The "backside" of aspirin (Acetylsalicylic acid): from molecular mechanism to pharmacopoeial impurity spectrum

2026-09-03 · CATO · CATO Standards

From willow bark to aspirin: The birth of acetylation modification

Using willow bark to relieve pain and reduce fever is an ancient thing. The real turning point occurred in the 19th century: chemists isolated salicylic acid from willow bark. It was indeed effective, but it was irritating to the stomach and tasted very bad, making it difficult to take for a long time.

In 1853, French chemist Charles Frédéric Gerhardt synthesized acetylsalicylic acid for the first time by reacting salicylic acid with acetic anhydride. However, the product was impure and unstable, and this work did not attract attention at the time.

The person who really turned it into a "medicine" was Felix Hoffmann, a chemist at Bayer. According to Bayer’s official historical records,

On August 10, 1897, he acetylated salicylic acid with acetic anhydride to obtain chemically pure and stable acetylsalicylic acid (ASA) for the first time; it was launched under the trade name Aspirin in 1899.

The reaction itself is not complicated:

Salicylic acid + acetic anhydride → acetylsalicylic acid (aspirin) + acetic acid

This esterification reaction also laid the foundation for future impurity and stability issues.

How aspirin works: Mechanism of irreversible inhibition of COX

In 1971, British pharmacologist John Vane proposed that aspirin drugs work by inhibiting the synthesis of prostaglandins. This discovery earned him the 1982 Nobel Prize in Physiology or Medicine together with Sune Bergström and Bengt Samuelsson.

CAS No.:50-78-2

The molecular details of the mechanism are more interesting for technical people: aspirin inhibits cyclooxygenase (COX, prostaglandin H₂ synthase). Unlike most NSAIDs "reversible competition"

, aspirin covalently transfers its acetyl group to a serine residue (Ser-530 of COX-1) in the COX active site channel, causing steric hindrance and preventing the substrate arachidonic acid from entering - this is an irreversible inhibition.

(Source: J. Med. Chem. 2007, 50(7) review)

This also explains the antiplatelet logic of low-dose aspirin: platelets have no nuclei, cannot resynthesize COX-1, and once acetylated, their production of thromboxane A₂ is turned off throughout the platelet's life cycle. This is the molecular basis of "long-term low-dose administration of cardiovascular and cerebrovascular drugs".

Aspirin stability: hydrolytic degradation and influencing factors

That beautiful synthesis reaction in the first section,

In turn, this is the biggest stability weakness of aspirin - the ester bond will hydrolyze.

In a humid environment, solid aspirin will slowly hydrolyze to regenerate salicylic acid and acetic acid; salicylic acid is its main degradation product.

(Source: Decomposition of Aspirin in the Solid State and other studies)

Multiple stability studies have consistently stated:

Humidity is the most critical degradation inducement, followed by temperature and light; the content recovery rate can significantly decrease under high humidity conditions.
(Source: J. Pharm. Res. Int. 2025 stability evaluation)
The hydrolysis rate is pH-dependent: it is relatively stable in the acidic environment of the stomach and accelerates hydrolysis in alkaline conditions (such as the small intestine).
(Source: ECSOC-28 Conference Paper, MDPI, 2024)

For analytical chemistry, here is a classic detail:

Salicylic acid has significant UV absorption at about 300 nm, while aspirin absorbs almost nothing in this region, and this difference is the basis for the stability-indicating assay.

(Source: Aspirin Degradation Kinetics Experimental Lecture Notes)

For this reason, the pharmacopoeias of various countries list free salicylic acid as a key limit indicator for aspirin raw materials and preparations. To "measure accurately and control", the premise is that you must have a salicylic acid reference substance with clear identity and traceable value.

Quality control of aspirin: from pharmacopoeial standards to impurity analysis

Modern drug quality research is not satisfied with "whether the main ingredients are enough", but also needs to answer "what else is in it and how much of each". ICH and various pharmacopoeias have established a complete set of identification and limit systems for specified impurities. Each impurity must be independently characterized and quantified, which requires corresponding impurity reference standards.

Take the acetylsalicylic acid varieties in the European Pharmacopoeia (Ph. Eur.)/British Pharmacopoeia (BP) as an example.

The specific impurities specifically listed are A, B, C, D, E, and F.

(letter

Source: Ph. Eur. / BP Aspirin monograph)

In short, this table depicts the complete life cycle of a molecule from synthesis to degradation: impurities A/B originate from raw materials and processes, D/E/F originate from esterification and condensation between molecules, and impurity C (salicylic acid) has dual identities of process residue and storage degradation, and is a key indicator of aspirin quality.

CATO provides aspirin impurity control standards

CATO has built a relatively complete reference product line around acetylsalicylic acid, covering multiple levels from pharmacopoeial impurities to isotope internal standards, metabolites, salt forms and synthetic related substances:

① Pharmacopoeia Impurity Reference Standard (EP/BP system, full spectrum coverage from A to F)

② Process/degradation related impurities (including dimerization and condensation)

Covers aspirin impurity 8-17, 19-31 and other series, such as trisalicylic acid (C4X-140828, CAS 85531-17-5) and multiple dimerization/trimerization condensation impurities, meeting the needs of in-depth impurity spectrum research.

③ Stable isotope internal standard

Acetylsalicylic acid-D₃ (C1788333, CAS 921943-73-9)—an internal standard for LC-MS/MS quantification, is a common tool to improve accuracy in biological sample and assay determinations.

④ Metabolite reference substance

Acetylsalicylic acid acyl-D-glucuronate (CP129951, CAS 24719-72-0) - the glucuronic acid conjugation product of phase II metabolism of aspirin in vivo, serving metabolism and pharmacokinetics research.

⑤ Salt forms and synthetic related substances

DL-lysine acetylsalicylate (lysine acetylsalicylate, C4X-2211, CAS 62952-06-1), O-acetyl salicylic acid chloride (C1790086, CAS 5538-51-2), etc., covering water-soluble salt forms and synthetic intermediates.

As a supplier with ISO 17034 reference material producer qualification (CNAS RM0030 / ANAB AR-2832 dual accreditation), CATO can provide value traceability and supporting analysis data support for impurity reference standards.

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References

Bayer official website "Felix Hoffmann": https://www.bayer.com/en/history/felix-hoffmann
Science History Institute "Felix Hoffmann": https://www.sciencehistory.org/education/scientific-biographies/felix-hoffmann/
The Nobel Prize official website "The Nobel Prize in Physiology or Medicine 1982": https://www.nobelprize.org/prizes/medicine/1982/vane/facts/
Structural and Functional Basis of Cyclooxygenase Inhibition, J. Med. Chem. 2007, 50(7): https://pubs.acs.org/doi/pdf/10.1021/jm0613166
European Pharmacopoeia / British Pharmacopoeia Aspirin (acetylsalicylic acid) monograph (Impurities A–F): https://www.drugfuture.com/Pharmacopoeia/BP2012/data/8077.html
European Pharmacopoeia 7.0 Acetylsalicylic acid (Related Substances/Impurities Relatively Retained): https://www.drugfuture.com/Pharmacopoeia/EP7/DATA/0309E.PDF
Decomposition of Aspirin in the Solid State (Solid State Hydrolytic Degradation): https://pubmed.ncbi.nlm.nih.gov/3379588/
Stability Evaluation of Acetylsalicylic Acid ... under Different Storage Conditions, J. Pharm. Res. Int. 2025
The Effect of pH on the Hydrolysis of Acetylsalicylic Acid, ECSOC-28, MDPI, 2024: https://sciforum.net/paper/view/20185