Carbon-14 has been used as a tracer for decades, but there has been a fundamental shift in how chemists design, make and care for ¹⁴C-labelled compounds. The focus has moved from simply “getting a label in somewhere” to ensuring it stays in the right place, in the correct chemical form, throughout the lifetime of a study.
This article examines recent developments in the stability of carbon-14 labelled compounds, including synthetic methods and label placement, GMP storage, new drug modalities, and regulatory expectations.
What “stability” means for ¹⁴C-labelled compounds
When people talk about stability in this context, they are usually wrapping together three separate questions.
First is nuclear stability. ¹⁴C has a half-life of about 5,730 years and emits low-energy β radiation, so the isotope itself does not fade away on any practical timescale for ADME, environmental or mechanistic studies.
Second is chemical and radiochemical stability. The labelled active ingredient must remain chemically intact and have high radiochemical purity during synthesis, storage, formulation, and dosing. Peroxide formation, oxidation, hydrolysis, or β-induced degradation can generate side products and reduce specific activity if the material is mishandled.
Third is the metabolic stability of the label position. For human mass balance, QWBA, or environmental fate work, the key requirement is that the ¹⁴C atom remains covalently attached to drug-related material throughout absorption, distribution, and metabolism. Poor placement of the label can lead to early loss of ¹⁴CO₂ or a small fragment, undermining the interpretation of the whole study.
Recent developments touch all three dimensions, but the biggest gains have come from chemistry and study design rather than physics.
Late-stage labelling and isotope exchange
One of the most critical shifts has been the move towards late-stage ¹⁴C labelling. Traditional radiosynthesis often required building the whole molecule from a simple ¹⁴C synthon, such as barium carbonate or a labelled carboxylic acid, subjecting the radiolabel to many steps, high temperatures and long reaction times. This consumed expensive ¹⁴C, generated waste and could erode radiochemical purity.
The new methodology now allows chemists to introduce ¹⁴C at or near the final step. Babin and co-workers reviewed a range of late-stage carbon isotope labelling and isotope-exchange strategies, including palladium-catalysed carbonylation, decarboxylative carboxylation and direct carbonyl exchange at aryl or heteroaryl positions.
More recently, Batista and colleagues reported a palladium-catalysed electrocarboxylation platform that can use near-stoichiometric ¹⁴CO₂, generated in situ from Ba¹⁴CO₃, to label complex aryl electrophiles. This set-up is designed with ¹⁴C in mind: it tolerates pharmaceutical complexity and avoids the huge excess of CO₂ gas often used in non-radioactive methods.
From a stability perspective, late-stage approaches offer several advantages:
- The label spends less time under harsh synthetic conditions, cutting down the risk of degradation and side-product formation.
- The chemist can choose the label position once metabolism data are available, targeting robust aromatic carbons, core amides or heterocycles rather than labile termini.
- Because the label is installed on the final scaffold, there is no need to re-optimise an entire synthetic route just to accommodate ¹⁴C, which reduces development time and waste.
Isotope-exchange methods push this even further. For some scaffolds, it is now possible to replace a non-radioactive carbon atom with ¹⁴C in a single step, without altering the rest of the molecule. This is particularly attractive for back-labelling clinical candidates that are already in development pipelines.
Smarter label placement and the move to multiple labels
Parallel to advances in synthesis, there has been a clear trend towards more systematic label placement. Guidance from regulators and industry groups emphasises that the radionuclide position should be both chemically and metabolically stable, so that parent drug and metabolites relevant for safety assessments carry the label.
A recent recommendations paper on the use of multiple labels in human mass balance studies outlines strategies for cases where a single label is insufficient. Cuyckens and co-workers discuss workflows in which ¹⁴C is combined with ³H, or two ¹⁴C atoms are positioned in different parts of the molecule to follow distinct biotransformation routes.
Several themes have emerged:
- For small molecules, the preferred positions are often aromatic carbons or “hard” sp²/sp³ carbons within the core, away from obvious metabolic soft spots such as terminal carboxylates, benzylic positions or labile linkers.
- For drugs that undergo extensive cleavage, a single label on a side chain may end up on a minor metabolite or be lost as CO₂. In such cases, adding a second label on a different ring or on the backbone can preserve a reliable signal for total drug-related material.
- For very long-acting drugs and depot formulations, metabolic robustness is essential: if sampling lasts for months, any tendency for the label to be released as small gaseous or one-carbon species will distort the apparent terminal half-life.
This way of thinking turns label placement into an integral part of DMPK study design, rather than a late technical detail.
Radiochemical stability, GMP practice and storage
Once a suitable label has been installed, maintaining its integrity during storage and use is the next challenge. Contract manufacturers and pharma isotope groups have published more data and guidance in the last few years on radiochemical stability of ¹⁴C APIs.
A recent technical report examines how various structural motifs behave under different storage temperatures (2–8 °C, −20 °C, −80 °C) and how factors such as oxygen exposure and light contribute to degradation. Some functionalities, including activated aromatics and electron-rich heterocycles, are more susceptible to oxidative or radiolytic processes, making colder conditions and an inert atmosphere particularly beneficial.
A recent review outlines broader considerations for ¹⁴C-labelled APIs in a GMP setting, covering how radiolysis, hydrolysis, and oxidative processes interact with formulation choices, and how analytical techniques such as HPLC with radiometric detection, LC–MS, and specific-activity measurements are used to confirm label integrity over time.
A common pattern in current practice is:
- Storage of solids at low temperature in amber glass, often under nitrogen or argon.
- Minimising time in solution; preparing dosing formulations shortly before use rather than keeping thawed solutions for long periods.
- Periodic retesting of material that has been on the shelf, with repurification by preparative HPLC or recrystallisation if radiochemical purity drifts below pre-set limits.
Quality standards documents for ¹⁴C APIs used in human studies stress that there is no separate pharmacopoeial monograph for these materials. Instead, acceptable stability and purity specifications must be defined on a case-by-case basis and agreed upon in quality agreements between the synthesis site and the clinical manufacturing unit.
Extending stable labelling to new modalities
Small-molecule drugs are no longer the only focus. The rapid rise of oligonucleotides, peptides, antibody–drug conjugates (ADCs) and other large modalities has pushed radiochemists to think differently about stability.
Edelmann’s review on radiolabelling small and biomolecules describes the challenges of incorporating ¹⁴C, ³H, and other radionuclides into peptides, proteins, and oligonucleotides without disrupting their function. Later work from the same author and others examines nucleic acid-based therapeutics in more detail, highlighting how backbone modifications and sugar chemistry influence both metabolic stability and label retention.
For oligonucleotides, the favoured approach is often to place ¹⁴C in a stable base or sugar unit that survives exonuclease trimming long enough to inform on distribution and clearance, or in a ligand such as GalNAc used for targeted delivery to the liver.
In the ADC field, researchers have outlined strategies for labelling both payloads and antibodies to monitor their distinct in vivo routes. When analytical sensitivity allows, ¹⁴C is increasingly chosen for payloads because it offers a chemically and metabolically dependable tag with relatively low radiolysis risk. Method-development studies on ¹⁴C-labelled IgG antibodies, reported in 2024, examine storage conditions and formulation parameters that help maintain radiochemical integrity while preserving the native protein structure.
Beyond therapeutics, ¹⁴C labelling has also been applied to study polymer degradation and environmental distribution of complex materials, where the label often needs to remain attached through lengthy weathering or biodegradation protocols. Once again, the design challenge is to anchor the isotope in a part of the polymer that best represents the material’s long-term fate.
¹⁴C versus tritium: stability and data quality
Tritium labelling still has an important role, particularly when extremely high specific activity or very low doses are required. However, experience has shown that ³H is more prone to exchange with solvent or biological media, and that β radiation from tritium can trigger degradation at sensitive positions.
For many applications, this has reinforced ¹⁴C as the workhorse isotope:
- It provides a clear, non-exchangeable tag when placed on carbon atoms that do not undergo rapid metabolic transformations.
- Its low β energy reduces radiolytic issues, especially when combined with appropriate storage and packaging.
- The limitation of lower specific activity can be overcome in human microtracer studies by using accelerator mass spectrometry (AMS), allowing doses in the microcurie range while still obtaining precise concentration-time profiles.
Current clinical pharmacology guidance supports these trends, encouraging the use of well-designed ¹⁴C human ADME studies early in development, sometimes even before extensive animal work, provided dosimetry and safety limits are respected.
Where things are heading
Taken together, these developments paint a clear picture of how work with ¹⁴C-labelled compounds is evolving.
Late-stage labelling and isotope exchange are making synthesis more efficient, flexible and kinder to the label. Method developers are now designing catalytic reactions and electrochemical set-ups specifically with ¹⁴C use in mind, rather than simply adapting cold chemistry.
At the same time, study designers are treating the label position as a critical aspect of ADME and safety strategy. Guidance on multiple labels, human-first ADME approaches, and acceptable recovery thresholds is tightening the link between radiochemistry and regulatory expectations.
On the manufacturing side, isotope laboratories are publishing more stability data, setting clearer expectations for storage, retest intervals and repurification routes for ¹⁴C APIs and drug products used in clinical trials.
Finally, the expansion into oligonucleotides, peptides, biologics and advanced materials is forcing radiochemists to consider structural biology, linker design and macromolecular stability alongside classic small-molecule concerns.
Disclaimer
The information presented in this article is intended for scientific, educational, and informational purposes only. It does not constitute professional advice on radiochemical synthesis, regulatory compliance, manufacturing practice, safety assessment, or any related activity. Procedures, methods, and approaches described here may not be appropriate for every laboratory, organisation, or project, and outcomes can vary depending on equipment, infrastructure, technical expertise, and regulatory requirements.
Readers should not rely solely on this article when making decisions about the design, preparation, handling, storage, analysis, or application of carbon-14 labelled compounds. All work involving radioactive materials must follow applicable laws, guidance, institutional policies, and health and safety standards, and should be carried out by trained personnel using appropriate facilities and controls.
The authors and publisher accept no responsibility for any loss, damage, or adverse outcome arising from the use of the information discussed. Users are strongly encouraged to consult qualified specialists, regulatory authorities, and up-to-date technical references before undertaking any radiochemical or regulatory activity.
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