According to the FDA, ensuring data integrity is an important component of industry’s responsibility to ensure the safety, efficacy, and quality of drugs. ARL Bio Pharma has GMP software that controls our laboratory’s instruments and generated data. This software complies with 21CFR Part 11 as it relates to electronic records. All data is locked down and backed up so ARL can retrieve information in its native format with tracked changes and modifications. Our validated software meets FDA’s expectation for GMP Compliance to provide our clients with data integrity critical to the GMP data lifecycles.

To learn more about Data Integrity and Compliance with Drug cGMP, click here. 

How are endotoxin limits calculated? How do my endotoxin test results relate to those limits?

Endotoxin limits for drug products specify the amount of endotoxin that can safely be present. Limits are based on human tolerance and generally specified in the USP monograph or if no monograph exists, calculated via USP <85> Bacterial Endotoxin Test. The testing limit calculation considers:

  1. Route of administration
  2. Dose of the product per kilogram of body weight (which should include the maximum bolus dose and the minimum patient weight)
  3. Duration (time) of administration

Once a limit is determined, USP <85> allows a testing facility flexibility to determine the appropriate test dilution to reduce drug sample properties that interfere with the test. Using the endotoxin limit and the assay’s sensitivity, a maximum valid dilution (MVD) is calculated. The MVD provides the maximum dilution possible to obtain a result and maintain the sensitivity necessary to determine a pass or fail. For example, if a sample is submitted with an endotoxin test limit of NMT 14 EU/mL and the MVD is determined to be 1400, then the product concentration can be diluted no further than 1:1400 and still generate a result with the appropriate sensitivity.

The test result is calculated by the assay software using the test dilution and assay sensitivity. In a case where no endotoxins are present, for example, a 1:1000 dilution was used and no presence of endotoxin was detected, a result of <10 EU/mL is reported. A result which includes a “less than” value does not indicate that the actual endotoxin value is near the number reported, only that the dilution factor used allowed for that level of sensitivity in results calculation. This means that results of <10 EU/mL and <4 EU/mL both show no presence of endotoxin at the dilution with which they were tested; and neither result offers any indication of the exact level of endotoxin present below the values reported.

If you would like to always receive the same result when endotoxins are not detected, a validation of the endotoxin test method is required.

Contact ARL at info@arlok.com or 800-393-1595 to schedule your endotoxin test method validation.

Hand sanitizers are regulated as over the counter (non-prescription) drugs by the U.S. Food and Drug Administration. The FDA has issued warnings to consumers about hand sanitizer products containing methanol. According to the CDC, Alcohol-based hand sanitizers should only contain ethanol or isopropanol, but some products have been found to contain methanol. Ethanol is a valid active ingredient, but methanol, even in small amounts, can lead to patient adverse events.

Hand Sanitizer Testing Requirements

503A pharmacies and 503B outsourcing facilities that procure hand sanitizer API (ethanol or isopropanol) must perform testing before compounding regardless of what is on the API vendor’s certificate of analysis. This requirement falls under the enforcement discretion described in the FDA’s guidance updated on August 7, 2020, to ensure compounded hand sanitizer is free of contamination.

According to the FDA, the new guidance clarifies that companies test each lot of the active ingredient (ethanol or isopropyl alcohol (IPA)) for methanol. FDA recommends using the test methods described in the USP monograph for alcohol (ethanol) and conducting the testing in a laboratory that has been previously inspected by FDA and is compliant with current good manufacturing practice (CGMP).

ARL Bio Pharma is inspected by the FDA, CGMP compliant, and provides testing on ethanol and IPA according to USP monographs.

Hand Sanitizer Formulations

The United States Pharmacopeia created a document in April 2020, outlining recommendations from its Compounding Expert Committee. This document includes three formulations for compounding hand sanitizer and appropriate ingredient substitutions based on shortage issues.

The starting ingredients for Formulations 2 and 3 provide alternatives to WHO’s formula using isopropyl alcohol 99.8%, due to shortage concerns.

Hand sanitizer should be prepared under conditions routinely used to compound similar nonsterile drugs (USP General Chapter <795>).

USP Resources:

For more information on testing ethanol or isopropanol API, contact info@arlok.com or 800-393-1595

Dr. Nicole Vu, Scientific Director, ARL

Pharmaceutical ingredients are controlled by cGMP regulations. Users of pharmaceutical materials should be informed regarding when the material can be re-tested for continued use.

ICH Q1A(R2)/FDA Definition:

Expiration date: The date placed on the container/labels of an API designating the time during which the API is expected to remain within established shelf-life specifications if stored under defined conditions and after which it should not be used.

Retest date: The date when a material should be reexamined to ensure that it is still suitable for use.

Retest period: The period of time during which the drug substance is expected to remain within its specification and, therefore, can be used in the manufacture of a given drug product, provided that the drug substance has been stored under the defined conditions. After this period, a batch of drug substance destined for use in the manufacture of a drug product should be retested for compliance with the specification and then used immediately. A batch of drug substance can be retested multiple times and a different portion of the batch used after each retest, as long as it continues to comply with the specification. For biotechnological/biological substances known to be labile, it is more appropriate to establish a shelf-life than a retest period. The same may be true for certain antibiotics.

Additional information (ICH Q7A: Good Manufacturing Practice Guide For Active Pharmaceutical Ingredients)

  1. An API expiry or retest date should be based on an evaluation of data derived from stability studies. Common practice is to use a retest date, not an expiration date (Section 11.6). Some very labile materials such as antibiotics and biotech materials may be required to be labeled with an expiration date. The retest date or the expiration date is listed either on the container or on the COA. 
  2. If the API material has an assigned expiration date: User of the API material cannot extend the manufacturer’s expiration date by means of re-testing.
  3. If the API is labeled with a retest date: The API material can be re-tested, and if retest results are in compliance with the material specification for critical attributes, such as content and purity (chemical & microbiological), the batch may be used immediately. Test results must be obtained with pharmacopeial procedures or stability indicating methods, using primary reference standard. Use immediately is not defined by the FDA. Common practice in the industry is to use within 30 days of the retest date.
  4. Under current rule, a new date for retesting beyond 30 days of immediate use should be based on current retest results and supporting stability data. Successive retest periods may not be longer than the original retest period assigned by the manufacturer of the API. 
  5. Section 19.8: Expiry and retest dating as defined in Section 11.6 applies to existing APIs used in clinical trials (i.e., established APIs). New APIs in the early stage of clinical trials (non-established APIs) are not required to have an expiration date nor a retest period.

Particulate matter contamination in injectable drug products, especially in large numbers, can cause harm to patients. Common particulate sources include: solvent impurities, drug precipitates, dust, glass, rubber, environmental contaminants, fibers and other insoluble materials. 

The size of particulate matter is an important factor when considering the potential risk to patients. Particles as small as 2 μm in diameter have been associated with microthrombi formation in patients. Dr. Michael Akers with the Food and Drug Administration notes that the smallest capillary blood vessels are considered to have a diameter of approximately 7 μm. Therefore, all particles having a size equal to or greater than 7 μm can conceivably become entrapped in and obstruct capillaries, increasing the potential for adverse effects. Simple visual inspection, which is required for compounded injections, may be adequate for large particles, but is inadequate for smaller particles. The lower limit of visibility of the naked eye is approximately 40 μm. Specialized testing methods are therefore necessary to adequately assess the total particulate burden of injections. 

The United States Pharmacopeia establishes procedures and standards to ensure the quality of intravenous injections, including particulate counts. Compounding pharmacists can reduce the incidence of adverse events in patients by assuring the quality of their preparations through filtration of intravenous preparations and analytical testing procedures.  

USP Chapter <788> Particulate Matter in Injections recommends two test procedures for determining it. These two tests are (1) light obscuration particle count test and (2) microscopic particle count test. The two procedures test for particulate matter in preparations and count particles that are equal to or greater than 10 μm and 25 μm. 

The USP guidelines limit particulate matter introduced into preparations and into the patient. USP testing methodologies can assure compounding pharmacists of the quality of their preparations and the quantification of subvisible particles. Pharmacists can use these tools to improve patient safety. 

To learn more about Particulate Matter in Injections, click here to read more. 

Visit ARL’s website to learn more about particulate matter testing services. To request a quote email: info@arlok.com. 

Today, FDA issued updated guidances to provide additional clarification on testing of alcohol used in hand sanitizers manufactured under FDA’s temporary policies to help ensure that harmful levels of methanol are not present in these products. This will help ensure widespread access to alcohol-based hand sanitizers that are free of contamination.

FDA has updated their guidances to provide clarification that companies test each lot of the active ingredient (ethanol or isopropyl alcohol (IPA)) for methanol if the ethanol or IPA is obtained from another source. FDA recommends using the test methods described in the USP monograph for alcohol (ethanol) and conducting the testing in a laboratory that has been previously inspected by FDA and is compliant with current good manufacturing practice (CGMP).

Additionally, any alcohol (ethanol) or IPA found to contain more than 630 ppm methanol does not fall within the policies described in the temporary guidances and may be considered evidence of substitution and/or contamination. Alcohol-based hand sanitizers that are contaminated with methanol are subject to adulteration charges under the FD&C Act. The alcohol (ethanol) or IPA should be destroyed following guidelines for hazardous waste and the manufacturer or compounder should contact FDA regarding the test results and the alcohol’s source.

The temporary guidances have also been updated to provide adverse event reporting guidelines for state-licensed pharmacies and outsourcing facilities.

The agency also included an additional denaturant formula in the temporary guidances. Denaturing alcohol in hand sanitizers is critical to deter children from unintentional ingestion. Consumer and health care professional safety is a top priority for FDA, and an important part of FDA’s mission is to protect the public from harm, especially as we seek to help increase hand sanitizer supply.

Contact ARL at 800-393-1595 for hand sanitizer testing.

FDA expands hand sanitizer warnings to include 1-propanol contamination 

Evelyn Orona, ARL Bio Pharma Associate Microbiology Laboratory Supervisor

Andrew Taylor, ARL Bio Pharma Microbiology Laboratory Supervisor

ARL Bio Pharma utilizes the kinetic turbidimetric method described in USP <85> Bacterial Endotoxins Test to provide results. An endotoxin test detects toxins that are released from the cell wall of disrupted gram-negative bacteria. This is a reaction over time test that quantitates the amount of endotoxin in a sample compared to a standard curve. This article discusses USP <85> challenges and why method validation is important to provide consistent results.

The most common challenge encountered while performing endotoxin testing is that many drug products cause the assay reaction to slow down (inhibition) or speed up (enhancement) during the testing process, which does not allow for an accurate result to be obtained. Those properties which cause the inhibition or enhancement must be overcome to obtain a reliable and repeatable test result. Microbiologists performing testing must determine the degree in which inhibition or enhancement is occurring. This is done by adding known amounts of endotoxin to test samples and comparing the results to those known values. If there is no inhibition or enhancement, results from the test samples spiked with a known amount of endotoxin will be the same as the expected results. Inhibition would artificially lower results for the test sample, and enhancement would increase results for the test sample, relative to the expected spike concentration. 

Once inhibition or enhancement is understood, microbiologists can develop a plan of action to overcome those sample properties.

For sample testing, overcoming inhibition or enhancement typically begins with dilution of the sample in endotoxin-free water. A test sample cannot be diluted so much that the test results obtained become an inaccurate representation of any endotoxin that is present. A sample’s Maximum Valid Dilution calculated from the sample’s endotoxin limit (provided by the client or calculated using the route of administration, the maximum dose per hour, and the average patient weight) allows microbiologists to design endotoxin testing using the appropriate sample dilution. Additional reagents may also be added to overcome inhibition or enhancement if an appropriate method cannot be obtained through dilution alone.

Endotoxin validation testing provides a more robust assessment of the test method. Briefly, inhibition and enhancement testing are performed using multiple dilutions/diluents simultaneously. The results are then assessed to see which method generates the results closest to the ideal spike recovery percentage and sample pH. Once the appropriate sample preparation steps are determined, the sample is tested in triplicate using the chosen method. Should this method provide accurate and precise test results, it is considered validated and retained in ARL’s method suitability database for future submissions of the product. This ensures this method will be used for every submission of a particular formulation.

Once an appropriate method is determined, the lowest value on the standard curve is used to calculate the concentration of endotoxin in a sample, should any be present. For samples showing the presence of endotoxin, the software used by ARL (EndoScan-V by Charles River) will calculate a numerical value that will be reported. For samples showing no presence of endotoxin, the software generates a “less than” numerical value. This “less than” numerical value is calculated by multiplying the dilution factor used by the lowest point on the standard curve (for samples using mL units) or by dividing the lowest point on the standard curve by the test concentration (for samples in all other units). Where no presence of endotoxin is detected, the software cannot calculate an exact numerical value, thus a “less than” value is reported.

When no validated method is on file, microbiologists have the discretion to choose a sample dilution to use for each sample test. This creates the potential for different dilutions to be used between sample submissions, which will lead to different results being reported. While this complies with USP <85> guidelines, many clients choose to perform Endotoxin Validation on formulations submitted for endotoxin testing to have consistent testing methods and results.

For more information on USP <85> Endotoxin Testing and Method Validation, contact 800-393-1595 or info@arlok.com.

Resources:

Contamination of pharmaceuticals with microorganisms may lead to deleterious effects on therapeutic properties of the drug, and may potentially cause injuries to intended recipients. Cases of contaminated nonsterile products have been reported in increasing numbers, and often associated with the presence of objectionable microorganisms. Methods for detection of these organisms are described in three major Pharmacopeias.

USP <61> Microbiological Examination of Non-Sterile products: Microbial Enumeration provides tests used to determine bioburden present in raw materials, during production, and in the finished product. This microbiological test determines the total population of aerobic bacteria and yeast and molds that might be present in pharmaceutical ingredients and finished products.

USP <62>  Microbiological Examination of Non-Sterile products: Tests for Specified Organisms
provides tests used to check for the presence of objectionable microorganisms that could cause the patient harm based on variability in the route of administration. This microbiological test verifies the absence of objectionable microorganisms in drug products and raw materials based on route of administration.

USP <1111> Microbiological Examination of Non-Sterile products: Acceptance criteria for Pharmaceutical Preparations and Substances for Pharmaceutical Use lists acceptance criteria for microbiological quality of nonsterile dosage forms including: route of administration, total aerobic microbial count, total yeast and mold count, and absence of specified microorganisms.

The microbial limit for nonsterile products must be within an acceptable range that does not pose health hazards to intended patient groups or diminish product stability. 

To learn more about Microbial Limits for Nonsterile Pharmaceuticals, visit our article links below. 

“Microbial Limit Tests for Nonsterile Pharmaceuticals, Part 1” – This article provides a valuable introduction on microbial limits for nonsterile products and includes an overview of USP <61> and USP <1111>.

“Microbial Limit Tests for Nonsterile Pharmaceuticals, Part 2” – This article provides an overview of USP <62>, stability study considerations for nonsterile products, and water activity of nonsterile products relationship to microbiological quality.

Visit ARL’s website to learn more about the testing services available for nonsterile products. To request a quote email:  info@arlok.com.

Nora Talley, ARL Bio Pharma Analytical Associate Supervisor

Jessica Munson, M.S., ARL Bio Pharma Analytical Supervisor

This article outlines the importance of sample amounts submitted for potency testing and the impact it can have on potency results. Formulations require different sample amounts to allow for an accurate result representative of the compounded preparation. The table below can also be found in ARL’s Sample Submission Guide.

Raw Powders50 mg
Triturates/Granulations/Blend Powders1 g
Solid Dose Forms (Tablets/Capsules/Pellets/Troches/Suppositories)5 units
Suspensions10 mL
Solutions2 mL
Semi-Solid Dose Forms (Creams/Lotions/Gels)3 g

Powders

Raw powders require 50 mg to meet USP balance requirements and to allow for loss due to powder sticking to the container. Triturates, granulations, and blend powders differ from raw powders as they are a dilution of the active pharmaceutical ingredient (API). One (1) gram is requested for these sample types to allow for adequate API sampling and to reduce the effects of content uniformity.

Solid Dose Forms

Solid dosage forms such as capsules, suppositories, troches, pellets, tablets, and lozenges can be tested as single units for content uniformity assessment or by combining multiple units for a potency test. Combining multiple units collected throughout the batch is the best way to obtain an accurate potency result, which is representative of the overall batch, for solid dosage forms.

Suspensions

A minimum of 10 mL is requested for suspensions so proper mixing can occur. When the suspension is not mixed sufficiently to suspend the API, a test sample that truly represents the potency of the compounded preparation may not be obtained. This can cause both high- and low-test results.

Note: the container used for the sample (if not sent in the final package) should not be overly large, ideally not more than two times (2x) the size of the sample, as this can also cause sampling problems.

Solutions

Solutions are by definition uniform and are simple to sample. 2 mL of solutions are requested so 1 mL can be collected accurately for the potency test. Exceptions can be made for highly priced drugs.

Semi-Solid Dose Forms

In semi-solid dosage forms such as creams, lotion, gels and ointments, it is essential that the ingredients of the semi solid compound are evenly distributed throughout the sample. To evenly distribute the API, three (3) grams need to be provided so that when necessary the sample can be mixed evenly with in its container with a spatula.

The amounts ARL requests are not minimum amounts required for a potency test. 

Instead, these are amounts that ARL recommends to provide a potency test result that truly represents the sample.

For more information on potency test sample amounts, contact ARL at 800-393-1595 or info@arlok.com.

Growth Promotion demonstrates that media used in environmental monitoring programs, media fills, or personnel qualification are capable of supporting microorganism growth. 

Benefits of growth promotion testing: