Bacterial Endotoxin Testing: A Mandatory Requirement for Key Pharmaceutical Products

2026-02-05

The primary objective of Bacterial Endotoxin Testing (BET) is to control the risk of pyrogens in pharmaceutical products and to prevent serious adverse reactions in humans, such as fever and shock. According to major pharmacopoeial standards, including the United States Pharmacopeia (USP) and the European Pharmacopoeia (Ph. Eur.), medicinal products that directly enter the bloodstream, come into contact with mucous membranes or damaged tissues, or are used in surgical or implantable procedures are subject to mandatory endotoxin testing.

1. Injectable Drugs (Core Mandatory Testing Category)

Injectable drugs are administered directly into the human circulatory system. As a result, excessive endotoxin levels present the most immediate and significant safety risk. Therefore, injectable products represent a primary focus of Bacterial Endotoxin Testing (BET) and include all dosage forms intended for parenteral administration.

Intravenous Injections
All intravenous preparations are subject to mandatory endotoxin testing, including:
Large-volume parenterals (LVPs, ≥50 mL)
Small-volume parenterals (SVPs, <50 mL)

Typical examples include antibiotic injections, vitamin

injections, electrolyte solutions for infusion, and injectable chemotherapeutic agents.

Intramuscular and Subcutaneous Injections
Most intramuscular and subcutaneous preparations require endotoxin testing, particularly those intended for:
Treatment of severe or critical conditions
Long-term or repeated administration
High-concentration formulations

Examples include insulin injections and vaccine adjuvant preparations.

Injectable Sterile Powders and Lyophilized Preparations
Sterile powders and lyophilized injectable products must be tested for endotoxins after reconstitution, such as injectable cephalosporin antibiotics and lyophilized biological products.

2. Ophthalmic Preparations (Mucosal Contact Category)

Ocular mucosal tissues are highly sensitive; therefore, stringent endotoxin control requirements apply to ophthalmic products intended for intraocular administration or surgical use.

Intraocular Injections
Preparations intended for intravitreal or anterior chamber administration must undergo endotoxin testing, such as injectable anti-angiogenic therapies.
Ophthalmic Preparations for Surgical Use
Irrigation solutions, viscoelastic agents, and related auxiliary products used in cataract and glaucoma surgery are required to comply with BET requirements.
Ophthalmic Solutions (Eye Drops)
Conventional eye drops must also be tested for endotoxins if they are labeled “for use in corneal injury”or classified as sterile preparations.

3. Blood Products and Biological Products

Due to their complex raw material sources—such as human plasma, recombinant proteins, and microbial fermentation–derived materials—these products present a high inherent risk of endotoxin contamination and are therefore subject to mandatory bacterial endotoxin testing.

Blood Products
Including human serum albumin, immunoglobulins, coagulation factors, and related plasma-derived products.
Vaccines
Including inactivated vaccines, live attenuated vaccines, and recombinant vaccines. Endotoxin levels must be controlled in both raw materials and finished products.
Recombinant Biological Drugs
Including monoclonal antibodies, cytokines, enzyme-based preparations, and other recombinant biotherapeutics.

4. Dialysis and Perfusion Preparations

These preparations are used for direct substance exchange with the bloodstream. Excessive endotoxin levels may induce dialysis-related febrile reactions; therefore, such products are subject to mandatory bacterial endotoxin testing.

Dialysis Solutions
Including hemodialysis solutions and peritoneal dialysis solutions.
Perfusion and Extracorporeal Circulation Fluids
Including organ preservation and transplant perfusion solutions, as well as extracorporeal circulation fluids.

5. Medications Used in Conjunction with Implantable or Interventional Medical Devices

Pharmaceutical products used in conjunction with medical devices that remain in long-term contact with human tissues or are introduced into the body require concurrent control of endotoxin levels.

Excipients for Implantable Products
Such as sustained-release microspheres and solvents used with medical adhesives.
Irrigation Solutions for Interventional Procedures
Including catheter flushing and irrigation solutions used in vascular and neurointerventional procedures.

6. Other High-Risk Preparations

Topical Preparations for Damaged Skin or Mucous Membranes
Including sterile solutions used with burn ointments, ulcer irrigation solutions, and oral rinses.
Cavity-Use Preparations
Including sterile enema solutions and uterine irrigation solutions.

Supplementary Notes

Conventional Topical and Oral Preparations
Topical formulations intended for intact skin (e.g., ointments and creams) and oral dosage forms (e.g., tablets and capsules) generally do not require endotoxin testing, unless there is a specific risk of contamination arising from raw materials or the manufacturing process.
Endotoxin Limits According to Pharmacopoeias
Different pharmacopoeias specify limits for endotoxin levels (e.g., EU/mL, EU/mg) for various types of pharmaceutical products. These limits must be interpreted and applied in accordance with the route of administration and the intended dosage.

Endotoxin Limit Value Comparison Table for Common Mandatory-Test Pharmaceuticals

Pharmaceutical Categories	Typical Variety	Endotoxin Limits	Units	Core Basis / Application Notes
Water for Injection	Water for Injection, Sterile Water for Injection	≤0.25	EU/mL	· General Chapters of Chinese Pharmacopoeia as the baseline endotoxin limit for injection solvents and the internal control threshold for most injection preparations.Issued by the Pharmacopeial Convention
Large-Volume Intravenous Infusions (≥100 mL)	Sodium Chloride Injection, Glucose Injection, Compound Electrolyte Infusion	≤0.50	EU/mL	· Specified in Pharmacopoeia: This limit shall not be exceeded for large-volume infusions (≥ 100 mL), to ensure the safety of high-dose administration.Issued by the Pharmacopeial Convention
Small-Volume Intravenous Injections (＜100 mL)	Cephalosporin Antibiotics for Injection, Vitamin C Injection	≤0.50	EU/mL	· Calculated per the formula L=K/M (where K=5 EU/(kg·h)). The limit is established in combination with the maximum clinical dosage, with stricter requirements for some high-risk formulations.
Sterile Powders for Injection	Piperacillin Sodium and Tazobactam Sodium for Injection, Lyophilized Human Serum Albumin	≤0.25-0.50	EU/mg	· Tested after reconstitution. The limit is converted based on the dosage of active pharmaceutical ingredients (APIs) / finished preparations. For biological products, the limit is typically controlled at 0.25 EU/mg.
Intramuscular/ Subcutaneous Injections	Insulin Injection, Thymopeptide for Injection	≤0.50-2.0	EU/mL	· Adjusted according to administration frequency and dosage. Stricter limits apply to long-term administration / high-concentration formulations.
Intraocular Injections	Anti-VEGF Intravitreal Injection, Intraocular Perfusion Solution	≤0.20	EU/mL	· Given the high sensitivity of intraocular mucous membranes, the limit refers to the standards for ophthalmic preparations in ChP. For some formulations, the limit is controlled at 0.1 EU/mL.
Blood Products	Human Serum Albumin, Human Immunoglobulin for Intravenous Injection	≤0.50	EU/mL	· In accordance with the General Principles for Biological Products in Pharmacopoeia, the limit is calculated based on the maximum dosage per kilogram of body weight per hour to ensure infusion safety.
Vaccines	Inactivated Influenza Vaccine, Recombinant Hepatitis B Vaccine	≤0.10-0.25	EU/mL	· Balancing immunogenicity and safety, the limit for vaccine adjuvant products is typically ≤ 0.1 EU/mL.
Dialysate	Hemodialysis Solution, Peritoneal Dialysis Solution	≤0.50	EU/mL	· To avoid dialysis-related pyrogen reactions, the internal control limit for some products is ≤ 0.25 EU/mL.
Intrathecal/ Intraspinal Injections	Spinal Anesthetics, Intrathecal Chemotherapeutic Agents	≤0.20	EU/mL	· Due to the high risk to the central nervous system (CNS), the K value is set at 0.2 EU/(kg·h), making the limit far stricter than that for intravenous administration.