Qualification, Validation, and Verification 4

"If clearly visible growth of microorganisms is obtained after the incubation, visually comparable to that in the control vessel without product, either the product possesses no antimicrobial activity under the conditions of the test or such activity has been satisfactorily eliminated. The test for sterility may then be carried out without further modification."
It may be advantageous, and more consistent, for the text in Chapter ‹71› to be changed to "Suitability of the Test Method," if not to "Verification of the Test Method." The latter change also may be appropriate for Chapters ‹61› and ‹62›, given that what is being assessed is the verification that the actual test conditions relative to those established during the validation permits the proper functioning of the method. Given the harmonized status of these three chapters, such changes, although possible, would certainly take longer to become official.
The same cautions provided at the end of the section on validation are applicable here. If a method in use previously was derived from a pharmacopeial method but used for a purpose other than satisfying monograph requirements, it is not necessary to adopt a revised method in the pharmacopeia when it becomes official. It is therefore not necessary to reverify the suitability of your test article to the revised method. Likewise, the use of a nonpharmacopeial method for purposes other than satisfying a monograph requirement when a pharmacopeial method exists of potential relevance does not necessitate reverification.
General requirements for validationThere are numerous documents that describe the general approach to a validation process. They describe several characteristics (data elements in Chapter ‹1225›) that may be examined during validation, with specific sets selected based upon the nature of the test method. A brief description of these characteristics is provided herein using the characteristics as outlined in the IC Harmonization Harmonized Tripartite Guideline, Validation of Analytical Procedures: Text and Methodology.
Accuracy is a determination of how close the measured value is (in the case of an analytical method) to the true value. As such, one might define accuracy of method as equal to true value plus error. Error may contain both the systematic error (bias) and imprecision of measurement. With the potential error possible, it is important to include a means of reflecting the "true value" as closely as possible. For many compendial tests, this involves the use of a reference standard. Because a method is expected to be useful over a range of true values, the accuracy should be assessed over the expected range of values to which the method is to be applied. As stated previously, the validation should also state the conditions under which the accuracy was determined. Because it is not possible to determine all possible sets of conditions for which a compendial assay might be applicable, accuracy may need to be verified before use of a validated method. The concept of accuracy is more problematic for microbiological assays.
The precision of a method determined during validation should be representative of the repeatability (reproducibility) of the method. As was the case for the determination of accuracy, it should be determined over the expected range of articles to be measured, and the conditions used during the validation should be clearly stated. As for accuracy, the use of reference standards is common because the goal of the assessment of precision is to determe method repeatability without introducing unknown variance as a result of different test articles or test articles drawn from a heterogeneous source. The latter point also complicates the validation of microbiological assays.
Specificity refers to the ratio of false positives to false negatives. A highly specific method would have a very low ratio, given that it should be able to detect the article of interest present in very low quantities in the presence of much higher quantities of similar but not identical articles. As stated previously, specificity should be determined over the expected range of usage for the method, and conditions used during the validation should be clearly stated.
Linearity, in essence, refers to the existence of a direct relationship between the quantity of article contained in the sample being analyzed and the measured value resulting from the analysis. It is not the purpose of this article to delve into statistical intricacies pertaining to data transformation, the use of linear or nonlinear regression techniques, residual analysis, and so forth. Currently, it is sufficient that an assay purporting to be quantitative in nature must have a demonstrable quantitative relationship between the quantity of material of interest contained in the sample and the measured response.

Qualification, Validation, and Verification 3

By: David A. Porter
Therefore, given the following from FDA's Guide to Inspections of Pharmaceutical Quality Control Laboratories: "Methods appearing in the USP are considered validated and they are considered validated if part of an approved ANDA" (8), the use of Method 3 would be valid if the conditions stated are met in testing the material of interest. The same FDA document states "For compendial methods, firms must demonstrate that the method works under the actual conditions of use," which, for the sake of this article, will be considered verification. Chapter ‹1047› provides several other procedures, all also validated, that could be considered given test material that does not satisfy the conditions for Method 3.
Remember the purpose. It is important to bear in mind the purpose of the method to be validated. If the method is intended to serve as an alternative to a pharmacopeial method, then one must establish its equivalence to the pharmacopeial method in terms of the end result. Remember that the purpose of a method in the pharmacopeia is to determine whether the pharmacopeial article (for which a monograph exists in the pharmacopeia) satisfies the requirements in the monograph. If instead the purpose behind the use of a pharmacopeial method is for a purpose other than demonstrating that the article complies with monograph requirements (for example, imagine that total organic carbon is to be determined using Chapter ‹643› "Total Organic Carbon"), it is not necessary to perform the validation relative to the pharmacopeial results. This means that the validation should be conducted relative to the specific purpose for which it is intended. Also implicit in this is the use of a nonpharmacopeial method to determine something for which a pharmacopeial method exists, but again for purposes unrelated to satisfying a monograph requirement. In such a case, it is unnecessary to consider validating the method relative to that in the pharmacopeia.
Verification
If the use of the term validation is restricted to mean the demonstration of suitability of a method or process for its intended purpose, and the term verification for the demonstration that the previously validated method is suitable for use given specific experimental conditions that may or may not be appropriate given the conditions present during the validation, the terminological situation may be clarified.

Figure 1

These actual conditions include specific ingredients or products, specific laboratory personnel, equipment, and reagents. There are, however, instances in the literature where this distinction is not maintained. Consider the dictionary definition given previously for validation and its use of verification as a synonym for validation. Further muddying of the waters occurs when phrases such as "system suitability tests" (see Figure 1 and the system-suitability section in Chapter ‹621› "Chromatography"). The phrase also appears in the "Suitability of the Counting Method in the Presence of Product" section of Chapter ‹61› "Microbiological Examination of Nonsterile Products: Microbial Enumeration Tests", the "Suitability of the Test Method" section of Chapter ‹62› "Microbiological Examination of Nonsterile Products: Tests for Specified Microorganisms", and the "Validation Test" section of Chapter ‹71› "Sterility Tests." (1). In all cases, the intention is to ensure that the validated method will work under the specific conditions the analyst plans to use. This means that a chromatographic system can deliver resolution and reproducibility on par with the system used during validation. For the two microbiology test chapters for nonsterile products, one must show that microbial growth in the presence of the article to be tested is not hindered. This is because the method depends on unencumbered microbial growth for it to work. In other words, a condition established in validating the method initially was unhindered microbial growth. The use of "validation test" in Chapter ‹71› is unfortunate because the intention was again to demonstrate that microbial growth is not hindered, as indicated in the following text:

Qualification, Validation, and Verification 2

By: David A. Porter
FDA provides a definition of validation in numerous documents. One such document, Guidance for Industry: Analytical Procedures and Methods Validation Chemistry, Manufacturing, and Controls Documentation says "methods validation is the process of demonstrating that analytical procedures are suitable for their intended use" (5). There also are numerous documents defining validation within the context of processes. From FDA's Guideline on General Principles of Process Validation:
"Validation—Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its predetermined specifications and quality attributes (6)."
The same definition is provided in other FDA documents, such as Guideline on Sterile Drug Products Produced by Aseptic Processing. FDA document Guidance for Industry: Quality Systems Approach to Pharmaceutical Current Good Manufacturing Practice Regulations provides this definition:
"With proper design (see section IV.C.1), and reliable mechanisms to transfer process knowledge from development to commercial production, a manufacturer should be able to validate the manufacturing process. In a quality system, process validation provides initial proof, through commercial batch manufacture, that the design of the process produces the intended product quality (7). "The remainder of the discussion about validation in this article will be restricted to a discussion of method validation.
Does it suit its purpose? The foregoing is clearly not an exhaustive list of the manners in which validation has been defined. It does appear that a recurring theme among the various definitions pertains to demonstrating that the method or process is suitable for its intended use. In this article, consider validation to be the demonstration that a method or process is suitable for its intended purpose. Accepting that, it is imperative that the intended purpose of a method or process is clearly stated at the outset of the validation. An example of the importance of such a statement can be found in Chapter ‹71› "Sterility Tests" (1). It states that "the following procedures are applicable for determining whether a Pharmacopeial article purporting to be sterile complies with the requirements set forth in the individual monograph with respect to the test for sterility." The next paragraph states
"These Pharmacopeial procedures are not by themselves designed to ensure that a batch of product is sterile or has been sterilized. This is accomplished primarily by validation of the sterilization process or of the aseptic processing procedures."
During the years there has been concern that the tests for sterility as provided in Chapter ‹71› are not adequate to prove that a batch of product is sterile. As stated previously, the tests in Chapter ‹71› were intended only to show that a Pharmacopeial article is sterile. Such a demonstration constitutes a necessary but not sufficient condition for sterile pharmacopeial articles. If one were to validate an alternative procedure for that in Chapter ‹71›, it would not be necessary to develop one that is intended to demonstrate sterility of an entire lot of product.
In addition, it is appropriate that the conditions are provided under which the validation was performed. Given that there are essentially countless variations on experimental conditions, product matrix effects, and so forth, a validation cannot reasonably expect to address all such permutations. For example, Method 3 in the section of Chapter ‹1047› "Biotechnology-Derived Articles—Tests", which addresses assays for total protein, indicates in a note:
"[Do not use quartz (silica) spectrophotometer cells: the dye binds to this material. Because different protein species may give different color response intensities, the standard protein and test protein should be the same.] There are relatively few interfering substances, but detergents and ampholytes in the test specimen should be avoided. Highly alkaline specimens may interfere with the acidic reagent (1)."

Qualification, Validation, and Verification 1

By: David A. Porter
The terms qualification, validation, and verification occur numerous times in US Pharmacopeia 29 (1). Qualification is found in Chapters ‹1035› "Biological Indicators for Sterilization," ‹1043› "Ancillary Materials Cell, Gene, and Tissue-Engineered Products," ‹1046› "Cell and Gene Therapy Products," and ‹1119› "Near-Infrared Spectrophotometry," among others. Validation appears in ‹1225› "Validation of Compendial Procedures," ‹1223› "Validation of Alternative Microbiological Methods," ‹1010› "Analytical Data—Interpretation and Treatment," ‹1043› "Ancillary Materials for Cell-, Gene-, and Tissue-Engineered Products," ‹1117› "Microbiological Best Laboratory Practices," ‹1120› "Raman Spectrophotometry," and many others. Verification appears in ‹1010› "Analytical Data—Interpretation and Treatment," ‹1035› "Biological Indicators for Sterilization," ‹1035› "Biological Indicators for Sterilization," ‹85› "Bacterial Endotoxins Test," and others. The terms also are present in documents from the US Food and Drug Administration, the Environmental Protection Agency (EPA), and the International Conference on Harmonization (ICH). Given the numerous definitions for the three terms, this article in part is intended to provide an approach to fostering more consistency in the usage of the terms. A recent issue of the Pharmacopeial Forum (2) had a diagram from a proposed General Chapter ‹1058› "Analytical Instrument Qualification" that was intended to show "...four critical components involved in the generation of reliable and consistent data (quality data)." From most to least critical, the components were quality-control check samples, system suitability tests, analytical methods validation, and analytical instrument qualification. In this article, consider system-suitability tests to be the same as verification. Why this usage should be acceptable will be explained. This article will consider validation and verification in detail. Reference to analytical instrument qualification is made. For further discussion of the top tier, "Quality Control Check Samples," refer to Chapter ‹1058› "Analytical Instrument Qualification" (1).
Qualification
Qualification of analytical instrumentation is essential for accurate and precise measurement of analytical data. If the instrumentation is not qualified, ensuring that the results indicated are trustworthy, all other work based upon the use of that instrumentation is suspect. For the purposes of this article, the assumption will be made that the foundation of validation and verification work to follow is based solidly upon well-qualified instrumentation. Validation
Definitions. Numerous documents provide definitions of validation. A dictionary definition (3) of validation includes "...the process of determining the degree of validity of a measuring device," and for validate: "to make legally valid," with synonyms "verify, substantiate." Clearly, the synonyms do not distinguish between validation and verification, so let us now turn to definitions provided by other sources. USP chapter ‹1225› "Validation of Compendial Procedures" provides the following:
"Validation of an analytical procedure is the process by which it is established, by laboratory studies, that the performance characteristics of the procedure meet the requirements for the intended analytical applications."
From the ICH document Validation of Analytical Procedures: Text and Methodology:
However, it is important to remember that the main objective of validation of an analytical procedure is to demonstrate that the procedure is suitable for its intended purpose

Equipment Hold-Time for Cleaning Validation 3

By: Richard J. Forsyth

Table V: Wet granulation equipment train—clean-hold validation (Acceptable residue limit [ARL] = 100 cfu/swab) An examination of the clean-hold time data supports the more aggressive approach. The data were consistent for both the wet- and dry-granulation equipment. The average bioburden level for the 180 samples taken was 1.1 cfu/swab. There were 128 samples with no detectable bioburden and only nine with a bioburden greater than 10 cfu/swab. Although the majority of samples were taken shortly after cleaning, samples were taken at 1, 2, 5, and 8 mo and at 2 yrs, 5 mo with no discernable increase in bioburden. With a bioburden limit of 100 cfu/swab, clean-hold time is not an issue for cleaned equipment that is dried, covered, and stored appropriately. Table VI: Equipment clean-hold time The dirty-hold-time study needed to answer two questions. Does the soil become harder to clean the longer it sits, and what is the possibility of microbial proliferation on soiled equipment? Soils can be more difficult to clean when they are wet and allowed to dry onto the surface, or when the soil is hygroscopic and transforms into a pasty material or subsequently dries. A high-shear granulator is the only equipment that carries out wet granulation at the conclusion of unit operation. The dirty-hold time for the high-shear granulator (196 h) was lengthy enough to allow any wet material to dry. The controlled humidity of the pilot plant prevented any moisture uptake by residual granulation. All other equipment in the validation studies resulted in a dry granulation at the conclusion of unit operation. Microbial proliferation was not a realistic possibility, which was corroborated by the clean-hold time data. Figure 1. High-shear granulator prepared for cleaning. (FIGURES ARE COURTESY OF THE AUTHOR) Subsequent to the validation studies, the gross cleaning of the equipment, including scraping and vacuuming the equipment was shifted from the equipment-cleaning process to the manufacturing process, which effectively shortened dirty-hold times. Because of environmental considerations for residue disposal, equipment operators scrape and vacuum accumulated residue from equipment surfaces. Operators then wipe equipment surfaces with alcohol to remove as much of the residue as possible to minimize the amount of residue discharge to the municipal sewer system. An example of a typical soiled equipment surface prepared for cleaning is shown in Figures 1 and 2. The steps taken for environmental concerns effectively shorten the dirty-hold time. The alcohol wipe dries within minutes, leaving no wet material to dry and become harder to clean. The dry soiled surfaces do not have sufficient water activity to support microbial proliferation. There is no sufficient residue remaining for hygroscopic residues to be a concern. The dirty-hold time data, which measured cleaning effectiveness out to 9 days, demonstrated a worst-case scenario for the pilot plant facility. The dirty-hold time is not of significant concern for soiled equipment awaiting cleaning. Figure 2. Granulator exit chute prepared for cleaning. Under the operating conditions tested as part of the cleaning validation studies, the clean- and dirty-hold times have little impact on the ongoing operations of the pilot-plant facility. In addition, routine verification of adherence to these parameters adds little value to a firm's ability to produce quality formulations. The risk, therefore, tied to not monitoring hold times should be low for validated cleaning and storage conditions.

Conclusion
If clean- and dirty-equipment hold times are established during validation and maintained under properly defined and controlled conditions, the need to monitor clean- or dirty-hold times is not necessary. Avoiding these steps can result in savings of time and resources as well as potential regulatory exposure.
Richard J. Forsyth is an associate director of worldwide GMP quality with Merck & Co., Inc, WP53C-307, West Point, PA. 19486, tel. 215.652.7462, fax 215.652.7106, richard_forsyth@merck.com
References
1. FDA, Guide to Inspection of Validation of Cleaning Processes, Division of Field Investigations, Office of Regional Operations, Office of Regulatory Affairs (Rockville, MD), July 1993.
2. EU, Annex 15, European Union Guide to Good Manufacturing Practice, Working Party on Control of Medicines and Inspections, European Commission (Brussels, Belgium), July 2001.
3. Health Canada, Cleaning Validation Guidelines (Guide-0028), Drug GMP Inspection Unit (Ottawa, Ontario) May 2000.
4. "Recommendations on Validation Master Plan Installation and Operational Qualification; Non-Sterile Process Validation; and Cleaning Validation," in proceedings of the PIC/S (PIC/S, July 2004).
5. R. J. Forsyth and D. Haynes, "Cleaning Validation in a Pharmaceutical Research Facility," Pharm. Technol. 22 (9), 104–112, 1998.
6. J. A. Morales Sanchez, "Equipment Cleaning Validation Within a Multi-Product Manufacturing Facility," BioPharm Inter. 31 (2), 38– 49, 2006.
7. A. H. Mollah, "Risk-Based Cleaning Validation in Biopharmaceutical API Manufacturing," BioPharm Inter. 30 (11), 54–68, 2005.
8. T. Fugate, "Hold Time Studies: A Lost Parameter for Cleaning Validation," J. Val. Technol. 13 (3), 206–209, 2007.

Equipment Hold-Time for Cleaning Validation 2

By: Richard J. Forsyth
Validation studies

Table II: Wet granulation equipment train—dirty-hold validation (Acceptable residue limit [ARL] = 100 µg/swab)

As part of the cleaning-validation study in a pilot plant (5), soiled equipment was held after processing for an extended period of time before cleaning. The hold times for the three validation trials held ranged from 2 h–217 h or 9 days. Data from the validation study including dirty-hold times are shown in Tables I, II, and III. The results include data from a typical dry-granulation equipment train and a wet-granulation equipment train. The majority of the data (187 of 231 swabs) showed no detectable residue. All results were far below the acceptable residue limit (ARL) of 100 µg/swab. The tests showed that over the time span examined, the dirty-hold time had no discernable impact on the ability of the cleaning process to effectively remove the soil from the manufacturing equipment.

Table III: Equipment dirty-hold time

The clean-hold time validation study was conducted independently. After cleaning, the equipment was wiped or sprayed with alcohol to remove residual water, dried, and covered to prevent any dust or particulate accumulation. The validation study consisted of three trials; one trial extended the clean-hold time. The clean-hold times for the three trials ranged from same-day cleaning to a hold time of 2 yrs and 5 mo. Storage conditions included both the clean-equipment hold area in the pilot plant and a storage room outside the pilot plant but in the same building. Data from the clean-hold time study are shown in Tables IV, V, and VI. The majority of the data (128 of 180 swabs) showed no detectable bioburden and all results were far below the ARL of 100 colony forming units (cfu)/swab. The results include data from a typical dry-granulation equipment train and a wet granulation equipment train. The results demonstrate that bioburden was not present immediately after cleaning and was not a cause for concern during storage of properly cleaned, dried, and covered equipment. Discussion

Table IV: Dry granulation equipment train—clean-hold validation (Acceptable residue limit [ARL] = 100 cfu/swab)

Cleaning-validation studies have established equipment dirty-hold times and clean-hold times for pharmaceutical manufacturing equipment. The ongoing expectation is that equipment cleaning documentation verifies compliance with the validated hold times. A conservative approach uses either the established time for each group of equipment, which makes recordkeeping difficult, or the shortest established extended hold time for all equipment. Using this approach, the dirty-hold time is limited to 7 days and the clean-hold time to several weeks. A more aggressive approach uses the longest hold-time data. This gives a maximum dirty-hold time of 9 days and a clean-hold time of more than 2 yrs.

Equipment Hold-Time for Cleaning Validation 1

By: Richard J. Forsyth

The concepts of "clean-hold time" and "dirty-hold time" have been part of cleaning validation since its inception. Clean hold time is generally considered to be the time between the completion of cleaning and the initiation of the subsequent manufacturing operation. Dirty hold time can begin when the clean equipment is initially soiled, but more often is defined as the time between the end of manufacturing and the beginning of the cleaning process. Intuitively, it makes sense to be concerned about both hold times. Dirty equipment is harder to clean the longer the hold time, and clean equipment has a greater chance of becoming soiled as hold time increases.
Background
In its Guide to Inspection of Validation of Cleaning Processes, the US Food and Drug Administration considers identifying and controlling the length of time between the end of processing and each cleaning step to be critical elements of the cleaning processes (1). FDA also expects pharmaceutical companies to demonstrate that routine cleaning and storage of equipment does not allow for microbial proliferation. The European Union expects companies to provide a validation master plan with clearly defined and documented validation program elements (2). Health Canada looks for companies to describe the interval between the end of production and the beginning of the cleaning procedures as well timeframes and conditions for the storage of clean equipment that do not allow for microbial proliferation (3). Finally, the Pharmaceutical Inspection Convention and Pharmaceutical Inspection Co-operation Scheme (PIC/S) guideline looks for documentation of both dirty- and clean-hold times (4). The general practice among industry is to routinely document and track equipment-hold times to ensure ongoing compliance.
Although regulatory agencies expect manufacturers to document and address hold times, they do not describe a process for establishing hold times. In this validation study, a dirty-hold time was established but ongoing implications were not examined (5). Several articles define both clean- and dirty-hold times and how to establish them but do not mention a strategy to guide the experiments (6, 7). A more recent article, which referred to hold-time studies as "the lost parameter for cleaning validation," explored several issues associated with hold-time studies (8). Issues included storage conditions, test locations, testing methodology, and the length of hold-time studies. The concern with clean-hold times is that clean equipment will not stay clean indefinitely despite using appropriate storage conditions. Holding soiled equipment makes it more difficult to remove pharmaceutical soil and allows biological contamination to proliferate. To address these concerns, the author extended clean-hold time testing for more than 2 yrs and extended dirty-hold time studies for up to 9 days. After identifying clean- and dirty-hold time, ongoing control of the hold times became difficult. Every time a piece of equipment is used, the operator needs to confirm and document that the actual clean-hold time does not exceed the established clean-hold time. And before washing a piece of equipment, the washer needs to confirm and document that the actual dirty-hold time does not exceed the established dirty-hold time.

Table I: Dry granulation equipment train—dirty-hold validation (Acceptable residue limit [ARL] = 100 µg/swab)

This study suggests that if clean- and dirty-hold time issues are addressed during the validation study that the severity of exceeding the established hold times diminishes to a near-acceptable level.

Applying Quality by Design to Sterile Manufacturing Processes 5

By: Warren Charlton

Quality risk-management review

Table I: Sample risk-analysis evaluation.

Sterile-filling equipment and product delivery systems support processes (e.g., CIP/SIP, sterile filtration) and are well defined in industry. Use risk-assessment tools to review automation software, controls, alarms, and to define PAT needs using a QbD approach. Table I includes instructions and a key for severity, occurrence, and detection for use in Table II. Table II includes three examples to demonstrate the use of risk assessment as described below. (This methodology provides a number [e.g., ≥50 fails], which makes a result a clear decision, requiring modifications either to the process or finding a PAT alternative.)

Table II: Sample risk-analysis worksheet.

Isolators on a filling line. Risk assessment in this case can highlight and quantify benefits. Assessment involves product and personnel safety and requires process modifications to be successful.
Manual loading of filled vials into a freeze dryer as compared with automatic loading. It is possible to eliminate the need for using trays if the design includes the capability to transfer the freeze-dried vial after processing in the freeze drier directly to a capper. Automation software used in conjunction with unit operations and equipment. This risk assessment example involves a sterile filler designed for 10% check weighing and compares it with 100% check-weighing capability (6). Risk assessment can help demonstrate the importance of good CQA choices to support management approvals.
Draft guidance questions and concerns
There are some remaining questions despite the thoroughness of the draft guidance. Below are a few key issues.

• What is required for final PQ approval? The final guidance should include clarification on what constitutes validation. This clarification is critical because the common practice of using three batches to verify validation no longer applies (3).
• In the case of a PAT strategy, will the approach to process qualification be different from other process designs? The final guidance needs to include more specifics with regard to what degree of PAT is required to positively impact validation and approvals (3). This clarification is especially important because often, the more PAT involved, the more investment and validation required.
• Industry needs to have a sense of which statistical methods and levels of sampling FDA recommends. Reference to specific documents and testing level(s) may suffice (3).
• The final guidance should discuss the impact of the new guidance on existing products and processes and how to integrate them into the new approach (3).
• The final guidance should discuss potential impact on current and future new drug and abbreviated new drug applications (NDAs and ANDAs) and their site of manufacture. For example, is there an expected date to have the new process validation requirements implemented in applications? For NDAs, additional time may be needed and patents may be affected. For ANDAs, shouldn't companies be required to demonstrate equivalency of control as the innovator process?
• Finally, there is a concern that product development information could become available though freedom of information, thus revealing data that have significant confidential information about the process. How will this be handled? 

In conclusion, the 2008 Draft Guidance for Industry on Process Validation: General Principles and Practices brings the product life-cycle approach to process development. The document provides insight into the "how" with regard to holding discussions on project management, documentation, verification, reviews, and methods (e.g., QbD, DOE, risk assessment) and reviews expectations from development to commercial processes for testing, validation, verification, and release. It is important that all pharmaceutical companies review the draft guidance and understand how it may affect their sterile manufacturing processes. The impact on confidentiality may be industry's biggest concern because process design and associated technical knowledge are what define a company's success.
Warren Charlton is a consultant at WHC Bio Pharma Technical Services, PO Box 20309, Greenville, NC 27858, tel. 252.327.4733, fax 252.756.4733, warren@whcbiopharma.com
References
1. ICH, Q8(R1) Pharmaceutical Development (Geneva, Switzerland, Nov. 10, 2005; Rev. 2008). 2. ICH, Q9 Quality Risk Management (Geneva, Switzerland, Nov. 9. 2005).
3. J. Agalloco et al., "FDA's Guidance for Industry: Process Validation: General Principles and Practices," presented at PDA, Jan. 14, 2009.
4. FDA, Draft Guidance for Industry—Process Validation: General Principles and Practices (Rockville, MD, Nov. 2008).
5. W. Charlton, T. Ingallinera, and D. Shive, "Validation of Clinical Manufacturing," and Validation Chapter, in Validation of Pharmaceutical Process, J. Agalloco and F. Carleton, eds. (Informa Healthcare, New York, 3rd ed., 2008), pp. 542–544.
6. Bausch & Stroebel Risk Analysis System.

Applying Quality by Design to Sterile Manufacturing Processes 4

By: Warren Charlton
Environmental testing and validation
Environmental testing is designed to check facility, process, personnel and environmental cleaning methods to determine: (A) that after being in a state of shutdown that includes relaxed gowning, the facility and equipment can be cleaned, sanitized, and in a state of microbial control appropriate for pharmaceutical operations; and (B) that the facility can maintain a level of environmental control during normal processing operations.
Critical testing required before startup include: high-efficiency particulate air (HEPA) filter testing, room differential verifications (e.g., air flows, alarms), cleanroom smoke studies, and environmental mapping during static and dynamic conditions (i.e., nonoperational and operational conditions). Note that any physical change(s) made in aseptic processing (e.g., facilities, equipment, layout) when adding a new process requires revalidation.
Aseptic processingAseptic processing includes sterile manufacturing, aseptic filling, lyophilization, stoppering, sealing, and spray-drying. Examples of facility and processing-type CQAs that enhance overall process control from outside influence are outlined below.

• Aseptic operations should be protected from potential leaks (e.g., roof leaks and process leaks, including condensation from heat, ventilation, and air conditioning [HVAC] systems). Some processing equipment (e.g., restricted access barrier systems [RABS]) may also contain HVAC or refrigeration that create moisture issues.
• Critical operations should be visible without environmental impact. The objective is to keep personnel out of the critical processing areas while still being able to view operations for documentation and training.
• Gowning areas, including barriers such as the step-over for applying booties, are typically dirty due to employee activity. Consider airflow turns higher than normally recommended (at least 80 air turns is preferred).
• Cleanroom airlocks should be designed at the same classification as the area they support. Consider having sweeping airflow from the clean side to the dirty side with the return close to the dirty side's entry door.
• Cleanrooms should be designed to take up just one classification level, thereby eliminating confusion (e.g., no Class A (ISO 5] and Class B [ISO 6] in the same room).
• Filling lines and critical processes should be physically separated from operators. Equipment within isolators or RABS can help to significantly control contamination.
• When using isolators with VHP (H₂O₂), consider the location of HVAC room inlets and returns to avoid potential cooling impact on the sterilization process.
• Cleanroom access should be limited to those personnel essential to the operation, including quality assurance personnel. It is amazing to see how the industry has added nonessential personnel to critical areas in the guise of quality when people are the number one environment problem.
• Each processing area should have its own separate gowning area and exit, without overlapping pathways.
• Environmental-monitoring (EM) data control should be automated. Software systems are available that effectively track and manage all environmental data based on the latest regulatory guidelines.
• Design equipment to limit product exposure to personnel and the environment, including any environmental monitoring.
• Include capability to clean-in-place and sterilize-in-place (CIP/SIP).
• Consider 100% check weighing.
• Check preventative maintenance operations capability from outside the aseptic area. Determine whether there is a need for process-piping temperature control from the manufacturing process through filling. Does the piping travel through noncontrolled environments?
• Determine whether there is a maximum time for filled product to be removed from cold storage. These operations should be considered as automation of process.
• Use of disposables and presterilized items can be positive, but transfers into Class-A areas without a verifiable method of sterilizing the bagged sterile items can be troublesome. Use VHP or CLO₂ to mitigate this concern.

Applying Quality by Design to Sterile Manufacturing Processes 3

By: Warren Charlton
Technology transfer
Assuming that the manufacturing process design space has been properly developed and CQAs and DOEs have been verified, technology transfer, as well as scale-up, should go smoothly. As with most typical API-manufacturing operations, process design and controls ensure a state of control. When these approaches and technologies are applied to a batch-based system, they may, in the future, lead to more continuous sterile processing due to the increased level of control.
The following sterile-process technologies are well defined in the literature: sterile filtration (F), autoclaves (A), steam cycles, dry-heat (DH) ovens and tunnels, gas sterilization (ETO), chlorine dioxide (ClO₂), ionization radiation (ebeam, gamma), and terminal sterilization (steam, gamma radiation). These technologies must be validated with the specific component, drug product, or commodity.
Sterilization technology review. Sterile filtration. Filter validation is normally done by filter manufacturers or outside laboratories but the responsibility remains with the drug manufacturer (5). Steam sterilization. Steam sterilization (i.e., autoclaves, sterilize-in-place [SIP]), is the method of choice whenever possible due to data available and capability to assure sterility. This method is used for transfers into the aseptic area, process sterilization, SIP of the product delivery system, and product contact-component sterilization. Steam sterilization is limited by its temperature and pressure impact. Many plastic items, therefore, require other methods of sterilization. Control is typically ≥121.1 °C. Product contact components must have had prior pyrogen removal steps to ensure expectation of a minimum 3-log reduction.
Dry-heat sterilization. The DH approach is mainly used for glass components because of the temperature during processing (a dry-heat oven is typically controlled ≥200–250 °F and dry-heat tunnels are typically controlled at 275–350 °F). Dry-heat sterilization can also be validated for a 3-log or greater pyrogen reduction.
ETO. Gas sterilization or ETO, is used for product contact plastics and commodity transfers. The method is not used in processes and operations due to safety issues. Product contact components require ETO degassing after the cycle is completed. ETO is a toxic and hazardous chemical. Cycle control includes ETO concentration, humidity, and pressure and similar to steam sterilization, requires prior pyrogen removal steps.
Ebeam. Ebeam is easy to define and makes it easy to control sterilization of the surface and, to some extent, the depth of exposure and microbial kill. Recently, the ebeam method has been used to sterilize the lids of syringe bulk containers before filling. Because ebeam is not currently used for product contact components, pyrogens are not an issue. The sterilization dose for radiation processes (i.e., ebeam and gamma) are 25 kGy (2.5 Mrad).
Vaporized hydrogen peroxide (VHP). VHP is currently the method of choice for isolator decontamination. Sterility is sometimes claimed. Because VHP is not a true gas, it can be affected by cold spots. VHP also can be used to sanitize transfer items into aseptic-filling operations, but cannot be used to sterilize or depyrogenate components. Cycle requires control of concentration of H₂O₂, relative humidity, and temperature. Circulation fans within an isolator are often used to help provide constant conditions within the isolator (5).
Chlorine Dioxide (CD). CD is currently the least used method throughout industry for sterilization, but provides significant opportunities because it is a true gas and can be validated for sterility. Areas of opportunity include isolators, transfers to the aseptic area, and processing equipment. CD is widely used in the food industry for sanitization and disinfection. Concentration can be monitored and controlled. Aeration is repeatable. Passivation of stainless steel may be required. Controls consists of concentration, humidity, temperature, and pressure.
Other sterilization technologies include gamma radiation, which is used for product terminal sterilization and component sterilization by contract manufacturing organizations (CMOs). Gamma-radiated presterilized syringes are a common usage of this technology in the industry. Transfer of these types of presterilized components to aseptic-filling areas can be troublesome, however, without a defined transfer sterilization method. Recently, some equipment suppliers have included ebeam sterilization to improve this transfer. Ozone (O₃) technology has potential as a sterilization method as well, but is not currently commercially available. Peracetic acid as a sterilization technology may still be used by a few companies, but most have switched to VHP. Other forms of sterilization or environmental control have limited support data and therefore represent an increased risk requiring in-house technical knowledge (e.g., high-intensity light, ultraviolet light).

Applying Quality by Design to Sterile Manufacturing Processes 2

By: Warren Charlton
Design of experiment and risk analysis. Design of experiment (DOE) and risk analysis are recommended in the draft guidance to provide data that support process design. The emphasis should be to reveal relationships between variable inputs (e.g., component characteristics, processing parameters) and resulting outputs (e.g., in-process material, intermediates, or the final product).
Early-stage product assurance. The draft guidance states "early process design experiments do not need to be performed under CGMP conditions." Emphasis during early stage development are verification rather than validation. "Decisions and justifications of the controls should be sufficiently documented and internally reviewed," according to the draft guidance. Some examples of early-stage product assurance are outlined below (4, 5).

• Viral and impurity clearance studies are required because they have a direct impact on product quality
• Cleaning verification would replace validation during the early stages of development
• Sterility must be ensured but may not be completely validated (e.g., if performing sterile filtration on a water-based product, a filter integrity test may be performed using water-for-injection to demonstrate that the filter still meets its manufacturer's integrity test value)
• Preliminary specifications, tests, acceptance criteria, and limits should be operational.

Overall, the further along a company is in its product development, the more verification and validation are expected. Process controls that address variability can help to ensure the product's quality. Models. "It is important to understand the degree to which models represent the commercial process including differences that may exist," states the guidance (4). The significance of understanding the process increases with a model that properly reflects all the variants of the process and product. A good model can be used as a tool for process and equipment design, process control development, estimating variablity, and training personnel.
Process analytical technology and qualification. Process analytical technology (PAT) "uses timely analysis and control loops to adjust processing conditions so that the output remains constant," states the guidance (4). As a result, there is a higher degree of process control. It is unclear, however, whether maintaining the process within the design space using PAT reduces process-validation testing. The draft guidance places added emphasis on design (qualification = design plus verification). There are requirements, for example, to challenge the process under load, to test interventions, and to test stoppage and start-up routines as expected for production.
The common industry practice of testing three batches for PQ may no longer apply. Specific requirements regarding when batches can be released and the information needed to begin commercial distribution are included in the draft guidance. PQ must have a higher level of sampling, testing, and scrutiny of process performance to confirm the product's quality during batch processing.
According to the draft guidance, "In the case of PAT strategy, the approach to process qualification will be different from that for other process designs," but the agency did not explain how the process will be different (4). This difference is important because additional use of PAT leads to additional validation to verify the technology's capability. Industy will assume this means an easier validation approach because conformation of control would be readily available, but there are no examples provided in the draft guidance.
With regard to continued process verification, the goal is to ensure the process remains in a state of control during commercial operations (3, 4). The draft guidance recommends manufacturers include intra-batch as well as inter-batch variation as part of their continued verification program. PQ sampling levels should continue until variability is assessed.
The draft guidance suggests that a statistician be involved in the development of the company's data collection plan and in the selection of statistical methods to evaluate process stability and capability. Included in this review should be quantitative statistical methods where feasible. Process flow diagrams for commercial manufacturing, for example, should be completed as follows:

• Describe each unit operation and placement in the process
• Describe process monitoring and control points
• Describe components and other processing material inputs
• Describe expected outputs (e.g., in-process and finished products).

In addition, the flow diagrams should preserve a life-cycle approach to facilitate comparison and decision-making regarding their comparability.

Applying Quality by Design to Sterile Manufacturing Processes 1

By: Warren Charlton
In November 2008, the US Food and Drug Administration issued a much needed document titled, Draft Guidance for Industry on Process Validation: General Principles and Practices. The draft guidance clarifies the quality-by-design (QbD) approach to processing human and veterinary drugs, including biologics, active pharmaceutical ingredients (APIs), and medical devices. Although the document may have been better titled "process design," it has addressed many industry concerns regarding how to take a life-cycle approach and how to meet regulatory expectations with regard to validation. Many questions still remain, however, and industry must grasp and integrate the proposed guidance concepts along with the International Conference on Harmonization's guidelines Q8 Pharmaceutical Development and Q9 Quality Risk Management (1, 2).

Quality-by-Design overview

Individuals working in pharmaceutical sterile development and manufacturing will undoubtedly be the most affected by regulations associated with QbD because of the unique associated technologies, processes, and products currently in use and development (see sidebar, "Quality-by-Design Overview"). Many of these technologies require new manufacturing processes that support existing traditional fill-finish operations. Many facilities, utilities, processes, and equipment may need to be modified (e.g., improving controls to reduce variability). This article provides an overview of FDA's draft guidance on process validation and QbD's impact on sterile manufacturing operations. Manufacturing process

Figure 1. (ALL FIGURES ARE COURTESY OF THE AUTHOR)

The following sections discuss how the draft guidance may influence the manufacturing process and product development. Processes addressed include manufacturing, aseptic manufacturing, lyophilization (freeze drying), and others that depend on multiple-unit operations (see Figure 1). Process design is typically the biggest challenge for a company's process development team and has tremendous impact on the success of the product. Sterile manufacturing has become increasingly more complex because of the increase in the number of poorly stable compounds, new technologies, unit operations, and controls. Numerous biotechnology companies, for example, use unique freeze–thaw systems to support dispensing and bulk-drug storage at temperatures often ≤ –80 °C that provide complete control of bulk-drug thawing, dispensing , freezing, and storage.
FDA's draft guidance suggests that the process development team design a process suitable for routine commercial manufacturing that can consistently deliver a product that meets its critical quality attributes (CQAs). The team's objectives are to: understand the sources of variation, detect presence and degree of variations, understand the impact of variation on the process and product attributes, and control variation in a manner that is commensurate and proportionate to the risks presented to the process and product. FDA further recommends an integrated team approach, with members representing multiple disciplines. The team should have solid project-management skills and archiving capabilities, be able to capture scientific knowledge and maintain project plans, and have full senior-management support. The latter may involve regular reports or presentations to senior management (3, 4).
In addition, the draft guidance calls for process decisions and justifications of control to be documented, internally reviewed, and preserved for later use in the product life cycle. Verification and mapping of the process design through to commercial documentation is required and planned changes should be documented and justified. Finally, after the performance qualification (PQ), a report documenting and cross-referencing results, data, issues, nonconformances, corrective actions, and overall conclusions regarding whether the process is in an adequate state of control must undergo management review and approval.

Cleaning Verification: Method Development and Validation Using Ion Mobility Spectrometry2

By: Elizabeth Galella, Scott Jennings, Madhavi Srikoti, Elizabeth Bonasso
IMS distinguishes ions of a given compound on the basis of their velocities through a drift tube under the influence of a weak electric field. Ion velocity (v) is proportional to the applied electric field (E)
in which K is the ion mobility in units of cm² /Vs. Drift time t is proportional to 1/K , ion mass m, and collisional cross section C. Reduced mobility K_o for an analyte is calculated by normalizing its drift time to that of the internal calibrant as in the following equation: in which (K_o^C ) is the reduced mobility of the calibrant and t C is the drift time of the calibrant.

Figure 3: Illustration of ions of varying sizes migrating toward the detector through a constant flow of air.

These velocities, or ion mobilities, are determined and selective for a given compound. The high sensitivity of the instrument (nanogram to picogram range) provides for an excellent technique to quickly verify equipment cleanliness. Figure 3 illustrates the separation process described in the theory above. This technique allows for positive or negative mode of detection. Each mode has a unique internal calibrant used for calculation of the K_o. In the positive mode the internal calibrant is nicotinamide. In the negative mode the internal calibrant is methyl salicylate. 
Advantages of IMS

Figure 4: Efficiency gained by ion mobility spectrometry (IMS) versus high-performance liquid chromatography (HPLC).

Ion mobility has unique advantages over the conventional HPLC technique for cleaning verification. Although the principles of IMS technology have been well established, its uses in the pharmaceutical industry have, until recently, been limited. This has been changed by the development of easy-to-use commercial instruments of moderate cost, with small footprint and high sensitivity (4). As shown in Figure 4, when compared with the typical HPLC method, the IMS technique can save a significant amount of time in the analyzing cleaning verification samples. Furthermore, a quick response time in getting results can reduce down time of key manufacturing and packaging equipment, ultimately leading to significant cost savings and increased productivity for the company (1,3). Method development
As with most analytical methods, the IMS instrument parameters must be examined and optimized for each compound as part of method development. These parameters include ionization mode, desorber temperature, injection volume, post-injection delay, drift flow velocity, and analysis time. The development process begins by examining the selectivity of the target compound in both positive and negative modes. In the authors' experience, most pharmaceutical compounds respond better in the positive mode because of the presence of basic functional groups within the molecule. The desorber temperature for compounds analyzed in the positive mode is typically set at ~290 °C. This temperature should be hot enough to effectively desorb the sample off the substrate but not so hot as to thermally degrade the compound. Typical sample volumes are 1 µL, injected using a 10-µL syringe. The post-injection delay is dependent on this volume and the type of solvent being used. A large injection volume of a solvent of low volatility requires a longer post-injection delay because it will take more time for the solvent to evaporate. Finally, the analysis time and drift flow velocity settings are dependent on the IMS response. A sufficient analysis time and drift flow are necessary to ensure depletion of all sample ions in the drift tube. In a case where the analysis time is too short, or drift flow velocity is too low, sample carryover may be an issue.

Figure 5: Second-order polynomial calibration curve.

The generally accepted practice is that an analytical method must exhibit sufficient sensitivity to measure the concentration at the ARL of the active agent or degradants being investigated in the swab and rinse samples. Typically, a limit test is used to establish a pass–fail criterion for swab samples and determine whether the manufacturing equipment is clean. Determining the pass–fail limit is accomplished by evaluating the instrument response curve and the system's precision. It allows analysts to capture the ARL within a linear calibration range, therefore providing an accurate means of determining the cleanliness of equipment parts. In the response curve, the "target level" represents the highest point within linear range (see Figure 5). The "action level" represents the resulting pass–fail level after adjusting the target level for instrument response variability. Any sample that responds above this action level registers as a failure. Any sample that responds below this action level registers as passing. Consequently, there is an area of uncertainty between the target and action level within which false-positive results may occur. However, these are generally infrequent occurrences. 
Method validation
Typical IMS method validation parameters to be considered include selectivity, linearity, reproducibility, recovery and solution stability. Selectivity of the analyte is determined based on evaluation of the molecular structure dictating the mode of detection. Selectivity from the sample matrix is based on minimizing the interference from product excipients and cleaning detergents, and should be examined before performing the recovery experiment. In addition, the linearity is based on a second-order polynomial curve obtained from the response versus the amount introduced, as described previously. Reproducibility must be used to determine the action level. Therefore, these parameters must be considered during method development and verified during validation.

Table I: Recovery of active from factory-finished, stainless-steel plates.

Recovery. After determination of the action level, sample recovery is determined from a 10-in.² stainless-steel coupon. A stock sample solution is prepared at 100% of the API concentration at the target level. The solution is then spiked onto three separate stainless steel coupons. The surfaces are allowed to dry before swabbing with a clean Texwipe alpha polyester swab. According to industry guidelines, the swabbing technique includes horizontal strokes to swab the entire designated area with one side of the swab head and then vertical strokes with the other side of the swab head. Each swab is then placed back into the vial containing sample diluent, extracted, and later analyzed for active content. Table I shows the recovery values from stainless-steel surfaces and corresponding percent relative standard deviation (%RSD) at the action level for a novel pharmaceutical compound under development. Table I shows the recovery results, which are typical of those generated by swabbing three different stainless-steel coupons. The observed variation is primarily a result of the inherent variability in the surface finish of the coupon. These values are independent of the analysis technique and may be optimized by adjusting the swabbing procedure and/or materials. However, application of a predetermined correction factor to swab assay results can be used to compensate for recovery values as low as 50% during method validation. Solution stability. Sample and standard solution stability studies are typically performed to cover 24 h at room temperature. Additional validation studies, such as robustness and intermediate precision, are not included in the authors' validation plan for a limit test. All of the parameters described previously must meet pre-established criteria as defined in the validation protocol. The criteria and validation protocol must conform to International Conference on Harmonization guidelines as well as internal standard operating procedures.
Conclusion
The ease of use and the small footprint of ion mobility spectrometry instrument allows for the system to be implemented in various work environments, such as quality control, and to report results with great sensitivity (nanogram to picogram range). Moreover, data reduction software package upgrade ensures 21 CFR Part 11 compliance. The use of this software, in addition to a limit test, has simplified the process of data manipulation, resulting in high-confidence passes for each clean sample analyzed.
Elizabeth Galella* is a research scientist, Scott Jennings is a senior research scientist, Madhavi Srikoti is an associate research scientist, Elizabeth Bonasso is a research scientist, all at the analytical research and development unit of the Pharmaceutical Research Institute, Bristol-Myers Squibb, One Squibb Drive, New Brunswick, NJ 08903, elizabeth.galella@bms.com
References
1. G. Walia et al., "Using Ion Mobility Spectrometry for Cleaning Verification in Pharmaceutical Manufacturing," Pharm. Technol. 27 (4), 72–78 (2002).
2. G. Walia et al., "Implementing Ion Mobility Spectrometry as a Cleaning Verification Method," Pharm. Technol.27 (3), 22–25 (2002).
3. D. Brand, X. Li, and T. Wortley, "Ion Trap Mobility Spectrometry: Reducing Downtime in Cleaning Validation and Verification," GE Sensing, http://www.gesensing.com/toolsupport/whitepaper.htm accessed June 16, 2009.
4. D. Brand et al., "Direct Swabbing and Surface Recovery with Ion Trap Mobility Spectrometry," GE Sensing, http://www.gesensing.com/products/resources/datasheets/GEsensing_whitepaperfinal.pdf accessed June 2, 2009.
5. R. DeBono, "Ion Mobility Spectrometry: A Fast, Sensitive, and Robust HPLC Alternative," Appl. in Chrom. March, 20–23 (2002), http://trace.smithsdetection.com/Documents/LifeSciences/LE203sBarringer.pdf.
6. G.A. Eiceman, "Advances in Ion Mobility Spectrometry," Crit. Rev. Anal. Chem.22 (1, 2), 471–490 (1990).

Cleaning Verification: Method Development and Validation Using Ion Mobility Spectrometry1

The cleanliness of manufacturing and packaging equipment in the pharmaceutical industry is critical to ensure product efficacy, personal safety, and the absence of unwanted active pharmaceutical ingredient (API) before the introduction of a different compound. Upon completion of processing a drug substance or tablet formulation, a detergent cleans each part of the pharmaceutical equipment. The cleaning verification process in the pharmaceutical industry involves several steps. First, the clinical supply or manufacturing unit of a company submits samples for analysis following the equipment cleaning process. The sample analysis is completed by an analytical department or quality control (QC) personnel. The results are reported back to the clinical supply unit, and the process is repeated until all samples have met the acceptable residual limit (ARL). The subject exposure limit (SEL) (in units of mg/subject/day) for a given compound is provided by a drug-safety evaluation group and is an industry-wide practice. The ARL of the API in each swab sample is a set at one-tenth of the SEL.
These types of samples have been analyzed by high-performance liquid chromatography (HPLC). In an effort to significantly reduce sample turnaround time, which can take as long as several days, ion mobility spectrometry (IMS) has been evaluated as an alternative method for analysis (1–4). The time taken for analysis is important because the faster that cleaning can be confirmed, the faster the plant can return to operation. This article discusses the theory of IMS, the benefit of IMS over HPLC analysis, and experimental considerations for method development and validation.
Theory

Figure 1: The IonScan-LS ion mobility spectometer with the Cobra autosampler.(FIGURES COURTESY OF THE AUTHORS)

IMS is a type of separation technique, similar to time-of-flight mass spectrometry, that distinguishes ions of a given compound based on their velocities through a drift tube under the influence of a weak electric field. The technique characterizes chemical substances based on their gas-phase ion mobilities and provides detection and quantitation of trace analytes (1). There are several IMS instrument vendors. Figure 1 shows the IonScan-LS ion mobility spectrometer with the Cobra autosampler (Smiths Detection, Warren, NJ), which was used in the studies described in this article.

Figure 2: Cross-sectional view of the ion mobility spectrometer sample inlet, ionization chamber, drift tube, and detector.

To perform the IMS analysis, a sample solution containing a compound of interest is injected by the autosampler onto a PTFE substrate and allowed to dry. Figure 2 shows the sample inlet, ionization chamber, drift tube, and detector (5). Sample is introduced by heating the substrate to ~290 °C, which results in desorption or vaporization of the sample into an inlet tube. Primary ion formation occurs through atmospheric pressure chemical ionization (APCI) using nickel-63 as the radioactive source. Following many collisions, product ions are formed and gated into the drift tube. These ions-based on size, shape, and charge-travel through the drift tube toward the detector at different velocities. In contrast to mass spectrometers, separation of ions is based on a size–charge relationship rather than the mass–charge ratio (6).