnings regarding the tests, substantial fines that could end careers, but ultimately no actual possibility of punishment due to the absence of post-race positive samples. Yes, go figure.

Things will truly get interesting if penalties of this magnitude are actually dished out, because there will be areas of contention and grounds for legal objections raised in terms of the applicable thresholds, testing procedures, logistical mistakes, victimisation of specific trainers and the reliability of the testing equipment.

A closer look at ‘milkshaking’ presents some sobering scientific views from a number of recognised veterinary authorities.

Dr. Michael I. Lindinger, PhD, the President of the Nutraceutical Alliance, known for its comprehensive research capabilities, wrote in a 2021 edition of Canadian Thoroughbred Magazine: “The threshold for the TCO2 test set by the IFHA is not based on physiological data, but rather statistically manipulated data obtained from thousands of racehorses on race day, in varying conditions of health, and in varying conditions of hydration. Most of these studies assume, as opposed to actual knowledge, that horses that tested with a TCO2 greater than 35 mmol/l had been administered an alkalizing substance. This is only an assumption, not fact. This is a very poor justification for a threshold test of a naturally occurring substance. It is wishful thinking, not science.”

Lindinger’s assessment is supported by The, who works closely with university and research institute experts. In their article, Put the baking soda back in the bottle: banned sodium bicarbonate ‘milkshakes’ don’t make racehorses faster, they wrote: “Our analysis included data from eight experimental trials featuring 74 horses. Overall, sodium bicarbonate administration in the hours before treadmill tests or simulated race trials did not improve horses’ running performance in either type of test. In fact, in treadmill exercise tests in which horses were not ridden by jockeys, sodium bicarbonate actually had a very small negative effect on running performance, albeit not a statistically significant one.

“Whereas human athletes might gain a placebo effect from sodium bicarbonate, this is unlikely to apply to horses who don’t understand the intended point of the milkshake. And while some racehorse trainers may be educated in exercise physiology and the importance of blood pH, others may believe they work simply because received wisdom and racing folklore say so.

“The controversial and banned practice of giving horses baking soda ‘milkshakes’ before a race doesn’t work, according to our analysis of the available research. Racing folklore says sodium bicarbonate milkshakes can boost racehorses’ endurance because the alkalinity of the baking soda helps counter the buildup of lactic acid in the blood when running. But our systematic research review, published in the Journal of Equine Veterinary Science reveals milkshakes don’t boost horses’ athletic performance.”

***Testing procedures for TC02 came under scrutiny in the USA in 2013, when the Texas Racing Commission dismissed its case against then leading trainer, Karl Broberg, for an alleged TCO2 violation at Sam Houston Race Park. A second TCO2 complaint against an unnamed trainer whose horse tested above the then 37.0 limit that same night was also dismissed.

TCO2 values vary from instrument to instrument depending on the analytical approach used. Both of the alleged violations were dismissed because the method of collection and handling of blood samples was not compatible with the type of equipment being used to measure total carbon dioxide. The Texas A&M Medical Diagnostic Laboratory, the state’s testing lab, was using a portable blood-gas analyzer.

TCO2 levels are more accurately measured with TCO2 analyzers, rather than blood-gas analyzers. Those machines can give elevated TCO2 readings if the blood cells are not separated from serum through centrifugation (spinning a sample at high speeds to separate blood components) within an hour of collection.

If the sample is not centrifuged, the CO2 produced inside the blood cells can diffuse into the serum. In the Broberg case, the sample that resulted in a 42.9 TCO2 reading (well above the 37.0 legal threshold) also had a potassium ion level that was off the charts, suggesting sample degradation.

The NHA’s CEO, Vee Moodley, said in response to a query that the NHA are using “approved and validated hand-held blood gas analyzers which measure the total carbon dioxide”. The samples are not centrifuged. Blood collected for on course TCO2 analysis is analysed as whole blood within 10 minutes of collection.

International testing for TCO2 varies widely. Ireland changed their protocols in 2021. They took 5966 samples in 2023 (of which 569 were pre-race blood tests). Of the entire total, six (0,1%) race day positives were brought before the Irish Horseracing Regularity Board (IRHB), with no TC02 positives stipulated in the IHRB’s Year-End report. Testing is not performed at all in Japan, where horses are under surveillance and officials believe they are not likely to be ‘milkshaked’.

In Singapore there has been one TC02 penalty imposed in the last 18 years. In August 2022, trainer HW Tan was found guilty on a pre-race TC02 test at Kranji (37,35 mmol/L). His runner actually competed and finished second. He pleaded guilty and was fined $20,000. Singapore’s Chief Stipendiary Steward Scott Kelly said that post-race analysing of pre-race tests is their standard practice at present. Pre-race samples are not immediately analysed.

Natalie Voss reported in an article published on The Paulick Report: “A study published in the Equine Veterinary Journal in 2006 showed a number of other factors associated with TCO2 levels, including gender (male horses tended to have higher TCO2 levels), class of race (maidens tended to have lower TCO2), finish position (horses hitting the board had higher TCO2 than those that did not), and oddly, cloudy weather.

It’s unclear what caused these differences, and whether TCO2 levels explained a horse’s performance (with a better performance being a result of higher TCO2) or if TCO2 readings were more a marker of a given horse’s fitness. The study also found trainer- and horse-specific trends, which doesn’t necessarily suggest trainers in the study were intentionally manipulating TCO2 levels.

It is possible that different barns might create different environments for TCO2 readings given their combination of feed and exercise programs.

From the results of a 2022 study, updated in 2023, the Journal of Veterinary Science assessed: “From the statistical analysis of TC02 measurements taken of screening samples from Thoroughbred and Quarter Horses in Louisiana during a 12-month period, a clear seasonal variation has been identified, with higher concentrations of TC02 being reported in winter and spring months in comparison to summer months.”

Dr. Lindinger wrote in another article published in the Journal: “Racing jurisdictions have attempted to justify the use of a single measurement, namely TCO2, as a means of positively and definitively identifying the administration of alkalinizing substances to racehorses. However, there is by no means consensus amongst equine researchers with the majority of equine physiologists advocating the need for multiple measurements and other controls.”


Vee Moodley and Dr Mathew Bawden answered further questions presented – reproduced below as given and replied to, with overlapping issues condensed for readability:

You are quoted as saying that the ultimate goal would be for all horses in South Africa to be tested pre-race. Will that require extra staff, is it realisable and logistically foolproof?
VM: Yes, it is definitely possible, we have done approximately 60% coverage and plans are at an advanced stage to achieve 100% over time, in all centres. We are currently in the launch phase for on-course testing and all centres have been tested historically.

How much blood is taken from a horse in this procedure, and would the amount of blood drawn affect a horse’s performance? I’m thinking in this regard of a human blood test where the doctors always warn of things like dizziness and nausea.
MB: Only 4ml of blood is collected pre-race for on course TCO2 analysis, this equates to 0.01% of the total blood volume (A normal 500kg horse has a blood volume of approximately 40L).

In regards traditionally highly-strung horses, for example last Saturday’s pre-race test subject Zeus – can the commotion, the prolonged activity around certain horses affect their performances negatively thereafter?
MB: Most horses, including Zeus, are familiar with the environment and are fairly calm at the time of pre-race specimen collection. The time from when they enter the specimen collection area to when they leave is approximately 2 minutes.

What are the fines to be imposed on offenders or serial offenders?
VM: At this juncture, scratchings are implemented based on our TC02 Standard Operating Procedures (SOPs).

For clarity: Are the trainers of horses who are scratched pre-race, subject to warnings or fines?
No answers given, on several requests made, subsequent to first questions.

If all horses were to be tested before a race day, how many veterinarians would be required to make the procedures run smoothly? This goes to a question of costs – will full-on testing of all horses require significant extra costs?
MB: We have a minimum of two veterinarians present at all race meetings and will add a third if and when necessary, dependent on the number of carded runners on the day.

I have spoken to any number of trainers, some don’t want to comment, several others say that there should rather be tests for substances like clenbuterol, seen as the substance being abused by many. Will it not be more effective to focus on tests for substances like this, instead of the TC02 tests?

MB: We screen all blood and urine specimens obtained pre- and post-race as well as out of competition very sensitively for a wide range of prohibited substances including clenbuterol. We have also put in place new IFHA mandated requirements regarding the use of clenbuterol which will come into effect on the 1st May 2024.

Here is a quote from Dr Sams of the HFL Sports Science Laboratory in the United States, from an article on TC02: “I think the trainers have made the decision it (milkshaking) just isn’t worth it. The benefit doesn’t outweigh the risk. I think maybe more importantly, I think trainers here realize that others aren’t gaining an advantage by using bicarbonate. That’s probably the greatest disincentive.”

 This quote suggests that the hype around TC02 is overblown, that is it not that much of an advantage, if at all. Your views?
VM: Alkalization of a horse on a race day is a IFHA prohibited practice regardless of causation. This is due to the potential performance enhancing effects it may have.

The substance EPO has been mentioned as something that ‘cannot be detected’ but may well be in use. Is this true, and if it is, then how will we ever stop it?
MB: EPO can be detected by our laboratory and we are currently screening for its use.

 ‘Designer Drugs’ are said to be quite easily obtainable on the internet and can be delivered to front doors. Is this a problem worthy of more focus, or are you on top of trends?
MB: We do have reference materials to routinely screen for these “designer drugs”, however upon analysis most of these preparations don’t contain what is claimed on the label. Any preparations found and confiscated which contains a prohibited substance and is not prescribed by a veterinarian or registered locally in terms of the Fertilizers, Farm Feeds, Seeds and Remedies Act 36 of 1947 or Medicines and Related Substances Act 101 of 1965 will incur severe penalties and be handed over to the authorities for further investigation.

There is a perception from trainers at Turffontein that they are in the ‘Lions Den’ due to their close proximity to the NHA. Is there fairness across the board and at all centres, or would a system of ‘satellite offices’ at training centres be a solution?
MB: Out of competition and race day specimen collection procedures are implemented fairly and consistently across all racing centres.


While the NHA’s proactive stance has been lauded in most quarters, it has sparked uncertainty among owners, trainers, and the racing public. One owner observed: “The stipes had a big security guard walking with them at the Turffontein meeting last Saturday. What are they scared of? What is our industry coming to? The stabling area is a war zone now. What has happened to sportsmanship and gentlemanliness that racing was known for?”

Not one of the 12 trainers spoken to were willing to have their names quoted – in fear of being accused of fingering fellow-trainers on the one hand; victimisation from unscrupulous NHA officials on the other. It needs to be said that all were in favour of pre-race TC02 testing. There were some that felt that the NHA were targeting specific stables as a result of pressure from ‘powerful sources’ outside of Johannesburg. Half of them felt that the hype around bi-carb was a storm in a teacup and agreed with research which plays down its effectivity.

The other half was skeptical of the research mentioned, felt that more research was needed and one trainer commented: “Milkshaking, TC02 increases the blood buffering capacity of a horse and increases the clearance of lactate from its muscles. This means that it delays fatigue in a horse during a race.

We need to know exactly what the effects are, second by split second. Years ago, we had the Lasix trials, maybe something similar should be arranged for TC02.

Think about it. An advantage of even just a split second in a race, gained by delayed fatigue, can be equivalent to a length or more at the winning post. This means the cheater gets the first prize, and over time cheating adds up in financial terms and in log positions!”

Another said: “There should be more testing of horses out of competition, which is when anabolic steroids are used. Also, the activities of veterinarians should be closely monitored. There are therapeutic steroids in use – legal within prescribed periods – but there are trainers who abuse them by ordering steroidal injections for horses who are not injured. The vets just play along.”

As can be seen from the research and comments above, and the NHA’s vague answers to some pertinent questions, the TC02 issue is complex, subject to examination and has potential legal implications for the NHA, who must be aware of this. As things stand at this writing, no punishments have been declared for positive pre-race tests and, with no post-race tests planned, there cannot possibly be positive samples.

But despite this apparent Catch 22, it is conceivable (and even probable) that the NHA’s emphasis on TCO2 testing is intentionally crafted to dissuade trainers who have employed milkshaking as a means to conceal other substances, from using bicarbonate altogether.

The masking effect seems to be most effective for drugs such as lidocaine, procaine and cocaine, but as per Dr Bawden the NHA are aware of, and hot on the heels of other prohibited substances. Eliminating milkshaking will curb or eliminate, by implication, a range of dangerous and harmful substances.

Dr Lindinger’s conclusive view supports this: “A TCO2-only test is not capable of being used to identify horses that are health-optimized versus those that have been intentionally manipulated to enhance performance. Such testing does exist and is straightforward to implement.”

The Conversation’s final assessment will serve as our own conclusion: “(TC02) is an inappropriate test and provides no recourse for those horsemen that have not done anything wrong. Milkshakes are already banned. Our research shows they deliver no benefit anyway. Trainers who are happy to continue this illicit practice and run the gauntlet of potential sanctions should consider whether it is worth it at all, and whether instead they should reconsider on moral, medical and scientific grounds.”


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