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Residual Moisture Content in Lyophilized Drugs

Residual moisture is a critical parameter for stability of the API in lyophilized drug products, and well-known quality attribute (CQA) of lyophilized drug. The amount of residual moisture content is related to the stability of the formulation over the shelflife of the product. The traditional methods that are used to measure residual moisture are generally destructive and low throughput. As a result, there is a significant interest in non-destructive testing of residual moisture as part of development, scale up and manufacturing.


To determine the residual moisture content then is critical and a number of methods have been used including:

  1. Karl Fischer Titration

  2. Thermogravimetric Analysis

  3. Near Infra-red Spectroscopy

  4. Headspace Moisture Analysis


1. The KF method is most widely used and is generally considered to measure the total water within the freeze-dried vial, assuming that the sample is wholly soluble in the KF medium. The KF method is time-consuming, requires operator expertise and careful sample handling to avoid contamination by environmental moisture, and is destructive (meaning that the sample is destroyed during analysis).


2. Thermogravimetric analysis is widely used to determine the weight loss associated with the heating event to drive off the residual moisture. This method measures not only the water but also any other volatiles that are produced as a result of heating. Therefore, the composition of the lyophilized material must be well understood for this method to be used to accurately measure the residual water content. This method also suffers from potential artifacts due to environmental moisture and is also a destructive method.


3. Near Infrared Spectroscopy is a method that can be used for nondestructive residual moisture analysis. The basic principle involves shining a beam through the bottom of the vial up into the lyophilized cake. A model can be developed using Karl Fischer as a basis. Once the model is developed, lyophilized vials may be scanned by NIR and the moisture model applied to determine the residual moisture based on peak amplitude. NIR has potential application as a process analytical technology (PAT) as this method has the potential to be set up in-line on a commercial scale, thereby making 100% in-line residual moisture content analysis across a batch very much a possibility.


4. The frequency modulated spectroscopy solution, in which moisture is measured by laser absorption in the gas headspace of a vial through the glass container. The laser is tuned to match the internal absorption frequency of water molecules so that the amount of laser light absorbed is correlated to the amount of water in the headspace. The measurement is rapid and non-destructive. Measurements can be made in less than a few seconds and the sample remains intact.

The applications of headspace moisture analysis take advantage of the fact that it is straightforward to analyze the headspace moisture content of all vials in a statistically relevant sample set. In fact, it is even possible to perform 100% headspace moisture analysis of a commercial batch of freeze-dried pharmaceutical product using automated headspace moisture inspection machines.


The industry is moving quickly towards measurement of residual moisture by non-destructive means. The benefits of moving to nondestructive testing include:


  • Efficiency – sample analysis is done in seconds; the sample population for testing can be significantly larger and 100% inspection is feasible

  • Non-destructive – the product may be used for other testing and possibly patient supply

  • Possibility of in-line or at scale

  • Application to map entire development lyophilizer

  • Applicable to commercial manufacturing with no vial loss

  • Value to batch release in the event of a deviation in the cycle

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