Within the pharmaceutical, health care, and numerous other industries, ensuring a product meets the quality standards outlined by the FDA is paramount to ensuring safety and efficacy. In the case of injectable products, the United States Pharmacopeia (USP) outlines necessary product quality tests required for compliance in USP <1>: Injections and Implanted Drug Products (Parenterals)-Product Quality Tests.
One crucial aspect of quality testing that particularly demands attention is the presence of particulate matter within injectable solution products. This concern is specifically addressed in the chapter USP <788>: Particulate Matter in Injections, which meticulously details the procedures and criteria required to ensure that the level of particulate matter in an injectable product meets the stringent standards set by the FDA.
So, what samples are subject to the USP <788> criteria? What analyses are used to test for compliance with USP <788> requirements? And how can these procedures and criteria be related to other cleanliness tests?
Understanding the Purpose
USP <788> serves the vital purpose of determining the level of particulate matter in injections by utilizing two primary procedures: a light obscuration particle count test (Method I) and/or a microscopic particle count test (Method II). The chapter applies to solutions meant for injection, particularly those administered intramuscularly or subcutaneously. However, exceptions to the conformance criteria might be considered based on specific circumstances such as if the product is for veterinary use or is a radiopharmaceutical preparation.
The presence of foreign particulate matter is vital to enumerate due to the potential consequences that stem from high concentrations of particulate or large particles within a given product. Large particles, for instance, can potentially clog narrow blood vessels when injected into the blood stream. Additionally, it is important to determine if a product precipitates over time or in certain conditions as this could affect a drug’s efficacy, stability, and potentially, the overall safety. It would also be valuable to enumerate the extent to which this occurs to determine a course of action to limit precipitation in the product for future reference.
Particulate matter, within the scope of this context, encompasses extraneous mobile undissolved particles that are unintentionally found in solutions, with the exception of gas bubbles. These particles can be categorized into three distinct classifications depending on their origin: extrinsic, intrinsic, or inherent. Extrinsic particulate matter originates externally, often introduced during the manufacturing or handling processes. Intrinsic particulate matter arises from within the solution, stemming from factors such as impurities or interactions between solution components. Inherent particulate matter is inherently present in the solution due to its composition or nature.
This classification system helps to discern the source and nature of particulate matter, facilitating a comprehensive understanding of its presence in various solutions and identifying where in the manufacturing process particulate could be entering or forming in the product.
Figure #1 (Below):
This image shows an example of particulate matter that has been captured on a filter membrane for analysis by the microscopic particle count test (USP <788> Method II)
Deciding on Samples to Analyze
Determining which samples to subject to analysis is crucial. When analyzing a smaller group of units to infer findings for a bulk lot, a robust sampling plan is imperative to ensure the relative accuracy and reliability of extrapolated results. For samples that have a relatively small volume (<25 mL), it is paramount that multiple units are combined for analysis in order to get a representative particle concentration value that can be extrapolated to a bulk lot.
In addition to considering the volume and number of units of product to sample, it is just as important to consider whether the product is applicable to USP <788> or if might be more applicable to one of the orthogonal USP chapters. For example, products considered to be ophthalmic solutions or therapeutic protein injections have their own respective USP chapters outlining procedure and criteria for foreign particulate matter analysis (USP <789>: Particulate Matter in Ophthalmic Solutions and USP <787>: Subvisible Particulate Matter in Therapeutic Protein Injections respectively).
Though the chapters share some similarities in how the samples are prepared and analyzed, subtle differences in the product and its application could mean slightly different procedural steps or conformance requirements for particulate matter within a sample.
Analytical Procedure
USP <788> provides two accepted procedures for analysis. The light obscuration particle count test, or Method I, involves passing a relatively dilute liquid sample through a laser and detector. Based on the amount of light blockage (obscuration) and application of a calibration curve, a circular equivalent particle size is assigned to a particle.
On the other hand, the microscopic particle count test, or Method II, entails filtering the sample onto a membrane filter and manually counting the particulate matter using a specialized USP microscope setup. Particle sizing in Method II is facilitated by a USP-verified graticule and micrometer for particles larger than or equal to 10 and 25 microns.
Generally, Method I is the preferred technique, but Method II can be a useful application in instances where the sample may not be suitable for Method I, such as high viscosity samples and samples with reduced clarity or the presence of emulsion droplets. Though analysis by Method I and Method II involve different techniques and principals, both procedures include considerations in order to properly prepare samples for analysis and limit external contamination before analysis.
Figure #2 (Below): Particle Sizing Systems AccuSizer A7000 SIS
AccuSizer A7000 SIS particle counter used for USP <788> Method I analyses. This instrument uses the light obscuration principle to count and size particulate suspended in a carrier fluid and pulled through a sensor using a syringe injection system.
Figure #3 (Below): Nikon Eclipse Ci POL Microscope
Nikon Eclipse Ci POL Microscope used for USP <788> Method II analyses. This microscope is equipped with a USP verified graticule to size particulate in relation to 10- and 25-microns reference circles and a micrometer.
Pros and Cons of Method I
Though light obscuration (Method I) is the preferred technique per USP <788>, there are pros and cons for both light obscuration and the microscopic particle count test (Method II) that may skew favor for the use of one technique over the other.
Light obscuration is preferred when possible as it not only provides more in-depth statistics on particle size and concentration, but is also typically more expedient than the microscopic particle count test. The instrument works to count and size particulate as they pass through a laser one-by-one based on a calibration constructed using spherical reference materials of various sizes. This allows for reproduceable, automated, and unbiased analysis and results, but it does carry some of its own drawbacks.
For instance, as the instrument relies on optical properties of the suspended sample as it passes through the laser, structures such as fibers, emulsion droplets, and bubbles can all be interpreted as particulate by the instrument. This can be cause for concern for emulsions, particularly viscous, or bubbly products. Other factors that may cause concern for analysis of certain products are its relatively limited compatibility with certain solvents and carriers as well as its viscosity as samples that are too viscous can have trouble traveling through the sensor via the syringe injection system.
Pros and Cons of Method II
Though the microscopic particle count test is not the preferred analytical technique for USP <788>, it does provide more relative flexibility for which samples can be analyzed. For the most part, samples analyzed by light obscuration would likely also be compatible with the technique. For samples containing emulsion structures, bubbles, or that are particularly viscous, as long as the product can be successfully filtered, the unwanted bubbles and emulsions can pass through the pores of the filter, leaving behind just the foreign particulate matter. So, these unwanted structures are not counted in the analysis as with light obscuration.
Additionally, the technique is able to provide more information on morphology and general particle characteristics as particulate can be visually observed under the microscope. This can be valuable for products containing elongated particulate or multiple populations of distinct particulate as they can be visually discerned and sized respectively. Also, if unknown particles of a distinct morphology are observed that weren’t originally expected within the sample, the filter can be retained and analyzed to try to identify the unknown particulate by techniques such as Raman spectroscopy or energy dispersive x-ray spectroscopy (EDS/EDX). This can be particularly beneficial to determine if particulate is entering the system and contaminating the sample in the manufacturing process of the product.
The main disadvantages of this technique revolve around the inclusion of human error in the sizing and counting of particulate and that the technique is inherently more time-consuming. As particles are sized based on comparisons to the USP verified reference circles or micrometer within the graticule, there is more room for human error in the sizing of particulate than with the light obscuration instrument.
Figure #4 (Below):
This image shows an example of the USP verified graticule containing 10- and 25-micron reference circles as well as a micrometer used to count and size particulate matter on the surface of a filter membrane.
While Method I is generally preferred due to its efficiency, Method II comes into play when Method I isn’t feasible—for instance, when the sample exceeds the coincidence/concentration limit of the light obscuration instrument, or if it is too viscous or incompatible with the system. If Method I fails to meet specifications, Method II can be employed to validate the results obtained.
Expanded Applications of USP <788>
Interestingly, the implications of USP <788> can be extended beyond merely assessing injectable products. Its guidance has also been applied to evaluate the cleanliness of medical devices and solutions not necessarily categorized as injectables due to a lack of other specific cleanliness guidelines for such samples. Although claiming full compliance with the chapter might not be possible in these scenarios, it still offers a foundational benchmark for assessing general cleanliness and provides valuable information regarding particle size distribution and count.
Furthermore, analogous chapters such as USP <787> and USP <789> address subvisible particulate matter in therapeutic protein injections and particulate matter in ophthalmic solutions, respectively, offering slightly different procedures and acceptance criteria while aligning with the overarching goal of ensuring product safety and quality.
In essence, USP <788> stands as a meticulous guide, ensuring that injections meet rigorous standards regarding particulate matter. Its methodologies, criteria, and applications not only help to guarantee the safety of injectables but also serve as a benchmark for cleanliness evaluation across various pharmaceutical and general domains, fortifying the commitment to delivering safe and effective products to people worldwide.
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