Cleaning Validation in Pharmaceutical Manufacturing: Limits, Sampling and Regulations

Cleaning validation is the documented process of proving that a cleaning procedure reliably removes residues from pharmaceutical manufacturing equipment to a safe, predetermined level. The residues of concern include the active ingredient of the product just made, its excipients and degradation products, the detergents and solvents used for cleaning, and microbial contamination. The aim is to make sure that nothing left behind on shared equipment can contaminate the next product made on it and harm a patient.

In the European Union's GMP guidance, cleaning validation is defined as "documented evidence that an approved cleaning procedure will reproducibly remove the previous product or cleaning agents used in the equipment below the scientifically set maximum allowable carryover level." It is a core requirement of good manufacturing practice (GMP) wherever equipment is shared between products.

This article explains why cleaning validation exists, how the main regulators approach it (the US Food and Drug Administration, the European Medicines Agency and the European Commission, and the Pharmaceutical Inspection Co-operation Scheme, PIC/S), how acceptance limits are set, including the modern health-based approach using permitted daily exposure (PDE) and maximum allowable carryover (MACO) calculations, how samples are taken and analysed, and the most common pitfalls. A fully worked, hypothetical calculation is included.

This is a general educational overview. Anyone designing or reviewing a real cleaning validation programme should work from the current text of the applicable regulations and guidance, with qualified toxicological and quality input.

Why cleaning validation matters

The risk of cross-contamination

Most pharmaceutical plants make more than one product on the same equipment. A tablet press, blender, granulator, reactor or filling line may be used for one product in the morning and a different one in the afternoon. If residues of the first product remain, they can end up in the second. This is cross-contamination, and its consequences depend on what the residue is.

A trace of a harmless excipient may not matter. A trace of a potent hormone, a cytotoxic cancer drug, a highly sensitising antibiotic such as a penicillin, or a substance that harms an unborn child could cause real injury to a patient who never chose to take it. Because the patient taking the second product receives no benefit from the contaminant, the EMA's guideline on health-based exposure limits notes that such residues "provide no benefit to the patient or target animal and may even pose a risk."

Historical incidents that shaped regulation

The FDA's 1993 Guide to Inspections: Validation of Cleaning Processes describes how the agency's concerns developed. Early attention focused on gross insanitation and on specific high-risk cross-contamination, especially penicillin contaminating non-penicillin products and potent steroids or hormones contaminating other products.

Two cases from the late 1980s and early 1990s raised the profile of cleaning validation:

• The 1988 Cholestyramine Resin recall. A bulk pharmaceutical chemical was contaminated with low levels of intermediates and degradation products from the manufacture of agricultural pesticides. The likely cause, according to the FDA guide, was the reuse of drums of recovered solvent without adequate controls, testing or validated cleaning. Contaminated bags used at a second facility then spread the contamination to lots made there.
• A 1992 import alert. The FDA took action against a foreign bulk drug manufacturer that made potent steroid and non-steroidal products on common equipment. The firm had recently validated its cleaning, but the FDA found the validation inadequate because it only tested for the absence of the previous compound itself. Thin-layer chromatography of rinse water revealed reaction by-products and degradants that the firm had not looked for.

These cases showed two things that remain central today: residues can come from unexpected sources, and testing only for the obvious compound can miss the real hazard.

Cleaning is a risk-reducing measure

The EMA describes cleaning as "a risk reducing measure." It sits alongside other controls, such as facility design, airflow, gowning and segregation, in a wider strategy for preventing cross-contamination. Cleaning validation provides the evidence that this particular control works.

The regulatory framework

United States: 21 CFR 211.67 and the FDA guide

In the United States, the legal requirement for cleaning comes from the current good manufacturing practice regulations for finished pharmaceuticals. Section 211.67(a) of Title 21 of the Code of Federal Regulations states that "Equipment and utensils shall be cleaned, maintained, and, as appropriate for the nature of the drug, sanitized and/or sterilized at appropriate intervals to prevent malfunctions or contamination that would alter the safety, identity, strength, quality, or purity of the drug product beyond the official or other established requirements."

Section 211.67(b) requires written procedures for cleaning and maintenance, including assignment of responsibility, cleaning schedules, a detailed description of methods, equipment and materials (including how to dismantle and reassemble equipment), removal of previous batch identification, protection of clean equipment from contamination before use, and inspection for cleanliness immediately before use. Section 211.67(c) requires records of cleaning and inspection.

The regulation does not itself use the word "validation" for cleaning, but the FDA's Guide to Inspections: Validation of Cleaning Processes, dated July 1993, sets out what investigators expect to see. Its key expectations include:

• Written procedures for each cleaning process, with separate procedures where needed for different situations, such as batch-to-batch cleaning versus product changeover, or water-soluble versus non-water-soluble residues.
• General validation procedures that state who is responsible for performing and approving validation studies, the acceptance criteria, and when revalidation is needed.
• Specific written protocols, prepared in advance for each system or piece of equipment, covering sampling procedures, analytical methods and their sensitivity.
• Studies carried out according to protocol and documented.
• A final report approved by management stating whether the cleaning process is valid, with data supporting the conclusion that residues have been reduced to an "acceptable level."

The guide also says that between batches of the same product, a "visibly clean" standard may be enough and validation is not required, and that equipment that is very hard to clean, such as fluid bed dryer bags or equipment handling tarry residues, may be dedicated to a single product.

Importantly, the FDA does not set specific acceptance limits or prescribe how they must be calculated. The guide explains that the variety of equipment and products is too wide for a single standard, and instead expects each firm's rationale to be "logical, practical, achievable, and verifiable" and based on knowledge of the materials involved. The Wikipedia summary of the topic notes that current FDA expectations also include risk management principles and health-based exposure limits as the basis for limits on active ingredients.

European Union: Annex 15 and the HBEL guideline

In the EU, cleaning validation is covered mainly by three documents.

EudraLex Volume 4, Annex 15: Qualification and Validation. The revised Annex 15, published by the European Commission on 30 March 2015 with a deadline for coming into operation of 1 October 2015, contains a dedicated section on cleaning validation (section 10). Its main requirements are summarised in the table below.

Annex 15 also defines cleaning verification as "the gathering of evidence through chemical analysis after each batch/campaign to show that the residues of the previous product or cleaning agents have been reduced below the scientifically set maximum allowable carryover level." Validation proves a process works in general; verification checks a specific cleaning event.

EU GMP Chapters 3 and 5. These chapters, on premises and equipment and on production, were revised to promote a science- and risk-based approach to preventing cross-contamination and to refer to a "toxicological evaluation" for setting threshold values.

The EMA guideline on health-based exposure limits. The Guideline on setting health based exposure limits for use in risk identification in the manufacture of different medicinal products in shared facilities (EMA/CHMP/CVMP/SWP/169430/2012) was adopted by the Committee for Medicinal Products for Human Use on 20 November 2014 and came into effect on 1 June 2015, with phased implementation for existing products. It describes how to derive a permitted daily exposure (PDE) for each active substance, explained in detail below. Annex 15 paragraph 10.6 refers to it directly.

The guideline's implementation was phased:

The EMA question-and-answer document (2018). On 19 April 2018 the EMA published Questions and answers on implementation of risk-based prevention of cross-contamination in production and the HBEL guideline (EMA/CHMP/CVMP/SWP/246844/2018). Its answers are some of the most practically important statements in this field and are discussed throughout this article. The first answer is unambiguous: "Yes, HBELs should be established for all medicinal products."

PIC/S

The Pharmaceutical Inspection Co-operation Scheme is an international arrangement between GMP inspectorates. For many years its main cleaning validation guidance was PI 006-3, Recommendations on Validation Master Plan, Installation and Operational Qualification, Non-Sterile Process Validation, Cleaning Validation, dated 25 September 2007. According to secondary summaries, it described acceptance criteria based on the most stringent of three tests: visually clean, no more than 10 ppm of one product in another, and no more than 0.1 percent of the normal therapeutic dose of one product in the maximum daily dose of the next.

PIC/S has since replaced this with PI 006-4, Recommendations on Qualification and Validation, which, according to the industry publication GMP Insiders, came into force on 1 October 2026. Its cleaning validation section is reported to be rewritten around health-based exposure limits, with statements such as "Visually clean is the minimum standard, not the standard" and that "limits are not to be set at the calculated HBEL level." It also reportedly prefers swab sampling to rinse sampling where feasible and requires cumulative carryover across shared equipment trains to be considered. PIC/S also published PI 053-1 (2020), a transposition of the EMA's 2018 question-and-answer document.

Industry guidance

Alongside regulatory documents, industry bodies publish detailed technical guidance. Examples include the International Society for Pharmaceutical Engineering's Guide: Cleaning Validation Lifecycle (August 2020) and its Baseline Guide Volume 7 on risk-based manufacture of pharmaceutical products (second edition, July 2017), which the EMA's 2018 Q&A cites, and the Parenteral Drug Association's Technical Report 29 on cleaning validation (1998, revised 2012). These are not legally binding but are widely used.

Setting acceptance limits: from 10 ppm to health-based limits

The central question in cleaning validation is: how clean is clean enough? The answer has changed significantly over the past three decades.

The traditional criteria

For many years, industry used three main criteria, which the FDA's 1993 guide lists as limits cited in industry literature and presentations:

• An analytical detection level, such as 10 parts per million (ppm). No more than 10 ppm of the previous product's active ingredient should appear in the next product.
• A biological activity level, such as 1/1000 of the normal therapeutic dose. No more than one-thousandth of the smallest therapeutic dose of the previous product should appear in the largest daily dose of the next product.
• An organoleptic level, meaning no visible residue. Equipment should look clean.

Companies usually applied whichever criterion gave the strictest limit. A further traditional method, used mainly when no therapeutic dose existed (for cleaning agents or intermediates), derived an acceptable daily intake from the acute toxicity measure LD50, the dose that kills half of test animals.

These approaches had the advantage of simplicity, but they were not tied to the actual hazard of the residue. A 1/1000 factor applied to a mild drug and to a highly toxic one treats them as if a thousandth of a dose were equally safe for both. The 10 ppm rule has no toxicological basis at all.

Why the traditional limits were abandoned

The EMA guideline states the problem directly: the variety of approaches used to set carryover limits "often do not take account of the available pharmacological and toxicological data," so "a more scientific case by case approach is warranted." In consultation on its 2018 Q&A, the EMA's implementation team accepted stakeholder proposals not to pursue traditional approaches such as 1/1000th of a dose. The final Q&A instead requires HBELs for all medicinal products.

The 2018 Q&A also rules out the LD50: "No, LD50 is not an adequate point of departure to determine a HBEL for drug products." Lethal-dose data describe acute death, not the subtle effects of small daily exposures over time.

One industry publication summarises the current European position by saying that "the previously used criteria 1/1000 dose and 10 ppm are not based on scientific facts" and should no longer be used as acceptance criteria.

The health-based approach

Under the health-based approach, the starting point is a substance-specific health-based exposure limit (HBEL). In the EMA framework the HBEL is usually expressed as a permitted daily exposure (PDE), defined as "a substance-specific dose that is unlikely to cause an adverse effect if an individual is exposed at or below this dose every day for a lifetime." In other frameworks a similar concept is called the acceptable daily exposure (ADE).

The HBEL is then used to calculate how much residue can be tolerated on the equipment, the maximum allowable carryover (MACO), and from that the residue limits on surfaces and in swab or rinse samples.

Calculating the permitted daily exposure

The steps

The EMA guideline sets out four steps for deriving a PDE, based on Appendix 3 of the ICH Q3C guideline on residual solvents:

1. Hazard identification: review all relevant data, including non-clinical pharmacodynamic data, repeat-dose toxicity, carcinogenicity, genotoxicity, reproductive and developmental toxicity, and clinical data on therapeutic and adverse effects.
2. Identification of critical effects: the most sensitive indicators of adverse effects in non-clinical studies (unless shown not to be relevant to humans), plus any clinical therapeutic or adverse effect.
3. Determination of the NOAEL: the no-observed-adverse-effect level, defined as "the highest tested dose at which no 'critical' effect is observed." If several studies show the critical effect, the lowest NOAEL is used. If no NOAEL is available, the lowest-observed-adverse-effect level (LOAEL) may be used.
4. Application of adjustment factors to account for uncertainty.

The formula

The PDE is calculated as:

PDE = (NOAEL × Weight Adjustment) / (F1 × F2 × F3 × F4 × F5)

The adjustment factors defined in the EMA guideline are:

The guideline refers to Appendix 3 of ICH Q3C for further guidance on choosing F1 and F4, allows additional modifying factors if well supported, and accepts deviations from the defaults if they are scientifically justified. Where the PDE is based on human data, it may be acceptable to use only F2 and possibly F5. If a daily dose is expressed per patient rather than per kilogram, a standard body weight of 50 kg is used for human medicines. Alternatives to the NOAEL, such as the benchmark dose, are also allowed.

Special cases

The EMA guideline addresses several situations where the standard PDE approach needs modification:

• Genotoxic substances without a threshold. For substances that damage DNA with no safe threshold, the guideline allows a threshold of toxicological concern (TTC) of 1.5 micrograms per person per day, drawn from the EMA guideline on genotoxic impurities. Where carcinogenicity data are sufficient, a compound-specific assessment should be used instead.
• Highly sensitising substances. EU GMP Chapter 3 requires dedicated facilities for products with a high sensitising potential where scientific data do not support an acceptable exposure level or the risk cannot be adequately controlled.
• Therapeutic macromolecules and peptides. Proteins and peptides tend to degrade and become inactive under the pH extremes and heat used in cleaning, so a PDE for the intact active may not be required.
• Lack of reproductive toxicity data. Where such data are missing, an additional adjustment factor (for example, 10) may be applied if justified.
• Early investigational products. Where data are limited, default categories based on expected potency and toxicity, similar to tiered TTC approaches, may be used.
• Route of administration. If the next product is given by a different route, with clearly different bioavailability (for example, more than 40 percent different), correction factors should be applied.

Who calculates the PDE

The 2018 EMA Q&A states that HBELs should be determined by a person with "adequate expertise and experience in toxicology/pharmacology," familiarity with pharmaceuticals, and experience in setting exposure limits. It is not acceptable to "purchase" HBEL assessments without checking that the provider is a qualified contractor. The guideline also expects a documented literature search, an expert discussion of the critical effects, and a summary page for inspectors.

From PDE to MACO and surface limits

The MACO formula

Once the PDE of the previous product's active ingredient is known, the maximum allowable carryover into the next product can be calculated. The general form, as presented in ISPE training material, is:

MACO = (PDE of previous product × minimum batch size of next product) / maximum daily dose of next product

where:

• PDE is the permitted daily exposure of the active ingredient in the previous product (for example, in micrograms per day);
• minimum batch size is the smallest batch of the next product that could be made on the equipment (in milligrams or grams);
• maximum daily dose is the largest amount of the next product a patient could take in a day (in the same mass unit).

The logic is that the residue will be spread through the next batch. If the batch is small and the patient takes a lot of the next product each day, more of the residue reaches each patient, so the allowable carryover is lower.

The traditional dose-based formula had the same structure, with the minimum therapeutic dose of the previous product divided by a safety factor in place of the PDE. ISPE training material lists the commonly used, now obsolete, safety factors:

Where several products share equipment

When several products share equipment, the calculation is usually performed for every possible pair of previous and next products, and the most restrictive result is used. ISPE training material describes a simplified approach using the lowest PDE among the possible previous products and the smallest batch size among the possible next products.

Surface limits

The MACO is a total mass for the whole equipment train. To compare it with sampling results, it is converted to a limit per unit area, sometimes called the surface area limit (SAL):

SAL = MACO / shared surface area

The shared surface area is the total product-contact surface common to both products. Annex 15 paragraph 10.6 also requires acceptance criteria to consider "the potential cumulative effect of multiple items of equipment in the process equipment train," and PIC/S PI 006-4 is reported to warn against assuming that contamination is spread uniformly over all surfaces.

Swab and rinse limits

For swab sampling, the surface limit is multiplied by the area swabbed to give the maximum allowable amount in one swab. If the swab is extracted into a known volume of solvent, the limit can be expressed as a concentration in the extract. For rinse sampling, the limit is the amount allowed in the rinse volume that contacted a known surface area.

Alert limits and process capability

A crucial point in modern guidance is that the cleaning limit is not the same as the HBEL-derived MACO. The 2018 EMA Q&A says the HBEL guideline "is not intended to be used to set cleaning limits at the level of the calculated HBEL." For existing products, historically used cleaning limits "should be retained and can be considered alert limits" provided that, taking process capability into account, they give sufficient assurance that the HBEL will not be exceeded. Results above an alert limit should trigger investigation, and repeated excursions are not acceptable where they show the process is not in control. Statistical methods may be used to judge control.

In other words, the health-based calculation tells you the boundary of safety. The actual operating limits should sit well inside it, set by what a capable, well-controlled cleaning process actually achieves.

A worked example (hypothetical)

All values below are invented for illustration. They do not describe any real drug, product or facility, and the adjustment factor choices are simplified.

Step 1: derive the PDE for the previous product's active ingredient

Suppose the active ingredient in Product A has a critical effect identified in a long-term rat study, with a NOAEL of 0.05 mg/kg/day.

• Weight adjustment: 50 kg (standard human body weight in the EMA guideline).
• F1 (rat to human): assume 5, within the guideline's range of 2 to 12.
• F2 (individual variability): 10.
• F3: the hypothetical study lasts longer than 4 weeks, so the 10-fold short-duration factor does not apply; for simplicity this example assumes 1.
• F4 (severe toxicity): assume 1, as no severe toxicity is assumed.
• F5: 1, because a NOAEL was established.

PDE = (0.05 mg/kg/day × 50 kg) / (5 × 10 × 1 × 1 × 1) = 2.5 / 50 = 0.05 mg/day = 50 micrograms per day.

Step 2: calculate the MACO into the next product

Product B is a tablet made on the same equipment.

• Minimum batch size of Product B: 100 kg = 100,000,000 mg.
• Maximum daily dose of Product B: 1,000 mg of tablets per day.

MACO = (50 micrograms/day × 100,000,000 mg) / 1,000 mg/day = 50 × 100,000 = 5,000,000 micrograms = 5 g.

So up to 5 g of Product A's active ingredient could, in theory, be spread through a 100 kg batch of Product B without any patient taking more than 50 micrograms a day of it.

Step 3: convert to a surface limit

Suppose the shared product-contact surface area of the equipment train is 50,000 cm² (5 m²).

SAL = 5,000,000 micrograms / 50,000 cm² = 100 micrograms per cm².

Step 4: convert to a swab limit

Suppose each swab covers 25 cm² and is extracted into 10 mL of solvent.

• Maximum amount per swab = 100 micrograms/cm² × 25 cm² = 2,500 micrograms.
• Maximum concentration in the extract = 2,500 micrograms / 10 mL = 250 micrograms per mL.

Step 5: account for recovery

Suppose recovery studies show the swab method recovers 80 percent of residue from stainless steel. A measured result therefore understates the true residue. One way to account for this is to divide each measured result by 0.80 before comparing it with the limit. Equivalently, the measured amount on a swab must not exceed 2,500 × 0.80 = 2,000 micrograms.

Step 6: compare with the traditional criteria

For comparison, assume Product A's minimum therapeutic dose is 10 mg/day.

• 1/1000 of dose criterion: allowable daily intake = 10 mg / 1,000 = 0.01 mg = 10 micrograms per day. MACO = (10 micrograms × 100,000,000 mg) / 1,000 mg = 1,000,000 micrograms = 1 g.
• 10 ppm criterion: 10 ppm of a 100,000,000 mg batch = 100,000,000 × 10 / 1,000,000 = 1,000 mg = 1 g.

In this hypothetical case the health-based limit is less strict than the traditional ones. ISPE training material notes that in most cases PDE values are above the old criteria, and that sometimes they are so high that visible residues could remain while the calculated limit is met. That is exactly why visual cleanliness remains a minimum requirement, and why the EMA says cleaning limits should not simply be set at the HBEL. For a highly potent product with a very low PDE, the opposite would happen: the health-based limit could be far stricter than 1/1000 of a dose, and possibly below what can be seen or even measured.

Step 7: set operating limits

In practice, the company would set its alert and action limits based on what its validated cleaning process consistently achieves, provided that these limits are well inside the 5 g MACO. If historical data show the process typically leaves well under 1 percent of the MACO, an alert limit far below the health-based limit would be appropriate and would detect a deteriorating process long before patients were at risk.

Sampling methods

Direct surface sampling (swabbing)

Swabbing means wiping a defined area of equipment surface with a swab moistened with a suitable solvent, then extracting the swab and analysing the extract. The FDA's 1993 guide describes direct surface sampling as the preferred method, because:

• areas that are hardest to clean and reasonably accessible can be sampled directly;
• results can be expressed as residue per unit area;
• residues that are dried on or insoluble can be physically removed by wiping.

The swab material and extraction solvent must not interfere with the analysis. Sampling locations should be chosen because they are worst cases: places where residue is most likely to remain, such as gaskets, valve seats, corners, discharge chutes and areas that are hard to reach. Annex 15 paragraph 10.11 requires the protocol to state the sampling locations and the rationale for choosing them.

Rinse sampling

Rinse sampling means passing a known volume of solvent or water over the equipment surfaces and analysing it. It can cover large areas and internal surfaces that cannot be reached with a swab, such as long pipes. Its weakness, according to the FDA guide, is that residues may be insoluble or physically trapped, so they do not dissolve into the rinse. The guide makes this point with an analogy: to check whether a dirty pot is clean, you inspect the pot, not the rinse water.

The FDA also warns that testing rinse water only against general water quality standards is not enough; the rinse must be tested for the actual contaminant. Reported summaries of PIC/S PI 006-4 note that rinse sampling tends to average out results, which is why swabbing is preferred where feasible.

Placebo sampling

Some firms have run a placebo batch through cleaned equipment and tested it for residue. The FDA guide lists significant limitations: residue may not be distributed evenly through the placebo (for example, residue on a blender discharge valve may end up concentrated in the first portion of the batch), larger particles may not disperse evenly, residue may not wear off surfaces uniformly, and dilution reduces sensitivity. Placebo methods should therefore be used only alongside swab and/or rinse sampling.

Visual inspection

Visual inspection is required but not sufficient on its own. Annex 15 calls a visual check "an important part of the acceptance criteria" but says "it is not generally acceptable for this criterion alone to be used." The 2018 EMA Q&A sets out detailed expectations where visual inspection is relied upon:

• establish the level at which a product residue becomes readily visible, taking into account real lighting conditions and viewing distances;
• inspect all product-contact surfaces where contamination may be held, including those requiring dismantling or tools such as mirrors, lights or borescopes;
• include non-product-contact surfaces that could shed residue into later batches;
• maintain written instructions and records confirming all inspections;
• train inspectors specifically, test their eyesight periodically, and prove competence by practical assessment.

Analytical testing at changeover

The 2018 EMA Q&A states that, even after cleaning validation is complete, "analytical testing is expected at each changeover unless justified otherwise" through a documented quality risk management process. That process must consider at least the repeatability of the cleaning (noting that manual cleaning is generally less repeatable than automated cleaning), the hazard posed by the product, and whether visual inspection can reliably detect residues at the HBEL-justified limit.

Analytical methods

Specific methods

The most direct approach is to measure the specific residue with a method such as high-performance liquid chromatography (HPLC), which separates and quantifies individual compounds. Specific methods are needed where the hazard is a particular active ingredient, and where degradants or by-products may matter, as in the 1992 import alert case. The FDA guide notes that bulk processes may contain unidentified partial reactants and by-products and that screening, for example by thin-layer chromatography, may be needed, particularly for potent compounds on shared equipment.

Non-specific methods

Annex 15 allows representative parameters where specific testing is not feasible, naming total organic carbon (TOC) and conductivity. TOC measures all carbon-containing material in a sample and was described for cleaning validation use in a 1996 article by Jenkins and colleagues in the PDA Journal of Pharmaceutical Science and Technology. Conductivity is useful for detecting ionic cleaning agents. Reported summaries of PIC/S PI 006-4 state that non-specific methods are acceptable with justification, on the conservative assumption that the whole result is attributable to the target residue.

The FDA guide also allows indirect tests such as conductivity for routine monitoring once a process is validated, provided the test has been shown to correlate with equipment cleanliness; during validation, uncleaned equipment must be shown to fail the test.

Method validation and recovery

An analytical method is only useful if it can detect residues at the relevant level and if the sampling actually picks them up. The FDA guide requires firms to establish the specificity and sensitivity of their methods and to test recovery from equipment surfaces, giving examples such as 50 percent or 90 percent recovery. It warns that a negative result "may also be the result of poor sampling technique," and so does not prove absence. Annex 15 paragraph 10.12 requires recovery to be shown "from all product contact materials sampled in the equipment with all the sampling methods used." Different surfaces, such as stainless steel, glass, plastics and gasket materials, can give very different recoveries.

Designing a cleaning validation programme

The lifecycle approach

Modern guidance treats cleaning validation as a lifecycle rather than a one-off exercise. Reported summaries of PIC/S PI 006-4 describe three stages: developing the cleaning process and identifying critical cleaning parameters; a validation study consisting of a predetermined number of cleaning verification assessments; and ongoing evaluation after the study is complete. ISPE's 2020 Cleaning Validation Lifecycle guide follows the same broad idea.

Worst-case and grouping approaches

Testing every product on every piece of equipment can be impractical, so firms often group products or equipment and test a worst case. Annex 15 paragraph 10.10 requires a scientific rationale for the worst-case product and an assessment of the impact of new products. It lists solubility, cleanability, toxicity and potency as possible criteria. Where similar equipment is grouped, paragraph 10.1 requires justification of the specific equipment chosen.

Hold times and campaigns

Residues become harder to remove the longer they dry. The FDA guide expects firms to control the time between the end of processing and cleaning. Annex 15 paragraph 10.8 requires defined "dirty hold times" (from end of manufacture to cleaning) and "clean hold times" (from cleaning to next use), the latter because clean equipment can become contaminated, including microbiologically, during storage. Where products are made in campaigns, paragraph 10.9 requires validation to be based on the maximum campaign length, since residue may build up over many batches.

Microbiological aspects

Cleaning must control microorganisms as well as chemicals. The FDA guide stresses drying equipment before storage and preventing stagnant water, and notes that good cleaning supports later sterilisation and pyrogen control. Annex 15 paragraph 10.7 requires microbial and endotoxin risks to be considered in protocols.

How many runs?

Annex 15 does not fix a number; it says the cleaning procedure should be performed "an appropriate number of times based on a risk assessment." One industry publication notes that, as a rule, the procedure must succeed three times in succession before it is considered validated, reflecting common practice. The justification should come from risk and from data on process variability.

Manual versus automated cleaning

Manual cleaning depends heavily on the operator and is generally less repeatable than automated clean-in-place systems. Annex 15 paragraph 10.15 requires the effectiveness of manual cleaning to be confirmed at a justified frequency, and the 2018 EMA Q&A lists repeatability as a key factor in deciding whether changeover testing can be reduced. For products with very low HBELs, reported summaries of PIC/S PI 006-4 say the necessary consistency may not be achievable manually, and dedicated parts, automation or changeover verification should be considered.

When to dedicate equipment

If cleaning cannot reliably achieve safe residue levels, equipment or facilities should be dedicated. Annex 15 paragraph 10.14 requires dedicated equipment or other measures where cleaning is ineffective or inappropriate, and the 2018 EMA Q&A says that where controls cannot consistently keep contamination below the HBEL, the products "should be manufactured in dedicated facilities." The Q&A also says simply grouping products of the same therapeutic class in a dedicated area does not solve the problem, because cross-contamination within the class can still harm patients.

Common pitfalls

Regulatory guidance identifies many recurring problems.

Testing until clean

Some manufacturers have repeatedly cleaned, sampled and retested equipment until a passing result appears. The FDA guide calls this "testing until clean" and says it is generally unacceptable for a validated process, except in rare cases, because repeated retesting shows that residues are present and that the cleaning process is not validated. Annex 15 paragraph 10.2 says the same: "Repeated cleaning and retesting until acceptable residue results are obtained is not considered an acceptable approach."

Testing only for the obvious residue

The 1992 import alert case showed the danger of testing only for the previous compound. Degradation products, reaction by-products and cleaning agents may all matter.

Ignoring cleaning agents

Detergents are residues too. The FDA guide notes that suppliers often do not disclose detergent composition, which makes residue evaluation harder, and expects detergent residues to be essentially absent after cleaning. It recommends choosing detergents that are easy to remove. Annex 15 paragraph 10.6 requires limits for cleaning agents.

Inadequate recovery studies

Sampling methods that recover only a fraction of residue, without correction, will underestimate contamination. Recovery must be demonstrated on each material of construction.

Relying on visual inspection alone

Visual cleanliness is necessary but not sufficient, and for potent products the HBEL-based limit may be below what can be seen.

Setting limits at the HBEL

Using the calculated MACO as the operating limit leaves no margin and ignores process capability. The EMA Q&A explicitly says the HBEL guideline is not intended for setting cleaning limits at that level.

Common misconceptions

"The FDA requires a 10 ppm limit"

It does not. The FDA's 1993 guide lists 10 ppm, 1/1000 of a dose and visual cleanliness as examples cited in industry literature, but it explicitly states that the FDA does not set specific limits. Each firm must justify its own.

"If it looks clean, it is clean"

Not necessarily. Highly potent residues may be harmful at levels well below visibility, and both Annex 15 and PIC/S guidance treat visual cleanliness as a minimum rather than the whole standard.

"Health-based limits are always stricter"

Not always. For many products the PDE-based MACO is higher than traditional limits, as the worked example shows. For highly potent or hazardous products it can be much lower. Its advantage is that it reflects actual hazard, not that it is always tighter.

"Once validated, always validated"

Cleaning validation is a lifecycle activity. Changes to products, equipment, detergents, batch sizes, procedures or toxicological knowledge can require re-evaluation, and the EMA Q&A notes that the data underlying an HBEL need periodic reassessment. The FDA guide expects procedures to say when revalidation is needed.

"Biologics need PDE limits for the intact protein"

Not always. Both the EMA guideline and Annex 15 recognise that proteins and peptides are usually degraded and inactivated by cleaning conditions, so a toxicological evaluation of the intact molecule may not be applicable, though other contamination routes must still be considered.

Frequently asked questions

What is the difference between cleaning validation and cleaning verification?

Validation is documented evidence that a cleaning procedure reliably removes residues below the acceptable level in general. Verification is the chemical analysis after a specific batch or campaign to confirm that this particular cleaning worked. EU Annex 15 defines both.

What is MACO?

Maximum allowable carryover: the largest total amount of the previous product's active ingredient that can be carried into the next product's batch without any patient exceeding the safe daily exposure. It is commonly calculated as PDE multiplied by the minimum batch size of the next product, divided by the maximum daily dose of the next product.

What is a PDE?

Permitted daily exposure: a substance-specific dose unlikely to cause any adverse effect if taken every day for a lifetime. Under the EMA guideline it is calculated from the NOAEL, adjusted for body weight and divided by uncertainty factors F1 to F5.

Are the 1/1000 dose and 10 ppm criteria still acceptable?

In Europe, the EMA's 2018 Q&A requires health-based exposure limits for all medicinal products, and traditional criteria are no longer accepted as the scientific basis for limits, though historical limits may be retained as alert limits if they keep exposure safely below the HBEL. PIC/S PI 006-4 has also moved to HBEL-based limits. In the United States, the FDA has never mandated a specific numerical limit.

How many cleaning runs are needed for validation?

Annex 15 requires a number based on risk assessment. Three consecutive successful runs are common industry practice, but the number should be justified.

Sources

• Cleaning validation (Wikipedia)
• FDA: Validation of Cleaning Processes (7/93), Guide to Inspections
• 21 CFR 211.67 Equipment cleaning and maintenance (Cornell LII)
• EMA: Guideline on setting health based exposure limits for use in risk identification in the manufacture of different medicinal products in shared facilities
• EMA: Guideline text, EMA/CHMP/CVMP/SWP/169430/2012 (PDF)
• EMA: Questions and answers on implementation of risk-based prevention of cross-contamination, EMA/CHMP/CVMP/SWP/246844/2018 (PDF)
• European Commission: EudraLex Volume 4, Annex 15 Qualification and Validation (2015)
• GMP Insiders: PIC/S issues PI 006-4 on Qualification and Validation
• ECA: PIC/S PI 006-3 (September 2007)
• Pierre Devaux, "Defining Limits and Doing MACO Calculations", ISPE Nordic (PDF)
• GMP Publishing: Basis of cleaning validation, setting of PDE limits
• EMA consultation comments on the Q&A (PDF)