Doping control analysis of TB-500, a synthetic version of an active region of thymosin β₄, in equine urine and plasma by liquid chromatography-mass spectrometry.
Ho. Emmie N M EN; Kwok. W H WH; Lau. M Y MY; Wong. April S Y AS; Wan. Terence S M TS; Lam. Kenneth K H KK; Schiff. Peter J PJ; Stewart. Brian D BD
Key Findings
- A liquid chromatography‑mass spectrometry method can reliably detect N‑acetylated LKKTETQ (TB‑500) in equine plasma at 0.02 ng/mL and urine at 0.01 ng/mL.
- The study identified the main metabolites of TB‑500 in vitro and confirmed their presence in samples from treated horses.
- Detection was successful after a single 10 mg dose, showing the test can catch even low‑level, one‑time use.
Practical Outcomes
- For biohackers, the study mainly shows that TB‑500 can be traced in biological fluids at very low levels, which is useful for anti‑doping labs but offers little direct guidance on how to use the peptide for health or performance. It suggests that if you use TB‑500, it could be detectable in urine/plasma, potentially affecting sports eligibility or future testing.
Summary
Scientists created a very sensitive lab test that can spot the peptide TB-500 (a synthetic piece of thymosin β4) and its breakdown products in horse urine and blood. They can detect it at tiny amounts (0.01‑0.02 ng/mL) after a single 10 mg dose. This is the first time the drug has been proven detectable in horses after use.
Abstract
A veterinary preparation known as TB-500 and containing a synthetic version of the naturally occurring peptide LKKTETQ has emerged. The peptide segment (17)LKKTETQ(23) is the active site within the protein thymosin β(4) responsible for actin binding, cell migration and wound healing. The key ingredient of TB-500 is the peptide LKKTETQ with artificial acetylation of the N-terminus. TB-500 is claimed to promote endothelial cell differentiation, angiogenesis in dermal tissues, keratinocyte migration, collagen deposition and decrease inflammation. In order to control the misuse of TB-500 in equine sports, a method to definitely identify its prior use in horses is required. This study describes a method for the simultaneous detection of N-acetylated LKKTETQ and its metabolites in equine urine and plasma samples. The possible metabolites of N-acetylated LKKTETQ were first identified from in vitro studies. The parent peptide and its metabolites were isolated from equine urine or plasma by solid-phase extraction using ion-exchange cartridges, and analysed by liquid chromatography-mass spectrometry (LC/MS). These analytes were identified according to their LC retention times and relative abundances of the major product ions. The peptide N-acetylated LKKTETQ could be detected and confirmed at 0.02 ng/mL in equine plasma and 0.01 ng/mL in equine urine. This method was successful in confirming the presence of N-acetylated LKKTETQ and its metabolites in equine urine and plasma collected from horses administered with a single dose of TB-500 (containing 10mg of N-acetylated LKKTETQ). To our knowledge, this is the first identification of TB-500 and its metabolites in post-administration samples from horses.
Study Information
pubmed
2012
2012-09-23T00:00:00.000Z
10.1016/j.chroma.2012.09.043
28
20