Use of Drugs and Medical Materials in Research Animals

Background

Federal animal welfare regulations1 require that animals used for research receive adequate veterinary care2.  This includes using drugs, experimental diets, and medical materials before they expire and using pharmaceutical–grade drugs when available.  Policy statements and guidance have been developed to address specific issues including but not limited to:

  • Expiration Dates and Beyond-Use Dates
  • Non-Pharmaceutical Grade Substances
  • Controlled Substances
  • Experimental Diets
  • Recordkeeping Requirements
  • Drug Inventory Management
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Definitions

Expiration Date – Manufacturer-assigned date indicating the period during which an unopened product is expected to remain stable and suitable for use when stored according to labeled conditions.

Beyond-Use Date (BUD) – A type of expiration date assigned after a drug or preparation has been opened, punctured, diluted, reconstituted, compounded, or otherwise altered.  The BUD establishes the period during which the preparation may be used under specified storage conditions.  See Institutional BUD Guidance below (link)

Pharmaceutical Grade - A drug, biologic or reagent manufactured under Good Manufacturing Practices (GMP) and approved, conditionally approved, or indexed by the U.S. Food and Drug Administration (FDA), or for which a chemical purity standard has been established by the United States Pharmacopeia-National Formulary or British Pharmacopeia. Pharmaceutical-grade agents are intended for use in human or veterinary medicine and meet the highest standards for purity and bioavailability.3

Non-Pharmaceutical-Grade (NPG) - A drug, chemical, biologic, reagent, or other substance that does not meet the definition of a pharmaceutical-grade compound. Examples include laboratory-grade chemicals, research compounds, and substances prepared from bulk chemical ingredients.

Compounded or In-House Prepared Drugs – any substance that has been mixed, diluted, reconstituted, transferred to another container, or otherwise modified from its original form.

Acute/terminal procedure - A surgical or non-surgical procedure for which an animal receives an anesthetic agent and is euthanized without recovering from anesthesia.

Survival procedure - A surgical or non-surgical procedure performed on an animal that is subsequently allowed to recover from anesthesia.

Experimental Diets – Any non-standard feed including medicated diets, purified ingredient diets, high-fat diets, irradiated diets or researcher-defined formulations.

Medical materials - Items used clinically or for research purposes that may have an expiration date assigned by the manufacturer for the purpose of guaranteeing sterility.

Policy

In accordance with the Animal Welfare Act Regulations, PHS Policy, The Guide for the Care and Use of Laboratory Animals, and accepted veterinary medical practice, individuals involved in animal research must follow the requirements below to ensure appropriate use of drugs, chemicals, diets, and medical materials. These requirements are intended to protect animal welfare, ensure adequate veterinary care, and maintain the integrity of research outcomes.

Individuals involved in animal care and use must comply with the following requirements:

  1. Expired Drugs and Medical Materials

Drugs and medical materials must not be used after their expiration date or beyond-use date, whichever occurs first, unless this policy specifically permits such use.

Anesthetics, analgesics, euthanasia agents, therapeutic drugs, and emergency medications must not be used after their expiration date or beyond-use date, whichever occurs first, under any circumstances, including terminal procedures.

Expired drugs and medical materials (excluding anesthetics, analgesics, euthanasia agents, therapeutic drugs, and emergency medications) may only be used in terminal procedures when their use is not expected to:

    • Adversely affect animal welfare;
    • Compromise anesthesia or euthanasia;
    • Interfere with clinical care; or
    • Negatively affect the scientific objectives of the study.

If uncertainty exists regarding the suitability or potential impact of any expired drug or medical material, consult with a ULAR veterinarian.

  1. Controlled Substances

Controlled Substances administered to research animals must comply with the preceding IACUC policies.  Failure to follow DEA requirements could result in the loss of the institutional license to purchase these agents. In addition, Controlled Substances are subject to additional policies regarding their purchase, storage, use, records/documentation, and disposal.

Review the EHS Controlled Substances institutional requirements. More information is also available at the IACUC Requirements for Use of Controlled Substances page.

  1. Pharmaceutical-Grade Drugs

Pharmaceutical-grade medications, chemicals, and other substances must be used whenever they are available and appropriate for use in animals, including in terminal procedures.

  1. Non-Pharmaceutical-Grade (NPG) Drugs

Use of non-pharmaceutical-grade drugs or compounds in animals must be reviewed and approved by the IACUC before use.

The IACUC may approve such use when the protocol includes scientific justification or demonstrates that no acceptable veterinary- or human-pharmaceutical-grade alternative is available. Cost savings alone do not justify the use of NPG compounds.

Reconstitution, dilution, aliquoting, or routine mixing of pharmaceutical-grade products using accepted aseptic techniques does not, by itself, render a preparation NPG. However, preparations may be considered NPG when personnel prepare them from bulk chemical ingredients, incorporate non-pharmaceutical-grade components, create novel formulations for which quality or stability cannot be adequately established, or otherwise substantially modify the original product in a manner that introduces uncertainty regarding sterility, stability, purity, potency, or safety.

Protocol submissions requesting use of NPG compounds must include sufficient detail to evaluate animal welfare and scientific impact, including:

    • Source and chemical composition of the compound;
    • Rationale for use in lieu of a pharmaceutical-grade alternative;
    • Procedures for preparation, dilution, or formulation;
    • Sterilization methods, when applicable; and
    • Storage conditions, including container type (e.g., glass or plastic), protection from light or other environmental conditions, whether stored in single- or multi-use containers (e.g., multi-dose injection vials), and conditions required to maintain stability, sterility, and integrity, including temperature and duration of storage.

  1. Avertin® (Tribromoethanol)

    Tribromoethanol (TBE), formerly available commercially as Avertin®, is no longer available in a pharmaceutical grade form and can only be prepared using chemical-grade ingredients. Multiple reports in the scientific literature associate Avertin® with significant side effects including peritonitis, ileus, and death, particularly following repeated administration5. Furthermore, anesthesia duration associated with tribromoethanol may be inconsistent. For most procedures, pharmaceutical-grade anesthetics are recommended as an alternative to tribromoethanol. IACUC may approve the use of tribromoethanol as a non-pharmaceutical grade drug (NPG) according to the policies on this page.

  2. Experimental Diets (e.g., medicated feed, high-fat diets)

    Experimental diets, including but not limited to medicated diets, high-fat diets, and purified diets, must not be used past the manufacturer’s expiration date or before six (6) months after the mill date, unless longer use is specifically recommended or advised by the manufacturer.

    Manufacturer’s instructions for storage and handling of diets must be followed unless otherwise approved by the IACUC or Attending Veterinarian based on scientific justification.

  1. Medical Materials and Surgical Supplies

    Medical materials and surgical supplies must be stored, handled, and used in a manner that maintains sterility, functionality, and suitability for the intended procedure.

    Medical materials include, but are not limited to, sutures, surgical drapes, gloves, gauze, catheters, needles, syringes, wound clips, and other disposable or reusable items used in animal procedures.

    Medical materials must not be used in animals when their sterility, integrity, or functionality has been compromised in a way that could adversely affect animal welfare or study outcomes.  Expired or non-sterile medical materials may be used for terminal procedures as long as their use does not affect study outcomes.

    Investigators must consult with the Attending Veterinarian or designee when uncertainty exists regarding the suitability of any expired or compromised medical material.

  1. Recordkeeping for Drug Inventories

    Principal Investigators (PIs) are responsible for maintaining drug inventories for any drugs or agents used in live animals. Records must include expiration dates for all protocol-approved drugs/agents. See sample drug inventory log template.

    Drugs and agents used for research in live animals must be appropriately labeled, including those removed from original packaging for mixing or preparation. This requirement applies to all compounded and diluted drugs, regardless of whether their components are pharmaceutical grade. At minimum, storage vials must be labeled with:

    • Name of the compound,
    • Concentration(s),
    • Date it was mixed,
    • Expiration date(s) of the original pharmaceutical grade drug(s), if applicable,
    • Beyond Use Date (BUD), or use by date, as approved in the AUP, or based on institutional guidance (above),
    • Name of the individual who prepared the mixture.

Guidance and Additional Information

Beyond-Use Date (BUD)

Manufacturer Beyond-Use Date (BUD) information may be in the package insert or on the vial labels, indicating the period that the drug can be used after opening or puncturing the vial, after a certain number of vial punctures, or other criteria.

A BUD should be established for each NPG substance and described in the approved animal use protocol.  Consideration should be given to factors that affect stability and sterility of the drug under specific storage conditions including type of container and storage temperature.

Unless manufacturer instructions or peer-reviewed published stability data support a different timeframe, the following institutional BUDs apply:

Preparation Type Institutional Default BUD
Multi-dose injection vials after first puncture (when no manufacturer BUD guidance exists) 30 days
Compounded injectable mixtures without published stability data Prepare in the smallest practical volume and use within a short period consistent with aseptic handling. Use within 72 hours.

 

Examples of peer-reviewed published stability data for selected drugs:

Preparation Published BUD
Diluted buprenorphine HCl Stable up to 180 d after dilution in sterile, bacteriostatic saline and stored in sterile glass multidose injection vials. Not stable if stored in plastic containers.6
Diluted carprofen Stable up to 180 d refrigerated after dilution in sterile non-bacteriostatic saline and stored in sterile glass multidose injection vials.7
Ketamine-xylazine diluted in sterile saline Stable up to 3 months stored at room temperature in sterile glass multidose injection vials.8

 

Drug Inventory Management

  • The IACUC recommends that PIs discuss drug inventory tracking during lab group meetings.
  • The expiration or BUD label should be visually prominent (colored, highlighted, etc.).
  • Distinguish expired items from in-date items (e.g., mark expired items with “Not For Use in Animals” label, designate different shelves within the same drug locker, place expired items into paper bag within the same storage container – Consult with EHS to ensure you follow Controlled Substance storage requirements.

Disposal of Expired Drugs

Disposal of expired drugs is the responsibility of the PI.  Follow instructions from the EHS Hazardous Waste page, for Pharmaceutical Waste and Controlled Substance Waste as applicable.

Noncompliance

Use of expired drugs, diets, or medical materials in live animals, unless specifically allowed in this policy, will be treated as noncompliance, and handled in accordance with the Policy for Resolving Regulatory Noncompliance. Additionally, the Lead Researcher/Principal Investigator may be subject to IACUC sanctions and/or protocol suspension and may be held financially responsible for any fines imposed on the institution by USDA.

References

  1. U.S. Department of Agriculture (USDA). (n.d.). Animal Welfare Regulations (9 CFR Chapter 1, Subchapter A, Part 2, Subpart C, §2.33(b)(1–5)).
  2. U.S. Department of Agriculture (USDA). (2011, March 25). Animal Welfare Policy #3: Veterinary Care.
  3. National Research Council. (2011). Guide for the care and use of laboratory animals (8th ed.). National Academies Press.
  4. Office of Laboratory Animal Welfare (OLAW). (n.d.). Frequently asked questions: PHS Policy on humane care and use of laboratory animals—Use of non-pharmaceutical-grade compounds in animals. National Institutes of Health.
  5. Avertin® (tribromoethanol) references:
    • Lieggi, C. C., et al. (2005). Efficacy and safety of stored and newly prepared tribromoethanol in ICR mice. Contemporary Topics in Laboratory Animal Science, 44(1), 17–22.
    • Lieggi, C. C., et al. (2005). An evaluation of preparation methods and storage conditions of tribromoethanol. Contemporary Topics in Laboratory Animal Science, 44(1), 11–16.
    • Meyer, R. E., & Fish, R. E. (2005). A review of tribromoethanol anesthesia for production of genetically engineered mice and rats. Lab Animal (NY), 34(10), 47–52.
    • Zeller, W., et al. (1998). Adverse effects of tribromoethanol as used in the production of transgenic mice. Laboratory Animals, 32(4), 407–413.
    • Koizumi, T., Maeda, H., & Hioki, K. (2002). Sleep-time variation for ethanol and the hypnotic drugs tribromoethanol, urethane, pentobarbital, and propofol within outbred ICR mice. Experimental Animals, 51(2), 119–124.
    • Cold Spring Harbor Laboratory. (n.d.). Tribromoethanol (Avertin). Cold Spring Harbor Protocols.
  6. DenHerder, J. M., et al. (2017). Effects of time and storage conditions on the chemical and microbiologic stability of diluted buprenorphine for injection. Journal of the American Association for Laboratory Animal Science, 56(4), 457–461.
  7. Xu, J., et al. (2021). Strength and sterility of stock and diluted carprofen over time. Journal of the American Association for Laboratory Animal Science, 60(4), 470–474.
  8. Dodelet-Devillers, A., et al. (2016). Assessment of stability of ketamine-xylazine preparations with or without acepromazine using high-performance liquid chromatography–mass spectrometry. Canadian Journal of Veterinary Research, 80, 86–89.